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Abstract: FR-PO205

Effect of Exosomes from High Glucose-Treated Mesangial Cells on Healthy Podocytes

Session Information

Category: Diabetic Kidney Disease

  • 601 Diabetic Kidney Disease: Basic

Authors

  • Novaes, Antonio S., Federal University of São Paulo, São Paulo, Brazil
  • Hosni, Nicole Dittrich, Federal University of São Paulo, São Paulo, Brazil
  • Felizardo, Raphael F., Federal University of São Paulo, São Paulo, Brazil
  • Camara, Niels Olsen Saraiva, Federal University of São Paulo, São Paulo, Brazil
  • Boim, Mirian A., Federal University of São Paulo, São Paulo, Brazil
Background

Mesangial cells can communicate with podocytes in vivo, contributing to podocytes damage in the diabetic environment. This paracrine communication may have a relevant role in the diabetic nephropathy (DN). This study investigated whether exosomes secreted by high glucose-treated mouse mesangial cells (MMC) are able to induce dysfunction in normal podocytes.

Methods

MMC were cultured under standard (SG, 5 mM) or high glucose concentrations (HG, 30 mM) for 24 hr. Exosomes (Exos) secreted to the culture medium (SG-Exos or HG-Exos) were purified by ultracentrifugation. The vesicles size/concentration ratio was estimated by the particle tracking (NanoSigth). Vesicles characterization was performed by the presence of markers CD63 and CD81 by western blot. Podocytes in culture were exposed to either SG-Exos or HG-Exos for 24hr. In parallel, HG-MMC and podocytes were co-cultured using a transwell system. Exos secretion by MMC was inhibited by GW4869. Expressions of podocytes makers (actinin IV, p-cadherin and synaptopodin) and profibrotic markers (desmin, TGF-β1 and collagen IV) were analyzed by qPCR. Levels of synaptopodin, desmin, vimentin, podocin, alpha-actinin 4 and nephrin was determined by western blot. Expressions of ZO-1 and Nephrin were evaluated by immunofluorescence.

Results

HG stimulus induced a change in the amount, but not in the size of Exos released by MMC. HG-Exos induced phenotypic transition of podocytes that underwent epithelial mesenchymal transition, demonstrated by a downregulation of actinin 4, p-cadherin, synaptopodin together with an upregulation of desmin and TGF-β1. HG-Exos induced increases in vimentin and desmin proteins while protein expressions of nephrin, synaptopodin, alpha-actinin 4 and podocin were decreased. Levels of ZO-1 and nephrin were decreased in podocytes treated with HG-Exos by immunofluorescence. The co-culture experiments showed that the inhibitor of exosome secretion, attenuated the effects of HG-Exos on the expression of nephrin, synaptopodin, alpha-actinin 4 and podocin. These data suggest that Exos secreted by HG-MMC can cause dysfunction in health podocytes.

Conclusion

Results in vitro, demonstrated that exosomes can mediate the paracrine communication between MMC and podocytes, and suggest that high glucose stimulus in MMC can modified podocytes function contributing to DN.

Funding

  • Government Support - Non-U.S.