Background/Purpose: Sjögren’s Disease (SjD) is a systemic rheumatic disorder characterized by lymphocytic infiltration of exocrine glands and immune dysregulation. Heterogeneity exists among patients in terms of disease activity and presence of anti-Ro/SS-A antibodies, which are absent in up to 40% of patients. These observations highlight the need to define subsets of SjD patients with unique characteristics. We aimed to characterize peripheral T cell immunophenotypes in SjD patients meeting the 2016 ACR/EULAR criteria using multiparametric flow cytometry and to explore their relationship to Ro/SS-A serologic status and clinical measures of disease activity.

Methods: Cryopreserved peripheral blood mononuclear cells from SjD patients and healthy controls (HC) were analyzed using an adaptive immunity-focused 37-plex multiparametric flow cytometry panel. A total of 93 Ro⁺ SjD, 63 Ro^– SjD, and 39 healthy controls (HC) were included. Differences between patient groups were first assessed by Cluster Identification, Trajectory, Regression, and Unsupervised Stratification (CITRUS, Cytobank) followed by manual gating using FlowJo. Cellular proportions were analyzed as % of the CD3⁺ population. Inter-group comparisons were evaluated by Kruskal-Wallis tests with Dunn’s post-test corrections. Intra-group comparisons were evaluated using Mann-Whitney U tests. Correlations were analyzed using a Spearman correlation matrix. Statistical analyses were completed using GraphPad Prism.

Results: SjD patients had increased frequencies of a terminally differentiated cytotoxic CD8 T cell population (SLAMF7⁺CD57⁺CD28^–) (Ro⁺ vs. HC, p=0.054; Ro^– vs. HC, p=0.044) and reduced frequencies of CD161⁺CCR5⁺ effector memory (EM) CD4 and CD8 T cells compared with HC (Ro⁺ vs. HC, p< 0.0001; Ro^– vs. HC, p< 0.0001). CD38⁺ EM CD4⁺ T cells were preferentially expanded in Ro⁺ participants compared to Ro^– (p=0.020). Correlation analysis identified associations between multiple T cell subsets and clinical measures of disease, highlighting their potential relevance as indicators of immune activity in SjD.

Conclusion: These findings support both shared and subgroup-specific T cell alterations in SjD. SLAMF7⁺CD57⁺CD28^– cytotoxic CD8 T cells, previously described in IgG4 related disease¹, were increased in both Ro+ and Ro^– SjD patients, suggesting a common cytotoxic signature.  CD161⁺CCR5⁺ EM T cells  were depleted in SjD patients regardless of serostatus, potentially reflecting migration to exocrine glands. CD38⁺ EM CD4 T cells were selectively expanded in Ro⁺ SjD and may reflect enhanced immune activation within this subgroup. These observations identify immune phenotypes that contribute to SjD heterogeneity.

1 Perugino CA, et al. J Allergy Cin Immunol. 2021;147(1):368-382.

Funding: Supported by NIH/NIAMS UC2 AR081032-05S1, UC2 AR081032, and P30 AR073750

Figure 1. Heatmap of T cell subset median frequencies of HC, Ro+ SjD, and Ro- SjD Groups. Frequencies were z-score transformed within each population, with higher/lower values indicating relative expansion or reduction.

Figure 2. Association of T cell subset frequencies with clinical and laboratory features of disease in Ro+ SjD and Ro- SjD Groups. Clustering of Spearman r-values in Ro+ SjD (upper panel) and Ro- SjD (lower panel) participants. Clinical variable abbreviations: RF=rheumatoid factor; FS=minor salivary gland biopsy focus score; ESR=erythrocyte sedimentation rate; ESSDAI=EULAR Sjogrens Syndrome Disease Activity Index; Criteria Points=2016 ACR/EULAR classification criteria weighted score; WUSF=whole unstimulated salivary flow rate; vBS=maximum vanBijsterveld corneal damage score.

ACR Meeting Abstracts - https://acrabstracts.org/abstract/peripheral-t-cell-immunophenotyping-identifies-terminally-differentiated-cytotoxic-cd8-t-cell-expansion-effector-memory-t-cell-depletion-and-anti-ro-associated-cd38%e2%81%ba-cd4-t-cell-activation-in/