Published online before print September 29, 2005, doi: 10.1161/01.ATV.0000188511.84138.9b
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© 2005 American Heart Association, Inc.
Vascular Biology |
Role of Focal Adhesion Kinase in Flow-Induced Dilation of Coronary Arterioles
From the Department of Physiology, Louisiana State University Health Sciences Center, New Orleans.
Correspondence to William M. Chilian, PhD, Louisiana State University, 1201 Perdido St, New Orleans, LA 70112. E-mail chilian{at}lsuhsc.edu
Backgound— Flow-induced regulation of endothelial NO synthase (eNOS) depends on integrin signaling and tyrosine kinase activation. Integrins cluster in focal adhesion complexes, where the extracellular matrix is connected to the cytoskeleton and where focal adhesion kinase (FAK) is located. FAK plays a central role in integrin signaling and Src activation. Accordingly, we hypothesized that FAK plays an important role in flow-induced dilation (FID).
Methods and Results— To inactivate FAK-dependent signaling, anti-FAK, phosphospecific (Tyr397) antibody (FAKab), which binds against the FAK autophosphorylation site, was incorporated into endothelium of rat coronary arterioles using liposomal transfection. The responses to flow, acetylcholine (Ach), or the NO donor MAHAMANONOate (NOC-9) were observed before and after FAKab. In control and vehicles (denatured antibody or transfecting reagent alone), flow produced progressive dilation to a maximal value of 35% increase in diameter, which was inhibited by N
-nitro-L-arginine methyl ester (L-NAME). However, FAKab prevented FID (P<0.01 versus control). Combined treatment with FAKab and L-NAME did not produce inhibition greater than FAKab alone. FAKab did not blunt Ach- or NOC-9–induced dilation. Western analysis demonstrated that FAKab prevented flow-induced phosphorylation of FAK (pY397-FAK), Akt (pS473-Akt), and eNOS (pS1179-eNOS).
Conclusion— Our study demonstrates the pivotal role of FAK in NO-mediated FID. Inhibition of FAK signaling with FAKab impaired FID and phosphorylation of Akt and eNOS. Our data suggest that the activation of FAK is central to the mechanotransduction of FID via regulation of activation of Akt and eNOS.
Although NO-mediated flow-induced dilation (FID) has been observed by many groups, the signal transduction pathway is still not totally resolved. We sought to determine the role of focal adhesion kinase in FID by loading endothelial cells of intact, isolated coronary resistance vessels with an anti-FAK that prevented its downstream signaling. Anti-FAK blocked FID but not dilation to endothelium-dependent and -independent agonists. Anti-FAK also prevented the flow-dependent phosphorylation of Akt and eNOS, 2 enzymes essential for FID. We conclude that FAK plays a pivotal role in NO-mediated FID.
Key Words: coronary microcirculation • resistance vessels • nitric oxide • Akt
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