Hyperosmolarity induces caveolae disruption impairing human extravillous trophoblast differentiation

REPPETTI J.; MEDINA Y.; FARINA M.; DAMIANO A. E.; MARTINEZ NORA.

Congreso; International Federation of Placenta Associations 2021; 2021

HYPEROSMOLARITY INDUCES CAVEOLAE DISRUPTION IMPAIRING HUMAN EXTRAVILLOUS TROPHOBLAST DIFFERENTIATIONReppetti Julieta1, Medina Yollyseth1, Farina Mariana2, Damiano Alicia E.1,3, Martínez Nora1.1 Laboratorio de Biología de la Reproducción, Instituto de Fisiología y Biofísica Bernardo Houssay (IFIBIO Houssay) UBA-CONICET. Facultad de Medicina, Universidad de Buenos Aires. Buenos Aires, Argentina. 2 Laboratorio de Fisiopatología Placentaria, CEFYBO UBA-CONICET, Facultad de Medicina, Universidad de Buenos Aires. Buenos Aires, Argentina.3 Cátedra de Biología Celular y Molecular. Departamento de Ciencias Biológicas, Facultad de Farmacia y Bioquímica, Universidad de Buenos Aires. Buenos Aires, Argentina.A hyperosmotic microenvironment is related to cell differentiation in many tissues. Emerging evidence supports, that hyperosmolarity induces the internalization of caveolae into the cytoplasm and accelerates their turnover. Caveolae are membrane domains involved in vesicular trafficking and signal transduction.Many signaling pathways associated with the regulation of trophoblast differentiation are localized in caveolae. Previously, we demonstrated that caveolae are involved in the migration and the endovascular differentiation of extravillous trophoblast (EVT). However, up to now, the molecular mechanism is unknown.The aim of this study was to explore the effect of hyperosmolarity on the caveolae localization and its participation in the differentiation of the EVT.Methods: Swan-71 cell line (human extravillous trophoblast cells) was cultured in complete DMEM-F12 and 100mM of sucrose was added to generate the hyperosmolar condition. Cell viability was evaluated by 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) assay. The localization of caveolae was analyzed by Transmission Electron Microscopy (TEM). Cav-1 expression was determined by WB in different conditions (isoosmolarity or control and hyperosmolarity, with or without MG-132- a proteasome inhibitor- and NH4Cl- a lysosomal inhibitor). Migration was assessed by wound healing assay, the activity of metalloproteinases (MMPs) by zymography, and invasion was evaluated in chambers coated with Matrigel®. Endovascular differentiation was analyzed by the formation of tube-like structures in plates coated with Matrigel®.Results: In all the conditions, cell viability was not modified. Hyperosmolarity induced the internalization of caveolae and Cav-1 degradation by lysosomal proteolysis (p