Vitamin D attenuates TGF-β1-induced lung fibroblast proliferation and migration through repression of RasGRP3

Background:Transforming growth factor-β1(TGF-β1)is a pleiotropic cytokine that plays a central role in the pathogenesis of idiopathic pulmonary fibrosis(IPF).While previous studies have revealed a cross-talk between vitamin D and TGF-β1 signaling,it is still unclear how they interact with each other to regulate the progression of IPF.Methods:In this work,we searched for a novel mediator of TGF-β1 activity in lung fibroblasts and examined its regulation by vitamin D.In addition,we investigated the mechanism underlying the interaction between vitamin D and TGF-β1 signaling in lung fibroblast activation.Bioinformatic analysis was performed to identify TGF-β1 downstream target genes.Knockdown and overexpression expression experiments were conducted to determine gene function in the regulation of lung fibroblast proliferation and migration.Results:Analysis of publicly available datasets revealed that RAS guanyl releasing protein 3(RasGRP3)was upregulated in TGF-β1-treated lung fibroblasts and lung tissues from IPF patients relative to healthy controls.Our data confirmed the upregulation of RasGRP3 by TGF-β1 in human MRC5 lung fibroblasts.Overexpression of RasGRP3 enhanced MRC5 cell proliferation and migration.Knockdown of RasGRP3 blocked TGF-β1-induced MRC5 proliferation and migration.Vitamin D abolished TGF-β1-induced RasGRP3 upregulation,which was reversed by inhibition of the vitamin D receptor(VDR).Mechanistically,vitamin D promoted VDR enrichment and prevented mothers against decapentaplegic homolog(SMAD)2 and 3 occupancy at the promoter of RasGRP3.Additionally,overexpression of RasGRP3 reversed the suppressive effect of vitamin D on MRC5 cell proliferation and migration.Conclusion:In conclusion,vitamin D antagonizes TGF-β1-induced lung fibroblast activation by repressing RasGRP3 transcription.