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

Abstract

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-␣ (TNF␣). 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␣. Mechanistically, our data indicated that, in response to TNF␣, NFB/p65 phosphorylation and translocation as well as IB␣ phosphorylation and degradation were unchanged, suggesting that Lnk does not modulate NFB 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 Lnkmediated activation of PI3K in TNF␣-activated ECs correlated with the inhibition of ERK1/2 phosphorylation, whereas phosphorylation of p38 and c-Jun NH 2 -terminal kinase (JNK) mitogen-activated protein kinases (MAPKs) was unchanged. ERK1/2 inhibition decreases VCAM-1 expression in TNF␣-treated ECs. Collectively, our results identify the adaptor Lnk as a negative regulator in the TNF␣-signaling pathway mediating ERK inhibition and suggest a role for Lnk in the interplay between PI3K and ERK triggered by TNF␣ in ECs.

Lnk Deletion Reinforces the Function of Bone Marrow Progenitors in Promoting Neovascularization and Astrogliosis Following Spinal Cord Injury

“…Lnk is furthermore considered to negatively regulate endothelial cell derived hematopoiesis during embryonic development [30]. In addition, Lnk is expressed in endothelial cells and played a role in the regulation of vascular cell adhesion molecule-1 expression in response to tumor necrosis factor-a through inhibition of the extracellular-related kinase 1/2 pathway [31,32]. On the basis of these data, we speculate that Lnk might be involved in the differentiation and functional regulation of endothelial cells as well as endothelial progenitor cells [33].…”

Tribbles-1 as a Novel Biomarker of Chronic Antibody-Mediated Rejection

“…For monocyte activation tests, CD14 ϩ cells were cultured (1 ϫ 10 6 cells/ml) as previously for 24 h alone or in the presence of 1 g/ml LPS or the previous proinflammatory cocktail. Human renal artery-derived EC were cultured and activated with recombinant human TNF-␣ as described previously 43 for 6, 12, and 24 h. At least 10 6 monocytes, DC, and EC were washed and resuspended in the appropriate volume of TRIzol reagent and stored at Ϫ80°C until use.…”

Negative Regulation of Haematopoiesis: Role of Inhibitory Adaptors

“…Moreover, Lnk expression is upregulated by certain cytokines important for the development and function of these haematopoietic cells, such as SCF, thrombopoietin [TPO], and erythropoietin [EPO] (Kent et al, 2008;Buza-Vidas et al, 2006;Gerry et al, 2009aGerry et al, , 2009bBaran-Marszak et al, 2010). Interestingly, Lnk is also highly expressed in endothelial cells and its expression is also induced by Tumor Necrosis Factor (TNF)- (Fitau et al, 2006;Kwon et al, 2009). These findings suggest the implication of Lnk adaptor in a negative feedback loop for the regulation of these cytokine pathways.…”

“…However, APS can also function as negative regulator in the BCR and JAK2 signalling pathways (Yokouchi et al, 1997;Wakioka et al, 1999). Conversely, Lnk is considered as a negative regulator of growth factor and cytokine receptor-induced proliferation and migration (Takaki et al, 2000Velazquez et al, 2002;Tong & Lodish, 2004;Tong et al, 2005;Fitau et al, 2006;Simon et al, 2008;. Nonetheless, Lnk seems to play a positive role in mouse platelets for the stabilization of thrombus through the integrin IIb3 outside-in signalling and in human vascular endothelial cells via the PI3K/Akt pathway activated by TNF- (Takizawa et al, 2010;Fitau et al, 2006).…”

“…Conversely, Lnk is considered as a negative regulator of growth factor and cytokine receptor-induced proliferation and migration (Takaki et al, 2000Velazquez et al, 2002;Tong & Lodish, 2004;Tong et al, 2005;Fitau et al, 2006;Simon et al, 2008;. Nonetheless, Lnk seems to play a positive role in mouse platelets for the stabilization of thrombus through the integrin IIb3 outside-in signalling and in human vascular endothelial cells via the PI3K/Akt pathway activated by TNF- (Takizawa et al, 2010;Fitau et al, 2006). Together with data on in vivo ablation of these adaptors, these findings demonstrate that these adaptor proteins can function as positive and/or negative regulators depending on their cell expression and on the growth factor or cytokine receptor-mediated pathway.…”