Cardiomyocyte Derived miR-328 Promotes Cardiac Fibrosis by Paracrinely Regulating Adjacent Fibroblasts

Background/Aims: In our previous study, we demonstrated that elevated expression of miR-328 is a potent determinant of cardiac fibrosis during myocardial infarction (MI). In the present study, histological examination revealed progressive fibrosis in transgenic mice overexpressing cardiomyocyte-specific miR-328. This study investigated whether the transfer of miR-328 from cardiomyocytes (CMs) to cardiac fibroblasts (CFs) in a paracrine manner contributes to myocardial fibrosis. Methods: Myocardial infarction was established by the occlusion of the left coronary artery. Masson’s trichrome staining and collagen assays were used to evaluate the progression of fibrosis. The vesicles and translocation of miR-328 in a co-culture assay system were respectively observed using transmission electron microscopy (TEM) and immunofluorescence staining (IF). Real-time PCR was employed to detect the level of miR-328, Col1α1 and Col3α1. The protein expression of Col1α1, TGF-βRIII, p-smad2/3 (phosphorylated-smad2/3) and TGF-β1 were probed using western blot analysis. Results: Cardiomyocyte-specific miR-328 overexpressing transgenic (TG) mice showed enhanced collagen deposition and provoked cardiac fibrosis by the activation of the TGF-β1 pathway, and this effect was abrogated after knockdown of endogenous miR-328 in mice. Correspondingly, the expression of miR-328 was increased in CFs co-cultured with CMs transfected with miR-328 mimics, likely in a paracrine manner. The cardiomyocyte-mediated augmentation of miR-328 contributes to fibrogenesis in CFs, and this pro-fibrotic effect was reversed after the transfection of miR-328 inhibitor in CFs. Conclusion: A novel molecular mechanism for miR-328 derived from CMs as a paracrine signaling mediator of cardiac fibrogenesis further demonstrates that miR-328 is a potential therapeutic target.