Protective Effect of Fasudil on Hydrogen Peroxide-Induced Oxidative Stress Injury of H9C2 Cardiomyocytes.

Objective: Oxidative damage is a pathological factor that causes cardiovascular damage in the clinic and is increasingly serious. This study focused on the effect of fasudil on H ₂ O ₂ -induced oxidative damage in cardiomyocytes.
Materials and Methods: H9C2 cardiomyocytes were cultured in vitro and divided into three groups: control group (Con group), H ₂ O ₂ treatment (H ₂ O ₂ group), and fasudil and H ₂ O ₂ cotreatment (H ₂ O ₂ +fasudil group). The content levels of LDH and MDA in the supernatant were detected, and the morphology of H9C2 cardiomyocytes was observed by light microscopy. 8-OHdG staining was observed by a fluorescence inversion microscope. Cell Counting Kit (CCK-8), western blotting, real-time polymerase chain reaction (RT-PCR), and enzyme-linked immunosorbent assay (ELISA) were used to investigate the effect of fasudil on the Rho/ROCK signaling pathway.
Results: Our results showed that after H ₂ O ₂ treatment, the H9C2 cardiomyocytes were irregular in shape and elliptical. But the morphology of the H ₂ O ₂ +fasudil group was similar to that of the Con group. The green fluorescence of the H ₂ O ₂ group was significantly enhancer than that of the Con group, while the green fluorescence of the H ₂ O ₂ +fasudil group was weaker than those of the H ₂ O ₂ group. By detecting the supernatant, it was found that the contents of LDH were significantly increased, and the contents of SOD and CAT in the H ₂ O ₂ group were significantly decreased. And the expression of antioxidant indicators in the H ₂ O ₂ group was significantly decreased by western blotting. The results of RT-PCR showed that SOD1 and SOD2 mRNA in the H ₂ O ₂ group was significantly reduced, and the contents of GPX1 and GPX3 in the H ₂ O ₂ group were significantly decreased by enzyme-linked immunosorbent assay (ELISA). The expression of ROCK1, ROCK2, and downstream phosphorylation of myosin phosphatase target subunit-1 (p-MYPT-1) was significantly increased in the H ₂ O ₂ group, while fasudil inhibited the increase of ROCK1, ROCK2, and p-MYPT-1.
Conclusions: Fasudil can inhibit the Rho/ROCK signaling pathway induced by H ₂ O ₂ and reduce oxidative stress response, inhibit apoptosis, and improve antioxidant enzyme activity in H9C2 cardiomyocytes thereby delaying cell senescence.
Competing Interests: The authors declared no conflict of interest.
(Copyright © 2021 Yu Zhang et al.)