Urotensin II-induced collagen synthesis in cultured smooth muscle cells from rat aortic media and a possible involvement of transforming growth factor-β1/Smad2/3 signaling pathway.

BACKGROUND

Recent studies suggest that urotensin II (UII) and transforming growth factor-β1 (TGF-β1) both have critical roles in vascular remodeling. UII is a recently discovered vasoconstrictive peptide that is involved in the pathogenesis of atherosclerosis, restenosis and hypertension. TGF-β1 is an important factor that has a pivotal role in vascular fibrosis. This study aimed to explore whether TGF-β1 is involved in UII-induced collagen synthesis in rat aortic vascular smooth muscle cells (VSMCs) and examined the effects and mechanisms of UII on collagen synthesis and secretion in VSMCs.

METHODS

VSMCs were prepared by the explant culture method. TGF-β1 and collagen I secretions from the cells were determined by enzyme-linked immunosorbent assay (ELISA). The mRNA and protein expressions of TGF-β1, collagen I, Smad2 and Smad3 were determined using Real-time RT-PCR and Western blotting.

RESULTS

UII dose-dependently promoted TGF-β1 protein expression and secretion from VSMCs, with maximal effect at 10(-8) mol/l at 24 h for protein expression and 10(-7) mol/l at 24 h for protein secretion (both P<0.01). Moreover, UII dose-dependently promoted Smad2 and Smad3 mRNA expression in VSMCs, with maximal effect at 10(-8) mol/l for 12 h (both P<0.01). The effects of UII were significantly inhibited by its receptor antagonists urantide (10(-6) mol/l) or SB-710411 (10(-6) mol/l), and by the mitogen-activated protein kinase (MAPK/ERK) inhibitor PD98059 (10(-6) mol/l). UII dose-dependently promoted collagen I mRNA expression and protein secretion in VSMCs, with maximal effect at 10(-8) mol/l at 12h for mRNA expression and 10(-6) mol/l at 24 h for protein secretion (both P<0.01). Collagen synthesis and secretion from VSMCs induced by UII were inhibited significantly by a TGF-β1-specific neutralizing antibody, SB-431542 (an antagonist of the TGF-β1 type II receptor) and PD98059 (all P<0.01).

CONCLUSIONS

This study suggests that UII could induce collagen synthesis and secretion through upregulation of TGF-β1 expression and secretion in VSMCs, and that TGF-β1/Smad2/3 signaling might be one of the important pathways by which UII is involved in vascular fibrosis.