Abstract 14432: Electrical Stimulation Induces Cytoprotective Effects on Stem Cells By Connective Tissue Growth Factor Through FAK/AKT Signaling and microRNAs
Background: The survival of stem cells following transplantation in the infarcted myocardium is important in stem cell based therapy and preconditioning (PC) of stem cells is one of the most powerful stimulus for cytoprotection. In this study, we investigated whether PC by electrical stimulation (EleS) induces cytoprotective effect on cardiac stem cells (CSC) and determine its underlying molecular mechanisms.
Methods and results: Sca-1+ CSCs were isolated from male C57BL6 mice (12 weeks) hearts as described (PLoS One. 2011;6(9):e25265). PC of CSCs with EleS was carried out for 3 hr at 1.5 V followed by exposure to 300 µM H2O2 for 5 hrs . Cell damage was assessed by TUNEL assay and lactate dehyrogenase release which was reduced by 26.68±3.68 % and 22.18±6.5% resp compared to non-EleS. Increased phosphorylation of AKT, FAK and GSK3β, and decreased pro-apoptotic protein caspase-3 expression were observed in PC CSCs. Pretreatment with PI3-K inhibitor, Wortmannin (5 uM) and FAK inhibitor-14 (25 uM) abolished the pro-survival effects of Eles concomitant with reduction in AKT and FAK phosphorylation. RT-PCR based array for adhesion molecules showed significant increase in connective tissue growth factor (Ctgf) by EleS. CTGF upregulation by EleS was validated by RT-PCR, immunocytochemistry, and Western blotting. Transplantation of PC male CSCs in the infarcted heart of female mice resulted in increased survival as evaluated by Sry gene expression (1.6 fold increase vs. non-EleS). Transplantation of PC CSCs with Ctgf siRNA abolished prosurvival effect of PC. We further found that miR-322, -199a-5p, -30 a,b,e, 107, -199a-3p, -103, -145, -10a, and -15b were significantly downregulated by EleS. Computational analysis also indicated these miRNAs directly bind to 3’ untranslated region of Ctgf gene, suggesting these miRNAs may participate in Ctgf expression.
Conclusions: This study demonstrated that EleS enhances CSC survival both under in vitro and in vivo models by CTGF/FAK signaling and AKT prosurvival pathway. Specific miRNAs targeting CTGF may play a significant role in CSC survival. It is concluded that EleS is a promising approach for preconditioning of stem cells to significantly enhance their survival.
- © 2012 by American Heart Association, Inc.