Cathepsin S-mediatcd Fibroblast Trans-difflerentiation Regulates Left Ventricuiar Remodeling After Myocardial Infarction

Background:Although huge progresses of cardiovascular prevention and treatment have been made during the past decades, myocardial infarction (MI) is still one of the leading causes of mortality and morbidity all around the world. Left ventricular (LV) remodeling, which including the structural, physiological and neurohormonal changes of left ventricular wall, is a major event following MI. Adequate remodeling helps reserve cardiac function, while adverse remodeling usually associates with poor prognosis. Lots of factors participate in LV remodeling, such as inflammatory response, hemodynamic load, neurohormonal activation, cytokine production and extracellular matrix (ECM) turnover. Both degradation and synthesis of matrix proteins contribute to ECM turnover. Multiple proteases participate in matrix breakdown, such as the dynamics balance of matrix metalloproteinases (MMPs) and tissue inhibitors of metalloproteinases (TIMPs) plays important role in regulating ECM components. Besides MMPs, however, other proteases also have been demonstrated to engage in ECM catabolism. Lysosomal cysteinyl cathepsins contribute to ECM catabolism in arterial diseases such as atherosclerosis and abdorminal aortic aneurysm, but their function in post-MI LV remodeling remains unknown.Objectives:This study is designed to investigate whether cysteinyl cathepsins participate in post-MI LV remodeling, and explored the mechanisms by which these proteases might regulate myocardial remodeling after MI.Methods:Mouse MI model was induced by permanent left anterior descending coronary artery ligation. Sham operated ones underwent the same procedure but ligation. Both infarct and remote myocardium from sham and post-MI1,2,3,7and28days were homogenized to evaluate mRNA expressions of cysteinyl cathepsins by quantitative real-time polymerase chain reaction (qPCR), and detect their activities by labeling active site with biotin-conjugated JPM probe. To further investigate the role of cathepsins in post-MI LV remodeling process, a non-selective cysteinyl cathepsin inhibitor E64d was administrated intraperitoneally within the first7days post-MI. Cardiac functions were analyzed by echocardiography at baseline,7and28days post-MI. Mice were sacrifice at7and28days post-MI. The effects of E64d treatment on post-MI inflammatory level, fibrosis, apoptosis and angiogenesis were evaluated by immunostaining and qPCR, and MMPs activities were detected by zymography. Immunostaing, Western Blot and qPCR were applied to study the role of cathepsins in cardiac fibroblast (CF) trans-differentiate into myofibroblast. The effects of non-selective cathepsin inhibitor, selective Cathepsin S (CatS) inhibitor and genetically-determined CatS deficiency were tested in transforming growth factor-P (TGF-β) induced in vitro myofibroblast trans-differentiation model. To further understand the role of CatS in post-MI LV remodeling, CatS-deficient Ctss-/-mice were introduced to MI model. Post-MI fibrosis level, myofibroblast trans-differentiation, extra domain A (ED-A) fibronectin expression, Smad2and Smad3phosphorylation were detected by immunostaining and Western Blot.Results:Cathespins expression and activity changed both in infarct and remote areas. CatS showed the most dramatic increase in the infarct area with a peak (>30folds) at7days post-MI, in company with its activity. E64d administration aggravated reduced left ventricular function at7and28days post-MI. Histological analysis demonstrated that E64d increased post-MI inflammatory cell accumulation and cytokine expression, inhibited the activation of the TGF-β1signaling molecules Smad2and Smad3, altered collagen and fibronectin deposition, and suppressed the expression of ED-A fibronectin, an alternatively spliced fibronectin variant that promotes myofibroblast trans-differentiation, and of myofibroblast α-smooth muscle actin (a-SMA), but did not affect apoptosis or angiogenesis in the infarcted myocardium. Selective inhibition or genetically-determined deficiency of CatS reduced myocardial Smad2and Smad3activation and ED-A fibronectin expression, thus suppressing TGF-β1-induced fibroblast trans-differentiation and a-SMA expression and resulted in adverse collagen turnover, greater LV dilation, and worsened cardiac functions, recapitulating the findings in mice treated with general cysteinyl proteinase inhibitor E64d. Humans with coronary heart disease (CHD) had significantly higher plasma levels of mature CatS than did individuals without CHD.Conclusions:Cathepsins play important role in regulating post-MI LV remodeling, especially ECM turnover. CatS limits adverse LV remodeling and preserves LV function after experimental MI by promoting myofibroblast trans-differentiation via TGF-(31signaling and fibronectin ED-A production.