Abstract: TH-PO695

Prolonged Exposure of Podocytes to Insulin Induces Insulin Resistance through Lysosomal and Proteasomal Degradation of the Insulin Receptor

Session Information

Category: Diabetes

  • 501 Diabetes Mellitus and Obesity: Basic - Experimental

Authors

  • Lay, Abigail Charlotte, University of Bristol, Bristol, United Kingdom
  • Hurcombe, Jenny, University of Bristol, Bristol, United Kingdom
  • Østergaard, Mette Viberg, University of Bristol, Bristol, United Kingdom
  • Barrington, Fern, University of Bristol, Bristol, United Kingdom
  • Lennon, Rachel, University of Manchester, Manchester, United Kingdom
  • Welsh, Gavin Iain, University of Bristol, Bristol, United Kingdom
  • Coward, Richard, University of Bristol, Bristol, United Kingdom
Background

Podocytes are insulin responsive cells of the glomerular filtration barrier and are key in preventing albuminuria, a hallmark feature of diabetic nephropathy. While there is evidence that a loss of insulin signalling to podocytes is detrimental, the molecular mechanisms underpinning the development of podocyte insulin resistance in diabetes remain unclear. Thus, we aimed to further investigate podocyte insulin responses early in the context of diabetic nephropathy.

Methods

Conditionally immortalised human and mouse podocyte cell lines and glomeruli isolated from db/db DBA2J mice were studied. Podocyte insulin responses were investigated with western blotting, cellular glucose uptake assays and automated fluorescent imaging of the actin cytoskeleton. Q-PCR was employed to investigate changes in mRNA. Human cell lines stably overexpressing the IR and nephrin were also generated, using lentiviral constructs.

Results

Podocytes exposed to a diabetic environment (high glucose, high insulin and the pro-inflammatory cytokines TNF-α and IL-6) become insulin resistant with respect to glucose uptake and activation of PI3K and MAPK signalling. These podocytes lose expression of the insulin receptor (IR) as a direct consequence of prolonged exposure to high insulin concentrations, which causes an increase in IR protein degradation via a proteasome-dependent and bafilomycin-sensitive pathway. Reintroducing the IR into insulin resistant human podocytes rescues upstream phosphorylation events, but not glucose uptake. Stable expression of nephrin is also required for the insulin-stimulated glucose uptake response in podocytes and for efficient insulin-stimulated remodelling of the actin cytoskeleton.

Conclusion

Together these results suggest that IR degradation, caused by high levels of insulin, drives early podocyte insulin resistance and that both the IR and nephrin are required for full insulin sensitivity of this cell. This could be highly relevant for the development of nephropathy in diabetic patients and patients with the metabolic syndrome who are commonly hyperinsulinaemic in the early phases of their disease.

Funding

  • Government Support - Non-U.S.