Culturing Rat Vascular Smooth Muscle Cells

rVSMC primary cells (Lonza R-ASM-580; 3001900115) were cultured following the procedures described by others with slight modifications [12 , 14 (link)]. Cells were cultured in a humidified atmosphere at 37°C and 5% CO₂. Standard culture medium consisted of DMEM (Gibco 1 1966- 500), streptomycin/penicillin (Gibco 1 5630-080), and 0.3% DSMO (Sigma-Aldrich D2650), supplemented with 20% fetal bovine serum (FBS, SERADIGM 1 500-500). Cell layers were detached with 0.05% trypsin/0.02% EDTA solution (GIBCO 25200). Cells were maintained at 70–80% confluence by passing as needed. A portion of the rVSMCs was stored for later use under frozen conditions using 90% FBS (Seradigm 1 500-500) and 10% DMSO (Sigma D2650).