Figure 3

Effects of knockdown or overexpression of FASN on death of A549 alveolar epithelial cells and in the lungs of FASN transgenic mice. Stable FASN-overexpressing or FASN-knockdown cells were treated with BLM (10 μg/mL) for 24 h in serum-free medium. (A) Apoptosis was defined by annexin V+/both PI+ and PI− staining. Necrosis was defined by PI+/both Annexin V+ and Annexin V− staining. Quantification of the apoptotic and necrotic cell percentage. The data are expressed as the mean ± SEM (n = 4). *P < 0.05, vs. scramble or pCMV. # P < 0.05, vs. scramble + BLM or pCMV + BLM. (B) Protein expression of the total and active forms of caspase-3 was measured by immunoblotting. The intensity of the active form of caspase-3 was quantified by densitometry, and the data were normalized to β-actin. Cropped images are displayed, uncropped blots are displayed in Supplementary Fig. 5. The data are expressed as the mean ± SEM. Error bars represent mean ± SEM (n = 4) *P < 0.05, vs. scramble or pCMV. # P < 0.05, vs. scramble + BLM or pCMV + BLM. (C) Tissues stained by the TUNEL method were observed at × 100 magnification, and the number of TUNEL-positive cells in 20 fields per lung was counted (n = 8/group). The data are expressed as the mean ± SEM. * P < 0.05, vs doxy(−)/BLM (+).