Labial Salivary Glands Research Articles

Type I Interferon Dependent hsa-miR-145-5p Downregulation Modulates MUC1 and TLR4 Overexpression in Salivary Glands From Sjögren's Syndrome Patients.

Sjögren’s syndrome (SS) is an autoimmune disease that mainly affects salivary glands (SG) and is characterized by overactivation of the type I interferon (IFN) pathway. Type I IFNs can decrease the levels of hsa-miR-145-5p, a miRNA with anti-inflammatory roles that is downregulated in SG from SS-patients. Two relevant targets of hsa-miR-145-5p, mucin 1 (MUC1) and toll-like receptor 4 (TLR4) are overexpressed in SS-patients and contribute to SG inflammation and dysfunction. This study aimed to evaluate if hsa-miR-145-5p modulates MUC1 and TLR4 overexpression in SG from SS-patients in a type I IFN dependent manner. Labial SG (LSG) biopsies from 9 SS-patients and 6 controls were analyzed. We determined hsa-miR-145-5p levels by TaqMan assays and the mRNA levels of MUC1, TLR4, IFN-α, IFN-β, and IFN-stimulated genes (MX1, IFIT1, IFI44, and IFI44L) by real time-PCR. We also performed in vitro assays using type I IFNs and chemically synthesized hsa-miR-145-5p mimics and inhibitors. We validated the decreased hsa-miR-145-5p levels in LSG from SS-patients, which inversely correlated with the type I IFN score, mRNA levels of IFN-β, MUC1, TLR4, and clinical parameters of SS-patients (Ro/La autoantibodies and focus score). IFN-α or IFN-β stimulation downregulated hsa-miR-145-5p and increased MUC1 and TLR4 mRNA levels. Hsa-miR-145-5p overexpression decreased MUC1 and TLR4 mRNA levels, while transfection with a hsa-miR-145-5p inhibitor increased mRNA levels. Our findings show that type I IFNs decrease hsa-miR-145-5p expression leading to upregulation of MUC1 and TLR4. Together, this suggests that type I interferon-dependent hsa-miR-145-5p downregulation contributes to the perpetuation of inflammation in LSG from SS-patients.

Gene expression alterations in salivary gland epithelia of Sj\xf6gren\u2019s syndrome patients are associated with clinical and histopathological manifestations

Sjögren’s syndrome (SS) is a complex autoimmune disease associated with lymphocytic infiltration and secretory dysfunction of salivary and lacrimal glands. Although the etiology of SS remains unclear, evidence suggests that epithelial damage of the glands elicits immune and fibrotic responses in SS. To define molecular changes underlying epithelial tissue damage in SS, we laser capture microdissected (LCM) labial salivary gland epithelia from 8 SS and 8 non-SS controls for analysis by RNA sequencing (RNAseq). Computational interrogation of gene expression signatures revealed that, in addition to a division of SS and non-SS samples, there was a potential intermediate state overlapping clustering of SS and non-SS samples. Differential expression analysis uncovered signaling events likely associated with distinct SS pathogenesis. Notable signals included the enrichment of IFN-γ and JAK/STAT-regulated genes, and the induction of genes encoding secreted factors, such as LTF, BMP3, and MMP7, implicated in immune responses, matrix remodeling and tissue destruction. Identification of gene expression signatures of salivary epithelia associated with mixed clinical and histopathological characteristics suggests that SS pathology may be defined by distinct molecular subtypes. We conclude that gene expression changes arising in the damaged salivary epithelia may offer novel insights into the signals contributing to SS development and progression.

A biomarker for lymphoma development in Sjogren's syndrome: Salivary gland focus score

The aim of this study is to explore the role of labial minor salivary gland (LMSG) focus score (FS) in stratifying Sjögren's Syndrome (SS) patients, lymphoma development prediction and to facilitate early lymphoma diagnosis. Ιn an integrated cohort of 1997 patients, 618 patients with FS ≥ 1 and at least one-year elapsing time interval from SS diagnosis to lymphoma diagnosis or last follow up were identified. Clinical, laboratory and serological features were recorded. A data driven logistic regression model was applied to identify independent lymphoma associated risk factors. Furthermore, a FS threshold maximizing the difference of time interval from SS until lymphoma diagnosis between high and low FS lymphoma subgroups was investigated, to develop a follow up strategy for early lymphoma diagnosis. Of the 618 patients, 560 were non-lymphoma SS patients while the other 58 had SS and lymphoma. FS, cryoglobulinemia and salivary gland enlargement (SGE) were proven to be independent lymphoma associated risk factors. Lymphoma patients with FS ≥ 4 had a statistically significant shorter time interval from SS to lymphoma diagnosis, compared to those with FS < 4 (4 vs 9 years, respectively, p = 0,008). SS patients with FS ≥ 4 had more frequently B cell originated manifestations and lymphoma, while in patients with FS < 4, autoimmune thyroiditis was more prevalent. In the latter group SGE was the only lymphoma independent risk factor. A second LMSG biopsy is patients with a FS ≥ 4, 4 years after SS diagnosis and in those with FS < 4 and a history of SGE, at 9-years, may contribute to an early lymphoma diagnosis. Based on our results we conclude that LMSG FS, evaluated at the time of SS diagnosis, is an independent lymphoma associated risk factor and may serve as a predictive biomarker for the early diagnosis of SS-associated lymphomas.

POS0778 ULTRA HIGH-RESOLUTION ULTRASOUND (UHFUS) OF LABIAL GLANDS IS A STRONG PREDICTOR OF SALIVARY GLAND HISTOPATHOLOGY IN SJÖGREN’S SYNDROME (PSS)

Background:Last-generation ultra high-resolution ultrasound (UHFUS) transducers, producing frequencies up to 70 MHz and achieving tissue resolution up to 30 μm, are opening up new possibilities for the study of labial salivary glands (LSG) in patients clinically suspected with primary Sjögren’s syndrome (pSS).Objectives:To explore the value of LSG-UHFUS as a predictor of the intensity of the histological inflammation in LSG biopsy in an inception cohort of patients with sicca symptoms derived from daily clinical practice.Methods:Consecutive patients undergoing a LSG for clinically suspected pSS were included in this study from January 2018 to October 2020. UHFUS of LSG was performed by using VEVO MD, equipped with a 70 MHz probe, scanning first the central compartment of the inferior lip, and then both peripheral compartments. Parenchymal homogeneity was graded on a scale of 0 (normal) to 3 (evident). UHFUS imaging was used to locate the LSG for the US-guided biopsy. For each of the LSG specimens the area of the glandular tissue, the presence and number of foci and the focus score (FS) were assessed. Immunostaining for CD21 was performed to evaluate the presence of follicular dendritic cells (FDC) in foci.Results:We included a total of 171 patients with suspected pSS: out of them, 83 (48.5%) received a diagnosis of pSS (ACR 2016 criteria) and 88 (51.5 %) were diagnosed as no-SS sicca controls. Out of the 171 LSG biopsies, 73/171 (42.7%) were characterized by a nonspecific chronic sialadenitis (NSCS) whereas a focal lymphocytic sialadenitis (FLS) was described in the remaining 98/171 (57.3%). UHFUS-LSG scores were differently distributed in pSS patients with respect to no-SS sicca controls: a score 0 was detected in 16/88 no-SS sicca controls and in none of the 83 pSS patients, whereas a score 3 was detected in 12/83 pSS and in none of no-SS sicca controls. (p&lt;0.001). The higher was the UHFUS grading, the higher was the number of foci (from a mean (S.D) = 0.38 (0.72) in UHFUS-score 0 to 4.58 (3.11) in UHFUS-score 3, p&lt;0.001) and the focus score (from 0.17 (0.32) in UHFUS-score 0 to 1.88 (0.90) in UHFUS-score 3, p&lt;0.001) of the samples. Overall, out of the 16 no-SS controls with LSG-UHFUS score 0, only 4/16 showed histological features of focal lymphocytic infiltrate; however, the mean FS of those 4/16 samples was 0.17 (0.32). In fact, considering a FS≥1, negative UHFUS-LSG was highly predictive of a negative histology (100 %). By contrast, out of the 12 pSS patients with the highest score at LSG-UHFUS (score 3), 10/12 presented FDC networks in their biopsies, with a positive predictive value of 83% of the severity of the infiltrate.Conclusion:UHFUS of LSG appeared feasible and sensitive in pSS; due to its striking negative predictive value this novel tool can help to identify negative patients on subsequent lip biopsy, thus avoiding invasive procedures in selected cases. Besides its diagnostic role, considering the association with the presence of FDC networks, UHFUS-LGS may also have a role in patients prognostic stratification.Disclosure of Interests:None declared

A novel diagnostic technique of measuring labial minor salivary gland secretions using sodium fluorescein dye: Implications for patients with dry eyes

ABSTRACT Purpose: To evaluate the lower labial minor salivary glands (MSGs) flow rate using fluorescein dye in healthy individuals and patients with Sjögren’s syndrome (SS) and Stevens–Johnson syndrome (SJS). Methods: Thirty consecutive patients with SS (N = 15; mean age 35.7 years) and SJS (N = 15; 57.7 years), and their age- and sex-matched healthy controls (N = 40; mean ocular surface disease index (OSDI) = 6.1) had ocular examination and whole lower labial mucosa evaluation after ophthalmic fluorescein strip application under cobalt blue light. Analyzed parameters include average labial distribution of functional MSGs, their secretory flow rate, and comparison between healthy and diseased groups. Results: The mean salivary flow rate from the lower labial MSGs of normal individuals showed decade-wise decline (2.7 ± 0.29 l/min, 2.6 ± 0.39 l/min, 2.3 ± 0.6 l/min, 1.9 ± 0.95 l/min) with similar trend for the number of secreting MSG openings (38.7, 37.2, 30, 22). The mean OSDI score was 45.3 in SJS and 28.2 in SS group. Thirty eyes in each SS and SJS group had mean Schirmer I value of 1.5 ± 2.2 mm and 5.1 ± 4.4 mm. The mean lower labial salivary flow rate in SS group was 0.5 ± 0.28 l/min, and 2.0 ± 0.95 l/min in SJS group. When compared to normal, SS patients had a significant reduction in the flow rate (p < .00001); however, it was not significant for the SJS group (p = .28). The mean number of secreting MSG openings was reduced in both the groups compared to normal (SJS, 20.5; p = .01 and SS, 12; p < .0001). There was a significant difference between SS and SJS groups in terms of flow rate (p < .00001) and number of MSG openings (p = .001). The MSG flow rate and Schirmer values did not show any correlation in SS or SJS patients. Conclusion: The fluorescein-assisted evaluation of the lower lip can be used as a quantitative method for measuring the labial salivary flow rate in SS and SJS patients.

Inhibitory effect of HTLV-1 infection on the production of B-cell activating factors in established follicular dendritic cell-like cells.

IntroductionThe low frequency of ectopic germinal center in labial salivary glands of patients with human T‐cell leukemia virus type 1 (HTLV‐1) antibody‐positive Sjögren's syndrome (SS) suggests that HTLV‐1 has some effects on follicular dendritic cells (FDCs).MethodsWe used flow cytometry, immunofluorescence, and enzyme‐linked immunosorbent assays (ELISAs) to investigate the direct effect of HTLV‐1 on B‐cell activating factors produced by established FDC like cells obtained from excised human tonsils. We then measured the serum B‐cell activating factor (BAFF) and C‐X‐C motif ligand (CXCL) 13 concentrations of the HTLV‐1‐seropositive SS patients and the HTLV‐1‐seronegative SS patients by ELISA.ResultsAmong the 31 isolated specimens, 22 showed morphological characteristics of FDCs. Day 2‐cultured specimens showed expressions of CD14, CD23, CD40, intracellular adhesion molecule‐1 (ICAM‐1), and vascular cell adhesion molecule‐1. After 2 weeks, 12 of these specimens expressed ICAM‐1, FDC, and fibroblast cell marker. Intracellular BAFF and CXCL13 were constitutively expressed regardless of stimulation. After direct coculture with the HTLV‐1‐infected T‐cell line HCT‐5 or MT‐2, the BAFF and CXCL13 expressions on the FDC‐like cells were decreased in accord with the increased number of HCT‐5 and MT‐2 cells with styliform change and without HTLV‐1 Gag protein expression. Interferons upregulated the concentration of BAFF (but not CXCL13) in the culture supernatant, which showed a declining trend under the presence of HCT‐5 or MT‐2. The serum concentrations of BAFF and CXCL13 in the HTLV‐1‐seropositive SS patients were lower than those of the HTLV‐1 seronegative SS patients.ConclusionsHTLV‐1 partially inhibited the BAFF and CXCL13 expressions of established FDC‐like cells.

Hypomethylation mediates genetic association with the major histocompatibility complex genes in Sjögren's syndrome.

Differential methylation of immune genes has been a consistent theme observed in Sjögren's syndrome (SS) in CD4+ T cells, CD19+ B cells, whole blood, and labial salivary glands (LSGs). Multiple studies have found associations supporting genetic control of DNA methylation in SS, which in the absence of reverse causation, has positive implications for the potential of epigenetic therapy. However, a formal study of the causal relationship between genetic variation, DNA methylation, and disease status is lacking. We performed a causal mediation analysis of DNA methylation as a mediator of nearby genetic association with SS using LSGs and genotype data collected from 131 female members of the Sjögren's International Collaborative Clinical Alliance registry, comprising of 64 SS cases and 67 non-cases. Bumphunter was used to first identify differentially-methylated regions (DMRs), then the causal inference test (CIT) was applied to identify DMRs mediating the association of nearby methylation quantitative trait loci (MeQTL) with SS. Bumphunter discovered 215 DMRs, with the majority located in the major histocompatibility complex (MHC) on chromosome 6p21.3. Consistent with previous findings, regions hypomethylated in SS cases were enriched for gene sets associated with immune processes. Using the CIT, we observed a total of 19 DMR-MeQTL pairs that exhibited strong evidence for a causal mediation relationship. Close to half of these DMRs reside in the MHC and their corresponding meQTLs are in the region spanning the HLA-DQA1, HLA-DQB1, and HLA-DQA2 loci. The risk of SS conferred by these corresponding MeQTLs in the MHC was further substantiated by previous genome-wide association study results, with modest evidence for independent effects. By validating the presence of causal mediation, our findings suggest both genetic and epigenetic factors contribute to disease susceptibility, and inform the development of targeted epigenetic modification as a therapeutic approach for SS.

Is it possible to not perform salivary gland biopsy in targeted patients according to unstimulated salivary flow results in patients with suspected Sj\xf6gren's syndrome?

Introduction/objectiveXerostomia is one of the main symptoms of primary Sjögren's syndrome (pSS). The unstimulated salivary flow (UWS) test is one of the objective Sjögren's syndrome classification criteria used to assess xerostomia's severity. The study’s objective was to evaluate UWS rate measurements (with a threshold rate of 0.1 mL/min) in the screening of patients suspected with pSS, presenting with xerostomia in whom labial salivary gland biopsy (LSGB) should be performed. We will try to answer whether it is possible not to perform LSGB in targeted patients according to UWS results? We analyze the correlation between UWS value and focus score (FS) and anti-SSA antibodies.MethodsThe study group consisted of subjects above 18 years of age with a subjective feeling of oral dryness.ResultsA total of 105 subjects were qualified for the study. The final diagnosis of pSS was made in 44 patients according to the classification criteria from 2016. No age differences were identified between pSS patients and control group subjects (patients with dry mouth without autoimmune background). UWS rates were significantly lower in pSS patients than in the control group. No association was identified between UWS and focus score (FS) ≥ 1 in LSGB. No differences were observed between anti-SSA-positive and anti-SSA-negative patients in terms of age, UWS rates, FS.ConclusionLSGB should be performed in all suspected pSS cases regardless of the UWS rate value, particularly in subjects without specific anti-SSA antibodies. In patients with suspected pSS, only less than one-half of the UWS measurements are below the value of 0.1 mL/min adopted as the threshold in the classification criteria for pSS.

Labial salivary gland assessment in idiopathic pulmonary fibrosis patients with sicca symptoms

Patients with idiopathic pulmonary fibrosis (IPF) commonly present with sicca symptoms. This study aimed to assess labial minor salivary glands (LMSGs) in those patients to rule out Sjögren's syndrome (SS), in which sicca symptoms are the clinical hallmark. Cases of patients with IPF with sicca symptoms referred to the oral medicine clinic at the University of Florida within the last 13 years were selected with institutional review board approval. Demographic characteristics, clinical findings, laboratory results, and histomorphologic parameters were retrospectively analyzed. A total of 12 patients (9 men and 3 women, ages 55-76 years) were identified. History of exposure to asbestos or chemicals, smoking, and medication information was obtained. All patients reported sicca symptoms with 57% of those exhibiting objective or borderline dryness. Anti-SSA/Ro and anti-SSB/La were positive in 25% and 8% of the cases, respectively. Microscopically, 1 out of 12 patients was biopsy positive in the absence of anti-SSA/Ro, fulfilling the 2016 SS criteria with positive sialometry. A LMSG biopsy is critical to identify SS in patients with diagnosed IPF and present sicca symptoms, especially those with negative serology, as revealed in our study.

Regulation of STAT4 by ETS1 in Sjögren syndrome pathogenesis

<h3>Objectives</h3> Primary Sjögren syndrome (pSS) is a chronic autoimmune disease mainly affecting women over the age of 40, causing dry mouth and eyes. The etiology of pSS is complex and includes the overexpression of inflammatory cytokines from affected salivary gland epithelial cells (SGECs). Our group has previously identified candidate biomarkers overexpressed in salivary gland tissues of patients with pSS. We showed that ETS proto-oncogene 1 (ETS1) had higher mRNA and protein expression in pSS salivary glands that correlated with higher levels of lymphocytic infiltration. It has also been shown that the signal transducer and activator of transcription 4 (STAT4) gene contains pSS susceptibility polymorphisms and is a significant mediator of cytokine (ie, interferon, tumor necrosis factor-α) production. Additionally, in previous studies unrelated to pSS, ETS1 was shown to regulate STAT4 expression at the transcription level. Thus, we hypothesized that ETS1 plays a significant role in regulating STAT4 transcription within SGECs of patients with pSS. Our objectives were as follows: (1) to determine whether downregulation of ETS1 mRNA by siRNA knockdown results in decreased mRNA and protein expression of STAT4 within the salivary gland cell line A253 and (2) to demonstrate the regulation of STAT4 mRNA expression by ETS1 siRNA knockdown in cultured SGECs from control patients with pSS and non-pSS sicca. <h3>Methods</h3> The salivary gland cell line, A253, was transfected with siRNA targeting ETS1 or a scrambled siRNA control. After 48 hours, mRNA and protein were isolated. cDNA synthesis was performed using 1 µg of mRNA. Expression of ETS1 and STAT4 mRNA was quantified by quantitative reverse transcription–polymerase chain reaction using the delta cycle threshold method to determine the relative fold changes normalized to glyceraldehyde 3-phosphate dehydrogenase in triplicate experiments. In addition, downregulation of STAT4 protein by ETS1 siRNA was determined by Western blot analysis and semiquantified by densitometry using cofilin as the loading control. SGECs were cultured from labial salivary gland biopsies of 2 patients with pSS (focus score [FS] ≥1) and 4 patients with non-pSS sicca (FS = 0, n = 2; 0 < FS < 1; n = 2) and transfected with siRNA targeting ETS1 or a scrambled siRNA control. After 48 hours, mRNA was isolated for cDNA synthesis and used to determine the mRNA expression of ETS1 and STAT4 as described above. <h3>Results</h3> In A253 cells, STAT4 mRNA expression was changed by −2.15-fold, on average, when treated with siRNA targeting ETS1. STAT4 protein expression was measured by Western blot analysis 48 hours post-ETS1 siRNA knockdown showing an average −1.25-fold change in A253 total cell lysate. SGECs cultured from patients with lymphocytic infiltration (ie, FS >0) showed a significantly higher expression of STAT4 (<i>P</i> = .0485) than patients with non-pSS sicca with FS = 0. However, we did not find a significant change in STAT4 mRNA expression (average, +1.14-fold) in cultured SGECs treated with siRNA targeting ETS1. <h3>Conclusions</h3> STAT4 is overexpressed by cultured SGECs in patients with pSS and those without pSS with focal lymphocytic infiltrates. STAT4 overexpression in SGECs of patients with pSS and those without pSS could be a result of inflammatory cytokine exposure by neighboring cells or effects of infiltrating lymphocyte exposure. Thus, our results suggest a separate regulatory mechanism of STAT4 in pSS bypassing ETS1 regulation, which might be due to pathogenic pathways involving other transcriptional regulation.

Neuropathic Pain in the Eyes, Body, and Mouth: Insights from the Sjögren's International Collaborative Clinical Alliance.

To evaluate how ocular, oral, and bodily neuropathic pain symptoms, which characterize small fiber neuropathies, are associated with Sjögren's syndrome (SS) classification based on the American College of Rheumatology/European League Against Rheumatism (ACR/EULAR) criteria. Participants enrolled in the Sjögren's International Collaborative Clinical Alliance (SICCA) registry had ocular, rheumatologic, oral, and labial salivary gland (LSG) biopsy examinations, blood and saliva samples collected, and completed questionnaires at baseline. We used mixed effects modeling with age, country, gender, and depression being fixed effects and study site, a random effect, to determine if neuropathic pain indicators (assessed via questionnaires) were associated with being classified as SS. A total of 3,514 participants were enrolled into SICCA, with 1,541 (52.9%) meeting the 2016 ACR/EULAR classification criteria for SS. There was a negative association between being classified as SS and experiencing bodily neuropathic pain features of needle-like pain, prickling/tingling sensation, ocular neuropathic pain of constant burning, and constant light sensitivity, and having a presumptive diagnosis of neuropathic oral pain. We found that those classified as SS had lower scores/reports of painful neuropathies compared with those classified as non-SS. Non-SS patients with dry eye disease or symptoms could benefit from pain assessment as they may experience painful small-fiber neuropathies (SFNs). Pain questionnaires may help identify pain associated with SFNs in patients with SS and non-SS dry eye. Future studies would be helpful to correlate self-reports of pain to objective measures of SFNs in those with SS, non-SS dry eye, and healthy controls.

Ultra-high frequency ultrasonography (UHFUS)-guided minor salivary gland biopsy: A promising procedure to optimize labial salivary gland biopsy in Sjögren's syndrome.

Sjögren's syndrome (SS) is an autoimmune disease characterized by an inflammatory infiltrate of exocrine salivary and lachrymal glands. Diagnosis is complex, and minor salivary gland biopsy and subsequent focus score (FS) calculation appear of extreme importance in the diagnostic work-up of the disease. Ultra-high frequency ultrasonography (UHFUS) is a recently introduced diagnostic technique, which is gaining an increasingly important role in intraoral imaging. This study aims at exploring the usefulness of UHFUS for obtaining valuable labial salivary gland samples to assess the histopathological features of SS patients. Patients with clinical suspect of SS and eligible for minor salivary gland biopsy were enrolled. UHFUS scan of the lower lip was performed. Glandular echostructure was classified according to Outcome Measures in Rheumatology (OMERACT) scoring system. The glands to be sampled were selected on the basis of UHFUS evaluation and biopsied. The areas of the samples were recorded and compared with those obtained without UHFUS guidance. The correlation between UHFUS grade and labial gland FS was also assessed. The areas of the samples obtained with UHFUS guidance were significantly higher (7.25±3.98mm2 ) than those obtained by conventional procedures (5.79±3.49mm2 , P=.02). UHFUS correlated significantly with the salivary gland FS (r=.532, P=.001). UHFUS seems a promising tool in SS diagnostic algorithm, being able to provide a valuable support to the biopsy procedure. Further studies are mandatory to confirm the role of UHFUS in SS.

Accuracy of minor salivary gland biopsy in the diagnosis of Sjögren syndrome.

The aim of this study was to find out the correlation and evaluate the accuracy of labial minor salivary gland biopsy as a diagnostic tool in the multidisciplinary management of patients with Sjögren syndrome. Patients, referred to our outpatient office between January 2014 and December 2018 from a rheumatologist for biopsy examination, as part of the complex diagnostic plan for suspected Sjögren syndrome, were included in the current study. Each specimen was examined histomorphometrically by the pathologist to calculate the focus score describing the degree of salivary gland inflammatory infiltration. Fifty patients met the inclusion criteria. From the total number of patients, 39 presented with an established Sjögren syndrome by fulfilling the revised American-European criteria. From those, 27 had a positive lip biopsy. The remaining 12 patients from the total group, who were diagnosed with Sjögren syndrome based on the same criteria, had a negative lip biopsy. The labial minor salivary gland biopsy is a valuable diagnostic tool to establish the diagnosis of Sjögren syndrome. However, a positive biopsy result must always be correlated with all the other diagnostic criteria to prove the exact diagnosis (Tab. 1, Fig. 4, Ref. 49). Text in PDF www.elis.sk Keywords: Sjögren syndrome, keratoconjunctivitis sicca, xerostomia, labial minor salivary glands, biopsy, focal lymphocytic infiltration, focus score.

Normal salivary gland ultrasonography could rule out the diagnosis of Sjögren’s syndrome in anti-SSA-negative patients with sicca syndrome

ObjectiveTo evaluate the relevance of salivary gland ultrasound (SGUS) and its place in the diagnostic algorithm in patients referred with dry syndrome (DS) for a suspicion of Sjögren’s syndrome (SS).MethodsWe...

Ultrasonography predicts the results of labial salivary gland biopsy in patients with suspected Sjögren's syndrome: a matrix risk model.

Objective:Although a positive result of labial salivary gland biopsy (LSGB) is critical for the diagnosis of Sjögren’s syndrome, rheumatologists prefer assessing the non-invasive objective items and hope to learn the predicted probability of positive LSGB before referring patients with suspected Sjögren’s syndrome to receive biopsy. This study aimed to explore the predictive value of combined B-mode ultrasonography (US) and shear-wave elastography (SWE) examination on LSGB results.Methods:A derivation cohort and later a validation cohort of patients with suspected Sjögren’s syndrome were recruited. All participants received clinical assessments, B-mode US and SWE examination on bilateral parotid and submandibular glands before LSGB. Positive LSGB was defined by a focus score ⩾1 per 4 mm2 of glandular tissue.Results:In the derivation cohort of 91 participants, either the total US scores or the total SWE values of four glands significantly distinguished patients with positive LSGB from those with negative results (area under the curve (AUC) = 0.956, 0.825, both p < 0.001). The positive predictive value (PPV) was 100% in patients with total US scores ⩾9 or with total SWE values ⩾33 kPa. The negative predictive value (NPV) was 100% in patients with total US scores <5, but 68% in patients with total SWE values <27 kPa. A matrix risk model was derived based on the combination of total US scores and total SWE values. Patients can be stratified into high, moderate, and low risk of positive LSGB. In the validation cohort of 52 participants, the PPV was 94% in the high-risk subpopulation and the NPV was 93% in the low-risk subpopulation.Conclusion:A novel matrix risk model based on the combined B-mode US and SWE examination can help rheumatologists to make a shared decision with suspected Sjögren’s syndrome patients on whether the invasive procedure of LSGB should be performed.

The complement system in primary Sjögren's syndrome: the expression of certain cascade and regulatory proteins in labial salivary glands - observational study.

IntroductionThe complement cascade and regulatory proteins are involved in the pathogenesis of the Sjögren’s syndrome and other autoimmune diseases. The complement activation via the alternative pathway was recognized as a major pathogenic mechanism in autoimmune conditions. The aim of this study was to assess expression of complement cascade components and regulatory proteins in minor salivary glands in patients with primary Sjögren’s syndrome (pSS).Materials and methodsThe expression of C1q and C5b-9 – membrane attack complex and regulatory proteins such as: membrane cofactor protein (MCP), decay-accelerating factor (DAF) and protectin were examined using immunochemistry method in specimens from biopsy of minor salivary glands in pSS patients. The biopsy material was obtained from 20 pSS patients, 5 patients with non-specific sialadenitis and from 5 patients with suspicion of dryness syndrome without sialadenitis confirmation.ResultsNone of the examined samples showed the expression of C1q or the effector C5b-9. Membrane cofactor protein expression was lower in pSS group than in both non-specific sialadenitis and noninflamed salivary glands. The inflammatory cells in pSS samples partially expressed MCP. There were differences in the sites and intensity of membrane protectin expression exclusively on the luminal surfaces in pSS; on the luminal and, partially, antiluminal surface in non-specific inflammation, and on the entire cell surface in unaffected salivary glands. There were no DAF expression in salivary gland tissue in biopsy specimens in all studied subjects.ConclusionsThe study demonstrated the absence of complement-cascade proteins (C1q, MAC) in the salivary glands of pSS patients, which may indicated a lack of local complement activation via the classical pathway and the observed gland tissue damage being due to a mechanism other than MAC-induced cytolysis. The differences in the expression of complement regulatory proteins between pSS, non-specific sialadenitis, and normal salivary glands may indicate that alternative functions of these regulatory proteins may be of greater significance in pSS. Low MCP expression in pSS in comparison with non-specific sialadenitis and normal salivary glands, may suggest altered modulation of cell-mediated immunity in pSS. The differences in the location and intensity of protectin (CD59) expression indicates a possibility of reducing the proinflammatory effect of protectin in pSS.

The relationship between Sjogren's syndrome, systemic sclerosis and lymphoproliferative diseases

To study demographic, clinical, laboratory and immunological characteristics of patients with a combination of primary Sjogrens syndrome (pSS) and SSc and diagnosed lymphoproliferative diseases (LPDs); to characterize morphological/immunomorphological variants and course of non-Hodgkins lymphomas (NHL), developing in patients with these rheumatic diseases (RDs). In 19982018 at the Nasonova Research Institute of Rheumatology, 13 patients with clinical and laboratory manifestations of pSS (12) and SSc (13) were diagnosed with various lymphoproliferative diseases (LPDs). In 3 cases, an induced RD was observed: 1 case of a diffuse, rapidly progressive form of SSc, 2 cases of pSS in combination with a limited form of SSc after chemotherapy and radiation therapy of Hodgkins lymphoma (1), B-cell NHL (1) and CR of the breast (1) respectively. The first 2 cases were excluded from the analysis, since the development of lymphomas is not pathogenetically associated with RD. Of 11 patients with LPDs, 10 after a long course of RDs were diagnosed with NHL [MALT lymphoma of the parotid salivary glands 7, disseminated MALT lymphoma 2, disseminated MALT lymphoma with transformation into diffuse large B-cell lymphoma (DLBCL) 1]. RDs debuted with Raynauds phenomenon (RP) in 64.5% and pSS manifestations in 45.5% of patients. Stomatological manifestations of pSS were characterized by recurrent parotitis in 36%, significant parotid gland enlargement with massive infiltration of labial salivary glands (focus score 4) in 100%, severe xerostomia in 70%, extraglandular manifestations and lymphadenopathy in 50% of patients. The course of the SSc was characterized by mild RP with various types of capillaroscopic changes and mild lung changes and non-significant progression during long-term follow-up (median 22 years). The entire spectrum of SSс specific antibodies (anticentromere antibodies 60%, antibodies to ribonucleoprotease III 30%, Pm/Scl 10%), excepting antibodies to topoisomerase I, as well as pSS specific autoantibodies (antiRo/La 70%, RF (rheumatoid factor) 90%), were detected in patients with a combination of these RDs. pSS is often combined with a limited form of SSc regardless of the type of autoantibodies detected. The presence of pSS, rather than SSc, is a high-risk factor for the development of NHL in this group of patients. The patients with pSS and SSc are characterized by a steady progression of pSS with a slow and mild course of SSc throughout the observation period. The development of severe stomatological manifestations and high immunological activity of pSS contribute to the development of localized MALT lymphomas (70%) and disseminated MALT lymphomas (30%) with primary lesions of the salivary glands and transformation into DLBCL in case of their late diagnosis. The optimal method for preventing the development of NHL in this group of patients is the early diagnosis of pSS, the appointment of alkylating cytotoxic agents and/or anti-B-cell therapy in the early stages of pSS. Given the possibility of transformation of localized NHL into DLBCL, for early diagnosis, minimally invasive surgical biopsies of significantly enlarged parotid salivary glands should be performed before glucocorticoids are prescribed. Detection of positive B-cell clonality and lymphoepithelial lesions in the parotid salivary gland is considered a predictor of MALT lymphoma development during follow-up. Localized and disseminated MALT lymphomas in patients with pSS and SSc respond well to therapy, in contrast to MALT lymphomas transformed into DLBCL.

Histomorphological investigations on the lips of Rahmani sheep (Ovis aries): A scanning electron and light microscopic study.

This study was conducted to provide comprehensive information on the anatomical and histological features of the upper (UL) and lower (LL) lips of Rahmani sheep by gross examination, morphometric analysis in addition to Light and scanning electron microscope (SEM). Samples from normal healthy adult male sheep heads were collected directly after slaughtering. UL and LL were dissected, and specimens were collected for both light and SEM. The UL was longer approximately by one-fold and thicker by one-fold at the median and the oral angle areas, and by one- and half-folds at the paramedian area than the LL. The free border of both lips was characterized rostrally by the presence of labial projections. By SEM the edges of the inner aspect of the lips and of the philtrum were distinguished by labial projections. These projections which surrounding the philtrum subdivided into polygonal areas with numerous keratinized scales especially at the apical parts which increased dorsally toward the nostril. Most of the openings of the upper labial salivary glands were volcanic crater-shaped while that of the lower labial salivary glands were various shapes; round folded margin, rosette and whirlpool shaped. Histologically, the shape of the projection or papillae differs at the median and paramedian areas of the UL than the LL. However, there was no differences near the oral angle area. In conclusion, the shape, size and amount of keratinization of the papillae may offer efficiency to the lips during feeding process.

Proteomics-based analysis indicating α-enolase as a potential biomarker in primary Sjögren's syndrome.

Primary Sjögren's syndrome (pSS) is a chronic autoimmune disease. Its etiology is not well understood. Salivary glands are the main target organ in pSS, investigating the changes of salivary protein in pSS patients may not only be a valuable way of identifying new biomarkers/antigens for pSS, but also of revealing the pathogenesis underlying this autoimmune disease. In the present study, we aimed to investigate new biomarkers and explore their potential role in pSS. In this study, α-enolase (ENO1) was found to be overexpressed in pSS by 1D gel electrophoresis/mass spectrometry. The finding was verified by Western blots, immunohistochemistry (IHC), and polymerase chain reaction (PCR) results in both saliva and labial salivary glands. The expression level of immunoglobulin G (IgG) antibody to ENO1 was then tested by enzyme-linked immunosorbent assay (ELISA). ENO1 autoantibody was found to be overexpressed in pSS compared with healthy controls. The effects of ENO1 overexpression on rat submandibular gland cell line SMG-C6 was investigated in vitro. The expressions of proteins related to saliva secretion and immunomodulatory were upregulated in ENO1 overexpressed SMG-C6 cells. Both ENO1 and anti-ENO1 autoantibody are overexpressed in pSS patients. Nevertheless, their potential role in the pathogenesis of pSS warrants further study.

15896 Utility of the labial salivary gland punch biopsy for the diagnosis of Sjögren syndrome

Background: Several labial salivary gland biopsy techniques exist, each with varying degrees of complexity, efficacy and potential omplications. Few studies evaluate the utility of the labial salivary gland punch biopsy (LSGPB) for the diagnosis of Sjögren syndrome (SS).