SAT651 Human Adipose-derived Extracellular Vesicles (AdEVs) Increase Proinflammatory Markers In Renal And Endothelial Cells: An In Vitro Preliminary Study

Abstract

Abstract Disclosure: C.A. Carvajal: None. M.P. Hernandez: None. P. Carrion: None. A. Tapia: None. J.A. Lopez: None. A. Vecchiola: None. C.E. Fardella: None. A. Sandoval-Borquez: None. During obesity, white adipose tissue (WAT), undergoes hypertrophic and hyperplastic changes by differentiation of preadipocytes into adipocytes, which increase the expression of PPARγ, FASN, FABP4, and adiponectin genes. These adipokines play a key role in obesity progression and in cell communication. WAT also causes a chronic inflammatory state that modifies the gene expression and secretome, including releasing of adipose-derived extracellular vesicles (AdEVs) that carry a specific cargo thought to modify different signaling pathways in target cells. Aim: To evaluate the effect of AdEVs in renal and endothelial cells and its inflammatory phenotype. Methods: Human SW872 preadipocytes and differentiated adipocytes were cultured and characterized by optical microscopy and Oil-Red-O stain. EVs from both preadipocytes and adipocytes were isolated by ultracentrifugation and characterized by nanoparticle tracking analysis, electron microscopy and w-blot. AdEVs (1x10^3) were added to renal HCD and endothelial EaHy926 cells for 24 hours. Expression of adipogenic differentiation genes in preadipocytes, adipocytes and their EVs, as well as the expression of inflammatory markers (IL-6 and IL-1B) were perfomed by RT-qPCR. Results: SW872 differentiated cells showed a classical adipocyte morphology and important accumulation of lipid-droplets. Isolated AdEVs have a donut shape morphology and size (50 - 150 nm) in accordance to the MISEV2018 guidelines, including also the EVs markers CD9 and Tsg101. A higher EVs concentration released from preadipocytes compared to adipocytes was observed (5.21x10^10 vs 0.44x10^10 particles/mL, p ≤0.05). The analysis of relative gene expression (2^-ΔΔCT), showed that adipocytes increase FAPB4 and adiponectin and decrease in PPARγ and FASN gene expression, with respect to preadipocytes (p ≤0.05). EVs from preadipocytes and adipocytes present similar relative expression of adiponectin, PPARγ and FASN than their parental cells. Finally, both renal or endothelial cells treated with AdEVs express higher levels of IL-6 and IL-1B that untreated cells (p≤ 0.05). Conclusion: EVs released from SW872 adipocytes and preadipocytes have similar gene expression as their parental cells. Treatment of both renal and endothelial cells with AdEVs showed an increase in inflammatory markers, as IL-6 and IL-1B. These preliminary results reveals novel insights about the impact of AdEVs in renal collecting duct and endothelial cells, which support novel roles of AdEVs -and its cargo- in human pathophysiology associated to obesity. Acknowledgements: This study was supported by grants ANID-FONDECYT 1212006, 3200646; CONICYT-FONDEQUIP EQM150023, SOCHED & CETREN UC. Presentation: Saturday, June 17, 2023