THU0234 Epigenetic cell counting: a novel tool to quantify immune cells in salivary glands detects robust correlations of tfh cells with immunopathology

Background Histological analysis of salivary glands for decades has been a valuable tool in the characterization of patients with primary Sjogren9s syndrome (pSS) and non-Sjogren9s sicca (nSS) patients. Importantly, it has helped in understanding the immunopathology of sicca patients. Nonetheless, standardization of histological assessments, e.g. to quantify lymphocytic foci or germinal centers is lacking, contributing to improper classification of disease and assessment of risk of lymphoma for example. Also, detailed and reproducible quantification of the heterogeneity of inflammatory cells and their contribution to immunopathology is lacking. Recent progress in epigenetics has revealed that cell-specific DNA methylation profiles can be applied to reliably quantify numbers of cells in blood and tissues. Objectives To investigate whether epigenetic cell counting can serve as a novel reliable tool to quantify immune cells in salivary glands of sicca patients. Methods DNA was isolated from frozen tissue sections of 13 nSS, 12 probable SS, 29 pSS and 7 overlap SS patients. Bisulfite conversion of demethylated DNA sites was followed by cell specific qPCR that was used to calculate the percentage of cell subsets related to the total number of cells quantified by housekeeping gene expression. Percentages of epigenetically counted cells were correlated to gene expression generated by RNA-Seq analysis of matched salivary gland tissue and histological and clinical parameters (LFS, %IgA+ plasma cells, serum IgG, SSA positivity). Results Strongly increased percentages of epigenetically quantified percentages of CD3, CD4, CD8, B cells, T follicular helper (Tfh) cells and Treg cells in pSS vs nSS patients were observed (all p Conclusions Epigenetic cell counting is a promising novel tool to reproducibly and easily quantify immune cells in the (inflamed) labial salivary gland of sicca patients with relatively low amount of tissue needed ( 3 ). Considering the potential of this technique to include a huge number of (cell-specific) biomarkers we believe this opens up new standardized ways for salivary gland analysis with high relevance for patient classification, understanding of immunopathology and clinical trials. Disclosure of Interest None declared