| Background:Bone morrow-derived mesenchymal stem cells (BM-MSCs) have been the optimal candidate cells for cardiovascular regeneration in recent years; however, the poor survival of the implanted cells because of the harsh environment hampered the progress of the treatment. Our previous studies have displayed statins could improve the post-infarct microenvironment to enhance the survival of implanted bone marrow-MSCs, however the exact mechanism remain to be investigated. There has been evidence demonstrating that statins could protect cells from hypoxia and serum-free (H/SF) through the increase in endothelial nitric oxide synthase (eNOS) activity. AMP-activated protein kinase (AMPK) is essential to keep the balance of energy production and metabolism in various tissues, which is the dominant factor to modulate apoptosis. Based on these, we hypothesized that atorvastatin could protect MSCs against H/SF injury through AMPK-eNOS pathway.Methods:BM-MSCs separated from Chinese mini-swine were exposed to hypoxia and serum-free (H/SF). Treated with different concentration of atorvastatin (Ator, 0.001μM-10μM) or the inhibitor of AMPK (compound C, CC, and 10μM), the apoptosis rate of BM-MSCs and protein pathways were analyzed.Results:Staining with AnnexinV/propidine iodine (PI), showed atorvastatin (0.001μM-10μM) inhibited apoptosis of porcine bone marrow-derived MSCs induced by H/SF condition; however, which was blocked by compound C, an inhibitor of AMPK. The similar trend was observed as to bax protein expression by western blotting; whereas the bcl-2 protein, an anti-apoptosis protein, increased in atorvastatin group. Meanwhile, phosphorylation of AMPK and eNOS in MSCs treated with atorvastatin was shown to increase, which was also inhibited by compound C.Conclusions:Atorvastatin can activate AMPK, thus resulting in activation of eNOS, which provides a novel explanation for the protective effects of statins on BM-MSCs. Aims:To investigate whether AMP-activated protein kinase (AMPK) involved in the effects of atorvastatin (Ator) improving the effect of BM-MSCs transplantation in treating acute myocardial infarction (AMI).Methods:Forty-two Chinese mini-porcine were randomized into 6 groups of sham (n=7), AMI(control; n=7), Ator(atorvastatin; n=7), MSCs(BM-MSCs transplantation; n=7), Com(combined treatment; n=7) and L-NNA(Ator+MSCs+L-NNA; n=7).90 minutes after temporary coronary ligation, animalsin groups of MSCs, Com and L-NNA received autologous BM-MSCs transplantation through intra-myocardial injection into the peri-infarcted areas and followed 4 weeks for functional and structural end points.Results:As evaluated by positron emission topography (PET), compared with AMI group, combined treatment reduced the infarct size by 10.71cm2 [95% confidence interval (CI),4.88-16.55cm2, P=0.001]. As analyzed by magnetic resonance imaging(MRI), combined treatment improved the left ventricular ejection fraction(LVEF) by 13.54%[95% CI,9.30-17.78%, P<0.001] and left ventricular stroke volume (LVSV) by 8.88ml/beat [95% CI,5.88-11.87ml/beat, P<0.001] compared with controls. Although MSCs transplantation therapy reduced myocardial lesion area by 5.33% [95% CI,1.87-8.80%, P=0.004], LVEF increased by 5.49% [95% CI,1.25-9.73%, P=0.013], Ator treatment had no effect on the improvement of the infarct size and LVEF (P>0.05). Tissue chemistry analyzation displayed that Ator increased the activity of No and NOS in the infarcted arounding area (P<0.05). As analyzed by western blotting, phosphorylation of AMPK increased in Ator treated animals comparing with untreated animals, especially in the Com group (P<0.001). It showed a positive relationship between phosphorylation of AMPK and cardiac function (Pearson Correlation:0.54, P=0.002). Meanwhile, Ator increased the phorsphoralytion of Akt, and it had positive relationship with the phosphorylation of eNOS (Pearson Correlation:0.479, P=0.018). Similarly, Ator raised the c-Jun and phorsphoralytion of JNK, and both of they showed a positive relationship (Pearson Correlation:0.439, P=0.032). Although Ator increased p53 level, it had no significently corelationship with either AMPK phosphoralytion or phorsphoralytion of JNK (Pearson Correlation:0.360, P=0.051; Pearson Correlation:0.204, P=0.279).Conclusion:Atorvastatin improved the effecacy of BM-MSCs transplantation in treating AMI. The mechanisms of this cardioprotective effect of Ator involved the AMPK/eNOS, Akt/eNOS and JNK/c-Jun/p53 pathways. This provides a novel explanation for the statins pleiotropic efficiency on cardiovascular system.
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