Figure 1.

Induction of collagen gel contraction by ET-1 in primary normal lung fibroblasts. (A) Floating (FLO) gels. Normal lung fibroblasts were seeded into a collagen gel matrix, and after gel polymerization, the gel was detached from the tissue culture plate and incubated in the presence or absence of ET-1 (100 nM) for 24 h. Cycloheximide was added to the gels 1 h before addition of ET-1. ET-1 promotes contraction of floating, but not fixed, gels. ET-1–mediated contraction of floating gels required protein synthesis, because the ability of ET-1 to promote contraction was blocked by cycloheximide. In addition, at the end of the experiment, cells treated were subjected to Western blot analysis to detect α-SMA protein, which was potently induced by ET-1 treatment. Thus, ET-1 promotes ECM contraction through the protein synthesis. (B) Fixed (FIX) gels. Normal lung fibroblasts were seeded into a collagen gel matrix, and the mixture remained attached to the tissue culture plate, and after 24 h, the gel was then detached, and incubated in the presence or absence of ET-1 (100 nM) for 1 h. ET-1 induces α-SMA protein expression in floating gels. Cells treated as in were subjected to Western blot analysis to detect α-SMA protein. Expression of α-SMA is elevated in fixed, mechanically stressed gels was markedly elevated, even in the absence of added ET-1. Thus, ET-1 does not directly promote mechanocontraction of already formed α-SMA stress fibers.