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miR-200a Prevents renal fibrogenesis through repression of TGF-β2 expression.

Wang B, Koh P, Winbanks C, Coughlan MT, McClelland A, Watson A, Jandeleit-Dahm K, Burns WC, Thomas MC, Cooper ME, Kantharidis P - Diabetes (2010)

Bottom Line: TGF-β1 and TGF-β2 also downregulated expression of miR-200a.The renal expression of miR-141 and miR-200a was also reduced in mouse models representing early and advanced kidney disease. miR-200a and miR-141 significantly impact on the development and progression of TGF-β-dependent EMT and fibrosis in vitro and in vivo.These miRNAs appear to be intricately involved in fibrogenesis, both as downstream mediators of TGF-β signaling and as components of feedback regulation, and as such represent important new targets for the prevention of progressive kidney disease in the context of diabetes.

View Article: PubMed Central - PubMed

Affiliation: JDRF Danielle Alberti Memorial Centre for Diabetes Complications, Melbourne, Australia.

ABSTRACT

Objective: Progressive fibrosis in the diabetic kidney is driven and sustained by a diverse range of profibrotic factors. This study examines the critical role of microRNAs (miRNAs) in the regulation of the key fibrotic mediators, TGF-β1 and TGF-β2.

Research design and methods: Rat proximal-tubular epithelial cells (NRK52E) were treated with TGF-β1 and TGF-β2 for 3 days, and expression of markers of epithelial-to-mesenchymal transition (EMT) and fibrogenesis were assessed by RT-PCR and Western blotting. The expression of miR-141 and miR-200a was also assessed, as was their role as translational repressors of TGF-β signaling. Finally, these pathways were explored in two different mouse models, representing early and advanced diabetic nephropathy.

Results: Both TGF-β1 and TGF-β2 induced EMT and fibrogenesis in NRK52E cells. TGF-β1 and TGF-β2 also downregulated expression of miR-200a. The importance of these changes was demonstrated by the finding that ectopic expression miR-200a downregulated smad-3 activity and the expression of matrix proteins and prevented TGF-β-dependent EMT. miR-200a also downregulated the expression of TGF-β2, via direct interaction with the 3' untranslated region of TGF-β2. The renal expression of miR-141 and miR-200a was also reduced in mouse models representing early and advanced kidney disease.

Conclusions: miR-200a and miR-141 significantly impact on the development and progression of TGF-β-dependent EMT and fibrosis in vitro and in vivo. These miRNAs appear to be intricately involved in fibrogenesis, both as downstream mediators of TGF-β signaling and as components of feedback regulation, and as such represent important new targets for the prevention of progressive kidney disease in the context of diabetes.

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Changes in miR-141/200a expression in the kidney in the adenine-induced renal fibrosis model. A: Trichrome staining of tissue sections from renal cortex from control and adenine-fed C57bl6 mice after 4 weeks of treatment. Blue staining indicates high levels of collagen in the adenine-fed mouse kidney compared with control. B: mRNA was extracted from the renal cortex of control and adenine-fed C57bl6 mice (n = 4 per group). Gene expression was assessed by real-time QPCR, revealing significantly increased expression of collagen I, collagen III, fibronectin, vimentin, TGF-β1, and TGF-β2. C: The increased expression of TGF-β1, TGF-β2, and collagen was associated with decreased expression of miR-141 and miR-200a but not the appropriate control, RNU6B, in kidney cortex (*P < 0.05 compared with control). (A high-quality color representation of this figure is available in the online issue.)
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Figure 8: Changes in miR-141/200a expression in the kidney in the adenine-induced renal fibrosis model. A: Trichrome staining of tissue sections from renal cortex from control and adenine-fed C57bl6 mice after 4 weeks of treatment. Blue staining indicates high levels of collagen in the adenine-fed mouse kidney compared with control. B: mRNA was extracted from the renal cortex of control and adenine-fed C57bl6 mice (n = 4 per group). Gene expression was assessed by real-time QPCR, revealing significantly increased expression of collagen I, collagen III, fibronectin, vimentin, TGF-β1, and TGF-β2. C: The increased expression of TGF-β1, TGF-β2, and collagen was associated with decreased expression of miR-141 and miR-200a but not the appropriate control, RNU6B, in kidney cortex (*P < 0.05 compared with control). (A high-quality color representation of this figure is available in the online issue.)

Mentions: In further experiments we investigated the expression of ECM genes and miR-141/200a in the advanced renal disease associated with adenine-induced renal fibrosis. As previously described, exposure to adenine resulted in marked tubulointerstitial fibrosis (Fig. 8A, right panel) associated with a massive upregulation in the expression of ECM genes as well as the fibrogenic mediators TGF-β1 and TGF-β2 (Fig. 8B). Consistent with our earlier in vitro and in vivo observations, miR-200a was significantly decreased (Fig. 8C). miR-141 was also significantly reduced in this model.


miR-200a Prevents renal fibrogenesis through repression of TGF-β2 expression.

Wang B, Koh P, Winbanks C, Coughlan MT, McClelland A, Watson A, Jandeleit-Dahm K, Burns WC, Thomas MC, Cooper ME, Kantharidis P - Diabetes (2010)

Changes in miR-141/200a expression in the kidney in the adenine-induced renal fibrosis model. A: Trichrome staining of tissue sections from renal cortex from control and adenine-fed C57bl6 mice after 4 weeks of treatment. Blue staining indicates high levels of collagen in the adenine-fed mouse kidney compared with control. B: mRNA was extracted from the renal cortex of control and adenine-fed C57bl6 mice (n = 4 per group). Gene expression was assessed by real-time QPCR, revealing significantly increased expression of collagen I, collagen III, fibronectin, vimentin, TGF-β1, and TGF-β2. C: The increased expression of TGF-β1, TGF-β2, and collagen was associated with decreased expression of miR-141 and miR-200a but not the appropriate control, RNU6B, in kidney cortex (*P < 0.05 compared with control). (A high-quality color representation of this figure is available in the online issue.)
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Related In: Results  -  Collection

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Figure 8: Changes in miR-141/200a expression in the kidney in the adenine-induced renal fibrosis model. A: Trichrome staining of tissue sections from renal cortex from control and adenine-fed C57bl6 mice after 4 weeks of treatment. Blue staining indicates high levels of collagen in the adenine-fed mouse kidney compared with control. B: mRNA was extracted from the renal cortex of control and adenine-fed C57bl6 mice (n = 4 per group). Gene expression was assessed by real-time QPCR, revealing significantly increased expression of collagen I, collagen III, fibronectin, vimentin, TGF-β1, and TGF-β2. C: The increased expression of TGF-β1, TGF-β2, and collagen was associated with decreased expression of miR-141 and miR-200a but not the appropriate control, RNU6B, in kidney cortex (*P < 0.05 compared with control). (A high-quality color representation of this figure is available in the online issue.)
Mentions: In further experiments we investigated the expression of ECM genes and miR-141/200a in the advanced renal disease associated with adenine-induced renal fibrosis. As previously described, exposure to adenine resulted in marked tubulointerstitial fibrosis (Fig. 8A, right panel) associated with a massive upregulation in the expression of ECM genes as well as the fibrogenic mediators TGF-β1 and TGF-β2 (Fig. 8B). Consistent with our earlier in vitro and in vivo observations, miR-200a was significantly decreased (Fig. 8C). miR-141 was also significantly reduced in this model.

Bottom Line: TGF-β1 and TGF-β2 also downregulated expression of miR-200a.The renal expression of miR-141 and miR-200a was also reduced in mouse models representing early and advanced kidney disease. miR-200a and miR-141 significantly impact on the development and progression of TGF-β-dependent EMT and fibrosis in vitro and in vivo.These miRNAs appear to be intricately involved in fibrogenesis, both as downstream mediators of TGF-β signaling and as components of feedback regulation, and as such represent important new targets for the prevention of progressive kidney disease in the context of diabetes.

View Article: PubMed Central - PubMed

Affiliation: JDRF Danielle Alberti Memorial Centre for Diabetes Complications, Melbourne, Australia.

ABSTRACT

Objective: Progressive fibrosis in the diabetic kidney is driven and sustained by a diverse range of profibrotic factors. This study examines the critical role of microRNAs (miRNAs) in the regulation of the key fibrotic mediators, TGF-β1 and TGF-β2.

Research design and methods: Rat proximal-tubular epithelial cells (NRK52E) were treated with TGF-β1 and TGF-β2 for 3 days, and expression of markers of epithelial-to-mesenchymal transition (EMT) and fibrogenesis were assessed by RT-PCR and Western blotting. The expression of miR-141 and miR-200a was also assessed, as was their role as translational repressors of TGF-β signaling. Finally, these pathways were explored in two different mouse models, representing early and advanced diabetic nephropathy.

Results: Both TGF-β1 and TGF-β2 induced EMT and fibrogenesis in NRK52E cells. TGF-β1 and TGF-β2 also downregulated expression of miR-200a. The importance of these changes was demonstrated by the finding that ectopic expression miR-200a downregulated smad-3 activity and the expression of matrix proteins and prevented TGF-β-dependent EMT. miR-200a also downregulated the expression of TGF-β2, via direct interaction with the 3' untranslated region of TGF-β2. The renal expression of miR-141 and miR-200a was also reduced in mouse models representing early and advanced kidney disease.

Conclusions: miR-200a and miR-141 significantly impact on the development and progression of TGF-β-dependent EMT and fibrosis in vitro and in vivo. These miRNAs appear to be intricately involved in fibrogenesis, both as downstream mediators of TGF-β signaling and as components of feedback regulation, and as such represent important new targets for the prevention of progressive kidney disease in the context of diabetes.

Show MeSH
Related in: MedlinePlus