Figure 7.

Induction of apoptosis by TM5275 in human lung fibroblasts treated with or without TGF-β1. CCL-210 cells were treated with 1 ng/ml of TGF-β1 in the presence or absence of 75 μM of TM5275 for 24 hours. (A) Apoptotic cell death was analyzed by flow cytometry techniques, using an Alexa Fluor 488 Annexin V/Dead Cell Apoptosis Kit from Invitrogen. Early and late stages of apoptotic cells were calculated separately or in combination. The results are expressed as percentages of total cell number. (B) The activity of caspase-3/7 was determined with an Apo-ONE Homogenous Caspase-3/7 Assay Kit from Promega. (C) Concentrations of p53 protein were determined by Western blot analysis, and the results were semiquantified with Image J software (D). ^aSignificantly different from corresponding untreated control group. ^bSignificantly different from corresponding group treated with TM5275 alone (P < 0.05, n = 7–8). RFU, relative fluorescence units.