Evidence of Caveolin-1/Caveolae-mediated migration during early stages of placentation

RECA ALEJANDRA; REPPETTI JULIETA; IBARRA CRISTINA; MARTÍNEZ NORA; DAMIANO ALICIA E

Oregón

Congreso; International Federation of Placenta Associations; 2016

Migration plays key roles in many physiological and pathological processes including development, angiogenesis, tissue regeneration and metastasis. Caveolin-1 (Cav-1) and caveolae constitute domains that compartmentalize intracellular signalling pathways to orchestrate different cellular events. During placentation the human extravillous trophoblast (EVT) cells invade the endometrium and maternal vasculature within the uterus. Cytotrophoblasts express Cav-1 and differentiation into syncytium results in a decrease of its expression and a marked reduction of caveolae. Objective: To evaluate the role of Cav-1 in the migration and invasion of trophoblastic cellsMethods: Trophoblastic cells (Swan 71 cell line, derived by telomerase-mediated transformation of a 7 week cytotrophoblast isolate) were cultured in complete DMEM-F12. Cav-1 expression was examined by immunocytochemistry and western blot. Cells were treated with 10 mM methyl-β-cyclodextrin (MCD) for 60 min to disrupt caveolae. Migration was assessed by wound healing assay. TScratch® software was used to quantify the data and scratch distance at 0 h was considered as 0 % of gap closure. Representative images were taken after 8, 18 and 24 h. To evaluate invasion, trophoblast cells were seeded in the top of transwell inserts of 8-µm pore precoated with Matrigel (0.3 mg/ml) and housed in a 24-well plate. After 24 h, cells located on the undersurface were fixed with methanol and stained with hematoxylin. Results: Wound healing assay: the closure of the gap expressed as Control vs. MCD at each time point was: 8 h (32.78 ± 1.14 vs 17.78 ± 0.5561), 18 h (66.74 ± 1.434 vs 51.79 ± 1.158), 24 h (84.45 ± 1.160 vs 64.95 ± 0.7165). Invasion assay: No differences were observed between control and MCD treated cells. Conclusion: These results show that lipid raft disruption significantly attenuates migration (but no invasion) in Swan 71 cells, suggesting a role for Cav-1/caveolae during the early stages of placental development.