M3-mAChR Stimulation Exerts Anti-apoptotic Effect Via Activating the HIF-1α/HO-1/VEGF Signaling Pathway in H9c2 Rat Ventricular Cells

Background: The protective role of M-3-mAChR against apoptosis has been identified previously. However, the underlying mechanisms remain unclear. This study was performed to clarify the signaling pathways of the anti-apoptotic effect mediated by activation of M-3-mAChR in cultured cardiac H9c2 cells. Methods: Both H9c2 rat ventricular cells and H9c2 cells with stable expression of M-3-mAChR were used. Results: Activation of M-3-mAChR by cabarchol produced protective effect on etoposide-induced apoptosis in H9c2 cells. Forced overexpression of M-3-mAChR in H9c2 cells further enhanced this effect. Application of 4-diphenyl-acetoxy-N-methyl-piperidine methiodide (inhibitor of M-3-mAChR), YC-1 [inhibitor of hypoxia-inducible factor 1, (HIF-1], or ZnPP (inhibitor of heme oxygenase-1) abrogated carbacol-induced cardioprotection, respectively. Moreover, the expression of HIF-1 alpha, HO-1, and vascular endothelial growth factor (VEGF) were enhanced after the activation of M-3-mAChR, and the induction of HO-1 and VEGF was reversed by HIF-1 alpha inhibitor YC-1. Conclusions: These findings indicated that M-3-mAChR upregulates HO-1 and VEGF expression likely through induction of HIF-1 alpha, which at least partly underlies the cytoprotection of M-3-mAChR activation in H9c2 cells.