N ε-(Carboxymethyl)Lysine Adducts of Proteins Are Ligands for Receptor for Advanced Glycation End Products That Activate Cell Signaling Pathways and Modulate Gene Expression

Abstract

Recent studies suggested that interruption of the interaction of advanced glycation end products (AGEs), with the signal-transducing receptor receptor for AGE (RAGE), by administration of the soluble, extracellular ligand-binding domain of RAGE, reversed vascular hyperpermeability and suppressed accelerated atherosclerosis in diabetic rodents. Since the precise molecular target of soluble RAGE in those settings was not elucidated, we tested the hypothesis that predominant specific AGEs within the tissues in disorders such as diabetes and renal failure, N ⑀ -(carboxymethyl)lysine (CML) adducts, are ligands of RAGE. We demonstrate here that physiologically relevant CML modifications of proteins engage cellular RAGE, thereby activating key cell signaling pathways such as NF-B and modulating gene expression. Thus, CML-RAGE interaction triggers processes intimately linked to accelerated vascular and inflammatory complications that typify disorders in which inflammation is an established component.Receptor for AGE 1 (RAGE), a member of the immunoglobulin superfamily, was first described as a cell surface interaction site for advanced glycation end products (AGEs), products of glycation and oxidation of proteins and lipids (1-2). AGEs are a heterogeneous class of compounds, whose accumulation in disorders such as diabetes, renal failure, Alzheimer's disease, and, indeed, natural aging, albeit to a lesser degree, has suggested their potential contribution to the pathogenesis of complications that typify these conditions (3-7). Our previous studies demonstrated that both in vitro and in vivo derived heterogeneous AGEs ligate cell surface RAGE on endothelium (ECs), mononuclear phagocytes (MPs), vascular smooth muscle (VSMC), and neurons to activate cell signaling pathways such as ERK1/ERK2 kinases and NF-B (8 -9), thereby redirecting cellular function in a manner linked to expression of inflammatory and prothrombotic genes important in the pathogenesis of chronic disorders as apparently diverse as diabetic macrovascular disease and amyloidosis (10 -20).Our recent studies suggested that interruption of the interaction of AGEs with RAGE in vivo, by administration of soluble RAGE (sRAGE), the extracellular ligand-binding domain of RAGE, reversed vascular hyperpermeability and suppressed accelerated atherosclerotic lesion development and complexity in diabetic rodents (19 -20). In the latter studies, analysis of plasma demonstrated evidence of an sRAGE⅐AGE complex; immunoprecipitation of plasma obtained from diabetic sRAGEtreated mice with anti-RAGE IgG yielded species immunoreactive with both anti-RAGE IgG or affinity purified anti-AGE IgG, suggesting that sRAGE might bind up AGEs and limit their interaction with and activation of cell surface RAGE. The beneficial effects of sRAGE were independent of alterations in other risk factors, such as hyperglycemia and hyperlipidemia, implicating a role for AGE-RAGE interaction in the development of vascular dysfunction in diabetes (20).These past studies, however, did not elucidate ...

Nϵ‐Carboxymethyllysine Increases the Expression of miR‐103/143 and Enhances Lipid Accumulation in 3T3‐L1 Cells

“…We did, however, observe a down‐regulation of INSR after 90 min and a differential regulation of the insulin‐regulated glucose transporter 4 ( GLUT4 ) after 30 and 60 min of treatment with 500 μM CML, indicating an impact of CML treatment on glucose homeostasis on a genetic level. CML has been identified as the major epitope of AGE‐modified proteins and activation of RAGE leading to a modulation of gene expression has been shown [Ikeda et al, 1996; Kislinger et al, 1999], yet, free CML has been demonstrated to not stably bind to RAGE [Xie et al, 2008]. However, a study on HEK‐293 cells expressing full length RAGE and HEK‐293 cells expressing a C‐terminally truncated version of the receptor showed casein‐CML, as well as free CML to increase p38 MAP kinase activation [Zill et al, 2003].…”

“…Exogenous AGEs, typically encountered in a Western diet, are at least partially absorbed into circulation where they add to the endogenous AGE level [Uribarri et al, 2005; Somoza et al, 2006; Sebekova et al, 2008; Birlouez‐Aragon et al, 2010]. Among the group of AGEs, N‐ϵ‐carboxymethyl‐ l ‐lysine (CML), a well‐characterized, stable [Ahmed et al, 1986; Kislinger et al, 1999; Uribarri et al, 2010] member, and major epitope of AGE‐modified proteins [Ikeda et al, 1996], has been detected in a wide range of foods, its concentration reaching up to 84.8 mg per average portion size in meat dishes [Hull et al, 2012]. In a study on the dose‐dependent utilization of CML in rats, 30% of dietary CML has been shown to enter the blood stream [Somoza et al, 2006].…”

Levels of soluble receptor for AGE are cross-sectionally associated with cardiovascular disease in type 1 diabetes, and this association is partially mediated by endothelial and renal dysfunction and by low-grade inflammation: the EURODIAB Prospective Complications Study

“…The current view is that the activation of RAGE, a 55 kDa protein and member of the immunoglobulin superfamily of cell-surface receptors, through ligand binding activates the endothelial cell and triggers multiple signalling cascades [8,9]. This results in activation and translocation of nuclear transcription factors and transcription of the target genes, including those for VCAM-1 [9], E-selectin [10] and proinflammatory cytokines [11].…”

“…The functional role of these soluble forms of RAGE in the circulation remains unclear, but they may reflect the activity of the AGE-RAGE axis. The ligation of RAGE activates the endothelial cell and triggers multiple signalling cascades [8,9], resulting in activation and translocation of nuclear transcription factors and transcription of the target genes, including adhesion molecules [9,10] and proinflammatory cytokines [11]. The activation of RAGE may also lead to nephropathy [12].…”

How RAGE turns in rage

“…8 Signaling studies revealed that ligation of cell surface RAGE activates cell signaling pathways such as p21 Ras , ERK1/ERK2 kinases and NF-B, thereby activating an inflammatory gene expression profile that typifies chronic inflammatory disorders. 9 The EN-RAGEs S100A8 (myeloid related protein 8 (MRP)) and S100A9 (MRP14) are specifically secreted by monocytes upon interaction with inflammation activated endothelium. 10 Hence, the subsequent interaction of the secreted EN-RAGEs with cell surface RAGE is expected to further amplify the inflammatory response and suggests a pivotal role for EN-RAGEs and RAGE in chronic cellular activation.…”