High glucose dmem f12 medium

Segmental bronchi were obtained from lung cancer patients undergoing pulmonary lobectomy or pneumonectomy in Zhongnan Hospital of Wuhan University. Informed consents were acquired from all patients and the study was approved by Medical Ethics Committee of Zhongnan Hospital (approval number: 2019044). Bronchial smooth muscle was dissected out and cut into small pieces, which were then placed into 25 cm 2 culture asks and cultured in RPMI-1640 medium containing L-glutamine (2 mM), penicillin (50 U/mL) and streptomycin (50 mg/mL) and supplemented with 10% fetal bovine serum (FBS). After reaching ~ 80% con uence, cells were passaged by tyrosination with 0.05% trypsin-EDTA and then grown in high glucose DMEM/F12 medium (Gibco-BRL, Carlsbad, CA, USA) supplemented with 20% FBS (Gibco-BRL). Typical hBSMCs exhibited a hill-and-valley pattern when reaching con uence. The purity of hBSMCs was con rmed by immunocytochemistry with mouse anti-human α-smooth muscle actin (SMA) antibody, and 95% cells were positively stained with anti-α-SMA (Fig. S1). hBSMCs before passage 5 were used for the experiments. In our experiments, hBSMCs were seeded in 6-well plates at a density of 5×10 4 cells/ml and cultured in DMEM/F12 with 10% FBS for 24 h. At ~ 70% con uence, cells were treated accordingly to the purpose of the experiments.