上海科技大学知识管理系统

Background: Myocardial infarction (MI) dramatically changes the mechanical stress, which is intensified by the fibrotic remodeling. Integrins, especially the alpha V subunit, mediate mechanical signal and mechanoparacrine of transforming growth factor beta 1 (TGF-beta 1) in various organ fibrosis by activating CFs into myofibroblasts (MFBs). We investigated a possible role of integrin alpha V mediated mechanoparacrine of TGF-beta 1 in MFBs activation for fibrous reparation in mice with MI.Methods: Heart samples from MI, sham, or MI plus cilengitide (14 mg/kg, specific integrin alpha V inhibitor) treated mice, underwent functional and morphological assessments by echocardiography, and histochemistry on 7, 14 and 28 days post-surgery. The mechanical and ultrastructural changes of the fibrous scar were further evaluated by atomic mechanics microscope (AFM), immunofluorescence, second harmonic generation (SHG) imaging, polarized light and scanning electron microscope, respectively. Hydroxyproline assay was used for total collagen content, and western blot for protein expression profile examination. Fibroblast bioactivities, including cell shape, number, Smad2/3 signal and expression of extracellular matrix (ECM) related proteins, were further evaluated by microscopic observation and immunofluorescence in polyacrylamide (PA) hydrogel with adjustable stiffness, which was re-explored in fibroblast cultured on stiff matrix after silencing of integrin alpha V. The content of total and free TGF-beta 1 was tested by enzyme-linked immunosorbent assay (ELISA) in both infarcted tissue and cell samples.Result: Increased stiffness with heterogeneity synchronized with integrin alpha V and alpha smooth muscle actin (alpha-SMA) positive MFBs accumulation in those less mature fibrous areas. Cilengitide abruptly reduced collagen content and disrupted collagen alignment, which also decreased TGF-beta 1 bioavailability, Smad2/3 phosphorylation, and alpha-SMA expression in the fibrous area. Accordingly, fibroblast on stiff but not soft matrix exhibited obvious MFB phenotype, as evidenced by enlarged cell, hyperproliferation, well-developed alpha-SMA fibers, and elevated ECM related proteins, while silencing of integrin alpha V almost abolished this switch via attenuating paracrine of TGF-beta 1 and nuclear translocation of Smad2/3.Conclusion: This study illustrated that increased tissue stiffness activates CFs into MFBs by integrin alpha V mediated mechanoparacrine of TGF-beta 1, especially in immature scar area, which ultimately promotes fibrous scar maturation.