Abstract: FR-PO712

Exosomes from High Glucose-Treated Human Mesangial Cells Activate Renin Angiotensin System in Healthy Mesangial Cells

Session Information

Category: Glomerular

  • 1002 Glomerular: Basic/Experimental Pathology

Authors

  • Novaes, Antonio S., Federal University of São Paulo, São Paulo, SÃO PAULO, Brazil
  • Borges, Fernanda Teixeira, UNIFESP, São Paulo, Brazil
  • Boim, Mirian A., UNIFESP, São Paulo, Brazil
Background

High glucose (HG) induced-intracellular angiotensin II (Ang II) accumulation is correlated with upregulation of Ang II target genes, such as pro-fibrotic cytokines. This effect can be propagated via microRNAs and peptides transferred to other cells via exosomes (Ex), which play a key role in cellular communication under physiological and pathological conditions.

Methods

To verify whether exosomal signaling initiated in HG stimulated human mesangial cells (HCM) would affect control cells function, HMC were cultured under standard (5 mM) or HG (30 mM) concentrations for 24 hr. Ex secreted to culture medium were purified by ultracentrifugation and analyzed by electron microscopy. The vesicles size/concentration ratio was determined by the particle tracking using a nanoparticles analyzer and the Ex were characterized by the presence of CD63 and CD81 by western blot. Ex from control (C-Ex) or HG stimulated-HMC (Ex-HG) were labeled with PKH26 and added in normal HMC. The Ex internalization was evaluated by confocal microscopy (CM). The presence angiotensinogen (AGT), renin and ACE in the Ex was analyzed by western blot. The bioactivity of the Ex was evaluated in Chinese Hamster Ovary cells (CHO-K1), which do not express components of RAS, and in CHO-K1 transfected ECA (ECA-CHO) in the presence of C-Ex or HG-Ex. The synthesis of Ang II in ECA-CHO was analyzed by CM. Expressions of fibronectin, AGT, renin, AT1, AT2 receptors and proliferation were used to assess the cellular response to signal transferred through the Ex in control HMC exposed to C-Ex or HG-Ex.

Results

HG stimulus induced a change in the amount, but not in the size of Ex. HG-Ex are internalized by normal HMC. C-Ex contained AGT and renin proteins, whose expressions were increased in cells exposed to HG-Ex. ACE was not detected in C-Ex and HG-Ex. The exposure of HG-Ex to ECA-CHO resulted in Ang II formation in these cells. The expression levels of fibronectin, AGT, renin, AT1 and AT2 were higher in control HMC treated with HG-Ex compared with those treated with C-Ex, indicating that HG-Ex can modify the function of target control HMC.

Conclusion

These results suggest that the intercellular communication through the exosomes may have pathophysiological implications in the diabetic kidney.

Funding

  • Government Support - Non-U.S.