Submitted on September 20, 2008
Revised on April 6, 2009
Accepted on April 6, 2009
Visfatin (nicotinomide phosphoribosyltransferase/pre-B cell colony-enhancing factor) activates eNOS via Akt and MAP kinases and improves endothelial function
Fina Lovren1, Yi Pan1, Praphulla C Shukla1, Adrian Quan1, Hwee Teoh2, Paul Edward Szmitko1, Mark D Peterson2, Mohammed Al-Omran3, and Subodh Verma2*
1 University of Toronto
2 St Michael's Hospital
3 College of Medicine and King Khalid University Hospital
* To whom correspondence should be addressed. E-mail: subodh.verma{at}sympatico.ca.
Improving eNOS bioactivity and endothelial function is important to limit native, vein graft and transplant atherosclerosis. Visfatin, a NAD biosynthetic enzyme, regulates the activity of the cellular survival factor, Sirt1. We hypothesized that visfatin may improve eNOS expression, endothelial function and post-natal angiogenesis. In human umbilical vein (HUVEC) and coronary artery endothelial cells, we evaluated the effects of recombinant human visfatin on eNOS expression and mRNA stability, in the presence and absence of visfatin RNA silencing. We also assessed visfatin-induced Akt activation and its association with src-tyrosine kinases, phosphorylation of Ser-1177 within eNOS in the presence and absence of PI3-kinase inhibition with LY294002, and evaluated the contributory role of ERK1/2. Finally, we determined the impact of visfatin on HUVEC migration, proliferation, inflammation-induced permeability and in-vivo angiogenesis. Visfatin (100 ng/ml) upregulated and stabilized eNOS mRNA, and increased the production of NO and cGMP. Visfatin-treated HUVEC demonstrated greater proliferation, migration, and capillary-like tube formation but less TNF
-induced permeability; these effects were decreased in visfatin gene silenced cells. Visfatin increased total Akt and Ser-473-phospho-Akt expression with concomitant rises in eNOS phosphorylation at Ser-1177; these effects were blocked by LY2940002. Studies with PP2 showed that the non receptor tyrosine kinase, src, is an upstream stimulator of the PI3 kinase-Akt pathway. Visfatin also activated MAP kinase through PI3 kinase, and MEK inhibition attenuated visfatin-elicted Akt and eNOS phosphorylation. Visfatin-filled matrigel implants showed an elevated number of infiltrating vessels, and visfatin treatment produced significant recovery of limb perfusion following hindlimb ischemia. These results indicate a novel effect of visfatin to stimulate eNOS expression and function in endothelial cells, via a common upstream, src-mediated signaling cascade, which leads to activation of Akt and MAP kinases. Visfatin represents a translational target to limit endothelial dysfunction, native, vein graft and transplant atherosclerosis, and improve post-natal angiogenesis.
