Role of extracellular signal-regulated kinases in angiotensin II-Stimulated contraction of smooth muscle cells from human resistance arteries

Backgronnd-We assessed the role of extracellular signal-regulated kinases ( ERKs) in Ang II-stimulated contraction and associated signaling pathways in vascular smooth muscle cells (VSMCs) from human small arteries. Methods and Results-VSMCs derived from resistance arteries (<300 mu m in di ameter) from subcutaneous gluteal biopsies of healthy subjects (n=8) were u sed to assess Ang II-stimulated [Ca2+](i), pH(i), and contractile responses . [Ca2+](i) and pH(i) were measured with fura 2-AM and BCECF-AM, respective ly, and contraction was measured photomicroscopically in cells grown on Mat rigel matrix. To determine whether tyrosine kinases and ERKs influence Ang II-stimulated responses, cells were pretreated with 10(-5) mol/L tyrphostin A-23 (tyrosine kinase inhibitor) and PD98059 (MEK inhibitor). Ang II-stimu lated MEK activity was determined by tyrosine phosphorylation of ERKs. The angiotensin receptor subtypes (AT(1) and AT(2)) were assessed with [Sar(1), Ile(8)]Ang II (a nonselective subtype antagonist), losartan (a selective AT (1) antagonist), and PD123319 (a selective AT(2) antagonist). Ang II dose-d ependently increased [Ca2+](i) (pD(2)=8.4+/-0.36, E-max=541+/-55 nmol/L), p H(i) (pD(2)=9.4+/-0.29, E-max=7.19+/-0.01), and contraction (pD(2)=9.2+/-0. 21, E-max=36+/-2.2%). Ang II induced rapid tyrosine phosphorylation of ERKs , which was inhibited by PD98059. Tyrphostin A-23 and PD98059 attenuated (P <0.05) Ang II-stimulated second messengers, and PD98059 reduced Ang II-indu ced contraction by >50%. [Sar(1),Ile(8)]Ang II and losartan, but not PD1233 19, blocked Ang II-stimulated responses. Conclusions-These data demonstrate that in VSMCs from human peripheral resi stance arteries, functional Ang II receptors of the AT(2) subtype are coupl ed to signaling cascades involving Ca2+ and pH(i) pathways that are partial ly dependent on tyrosine kinases and ERKs. ERKs, the signaling cascades cha racteristically associated with cell growth, may play an important role in Ang II-stimulated contraction of human VSMCs.