(A) MBP immunostaining showed that the CNS delivery of IFN-γ protected against EAE-induced demyelination in the lumbar spinal cords of mice on a Perk^+/+ background at PID17. In contrast, there was more severe demyelination in the lumbar spinal cord of IFN-γ^CNS+;Perk^+/– mice at PID17 compared with control mice. Scale bar: 50 μm. (B and C) Toluidine blue staining revealed that the myelin and axons in the spinal cords of IFN-γ^CNS+;Perk^+/+ mice remained almost intact at PID17. In contrast, CNS delivery of IFN-γ did not prevent demyelination and axon damage in the lumbar spinal cord of mice on a Perk^+/–background at PID17. Scale bar: 10 μm. (D and E) CC1 immunostaining showed that oligodendrocytes in the lumbar spinal cords of IFN-γ^CNS+;Perk^+/+ mice remained almost intact at PID17. In contrast, similar to control mice, IFN-γ^CNS+;Perk^+/– mice lost the majority of oligodendrocytes in the demyelinated lesions of their lumbar spinal cords at PID17. Scale bar: 25 μm. (F) Real-time PCR analysis of the mRNA level of MBP in the indicated spinal cords at PID17 relative to that in the spinal cords of age-matched naive mice. Error bars represent SD. n = 3. *P < 0.05 versus IFN-γ^CNS–;PERK^+/+ (C) or naive mice (E and F).

(A) CNS delivery of IFN-γ did not significantly affect the expression of iNOs. (B) CNS delivery of IFN-γ did not significantly affect the expression of TNF-α. (C) CNS delivery of IFN-γ decreased the expression of IL-2 in the spinal cords of mice on a Perk^+/+ background, but did not change IL-2 expression in mice on a Perk^+/– background. (D) CNS delivery of IFN-γ decreased the expression of IL-12 in the spinal cords of mice on a Perk^+/+ background, but did not change IL-12 expression in mice on a Perk^+/– background. (E) CNS delivery of IFN-γ decreased the expression of IL-23 in the spinal cords of mice on a Perk^+/+ background, but did not change IL-23 expression in mice on a Perk^+/– background. (F) CNS delivery of IFN-γ did not significantly affect the expression of IL-5. (G) CNS delivery of IFN-γ did not significantly affect the expression of IL-10. (H) CNS delivery of IFN-γ did not significantly affect the expression of IL-17. Error bars represent SD. n = 3. *P < 0.05 versus IFN-γ^CNS–;Perk^+/+.

(A) Expression pattern of total IFN-γ in the course of EAE (n = 3). (B) Expression pattern of transgenic IFN-γ in the course of EAE (n = 3). (C) Mean clinical score (n = 25). Error bars represent SD. *P < 0.05, **P < 0.01 versus IFN-γ^CNS–;Perk^+/+; Student’s t test.

(A and C) CD3 immunostaining showed that CNS delivery of IFN-γ reduced T cell infiltration in the lumbar spinal cords of mice on a Perk^+/+ background at PID17, but did not significantly affect T cell infiltration in mice on a Perk^+/– background. (B and C) CD11b immunostaining revealed that CNS delivery of IFN-γ did not significantly change the numbers of CD11b-positive microglia/macrophages in the lumbar spinal cord of mice on a Perk^+/+ or Perk^+/– background at PID17 (n = 3). (D) Real-time PCR analysis of the relative mRNA levels of CD3 and CD11b in the spinal cord at PID17 (n = 3). (E, F, and I) CD3 immunostaining showed that CNS delivery of IFN-γ did not affect T cell infiltration in lumbar spinal cord at PID14. (G, H, and I) CD11b immunostaining showed that CNS delivery of IFN-γ did not significantly change the numbers of CD11b-positive microglia/macrophages in the lumbar spinal cord at PID14 (n = 3). (J) Real-time PCR analysis of the relative mRNA levels of CD3 and CD11b in the spinal cord at PID14 (n = 4). (K) Real-time PCR analysis for the expression pattern of cytokines in the spinal cord at PID14 (n = 4). Scale bars: 50 μm. Error bars represent SD. *P < 0.05 versus IFN-γ^CNS–;Perk^+/+.

(A and B) CC1 and p-PERK double labeling showed modest activation of PERK in a few oligodendrocytes in the lumbar spinal cords of control mice at PID14, and CNS delivery of IFN-γ strongly activated PERK in the majority of oligodendrocytes. (C and D) CC1 and p-eIF2α double labeling showed that modest activation of eIF2α in a few oligodendrocytes in the lumbar spinal cord of control mice at PID14, and CNS delivery of IFN-γ markedly activated eIF2α in the majority of oligodendrocytes. Scale bar: 10 μm. (E) Quantitative analysis showed CNS delivery of IFN-γ significantly increased the percentage of double-positive CC1/p-PERK and CC1/p-eIF2α cells in the spinal cords of IFN-γ^CNS+;Perk^+/+ mice compared with IFN-γ^CNS–;Perk^+/+ mice at PID14. Error bars represent SD. n = 3. **P < 0.01 versus IFN-γ^CNS–;Perk^+/+.