Shengshou Hu

MicroRNA-24 regulates cardiac fibrosis after myocardial infarction

Cardiac fibrosis after myocardial infarction (MI) has been identified as a key factor in the development of heart failure. Although dysregulation of microRNA (miRNA) is involved in various pathophysiological processes in the heart, the role of miRNA in fibrosis regulation after MI is not clear. Previously we observed the correlation between fibrosis and the miR‐24 expression in hypertrophic hearts, herein we assessed how miR‐24 regulates fibrosis after MI. Using qRT‐PCR, we showed that miR‐24 was down‐regulated in the MI heart; the change in miR‐24 expression was closely related to extracellular matrix (ECM) remodelling. In vivo, miR‐24 could improve heart function and attenuate fibrosis in the infarct border zone of the heart two weeks after MI through intramyocardial injection of Lentiviruses. Moreover, in vitro experiments suggested that up‐regulation of miR‐24 by synthetic miR‐24 precursors could reduce fibrosis and also decrease the differentiation and migration of cardiac fibroblasts (CFs). TGF‐β (a pathological mediator of fibrotic disease) increased miR‐24 expression, overexpression of miR‐24 reduced TGF‐β secretion and Smad2/3 phosphorylation in CFs. By performing microarray analyses and bioinformatics analyses, we found furin to be a potential target for miR‐24 in fibrosis (furin is a protease which controls latent TGF‐β activation processing). Finally, we demonstrated that protein and mRNA levels of furin were regulated by miR‐24 in CFs. These findings suggest that miR‐24 has a critical role in CF function and cardiac fibrosis after MI through a furin–TGF‐β pathway. Thus, miR‐24 may be used as a target for treatment of MI and other fibrotic heart diseases.

Isolated Coronary Artery Bypass Graft Combined With Bone Marrow Mononuclear Cells Delivered Through a Graft Vessel for Patients With Previous Myocardial Infarction and Chronic Heart Failure: A Single-Center, Randomized, Double-Blind, Placebo-Controlled Clinical Trial

OBJECTIVES This study aimed at examining the efficacy of bone marrow mononuclear cell (BMMNC) delivery through graft vessel for patients with a previous myocardial infarction (MI) and chronic heart failure during coronary artery bypass graft (CABG). BACKGROUND Little evidence exists supporting the practice of BMMNC delivery through graft vessel for patients with a previous MI and chronic heart failure during CABG. METHODS From November 2006 to June 2009, a randomized, placebo-controlled trial was conducted to test the efficacy and safety of CABG for multivessel coronary artery disease combined with autologous BMMNCs in patients with congestive heart failure due to severe ischemic cardiomyopathy. Sixty-five patients were recruited, and 60 patients remained in the final trial and were randomized to a CABG + BMMNC group (n = 31) and a placebo-control group (i.e., CABG-only group, n = 29). All patients discharged received a 6-month follow-up. Changes in left ventricular ejection fraction from baseline to 6-month follow-up, as examined by magnetic resonance imaging, were of primary interest. RESULTS The overall baseline age was 59.5 ± 9.2 years, and 6.7% were women. After a 6-month follow-up, compared with the placebo-control group, the CABG + BMMNC group had significant changes in left ventricular ejection fraction (p = 0.029), left ventricular end-systolic volume index (p = 0.017), and wall motion index score (p = 0.011). Also, the changes in the distance on the 6-min walking test as well as B-type natriuretic peptide were significantly greater in the CABG + BMMNC group than in the control group. CONCLUSIONS In summary, patients with a previous MI and chronic heart failure could potentially benefit from isolated CABG (i.e., those who received CABG only) combined with BMMNCs delivered through a graft vessel. (Stem Cell Therapy to Improve Myocardial Function in Patients Undergoing Coronary Artery Bypass Grafting [CABG]; NCT00395811).

Lovastatin protects mesenchymal stem cells against hypoxia- and serum deprivation-induced apoptosis by activation of PI3K/Akt and ERK1/2.

Cell therapy with bone marrow‐derived mesenchymal stem cells (MSCs) has been shown to have great promises in cardiac repair after myocardial infarction. However, poor viability of transplanted MSCs in the infracted heart has limited the therapeutic efficacy. Our previous studies have shown in vitro that rat MSCs undergo caspase‐dependent apoptosis in response to hypoxia and serum deprivation (Hypoxia/SD). Recent findings have implicated statins, an established class of cholesterol‐lowering drugs, enhance the survival of cells under various conditions. In this study, we investigated the effect of lovastatin on rat MSCs apoptosis induced by Hypoxia/SD, focusing in particular on regulation of mitochondrial apoptotic pathway and the survival signaling pathways. We demonstrated that lovastatin (0.01–1 µM) remarkably prevented MSCs from Hypoxia/SD‐induced apoptosis through inhibition of the mitochondrial apoptotic pathway, leading to attenuation of caspase‐3 activation. The loss of mitochondrial membrane potential and cytochrome‐c release from mitochondria to cytosol were significantly inhibited by lovastatin. Furthermore, the antiapoptotic effect of lovastatin on mitochondrial apoptotic pathway was effectively abrogated by both PI3K inhibitor, LY294002 and ERK1/2 inhibitor, U0126. The phosphorylations of Akt/GSK3β and ERK1/2 stimulated by lovastatin were detected. The activation of ERK1/2 was inhibited by a PI3K inhibitor, LY294002, but U0126, a ERK1/2 inhibitor did not inhibit phosphorylation of Akt and GSK3β. These data demonstrate that lovastatin protects MSCs from Hypoxia/SD‐induced apoptosis via PI3K/Akt and MEK/ERK1/2 pathways, suggesting that it may prove a useful therapeutic adjunct for transplanting MSCs into damaged heart after myocardial infarction. J. Cell. Biochem. 103: 256–269, 2008.

Stenting technique, gender, and age are associated with cardioprotection by ischaemic postconditioning in primary coronary intervention: a systematic review of 10 randomized trials

AIMS We sought to perform a systematic review and meta-analysis to evaluate the potential factors affecting ischaemic postconditioning (IPoC) for patients with ST-segment elevation acute myocardial infarction (STEMI) in primary percutaneous coronary intervention (PCI). METHODS AND RESULTS Ten randomized controlled trials (RCTs) on IPoC reporting myocardial enzyme levels or left ventricular ejection fraction (LVEF) in a total of 560 STEMI patients were identified in PubMed, EMBase, and Cochrane Library (up to February 2012). Compared with controls, IPoC significantly reduced elevated cardiac enzyme levels [standardized mean difference = -0.84; 95% confidential interval (CI): -1.26 to -0.43; P < 0.00001; heterogeneity test, I(2) = 81.0%] and improved LVEF [weighted mean difference (WMD) = 3.98%; 95% CI: 1.27-6.70%; P = 0.004; heterogeneity test, I(2) = 87.1%]. The effect on LVEF remained significant after 1 year (WMD = 5.04%; 95% CI: 4.20-5.88%; P < 0.00001; heterogeneity test, I(2) = 0.0%). Univariate meta-regression analysis suggested that the major sources of significant heterogeneity (P < 0.1) were the use of direct-stenting technique (%) (coefficient = -0.886; P = 0.069; adjusted R(2) = 0.34) and male proportion (%) (coefficient = -0.022; P = 0.098; adjusted R(2) = 0.28) for myocardial enzyme levels, and age (coefficient = -1.34; P = 0.025; adjusted R(2)= 0.55) for LVEF (%). Subsequent multivariate regression and subgroup analysis confirmed these results. CONCLUSION Available evidence from this systematic review and meta-analysis of 10 RCTs suggests that IPoC may confer cardioprotection in terms of myocardial enzyme levels and LVEF for STEMI during primary PCI. These effects are more pronounced among young and male patients, and those in whom direct-stenting techniques were used. Future studies should focus on the mortality in high-quality, large-scale clinical trials with long-term follow-up.

Extracorporeal membrane oxygenation for treatment of cardiac failure in adult patients

This report reviews our experience in venoarterial extracorporeal membrane oxygenation (ECMO) support treatment in adult patients with cardiac failure, as well as analysis of the risk factors of early mortality. From February 2005 to June 2008, 45 patients undergoing cardiogenic shock required temporary ECMO support. They were divided into three groups: post-cardiotomy (n=31) and post-transplantation (n=5) heart failure, decompensated heart failure (n=9). ECMO implantation was performed through the femoral vessels, or axillary artery, or through the right atrium and ascending aorta. Average support duration was 126.7+/-104.3 h. Twenty-seven patients could be successfully weaned from support (60%); additionally, five were bridged to heart transplantation. The in-hospital mortality was 42% (19/45). Twenty-six patients (58%) could be successfully discharged. Additional intra-aortic balloon pumps were used in 11 patients, and six of them were successfully discharged. The mortality rate was obviously high for patients with acute renal failure treated by continuous renal replacement therapy (CRRT) under ECMO support (7/9 patients). The dominant mode of death was multisystem organ failure (9/19). ECMO offers effective cardiopulmonary support in adults. The better outcome requires a multidisciplinary approach to prevent complications unique to itself and limit organ injury before and during this support.

Expression of MicroRNA-1 and MicroRNA-21 in Different Protocols of Ischemic Conditioning in an Isolated Rat Heart Model

Background: ‘Conditioning’ [ischemic preconditioning (IPC), ischemic postconditioning (IPO) and remote ischemic preconditioning (RIPC)] the heart to render it more resistant to an episode of acute myocardial ischemia-reperfusion (I/R) injury is an endogenous cardioprotective strategy. There are several mechanisms proposed for ‘conditioning’, such as endogenous mediators or cytoprotective proteins. In recent reports, microRNAs (miRNAs) were involved in controlling the expression of myocardial ischemia-related genes. Some studies have demonstrated that cardiac miRNA-1 and miRNA-21 were significantly increased by late IPC with an increase in their target proteins [endothelial nitric oxide synthase and heat shock protein 70 (HSP70)], but their expression levels in ‘conditioning’ strategies are currently unknown. Methods: In the current study, Langendorff-perfused Sprague-Dawley rat hearts were randomly assigned to one of four groups [control group (CON group, n = 12), IPC group (n = 12), IPO group (n = 12) and RIPC group (n = 12)]. Cardiac function was digitalized and analyzed. The expression of miRNA-1 and miRNA-21 was detected by real-time reverse transcription polymerase chain reaction. The expression of HSP70, programmed cell death protein 4 (PDCD4), B-cell lymphoma/leukemia-2 (Bcl-2) and Bcl-2-associated X protein (Bax) was detected by Western blot. Cardiac infarct size and myocardial apoptosis were determined using the 2,3,5-triphenyltetrazolium chloride assay and terminal deoxynucleotidyl transferase dUTP nick end labeling assay, respectively. Results: The results revealed that miRNA-1 (233 ± 45%) and miRNA-21 (356 ± 33%) expression was up-regulated in the IPC group, but the expression of miRNA-1 was down-regulated in the RIPC (61 ± 16%) group and IPO group (61 ± 13%). The expression of PDCD4 [IPC (74 ± 11%), RIPC (81 ± 16%), IPO (83 ± 12%)], HSP70 [IPC (74 ± 5%), RIPC (81 ± 6%), IPO (67 ± 11%)] and Bax [IPC (27 ± 6%), RIPC (21 ± 3%), IPO (27 ± 4%)] was down-regulated in the conditioning groups compared with the CON group [PDCD4 (130 ± 11%), HSP70 (121 ± 11%) and Bax (63 ± 8%)]. In the conditioning hearts, infarct size [IPC (31.7 ± 4.1%), RIPC (29.6 ± 6.19%) and IPO (32.8 ± 4.71%)] and myocardial apoptosis [IPC (15.2 ± 4.21%), RIPC (17.2 ± 1.92%) and IPO (15.6 ± 4.04%)] were significantly decreased compared with the CON group (infarct size: 51.77 ± 4.3%, myocardial apoptosis: 32.8 ± 3.96%). Conclusion: We concluded that miRNA-1 and miRNA-21 expression differed in IPC, RIPC and IPO groups, and their target proteins were not inversely correlated with the miRNAs in all the conditioning groups, which revealed that the miRNAs were regulated but complicated by the different conditioning protocols.

In Vitro Effects of Pirfenidone on Cardiac Fibroblasts: Proliferation, Myofibroblast Differentiation, Migration and Cytokine Secretion

Cardiac fibroblasts (CFs) are the primary cell type responsible for cardiac fibrosis during pathological myocardial remodeling. Several studies have illustrated that pirfenidone (5-methyl-1-phenyl-2-[1H]-pyridone) attenuates cardiac fibrosis in different animal models. However, the effects of pirfenidone on cardiac fibroblast behavior have not been examined. In this study, we investigated whether pirfenidone directly modulates cardiac fibroblast behavior that is important in myocardial remodeling such as proliferation, myofibroblast differentiation, migration and cytokine secretion. Fibroblasts were isolated from neonatal rat hearts and bioassays were performed to determine the effects of pirfenidone on fibroblast function. We demonstrated that treatment of CFs with pirfenidone resulted in decreased proliferation, and attenuated fibroblast α-smooth muscle actin expression and collagen contractility. Boyden chamber assay illustrated that pirfenidone inhibited fibroblast migration ability, probably by decreasing the ratio of matrix metalloproteinase-9 to tissue inhibitor of metalloproteinase-1. Furthermore, pirfenidone attenuated the synthesis and secretion of transforming growth factor-β1 but elevated that of interleukin-10. These direct and pleiotropic effects of pirfenidone on cardiac fibroblasts point to its potential use in the treatment of adverse myocardial remodeling.

MicroRNA-193 pro-proliferation effects for bone mesenchymal stem cells after low-level laser irradiation treatment through inhibitor of growth family, member 5.

The enhanced proliferation of mesenchymal stem cells (MSCs) can be helpful for the clinical translation of cell therapy. Low-level laser irradiation (LLLI) has been demonstrated as regulating MSC proliferation. MicroRNAs (miRNAs) are involved in various pathophysiologic processes in stem cells, but the role of miRNAs in the LLLI-based promotion of MSC proliferation remains unclear. We found that the proliferation level and cell cycle-associated genes in MSCs were increased after LLLI treatment in a time-dependent manner. Microarray assays revealed subsets of miRNAs to be differentially regulated, and these dynamic changes were confirmed by quantitative real-time polymerase chain reaction (qRT-PCR) after LLLI. miR-193 was the most highly up-regulated miRNA, and the change in it was related with the proliferation level. Gain-loss function experiments demonstrated that miR-193 could regulate the proliferation of MSCs, including humans and rats, but could not affect the apoptosis and differentiation level. Blockade of miR-193 repressed the MSC proliferation induced by LLLI. By qRT-PCR, we found that miR-193, in particular, regulated cyclin-dependent kinase 2 (CDK2) expression. Bioinformatic analyses and luciferase reporter assays revealed that inhibitor of growth family, member 5 (ING5) could be the best target of miR-193 to functionally regulate proliferation and CDK2 activity, and the mRNA and protein level of ING5 was regulated by miR-193. Furthermore, the ING5 inhibited by small interfering RNA (siRNA) could up-regulate the proliferation of MSCs and the expression of CDK2. Taken together, these results strongly suggest that miR-193 plays a critical part in MSC proliferation in response to LLLI stimulation, which is potentially amenable to therapeutic manipulation for clinical application.

Increasing Long-Term Major Vascular Events and Resource Consumption in Patients Receiving Off-Pump Coronary Artery Bypass A Single-Center Prospective Observational Study

Background— Despite its widespread use and short-term efficacy, substantial uncertainty remains about the long-term outcomes and cost-effectiveness of off-pump coronary artery bypass (OPCAB). Methods and Results— A retrospective review of prospectively collected data was conducted of 6665 consecutive patients undergoing isolated coronary artery bypass graft (CABG) at our institution during 1999 to 2006. All patients were followed up until September 30, 2008. Short- and long-term outcomes were compared between OPCAB and conventional CABG. The 2 main long-term outcome measures were repeat revascularization and the composite outcome of major vascular events. Cost comparison at 2 years in a propensity-matched sample during follow-up was also a study interest. The overall mean baseline age was 60.3±8.6 years, and 17.0% were women. Compared with conventional CABG, patients who underwent OPCAB had lower rates of atrial fibrillation (P=0.003) and requirements for blood transfusion (P=0.03) and ventilation time >24 hours (P<0.001). After an average of 4.5 years of follow-up, the rates of repeat revascularization (adjusted hazard ratio, 1.40; 95% confidence interval, 1.03 to 1.89) and major vascular events (adjusted hazard ratio, 1.23; 95% confidence interval, 1.09 to 1.39) were significantly higher in the OPCAB than the conventional CABG group. At 2 years, OPCAB was associated with increased additional direct costs per patient compared with conventional CABG and had a similar survival rate. Conclusions— Compared with conventional CABG, OPCAB is associated with small short-term gain but increased long-term risks of repeat revascularization and major vascular events, especially among high-risk patients. Moreover, OPCAB consumes more resources and is less cost-effective in the long run.

MicroRNA: Novel Regulators Involved in the Remodeling and Reverse Remodeling of the Heart

Background: MicroRNAs (miRNAs) may serve as potential diagnostic biomarkers and therapeutic targets in cardiovascular research. However, the association between miRNAs and heart remodeling/reverse remodeling has not been evaluated. Methods: Lewis rats were divided into three groups: control animals, animals subjected to abdominal aortic constriction (AAC) and those with heterotopic transplantation of abdominal aortic constriction (AAC-HT), respectively. The cardiomyocyte cross-sectional area and changes in the heart cavity were determined. miRNA microarray was used to search for changes in miRNAs during hypertrophy and in the unloading heart, which was also verified using real-time quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR). Results: The AAC-HT group exhibited a 54% decrease in the cardiomyocyte cross-sectional area compared to the AAC group. In the two test groups, miRNA microarrays revealed changes in 293 miRNAs, among which 40 miRNAs changed >2-fold. Some major changes were also confirmed using qRT-PCR primers. The results indicated that changes in miR-23a and miR-29a were most significant, thus suggesting that these miRNAs may play important roles in heart remodeling and reverse remodeling. Conclusions: The changes observed in miRNA expression during hypertrophy and reverse remodeling may indicate possibly meaningful targets for regulating the remodeling or reverse remodeling of the heart.