Silencing Rac1 Protects H9c2 Cells Against Hypoxia-Reoxygenation Injury By Inactivation Of Notch Pathway

This study was aimed to determine the regulatory landscape of ectopic Ras-related C3 botulinum toxin substrate 1 (Rac1) in cardiomyocytes with hypoxia-reoxygenation (H/R) injury. We constructed a cell model with H/R injury in H9c2 cardiomyocytes to simulate myocardial Ischemia reperfusion injury (MIRI). When Rac1 was overexpressed or silenced, 3-(4,5-dimethylthiazol -2-yl)-2,5-diphenyltetrazolium bromide (MTT) colorimetric assay and flow cytometry were used to detect cell viability, reactive oxygen species (ROS) content and apoptosis in transformed cells. Key proteins Notch 1, Notch 2 and Jagged 1 (JAG1) of the Notch signaling pathway and apoptosis mediators B-cell lymphoma-2 (Bcl-2) and Caspase-3 were observed. Reoxygenation treatment provoked decline in the H9c2 cell viability in a time-dependent manner and about 54.67% viability was identified for 12 h. Thereby, 12 h reoxygenation duration was determined in further studies. Silencing Rac1 could remarkably raise cell viability (P < 0.05), suppress apoptosis (P < 0.05), scavenge ROS (P < 0.01) and inhibit expressions of key proteins of Notch signaling pathway, while overexpressing Rac1 had opposite effects on H/R-treated H9c2 cardiomyocytes (P < 0.05). Furthermore, silencing Rac1 suppressed H9c2 cell apoptosis by increasing Bcl-2 and decreasing cleaved Caspase-3. Collectively, our findings suggest that Rac1 acts as a key mediator of cardiomyocytes survival and responding oxidant stress after H/R injury. This study provides novel insight into the MIRI suppressive capacity of silencing Rac1 by inactivation of Notch signaling pathway, which holds great clinical potential in the cardioprotection against MIRI.