FIGURE 1.

10 ng/ml 72-h TGF-β1 induces stable myofibroblast differentiation, and prevention/reversal of myofibroblast phenotype is induced by 400 ng/ml 72-h BMP7. A–C, fibroblasts were grown to ∼70% confluence before undergoing growth arrest in serum-free medium for 48 h. A, medium was replaced with serum-free medium containing 0, 5, 10, 20, 30, and 50 ng/ml TGF-β1 and the incubations continued for 72 h. B, medium was replaced with serum-free medium containing 10 ng/ml TGF-β1 for 0, 8, 12, 24, 48, and 72 h. C, medium was then replaced with serum-free medium containing 10 ng/ml TGF-β1 for 72 h and then subsequently again replaced with serum-free medium, and the incubations continued for a further 0, 24, 48, and 72 h. The cells were then fixed and stained for α-SMA and viewed under UV light. D–F, fibroblasts were grown to ∼70% confluence before undergoing growth arrest in serum-free medium for 48 h. D, cells were treated with 0, 50, 100, 200, 400, and 800 ng/ml BMP7 and the incubations continued for 72 h; then medium was replaced with serum-free medium containing 10 ng/ml TGF-β1 for 72 h. E, cells were treated with 10 ng/ml TGF-β1 for 72 h before undergoing treatment with 0, 50, 100, 200, 400, and 800 ng/ml BMP7 for a further 72 h. F, cells were treated with 400 ng/ml BMP7 for 0, 8, 12, 24, 48, and 72 h; then medium was replaced with serum-free medium containing 10 ng/ml TGF-β1 for a further 72 h. RNA extraction and RT-QPCR assessment for α-SMA mRNA expression was subsequently performed. Control experiments included cells treated with serum-free medium alone, cells treated with 10 ng/ml TGF-β1 alone, and cells treated with 400 ng/ml BMP7 alone. White scale bars represent 20 micrometers.