Key Role of Src Kinase in S100B-induced Activation of the Receptor for Advanced Glycation End Products in Vascular Smooth Muscle Cells

Abstract

The receptor for advanced glycation end products (RAGE) and its ligands have been implicated in the activation of oxidant stress and inflammatory pathways in vascular smooth muscle cells (VSMCs) leading to the initiation and augmentation of atherosclerosis. Here we report that non-receptor Src tyrosine kinase and the membrane protein caveolin-1 (Cav-1) play a key role in the activation of RAGE by S100B in VSMCs. S100B increased the activation of Src kinase and tyrosine phosphorylation of caveolin-1 in VSMCs. A RAGEspecific antibody blocked both these effects. An inhibitor of Src kinase, PP2, significantly blocked S100B-induced activation of Src kinase, mitogen-activated protein kinases, transcription factors NF-B and STAT3, superoxide production, tyrosine phosphorylation of Cav-1, VSMC migration, and expression of the pro-inflammatory genes monocyte chemotactic protein-1 and interleukin-6. Cholesterol depletion also inhibited S100B-induced effects indicating the requirement for intact caveolae in RAGE-specific signaling. Nucleofection of either a Src dominant negative mutant, or a Cav-1 mutant lacking the scaffolding domain, or Cav-1 short hairpin RNA significantly reduced S100B-induced inflammatory gene expression in VSMCs. Furthermore, VSMCs derived from insulin-resistant and diabetic db/db mice displayed increased RAGE expression, Src activation, and migration compared with those from control db/؉ mice. The RAGE antibody blocked enhanced migration in db/db cells. These studies demonstrate for the first time that, in VSMCs, Src kinase and Cav-1 play important roles in RAGE-mediated inflammatory gene expression and migration, key events associated with diabetic vascular complications. Vascular smooth muscle cell (VSMC)2 proliferation, migration, and inflammatory gene expression play important roles in the development of atherosclerotic lesions. Diabetic conditions have been shown to enhance these processes, which lead to accelerated atherosclerosis (1-4). Evidence shows that the accumulation of advanced glycation end products (AGEs) and activation of the receptor for AGEs (RAGE) are key factors mediating these events (5-8). RAGE is a member of the immunoglobulin superfamily of cell-surface molecules and is expressed in many cell types, including VSMCs. It can be activated by multiple ligands such as amphoterin, ␤-amyloid peptide, and several short peptides belonging to the S100/calgranulin family, which includes S100B (9, 10). Recent studies using animal models showed that diabetes-induced accelerated atherosclerosis in apoE null mice is associated with enhanced accumulation of RAGE ligands and increased expression of RAGE itself (11, 12). The expression of RAGE as well as its ligands, including S100B, was increased in neointimal and medial cells. Administration of sRAGE could reduce neointimal thickening as well as VSMC proliferation in these animal models. Furthermore, RAGE null mice showed reduced arterial injury responses, whereas transgenic mice expressing DN-RAGE specifically in smooth muscle cells displa...

Enhanced Proatherogenic Responses in Macrophages and Vascular Smooth Muscle Cells Derived From Diabetic db/db Mice

“…Increases in IL-12 levels have recently been reported in mouse macrophages stimulated with glucose in vitro as well in macrophages isolated from streptozotocin-induced diabetic mice and db/db mice (49). In another very recent study, we reported that VSMCs from db/db mice displayed increased AGE receptor expression, Src kinase activation, and migration (50). Thus, our new data along with those of others indicate that vascular cells including VSMCs, endothelial cells, and macrophages are in a preactivated state in db/db mice, and this could be a major underlying cause for the reported predisposition of db/db mice to develop accelerated atherosclerosis (33).…”

KCa3.1 channels mediate the increase of cell migration and proliferation by advanced glycation endproducts in cultured rat vascular smooth muscle cells

“…Previous studies showed that the interaction of AGEs with RAGE in rat VSMCs leads to an increased phosphorylation of ERK1/2, P38-MAPK, 9,4 Src kinases 10 and promotes cell proliferation and migration. The phosphorylation of PI3K is also essential for RAGE ligand-induced VSMC migration and proliferation.…”

“…Recent studies have demonstrated that AGEs and the receptors of AGEs (RAGE) are upregulated in atherosclerotic plaques in diabetic subjects, particularly in intimal macrophages and smooth muscle cells. [4][5][6][7] Accumulation of AGEs and activation of RAGE are found to mediate proliferation, migration, and inflammatory gene expression in vascular smooth muscle cells (VSMCs), which are believed to accelerate the formation of atherosclerosis in diabetes; [8][9][10] however, the detailed mechanisms underlying atherosclerosis are not fully understood.…”

S100B and S100A6 Differentially Modulate Cell Survival by Interacting with Distinct RAGE (Receptor for Advanced Glycation End Products) Immunoglobulin Domains

“…Indeed although RAGE has been shown to transduce cellular signaling via its C-terminal tail (7,41,45,47,65), a recent report indicated that S100B-induced nitric oxide production in microglia does not require RAGE transducing activity but depends on RAGE extracellular domains (66). In addition, in vascular smooth muscle cells, S100B-dependent RAGE signaling has been shown to occur via the non-receptor Src tyrosine kinase used by several receptors lacking intrinsic tyrosine kinase activity to transduce intracellular signals (67). This suggests that RAGE-dependent signaling could occur via different mechanisms and be part of a multiprotein signaling complex.…”