title: Deletion of capn4 Protects the Heart Against Endotoxemic Injury by Preventing ATP Synthase Disruption and Inhibiting Mitochondrial Superoxide Generation.
abstract: BACKGROUND
  Our recent study has demonstrated that inhibition of calpain by transgenic overexpression of calpastatin reduces myocardial proinflammatory response and dysfunction in endotoxemia. However, the underlying mechanisms remain to be determined. In this study, we used cardiomyocyte-specific capn4 knockout mice to investigate whether and how calpain disrupts ATP synthase and induces mitochondrial superoxide generation during endotoxemia.
  
  
  RESULTS
  Cardiomyocyte-specific capn4 knockout mice and their wild-type littermates were injected with lipopolysaccharides. Four hours later, calpain-1 protein and activity were increased in mitochondria of endotoxemic mouse hearts. Mitochondrial calpain-1 colocalized with and cleaved ATP synthase-α (ATP5A1), leading to ATP synthase disruption and a concomitant increase in mitochondrial reactive oxygen species generation during lipopolysaccharide stimulation. Deletion of capn4 or upregulation of ATP5A1 increased ATP synthase activity, prevented mitochondrial reactive oxygen species generation, and reduced proinflammatory response and myocardial dysfunction in endotoxemic mice. In cultured cardiomyocytes, lipopolysaccharide induced mitochondrial superoxide generation that was prevented by overexpression of mitochondria-targeted calpastatin or ATP5A1. Upregulation of calpain-1 specifically in mitochondria sufficiently induced superoxide generation and proinflammatory response, both of which were attenuated by ATP5A1 overexpression or mitochondria-targeted superoxide dismutase mimetics.
  
  
  CONCLUSIONS
  Cardiomyocyte-specific capn4 knockout protects the heart against lipopolysaccharide-induced injury in endotoxemic mice. Lipopolysaccharides induce calpain-1 accumulation in mitochondria. Mitochondrial calpain-1 disrupts ATP synthase, leading to mitochondrial reactive oxygen species generation, which promotes proinflammatory response and myocardial dysfunction during endotoxemia. These findings uncover a novel mechanism by which calpain mediates myocardial dysfunction in sepsis.
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Rui
authors.initials: R
authors.lastname: Ni
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Dong
authors.initials: D
authors.lastname: Zheng
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Qiang
authors.initials: Q
authors.lastname: Wang
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Yong
authors.initials: Y
authors.lastname: Yu
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Ruizhen
authors.initials: R
authors.lastname: Chen
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Tao
authors.initials: T
authors.lastname: Sun
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Wang
authors.initials: W
authors.lastname: Wang
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Guo-Chang
authors.initials: GC
authors.lastname: Fan
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Peter A
authors.initials: PA
authors.lastname: Greer
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.).
authors.forename: Richard B
authors.initials: RB
authors.lastname: Gardiner
authors.affiliation: From the Jiangsu Key Laboratory of Infection and Immunity, Institutes of Biology and Medical Sciences, Soochow University, Suzhou, Jiangsu, China (R.N., D.Z., T.P.); Lawson Health Research Institute, Departments of Medicine (R.N., D.Z., T.S., T.P.), Pathology (R.N., D.Z., T.P.) and Biology (R.B.G.), University of Western Ontario, London, Ontario, Canada; Department of Dermatology (Q.W.) and Institute of Cardiovascular Diseases (Y.Y., R.C.), Zhongshan Hospital, Fudan University, Shanghai, China; Mitochondria and Metabolism Center, Departments of Anesthesiology and Pain Medicine, University of Washington, Seattle (W.W.); Department of Pharmacology and Cell Biophysics, University of Cincinnati College of Medicine, OH (G.-C.F.); and Division of Cancer Biology and Genetics, Queen's University Cancer Research Institute, Department of Pathology and Molecular Medicine, Queen's University, Kingston, Ontario, Canada (P.A.G.). tqpeng@suda.edu.cn.
authors.forename: Tianqing
authors.initials: T
authors.lastname: Peng
mesh_terms: D000818:Animals; D002154:Calpain / Q000096:biosynthesis / Q000235:genetics*; D004247:DNA / Q000235:genetics; D004195:Disease Models, Animal; D019446:Endotoxemia / Q000150:complications* / Q000235:genetics / Q000378:metabolism; D005260:Female; D017353:Gene Deletion; D005786:Gene Expression Regulation; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008929:Mitochondria, Heart / Q000378:metabolism* / Q000473:pathology; D025261:Mitochondrial Proton-Translocating ATPases / Q000096:biosynthesis / Q000235:genetics*; D015428:Myocardial Reperfusion Injury / Q000235:genetics* / Q000378:metabolism / Q000503:physiopathology; D009206:Myocardium / Q000378:metabolism / Q000473:pathology; D032383:Myocytes, Cardiac / Q000378:metabolism*; D010086:Oxidative Phosphorylation Coupling Factors / Q000096:biosynthesis / Q000235:genetics*
chemical_list: D010086:Oxidative Phosphorylation Coupling Factors; D004247:DNA; D002154:Calpain; C410233:Capns1 protein, mouse; C000628217:F(6) ATPase; D025261:Mitochondrial Proton-Translocating ATPases
keywords: calpain; heart; lipopolysaccharides; mitochondria; sepsis; superoxides