POS0454 DIGITAL IMAGE ANALYSIS OF INTRAEPITHELIAL B-LYMPHOCYTES AS AN OBJECTIVE ALTERNATIVE TO ASSESS LYMPHOEPITHELIAL LESIONS IN SALIVARY GLANDS OF SJÖGREN’S SYNDROME PATIENTS

Abstract

BackgroundSalivary glands of primary Sjögren’s syndrome (pSS) patients characteristically contain periductal lymphocytic infiltrates. Associated with these infiltrates, lymphoepithelial lesions (LELs) can be formed. LELs are composed of hyperplastic ductal epithelium with infiltrating lymphocytes and may assist in the diagnostic process of pSS1. However, manual identification of LELs on H&E staining is a subjective process and can be challenging. A more objective histological parameter that could assist in identification of LELs is the presence of intraepithelial lymphocytes within striated ducts, since we previously showed an association between intraepithelial B-lymphocytes and hyperplasia of the ductal epithelium2. These results, however, warrant validation in a larger cohort. Furthermore, associations between clinical parameters of pSS patients and presence of intraepithelial lymphocytes are not yet known.ObjectivesTo investigate if detection of intraepithelial lymphocytes using digital image analysis can be used as an objective alternative to assess LELs, and to correlate presence of intraepithelial lymphocytes to clinical parameters of pSS patients.MethodsPaired labial and parotid salivary gland biopsies of 109 patients clinically suspected for pSS were serially sectioned and stained for CD3 (T-lymphocytes), high molecular weight cytokeratin (striated ducts) and CD20 (B-lymphocytes). We developed a virtual triple staining digital image analysis (DIA) algorithm using the Visiopharm Integrator System (Visiopharm, Hørsholm, Denmark) to detect intraepithelial T- and B-lymphocytes within regions of interest comprising striated ducts. A maximum of 10 regions of interests was selected per biopsy. Presence of ductal hyperplasia was assessed on consecutive H&E stained slides. Patients were classified as pSS or non-SS according to the ACR-EULAR criteria.ResultsThe DIA algorithm presented in this study was an accurate and objective method to detect intraepithelial lymphocytes within striated ducts in salivary gland tissue. Presence of intraepithelial B-lymphocytes, in contrast to intraepithelial T-lymphocytes, was a specific finding in salivary gland biopsies of pSS patients. In total, 59% of labial and 68% of parotid gland biopsies of pSS patients contained intraepithelial B-lymphocytes, against only 2% of patients classified as non-SS. Intraepithelial B-lymphocytes were not only detected within striated ducts with hyperplasia (LELs), but also in around 25% of analyzed striated ducts without hyperplasia of pSS patients (precursor stage of LEL). PSS patients with presence of intraepithelial B-lymphocytes in the labial gland showed lower stimulated whole salivary flow (p=0.011) and higher ocular staining scores (p=0.048), compared to patients without intraepithelial B-lymphocytes. Serological parameters, such as, rheumatoid factor-, IgG- and ESR levels were significantly higher in pSS patients with presence of B-lymphocyte containing ducts, irrespective whether the labial or parotid gland was taken into account.ConclusionPresence of B-lymphocyte containing ducts is a specific finding in salivary gland biopsies of pSS patients and is associated with clinical parameters of pSS. Together, identification of B-lymphocyte containing ducts by using DIA could be used as an objective marker in the diagnostic histopathological work-up of patients suspected of pSS.