Sustained over-expression of calpain-2 induces age-dependent dilated cardiomyopathy in mice through aberrant autophagy

Xiao-yun Ji1,2,3, Dong Zheng4, Rui Ni2,3, Jin-xi Wang5, Jian-qiang Shao6, Zer Vue7, Antentor Hinton Jr.7, Long-Sheng Song5, Guo-Chang Fan8, Subrata Chakrabarti3, Zhao-liang Su1, Tian-qing Peng2,3,9
1 International Genome Center, Jiangsu University, Zhenjiang 212013, China
2 Lawson Health Research Institute, London Health Sciences Centre, London, ON N6A 5W9, Canada
3 Department of Pathology and Laboratory Medicine, Western University, London, ON N6A 5C1, Canada
4 Centre of Clinical Laboratory, the First Affiliated Hospital of Soochow University, Suzhou 215006, China
5 Division of Cardiovascular Medicine, Department of Internal Medicine, Abboud Cardiovascular Research Center, Carver College of Medicine, University of Iowa, Iowa City, IA 52242, USA
6 Central Microscopy Research Facility, University of Iowa, Iowa City, IA 52242, USA
7 Department of Molecular Physiology and Biophysics, Vanderbilt University, Nashville, TN 37232, USA
8 Department of Pharmacology and Systems Physiology, University of Cincinnati College of Medicine, Cincinnati, OH 45267, USA
9 Department of Medicine, Western University, London, ON N6A 5W9, Canada
Correspondence to: Zhao-liang Su:, Tian-qing Peng:,
DOI: 10.1038/s41401-022-00965-9
Received: 27 December 2021
Accepted: 24 July 2022
Advance online: 19 August 2022


Calpains have been implicated in heart diseases. While calpain-1 has been detrimental to the heart, the role of calpain-2 in cardiac pathology remains controversial. In this study we investigated whether sustained over-expression of calpain-2 had any adverse effects on the heart and the underlying mechanisms. Double transgenic mice (Tg-Capn2/tTA) were generated, which express human CAPN2 restricted to cardiomyocytes. The mice were subjected to echocardiography at age 3, 6, 8 and 12 months, and their heart tissues and sera were collected for analyses. We showed that transgenic mice over-expressing calpain-2 restricted to cardiomyocytes had normal heart function with no evidence of cardiac pathological remodeling at age 3 months. However, they exhibited features of dilated cardiomyopathy including increased heart size, enlarged heart chambers and heart dysfunction from age 8 months; histological analysis revealed loss of cardiomyocytes replaced by myocardial fibrosis and cardiomyocyte hypertrophy in transgenic mice from age 8 months. These cardiac alterations closely correlated with aberrant autophagy evidenced by significantly increased LC3BII and p62 protein levels and accumulation of autophagosomes in the hearts of transgenic mice. Notably, injection of 3-methyladenine, a well-established inhibitor of autophagy (30 mg/kg, i.p. once every 3 days starting from age 6 months for 2 months) prevented aberrant autophagy, attenuated myocardial injury and improved heart function in the transgenic mice. In cultured cardiomyocytes, over-expression of calpain-2 blocked autophagic flux by impairing lysosomal function. Furthermore, over-expression of calpain-2 resulted in lower levels of junctophilin-2 protein in the heart of transgenic mice and in cultured cardiomyocytes, which was attenuated by 3-methyladenine. In addition, blockade of autophagic flux by bafilomycin A (100 nM) induced a reduction of junctophilin-2 protein in cardiomyocytes. In summary, transgenic over-expression of calpain-2 induces age-dependent dilated cardiomyopathy in mice, which may be mediated through aberrant autophagy and a reduction of junctophilin-2. Thus, a sustained increase in calpain-2 may be detrimental to the heart.
Keywords: calpain-2; dilated cardiomyopathy; heart dysfunction; autophagy; junctophilin-2; 3-methyladenine

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