High-Dimensional Profiling Reveals Heterogeneity of the Th17 Subset and Its Association With Systemic Immunomodulatory Treatment in Non-infectious Uveitis

Abstract

Background: Non-infectious uveitis (NIU) is a severe intra ocular inflammation, which frequently requires prompt systemic immunosuppressive therapy (IMT) to halt the development of vision-threatening complications. IMT is considered when NIU cannot be treated with corticosteroids alone, which is unpredictable in advance. Previous studies have linked blood cell subsets to glucocorticoid sensitivity, which suggests that the composition of blood leukocytes may early identify patients that will require IMT.Objective: To map the blood leukocyte composition of NIU and identify cell subsets that stratify patients that required IMT during follow-up.Methods: We performed controlled flow cytometry experiments measuring a total of 37 protein markers in the blood of 30 IMT free patients with active non-infectious anterior, intermediate, and posterior uveitis, and compared these to 15 age and sex matched healthy controls. Results from manual gating were validated by automatic unsupervised gating using FlowSOM.Results: Patients with uveitis displayed lower relative frequencies of Natural Killer cells and higher relative frequencies of memory T cells, in particular the CCR6+ lineages. These results were confirmed by automatic gating by unsupervised clustering using FlowSOM. We observed considerable heterogeneity in memory T cell subsets and abundance of CXCR3-CCR6+ (Th17) cells between the uveitis subtypes. Importantly, regardless of the uveitis subtype, patients that eventually required IMT in the course of the study follow-up exhibited increased CCR6+ T cell abundance before commencing therapy.Conclusion: High-dimensional immunoprofiling in NIU patients shows that clinically distinct forms of human NIU exhibit shared as well as unique immune cell perturbations in the peripheral blood and link CCR6+ T cell abundance to systemic immunomodulatory treatment.

cDC2 and plasmacytoid dendritic cells diminish from tissues of patients with non‐Hodgkin orbital lymphoma and idiopathic orbital inflammation

“…Similar to our results, a decrease in blood DCs is observed in inflammatory disorders, such as systemic lupus erythematosus and the eye condition uveitis . Also, a reduction in DC numbers in blood has been documented in patients with leukemia , multiple myeloma , classical Hodgkin lymphoma , and other types of cancer .…”

“…Flow cytometry data were visualized with SOM clustering and Minimal Spanning Trees using the FlowSOM , as described previously . Briefly, single cell data (FSC‐A versus FSC‐H gates) were transformed using the logicleTransform function of the flowCore package .…”

Computational Analysis of Multiparametric Flow Cytometric Data to Dissect B Cell Subsets in Vaccine Studies

“…FlowSOM is considered one of the best high‐performance algorithms in automated identification of cell subsets showing an extremely fast runtime . It has also been used for characterizing both the cell phenotype and the cellular functionality, such as the simultaneous production of intracellular cytokine and degranulation . The FlowSOM algorithm is based on a self‐organizing map (SOM), where similar cells are assigned to the same node.…”

Imbalance in PB IL-17-Secreting and Regulatory Cells in Pars Planitis Is Associated with Dysregulation of IFN-γ-Secreting Cells, Especially in Patients with Clinical Complications

“…Similar findings were obtained in patients with uveitis related to systemic autoimmune inflammation or noninfectious uveitis [ 23 , 30 – 32 ]. Although there are conflicting results on the changes in the number of Tregs in active disease [ 30 , 31 , 33 , 34 ], most reports consistently emphasize their role in uveitis resolution and maintenance of the remission phase [ 30 , 33 , 35 , 36 ]. The Th17/Treg imbalance found in our study suggests that all patients lose their immunomodulatory properties and can no longer maintain an equilibrium between proinflammatory and anti-inflammatory cells, including within educational gates (EGs).…”