Figure 3.

The association of ascorbic acid, deferoxamine and N-acetylcysteine reduced apoptosis induced by simulated ischemia/reperfusion in CF. Cells were exposed to 6 h simulated ischemia, followed by 16 h simulated reperfusion (sI/R). Cells were treated with the association of ascorbic acid, deferoxamine, and n-acetylcysteine (A/D/N) using 10 μM of each antioxidant at the onset of simulated reperfusion. (A) The percentage (%) of necrotic cells was quantified by flow cytometry using propidium iodide (right panel), with representative histograms of each experimental group (left panel; n = 4). (B) The percentage (%) of the sub-G1 population was quantified by flow cytometry using propidium iodide (right panel), with representative histograms of each experimental group (left panel; n = 5). (C–E) show representative Western blots (upper panel) and densitometric analysis (lower panel) of pro-caspase 9 (n = 4), pro-caspase-3 (n = 5), and Bcl-xl/Bax ratio (n = 3), respectively. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as a loading control. The results are expressed as mean ± S.E.M. *** p < 0.001 and * p < 0.05 vs. C22 (control cells after 22 h normoxia); ### p < 0.001 and # p < 0.05 vs. sI/R.