Caloric Restriction and ALS Figure 13. Apoptosis regulatory proteins content in the quadriceps of CR vs. AL G93A mice. (A) Bax was significantly higher in CR vs. AL mice (41%, P = 0.027; main effect of diet). Bax was increased in CR vs. AL females (52%, P = 0.048). (B) There was no significant difference in Bcl-2 protein content. (C) Bax/Bcl-2 was significantly higher in CR vs. AL mice (68%, P = 0.040; main effect of diet). Bax/Bcl-2 was increased in CR vs. AL females (2.3-fold, P = 0.029). No significant changes were observed in the protein content of: (D) caspase 9, (E) cleaved caspase 9, and (F) the ratio of cleaved caspase 9/caspase 9 for diet or sex. Data are presented as means 6 SEM. n = 27; AL, 7 males and 7 females; CR, 7 males and 6 females. Asterisks denote significant changes (P#0.05 vs. AL). doi:10.1371/journal.pone.0009386.g013 expression (MnSOD, 2.7-fold; and GPx, 2.6-fold) [47] compared with males, and that this sexual dimorphism is conserved under CR when analyzing enzyme activity (MnSOD, 1.4-fold; and GPx, 1.3-fold) [49]. However, in spite of the compensatory increase in the protein content of these enzymes, the CR mice in our study reached clinical onset faster than AL mice. This may indicate that the protective antioxidant response was incomplete (i.e. partial), and hence insufficient to mitigate the CR-induced increase in lipid peroxidative damage. Alternatively, perhaps the increase in these enzymes is a potential reason why PC levels were decreased under CR [49,51]. Moreover, since basal enzyme activity of MnSOD (4–5-fold), Cu/Zn-SOD (7–10-fold) and catalase (2-fold) are substantially elevated in response to increased basal levels of oxidative stress in G93A mice, any further upregulation in these enzymes may be minimal, and hence may not confer a significant benefit [32]. The antioxidant enzyme activity of MnSOD in rat liver mitochondria increased by 1.4-fold in females vs. males under CR (60% of AL) [49], while cardiac Cu/Zn-SOD activity increased by 1.3-fold in rats under CR (60% of AL) [52]. In male type 2 diabetic fatty rats, CR (30% of AL) increased total SOD activity by 2.3-fold vs. AL rats [53]. This increase is somewhat similar to what we observed in PLoS ONE | www.plosone.org 13 February 2010 | Volume 5 | Issue 2 | e9386