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 8

Hyperactivation of mTORC1 induces ER stress and EMT-like phenotype in podocytes.

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Hyperactivation of mTORC1 induces ER stress and EMT-like phenotype in po...
(A) EMT-like phenotypic changes were induced in PcTSC1KO podocytes. Double staining with desmin and WT1 antibodies was performed in renal tissues from the indicated animals and analyzed by confocal microscopy (top row). Arrowheads indicate desmin expression in the podocytes. Rapamycin treatment was performed for a week. ZO-1 expression was analyzed by confocal microscopy (bottom row). Arrowheads indicate the liner expression pattern of ZO-1. (B) Detection of nephrin and podocin mRNA in PcTSC1KO urinary sediments. Twenty-four-hour urine samples were collected from the indicated animals. RT-PCR was performed using purified mRNA from the sediments. Glomerular mRNA was used as a positive control (p). (C) Accumulation of GRP78 in PcKOTsc1 podocytes (3 weeks of age). Double staining using anti-GRP78 and anti-synaptopodin antibodies in the indicated glomeruli is shown. Rapamycin treatment was performed for 1 week. (D) PBA treatment protects podocyte loss in PcKOTsc1 mice. Staining with GRP78 plus synaptopodin, nephrin, H&E, and WT1 is shown in the indicated animals (8 weeks of age). PBA treatment (400 mg/kg/d, i.p.) was performed for 6 weeks. (E) Podocyte number in the indicated glomerular sections from 8-week-old animals was measured. The ratio (number of WT1-positive cells/glomerular tuft area [μm2]) was determined in 15~30 glomeruli from the indicated animals as in Figure 3E. *P < 0.001 versus other groups, mean ± SEM, n = 4~8. (F) Urinary albumin concentrations were measured in the indicated animals. *P < 0.001 versus other groups; no statistical significance was observed between KO and KO with PBA treatment; mean ± SEM, n = 4~8. Original magnification, ×400 (A, C, and D).