T helper 17.1 cells associate with multiple sclerosis disease activity: perspectives for early intervention

Abstract

Interleukin-17-expressing CD4+ T helper 17 (Th17) cells are considered as critical regulators of multiple sclerosis disease activity. However, depending on the species and pro-inflammatory milieu, Th17 cells are functionally heterogeneous, consisting of subpopulations that differentially produce interleukin-17, interferon-gamma and granulocyte macrophage colony-stimulating factor. In the current study, we studied distinct effector phenotypes of human Th17 cells and their correlation with disease activity in multiple sclerosis patients. T helper memory populations single- and double-positive for C-C chemokine receptor 6 (CCR6) and CXC chemokine receptor 3 (CXCR3) were functionally assessed in blood and/or cerebrospinal fluid from a total of 59 patients with clinically isolated syndrome, 35 untreated patients and 24 natalizumab-treated patients with relapsing-remitting multiple sclerosis, and nine patients with end-stage multiple sclerosis. Within the clinically isolated syndrome group, 23 patients had a second attack within 1 year and 26 patients did not experience subsequent attacks during a follow-up of >5 years. Low frequencies of T helper 1 (Th1)-like Th17 (CCR6+CXCR3+), and not Th17 (CCR6+CXCR3-) effector memory populations in blood strongly associated with a rapid diagnosis of clinically definite multiple sclerosis. In cerebrospinal fluid of clinically isolated syndrome and relapsing-remitting multiple sclerosis patients, Th1-like Th17 effector memory cells were abundant and showed increased production of interferon-gamma and granulocyte macrophage colony-stimulating factor compared to paired CCR6+ and CCR6-CD8+ T cell populations and their blood equivalents after short-term culturing. Their local enrichment was confirmed ex vivo using cerebrospinal fluid and brain single-cell suspensions. Across all pro-inflammatory T helper cells analysed in relapsing-remitting multiple sclerosis blood, Th1-like Th17 subpopulation T helper 17.1 (Th17.1; CCR6+CXCR3+CCR4-) expressed the highest very late antigen-4 levels and selectively accumulated in natalizumab-treated patients who remained free of clinical relapses. This was not found in patients who experienced relapses during natalizumab treatment. The enhanced potential of Th17.1 cells to infiltrate the central nervous system was supported by their predominance in cerebrospinal fluid of early multiple sclerosis patients and their preferential transmigration across human brain endothelial layers. These findings reveal a dominant contribution of Th1-like Th17 subpopulations, in particular Th17.1 cells, to clinical disease activity and provide a strong rationale for more specific and earlier use of T cell-targeted therapy in multiple sclerosis.

Induction of brain‐infiltrating T‐bet–expressing B cells in multiple sclerosis

“…20 Mononuclear cells were isolated from buffy coats using Ficoll-Paque Plus (GE Healthcare, Freiburg, Germany) and density gradient centrifugation. PBMCs were frozen and stored in liquid nitrogen until use as previously described.…”

“…PBMCs were frozen and stored in liquid nitrogen until use as previously described. 20 Mononuclear cells were isolated from buffy coats using Ficoll-Paque Plus (GE Healthcare, Freiburg, Germany) and density gradient centrifugation. Blood and CSF samples from MS brain donors were acquired postmortem through heart puncture and ventricle drainage, respectively.…”

“…Blood and CSF samples from MS brain donors were acquired postmortem through heart puncture and ventricle drainage, respectively. 20 Heart blood mononuclear cells were isolated as described for buffy coat material. Collection tubes with CSF were centrifuged for 10 minutes at 500 × g. CSF and blood mononuclear cell fractions were resuspended in RPMI 1640 (Lonza, Verviers, Belgium) containing 10% heat-inactivated human AB serum (Sanquin, Rotterdam, the Netherlands) and 1% penicillin/streptomycin (Lonza) and left to rest at 37 C until further use.…”

“…Sorted memory B cells from healthy donor blood were assessed for selective in vitro transmigration toward CXCL10. high IL-17 low ) cells in MS patients (see Fig 4 and data not shown) 20. Percentages of subsets within the total memory pool were compared before and after migration, both to medium and to CXCL10 (−BEC, n = 8; +BEC, n = 6).…”

The role of chemokines and chemokine receptors in multiple sclerosis

“…After the secreted substances enter the central nervous system, they activate astrocytes and microglia, produce a large number of chemokines and cytokines, and recruit peripheral immune cells to sites of inflammation [123]. The use of CXCR3 and CCR6 as markers of identification of Th17 cells not only reflects their pro-inflammatory status but also reflects their ability to migrate to localized sites of inflammation [118]. The initiation of T cell entry into the CNS is governed by integrin-dependent vascular adhesion and chemokine-driven trans-blood-brain barrier [1].…”