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线粒体TRPV3参与压力超负荷诱导的大鼠心脏肥大

朱美萍^1,2, 张秉宜³, 连婷^1,2,4, 谈远佳^1,2,5, 常林林^1,2, 徐盼^1,2,6, 张瑾怡^1,2, 杜燕欢^1,2, 熊振宇^1,2, 杜琼⁷, 张世忠^1,2,*

¹三峡大学国家中医药管理局中药药理科研三级实验室，宜昌 443002；²三峡大学肿瘤微环境与免疫治疗湖北省重点实验室，宜昌 443002；³宜昌市中心人民医院超声科，宜昌 443002；⁴松滋市人民医院消化内科，松滋 434200；⁵枝江市人民医院药学部，枝江 44320；⁶江西警察学院侦查学院，南昌 330199；⁷宜昌市夷陵人民医院神经内科，宜昌 443002

摘要

线粒体在压力负荷增高诱导的心脏肥大中具有重要作用。本文旨在研究线粒体瞬时受体电位香草素受体3 (transient receptor potential vanilloid 3, TRPV3)在心脏肥大中的作用。用直径0.7 mm U形银夹钳夹住Sprague-Dawley (SD)大鼠腹主动脉，建立腹主动脉缩窄(abdominal aortic constriction, AAC)动物模型；用血管紧张素II (angiotensin II, Ang II)处理大鼠心肌H9C2细胞，建立肥大心肌细胞模型，并用TRPV3小干扰RNA (small interfering RNA, siRNA)敲低TRPV3表达。用JC-1探针检测线粒体膜电位(mitochondrial membrane potential, MMP)，用DHE探针检测ROS生成，用试剂盒检测线粒体呼吸链复合体I和III酶活性以及ATP生成，用免疫荧光染色法检测H9C2细胞TRPV3表达，用Western blot检测β-肌球蛋白重链(β-myosin heavy chain, β-MHC)、线粒体TRPV3和线粒体NOX4蛋白表达水平。结果显示，在大鼠AAC模型心脏组织和Ang II处理的H9C2细胞上，β-MHC、线粒体TRPV3和线粒体NOX4蛋白表达水平上调，MMP降低，ROS生成增多，线粒体呼吸链复合体I和III酶活性降低，ATP生成减少。线粒体TRPV3敲低后，H9C2细胞β-MHC和线粒体NOX4蛋白表达水平下调，MMP升高，ROS生成减少，线粒体呼吸链复合体I和III酶活性升高，ATP生成增多。以上结果提示，心肌细胞线粒体TRPV3可能通过上调NOX4加重线粒体功能障碍，从而参与压力负荷增高诱导的心脏肥大过程。

关键词： 心脏肥大; 压力超负荷; 线粒体; TRPV3/NOX4

Involvement of mitochondrial TRPV3 in cardiac hypertrophy induced by pressure overload in rats

ZHU Mei-Ping^1,2, ZHANG Bing-Yi³, LIAN Ting^1,2,4, TAN Yuan-Jia^1,2,5, CHANG Lin-Lin^1,2, XU Pan^1,2,6, ZHANG Jin-Yi^1,2, DU Yan-Huan^1,2, XIONG Zhen-Yu^1,2, DU Qiong⁷, ZHANG Shi-Zhong^1,2,*

¹Third-grade Pharmacological Laboratory on Traditional Chinese Medicine Approved by State Administration of Traditional Chinese Medicine, China Three Gorges University, Yichang 443002, China；²Hubei Key Laboratory of Tumor Microenvironment and Immunotherapy, China Three Gorges University, Yichang 443002, China；³Department of Gastroenterology, Yichang Central People’s Hospital, Yichang 443002, China；⁴Department of Gastroenterology, Songzi People’s Hospital, Songzi 434200, China；⁵Department of Pharmacy, Zhijiang People’s Hospital, Zhijiang 443200, China；⁶Investigation School of Jiangxi Police College, Nanchang 330199, China；⁷Department of Neurology, Yiling People’s Hospital of Yichang, Yichang 443002, China

Abstract

Mitochondria play an important role in pressure overload-induced cardiac hypertrophy. The present study aimed to investigate the role of mitochondrial transient receptor potential vanilloid 3 (TRPV3) in myocardial hypertrophy. A 0.7 mm diameter U-shaped silver clip was used to clamp the abdominal aorta of Sprague Dawley (SD) rats and establish an animal model of abdominal aortic constriction (AAC). Rat H9C2 myocardial cells were treated with angiotensin II (Ang II) to establish a hypertrophic myocardial cell model, and TRPV3 expression was knocked down using TRPV3 small interfering RNA (siRNA). JC-1 probe was used to detect mitochondrial membrane potential (MMP). DHE probe was used to detect ROS generation. Enzyme activities of mitochondrial respiratory chain complex I and III and ATP production were detected by assay kits. Immunofluorescence staining was used to detect TRPV3 expression in H9C2 cells. Western blot was used to detect the protein expression levels of β-myosin heavy chain (β-MHC), mitochondrial TRPV3 and mitochondrial NOX4. The results showed that, in the rat AAC model heart tissue and H9C2 cells treated with Ang II, the protein expression levels of β-MHC, mitochondrial TRPV3 and mitochondrial NOX4 were up-regulated, MMP was decreased, ROS generation was increased, mitochondrial respiratory chain complex I and III enzyme activities were decreased, and ATP production was reduced. After knocking down mitochondrial TRPV3 in H9C2 cells, the protein expression levels of β-MHC and mitochondrial NOX4 were down-regulated, MMP was increased, ROS generation was decreased, mitochondrial respiratory chain complex I and III enzyme activities were increased, and ATP production was increased. These results suggest that mitochondrial TRPV3 in cardiomyocytes exacerbates mitochondrial dysfunction by up-regulating NOX4, thereby participating in the process of pressure overload-induced myocardial hypertrophy.

Key words: cardiac hypertrophy; pressure overload; mitochondrion; TRPV3/NOX4

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通讯作者：张世忠　　E-mail:

引用本文：

朱美萍, 张秉宜, 连婷, 谈远佳, 常林林, 徐盼, 张瑾怡, 杜燕欢, 熊振宇, 杜琼, 张世忠. 线粒体TRPV3参与压力超负荷诱导的大鼠心脏肥大[J]. 生理学报 2024; 76 (5): 703-716.

ZHU Mei-Ping, ZHANG Bing-Yi, LIAN Ting, TAN Yuan-Jia, CHANG Lin-Lin, XU Pan, ZHANG Jin-Yi, DU Yan-Huan, XIONG Zhen-Yu, DU Qiong, ZHANG Shi-Zhong. Involvement of mitochondrial TRPV3 in cardiac hypertrophy induced by pressure overload in rats. Acta Physiol Sin 2024; 76 (5): 703-716