| Background:Osteopontin (OPN) is a pleiotropic cytokine which showed a close relationship with cardiac f ibrosis in previous studies. The expression of OPN has a significant increase in idiopathic dilated cardiomyopathy (DCM) and dilated cardiomyopathy caused by pressure overload. Recent research shows overexpression of OPN in cardiomyocytes can induce dilated cardiomyopathy (DCM). Focal adhesion kinase (FAK) is a cytoplasmic tyrosine kinase which plays a pivotal role in cell migration, proliferation and survival, and is also involved in OPN participated signal pathways. To gain a deep understanding of OPN and its relationship with FAK in DCM, our experiment aimed to testify the pathophysiological role of OPN in myocardial fibrosis and ventricular remodeling process of DCM and the effect of FAK during this development.Method:Eight-week-old cTnTR14IW transgenic mouse of DCM were used in this experiment. Animals were injected with OPN-shRNA in left ventricular free wall to suppress OPN expression. Animals were random divided into 3 groups: sham group, just underwent open thoracic surgery and inject nothing; vehicle group, were injected of control virus with no biological activities; OPN-shRNA group, were injected of OPN-shRNA which can inhibit the expression of OPN.4 weeks later, echocardiographic examination, western blot and Masson staining were performed to identify left ventricle function, ventricular remodeling, severity of fibrosis and their relevance with FAK four weeks later.Results:Echocardiographic examination and Masson assay shows that OPN-shRNA group mice holds an improved cardiac function and remodeling, including higher ejection fraction, fractional shortening, LV free wall thickness and decreased cardiac fibrosis. Western blot and PCR assay shows a decrease of expression of α-SMA, collagen-I, phosphor-FAK and phosphor-Akt in OPN-shRNA group vs. other two groups.Conclusion:Inhibition of OPN can reduce ventricular remodeling and dysfunction in DCM mice, which may attribute to the suppression of collagen-I secretion and differentiation of fibroblast, Akt phosphorylation is necessary in this process in a FAK dependent way.Focal Adhesion Kinase (FAK) located at structures known as focal adhesions mainly act on cell motility and survival. Recent researches showed that FAK also concerned with organ fibrosis. we hypothesized that osteopontin (OPN), an extracellular matrix cytokine closely related to fibrosis, could drive fibrogenesis through FAK dependent signaling pathway. Therefore, we designed this experiment to test our hypothesis. Mouse cardiac fibroblasts were cultured treated with OPN, preincubation of FAK inhibitors were performed 2 hours before OPN treatment. Collagen-I was detected via measuring cytoplasmic protein by western blot and cellular supernatant by ELISA. Expression of alpha-sma was identified by mmunofluorescence staining. The results showed that treatment with OPN can result in increased collagen-I, α-SMA expression indicating aggravated cell fibrosis while this process can be reversed by FAK inhibitor with repressed Akt phosphorylation. We concluded that FAK plays a key role in the signaling pathway of the synthesis of collagen-I caused by OPN and takes an important part in the transformation from cardiac fibroblast into myofibroblast, which may contribute to the target selection of drug therapy of cardiac fibrosis inhibition. |
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