Allan Gaw

Background Although statins reduce coronary and cerebrovascular morbidity and mortality in middle-aged individuals, their efficacy and safety in elderly people is not fully established. Our aim was to test the benefits of pravastatin treatment in an elderly cohort of men and women with, or at high risk of developing, cardiovascular disease and stroke. Methods We did a randomised controlled trial in which we assigned 5804 men (n=2804) and women (n=3000) aged 70–82 years with a history of, or risk factors for, vascular disease to pravastatin (40 mg per day; n=2891) or placebo (n=2913). Baseline cholesterol concentrations ranged from 4·0 mmol/L to 9·0 mmol/L. Follow-up was 3·2 years on average and our primary endpoint was a composite of coronary death, non-fatal myocardial infarction, and fatal or non-fatal stroke. Analysis was by intention-to-treat. Findings Pravastatin lowered LDL cholesterol concentrations by 34% and reduced the incidence of the primary endpoint to 408 events compared with 473 on placebo (hazard ratio 0·85, 95% Cl 0·74–0·97, p=0·014). Coronary heart disease death and non-fatal myocardial infarction risk was also reduced (0·81, 0·69–0·94, p=0·006). Stroke risk was unaffected (1–03, 0·81–1·31, p=0·8), but the hazard ratio for transient ischaemic attack was 0·75 (0·55–1·00, p=0·051). New cancer diagnoses were more frequent on pravastatin than on placebo (1·25, 1·04–1·51, p=0·020). However, incorporation of this finding in a meta-analysis of all pravastatin and all statin trials showed no overall increase in risk. Mortality from coronary disease fell by 24% (p=0–043) in the pravastatin group. Pravastatin had no significant effect on cognitive function or disability. Interpretation Pravastatin given for 3 years reduced the risk of coronary disease in elderly individuals. PROSPER therefore extends to elderly individuals the treatment strategy currently used in middle aged people. Published online Nov 19, 2002 http://image.thelancet.com/extras/02art8325web.pdf

The National Cholesterol Education Program (NCEP) recently proposed a simple definition for metabolic syndrome. Information on the prospective association of this definition for coronary heart disease (CHD) and type 2 diabetes is currently limited.We used a modified NCEP definition with body mass index in place of waist circumference. Baseline assessments in the West of Scotland Coronary Prevention Study were available for 6447 men to predict CHD risk and for 5974 men to predict incident diabetes over 4.9 years of follow-up. Mean LDL cholesterol was similar but C-reactive protein was higher (P<0.0001) in the 26% of men with the syndrome compared with those without. Metabolic syndrome increased the risk for a CHD event [univariate hazard ratio (HR)=1.76 (95% CI, 1.44 to 2.15)] and for diabetes [univariate HR=3.50 (95% CI 2.51 to 4.90)]. Metabolic syndrome continued to predict CHD events (HR=1.30, 95% CI, 1.00 to 1.67, P=0.045) in a multivariate model incorporating conventional risk factors. Men with 4 or 5 features of the syndrome had a 3.7-fold increase in risk for CHD and a 24.5-fold increase for diabetes compared with men with none (both P<0.0001). C-reactive protein enhanced prognostic information for both outcomes. With pravastatin, men with the syndrome had similar risk reduction for CHD as compared with those without (HR, 0.73 and 0.69; pravastatin versus placebo).A modified NCEP metabolic syndrome definition predicts CHD events, and, more strikingly, new-onset diabetes, and thus helps identify individuals who may receive particular benefit from lifestyle measures to prevent these diseases.

We examined the development of new diabetes mellitus in men aged 45 to 64 years during the West of Scotland Coronary Prevention Study.Our definition of diabetes mellitus was based on the American Diabetic Association threshold of a blood glucose level of >/=7.0 mmol/L. Subjects who self-reported diabetes at baseline or had a baseline glucose level of >/=7.0 mmol/L were excluded from the analyses. A total of 5974 of the 6595 randomized subjects were included in the analysis, and 139 subjects became diabetic during the study. The baseline predictors of the transition from normal glucose control to diabetes were studied. In the univariate model, body mass index, log triglyceride, log white blood cell count, systolic blood pressure, total and HDL cholesterol, glucose, and randomized treatment assignment to pravastatin were significant predictors. In a multivariate model, body mass index, log triglyceride, glucose, and pravastatin therapy were retained as predictors of diabetes in this cohort.We concluded that the assignment to pravastatin therapy resulted in a 30% reduction (P:=0.042) in the hazard of becoming diabetic. By lowering plasma triglyceride levels, pravastatin therapy may favorably influence the development of diabetes, but other explanations, such as the anti-inflammatory properties of this drug in combination with its endothelial effects, cannot be excluded with these analyses.

Leptin plays a role in fat metabolism and correlates with insulin resistance and other markers of the metabolic syndrome, independent of total adiposity. Therefore, we hypothesized that raised leptin levels may identify men at increased risk of a coronary event in the West of Scotland Coronary Prevention Study (WOSCOPS). Methods and Results- Plasma leptin levels were measured at baseline in 377 men (cases) who subsequently experienced a coronary event and in 783 men (controls) who remained free of an event during the 5-year follow-up period of the study. Controls were matched to cases on the basis of age and smoking history and were representative of the entire WOSCOPS cohort. Leptin levels were significantly higher in cases than controls (5.87+/-2.04 ng/mL versus 5.04+/-2.09 ng/mL, P<0.001). In univariate analysis, for each 1 SD increase in leptin, the relative risk (RR) of an event increased by 1.25 (95% confidence interval [CI], 1.10 to 1.43; P<0.001). There was minimal change in this RR with correction for body mass index (RR, 1.24; 95% CI, 1.06 to 1.45; P=0.006) or with further correction for classic risk factors, including age, lipids, and systolic blood pressure (RR, 1.20; 95% CI, 1.02 to 1.42; P=0.03). Leptin correlated with C-reactive protein (r=0.24, P<0.001) and, even with this variable added to the model, leptin retained significance as a predictor of coronary events (RR, 1.18; 95% CI, 1.00 to 1.39; P=0.05) at the expense of C-reactive protein.We show, for the first time, in a large prospective study that leptin is a novel, independent risk factor for coronary heart disease.

Accumulating evidence implicates inflammation as a potential pathway in the pathogenesis of type 2 diabetes. The objective of the present study was to assess the ability of C-reactive protein (CRP) to predict the development of diabetes in middle-aged men in the West of Scotland Coronary Prevention Study. Baseline plasma samples for CRP measurement were available for 5,245 men of whom 127 were classified as having a transition from normal glucose control to overt diabetes during the study, based on American Diabetes Association criteria. Baseline CRP was an important predictor of the development of diabetes in univariate analysis (hazard ratio [HR] for an increase of 1 SD = 1.55; 95% CI 1.32-1.82; P < 0.0001). In multivariate analysis, CRP remained a predictor of diabetes development (HR 1.30; 95% CI 1.07-1.58; P = 0.0075) independent of other clinically employed predictors, including baseline BMI and fasting triglyceride and glucose concentrations. Moreover, there was a graded increase in risk across CRP quintiles throughout the study, evident at even 1 year of follow-up. The highest quintile (CRP >4.18 mg/l) was associated with a greater than threefold risk of developing diabetes (HR 3.07; 95% CI 1.33-7.10) in a multivariate analysis at 5 years. Thus, CRP predicts the development of type 2 diabetes in middle-aged men independently of established risk factors. Because CRP, the most commonly used acute-phase protein in clinical practice, is very stable in serum, our observations have clinical potential in helping to better predict individuals destined to develop type 2 diabetes. They also add to the notion that low-grade inflammation is important in the pathogenesis of type 2 diabetes.

BackgroundInflammation and oxidative stress play critical roles in patients with chronic obstructive pulmonary disease (COPD). Mitochondrial oxidative stress might be involved in driving the oxidative stress–induced pathology.ObjectiveWe sought to determine the effects of oxidative stress on mitochondrial function in the pathophysiology of airway inflammation in ozone-exposed mice and human airway smooth muscle (ASM) cells.MethodsMice were exposed to ozone, and lung inflammation, airway hyperresponsiveness (AHR), and mitochondrial function were determined. Human ASM cells were isolated from bronchial biopsy specimens from healthy subjects, smokers, and patients with COPD. Inflammation and mitochondrial function in mice and human ASM cells were measured with and without the presence of the mitochondria-targeted antioxidant MitoQ.ResultsMice exposed to ozone, a source of oxidative stress, had lung inflammation and AHR associated with mitochondrial dysfunction and reflected by decreased mitochondrial membrane potential (ΔΨm), increased mitochondrial oxidative stress, and reduced mitochondrial complex I, III, and V expression. Reversal of mitochondrial dysfunction by the mitochondria-targeted antioxidant MitoQ reduced inflammation and AHR. ASM cells from patients with COPD have reduced ΔΨm, adenosine triphosphate content, complex expression, basal and maximum respiration levels, and respiratory reserve capacity compared with those from healthy control subjects, whereas mitochondrial reactive oxygen species (ROS) levels were increased. Healthy smokers were intermediate between healthy nonsmokers and patients with COPD. Hydrogen peroxide induced mitochondrial dysfunction in ASM cells from healthy subjects. MitoQ and Tiron inhibited TGF-β–induced ASM cell proliferation and CXCL8 release.ConclusionsMitochondrial dysfunction in patients with COPD is associated with excessive mitochondrial ROS levels, which contribute to enhanced inflammation and cell hyperproliferation. Targeting mitochondrial ROS represents a promising therapeutic approach in patients with COPD. Inflammation and oxidative stress play critical roles in patients with chronic obstructive pulmonary disease (COPD). Mitochondrial oxidative stress might be involved in driving the oxidative stress–induced pathology. We sought to determine the effects of oxidative stress on mitochondrial function in the pathophysiology of airway inflammation in ozone-exposed mice and human airway smooth muscle (ASM) cells. Mice were exposed to ozone, and lung inflammation, airway hyperresponsiveness (AHR), and mitochondrial function were determined. Human ASM cells were isolated from bronchial biopsy specimens from healthy subjects, smokers, and patients with COPD. Inflammation and mitochondrial function in mice and human ASM cells were measured with and without the presence of the mitochondria-targeted antioxidant MitoQ. Mice exposed to ozone, a source of oxidative stress, had lung inflammation and AHR associated with mitochondrial dysfunction and reflected by decreased mitochondrial membrane potential (ΔΨm), increased mitochondrial oxidative stress, and reduced mitochondrial complex I, III, and V expression. Reversal of mitochondrial dysfunction by the mitochondria-targeted antioxidant MitoQ reduced inflammation and AHR. ASM cells from patients with COPD have reduced ΔΨm, adenosine triphosphate content, complex expression, basal and maximum respiration levels, and respiratory reserve capacity compared with those from healthy control subjects, whereas mitochondrial reactive oxygen species (ROS) levels were increased. Healthy smokers were intermediate between healthy nonsmokers and patients with COPD. Hydrogen peroxide induced mitochondrial dysfunction in ASM cells from healthy subjects. MitoQ and Tiron inhibited TGF-β–induced ASM cell proliferation and CXCL8 release. Mitochondrial dysfunction in patients with COPD is associated with excessive mitochondrial ROS levels, which contribute to enhanced inflammation and cell hyperproliferation. Targeting mitochondrial ROS represents a promising therapeutic approach in patients with COPD.

The PROspective Study of Pravastatin in the Elderly at Risk (PROSPER) is a randomized, double-blind, placebo-controlled trial designed to test the hypothesis that treatment with pravastatin will diminish risk of subsequent major vascular events in a cohort of men and women (70 to 82 years old) with preexisting vascular disease or significant risk of developing this condition. Five thousand eight hundred four men and women in addition to receiving advice on diet and smoking, have been randomized equally to treatment with 40 mg pravastatin/day or matching placebo in 3 centers (Cork, Ireland, Glasgow, Scotland, and Leiden, The Netherlands). Following an average 3.5-year intervention period, a primary assessment will be made of the influence of this therapy on major vascular events (a combination of coronary heart disease, death, nonfatal myocardial infarction, and fatal and nonfatal stroke). A number of additional analyses will also be conducted on the individual components of the primary end point, on men, on women, and on subjects with and without previous evidence of vascular disease. Finally, an assessment will be made of the effects of treatment on cognitive function, disability, hospitalization or institutionalization, vascular mortality, and all-cause mortality.

BJOG: An International Journal of Obstetrics & GynaecologyVolume 103, Issue 7 p. 614-620 Potential pathogenic roles of aberrant lipoprotein and fatty acid metabolism in pre-eclampsia Naveed Sattar, Naveed Sattar Registrar in Pathological Biochemistry Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this authorAllan Gaw, Allan Gaw British Heart Foundation Research Fellow Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this authorChris J. Packard, Chris J. Packard Professor in Pathological Biochemistry Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this authorIan A. Greer, Ian A. Greer Professor of Obstetrics and Gynaecology Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this author Naveed Sattar, Naveed Sattar Registrar in Pathological Biochemistry Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this authorAllan Gaw, Allan Gaw British Heart Foundation Research Fellow Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this authorChris J. Packard, Chris J. Packard Professor in Pathological Biochemistry Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this authorIan A. Greer, Ian A. Greer Professor of Obstetrics and Gynaecology Department of Pathological Biochemistry, Royal Infirmary, GlasgowSearch for more papers by this author First published: July 1996 https://doi.org/10.1111/j.1471-0528.1996.tb09827.xCitations: 90Read the full textAboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume103, Issue7July 1996Pages 614-620 RelatedInformation

OBJECTIVE: To determine the influence of apolipoprotein E on cognitive decline in a cohort of elderly men and women. DESIGN: Prospective study. SETTING: Scotland, Ireland, and the Netherlands. PARTICIPANTS: Five thousand eight hundred four subjects aged 70 to 82 from the Prospective Study of Pravastatin in the Elderly at Risk (PROSPER). MEASUREMENTS: Subjects were assessed at baseline and over a mean 3.2‐year (range 0.7–4.2) follow‐up for memory (Picture‐Word Recall), speed of information processing (Stroop and Letter‐Digit Coding), global cognitive function (Mini‐Mental State Examination), and activities of daily living. RESULTS: At baseline, subjects with apolipoprotein E 4 versus those without E 4 had poorer memory performance (mean score difference −0.20 (95% confidence interval (CI)=−0.31 to −0.09) for immediate recall and −0.32 (95% CI=−0.48 to −0.16) for delayed recall and slower information processing (difference in Stroop, 2.79 seconds, (95% CI=1.20–4.28); Letter‐Digit score, −0.36, (95% CI=−0.77–0.05). Subjects with apolipoprotein E 4 showed a greater decline in immediate (−0.22, 95% CI=−0.33 to −0.11) and delayed (−0.30, 95% CI=−0.46 to −0.15) memory scores but no significant change in speed of information processing (Stroop, P =.17; Letter‐Digit, P =.06). Memory scores decreased 2.5% from baseline in those without E 4 , 4.3% in E 4 heterozygotes ( P =.01 for immediate and P =.03 for delayed, vs no E 4 ) and 8.9% to 13.8% in E 4 homozygotes ( P =.04 for immediate and P =.004 for delayed, vs heterozygotes). Apolipoprotein E 4 was associated with greater decline in instrumental activities of daily living ( P &lt;.001). Cognitive decline was not associated with lipoprotein levels. CONCLUSION: Findings in PROSPER indicate that E 4 is associated with more‐rapid cognitive decline and may, therefore, predispose to dementia.

To examine metabolic changes (lipids, liver enzymes, blood pressure, and weight) potentially associated with conversion to diabetes, we analyzed serial glucose and other metabolic measures obtained every 6 months within the West of Scotland Coronary Prevention Study trial. Changes in parameters for 86 men who converted to new-onset diabetes (“converters”: two consecutive glucose levels ≥7 mmol/l) were compared with 860 “nonconverters” matched for age and treatment allocation. Eighteen months before the diagnosis, converters to diabetes had elevated (P &amp;lt; 0.01) fasting glucose, weight, triglyceride, alanine aminotransferase (ALT), blood pressure, and white cell count and lower HDL cholesterol compared with nonconverters. The mean (SD) increase in fasting glucose over 18 months in converters was 1.80 (1.52) mmol/l, compared with 0.10 (0.57) in nonconverters. Of parameters measured, only ALT (P = 0.0005) and triglyceride (P = 0.030) increased significantly more over the 18 months in converters compared with nonconverters, but neither parameter increased significantly in nonconverters with high baseline glucose concentrations (&amp;gt;6.1 mmol/l). Finally, only sustained increases in ALT predicted a higher risk for diabetes. We conclude that a relatively rapid rise in fasting glucose levels is frequent in converters to diabetes and that associated increases over time in ALT and potentially triglyceride suggest hepatic fat accumulation as a contributing factor for conversion to diabetes in men at risk.

The role of C-reactive protein (CRP) in predicting vascular events and response to statin therapy remains uncertain. Additional large prospective studies are required.Baseline CRP was related to risk over 3.2 years for primary a combined end point (definite or suspected death from coronary heart disease, nonfatal myocardial infarction, and fatal or nonfatal stroke; n=865 events) and secondary (coronary heart disease events or stroke alone) and tertiary (stroke plus transient ischemic attack) end points in the Prospective Study of Pravastatin in the Elderly at Risk (n=5804 men and women; age, 70 to 82 years). CRP levels were higher in subjects who had a subsequent primary end-point event compared with those who did not (geometric mean; 3.64 mg/L [SD, 3.08 mg/L] versus 3.01 mg/L [SD, 3.05 mg/L]; P<0.0001). CRP correlated positively with body mass index and smoking status and negatively with high-density lipoprotein cholesterol. The unadjusted hazard ratio for the primary end point was 1.48 (95% CI, 1.26 to 1.74) in a comparison of top and bottom thirds for CRP, falling to 1.36 (95% CI, 1.15 to 1.61) with adjustment for established predictors and body mass index. Similar results were obtained for other end points or when results were examined separately by history of vascular disease. However, baseline CRP added minimally to risk prediction beyond conventional predictors and did not relate to the magnitude of pravastatin benefit.Elevated CRP minimally enhances cardiovascular disease prediction beyond established vascular risk factors and does not predict response to statin therapy in elderly subjects at risk. These data suggest that CRP has limited clinical value in cardiovascular disease risk stratification or predicting response to statin therapy in elderly people.

Statins are important in vascular disease prevention in the elderly. However, the best method of selecting older patients for treatment is uncertain. We assessed the role of plasma lipoproteins as predictors of risk and of treatment benefit in the PROspective Study of Pravastatin in the Elderly at Risk (PROSPER).The association of LDLc and HDLc with risk was examined in the 5804 70- to 82-year-old subjects of PROSPER. Baseline LDLc showed no relation to risk of the primary end point in the placebo group (P=0.27), nor did on-treatment LDLc in the pravastatin group (P=0.12). HDLc was inversely associated with risk in subjects on placebo (P=0.0019) but not in those on pravastatin (P=0.24). Risk reduction on pravastatin treatment was unrelated to baseline LDLc (P=0.38) but exhibited a significant interaction with HDLc (P=0.012). Subjects in the lowest 2 quintiles of HDLc (<1.15 mmol/L) had a risk reduction of 33% (hazard ratio, 0.67; 95% confidence limits, 0.55, 0.81; P<0.0001), whereas those with higher HDLc showed no benefit (RR, 1.06; 95% confidence limits, 0.88, 1.27; P=0.53). During follow-up, there was no relation between achieved level of LDLc or HDLc and risk. However, the change in the LDLc/HDLc ratio on statin treatment appeared to account for the effects of therapy.In people >70 years old, HDLc appears to be a key predictor of risk and of treatment benefit. Findings in PROSPER suggest that statin therapy could usefully be targeted to those with HDLc <1.15 mmol/L or an LDLc/HDLc ratio >3.3.

In normal physiologic responses to injury and infection, inflammatory cells enter tissue and sites of inflammation through a chemotactic process regulated by several families of proteins, including inflammatory chemokines, a family of small inducible cytokines. In neutrophils, chemokines chemokine (CXC motif) ligand 1 (CXCL1) and CXCL8 are potent chemoattractants and activate G protein-coupled receptors CXC chemokine receptor 1 (CXCR1) and CXCR2. Several small-molecule antagonists of CXCR2 have been developed to inhibit the inflammatory responses mediated by this receptor. Here, we present the data describing the pharmacology of AZD5069 [N-(2-(2,3-difluorobenzylthio)-6-((2R,3S)-3,4-dihydroxybutan-2-yloxy)[2,4,5,6-(13)C4, 1,3-(15)N2]pyrimidin-4-yl)azetidine-1-sulfonamide,[(15)N2,(13)C4]N-(2-(2,3-difluoro-6-[3H]-benzylthio)-6-((2R,3S)-3,4-dihydroxybutan-2-yloxy)pyrimidin-4-yl)azetidine-1-sulfonamide], a novel antagonist of CXCR2. AZD5069 was shown to inhibit binding of radiolabeled CXCL8 to human CXCR2 with a pIC50 value of 9.1. Furthermore, AZD5069 inhibited neutrophil chemotaxis, with a pA2 of approximately 9.6, and adhesion molecule expression, with a pA2 of 6.9, in response to CXCL1. AZD5069 was a slowly reversible antagonist of CXCR2 with effects of time and temperature evident on the pharmacology and binding kinetics. With short incubation times, AZD5069 appeared to have an antagonist profile with insurmountable antagonism of calcium response curves. This behavior was also observed in vivo in an acute lipopolysaccharide-induced lung inflammation model. Altogether, the data presented here show that AZD5069 represents a novel, potent, and selective CXCR2 antagonist with potential as a therapeutic agent in inflammatory conditions.

Knowledge about the pathogenesis and pathophysiology of chronic obstructive pulmonary disease (COPD) has advanced dramatically over the last 30 years. Unfortunately, this has had little impact in terms of new treatments. Over the same time frame, only one new class of medication for COPD has been introduced. Even worse, the rate at which new treatments are being developed is slowing. The development of new tools for the assessment of new treatments has not kept pace with understanding of the disease. In part, this is because drug development tools require a regulatory review, and no interested party has been in a position to undertake such a process. In order to facilitate the development of novel tools to assess new treatments, the Food and Drug Administration, in collaboration with the COPD Foundation, the National Heart Lung and Blood Institute and scientists from the pharmaceutical industry and academia conducted a workshop to survey the available information that could contribute to new tools. Based on this, a collaborative project, the COPD Biomarkers Qualification Consortium, was initiated. The Consortium in now actively preparing integrated data sets from existing resources that can address the problem of drug development tools for COPD.

Seven moderately hypercholesterolemic subjects were studied before and after 10 weeks of simvastatin therapy (20 mg/day). Therapy reduced low density lipoprotein (LDL) cholesterol by 39% (p < 0.001), whereas high density lipoprotein and very low density lipoprotein (VLDL) cholesterol were unchanged. Apolipoprotein (apo) B-containing lipoproteins were divided into VLDL1 (Sf 60-400), VLDL2 (Sf 20-60), intermediate density lipoprotein (IDL) (Sf 12-20), and LDL (Sf 0-12), and metabolic changes were sought in dual-tracer VLDL1 and VLDL2 turnover studies. VLDL1 apoB pool size was unaltered by therapy, as were its rates of synthesis, catabolism, and delipidation to VLDL2. Similarly, the VLDL2 apoB pool size was unchanged, but its metabolic fate was altered. The IDL pool size fell significantly (27%, p < 0.01) due entirely to an increased fractional catabolism of the lipoprotein. In our subjects, the circulating mass of LDL apoB decreased (49%, p < 0.01) primarily due to a reduction in its synthesis. Before therapy, 30% of the apoB entering the delipidation cascade in these hyperlipidemic subjects was converted to LDL. On therapy the input remained the same, but direct catabolism from VLDL2 and IDL was increased (p < 0.05), and as a result only 16% eventually appeared in LDL. These kinetic changes were associated with a fall in particle cholesteryl ester content throughout the delipidation cascade. We also observed a link between LDL kinetics and its subfraction distribution. Simvastatin influences the metabolism of LDL, IDL, and VLDL2 but not VLDL1.

Plasma levels of soluble intercellular adhesion molecule 1 (sICAM-1) and monocyte chemoattractant protein 1 (sMCP-1) are associated with increased risk for future coronary events. However, the effect of statins on these inflammatory markers has hardly been studied. We analyzed whether treatment with the different doses of atorvastatin affects sICAM-1 and sMCP-1 plasma levels in subjects at high cardiovascular risk.Achieve Cholesterol Targets Fast with Atorvastatin Stratified Titration was a 12-week, prospective, multicenter, open-label trial that enrolled a total of 2117 subjects with coronary heart disease (CHD), CHD equivalent (defined as diabetes, peripheral vascular disease, or cerebrovascular disease), or a 10-year CHD risk >20%. Subjects with low-density-lipoprotein cholesterol between 100 and 220 mg/dL (2.6-5.7 mmol/L) and triglycerides <600 mg/dL (6.8 mmol/L) were assigned to atorvastatin (10-80 mg/d) based on low-density-lipoprotein cholesterol at screening. The Atorvastatin on Inflammatory Markers study included statin-free patients (N = 1078).At baseline, 52%, 14%, 12%, and 22% of subjects were assigned to doses of 10, 20, 40, and 80 mg, respectively. Levels of sICAM-1 [geometric mean (95% confidence interval); 283.8 (278.1-289.6) vs 131.9 (127.2-136.6) ng/mL, P < .0001] and sMCP-1 [164.1 (159.9-168.2) vs 131.1 (123.1-139.6 pg/mL, P < .0001] were increased in subjects at high cardiovascular risk compared to healthy subjects (n = 130). In the whole population, sICAM-1 and sMCP-1 levels were reduced by atorvastatin [% change (95% confidence interval); -2.2 (-3.8 to -0.6); -4.1 (-6.1 to -2); P = .006 and P = .0002, respectively]. All doses of atorvastatin diminished sICAM-1 and sMCP-1 levels in the highest quartile.Short treatment with atorvastatin reduced sICAM-1 and sMCP-1 plasma levels showing anti-inflammatory effects in subjects at high cardiovascular risk.

1. Perivascular nerves of the sheep middle cerebral artery show immunoreactivity for both vasoactive intestinal polypeptide (VIP) and calcitonin gene-related peptide (CGRP). 2. Rings of endothelium-denuded sheep middle cerebral artery precontracted with 5-hydroxytryptamine were relaxed by CGRP (maximum relaxation = 87.8 +/- 8.1%, pD2 = 7.81 +/- 0.12, n = 12) and by VIP (maximum relaxation = 55.1 +/- 4.1%, pD2 = 7.65 +/- 0.04, n = 18). Rings of endothelium-denuded cat middle cerebral artery precontracted with U46619 were also relaxed by vasoactive intestinal polypeptide (maximum relaxation = 53.1 +/- 6.1%, pD2 = 7.82 +/- 0.11, n = 6). 3. Haemolysate (1 microliters ml-1) inhibited VIP-induced relaxation in endothelium-denuded sheep and cat middle cerebral artery (n = 6) but had no effect on the CGRP-induced relaxation of the sheep middle cerebral artery (n = 6). 4. The relaxant response to VIP in endothelium-denuded sheep middle cerebral artery was inhibited by methylene blue (10 microM) and augmented by either M&B 22948 (10 microM) or superoxide dismutase (150 units ml-1). Indomethacin (1 microM) had no effect. 5. The addition of L-NG-monomethyl arginine (100 microM) inhibited both neurogenic and VIP-induced relaxation of endothelium-denuded sheep MCA by 56 +/- 6% and 60 +/- 6% (n = 5) respectively. The CGRP-induced relaxation was unaffected. 6. It is concluded that neurally mediated vasodilatation in the sheep middle cerebral artery is mediated largely by VIP through a direct action on smooth muscle through a cyclic-GMP-mediated mechanism that appears to involve synthesis of nitric oxide from L-arginine. Vasodilatation by CGRP, which is also contained in perivascular nerves, does not utilize this pathway.

The chemokine receptors CXCR1 and CXCR2 are G-protein-coupled receptors (GPCRs) implicated in mediating cellular functions associated with the inflammatory response. Potent CXCR2 receptor antagonists have been discovered, some of which have recently entered clinical development. The aim of this study was to identify key amino acid residue differences between CXCR1 and CXCR2 that influence the relative antagonism by two compounds that have markedly different chemical structures. By investigating the effects of domain switching and point mutations, we found that the second extracellular loop, which contained significant amino acid sequence diversity, was not important for compound antagonism. We were surprised to find that switching the intracellular C-terminal 60 amino acid domains of CXCR1 and CXCR2 caused an apparent reversal of antagonism at these two receptors. Further investigation showed that a single amino acid residue, lysine 320 in CXCR2 and asparagine 311 in CXCR1, plays a predominant role in describing the relative antagonism of the two compounds. Homology modeling studies based on the structure of bovine rhodopsin indicated a potential intracellular antagonist binding pocket involving lysine 320. We conclude that residue 320 in CXCR2 forms part of a potential allosteric binding pocket on the intracellular side of the receptor, a site that is distal to the orthosteric site commonly assumed to be the location of antagonist binding to GPCRs. The existence of a common intracellular allosteric binding site at GPCRs related to CXCR2 may be of value in the design of novel antagonists for therapeutic intervention.

Inflammation is thought to play an important role in the development of cognitive decline and dementia in old age. The interleukin-1 signalling pathway may play a prominent role in this process. The gene encoding for interleukin-1 beta-converting enzyme (ICE) is likely to influence IL-1 beta levels. Inhibition of ICE decreases the age-related increase in IL-1 beta levels and may therefore improve memory function. We assessed whether genetic variation in the ICE gene associates with cognitive function in an elderly population. All 5804 participants of the PROspective Study of Pravastatin in the Elderly at Risk (PROSPER) were genotyped for the 10643GC, 9323GA, 8996AG and 5352GA polymorphisms in the ICE gene. Cross-sectional associations between the polymorphisms and cognitive function were assessed with linear regression. Longitudinal associations between polymorphisms, haplotypes and cognitive function were assessed with linear mixed models. All associations were adjusted for sex, age, education, country, treatment with pravastatin and version of test where appropriate. Subjects carrying the variants 10643C and 5352A allele had significantly lower IL-1 beta production levels (P < 0.01). Furthermore, we demonstrated that homozygous carriers of the 10643C and the 5352A allele performed better on all executive function tests at baseline and during follow-up compared to homozygous carriers of the wild-type allele (all P < 0.02). The haplotype with two variants present (10643C and 5352A) was associated with better executive function (all P < 0.02) compared to the reference haplotype without variants. For memory function the same trend was observed, although not significant. Genetic variation in the ICE gene is associated with better performance on cognitive function and lower IL-1 beta production levels. This suggests that low levels of IL-1 beta are protective for memory and learning deficits. Inhibition of ICE may therefore be an important therapeutic target for maintaining cognitive function in old age.

The regulation of tissue turnover requires the coordinated activity of both local and systemic factors. Nucleotides exist transiently in the extracellular environment, where they serve as ligands to P2 receptors. Here we report that the localized release of these nucleotides can sensitize osteoblasts to the activity of systemic factors. We have investigated the ability of parathyroid hormone (PTH), a principal regulator of bone resorption and formation, to potentiate signals arising from nucleotide stimulation of UMR-106 clonal rat osteoblasts. PTH receptor activation alone did not lead to [Ca2+]i elevation in these cells, indicating no Gq coupling, however, activation of Gq-coupled P2Y1 receptors resulted in characteristic [Ca2+]i release. PTH potentiated this nucleotide-induced Ca2+ release, independently of Ca2+ influx. PTH-(1–31), which activates only Gs, mimicked the actions of PTH-(1–34), whereas PTH-(3–34), which only activates Gq, was unable to potentiate nucleotide-induced [Ca2+]i release. Despite this coupling of the PTHR to Gs, cAMP accumulation or protein kinase A activation did not contribute to the potentiation. 3-Isobutyl-1-methylxanthine, but not forskolin effectively potentiated nucleotide-induced [Ca2+]i release, however, further experiments proved that cyclic monophosphates were not involved in the potentiation mechanism. Costimulation of UMR-106 cells with P2Y1agonists and PTH led to increased levels of cAMP response element-binding protein phosphorylation and a synergistic effect was observed on endogenous c-fos gene expression following costimulation. In fact the calcium responsive Ca/cAMP response element of the c-fos promoter alone was effective at driving this synergistic gene expression. These findings demonstrate that nucleotides can provide a targeted response to systemic factors, such as PTH, and have important implications for PTH-induced signaling in bone. The regulation of tissue turnover requires the coordinated activity of both local and systemic factors. Nucleotides exist transiently in the extracellular environment, where they serve as ligands to P2 receptors. Here we report that the localized release of these nucleotides can sensitize osteoblasts to the activity of systemic factors. We have investigated the ability of parathyroid hormone (PTH), a principal regulator of bone resorption and formation, to potentiate signals arising from nucleotide stimulation of UMR-106 clonal rat osteoblasts. PTH receptor activation alone did not lead to [Ca2+]i elevation in these cells, indicating no Gq coupling, however, activation of Gq-coupled P2Y1 receptors resulted in characteristic [Ca2+]i release. PTH potentiated this nucleotide-induced Ca2+ release, independently of Ca2+ influx. PTH-(1–31), which activates only Gs, mimicked the actions of PTH-(1–34), whereas PTH-(3–34), which only activates Gq, was unable to potentiate nucleotide-induced [Ca2+]i release. Despite this coupling of the PTHR to Gs, cAMP accumulation or protein kinase A activation did not contribute to the potentiation. 3-Isobutyl-1-methylxanthine, but not forskolin effectively potentiated nucleotide-induced [Ca2+]i release, however, further experiments proved that cyclic monophosphates were not involved in the potentiation mechanism. Costimulation of UMR-106 cells with P2Y1agonists and PTH led to increased levels of cAMP response element-binding protein phosphorylation and a synergistic effect was observed on endogenous c-fos gene expression following costimulation. In fact the calcium responsive Ca/cAMP response element of the c-fos promoter alone was effective at driving this synergistic gene expression. These findings demonstrate that nucleotides can provide a targeted response to systemic factors, such as PTH, and have important implications for PTH-induced signaling in bone. intracellular [Ca2+] parathyroid hormone, where not specified PTH = PTH-(1–34) fluorescent imaging plate reader basal salt solution 2-methylthioadenosine 5′-triphosphate 2-methylthioadenosine 5′-diphosphate 3-isobutyl-1-methylxanthine N 2,2′-O-dibutyrylguanosine 3′,5′-cyclic monophosphate cAMP response element-binding protein calcium and cAMP response element Dulbecco's modified Eagle's medium Nucleotides exist in the extracellular microenvironment of bone due to release from damaged cells at sites of injury, or via more controlled nonlytic release (1Bowler W.B. Tattersall J.A. Hussein R. Dixon C.J. Cobbold P.H. Gallagher J.A. Bone. 1998; 22: 3SCrossref Scopus (51) Google Scholar), where they differentially activate cell surface P2 receptors. Activation of Gq-coupled P2Y receptors leads to [Ca2+]i 1mobilization which has ultimately been found to induce processes including proliferation (2Dixon J.C. Bowler W.B. Littlewood-Evans A. Dillon J.P. Bilbe G. Sharpe G.R. Gallagher J.A. Br. J. Pharmacol. 1999; 127: 1680-1686Crossref PubMed Scopus (100) Google Scholar) and differentiation (3Koshiba M. Apasov S. Sverdlov V. Chen P. Erb L. Turner J.T. Weisman G.A. Sitkovsky M.V. Proc. Natl. Acad. Sci. U. S. A. 1997; 94: 831-836Crossref PubMed Scopus (77) Google Scholar) and to modulate osteoblast-induced bone formation (4Jones S.J. Gray C. Boyde A. Burnstock G. Bone. 1997; 21: 393-399Crossref PubMed Scopus (52) Google Scholar). The bone matrix synthesizing osteoblasts are thought to express multiple P2Y receptor subtypes (5Reimer W.J. Dixon S.J. Am. J. Physiol. 1992; 263: C1040-C1048Crossref PubMed Google Scholar), with expression profile changing as osteoblasts differentiate (6Dixon J.C. Bowler W.B. Walsh C.A. Gallagher J.A. Br. J. Pharmacol. 1997; 120: 777-780Crossref PubMed Scopus (45) Google Scholar). The mechanisms that determine focal responsiveness to systemic factors in bone to allow a local tissue turnover, characteristic of the remodeling process, remain unclear. However, our previous observations that coapplication of nucleotides and parathyroid hormone (PTH) can result in synergistic responses in osteoblasts (7Bowler W.B. Dixon C.J. Halleux C. Maier R. Bilbe G. Fraser W.D. Gallagher J.A. Hipskind R.A. J. Biol. Chem. 1999; 274: 14315-14324Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar) led us to hypothesize that locally released extracellular nucleotides could sensitize surrounding cells to the action of circulating hormones. Parathyroid hormone is one of the most important systemic regulators of bone and mineral homeostasis. The action of PTH on skeletal cells is complex and can result in the stimulation of both resorption and new bone formation (8Raisz L.G. Nature. 1965; 197: 1015-1017Crossref Scopus (68) Google Scholar, 9Dempster D.W. Cosman F. Parisien M. Shen V. Endocrine Rev. 1993; 14: 690-709Crossref PubMed Scopus (671) Google Scholar). This ability to stimulate coupled, but opposing processes, has been attributed to the nature of the receptor that transduces signals arising from PTH stimulation. The PTH1 receptor belongs to a subgroup of seven transmembrane receptors that include those responsive to calcitonin, secretin, and VIP (10Abou-Samra A.B. Juppner H. Force T. Freeman M.W. Kong X-F. Schipani E. Urena P. Richards J. Bonventre J.V. Potts Jr., J.T. Kronenberg H.M. Segre G.V. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 2732-2736Crossref PubMed Scopus (1007) Google Scholar). These receptors are distinct in their ability to couple to both Gs and Gq and hence activate dual signal transduction pathways leading to both cyclic AMP formation and release of Ca2+from intracellular stores ([Ca2+]i). The coupling of the PTH receptor to Gs in osteoblasts is well characterized. Activation of the cAMP signaling pathway is responsible for a number of PTH-induced downstream responses, including expression of the c-fos proto-oncogene (11Tyson D.R. Swarthout J.T. Partridge N.C. Endocrinology. 1999; 140: 1255-1261Crossref PubMed Google Scholar). In addition, the use of truncated PTH fragments in vivo has confirmed that complete N-terminal sequence (vital for Gs activation) is essential to drive the anabolic skeletal effects of PTH (12ArmamentoVillareal R. Ziambaras K. AbbasiJarhomi S.H. Dimarogonas A. Halstead L. Fausto A. Avioli L.V. Civitelli R. J. Bone Min. Res. 1997; 12: 384-392Crossref PubMed Scopus (46) Google Scholar) and cAMP accumulation is often used as a measure of PTH receptor activation (13Du P.F. Seitz P.K. Cooper C.W. Endocrine. 2000; 12: 25-33Crossref PubMed Google Scholar). However, the nature of PTH receptor/Gq-coupling and subsequent downstream responses in osteoblasts following activation remains controversial. Both inositol 1,4,5-trisphosphate-dependent and independent mechanisms for PTH-induced [Ca2+]i release have been reported (14Dunlay R. Hruska K. Am. J. Physiol. 1990; 258: F223-F231PubMed Google Scholar, 15Tong Y. Zull J. Yu L. J. Biol. Chem. 1996; 271: 8183-8191Abstract Full Text Full Text PDF PubMed Scopus (16) Google Scholar). To further complicate the issue of PTH-induced [Ca2+]i release, recent studies in cell types other than osteoblasts have demonstrated that while PTH alone is unable to induce [Ca2+]i release, it can strongly potentiate the [Ca2+]i elevations induced by agonists acting at other Gq-coupled G-protein-coupled receptor. The mechanism behind this potentiation remains unclear but has been attributed to G protein subunit interaction or shuttling of calcium between intracellular stores (16Short A.D. Taylor C.T. J. Biol. Chem. 2000; 275: 1807-1813Abstract Full Text Full Text PDF PubMed Scopus (30) Google Scholar). In a study by Kaplan et al. (17Kaplan A.D. Reimer W.J. Feldman R.D. Dixon S.J. Endocrinology. 1995; 136: 1674-1684Crossref PubMed Google Scholar) using rat UMR-106 cells, PTH alone was shown to produce small [Ca2+]i elevations and also potentiated nucleotide-induced [Ca2+]i elevations. However, in contrast to these findings, we previously demonstrated that PTH alone did not elevate [Ca2+]i and was ineffective at potentiating nucleotide-induced responses in the human osteosarcoma cell line SaOS-2 (7Bowler W.B. Dixon C.J. Halleux C. Maier R. Bilbe G. Fraser W.D. Gallagher J.A. Hipskind R.A. J. Biol. Chem. 1999; 274: 14315-14324Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar). In the same study, however, PTH and nucleotide costimulation of SaOS-2 cells did result in synergistic induction of gene expression. Considering the observations described above, the current study has addressed: (i) the controversy concerning the ability of PTH to effect [Ca2+]i release in the rat osteosarcoma cell line UMR-106, (ii) the mechanisms by which PTH can potentiate [Ca2+]i release induced by locally released nucleotides, and (iii) whether this potentiated [Ca2+]i response can ultimately drive downstream cellular responses in osteoblasts. Dulbecco's modified Eagle's medium (DMEM), α-modified Eagle's medium (α-MEM), Ham's F-12, and fetal calf/bovine serum were purchased from Life Technologies (United Kingdom or France). Human parathyroid hormones (PTH) 1–34 and 1–31, and bovine PTH-(3–34) were purchased from Peninsula Laboratories (UK). Nucleotides, bovine serum albumin, fluo-3 AM, potato apyrase, H-89, IBMX, dibutyryl-cGMP, SKF96365, nitrocellulose membranes, and peroxidase-coupled goat anti-rabbit antibodies were obtained from Sigma (UK). Phospho-CREB and CREB specific antisera were obtained from New England Biolabs (UK). Nitrocellulose membranes and enhanced chemiluminescence reagents were acquired from Amersham Pharmacia Biotech (UK) or PerkinElmer Life Sciences (Belgium). Zetabind hybridization membrane was purchased from Cuno (Meriden, CT). Luciferase lysis reagent and luciferase reagent were purchased fromPromega (UK). UMR-106 cells were kindly provided by T. J. Martin, Melbourne, Australia. UMR-106 rat osteoblast-like cells were cultured in α-MEM supplemented with 10% fetal calf serum, 100 μg/ml streptomycin, 100 units/ml penicillin, and 2 mml-glutamine. The cells were maintained at 37 °C in a humidified atmosphere of 95% air and 5% CO2. Confluent cells were either induced for subsequent preparation of whole cell extracts or RNA isolation, or harvested upon confluence for Ca2+ release measurements in the FLIPR system (18Kuntzweiler T.A. Arneric S.P. Donnelly-Roberts D.L. Drug Dev. Res. 1998; 44: 14-20Crossref Scopus (39) Google Scholar). The UMR-106 multi CRE c-fos reporter cell line was cultured in Ham's F-12 and DMEM (1:1) under similar conditions. Cells were grown in 225-cm2 vented cap flasks, harvested upon confluence, and resuspended in basal salt solution (BSS) composition (mm): NaCl (125), KCl (5Reimer W.J. Dixon S.J. Am. J. Physiol. 1992; 263: C1040-C1048Crossref PubMed Google Scholar), MgCl2 (1Bowler W.B. Tattersall J.A. Hussein R. Dixon C.J. Cobbold P.H. Gallagher J.A. Bone. 1998; 22: 3SCrossref Scopus (51) Google Scholar), CaCl2 (1.5), HEPES (25Donahue H.J. Fryer M.J. Erikson E.F. Heath H. J. Biol. Chem. 1988; 263: 13522-13527Abstract Full Text PDF PubMed Google Scholar), glucose (5Reimer W.J. Dixon S.J. Am. J. Physiol. 1992; 263: C1040-C1048Crossref PubMed Google Scholar), and 1 mg/ml bovine serum albumin, pH 7.3. Cells were loaded with 17 μmfluo-3-AM for 1 h at 37 °C with agitation and in the presence of apyrase (2 units/ml), after which cells were washed 3 times in BSS and aliquoted into 96-well black wall, clear base plates at a density of 2.5 × 105 cells/well. These were then centrifuged to obtain a confluent layer of cells on the base of the plate and cells were subsequently washed 4 times in BSS. Ca2+ flux was measured in all 96 wells simultaneously and in real time using a Fluorescent Imaging Plate Reader (FLIPR). When effects of PTH and nucleotides together were studied the cells were incubated for 60 s with PTH prior to exposure to nucleotide. The effects of IBMX and forskolin were assessed by their introduction to the cells 6 min prior to nucleotide addition, H-89 was introduced 10 min before the addition of nucleotides and dibutyryl-cGMP 30 min before nucleotide addition. In all cases [Ca2+]i release was measured for 1 min after nucleotide addition, a measurement of fluorescence being taken every second. Whole cell extracts were prepared as previously described (19Hipskind R.A. Baccarini M. Nordheim A. Mol. Cell. Biol. 1994; 14: 6219-6231Crossref PubMed Scopus (137) Google Scholar). At the appropriate time point, plates were placed on ice and the cell layer quickly washed twice with ice-cold phosphate-buffered saline (140 mm NaCl, 10 mm NaPO4, pH 7.3) containing 10 mmNaF and 100 μm Na3VO4. Cells were solubilized in 10 mm Tris-HCl, pH 7.05, 50 mmNaCl, 30 mm sodium pyrophosphate, 50 mm NaF, 5 mm ZnCl2, 1% (v/v) Triton X-100, 100 μm Na3VO4, 20 mmβ-glycerophosphate, 10 mm 4-nitrophenyl phosphate, 1 mm dithiothreitol, 0.5 mm benzamidine, 2.5 μg/ml aprotinin, leupeptin, pepstatin, 0.2 mmphenylmethylsulfonyl fluoride, and 200 nm okadaic acid. The cell layer was collected by scraping and lysis completed by vortexing for 60 s. The lysate was clarified by centrifugation at 14,000 × g at 4 °C for 15 min and the protein concentration determined with Bradford's reagent using bovine serum albumin as standard. An aliquot for Western analysis was denatured immediately in 2% SDS, 2% glycerol, 50 mm Tris-HCl, pH 6.8, 1% 2-mercaptoethanol and the remaining supernatant stored at −70 °C. 15 μg of whole cell extracts were separated on an 8.5% SDS-polyacrylamide electrophoresis minigel and transferred electrophoretically to a nitrocellulose membrane. The filter was blocked for 1 h at room temperature in 5% (w/v) low fat milk powder-TBST (10 mm Tris-HCl, pH 7.5, 100 mm NaCl, 0.1% Tween 20), followed by incubation overnight at 4 °C with the specific antiserum indicated below in blocking buffer. After washing in TBST, the blots were incubated for 1 h at room temperature with peroxidase-coupled goat anti-rabbit antibody, diluted 1/1500 in 5% (w/v) low fat milk powder-TBST. After washing, the immune complexes were visualized using enhanced chemiluminescence. The antisera used were: anti-CREB and anti-phospho Ser133CREB, diluted 1:1000. Total RNA was extracted from control and stimulated cells according to the protocol of Chomczynski and Sacchi (20Chomczynski P. Sacchi N. Anal. Biochem. 1987; 162: 156-161Crossref PubMed Scopus (63232) Google Scholar). Briefly, cells were lysed in 4 m guanidine thiocyanate, 0.5% Sarkosyl, 0.1 m mercaptoethanol, 25 mm sodium citrate, pH 7.0, followed by acid phenol/chloroform extraction. RNA was stored as a precipitate at −70 °C. 10 μg of total RNA was denatured and electrophoresed through a 0.8% (w/v) agarose gel containing 3.7% formaldehyde (v:v), followed by transfer to Zetabind hybridization membrane. Blots were prehybridized at 65 °C in 50% formamide, 5 × SSC, 5 × Denhardt's reagent, 1% SDS, 50 mm sodium phosphate buffer, pH 6.8, 5 mg/ml total calf liver RNA, 200 μg of tRNA, and probed with a c-fosriboprobe spanning exons 3 and 4 of the human c-fos gene mixed with a riboprobe derived from the rat gapdh cDNA as previously described (19Hipskind R.A. Baccarini M. Nordheim A. Mol. Cell. Biol. 1994; 14: 6219-6231Crossref PubMed Scopus (137) Google Scholar). Membranes were washed for 30 min at 65 °C with 0.2 × SSC, 1% SDS solution and mRNAs visualized using phosphorstorage technology and autoradiography with Kodak X-AR film and intensifying screens at −70 °C. RNA loading and integrity were measured by UV shadowing of the filter prior to hybridization. Reporter cells were seeded into white 96-well plates (Dynex) at a density of 96,000 cells/well in 50% DMEM, 50% Ham's F-12 containing 10% fetal calf serum. Cells were incubated for 24 h, after which time medium was replaced by 50% DMEM, 50% Ham's F-12 medium containing 0.1% bovine serum albumin and incubated for a further 24 h. Cells were induced for 4 h with ×10 concentration of agonist stock solution prepared in serum-free medium to the final concentrations indicated in Fig. 9. After induction, cells were washed twice in cold phosphate-buffered saline and lysed in luciferase cell culture lysis reagent for 15 min at room temperature (10 μl/well). A microtiter plate luminometer (ML 3000, Dynex Technologies) was set up in the enhanced flash mode (maximum sensitivity, delay time = 2 s, integration time = 10 s). Luciferase reagent (100 μl) was automatically added and light emission was measured every 10 ms during a period of 10 s. Data was recorded as the peak value of relative light units. Treatments were performed in triplicate and the mean fold increase in luciferase activity was calculated relative to mock-induced cells, which was taken as 1-fold. Since osteoblasts have been described to express numerous subtypes of the P2Y family, and previous studies to receptor profile UMR-106 cells have been incomplete, we used the FLIPR system to fully investigate P2Y receptor expression by these cells. The effects of a series of known P2 agonists on [Ca2+]i elevation were measured in fluo-3-loaded cells. The agonist potency order (p[A]50) was as follows: 2-MeSADP (5.27 ± 0.09) > 2-MeSATP (4.89 ± 0.15) > ADP (4.60 ± 0.15) > ATP (4.57 ± 0.11), suggesting that P2Y1is the predominant P2 receptor expressed by UMR-106 cells (Fig.1). While AMP, αβMeATP, and UDP were inactive at concentrations between 0.1 and 100 μm (data not shown), UTP evoked a small [Ca2+]i elevation (Fig. 1). This profile differs from that suggested by Kaplan et al. (17Kaplan A.D. Reimer W.J. Feldman R.D. Dixon S.J. Endocrinology. 1995; 136: 1674-1684Crossref PubMed Google Scholar) in an earlier study, where predominant functional effects were elicited by P2Y2/Y4 receptor agonists. We have previously reported heterogeneity of P2 receptor expression in populations of primary and clonal osteoblasts (7Bowler W.B. Dixon C.J. Halleux C. Maier R. Bilbe G. Fraser W.D. Gallagher J.A. Hipskind R.A. J. Biol. Chem. 1999; 274: 14315-14324Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar), a process that appears to be differentiation-dependent (6Dixon J.C. Bowler W.B. Walsh C.A. Gallagher J.A. Br. J. Pharmacol. 1997; 120: 777-780Crossref PubMed Scopus (45) Google Scholar). Diverse experimental culture methods or passage number may therefore account for the apparent differences in receptor profile between cells in this study and those used in the study of Kaplan et al. (17Kaplan A.D. Reimer W.J. Feldman R.D. Dixon S.J. Endocrinology. 1995; 136: 1674-1684Crossref PubMed Google Scholar). In a previous study using UMR-106 cells PTH potentiated [Ca2+]i elevations induced by certain nucleotide agonists (17Kaplan A.D. Reimer W.J. Feldman R.D. Dixon S.J. Endocrinology. 1995; 136: 1674-1684Crossref PubMed Google Scholar). In one of our earlier studies, however, using the human osteosarcoma cell line SaOS-2, we observed no potentiation of nucleotide-induced [Ca2+]i release following costimulation with PTH (7Bowler W.B. Dixon C.J. Halleux C. Maier R. Bilbe G. Fraser W.D. Gallagher J.A. Hipskind R.A. J. Biol. Chem. 1999; 274: 14315-14324Abstract Full Text Full Text PDF PubMed Scopus (108) Google Scholar). To clarify these differences, the effects of human PTH-(1–34) (100 ng/ml), in combination with known nucleotide agonists, on [Ca2+]i were investigated in UMR-106 cells using the FLIPR. In contrast to the earlier study of Kaplan et al. (17Kaplan A.D. Reimer W.J. Feldman R.D. Dixon S.J. Endocrinology. 1995; 136: 1674-1684Crossref PubMed Google Scholar), we repeatedly saw no elevation of [Ca2+]i in response to PTH (0.1–500 ng/ml, data not shown). We did, however, observe a striking potentiation to the [Ca2+]i response with all nucleotides tested (Fig.2 shows just ADP for clarity). A potentiated calcium response was observed when cells were stimulated with UTP and PTH, therefore confirming that P2Y2 receptors are expressed by these cells, albeit at low levels. To begin to investigate the mechanism behind this observed potentiation we wondered whether calcium influx contributed to this response. Again using the FLIPR we found that in the absence of extracellular Ca2+, ADP and PTH costimulation induced a [Ca2+]i response approximately six times greater than that induced by ADP alone (Fig.3 A). However, this response was depressed in comparison to that resulting from the same experiment performed in the presence of extracellular Ca2+ (Fig.3 A). Interestingly, introduction of the receptor-activated calcium channel blocker SKF96365 resulted in an increase in the observed potentiation between ADP and PTH (Fig. 3 B). Thus calcium influx is not the major component in the potentiation we observe. To further elucidate the mechanisms behind the observed potentiated [Ca2+]i release we used truncated forms of the PTH peptide, which have previously been reported to have selective stimulatory effects upon the G-proteins that couple second messenger signaling pathways to the PTH receptor. It has been reported that stimulation of the PTH receptor by PTH-(1–34) results in activation of both Gq and Gs and therefore utilizes both PKC and adenylyl cyclase to initiate signaling cascades (10Abou-Samra A.B. Juppner H. Force T. Freeman M.W. Kong X-F. Schipani E. Urena P. Richards J. Bonventre J.V. Potts Jr., J.T. Kronenberg H.M. Segre G.V. Proc. Natl. Acad. Sci. U. S. A. 1992; 89: 2732-2736Crossref PubMed Scopus (1007) Google Scholar). We report here that PTH-(1–31) (100 ng/ml), which stimulates adenylyl cyclase but not PKC (21Jouishomme H. Whitfield J.F. Gagnon L. Maclean S. Isaacs R. Chakravarthy B. Durkin J. Neugebauer W. Willick G. Rixon R.H. J. Bone Min. Res. 1994; 9: 943-949Crossref PubMed Scopus (124) Google Scholar), is able to potentiate ADP-induced Ca2+responses as effectively as the full-length peptide (PTH-(1–34)) (Fig.4 A). In contrast, PTH-(3–34) (100 ng/ml) which does not activate adenylyl cyclase (22Goltzman D. Peytremann A. Callahan E. Tregear G.W. Potts Jr., J.T. J. Biol. Chem. 1975; 250: 3199-3203Abstract Full Text PDF PubMed Google Scholar) was unable to potentiate the [Ca2+]i release induced by ADP (Fig. 4 A). To confirm that this peptide was functionally interacting with the PTH receptor it was introduced in combination with ADP and the active peptides, PTH-(1–34) or -(1–31). PTH-(3–34) was shown to act as a functional antagonist at the PTH receptor as it depressed the [Ca2+]i release induced by ADP in combination with either PTH-(1–34) or PTH-(1–31) (Fig.4 B). As a main downstream target for activated adenylyl cyclase pathways, we next investigated the potential role of PKA in driving potentiated [Ca2+]i release in UMR-106 cells. Preincubation of UMR-106 cells with the PKA inhibitor H-89 (1–100 μm) did not affect potentiated [Ca2+]i release induced by ADP and PTH, indicating that PKA activity is not necessary to initiate the potentiation (Table I). H-89 was inhibitory in these cells, since it blocked PTH-induced phosphorylation of CREB on Ser133, the residue phosphorylated by active PKA (Fig.5).Table IH-89 does not depress ADP/PTH-induced potentiated Ca2+ elevationsAgonist(s)Response of ADP (10−4m) + PTH (100 ng/ml)ADP22%ADP + PTH100%ADP + PTH + H89104%H-89 (1–100 μm) was introduced to fluo-3-loaded UMR-106 cells 10 min before ADP (100 μm) and PTH (100 ng/ml) addition. Ca2+ release was measured according to fluorescence every 30 s for 10 min after H-89 addition, then every second for a further 1 min after ADP/PTH addition. Fluorescence is expressed as a percentage of the response to 100 μm ADP + 100 ng/ml PTH. H-89 had no effect on the potentiation at any concentration tested, figures show effects of 10 μm H-89 (n = 3). Open table in a new tab H-89 (1–100 μm) was introduced to fluo-3-loaded UMR-106 cells 10 min before ADP (100 μm) and PTH (100 ng/ml) addition. Ca2+ release was measured according to fluorescence every 30 s for 10 min after H-89 addition, then every second for a further 1 min after ADP/PTH addition. Fluorescence is expressed as a percentage of the response to 100 μm ADP + 100 ng/ml PTH. H-89 had no effect on the potentiation at any concentration tested, figures show effects of 10 μm H-89 (n = 3). The results above showed that potentiation occurs with a PTH peptide that activates a signaling pathway involving adenylyl cyclase but not PKA. To assess whether elevated cAMP could drive potentiated [Ca2+]i release, cells were costimulated with ADP and the cell permeable adenylyl cyclase activator, forskolin (50 μm), in the absence of extracellular calcium. Forskolin was unable to potentiate ADP-induced calcium responses (TableII). Forskolin alone was shown to elevate intracellular cAMP levels in these cells and to induce CREB phoshorylation via PKA phosphorylation (data not shown). We also tested the ability of the nonspecific phosphodiesterase inhibitor, IBMX (1 mm) to potentiate ADP-induced [Ca2+]i release. In contrast to forskolin, IBMX effectively potentiated ADP-induced [Ca2+]i release (Table II).Table IIIBMX, but not forskolin can potentiate nucleotide-induced [Ca2+]i releaseAgonist(s)Response of ADP (10−4m) + PTH (100 ng/ml)ADP65%ADP + PTH100%ADP + forskolin64%ADP + IBMX96%Forskolin (50 μm), IBMX (1 mm), and PTH (100 ng/ml) were introduced to fluo-3-loaded UMR-106 cells 6 min before ADP addition (ADP concentration range, 0.3 μm to 1 mm). Ca2+ release was measured according to fluorescence every second for 1 min after ADP addition. Fluorescence is expressed as a percentage of the response to 100 μmADP + 100 ng/ml PTH (n = 3). Open table in a new tab Forskolin (50 μm), IBMX (1 mm), and PTH (100 ng/ml) were introduced to fluo-3-loaded UMR-106 cells 6 min before ADP addition (ADP concentration range, 0.3 μm to 1 mm). Ca2+ release was measured according to fluorescence every second for 1 min after ADP addition. Fluorescence is expressed as a percentage of the response to 100 μmADP + 100 ng/ml PTH (n = 3). Since the results obtained with forskolin and IBMX suggested that accumulation of cGMP, downstream of the activated PTH receptor, may be causing the potentiation of ADP-induced [Ca2+]i levels, we costimulated cells with ADP and the cell permeable cGMP analogue, dibutyryl-cGMP (300 μm), in the absence of extracellular calcium. Dibutyryl-cGMP was unable to potentiate ADP-induced [Ca2+]i elevations (Fig.6), eliminating accumulation of this cyclic monophosphate as a possible mechanism of potentiation. Dibutyryl-cGMP alone produced no [Ca2+]i elevation (data not shown). Phosphorylation of CREB on Ser133 is strongly linked to signaling-induced transcriptional activation. Ser133 is a substrate for PKA but also for kinases activated by elevated intracellular calcium. We investigated whether ADP and PTH alone or in combination induced CREB phosphorylation in UMR-106 cells. We performed Western analysis of whole cell extracts using antisera directed against CREB phosphorylated on Ser133. The P2Y1 agonist ADP (100 μm) induced low CREB activation following 15 min stimulation, while PTH-(1–34) (100 ng/ml) gave rise to a robust transcription factor phosphorylation. In combination, however, agonists induced levels of CREB activation greater than those seen with either agonist alone. Quantitative values for CREB and phospho-CREB were obtained by densitometric analysis (Fig.7). To determine the impact of the observed potentiated calcium response on gene expression we measured the effects of P2Y1 agonists and PTH on the levels of endogenous c-fos transcription in UMR-106 cells. This proto-oncogene is of particular relevance when studying osteoblasts as it has been implicated in many of the processes that govern skeletal tissue remodeling (23Grigoriadis A.E. Wang Z-Q. Cecchini M.G. Hofstetter W. Felix R. Fleish H.A. Wagner E.F. Science. 1994; 266: 443-448Crossref PubMed Scopus (1082) Google Scholar). Stimulation of quiescent UMR-106 cells with the P2Y1 agonists ADP and 2-MeSATP (10 μm) for 45 min had a very weak effect on c-fos mRNA expression, as assessed by Northern analysi

An analysis of apolipoprotein (apo) R turnovers conducted in subjects with moderate hypercholesterolemia was performed to discover relationships that may exist between apoR ki- netic parameters and plasma lipid and lipoprotein levels.A group of 21 subjects with plasma cholesterol in the range 250-300 mg/dl and triglyceride < 265 mg/dl were injected with tracers of '3'I-labeled very low density lipoprotein 1 (VLDL,, SI 60-400) and 1251-labeled VLDL, (SI 20-60) prepared by cumulative flotation ultracentrifugation.The metabolism of apoR in these fractions was followed through intermediate density (IDL, Sr 12-20) to low density (LDL, SI 0-12) lipoprotein.The most consistent feature giving rise to the higher apoR I&ls that occurred in VLDL?, IDL, and LDL in the hypercholesterolemic group was increased input of VLDL2 (787 * 607 (SD) mg/day vs. 349 * 213 in normals, P < 0.01).VLDLl apoR input was variably affected and not si.gnificantlydifferent from normal.However, the plasma residence time of this subfraction was increased (0.15 * 0.07 days vs. 0.08 * 0.03 days in normals, P < 0.001) due to a decreased fractional rate of direct catabolism.Fractional transfer rates (FTR) down the delipidation cascade and other fractional rates of direct catabolism were, overall, not significantly different from normal.The plasma residence time of VLDL, apoR and LDL apoB was similar in hypercholesterolemic and normal subjects, while that of IDL apoR was slightly increased.Variation in LDL apoR mass within the hypercholesterolemic group correlated with VLDL, apoR input (r = 0.58, P = 0.006), the fractional rate of transfer from IDL to LDL (r = 0.61, P = 0.003).and direct LDL input (r = 0.64, P -0.002).The proportion of LDL apoR mass derived by direct, i.e., VLDL-independent input, varied from 5 to 50% and was inversely correlated with plasma triglyceride ( r = -0.53,P -0.014) and positively with HDL2 ( r -0.66, P = 0.002).In addition, the amount of direct LDL input was related to the amount of VLDL, removed by direct catabolism (r -0.53, P -0.013).a The analysis indicated that moderate hypercholesterolemia arose principally from overproduction of small VLDL, while variation in VLDLl input and the IDL to LDL conversion rate (presumably hepatic lipase-mediated) modulated the extent of the elevation in LDL apoR.-Gaw,A.,

Increasing evidence indicates that the Fas/Fas ligand interaction is involved in atherogenesis. We sought to analyze soluble Fas (sFas) and soluble Fas ligand (sFasL) concentrations in subjects at high cardiovascular risk and their modulation by atorvastatin treatment.ACTFAST was a 12-week, prospective, multicenter, open-label trial which enrolled subjects (statin-free or statin-treated at baseline) with coronary heart disease (CHD), CHD-equivalent, or 10-year CHD risk > 20%. Subjects with LDL-C between 100 to 220 mg/dL (2.6 to 5.7 mmol/L) and triglycerides < or = 600 mg/dL (6.8 mmol/L) were assigned to a starting dose of atorvastatin (10 to 80 mg/d) based on LDL-C at screening. Of the 2117 subjects enrolled in ACTFAST, AIM sub-study included the 1078 statin-free patients. At study end, 85% of these subjects reached LDL-C target. Mean sFas levels were increased and sFasL were reduced in subjects at high cardiovascular risk compared with healthy subjects. Atorvastatin reduced sFas in the whole population as well as in patients with metabolic syndrome or diabetes. Minimal changes were observed in sFasL.sFas concentrations are increased and sFasL are decreased in subjects at high cardiovascular risk, suggesting that these proteins may be novel markers of vascular injury. Atorvastatin reduces sFas, indicating that short-term treatment with atorvastatin exhibits antiinflammatory effects in these subjects.

Routinely collected electronic patient records are already widely used in epidemiological research. In this work we investigated the potential for using them to identify endpoints in clinical trials.The events recorded in the West of Scotland Coronary Prevention Study (WOSCOPS), a large clinical trial of pravastatin in middle-aged hypercholesterolaemic men in the 1990s, were compared with those in the record-linked deaths and hospitalisations records routinely collected in Scotland.We matched 99% of fatal study events by date. We showed excellent matching (97%) of the causes of fatal endpoint events and good matching (>80% for first events) of the causes of nonfatal endpoint events with a slightly lower rate of mismatching of record linkage than study events (19% of first study myocardial infarctions (MI) and 4% of first record linkage MIs not matched as MI). We also investigated the matching of non-endpoint events and showed a good level of matching, with >78% of first stroke/TIA events being matched as stroke/TIA. The primary reasons for mismatches were record linkage data recording readmissions for procedures or previous events, differences between the diagnoses in the routinely collected data and the conclusions of the clinical trial expert adjudication committee, events occurring outside Scotland and therefore being missed by record linkage data, miscoding of cardiac events in hospitalisations data as 'unspecified chest pain', some general miscoding in the record linkage data and some record linkage errors.We conclude that routinely collected data could be used for recording cardiovascular endpoints in clinical trials and would give very similar results to rigorously collected clinical trial data, in countries with unified health systems such as Scotland. The endpoint types would need to be carefully thought through and an expert endpoint adjudication committee should be involved.

Background: over the next 20 years it is anticipated that there will be a significant increase in those aged 75 and over, and a consequent increase in cardiovascular disease, cancer and chronic illness.As this shift takes effect, there will be an increased need for treatment strategies that are of known benefit to this age group and a consequent rise in demand for clinical trials that are conducted specifically with the older population.Because factors that motivate older individuals to participate in clinical trials may differ from those that influence younger adults, it is important to evaluate the strategies used to encourage recruitment and retention and to determine how appropriate these are.Aim: evaluation of the reasons why subjects agree to participate in a controlled clinical trial of vascular disease prevention and the strategies used to improve compliance and protocol adherence.Setting: Scotland.Subjects: 2,520 Prospective Study of Pravastatin in the Elderly at Risk participants, aged 70-82 with either pre-existing vascular disease or at least one major vascular risk factor (hypertension, cigarette smoking, or diabetes mellitus).Design of study: two-stage iterative survey.Stage I was exploratory.Results: curiosity, or an interest in finding out more about the study, 'a desire to support research', and anticipated personal benefits, such as health screening, were the most important motivators for generating initial interest in the trial.Ongoing health monitoring was the most important recruitment and retention motivator (P = 0.001).Conclusions: curiosity, self interest and altruism may act as motivators at different points in the study time-line.However, fostering positive relationships between staff and recruits, and keeping recruits informed about the progress of the study are likely to maximise the retention of older subjects to long-term trials.

The flow-induced relaxation of a branch of the rabbit middle cerebral artery was examined to determine if an endothelial-independent as well as -dependent component occurs in pial as well as systemic small arteries and the possible role of products of the cyclooxygenase and the L-arginine nitric oxide synthase pathways. Intraluminal flow was achieved by the infusion of a tissue bath solution into isometrically mounted rabbit pial arteries in a resistance artery myograph through a small pipette. Intraluminal flow caused relaxation of the artery segment precontracted with 10 microM histamine. Treatment of endothelium-intact vessels with the nitric oxide synthase inhibitors NG-nitro-L-arginine (L-NNA) (100 microM) or NG-nitro-L-arginine methyl ester (L-NAME) (0.3 mM) significantly reduced the relaxation at flow rates of 5-30 microliters/min. This effect was partially reversed by 1 mM L-arginine. These inhibitors had no effect on the flow-induced relaxation of endothelium-denuded vessels. L-NNA did not influence the relaxation to 1 and 3 microM papaverine. Exposure to 10 microM aspirin, 10 microM indomethacin, or 300 nM tetrodotoxin had no effect on the flow-induced relaxation of either endothelium-intact or -denuded vessels (n = 6). Flow-induced relaxation was attenuated, but not abolished, by removal of the cerebrovascular endothelium. This reduction was not statistically significant. These results show that intraluminal flow caused relaxation of a branch of the rabbit middle cerebral artery, in part through a mechanism sensitive to inhibitors of nitric oxide synthase, most likely the generation of nitric oxide from the vascular endothelium. The major component of the relaxant response is independent of the endothelium and of nitric oxide synthesis through an L-NNA- or L-NAME-sensitive mechanism. The relaxation does not involve cyclooxygenase products nor neurogenic mediators. These results suggest that pial arteries, like those of the rabbit ear, exhibit a novel mechanism for the flow-induced relaxation of agonist-induced tone that is intrinsic to the tissues of the vascular wall subjacent to the endothelium.

Abstract : Proinflammatory cytokines, like interleukin‐6 (IL‐6) and tumor necrosis factor‐alpha (TNF‐α), are implicated in the development of atherosclerosis. The role of anti‐inflammatory cytokines, like IL‐10, is largely unknown. We investigated the association of four single nucleotide polymorphisms (SNPs) in the promoter region of the IL‐10 gene (4259AG, −1082GA, −592CA, and −2849GA), with coronary and cerebrovascular disease in participants of the PROspective Study of Pravastatin in the Elderly at Risk (PROSPER) trial. All associations were assessed with Cox proportional hazards models adjusted for sex, age, pravastatin use, and country. Haplotype analysis of the four SNPs showed a significant association between haplotype 4 (containing the −592A variant allele) and risk of coronary events ( P = 0.019). Moreover, analysis of separate SNPs found a significant association between −2849AA carriers with incident stroke (HR (95%CI) 1.50 (1.04–2.17), P value = 0.02). Our study suggests that not only proinflammatory processes contribute to atherosclerosis, but that also anti‐inflammatory cytokines may play an important role.

Adiponectin can suppress atherogenesis by inhibiting the adherence of monocytes, reducing their phagocytic activity, and suppressing the accumulation of modified lipoproteins in the vascular wall. Contradictory data have been reported about the effect of statins on adiponectin plasma levels. In this work, adiponectin plasma levels were measured in 102 statin-free subjects from the Spanish population of the Achieve Cholesterol Targets Fast with Atorvastatin Stratified Titration (ACTFAST) study, a 12-week, prospective, multi-centre, open-label trial which enrolled subjects with coronary heart disease, coronary heart disease-equivalent or a 10-year coronary heart disease risk > 20%. Subjects were assigned to atorvastatin (10–80 mg/day) based on low-density lipoprotein (LDL)-cholesterol concentration at screening. For comparison, age and gender-matched blood donors (N = 40) were used as controls. Control subjects did not present hypertension, hypercholesterolemia, diabetes, metabolic syndrome and history of cardiovascular diseases. Adiponectin levels were diminished in patients at high cardiovascular risk compared with control subjects [4166 (3661–4740) vs 5806 (4764–7075) ng/ml respectively; geometric mean (95% CI); P < 0.0001]. In the whole population, atorvastatin treatment increased adiponectin levels [9.7 (3.2–16.7);% Change (95% CI); P = 0.003]. This increment was in a dose-dependent manner; maximal effect observed with atorvastatin 80 mg/d [24.7 (5.7–47.1); P = 0.01]. Adiponectin concentrations were positively correlated with high-density lipoprotein-cholesterol both before and after atorvastatin treatment. No association was observed between adiponectin and LDL-cholesterol before and after atorvastatin treatment. In conclusion, atorvastatin increased adiponectin plasma levels in subjects at high cardiovascular risk, revealing a novel anti-inflammatory effect of this drug.

A multicompartmental model has been devised to explain apolipoprotein B (apoB) kinetics in very low density lipoprotein subfractions (VLDL1 Sf 60-400 and VLDL2 Sf 20-60), intermediate density (IDL Sf 12-20) and low density lipoproteins (LDL Sf 0-12). Normal and hyperlipemic subjects were given tracer doses of 131I-labeled VLDL1 and 125I-labeled VLDL2 and the metabolism of apoB in VLDL1, VLDL2, IDL, and LDL was followed over a period of 13 days. VLDL1 apoB and VLDL2 apoB clearance curves had an initial shoulder, a rapid decay, and a 'tail' of slowly metabolized lipoprotein. ApoB derived from VLDL1 appeared in IDL over 10-50 h and exhibited bi-exponential decay that was attributed to the presence of two metabolically distinct species. A further compartment was required to explain the observation that a substantial proportion of apoB from VLDL2 appeared and disappeared from the IDL density range faster than apoB derived from VLDL1 delipidation. Both of the more rapidly removed IDL species gave rise to LDL apoB that was also modeled as a heterogeneous entity with two plasma compartments. The final model, which has much in common with previous versions (M. Berman et al. 1978. J. Lipid Res. 19: 38-56), a multi-step delipidation pathway and slowly metabolized remnant compartments in VLDL, incorporates parallel delipidation routes in VLDL2, IDL, and LDL. These parallel pathways linked kinetic heterogeneity in VLDL with that in IDL and LDL.

Health Care Practitioners' attempts to implement secondary prevention targets for coronary heart disease (CHD) may be restricted by low rates of persistence with statin therapy. There is a need to understand why some patients, despite having established CHD and elevated cholesterol, do not comply with their prescribed statin regimen.To explore patients' perspectives on compliance with statin therapy.Primary care, West of Scotland.The research approach was qualitative. Thirty-three patients prescribed statin therapy and identified as having different patterns of compliance (poor moderate and good) were interviewed. The in-depth interviews were conducted on a one to one basis. Patients prescribed statin therapy for less than three months were excluded. Data were analysed thematically with the assistance of QSR Nudist.From analysis of the narrative data, two broad categories, i.e. 'Patient-health care provider communication' and 'Health beliefs' were identified. These categories encompassed six main themes: 'Initiation of therapy'; 'Subsequent feedback'; 'Sources of misconceptions'; 'Unconditional acceptance'; 'Conditional acceptance'; 'Deferment and Rejection'. Acceptance of and compliance with statin therapy appeared to be associated with the provision, interpretation and feedback of information during patient-practitioner consultations, and patients' beliefs about personal health status, cholesterol, and recommended cholesterol-lowering strategies.Patients' beliefs and understanding about cholesterol, and the role of cholesterol modifying strategies should be determined prior to the initiation of therapy and at appropriate intervals thereafter.

Important advances have been made in the past year in our understanding of the genetics of lipoprotein (a) [Lp(a)]. The apolipoprotein (a) [APO(a)] gene has been cloned and mice have been engineered to express apo(a) and Lp(a). These developments have provided the tools to answer many fundamental questions concerning the genetics, metabolism, and atherogenicity of Lp(a).

BACKGROUND: PROSPER was designed to investigate the benefits of treatment with pravastatin in elderly patients for whom a typical doctor might consider the prescription of statin therapy to be a realistic option. METHODS: The PROspective Study of Pravastatin in the Elderly at Risk (PROSPER) is a randomised, double blind, placebo-controlled trial to test the hypothesis that treatment with pravastatin (40 mg/day) will reduce the risk of coronary heart disease death, non-fatal myocardial infarction, and fatal or non-fatal stroke in elderly men and women with pre-existing vascular disease or with significant risk of developing this condition. RESULTS: In Scotland, Ireland, and the Netherlands, 23,770 individuals were screened, and 5,804 subjects (2,804 men and 3,000 women), aged 70 to 82 years (average 75 years) and with baseline cholesterol 4.0-9.0 mmol/l, were randomised. Randomised subjects had similar distributions with respect to age, blood pressure, and body mass index when compared to the entire group of screenees, but had a higher prevalence of smoking, diabetes, hypertension, and a history of vascular disease. The average total cholesterol level at baseline was 5.4 mmol/l (men) and 6.0 mmol/l (women). CONCLUSIONS: Compared with previous prevention trials of cholesterol-lowering drugs, the PROSPER cohort is significantly older and for the first time includes a majority of women. The study, having achieved its initial goal of recruiting more than 5,500 elderly high-risk men and women, aims to complete all final subject follow-up visits in the first half of 2002 with the main results being available in the fourth quarter of 2002.

Using analyses of the large cohort (n=5732) from the prospective study of pravastatin in the elderly at risk (PROSPER), we tested the hypothesis that Lp(a) concentration is an independent predictor of major vascular events and cognitive impairment in the elderly. Baseline Lp(a) levels were measured on fresh samples from 5732 subjects aged 70-82, who were followed for 3.2 years on average. Lp(a) levels were not significantly different across the age range in PROSPER, but were significantly higher in women (geometric mean 14.8 versus 12.4 mg/dl, P<0.0001). Those with a history of vascular disease had significantly higher Lp(a) levels, which remained after adjustment (P<0.0001). There was no statistically significant association between baseline Lp(a) and the risk of the primary endpoint (CHD death, non-fatal MI and fatal or non-fatal stroke) (hazard ratio 1.05, 95% CI 1.00-1.11, P=0.077), but after adjustment for baseline risk factors this did achieve statistical significance (1.06, 1.005-1.12, P=0.032). Finally, there was no statistically or clinically significant association between any adjusted baseline or dynamic cognition variables and Lp(a), and nor was there any significant association between Lp(a) and indices of disability throughout the study. This is the first study of the association between Lp(a) and a range of cardiovascular endpoints including cognitive and disability indices in the elderly. The main finding is that Lp(a) level, while influenced by a number of baseline characteristics, is not a significant predictor of cognitive function or levels of disability, but is a predictor of combined cardiovascular events over an average 3.2 year follow-up.

Objective Lipoprotein-associated phospholipase A2 (Lp-PLA2) is an inflammatory biomarker that circulates mainly bound to LDL. We evaluated the association of Lp-PLA2 with vascular events in the elderly where the importance of LDL is diminished as a risk factor for coronary disease. Methods Mass and activity of Lp-PLA2 were related to risk over 3.2 years for vascular events (definite or suspected death from CHD, non-fatal MI, fatal or non-fatal stroke) in the 2804 men and 3000 women age 70–82 years in the Prospective Study of Pravastatin in the Elderly (PROSPER). Results Lp-PLA2 showed a moderate, positive association with risk of a vascular event with hazard ratios of 1.25 (confidence interval (CI) 1.02–1.54) for mass and 1.39 (CI 1.14–1.70) for activity for top versus bottom quartile. Risk associations were attenuated when classical risk factors, lipids and inflammatory markers – C-reactive protein and white cell count – were included in the models. Lp-PLA2 was unrelated to stroke risk. Inclusion of all three inflammatory markers in multivariate models negated the association of HDL cholesterol with risk (hazard ratio 0.98; CI 0.88–1.10) and increased prediction of coronary events; the C statistic rose from 63.2% to 64.4% (P < 0.001). Conclusion In elderly people Lp-PLA2, alongside other inflammatory indices, is a potential biomarker for vascular events, particularly CHD.

The metabolism of apolipoprotein B-100 was studied in three patients with familial hyperchylomicronemia (type I hyperlipoproteinemia) using a very low density lipoprotein (VLDL) dual-tracer technique. Radioiodinated VLDL1 (Sf 60-400) and VLDL2 (Sf 20-60) were injected and their catabolism and rate of the transfer of apoB into VLDL2, intermediate density lipoprotein (IDL) (Sf 12-20), and low density lipoprotein (LDL) (Sf 0-12) were compared in patients and in five normolipidemic controls. The rates of delipidation of large triglyceride-rich VLDL1 to VLDL2 (0.26-0.54 pools/day vs. 2.5-5.2 pools/day in controls) and VLDL1 direct catabolism (0.33-0.92 pools/day vs. 4.2-14.7 pools/day in controls) were found to be significantly reduced in type I patients resulting in a tenfold increase of VLDL1 pool size. ApoB synthesis into this density interval was, however, normal as was that into smaller VLDL2. the circulating apoB mass in VLDL2 was not increased. In fact, apart from a modest decrease in the rate of VLDL2 delipidation to IDL and LDL, the behavior of apoB in this density interval was similar in hyperchylomicronemic and normal subjects. Likewise, the transfer of apoB through the IDL and LDL density ranges was not significantly different from normal. Pool sizes of these fractions, however, were reduced, the latter significantly (354-491 mg vs. 1,160-2,505 mg in controls) due to increased direct catabolism in hyperchylomicronemic patients. The results of this study indicate that lipoprotein lipase deficiency primarily affects VLDL1 metabolism, both its delipidation and direct removal from plasma. Lipolysis further down the delipidation cascade is not dependent on this enzyme. Hypercatabolism rather than a failure of synthesis of IDL and LDL was responsible for the decreased pools for both lipoproteins.

Atherosclerotic diseases are responsible for the majority of deaths in the elderly, and they can also increase the risk of disability. Statins are first-line therapies for lowering lipid levels and have been shown to reduce the risk of cardiovascular events in large-scale clinical trials. There is a growing body of evidence that statins are as efficacious at lowering lipid levels and reducing the risk of coronary heart disease (CHD) in elderly patients as in younger individuals. Furthermore, as this population is at a greater absolute risk of CHD, they may receive greater absolute benefits from treatment. However, despite these benefits, many elderly individuals at risk of CHD and stroke are not receiving adequate lipid-lowering therapy, which could help them to maintain their health and independence. Further, prospective randomised trials are required to guide physicians in the treatment of elderly patients at risk of atherosclerotic disease, thereby resolving the current undertreatment.

Aims To investigate whether selecting the starting dose of atorvastatin according to baseline and target (<2.6 mmol/L) LDL-cholesterol (LDL-C) values would allow high-risk subjects to achieve target LDL-C concentration within 12 weeks, with the initial dose or a single uptitration. Methods and results Twelve-week, prospective, open-label trial that enrolled 2117 high-risk subjects (statin-free [SF] or statin-treated [ST]). Subjects with LDL-C >2.6 mmol/L (100 mg/dL) but ≤5.7 mmol/L (220 mg/dL) were assigned a starting dose of atorvastatin (10, 20, 40 or 80 mg/day) based on LDL-C and status of statin use at baseline, with a single uptitration at 6 weeks, if required. There was no washout for ST subjects. At study end, 80% of SF (82%, 82%, 83% and 72% with 10, 20, 40 and 80 mg, respectively) and 59% of ST (60%, 61% and 51% with 20, 40 and 80 mg, respectively) subjects reached LDL-C target. In the ST group, an additional 21–41% reduction in LDL-C was observed over the statin used at baseline. Atorvastatin was well tolerated. Conclusion This study confirms that individualizing the starting dose of atorvastatin according to baseline and target LDL-C values (i.e. the required LDL-C reduction), allows a large majority of high-risk subjects to achieve target safely, within 12 weeks, with the initial dose or with a single titration.

Venous thromboembolic events (VTE), including deep venous thrombosis and pulmonary embolism, are common in older age. It has been suggested that statins might reduce the risk of VTE however positive results from studies of middle aged subjects may not be generalisable to elderly people. We aimed to determine the effect of pravastatin on incident VTE in older people; we also studied the impact of clinical and plasma risk variables.This study was an analysis of incident VTE using data from the Prospective Study of Pravastatin in the Elderly at Risk (PROSPER), a randomized, double-blind, placebo-controlled trial of pravastatin in men and women aged 70-82. Mean follow-up was 3.2 years. Risk for VTE was examined in non-warfarin treated pravastatin (n = 2834) and placebo (n = 2865) patients using a Cox's proportional hazard model, and the impact of other risk factors assessed in a multivariate forward stepwise regression analysis. Baseline clinical characteristics, blood biochemistry and hematology variables, plasma levels of lipids and lipoproteins, and plasma markers of inflammation and adiposity were compared. Plasma markers of thrombosis and hemostasis were assessed in a nested case (n = 48) control (n = 93) study where the cohort was those participants, not on warfarin, for whom data were available.There were 28 definite cases (1.0%) of incident VTE in the pravastatin group recipients and 20 cases (0.70%) in placebo recipients. Pravastatin did not reduce VTE in PROSPER compared to placebo [unadjusted hazard ratio (95% confidence interval) 1.42 (0.80, 2.52) p = 0.23]. Higher body mass index (BMI) [1.09 (1.02, 1.15) p = 0.0075], country [Scotland vs Netherlands 4.26 (1.00, 18.21) p = 0.050 and Ireland vs Netherlands 6.16 (1.46, 26.00) p = 0.013], lower systolic blood pressure [1.35 (1.03, 1.75) p = 0.027] and lower baseline Mini Mental State Examination (MMSE) score [1.19 (1.01, 1.41) p = 0.034] were associated with an increased risk of VTE, however only BMI, country and systolic blood pressure remained significant on multivariate analysis. In a nested case control study of definite VTE, plasma Factor VIII levels were associated with VTE [1.52 (1.01, 2.28), p = 0.044]. However no other measure of thrombosis and haemostasis was associated with increased risk of VTE.Pravastatin does not prevent VTE in elderly people at risk of vascular disease. Blood markers of haemostasis and inflammation are not strongly predictive of VTE in older age however BMI, country and lower systolic blood pressure are independently associated with VTE risk.Not applicable when study undertaken.

A new method for the rapid genotyping of low density lipoprotein receptor knockout mice is described. This method using a polymerase chain reaction (PCR) technique may be performed on small tissue biopsies, and represents a significant advantage in time over conventional genotyping by Southern blot analyses.

The aim of this study was to determine the effects of atorvastatin in patients of South Asian versus European origin who participated in the Achieve Cholesterol Targets Fast with Atorvastatin Stratified Titration (ACTFAST) study. ACTFAST was a 12‐week prospective, open‐label study in patients at high risk for atherosclerosis (European origin, n = 1978; South Asian origin, n = 64). Compared with patients of European origin, patients of South Asian origin were younger, were less likely to smoke, and had lower body mass index, systolic blood pressure, low‐density lipoprotein cholesterol (LDL‐C) and triglycerides. Because significant differences were observed in baseline characteristics between patient groups, case control propensity scores were used. In the unmatched analysis, South Asians had greater LDL‐C response to atorvastatin than patients of European origin. However, after propensity matching, atorvastatin lowered LDL‐C and high‐sensitivity C‐reactive protein (hs‐CRP) to a similar degree in both groups, with no differences in safety profile. The authors observed no correlation between change in hs‐CRP and LDL‐C concentrations in either population. In conclusion, atorvastatin lowered both LDL‐C and hs‐CRP to a similar degree in patients of South Asian or European origin, suggesting usual starting doses of atorvastatin (with appropriate monitoring), rather than lower starting doses as has been advocated by some, may be used in patients of South Asian origin.

Antagonism of the chemokine receptor CXCR2 has been proposed as a strategy for the treatment of inflammatory diseases such as arthritis, chronic obstructive pulmonary disease and asthma. Earlier series of bicyclic CXCR2 antagonists discovered at AstraZeneca were shown to have low solubility and poor oral bioavailability. In this Letter we describe the design, synthesis and characterisation of a new series of monocyclic CXCR2 antagonists with improved solubility and good pharmacokinetic profiles.

Plasma lipoprotein(a) [Lp(a)] levels are believed to be controlled predominantly by the apolipoprotein(a) [APO(a)] gene, which encodes the apo(a) glycoprotein, a key constituent of the Lp(a) particle. Previously, it has been accepted that the plasma Lp(a) level is inversely proportional to apo(a) length. To examine this relationship in greater detail, 1500 unrelated, homogeneous (sex, race, age, plasma lipids) subjects were studied, from which 769 were identified with a single-expressing APO(a) allele. A bimodal frequency distribution of apo(a) isoforms was observed. As expected, there was a general inverse relationship between apo(a) isoform size and Lp(a) level. However, when groups with equivalent single-expressing apo(a) isoforms were studied, it was clear that although smaller isoforms were associated on average with higher levels, they were also associated with the greatest variability in level. After logarithmic transformation of Lp(a) data, the overall contribution of the apo(a) length polymorphism was calculated to be 38%. However, in subjects with apo(a) isoforms of </=20 kringle-4 (K-4) repeats, only 9% of the variability in Lp(a) concentration is explicable on the basis of the apo(a) length polymorphism. In those with apo(a) isoforms of >20 K-4 repeats, the corresponding contribution is 10%. We conclude that the contribution of the apo(a) isoform size to the control of plasma Lp(a) level is considerably lower than previously calculated, because the variability in plasma Lp(a) concentration is not uniform across the apo(a) size spectrum.

In the sheep cerebral vasculature 5-hydroxytryptamine (5-HT) caused a contraction of which ketanserin was found to be an effective antagonist (Basilar artery, pA2 = 9.33 ± 0.16; middle cerebral artery, pA2 = 9.19 ± 0.16; pial artery, pA2 = 9.47 ± 0.12). Sumatriptan (GR 43175), a selective 5-HT1-like receptor agonist, was also found to have a small contractile effect on the sheep cerebral vasculature (Basilar artery, pD2 = 6.26 ± 0.11; middle cerebral artery, pD2 = 6.25 ± 0.10; pial artery, pD2 = 6.13 ± 0.15). The contractile effect of sumatriptan was not antagonised by either ketanserin (1 μM) or MDL 72222 (1 μM). 5-HT therefore appears to cause contraction by stimulation of a mixed receptor population of 5-HT1-like and 5-HT2 receptors. In the sheep middle cerebral artery the addition of haemolysate was found to cause a contractile response and also to augment the contractile effects of both noradrenaline and 5-HT but only in the presence of a functional endothelium. However, 5-HT was never found to relax precontracted rings of the middle cerebral artery in either the presence or absence of a functional endothelium. These results indicate a basal release of EDRF in cerebral arteries that attenuates the effects of various constrictor agents.

Background The gold standard clinical trial design is the double-blind, randomized, controlled trial. No standard practice exists for the “unblinding” of trial participants and no legal obligation is placed on investigators to inform participants of their treatment allocation or study results at the end of a trial. Here we document our experiences of unblinding the 2520 Scottish participants in the Prospective Study of Pravastatin in the Elderly at Risk (PROSPER). Methods The objectives of the PROSPER unblinding process were to provide all study participants with their study medication status and on-trial cholesterol levels and to respect the rights of participants not to be unblinded. It was considered imperative by the study executive that the blind was maintained until the presentation and publication of the results. Staff therefore remained “blinded” throughout the unblinding process. Inappropriate contact with the PROSPER participants was avoided by confirming their current vital status and health status. Results To coincide with the presentation of the PROSPER results, all participants, for whom it was deemed appropriate, were sent a summary of the results and were offered the opportunity to be advised of their treatment allocation and on-trial lipid profiles. The majority of participants opted for telephone unblinding. All primary care physicians who had patients randomised to the study were also sent a summary of the study results and sealed documents detailing the treatment allocation and lipid profiles for each patient. Relocated patients were traced and the information forwarded to their new primary care physicians. Conclusion The dissemination of study results and treatment allocation to study participants is an integral part of the research process and should be included in the design of any clinical trial.

The CD40/CD40 ligand plays a role in the inflammatory and prothrombotic processes in atherosclerosis. We analyzed whether short-term treatment with atorvastatin affects soluble CD40 ligand (sCD40L) plasma levels in subjects at high cardiovascular risk. sCD40L plasma concentrations were measured in 852 subjects from the Atorvastatin on Inflammatory Markers (AIM) Study, a 12-week prospective multicenter, open-label trial which enrolled statin-free subjects with coronary heart disease (CHD), CHD-equivalent (diabetes, peripheral vascular disease, or cerebrovascular disease), or a 10-year CHD risk >20%. Subjects were assigned to atorvastatin (10-80 mg/day) based on LDL-C at screening. Overall, sCD40L levels were not different in patients at high cardiovascular risk compared with healthy subjects. When sCD40L levels were divided in quartiles, patients in the highest quartile (N=213) had higher sCD40L concentrations than age- and gender-matched healthy subjects (N=29) (P<0.0001). Interestingly, all doses of atorvastatin significantly diminished sCD40L levels in subjects at the highest quartile. Furthermore, atorvastatin treatment decreased sCD40L more markedly in subjects with metabolic syndrome compared with those without metabolic syndrome. In conclusion, atorvastatin diminishes sCD40L plasma levels, more markedly so in subjects with metabolic syndrome. Our results indicate that short-term treatment with atorvastatin exhibits anti-inflammatory and antithrombotic effects in subjects at high cardiovascular risk.

Henry K. Beecher and Maurice H. Pappworth were the 2 most prominent medical whistleblowers in research ethics of the 20th century. Independently, both wrote highly controversial and ultimately influential articles and books. Although their work is now well-known in clinical research circles, their collaboration is not. Pappworth's article “Human Guinea Pigs: A Warning” was published in 1962; in it, he discussed a series of published studies that he considered unethical. Beecher read it and wrote to Pappworth seeking help. The current article reconstructs, from Beecher and Pappworth's correspondence in 1965–1966, an important juncture in the genesis of modern clinical research ethics. Although they shared much in common, they differed radically in the strategies they adopted: Beecher chose to conceal the identities of individuals, whereas Pappworth believed that only by naming and shaming could any exposé act as a deterrent. Their correspondence reveals how the 2 men shared their ideas and their material and provided each other with much-needed support. It also tracks the development of Beecher's shift from a position initially indistinguishable from Pappworth's toward the one he adopted when his seminal article of 1966 was published.

1. The cardiovascular actions of cholecystokinin and related peptides were investigated in the pithed rat. The receptors and the mechanisms involved in these experiments were characterized. 2. Sulphated cholecystokinin octapeptide (sCCK-8, 0.1-100 nmol kg-1, i.v.) elicited a dose-dependent bradycardia and increase in mean arterial blood pressure. Neither gastrin-17 nor pentagastrin had any effect at concentrations up to 100 nmol kg-1. 3. Both the pressor response and bradycardia elicited by sCCK-8 were reduced by the selective CCKA receptor antagonists, devazepide (0.5-50 nmol kg-1) and lorglumide (1-7 mumol kg-1). The selective CCKB receptor antagonists, CI-988 (1 mumol kg-1) and L-365,260 (15 mumol kg-1) did not inhibit the effects of sCCK-8. 4. The pressor response induced with sCCK-8 was reduced by treatment with either phentolamine (3 mumol kg-1) or guanethidine (2 mumol kg-1) and was unaffected by treatment with propranolol, atropine or hexamethonium. The pressor response also persisted following bilateral adrenalectomy. 5. The bradycardia induced with sCCK-8 was unaffected by treatment with phentolamine, propranolol, guanethidine, atropine, hexamethonium or bilateral adrenalectomy. 6. The tetrapeptide of cholecystokinin (CCK-4) elicited a dose-dependent pressor response but did not induce bradycardia. The pressor response was unaffected by devazepide (50 nmol kg-1), L-365260 (15 mumol kg-1) or phentolamine (3 mumol kg-1). 7. In the pithed rat, sCCK-8 acted via CCKA receptors to increase arterial blood pressure indirectly, at least in part, through activation of alpha-adrenoceptors. The observed bradycardia was also mediated byCCKA receptors but possibly through a direct action on the heart.

The effects of colestipol therapy alone (20 g/d) or combined with simvastatin (20 mg/d) were examined in a group of eight male patients with primary moderate hypercholesterolemia (total cholesterol > or = 6.5 mmol/L [> or = 250 mg/dL]) who had undergone coronary artery bypass grafting more than 3 months previously. Colestipol therapy decreased total cholesterol by 14% (P < .001) and LDL cholesterol (LDL-C) by 23% (P < .001), while dual therapy decreased total cholesterol by 38% and LDL-C by 52% (both P < .001 versus baseline). No significant changes were observed in plasma triglyceride, VLDL cholesterol, or HDL cholesterol levels. VLDL subfraction turnovers were conducted at baseline and again on each regimen. ApoB kinetic parameters derived from a multicompartmental model suggested that colestipol therapy resulted in an expansion of the total VLDL apoB pool (36%, P < .05) that was largely due to a fall in the clearance rate of VLDL1 apoB (49%), while the LDL apoB pool decreased 23% as a result of diminished direct LDL input. The model used also revealed that addition of simvastatin to the resin therapy caused increases in the fractional transfer rates of VLDL2 to IDL and IDL to LDL together with a 37% increment in the LDL apoB fractional catabolic rate. Compared with baseline, combined therapy generated falls in both IDL (35%, P = .01) and LDL (37%, P < .04) apoB pools due to enhanced clearance of IDL (214%, P < .03) and reduced total input of LDL (39%, P < .003).

Elevated C-reactive protein (CRP) concentration is a risk factor for cardiovascular events that may add prognostic information. Statin treatment is associated with significant reductions in CRP concentrations, which appear to be unrelated to the magnitude of LDL-cholesterol reduction. We investigated the effect of atorvastatin, across its dose range, on high sensitivity (hs)CRP in subjects at high cardiovascular risk.ACTFAST was a 12 week, prospective, multicenter, open-label trial in which high-risk subjects were assigned a starting dose of atorvastatin (10, 20, 40 or 80 mg/d) based on LDL-C and status of statin use at screening (1345 statin-free [SF] and 772 previously statin-treated [ST]).At baseline, ST subjects had significantly lower hsCRP levels than SF subjects (ST group 2.31, 95% CI 2.15, 2.48 mg/L vs. SF group 3.16, 95% CI 2.98, 3.34 mg/L, p<0.05). In the SF group, atorvastatin 10 to 80 mg significantly (p<0.01) reduced hsCRP levels in a dose dependent-manner. In ST group, additional hsCRP reductions were observed over the statin used at baseline, which were not dose-dependent. Atorvastatin significantly decreased hsCRP concentrations in subjects with or without diabetes or the metabolic syndrome.Atorvastatin treatment at different doses, particularly 80 mg, significantly reduced hsCRP serum concentrations. This reduction was observed in both SF and ST groups and was independent of the presence of metabolic syndrome and/or diabetes. The beneficial effect of atorvastatin was evident at 6 weeks, supporting the practice of early introduction of higher doses of atorvastatin in high-risk patients.

Fibric acid derivatives are an important and powerful group of lipid-lowering drugs which have found a key place in our lipid-regulating formulary. We review recent research on aspects of the mechanism of fibrate action, novel applications of established fibrates, development of new drugs and further evaluation of existing compounds.

Preface. Coronary heart disease: epidemiology, pathology and diagnosis. Risk factor assessment. Lipids and lipid-lowering drugs. Hypertension and anti-hypertensive therapy. Lifestyle management: smoking. Lifestyle management: diet. Lifestyle management: exercise. Lifestyle management: facilitating behavioural change. Medical management of coronary heart disease. Cardiac rehabilitation.Women and coronary heart disease. Type 2 diabetes and coronary heart disease.

Physicians often fail to achieve recommended low-density lipoprotein (LDL) cholesterol goals for their patients using lipid-lowering therapies in the primary care setting. A variety of factors may contribute to this failure, including inadequate effectiveness of lipid-lowering drugs in reducing LDL cholesterol at commonly used doses. In the Lipid Assessment Treatment Project (L-TAP), for example, the success rates for lipid-regulating therapies and treatments according to National Cholesterol Education Program (NCEP) Adult Treatment Panel II (ATP II) LDL cholesterol goals were 43% for bile acid sequestrants, 39% for niacin, 32% for gemfibrozil, 28% for psyllium fiber, 40% for statins, and 40% for combination therapy. Rosuvastatin is a new statin that has been shown to achieve significantly greater reductions in LDL cholesterol compared with pravastatin, simvastatin, and atorvastatin in primary hypercholesterolemia and enabled greater proportions of patients to achieve LDL cholesterol goals. Similarly, rosuvastatin proved superior to atorvastatin in lowering LDL cholesterol in patients with familial hypercholesterolemia, with more patients achieving LDL cholesterol goals. Data from these trials suggest that rosuvastatin is as safe and well tolerated as other statins. The availability of lipid-lowering drugs with greater LDL cholesterol-lowering effects could simplify the approach to coronary heart disease risk reduction in primary care.

Objectives: ACTFAST-2 was designed to match the starting dose of a statin to the baseline low density lipoprotein-cholesterol (LDL-C) value, to allow high-risk European subjects to achieve LDL-C targets within 12 weeks with the initial dose or one up-titration.Research design and methods: This was a 12-week, prospective, open-label trial that enrolled 610 high-risk subjects from 8 European countries. Subjects with LDL-C > 2.6 mmol/L (> 100 mg/dL),but ≤ 5.7 mmol/L (≤ 220 mg/dL) were assigned a starting dose of atorvastatin (10, 20, 40, 80 mg/day) according to LDL-C level and status of statin use at baseline (either statin-free or statin-treated), with a single up-titration at 6 weeks if needed.Results: At 12 weeks, 68.0% of subjects overall, including 73.5% of statin-free and 60.5% of statin-treated subjects, achieved LDL-C target (< 2.6 mmol/L (< 100 mg/dL). The total cholesterol/high density lipoprotein-cholesterol (TC/HDL-C) ratio target was achieved by 75.2% of subjects overall, including 78.1% of statin-free and 71.2% of statin-treated subjects. In the statin-free group, LDL-C decreased by a mean of 42%. In the statin-treated group, atorvastatin led to an additional 31% reduction in LDL-C over the statin used at baseline. Mean decreases in TC/HDL-C ratio were 30% and 20% in the statin-free and statin-treated groups, respectively. The incidence of AST/ALT greater than 3 times of upper limit of normal range in all patients was 0.8% and no rhabdomyolysis was reported.Conclusion: This study confirms that use of a flexible starting dose of atorvastatin allows the large majority of high-risk subjects to achieve their LDL-C target safely within 12 weeks with an initial dose or just a single up-titration.

Maurice H Pappworth (1910–1994) was an English physician who had developed a deep concern for the ethics of human research. In 1966, while researching his book – Human Guinea Pigs, which would be published the following year1 – he wrote to Ivy regarding experimentation on prisoners. Andrew C Ivy (1893–1978) was an eminent physician and physiologist and according to Moreno2 ‘by the end of the war he was probably…the most famous doctor in the country [US]…the prototype of today’s media medical expert’. Perhaps because of this status, and the fact that he had been actively involved in research involving prison inmates, he had been invited to serve as the American Medical Association’s (AMA) adviser to the Nuremberg prosecutors (Figure 1). Ivy responded to Pappworth’s enquiry3 on 6 April 1966 and began by stating: I was considering the subject of your letter before I testified at Nuremberg and gave the Judges my Version of the Code thus indicating from the outset his involvement. In his letter, he told Pappworth that he had been interested in research ethics since 1917 and that he had always tried to apply the ‘Golden Rule’ as a guiding principle in all his own research, that is that you should only do to others that which you would allow them to do to you. He indicated finally his belief that the natural extension of this principle was that you should always take your ‘own “Medicine” first’.3 This allusion to self-experimentation as a preliminary and necessary step in research becomes relevant when we consider Ivy’s appendix to the letter. Open in a separate window Figure 1. Andrew C Ivy being sworn in at the Nuremberg Doctors’ Trial, June 1947. Source: Photo courtesy of the National Archives RG 238 OMT-1-W-60.

Statin therapy has been conclusively shown to offer patients clinical benefit, virtually irrespective of their baseline risk status. However, the absolute risk reductions observed in different clinical trials, which have recruited patients across a spectrum of lipid levels and vascular disease states, show that baseline global risk determines the absolute benefit gained and in turn will specify the number of patients needed to be treated in order to realize this benefit. Global risk assessment is therefore central to the clinically meaningful use of statin therapy, and a strong case is now argued in the literature for a high-risk primary prevention strategy that goes hand in hand with standard secondary prevention. The routine use of Framingham-based risk assessment tools is advocated because these are the most widely evaluated and have been repeatedly shown to predict the risk of coronary heart disease accurately in western populations. The risk threshold in primary prevention that should determine pharmacological intervention is the subject of controversy. The currently used annual risk figure of 3% would clearly capture all very high-risk individuals but would also deny treatment to many individuals who will subsequently die from their first coronary event. Although a 1.5% annual risk threshold is economically untenable in the present UK health system, a level of 2% is, we believe, both achievable and affordable.

The clinical utility of a new assay for plasma lipoprotein(a)-cholesterol (Lp(a)-C) was assessed in parallel with our routine Lp(a) mass measurements in a nested-case control study of subjects within the placebo arm of the West of Scotland Coronary Prevention Study (WOSCOPS). A total of 238 control patients and 108 patients who had suffered a serious vascular event during the course of the WOSCOPS were examined. Lp(a) mass was assessed within 2 years of sampling by an ELISA method on baseline EDTA plasma samples which had been stored at -70 degrees C. Subsequently, the Lp(a) mass was re-measured by an immunoturbidimetric assay approximately 8 years after sampling. On the same stored aliquot the Lp(a)-C was measured. These analyses allowed us to assess whether the Lp(a)-C assay could provide any additional information over and above that which would be obtained from our Lp(a) mass assays. In addition the apo(a) isoform sizes of these subjects were measured using a high resolution immunoblotting system. The Lp(a)-C and Lp(a) mass measurements provided exactly the same information in the study, as they were equally non-discriminatory between cases and controls. The only difference between the two patient groups was the percentage of 'null' apo(a) alleles (control: 25.6% versus cases: 19.4%). We conclude that these results reinforce the concordance of the two assay systems and confirm that the Lp(a)-C assay provides no added information over and above that gained from traditional Lp(a) mass assays, which may be faster and less expensive.

The accuracy with which patients recall their cardiac symptoms prior to aorta-coronary artery bypass grafting is assessed approximately one year after surgery together with patient-related factors potentially influencing accuracy of recall.This is a novel investigation of patient's rating of preoperative symptom severity before and approximately one year following aorta-coronary artery bypass grafting.Patients undergoing aorta-coronary artery bypass grafting (n = 208) were recruited preoperatively and 177 of these were successfully followed up at 16.4 (SD 2.1) months after surgery and asked to describe current and recalled preoperative symptoms using a 15-point numerical scale.Accuracy of recall was measured and correlated (Pearson's correlation) with current and past symptoms, health-related quality of life and coronary artery disease risk factors. Hypothesis tests used Student's t-test and the chi-squared test.Respective angina and breathlessness scores were recalled accurately by 16.9% and 14.1% while 59% and 58% were inaccurate by more than one point. Although the mean preoperative and recalled scores for severity of both angina and breathlessness and were not statistically different, patients who recalled most accurately their preoperative scores had, on average, significantly higher preoperative scores than those with less accurate recall. Patients whose angina and breathlessness symptoms were relieved by operation had significantly better accuracy of recall than patients with greater levels of symptoms postoperatively.Patient's rating of preoperative symptom severity before and one year following aorta-coronary artery bypass grafting was completely accurate in approximately one sixth of patients with similar proportions of the remaining patients overestimating and underestimating symptoms. The extent to which angina and breathlessness was relieved by operation was a significant factor in improving accuracy of recall.Factors associated with accuracy of recall of symptoms provide useful insights for clinicians when interpreting patients' views of the effectiveness of aorta-coronary artery bypass grafting for the relief of symptoms associated with coronary heart disease.

The UK Clinical Trial Regulations and Good Clinical Practice guidelines specify that the study sponsor must ensure clinical trial data are accurately reported, recorded and verified to ensure patient safety and scientific integrity. The methods that are utilised to assess data quality and the results of any reviews undertaken are rarely reported in the literature. We have recently undertaken a quality review of trial data submitted to a Clinical Endpoint Committee for adjudication. The purpose of the review was to identify areas that could be improved for future clinical trials. The results are reported in this paper.Throughout the course of the study, all data queries were logged. Following study close out, queries were coded and categorised. A descriptive and comparative analysis was conducted to determine the frequency of occurrence for each category by country of origin.From 1595 endpoint packages reviewed, 782 queries were generated. No source data queries were generated for countries with ≤ 25 recruited subjects, but both low recruiting and high recruiting countries had a high number of queries relating to subject identifiers.The implementation of some simple measures could help improve data quality and lead to significant savings.

The responsiveness to noradrenaline was characterized in cerebral arteries from the sheep, since this species was large enough to permit a comparison of arteries from different parts of the cerebral vasculature. Noradrenaline caused contraction of the basilar artery, middle cerebral artery and small pial arteries by stimulation of α 1 ‐adrenoceptors. The maximum contraction to noradrenaline was small in the basilar artery (28% of the 5‐hydroxytryptamine (5‐HT) maximum) but larger in the middle cerebral artery (78% of the 5‐HT maximum) and pial artery (92% of the 5‐HT maximum) of the sheep. Cocaine (10 μ m ) potentiated noradrenaline‐induced contractions in the sheep middle cerebral artery but not in the sheep basilar artery. The noradrenaline contraction, relative to the 5‐HT contraction, was not affected by removal of the endothelium in either the sheep basilar or middle cerebral artery. The results showed a variation within the sheep cerebral vasculature in the response to noradrenaline which cannot be explained by regional differences in α‐adrenoceptor subtypes, noradrenaline uptake mechanisms or endothelial function.

Inflammation plays a prominent role in the development of atherosclerosis, which is the most important risk factor for vascular events. Lymphotoxin-alpha (LTA) is a pro-inflammatory cytokine and is found to be expressed in atherosclerotic lesions. We investigated the association between the C804A polymorphism within the LTA gene and coronary and cerebrovascular events in 5804 participants of the PROspective Study of Pravastatin in the Elderly at Risk (PROSPER). The primary endpoint was the combined endpoint of death from coronary heart disease, non-fatal myocardial infarction, and clinical stroke. Secondary endpoints were the coronary and cerebrovascular components separately. All associations were assessed with a Cox-proportional hazards model adjusted for sex, age, pravastatin use, and country. Our overall analysis showed a significant association between the C804A polymorphism and the primary endpoint (p = 0.03). After stratification for gender, this association was found only in males. Furthermore, we found that the association between the C804A polymorphism and the primary endpoint was mainly attributable to clinical strokes (p = 0.02). The C804A polymorphism in the LTA gene associates with clinical stroke, especially in men. But further research is warranted to confirm our results.

An elevated plasma lipoprotein(a) (Lp(a)) concentration is an independent risk factor for coronary heart disease (CHD). Plasma Lp(a) levels are believed to be predominantly controlled by the APO(a) gene, which encodes the apo(a) glycoprotein moiety of the Lp(a) particle. However, other parameters in the lipoprotein profile as well as co-existing disease states or personal traits have been proposed as co-varieties. In order to examine these potential controlling factors in greater detail than previously possible, 1760 unrelated Caucasian subjects were studied, from which were identified 907 with a single expressing APO(a) allele. This strategy was followed to obviate the difficulty in dealing with the co-expression of different apo(a) isoforms and the resulting compound plasma Lp(a) level. After cube-root transformation of the plasma Lp(a) levels to normalise their distribution, a series of correlates were computed. There was no good correlation between Lp(a) concentration and any other measured lipid or lipoprotein in the lipid profile or with any other variable examined, with the important exception of the length of the expressed apo(a) isoform (r=−0.491, P=0.0001). We conclude that in this population the plasma Lp(a) concentration is not predicted by the plasma lipid profile, alcohol intake, or smoking status but is predicted, albeit incompletely, by the length polymorphism of the APO(a) gene.

Abstract. Two independent studies were designed to investigate the separate and combined effects of acipimox and cholestyramine on plasma low density lipoprotein subfractions. In the first study, normolipidaemic subjects were given cholestyramine (16 g day ‐1 , 4 weeks), followed, after an 8‐week wash‐out period, by acipimox (750mg day ‐1 , 4 weeks). In the second study, moderately hypercholesterolaemic subjects were prescribed acipimox (1250mg day ‐1 , 10 weeks), followed by acipimox in combination with low dose cholestyramine (12g day ‐1 ) for a further 10 weeks. In the normal subjects, cholestyramine decreased total LDL mass (density (d)= 1.019–1.063g ml ‐1 ) by selectively reducing the largest, least dense LDL‐I(d 1.025–1.034 gml ‐1 , P &lt; 0.05) and LDL‐II (d 1.034–1.044 g ml ‐1 , P &lt;0.005) subfractions. The small, dense LDL‐III (d 1.044–1060 g ml ‐1 ) showed a variable response to the resin. In the same subjects acipimox produced no overall change in total LDL mass but showed a tendency to redistribute LDL towards LDL‐I (+10%) and LDL‐II (+10%) in a manner related to the changes in plasma triglyceride (TG) (TG vs. LDL‐III r = 0.75, P &lt;0.05). In the hypercholesterolaemic subjects acipimox induced a substantial redistribution of LDL subfractions (LDL‐1 + 84% P &lt;0.05; LDL‐III‐50%) without affecting total LDL mass. The addition of cholestyramine produced a significant decrease in total LDL mass which was again confined to the LDL‐I (‐28%) and LDL‐II (‐23%) subfractions. These consistent and complementary changes in discrete LDL subfractions may, in part, explain the hypolipidaemic potency of acipimox in combination with low dose cholestyramine. In view of the association between LDL subfrac‐tion profile and coronary heart disease risk, this particular combination of drugs may represent an effective regimen for the modification of an athero‐genic lipoprotein phenotype.

Purpose of review The clinical efficacy and safety of statin therapy have been well established from a series of large-scale, randomized controlled trials. These trials, however, have predominantly recruited patients under the age of 70 years. As a consequence, the use of statins in older patients has remained controversial. Recent findings The results of the first trial to look exclusively at the elderly - the Prospective Study of Pravastatin in the Elderly at Risk - have added enormously to our understanding of the use of statins in the elderly. These findings, together with those from the large elderly cohort within the Heart Protection Study and the smaller elderly subgroups within the other major statin trials, have forced us to re-evaluate any systematic exclusion of elderly patients from statin therapy. Summary The collective evidence now strongly supports the use of statins in the at-risk elderly population.

EDITOR—The distinction between giving a general summary of trial results to study participants and providing them with their own trial results is not made clear in the paper by Dixon-Woods et al or the accompanying editorial.1 2 Investigators may provide patients with a summary of the trial results, but even if they do not they should be aware that study participants may access these results elsewhere. The provision of personalised …

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In ring sections of the sheep middle cerebral artery, electrical field stimulation elicits a complex response due to the simultaneous release of vasodilator and vasoconstrictor neurotransmitters. Haemolysate abolishes the relaxant effects of the vasodilator neurotransmitter and causes a marked augmentation of the contractile response in both the presence (448 +/- 191%) and absence (409 +/- 134%) of an intact endothelium. The haemolysate also reverses relaxation induced by sodium nitroprusside or sodium nitrite but has no effect on relaxation induced by 8-Br-cGMP. The vasodilator neurotransmitter therefore appears to act directly on the smooth muscle to cause relaxation by the stimulation of guanylate cyclase. The vasoconstrictor neurotransmitters that are released are antagonised by prazosin (100 nM), ketanserin (100 nM) and atropine (100 nM), which suggests that the transmitters involved are noradrenaline, 5-hydroxytryptamine (5-HT), and acetylcholine, respectively. In the presence of these three antagonists at 10 microM, there was 86.9 +/- 4.8% inhibition. Incubation with 5-HT (10 microM) causes a marked augmentation of the contractile response (267 +/- 56%) to field stimulation that can be reduced by pretreatment with either desipramine or citalopram, inhibitors of noradrenergic and serotoninergic uptake mechanisms, respectively. The 5-HT appears to be taken up into noradrenergic nerves and released as an alternative neurotransmitter upon subsequent stimulation. These actions of haemolysate and 5-HT may be involved in the cerebral vasospasm observed following subarachnoid haemorrhage.

Developed countries show a change in population demographics as increasing life expectancy leads to a proportionally older population and a greater incidence of chronic disease and comorbidities. Those future capability and capacity demands on healthcare providers are a major societal and economic challenge requiring new models of health and care delivery for sustainability.1,2 Innovate UK Health and Care have invested through their Stratified Medicine and Assisted Living Innovation Platforms to improve patient care and treatment provision, ensuring patients receive the right treatment at the right time and are provided for appropriately, whether in primary or secondary care.3,4In an ageing population, the development of chronic disease and comorbidities such as cardiovascular and respiratory disease, cancer, dementia and diabetes can be attributed to a certain extent by lifestyle choices such as smoking, obesity, alcohol use and lack of exercise in earlier life. However, the impact of each or all of these choices on an individual's health status is complex and dependent upon that person's genetic and physiological predisposition.Although family history is still a good indicator of health risk, the development of new technologies that allow earlier and more accurate measurement and detection of disease such as proteomics, characterisation of the human genome, diagnostic tests and clinical imaging has led to a greater understanding of the underlying molecular causes of disease. Differences in an individual's ability to metabolise drugs can also have major implications for side effects, especially in the older patient with comorbidities who will be subject to several treatment regimes. This molecular understanding causes stratification of patients into specific subgroups, allowing the application of precision medicine to ensure that patients benefit from a more personalised approach of targeted treatments and therapeutic interventions, improving patient outcomes and decreasing risk of adverse events.5The United Kingdom has a leading capability in the understanding and implementation of precision (or personalised) medicine as a consequence of its academic, industrial and clinical science base; however, in relation to implementing improvements in citizen health for the future, it is imperative that patients are actively involved.6 The Academy of Medical Sciences has recently proposed extending precision medicine to enable greater prediction and prevention of disease through increasing patient participation (P4 medicine).7One way to drive this P4 approach is through the use of digital technologies. Increasingly large quantities of data relevant to health are becoming available. UK investments in infrastructure such as the 100,000 Genome Project, UK Biobank, the MRC Farr Institute and Clinical Practice Research Datalink (CPRD) and Health and Social Care Information Centre (HSCIC) clinical and care databases are allowing the United Kingdom to lead the way in predictive and precision medicine. This linkage of molecular information - genetic mutations related to rare disease or cancer - to electronic health records will lead to better diagnosis and prediction of health risk and will inform future treatment pathways within the National Health Service (NHS). The development of mobile health and wearable technologies such as mobile health apps, fitness monitors, glucose, heart rate and movement monitors allows longitudinal studies and more accurate monitoring to occur in real life, with patients better able to monitor and self-manage their own illness. One example is a mobile phone app where patients track and monitor their symptoms of psychosis, obtaining early intervention and support to improve their own disease management with a potential saving to each NHS trust of £1 million per year.8The use of patient data in the NHS is of key public interest, and there is considerable effort to guarantee the governance, security and privacy of patient data. …

There is now a large body of evidence from multiple clinical trials confirming that lowering plasma low density lipoprotein (LDL)-cholesterol results in a clinically significant reduction in coronary heart disease (CHD) risk. These include the Scandinavian Simvastatin Survival Study and the West of Scotland Study. However, further analyses of the Helsinki Heart Study (HHS) have provided additional detail on the relationship between other lipid changes and incidence of CHD. In the HHS, the reduction of CHD events was related not only to the LDL-cholesterol lowering ability of the drug used, but also to its HDL-cholesterol raising and triglyceride lowering effects. Furthermore, a recent comparison of the levels of atheroma regression associated with different drug classes reveals that, while statins produce much greater changes in total and LDL-cholesterol, fibrates have the most marked effect on coronary artery minimum lumen diameter changes. In practice, the overall CHD risk of patients should be reduced by a co-ordinated approach to management of all the correctable risk factors. As part of this approach, the lipid profile should be assessed and normalised initially through the instigation of lifestyle changes, and if necessary, the adjunctive use of lipid regulating drugs. While the lowering of LDL-cholesterol is clearly important, the significance of lowering plasma triglyceride should not be overlooked.

SUMMARY Aims : To examine the effects of a pepsin inhibitor, pepstatin‐A, a long acting H 2 ‐receptor blocker, loxtidine, exogenous pepsin and exogenous acid against indomethacin‐induced antral ulceration in the rat. Results : Indomethacin (60 mg/kg s.c.) caused antral ulceration in fasted/re‐fed rats over a period of 4 h. Ulceration was prevented in a dose‐dependent manner by treatment with pepstatin‐A (0.1–1 mg.kg hourly) or loxtidine (3 mg/kg) given orally. Acidified methylcellulose (1 mL hourly per os ) enhanced damage and also prevented protection by loxtidine (3 mg/kg per os ). The protection by pepstatin‐A was not altered by treatment with acidified methylcellulose but was reversed by treatment with a 10‐fold excess of pepsin. Conclusion : These studies suggest that mucosal degradation by pepsin, rather than direct damage by luminal acid, was the major factor in the development of indomethacin‐induced antral ulceration in the rat.

This article describes how the first nurse-led national telephone helpline for the prevention of coronary heart disease (CHD) was set up and presents the results after the first year of operation. The free service was well received. /Most callers were seeking advice on CHD prevention for the first time and traditional risk factors and general healthy lifestyle advice remained major areas of concern

Little information exists to quantify the functional status and economic consequences of lipid-lowering therapy in elderly patients. We describe the design of the cost-effectiveness component of the first large, randomized, placebo-controlled trial of lipid-lowering therapy in subjects aged 70 years or older. The PROspective Study of Pravastatin in the Elderly at Risk has randomized 5804 men and women 70-82 years old, with existing vascular disease or related risk factors, to receive 40 mg/d of pravastatin or placebo. The cost-effectiveness study will be based on within-trial observations of differences between the two study arms in rates of myocardial infarction, stroke and related vascular disease outcomes (including vascular dementia). In addition to comparing within-trial clinical outcomes, we will model the projected changes in life expectancy and cardiovascular outcome rates that would be seen with lifelong use of the drug, based on the risk reduction rates seen in the trial as well as baseline observational data from the three countries where the trial is being conducted (Scotland, Ireland and the Netherlands). A state transition (Markov) model will be constructed to estimate the likely states of health, functional status and health care resource utilization (including lipid-lowering drug costs) in a cohort of elderly patients with versus without pravastatin therapy over a series of 1-year cycles until death. In addition, a standard measure of utility (the Health Utilities Index) will be administered to all study subjects to permit calculation of quality-adjusted life-years gained with this regimen. This approach will make it possible to go beyond the calculation of a single endpoint for each subject, and to translate the trial findings into definitions of effectiveness and outcomes that will be relevant to the ongoing debate concerning how best to relate the benefits of such medications to their costs.

Relatively little has been written about the practicalities of the closeout of large, multi-centre clinical trials, but this aspect of trial conduct and design is both important and requires careful planning in order to be accomplished in a timely and orderly fashion. Here, we document our recent experiences of closing down the Prospective Study of Pravastatin in the Elderly at Risk (PROSPER). This five-year study with over 5800 subjects was closed down and published in 2002. We describe the methods used to ensure the speedy progression from the start of the closeout period through data lock to publication and presentation of the results. We discuss the strategic planning of all aspects of the closeout process, the training of staff for final visits, the methods used to follow-up all study participants including those used in dealing with “difficult to contact” and defaulted patients, and the strategies employed to ensure that study participants were left with positive feelings about the study. We also detail the methods employed to expedite the throughput of study paperwork and endpoints and the cleaning of data in preparation for data lock and subsequent publication and presentation of the results. Based on our experiences we summarize the most important aspects of closeout design and make recommendations for future studies, the most important of which is that a well-planned and well-managed closeout is a key feature of any large scale clinical trial and a coherent and practicable closeout strategy should be an integral part of the design.

The fibric acid derivatives, or fibrates, have, until recently, been the most widely used lipid-lowering drugs in clinical practice. In the UK, for the year ending June 1990, the fibrates available at that time (bezafibrate, gemfibrozil and clofibrate) constituted 62% of all prescribed items for hyperlipidaemia. In the same period, bezafibrate was the most widely prescribed lipid-lowering drug (O’Brien 1991). In the US, from 1984–1987, gemfibrozil held first place in the ranking of lipid-lowering drugs in terms of number of prescriptions written and was only overtaken in 1988, after the introduction of lovastatin (Wysowski et al. 1990).

To investigate whether using an algorithm to select the starting dose of a statin according to baseline and target LDL-cholesterol (LDL-C) values would facilitate achieving lipid targets in patients with diabetes or the metabolic syndrome.Two 12-week, prospective, open-label trials enrolled 2717 high-risk subjects, of whom 1024 had diabetes and 1251 had metabolic syndrome. Subjects with LDL-C between 100 and 220 mg/dL (2.6-5.7 mmol/L) were assigned a starting dose of atorvastatin (10, 20, 40, or 80 mg/d) based on LDL-C level and status of statin use at baseline (statin-free [SF] or statin-treated [ST]), with a single uptitration at 6 weeks, if required.Among patients with diabetes, 81% of SF subjects (82%, 84%, 82%, and 76% with 10, 20, 40, and 80 mg, respectively) and 60% of ST subjects (61%, 68%, and 47% with 20, 40, and 80 mg, respectively) achieved LDL-C target. Among patients with metabolic syndrome, 78% of SF subjects (81%, 84%, 82%, and 66% with 10, 20, 40, and 80 mg, respectively) and 57% of ST subjects (58%, 70%, and 47% with 20, 40, and 80 mg, respectively) achieved LDL-C target. Among ST subjects, we observed reductions in LDL-C with atorvastatin beyond those achieved with other statins used at baseline in patients with diabetes and patients with metabolic syndrome. Atorvastatin was well tolerated.The ACTFAST studies confirm that a targeted starting dose of atorvastatin allows most patients with type 2 diabetes or the metabolic syndrome to achieve their LDL-C target safely with the initial dose or just a single titration. This therapeutic strategy may help overcome the treatment gap still observed in the treatment of lipids in diabetes.

The implementation of a pharmacovigilance service compliant with the legal and regulatory responsibilities of clinical trial sponsors presents particular challenges for sponsors in a non-commercial setting.In this paper we examine these challenges in detail. We identify and discuss the key steps in the development of a pharmacovigilance service within a public health service and university setting in the United Kingdom. We describe how we have established a central Pharmacovigilance Office with dedicated staff and resources within our organisation. This office is supported by an electronic pharmacovigilance reporting infrastructure developed to facilitate the receipt and processing of safety information, the onward reporting in compliance with legislation and the provision of sponsor institution oversight of clinical trial participant safety. An education and training programme has also been set up to ensure that all relevant staff in the organisation are fully aware of the pharmacovigilance service and are appropriately trained in its use.We discuss possible alternatives to this approach and why we consider our solution to be the most appropriate to ensure that a non-commercial sponsor organisation and investigators are operating in a fully compliant way.

Lipoprotein turnover studies continue to be used as an important research tool in the investigation of normal lipoprotein metabolism, dyslipidaemic states and the effects of pharmacological intervention. The introduction of stable isotopic endogenous labelling has served to complement existing radioisotope protocols while the use of improved software has facilitated new, improved mathematical models to explain the data.

With disbelief the world received news of medical experiments performed on the inmates of concentration camps in Nazi Germany.1 The subsequent trial of 23 defendants, 20 of whom were medically qualified, in Nuremberg yielded 16 guilty verdicts and seven death sentences. What was perhaps so incredulous was not that such atrocities could be perpetrated by Nazis, but that they could have involved doctors. In his opening remarks at the trial Brigadier General Telford Taylor, the American Chief of Council, noted: `To kill, to maim, and to torture is criminal under all modern systems of law... yet these [physician] defendants, all of whom were fully able to comprehend the nature of their acts... are responsible for wholesale murder and unspeakably cruel tortures.'2 During the eight-month trial a detailed catalogue of medical experimentation was presented to the court. For example, at Dachau between 1942-1944 there was a series of experiments designed to examine aspects of aviation medicine, involving high-altitude experiments, the effects of freezing water baths and the enforced drinking of sea-water. In Buchenwald and Ravensbruck throughout the war, a series of studies examined the simulation and treatment of battlefield injuries, such as exposure to mustard gas, phosphorus burns, bone transplantation, and sulfanilimide treatments. In Dachau, Buchenwald and Sachsenhausen during 1941-1945 there were a variety of infectious disease studies involving the deliberate infection of inmates with malaria, epidemic jaundice and typhus. Finally, in Auschwitz, Buchenwald and Ravensbruck there were a number of eugenic and lethal experiments involving sterilization and poisoned bullets. Similar, though less widely acknowledged, were the medical experiments performed in Japanese facilities during the Second World War.3 Nuremberg may have been the first major public exposition of how doctors can become instruments in unethical medical experimentation, but it is tragically not the last, nor regrettably the most recent. During the Doctors' Trial in Nuremberg the defendants cited similar forms of human experimentation that had taken place in the USA.4 Names were named and details listed, but little note was taken by the American prosecutors. Indeed the medical representative of the American Medical Association present at the trial, Andrew C. Ivy, stated that no American prisoner had ever been experimented on against his will.5 However, despite their heinous crimes against humanity, on this point the defendants were correct—and Ivy was wrong. The USA before, during and after the war was the host to a series of medical experiments that bore some striking similarities. Furthermore, although the 10 principles of ethical research that became known as the Nuremberg Code were drafted by the American team in Germany, it is notable that the Code was never taken to heart by American physicians, who preferred to view the document as applying not to them, but to the barbaric physicians in Nazi Germany.5 Then, as now, a utilitarian approach to biomedical research was commonly upheld as justification. `This is done for the greater good' possibly thought the New York team that from 1963 deliberately infected the mentally retarded with hepatitis at Willowbrook, an institution on Staten Island, so that experimental vaccines could be tested.6 The `benefit to society' may have been the touchstone of the Tuskegee investigators who between 1932 and 1972 followed a group of poor black Alabama farm workers to learn about the natural history of syphilis.7 In the process, and to avoid disruption of their experiment, they denied the men the benefits of penicillin therapy and subjected them to various investigations including regular lumbar punctures, which they passed off as treatments for so-called `bad blood'. The `advance of science' may have been the vision of those at the Jewish Chronic Disease Hospital in Brooklyn, who in the early 1960s injected live cancer cells into elderly debilitated cancer patients without their consent.8 These abuses of human research ethics were brought to light often not by our own colleagues, but by the media.9

The number of statins available to physicians continues to grow, leading to the question: Are all statins alike? Comparisons of side effects and safety profiles and the dose-response relationship among the different drugs show similar results. The cholesterol-lowering action of each depends on its ability to lower low-density lipoprotein cholesterol (LDL-C). On the other hand, the molecular structures of the newer statins are not similar and could have an effect on the mechanism of action of the compounds. Differences in metabolism also suggest the possibility of serious drug-drug interactions, and differences in levels of lipid reduction at varying dosages among the statins point to clinical variation as well.

The significance of the link between plasma cholesterol concentrations and risk of cerebrovascular disease (CVD) in the elderly is still debated. In a case-control study we found that elderly patients with ischaemic CVD have an atherogenic lipid profile, despite having low or normal total cholesterol concentrations.

Nursing practice is informed by research, and all nurses, midwives and care staff should be research aware even if they do not consider themselves research active. The principles of research ethics, based on respect for individuals, should not be confined to research. All nurses must be aware of these principles, and practise in accordance with them.

Biokimia Klinis: Teks Bergambar merupakan buku yang sangat penting untuk kurikulum kedokteran masa kini yang terus berubah. Setiap pokok bahasan disajikan dalam serangkaian &ldquounit pelajaran&rdquo dalam dua-halaman, dan masing-masing unit mencakup satu aspek khusus biokimia klinis. Keempat bagian buku ini memberikan suatu dasar yang pokok mengenai bidang ilmu ini: - Pengenalan terhadap biokimia klinis memberikan pengetahuan dasar untuk berbagai pekerjaan laboratorium di rumah sakit modern dan bagaimana menginterpretasikan hasil pemeriksaan - Biokimia dasar meliputi segala analisis rutin yang dikerjakan dan relevansinya dalam situasi klinis - Endokrinologi membahas pemeriksaan fungsi tiroid, adrenal, hipofisis, dan gonad - Pemeriksaan khusus memberikan gambaran umum tentang pemeriksaan yang jarang diminta namun tetap penting. Perubahan utama untuk edisi keempat ini adalah: - Materi yang ditulis ulang: Hipokalemia Hiperkalemia Cairan serebrospinal dan cairan tubuh lainnya - Materi tentang Pemeriksaan fungsi ginjal dan Hipertensi direvisi total - Ditambahkan daftar sumber yang tersedia di situs web Table Of Content: - Pengenalan Biokimia Klinis - Biokimia Dasar - Endokrinologi - Pemeriksaan Khusus - Penjelasan Riwayat Kasus - Sumber Web

Abstract High resolution apo(a) phenotyping has been used in conjunction with apo(a) genotyping to elucidate underlying mechanisms of plasma lipoprotein(a) (Lp(a)) level control. With increasing interest in this field more laboratories are now establishing such methods. High resolution apo(a) phenotyping has previously been reported with in-house monoclonal antibodies. Here the effective use of a commercially available monoclonal antibody is demonstrated. Two primary antibodies were compared and apo(a) isoform analysis was performed using an immunobloltting method. When equal quantities of Lp(a) were loaded approximately equal signal intensities were obtained, irrespective of apo(a) isoform size. Serial dilutions of samples with known Lp(a) level and a single expressing apo(a) isoform were used to determine the detection limit of the system. With both monoclonals it was possible to detect 0.05 mg/dl Lp(a).

Acute coronary syndromes contribute significantly to the patterns of morbidity and mortality in the elderly. A meta-analysis of the pathology of acute myocardial infarction has shown that 80-90% of episodes result from the rupture of small, unstable lesions that cause <70% diameter stenosis. Statins have been shown to stabilize the architecture of atherosclerotic plaques in humans and in animals. Statin treatment has also been shown to restore endothelial function, inhibit platelet thrombus formation and exert an anti-inflammatory effect. Collectively, these data support the use of statin therapy in those at risk for acute coronary syndromes, of whom the elderly are foremost.

Treatment of hyperlipidemia to reduce the risk of ischemic heart disease was, prior to the statin era, disappointingly limited in its ability to yield the benefits expected from the strong relationship known to exist between serum cholesterol and coronary death. Three primary prevention trials, using clofibrate, cholestyramine and gemfibrozil, had achieved modest reductions in fatal and nonfatal coronary events but none was able to extend life overall or even to reduce cardiovascular mortality; and combined analyses of the three raised disquiet over potential links between cholesterol reduction, cancer and aggressive or violent behaviour. The time was therefore ripe to determine whether statins could help prevent that first and all important myocardial infarction.The West of Scotland Coronary Prevention Study recruited 6,595 45- to 64-year-old men with no history of prior myocardial infarction and with low-density lipoprotein cholesterol in the range 4-6 mmol/l (155-232 mg/dl). Subjects who had undergone coronary revascularisation or had been hospitalised for angina pectoris in the previous 12 months were excluded, as were those with significant electrocardiographic abnormalities. Participants were randomised to receive pravastatin 40 mg/day or matching placebo and were followed for an average of 4.9 years.Treatment with pravastatin reduced the combined risk of fatal and nonfatal myocardial infarction by 31%. Cardiovascular death overall fell by 32% and the need for coronary revascularisation procedures was reduced by 37%. All of these endpoint benefits were statistically significant. Because there was no increase in non-cardiovascular mortality, the reduction in death from any cause also proved to be statistically significant (p = 0.051 by log rank test and p = 0.037 after adjustment for baseline risk factors).The West of Scotland Coronary Prevention Study is the first to show that cholesterol reduction with pravastatin helps avoid the first myocardial infarction, reduces coronary mortality and extends life. Ongoing exploration of the study database continues to unearth additional surprisingly beneficial effects of the treatment and permits authoritative decision-making on the effective use of lipid-lowering drugs.

As the roles of practice nurses develop, methods of offering them continuing education are being examined. A postal survey of 94 practice nurses in the Glasgow area was conducted to gain an impression of their knowledge and needs related to coronary heart disease (OLD) prevention. On the basis of the responses, a series of study days was designed and implemented, and a biannual newsletter on CHD risk factor assessment published and distributed. Both initiatives have been welcomed and, the authors claim, fit hi with the UKCC’s requirements on continuing education.