Mir‐1976 Downregulation and Phosphorylation of LARP6 are Associated with Insulin‐like Growth Factor‐1 (IGF1)‐induced Collagen Upregulation in Vascular Smooth Muscle Cells

Abstract

Collagen content in atherosclerotic plaque is a hallmark of plaque stability. We have shown that insulin‐like growth factor‐1 (IGF1) increased collagen content in atherosclerotic plaques of Apoe−/− mice. We have identified LARP6 (La Ribonucleoprotein Domain Family, Member 6), a collagen mRNA binding protein, as a mediator of IGF1‐dependent collagen I upregulation and increased mature fibril formation in smooth muscle cells (SMCs), however specific mechanisms have not been elucidated yet. A total of six micro RNAs (miRs), including miR‐1976, have been shown to bind to LARP6 mRNA. In fact, miR‐1976 mimic (20 nM) decreased LARP6 by 67% and reduced procollagen I by 85% (both are P<0.05), indicating that miR‐1976 negatively regulates LARP6 expression. We found that IGF1 significantly downregulated miR‐1976 by 41±5% in SMCs after 3h of exposure. Moreover, miR‐1976 mimic inhibited IGF1‐induced LARP6 upregulation, suggesting that the downregulation of miR‐1976 mediates IGF1 upregulation of LARP6.It has been reported that LARP6 phosphorylation is critical for regulating translation of collagen polypeptides. To investigate whether IGF1 induces LARP6 phosphorylation, we assessed LARP6 electrophoretic mobility shift by SDS‐PAGE and immunoblotting. SMCs were serum‐starved and treated for 0–48h either with 10 ng/ml IGF1, or 2 ng/ml IFNy, or 2 ng/ml IL‐1b, or 2 ng/ml TNFa, or 10 ng/ml IL‐6. LARP6 protein was detected on immunoblots as a single 65kDa band in untreated SMCs. However, additional 67kDa LARP6 band was evident after 6h of cell treatment with IGF1 or with IL‐6. The density of 67kDa LARP6 band was gradually increased up to 18h of treatment. IGF1 and IL‐6 also increased expression levels of procollagen I and collagen I in a time‐dependent manner, with a peak at 18h. IFNy, IL‐1b, and TNFa did not induce 67kDa LARP6 band or increase in expression level of collagen in SMCs.Taken together, our data suggest that IGF1 promotes collagen expression by upregulating LARP6 levels via a downregulation of miR‐1976 and also by inducing phosphorylation of LARP6. Suggested mechanisms by which IGF1 induces collagen expression in vascular SMCs and thus potentially in atherosclerotic plaques would provide novel insights for a therapeutic approach increasing plaque stability.Support or Funding InformationNIH/NHLBI R01HL070241 (PD)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.