Mice were perfused with phosphate-buffered saline (PBS) (pH 7.4) and then with 4% (w/v) paraformaldehyde solution in PBS followed by dissection of cerebellum and whole spinal cord. The tissues were further fixed and then divided into halves: One-half was used for histological staining, whereas the other half was used for myelin staining as described earlier (1618, 41). For histological analysis, routine histology was performed to obtain perivascular cuffing and morphological details of CNS tissues. Paraformaldehyde-fixed tissues were embedded in paraffin, and serial sections (4 μm) were cut. Sections were stained with conventional H&E staining method. Digital images were collected under bright-field setting using a 40× objective. Slides were assessed in a blinded fashion by three examiners for inflammation in different anatomical compartments (meninges and parenchyma). Inflammation was scored using the following scale as described: for meninges and parenchyma: 0, no infiltrating cells; 1, few infiltrating cells; 2, numerous infiltrating cells; and 3, widespread infiltration. For vessels: 0, no cuffed vessel; 1, one or two cuffed vessels per section; 2, three to five cuffed vessels per section; and 3, more than five cuffed vessels per section. For scoring, we used at least six serial sections of each spinal cord from each of five mice per group.

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