Figure 6

Repetitive hypoxic preconditioning treatment modulates the peripheral B-cell immune compartment prior to central nervous system injury. (A) Transitional state of CD19⁺CD93⁺ B-cells were quantified and measured as T1 (IgM⁺CD23^-), T2 (IgM⁺CD23⁺) or T3 (IgM^-CD23⁺) cells. Repetitive hypoxic preconditioning (RHP) suppresses the maturation of B-cells from T2 to T3 stage. (B) Ex vivo splenic B-cell activation status was analyzed by quantifying the level of early (IgM⁺IgD^-), mid (IgM⁺IgD⁺) or late (IgM^-IgD⁺) CD19⁺ B-cells. RHP inhibits fully activated B-cell status in the resident B-cells. (C) Conventional B-cell subtypes such as marginal zone (MZ) and follicular B-cells (FOB) were quantified within the CD19⁺ CD93^- populations and not affected by RHP. (A-C) n = 6/group; two independent experiments. (D) In vitro polyclonal B cell responses were assessed using carboxyfluorescein succinimidyl ester (CFSE) dilution assay with lipopolysaccharide (LPS) stimulation. Delta proliferation fraction (dPF) is the percentage of CFSE low cells in the test condition (stimulated) minus the background (non-stimulated condition). Data is representative of two independent experiments with n = 4 per condition. Mean percentages ± SD are shown. 21 %, Untreated cohorts; PMA, phorbol myristate acetate.