Mechanical Compression and TGF-β Signaling in HBE Cells

Poly(I:C) (Invivogen, San Diego, CA) or recombinant human (rh) TGF-β1 (10 ng/ml, Cell Signaling Technologies, Danvers, MA) was spiked into the basolateral media of HBE cells in ALI cultures [26 (link), 27 (link)]. PBS for Poly(I:C) or sodium citrate for rhTGF-β1 was used as vehicle control. As previously described [23 (link), 24 (link), 32 (link)], HBE cells were subjected to mechanical compression at a magnitude of 30 cm H₂O pressure for three hours. Time-matched controls were subjected to 0 cm H₂O pressure. In experiments where a pharmacological inhibitor of TGF-β receptor 1, SB431542 was used (10 μM, Cell Signaling Technology), this was spiked to the basolateral medium at a final concentration of 10 μM, one hour prior to exposure to either stimulation. As a vehicle control for SB431542, 0.1% DMSO was used.