mTORC1 activation in podocytes is a critical step in the development of diabetic nephropathy in mice
Ken Inoki, Hiroyuki Mori, Junying Wang, Tsukasa Suzuki, SungKi Hong, Sei Yoshida, Simone M. Blattner, Tsuneo Ikenoue, Markus A. Rüegg, Michael N. Hall, David J. Kwiatkowski, Maria P. Rastaldi, Tobias B. Huber, Matthias Kretzler, Lawrence B. Holzman, Roger C. Wiggins, Kun-Liang Guan
Ken Inoki, Hiroyuki Mori, Junying Wang, Tsukasa Suzuki, SungKi Hong, Sei Yoshida, Simone M. Blattner, Tsuneo Ikenoue, Markus A. Rüegg, Michael N. Hall, David J. Kwiatkowski, Maria P. Rastaldi, Tobias B. Huber, Matthias Kretzler, Lawrence B. Holzman, Roger C. Wiggins, Kun-Liang Guan
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Research Article

mTORC1 activation in podocytes is a critical step in the development of diabetic nephropathy in mice

Abstract

Diabetic nephropathy (DN) is among the most lethal complications that occur in type 1 and type 2 diabetics. Podocyte dysfunction is postulated to be a critical event associated with proteinuria and glomerulosclerosis in glomerular diseases including DN. However, molecular mechanisms of podocyte dysfunction in the development of DN are not well understood. Here we have shown that activity of mTOR complex 1 (mTORC1), a kinase that senses nutrient availability, was enhanced in the podocytes of diabetic animals. Further, podocyte-specific mTORC1 activation induced by ablation of an upstream negative regulator (PcKOTsc1) recapitulated many DN features, including podocyte loss, glomerular basement membrane thickening, mesangial expansion, and proteinuria in nondiabetic young and adult mice. Abnormal mTORC1 activation caused mislocalization of slit diaphragm proteins and induced an epithelial-mesenchymal transition–like phenotypic switch with enhanced ER stress in podocytes. Conversely, reduction of ER stress with a chemical chaperone significantly protected against both the podocyte phenotypic switch and podocyte loss in PcKOTsc1 mice. Finally, genetic reduction of podocyte-specific mTORC1 in diabetic animals suppressed the development of DN. These results indicate that mTORC1 activation in podocytes is a critical event in inducing DN and suggest that reduction of podocyte mTORC1 activity is a potential therapeutic strategy to prevent DN.

Authors

Ken Inoki, Hiroyuki Mori, Junying Wang, Tsukasa Suzuki, SungKi Hong, Sei Yoshida, Simone M. Blattner, Tsuneo Ikenoue, Markus A. Rüegg, Michael N. Hall, David J. Kwiatkowski, Maria P. Rastaldi, Tobias B. Huber, Matthias Kretzler, Lawrence B. Holzman, Roger C. Wiggins, Kun-Liang Guan

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Figure 2

Characterization of PcKOTsc1 mice.

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Characterization of PcKOTsc1 mice. (A) Enhanced S6 phosphorylation, podo...
(A) Enhanced S6 phosphorylation, podocyte hypertrophy, and mesangial expansion in PcKOTsc1 glomeruli. Renal tissues from 4-week-old mice of the indicated genotypes were stained with phospho-S6, WT1, H&E, and PAS as indicated. Representative glomeruli from wild-type, PcKOTsc1 (KO), and rapamycin-treated PcKOTsc1 (KO+rapa) are shown. Rapamycin treatment (1 mg/kg, i.p., 3 times/wk) was performed from 2 to 4 weeks of age. (B) S6 but not Akt phosphorylation was enhanced in PcKOTsc1 glomeruli. Levels of phospho-S6, S6, phospho-Akt (Ser473), and Akt in glomeruli from mice of the indicated genotypes were determined by Western blot. The levels of phosphorylation of each protein were quantified. Data are expressed as fold increase in the ratio of phospho-protein to total protein. *P < 0.001 versus other groups; mean ± SEM, n = 3. (C) Enhanced matrix protein mRNAs in PcKOTsc1 glomeruli. The expression levels of fibronectin (FN) and type IV collagen (COL4) mRNAs in the indicated mouse glomeruli were determined by quantitative RT-PCR. Data were normalized to Hprt1. *P < 0.001 versus other groups; mean ± SEM, n = 7. (D) Enhanced deposition of the matrix proteins in PcKOTsc1 glomeruli. Representative glomerular images of staining for fibronectin and type IV collagen in the indicated animals are shown. The ratio of positive staining area to glomerular area in the indicated animals was determined (10 glomeruli/mouse, 3 mice/group). *P < 0.001 versus other groups, mean ± SEM. Original magnification, ×400 (A and D).