TGF-β Inhibits Ang II-Induced MAPK p44/42 Signaling in Vascular Smooth Muscle Cells by Ang II Type 1 Receptor Downregulation

Abstract

Vascular changes in diabetes are characterized by reduced vasoconstriction and vascular remodeling. Previously, we demonstrated that TGF-β1 impairs Ang II-induced contraction through reduced calcium mobilization. However, the effect of TGF-β1 on Ang II-induced vascular remodeling is unknown. Therefore, we investigated the effect of TGF-β1 on Ang II-induced activation of the MAPK p44/42 pathway in cultured rat aortic smooth muscle cells (RASMC). Activation of MAPK p44/42 was determined with a phospho-specific antibody. Angiotensin type 1 receptor (AT₁) and AT₁ mRNA levels were measured by [³H]candesartan-binding and real-time PCR, respectively. AT₁ gene transcription activity was assessed using AT₁ promoter-reporter constructs and by a nuclear runoff assay. In TGF-β1-pretreated cells, Ang II-induced phosphorylation of MAPK p44/42 was inhibited by 29 and 46% for p42 and p44, respectively, and AT₁ density was reduced by 31%. Furthermore, pretreatment with TGF-β1 resulted in a 64% reduction in AT₁ mRNA levels and decreased AT₁ mRNA transcription rate by 42%. Pretreatment with TGF-β1 blocked Ang II-induced proliferation of RASMC, while stimulating Ang II-induced upregulation of plasminogen activator inhibitor-1. In conclusion, TGF-β1 attenuates Ang II-mediated MAPK p44/42 kinase signaling in RASMC through downregulation of AT₁ levels, which is mainly caused by the inhibition of transcription of the AT₁ gene.