Jiahui Yu1, Yuhong Li2, Xinyan Liu1, Zhe Ma1, Sarhene Michael2, John O Orgah1, Guanwei Fan3, Yan Zhu4. 1. Tianjin State Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China; Research and Development Center of CM, Tianjin International Joint Academy of Biotechnology & Medicine, Tianjin, China. 2. Tianjin State Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China. 3. Tianjin State Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China; First Teaching Hospital of Tianjin University of Traditional Chinese Medicine, Tianjin Key Laboratory of Translational Research of TCM Prescription and Syndrome, Tianjin, China. Electronic address: fgw1005@hotmail.com. 4. Tianjin State Key Laboratory of Modern Chinese Medicine, Tianjin University of Traditional Chinese Medicine, Tianjin, China; Research and Development Center of CM, Tianjin International Joint Academy of Biotechnology & Medicine, Tianjin, China. Electronic address: yanzhu.harvard@iCloud.com.
Abstract
ETHNOPHARMACOLOGICAL RELEVANCE: Shenmai injection (SMI) is a CFDA-approved and widely prescribed herbal medicine injection in China for treating cardiac dysfunction, especially myocardial ischemia and reperfusion (I/R) injury. However, despite of its known clinical efficacy, the cardioprotective mechanisms of SMI remain to be established. AIM OF STUDY: The present study aimed to investigate the role of SMI on mitophagy and mitochondrial dynamics in cardiomyocytes with a hypoxia/reperfusion (H/R) injury setting. MATERIALS AND METHODS: H9c2 cardiomyocytes were subjected to 12 h of hypoxia followed by 2 h of reoxygenation to induce cellular injury. Multi-parameter imaging analysis was performed using Operetta High Content Imaging System to detect changes in mitochondrial function and morphological texture. The mPTP opening was directly assessed by analyzing mitochondrial calcein release in H9c2 and by Ca2+-induced swelling of isolated cardiac mitochondria. Mitochondrial respiration was measured by XF 24 analyzer of Seahorse Bioscience. RT-PCR and Western blotting analyses were used to detect mitophagy, mitochondrial fusion and fission biomarkers at the gene and protein levels. RESULTS: Pretreatment of SMI significantly improved myocardial cell survival and protected against H/R-induced deterioration of mitochondrial structure and function, as evidenced by decreased mitochondrial mass and cytosolic Ca2+, increased mitochondrial membrane potential (ΔΨm) and mitochondrial morphology by SER Texture analysis, inhibited mPTP opening in H9c2 cells and isolated cardiac mitochondria, and alleviated severely impaired mitochondrial respiration. Mechanistically, SMI attenuated H/R injury by inducing mitophagy and then modulated mitochondrial dynamics as indicated by a significantly increased expression of LC3, Beclin 1, Parkin and Pink, and the inhibition of excessive mitochondria fission and increased mitochondrial fusion. Finally, the cardioprotective effect of SMI was confirmed in a LAD-induced cardiac dysfunction model in vivo. CONCLUSION: We found that alleviation of H/R injury by pretreatment with SMI may be attributable to inducing mitophagy and modulating mitochondrial dynamics in cardiomyocytes, thereby providing a rationale for future clinical applications and potential mitoprotective therapy for MI/R injury.
ETHNOPHARMACOLOGICAL RELEVANCE: Shenmai injection (SMI) is a CFDA-approved and widely prescribed herbal medicine injection in China for treating cardiac dysfunction, especially myocardial ischemia and reperfusion (I/R) injury. However, despite of its known clinical efficacy, the cardioprotective mechanisms of SMI remain to be established. AIM OF STUDY: The present study aimed to investigate the role of SMI on mitophagy and mitochondrial dynamics in cardiomyocytes with a hypoxia/reperfusion (H/R) injury setting. MATERIALS AND METHODS: H9c2 cardiomyocytes were subjected to 12 h of hypoxia followed by 2 h of reoxygenation to induce cellular injury. Multi-parameter imaging analysis was performed using Operetta High Content Imaging System to detect changes in mitochondrial function and morphological texture. The mPTP opening was directly assessed by analyzing mitochondrial calcein release in H9c2 and by Ca2+-induced swelling of isolated cardiac mitochondria. Mitochondrial respiration was measured by XF 24 analyzer of Seahorse Bioscience. RT-PCR and Western blotting analyses were used to detect mitophagy, mitochondrial fusion and fission biomarkers at the gene and protein levels. RESULTS: Pretreatment of SMI significantly improved myocardial cell survival and protected against H/R-induced deterioration of mitochondrial structure and function, as evidenced by decreased mitochondrial mass and cytosolic Ca2+, increased mitochondrial membrane potential (ΔΨm) and mitochondrial morphology by SER Texture analysis, inhibited mPTP opening in H9c2 cells and isolated cardiac mitochondria, and alleviated severely impaired mitochondrial respiration. Mechanistically, SMI attenuated H/R injury by inducing mitophagy and then modulated mitochondrial dynamics as indicated by a significantly increased expression of LC3, Beclin 1, Parkin and Pink, and the inhibition of excessive mitochondria fission and increased mitochondrial fusion. Finally, the cardioprotective effect of SMI was confirmed in a LAD-induced cardiac dysfunction model in vivo. CONCLUSION: We found that alleviation of H/R injury by pretreatment with SMI may be attributable to inducing mitophagy and modulating mitochondrial dynamics in cardiomyocytes, thereby providing a rationale for future clinical applications and potential mitoprotective therapy for MI/R injury.
Authors: Yan-Qin Li; Fan Zhang; Li-Ping Yu; Jian-Kang Mu; Ya-Qin Yang; Jie Yu; Xing-Xin Yang Journal: Biomed Res Int Date: 2021-10-30 Impact factor: 3.411
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