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Abstract: PO0685

miR299a-5p Is a Novel Mediator of Fibrosis in Diabetic Kidney Disease

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Nmecha, Ifeanyi Kennedy/k, McMaster University Faculty of Health Sciences, Hamilton, Ontario, Canada
  • Gao, Bo, McMaster University Faculty of Health Sciences, Hamilton, Ontario, Canada
  • Krepinsky, Joan C., McMaster University Faculty of Health Sciences, Hamilton, Ontario, Canada
Background

Diabetic kidney disease (DKD) is the leading cause of kidney failure in North America, characterized by glomerular accumulation of extracellular matrix (ECM) proteins. High glucose (HG) induction of glomerular mesangial cell (MC) profibrotic responses, mediated by the cytokines TGFβ1 and activins, plays a central role in its pathogenesis. We recently showed that TGFβ1 upregulation of microRNA (miR) 299a-5p promoted its profibrotic responses in MC. Here we studied the role of this miR in DKD.

Methods

Primary mouse MC were treated with HG at 30 mM. miR299a-5p was detected by qPCR or ISH. miR overexpression and inhibition plasmids were transfected by electroporation. TGFβ1 and activin signaling was assessed by activity of their downstream mediator Smad3 using the CAGA12 reporter. ECM production was assessed using immunoblotting and activity of the COL1α1 promoter luciferase reporter.

Results

HG increased the expression of miR299a-5p in MC. This was also increased in type 1 Akita diabetic kidneys in both glomeruli and tubules, as assessed by ISH. In MC, miR299a-5p overexpression increased Smad3 activation and COL1α1 promoter activity. Conversely, miR299a-5p inhibition attenuated HG-induced COL1α1 promoter and Smad3 activation, as well as upregulation of ECM proteins. miR299a-5p is predicted to target the TGFβ1 inhibitor Cripto-1 (CR-1), and we previously showed that it targeted the activin inhibitor follistatin (FST). Here we show that HG decreased expression of both CR-1 and FST. This was similarly seen with miR299a-5p overexpression. CR-1 or FST treatment individually attenuated the increased COL1α1 promoter and Smad3 activity seen with miR299a-5p overexpression, and together showed an additive inhibitory effect.

Conclusion

These data support an important role for miR299a-5p in regulation of the profibrotic response to HG. Through suppression of two important antifibrotic proteins, CR-1 and FST, miR299a-5p potentiates the action of TGFβ family profibrotic cytokines. Future studies will determine whether inhibition of this miR can attenuate DKD.

Funding

  • Government Support – Non-U.S.