Recombinant tgf β1

For Transwell coculture experiments, 2 × 10⁵ MSCs were planted in the upper chambers of 0.4-um pore size Transwells (Corning, Lowell, MA). Afterward, 2 × 10⁵ mouse lung fibroblasts (NIH3T3 cells) or mouse lung epitheliums (MLE-12 cells) were placed in six-well culture plates. MSCs were treated with 3 mM NMDA for 24 h. NIH3T3 cells or MLE-12 cells were treated with or without 10 ng/ml of recombinant TGF-β1 (MyBioSource, San Diego, CA) for 24 h. After the medium was reomved and the cells were washed with PBS, the coculture experiments of MSCs with NIH/3T3 cells or MLE-12 cells were conducted in DMEM/F-12 with 2% FBS, 1% l-glutamine, and 1% solution of penicillin-streptomycin in a humidified CO₂ incubator at 37°C for 24 h. During coculture, 10 ng/ml recombinant hepatocyte growth factor (HGF) from R&D Systems (Abingdon, UK) were used to treat NIH/3T3 cells or MLE-12 cells.

Tunicamycin (TN), thapsigargin (TG), mouse anti-vimentin Cy 3 conjugated (#C5992), and mouse anti-pan cytokeratin FITC conjugated antibody (#C9080) were obtained from Sigma-Aldrich (St. Louis, MO). Anti-HGF affinity purified polyclonal antibody (#AF-294-SP) and recombinant HGF was purchased from R&D Systems (Abington U.K), and recombinant TGF-β1 was from MyBiosource Inc, (San Diego, USA).