Figure 3. Appropriate levels of MnSOD activity are required for quiescent fibroblasts’ entry into the proliferative cycle.

(A) MEFs were cultured continuously for 10 and 30d of quiescent growth with change in media every 3d. Ethanol-fixed cells were stained with PI and cell cycle phase distributions analyzed by flow cytometry. (B) Cells from replicate dishes were re-plated at lower density and cell numbers counted at 2, 4, and 6d post-replating. Asterisk indicates significant difference between cell lines at the indicated time points, n=3, p<0.05. (C) Quiescent MnSOD (+/+) MEFs were mock-infected, infected with 50 MOI of AdBgl II and a dominant negative mutant form of MnSOD (AdmMnSOD) (Zhang et al. 2006). Quiescent control and infected cells were harvested for flow cytometry measurements of DHE-fluorescence. Cells from replicate dishes were trypsinized and re-plated at a lower density. Re-plated cells were continued in culture for 48 h and harvested for flow cytometry measurements of DNA content. The percentage of G₁ calculated using CellQuest software. (**) indicates significant difference in AdmMnSOD infected cells compared to control and AdBgl II infected MnSOD (+/+) MEFs, n=3, p < 0.05.