Ozone induces tolerance against cardiomyocytes oxygen‑glucose deprivation/reperfusion through inhibition of autophagy pathway

Xu,Lingshan; Zhu,Liang; Liu,Ping; Wu,Zhouquan; Zou,Zhiqing

doi:10.3892/etm.2021.10301

Ozone induces tolerance against cardiomyocytes oxygen‑glucose deprivation/reperfusion through inhibition of autophagy pathway

Authors:
- Lingshan Xu
- Liang Zhu
- Ping Liu
- Zhouquan Wu
- Zhiqing Zou
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Affiliations: Department of Anesthesiology. Nanjing Medical University, Jiangning, Nanjing 211166, P.R. China, Department of Anesthesiology, The Affiliated Changzhou Second People's Hospital of Nanjing Medical University, Changzhou, Jiangsu 213000, P.R. China
Published online on: June 13, 2021 https://doi.org/10.3892/etm.2021.10301
Article Number: 869
Copyright: © Xu et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

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Abstract

Previous studies have reported that excess activation of autophagy in cardiomyocytes is associated with an increase in myocardial oxygen‑glucose deprivation/reperfusion (OGD/R) injury. Ozone therapy affords significant cardio‑protection against myocardial OGD/R injury. The present study was designed to determine whether ozone‑induced tolerance to myocardial OGD/R injury was mediated by inhibiting autophagy. Subsequently, the rat cardio myoblast H9C2 cell line was used in the present study. A model of H9C2 cells under OGD/R was established. The cells were incubated with different concentrations of ozone (10‑60 µg/ml) during reperfusion. Furthermore, to investigate the role of autophagy in OGD/R‑induced injury, the autophagy inducer and inhibitor were applied. Cell viability was detected by Cell Counting kit‑8 assay. Cell apoptosis was evaluated by flow cytometry. Oxidative stress was examined by superoxide dismutase, lactate dehydrogenase and malondialdehyde levels. The expressions of apoptosis regulator B‑cell lymphoma‑2 (Bcl‑2), Bcl‑2 associated X protein (BAX), cleaved caspase‑3, markers of autophagy microtuble‑associated protein 1 light chain 3 (LC3), autophagy‑related protein 5 (Atg5) and Beclin‑1 were measured by western blot analysis. As a result, OGD/R notably decreased cell viability and induced apoptosis in H9C2 cells, while ozone (10‑40 µg/ml) reversed the noxious effects of OGD/R on H9C2 cells, and 20 µg/ml ozone was the most effective. Ozone inhibited the decrease in the ratio of Bcl‑2/BAX and the expression of cleaved caspase‑3, and inhibited the increase in the ratio of LC3‑II/LC3‑I and the expression of Atg5 and Beclin‑1 elicited by OGD/R, as well as dose‑dependently preventing OGD/R‑induced oxidative stress. Furthermore, rapamycin markedly reversed the effects of ozone (20 µg/ml) on OGD/R‑induced expression of autophagy marker proteins and 3‑methyladenine further improved the effect of ozone. Taken together, the results of the present study provided a credible mechanism by which ozone treatment at low concentrations could protect the myocardium from OGD/R‑induced injury by inhibiting autophagy.

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