Role of oxidative stress in advanced glycation end product-induced mesangial cell activation.
BACKGROUND
Levels of advanced glycation end products (AGE) are elevated in individuals with advancing age, renal failure, and diabetes, and accumulation of these molecules may contribute to disease progression. The mechanism by which AGE proteins alter glomerular mesangial cell function, however, is not completely understood. The present study assessed the involvement of oxidative stress in AGE-dependent mesangial cell signaling events.


METHODS
Primary cultures of rat renal mesangial cells were exposed to in vitro AGE-BSA and H2O2. Nuclear factor-kappaB (NF-kappaB) and protein kinase C (PKC) isoform activation were studied using confocal microscopy and Western blotting. Quantitative polymerase chain reaction (PCR) was used to measure transforming growth factor-beta1 (TGF-beta1) levels. The involvement of oxidative stress was assessed by supplementing or compromising cellular antioxidant capacity.


RESULTS
NF-kappaB was dose-dependently activated by AGE. PKC activation was not involved in this response, but analysis of PKC-beta1 activation showed a stimulatory effect of AGE proteins on this isoform. Transcription of TGF-beta1 was stimulated by AGE and was prevented by PKC inhibition. Challenge with H2O2 had similar downstream effects on mesangial cell signaling. Antioxidants, vitamin E and nitecapone, prevented AGE-dependent NF-kappaB activation and normalized PKC activity and associated TGF-beta1 transcription. Depletion of the intracellular antioxidant, glutathione, effectively lowered the AGE concentration needed for mesangial cell activation of NF-kappaB and PKC-beta1. Treatment with a suboptimal AGE dose, under glutathione-depleted conditions, revealed a synergistic effect on both parameters.


CONCLUSIONS
The results support a central role for oxidative stress in AGE-dependent mesangial cell signaling and emphasize the importance of ROS in determining cell responsiveness.
Lal
Mark A
MA
Department of Woman and Child Health, Pediatric Unit, Karolinska Institutet, Stockholm, Sweden.
Brismar
Hjalmar
H
Eklöf
Ann-Christine
AC
Aperia
Anita
A
D000818:Animals; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000502:physiology; D005920:Glomerular Mesangium / Q000166:cytology / Q000187:drug effects* / Q000502:physiology*; D005978:Glutathione / Q000037:antagonists & inhibitors; D017127:Glycation End Products, Advanced / Q000494:pharmacology*; D006861:Hydrogen Peroxide / Q000494:pharmacology; D008297:Male; D008298:Maleates / Q000494:pharmacology; D016328:NF-kappa B / Q000502:physiology; D016877:Oxidants / Q000494:pharmacology; D018384:Oxidative Stress / Q000502:physiology*; D011493:Protein Kinase C / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D017382:Reactive Oxygen Species / Q000378:metabolism; D012710:Serum Albumin, Bovine / Q000494:pharmacology; D016212:Transforming Growth Factor beta / Q000502:physiology; D053773:Transforming Growth Factor beta1
D017127:Glycation End Products, Advanced; D008298:Maleates; D016328:NF-kappa B; D016877:Oxidants; D017382:Reactive Oxygen Species; C508962:Tgfb1 protein, rat; D016212:Transforming Growth Factor beta; D053773:Transforming Growth Factor beta1; C081628:advanced glycation end products-bovine serum albumin; D012710:Serum Albumin, Bovine; D006861:Hydrogen Peroxide; D011493:Protein Kinase C; C014476:diethyl maleate; D005978:Glutathione