Osteopontin is involved in urotensin II-induced migration of rat aortic adventitial fibroblasts.

Recent studies suggest that both osteopontin and urotensin II (UII) play critical roles in vascular remodeling. We previously showed that UII could stimulate the migration of aortic adventitial fibroblasts. In this study, we examined whether osteopontin is involved in UII-induced migration of rat aortic adventitial fibroblasts and examined the effects and mechanisms of UII on osteopontin expression in adventitial fibroblasts. Migration of adventitial fibroblasts induced by UII could be inhibited significantly by osteopontin antisense oligonucleotide (P<0.01) but not sense or mismatch oligonucleotides (P>0.05). Moreover, UII dose- and time-dependently promoted osteopontin mRNA expression and protein secretion in the cells, with maximal effect at 10(-8)mol/l at 3h for mRNA expression or at 12h for protein secretion (both P<0.01). Furthermore, the UII effects were significantly inhibited by its receptor antagonist SB710411 (10(-6)mol/l), and Ca(2+) channel blocker nicardipine (10(-5)mol/l), protein kinase C (PKC) inhibitor H7 (10(-5)mol/l), calcineurin inhibitor cyclosporine A (10(-5)mol/l), mitogen-activated protein kinase (MAPK) inhibitor PD98059 (10(-5)mol/l) and Rho kinase inhibitor Y-27632 (10(-5)mol/l). Thus, osteopontin is involved in the UII-induced migration of adventitial fibroblasts, and UII could upregulate osteopontin gene expression and protein synthesis in rat aortic adventitial fibroblasts by activating its receptor and the Ca(2+) channel, PKC, calcineurin, MAPK and Rho kinase signal transduction pathways.

Transforming growth factor-ß1 involved in urotensin II-induced phenotypic differentiation of adventitial fibroblasts from rat aorta.

BACKGROUND: Urotensin II (UII) is a new vasoconstrictive peptide that may activate the adventitial fibroblasts. Transforming growth factor-ß1 (TGF-ß1) is an important factor that could induce the phenotypical transdifferentiation of adventitial fibroblasts. This study aimed to explore whether TGF-ß1 is involved in UII-induced phenotypic differentiation of adventitial fibroblasts from rat aorta. METHODS: Adventitial fibroblasts were prepared by the explant culture method. TGF-ß1 protein secretion from the cells was determined by enzyme-linked immunosorbent assay (ELISA). The mRNA and protein expression of α-smooth nuscle actin (α-SM-actin), the marker of phenotypic differentiation from fibroblasts to myofibroblasts, were determined using real-time quantitative RT-PCR (real-time RT-PCR) and Western blotting, respectively. RESULTS: UII stimulated the secretion of TGF-ß1 in cultured adventitial fibroblasts in a time-dependent manner. The secretion reached a peak at 24 hours, was higher by 69.8% (P < 0.01), than the control group. This effect was also concentration dependent. Maximal stimulation was reached at 10(-8) mol/L of UII (P < 0.01), which was increased by 59.9%, compared with in the control group (P < 0.01). The secretion of TGF-ß1 induced by UII was significantly blocked by SB-710411 (10(-7) mol/L), a specific antagonist of UII receptor. In addition, both UII (10(-8) mol/L) and TGF-ß1 significantly stimulated α-SM-actin mRNA and protein expression. Moreover, the α-SM-actin induced by UII was inhibited by the specific neutralizing antibody (20 µg/ml) of TGF-ß1, while the α-SM-actin expression stimulated by TGF-ß1 (20 ng/ml) was inhibited by SB-710411 (10(-7) mol/L), the UII receptor antagonist. CONCLUSION: This study suggests that UII could induce TGF-ß1 secretion in adventitial fibroblasts via UT activation, and TGF-ß1 might be involved in phenotypic differentiation from adventitial fibroblasts into myofibroblasts induced by UII, and TGF-ß1 signaling might be one of the important pathways by which UII is involved in vascular fibrosis.