Abstract
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 H2O2-induced oxidative damage in cardiomyocytes. Materials and Methods H9C2 cardiomyocytes were cultured in vitro and divided into three groups: control group (Con group), H2O2 treatment (H2O2 group), and fasudil and H2O2 cotreatment (H2O2+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 H2O2 treatment, the H9C2 cardiomyocytes were irregular in shape and elliptical. But the morphology of the H2O2+fasudil group was similar to that of the Con group. The green fluorescence of the H2O2 group was significantly enhancer than that of the Con group, while the green fluorescence of the H2O2+fasudil group was weaker than those of the H2O2 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 H2O2 group were significantly decreased. And the expression of antioxidant indicators in the H2O2 group was significantly decreased by western blotting. The results of RT-PCR showed that SOD1 and SOD2 mRNA in the H2O2 group was significantly reduced, and the contents of GPX1 and GPX3 in the H2O2 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 H2O2 group, while fasudil inhibited the increase of ROCK1, ROCK2, and p-MYPT-1. Conclusions Fasudil can inhibit the Rho/ROCK signaling pathway induced by H2O2 and reduce oxidative stress response, inhibit apoptosis, and improve antioxidant enzyme activity in H9C2 cardiomyocytes thereby delaying cell senescence.
Highlights
The rising trend of cardiovascular disease leading to human death is one of the main causes of sudden death in humans [1]
More studies have shown that oxidative stress- (OS-) induced cardiomyocyte apoptosis plays an important role in the development of cardiovascular diseases [4]
When the cells were at the appropriate density, the H2O2 group and the fasudil group were treated with 200 μmol/L H2O2, and the Con group was added with the same amount of Dulbecco’s modified Eagle’s medium (DMEM) for 24 h
Summary
The rising trend of cardiovascular disease leading to human death is one of the main causes of sudden death in humans [1]. As people’s quality of life improves, the incidence of many underlying diseases increases year by year, including diabetes, high blood pressure, and myocardial ischemia [2]. The incidence and mortality of cardiovascular diseases have remained at the leading level [3]. More studies have shown that oxidative stress- (OS-) induced cardiomyocyte apoptosis plays an important role in the development of cardiovascular diseases [4]. Cardiomyocytes are a type of highly differentiated cells. The search for new drugs to inhibit OS and reduce myocardial cell apoptosis is currently a hot spot in the treatment of cardiovascular diseases
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