Yuyan Xiong

Abstract Aims Naturally secreted nanovesicles, known as exosomes, play important roles in stem cell-mediated cardioprotection. We have previously demonstrated that atorvastatin (ATV) pretreatment improved the cardioprotective effects of mesenchymal stem cells (MSCs) in a rat model of acute myocardial infarction (AMI). The aim of this study was to investigate if exosomes derived from ATV-pretreated MSCs exhibit more potent cardioprotective function in a rat model of AMI and if so to explore the underlying mechanisms. Methods and results Exosomes were isolated from control MSCs (MSC-Exo) and ATV-pretreated MSCs (MSCATV-Exo) and were then delivered to endothelial cells and cardiomyocytes in vitro under hypoxia and serum deprivation (H/SD) condition or in vivo in an acutely infarcted Sprague-Dawley rat heart. Regulatory genes and pathways activated by ATV pretreatment were explored using genomics approaches and functional studies. In vitro, MSCATV-Exo accelerated migration, tube-like structure formation, and increased survival of endothelial cells but not cardiomyocytes, whereas the exosomes derived from MSCATV-Exo-treated endothelial cells prevented cardiomyocytes from H/SD-induced apoptosis. In a rat AMI model, MSCATV-Exo resulted in improved recovery in cardiac function, further reduction in infarct size and reduced cardiomyocyte apoptosis compared to MSC-Exo. In addition, MSCATV-Exo promoted angiogenesis and inhibited the elevation of IL-6 and TNF-α in the peri-infarct region. Mechanistically, we identified lncRNA H19 as a mediator of the role of MSCATV-Exo in regulating expression of miR-675 and activation of proangiogenic factor VEGF and intercellular adhesion molecule-1. Consistently, the cardioprotective effects of MSCATV-Exo was abrogated when lncRNA H19 was depleted in the ATV-pretreated MSCs and was mimicked by overexpression of lncRNA H19. Conclusion Exosomes obtained from ATV-pretreated MSCs have significantly enhanced therapeutic efficacy for treatment of AMI possibly through promoting endothelial cell function. LncRNA H19 mediates, at least partially, the cardioprotective roles of MSCATV-Exo in promoting angiogenesis.

Arginase is a key hydrolase in the urea cycle that hydrolyses L-arginine to urea and L-ornithine. Increasing number of studies in recent years demonstrate that two mammalian arginase isoforms, arginase 1 (ARG1) and arginase 2 (ARG2), were aberrantly upregulated in various types of cancers, and played crucial roles in the regulation of tumor growth and metastasis through various mechanisms such as regulating L-arginine metabolism, influencing tumor immune microenvironment, etc. Thus, arginase receives increasing focus as an attractive target for cancer therapy. In this review, we provide a comprehensive overview of the physiological and biological roles of arginase in a variety of cancers, and shed light on the underlying mechanisms of arginase mediating cancer cells growth and development, as well as summarize the recent clinical research advances of targeting arginase for cancer therapy.

Background Macrophage‐mediated chronic inflammation is mechanistically linked to insulin resistance and atherosclerosis. Although arginase I is considered antiinflammatory, the role of arginase II (Arg‐ II ) in macrophage function remains elusive. This study characterizes the role of Arg‐ II in macrophage inflammatory responses and its impact on obesity‐linked type II diabetes mellitus and atherosclerosis. Methods and Results In human monocytes, silencing Arg‐ II decreases the monocytes' adhesion to endothelial cells and their production of proinflammatory mediators stimulated by oxidized low‐density lipoprotein or lipopolysaccharides, as evaluated by real‐time quantitative reverse transcription‐polymerase chain reaction and enzyme‐linked immunosorbent assay. Macrophages differentiated from bone marrow cells of Arg‐ II –deficient (Arg‐ II −/−) mice express lower levels of lipopolysaccharide‐induced proinflammatory mediators than do macrophages of wild‐type mice. Importantly, reintroducing Arg‐ II cDNA into Arg‐ II −/− macrophages restores the inflammatory responses, with concomitant enhancement of mitochondrial reactive oxygen species. Scavenging of re‐ active oxygen species by N ‐acetylcysteine prevents the Arg‐ II –mediated inflammatory responses. Moreover, high‐fat diet–induced infiltration of macrophages in various organs and expression of proinflammatory cytokines in adipose tissue are blunted in Arg‐ II −/− mice. Accordingly, Arg‐ II −/− mice reveal lower fasting blood glucose and improved glucose tolerance and insulin sensitivity. Furthermore, apolipoprotein E (ApoE)–deficient mice with Arg‐ II deficiency (ApoE −/− Arg‐ II −/−) display reduced lesion size with char‐ acteristics of stable plaques, such as decreased macrophage inflammation and necrotic core. In vivo adoptive transfer experiments reveal that fewer donor ApoE −/− Arg‐ II −/− than ApoE −/− Arg‐ II +/+ monocytes infiltrate into the plaque of ApoE −/− Arg‐ II +/+ mice. Conversely, recipient ApoE −/− Arg‐ II −/− mice accumulate fewer donor monocytes than do recipient ApoE −/− Arg‐ II +/+ animals. Conclusions Arg‐ II promotes macrophage proinflammatory responses through mitochondrial reactive oxygen species, contributing to insulin resistance and atherogenesis. Targeting Arg‐ II represents a potential therapeutic strategy in type II diabetes mellitus and atherosclerosis. ( J Am Heart Assoc. 2012;1:e000992 doi: 10.1161/ JAHA .112.000992.)

Abstract Background Bone marrow mesenchymal stem cells (MSCs) are among the most common cell types to be used and studied for cardiac regeneration. Low survival rate and difficult retention of delivered MSCs in infarcted heart remain as major challenges in the field. Co-delivery of stem cell-derived exosomes (Exo) is expected to improve the recruitment and survival of transplanted MSCs. Methods Exo was isolated from MSCs and delivered to an acute myocardial infarction (AMI) rat heart through intramyocardial injection with or without intravenous infusion of atrovastatin-pretreated MSCs on day 1, day 3, or day 7 after infarction. Echocardiography was performed to evaluate cardiac function. Histological analysis and ELISA test were performed to assess angiogenesis, SDF-1, and inflammatory factor expression in the infarct border zone. The anti-apoptosis effect of Exo on MSCs was evaluated using flow cytometry and Hoechst 33342 staining assay. Results We found that intramyocardial delivery of Exo followed by MSC transplantation (in brief, Exo+MSC treatment) into MI hearts further improved cardiac function, reduced infarct size, and increased neovascularization when compared to controls treated with Exo or MSCs alone. Of note, comparing the three co-transplanting groups, intramyocardially injecting Exo 30 min after AMI combined with MSCs transplantation at day 3 after AMI achieved the highest improvement in heart function. The observed enhanced heart function is likely due to an improved microenvironment via Exo injection, which is exemplified as reduced inflammatory responses and better MSC recruitment and retention. Furthermore, we demonstrated that pre-transplantation injection of Exo enhanced survival of MSCs and reduced their apoptosis both in vitro and in vivo. Conclusions Combinatorial delivery of exosomes and stem cells in a sequential manner effectively reduces scar size and restores heart function after AMI. This approach may represent as an alternative promising strategy for stem cell-based heart repair and therapy.

Acute myocardial infarction (AMI) is one of the leading causes of mortality around the world, and the inflammatory response plays a pivotal role in the progress of myocardial necrosis and ventricular remodeling, dysfunction and heart failure after AMI.Therapies aimed at modulating immune response after AMI on a molecular and cellular basis are urgently needed.Exosomes are a type of extracellular vesicles which contain a large amount of biologically active substances, like lipids, nucleic acids, proteins and so on.Emerging evidence suggests key roles of exosomes in immune regulation post AMI.A variety of immune cells participate in the immunomodulation after AMI, working together to clean up necrotic tissue and repair damaged myocardium.Stem cell therapy for myocardial infarction has long been a research hotspot during the last two decades and exosomes secreted by stem cells are important active substances and have similar therapeutic effects of immunomodulation, anti-apoptosis, anti-fibrotic and angiogenesis to those of stem cells themselves.Therefore, in this review, we focus on the characteristics and roles of exosomes produced by both of endogenous immune cells and exogenous stem cells in myocardial repair through immunomodulation after AMI.

Cardiac fibrosis, a common pathophysiologic process in most heart diseases, refers to an excess of extracellular matrix (ECM) deposition by cardiac fibroblasts (CFs), which can lead to cardiac dysfunction and heart failure subsequently. Not only CFs but also several other cell types including macrophages and endothelial cells participate in the process of cardiac fibrosis via different molecular pathways. Exosomes, ranging in 30–150 nm of size, have been confirmed to play an essential role in cellular communications by their bioactive contents, which are currently a hot area to explore pathobiology and therapeutic strategy in multiple pathophysiologic processes including cardiac fibrosis. Cardioprotective factors such as RNAs and proteins packaged in exosomes make them an excellent cell-free system to improve cardiac function without significant immune response. Emerging evidence indicates that targeting selective molecules in cell-derived exosomes could be appealing therapeutic treatments in cardiac fibrosis. In this review, we summarize the current understandings of cellular effectors, molecular pathways, and exosomal roles in cardiac fibrosis.

Bone marrow cells (BMCs), especially mesenchymal stem cells (MSCs), have shown attractive application prospects in acute myocardial infarction (AMI). However, the weak efficacy becomes their main limitation in clinical translation. Based on the anti-inflammation and anti-apoptosis effects of a Chinese medicine-Tongxinluo (TXL), we aimed to explore the effects of TXL-pretreated MSCs (MSCsTXL) in enhancing cardiac repair and further investigated the underlying mechanism.MSCsTXL or MSCs and the derived exosomes (MSCsTXL-exo or MSCs-exo) were collected and injected into the infarct zone of rat hearts. In vivo, the anti-apoptotic and anti-inflammation effects, and cardiac functional and histological recovery were evaluated. In vitro, the apoptosis was evaluated by western blotting and flow cytometry. miRNA sequencing was utilized to identify the significant differentially expressed miRNAs between MSCsTXL-exo and MSCs-exo, and the miRNA mimics and inhibitors were applied to explore the specific mechanism.Compared to MSCs, MSCsTXL enhanced cardiac repair with reduced cardiomyocytes apoptosis and inflammation at the early stage of AMI and significantly improved left ventricular ejection fraction (LVEF) with reduced infarct size in an exosome-dependent way. Similarly, MSCsTXL-exo exerted superior therapeutic effects in anti-apoptosis and anti-inflammation, as well as improving LVEF and reducing infarct size compared to MSCs-exo. Further exosomal miRNA analysis demonstrated that miR-146a-5p was the candidate effector of the superior effects of MSCsTXL-exo. Besides, miR-146a-5p targeted and decreased IRAK1, which inhibited the nuclear translocation of NF-κB p65 thus protecting H9C2 cells from hypoxia injury.This study suggested that MSCsTXL markedly facilitated cardiac repair via a new mechanism of the exosomal transfer of miR-146a-5p targeting IRAK1/NF-κB p65 pathway, which has great potential for clinical translation.

Extracellular vesicles (EVs) derived from bone marrow mesenchymal stem cells (MSCs) pretreated with atorvastatin (ATV) (MSCATV-EV) have a superior cardiac repair effect on acute myocardial infarction (AMI). The mechanisms, however, have not been fully elucidated. This study aims to explore whether inflammation alleviation of infarct region via macrophage polarization plays a key role in the efficacy of MSCATV-EV.MSCATV-EV or MSC-EV were intramyocardially injected 30 min after coronary ligation in AMI rats. Macrophage infiltration and polarization (day 3), cardiac function (days 0, 3, 7, 28), and infarct size (day 28) were measured. EV small RNA sequencing and bioinformatics analysis were conducted for differentially expressed miRNAs between MSCATV-EV and MSC-EV. Macrophages were isolated from rat bone marrow for molecular mechanism analysis. miRNA mimics or inhibitors were transfected into EVs or macrophages to analyze its effects on macrophage polarization and cardiac repair in vitro and in vivo.MSCATV-EV significantly reduced the amount of CD68+ total macrophages and increased CD206+ M2 macrophages of infarct zone on day 3 after AMI compared with MSC-EV group (P < 0.01-0.0001). On day 28, MSCATV-EV much more significantly improved the cardiac function than MSC-EV with the infarct size markedly reduced (P < 0.05-0.0001). In vitro, MSCATV-EV also significantly reduced the protein and mRNA expressions of M1 markers but increased those of M2 markers in lipopolysaccharide-treated macrophages (P < 0.05-0.0001). EV miR-139-3p was identified as a potential cardiac repair factor mediating macrophage polarization. Knockdown of miR-139-3p in MSCATV-EV significantly attenuated while overexpression of it in MSC-EV enhanced the effect on promoting M2 polarization by suppressing downstream signal transducer and activator of transcription 1 (Stat1). Furthermore, MSCATV-EV loaded with miR-139-3p inhibitors decreased while MSC-EV loaded with miR-139-3p mimics increased the expressions of M2 markers and cardioprotective efficacy.We uncovered a novel mechanism that MSCATV-EV remarkably facilitate cardiac repair in AMI by promoting macrophage polarization via miR-139-3p/Stat1 pathway, which has the great potential for clinical translation.

In recent decades, the occurrence of heart failure with preserved ejection fraction (HFpEF) has outweighed that of heart failure with reduced ejection fraction by degrees, but few drugs have been demonstrated to improve long-term clinical outcomes in patients with HFpEF. Levosimendan, a calcium-sensitizing cardiotonic agent, improves decompensated heart failure clinically. However, the anti-HFpEF activities of levosimendan and underlying molecular mechanisms are unclear.In this study, a double-hit HFpEF C57BL/6N mouse model was established, and levosimendan (3 mg/kg/week) was administered to HFpEF mice aged 13 to 17 weeks. Different biological experimental techniques were used to verify the protective effects of levosimendan against HFpEF.After four weeks of drug treatment, left ventricular diastolic dysfunction, cardiac hypertrophy, pulmonary congestion, and exercise exhaustion were significantly alleviated. Junction proteins in the endothelial barrier and between cardiomyocytes were also improved by levosimendan. Among the gap junction channel proteins, connexin 43, which was especially highly expressed in cardiomyocytes, mediated mitochondrial protection. Furthermore, levosimendan reversed mitochondrial malfunction in HFpEF mice, as evidenced by increased mitofilin and decreased ROS, superoxide anion, NOX4, and cytochrome C levels. Interestingly, after levosimendan administration, myocardial tissue from HFpEF mice showed restricted ferroptosis, indicated by an increased GSH/GSSG ratio; upregulated GPX4, xCT, and FSP-1 expression; and reduced intracellular ferrous ion, MDA, and 4-HNE levels.Regular long-term levosimendan administration can benefit cardiac function in a mouse model of HFpEF with metabolic syndromes (namely, obesity and hypertension) by activating connexin 43-mediated mitochondrial protection and sequential ferroptosis inhibition in cardiomyocytes.

Rationale: The crosstalk between cardiac microvascular endothelial cells (CMECs) and cardiomyocytes (CMs) has emerged as a key component in the development of, and protection against, cardiac diseases. For example, activation of endothelial nitric oxide synthase (eNOS) in CMECs, by therapeutic strategies such as ischemic preconditioning, plays a critical role in the protection against myocardial ischemia/reperfusion (I/R) injury. However, much less is known about the signals produced by CMs that are able to regulate CMEC biology. Here we uncovered one such mechanism using Tongxinluo (TXL), a traditional Chinese medicine, that alleviates myocardial ischemia/reperfusion (I/R) injury by activating CMEC eNOS. The aim of our study is to identify the signals produced by CMs that can regulate CMEC biology during I/R. Methods: Ex vivo, in vivo, and in vitro settings of ischemia-reperfusion were used in our study, with the protective signaling pathways activated in CMECs identified using genetic inhibition (p70s6k1 siRNA, miR-145-5p mimics, etc.), chemical inhibitors (the eNOS inhibitor, L-NNA, and the small extracellular vesicles (sEVs) inhibitor, GW4869) and Western blot analyses. TritonX-100 at a dose of 0.125% was utilized to inactivate the eNOS activity in endothelium to investigate the role of CMEC-derived eNOS in TXL-induced cardioprotection. Results: We found that while CMEC-derived eNOS activity was required for the cardioprotection of TXL, activation of eNOS in CMECs by TXL did not occur directly. Instead, eNOS activation in CMECs required a crosstalk between CMs and CMECs through the uptake of CM-derived sEVs. We further demonstrate that TXL induced CM-sEVs contain increased levels of Long Intergenic Non-Protein Coding RNA, Regulator Of Reprogramming (Linc-ROR). Upon uptake into CMECs, linc-ROR downregulates its target miR-145-5p leading to activation of the eNOS pathway by facilitating the expression of p70s6k1 in these cells. The activation of CMEC-derived eNOS works to increase survival in both the CMECs and the CMs themselves. Conclusions: These data uncover a mechanism by which the crosstalk between CMs and CMECs leads to the increased survival of the heart after I/R injury and point to a new therapeutic target for the blunting of myocardial I/R injury.

Bone marrow-derived mesenchymal stem cells (MSCs), which possess immunomodulatory characteristic, are promising candidates for the treatment of acute myocardial infarction (AMI). However, the low retention and survival rate of MSCs in the ischemic heart limit their therapeutic efficacy. Strategies either modifying MSCs or alleviating the inflammatory environment, which facilitates the recruitment and survival of the engrafted MSCs, may solve the problem. Thus, we aimed to explore the therapeutic efficacy of sequential transplantation of exosomes and combinatorial pretreated MSCs in the treatment of AMI.Exosomes derived from MSCs were delivered to infarcted hearts through intramyocardial injection followed by the intravenous infusion of differentially pretreated MSCs on Day 3 post-AMI. Enzyme linked immunosorbent assay (ELISA) was performed to evaluate the inflammation level as well as the SDF-1 levels in the infarcted border zone of the heart. Echocardiography and histological analysis were performed to assess cardiac function, infarct size, collagen area and angiogenesis.Sequential transplantation of exosomes and the combinatorial pretreated MSCs significantly facilitated cardiac repair compared to AMI rats treated with exosomes alone. Notably, compared to the other three methods of cotransplantation, combinatorial pretreatment with hypoxia and Tongxinluo (TXL) markedly enhanced the CXCR4 level of MSCs and promoted recruitment, which resulted in better cardiac function, smaller infarct size and enhanced angiogenesis. We further demonstrated that exosomes effectively reduced apoptosis in MSCs in vitro.Sequential delivery of exosomes and pretreated MSCs facilitated cardiac repair post-AMI, and combined pretreatment with hypoxia and TXL better enhanced the cardioprotective effects. This method provides new insight into the clinical translation of stem cell-based therapy for AMI.

The development of polymeric systems that can integrate individual basic logic gates into combinational circuits has been extremely interesting as smart materials. A novel thermometer fluorescent sensor poly(NIPMAM-co-MDCPDP) consisting of N-isopropylmethacrylamide (NIPMAM) as a thermoresponsive unit and dicyanomethylene-4H-pyran derivative as a dipolar-sensitive fluorophore unit was designed. Poly(NIPMAM-co-MDCPDP) performs fluorescence quenching merely by coordination with Cu2+ ions or increasing temperature in neutral or acid aqueous solution. The ON−OFF fluorescence response of poly(NIPMAM-co-MDCPDP) is driven by a temperature-induced phase transition from coil to globule and the capture of Cu2+ ions, resulting in a decrease of the ICT efficiency in neutral solution. The combinational serial NOR logic operation as well as two INHIBIT logic gates was constructed with three inputs: various pH, temperature change, and Cu2+ ions. The proposed combinational logic circuits play a key role in mimicking comprehensive arithmetic operations at the nanoscale level.

Abstract Aims Interleukin (IL)-5 mediates the development of eosinophils (EOS) that are essential for tissue post-injury repair. It remains unknown whether IL-5 plays a role in heart repair after myocardial infarction (MI). This study aims to test whether IL-5-induced EOS population promotes the healing and repair process post-MI and to reveal the underlying mechanisms. Methods and results MI was induced by permanent ligation of the left anterior descending coronary artery in wild-type C57BL/6 mice. Western blot and real-time polymerase chain reaction revealed elevated expression of IL-5 in the heart at 5 days post-MI. Immunohistostaining indicated that IL-5 was secreted mainly from macrophages and CD127+ cells in the setting of experimental MI. External supply of recombinant mouse IL-5 (20 min, 1 day, and 2 days after MI surgery) reduced the infarct size and increased ejection fraction and angiogenesis in the border zone. A significant expansion of EOS was detected in both the peripheral blood and infarcted myocardium after IL-5 administration. Pharmacological depletion of EOS by TRFK5 pretreatment muted the beneficial effects of IL-5 in MI mice. Mechanistic studies demonstrated that IL-5 increased the accumulation of CD206+ macrophages in infarcted myocardium at 7 days post-MI. In vitro co-culture experiments showed that EOS shifted bone marrow-derived macrophage polarization towards the CD206+ phenotypes. This activity of EOS was abolished by IL-4 neutralizing antibody, but not IL-10 or IL-13 neutralization. Western blot analyses demonstrated that EOS promoted the macrophage downstream signal transducer and activator of transcription 6 (STAT6) phosphorylation. Conclusion IL-5 facilitates the recovery of cardiac dysfunction post-MI by promoting EOS accumulation and subsequent CD206+ macrophage polarization via the IL-4/STAT6 axis.

Exosomes derived from bone marrow mesenchymal stem cells (MSC-exo) have been considered as a promising cell-free therapeutic strategy for ischemic heart disease. Cardioprotective drug pretreatment could be an effective approach to improve the efficacy of MSC-exo. Nicorandil has long been used in clinical practice for cardioprotection. This study aimed to investigate whether the effects of exosomes derived from nicorandil pretreated MSC (MSCNIC-exo) could be enhanced in facilitating cardiac repair after acute myocardial infarction (AMI).MSCNIC-exo and MSC-exo were collected and injected into the border zone of infarcted hearts 30 minutes after coronary ligation in rats. Macrophage polarization was detected 3 days post-infarction, cardiac function as well as histological pathology were measured on the 28th day after AMI. Macrophages were separated from the bone marrow of rats for in vitro model. Exosomal miRNA sequencing was conducted to identify differentially expressed miRNAs between MSCNIC-exo and MSC-exo. MiRNA mimics and inhibitors were transfected to MSCs or macrophages to explore the specific mechanism.Compared to MSC-exo, MSCNIC-exo showed superior therapeutic effects on cardiac functional and structural recovery after AMI and markedly elevated the ratio of CD68+ CD206+/ CD68+cells in infarcted hearts 3 days post-infarction. The notable ability of MSCNIC-exo to promote macrophage M2 polarization was also confirmed in vitro. Exosomal miRNA sequencing and both in vivo and in vitro experiments identified and verified that miR-125a-5p was an effector of the roles of MSCNIC-exo in vivo and in vitro. Furthermore, we found miR-125a-5p promoted macrophage M2 polarization by inhibiting TRAF6/IRF5 signaling pathway.This study suggested that MSCNIC-exo could markedly facilitate cardiac repair post-infarction by promoting macrophage M2 polarization by upregulating miR-125a-5p targeting TRAF6/IRF5 signaling pathway, which has great potential for clinical translation.

Despite several recent advances, current therapy and prevention strategies for myocardial infarction are far from satisfactory, owing to limitations in their applicability and treatment effects.Nanoparticles (NPs) enable the targeted and stable delivery of therapeutic compounds, enhance tissue engineering processes, and regulate the behaviour of transplants such as stem cells.Thus, NPs may be more effective than other mechanisms, and may minimize potential adverse effects.This review provides evidence for the view that functionoriented systems are more practical than traditional material-based systems; it also summarizes the latest advances in NP-based strategies for the treatment and prevention of myocardial infarction.

Type 2 immunity participates in the pathogeneses of helminth infection and allergic diseases. Emerging evidence indicates that the components of type 2 immunity are also involved in maintaining metabolic hemostasis and facilitating the healing process after tissue injury. Numerous preclinical studies have suggested regulation of type 2 immunity-related cytokines, such as interleukin-4, -13, and -33, and cell types, such as M2 macrophages, mast cells, and eosinophils, affects cardiac functions after myocardial infarction (MI), providing new insights into the importance of immune modulation in the infarcted heart. This review provides an overview of the functions of these cytokines and cells in the setting of MI as well as their potential to predict the severity and prognosis of MI.

Over the past decade, histone deacetylases (HDACs) has been proven to manipulate development and exacerbation of cardiovascular diseases, including myocardial ischemia/reperfusion injury, cardiac hypertrophy, ventricular remodeling, and myocardial fibrosis. Inhibition of HDACs, especially class-I HDACs, is potent to the protection of ischemic myocardium after ischemia/reperfusion (I/R). Herein, we examine whether mocetinostat (MGCD0103, MOCE), a class-I selective HDAC inhibitor in phase-II clinical trial, shows cardioprotection under I/R in vivo and in vitro, if so, reveal its potential pharmacological mechanism to provide an experimental and theoretical basis for mocetinostat usage in a clinical setting. Human cardiac myocytes (HCMs) were exposed to hypoxia and reoxygenation (H/R), with or without mocetinostat treatment. H/R reduced mitochondrial membrane potential and induced HCMs apoptosis. Mocetinostat pretreatment reversed these H/R-induced mitochondrial damage and cellular apoptosis and upregulated CREB, p-CREB, and PGC-1α in HCMs during H/R. Transfection with small interfering RNA against PGC-1α or CREB abolished the protective effects of mocetinostat on cardiomyocytes undergoing H/R. In vivo, mocetinostat was demonstrated to protect myocardial injury posed by myocardial I/R via the activation of CREB and upregulation of PGC-1α. Mocetinostat (MGCD0103) can protect myocardium from I/R injury through mitochondrial protection mediated by CREB/PGC-1α pathway. Therefore, activation of the CREB/PGC-1α signaling pathway via the inhibition of Class-I HDACs may be a promising new therapeutic strategy for alleviating myocardial reperfusion injury.

Excessive free fatty acids (FFAs) accumulation is a leading risk factor for the pathogenesis of insulin resistance (IR) in metabolic tissues, including the liver. Ethanolamine-phosphate phospho-lyase (ETNPPL), a newly identified metabolic enzyme, catalyzes phosphoethanolamine (PEA) to ammonia, inorganic phosphate, and acetaldehyde and is highly expressed in hepatic tissue. Whether it plays a role in regulating FFA-induced IR in hepatocytes has yet to be understood. In this study, we established an in vitro palmitic acid (PA)-induced IR model in human HepG2 cells and mouse AML12 cells with chronic treatment of PA. Next, we overexpressed ETNPPL by using lentivirus-mediated ectopic to investigate the effects of ETNPPL per se on IR without PA stimulation. We show that ETNPPL expression is significantly elevated in PA-induced IR and that silencing ETNPPL ameliorates this IR in hepatocytes. Inversely, overexpressing ETNPPL under normal conditions without PA promotes IR, reactive oxygen species generation, and ARG2 activation in both HepG2 and AML12 cells. Moreover, ETNPPL depletion markedly down-regulates ARG2 expression in hepatocytes. Besides, silencing ARG2 prevents ETNPPL-induced ROS accumulation and inhibition of autophagic flux and IR in hepatocytes. Finally, we found that phytopharmaceutical disruption of ETNPPL by quercetin ameliorates PA-induced IR in hepatocytes. Our study discloses that ETNPPL inhibiting autophagic flux mediates insulin resistance triggered by PA in hepatocytes via ARG2/ROS signaling cascade. Our findings provide novel insights into elucidating the pathogenesis of obesity-associated hepatic IR, suggesting that targeting ETNPPL might represent a potential approach for T2DM therapy.

Acute myocardial infarction (AMI) is an event of ischemic myocardial necrosis caused by acute coronary artery occlusion, which ultimately leads to a large loss of cardiomyocytes. The prerequisite of salvaging ischemic myocardium and improving cardiac function of patients is to provide adequate blood perfusion in the infarcted area. Apart from reperfusion therapy, it is also urgent and imperative to promote angiogenesis. Recently, growing evidence based on promising preclinical data indicates that mesenchymal stem cells (MSCs) can provide therapeutic effects on AMI by promoting angiogenesis. Extracellular vesicles (EVs), membrane-encapsulated vesicles with complex cargoes, including proteins, nucleic acids, and lipids, can be derived from MSCs and represent part of their functions, so EVs also possess the ability to promote angiogenesis. However, poor control of the survival and localization of MSCs hindered clinical transformation and made scientists start looking for new approaches based on MSCs. Identifying the role of MSCs and their derived EVs in promoting angiogenesis can provide a theoretical basis for improved MSC-based methods, and ultimately promote the clinical treatment of AMI. This review highlights potential proangiogenic mechanisms of transplanted MSCs and the derived EVs after AMI and summarizes the latest literature concerning the novel methods based on MSCs to maximize the angiogenesis capability.

Melanoma is a malignant and aggressive cancer, and its progression is greatly affected by interactions between melanoma cells and their surroundings. Exploration on mechanism of melanoma and improved diagnostic and therapeutic strategies are becoming increasingly important. Unlike extracellular messengers that mainly work on targeted cells through corresponding receptors, exosomes are essential intercellular messengers that deliver biologically active substances such as nucleic acids and proteins to target cells for cell–cell communication. Of them, microRNAs (miRNAs) are common and important exosomal components that can regulate the expression of a wide range of target genes. Accordingly, exosome-derived miRNAs play a significant role in melanoma progression, including invasion and metastasis, microenvironment establishment, angiogenesis, and immune escape. MiRNA signatures of exosomes are specific in melanoma patients compared to healthy controls, thus circulating miRNAs, especially exosomal miRNAs, become potential diagnostic markers and therapeutic targets for melanoma. This review aims to summarize recent studies on the role of exosomal miRNAs in melanoma as well as ongoing efforts in melanoma treatment.

Aim: To determine the efficacy and safety of intracoronary infusion of autologous bone marrow mesenchymal stem cells (MSC INJ ) in combination with intensive atorvastatin (ATV) treatment for patients with anterior ST-segment elevation myocardial infarction-elevation myocardial infarction. Patients &amp; methods: The trial enrolls a total of 100 patients with anterior ST-elevation myocardial infarction. The subjects are randomly assigned (1:1:1:1) to receive routine ATV (20 mg/d) with placebo or MSCs INJ and intensive ATV (80 mg/d) with placebo or MSCs INJ . The primary end point is the absolute change of left ventricular ejection fraction within 12 months. The secondary end points include parameters in cardiac function, remodeling and regeneration, quality of life, biomarkers and clinical outcomes. Results &amp; conclusion: The trial will implicate the essential of cardiac micro-environment improvement (‘fertilizing’) for cell-based therapy. Clinical Trial Registration: NCT03047772.

The SDF-1/CXCR4 signaling plays a critical role in the trafficking of mesenchymal stem cells (MSCs) to the sites of tissue damage. Our recent study demonstrated that atorvastatin (ATV) treatment improved the survival of MSCs, and ATV pretreated MSCs (ATV-MSCs) exhibited enhanced engraftment to injured myocardium. In this study, we investigated whether combined treatment with ATV and ATV-MSCs enhances cardiac repair and regeneration by activating SDF-1/CXCR4 signaling in a rat model of acute myocardial infarction. Rats were randomized into eight groups: the Sham, AMI control and 6 other groups that were subjected to AMI followed by treatment with MSCs, ATV, ATV+MSCs, ATV-MSCs, ATV+ATV-MSCs, ATV+ATV-MSCs+AMD3100 (SDF-1/CXCR4 antagonist), respectively. ATV+ATV-MSCs significantly potentiated targeted recruitment of MSCs to peri-infarct myocardium and resulted in further improvements in cardiac function and reduction in scar size compared with MSCs treatment alone at 4-week after AMI. More importantly, the cardioprotective effects conferred by ATV+ATV-MSCs were almost completely abolished by AMD3100 treatment. Together, our study demonstrated that ATV+ATV-MSCs significantly enhanced the targeted recruitment and survival of transplanted MSCs, and resulted in subsequent cardiac function improvement by augmenting SDF-1/CXCR4 signaling.

Objective To test whether intensive atorvastatin (ATV) increases the efficacy of transplantation with autologous bone marrow mononuclear cells (MNCs) in patients suffering from anterior ST-elevated myocardial infarction (STEMI). Methods This clinical trial was under a 2×2 factorial design, enrolling 100 STEMI patients, randomly into four groups of regular (RA) or intensive ATV (IA) with MNCs or placebo. The primary endpoint was the change of left ventricular ejection fraction (LVEF) at 1-year follow-up from baseline, primarily assessed by MRI. The secondary endpoints included other parameters of cardiac function, remodelling and regeneration determined by MRI, echocardiography, positron emission tomography (PET) and biomarkers. Results All the STEMI patients with transplantation of MNCs showed significantly increased LVEF change values than those with placebo (p=0.01) with only in the IA+MNCs patients group demonstrating significantly elevation of LVEF than in the IA+placebo group (+12.6% (95%CI 10.4 to 19.3) vs +5.0% (95%CI 4.0 to 10.0), p=0.001), pointing to a better synergy between ATV and MNCs (p=0.019). PET analysis revealed significantly increased viable areas of myocardium (p=0.015), while the scar sizes (p=0.026) and blood aminoterminal pro-B-type natriuretic peptide (p&lt;0.034) reduced. All these above benefits of MNCs were also attributed to IA+MNCs instead of RA+MNCs group of patients with STEMI. Conclusions Intensive ATV treatment augments the therapeutic efficacy of MNCs in patients with anterior STEMI at the convalescent stage. The treatment with the protocol of intensive ATV and MNC combination offers a clinically essential approach for myocardial infarction. Trial registration number NCT00979758 .

Ferroptosis, a newly defined and iron-dependent cell death, morphologically and biochemically differs from other cell deaths. Melanoma is a serious type of skin cancer, and the poor efficacy of current therapies causes a major increase in mortality. Sorafenib, a multiple kinase inhibitor, has been evaluated in clinical phase trials of melanoma patients, which shows modest efficacy. Emerging evidence has demonstrated that arginase 2 (Arg2), type 2 of arginase, is elevated in various types of cancers including melanoma. To investigate the role and underlying mechanism of Arg2 in sorafenib-induced ferroptosis in melanoma, reverse transcriptase-quantitative polymerase chain reaction, western blot analysis, adenovirus and lentivirus transduction, and in vivo tumor homograft model experiments were conducted. In this study, we show that sorafenib treatment leads to melanoma cell death and a decrease in Arg2 at both the mRNA and protein levels. Knockdown of Arg2 increases lipid peroxidation, which contributes to ferroptosis, and decreases the phosphorylation of Akt. In contrast, overexpression of Arg2 rescues sorafenib-induced ferroptosis, which is prevented by an Akt inhibitor. In addition, genetic and pharmacological suppression of Arg2 is able to ameliorate the anticancer activity of sorafenib in melanoma cells in vitro and in tumor homograft models. We also show that Arg2 suppresses ferroptosis by activating the Akt/GPX4 signaling pathway, negatively regulating sorafenib-induced cell death in melanoma cells. Our study not only uncovers a novel mechanism of ferroptosis in melanoma but also provides a new strategy for the clinical applications of sorafenib in melanoma treatment.

Revascularization therapies for culprit arteries, regardless of percutaneous coronary intervention and coronary artery bypass grafting, are considered the best strategy for improving the clinical prognosis of patients with acute coronary syndrome (ACS). Nonetheless, myocardial reperfusion following effective revascularization can trigger significant cardiomyocyte death and coronary endothelial collapse, known as myocardial ischemia/reperfusion injury (MIRI). Usually, endothelial cells and their intercellular tight junctions cooperatively maintain the microvascular endothelial barrier and its relatively low permeability but fail in reperfusion areas. Microvascular endothelial hyperpermeability induced by ischemia/reperfusion (IR) contributes to myocardial edema, increased infiltration of pro-inflammatory cells, and aggravated intramyocardial hemorrhage, which may worsen the prognosis of ACS. The tracer used in this study-70,000 Da FITC-dextran, a branched glucose molecule labeled by fluorescein isothiocyanate (FITC)-appears too large to infiltrate the cardiac microvascular endothelium in normal conditions. However, it is capable of infiltrating a broken barrier after MIRI. Thus, the higher the endothelial permeability is, the more FITC-dextran accumulates in the extravascular intercellular space. Thus, the intensity of fluorescence from FITC can indicate the permeability of the microvascular endothelial barrier. This protocol takes advantage of FITC-dextran to evaluate the cardiac microvascular endothelial barrier functionally, which is detected by an automated quantitative pathology imaging system.

Retraction of ‘Direct Sb–S cross-coupling of halostibines with thiols and disulfides at room temperature’ by Wei Li et al. , Org. Biomol. Chem. , 2021, Accepted Manuscript, https://doi.org/10.1039/d1ob00959a.

Gastric cancer (GC) is one of the most prevalent cancers worldwide. Ferroptosis and the immune status of tumor tissue play vital roles in the initiation and progression of GC. However, the role and functional mechanisms of ferroptosis- and immunity-related genes (FIRGs) in GC pathogenesis and their correlations with GC prognosis have not been elucidated. We aim to establish a prognostic prediction model based on the FIRGs signature for GC patients. Differentially expressed genes were screened from the Cancer Genome Atlas (TCGA) GC cohorts. The least absolute shrinkage and selection operator (LASSO) regression was performed to establish a FIRGs-based risk model. This gene signature with 7 FIRGs was identified as an independent prognostic factor. A nomogram incorporating clinical parameters and the FIRG signature was constructed to individualize outcome predictions. Finally, we provided in vivo and in vitro evidence to verify the reliability of FIRG signature for GC prognosis, and validate the expression and function of FIRGs contributing to the development and progression of GC. Herein, our work represents great therapeutic and prognostic potentials for GC.

Osteonecrosis of the femoral head (ONFH) is a disease with a high disability rate, and genetic factors are closely related to its pathogenesis. This study aimed to investigate the possible correlation between ESR1 and APOE gene polymorphisms and the risk of ONFH.In this case-control study, the potential association between three genetic variants (rs2982573 C < T, rs10872678 C < T, and rs9322332 A < C) of the ESR1 gene and two genetic variants (rs7259620 A < G and rs769446 C < T) of the APOE gene with the risk of ONFH was investigated. Correlations between gene polymorphisms and ONFH risk were assessed using logistic regression analysis, with calculation of odds ratios (ORs) and 95% confidence intervals (CIs).The overall analysis demonstrated that rs9322332 in the ESR1 gene exhibited a correlation with a decreased risk of ONFH under the homozygous (AA vs.CC: OR = 0.69, 95% CI [0.53-0.90], p = 0.006), dominant (CA + AA vs. CC: OR = 0.70, 95% CI [0.54-0.90], p = 0.006), and additive (OR = 0.79, 95% CI [0.66-0.95], p = 0.013) models. The stratification analysis revealed that rs9322332 was linked to a lower risk of ONFH in subgroups characterized by individuals aged over 51 years and non-smokers. Nevertheless, there were no notable correlations found between ESR1 rs2982573 and rs10872678, as well as APOE rs7259620 and rs769446, with the risk of ONFH.ESR1-rs9322332 is closely linked to a decreased risk of ONFH, thereby enhancing our understanding of the relationship between gene polymorphisms and ONFH.

With the development of stem cell therapy in translational research and regenerative medicine, bone marrow mesenchymal stem cells (BM-MSCs), as a kind of pluripotent stem cells, are favored for their instant availability and proven safety. It has been reported that transplantation of BM-MSCs is of great benefit to repairing injured tissues in various diseases, which might be related to modulating the immune and inflammatory responses via paracrine mechanisms. Extracellular vesicles (EVs), featuring a double-layer lipid membrane structure, are considered to be the main mediators of the paracrine effects of stem cells. Recognized for their crucial roles in cell communication and epigenetic regulation, EVs have already been applied in vivo for immunotherapy. However, similar to its maternal cells, most of the studies on the efficacy of transplantation of EVs still remain at the level of small animals, which is not enough to provide essential evidence for clinical translation. Here, we use density-gradient centrifugation to isolate bone marrow cells (BMC) from porcine bone marrow at first, and get porcine BM-MSCs (pBM-MSCs) by cell culture subsequently, identified by the results of observation under the microscope, induced differentiation assay, and flow cytometry. Furthermore, we isolate EVs derived from pBM-MSCs in cell supernatant by ultracentrifugation, proved by the techniques of transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blotting successfully. Overall, pBM-MSCs and their derived EVs can be isolated and identified effectively by the following protocols, which might be widely used in pre-clinical studies on the transplantation efficacy of BM-MSCs and their derived EVs.

Background: Eosinophils have been commonly recognized as one of the major participants in helminth infection and allergic diseases. Recent studies suggest that these innate immune cells mediate tissue repairs after muscle, hepatic, or endometrial injuries. Yet, the precise role of eosinophils in myocardial infarction (MI) remains unclear. Interleukin (IL)-5 is the most important cytokines that are responsible for the development, activation, and survival of eosinophils. Here, we report a role of IL-5 in infarcted heart. Methods and Results: MI was induced by permanent ligation of left anterior descending coronary artery in wild-type C57BL/6 mice. Western blot show that infarcted heart IL-5 expression is increased by 2.35-fold (P&lt;0.01) higher than their expression in the heart of sham mice at day 5 after surgery. External supply of recombinant mouse IL-5 (100 μg/kg/day for 3 days) reduces the infarct size (by 13.38%, P&lt;0.01), increases ejection fraction (from 32.41% to 43.93%, P&lt;0.01), and increases angiogenesis (CD31 area from 3.35% to 4.14%, P=0.03) in the peri-infarct zone. An expansion of eosinophils is observed in both the peripheral blood and infarcted myocardium after IL-5 administration. Pharmacological depletion of eosinophils by TRFK5 pretreatment mutes the beneficial effects of IL-5 in MI mice. Mechanistic studies also demonstrate that IL-5 increases the accumulation of M2 macrophages in infarcted myocardium at day 7 post-MI. In vitro co-culture experiments reveal that eosinophils shift bone marrow-derived macrophage polarization towards the M2 phenotypes. This effect is abolished by IL-4 neutralizing antibody, but not IL-10 or IL-13 neutralization. Western blot analyses show that eosinophils promote the macrophage downstream STAT6 phosphorylation. Conclusions: We demonstrate here that IL-5 facilitates the recovery of post-MI cardiac function by promoting eosinophils accumulation and subsequent polarization of M2 macrophages via the IL-4/STAT6 axis.

A facile and efficient protocol has been developed for preparing thiostibines via direct Sb–S cross-coupling of halostibines containing azastibocine frameworks with thiols under additive-free conditions. Additionally, a zinc(0)-mediated transformation of...

Endothelium-derived nitric oxide (NO) produced from endothelial NO-synthase (eNOS) is one of the most important vasoprotective molecules in cardiovascular physiology. Dysfunctional eNOS such as uncoupling of eNOS leads to decrease in NO bioavailability and increase in superoxide anion (O2.−) production, and in turn promotes cardiovascular diseases. Therefore, appropriate measurement of NO and O2.− levels in the endothelial cells are pivotal for research on cardiovascular diseases and complications. Because of the extremely labile nature of NO and O2.−, it is difficult to measure NO and O2.− directly in a blood vessel. Numerous methods have been developed to measure NO and O2.− production. It is, however, either insensitive, or non-specific, or technically demanding and requires special equipment. Here we describe an adaption of the fluorescence dye method for en face simultaneous detection and visualization of intracellular NO and O2.− using the cell permeable diaminofluorescein-2 diacetate (DAF-2DA) and dihydroethidium (DHE), respectively, in intact aortas of an obesity mouse model induced by high-fat-diet feeding. We could demonstrate decreased intracellular NO and enhanced O2.− levels in the freshly isolated intact aortas of obesity mouse as compared to the control lean mouse. We demonstrate that this method is an easy technique for direct detection and visualization of NO and O2.− in the intact blood vessels and can be widely applied for investigation of endothelial (dys)function under (physio)pathological conditions.

Abstract Purpose Recent decades have been witnessing that heart failure with preserved ejection fraction (HFpEF) outweighs heart failure with reduced ejection fraction by degrees, but few drugs were proven to improve long-term clinical outcomes in patients with HFpEF. Levosimendan, a calcium sensitizing cardiotonic agent, has been found to improve decompensated heart failure clinically. However, the protective activities and underlying molecular mechanisms of levosimendan on HFpEF have not been revealed. Methods The double-hit HFpEF C57BL/6N mouse model was established, and levosimendan (3 mg/kg/week) was administered to HFpEF mice aged from 13 to 17 weeks to verify its protective effects on HFpEF. The myocardium was biochemically evaluated by western blot, immunofluorescence, flow cytometry, etc. Results After four-week administration, cardiac hypertrophy, pulmonary congestion, and exercise exhaustion were significantly alleviated. Meanwhile, junction proteins located in endothelial barrier and between cardiomyocytes were improved by levosimendan treatment. Among the gap junction channel proteins concerned, connexin 43, especially expressed on cardiomyocytes, could conduct mitochondrial protection. Furthermore, levosimendan indeed reversed mitochondrial malfunction in HFpEF mice, evidenced by increased mitofilin and decreased ROS, superoxide anion, NOX4 and cytochrome C levels. Interestingly, after levosimendan treatment, myocardium from HFpEF mice showed restricted ferroptosis, indicated by upregulated GSH/GSSG ratio, GPX4, xCT and FSP-1 expression with down-regulated intracellular ferrous ion, MDA and 4-HNE. Conclusion This study confirmed that regular long-term levosimendan administration could benefit HFpEF individuals, particularly those with metabolic syndrome, such as obesity and hypertension, by activating connexin 43-conducted mitochondrial protection and sequential ferroptosis inhibition in cardiomyocytes.

Abstract The authors have requested that this preprint be removed from Research Square.

Tongxinluo (TXL), a widely used traditional Chinese medicine in cardiovascular diseases, has anti-inflammatory and anti-apoptosis effects. However, no studies have explored whether translanted bone-marrow-derived mesenchtmal stem cells pretreated by TXL (MSCTXL) can enhance survival of MSCs thus

Abstract Backgrounds : Acute myocardial infarction (AMI) is the predominant cause of cardiac death and ischemic heart failure (IHF) worldwide in coronary artery disease (CAD). Although it results from coronary acute occlusion, we in the study explored some key genes involved in the development of AMI and consequent IHF using bioinformatics analysis. Methods Utilizing expression data of 52 patients with AMI and 53 controls from GSE66360 and GSE97320 datasets, we screened shared differentially expressed genes (DEGs) in the independent datasets. Functional enrichment analysis and protein-protein interaction (PPI) network were employed. GSE58967 of 111 AMI patients and 46 controls was used to validate the shared DEGs and further analyzed to identify the DEGs in AMI patients with and without heart failure (HF) with the dynamic changes also being evaluated. The receiver operating characteristic (ROC) curves and area under the curve (AUC) were used to validate the diagnostic efficiency. Results In the comparison of AMI patients with controls, we identified 105 shared DEGs. Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis showed the shared DEGs mainly enriched in immune-related inflammation process and pathways. Filtered with PPI network, 5 genes of CXCL8 , CXCL1 , MMP9 , FPR1 and TLR2 were considered as hub genes, which were further validated in GSE59867. Compared with the genes in AMI patients without HF, those of TNFAIP6 , ADM , TRIB1 , AQP9 and IL1R2 associated with ventricular remodeling were found to be significantly high expressed in patients with HF on admission with the AUC of ROC curves was 0.792–0.847 (all p &lt; 0.05), which can be used as the potential biomarkers for early prediction of HF after AMI. Conclusions These findings based on integrated bioinformatic analysis provide new insights into the important roles of genes to play in the patients with AMI and consequent HF.

Abstract Background Despite the improved survival observed in PD-1/PD-L1 blockade therapy, there still is a lack of response to the anti-PD1 therapy for a large proportion of cancer patients across multiple indications, including non-small cell lung cancer (NSCLC) Methods Transcriptomic profiling was performed on 57 whole blood samples from 31 NSCLC patients and 5 healthy donors, including both responders and non-responders received anti-PD-1 Tislelizumab plus chemotherapy, to characterize differentially expressed genes (DEGs), signature pathways, and immune cell subsets regulated during treatment. Mutations of oncogenic drivers were identified and associated with therapeutic outcomes in a validation cohort with 1661 cancer patients. These multi-level biomarkers were validated and compared across different methods, external datasets and multiple computational tools. Results NSCLC patients examined and achieved pathological complete response (pCR) were considered as responders or non-responders otherwise. Expression of hundreds DEGs (FDR p&lt;0.05, fold change&lt;-2 or &gt;2) was changed in blood during neoadjuvant anti-PD-1 treatment, as well as in lung cancer tissue as compared to normal samples. Enriched PD-1-mediated pathways and elevated cell abundances of CD8 T cells and regulatory T cells were exclusively observed in responder blood samples. In an independent validation cohort of 1661 pan-cancer patients, a panel of 4 top ranked genetic alterations (PTCH1, DNMT3A, PTPRS, JAK2) identified from responders in discovery cohort were found positively associated with the overall survival (p&lt;0.05). Conclusion These findings suggest peripheral blood-based biomarkers and cell subsets could be utilized to define the response to neoadjuvant PD-1 blockade in NSCLC patients and a set of novel gene mutations is strongly associated with the therapeutic outcome of cancer immunotherapy.