The adaptor molecule Lnk negatively regulates tumor necrosis factor-α-dependent VCAM-1 expression in endothelial cells through inhibition of the ERK1 and -2 pathways

Lnk, with APS and SH2-B (Src homology 2-B), belongs to a family of SH2-containing proteins with potential adaptor functions. Lnk regulates growth factor and cytokine receptor-mediated pathways implicated in lymphoid, myeloid, and platelet homeostasis. We have previously shown that Lnk is expressed and up-regulated in vascular endothelial cells (ECs) in response to tumor necrosis factor-alpha(TNF alpha). In this study, we have shown that, in ECs, Lnk down-regulates the expression, at both mRNA and protein levels, of the proinflammatory molecules VCAM-1 and E-selectin induced by TNF alpha. Mechanistically, our data indicated that, in response to TNF alpha, NF kappa B/p65 phosphorylation and translocation as well as I kappa B alpha phosphorylation and degradation were unchanged, suggesting that Lnk does not modulate NF kappa B activity. However, Lnk activates phosphatidylinositol 3-kinase (PI3K) as reflected by Akt phosphorylation. Our results identify endothelial nitric-oxide synthase as a downstream target of Lnk-mediated activation of the PI3K/Akt pathway and HO-1 as a new substrate of Akt. We found that sustained Lnk-mediated activation of PI3K in TNF alpha-activated ECs correlated with the inhibition of ERK1/2 phosphorylation, whereas phosphorylation of p38 and c-Jun NH2-terminal kinase (JNK) mitogen-activated protein kinases (MAPKs) was unchanged. ERK1/2 inhibition decreases VCAM-1 expression in TNF alpha-treated ECs. Collectively, our results identify the adaptor Lnk as a negative regulator in the TNF alpha-signaling pathway mediating ERK inhibition and suggest a role for Lnk in the interplay between PI3K and ERK triggered by TNF alpha in ECs.