Figure 2: NAD⁺ protects against EAE through IL-10.

(a) C57BL/6 mice were subjected to EAE by MOG immunization and treated daily with intraperitoneal injection of 60 mg of NAD⁺ or a placebo solution (PBS). After 18 days, mice were euthanized and CD4⁺ T cells were isolated from spleens, total cell number as well as frequencies of CD4⁺CD25⁺Foxp3⁺, CD4⁺IFNγ⁺, CD4⁺IFNγ⁺IL-10⁺, CD4⁺IL-23R⁺IL-17A⁺, CD4⁺IL-17A⁺GM-CSF⁺, CD4⁺IL-17A⁺ TGFβ⁺ cells were analysed by flow cytometry. (b) mRNA was extracted from the spinal cord of treated NAD⁺ and control group animals and expression of IFN-γ and IL-17A was measured by real-time PCR. (c) Disease scores of EAE in IL-10^−/− (C57BL/6 background) and wild-type (C57BL/6 background) mice treated with NAD⁺ or a placebo solution (PBS). (d) Absolute number of lymphocytes in the blood and in the spleen (n=8–11 per group, representative plots shown). Data derived from two independent experiments; data represent mean±s.d. *P<0.05; **P<0.01; ***P<0.001; NS, not significant; ND, non-detectable. Student’s t-test and Mann–Whitney U-test were used accordingly to compare between groups.