Human in vitro-induced IL-17A+ CD8+ T-cells exert pro-inflammatory effects on synovial fibroblasts.

IL-17A+ CD8+ T-cells, termed Tc17 cells, have been identified at sites of inflammation in several immune-mediated inflammatory diseases. However, the biological function of human IL-17A+ CD8+ T-cells is not well characterized, likely due in part to the relative scarcity of these cells. Here, we expanded IL-17A+ CD8+ T-cells from healthy donor PBMC or bulk CD8+ T-cell populations using an in vitro polarization protocol. We show that T-cell activation in the presence of IL-1ß and IL-23 significantly increased the frequencies of IL-17A+ CD8+ T-cells, which was not further enhanced by IL-6, IL-2, or anti-IFNγ mAb addition. In vitro-generated IL-17A+ CD8+ T-cells displayed a distinct type-17 profile compared with IL-17A- CD8+ T-cells, as defined by transcriptional signature (IL17A, IL17F, RORC, RORA, MAF, IL23R, CCR6), high surface expression of CCR6 and CD161, and polyfunctional production of IL-17A, IL-17F, IL-22, IFNγ, TNFα, and GM-CSF. A significant proportion of in vitro-induced IL-17A+ CD8+ T-cells expressed TCRVα7.2 and bound MR1 tetramers indicative of MAIT cells, indicating that our protocol expanded both conventional and unconventional IL-17A+ CD8+ T-cells. Using an IL-17A secretion assay, we sorted the in vitro-generated IL-17A+ CD8+ T-cells for functional analysis. Both conventional and unconventional IL-17A+ CD8+ T-cells were able to induce pro-inflammatory IL-6 and IL-8 production by synovial fibroblasts from patients with psoriatic arthritis, which was reduced upon addition of anti-TNFα and anti-IL-17A neutralizing antibodies. Collectively, these data demonstrate that human in vitro-generated IL-17A+ CD8+ T-cells are biologically functional and that their pro-inflammatory function can be targeted, at least in vitro, using existing immunotherapy.

Polyfunctional, Proinflammatory, Tissue-Resident Memory Phenotype and Function of Synovial Interleukin-17A+CD8+ T Cells in Psoriatic Arthritis.

OBJECTIVE: Genetic associations imply a role for CD8+ T cells and the interleukin-23 (IL-23)/IL-17 axis in psoriatic arthritis (PsA) and other spondyloarthritides (SpA). IL-17A+CD8+ (Tc17) T cells are enriched in the synovial fluid (SF) of patients with PsA, and IL-17A blockade is clinically efficacious in PsA/SpA. This study was undertaken to determine the immunophenotype, molecular profile, and function of synovial Tc17 cells in order to elucidate their role in PsA/SpA pathogenesis. METHODS: Peripheral blood (PB) and SF mononuclear cells were isolated from patients with PsA or other types of SpA. Cells were phenotypically, transcriptionally, and functionally analyzed by flow cytometry (n = 6-18), T cell receptor ß (TCRß) sequencing (n = 3), RNA-Seq (n = 3), quantitative reverse transcriptase-polymerase chain reaction (n = 4), and Luminex or enzyme-linked immunosorbent assay (n = 4-16). RESULTS: IL-17A+CD8+ T cells were predominantly TCRαß+ and their frequencies were increased in the SF versus the PB of patients with established PsA (P < 0.0001) or other SpA (P = 0.0009). TCRß sequencing showed that these cells were polyclonal in PsA (median clonality 0.08), while RNA-Seq and deep immunophenotyping revealed that PsA synovial Tc17 cells had hallmarks of Th17 cells (RORC/IL23R/CCR6/CD161) and Tc1 cells (granzyme A/B). Synovial Tc17 cells showed a strong tissue-resident memory T (Trm) cell signature and secreted a range of proinflammatory cytokines. We identified CXCR6 as a marker for synovial Tc17 cells, and increased levels of CXCR6 ligand CXCL16 in PsA SF (P = 0.0005), which may contribute to their retention in the joint. CONCLUSION: Our results identify synovial Tc17 cells as a polyclonal subset of Trm cells characterized by polyfunctional, proinflammatory mediator production and CXCR6 expression. The molecular signature and functional profiling of these cells may help explain how Tc17 cells can contribute to synovial inflammation and disease persistence in PsA and possibly other types of SpA.

IL-17 in the immunopathogenesis of spondyloarthritis.

Spondyloarthritis (SpA) is a term that refers to a group of inflammatory diseases that includes psoriatic arthritis, axial SpA and nonradiographic axial SpA, reactive arthritis, enteropathic arthritis and undifferentiated SpA. The disease subtypes share clinical and immunological features, including joint inflammation (peripheral and axial skeleton); skin, gut and eye manifestations; and the absence of diagnostic autoantibodies (seronegative). The diseases also share genetic factors. The aetiology of SpA is still the subject of research by many groups worldwide. Evidence from genetic, experimental and clinical studies has accumulated to indicate a clear role for the IL-17 pathway in the pathogenesis of SpA. The IL-17 family consists of IL-17A, IL-17B, IL-17C, IL-17D, IL-17E and IL-17F, of which IL-17A is the best studied. IL-17A is a pro-inflammatory cytokine that also has the capacity to promote angiogenesis and osteoclastogenesis. Of the six family members, IL-17A has the strongest homology with IL-17F. In this Review, we discuss how IL-17A and IL-17F and their cellular sources might contribute to the immunopathology of SpA.

Upregulation of Glucose Uptake and Hexokinase Activity of Primary Human CD4+ T Cells in Response to Infection with HIV-1.

Infection of primary CD4+ T cells with HIV-1 coincides with an increase in glycolysis. We investigated the expression of glucose transporters (GLUT) and glycolytic enzymes in human CD4+ T cells in response to infection with HIV-1. We demonstrate the co-expression of GLUT1, GLUT3, GLUT4, and GLUT6 in human CD4+ T cells after activation, and their concerted overexpression in HIV-1 infected cells. The investigation of glycolytic enzymes demonstrated activation-dependent expression of hexokinases HK1 and HK2 in human CD4+ T cells, and a highly significant increase in cellular hexokinase enzyme activity in response to infection with HIV-1. HIV-1 infected CD4+ T cells showed a marked increase in expression of HK1, as well as the functionally related voltage-dependent anion channel (VDAC) protein, but not HK2. The elevation of GLUT, HK1, and VDAC expression in HIV-1 infected cells mirrored replication kinetics and was dependent on virus replication, as evidenced by the use of reverse transcription inhibitors. Finally, we demonstrated that the upregulation of HK1 in HIV-1 infected CD4+ T cells is independent of the viral accessory proteins Vpu, Vif, Nef, and Vpr. Though these data are consistent with HIV-1 dependency on CD4+ T cell glucose metabolism, a cellular response mechanism to infection cannot be ruled out.

IL-17+ CD8+ T cells: Differentiation, phenotype and role in inflammatory disease.

The pro-inflammatory cytokine interleukin-17A (IL-17) has been the subject of research by many groups worldwide. IL-17 expression is often associated with a specific subset of CD4+ T cells (the so-called Th17 cells); however various other immune cell subsets can also synthesise and express IL-17, including CD8+ T cells. Here we review recent data regarding the presence of IL-17+ CD8+ T cells (also known as Tc17 cells) in human inflammatory disease, discuss current knowledge regarding the culture conditions required for the differentiation of these cells in humans and mice, and describe key phenotypic and functional features. Collectively, this information may shed light on the potential pathogenic role that IL-17+ CD8+ T cells may play in human inflammatory disease.