A Profibrotic Phenotype in Naïve and in Fibrotic Lung Myofibroblasts Is Governed by Modulations in Thy-1 Expression and Activation

Figure 5

Proliferation rates of Thy1⁺ and Thy1⁻ myofibroblasts isolated from the fibrotic lungs. Lung cells were removed from bleomycin-treated mice at 14-day post-IT. Primary lung myofibroblast cultures were obtained and maintained in RPMI medium. The cultures were sorted by FACS using PE-conjugated anti-Thy1 (CD90) mAb to isolate Thy1⁻ and Thy1⁺ myofibroblasts. (a) Cell mass of Thy1⁺ and Thy1⁻ myofibroblasts measured after 24 h, by methylene blue staining followed by colorimetric assay. Results are presented as fold change from day 0 (). (b) CFSE staining primary cultures of Thy1⁺ and Thy1⁻ myofibroblasts were stained with CFSE. After 72 h of incubation, intensity of CFSE labeling was measured by flow cytometry. The percentage of the cells that lost CFSE labeling at days 0 and 3 is noted. (c) BrdU uptake Thy1⁺ and Thy1⁻ myofibroblasts were treated with 20 μM BrdU for 6 h. BrdU uptake was assessed by anti-BrdU-FITC-conjugated mAb staining and subjected to confocal microscopy analysis.

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