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 H-2 O-2 . 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 H-2 O-2 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. Conclusion. 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.

Nyckelord

AGE

diabetes

oxidative stress

NF-kappa B

protein kinase C

TGF-beta 1

d-alpha-tocopherol

diabetic rats

endothelial-cells

gene-expression

oxidant stress

growth-factor

antioxidant

complications

glycosylation

proteins

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