Cc 2935

MCF7 cells (HTB-22, ATCC) and HeLa cells (CCL-2, ATCC) were cultured in DMEM media (ATCC) supplemented with 10% fetal bovine serum (FBS, VWR) and 1% penicillin/streptomycin (Thermo Fisher Scientific). Human umbilical vein endothelial cells (HUVEC, CC-2935, Lonza) were cultured in EGM⁺ media (Lonza) supplemented with 1% penicillin/streptomycin using collagen-coated flasks (Corning). All cells were housed in 5% CO₂ humidified atmosphere at 37 °C. Cell lines were authenticated with STR profiling and tested negative for mycoplasma contamination prior to the study.

Trophoblast and endothelial cells were used to develop an in vitro 3D placental microfluidic culture system that would recapitulate the human placental microenvironment. The chorionic villi-derived first-trimester human placenta HTR8/SVneo trophoblast cell line (CRL-3271, ATCC, Manassas, VA, USA), commonly used as a trophoblast cell model (Graham et al. 1993 (link); Msheik et al. 2020 (link); Wong et al. 2019 (link)). While this cell line has been reported to include a heterogeneous cell population (Abou-Kheir et al. 2017 (link)), it continues to be used as the primary cell model for trophoblast cell invasion studies (Li et al. 2015 (link); Wong et al. 2019 (link); Zhao et al. 2018 (link)). Cells were proliferated by seeding them into 100 mm plates at 500,000 cells per dish and cultured in growth medium (DMEM/F12 supplemented with 1 % penicillin-streptomycin, 2 mM L-glutamine, 10 % FBS, and 10 mM HEPES) at 37 °C, 5 % CO₂. Human umbilical vein endothelial cells (HUVECs, CC-2935, Lonza, Minneapolis, MN, USA) were used as the endothelial cell layer and were grown in complete medium (basal medium (CC-3156, Lonza, Minneapolis, MN, USA) supplemented with EGM-2 Plus medium (CC-4176, Lonza, Minneapolis, MN, USA)). All reagents were purchased from Thermo Fisher unless otherwise stated.