Mol Ther. 2014 Apr;22(4):842-53. doi: 10.1038/mt.2013.235.

MicroRNA-29b inhibits diabetic nephropathy in db/db mice.

Chen HY1, Zhong X1, Huang XR2, Meng XM1, You Y1, Chung AC2, Lan HY3.

  • 1Department of Medicine and Therapeutics, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong, China.
  • 21] Department of Chemical Pathology, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong, China [2] CUHK Shenzhen Research Institute, Shenzhen, The Chinese University of Hong Kong, Hong Kong, China.
  • 31] Department of Medicine and Therapeutics, Li Ka Shing Institute of Health Sciences, The Chinese University of Hong Kong, Hong Kong, China [2] CUHK Shenzhen Research Institute, Shenzhen, The Chinese University of Hong Kong, Hong Kong, China.

 

ABSTRACT:

Inflammation and its consequent fibrosis are two main features of diabetic nephropathy (DN), but target therapy on these processes for DN remains yet ineffective. We report here that miR-29b is a novel therapeutic agent capable of inhibiting progressive renal inflammation and fibrosis in type 2 diabetes in db/db mice. Under diabetic conditions, miR-29b was largely downregulated in response to advanced glycation end (AGE) product, which was associated with upregulation of collagen matrix in mesangial cells via the transforming growth factor-β (TGF-β)/Smad3-dependent mechanism. These pathological changes were reversed by overexpressing miR-29b, but enhanced by knocking-down miR-29b. Similarly, loss of renal miR-29b was associated with progressive diabetic kidney injury, including microalbuminuria, renal fibrosis, and inflammation. Restored renal miR-29b by the ultrasound-based gene therapy was capable of attenuating diabetic kidney disease. Further studies revealed that inhibition of Sp1 expression, TGF-β/Smad3-dependent renal fibrosis, NF-κB–driven renal inflammation, and T-bet/Th1-mediated immune response may be mechanisms associated with miR-29b treatment in db/db mice. In conclusion, miR-29b may play a protective role in diabetic kidney disease and may have therapeutic potential for diabetic kidney complication.

PMID: 24445937

 

SUPPLEMENT:

Diabetic nephropathy is a leading cause of end-stage of kidney disease and is a major medical problem worldwide. However, treatment of diabetic nephropathy remains non-specific and non-effective. Increasing evidence shows that renal inflammation and its consequent fibrosis are a critical process leading to diabetic nephropathy. In this study, we identified that loss of miR-29b in the diabetic kidney enhanced this process with progressive renal injury in db/db mice, which was inhibited by miR-29b therapy. Findings from this study demonstrated a protective role and therapeutic potential of miR-29b for diabetic nephropathy.

As illustrated in Figure 1, there are three significant findings regarding the protective mechanisms and therapeutic actions of miR-29b, which are outlined below.

First, inhibition of TGF-β/Smad3-mediated renal fibrosis may be a mechanism associated with a protective effect of miR-29b on DN. It is well established that miR-29 directly targets the 3′UTRs of collagen I/III/IV, fibrillin, and elastin. Therefore, overexpression of miR-29 can repress the expression of collagens I and IV directly as shown in this study. We have previously demonstrated that Smad3 can physically interact with the miR-29b promoter and negatively regulates its expression in response to TGF-β1 and in turn miR-29 can also directly inhibit TGF-β1 transcription [1, 2]. Therefore, miR-29b may exert its protective effect on diabetic nephropathy by negatively regulating TGF-β/Smad3-mediated renal fibrosis. In addition, cooperation between Smad3 and Sp1 has been shown to enhance the TGF-β-induced fibrotic response [3]. Thus, inhibition of Sp1 expression, thereby blocking the Sp1-Smad3 interaction, may be also attributed to the inhibitory effect of miR-29b on TGF-β/Smad3-mediated renal fibrosis in the diabetic kidney of db/db mice.

Second, inhibition of Sp1/NF-κB–driven renal inflammation could be another mechanism whereby miR-29b inhibited diabetic kidney disease. It is now well accepted that diabetes mellitus is a low grade inflammatory disease. In the present study, we revealed that upregulation of proinflammatory cytokines and accumulation of inflammatory macrophages in the diabetic kidney were associated with a loss of miR-29b and activation of the Sp1/NF-κB pathway. Because miR-29 is negatively regulated by the NF-κB-YY1-miR-29 regulatory circuit [4], downregulation of miR-29b in the diabetic kidney may also be NF-κB–dependent. Since Sp1 functions as a key transcriptional factor to activate the NF-κB pathway and is also a target of miR-29b. Thus, miR-29b treatment suppressed renal inflammation via the Sp1/NF-κB/miR-29b autoregulatory loop.

Finally, we also found that inhibition of Th1-mediated immune response may be a new mechanism whereby miR-29b protects against DN. It has been reported that T cells do participate in immune response in diabetic kidneys of db/db mice. In this study, we also found upregulation of a Th1 transcriptional factor T-bet and an increase in IFN-γ in the diabetic kidney of db/db mice was associated with downregulation of miR-29b. Whereas, overexpression of renal miR-29b attenuated Th1-mediated immune response by inhibiting the T-bet-IFN-γ pathway. Thus, the therapeutic effect of miR-29b on DN may be attributed to its inhibitory effect on T-bet–dependent Th1-mediated renal injury.

In summary, as shown in the Figure 1, this study reveals that loss of renal miR-29b may contribute to progressive renal injury in db/db mice, which may be regulated by both TGF-β/Smad3 and NF-κB signaling pathways. Restored intrarenal levels of miR-29b by ultrasound-mediated gene therapy is capable of inhibiting DN by blocking TGF-β/Smad3-mediated renal fibrosis, NF-κB–driven renal inflammation, and T-bet–/IFN-γ–dependent immune response in db/db mice. Taken together, miR-29b may represent a novel therapeutic approach for DN.

Acknowledgements: This work was supported by grants from Major State Basic Research Development Program of China (973 program, No. 2012CB517700); The Shenzhen Basic Research Program (SZSITC, JC201104220290A); the Research Grant Council of Hong Kong (GRF469110, CUHK5/CRF09, and CUHK3/CRF/12R); NSFC/RGC Joint Research Scheme (N_CUHK404/10); and the Focused Investment Scheme A and B from the Chinese University of Hong Kong.

Contact
Professor Hui Y Lan, M.D., Ph.D.
Department of Medicine and Therapeutics, Li Ka Shing Institute of Health Sciences,
The Chinese University of Hong Kong,
Shatin, New Territories,
Hong Kong, China.
E-mail: hylan@cuhk.edu.hk
PowerPoint PresentationFigure 1. miR-29b inhibits diabetic nephropathy by blocking the TGF-β/Smad-Sp1/NF-κB-Th1 pathways. Activation of TGF-β/Smad3 and Sp1/NF-κB in the diabetic kidney negatively regulates miR-29b expression, while overexpression of miR-29b blocks TGF-b/Smad3-mediated renal fibrosis, Sp1/NF-kB-driven renal inflammation, and T-bet-dependent Th1-mediated immune injury in DN via the negatively regulatory loop.

 

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