Strikingly, both mTORC1 target genes and mTOR mRNA itself were induced in progressive diseases, whereas transcript levels were unchanged or even repressed in MCD compared with controls (Figure (Figure6A).6A). Consistent with this finding, immunofluorescence staining of biopsy samples for phosphorylated selleck kinase inhibitor S6 (pS6) as a marker of mTORC1 activity confirmed increased pS6 levels in podocytes of patients with diabetic nephropathy (Figure (Figure6,6, B and C). Profound activation of mTORC1 and phosphorylation of S6 was also observed by immunofluorescence staining and Western blot analysis of pS6 in an animal model of diabetic nephropathy (streptozotocin [STZ] model) (Figure (Figure6,6, D�CF), confirming the association of mTORC1 activation and diabetic nephropathy.

Figure 6 mTORC1 hyperactivation is a molecular signature of diabetic nephropathy. Podocyte-specific genetic inhibition of mTOR activation prevents progressive glomerular diseases. We next investigated whether interference with mTORC1 activation ameliorates the progression of renal disease involving the podocyte. Since complete deletion of mTORC1 in podocytes resulted in glomerular disease, we generated mice lacking only one Raptor allele in podocytes (RaptorHet podocyte) (Figure (Figure7A).7A). Western blot analysis from glomerular lysates confirmed a reduction of raptor protein and pS6 levels in RaptorHet podocyte mice (Figure (Figure7,7, B and C). To induce diabetic nephropathy, mice were injected with low-dose STZ at 8 weeks of age. Blood glucose levels, blood pressure, and glomerular macrophage infiltration were similar in RaptorHet podocyte and control mice (Supplemental Figure 4).

However, induction of diabetes resulted in a significant increase of mTORC1 activity, as evidenced by increased signals of pS6 in diabetic WT animals; pS6 levels were significantly lowered by deletion of 1 Raptor allele in RaptorHet podocyte mice (Figure (Figure7,7, D and E). While WT diabetic mice developed substantial proteinuria by 20 weeks of age, proteinuria was significantly reduced in diabetic RaptorHet podocyte mice (Figure (Figure7F).7F). Strikingly, diabetic RaptorHet podocyte mice displayed significantly reduced glomerulosclerosis scores and less mesangial matrix expansion (Figure (Figure7,7, G�CI).

Quantitative stereological analyses demonstrated that the mean podocyte volume in RaptorHet podocyte mice was reduced by about 30% compared with that in diabetic WT animals (Figure (Figure7,7, J Dacomitinib and K). It is worth noting that similar results were also observed in animal models of type 2 diabetes (36), underlining that mTORC1 deregulation is a major driving force for glomerular diseases such as diabetic nephropathy, which can be prevented by lowering mTOR activity in podocytes. Figure 7 Podocyte-specific genetic inhibition of mTOR hyperactivation prevents progressive glomerular diseases.