Cardiovascular Research Advance Access first published online on March 24, 2009
This version [Corrected Proof] published online on April 9, 2009
Cardiovascular Research, doi:10.1093/cvr/cvp100
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Over-expression of calpastatin inhibits calpain activation and attenuates myocardial dysfunction during endotoxaemia
1 Shanghai Institute of Cardiovascular Diseases, Zhongshan Hospital, Fudan University, Shanghai, China
2 Critical Illness Research, Lawson Health Research Institute, London Health Sciences Center, VRL 6th Floor, A6-140, 800 Commissioners Road, Ontario, Canada N6A 4G5
3 Departments of Pathology, University of Western Ontario, London, Ontario, Canada
4 Departments of Medicine, University of Western Ontario, London, Ontario, Canada
* Corresponding author. Tel: +1 519 6858300 X 55441, fax: +1 519 6858341.E-mail address: tpeng2{at}uwo.ca
Aims: Lipopolysaccharide (LPS) induces cardiomyocyte caspase-3 activation and proinflammatory factors, in particular tumour necrosis factor-alpha (TNF-
) production, both of which contribute to myocardial dysfunction during sepsis. The present study was to investigate the roles of calpain/calpastatin system in cardiomyocyte caspase-3 activation, TNF- expression, and myocardial dysfunction during LPS stimulation.
Methods and results: In cultured adult rat cardiomyocytes, LPS (1 µg/mL) induced calpain and caspase-3 activity, and up-regulated TNF- expression. These effects of LPS were abrogated by over-expression of calpastatin, an endogenous calpain inhibitor, transfection of calpain-1 siRNA, or various pharmacological calpain inhibitors. Furthermore, blocking gp91phox-NADPH oxidase prevented calpain and caspase-3 activation and decreased TNF- expression in LPS-stimulated cardiomyocytes. To investigate the role of calpastatin in endotoxaemia, transgenic mice with calpastatin over-expression (CAST-Tg) and wild-type mice were treated with LPS (4 mg/kg, i.p.) or saline in the presence of calpain inhibitor-III (10 mg/kg, i.p.) for 4 h, and their heart function was measured with a Langendorff system. Over-expression of calpastatin significantly attenuated myocardial dysfunction (P < 0.05). Consistently, calpain activity, caspase-3 activity, and TNF- expression were also reduced in CAST-Tg and calpain inhibitor-III compared with wild-type and vehicle-treated hearts, respectively.
Conclusion: gp91phox-NADPH oxidase-mediated calpain-1 activation induces caspase-3 activation and TNF- expression in cardiomyocytes during LPS stimulation. Over-expression of calpastatin inhibits calpain activation and improves myocardial function in endotoxaemia. The present study suggests that targeting calpain/calpastatin system may be a potential therapeutic intervention for septic hearts.
KEYWORDS Calpastatin; Calpain; Sepsis; TNF-alpha; Caspase-3; Heart; Myocardial dysfunction
Time for primary review: 46 days
The first two authors contributed equally to the study.