4.1. Cell Culture and Treatments

The mouse endothelial cell line also known as EOMA cells was purchased from ATCC (Manassas, VA, USA), which was isolated originally from mouse hemangioendothelioma. These EOMAs were cultured in Dulbecco’s modified Eagle’s medium (Gibco, Carlsbad, CA, USA) with 1% penicillin–streptomycin (Thermo Fischer Scientific, Carlsbad, CA, USA) and supplemented with 10% fetal bovine serum (Thermo Fischer Scientific), and were placed in a humidifier at 37 °C and supplied with 95% air and 5% CO₂ mixture. Cells were trypsinized with Trypsin (Trypsin/EDTA; Sigma, St. Louis, MO, USA). The EOMAs were further diluted in Dulbecco’s modified Eagle’s medium supplemented with 10% fetal bovine serum. Cells were treated with TMAO (10, 30, 100, and 300 μm) for the TMAO stimulation, and then incubated overnight [9]. In glycyrrhizin (gly) cell groups, cells were pretreated with gly (120 µM) for thirty minutes.