Catalpol protects AC16 cells from hypoxia/reoxygenation injury by regulating the miR-22-3p/DPP4 axis.

Abstract

Catalpol (CA) is widely used in the protection of cardiomyocytes. Nevertheless, the mechanism of CA in alleviating ischemia-reperfusion-induced injury of cardiomyocytes remains unclear. Human cardiomyocyte AC16 cells were subjected to hypoxia/reoxygenation (H/R) injury. Quantitative real-time polymerase chain reaction (qRT-PCR) and western blot analysis were applied to detect tumor necrosis factor-alpha (TNF-α) mRNA, interleukin-6 (IL-6) mRNA, interleukin-1beta (IL-1β) mRNA, microRNA-22-3p (miR-22-3p), dipeptidyl peptidase 4 (DPP4) mRNA, and DPP4 protein expressions. The cell viability and apoptosis were measured by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay and flow cytometry, respectively. Lactate dehydrogenase (LDH) and creatine kinase (CK-MB) were examined by enzyme-linked immunosorbent assay (ELISA) kits. A dual-luciferase reporter gene assay was performed to confirm the binding sequence between miR-22-3p and DPP4 mRNA 3'-untranslated region (3'UTR). CA promoted the viability and reduced cell apoptosis of AC16 cells and repressed the release of inflammatory cytokines TNF-α, IL-6, and IL-1β, and inhibited the leakage of myocardial injury markers LDH and CK-MB. Furthermore, CA enhanced the expression of miR-22-3p in cardiomyocytes, and DPP4 was validated to be the target gene of miR-22-3p. The inhibition of miR-22-3p and augmentation of DPP4 reversed the above effects of CA. CA protects A16 cells from H/R injury by regulating the miR-22-3p/DPP4 axis.