Circulating immune cells in multiple sclerosis

Abstract

Circulating T and B lymphocytes contribute to the pathogenesis of the neuroinflammatory autoimmune disease, multiple sclerosis (MS). Further progress in the development of MS treatments is dependent upon a greater understanding of the immunological disturbances that underlie the disease. Analyses of circulating immune cells by flow cytometry have revealed MS-associated alterations in the composition and function of T and B cell subsets, including temporal changes associated with disease activity. Disturbances in circulating immune populations reflect those observed in the central nervous system and include skewing towards proinflammatory CD4⁺ and CD8⁺ T cells and B cells, greater proportions of follicular T helper cells and functional defects in the corresponding T and B regulatory subsets. Utilizing the analytical power of modern flow cytometers, researchers are now well positioned to monitor immunological changes associated with disease activity or intervention, describe immunological signatures with predictive value and identify targets for therapeutic drug development. This review discusses the contribution of various T and B lymphocyte subsets to MS pathogenesis, provides current and relevant phenotypical descriptions to assist in experimental design and highlights areas of future research.

Keywords: B cells; T cells; flow cytometry; multiple sclerosis.

Figure 1

Intracellular cytokine staining to identify T helper type 1 (Th1), Th17 and Th17.1 cells. In comparison to a healthy control sample (a), the proportion of Th17 and Th17.1 cells can be increased substantially in samples from patients with clinically isolated syndrome (CIS), particularly in the setting of active disease (b). Plots represent CD3⁺CD4⁺ T cells gated on live, single‐cell peripheral blood mononuclear cells (PBMC), following a 4‐h stimulation with phorbol myristate acetate (PMA)/ionomycin in the presence of brefeldin A (BD leucocyte activation cocktail with GolgiPlug).

Figure 2

Peripheral blood follicular T cells. These cells are distinguished from conventional CD4⁺ T cells by the expression of CXCR5 and helper subsets categorized based on chemokine receptor, programmed death 1 (PD)‐1 and inducible T cell co‐stimulator (ICOS) expression. (a) Examining forkhead box protein 3 (FoxP3) and CXCR5 expression on live, single‐cell, CD3⁺CD4⁺ T cells reveals four distinct subsets (clockwise from top left quadrant): CCXR5^–FoxP3⁺ regulatory T cells (T_reg); CXCR5⁺FoxP3⁺ follicular T helper (Tfr); CXCR5⁺FoxP3^– Tfh; and CXCR5^–FoxP3^– conventional T cells. The absence of CXCR5 fluorochrome‐conjugated antibody in T_reg analysis would result in Tfr being analysed simultaneously with conventional T_reg. Simultaneously, FoxP3 fluorochrome‐conjugated antibodies (or other regulatory markers) are required to distinguish between Tfh and follicular regulatory T (Tfr) cells. (b) CXCR5⁺FoxP3^–CD45RA^– Tfh cells can be categorized as Tfh17 (CCR6⁺CXCR3^–), Tfh17.1 (CCR6⁺CXCR3⁺), Tfh1 (CCR6^–CXCR3⁺) and Tfh2 (CCR6^–CXCR3^–). CCR4 and CD161 expression may be used to refine Tfh subsets further (refer to Table 1). (c) While helper capacity of Tfh1 cells is restricted to the activated ICOS⁺PD‐1⁺⁺CCR7^lo subset, all Tfh2 and Tfh17 cells are capable of helping B cells with varying capacity. The intensity of the background greyscale reflects the capacity to provide help to B cells. This figure has been adapted from reference 51 with permission from the authors.

Figure 3

Mucosal‐associated invariant T (MAIT) cells possess a phenotype consistent with central nervous system (CNS) infiltration and proinflammatory cytokine production. (a) CD3⁺CD4^– MAIT cells (predominantly CD8⁺, not shown) express high levels of CD161, and are predominantly effector memory (EM) (CCR7^–CD45RA^–) (b) and CCR6⁺ cells (c). This phenotype is consistent with migration to inflamed tissue, the capacity to cross the blood–brain barrier (BBB) and exert effector function, including interleukin (IL)‐17 production.