Erlin A Haacke

B-cell hyperactivity in primary Sjögren's syndrome

Primary Sjögrens syndrome (pSS) is characterized by mononuclear inflammatory infiltrates and IgG plasma cells in salivary and lacrimal glands which lead to irreversible destruction of the glandular tissue and is accompanied by sensation of dryness of mouth and eyes. B cells play a central role in the immunopathogenesis and exhibit signs of hyperactivity. Hyperactivity of B cells is the consequence of the coordinated and integrated action of stimulation of the B-cell receptor, CD40 and toll-like receptors in the presence of appropriate cytokines. As discussed, overexpression of type I IFN and BAFF on one hand and IL-6 and IL-21 on the other hand are critically involved in the enhanced plasma cell formation in pSS patients. Hyperactivity of B cells results in secretion of autoantibodies and production of various cytokines. These insights in the role of B cells in the pathogenetic process of pSS offer ample targets for successful therapeutical intervention in pSS.

Towards personalised treatment in primary Sjögren's syndrome: baseline parotid histopathology predicts responsiveness to rituximab treatment.

Objectives The aims of this study were (1) to assess the effect of rituximab (RTX; anti-CD20) treatment in patients with primary Sjögrens syndrome (pSS) based on sequential parotid biopsies obtained in a placebo-controlled, randomised clinical trial, and (2) to assess the prognostic value of the histological characteristics of parotid gland tissue with regard to responsiveness to RTX treatment. Methods In a double-blinded, placebo-controlled trial, sequential parotid gland biopsies were taken from 20 RTX-treated and 10 placebo-treated patients with pSS, at baseline and 12 weeks after treatment. The relative amount of lymphocytic infiltrate (stained for CD45), absolute number of T cells and B cells per mm2 parenchyma (stained for CD3 and CD20, respectively), focus score, number of germinal centres and of lymphoepithelial lesions per mm2 in parotid gland parenchyma were assessed. Histopathological data were compared between clinical responders (decrease in European League Against Rheumatism Sjögrens Syndrome Disease Activity Index (ESSDAI) score of ≥3 at 12 weeks compared with baseline) and non-responders (change in ESSDAI<3) to RTX treatment. Results In RTX-treated patients, a significant reduction in the number of CD20+ B cells/mm2 parenchyma was observed, while no such reduction was observed in placebo-treated patients. The number of CD3+ T cells/mm2 in parenchyma did not change in either group. Furthermore, the number and the severity of lymphoepithelial lesions/mm2 and number of germinal centres/mm2 was significantly reduced in RTX-treated patients, but did not change in placebo-treated patients. When comparing the pretreatment characteristics of clinical responders with non-responders, the median number of CD20+ B cells/mm2 parenchyma at baseline was significantly higher in responders (1871 vs 353 cells/mm2, p<0.05). Other histopathological baseline characteristics were not predictive for response to RTX treatment. Conclusions RTX treatment in pSS leads to a major reduction of lymphocytic infiltration and to fewer B cells, germinal centres and lymphoepithelial lesions in parotid gland parenchyma. A high pretreatment number of CD20+ B cells/mm2 parotid gland parenchyma predicts better responsiveness of patients with pSS to RTX treatment. Pretreatment parotid gland histopathological characteristics could therefore contribute to a more personalised treatment approach to pSS.

Persistence of immunoglobulin-producing cells in parotid salivary glands of patients with primary Sjogren's syndrome after B cell depletion therapy

Objectives To assess the persistence of immunoglobulin-producing cell populations in the parotid salivary glands of patients with primary Sjögrens syndrome (pSS) after B cell depletion therapy with rituximab. Methods Thirteen patients with pSS and four control patients were included in this study. Patients with pSS were treated with rituximab or placebo. Sequence analysis was carried out on IgA- and IgG-encoding transcripts extracted from parotid salivary gland biopsy specimens taken before treatment and at 12–16 and 36–52 weeks after treatment. Results At baseline, many clonally related sequences were seen in patients with pSS. The number of clonal expansions was significantly higher in patients with pSS than in control patients. Clonal expansions were composed of IgA- and/or IgG-expressing cells. Rituximab did not significantly alter the degree of clonal expansions. Groups of clonally related cells had members which were shared between biopsy specimens taken before and after treatment. Mutation frequencies of immunoglobulin sequences from clonally related cells in patients with pSS were higher after treatment. Conclusions Rituximab treatment does not alter the characteristic features of increased clonal expansions seen in the parotid salivary glands of patients with pSS. The presence of clonally related immunoglobulin-producing cells before and after rituximab treatment strongly suggests that immunoglobulin-producing cells persist in salivary glands of patients with pSS despite B cell depletion. The presence of mixed isotype expression within groups of clonally related cells indicates local class switching in salivary glands of patients with pSS. Persistent immunoglobulin-producing cells may underlie disease relapse after treatment.

Standardisation of labial salivary gland histopathology in clinical trials in primary Sjögren's syndrome

Labial salivary gland (LSG) biopsy is used in the classification of primary Sjögrens syndrome (PSS) and in patient stratification in clinical trials. It may also function as a biomarker. The acquisition of tissue and histological interpretation is variable and needs to be standardised for use in clinical trials. A modified European League Against Rheumatism consensus guideline development strategy was used. The steering committee of the ad hoc working group identified key outstanding points of variability in LSG acquisition and analysis. A 2-day workshop was held to develop consensus where possible and identify points where further discussion/data was needed. These points were reviewed by a subgroup of experts on PSS histopathology and then circulated via an online survey to 50 stakeholder experts consisting of rheumatologists, histopathologists and oral medicine specialists, to assess level of agreement (0–10 scale) and comments. Criteria for agreement were a mean score ≥6/10 and 75% of respondents scoring ≥6/10. Thirty-nine (78%) experts responded and 16 points met criteria for agreement. These points are focused on tissue requirements, identification of the characteristic focal lymphocytic sialadenitis, calculation of the focus score, identification of germinal centres, assessment of the area of leucocyte infiltration, reporting standards and use of prestudy samples for clinical trials. We provide standardised consensus guidance for the use of labial salivary gland histopathology in the classification of PSS and in clinical trials and identify areas where further research is required to achieve evidence-based consensus.

B Cell Depletion Therapy Normalizes Circulating Follicular Th Cells in Primary Sjögren Syndrome

Objective. To assess the effect of B cell depletion therapy on effector CD4+ T cell homeostasis and its relation to objective measures of disease activity in patients with primary Sjögren syndrome (pSS). Methods. Twenty-four patients with pSS treated with rituximab (RTX) and 24 healthy controls (HC) were included. Frequencies of circulating effector CD4+ T cell subsets were examined by flow cytometry at baseline and 16, 24, 36, and 48 weeks after the first RTX infusion. Th1, Th2, follicular Th (TFH), and Th17 cells were discerned based on surface marker expression patterns. Additionally, intracellular cytokine staining was performed for interferon-γ, interleukin (IL)-4, IL-21, and IL-17 and serum levels of these cytokines were analyzed. Results. In patients with pSS, frequencies of circulating TFH cells and Th17 cells were increased at baseline compared with HC, whereas frequencies of Th1 and Th2 cells were unchanged. B cell depletion therapy resulted in a pronounced decrease in circulating TFH cells, whereas Th17 cells were only slightly lowered. Frequencies of IL-21–producing and IL-17–producing CD4+ T cells and serum levels of IL-21 and IL-17 were also reduced. Importantly, the decrease in circulating TFH cells was associated with lower systemic disease activity over time, as measured by the European League Against Rheumatism Sjögren’s Syndrome Disease Activity Index scores and serum IgG levels. Conclusion. B cell depletion therapy in patients with pSS results in normalization of the elevated levels of circulating TFH cells. This reduction is associated with improved objective clinical disease activity measures. Our observations illustrate the pivotal role of the crosstalk between B cells and TFH cells in the pathogenesis of pSS.

Germinal centres in diagnostic labial gland biopsies of patients with primary Sjogren's syndrome are not predictive for parotid MALT lymphoma development

Objective Patients with primary Sjögren’s syndrome (pSS) have an increased risk of developing non-Hodgkin’s lymphoma (NHL), particularly parotid gland mucosa-associated lymphoid tissue (MALT) lymphomas. Presence of germinal centres (GCs) in labial gland biopsies has been suggested as predictive factor for NHL. We assessed whether presence of GCs is increased in labial gland biopsies from patients with pSS who developed parotid MALT lymphoma, the dominant NHL-subtype in pSS, compared with patients with pSS who did not develop lymphoma. Methods Eleven labial gland biopsies from patients with pSS that were taken prior to parotid MALT lymphoma development were compared with biopsies of 22 matched pSS controls (1:2) who did not develop lymphoma. Biopsies were evaluated for GCs (H&E and Bcl6). Results Labial gland biopsies of pSS MALT lymphoma patients, revealed GCs in 2/11 (18%) H&E sections and 3/11 (27%) Bcl6 stained sections. In controls, GCs were present in 4/22 (18%) of H&E sections and 5/22 (23%) of Bcl6 stained sections. Conclusion Presence of GCs in labial gland biopsies does not differ between patients with pSS that develop parotid MALT lymphoma and patients with pSS who do not develop lymphoma. The presence of GCs in labial gland biopsies is therefore not a predictive factor for pSS-associated parotid MALT lymphomas.

In primary Sjögren's syndrome high absolute numbers and proportions of B cells in parotid glands predict responsiveness to rituximab as defined by ESSDAI, but not by SSRI

With great interest we have read the letter to the editor by Cornec et al 1 regarding our paper ‘Towards personalised treatment in primary Sjogrens syndrome (pSS): baseline parotid histopathology predicts responsiveness to rituximab treatment’.2 In essence, we showed in our paper that absolute numbers of CD20+ cells/mm2 of parenchyma of parotid gland tissue are predictive for the responsiveness of patients with pSS to rituximab (RTX) treatment. Cornec et al argue that there is a discrepancy in outcomes presented in their study and our study,1 as they observed that a high proportion of minor salivary gland B cells predict the absence of a clinical response to RTX.3 As we will show and explain here, there is no inconsistency between the two studies and most of the apparent discrepancy is likely the result of differences in how the tissues are analysed and how the disease activity is established. A major difference in the two studies is how B cells are assessed in tissue sections of salivary gland biopsies of patients with pSS before (and after) RTX treatment. We measured absolute numbers of CD20+ B cells/mm2 of parenchyma, while Cornec et al assessed the proportion of B cells.1 ,3 Obviously, even when there is a change in absolute numbers of B cells in the tissue, the B/T cell ratio still can remain the same. Thus, although higher numbers of B cells, do not need to be reflected per se in higher proportions of B cells, we also found in our study that patients with higher absolute numbers of B cells in the glandular tissue, had a higher B/B+T cell ratio. Furthermore, responders to RTX, as defined by a decrease in European League Against …

Need for consensus guidelines to standardise the assessment of germinal centres and other histopathological parameters in salivary gland tissue of patients with primary Sjögren's syndrome

We have read with great interest the letter to the editor by van Roon et al 1 commenting on our paper ‘Towards personalised treatment in primary Sjogrens syndrome: baseline parotid histopathology predicts responsiveness to rituximab treatment’.2 The authors argue that there is a need for standardisation of the histopathological characteristics of salivary gland tissue of patients with primary Sjogrens syndrome (pSS), in general, and of the presence of germinal centres (GCs), in particular. We fully agree with van Roon et al 1 and other authors about the need for consensus guidelines to standardise the histopathological evaluation of salivary gland biopsies in patients with pSS.3 A standardised scoring system may facilitate prognostication and stratification of patients with pSS and is needed for a valid evaluation of various clinical trials.3 In particular, histological definition of GCs in salivary gland tissue is warranted, since these structures can be difficult to detect in diagnostic H&E-stained tissue sections. Detection of GCs in the periductal lymphoid infiltrates of the salivary glands is clinically relevant, because the presence of these structures is associated with more severe disease.4 Importantly, the presence of GCs in minor salivary gland biopsies has been postulated to be a predictor of patients who are at risk of lymphoma development.5 …

Standardisation of the detection of germinal centres in salivary gland biopsies of patients with primary Sjögren’s syndrome is needed to assess their clinical relevance

We thank Alunno et al 1 for their comments, as a response to our recent publication2 in which we describe that, in contrast to the prevailing view, germinal centres in diagnostic labial gland biopsies are not predictive for the development of mucosa-associated lymphoid tissue (MALT) lymphoma in parotid salivary glands in patients with (primary) Sjogren’s syndrome (pSS). As we2 and others also noted before,3–6 Alunno et al 1 underpin in their comments the need for standardisation of the detection method of germinal centres in salivary gland biopsies in patients with pSS. Germinal centres are structures that arise in B-cell follicles of secondary lymphoid organs as a response to antigenic stimulation. In these structures, high-affinity memory B-cells are generated, as a consequence of an intimate interplay between B-cells, follicular helper T cells (TFH) and immune-complex presenting follicular dendritic cells (FDCs). Germinal centres can also be found within B-cell areas of ectopic (tertiary) lymphoid tissue that develops in target tissues of various rheumatic autoimmune diseases, including pSS.7 While detection in secondary lymphoid tissue is …

Parotid Gland Biopsy, the Alternative Way to Diagnose Sjogren Syndrome

Salivary gland biopsy is a technique broadly applied for the diagnosis of Sjögren syndrome (SS), lymphoma in SS, and connective tissue disorders (sarcoidosis, amyloidosis). In SS characteristic histology findings are found, including lymphocytic infiltration surrounding the excretory ducts in combination with destruction of acinar tissue. In this article the main techniques are described for taking labial and parotid salivary gland biopsies with respect to their advantages, postoperative complications, and usefulness for diagnostic procedures, monitoring disease progression, and evaluation of treatment.