Background/Purpose: Th-9 cells are distinct T cell subset secreting IL-9 and associated with allergic and parasitic diseases and produced by Th9 cells. Contribution of these cells autoimmune colitis (1, 2), uveitis (3) and experimental autoimmune encephalomyelitis (EAE) has ben reported recently. However, reports on their presence or enrichment at the pathologic sites and impact on the local inflammatory cascade in rheumatoid arthritis (RA) remain unknown. Functional relevance of newly described IL-9 producing CD4+ T cells is not yet been clearly described in RA. This study is an attempt to determine the status of Th9 cells in the disease site in RA patients. Here for the first time, we show that Th9 cells or IL-9 can modulate survival of neutrophils and T cell function and enhances the local inflammatory cascade in RA inflicted joints.

Methods: We recruited treatment naïve active RA patients (n=26) with DAS28-ESR >3.5. Polymorphonuclear (PMNs/Neutrophil) and mononuclear cells (MNCs) were isolated both from the peripheral blood (PBL) and synovial fluid (SF). Neutrophils were cultured in presence of recombinant IL-9 (rIL-9), synovial fluid of RA patients (RA SF), IL-9 neutralizing antibody (aIL-9). MNCs were cultured with plate bound anti-CD3 & anti-CD28 along with rIL-9, RA SF, aIL-9. To easure apoptosis neutrophils were stained for Annexin V and their lysate was immunobloted for expression of anti-apoptotic protein MCL-1. MNCs were stained with CD4 and IL-9 to determine the frequency and function of Th9 cells. Frequencies of other cytokine (IFN-γ, TNF-α, IL-17) producing CD4+ T cells were evaluated by intracellular cytokine staining. 

Results: Significant enrichment of Th9 cell was noted in synovial fluid (SF) of RA patients. Recombinant IL-9 (rIL-9) and RA SF reduced the spontaneous apoptosis of neutrophils along with higher expression of survival protein MCL-1. Blocking endogenous IL-9 of SF decreased their survival and rescued their apoptosis, suggesting the critical role of IL-9 in prolonged neutrophil survival in RA. IL-9 and RA SF also increased the frequencies of TNF-α+, IFN-γ+, IL-17+ CD4+ T cells. Interestingly, both synovial and peripheral Th9 cell frequency showed significant correlation with disease activity DAS-28 ESR score. 

Conclusion: Th9 cells or IL-9 plausibly mediate synovial inflammation by prolonging the neutrophil survival and enhancing inflammatory cytokine producing T cells in the RA joint. So blocking IL-9 by specific antibody could be a possible therapy in RA. Frequency of Th9 cell may serve as objective disease severity biomarker for joint inflammation in RA patients

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ACR Meeting Abstracts - https://acrabstracts.org/abstract/synovial-th9-cells-prolong-neutrophil-survival-and-functionally-enhance-the-infiltrating-t-cells-in-rheumatoid-arthritis/