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Abstract: TH-PO868

Diabetic Conditions Enhance the Phosphorylation of PACSIN2/Syndapin II in Podocytes

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Lehtonen, Sanna H., University of Helsinki, Helsinki, Finland
  • Dumont, Vincent, University of Helsinki, Helsinki, Finland
  • Lindfors, Sonja, University of Helsinki, Helsinki, Finland
  • Forsblom, Carol, Folkhalsan Research Center, Helsinki, Finland
  • Kawachi, Hiroshi, Department of Cell Biology, Kidney Research Center, Niigata University Graduate School of Medical and Dental Sciences, Niigata, Japan
  • Groop, Per-Henrik, University of Helsinki and Helsinki University Hospital, Helsinki, Finland
  • Suetsugu, Shiro, Nara Institute of Science and Technology, Nara, Japan
Background

We previously found that PACSIN2/syndapin II, which regulates endocytosis and intracellular trafficking, is elevated in glomeruli of Zucker Diabetic Fatty (ZDF) rats showing aberrant localization of nephrin and severe albuminuria. We also showed in vitro that PACSIN2 overexpression reduces nephrin insertion at the plasma membrane. PACSIN2 can be phosphorylated at serine 313 (pS313-PACSIN2) by Protein Kinase C alpha (PKCα), which in turn regulates the endocytic activity of PACSIN2. Importantly, PKCα is elevated in diabetes and associated with loss of nephrin expression. The aim of this study was to evaluate the phosphorylation status of PACSIN2 in glomeruli in diabetes. We also assessed the regulation of pS313-PACSIN2 and the presence of nephrin at the plasma membrane after treating cultured podocytes with palmitate, elevated in patients with diabetes.

Methods

PKCα, total PACSIN2 and pS313-PACSIN2 levels were assessed by Western blotting. On-Cell Western, a 96-well plate -based surface labeling assay, was used to determine the insertion of nephrin at the plasma membrane. Glomeruli were isolated from lean and obese ZDF rats by graded sieving. Cultured podocytes were incubated with palmitic acid or with sera from patients with T2D having normo- or microalbuminuria.

Results

PKCα, total PACSIN2 and pS313-PACSIN2 were elevated in glomeruli of obese and diabetic ZDF rats compared to lean controls. Also, the level of free fatty acids was increased in the sera of obese ZDF rats. Increased pS313-PACSIN2 and a trend for an increase of total PACSIN2 were observed when human podocytes were incubated with sera from patients with T2D with microalbuminuria compared to normoalbuminuric patients. Treating podocytes with palmitic acid resulted in the time-dependent phosphorylation of PACSIN2 on S313. Palmitic acid treatment also resulted in the reduction of nephrin inserted at the plasma membrane.

Conclusion

Our results indicate that PACSIN2 and its phosphorylation at serine 313 are elevated in glomeruli in a rat model of diabetes and diabetic kidney disease. This could be a consequence of elevated circulating palmitate and result in the reduction of nephrin at the plasma membrane, causing the destabilization of the slit diaphragm and albuminuria.

Funding

  • Private Foundation Support