And To Improve The Therapeutic Effect Of Acute Myocardial Infarction By Increasing The Homing And Survival Of Bone Marrow Mesenchymal Stem Cells

Combination of Loading dose of Atorvastatin and Atorvastatin-pretreated Mesenchymal Stem Cells Improves the Cardiac Performance after Acute Myocardial Infarction by Targeting Both SDF-1 and CXCR4Background:Stem cell therapy is a promising new approach for the treatment of ischemic heart diseases. The therapeutic potential of MSCs for cardiac repair is hampered partly due to their low engraftment rate within the ischemic myocardial. The interaction between stromal cell-derived factor 1 (SDF-1) and its receptor chemokine CXC motif receptor 4 (CXCR4) plays a vital role in the engraftment of mesenchymal stem cells (MSCs) in the injured myocardium. Our previous studies found that loading dose of atorvastatin (ATV) can enhance the survival of MSCs under ischemic microenvironment. Our recent findings showed that pretreatment of MSCs with ATV might improve the CXCR4 expressions. This study aims to investigate whether the combination of loading dose of ATV and ATV-pretreated MSCs can promote the engraftment of MSCs and enhance the cardiac performance in acute myocardial infarction (AMI) through SDF-1/CXCR4 axis.Methods:In the first part, female Sprague-Dawley were randomized into Sham, AMI control and loading dose of ATV group. To evaluate the expression change of SDF-1, lday, lweek and 2 weeks after AMI, expressions of SDF-1 in peri-infarcted heart were evaluated using RT-PCR, ELISA and immunohistochemistry. The time point with highest expression of SDF-1 was used in the second part for MSCs transplantation. In the second part, rats were randomized into eight groups:Sham, AMI control, MSCs, loadiong dose of ATV, loading dose of ATV+MSCs, ATV-pretreated MSCs (ATV-MSCs), loading dose of ATV+ATV-pretreated MSCs (ATV+ATV-MSCs), loading dose of ATV+ATV-pretreated MSCs+AMD3100 (SDF-1/CXCR4 antagonist) (ATV+ATV-MSCs+AMD3100). MSCs or ATV-pretreated MSCs labeled with CM-Dil were injected through the jugular vein. Cardiac functions were assessed using echocardiography and left heart catheterization. Recruitment of MSCs to the infarcted heart, inflammatory cells and fibrosis were evaluated with histopathology. Apoptosis was detected with TUNEL kit, and protein microarray was employed to measure the inflammatory cytokines. Arteriogenesis, angiogenesis, c-Kit+ stem cells numbers and differentiation of MSCs into cardiomyocytes were examined with immunofluorescence.Results:ATV significantly increased the SDF-1 expression in the peri-infarct area of the myocardium, reaching the peak at 1 week post AMI, which was used for MSCs transplantation. Cardiac performance was improved in ATV-MSCs and ATV+MSCs groups than MSCs group, evidenced by increased left ventricular ejection fraction (LVEF), left ventricular fractional shortening (LVFS) and dp/dt, as well as the decreased left ventricular end-diastolic diameter (LVEDd), left ventricular end-systolic diameter (LVESd) and left ventricular end-diastolic pressure (LVEDP). Compared with ATV-MSCs group, LVEF and LVFS were significantly improved in ATV+ATV-MSCs group. Changes in LVEDP and dp/dt in ATV+ATV-MSCs group were statistical significant compared with both ATV-MSCs and ATV+MSCs group, which were partly inhibited by AMD3100. Protein microarray analysis demonstrated that, compared with the AMI group, ATV+ATV-MSCs group significantly reduced the pro-inflammatory cytokines IL-1α、 IL-1β、IL-6 and TNF-a, while increased the anti-inflammatory cytokine IL-10. Histological analysis showed that compared with ATV-MSCs group, the left ventricular fibrotic area was markedly reduced in ATV+ATV-MSCs group, and the inflammatory cells were obviously attenuated. ATV combined with ATV-MSCs facilitated the recruitment of MSCs in the infarcted myocardium. This was accompanied by reduced apoptosis and enhanced arteriogenesis and angiogenesis. The administration of AMD3100 abolished the protective effect of combined use of loading dose of ATV and ATV-MSCs. Compared to the AMI group, transplantation of MSCs markedly increased the the number of c-Kit+ stem cells in the peri-infarcted myocardium, which was further upregulated with the combined use of loading dose of ATV and ATV-MSCs. Nevertheless, the differention of MSCs into cardiomyocytes did not differ between groups.Conclusions:Administration of loading dose of ATV combined with ATV-pretreated MSCs can increase the recruitment of MSCs and improve the cardiac performance by targeting SDF-1/CXCR4 axis. This may attribute to the inhibition of inflammation, apoptosis and fibrosis, the upregulation of c-Kit+ stem cells and the promotion of arteriogenesis and angiogenesis. These results suggest that combined use of loading dose of ATV and ATV pretreatment of donor MSCs is an effective way to promote cell therapeutic potential for AMI.Globular Adiponetin Inhibits the Apoptosis of Mesenchymal Stem Cells under Hypoxia and Serum Deprivation Conditions via the AdipoRl/AMPK PathwayBackground:Poor viability of transplanted mesenchymal stem cells (MSCs) within the ischemic heart has limited their therapeutic potential for cardiac repair. Hypoxia and serum deprivation (H/SD) was widely used to mimick the ischemic myocardium microenvironment and induced MSCs apoptosis through the mitochondrial apoptotic pathway. Adiponectin (APN) is an adipocyte-secreted adipokine with pleiotropic actions. It has recently been proposed to participate in tissue regeneration and promote the survival of several other stem cells. We embarked on investigating the role of APN in the prevention of MSCs against H/SD-induced apoptosis and elucidated the potential mechanisms involved.Methods:Rat bone marrow MSCs were exposed to H/SD with different concentrations (0,0.01,0.1,1 ug/ml) of globular APN (gAPN) in a sealed GENbox hypoxic chamber for 6 hours. MSCs were transfected with adiponectin receptors 1 or 2 (AdipoRl or AdipoR2) siRNAs to determine the major receptor type that mediated the actions of gAPN. For pathway studies, the cells were pre-incubated with AMP-activated protein kinase (AMPK) inhibitor Compound C. Apoptosis was detected by measuring Hoechst 33342 positive cells using a fluorescent microscope and Annexin V positive cells using flowcytometry. Caspase-3 activity was measured using a Caspase-3/CPP32 Colorimetric assay kit. Mitochondrial dysfunction was analyzed by assessing the red over green fluorescence intensity with JC-1 staining kit. The expressions of apoptosis-related proteins Bcl-2, Bax, Pro-caspase-3 and Cleaved-caspase-3 were examined using Western blot. The phosphorylation of AMPK was also evaluated with Western blot.Results:The data showed that H/SD-induced apoptosis was attenuated by gAPN in a concentration-dependent manner evidenced by reduced Annexin V+ cells and decreased caspase-3 activity. This protective effect of gAPN was mainly mediated through AdipoR1. Moreover, gAPN increased the phosphorylation of AMPK and Compound C effectively reversed the anti-apoptotic effect of gAPN. Furthermore, gAPN significantly decreased H/SD-induced Cytochrome C release and Bax translocation into mitochondria. The inhibition of the AdipoRl or AMPK pathway partly abolished the effect of gAPN on mitochondrial dysfunction.Conclusion:These data demonstrated that globular APN prevents H/SD - induced apoptosis and mitochondrial dysfunction in MSCs via adipoRl linked to the activation of downstream AMPK pathway. Our findings indicate that gAPN is a novel, potent survival factor for MSCs in the ischemic myocardium.