title: Binding of receptor for advanced glycation end products (RAGE) ligands is not sufficient to induce inflammatory signals: lack of activity of endotoxin-free albumin-derived advanced glycation end products.
abstract: OBJECTIVE
  Activation of the receptor for advanced glycation end products (RAGE) reportedly triggers cellular responses implicated in the pathogenesis of diabetes, such as increasing vascular cell adhesion molecule-1 (VCAM-1) expression on vascular endothelial cells and inducing TNF-alpha secretion by mononuclear cells. The objective of this study was to evaluate whether RAGE binding affinity of AGE-BSAs and cellular activation correlate.
  
  
  METHODS
  To produce AGEs with varying glycation, bovine albumin AGEs were prepared with 500 mmol/l of glucose, fructose or ribose at times of incubation from 1 to 12 weeks. In addition, AGE-BSA was generated using either glyoxylic acid or glycolaldehyde. Cellular binding of the AGE-BSAs and the effect on endothelial cell VCAM-1 expression were studied in RAGE-expressing human microvascular endothelial cell line-4 cells. Induction of TNF-alpha secretion was assessed using RAGE-expressing human peripheral blood mononuclear cells (PBMCs).
  
  
  RESULTS
  Cellular binding of the different AGE preparations correlated well with RAGE affinity. Interestingly, we found that the AGE preparations, which were essentially endotoxin free (< or =0.2 ng/mg protein), were incapable of inducing VCAM-1 or TNF-alpha secretion regardless of RAGE binding affinity, AGE concentration or incubation time. In contrast, the reported RAGE ligand S100b was confirmed to induce VCAM-1 expression on endothelial cells and TNF-alpha secretion by PBMCs after 24 h of treatment.
  
  
  CONCLUSIONS
  The results of this study suggest that AGE modification and high RAGE binding affinity are not sufficient to generate pro-inflammatory signalling molecules. Thus, RAGE binding affinity of AGE-BSAs does not seem to correlate with cellular activation. Our findings using AGEs with strong RAGE-binding properties indicate that AGEs may not uniformly play a role in cellular activation.
authors.affiliation: Novartis Institutes for BioMedical Research, Cambridge, Massachusetts, USA.
authors.forename: J V
authors.initials: JV
authors.lastname: Valencia
authors.forename: M
authors.initials: M
authors.lastname: Mone
authors.forename: C
authors.initials: C
authors.lastname: Koehne
authors.forename: J
authors.initials: J
authors.lastname: Rediske
authors.forename: T E
authors.initials: TE
authors.lastname: Hughes
mesh_terms: D000818:Animals; D002417:Cattle; D002478:Cells, Cultured; D004730:Endothelium, Vascular / Q000502:physiology*; D017127:Glycation End Products, Advanced / Q000378:metabolism*; D006801:Humans; D007231:Infant, Newborn; D007249:Inflammation / Q000503:physiopathology*; D007700:Kinetics; D007963:Leukocytes, Mononuclear / Q000502:physiology*; D008297:Male; D008833:Microcirculation; D011485:Protein Binding; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic / Q000378:metabolism*; D012710:Serum Albumin, Bovine / Q000378:metabolism*; D012867:Skin / Q000098:blood supply
chemical_list: D017127:Glycation End Products, Advanced; D000067759:Receptor for Advanced Glycation End Products; D011971:Receptors, Immunologic; C081628:advanced glycation end products-bovine serum albumin; D012710:Serum Albumin, Bovine