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Methylglyoxal-induced fibronectin gene expression through Ras-mediated NADPH oxidase activation in renal mesangial
Cheng Ho1, Pei-Hsien Lee, Wei-Jan Huang
1Division of Endocrinology and Metabolism, Chang Gung Memorial Hospital, Chiayi, Taiwan.
Background:
The formation of methylglyoxal (MGO), a highly reactive dicarbonyl compound, is accelerated under diabetic conditions. Although recent studies have suggested that apoptotic cell death is involved in diabetic nephropathy, the precise mechanism of MGO-induced renal fibrosis remains to be elucidated.
Methods:
Rat kidney mesangial cells with or without pretreatment with inhibitors, including superoxide dismutase, catalase, L-NAME, diphenylene iodonium, rotenone, allopurinol, PD98059, SB203580 and SP600125 were cultured in medium containing 100 microM MGO. In the MGO-treated cell culture system, fibrosis-related signalling pathway was assessed by enzyme-linked immunosorbent assay, reverse transcription-polymerase chain reaction and western blotting.
Results:
Expression of fibronectin induced by MGO was highest after 48 h treatment. Superoxide production rapidly increased after 2 h and remained at a high level for 24 h. Scavenging O(2) (-) reversed transforming growth factor beta 1 (TGF-beta1) and fibronectin mRNA level. Pretreatment with diphenylene iodonium significantly suppressed MGO-induced superoxide, TGF-beta1 expression and fibronectin gene expression, indicating that NADPH oxidase is responsible for inducing superoxide formation and subsequently induced renal fibrosis. High MGO rapidly enhanced Ras activation in 1 h and progressively increased cytosolic p38 activation. Additionally, SB203580 pretreatment reduced MGO promotion of fibronectin gene activation suggesting that cytosolic p38 activation might affect MGO-induced renal mesangial fibrosis. Inhibiting Ras activity with manumycin A significantly reduced the promoting effect of MGO on superoxide synthesis, and fibronectin expression.
Conclusion:
Induction of superxoide by Ras via p38 pathway activates fibrotic gene transcription of mesangial cells. Reduction of oxidative stress by scavenging superoxide may offer an alternative strategy for controlling MGO-induced renal fibrosis.
Insights
Methylglyoxal (MGO) accelerates diabetic nephropathy by inducing oxidative stress and renal fibrosis through the Ras-p38 pathway. Reducing oxidative stress may help manage MGO-induced kidney fibrosis.
Area of Science:
- Nephrology
- Diabetology
- Biochemistry
Background:
- Methylglyoxal (MGO), a reactive dicarbonyl, increases under diabetic conditions.
- Diabetic nephropathy involves apoptotic cell death, but MGO's role in renal fibrosis is unclear.
Purpose of the Study:
- To elucidate the mechanism of methylglyoxal-induced renal fibrosis in diabetic nephropathy.
- To investigate the signaling pathways involved in MGO-induced mesangial cell fibrosis.
Main Methods:
- Rat kidney mesangial cells were treated with MGO and various inhibitors.
- Fibrosis-related signaling pathways were analyzed using ELISA, RT-PCR, and Western blotting.
Main Results:
- MGO induced fibronectin expression and superoxide production, with NADPH oxidase mediating this effect.
- MGO activated Ras and p38 pathways, which were crucial for fibronectin gene activation and renal fibrosis.
- Inhibiting superoxide production or Ras/p38 pathways attenuated MGO-induced fibronectin expression.
Conclusions:
- Ras activation via the p38 pathway induces fibrotic gene transcription in mesangial cells.
- Scavenging superoxide offers a potential strategy to control MGO-induced renal fibrosis.
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