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Theranostics 2022; 12(3):1267-1285. doi:10.7150/thno.65716 This issue Cite

Research Paper

Overexpression of MFN2 alleviates sorafenib-induced cardiomyocyte necroptosis via the MAM-CaMKIIδ pathway in vitro and in vivo

Ziping Song^1,2, Haixu Song², Dan Liu², Bing Yan², Daowen Wang¹, Yan Zhang², Xiaojie Zhao², Xiaoxiang Tian², Chenghui Yan^1,2✉, Yaling Han^1,2✉

1. Division of Cardiology, Department of Internal Medicine, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
2. Department of Cardiology and Cardiovascular Research Institute of PLA, General Hospital of Northern Theater Command, Shenyang 110016, China.

✉ Corresponding authors: Yaling Han, MD, PhD, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Region, Shenyang 110016, China; phone: (0086)-24-28856123, Email: hanyalingnet. Chenghui Yan, MD, PhD, Cardiovascular Research Institute and Department of Cardiology, General Hospital of Northern Region, Shenyang 110016, China; phone: (0086)-24-28856123, Email: yanch1029com More

Abstract

Background: The continued success of oncological therapeutics is dependent on the mitigation of treatment-related adverse events, particularly cardiovascular toxicities. As such, there is an important need to understand the basic mechanisms of drug toxicities in the process of antitumor therapy. Our aim in this study was to elucidate the underlying mechanisms of sorafenib (sor)-induced cardiomyocyte damage.

Methods: Primary mouse cardiomyocytes were prepared and treated with sor and various other treatments. Cardiomyocyte necroptosis was detected by flow cytometry, western blotting, and CCK8 assays. Mitochondrial Ca²⁺ uptake was detected by the Rhod-2 probe using confocal imaging. Morphological changes in mitochondria and mitochondria-associated endoplasmic reticulum (ER) membranes (MAMs) were imaged using transmission electron microscopy (TEM) and confocal microscopy. Cardiac perfusion was performed to detect cardiac specific role of MFN2 overexpression in vivo.

Results: We reported that mitochondrial Ca²⁺ overload, the subsequent increase in calmodulin-dependent protein kinase II delta (CaMKIIδ) and RIP3/MLKL cascade activation, contributed to sor-induced cardiac necroptosis. Excess MAM formation and close ER-mitochondria contact were key pathogenesis of sor-induced Ca²⁺ overload. Sor mediated MFN2 downregulation in a concentration-dependent manner. Furthermore, we found that reduced mitofusin-2 (MFN2) level augmented sor-mediated elevated MAM biogenesis and increased mitochondria-MAM tethering in cardiomyocytes. Sor-induced Mammalian Target of Rapamycin (mTOR) inactivation, followed by the activation and nuclear translocation of Transcription Factor EB (TFEB), contributed to mitophagy and MFN2 degradation. In an in vivo model, mice subjected to sor administration developed cardiac dysfunction, autophagy activation and necroptosis; our investigation found that global and cardiac-specific overexpression of MFN2 repressed cardiac dysfunction, and sor-induced cardiomyocyte necroptosis via repressing the MAM-CaMKIIδ-RIP3/MLKL pathway.

Conclusion: Sorafenib mediated cardiomyocyte necroptosis through the MFN2-MAM-Ca²⁺-CaMKIIδ pathway in vitro and in vivo. The overexpression of MFN2 could rescue sor-induced cardiomyocyte necroptosis without disturbing the anti-tumor effects.

Keywords: Cardio-Oncology, Sorafenib, MFN2, Necroptosis, MAM

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