title: Nepsilon-(Carboxymethyl)lysine induces gamma-glutamylcysteine synthetase in RAW264.7 cells.
abstract: Advanced glycation end products (AGEs) play an important role in the development of angiopathy in diabetes mellitus and atherosclerosis. Here, we show that adducts of N(epsilon)-(carboxymethyl)lysine (CML), a major AGE, and bovine serum albumin (CML-BSA) stimulated gamma-glutamylcysteine synthetase (gamma-GCS), which is a key enzyme of glutathione (GSH) synthesis, in RAW264.7 mouse macrophage-like cells. CML-BSA stimulated the expression of gamma-GCS heavy subunit (h) time- and dose-dependently and concomitantly increased GSH levels. CML-BSA also stimulated DNA-binding activity of activator protein-1 (AP-1) within 3h, but the stimulatory effect decreased in 5h, and nuclear factor-kappaB (NF-kappaB) with a peak activity at 1h and the stimulatory effect diminished in 3h. Studies of luciferase activity of the gamma-GCSh promoter showed that deletion and mutagenesis of the AP-1-site abolished CML-BSA-induced up-regulation, while that of NF-kappaB-site did not affect CML-BSA-induced activity. CML-BSA also stimulated the activity of protein kinase C, Ras/Raf-1, and MEK/ERK1/2. Inhibition of ERK1/2 abolished CML-BSA-stimulated AP-1 DNA-binding activity and gamma-GCSh mRNA expression. Our results suggest that induction of gamma-GCS by CML adducts seems to increase the defense potential of cells against oxidative stress produced during glycation processes.
authors.affiliation: The Second Department of Internal Medicine, Nagasaki University School of Medicine, 852-8521, Nagasaki, Japan.
authors.forename: Yoshiyuki
authors.initials: Y
authors.lastname: Miyahara
authors.forename: Satoshi
authors.initials: S
authors.lastname: Ikeda
authors.forename: Takahiro
authors.initials: T
authors.lastname: Muroya
authors.forename: Chie
authors.initials: C
authors.lastname: Yasuoka
authors.forename: Yoshishige
authors.initials: Y
authors.lastname: Urata
authors.forename: Seikoh
authors.initials: S
authors.lastname: Horiuchi
authors.forename: Shigeru
authors.initials: S
authors.lastname: Kohno
authors.forename: Takahito
authors.initials: T
authors.lastname: Kondo
mesh_terms: D000818:Animals; D001483:Base Sequence; D015153:Blotting, Western; D002460:Cell Line; D017931:DNA Primers; D024202:Electrophoretic Mobility Shift Assay; D005721:Glutamate-Cysteine Ligase / Q000096:biosynthesis* / Q000235:genetics; D005978:Glutathione / Q000378:metabolism; D008239:Lysine / Q000031:analogs & derivatives / Q000494:pharmacology*; D020935:MAP Kinase Signaling System; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism; D011493:Protein Kinase C / Q000378:metabolism; D019908:Proto-Oncogene Proteins c-raf / Q000378:metabolism; D012333:RNA, Messenger / Q000235:genetics; D018808:Transcription Factor AP-1 / Q000378:metabolism
chemical_list: D017931:DNA Primers; D016328:NF-kappa B; D012333:RNA, Messenger; D018808:Transcription Factor AP-1; C048496:N(6)-carboxymethyllysine; D019908:Proto-Oncogene Proteins c-raf; D011493:Protein Kinase C; D005721:Glutamate-Cysteine Ligase; D005978:Glutathione; D008239:Lysine