Yoshifusa Aizawa

<b><i>Background and Objectives:</i></b> There is a substantial body of evidence assessing the effects of equine-assisted therapy on physiological and psychological aspects of individuals with disabilities. This study aimed to evaluate the physiological benefits of this alternative therapy for children with cerebral palsy (CP) by means of a systematic review and meta-analysis. <b><i>Methods:</i></b> This systematic review included all randomized and nonrandomized clinical trials of hippotherapy (HT), therapeutic horse riding (THR), and artificial saddle (AS) for the treatment of children with CP by a systematic search in Medline, Embase, Cochrane Library, and other databases up to November 2012. Articles were assessed for inclusion eligibility and quality by two independent reviewers. Any discordant case was re-reviewed and consensus was obtained after sufficient discussion. A random effects model of meta-analysis was applied to provide summary statistics for each outcome. <b><i>Results:</i></b> Seven randomized controlled trials (RCTs), 4 non-RCTs, and 7 self-controlled studies were included for quality assessment. Ten studies assessed the effect of HT, 5 evaluated THR, and 3 evaluated AS. The sample size differed from 3 to 72, and the quality ranged from low to moderate. Six studies were included in the meta-analysis, and there was a significant improvement in the 66-item Gross Motor Function Measure (GMFM-66), the GMFM-66/88 total score, and the dimension E of the GMFM. Although the asymmetry score tended to be reduced, it failed to reach statistical significance. <b><i>Conclusions:</i></b> HT, THR, and AS seem to improve the total score of the gross motor function via improvement of the walking, running, and jumping dimension. However, they are not likely to be of benefit to the symmetry of postural muscle activity. Studies included in this review lack high-quality RCTs with a sufficient number of subjects, which thus warrants further evaluations of these modalities using large-scale well-designed RCTs.

Early repolarization is a common electrocardiographic finding that is generally considered to be benign. Its potential to cause cardiac arrhythmias has been hypothesized from experimental studies, but it is not known whether there is a clinical association with sudden cardiac arrest.We reviewed data from 206 case subjects at 22 centers who were resuscitated after cardiac arrest due to idiopathic ventricular fibrillation and assessed the prevalence of electrocardiographic early repolarization. The latter was defined as an elevation of the QRS-ST junction of at least 0.1 mV from baseline in the inferior or lateral lead, manifested as QRS slurring or notching. The control group comprised 412 subjects without heart disease who were matched for age, sex, race, and level of physical activity. Follow-up data that included the results of monitoring with an implantable defibrillator were obtained for all case subjects.Early repolarization was more frequent in case subjects with idiopathic ventricular fibrillation than in control subjects (31% vs. 5%, P<0.001). Among case subjects, those with early repolarization were more likely to be male and to have a history of syncope or sudden cardiac arrest during sleep than those without early repolarization. In eight subjects, the origin of ectopy that initiated ventricular arrhythmias was mapped to sites concordant with the localization of repolarization abnormalities. During a mean (+/-SD) follow-up of 61+/-50 months, defibrillator monitoring showed a higher incidence of recurrent ventricular fibrillation in case subjects with a repolarization abnormality than in those without such an abnormality (hazard ratio, 2.1; 95% confidence interval, 1.2 to 3.5; P=0.008).Among patients with a history of idiopathic ventricular fibrillation, there is an increased prevalence of early repolarization.

Cardiac ion channelopathies are responsible for an ever-increasing number and diversity of familial cardiac arrhythmia syndromes. We describe a new clinical entity that consists of an ST-segment elevation in the right precordial ECG leads, a shorter-than-normal QT interval, and a history of sudden cardiac death.Eighty-two consecutive probands with Brugada syndrome were screened for ion channel gene mutations with direct sequencing. Site-directed mutagenesis was performed, and CHO-K1 cells were cotransfected with cDNAs encoding wild-type or mutant CACNB2b (Ca(v beta2b)), CACNA2D1 (Ca(v alpha2delta1)), and CACNA1C tagged with enhanced yellow fluorescent protein (Ca(v)1.2). Whole-cell patch-clamp studies were performed after 48 to 72 hours. Three probands displaying ST-segment elevation and corrected QT intervals < or = 360 ms had mutations in genes encoding the cardiac L-type calcium channel. Corrected QT ranged from 330 to 370 ms among probands and clinically affected family members. Rate adaptation of QT interval was reduced. Quinidine normalized the QT interval and prevented stimulation-induced ventricular tachycardia. Genetic and heterologous expression studies revealed loss-of-function missense mutations in CACNA1C (A39V and G490R) and CACNB2 (S481L) encoding the alpha1- and beta2b-subunits of the L-type calcium channel. Confocal microscopy revealed a defect in trafficking of A39V Ca(v)1.2 channels but normal trafficking of channels containing G490R Ca(v)1.2 or S481L Ca(v beta2b)-subunits.This is the first report of loss-of-function mutations in genes encoding the cardiac L-type calcium channel to be associated with a familial sudden cardiac death syndrome in which a Brugada syndrome phenotype is combined with shorter-than-normal QT intervals.

We examined the modulatory effects of autonomic nervous system and antiarrhythmic drugs on the ST segment in patients with Brugada syndrome to gain an insight into the mechanism of ST segment elevation.Right bundle branch block, ST segment elevation and ventricular tachyarrhythmias define a distinct clinical and electrocardiographic (ECG) syndrome (Brugada syndrome). However, the mechanism of ST segment elevation and the causes of this syndrome are unknown.The study included four patients in whom structural heart or coronary artery disease was excluded by noninvasive and invasive tests. High take-off ST segment elevation of either the coved or saddle-back type in precordial leads V1, V2 and V3 was seen in all patients. Three patients experienced recurrent episodes of syncope or aborted sudden cardiac death, and the remaining patient had palpitation. Autonomic receptor stimulation and blockade and intravenous administration of antiarrhythmic drugs were performed during sinus rhythm while the 12-lead ECG was recorded. Metaiodobenzylguanidine (MIBG) scanning and Holter monitoring were also performed.Beta-adrenoceptor stimulation by intravenous isoproterenol consistently reduced (> or = 0.1 mV) ST segment elevation at or 80 ms after the J point in all four patients. Selective alpha-adrenoceptor stimulation by intravenous norepinephrine in the presence of propranolol or by intravenous methoxamine consistently augmented, whereas alpha-adrenoceptor blockade reduced, ST segment elevation in three patients. Intracoronary acetylcholine or intravenous edrophonium or neostigmine augmented ST segment elevation without inducing coronary spasm in three of four patients. Class IA antiarrhythmic drugs also consistently augmented (three patients), whereas class IB drugs had no effect on (two patients) ST segment elevation. No abnormality was found on MIBG imaging or heart rate variability in three patients, suggesting that autonomic dysfunction is not a primary disease process. Class IA drugs had no effect on ST segment in three control patients, suggesting that the ST segment elevation seen in patients with Brugada syndrome in response to the drugs is not a nonspecific response.ST segment elevation in patients with Brugada syndrome was augmented by selective stimulation of alpha-adrenoceptors or muscarinic receptors or by class IA drugs but was mitigated by beta-adrenoceptor stimulation or alpha-adrenoceptor blockade. These responses might be explained by postulating the presence of an area of early repolarization or a local "depolarized" area in the ventricle causing ST segment elevation in this syndrome. Because only a small number of patients were studied, these possibilities need further evaluation.

The metabolic syndrome consists of a cluster of atherosclerotic risk factors, many of which also have been implicated in the genesis of atrial fibrillation (AF). However, the precise role of the metabolic syndrome in the development of AF is unknown.This prospective, community-based, observational cohort study was based on an annual health check-up program in Japan. We studied 28 449 participants without baseline AF. We used 2 different criteria for the metabolic syndrome--the guidelines of the National Cholesterol Education Program Third Adult Treatment Panel (NCEP-ATP III) and those of the American Heart Association/National Heart, Lung, and Blood Institute (AHA/NHLBI)--to study the risk of development of new-onset AF. The metabolic syndrome was present in 3716 subjects (13%) and 4544 subjects (16%) using the NCEP-ATP III and AHA/NHLBI definitions, respectively. During a mean follow-up of 4.5 years, AF developed in 265 subjects (105 women). Among the metabolic syndrome components, obesity (age- and sex-adjusted hazard ratio [HR], 1.64), elevated blood pressure (HR, 1.69), low high-density lipoprotein cholesterol (HR, 1.52), and impaired fasting glucose [corrected] (HR, 1.44 [NCEP-ATP III] and 1.35 [AHA/NHLBI]) showed an increased risk for AF. The association between the metabolic syndrome and AF remained significant in subjects without treated hypertension or diabetes by the NCEP-ATP III definition (HR, 1.78) but not by the AHA/NHLBI definition (HR, 1.28).The metabolic syndrome was associated with increased risk of AF. The metabolic derangements of the syndrome may be important in the pathogenesis of AF.

BackgroundThe mortality and morbidity rates of various cardiovascular diseases differ between Western countries and Japan. The age- and gender-specific prevalence rate of atrial fibrillation (AF) in the general population of Japan was determined using the data from periodic health examinations in 2003.MethodsData of 630,138 subjects aged 40 years or more (47% were men and 34% were employees of companies and local governments) were collected from northern to southern Japan. The prevalence of diagnosed AF in each 10-year age group of both men and women was determined. Based on these prevalence rates and the Registry of Residents, the number of people having AF in Japan was estimated.ResultsThe prevalence rate of AF increased as both male and female subjects aged, and it was 4.4% for men but only 2.2% for women aged 80 years or more (p<0.0001). As a whole, the AF prevalence of men was three times that of women (1.35 versus 0.43%, p<0.0001). There may be approximately 716,000 people (95% confidence interval (CI), 711,000–720,000) with AF in Japan, an overall prevalence of 0.56%. The number of people having AF was projected to be 1.034 (95% CI, 1.029–1.039) million, an overall prevalence of 1.09%, in 2050.ConclusionsThe prevalence of AF increased in Japan as the population aged, as in Western countries. The overall prevalence of AF in Japan is approximately two-thirds of that in the USA. The projected increase in the number of people having AF is modest in Japan in 2050.

The future burden of heart failure in Japan was projected to 2055 in order to prospectively estimate of the number of these patients.The statistics are based on prevalence data of left ventricular dysfunction (LVD) in Sado City using the Sado Heart Failure Study (2003) and population estimates from the Japanese National Institute of Population and Social Security Research Report (2006). The number of Japanese outpatients with LVD was 979,000 in 2005, and is predicted to increase gradually as the population ages, reaching 1.3 million by 2030.LVD is expected to precipitate a future epidemic of heart failure in Japan.

Afferent and efferent cardiac neurotransmission via the cardiac nerves intricately modulates nearly all physiological functions of the heart (chronotropy, dromotropy, lusitropy, and inotropy). Afferent information from the heart is transmitted to higher levels of the nervous system for processing (intrinsic cardiac nervous system, extracardiac-intrathoracic ganglia, spinal cord, brain stem, and higher centers), which ultimately results in efferent cardiomotor neural impulses (via the sympathetic and parasympathetic nerves). This system forms interacting feedback loops that provide physiological stability for maintaining normal rhythm and life-sustaining circulation. This system also ensures that there is fine-tuned regulation of sympathetic–parasympathetic balance in the heart under normal and stressed states in the short (beat to beat), intermediate (minutes to hours), and long term (days to years). This important neurovisceral/autonomic nervous system also plays a major role in the pathophysiology and progression of heart disease, including heart failure and arrhythmias leading to sudden cardiac death. Transdifferentiation of neurons in heart failure, functional denervation, cardiac and extracardiac neural remodeling has also been identified and characterized during the progression of disease. Recent advances in understanding the cellular and molecular processes governing innervation and the functional control of the myocardium in health and disease provide a rational mechanistic basis for the development of neuraxial therapies for preventing sudden cardiac death and other arrhythmias. Advances in cellular, molecular, and bioengineering realms have underscored the emergence of this area as an important avenue of scientific inquiry and therapeutic intervention.

Our purpose was to evaluate the efficacy of antiarrhythmic drugs (AADs) in recurrent ventricular fibrillation (VF) associated with inferolateral early repolarization pattern on the electrocardiogram. Although an implantable cardioverter-defibrillator is the treatment of choice, additional AADs may be necessary to prevent frequent episodes of VF and reduce implantable cardioverter-defibrillator shock burden or as a lifesaving therapy in electrical storms. From a multicenter cohort of 122 patients (90 male subjects, age 37 ± 12 years) with idiopathic VF and early repolarization abnormality in the inferolateral leads, we selected all patients with more than 3 episodes of VF (multiple) including those with electrical storms (≥3 VF in 24 h). The choice of AAD was decided by individual physicians. Follow-up data were obtained for all patients using monitoring with implantable defibrillator. Successful oral AAD was defined as elimination of all recurrences of VF with a minimal follow-up period of 12 months. Multiple episodes of VF were observed in 33 (27%) patients. Electrical storms (34 ± 47 episodes) occurred in 16 and were unresponsive to beta-blockers (11 of 11), lidocaine/mexiletine (9 of 9), and verapamil (3 of 3), while amiodarone was partially effective (3 of 10). In contrast, isoproterenol infusion immediately suppressed electrical storms in 7 of 7 patients. Over a follow-up of 69 ± 58 months, oral AADs were poorly effective in preventing recurrent VF: beta-blockers (2 of 16), verapamil (0 of 4), mexiletine (0 of 4), amiodarone (1 of 7), and class 1C AADs (2 of 9). Quinidine was successful in 9 of 9 patients, decreasing recurrent VF from 33 ± 35 episodes to nil for 25 ± 18 months. In addition, quinidine restored a normal electrocardiogram. Multiple recurrences of VF occurred in 27% of patients with early repolarization abnormality and may be life threatening. Isoproterenol in acute cases and quinidine in chronic cases are effective AADs.

Atrial fibrillation (AF) and chronic kidney disease share risk factors and pathophysiologic mechanisms, suggesting that two conditions have close relationships. This is a prospective community-based observational cohort study including 235,818 subjects based upon a voluntary annual health check-up program in Japan. We studied the association of kidney dysfunction at entry with subsequent new-onset AF and the association of AF at entry with the development of kidney disease. During a follow-up of 5.9 ± 2.4 years, AF developed in 2947 subjects (1.3%). Baseline serum creatinine and estimated glomerular filtration rate (GFR) were associated with risk of subsequent AF. The HRs (95% CI) for AF were 1.32 (1.08-1.62) and 1.57 (0.89-2.77) for GFR 30 to 59 and <30 mL/min per 1.73 m2, respectively. The effect of kidney disease on risk of new-onset AF remained significant in subjects without treated hypertension or diabetes. During the follow-up, 7791 subjects (3.3%) developed kidney dysfunction (GFR <60 mL/min per 1.73 m2), and 11 307 subjects (4.9%) developed proteinuria. Atrial fibrillation at entry was associated with development of kidney dysfunction (HRs [95% CI], 1.77 [1.50-2.10]) and proteinuria (HR [95% CI], 2.20 [1.92-2.52]). The association persisted in subjects without treated hypertension or diabetes. Kidney dysfunction increased the risk of new onset of AF, and AF increased the risk of development of kidney disease. This finding supports the concept that the two conditions share common abnormal molecular signaling pathways contributing to their pathogenesis.

The prognosis of patients with saddleback or noncoved type (non-type 1) ST-elevation in Brugada syndrome is unknown. The purpose of this study was to clarify the long-term prognosis of probands with non-type 1 ECG and those with coved (type 1) Brugada-pattern ECG.A total of 330 (123 symptomatic, 207 asymptomatic) probands with a coved or saddleback ST-elevation > or = 1 mm in leads V(1)-V(3) were divided into 2 ECG groups-type 1 (245 probands) and non-type 1 (85 probands)-and were prospectively followed for 48.7+/-15.0 months. The absence of type 1 ECG was confirmed by drug provocation test and multiple recordings. The ratio of individuals with a family history of sudden cardiac death (14%) was lower than previous studies. Clinical profiles and outcomes were not notably different between the 2 groups (annual arrhythmic event rate of probands with ventricular fibrillation; type 1: 10.2%, non-type 1: 10.6%, probands with syncope; type 1: 0.6%, non-type 1: 1.2%, and asymptomatic probands; type 1: 0.5%, non-type 1: 0%). Family history of sudden cardiac death at age <45 years and coexistence of inferolateral early repolarization with Brugada-pattern ECG were independent predictors of fatal arrhythmic events (hazard ratio, 3.28; 95% confidence interval, 1.42 to 7.60; P=0.005; hazard ratio, 2.66; 95% confidence interval, 1.06 to 6.71; P=0.03, respectively, by multivariate analysis), although spontaneous type 1 ECG and ventricular fibrillation inducibility by electrophysiological study were not reliable parameters.The long-term prognosis of probands in non-type 1 group was similar to that of type 1 group. Family history of sudden cardiac death and the presence of early repolarization were predictors of poor outcome in this study, which included only probands with Brugada-pattern ST-elevation.

Glycosuria produced by sodium–glucose co-transporter-2 (SGLT-2) inhibitors is associated with weight loss. SGLT-2 inhibitors reportedly might reduce the occurrence of cardiovascular events. Epicardial adipose tissue (EAT) is a pathogenic fat depot that may be associated with coronary atherosclerosis. The present study evaluated the relationship between an SGLT-2 inhibitor (dapagliflozin) and EAT volume. In 40 diabetes mellitus patients with coronary artery disease (10 women and 30 men; mean age of all 40 patients was 67.2 ± 5.4 years), EAT volume was compared prospectively between the dapagliflozin treatment group (DG; n = 20) and conventional treatment group (CTG; n = 20) during a 6-month period. EAT was defined as any pixel that had computed tomography attenuation of − 150 to − 30 Hounsfield units within the pericardial sac. Metabolic parameters, including HbA1c, tumor necrotic factor-α (TNF-α), and plasminogen activator inhibitor-1 (PAI-1) levels, were measured at both baseline and 6-months thereafter. There were no significant differences at baseline of EAT volume and HbA1c, PAI-1, and TNF-α levels between the two treatment groups. After a 6-month follow-up, the change in HbA1c levels in the DG decreased significantly from 7.2 to 6.8%, while body weight decreased significantly in the DG compared with the CTG (− 2.9 ± 3.4 vs. 0.2 ± 2.4 kg, p = 0.01). At the 6-month follow-up, serum PAI-1 levels tended to decline in the DG. In addition, the change in the TNF-α level in the DG was significantly greater than that in the CTG (− 0.5 ± 0.7 vs. 0.03 ± 0.3 pg/ml, p = 0.03). Furthermore, EAT volume significantly decreased in the DG at the 6-month follow-up compared with the CTG (− 16.4 ± 8.3 vs. 4.7 ± 8.8 cm3, p = 0.01). Not only the changes in the EAT volume and body weight, but also those in the EAT volume and TNF-α level, showed significantly positive correlation. Treatment with dapagliflozin might improve systemic metabolic parameters and decrease the EAT volume in diabetes mellitus patients, possibly contributing to risk reduction in cardiovascular events.

The genotype-phenotype correlation of SCN5A mutations as a predictor of cardiac events in Brugada syndrome remains controversial. We aimed to establish a registry limited to probands, with a long follow-up period, so that the genotype-phenotype correlation of SCN5A mutations in Brugada syndrome can be examined without patient selection bias.This multicenter registry enrolled 415 probands (n=403; men, 97%; age, 46±14 years) diagnosed with Brugada syndrome whose SCN5A gene was analyzed for mutations.During a mean follow-up period of 72 months, the overall cardiac event rate was 2.5%/y. In comparison with probands without mutations (SCN5A (-), n=355), probands with SCN5A mutations (SCN5A (+), n=60) experienced their first cardiac event at a younger age (34 versus 42 years, P=0.013), had a higher positive rate of late potentials (89% versus 73%, P=0.016), exhibited longer P-wave, PQ, and QRS durations, and had a higher rate of cardiac events (P=0.017 by log-rank). Multivariate analysis indicated that only SCN5A mutation and history of aborted cardiac arrest were significant predictors of cardiac events (SCN5A (+) versus SCN5A (-): hazard ratio, 2.0 and P=0.045; history of aborted cardiac arrest versus no such history: hazard ratio, 6.5 and P<0.001).Brugada syndrome patients with SCN5A mutations exhibit more conduction abnormalities on ECG and have higher risk for cardiac events.

The original guideline for non-pharmacological treatments (cardiac implantable electronic device, catheter ablation, and arrhythmia surgery) of arrhythmias (Japanese Circulation Society [JCS] Guideline on Non-pharmacotherapy of Cardiac Arrhythmias) was first published in 2001, and there have been two revisions thereafter (2006 and 2011). The “JCS Guideline on Indications and Procedures for Catheter Ablation” was published in 2012 to cover the rapid development and expansion of catheter ablation techniques. Advances in non-pharmacological treatment of arrhythmia have further accelerated since then, with the succeeding emergence of new functions, usefulness, and evidence. Against the background of these remarkable developments, the guidelines needed to undergo many changes and revisions. Therefore, the format has been revised again to include cardiac implantable electronic devices and catheter ablation therapies. Since 2011, there has been a succession of innovative devices and treatment methods, such as (1) implantable cardiac monitoring, (2) subcutaneous implantable cardioverter-defibrillators, (3) wearable cardioverter-defibrillators, (4) remote monitoring, (5) magnetic resonance imaging-compatible devices, (6) leadless pacemakers, (7) balloon technology for pulmonary vein isolation, (8) percutaneous lead extraction, and (9) left atrial appendage closure devices. Thus, this revision needed to add new sections to accommodate these developments. In addition, new content on existing treatment methods has been added as much as possible, including hardware improvements, new evidence, and the challenge of reducing radiation exposure. Selecting and summarizing suitable information from the vast amount of available data within a limited space could have been a daunting task for all team members; however, our efforts have culminated in this guideline revision containing carefully selected and essential information, thanks to everyone who collaborated on this project. Two related guidelines (JCS Guideline on Treatment of Acute and Chronic Heart Failure, and JCS Guideline on Treatment of Genetic Arrhythmia) were each revised in 2018.1, 2 Some working group members participated in the revision of both guidelines, as team members or observers, thus ensuring consistency between the guidelines. Non-pharmacotherapy in the broad sense includes external cardioversion for atrial fibrillation and sustained ventricular tachyarrhythmias, temporary intravenous pacing, and percutaneous pacing. However, for details of these therapies, refer to the JCS Guidelines on Pharmacotherapy of Atrial Fibrillation3 and the Japan Resuscitation Council Guidelines.4 Pacemaker treatment for bradyarrhythmia was first approved for national health insurance coverage in Japan in 1974, and its use rapidly became widespread thereafter. Approximately 40 years later, in 2017, the number of patients treated with this technology has increased to 60 137 (41 895 new cases and approximately 18 242 replacements).5 Capsule-shaped leadless pacemaker also became available in 2016, and this technology is being established as a new option. Non-pharmacological treatment of tachy-arrhythmia began in 1969 from when Will C. Sealy performed surgery in patients with Wolff-Parkinson-White (WPW) syndrome (Figure 1). Since then, the application of surgical treatment has expanded to conditions such as ventricular tachycardia (VT) and atrial fibrillation (AF), and surgery has been the pioneer of radical therapy for tachyarrhythmias. At the present time, many surgical methods have been replaced by catheter ablation; however, surgical treatment still remains an indispensable option for patients with a tachycardia resistant to other medical treatments. As with surgical treatment, catheter ablation was initially performed for supraventricular tachycardias such as WPW syndrome. However, the revolutionary discovery of pulmonary vein isolation (PVI) for AF and the advent of three-dimensional (3D) navigation systems have subsequently resulted in a tremendous increase in the number of cases treated with catheter ablation. In 2016, >74 000 catheter ablation procedures were performed in Japan, of which >45 000 were implemented for AF.6 In 2015, a PVI method using cryoballoon ablation was introduced in Japan. Later, new techniques such as hot balloon or laser balloon (endoscopic systems using laser irradiation) ablation technologies entered the market, and safer and easier treatment methods are currently being established. In addition, prevention of systemic embolism using a left atrial appendage closure (LAAC) device is being established as a breakthrough treatment for AF patients who have difficulty in continuing anticoagulation therapy. Because early implantable cardioverter-defibrillators (ICDs) were highly invasive owing to the requirement for a thoracotomy, the indications for this treatment were highly limited. However, the development of transvenous leads, the discovery of the biphasic shock method, and a reduction in both the size and weight of the generator have now enabled implantation using the same technique as for pacemakers, which has contributed to expanding the application of ICDs to primary prevention. In 2017, approximately 6691 devices (4288 new cases, 2403 replacements) were implanted in Japan.5 The subcutaneous implantable cardioverter-defibrillator (S-ICD) was developed in 2015, and its clinical usage is progressing. In 2004, cardiac resynchronization therapy (CRT) became available for patients with impaired cardiac function, and the usefulness of this treatment has been verified. especially in heart failure patients with complete left bundle branch block in Japan. Because patients with heart failure have a high risk of sudden death, an ICD with a biventricular pacing function (CRT defibrillator [CRT-D]) was also developed and approved in 2006. In 2017, CRT was newly administered in 3321 patients in Japan, 2399 of whom (72%) received the CRT-D, demonstrating that the treatment has been actively applied to prevent sudden death.5 A wearable cardioverter-defibrillator (WCD) was introduced in Japan in 2015, which can be used for candidates for ICD therapy as a bridge treatment until application of an ICD is possible. Many devices are also equipped with a remote monitoring function, which sends most of the biological information and device data to the medical facility while the patient is staying at home by enabling the early detection of abnormal findings. As described above, there have been remarkable developments in the non-pharmacotherapy of arrhythmias. However, problems still remain, including (1) the risk of complications associated with aging of patients, (2) the requirement for high-quality training of specialists and medical staff to enable them to handle the expanding indications and diversifying treatment methods, (3) the overflow of information and increasing complexity of management because of the sophisticated and multiple functionalities of the devices, and (4) the impact of the expanding indications of expensive devices in the setting of limited medical resources. In the future, it will be necessary to formulate evidence unique to Japan on the extent to which cutting-edge non-pharmacological treatments for arrhythmias improve the prognosis of patients. This guideline recommends indications for non-pharmacotherapy of arrhythmia based on the latest findings and evidence. There is an increasing variety of non-pharmacotherapies, and extensive progress is being made in this field. This guideline contains information on conventional cardiac implantable electronic devices (CIEDs), such as pacemakers, ICDs, and ICDs with biventricular pacing function, as well as new information on remote monitoring, magnetic resonance imaging-conditional CIEDs, leadless pacemakers, percutaneous lead extraction, implantable monitors, S-ICDs, and WCDs. Information on catheter ablation includes radiation exposure, new 3D mapping systems, balloon ablation for AF, bipolar ablation, and chemical ablation. In addition, this guideline discusses the LAAC device for the first time, which is not a treatment for arrhythmia itself but for preventing thromboembolism – a serious problem associated with AF. Non-pharmacotherapy of arrhythmia is expected to increase in the future, so there is a need to standardize all non-pharmacotherapy processes, including not only treatment indications but also their theoretical background, recommended procedures, necessary equipment and implementation system, and precautions that have to be taken before and after the procedure. The indications of non-pharmacological treatments of tachyarrhythmia in children differ from those in adults, so there are many cautionary points to note. Therefore, CIEDs and catheter ablation for children are described under independent chapters, as in previous guidelines. The information on surgical treatment for arrhythmia mainly focuses on surgical treatment for AF and VT. Surgery for supraventricular tachycardia has been omitted from this guideline because the number of surgical procedures has dramatically decreased in recent years. Nevertheless, surgery is still indicated for some patients with supraventricular tachycardias, including those with unsuccessful ablation. The aim of this guideline is to clarify the indications, results, and complications of non-pharmacological treatments for arrhythmias such as bradyarrhythmia, supraventricular tachycardia, AF, premature ventricular contractions, VT, and ventricular fibrillation, as well as treatment for the associated heart failure and thromboembolism. We are striving for standardized treatment by explicitly describing the procedures. Specific information on the procedures is also included, such as the knowledge, equipment, and doctor/facility conditions required to perform the procedure. The guideline has been created based on evidence and consensus at the time of publication and should be updated over time. This guideline describes the recommended indications and procedures as of 2018. Future technological advances will further expand the indications for non-pharmacotherapy of arrhythmia and make the procedures more reliable and convenient. This guideline is designed to be used as a reference by doctors diagnosing and treating diseases in clinical practice, and the final decision should be made by the attending physicians after ascertaining the patient's condition. Even when selecting a diagnosis or treatment that does not follow the guideline, the decision of the attending physicians should be prioritized in consideration of the individual patient's situation. In actual clinical settings, it is most important for the attending physicians to make the judgment after fully considering the clinical background and social situation of each patient while complying with the guideline. For this guideline, we first surveyed materials based on evidence from the USA and Europe, then further critically examined the level of evidence, collected information available in Japan, and examined all materials based on the experiences and opinions of members and collaborators in the joint working group. The recommendation classes and evidence levels used in this guideline conform to those of the American Heart Association (AHA), American College of Cardiology (ACC), and Heart Rhythm Society (HRS) guidelines.7 The recommendation class of indications for each diagnosis and treatment method is classified as I, IIa, IIb, and III, and the level of evidence is classified into levels A, B, and C (Tables 1,2). The guideline also states the class of recommendation and level of evidence based on the “MINDS Handbook for Clinical Practice Guideline Development 2007”,8 published by the Medical Information Network Distribution Service (MINDS) Evidence-based Medicine dissemination promotion project as a guideline preparation method (Tables 3,4). The MINDS grades of recommendation are comprehensively determined, taking into account the following factors: (1) level of evidence, (2) amount and variation of evidence, (3) extent of clinical effectiveness, (4) clinical applicability (physician ability, regional characteristics, medical resources, insurance system, etc), and (5) evidence on harm and cost. Not recommended as evidence indicates that the treatment is ineffective or even harmful The MINDS level of evidence (levels of evidence in literature on treatment) is a classification based on research design, and the highest level was adopted when multiple papers were considered. This guideline describes both the conventional AHA/ACC/HRS guideline classifications and the MINDS classification, whenever possible, for the content of each diagnosis and treatment. However, the MINDS grade of recommendation and level of evidence should be used only as a reference, as this system regards the evidence level in a fundamentally different manner. This revision adds new knowledge acquired from advances in diagnostic techniques and treatment methods, or recently reported important evidence, while considering consistency with each of the previously reported guidelines published by the JCS Joint Working Group. Pacemaker therapy for bradyarrhythmia became covered by insurance in Japan in 1974. Initially, pacemakers only had ventricular pacing function; however, at present, pacemakers with functions such as maintaining atrioventricular synchrony with dual-chamber pacing modes, as well as monitoring atrioventricular conductivity to suppress right ventricular pacing, have been developed, which has contributed to improving patients’ prognosis. In 2017, a capsule-type leadless pacemaker appeared on the market, and its clinical application is progressing. An implantable cardioverter-defibrillator (ICD) was introduced in Japan in 1996 to treat fatal arrhythmias (ventricular tachycardia [VT]/ventricular fibrillation [VF]). Early ICDs were a highly invasive treatment requiring a thoracotomy, severely limiting their indications. However, the subsequent development of transvenous leads, discovery of the biphasic shock method, and reduction in the size and weight of the generator enabled use of the same technique as that for pacemaker implantation, which has greatly contributed to the expansion of indications for improving life prognosis and for primary prevention. The year 2000 saw the appearance of the dual-chamber ICD, which contributed to a dramatic improvement in pacing function during bradycardia, equivalent to that of a pacemaker for bradycardia, and a diagnostic algorithm based on atrial signal detection. Furthermore, the subcutaneous ICD (S-ICD) was introduced in 2015 and has been actively used for patients without venous access and/or those who do not require pacing functions. In 2004, biventricular pacing, or cardiac resynchronized therapy (CRT), became available for patients with impaired cardiac function, and its utility has been confirmed, especially in patients with heart failure who have desynchronous contractions due to complete left bundle branch block (CLBBB). As patients with heart failure are at a high risk of sudden death, an ICD with a biventricular pacing function (CRT-D) was developed and approved in 2006. Improved pacing-site selectivity with quadrupolar left ventricular leads, functions utilizing self-right bundle conduction, and multipoint left ventricular pacing have been applied in clinical practice and may reduce the number of non-responders. As of 2017, CRT-D has been indicated for 72% of new cardiac resynchronization therapy (CRT) cases in Japan; thus, aggressive prevention of sudden death is ongoing.5 ICDs have limited effectiveness after an acute myocardial infarction and at the early stage after the diagnosis of heart failure. In 2015, wearable cardioverter-defibrillators (WCDs) came into use to prevent sudden death during the waiting period while the indications for ICD are being determined. WCD is also used as a bridging treatment until the next implantation for patients whose ICD has been removed because of infection or other reasons. Many devices are equipped with a remote monitoring function, which now enables the early detection of abnormal findings related to device functions and biological characteristics. Furthermore, although their use is conditional, magnetic resonance imaging (MRI)-compatible devices has become to be recognized as ordinary function, which is particularly useful in Japan where the rate of installation and using MRI are high. Non-pharmacological treatment of arrhythmia requires advanced medical technology, and progress is rapid in this field. Physician and facility requirements are extremely important for the application of this guideline. This section describes the current facility standards and practitioner standards; however, as these may be revised in the future, refer to the Japanese Circulation Society or Japanese Heart Rhythm Society (HRS) websites to obtain the latest information. Each non-pharmacotherapy must be applied effectively and safely, and a system for responding to emergencies (human resource development, establishment of a team medical system, use of fully maintained equipment) is required. The following institutional standards and practitioner standards were proposed by the Japanese HRS in 2017, considering the novelty of the leadless pacemaker and that this procedure requires cardiac access via the femoral vein using a large sheath9 (see “3.7 Leadless pacemakers” in this chapter). Facility standards Practitioner standards The following practitioner standards were proposed by the Japanese HRS in 2016 for S-ICD implantation.11 Practitioner standards Clinical use of a WCD requires appropriate selection of cases and understanding of the equipment, and the following practitioner standards were proposed by the Japanese HRS in 2017.12 Practitioner standards ICMs may be implanted in any facility that satisfies the facility standards for pacemakers, ICD, or CRT-P/CRT-D as a condition for insurance application (based on the 2015 Medical Fee Points Table). When deciding on the indications for treatments that require advanced medical technology such as CIEDs, it is essential that the patient provides voluntary consent after receiving sufficient information. The information should be provided using words that the patient can understand, pursuant to the provisions of Chapter 1, Article 1-4, paragraph 2 of the Medical Care Act: “In the delivery of medical care, a physician, dentist, pharmacist, nurse, or other medical care professional shall give appropriate explanations and endeavor to foster understanding in the recipients of medical care.” The content of the explanation is based on the judgment according to the knowledge and experience of each doctor; however, it is necessary to provide the following information to the patient: (1) information on the disease (type and severity of arrhythmia, underlying heart disease, etc); (2) aim and details of the treatment (including device model and manufacturer name), therapeutic effect and success rate, complications (types, severity, and incidence) during the acute phase and during long-term follow-up (requires not only general information but also information on the performance in the facility in question), and the reason for selecting the treatment; (3) treatments other than the treatment in question (pharmacotherapies, other non-pharmacotherapies [including treatment available at other facilities]) and the therapeutic effect of those treatments; (4) expected results with monitoring alone without the treatment in question (predicted outcome and probability thereof); (5) positioning of the treatment in question for various arrythmias and possible unexpected complications (short-term and long-term); (6) cost of the treatment (including the cost of this treatment and other treatments); and (7) assurance that consent can be withdrawn before and during treatment. After the provision of the above information, if the patient requests opinions from other doctors or medical institutions (second opinion), then it is essential to respond to the request. The patient is the main person in the decision-making process, and the right of self-determination of the patient is the most important factor when deciding on indications for non-pharmacotherapy. Basically, the consent of the patient and/or the family is required, based on their understanding of the explanation provided by the medical staff involved in testing and treatment. If the patient is unable to express their intention or is a minor, a family representative or legal representative will act on their behalf. Ultimately, the signatures of all attendees, including the medical staff, are obtained. Normally, 2 copies of the information sheet are prepared. The original copy is generally kept in the patient's medical record and another copy is given to the patient. Physicians must be fully cognizant that informed consent is an important opportunity for the patients to compare and consider the benefits and disadvantages of the treatment, and to enable selection of treatment that is truly beneficial for them. The information must also be specific and easy to understand for the patients and their families. Complications with CIED implantation are due to the device itself (generators and/or leads), or the implantation procedure. Generator complications include malfunctions such as recalls and resets due to electromagnetic interference. Lead complications include malfunctions such as recalls, lead dislodgement and aging, elevated pacing and sensing threshold,13 venous occlusion,14, 15 and tricuspid insufficiency.16-18 Appropriate implantation techniques can reduce complications (eg, infection, pneumothorax, lead fracture, lead insulation break, and perforation resulting from lead insertion technique). There are differences in the incidence of CIED infection among facilities, which is reported to range from 0.2% to 7%.19 Expansion of the ICD and CRT-D indications has resulted in increased implantation of the devices in high-risk patients (ie, elderly patients; patients with heart failure, renal failure, or diabetes; and patients taking steroids and/or antithrombotic agents), as well as an increased number of replacement operations due to long-term survival, which increases the incidence of infection.19-21 It is important to remember that implantation of a CIED is essentially a surgical operation; thus, it is necessary to fully understand the cause of infection and adopt preventive measures (maintaining the operating room environment,22, 23 hand washing, differentiating clean and unclean, surgical instrument disinfection and sterilization, surgical field disinfection, double gloving,24, 25 and use of antibiotics26). The Japanese Society of Chemotherapy and the Japan Society for Surgical Infection have proposed “Practical Guidelines for Proper Use of Antibiotics for Prevention of Postoperative Infection”,26 particularly with respect to the use of antibiotics. As hematoma formation is also an infection risk, intraoperative complete hemostasis is required.19 The depth of the pocket holding the CIED unit is also important. The pocket is created directly above the fascia of the pectoralis major muscle, not in the subcutaneous fat, to prevent compression necrosis of the skin.27 If the subcutaneous tissue containing the subcutaneous fat is thin, it is recommended to create a pocket under the pectoralis major muscle.27 Puncture for lead insertion can cause complications such as pneumothorax,28, 29 and arterial puncture. Therefore, methods such as angiography before puncture30 and ultrasound-guided puncture31 have been recommended to mitigate complications. Axillary vein puncture is prefer over subclavian vein puncture to prevent lead compression fracture by the subclavian muscle and costoclavicular ligament.32, 33 Meanwhile, the incision method is recommended not only to reduce the aforementioned complications but also to prevent lead fracture, prevent damage to the covering, and improve lead durability.34 When inserting the lead, care should be taken to avoid perforation of the venous wall and myocardial wall. The perforation rate is reported to be 0.4%–0.64%,28, 29 and perforation is caused by the use of hard stylets, excessive pressure on the lead, rotation of the lead body after lead placement, and lead traction. It is also important to understand the characteristics of the lead tip shape (passive or active). Before discharge, items including the wound's condition (dehiscence, signs of infection), position of the implanted lead, pacing threshold, and sensory potential amplitude should be checked. As an early postoperative complication, hematoma has a relatively high incidence. Patients who are taking anticoagulants or antiplatelet drugs due to atrial fibrillation (AF), mechanical valve replacement or arteriosclerotic disease are considered a high-risk group, and postoperative wound observation is particularly important in these patients. Hematoma not only causes postoperative pain but can also cause device infection at a remote phase because it leads to incision dehiscence and compression necrosis. When hematoma is discovered, it is important to determine the necessity of reoperation. If there is no discoloration of the skin surface and no sign of skin necrosis or infection, the hematoma is almost always reabsorbed and can be dealt with using recompression. However, hematoma removal and hemostasis should be considered if the pain intensifies because of hematoma distension or if wound dehiscence occurs. Removal of hematoma by puncture is never performed because it increases the risk of infection. There is no fixed trend in the onset of device infection, and risk factors include diabetes, renal impairment, heart failure, steroid use, non-use of preoperative antibiotics, postoperative hematoma and other wound complications, dialysis, chronic obstructive pulmonary disease, cerebrovascular disorders, external pacemaker placement, device replacement, and early reoperation.35, 36 Once device infection has occurred, it is necessary to remove the entire system, which poses a great risk. Careful consideration is required before, during, and after surgery. Chest X-ray (frontal and left lateral view) to check lead position abnormalities and movement, and simultaneous ECG monitoring to detect early pacing/sensing failure are also important. The intraoperative threshold and peak value of the intracardiac potential may change significantly after surgery. It is necessary to check the pacing threshold and the detected peak value of the intracardiac potential before discharge, and readjust the values as necessary. It is also recommended to conduct optimization before discharge of patients who underwent CRT. Guidance on daily life after discharge should also be provided. Electromagnetic interference can cause electrical noise, which can trigger pacing inhibition in pacemakers and inappropriate therapy in ICDs. A detailed list of electromagnetic interference sources has been published by the Japan Arrhythmia Device Industry Association.37 Household appliances can generally be used if the switch is not frequently turned on and off; however, patients must be instructed to maintain a distance of 50 cm from electromagnetic induction-heating rice cookers and 15 cm from mobile phones. Other than home appliances, use of low-frequency therapy equipment, powerful magnets, electric baths, etc, is prohibited. Caution is needed when using electric vehicle chargers (especially rapid chargers), wireless cards (contactless IC cards), electronic article surveillance equipment, and electronic tags (ie, RFID). The patient should be instructed to move away from the location if they experience dizziness, lightheadedness, palpitations, etc. X-ray and CT have been reported to have an effect, and medical care may be required to prevent interference. There are conditions to be satisfied for MRI, and patients with ICD/CRT-D are required by law to have a restricted driver's license (see Chapter VI, 1.2.1 Driving restrictions [particularly for patients with an ICD]). The recent emergence of new devices has made management more complicated. There is also a concern that the degree of understanding of device functions will decline as patients become older. It is important to provide thorough patient education using pamphlets and to offer information on appropriate measures for possible problems. CIED outpatient management is basically performed with device interrogation using the obtained parameters. It is necessary to ascertain not only the mechanical information of the device but also the general condition of the patient using various parameters. Specific information includes: (1) battery status; (2) lead information; (3) pacing settings; (4) intracardiac electrogram sensing and pacing threshold; (5) arrhythmia detection and treatment status; and (6) heart rate histogram and physical activity biometric information, etc The patient guidance management fee is calculated by looking at this information. In recent years, it has become possible to perform remote monitoring in most CIEDs. Remote monitoring has been shown to be as safe as conventional face-to-face examinations, and to enable the earlier diagnosis of arrhythmia and lead/device failures.38-41 Furthermore, shortened hospital stays,42, 43 and improved life prognosis have also been reported.44, 45 Therefore, remote monitoring for patients with CIEDs is highly useful, and it is recommended to introduce it as a standard management method; however, there are concerns about increased workload for hospital staff. There has been an increase in the number of patients implanted with CIEDs and in the aging population; therefore, efficiently providing outpatient services to patients with CIEDs is required. Doc

1 Current status of non‐pharmacotherapies in Japan Pacemaker treatment for bradyarrhythmia was first approved for national health insurance coverage in Japan in 1974, and its use rapidly became widespread thereafter. Approximately 40 years later, in 2017, the number of patients treated with this technology has increased to 60 137 (41 895 new cases and approximately 18 242 replacements). 5 Capsule‐shaped leadless pacemaker also became available in 2016, and this technology is being established as a new option. Non‐pharmacological treatment of tachy‐arrhythmia began in 1969 from when Will C. Sealy performed surgery in patients with Wolff‐Parkinson‐White (WPW) syndrome (Figure 1). Since then, the application of surgical treatment has expanded to conditions such as ventricular tachycardia (VT) and atrial fibrillation (AF), and surgery has been the pioneer of radical therapy for tachyarrhythmias. At the present time, many surgical methods have been replaced by catheter ablation; however, surgical treatment still remains an indispensable option for patients with a tachycardia resistant to other medical treatments. Open in a separate window FIGURE 1 History of non‐pharmacotherapy of cardiac arrhythmia

The peroxisome proliferator-activated receptor α (PPARα) is a member of the nuclear receptor superfamily and mediates the biological effects of peroxisome proliferators. To determine the physiological role of PPARα in cardiac fatty acid metabolism, we examined the regulation of expression of cardiac fatty acid-metabolizing proteins using PPARα-null mice. The capacity for constitutive myocardial β-oxidation of the medium and long chain fatty acids, octanoic acid and palmitic acid, was markedly reduced in the PPARα-null mice as compared with the wild-type mice, indicating that mitochondrial fatty acid catabolism is impaired in the absence of PPARα. In contrast, constitutive β-oxidation of the very long chain fatty acid, lignoceric acid, did not differ between the mice, suggesting that the constitutive expression of enzymes involved in peroxisomal β-oxidation is independent of PPARα.Indeed, PPARα-null mice had normal levels of the peroxisomal β-oxidation enzymes except the D-type bifunctional protein. At least seven mitochondrial fatty acid-metabolizing enzymes were expressed at much lower levels in the PPARα-null mice, whereas other fatty acid-metabolizing enzymes were present at similar or slightly lower levels in the PPARα-null, as compared with wild-type mice. Additionally, lower constitutive mRNA expression levels of fatty acid transporters were found in the PPARα-null mice, suggesting a role for PPARα in fatty acid transport and catabolism. Indeed, in fatty acid metabolism experiments in vivo, myocardial uptake of iodophenyl 9-methylpentadecanoic acid and its conversion to 3-methylnonanoic acid were reduced in the PPARα-null mice. Interestingly, a decreased ATP concentration after exposure to stress, abnormal cristae of the mitochondria, abnormal caveolae, and fibrosis were observed only in the myocardium of the PPARα-null mice. These cardiac abnormalities appeared to proceed in an age-dependent manner. Taken together, the results presented here indicate that PPARα controls constitutive fatty acid oxidation, thus establishing a role for the receptor in cardiac fatty acid homeostasis. Furthermore, altered expression of fatty acid-metabolizing proteins seems to lead to myocardial damage and fibrosis, as inflammation and abnormal cell growth control can cause these conditions. The peroxisome proliferator-activated receptor α (PPARα) is a member of the nuclear receptor superfamily and mediates the biological effects of peroxisome proliferators. To determine the physiological role of PPARα in cardiac fatty acid metabolism, we examined the regulation of expression of cardiac fatty acid-metabolizing proteins using PPARα-null mice. The capacity for constitutive myocardial β-oxidation of the medium and long chain fatty acids, octanoic acid and palmitic acid, was markedly reduced in the PPARα-null mice as compared with the wild-type mice, indicating that mitochondrial fatty acid catabolism is impaired in the absence of PPARα. In contrast, constitutive β-oxidation of the very long chain fatty acid, lignoceric acid, did not differ between the mice, suggesting that the constitutive expression of enzymes involved in peroxisomal β-oxidation is independent of PPARα.Indeed, PPARα-null mice had normal levels of the peroxisomal β-oxidation enzymes except the D-type bifunctional protein. At least seven mitochondrial fatty acid-metabolizing enzymes were expressed at much lower levels in the PPARα-null mice, whereas other fatty acid-metabolizing enzymes were present at similar or slightly lower levels in the PPARα-null, as compared with wild-type mice. Additionally, lower constitutive mRNA expression levels of fatty acid transporters were found in the PPARα-null mice, suggesting a role for PPARα in fatty acid transport and catabolism. Indeed, in fatty acid metabolism experiments in vivo, myocardial uptake of iodophenyl 9-methylpentadecanoic acid and its conversion to 3-methylnonanoic acid were reduced in the PPARα-null mice. Interestingly, a decreased ATP concentration after exposure to stress, abnormal cristae of the mitochondria, abnormal caveolae, and fibrosis were observed only in the myocardium of the PPARα-null mice. These cardiac abnormalities appeared to proceed in an age-dependent manner. Taken together, the results presented here indicate that PPARα controls constitutive fatty acid oxidation, thus establishing a role for the receptor in cardiac fatty acid homeostasis. Furthermore, altered expression of fatty acid-metabolizing proteins seems to lead to myocardial damage and fibrosis, as inflammation and abnormal cell growth control can cause these conditions. peroxisome proliferator-activated receptor 9-methylpentadecanoic acid 3-methylnonanoic acid p-iodophenyl acetic acid heart-type fatty acid-binding protein very long chain acyl-CoA dehydrogenase long chain acyl-CoA dehydrogenase long chain acyl-CoA synthetase mitochondrial trifunctional protein α and β subunit fatty acid transport protein fatty acid translocase medium chain acyl-CoA dehydrogenase short chain acyl-CoA dehydrogenase short chain 3-hydroxyacyl-CoA dehydrogenase carnitine palmitoyl-CoA transferase Long chain fatty acids are one of the major cardiac energy substrates, so understanding long chain fatty acid metabolism may help in elucidating the mechanisms of various heart diseases (1.Bremer J. Osmundsen H. Fatty Acid Metabolism and Its Regulation. Elsevier Science Publishers, New York1984Google Scholar, 2.Watanabe K. Ohta Y. Toba K. Ogawa Y. Hanawa H. Hirokawa Y. Kodama M. Tanabe N. Hirono S. Ohkura Y. Nakamura Y. Kato K. Aizawa Y. Fuse I. Miyajima S. Kusano Y. Nagatomo T. Hasegawa G. Naito M. Ann. Nucl. Med. 1998; 12: 261-266Crossref PubMed Scopus (45) Google Scholar, 3.Aoyama T. Souri M. Ushikudo S. Kamijo T. Yamaguchi S. Kelly R. Rhead W.J. Uetake K. Tanaka K. Hashimoto T. J. Clin. Invest. 1995; 95: 2465-2473Crossref PubMed Scopus (154) Google Scholar). Changes in peroxisomal and microsomal gene expression induced by peroxisome proliferators are mediated by the peroxisome proliferator-activated receptor α (PPARα),1 a member of the nuclear receptor superfamily (4.Isemann I. Green S. Nature. 1990; 347: 645-650Crossref PubMed Scopus (3131) Google Scholar, 5.Zhang B. Marcus S.L. Sajjadi F.G. Alvares K. Reddy J.K. Subramani S. Rachubinski R.A. Capone J.P. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 7541-7545Crossref PubMed Scopus (244) Google Scholar, 6.Gulick T. Cresci S. Caira T. Moore D.D. Kelly D.P. Proc. Natl. Acad. Sci. U. S. A. 1994; 91: 11012-11016Crossref PubMed Scopus (501) Google Scholar, 7.Lee S.S. Pineau T. Drago J. Lee E.J. Owens J.W. Kroetz D.L. Fernandez-Salguero P.M. Westphal H. Gonzalez F.J. Mol. Cell. Biol. 1995; 15: 3012-3022Crossref PubMed Scopus (1520) Google Scholar, 8.Braissant O. Foufelle F. Scotto C. Dauca M. Wahli W. Endocrinology. 1996; 137: 354-366Crossref PubMed Scopus (1737) Google Scholar). Three distinct PPARs have been found, PPARα, PPARβ (also called PPARδ), and PPARγ. The tissue distribution of each receptor is different, implying that each has unique functions (8.Braissant O. Foufelle F. Scotto C. Dauca M. Wahli W. Endocrinology. 1996; 137: 354-366Crossref PubMed Scopus (1737) Google Scholar). In rodents, PPARα is abundant in the liver, kidney, and heart, all of which have high rates of fatty acid metabolism (8.Braissant O. Foufelle F. Scotto C. Dauca M. Wahli W. Endocrinology. 1996; 137: 354-366Crossref PubMed Scopus (1737) Google Scholar). Little is known about the regulation of fatty acid metabolism in the heart, although such information may help to elucidate the regulatory systems and the physiological roles of PPARα in heart. Interest in the clinical use of iodine-123-labeled fatty acids is currently primarily focused on the use of iodine-123 15-(p-iodophenyl) pentadecanoic acid and modified fatty acid analogues such as 15-(p-iodophenyl)-3-R,S-methylpentadecanoic acid. which show delayed myocardial clearance, thus permitting single photon emission tomographic imaging (9.Chouraqui P. Maddahi J. Henkin R. Karesh S.M. Galie E. Berman D.S. J. Nucl. Med. 1991; 32: 447-452PubMed Google Scholar, 10.Knapp F.F. Kropp Jr., J. Eur. J. Nucl. Med. 1995; 22: 361-381Crossref PubMed Scopus (115) Google Scholar). Recently, 15-p-iodine-123 iodophenyl 9-methylpentadecanoic acid (9MPA), a new single photon agent, has been developed (9.Chouraqui P. Maddahi J. Henkin R. Karesh S.M. Galie E. Berman D.S. J. Nucl. Med. 1991; 32: 447-452PubMed Google Scholar, 10.Knapp F.F. Kropp Jr., J. Eur. J. Nucl. Med. 1995; 22: 361-381Crossref PubMed Scopus (115) Google Scholar, 11.Nakajima K. Taki J. Yamamoto W. Takata S. Shimizu M. Tonami N. Nucl. Med. Commun. 1998; 19: 839-847Crossref PubMed Scopus (3) Google Scholar). 9MPA is a modified long chain (15 carbons, C-15) fatty acid, which differs from iodophenyl pentadecanoic acid by a methyl branch at carbon 9. After venous injection, 9MPA is transferred to a myocardial triglyceride pool or undergoes β-oxidation three times in mitochondria, and a metabolite of 9-p-iodophenyl-3-methylnonanoic acid (3MNA) appears. Continuously, 3MNA undergoes α- and β-oxidation, to yield the metabolite of p-iodophenylacetic acid (PIPA). 9MPA is therefore well suited to studies into the abilities of fatty acid uptake and oxidation in vivo. In the present study, we analyzed cardiac fatty acid metabolism bothin vitro and vivo, using PPARα-null mice, finding a key role for PPARα in fatty acid metabolism and homeostasis. Additionally, we found that altered fatty acid metabolism, as well as inflammation and abnormal cell growth control, can cause cardiac tissue damage. [1-14C]Octanoic acid (2 GBq (54 mCi)/mmol), [1-14C]palmitic acid (2 GBq/mmol), and [1-14C]lignoceric acid (1.7 GBq/mmol) were purchased from American Radiolabeled Chemicals (St. Louis, MO).125I-Labeled 9MPA (7.4 GBq/mg), 125I-labeled 3MNA (7.4 GBq/mg), and 125I-labeled PIPA (7.4 GBq/mg) were donated by Daiichi Radioisotope Laboratories Ltd. (Tokyo, Japan). PPARα-null mice in an Sv/129 genetic background were produced as described (12.Aoyama T. Peters J.M. Iritani N. Nakajima T. Furihata K. Hashimoto T. Gonzalez F.J. J. Biol. Chem. 1998; 273: 5678-5684Abstract Full Text Full Text PDF PubMed Scopus (764) Google Scholar). Wild-type Sv/129 were used as controls in all experiments. The animals were housed five per cage and allowed free access to tap water and standard laboratory mouse chow (Oriental Japan Inc. Tokyo, Japan). Mice were housed in a temperature-controlled room (22 ± 2 °C) under a 12-h light/dark (7 p.m. to 7 a.m.) cycle. After determination of the heart rate and blood pressure by the tail cuff method (Softron Co., Tokyo, Japan) at the age of 16 or 32 weeks, the mice were killed. Mice at the age of 16 weeks were used in the experiments involving stresses. Mice were fasted for 48 h and then fed for 24 h. Mice again were fasted for 48 h and then used to analyze ATP, calcium, and magnesium concentrations in myocardium. Following starvation plus high temperature stress, the mouse was placed into a 50-ml plastic tube with many small holes and exposed at 33 °C for 1 h in the air incubator. It was then immediately used for the analysis. All mice survived the stresses, but most of the PPARα-null mice died of hyperthermia when exposed at 42 °C for 20–50 min. Fatty acid β-oxidation activity was measured by the method of Shindo et al. (13.Shindo Y. Osumi T. Hashimoto T. Biochem. Pharmacol. 1978; 27: 2683-2688Crossref PubMed Scopus (41) Google Scholar). Briefly, unfrozen myocards were homogenized in four volumes of 0.25m sucrose containing 1 mm EDTA in a Potter-Elvehjem homogenizer using a tightly fitting Teflon pestle. Approximately 500 μg of homogenate was incubated with the assay medium in 0.2 ml of 150 mm potassium chloride, 10 mm HEPES, pH 7.2, 0.1 mm EDTA, 1 mmpotassium phosphate buffer, pH 7.2, 5 mm Tris malonate, 10 mm magnesium chloride, 1 mm carnitine, 0.15% bovine serum albumin, 5 mm ATP, and 50 μmeach fatty acid (5.0 × 104 cpm of radioactive substrate). The reaction was run for 30 min at 25 °C and stopped by the addition of 0.2 ml of 0.6 n perchloric acid. The mixture was centrifuged at 2,000 × g for 10 min, and the unreacted fatty acid in the supernatant was removed with three extractions of 2 ml of n-hexane. Radioactive degradation products in the water phase were counted, and fatty acid β-oxidation activity was expressed as nanomoles/min/mg of protein. Myocardial extracts were subjected to 10% or 15% SDS-polyacrylamide gel electrophoresis and transferred to nitrocellulose membranes. The membranes were incubated with the primary antibody followed by alkaline phosphatase-conjugated goat anti-rabbit IgG. The origin of primary rabbit polyclonal antibodies was described elsewhere (12.Aoyama T. Peters J.M. Iritani N. Nakajima T. Furihata K. Hashimoto T. Gonzalez F.J. J. Biol. Chem. 1998; 273: 5678-5684Abstract Full Text Full Text PDF PubMed Scopus (764) Google Scholar). Rabbit polyclonal antibody against the heart-type fatty acid-binding protein (H-FABP) was prepared as described previously (14.Sakai K. Fujii H. Yamamoto T. Sakakibara J. Izumi T. Shibata A. Ono T. Eur. J. Biochem. 1995; 229: 201-206Crossref PubMed Scopus (15) Google Scholar). mRNA analysis was performed by Northern blotting. Total myocardial RNA was extracted, electrophoresed on 1.1 m formaldehyde-containing 1% agarose gels, and transferred to nylon membranes (15.Aoyama T. Ueno I. Kamijo T. Hashimoto T. J. Biol. Chem. 1994; 269: 19088-19094Abstract Full Text PDF PubMed Google Scholar). The membranes were incubated with32P-labeled cDNA probes and analyzed on a Fuji system analyzer (Fuji Photo Film Co., Tokyo, Japan). The cDNA probes used were for very long chain acyl-CoA dehydrogenase (VLCAD) (15.Aoyama T. Ueno I. Kamijo T. Hashimoto T. J. Biol. Chem. 1994; 269: 19088-19094Abstract Full Text PDF PubMed Google Scholar), long chain acyl-CoA dehydrogenase (LCAD) (16.Matsubara Y. Indo Y. Naito E. Ozasa H. Glassberg R. Vockley J. Ikeda Y. Kraus J. Tanaka K. J. Biol. Chem. 1989; 264: 16321-16331Abstract Full Text PDF PubMed Google Scholar), long chain acyl-CoA synthetase (LACS) (17.Suzuki H. Kawarabayashi Y. Kondo J. Abe T. Nishikawa K. Kimura S. Hashimoto T. Yamamoto T. J. Biol. Chem. 1990; 265: 8681-8685Abstract Full Text PDF PubMed Google Scholar), mitochondrial trifunctional protein β subunit (TPβ) (18.Kamijo T. Aoyama T. Miyazaki J. Hashimoto T. J. Biol. Chem. 1993; 268: 26452-26460Abstract Full Text PDF PubMed Google Scholar), fatty acid transport protein (FATP) (19.Schaffer J, A. Lodish H, F. Cell. 1994; 79: 427-436Abstract Full Text PDF PubMed Scopus (756) Google Scholar), and fatty acid translocase (FAT) (20.Abumrad N, A. El-Magharabi M, R. Amri E, Z. Lopez E. Grimaldi P, A. J. Biol. Chem. 1993; 268: 17665-17668Abstract Full Text PDF PubMed Google Scholar). Isotope with a long half-life is useful when radioactivity is assayed. 125I-Labeled 9MPA was used, because123I has a shorter (13 h) and 125I a longer (60 days) half-life. 125I-9MPA (741 KBq (20 μCi)) was intravenously injected into mice. At 3 and 10 min after injection, the mice were sacrificed. Lipid extraction from tissues was performed according to a modified version of the method developed by Folchet al. (21.Folch G. Lees M. Stanley G.H.S. J. Biol. Chem. 1957; 266: 497-509Abstract Full Text PDF Google Scholar, 22.Neely J.R. Rovetto M.J. Oram J.F. Prog. Cardiovasc. Dis. 1972; 15: 289-329Crossref PubMed Scopus (392) Google Scholar). Briefly, tissue specimens were homogenized and extracted twice with chloroform/methanol (2:1, v/v). The resulting organic, aqueous, and solid phase was separated. The myocardial radioactivity in each phase was calculated as a percentage of the administered dose per gram of tissue. Radioactivity distribution in the organic phase was assayed as described (21.Folch G. Lees M. Stanley G.H.S. J. Biol. Chem. 1957; 266: 497-509Abstract Full Text PDF Google Scholar, 22.Neely J.R. Rovetto M.J. Oram J.F. Prog. Cardiovasc. Dis. 1972; 15: 289-329Crossref PubMed Scopus (392) Google Scholar), by thin-layer chromatography on a reversed phase plate (C18 Silicagel Spotfilm; Tokyo Kasei Kogyo Co. Ltd., Tokyo, Japan), together with125I-labeled standard fatty acids (9MPA, 3MNA, and PIPA). Mouse heart, approximately 200 mg, was minced and mixed with 300 μl of 50 mm sodium phosphate buffer (pH 7.3). The mixture was well treated with a microsonicator (Powersonic model 50; Yamato, Tokyo, Japan). The lysate was centrifuged at 4,000 × g for 10 min. The pellet was mixed with 200 μl of the buffer and then sonicated and recentrifuged. The same operation was performed once more, and the resultant three supernatant fractions were combined. The proteins in the supernatant solution were extracted with water-saturated chloroform, and the final solution was treated with DIA reagent calcium® or DIA reagent magnesium® (Mitsubishi Chemicals, Tokyo, Japan). Calcium and magnesium concentrations were determined with an Olympus AU5200 (5232-01) and Olympus AU800 (802-01) autoanalyzer (Olympus, Tokyo, Japan), respectively. The ATP concentration was measured with ATP bioluminescence assay kit HS-II (Roche Molecular Biochemicals Japan, Tokyo, Japan), which contained cell lysis solution preventing ATP degradation. All tissues upon removal were rinsed with PBS and then treated either with liquid nitrogen for RNA preparation or with 10% formalin for histological examination. For light microscopy, the formalin-fixed and paraffin-embedded heart sections were stained with hematoxylin-eosin and Azan-Mallory. Using Azan-Mallory-stained specimens from the middle level of both ventricles, the area of myocardial fibrosis was quantified by color differences (blue fibrotic area as opposed to red myocardium) using a color image analyzer (CIA-102; Olympus, Tokyo, Japan). The degree of fibrotic area was scored using a 4-grade scale (none, 0%; mild, <10% of the area; moderate, 10–50% of the area; and severe, >50% of the area). For electron microscopy, the heart was fixed in 2.5% glutaraldehyde in 0.1 m phosphate buffer and post-fixed with 1.5% osmium tetroxide in phosphate buffer. After dehydration in graded ethanol and propylene oxide, the blocks were embedded in Epon 812 (E. Fullan, Latham, NY) epoxy resin. Ultrathin sections were observed under a Hitachi H-800 electron microscope (Hitachi, Tokyo, Japan) after staining with lead citrate. The degree of abnormal cristae in mitochondria and abnormal caveolae in endothelial cells (none, mild, moderate, and severe) was visually assessed by 3 physicians blinded to the PPARα-null and wild-type mice. To identify the level of specific fatty acid-metabolizing enzymes and H-FABP that might be influenced by PPARα, antibodies were used to measure protein levels by immunoblotting (Fig.1, and TableI). Constitutive expression levels of several enzymes (VLCAD, medium chain acyl-CoA dehydrogenase (MCAD), short chain acyl-CoA dehydrogenase (SCAD), mitochondrial trifunctional protein α subunit (TPα), short chain 3-hydroxyacyl-CoA dehydrogenase (SCHAD), LACS, carnitine palmitoyl-CoA transferase (CPT II), and peroxisomal D-type bifunctional protein) were much lower, 19–77%, in PPARα-null mice than wild-type mice. Some other mitochondrial, microsomal, and cytosolic fatty acid-metabolizing proteins examined and H-FABP were expressed at similar levels in PPARα-null and wild-type mice. When comparing the protein contents with those in liver, outstanding reductions in heart were observed for five mitochondrial β-oxidation enzymes (MCAD, SCAD, TPα, SCHAD, and CPT II) (Table I), indicating constitutive organ-specific expression in a limited number of β-oxidation enzymes. The protein contents of the hearts from 32-week-old mice were very similar to those from 16-week-old mice, described in Table I, (data not shown), suggesting the absence of age-dependent change in β-oxidation ability.Table IImmunoblot quantitation of cardiac fatty acid-metabolizing enzymes and fatty acid-binding protein in wild-type (+/+) and PPARα-null (−/−) miceProteinLocalizationIn heartpvalue aData compare (+/+) with (−/−) in heart.(−/−) bData are from Ref. 12. in liverp value cData compare (+/+) with (−/−) in liver.(+/+)(−/−)VLCADMt1.000.19 ± 0.01<0.0010.32 ± 0.040.000LCADMt1.001.10 ± 0.140.6780.40 ± 0.21 dA significant difference was found between heart and liver.0.039MCADMt1.000.70 ± 0.030.0091.05 ± 0.14 dA significant difference was found between heart and liver.0.652SCADMt1.000.34 ± 0.01<0.0010.87 ± 0.14 dA significant difference was found between heart and liver.0.248TPαMt1.000.68 ± 0.050.0070.98 ± 0.09 dA significant difference was found between heart and liver.0.810TPβMt1.000.87 ± 0.100.2231.03 ± 0.071.000MHMt1.000.95 ± 0.040.8021.12 ± 0.120.423SCHADMt1.000.42 ± 0.030.0083.90 ± 1.02 dA significant difference was found between heart and liver.0.008T1Mt1.000.89 ± 0.100.0990.59 ± 0.02 dA significant difference was found between heart and liver.0.001T2Mt/Cs1.000.81 ± 0.060.2150.98 ± 0.290.912LACSMt/Ms1.000.49 ± 0.040.0100.42 ± 0.060.001CPT IIMt1.000.23 ± 0.01<0.0010.90 ± 0.18 dA significant difference was found between heart and liver.0.574MTE IMt1.000.88 ± 0.120.1960.95 ± 0.070.567CTE IICs1.000.83 ± 0.110.080ND dA significant difference was found between heart and liver.AOXPs1.001.20 ± 0.070.5540.98 ± 0.140.850PHPs1.000.92 ± 0.020.6720.93 ± 0.200.653DBFPs1.000.77 ± 0.140.0490.64 ± 0.010.001PTPs1.001.12 ± 0.070.7051.06 ± 0.030.196VLACSPs/Ms1.000.95 ± 0.060.5930.86 ± 0.070.087H-FABPCs1.000.92 ± 0.040.557NETotal cardiac cell extract was subjected to electrophoresis and proteins were transferred to nitrocellulose membranes and screened with specific antibodies. The signals were quantified by scanning densitometry, and the values from (+/+) mice were assigned the number 1.00. Mt, mitochondria; Ms, microsomes; Ps, peroxisomes; Cs, cytosol; ND, not detected; NE, not examined. Results are the means ± S.D. of three determinations. AOX, acyl-CoA oxidase; PH, peroxisomal bifunctional protein; PT, peroxisomal thiolase; VLACS, very long chain acyl-CoA synthetase; T1, short chain specific 3-ketoacyl-CoA thiolase; T2, acetoacetyl-CoA thiolase; MH, mitochondrial short chain specific hydratase; MTE I, mitochondrial thioesterase; CTE II, cytosolic thioesterase; DBF, peroxisomal D-type bifunctional protein.a Data compare (+/+) with (−/−) in heart.b Data are from Ref. 12.Aoyama T. Peters J.M. Iritani N. Nakajima T. Furihata K. Hashimoto T. Gonzalez F.J. J. Biol. Chem. 1998; 273: 5678-5684Abstract Full Text Full Text PDF PubMed Scopus (764) Google Scholar.c Data compare (+/+) with (−/−) in liver.d A significant difference was found between heart and liver. Open table in a new tab Total cardiac cell extract was subjected to electrophoresis and proteins were transferred to nitrocellulose membranes and screened with specific antibodies. The signals were quantified by scanning densitometry, and the values from (+/+) mice were assigned the number 1.00. Mt, mitochondria; Ms, microsomes; Ps, peroxisomes; Cs, cytosol; ND, not detected; NE, not examined. Results are the means ± S.D. of three determinations. AOX, acyl-CoA oxidase; PH, peroxisomal bifunctional protein; PT, peroxisomal thiolase; VLACS, very long chain acyl-CoA synthetase; T1, short chain specific 3-ketoacyl-CoA thiolase; T2, acetoacetyl-CoA thiolase; MH, mitochondrial short chain specific hydratase; MTE I, mitochondrial thioesterase; CTE II, cytosolic thioesterase; DBF, peroxisomal D-type bifunctional protein. To determine whether the drop in the expression level of fatty acid-metabolizing enzymes is due to altered gene expression, myocardial mRNA levels were analyzed by Northern blotting (Fig. 2). Constitutive levels of mRNA for VLCAD and LACS in the PPARα-null mice were 32.7 ± 2.5% (p = 0.0002) and 21.6 ± 1.0% (p = 0.0004) of those in the wild-type mice, respectively. Those for LCAD and TPβ were 101.6 ± 3.2% (p = 0.2511) and 116.5 ± 6.9% (p= 0.055), respectively, similar between the two strains. These results are consistent with the protein measurements. It is noteworthy that myocardial levels of mRNA for FATP and FAT in the PPARα-null mice were 51.6 ± 4.4% (p = 0.0034) and 43.8 ± 5.6% (p = 0.0028) of those in the wild-type mice, respectively, demonstrating that the ability concerning fatty acid uptake in the PPARα-null mice is probably inferior to that in the wild-type mice. As shown in Table I, several enzymes involved in fatty acid β-oxidation had lower constitutive expression levels in the PPARα-null mice. To evaluate the significance of the altered activity levels of fatty acid β-oxidation enzymes, overall myocardial β-oxidation activity was measured, using octanoic acid (C-8), palmitic acid (C-16), and lignoceric acid (C-24) as substrates (Fig.3). The octanoic acid β-oxidation activity of the PPARα-null mice was lower than that of wild-type mice, which is consistent with the lower expression levels of MCAD, SCAD, and SCHAD, having higher catalytic activities for medium and short chain fatty acids, in the PPARα-null mice, respectively (TableI). The palmitic acid β-oxidation activity of the PPARα-null mice was very low, reflecting the reduced expression of long chain-specific mitochondrial fatty acid-metabolizing proteins (VLCAD, TPα, LACS, and CPT II) (Table I). Lignoceric acid β-oxidation activities of the two strains were nearly identical, reflecting the similar expression levels of very long chain-specific peroxisomal fatty acid-metabolizing proteins (acyl-CoA oxidase, peroxisomal bifunctional protein, peroxisomal thiolase, and very long chain acyl-CoA synthetase) (TableI). The overall β-oxidation activities in hearts from 32-week-old mice were similar to those from 16-week-old mice (Fig. 3), indicating the absence of age-dependent change, as expected. The uptake of125I-labeled compound (% dose/g tissue) in the heart is summarized in Table II. The myocardial initial uptake (at 3 min after injection) of 125I-9MPA was higher in wild-type than in the PPARα-null mice, suggesting that the myocardial initial uptake decreased due to lower levels of FATP and FAT in the PPARα-null mice (Fig. 2).Table IIDistribution of 125 I radioactivity in heart following intravenous injection of 125 I-labeled 9MPATime% dose/g tissue9MPA3MNAPIPA3MNA/9MPAmin%%%Wild-type mice 312.1 ± 1.93.7 ± 0.516.7 ± 2.24.7 ± 0.44.5 /1.0 1023.3 ± 2.52.1 ± 0.412.3 ± 2.012.9 ± 2.65.9 /1.0PPARα-null mice 37.9 ± 0.912.8 ± 2.2 ap < 0.01 between the wild-type and PPARα-null mice.16.5 ± 2.52.8 ± 0.31.3 /1.0 ap < 0.01 between the wild-type and PPARα-null mice. 1012.0 ± 1.8 ap < 0.01 between the wild-type and PPARα-null mice.6.3 ± 0.9 ap < 0.01 between the wild-type and PPARα-null mice.18.3 ± 2.85.4 ± 0.8 ap < 0.01 between the wild-type and PPARα-null mice.2.9 /1.0 ap < 0.01 between the wild-type and PPARα-null mice.Results are the means ± S.D. of three determinations.a p < 0.01 between the wild-type and PPARα-null mice. Open table in a new tab Results are the means ± S.D. of three determinations. Although 9MPA was rapidly metabolized to 3MNA and only a small amount of 9MPA remained at 3 min after injection in the wild-type mice, the conversion clearly decreased in the PPARα-null mice (Table II and Fig. 4). The 3MNA/9MPA ratios in wild-type and the PPARα-null mice were 4.5 and 1.3 (at 3 min after injection) and 5.9 and 2.9 (at 10 min after injection), respectively (Table II). The slower conversion in the PPARα-null mice is compatible with the very poor myocardial β-oxidation activity of palmitic acid (Fig. 3). The heart from wild-type mice at the age of 32 weeks seemed to be histologically normal (Fig.5 A (a andd), Table III). On the other hand, the heart from the PPARα-null mice at the age of 16 weeks showed a little focal fibrosis and myocardial degeneration associated with contraction band necrosis (Fig. 5 A (b ande), Table III). At the age of 32 weeks, diffuse fibrosis occupied one-third of the inner wall of the myocardium and marked myofibrillar fragmentation of the myocardium was observed (Fig.5 A (c and f), Table III). Inflammatory infiltrates were predominantly composed of macrophages with a few lymphocytes and neutrophils. These pathological findings were not specific for any myocardial disease and were therefore regarded as an unclassified cardiomyopathy. Electron microscopy revealed that the cristae of mitochondria increased in number and density in the myocardial cells of PPARα-null mice at 16 and 32 weeks (Fig.5 B, (B and C), Table III). The number of caveolae in the cardiac capillary endothelium of PPARα-null mice (Fig. 5 B, (E and F), Table III) was larger than that of wild-type mice (Fig. 5 B, (D), Table III). Interestingly, the cardiac abnormalities, the myocardial fibrosis and the degeneration, appear to proceed in an age-dependent manner.Table IIIHistological analysis of cardiac cells in the PPARα-null and wild-type miceWild-type micePPARα-null mice16 weeks32 weeks16 weeks32 weeksFibrosis−−+++Abnormal cristae−−+++++Abnormal caveolae−−+++++−, none; +, mild; ++, moderate; +++, severe. Open table in a new tab −, none; +, mild; ++, moderate; +++, severe. To examine the relation between the impaired fatty acid catabolic ability and the cardiac abnormality in the PPARα-null mice, we carried out experiments in which mice were exposed to the stresses, starvation, and high temperature. The former stress was adopted to enhance dependence on fatty acids/triglycerides as energy sources by reducing serum glucose and lactate concentrations, and the latter to increase the load to cardiac muscle. The cardiac palmitic acid β-oxidation activity after giving these stresses slightly decreased, 5–12% lower than the constitutive activity in the wild-type mice and 2–9% lower than that in the PPARα-null mice, respectively,

A series of hindered phenols were investigated as hypolipidemic and/or hypoglycemic agents with ability to inhibit lipid peroxidation. 1,3-Benzoxathioles (9 and 22), phenoxypentanoic acid (34), phenoxypentanol (35a), phenoxynonanol (35b), phenylchloropropionic acid having a chromanyl group (25), and a thiazolidine compound (27) derived from 25, all having a hindered phenol group, were prepared and examined. Compound 27 showed the expected biological properties in vivo and in vitro without any liver weight increase. Biological activities of the analogous thiazolidine compounds, 43-58, were compared. Thus, (+/-)-5-[4-[(6-hydroxy-2,5,7,8-tetramethylchroman-2-yl)methoxy]- benzyl]-2,4-thiazolidinedione (27) (CS-045) was found to have all of our expected properties and was selected as a candidate for further development as a hypoglycemic and hypolipidemic agent.

Background Although previous clinical trials demonstrated the non-inferiority of a rate control to rhythm control strategy for management of atrial fibrillation (AF), the optimal treatment strategy for paroxysmal AF (PAF) remains unclear. Methods and Results A randomized, multicenter comparison of rate control vs rhythm control in Japanese patients with PAF (the Japanese Rhythm Management Trial for Atrial Fibrillation (J-RHYTHM) study) was conducted. The primary endpoint was a composite of total mortality, symptomatic cerebral infarction, systemic embolism, major bleeding, hospitalization for heart failure, or physical/psychological disability requiring alteration of treatment strategy. In the study, 823 patients with PAF were followed for a mean period of 578 days. The primary endpoint occurred in 64 patients (15.3%) assigned to rhythm control and in 89 patients (22.0%) to rate control (P=0.0128). No significant differences between the treatment strategies were observed in the incidences of death, stroke, bleeding and heart failure. Meanwhile, significantly fewer patients requested changes of assigned treatment strategy in the rhythm control vs the rate control group, which was accompanied by improvement in AF-specific quality of life scores. Conclusion The J-RHYTHM study showed that rhythm control was associated with fewer primary endpoints than rate control. However, mortality and cardiovascular morbidity were not affected by the treatment strategy (umin-CTR No. C000000106).

Brugada syndrome is a genetic disease associated with sudden cardiac death that is characterized by ventricular fibrillation and right precordial ST segment elevation on ECG. Loss-of-function mutations in SCN5A, which encodes the predominant cardiac sodium channel alpha subunit NaV1.5, can cause Brugada syndrome and cardiac conduction disease. However, SCN5A mutations are not detected in the majority of patients with these syndromes, suggesting that other genes can cause or modify presentation of these disorders. Here, we investigated SCN1B, which encodes the function-modifying sodium channel beta1 subunit, in 282 probands with Brugada syndrome and in 44 patients with conduction disease, none of whom had SCN5A mutations. We identified 3 mutations segregating with arrhythmia in 3 kindreds. Two of these mutations were located in a newly described alternately processed transcript, beta1B. Both the canonical and alternately processed transcripts were expressed in the human heart and were expressed to a greater degree in Purkinje fibers than in heart muscle, consistent with the clinical presentation of conduction disease. Sodium current was lower when NaV1.5 was coexpressed with mutant beta1 or beta1B subunits than when it was coexpressed with WT subunits. These findings implicate SCN1B as a disease gene for human arrhythmia susceptibility.

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Long QT syndrome (LQTS) is an inheritable and life-threatening disease; however, it is often difficult to determine disease characteristics in sporadic cases with novel mutations, and more precise analysis is necessary for the successful development of evidence-based clinical therapies. This study thus sought to better characterize ion channel cardiac disorders using induced pluripotent stem cells (iPSCs).We reprogrammed somatic cells from a patient with sporadic LQTS and from controls, and differentiated them into cardiomyocytes through embryoid body (EB) formation. Electrophysiological analysis of the LQTS-iPSC-derived EBs using a multi-electrode array (MEA) system revealed a markedly prolonged field potential duration (FPD). The IKr blocker E4031 significantly prolonged FPD in control- and LQTS-iPSC-derived EBs and induced frequent severe arrhythmia only in LQTS-iPSC-derived EBs. The IKs blocker chromanol 293B did not prolong FPD in the LQTS-iPSC-derived EBs, but significantly prolonged FPD in the control EBs, suggesting the involvement of IKs disturbance in the patient. Patch-clamp analysis and immunostaining confirmed a dominant-negative role for 1893delC in IKs channels due to a trafficking deficiency in iPSC-derived cardiomyocytes and human embryonic kidney (HEK) cells.This study demonstrated that iPSCs could be useful to characterize LQTS disease as well as drug responses in the LQTS patient with a novel mutation. Such analyses may in turn lead to future progress in personalized medicine.

Background Strokes develop even in patients with low CHADS2 scores, and the left atrial appendage (LAA) is the embolic source 90% of the time. We focused on the LAA morphology as a new predictor of strokes. Objective To clarify the anatomical characteristics of the LAA for risk stratification of strokes in patients with nonvalvular atrial fibrillation (AF) who have low CHADS2 scores. Methods Among 80 patients who underwent catheter ablation of AF with contrast-enhanced computed tomography, the LAA characteristics were compared between 30 patients with histories of strokes and 50 age-matched controls. The LAA anatomy was classified into 4 types—“cactus,” “cauliflower,” “chicken wing,” and “windsock”—discriminated by the computed tomography measurements of the length, angle, and number of lobes of the LAA. Results The average CHADS2 score did not differ significantly between patients with stroke and controls (0.8±0.8 vs 0.6±0.7; P = .277). Eight (26.7%) patients with stroke had CHA2DS2-VASc scores of 0. The left atrial size, LAA flow velocity, left ventricular function, and serum brain natriuretic peptide level were also unable to predict strokes. However, a “cauliflower” LAA, defined as a main lobe of less than 4 cm long without forked lobes, was significantly more common in patients with stroke (odds ratio 3.857; 95% confidence interval 1.482–10.037; P = .005). The CHA2DS2-VASc score-adjusted logistic regression analysis revealed the cauliflower LAA as an independent predictor of a stroke (odds ratio 3.355; 95% confidence interval 1.243–9.055; P = .017). Conclusions The LAA anatomy might be useful for predicting strokes in patients with nonvalvular AF who have low CHADS2 scores. Strokes develop even in patients with low CHADS2 scores, and the left atrial appendage (LAA) is the embolic source 90% of the time. We focused on the LAA morphology as a new predictor of strokes. To clarify the anatomical characteristics of the LAA for risk stratification of strokes in patients with nonvalvular atrial fibrillation (AF) who have low CHADS2 scores. Among 80 patients who underwent catheter ablation of AF with contrast-enhanced computed tomography, the LAA characteristics were compared between 30 patients with histories of strokes and 50 age-matched controls. The LAA anatomy was classified into 4 types—“cactus,” “cauliflower,” “chicken wing,” and “windsock”—discriminated by the computed tomography measurements of the length, angle, and number of lobes of the LAA. The average CHADS2 score did not differ significantly between patients with stroke and controls (0.8±0.8 vs 0.6±0.7; P = .277). Eight (26.7%) patients with stroke had CHA2DS2-VASc scores of 0. The left atrial size, LAA flow velocity, left ventricular function, and serum brain natriuretic peptide level were also unable to predict strokes. However, a “cauliflower” LAA, defined as a main lobe of less than 4 cm long without forked lobes, was significantly more common in patients with stroke (odds ratio 3.857; 95% confidence interval 1.482–10.037; P = .005). The CHA2DS2-VASc score-adjusted logistic regression analysis revealed the cauliflower LAA as an independent predictor of a stroke (odds ratio 3.355; 95% confidence interval 1.243–9.055; P = .017). The LAA anatomy might be useful for predicting strokes in patients with nonvalvular AF who have low CHADS2 scores.

Background Short QT syndrome (SQTS) is characterized by an abnormally short QT interval and sudden death. Due to the limited number of cases, the characteristics of SQTS are not well understood. It has been reported recently that early repolarization is associated with idiopathic ventricular fibrillation and the QT interval is short in patients with early repolarization. Objective The purpose of this study was to study the association between early repolarization and arrhythmic events in SQTS. Methods The study consisted of three cohorts: SQTS cohort (N = 37), control cohort with short QT interval and no arrhythmic events (N = 44), and control cohort with normal QT interval (N = 185). ECG parameters were compared among the study cohorts. Results Heart rate, PR interval, and QRS duration were similar among the three study cohorts. Early repolarization was more common in the SQTS cohort (65%) than in the short QT control cohort (30%) and the normal QT control cohort (10%). Duration from T-wave peak to T-wave end was longer in the SQTS cohort than in the short QT control cohort, although QT and corrected QT intervals were similar. In the SQTS cohort, there were more males among patients with arrhythmic events than in those with a family history but without arrhythmic events. In multivariate models, early repolarization was associated with arrhythmic events in the SQTS cohort. ECG parameters including QT and QTc intervals were not associated with arrhythmic events in the SQTS cohort. Conclusion There is a high prevalence of early repolarization in patients with SQTS. Early repolarization may be useful in identifying risk of cardiac events in SQTS. Short QT syndrome (SQTS) is characterized by an abnormally short QT interval and sudden death. Due to the limited number of cases, the characteristics of SQTS are not well understood. It has been reported recently that early repolarization is associated with idiopathic ventricular fibrillation and the QT interval is short in patients with early repolarization. The purpose of this study was to study the association between early repolarization and arrhythmic events in SQTS. The study consisted of three cohorts: SQTS cohort (N = 37), control cohort with short QT interval and no arrhythmic events (N = 44), and control cohort with normal QT interval (N = 185). ECG parameters were compared among the study cohorts. Heart rate, PR interval, and QRS duration were similar among the three study cohorts. Early repolarization was more common in the SQTS cohort (65%) than in the short QT control cohort (30%) and the normal QT control cohort (10%). Duration from T-wave peak to T-wave end was longer in the SQTS cohort than in the short QT control cohort, although QT and corrected QT intervals were similar. In the SQTS cohort, there were more males among patients with arrhythmic events than in those with a family history but without arrhythmic events. In multivariate models, early repolarization was associated with arrhythmic events in the SQTS cohort. ECG parameters including QT and QTc intervals were not associated with arrhythmic events in the SQTS cohort. There is a high prevalence of early repolarization in patients with SQTS. Early repolarization may be useful in identifying risk of cardiac events in SQTS.

Background— Recently, we and others reported that early repolarization (J wave) is associated with idiopathic ventricular fibrillation. However, its clinical and genetic characteristics are unclear. Methods and Results— This study included 50 patients (44 men; age, 45±17 years) with idiopathic ventricular fibrillation associated with early repolarization, and 250 age- and sex-matched healthy controls. All of the patients had experienced arrhythmia events, and 8 (16%) had a family history of sudden death. Ventricular fibrillation was inducible by programmed electric stimulation in 15 of 29 patients (52%). The heart rate was slower and the PR interval and QRS duration were longer in patients with idiopathic ventricular fibrillation than in controls. We identified nonsynonymous variants in SCN5A (resulting in A226D, L846R, and R367H) in 3 unrelated patients. These variants occur at residues that are highly conserved across mammals. His-ventricular interval was prolonged in all of the patients carrying an SCN5A mutation. Sodium channel blocker challenge resulted in an augmentation of early repolarization or development of ventricular fibrillation in all of 3 patients, but none was diagnosed with Brugada syndrome. In heterologous expression studies, all of the mutant channels failed to generate any currents. Immunostaining revealed a trafficking defect in A226D channels and normal trafficking in R367H and L846R channels. Conclusions— We found reductions in heart rate and cardiac conduction and loss-of-function mutations in SCN5A in patients with idiopathic ventricular fibrillation associated with early repolarization. These findings support the hypothesis that decreased sodium current enhances ventricular fibrillation susceptibility.

Background: Target anticoagulation levels for warfarin in Japanese patients with non-valvular atrial fibrillation (NVAF) are unclear. Methods and Results: Of 7,527 patients with NVAF, 1,002 did not receive warfarin (non-warfarin group), and the remaining patients receiving warfarin were divided into 5 groups based on their baseline international normalized ratio (INR) of prothrombin time (≤1.59, 1.6–1.99, 2.0–2.59, 2.6–2.99, and ≥3.0). Patients were followed-up prospectively for 2 years. Primary endpoints were thromboembolic events (cerebral infarction, transient ischemic attack, and systemic embolism), and major hemorrhage requiring hospital admission. During the follow-up period, thromboembolic events occurred in 3.0% of non-warfarin group, but at lower frequencies in the warfarin groups (2.0, 1.3, 1.5, 0.6, and 1.8%/2 years for INR values of ≤1.59, 1.6–1.99, 2.0–2.59, 2.6–2.99, and ≥3.0, respectively; P=0.0059). Major hemorrhage occurred more frequently in warfarin groups (1.5, 1.8, 2.4, 3.3, and 4.1% for INR values ≤1.59, 1.6–1.99, 2.0–2.59, 2.6–2.99, and ≥3.0, respectively; P=0.0041) than in non-warfarin group (0.8%/2 years). These trends were maintained when the analyses were confined to patients aged ≥70 years. Conclusions: An INR of 1.6–2.6 is safe and effective at preventing thromboembolic events in patients with NVAF, particularly patients aged ≥70 years. An INR of 2.6–2.99 is also effective, but associated with a slightly increased risk in major hemorrhage. (UMIN Clinical Trials Registry UMIN000001569) (Circ J 2013; 77: 2264–2270)

Dyslipidemia, an important risk factor for cardiovascular disease, may be associated with atrial fibrillation (AF). Cross-sectional studies that have examined this association, however, have produced controversial results, and few longitudinal studies have been conducted.Using annual health examinations in Japan, the association between lipid profile and the risk of new-onset AF was investigated in the general population. A total of 28,449 individuals who did not have AF at baseline were included in the study. During a follow-up of 4.5±2.7 years, 265 individuals (0.9%) developed AF. In multivariate models, low high-density lipoprotein (HDL) cholesterol was associated with the development of AF in women (hazard ratio [HR], 2.86; 95% confidence interval [CI]: 1.49-5.50) but not in men (HR, 1.35; 95%CI: 0.77-2.38). Women had a 28% higher risk of AF with each 10% decrease in HDL cholesterol. Neither triglycerides nor lipid ratios were associated with AF. After excluding individuals with risk factors for AF, including those who were taking anti-hypertensive drugs, had diabetes, and structural heart disease, the association between low HDL cholesterol and AF remained significant in women.Low HDL cholesterol was associated with an increased risk of new-onset AF in women, but not in men.

This study evaluated the pause-dependency of the J-wave to characterize this phenomenon in idiopathic ventricular fibrillation (VF).The J-wave can be found in apparently healthy subjects and in patients at risk for sudden cardiac death, and risk stratification is therefore needed.Forty patients with J-wave-associated idiopathic VF were studied for J waves with special reference concerning pause-dependent augmentation. J waves were defined as those ≥0.1 mV above the isoelectric line and were compared with 76 non-VF patients of comparable age and sex.The J-wave was larger in patients with idiopathic VF than in the controls: 0.360 ± 0.181 mV versus 0.192 ± 0.064 mV (p = 0.0011). J waves were augmented during storms of VF (n = 9 [22.5%]), which was controlled by isoproterenol; they disappeared within weeks in 5 patients. In addition, sudden prolongation of the R-R interval was observed in 27 patients induced by benign arrhythmia, and 15 patients (55.6%) demonstrated pause-dependent augmentation (from 0.391 ± 0.126 mV to 0.549 ± 0.220 mV; p < 0.0001). In the other 12 experimental subjects and in the 76 control subjects, J waves remained unchanged. Pause-dependent augmentation of J waves was detected in 55.6% (sensitivity) but was specific (100%) in the patients with idiopathic VF with high positive (100%) and negative (86.4%) predictive values.Pause-dependent augmentation of J waves was confirmed in about one-half of the patients with idiopathic VF after sudden R-R prolongation. Such dynamicity of J waves was specific to idiopathic VF and may be used for risk stratification.

Atrial fibrillation (AF) is a common arrhythmia frequently associated with hypertension. This study was designed to test the hypothesis that lowering blood pressure by angiotensin II-receptor blockers (ARB) has more beneficial effects than by conventional calcium channel blockers (CCB) on the frequency of paroxysmal AF with hypertension. The Japanese Rhythm Management Trial II for Atrial Fibrillation (J-RHYTHM II study) is an open-label randomized comparison between an ARB (candesartan) and a CCB (amlodipine) in the treatment of paroxysmal AF associated with hypertension. Using daily transtelephonic monitoring, we examined asymptomatic and symptomatic paroxysmal AF episodes during a maximum 1 year treatment. The primary endpoint was the difference in AF frequency between the pre-treatment period and the final month of the follow-up. The secondary endpoints included cardiovascular events, development of persistent AF, left atrial dimension, and quality-of-life (QOL). The study enrolled 318 patients (66 years, male/female 219/99, 158 in the ARB group and 160 in the CCB group) treated at 48 sites throughout Japan. At baseline, the frequency of AF episodes (days/month) was 3.8 ± 5.0 in the ARB group vs. 4.8 ± 6.3 in the CCB group (not significant). During the follow-up, blood pressure was significantly lower in the CCB group than in the ARB group (P < 0.001). The AF frequency decreased similarly in both groups, and there was no significant difference in the primary endpoint between the two groups. There were no significant differences between the two groups in the development of persistent AF, changes in left atrial dimension, occurrence of cardiovascular events, or changes in QOL. In patients with paroxysmal AF and hypertension, treatment of hypertension by candesartan did not have an advantage over amlodipine in the reduction in the frequency of paroxysmal AF (umin CTR C000000427).

The intraperitoneal injection of 15 mg. puromycin every hour for 4 hr. into 250–300-g. rats reduced protein synthesis in vivo in uteri, liver, heart, kidney, and thymus to 10, 15, 36, 40, and 49% of controls, respectively. Under these same conditions ribonucleic acid (RNA) synthesis was not inhibited in any tissue studied except the thymus. The synthesis of phospholipid in the liver and uteri was not inhibited, but a stimulatory response to puromycin was observed. These results are discussed with regard to the use of puromycin in studying the role of protein synthesis in certain physiological processes.

Radiofrequency (RF) catheter ablation of the renal artery is therapeutic in patients with drug-refractory essential hypertension. This study was designed to examine the role of the renal autonomic nerves and of RF application from inside the renal artery in the regulation of blood pressure (BP). An open irrigation catheter was inserted into either the left or right renal artery in 8 dogs. RF current (17 ± 2 watts) was delivered to one renal artery. Electrical autonomic nerve stimulation was applied to each renal artery before and after RF ablation. BP, heart rate, indices of heart rate variability, and serum catecholamines were analyzed. Before RF ablation, electrical autonomic nerve stimulation of either renal artery increased BP from 150 ± 16/92 ± 15 to 173 ± 21/105 ± 16 mm Hg. After RF ablation, BP increased similarly when the nonablated renal artery was electrically stimulated, although the rise in BP was attenuated when the ablated renal artery was stimulated. Serum catecholamines and sympathetic nerve indices of heart rate variability increased when electrical autonomic nerve stimulation was applied before RF ablation and to the nonablated renal artery after RF ablation, although it changed minimally when the ablated renal artery was stimulated, suggesting interconnectivity between afferent renal nerve stimulation and systemic sympathetic activity. Renal artery angiogram showed no apparent injury after RF ablation. In conclusion, electrical stimulation of the renal arterial autonomic nerves increases BP via an increase in central sympathetic nervous activity. This response might be used to determine the target ablation site and end point of renal artery RF ablation.

Abstract Aims The prognostic value of genetic variants for predicting lethal arrhythmic events (LAEs) in Brugada syndrome (BrS) remains controversial. We investigated whether the functional curation of SCN5A variations improves prognostic predictability. Methods and results Using a heterologous expression system and whole-cell patch clamping, we functionally characterized 22 variants of unknown significance (VUSs) among 55 SCN5A mutations previously curated using in silico prediction algorithms in the Japanese BrS registry (n = 415). According to the loss-of-function (LOF) properties, SCN5A mutation carriers (n = 60) were divided into two groups: LOF-SCN5A mutations and non-LOF SCN5A variations. Functionally proven LOF-SCN5A mutation carriers (n = 45) showed significantly severer electrocardiographic conduction abnormalities and worse prognosis associated with earlier manifestations of LAEs (7.9%/year) than in silico algorithm-predicted SCN5A carriers (5.1%/year) or all BrS probands (2.5%/year). Notably, non-LOF SCN5A variation carriers (n = 15) exhibited no LAEs during the follow-up period. Multivariate analysis demonstrated that only LOF-SCN5A mutations and a history of aborted cardiac arrest were significant predictors of LAEs. Gene-based association studies using whole-exome sequencing data on another independent SCN5A mutation-negative BrS cohort (n = 288) showed no significant enrichment of rare variants in 16 985 genes including 22 non-SCN5A BrS-associated genes as compared with controls (n = 372). Furthermore, rare variations of non-SCN5A BrS-associated genes did not affect LAE-free survival curves. Conclusion In vitro functional validation is key to classifying the pathogenicity of SCN5A VUSs and for risk stratification of genetic predictors of LAEs. Functionally proven LOF-SCN5A mutations are genetic burdens of sudden death in BrS, but evidence for other BrS-associated genes is elusive.

Abstract Allogeneic bone marrow transplantation (allo-BMT) is associated with both graft-versus-host disease (GVHD) and graft-versus-leukemia (GVL) effect. In the present study, we examined the contribution of cytotoxic effector mechanisms, which are mediated by tumor necrosis factor- (TNF-), Fas ligand (FasL), or perforin, to GVHD and GVL effect in a murine BMT model. Bone marrow cells plus spleen cells (BMS) from wild-type, FasL-defective, or perforin-deficient donors were transferred into lethally irradiated recipients in the parent (C57BL/6) to F1 (C57BL/6 × DBA/2) BMT model with or without prior inoculation of DBA/2 leukemia L1210 or P815 mast cytoma cells. The effect of anti–TNF- antibody administration was also examined. Whereas the defect or blockade of each cytotoxic pathway could ameliorate lethal acute GVHD, the GVL effect was differentially affected. The wild-type BMS recipients died of acute GVHD within 50 days without residual leukemia cells. The FasL-defective BMS recipients showed 60%&amp;lt; survival over 80 days without acute GVHD or residual leukemia cells. Administration of anti–TNF- antibody resulted in early leukemia relapse and the recipients died within 25 days with massive leukemia infiltration in the liver. The perforin-deficient BMS recipients died within 60 days with residual leukemia cells. These results suggest that blockade of the Fas/FasL pathway could be used for ameliorating GVHD without impairing GVL effect in allo-BMT.

Abstract —The expression of coxsackievirus and adenovirus receptor (CAR) was dominant in the brains and hearts of mice until the newborn phase. There is no detailed information concerning the relation between the expression of CAR and development of hearts. It is also uncertain whether CAR is able to be induced in adult hearts after cardiac injury. We demonstrated that CAR was abundant in the hearts of newborn rats but was barely detectable in the hearts of adult rats. The expression of CAR in rat hearts with experimental autoimmune myocarditis, which was induced by immunization of purified cardiac myosin, was serially investigated. Active myocarditis was observed from day 15 after immunization. By immunohistochemistry, cardiomyocytes were strongly stained for CAR antibody from days 24 to 42. CAR mRNA was also detected from days 18 to 30 by using reverse transcription–polymerase chain reaction. In the next experiment, the induction of CAR on isolated cardiomyocytes was investigated. CAR was barely detectable in cultured cardiomyocytes by Western blot analysis after isolation. This molecule gradually appeared along with the creation of clusters and beating of cardiomyocytes. Furthermore, the induction of CAR in cultured cardiomyocytes increased after supplement with conditioned medium of rat splenocytes activated by concanavalin A. In conclusion, rat CAR is expressed strongly in the hearts of newborn rats and is suppressed in those of adult rats. The expression of CAR is enhanced during the active phase of experimental autoimmune myocarditis and is induced by inflammatory mediators. CAR may play a role in cell-to-cell contact and adhesion of cardiomyocytes.

Cardiac sarcoidosis is increasingly recognized and is associated with poor prognosis. Ventricular tachycardia (VT) associated with cardiac sarcoidosis is the most likely cause of sudden death in most patients, but the mechanism has not been well established.This study investigated the mechanisms and outcome of VT associated with cardiac sarcoidosis.The study included eight consecutive patients (five men, three women, aged 54 +/- 19 years) who had sustained monomorphic VT associated with cardiac sarcoidosis in our hospital.The average ejection fraction was 43 +/- 11%. Twenty-two VTs were observed in these patients, and mean heart rate during VT was 192 +/- 29 beats/min (range 144-259). The phenomenon of transient entrainment was documented in 10 of 22 (45%) VTs by ventricular pacing (eight in the active phase). Another five (23%) VTs could not be entrained, but could be initiated by programmed stimulation and terminated by rapid pacing, reproducibly. In 3 of the 22 (14%) VTs, cardioversion was required urgently because of the fast rate, while the remaining 4 (18%) could be induced during electrophysiologic study.In this study, there was a high possibility that the mechanism of 15 (68%) VTs was reentry. Reentrant substrate is formed not only in association with the healing of cardiac granulomas in the inactive phase of cardiac sarcoidosis but also in the active phase. Ventricular tachycardia with cardiac sarcoidosis, even if this mechanism is reentry, has different inducibility between the active and inactive phases in an electrophysiologic study. This makes the therapy for cardiac sarcoidosis (e.g., corticosteroids, antiarrhythmic agents, and catheter ablation) difficult. The implantable cardioverter-defibrillator is an effective treatment for ventricular tachyarrythmia with cardiac sarcoidosis.

The Center for Epidemiologic Studies Depression Scale (CES-D) is used at workplaces to screen depressive disorders. The aim of this study was to examine the validity of the CES-D for depression in a workplace.The CES-D was administered to 2,219 workers (84.2% men; age 21-68 years) at a manufacturing company in Japan. Concomitantly all workers had an interview with the Mini International Neuropsychiatric Interview (MINI) as a gold standard for diagnosing major depressive disorder (MDD). The validity was evaluated by a receiver operating characteristic (ROC) curve.The area under the ROC curve of the CES-D was 0.96 [95% Confidence Interval (CI): 0.94-0.99]. The optimal cut-off score of MDD was 19 for screening.The validity of CES-D is confirmed and it is a valid instrument for detecting MDD in working populations in Japan.

Background There have been a number of recent reports on the use of autologous bone marrow implantation (BMI) in the treatment of peripheral arterial disease, with a clinical response rate of approximately 70%. However, the factors that influence efficacy have not yet been clarified. We have analyzed the relationship between the number of implanted bone marrow cells and the clinical efficacy of BMI. Methods and Results Eight patients with arteriosclerosis obliterans were treated with BMI. Bone marrow was aspirated from the ilium (500-1,000 ml), the mononuclear cells were separated and then were implanted. The clinical effectiveness of BMI was evaluated by assessing changes in the ankle-brachial pressure index (ABI) and the transcutaneous oxygen pressure (TcO2) between the pre-treatment baseline, with follow-up testing at 4 weeks. These changes were defined as ΔABI and ΔTcO 2. The mean number of CD34-positive cells was 1.04±0.60 ×106 /kg body weight. There was a strong correlation between the number of CD34-positive cells and ΔABI (r=0.754, p=0.028). Conclusions It is likely that the number of implanted CD34-positive cells is one of the primary factors that influence the clinical efficacy of BMI. (Circ J 2004; 68: 1189 - 1193)

We sought to compare the arrhythmic risk and sensitivity to sympathetic stimulation of mutations located in transmembrane regions and C-terminal regions of the KCNQ1channel in the LQT1 form of congenital long QT syndrome (LQTS). The LQT1 syndrome is frequently manifested with variable expressivity and incomplete penetrance and is much more sensitive to sympathetic stimulation than the other forms. Sixty-six LQT1 patients (27 families) with a total of 19 transmembrane mutations and 29 patients (10 families) with 8 C-terminal mutations were enrolled from five Japanese institutes. Patients with transmembrane mutations were more frequently affected based on electrocardiographic (ECG) diagnostic criteria (82% vs. 24%, p < 0.0001) and had more frequent LQTS-related cardiac events (all cardiac events: 55% vs. 21%, p = 0.002; syncope: 55% vs. 21%, p = 0.002; aborted cardiac arrest or unexpected sudden cardiac death: 15% vs. 0%, p = 0.03) than those with C-terminal mutations. Patients with transmembrane mutations had a greater risk of first cardiac events occurring at an earlier age, with a hazard ratio of 3.4 (p = 0.006) and with an 8% increase in risk per 10-ms increase in corrected Q-Tend. The baseline ECG parameters, including Q-Tend, Q-Tpeak, and Tpeak-end intervals, were significantly greater in patients with transmembrane mutations than in those with C-terminal mutations (p < 0.005). Moreover, the corrected Q-Tend and Tpeak-end were more prominently increased with exercise in patients with transmembrane mutations (p < 0.005). In this multicenter Japanese population, LQT1 patients with transmembrane mutations are at higher risk of congenital LQTS-related cardiac events and have greater sensitivity to sympathetic stimulation, as compared with patients with C-terminal mutations.

Atrial fibrillation (AF) is the most common clinical arrhythmia and a major cause of cardiovascular morbidity and mortality. Among the gene defects previously associated with AF is a gain of function of the slowly activating delayed rectifier potassium current IKs, secondary to mutations in KCNQ1. Coexpression of KCNE5, the gene encoding the MiRP4 beta-subunit, has been shown to reduce IKs.The purpose of this study was to test the hypothesis that mutations in KCNE5 are associated with AF in a large cohort of patients with AF.One-hundred fifty-eight patients with AF were screened for mutations in the coding region of KCNE5.A missense mutation involving substitution of a phenylalanine for leucine at position 65 (L65F) was identified in one patient. This patient did not have a history of familial AF, and neither KCNQ1 nor KCNE2 mutations were found. Transient transfection of Chinese hamster ovary (CHO) cells expressing IKs(KCNQ1+KCNE1) with KCNE5 suppressed the developing and tail currents of IKs in a concentration-dependent manner. Transient transfection with KCNE5-L65F failed to suppress IKs, yielding a current indistinguishable from that recorded in the absence of KCNE5. Developing currents recorded during a test pulse to +60 mV and tail currents recorded upon repolarization to -40 mV both showed a significant concentration-dependent gain of function in IKs with expression of KCNE5-L65F vs KCNE5-WT.The results of this study suggest that a missense mutation in KCNE5 may be associated with nonfamilial or acquired forms of AF. The arrhythmogenic mechanism most likely is a gain of function of IKs.

On October 23, 2004, a major earthquake, which registered 6.8 on the Richter scale, occurred in Niigata Prefecture in Japan. Emotional stress is important as a trigger of transient left ventricular apical ballooning (so-called 'Takotsubo' cardiomyopathy), but its incidence and clinical profile immediately after a natural disaster have not been fully elucidated.'Takotsubo' cardiomyopathy was diagnosed in 16 patients (1 man, 15 women, mean age 71.5 years) within 1 month after the earthquake. Of them, 13 (81%) lived in areas where the Japan Meteorological Agency seismic intensity scale registered 6 or above, and 11 (69%) developed symptoms on the day of the earthquake. The incidence of 'Takotsubo' cardiomyopathy 1 month after the earthquake was approximately 24-fold higher near the epicenter than that before the earthquake.'Takotsubo' cardiomyopathy can occur on the day of the earthquake in elderly women living near the epicenter.

This study aimed to clarify detailed and serial electrocardiographic findings in patients with Takotsubo cardiomyopathy from onset to recovery. Nine consecutive women aged 65 to 84 years (mean 74) with Takotsubo cardiomyopathy were investigated. Standard 12-lead electrocardiograms were recorded during hospitalization and ST-segment elevation and T-wave inversion were manually measured daily in each patient. All 9 patients had 4 phases found electrocardiographically. Phase 1 was characterized by ST-segment elevation immediately after onset. Subsequently, T-wave inversion was observed from days 1 to 3 (phase 2), then inverted T waves improved transiently from days 2 to 6 (phase 3). After this phase, giant inverted T waves with QT prolongation appeared and persisted > or =2 months until recovery (phase 4). Serum creatine kinase levels were increased only at onset. Left ventricular wall motion abnormalities evaluated using echocardiography improved gradually after phase 3 in all patients. Second T-wave inversions (phase 4) were significantly deeper than those of the first one (phase 2; p <0.05). In conclusion, 4 electrocardiographic phases in patients with Takotsubo cardiomyopathy were shown. This observation may be helpful to understand the pathophysiologic process of Takotsubo cardiomyopathy.

Abstract IL-22 is one of several cytokines with limited homology to IL-10. However, the biological activities of IL-22 are mostly unknown. The purpose of this study was to evaluate the effect of IL-22 on rat experimental autoimmune myocarditis (EAM) and elucidate an aspect of the biological activities of IL-22. Rats were immunized on day 0; IL-22-Ig-treated rats were injected with pCAGGS-IL-22-Ig and control rats with pCAGGS-Ig using hydrodynamics-based gene delivery on day 1 or day 6. IL-22-Ig gene therapy administered on day 1 or day 6 after immunization was effective in controlling EAM as monitored by the heart weight to body weight ratio, and the myocarditis area in rats was sacrificed on day 17. Examination of the expression of IL-22-related genes in purified cells from EAM hearts suggested that IL-22-Ig acting target cells were noncardiomyocytic (NC) noninflammatory cells such as fibroblasts, smooth muscle cells, and endothelial cells. Therefore, we examined the effect of rIL-22 or serum containing IL-22-Ig on the expression of immune-relevant genes in IL-1-stimulated NC cells cultured from EAM hearts. Results showed that the expression of immunologic molecules (PGE synthase, cyclooxygenase-2, MIP-2, MCP-1, IL-6, and cytokine-induced neutrophil chemoattractant-2) in IL-1-stimulated NC cells was significantly decreased by rIL-22 or serum containing IL-22-Ig. EAM was suppressed by hydrodynamics-based delivery of plasmid DNA encoding IL-22-Ig, and the reason for this effectiveness may be that IL-22 suppressed gene expression of PG synthases, IL-6, and chemokines in activated NC noninflammatory cells.

Hypertrophic cardiomyopathy (HCM), which is inherited as an autosomal dominant trait, is the most prevalent hereditary cardiac disease. Although there are several reports on the systematic screening of mutations in the disease-causing genes in European and American populations, only limited information is available for Asian populations, including Japanese.Genetic screening of disease-associated mutations in 8 genes for sarcomeric proteins, MYH7, MYBPC3, MYL2, MYL3, TNNT2, TNNI3, TPM1, and ACTC, was performed by direct sequencing in 112 unrelated Japanese proband patients with familial HCM; 37 different mutations, including 13 novel ones in 5 genes, MYH7, MYBPC3, TNNT2, TNNI3, and TPM1, were identified in 49 (43.8%) patients. Among them, 3 carried compound heterozygous mutations in MYBPC3 or TNNT2. The frequency of patients carrying the MYBPC3, MYH7, and TNNT2 mutations were 19.6%, 10.7%, and 8.9%, respectively, and the most frequently affected genes in the northeastern and southwestern parts of Japan were MYBPC3 and MYH7, respectively. Several mutations were found in multiple unrelated proband patients, for which the geographic distribution suggested founder effects of the mutations.This study demonstrated the frequency and distribution of mutations in a large cohort of familial HCM in Japan.

Bradycardia is a trigger of ventricular arrhythmias in patients with arrhythmia including Brugada syndrome and long QT syndrome. The HCN4 channel controls the heart rate, and its mutations predispose to inherited sick sinus syndrome and long QT syndrome associated with bradycardia. We found a 4 base-insertion at the splice donor site of the HCN4 gene in a patient with idiopathic ventricular tachycardia, which was supposed to generate a truncated channel. To investigate the role of the HCN4 channel in ventricular arrhythmia, we introduced a ventricular action potential of I(f) channel produced by HCN4 in a computer simulation model and found that the I(f) channel generated a leaky outward current during the plateau phase of ventricular action potential. Currents through the I(f) channel were suggested to contribute to the shortening of the action potential duration and the prevention of early after-depolarization in bradycardia. These observations suggested that the HCN4 channel played a preventive role in triggering bradycardia-induced ventricular arrhythmias.

Background: Lipocalin-2/neutrophil gelatinase-B associated lipocalin (Lcn2/NGAL) is involved in the transport of iron and seems to play an important role in inflammation. A recent study has reported that it is also expressed in the failing heart and may be a biomarker not only for renal failure but also for heart failure. Because Lcn2/NGAL is thought to be induced by interleukin-1, it might be strongly induced in the presence of myocarditis. Methods and Results: This study investigated the expression of Lcn2/NGAL in rat experimental autoimmune myocarditis (EAM) and in human myocarditis. In EAM hearts, the expression of Lcn2/NGAL was markedly increased (>100-fold at an early stage), and in human myocarditis it was also highly expressed compared with non-inflammatory failing hearts. Lcn2/NGAL expressing cells in hearts with EAM and human myocarditis were identified as cardiomyocytes, vascular wall cells, fibroblasts and neutrophils. Lcn2/NGAL in EAM rats was also expressed in the liver. Plasma Lcn2/NGAL levels abruptly increased at an early stage of EAM, and high levels were initially sustained during the inflammatory stage, then decreased with recovery. In contrast, levels of B-type natriuretic peptide increased only slowly as the disease progressed. Conclusions: Cardiomyocytes, vascular wall cells and fibroblasts in myocarditis strongly express Lcn2/NGAL via proinflammatory cytokines. (Circ J 2010; 74: 523 - 530)

The beneficial effects of telmisartan on Angiotensin (Ang)-II mediated oxidative stress and renal fibrosis in streptozotocin (STZ)-induced diabetic nephropathy (DN) were studied. Thirty mice were divided into normal (NG), STZ-induced diabetic (DG) and telmisartan-treated diabetic (TG) groups. Compared with NG mice, DG mice showed significant up-regulations of AT-1R, TGF-β1, p-p38MAPK, p-MAPKAPK-2, p-Akt, p47phox, p67phox, gp91phox protein and collagen-III and all of these were significantly reversed in TG mice. The down-regulated protein expression of Ang-(1–7) mas receptor, ACE-2, PPAR-γ and PGC-1α were observed in DG mice and a significant up-regulation effect of telmisartan has been seen in the TG mice. Furthermore, TG mice showed reduced expression of fibronectin, production of superoxide radical as well as renal hypertrophy and fibrosis when compared with DG mice. These findings suggest that Ang-II plays a significant role in DN and telmisartan would be beneficial in reducing oxidative stress and fibrosis in STZ-induced DN.

AimsAccumulating data show the efficacy of catheter ablation (CA) for atrial fibrillation (AF); however, postoperative recurrence is not uncommon. The aim of this study was to identify predictors of AF recurrence in patients undergoing CA.

This study sought to characterize patients with idiopathic ventricular fibrillation (IVF) who develop electrical storms.Some IVF patients develop ventricular fibrillation (VF) storms, but the characteristics of these patients are poorly known.Ninety-one IVF patients (86% male) were selected after the exclusion of structural heart diseases, primary electrical diseases, and coronary spasm. Electrocardiogram features were compared between the patients with and without electrical storms. A VF storm was defined as VF occurring ≥3 times in 24 h and J waves >0.1 mV above the isoelectric line in contiguous leads.Fourteen (15.4%) patients had VF storms occurring out-of-hospital at night or in the early morning. J waves were more closely associated with VF storms compared to patients without VF storms: 92.9% versus 36.4% (p < 0.0001). VF storms were controlled by intravenous isoproterenol, which attenuated the J-wave amplitude. After the subsidence of VF storms, the J waves decreased to the nondiagnostic level during the entire follow-up period. Implantable cardioverter-defibrillator therapy was administered to all patients during follow-up. Quinidine therapy was limited, but the patients on disopyramide (n = 3), bepridil (n = 1), or isoprenaline (n = 1) were free from VF recurrence, while VF recurred in 5 of the 9 patients who were not given antiarrhythmic drugs.The VF storms in the IVF patients were highly associated with J waves that showed augmentation prior to the VF onset. Isoproterenol was effective in controlling VF and attenuated the J waves, which diminished to below the diagnostic level during follow-up. VF recurred in patients followed up without antiarrhythmic agents.

Electrical storms (ESs) in patients with Brugada syndrome (BrS) are rare though potentially lethal.We studied 22 men with BrS and ES, defined as ≥3 episodes/d of ventricular fibrillation (VF) and compared their characteristics with those of 110 age-matched, control men with BrS without ES. BrS was diagnosed by a spontaneous or drug-induced type 1 pattern on the ECG in the absence of structural heart disease. Early repolarization (ER) was diagnosed by J waves, ie, >0.1 mV notches or slurs of the terminal portion of the QRS complex. The BrS ECG pattern was provoked with pilsicainide. A spontaneous type I ECG pattern, J waves, and horizontal/descending ST elevation were found, respectively, in 77%, 36%, and 88% of patients with ES, versus 28% (P<0.0001), 9% (P=0.003), and 60% (P=0.06) of controls. The J-wave amplitude was significantly higher in patients with than without ES (P=0.03). VF occurred during undisturbed sinus rhythm in 14 of 19 patients (74%), and ES were controlled by isoproterenol administration. All patients with ES received an implantable cardioverter defibrillator and over a 6.0±5.4 years follow-up, the prognosis of patients with ES was significantly worse than that of patients without ES. Bepridil was effective in preventing VF in 6 patients.A high prevalence of ER was found in a subgroup of patients with BrS associated with ES. ES appeared to be suppressed by isoproterenol or quinidine, whereas bepridil and quinidine were effective in the long-term prevention of VF in the highest-risk patients.

Treatment guidelines for atrial fibrillation (AF) used in Western countries describe female gender as a risk factor for thromboembolic events in patients with nonvalvular AF (NVAF). The present study aimed to determine the impact of gender on prognosis of Japanese patients with NVAF. A subanalysis of 7,406 patients with NVAF (mean age 70 years) who were followed-up prospectively for 2 years was performed using data from the J-RHYTHM registry. The primary end points were thromboembolic events, major hemorrhaging, total mortality, and cardiovascular mortality. Compared with male subjects (n = 5,241), female subjects (n = 2,165) were older and displayed greater prevalences of paroxysmal AF, heart failure, and hypertension but less prevalences of diabetes, previous cerebral infarction, and coronary artery disease. Male and female patients had mean CHADS2 (Congestive heart failure, Hypertension, Age of 75 years or more, Diabetes mellitus and prior Stroke or transient ischemic attack) scores of 1.6 and 1.8, respectively (p <0.001). Warfarin was given to 87% of male patients and 86% of female patients (p = 0.760), and the 2 genders displayed similar mean international normalized ratio of prothrombin time values at baseline (1.91 vs 1.90, respectively, p = 0.756). Multivariate logistic regression analysis indicated that male gender was an independent risk factor for major hemorrhaging (odds ratio 1.59, 95% confidence interval 1.05 to 2.40, p = 0.027) and all-cause mortality (odds ratio 1.78, 95% confidence interval 1.25 to 2.55, p <0.002) but not for thromboembolic events (odds ratio 1.24, 95% confidence interval 0.83 to 1.86, p = 0.297) or cardiovascular mortality (odds ratio 0.96, 95% confidence interval 0.56 to 1.66, p = 0.893). In conclusion, female gender is not a risk factor for thromboembolic events among Japanese patients with NVAF who were treated mostly with warfarin. However, male gender is a risk factor for major hemorrhaging and all-cause mortality.

Objective Whether intracerebral hemorrhage (ICH) associated with non–vitamin K antagonist oral anticoagulants (NOAC‐ICH) has a better outcome compared to ICH associated with vitamin K antagonists (VKA‐ICH) is uncertain. Methods We performed a systematic review and individual patient data meta‐analysis of cohort studies comparing clinical and radiological outcomes between NOAC‐ICH and VKA‐ICH patients. The primary outcome measure was 30‐day all‐cause mortality. All outcomes were assessed in multivariate regression analyses adjusted for age, sex, ICH location, and intraventricular hemorrhage extension. Results We included 7 eligible studies comprising 219 NOAC‐ICH and 831 VKA‐ICH patients (mean age = 77 years, 52.5% females). The 30‐day mortality was similar between NOAC‐ICH and VKA‐ICH (24.3% vs 26.5%; hazard ratio = 0.94, 95% confidence interval [CI] = 0.67–1.31). However, in multivariate analyses adjusting for potential confounders, NOAC‐ICH was associated with lower admission National Institutes of Health Stroke Scale (NIHSS) score (linear regression coefficient = −2.83, 95% CI = −5.28 to −0.38), lower likelihood of severe stroke (NIHSS &gt; 10 points) on admission (odds ratio [OR] = 0.50, 95% CI = 0.30–0.84), and smaller baseline hematoma volume (linear regression coefficient = −0.24, 95% CI = −0.47 to −0.16). The two groups did not differ in the likelihood of baseline hematoma volume &lt; 30cm 3 (OR = 1.14, 95% CI = 0.81–1.62), hematoma expansion (OR = 0.97, 95% CI = 0.63–1.48), in‐hospital mortality (OR = 0.73, 95% CI = 0.49–1.11), functional status at discharge (common OR = 0.78, 95% CI = 0.57–1.07), or functional status at 3 months (common OR = 1.03, 95% CI = 0.75–1.43). Interpretation Although functional outcome at discharge, 1 month, or 3 months was comparable after NOAC‐ICH and VKA‐ICH, patients with NOAC‐ICH had smaller baseline hematoma volumes and less severe acute stroke syndromes. Ann Neurol 2018;84:702–712

Summary Background Patients with chronic hepatitis C are often complicated by chronic kidney disease ( CKD ). Aim To evaluate the efficacy, safety and pharmacokinetics of glecaprevir/pibrentasvir in patients with severe renal impairment. Methods In a prospective, multicentre study involving 35 medical institutions, 832 genotype 1‐3 patients were treated with glecaprevir/pibrentasvir. The efficacy and safety of glecaprevir/pibrentasvir were analysed for patients with CKD stage 4 or 5. Multivariate analysis was performed to identify the factors associated with the most frequently observed adverse event. In patients undergoing haemodialysis, a pharmacokinetic study was conducted to investigate the dialysability of the drugs: plasma samples were obtained from the arterial and venous sides of a dialyser to serially measure drug concentrations. Results The subjects comprised 141 patients (32 with CKD stage 4 and 109 with CKD stage 5), of whom 100 were undergoing haemodialysis. All but one stage 5 CKD patients undergoing haemodialysis achieved sustained virologic response (99.3%). Adverse events were observed in 39.7% of subjects: pruritus was the most frequent (30.5%), and was significantly associated with haemodialysis. In the pharmacokinetic study, no arterial‐venous differences in the plasma concentrations of glecaprevir/pibrentasvir were detected during the haemodialysis sessions. Conclusions Glecaprevir/pibrentasvir was highly effective and safe in chronic hepatitis C patients with severe renal impairment. Haemodialysis was associated with increased incidence of pruritus, which was the most frequent adverse event, but had little or no influence on the drug concentrations, which indicated that their dialysability is very low and that no dose modification is required in patients undergoing haemodialysis. ( UMIN registration no. 000032073).

Thyroid hormones (THs) are synthesized in the thyroid gland, and they circulate in the blood to regulate cells, tissues, and organs in the body. In particular, they exert several effects on the cardiovascular system. It is well known that THs raise the heart rate and cardiac contractility, improve the systolic and diastolic function of the heart, and decrease systemic vascular resistance. In the past 30 years, some researchers have studied the molecular pathways that mediate the role of TH in the cardiovascular system, to better understand its mechanisms of action. Two types of mechanisms, which are genomic and non-genomic pathways, underlie the effects of THs on cardiomyocytes. In this review, we summarize the current knowledge of the action of THs in the cardiac function, the clinical manifestation and parameters of their hemodynamics, and treatment principles for patients with hyperthyroid- or hypothyroid-associated heart disease. We also describe the cardiovascular drugs that induce thyroid dysfunction and explain the mechanism underlying the thyroid toxicity of amiodarone, which is considered the most effective antiarrhythmic agent. Finally, we discuss the recent reports on the involvement of thyroid hormones in the regulation of myocardial regeneration and metabolism in the adult heart.

Ribonuclease protection assay was used to demonstrate mRNA expression of several cytokines as well as inducible NO synthase (iNOS), constitutive endothelial NO synthase (cNOS) and perforin in the myocardium during the course of experimental autoimmune myocarditis (EAM) in rats. Interleukin 2 (IL-2) appeared in the initial inflammatory phase (day 14), subsided in the maximum inflammatory phase (day 19) and disappeared by the recovery phase (day 25). mRNA of IL-3 beta, interferon gamma INF-gamma and tumor necrosis factor alpha (TNF-alpha) were detected only in the maximum inflammatory phase and iNOS also appeared for several days at this time. In contrast. IL-10 mRNA was detected after the maximum inflammatory stage and persisted into the recovery phase (days 25-36). Although transforming growth factor beta 1 (TGF-beta 1) could be detected in all phases, the expression was markedly enhanced in the maximum inflammatory phase and gradually diminished (around day 36) to basal levels. Perforin mRNA was not detected at any point in the disease. Besides macrophages and CD4 T cells, a number of neutrophils were found in the myocardium especially at peak inflammatory stage. We suggest that antigen (Ag) primed Ag presenting cells or macrophages interact with T cells (Th1) to produce IL-2 and subsequent IFN-gamma, which further activates macrophages in the myocardium. Consequently, TNF-alpha and iNOS may inflict tissue damage to myocardium. It is also suggested that TGF-beta) and one representative Th2 cytokine, IL-10, help inhibit inflammation. These findings suggest that Th1 and Th2 cytokines are produced at different stages of EAM and modulate the inflammation and the course of EAM.

Hepatocellular carcinoma (HCC) occurs more frequently in patients with hepatitis C virus (HCV)-related chronic liver disease than those with hepatitis B virus-related disease. It is important to assess the factors affecting the development of HCC.A long term follow-up study involving patients with chronic HCV was performed retrospectively. A total of 153 patients diagnosed between June 1981 and November 1990 with chronic HCV with or without cirrhosis by liver biopsy were enrolled in a long term follow-up study (average, 99.4 months) and the cumulative incidence rate of HCC and factors affecting the appearance of HCC were examined.The 5-year cumulative incidence rate was 9%, the 10-year cumulative incidence rate was 23%, and the 15-year cumulative incidence rate was 42%. The annual rate of incidence increased as the follow-up period progressed. The authors selected ten variables and investigated their effect on the incidence rate of HCC, including age, gender, habitual heavy drinking, positivity of antibody against hepatitis B virus surface antigen, treatment with interferon (IFN) during the follow-up period, maximum and minimum serum alanine aminotransferase levels during the follow-up period, histologic staging, grading, and irregular regeneration of hepatocytes. Of the 10 variables, age (> 50 years), habitual heavy drinking, and histologic staging were determined to be independent risk factors according to multivariate Cox proportional hazards regression analysis. IFN therapy by itself was not found to be an independent factor affecting the appearance of HCC.In patients with chronic HCV, the annual incidence rate of HCC appeared to increase as the follow-up period progressed. According to the results of the current study, the factors that independently affected the development of HCC were age, habitual heavy drinking, and histologic staging.

Abstract Expression of CD134 (OX40) on activated CD4+ T cells has been observed in acute graft-versus-host disease (GVHD) after human and rat allogeneic bone marrow transplantation (BMT). We investigated the role of interaction between CD134 and CD134 ligand (CD134L) in a murine model of acute GVHD by using a newly established monoclonal antibody (mAb) against murine CD134L. Acute GVHD was induced by transfer of bone marrow cells and spleen cells into lethally irradiated recipients in a parent (C57BL/6) to first filial generation (C57BL/6 crossed with DBA/2) BMT. Administration of anti-CD134L mAb significantly reduced the lethality of acute GVHD and other manifestations of the disease, such as loss of body weight, hunched posture, diarrhea, and patchy alopecia. The survival rate 80 days after BMT in mice treated with the mAb was about 70%, whereas all mice treated with control antibodies died within 43 days. Histologic examinations revealed that inflammatory changes in target organs such as the liver, gut, and skin were also ameliorated in mice treated with the mAb compared with control mice. An in vitro assay of T-cell proliferation showed a marked hyporesponsiveness to host alloantigen in samples from mice treated with anti-CD134L mAb. In addition, low levels of interferon γ and transiently elevated levels of interleukin 4 and IgE in serum samples were found in mice treated with anti-CD134L mAb. These results suggest that CD134-CD134L interactions have an important role in the pathogenesis of acute GVHD.

Clinical manifestations of acute myocarditis, with distinct onset, vary from asymptomatic to fatal. The predictors of the course of the disease in patients with acute myocarditis at initial presentation have not yet been established. In this study, we examined the predictive values of various parameters in the disease course of patients with myocarditis.Twenty-one consecutive patients who had been diagnosed as having acute myocarditis by histological examinations were analyzed. The patients with myocarditis were divided into the survival group (n=13) and the fatal group (n=8). We examined the parameters of the clinical state, hemodynamic variables, required therapies, biochemical laboratory data, and cytokines. The control groups were composed of 23 patients with old myocardial infarction and 20 healthy volunteers. The fatal group had lower blood pressure and higher pulmonary capillary wedge pressure compared with those values in the survival group. Mechanical ventilation support was more frequently required in the fatal group. Serum levels of soluble Fas (sFas) and soluble Fas ligand (sFasL) were significantly higher in the myocarditis group than in the 2 control groups. Furthermore, levels were significantly higher in the fatal group than in the survival group for sFas (13.93+/-4.77 versus 3.77+/-0.52 ng/mL, respectively; P:<0.001) and sFasL (611.4+/-127.7 versus 269.5+/-37.3 pg/mL, respectively; P:<0.05). Other clinical states, hemodynamic variables, required therapies, and biochemical laboratory parameters were not different between the 2 groups.Elevation of sFas and sFasL levels at initial presentation appear to be a good serological marker to predict the prognosis of acute myocarditis.

Autoimmunity after viral myocarditis is considered to be one of the causes of dilated cardiomyopathy. Cytokines are assumed to play an important role in the pathogenesis. We recently reported that interleukin (IL)-2 and interferon (IFN)-gamma mRNA are expressed in the myocardium of rats with experimental autoimmune myocarditis (EAM). However, the role of cytokines in autoimmune myocardial injury in detail is still not clear. Reverse transcription-polymerase chain reaction identified IL-12 (p40) mRNA in antigen-presenting cells in the initial phase of EAM. Cardiac myosin-specific T lymphocytes (MSTLs) were cultured with cardiac myosin peptide (CMP) in the presence of IL-2 and/or IL-12 and were transferred to other naive rats. The results showed that EAM could be effectively induced by transfer of MSTLs cultured with IL-12, whereas transfer of MSTLs cultured with IL-2 was less effective. However, IL-2 acts synergistically with IL-12, and MSTLs cultured with both cytokines most efficiently induce EAM. In vitro experiments showed that MSTLs cultured with both IL-12 and IL-2 produced a much greater amount of IFN-gamma than did MSTLs cultured with either IL-12 or IL-2 alone. The amount of IFN-gamma production was correlated with pathogenicity of MSTLs. Transfer experiments after sorting further demonstrated that the transfer was affected by CD4+ helper T (Th) cells but not by CD8+ cytotoxic T lymphocytes. IL-12 and IL-2 synergistically enhance the pathogenicity of MSTLs. Furthermore, a type 1 Th (Th1) cytokine, IFN-gamma, which is a potent regulatory cytokine of autoimmunity, is produced by MSTLs. IL-12 and IL-2 potentiate the expansion of cardiac myosin-specific Th1 cells and play an important role in the development of autoimmune myocardial injury.

Information concerning the electrophysiological characteristics of the reentrant circuit is still limited. To understand the incidence and mechanism of pacing-induced interruption of ventricular tachycardia (VT), rapid pacing was performed to entrain VT, and the local electrogram at the VT origin and the surface electrocardiogram were analyzed.Among 25 patients, evidence of transient entrainment was confirmed in 20 patients, but the critical paced cycle length at which VT was interrupted was obtained in 13 patients when the paced cycle length was decreased in steps of 10 msec. During pacing at the critical cycle length (defined as block cycle length), changes in the local electrogram at VT origin were confirmed in all of the 13 patients; that is, 1) a change in morphology and 2) a change in the timing of activation: a sudden shortening in the stimulus to local electrogram time (third entrainment criterion by Waldo). The two changes mean that the exit is activated from a different direction (retrograde capture) because of an orthodromic block in the slow conduction zone. The QRS complex in the surface electrocardiogram showed a change in configuration from the fusion complex to the fully paced one at the same time when the exit was captured antidromically.Based on our observations in these patients, ventricular tachycardia interruption is very often associated with orthodromic block in the reentrant circuit at a critical cycle length of rapid pacing.

The genomic aberrations in extra nodal marginal zone B cell lymphoma vary according to their anatomical origin. This polarization is a reflection of the participation of different genes in the lymphomagenesis of marginal zone B cell lymphoma. We previously demonstrated by means of genome-wide array comparative genomic hybridization (CGH) that the genomic profile of ocular adnexal marginal zone B cell lymphoma is distinct from that of pulmonary or nodal marginal zone B cell lymphoma. The novel finding was a recurrent deletion of a 2.9-Mb region at chromosome band 6q23.3-q24.1, including homozygous loss, in ocular adnexal marginal zone B cell lymphoma. For a more detailed examination of the deletions of 6q23.3-24.1, we used contig bacterial artificial chromosome (BAC) array CGH, containing 24 BAC clones covering the 2.9-Mb region, to analyze nine cases with 6q23.3-q24.1 loss. We narrowed the minimal common region down to a length of 586 kb with two genes and four expressed sequence tags (ESTs). All of these genes and ESTs were subjected to RT-PCR and real-time quantitative RT-PCR. Correlation between genomic loss and expression level was found only for TNFAIP3, demonstrating that TNFAIP3 is a target gene of 6q deletion in ocular adnexal marginal zone B cell lymphoma. TNFAIP3 is an inhibitor of NF-kB signaling so that loss of this gene may play an important role in lymphomagenesis and suggests that TNFAIP3 may act as a tumor suppressor gene in ocular adnexal marginal zone B cell lymphoma.

Mammalian 14-3-3 proteins are dimeric phosphoserine-binding proteins that participate in signal transduction and regulate several aspects of cellular biochemistry. Diabetic cardiomyopathy is associated with increased oxidative stress and inflammation. In order to study the pathogenic changes underlying diabetic cardiomyopathy, we examined the role of 14-3-3 protein and apoptosis signal-regulating kinase 1 (Ask1) signaling by using transgenic mice with cardiac-specific expression of a dominant-negative 14-3-3η protein mutant (DN 14-3-3η) after induction of experimental diabetes. The elevation in blood glucose was comparable between wild type (WT) and DN 14-3-3η mice. However, a marked downregulation of thioredoxin reductase was apparent in DN 14-3-3η mice compared to WT mice after induction of diabetes. Significant Ask1 activation in DN 14-3-3η after diabetes induction was evidenced by pronounced de-phosphorylation at Ser-967 and intense immunofluorescence observed in left ventricular (LV) sections. Echocardiographic analysis revealed that cardiac functions were notably impaired in diabetic DN 14-3-3η mice compared to diabetic WT mice. Marked increases in myocardial apoptosis, cardiac hypertrophy, and fibrosis were observed with a corresponding up-regulation of atrial natriuretic peptide and galectin-3, as well as a downregulation of sarcoendoplasmic reticulum Ca2+ ATPase2 expression. Furthermore, diabetic DN 14-3-3η mice displayed significant reductions of platelet-endothelial cell adhesion molecule-1 staining as well as endothelial nitric acid synthase and vascular endothelial growth factor expression. In conclusion, our data suggests that enhancement of 14-3-3 protein could provide a novel therapeutic strategy against hyperglycemia-induced left ventricular dysfunction and can limit the progression of diabetic cardiomyopathy by regulating Ask1 signaling.

The p38 mitogen-activated protein kinase (MAPK) is activated during heart diseases that might be associated with myocardial damage and cardiac remodeling process. Diabetic cardiomyopathy is associated with increased oxidative stress and inflammation. The purpose of this study was to investigate the role of p38alpha MAPK after experimental diabetes by using transgenic (TG) mice with cardiac-specific expression of a dominant-negative mutant form of p38alpha MAPK. The elevation of blood glucose was comparable between the nontransgenic (NTG) and TG mice. The expression of phospho-p38 MAPK and phospho-MAPK-activated protein kinase 2 levels were significantly suppressed in TG mice heart than in NTG mice after diabetes induction. Left ventricular (LV) dimension in systole was smaller, and the percent fractional shortening was higher in diabetic TG mice compared with diabetic NTG mice. In addition, diabetic TG mice had reduced cardiac myocyte diameter, content of cardiac fibrosis, LV tissue expressions of atrial natriuretic peptide, transforming growth factor beta1, and collagen III compared with diabetic NTG mice. Moreover, LV expression of NADPH oxidase subunits, p22(phox), p67(phox), gp91(phox), and Nox4, reactive oxygen species and lipid peroxidation levels were significantly increased in diabetic NTG mice, but not in diabetic TG mice. Furthermore, myocardial apoptosis, the number of caspase-3-positive cells, and the downregulation of antiapoptotic protein Bcl-X(L) were less in diabetic TG mice compared with diabetic NTG mice. In conclusion, our data establish that p38alpha MAPK activity is required for cardiac remodeling after diabetes induction and suggest that p38alpha MAPK may promote cardiomyocyte apoptosis by downregulation of Bcl-X(L).

The effect of regular hemodialysis (HD) with dialyzate containing acetate was evaluated in 20 patients. After dialysis, a significant increase in limb blood flow was found (P less than 0.01) while the mean arterial blood pressure remained unchanged indicating a significant decrease in peripheral vascular resistance after HD (P less than 0.01). Cardiac function was evaluated using the ratio of the preejection period to left ventricular ejection time (PEP/ET); this value showed a significant increase after HD suggesting depression of cardiac function (P less than 0.001). The study was repeated substituting bicarbonate for acetate in 13 of the 20 patients. Under these conditions, limb blood flow and peripheral vascular resistance showed no significant change though mean arterial blood pressure decreased significantly (P less than 0.01). The ratio, PEP/ET, showed a significant increase after HD (P less than 0.01), but the value was significantly lower than that found after HD with dialyzate containing acetate (P less than 0.05). Changes in the serum levels of calcium, potassium, pH and body weight could not explain the differences found after HD with the two kinds of dialyzate. The results of the present study suggest strongly that acetate exerts a depressant action on the cardio-vascular system.

Physical and psychological stress induced by catastrophic events such as earthquakes can lead to sudden death, acute coronary syndrome, stroke, and other cardiovascular diseases. We investigated the impact of the earthquake that occurred in Niigata, Japan, on pulmonary embolism. Pulmonary embolism increased to 9 cases in the 4 weeks after the earthquake, compared to 1 case in the 4 weeks before the earthquake, 2 cases in the corresponding 8 weeks in 2003, and 1 case in 2002. The first case occurred two days after the initial earthquake and new cases were reported for 27 days thereafter. Six of 9 patients (67%) took refuge in their automobiles before the onset of pulmonary embolism. Sudden death also increased after the earthquake and 7 of 22 cases (32%) spend night(s) in automobile. In conclusion, pulmonary embolism should be attended after disasters and prolonged immobilization in automobiles may increase risk of pulmonary embolism and sudden death.

Diabetes mellitus increases the risk of heart failure independently of underlying coronary artery disease, and many believe that diabetes leads to cardiomyopathy. The underlying pathogenesis is partially understood. Several factors may contribute to the development of cardiac dysfunction in the absence of coronary artery disease in diabetes mellitus. There is growing evidence that excess generation of highly reactive free radicals, largely due to hyperglycemia, causes oxidative stress, which further exacerbates the development and progression of diabetes and its complications. Hyperglycemia-induced oxidative stress is a major risk factor for the development of micro-vascular pathogenesis in the diabetic myocardium, which results in myocardial cell death, hypertrophy, fibrosis, abnormalities of calcium homeostasis and endothelial dysfunction. Diabetes-mediated biochemical changes show cross-interaction and complex interplay culminating in the activation of several intracellular signaling molecules. Diabetic cardiomyopathy is characterized by morphologic and structural changes in the myocardium and coronary vasculature mediated by the activation of various signaling pathways. This review focuses on the oxidative stress and signaling pathways in the pathogenesis of the cardiovascular complications of diabetes, which underlie the development and progression of diabetic cardiomyopathy.

Erythropoietin (EPO) has been found to have anti-apoptotic and tissue protective effects on the myocardium. The aim of the present pilot study was to observe the safety and efficacy of EPO administration for patients with acute myocardial infarction (AMI).Patients admitted with AMI had all undergone successful percutaneous coronary intervention (PCI). Patients were randomly assigned to 2 groups (control and EPO groups), and given 12,000 IU EPO iv or saline after PCI. The primary endpoints were the difference between the acute phase and chronic phase (6 months after the attack) regarding left ventricular function as measured on electrocardiogram-gated single-photon emission computed tomography. Thirty-six patients (control 16, EPO 20) were eligible for analysis. Left ventricular ejection fraction (LVEF) significantly increased in the EPO group (from 51.0+/-19.6% to 58.5+/-15.0%, P=0.0238), but not in the control group. Further analysis was separately undertaken in patients with occlusion in the left anterior descending artery (LAD) and others (non-LAD). LVEF was <50% in most patients in the LAD subgroup, and LVEF significantly increased in the EPO group (37.5+/-13.0 to 52.7+/-15.8, P=0.0049), but not in the control group. EPO administration did not trigger any adverse clinical events.EPO administration is a promising treatment for AMI.

A multicenter, randomized, placebo-controlled, double-blind trial was conducted with patients with persistent atrial fibrillation (AF) to determine the dose-response effects and safety of bepridil, using every-day transtelephonic monitorings.A total of 90 patients were randomized to receive placebo, 100 mg/day and 200 mg/day of bepridil treatment for 12 weeks. After the treatment, those patients who converted to sinus rhythm was 3.4% in placebo, 37.5% in those who received 100 mg/day and 69.0% in those who received 200 mg/day, thus demonstrating a linear dose-response relationship for AF conversion. The conversion rate gradually reached a maximal value at approximately 6 weeks after initiation of bepridil. However, the AF recurrence rate was high (91.7% in those receiving 100 mg/day and 75.0% in those receiving 200 mg/day). Adverse events, presumably related to the drug, were also frequent: ventricular tachycardia in 2, QT prolongation in 4 and sinus bradycardia in 2 patients. In those patients treated with 200 mg/day group, 1 patient died suddenly because of ventricular tachycardia.This study demonstrated the dose response-relationships of bepridil for AF conversion to sinus rhythm. However, the high rate of AF recurrence and substantial drug-related adverse effects, including sudden death, raised caution about using bepridil to treat persistent AF. The balance between benefits and risks of the drug should be individualized.

Background Most patients with Takotsubo cardiomyopathy show a favorable outcome. Although several complications have been reported, the frequency of thromboembolism has not been clarified. Methods Clinical characteristics and complications of 21 consecutive patients (18 female, aged 72 years) with Takotsubo cardiomyopathy during the past 9 years were investigated. Results The most major complication was heart failure (52%). Thromboembolism was found in 3 patients (14%) and this was the second most frequent cardiovascular complication. One of the 3 patients showed left ventricular thrombus and the other 2 experienced cardioembolic stroke. All 3 patients visited the emergency department more than 48 h after initial chest pain occurred. Conclusions This study indicates that thromboembolism is a common complication in the acute phase of Takotsubo cardiomyopathy, and anticoagulation therapy should be performed in all patients until wall motion abnormalities improve. Takotsubo cardiomyopathy should be considered one of the important causes of cardioembolic stroke.

Daunorubicin (DNR) is one of the anthracycline anti-tumor agents widely used in the treatment of acute myeloid leukemia. However, the clinical use of DNR has been limited by its undesirable systemic toxicity, especially in the heart and kidney. This study was designed to test the effectiveness of carvedilol, a nonselective beta-blocker against DNR-induced cardiotoxicity and nephrotoxicity. Rats were treated with a cumulative dose of 9 mg/kg body weight DNR (i.v.). Carvedilol was administered orally every day for 6 weeks. DNR rats showed cardiac and nephrotoxicities as evidenced by worsening cardiac and kidney functions, which were evaluated by hemodynamic and echocardiographic studies, and by measuring protein in urine, levels of urea and creatinine in serum, lipid profiles, malondialdeyde level and the total level of glutathione peroxidase activity in both heart and kidney tissues. These changes were reversed by treatment with carvedilol, which resulted in significant improvement in the cardio-renal function. Furthermore, carvedilol down-regulated matrix metalloproteinase-2 expression in the heart, increased nephrin expression in the kidney, and attenuated the increased protein expression of NADPH oxidase subunits in heart and kidney. Moreover, carvedilol reduced myocardial and renal apoptosis and improved the histopathological changes in heart and kidney induced by DNR. In conclusion, the present study demonstrated a beneficial effect of carvedilol treatment in the prevention of DNR-induced cardiotoxicity and nephrotoxicity by reversing the oxidative stress and apoptosis.

The aim of this study was to investigate the relationship between J-wave dynamics and arrhythmias during myocardial ischaemia in patients with vasospastic angina (VSA).Sixty-seven consecutive patients diagnosed with VSA by a provocation test for coronary spasm were grouped according to whether they had a J wave in the baseline electrocardiograms or not (VSA-JW group, n = 14; VSA-non-JW group: n = 53). We retrospectively studied the associations between J-wave and ST-segment dynamics and induced ventricular fibrillations (VFs) during coronary spasm. In the VSA-JW group, 7 of the 14 patients showed changes in J-wave morphology and/or gains in J-wave voltage, followed by VF in 4 patients. Compared with patients without VF, the four patients with VF showed similar maximal voltage in the baseline J waves but a higher voltage during induced coronary spasms (0.57 ± 0.49 vs. 0.30 ± 0.11 mV; P = 0.011). In three patients with VF, J waves progressively increased and were accompanied by the characteristic coved-type or lambda-shaped ST-segment elevations. In the VSA-non-JW group, only four patients showed new appearances of J waves during coronary spasms and another patient without a distinct J wave developed VF. Ventricular fibrillations were induced more frequently in the VSA-JW group than in the VSA-non-JW group [4/14 (29%) vs. 1/53 (2%); P = 0.012].J-wave augmentations were caused by myocardial ischaemia during coronary spasms. The presence and augmentation of J waves, especially prominent J waves with the characteristic ST-elevation patterns, were associated with VF.

Hepatitis C virus (HCV) is a hepatotrophic virus and a major cause of chronic liver disease, including hepatocellular carcinoma, worldwide. The life cycle of HCV is closely associated with the metabolism of lipids and lipoproteins. The main function of lipoproteins is transporting lipids throughout the body. Triglycerides, free cholesterol, cholesteryl esters, and phospholipids are the major components of the transported lipids. The pathway of HCV assembly and secretion is closely linked to lipoprotein production and secretion, and the infectivity of HCV particles largely depends on the interaction of lipoproteins. Moreover, HCV entry into hepatocytes is strongly influenced by lipoproteins. The key lipoprotein molecules mediating these interactions are apolipoproteins. Apolipoproteins are amphipathic proteins on the surface of a lipoprotein particle, which help stabilize lipoprotein structure. They perform a key role in lipoprotein metabolism by serving as receptor ligands, enzyme co-factors, and lipid transport carriers. Understanding the association between the life cycle of HCV and lipoprotein metabolism is important because each step of the life cycle of HCV that is associated with lipoprotein metabolism is a potential target for anti-HCV therapy. In this article, we first concisely review the nature of lipoprotein and its metabolism to better understand the complicated interaction of HCV with lipoprotein. Then, we review the outline of the processes of HCV assembly, secretion, and entry into hepatocytes, focusing on the association with lipoproteins. Finally, we discuss the clinical aspects of disturbed lipid/lipoprotein metabolism and the significance of dyslipoproteinemia in chronic HCV infection with regard to abnormal apolipoproteins.

To clarify the influence of hypertension and blood pressure (BP) control on thromboembolism and major hemorrhage in patients with nonvalvular atrial fibrillation, a post hoc analysis of the J-RHYTHM Registry was performed.A consecutive series of outpatients with atrial fibrillation was enrolled from 158 institutions. Of 7937 patients, 7406 with nonvalvular atrial fibrillation (70.8% men, 69.8±10.0 years) were followed for 2 years or until an event occurred. Hypertension was defined as a systolic BP ≥140 mm Hg, a diastolic BP ≥90 mm Hg, a history of hypertension, and/or antihypertensive drug use. Hypertension was an independent risk factor for major hemorrhage (hazard ratio 1.52, 95% CI 1.05-2.21, P=0.027) but not for thromboembolism (hazard ratio 1.05, 95% CI 0.73-1.52, P=0.787). When patients were divided into quartiles according to their systolic BP at the time closest to the event or at the end of follow-up (Q1, <114; Q2, 114-125; Q3, 126-135; and Q4, ≥136 mm Hg), odds ratios for both events were significantly higher in Q4 than in Q1 (thromboembolism, odds ratio 2.88, 95% CI 1.75-4.74, P<0.001; major hemorrhage, odds ratio 1.61, 95% CI 1.02-2.53, P=0.041) after adjustment for components of CHA2DS2-VASc score, warfarin use, and antiplatelet use. A systolic BP of ≥136 mm Hg was an independent risk factor for thromboembolism and major hemorrhage.BP control appears to be more important than a history of hypertension and baseline BP values at preventing thromboembolism and major hemorrhage in patients with nonvalvular atrial fibrillation.URL: http://www.umin.ac.jp/ctr. Unique identifier: UMIN000001569.

Various computer experiments based on the numerical integration of dynamical equation of motion are their importance is emphasized in connection with ergodic theory. To do this a short review of the present status of the ergodic theories is also given and the interrelations among various results of ergodic theories are clarified. The inducition period discovered in the previous papers (H. Hirooka and N. Saitô, J. Phys. Soc. Japan 26 (1969), 624; N. Ooyama, H. Hirooka and N. Saitô, J. Phys. Soc. Japan 27 (1969), 815) is further examined in one-dimensional anharmonic lattices including exponential lattices. The apparent randomness which the system exhibits after elapsing the induction period is, strictly speaking, quasi-stochastic in the sence that the process is described completely as deterministic. The “true” stochasticity which seems required in the thermodynamical state will be realized in the thermodynamical limit.

Background Causative arrhythmias of sudden cardiac arrest (SCA) are changing in this age of improved coronary care. Objective The purpose of this study was to examine the frequency of terminal arrhythmias and the electrical events prior to SCA. Methods We analyzed 24-hour Holter recordings of 132 patients enrolled from 41 institutions who either died (n = 88) or had an aborted death (n = 44). The Holter recordings were obtained for diagnosing and evaluating diseases and arrhythmias in those without any episodes suggestive of SCA. Results In 97 patients (73%), SCA was associated with ventricular tachyarrhythmias and in 35 (27%) with bradyarrhythmias. The bradyarrhythmia-related SCA patients were older than those with a tachyarrhythmia-related SCA (70 ± 13 years vs 58 ± 19 years, P < .001). The most common arrhythmia for a tachyarrhythmia-related SCA was ventricular tachycardia degenerating to ventricular fibrillation (45%). The bradyarrhythmia-related SCA was caused by asystole (74%) or AV block (26%). Spontaneous conversion was observed in 37 patients (38%) with ventricular tachyarrhythmias. Of those, 62% of the patients experienced symptoms including syncope, chest pain, or convulsion. Multivariate logistic analysis revealed that independent predictors of mortality for tachyarrhythmia-related SCAs were advanced age (odds ratio 1.04, 95% confidence interval 1.02–1.08) and ST elevation within the hour before SCA (odds ratio 3.54, 95% confidence interval 1.07–13.5). In contrast, the presence of preceding torsades de pointes was associated with spontaneous conversion (odds ratio 0.20, 95% confidence interval 0.05–0.66). Conclusion The most frequent cause of SCA remains ventricular tachyarrhythmias. Advanced age and ST elevation before SCA are risk factors for mortality in tachyarrhythmia-related SCAs. Causative arrhythmias of sudden cardiac arrest (SCA) are changing in this age of improved coronary care. The purpose of this study was to examine the frequency of terminal arrhythmias and the electrical events prior to SCA. We analyzed 24-hour Holter recordings of 132 patients enrolled from 41 institutions who either died (n = 88) or had an aborted death (n = 44). The Holter recordings were obtained for diagnosing and evaluating diseases and arrhythmias in those without any episodes suggestive of SCA. In 97 patients (73%), SCA was associated with ventricular tachyarrhythmias and in 35 (27%) with bradyarrhythmias. The bradyarrhythmia-related SCA patients were older than those with a tachyarrhythmia-related SCA (70 ± 13 years vs 58 ± 19 years, P < .001). The most common arrhythmia for a tachyarrhythmia-related SCA was ventricular tachycardia degenerating to ventricular fibrillation (45%). The bradyarrhythmia-related SCA was caused by asystole (74%) or AV block (26%). Spontaneous conversion was observed in 37 patients (38%) with ventricular tachyarrhythmias. Of those, 62% of the patients experienced symptoms including syncope, chest pain, or convulsion. Multivariate logistic analysis revealed that independent predictors of mortality for tachyarrhythmia-related SCAs were advanced age (odds ratio 1.04, 95% confidence interval 1.02–1.08) and ST elevation within the hour before SCA (odds ratio 3.54, 95% confidence interval 1.07–13.5). In contrast, the presence of preceding torsades de pointes was associated with spontaneous conversion (odds ratio 0.20, 95% confidence interval 0.05–0.66). The most frequent cause of SCA remains ventricular tachyarrhythmias. Advanced age and ST elevation before SCA are risk factors for mortality in tachyarrhythmia-related SCAs.

We investigated various serum inflammatory markers to predict ablation responders who have no atrial fibrillation (AF) relapse after the initial ablation.Forty-four consecutive AF patients (age: 59 ± 8 years, paroxysmal: 31, CHADS₂: 1.1 ± 1.1) who underwent an initial pulmonary vein isolation were investigated. Various serum inflammatory markers, such as adiponectin, ANP, BNP, 1CTP, F1+2, hs-CRP, IL-6, intact P1NP, MDA-LDL, MMP-2, TGF-β, TIMP-2, and TNF-α, were evaluated prior to ablation. AF relapse was defined as AF documented in telemonitoring electrocardiograms twice a day during 9.7 ± 2.4 months of follow-up with three months of a blanking-period.A total of 29 patients (paroxysmal: 21) maintained sinus rhythm after the initial catheter ablation. These ablation responders had significantly lower MMP-2 (Sinus vs. Relapsed: 748 ± 132.7 vs. 841.2 ± 152.4 ng/mL, P=0.042) and TNF-α (1.1 ± 0.4 vs. 1.8 ± 1.7 pg/mL, P=0.046) levels prior to ablation. A BNP-adjusted Cox multivariate regression analysis revealed that the independent predictive factor for AF recurrence was high MMP-2 levels (>766 ng/mL) accompanied by high TNF-α levels (>1.2 pg/mL).The levels of MMP-2 and TNF-α might be useful for predicting initial AF catheter ablation responders.

It has never been possible to immediately evaluate heart rate variability (HRV) during exercise. We aimed to visualize the real-time changes in the power spectrum of HRV during exercise and to investigate its relationship to the ventilatory threshold (VT).Thirty healthy subjects (29.1±5.7 years of age) and 35 consecutive patients (59.0±13.2 years of age) with myocardial infarctions underwent cardiopulmonary exercise tests with an RAMP protocol ergometer. The HRV was continuously assessed with power spectral analyses using the maximum entropy method and projected on a screen without delay. During exercise, a significant decrease in the high frequency (HF) was followed by a drastic shift in the power spectrum of the HRV with a periodic augmentation in the low frequency/HF (L/H) and steady low HF. When the HRV threshold (HRVT) was defined as conversion from a predominant high frequency (HF) to a predominant low frequency/HF (L/H), the VO2 at the HRVT (HRVT-VO2) was substantially correlated with the VO2 at the lactate threshold and VT) in the healthy subjects (r=0.853 and 0.921, respectively). The mean difference between each threshold (0.65 mL/kg per minute for lactate threshold and HRVT, 0.53 mL/kg per minute for VT and HRVT) was nonsignificant (P>0.05). Furthermore, the HRVT-VO2 was also correlated with the VT-VO2 in these myocardial infarction patients (r=0.867), and the mean difference was -0.72 mL/kg per minute and was nonsignificant (P>0.05).A HRV analysis with our method enabled real-time visualization of the changes in the power spectrum during exercise. This can provide additional information for detecting the VT.

SCN5A is abundant in heart and has a major role in INa. Loss-of-function mutation in SCN5A results in Brugada syndrome (BrS), which causes sudden death in adults. It remains unclear why disease phenotype does not manifest in the young even though mutated SCN5A is expressed in the young. The aim of the present study is to elucidate the timing of the disease manifestation in BrS. A gain-of-function mutation in SCN5A also results in Long QT syndrome type 3 (LQTS3), leading to sudden death in the young. Induced pluripotent stem cells (iPSCs) were generated from a patient with a mixed phenotype of LQTS3 and BrS with the E1784K SCN5A mutation. Here we show that electrophysiological analysis revealed that LQTS3/BrS iPSC-derived cardiomyocytes recapitulate the phenotype of LQTS3 but not BrS. Each β-subunit of the sodium channel is differentially expressed in embryonic and adult hearts. SCN3B is highly expressed in embryonic hearts and iPSC-derived cardiomyocytes. A heterologous expression system revealed that INa of mutated SCN5A is decreased and SCN3B augmented INa of mutated SCN5A. Knockdown of SCN3B in LQTS3/BrS iPSC-derived cardiomyocytes successfully unmasked the phenotype of BrS. Isogenic control of LQTS3/BrS (corrected-LQTS3/BrS) iPSC-derived cardiomyocytes gained the normal electrophysiological properties.

To investigate the influence of interferon-free antivirus therapy on lipid profiles in chronic hepatitis C virus genotype 1b (HCV1b) infection.Interferon-free antiviral agents were used to treat 276 patients with chronic HCV1b infection, and changes in serum lipids of those who achieved sustained virologic response (SVR) were examined. The treatment regimen included 24 wk of daclatasvir plus asunaprevir (DCV + ASV) or 12 wk of sofosbuvir plus ledipasvir (SOF + LDV). SVR was achieved in 121 (85.8%) of 141 patients treated with DCV + ASV and 132 (97.8%) of 135 patients treated with SOF + LDV. In the two patient groups (DCV + ASV-SVR and SOF + LDV-SVR), serum total cholesterol (TC), low-density lipoprotein cholesterol (LDL-C), high-density lipoprotein cholesterol (HDL-C), and triglycerides were measured at baseline during treatment and at 4 and 12 wk after treatment. Then, longitudinal changes in lipid profiles were analyzed.Serum levels of TC, LDL-C, and HDL-C were significantly increased throughout the observation period in both the DCV + ASV-SVR and SOF + LDV-SVR groups. During antivirus treatment, the increases in TC and LDL-C were significantly greater in the SOF + LDV-SVR group than in the DCV + ASV-SVR group (P < 0.001). At 4 and 12 wk after the therapy, serum levels of TC and LDL-C were similar between the two groups and were significantly greater than those at baseline. Approximately 75%-80% of the increase in TC was derived from an increased LDL-C. In multiple regression analysis, the difference in therapy protocol (DCA + ASV or SOF + LDV) was an independent predictor that was significantly associated with the increase in TC and LDL-C at 4 wk of therapy.Serum cholesterol significantly increased during SOF + LDV treatment. After treatment, HCV elimination was associated with a similar increase in cholesterol regardless of the therapy protocol.

Autoimmune hepatitis (AIH) is a chronic progressive liver disease characterized by high levels of aminotransferases and autoantibodies, hypergammaglobulinemia, and interface hepatitis. AIH affects all races and all ages worldwide, regardless of sex, although a preponderance of females is a constant finding. The etiology of AIH has not been completely elucidated, but immunogenetic background and environmental parameters may contribute to its development. The most important genetic factor is human leukocyte antigens (HLAs), especially HLA-DR, whereas the role of environmental factors is not completely understood. Immunologically, disruption of the immune tolerance to autologous liver antigens may be a trigger of AIH. The diagnosis of classical AIH is fairly easy, though not without pitfalls. In contrast, the diagnosis of atypical AIH poses great challenges. There is confusion as to the definition of the disease entity and its boundaries in the diagnosis of overlap syndrome, drug-induced autoimmune hepatitis, and AIH with concomitant nonalcoholic fatty liver disease (NAFLD) or chronic hepatitis C. Centrilobular zonal necrosis is now included in the histological spectrum of AIH. However, the definition and the significance of AIH presenting with centrilobular zonal necrosis have not been examined extensively. In ~20% of AIH patients who are treated for the first time with standard therapy, remission is not achieved. The development of more effective and better tolerated novel therapies is an urgent need. In this review, we discuss the current challenges and the future prospects in relation to the diagnosis and treatment of AIH, which have been attracting considerable recent attention.

BackgroundElectrical storm (E-Storm), defined as multiple episodes of ventricular arrhythmias within a short period of time, is an important clinical problem in patients with an implantable cardiac defibrillator (ICD) including cardiac resynchronization therapy devices capable of defibrillation. The detailed clinical aspects of E-Storm in large populations especially for non-ischemic dilated cardiomyopathy (DCM), however, remain unclear.ObjectiveThis study was performed to elucidate the detailed clinical aspects of E-Storm, such as its predictors and prevalence among patients with structural heart disease including DCM.MethodsWe analyzed the data of the Nippon Storm Study, which was a prospective observational study involving 1570 patients enrolled from 48 ICD centers. For the purpose of this study, we evaluated 1274 patients with structural heart disease, including 482 (38%) patients with ischemic heart disease (IHD) and 342 (27%) patients with DCM.ResultsDuring a median follow-up of 28 months (interquartile range: 23 to 33 months), E-Storm occurred in 84 (6.6%) patients. The incidence of E-Storm was not significantly different between patients with IHD and patients with DCM (log-rank p = 0.52). Proportional hazard regression analyses showed that ICD implantation for secondary prevention of sudden cardiac death (p = 0.0001) and QRS width (p = 0.015) were the independent risk factors for E-storm. In a comparison between patients with and without E-Storm, survival curves after adjustment for clinical characteristics showed a significant difference in mortality.ConclusionE-Storm was associated with subsequent mortality in patients with structural heart disease including DCM.

Background Inappropriate doses of direct oral anticoagulants (DOACs) are often prescribed. This study evaluated the prevalence, outcomes, and predictors of the prescription of inappropriately low doses of 4 types of DOACs in patients with atrial fibrillation (AF). Methods We retrospectively analyzed prospectively collected data from a single-center registry with 2272 patients prescribed DOACs for AF (apixaban: 1014; edoxaban: 267; rivaroxaban: 498; dabigatran: 493). Patients were monitored for 2 years and classified into appropriate-dose (n = 1,753; including appropriate low doses), inappropriate-low-dose (n = 490) and inappropriate-high-dose groups (n = 29). Major bleeding (MB) and thromboembolic events (TEEs) were evaluated. Results The mean age was 72 ± 10 years. The CHADS2 and HAS-BLED scores were 1.95 ± 1.32 and 1.89 ± .96, respectively. Overall, the incidences of MB and TEE were 2.3 and 2.1 per 100-patinet year, respectively. The inappropriate-low-dose group had younger age, heavier body weight, and higher creatinine clearance value than the appropriate-dose group. Multiple logistic regression analyses demonstrated the following independent determinants of the prescription of an inappropriately low dose: apixaban: HAS-BLED score; edoxaban: age; rivaroxaban: age, creatinine clearance value, HAS-BLED score, CHADS2 score, and antiplatelet therapy; dabigatran: age. There were not significant differences in the incidence of major bleeding and stroke/systemic emboli among the inappropriate-low-dose group of 4 DOACs compared with the appropriate-dose group of 4 DOACs. Conclusions In a single-center registry, 23% of patients with AF treated with a DOAC received an inappropriate dose. Several clinical factors, such as age and the creatinine clearance value, can identify patients at risk of under-treatment with DOACs.

The clinical characteristics and outcomes of women and men with atrial fibrillation (AF) are reported to be different. However, whether sex-related differences extend to patients' symptom burden and perceived quality of life (QOL) or the management pattern of AF has been rarely studied, particularly in Asian countries.To assess the differences in symptoms, treatment, and QOL between Japanese female and male patients with AF.Retrospective cohort study using data from the multicenter outpatient registry Keio Interhospital Cardiovascular Studies-Atrial Fibrillation (KiCS-AF), which collects information regarding health status and the treatment of patients with newly diagnosed or referred AF. One-year follow-up data were available for 1534 patients at 11 referral centers in the Tokyo, Japan, area who were enrolled between September 2012 and December 2015. All data available up to the 1-year follow-up examination through July 31, 2017, were included.Sex, symptoms, AF treatment, and QOL as determined by Atrial Fibrillation Effect on Quality of Life (AFEQT) questionnaires at their initial visit and 1 year later.Of 1534 patients, 1076 (70.1%) were men. Compared with men, women were more likely to be older (median age, 73 years [interquartile range {IQR}, 65-78 years] vs 65 years [IQR, 57-73 years], P < .001) and have higher median brain-type natriuretic peptide levels (102.8 pg/mL [IQR, 47.3-235.5 pg/mL] vs 74.1 pg/mL [IQR, 28.5-150.5 pg/mL], P < .001). Women also had lower median AFEQT overall summary scores than men (75 [IQR, 61-85] vs 80 [IQR, 69-90]; P < .001) but similar treatment satisfaction at baseline. During follow-up, women were less likely to be treated with a rhythm control strategy (48.1% [n = 214] vs 58.0% [n = 621], P < .001), including catheter ablation of AF (adjusted hazard ratio, 0.77 [95% CI, 0.62-0.95]; P = .02). At 1-year follow-up, women and men had improved QOL scores, regardless of their baseline characteristics (eg, age or brain-type natriuretic peptide levels) or treatment strategies, yet the sex gap persisted and grew (adjusted change in AFEQT overall summary score during 12 months, 5.89 [95% CI, 2.24-9.54] in women vs 8.94 [95% CI, 5.59-12.30] in men; P = .02).In contemporary Japanese clinical practice among unselected patients with AF, women were initially seen with greater QOL impairment, and the sex gap grew 1 year after presentation. The present study underscores the need for focused efforts to better understand and close this observed sex gap over the initial year of treatment for patients with AF.

Epicardial adipose tissue (EAT) may be associated with arrhythmogenesis. P-wave indices such as P-wave dispersion and P-wave variation indicated a slowed conduction velocity within the atria. This study investigated the effect of dapagliflozin on EAT volume and P-wave indices.In the present ad hoc analysis, 35 patients with type 2 diabetes mellitus and coronary artery disease were classified into dapagliflozin group (n=18) and conventional treatment group (n=17). At baseline, EAT volume, HbA1c and plasma level of tumor necrotic factor-α (TNF-α) levels, echocardiography, and 12-lead electrocardiogram (ECG) were performed. EAT volume was measured using computed tomography. Using 12-lead ECG, P-wave indices were measured.At baseline, EAT volumes in the dapagliflozin and conventional treatment groups were 113±20 and 110±27 cm3, respectively. Not only HbA1c and plasma level of TNF-α but also echocardiography findings including left atrial dimension and P-wave indices were comparable between the two groups. After 6 months, plasma level of TNF-α as well as EAT volume significantly decreased in the dapagliflozin group only. P-wave dispersion and P-wave variation significantly decreased in the dapagliflozin group only (-9.2±8.7 vs. 5.9±19.9 ms, p=0.01; -3.5±3.5 vs. 1.7±5.9 ms, p=0.01). The change in P-wave dispersion correlated with changes in EAT volume and plasma level of TNF-α. In multivariate analysis, the change in EAT volume was an independent determinant of the change in P-wave dispersion.Dapagliflozin reduced plasma level of TNF-α, EAT volume, and P-wave indices, such as P-wave dispersion. The changes in P-wave indices were especially associated with changes in EAT volume.The number and date of registration: UMIN000035660, 24/Jan/2019.

Two major forms of inherited J-wave syndrome (JWS) are recognized: early repolarization syndrome (ERS) and Brugada syndrome (BrS).This study sought to assess the distinct features between patients with ERS and BrS carrying pathogenic variants in SCN5A.Clinical evaluation and next-generation sequencing were performed in 262 probands with BrS and 104 with ERS. Nav1.5 and Kv4.3 channels were studied with the use of patch-clamp techniques. A computational model was used to investigate the protein structure.The SCN5A+ yield in ERS was significantly lower than in BrS (9.62% vs 22.90%; P = 0.004). Patients diagnosed with ERS displayed shorter QRS and QTc than patients with BrS. More than 2 pathogenic SCN5A variants were found in 5 probands. These patients displayed longer PR intervals and QRS duration and experienced more major arrhythmia events (MAE) compared with those carrying only a single pathogenic variant. SCN5A-L1412F, detected in a fever-induced ERS patient, led to total loss of function, destabilized the Nav1.5 structure, and showed a dominant-negative effect, which was accentuated during a febrile state. ERS-related SCN5A-G452C did not alter the inward sodium current (INa) when SCN5A was expressed alone, but when coexpressed with KCND3 it reduced peak INa by 44.52% and increased the transient outward potassium current (Ito) by 106.81%.These findings point to SCN5A as a major susceptibility gene in ERS as much as it is in BrS, whereas the lower SCN5A+ ratio in ERS indicates the difference in underlying electrophysiology. These findings also identify the first case of fever-induced ERS and demonstrate a critical role of Ito in JWS and a higher risk for MAE in JWS probands carrying multiple pathogenic variants in SCN5A.

Abstract Introduction Most patients with Brugada syndrome (BrS) are first diagnosed in their 40s, with sudden cardiac death (SCD) often occurring in their 50s. Ventricular fibrillation (VF) may occur in some patients with BrS despite having been asymptomatic for a long period. This study aimed to assess the incidence and risk factors for late life‐threatening arrhythmias in patients with BrS. Methods Patients with BrS ( n = 523; mean age, 51 ± 13 years; male, n = 497) were enrolled. The risk of late life‐threatening arrhythmia was investigated in 225 patients who had experienced no cardiac events (CEs: SCD or ventricular tachyarrhythmia) for at least 10 years after study enrollment. The incidence of CEs during the follow‐up period was examined. Results During the follow‐up of the 523 patients, 59 (11%) experienced CEs. The annual incidences of CEs were 2.87%, 0.77%, and 0.09% from study enrollment to 3, 3−10, and after 10 years, respectively. Among 225 patients who had experienced no CEs for at least 10 years after enrollment, four patients (1.8%) subsequently experienced CEs. Kaplan–Meier analysis revealed significant differences in the incidence of late CEs between patients with and without a history of symptoms ( p = .032). The positive and negative predictive values of late CEs for the programmed electrical stimulation (PES) test were 2.9% and 100%, respectively. Conclusion Our results suggest that patients with BrS who are asymptomatic and have no ventricular tachycardia/VF inducibility by PES are at extremely low risk of experiencing late life‐threatening arrhythmias.

Abstract Background and Aims Brugada syndrome (BrS) is an inherited arrhythmia with a higher disease prevalence and more lethal arrhythmic events in Asians than in Europeans. Genome-wide association studies (GWAS) have revealed its polygenic architecture mainly in European populations. The aim of this study was to identify novel BrS-associated loci and to compare allelic effects across ancestries. Methods A GWAS was conducted in Japanese participants, involving 940 cases and 1634 controls, followed by a cross-ancestry meta-analysis of Japanese and European GWAS (total of 3760 cases and 11 635 controls). The novel loci were characterized by fine-mapping, gene expression, and splicing quantitative trait associations in the human heart. Results The Japanese-specific GWAS identified one novel locus near ZSCAN20 (P = 1.0 × 10−8), and the cross-ancestry meta-analysis identified 17 association signals, including six novel loci. The effect directions of the 17 lead variants were consistent (94.1%; P for sign test = 2.7 × 10−4), and their allelic effects were highly correlated across ancestries (Pearson’s R = .91; P = 2.9 × 10−7). The genetic risk score derived from the BrS GWAS of European ancestry was significantly associated with the risk of BrS in the Japanese population [odds ratio 2.12 (95% confidence interval 1.94–2.31); P = 1.2 × 10−61], suggesting a shared genetic architecture across ancestries. Functional characterization revealed that a lead variant in CAMK2D promotes alternative splicing, resulting in an isoform switch of calmodulin kinase II-δ, favouring a pro-inflammatory/pro-death pathway. Conclusions This study demonstrates novel susceptibility loci implicating potentially novel pathogenesis underlying BrS. Despite differences in clinical expressivity and epidemiology, the polygenic architecture of BrS was substantially shared across ancestries.

Dendritic cells (DC), the most potent 'professional' antigen-presenting cells, hold promise for improving the immunotherapy of cancer. In this study, we investigated the ability of normal donor DC pulsed ex vivo with 12 mer bcr-abl (b3a2) peptide to generate b3a2-specific autologous or HLA-identical sibling donor's cytotoxic T-lymphocytes (CTL). DC that were grown from normal peripheral blood adherent cells or purified DC precursors in the presence of GM-CSF and IL-4, were pulsed with b3a2-peptide then were induced to become mature and functional cells by the addition of TNF-α. These peptide-pulsed mature DC elicited a potent b3a2-specific CTL response in vitro. The b3a2-peptide pulsed DC-primed peripheral blood lymphocytes (PBL) displayed significantly higher cytotoxic activity compared with peptide non-pulsed DC-primed PBL against target cells, which are b3a2 positive marrow cells derived from HLA-identical sibling chronic myelogenous leukemia (CML) patient, or peptide-pulsed autologous macrophages (P < 0.001). in addition, the b3a2 peptide-pulsed dc-primed and non-pulsed dc-primed pbl showed no cytotoxic response against peptide non-pulsed autologous macrophages. these findings revealed that normal donor pbl pre-immunized with b3a2-peptide pulsed autologous dc could increase the graft-versus-leukemia effect without exaggerating graft-versus-host-disease. both cd8+ and CD4+ T lymphocytes were shown to be involved in the effector cell populations. The b3a2 peptide-pulsed DC-primed T cells were significantly superior in their production of GM-CSF and TNF-α compared with peptide non-pulsed DC-primed T cells. These intriguing preclinical results imply the feasibility of developing b3a2 peptide-DC based protocol for in vitro sensitization of normal donor leukocytes before donor leukocyte transfusions for patients with CML, who relapsed after HLA-matched sibling bone marrow transplantation.

The cardioprotective properties of carvedilol (a vasodilating beta-adrenoceptor blocking agent) were studied in a rat model of dilated cardiomyopathy induced by autoimmune myocarditis. Twenty-eight days after immunization, surviving Lewis rats (32/43=74%) were divided into three groups to be given 2 mg kg(-1) day(-1) (Group-C2, n=10) or 20 mg kg(-1) day(-1) (Group-C20, n=10) of carvedilol, or vehicle (0.5% methylcellulose, Group-V, n=12). After oral administration for 2 months, body weight, heart weight (HW), heart rate (HR), rat alpha-atrial natriuretic peptide (r-ANP) in blood, central venous pressure (CVP), mean blood pressure (mean BP), peak left ventricular pressure (LVP), left ventricular end-diastolic pressure (LVEDP), +/-dP dt(-1) and area of myocardial fibrosis were measured. Values were compared with those for normal Lewis rats (Group-N, n=10). Two out of 12 (17%) rats in Group-V died from day 28 to day 42 after immunization. No rat died in Groups-C2, -C20 and -N. Although the CVP, mean BP, LVP and +/-dP dt(-1) did not differ among the three groups, the HW, HR and r-ANP in Group-C2 (1.14+/-0.03, 339+/-16 and 135+/-31) and Group-C20 (1.23+/-0.04, 305+/-8 and 156+/-24) were significantly lower than those in Group-V (1.36+/-0.04 g, 389+/-9 beats min(-1) and 375+/-31 pg ml(-1), respectively). The LVEDP in Group-C2 was significantly lower than that in Group-V (7.4+/-1.4 and 12.2+/-1.2 mmHg, respectively, P<0. 05). The area of myocardial fibrosis in Group-C2 was smaller than that in Group-V (12+/-1 and 31+/-2%, P<0.01). These results indicate that a low dose of carvedilol has beneficial effects on dilated cardiomyopathy.

Although immunosuppressive therapy for myocarditis has attracted a great deal of attention, its effectiveness is controversial. Interleukin (IL)-10 has a variety of immunomodulatory properties. Among the nonviral techniques for gene transfer in vivo, the direct injection of plasmid DNA into muscle is simple, inexpensive, and safe.We examined the applicability of murine IL-10 (mIL-10) gene transfer to the treatment of rats with experimental autoimmune myocarditis. Nine-week-old Lewis rats were inoculated with pig myosin (day 0). A plasmid vector expressing mIL-10 cDNA (800 microgram per rat) was transferred into the tibialis anterior muscles by electroporation 3 times (5 days before immunization and at days 4 and 13); control rats received empty plasmid. Electroporation increased the serum mIL-10 levels to >250 pg/mL. The 21-day survival rate in rats treated with mIL-10 cDNA was higher (15 of 15; 100%) than that of the control group (9 of 15; 60%). Furthermore, mIL-10 treatment significantly attenuated myocardial lesions and improved hemodynamic parameters.These findings showed that gene transfer into muscle by electroporation in vivo is an effective means of delivery of IL-10 for the treatment of autoimmune myocarditis.

Some cytokines are believed to play a role in the development of acute and chronic GVHD after allo-hematopoietic stem cell transplantation. It has been reported that TNF-alpha and IL-10 gene polymorphisms are associated with the production of those cytokines and the development of graft failure after organ transplantation and systemic lupus erythematosus. We examined whether TNF-alpha and IL-10 gene polymorphisms affect the severity of acute GVHD (aGVHD) and chronic GVHD (cGVHD). Sixty-two and 54 patients were available for the analysis of aGVHD and cGVHD, respectively. We analyzed the gene polymorphisms derived from pre- and post-transplant blood cells. Donor-derived TNF2 allele (A) was more frequently detected in patients with aGVHD III/IV than those aGVHD 0-II (2/6 vs 2/56) (P = 0.04). The donors of the patients with cGVHD more frequently possessed a greater number of alleles (allele 13 or more which contain 26 or more CA repeats) in IL-10.G than those without (13/26 vs 5/28) (P = 0.02), and the patients with cGVHD had more CA repeats in donor-derived IL-10.G than those without (mean = 25.2 vs 23.4) (P = 0.01). Donor-derived TNF-308 and IL-10.G alleles may contribute to severe aGVHD and cGVHD, respectively, and will help us distinguish those patients at high risk for GVHD.

C-reactive protein (CRP) independently predicts cardiovascular disease (CVD) and is considered to be part of metabolic syndrome (MS). The concentration of CRP are proposed to be <1.0 mg/L as low risk, 1.0 to 3.0 mg/L as intermediate risk, and >3.0 mg/L as high risk for CVD in Western society.Apparently healthy 179 men and 166 women were categorized with modified National Cholesterol Education Program Adult Treatment Panel III criteria (body mass index > or = 25 in place of abdominal obesity) for defining MS. The cut-off points of CRP were evaluated for both MS defined by impaired fasting glucose criteria of > or = 110 mg/dl (MS110) and > or = 100 mg/dl (MS100), separately by sex. The optimal cut-off point of CRP was 0.65 mg/L in all subgroups. The sensitivity and specificity of this CRP value for male MS100, female MS100, male MS110, and female MS110 were 0.650 and 0.626, 1.000 and 0.771, 0.739 and 0.609, and 1.000 and 0.756, respectively.The optimal cut-off point of CRP for MS might be 0.65 mg/L in Japan and this value can be useful in routine clinical practice and studies on MS.

This study investigated the treatment of ventricular tachycardia (VT) after repair of tetralogy of Fallot or double outlet of the right ventricle. The ideal antiarrhythmic therapy for VT in patients after repair of congenital heart disease, especially without left ventricular dysfunction, has not yet been established. Seven consecutive patients (2 women and 5 men) with stable monomorphic sustained VT were investigated. The mean age was 25 ± 7 years (range, 16-35 years). Four patients had undergone surgical repair of tetralogy of Fallot, and 3 had surgical correction of double outlet of the right ventricle at the mean age of 18 ± 7 years (range, 9-27 years) before documentation of the arrhythmia. The mean ejection fraction of the left ventricle was 60% ± 8% (range, 50-72). Fourteen sustained monomorphic VTs were induced in 7 patients using programed electrical stimulation. The mean cycle length of tachycardia was 346 ± 77 milliseconds (range, 260-480 seconds). The site of the surgical correction of the right ventricle was associated with the origin of VT in all patients. Radiofrequency catheter ablation was attempted in 8 VTs in 7 patients: 7 clinical and 1 nonclinical VTs. In 6 patients, class III antarrhythmic agents were added because VT remained inducible after ablation. During a follow-up of 61 ± 29 months (range, 15-110 months), there were no recurrences of VT. In patients with drug-refractory VT originating from the right ventricle late after congenital heart disease, and when their left ventricular function do not deteriorate, combined therapy for radiofrequency catheter ablation with class III antiarrhythmic agents might effective and should be considered as a therapeutic option.

Summary Several reports have demonstrated a possible association of periodontal infections with coronary heart disease (CHD) by elevated antibody titre to periodontopathic bacteria in CHD patients compared with non-diseased controls. Although each periodontopathic bacterium may vary in virulence for periodontitis and atherosclerosis, antibody response to multiple bacteria in CHD patients has not been understood fully. Therefore, serum levels of antibody to 12 periodontopathic bacteria together with other atherosclerotic risk markers were compared among 51 patients with CHD, 55 patients with moderate to severe chronic periodontitis and 37 healthy individuals. The antibody response was the most prevalent for Porphyromonas gingivalis, a major causative organism, in CHD as well as periodontitis patients. However, antibody positivity was different between CHD and periodontitis if the response was analysed for two different strains of P. gingivalis, namely FDC381 and Su63. While periodontitis patients were positive for both P. gingivalis FDC381 and Su63, a high frequency of antibody positivity for P. gingivalis Su63 but not for FDC381 was observed in CHD patients. The results indicate that the presence of particular periodontopathic bacteria with high virulence may affect atherogenesis. Identifying the virulence factors of P. gingivalis Su63 may gain insight into the new therapeutic modality for infection-induced deterioration of atherosclerosis.

Tachycardia-induced cardiomyopathy is characterized by ventricular systolic dysfunction and congestive heart failure resulting from persistent or highly frequent tachyarrhythmias with uncontrolled heart rate. While reversible and often considered benign, few studies have examined the outcome of the disorder. The clinical characteristics, treatment, and long-term outcomes of 12 consecutive patients with tachycardia-induced cardiomyopathy (9 men, age, 51.9 +/- 17.6 years) were studied. The mean period between the occurrence of tachyarrhythmias and the development of congestive heart failure was 26.0 +/- 34.3 days. The mean heart rate on admission was 156.3 +/- 28.7 beats/min. All patients had severe heart failure with a NYHA functional class of 2.3 +/- 0.5, left ventricular ejection fraction of 0.32 +/- 0.10, and brain natriuretic peptide level of 505.7 +/- 449.1 pg/mL. In all patients, cardiac dysfunction recovered after 53.5 +/- 61.3 days. During the follow-up of 53 +/- 24 months, 2 patients had a recurrence of heart failure with uncontrolled tachyarrhythmia and 1 patient died suddenly. In tachycardia-induced cardiomyopathy, recurrent heart failure with uncontrollable tachyarrhythmia and sudden death were observed after recovery from cardiac dysfunction. A substrate for heart failure and/or life-threatening arrhythmia might persist, and careful, long-term follow-up seems required.

Andersen-Tawil syndrome (ATS) is a rare inherited disorder characterized by periodic paralysis, mild dysmorphic features, and QT or QU prolongation with ventricular arrhythmias in electrocardiograms (ECGs). Mutations of KCNJ2, encoding the human inward rectifying potassium channel Kir 2.1, have been identified in patients with ATS. We aimed to clarify the genotype-phenotype correlations in ATS patients. We screened 23 clinically diagnosed ATS patients from 13 unrelated Japanese families. Ten different forms of KCNJ2 mutations were identified in the 23 ATS patients included in this study. Their ECGs showed normal QTc intervals and abnormal U waves with QUc prolongation and a variety of ventricular arrhythmias. Especially, bidirectional ventricular tachycardia (VT) was observed in 13 of 23 patients (57%). Periodic paralysis was seen in 13 of 23 carriers (57%), dysmorphic features in 17 (74%), and seizures during infancy in 4 (17%). Functional assays for the two novel KCNJ2 mutations (c. 200G>A (p. R67Q) and c. 436G>A (p. G146S)) displayed no functional inward rectifying currents in a heterologous expression system and showed strong dominant negative effects when co-expressed with wild-type KCNJ2 channels (91% and 84% reduction at -50 mV respectively compared to wild-type alone). Immunocytochemistry and confocal imaging revealed normal trafficking for mutant channels. In our study, all of the clinically diagnosed ATS patients had KCNJ2 mutations and showed a high penetrance with regard to the typical cardiac phenotypes: predominant U wave and ventricular arrhythmias, typically bidirectional VT.

Although both hepatocyte growth factor (HGF) and vascular endothelial growth factor (VEGF) are potent angiogenic growth factors in animal models of ischemia, their characteristics are not the same in animal experiments and clinical trials. To elucidate the discrepancy between HGF and VEGF, we compared the effects of HGF and VEGF on endothelial progenitor cells under angiotensin II stimulation, which is a well-known risk factor for atherosclerosis. Here, we demonstrated that HGF, but not VEGF, attenuated angiotensin II-induced senescence of endothelial progenitor cells through a reduction of oxidative stress by inhibition of the phosphatidylinositol-3,4,5-triphosphate/rac1 pathway. Potent induction of neovascularization of endothelial progenitor cells by HGF, but not VEGF, under angiotensin II was also confirmed by in vivo experiments using several models, including HGF transgenic mice.

The computer experiments are carried out on the phenomena of self-synchronization in a many-mode system described by the van der Pol type equation. The results are successfully explained in terms of the perturbation theory based on a mean field approximation proposed in a previous article. The extension of the theory to further complicated phenomena of mode-locking is briefly discussed.