Journal: The Journal of Neuroscience

Article Title: Regulatory B Cells Normalize CNS Myeloid Cell Content in a Mouse Model of Multiple Sclerosis and Promote Oligodendrogenesis and Remyelination

doi: 10.1523/JNEUROSCI.2840-19.2020

Figure Lengend Snippet: Oligodendroglial lineage after adoptive transfer with B cells or Bregs in EAE mice. Mice were killed starting 2 weeks after adoptive cell transfer, and spinal cords were collected for analysis by flow cytometry and Western blotting. Naive mice and untreated EAE mice were used as control. A, Gating strategy: single cells in side and forward scatter, cells, live cells followed by CD45–/low gate. B, Expression of oligodendroglial markers 2 (and 3.5 weeks for Bregs** treated animals) after adoptive transfer with Bregs or B cells compared with WT and EAE-untreated mice: A2B5, CD140, GalC, and O1 in WT mice and EAE C57BL/6 treated either with B cells or Bregs. C, Statistical analysis of the oligodendroglial marker expression in the spinal cords and brains 2 weeks after adoptive transfer. D, Oligodendroglial phenotyping from spinal cords in WT, Bregs, and B-cell-treated mice: A2B5+CD140+, early OPCs, and O1+GALC+ mature oligodendrocytes. E, Western blotting analysis on the expression of MOG, MBP, and paired-related homeobox protein 1 (PRXX1) by spinal cord oligodendrocytes from EAE C57BL/6 mice untreated, treated with Bregs or B cells, and WT mice 2 weeks after adoptive transfer, load control GADPH. Data are representative of three independent experiments in vivo with n ≥ 6. Data are mean ± SEM. *p < 0.05; **p ≤ 0.01; ***p ≤ 0.001; compared with the corresponding control (Student's t test).

Article Snippet: For oligodendrocyte analysis, the following markers were used as per the manufacturer's recommendation: R&D Systems anti-A2B5 AlexaFluor-647 (clone #105), R&D Systems anti-oligodendrocyte marker O1 AlexaFluor-405-conjugated antibody, Sigma Millipore anti-galactocerebroside FITC antibody (clone mGalC), and Invitrogen anti-CD140 (PDGFRA) FITC (clone APA5).

Techniques: Adoptive Transfer Assay, Flow Cytometry, Western Blot, Expressing, Marker, In Vivo

Journal: The Journal of Neuroscience

Article Title: Regulatory B Cells Normalize CNS Myeloid Cell Content in a Mouse Model of Multiple Sclerosis and Promote Oligodendrogenesis and Remyelination

doi: 10.1523/JNEUROSCI.2840-19.2020

Figure Lengend Snippet: Model of repair process induced by Bregs and effect in lymphoid organs and CNS. EAE manifested with an increase in autoreactive T cells and inflammation in lymphoid organs and the CNS causing demyelination and axon loss occur. In lymphoid organs, autoreactive T cells (Th1 and Th17) are induced after MOG immunization, and they then migrate in the CNS. The inflammatory microenvironment of the CNS induced demyelination and loss of oligodendrocytes. Myeloid-derived cells, both resident microglia and blood-derived monocytes/macrophages, showed a proinflammatory phenotype in B-cell-treated mice compared with WT or Bregs-treated animals (left). Adoptive transfer of Bregs can decrease the frequency of autoreactive T cell in the periphery and the CNS with concomitant increased of IL-10-producing T cells, both Tr-1 and canonical Tregs. Moreover, in mice that received Bregs, myeloid-derived cells, both monocytes/macrophages and microglia, displayed a more immunosuppressive phenotype, and their frequency is lowered respect to EAE mice (right). Concomitantly, a maturation/differentiation of OPCs was evident in the spinal cords of mice that received Bregs concurrent with remyelination.

Article Snippet: For oligodendrocyte analysis, the following markers were used as per the manufacturer's recommendation: R&D Systems anti-A2B5 AlexaFluor-647 (clone #105), R&D Systems anti-oligodendrocyte marker O1 AlexaFluor-405-conjugated antibody, Sigma Millipore anti-galactocerebroside FITC antibody (clone mGalC), and Invitrogen anti-CD140 (PDGFRA) FITC (clone APA5).

Techniques: Derivative Assay, Adoptive Transfer Assay