RNA up-regulation of chemokines in the CNS of Glb1^-/- mice. (A) Real-time PCR was performed on total RNA extracted from the cerebellum of Glb1^+/+ and Glb1^-/- mice at 3 months (▦) and 4 months (▪) of age. SDF-1α and SDF-1β mRNAs were higher in Glb1^-/- mice than in Glb1^+/+ mice. The reactions were standardized to the level of GAPDH mRNA. (B) ELISA of SDF-1α protein in Glb1^-/- cerebellum extracts showed that relatively more SDF-1α was expressed at 3 months (▦) than at 4 months (▪) of age. (C-D) RNase protection assay revealed that the β-chemokines RANTES, MIP-1β, MIP-1α, IP-10, MCP-1, and TCA-3 were up-regulated in hindbrain, midbrain, and forebrain regions (HMF); brain stem (BS); cerebellum (CB); and spinal cord of 3- (C) and 4-month-old (D) Glb1^-/- mice compared with those of Glb1^+/+ controls. The relative levels of RNA induction were normalized against L32 RNA. The amount of chemokines expressed is reported as fold increase over that detected in age-matched Glb1^+/+ mice. (E) CSF from 3-month-old Glb1^-/- mice contained more white blood cells (WBCs) than did the CSF from PPCA^-/- mice or Glb1^+/+ littermates. (F) Transmigration assay demonstrated increased monocyte migration toward Glb1^-/- CSF than toward Glb1^+/+ CSF. Nonspecific cell migration toward serum-free medium was used as a negative control. Data are expressed as mean ± standard deviation (SD).