Published online before print August 22, 2002, doi: 10.1161/01.ATV.0000034475.40227.40
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© 2002 American Heart Association, Inc.
Vascular Biology |
Fluid Shear Stress Activates Proline-Rich Tyrosine Kinase via Reactive Oxygen Species–Dependent Pathway
From the Center for Cardiovascular Research (L.-K.T., J.A., C.Y., B.C.B.), University of Rochester, Rochester, NY, and the Division of Cardiovascular and Respiratory Medicine (M.O.), Kobe University Graduate School of Medicine, Kobe, Japan.
Correspondence to Bradford C. Berk, MD, PhD, University of Rochester, Department of Medicine, Box MED, Rochester, NY 14642. E-mail bradford_berk{at}urmc.rochester.edu
Objective— Fluid shear stress (flow) modulates endothelial cell (EC) function via specific signal transduction events. Previously, we showed that flow-mediated tyrosine phosphorylation of p130 Crk-associated substrate (Cas) required calcium-dependent c-Src activation. Because flow increases reactive oxygen species (ROS) production in ECs and because H2O2 increases tyrosine phosphorylation of proline-rich tyrosine kinase (PYK2), we hypothesized that flow may activate PYK2 via ROS.
Methods and Results— Exposure of bovine aortic ECs to flow stimulated PYK2 phosphorylation rapidly, with a peak at 2 minutes. The activation of PYK2 and phosphorylation of Cas induced by flow were inhibited by pretreatment with the antioxidant N-acetylcysteine. Flow-induced PYK2 phosphorylation was inhibited by BAPTA-AM, an intracellular calcium chelator. Bovine aortic ECs transfected with kinase-inactive PYK2 showed attenuated flow-stimulated Cas tyrosine phosphorylation. Although flow-induced Cas phosphorylation was inhibited by kinase-inactive Src, PYK2 activation induced by flow was not inhibited by overexpression of kinase-inactive Src.
Conclusions— These results show a redox-sensitive pathway for flow-mediated activation of nonreceptor tyrosine kinase activity that requires ROS and intracellular calcium, but not Src kinase.
Key Words: proline-rich tyrosine kinase • calcium • reactive oxygen species • shear stress
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