Inflammatory cytokine levels increased in the spinal cord of double knockout (DKO) mice with aging. (A) mRNAs from the spinal cord of individual mice were analyzed for the expression levels of cytokine genes with real-time reverse transcription-polymerase chain reaction (RT-PCR) and corrected by the mouse GAPDH gene. Compared with wild-type (WT) mice, expression levels of interleukin-1-alpha (IL-1α), IL-1β, and tumor necrosis factor-alpha (TNFα) in the spinal cord of the DKO mice were upregulated with aging (diamond, WT; circle, DKO). The numbers of mice examined were as follows: 4-, 15-, 28-, and 48-week-old WT (n = 3) and DKO (n = 3). Analysis of data by two-way analysis of variance (ANOVA) was performed as described in Figure 1D. For interaction of mouse groups and time courses, all genes were not statistically significant. In the comparison among time courses in mice, only TNFα in DKO mice turned out to be statistically significant (P = 0.0424). Comparison of WT and DKO with same age is shown in figure (*P <0.05; **P <0.01). (B) Protein levels of IL-1α, IL-1β, and TNFα in the spinal cord of 30- and 60-week-old WT and DKO mice were analyzed with enzyme-linked immunosorbent assay (ELISA). The numbers of mice examined were as follows: 30- and 60-week-old WT (n = 3) and DKO (n = 3); data are presented as mean ± standard deviation (SD) and were analyzed by Student t test (*P <0.05). ND, not detectable.