Interleukin-33 produced by M2 macrophages and other immune cells contributes to Th2 immune reaction of IgG4-related disease.
ABSTRACT: IgG4-related disease (IgG4-RD) is characterized by elevated serum IgG4 and marked infiltration of IgG4-positive cells in multiple organs. Interleukin-33 (IL-33) is a recently described cytokine that is secreted by damaged epithelial cells, macrophages, and dendritic cells, and potently activates helper T type 2 (Th2) immune responses, which have been suggested to play a major role in IgG4 production of IgG4-RD. Here, we assessed the expression of IL-33 and related molecules in the salivary glands (SGs) of patients with IgG4-RD versus that in patients with Sjögren's syndrome (SS) and controls. Expression of IL-33 and its receptor (ST2) was strongly detected around ectopic germinal centers (GCs) in the SGs from patients with IgG4-RD, whereas IL-33 was expressed only in epithelial cells in patients with SS and controls. Moreover, IL-33 and CD68+/CD163+ macrophages were mainly distributed around ectopic GCs in patients with IgG4-RD. Double immunofluorescence staining showed that IL-33 expression co-localized with CD68+/CD163+ macrophages. Finally, mRNA expression levels of IL-33 showed a positive correlation to those of Th2 cytokines (IL-4 and IL-13) in patients with IgG4-RD. Our data suggest that IL-33 produced by M2 macrophages might contribute to the pathogenesis of IgG4-RD via aberrant activation of Th2 immune responses.
Project description:Macrophage polarization is increasingly recognised as an important pathogenetic factor in inflammatory and neoplastic diseases. Proinflammatory M1 macrophages promote T helper (Th) 1 responses and show tumoricidal activity. M2 macrophages contribute to tissue repair and promote Th2 responses. CD68 and CD163 are used to identify macrophages in tissue sections. However, characterisation of polarised macrophages in situ has remained difficult. Macrophage polarisation is regulated by transcription factors, pSTAT1 and RBP-J for M1, and CMAF for M2. We reasoned that double-labelling immunohistochemistry for the detection of macrophage markers together with transcription factors may be suitable to characterise macrophage polarisation in situ. To test this hypothesis, we have studied conditions associated with Th1- and Th2-predominant immune responses: infectious mononucleosis and Crohn's disease for Th1 and allergic nasal polyps, oxyuriasis, wound healing and foreign body granulomas for predominant Th2 response. In all situations, CD163+ cells usually outnumbered CD68+ cells. Moreover, CD163+ cells, usually considered as M2 macrophages, co-expressing pSTAT1 and RBP-J were found in all conditions examined. The numbers of putative M1 macrophages were higher in Th1- than in Th2-associated diseases, while more M2 macrophages were seen in Th2- than in Th1 related disorders. In most Th1-related diseases, the balance of M1 over M2 cells was shifted towards M1 cells, while the reverse was observed for Th2-related conditions. Hierarchical cluster analysis revealed two distinct clusters: cluster I included Th1 diseases together with cases with high numbers of CD163+pSTAT1+, CD68+pSTAT1+, CD163+RBP-J+ and CD68+RBP-J+ macrophages; cluster II comprised Th2 conditions together with cases displaying high numbers of CD163+CMAF+ and CD68+CMAF+ macrophages. These results suggest that the detection of pSTAT1, RBP-J, and CMAF in the context of CD68 or CD163 expression is a suitable tool for the characterisation of macrophage polarisation in situ. Furthermore, CD163 cannot be considered a reliable M2 marker when used on its own.
Project description:IgG4-related disease (IgG4-RD) is a multi-organ autoimmune disease characterized by elevated serum IgG4 concentration. Although serum IgG4 concentration is widely used as a biomarker for IgG4-RD and type 1 autoimmune pancreatitis (AIP), a pancreatic manifestation of IgG4-RD, a significant number of patients have normal serum IgG4 levels, even in the active phase of the disease. Recently, we reported that the development of experimental AIP and human type 1 AIP is associated with increased expression of IFN-? and IL-33 in the pancreas. In this study, we assessed the utility of serum IFN-? and IL-33 levels as biomarkers for type 1 AIP and IgG4-RD. Serum IFN-? and IL-33 concentrations in patients who met the diagnostic criteria for definite type 1 AIP and/or IgG4-RD were significantly higher than in those with chronic pancreatitis or in healthy controls. Strong correlations between serum IFN-?, IL-33, and IgG4 concentrations were observed. Diagnostic performance of serum IFN-? and IL-33 concentrations as markers of type 1 AIP and/or IgG4-RD was comparable to that of serum IgG4 concentration, as calculated by the receiver operating characteristic curve analysis. Induction of remission by prednisolone treatment markedly decreased the serum concentration of these cytokines. We conclude that serum IFN-? and IL-33 concentrations can be useful as biomarkers for type 1 AIP and IgG4-RD.
Project description:IgG4-related disease (IgG4-RD) is a fibro-inflammatory disorder involving virtually every organ with a risk of organ dysfunction. Despite recent studies regarding B cell and T cell compartments, the disease's pathophysiology remains poorly understood. We examined and characterized subsets of circulating lymphocytes in untreated patients with active IgG4-RD. Twenty-eight consecutive patients with biopsy-proven IgG4-RD were included in a prospective, multicentric study. Lymphocytes' subsets were analyzed by flow cytometry, with analysis of TH1/TH2/TH17, TFH cells, and cytokine release by peripheral blood mononuclear cells. Results were compared to healthy controls and to patients with primary Sjögren's syndrome. Patients with IgG4-RD showed an increase of circulating T regulatory, TH2, TH17, and CD4+CXCR5+PD1+ TFH cell subsets. Accordingly, increased levels of IL-10 and IL-4 were measured in IgG-RD patients. TFH increase was characterized by the specific expansion of TFH2 (CCR6-CXCR3-), and to a lesser extent of TFH17 (CCR6+CXCR3-) cells. Interestingly, CD4+CXCR5+PD1+ TFH cells normalized under treatment. IgG4-RD is characterized by a shift of circulating T cells toward a TH2/TFH2 and TH17/TFH17 polarization. This immunological imbalance might be implicated in the disease's pathophysiology. Treatment regimens targeting such T cells warrant further evaluation.
Project description:IgG4 subclass antibodies are expressed in alternative Th2 environments featuring high IL-10 expression, including several solid tumors such as melanoma. To induce tolerance, allergen immunotherapy mediates antibody class switching from pro-inflammatory IgE to anti-inflammatory IgG4. We previously reported that IgG4 drives allergic M2 macrophages toward tolerogenic states. Here we assessed the roles of IgG4 and macrophage activation in colorectal cancer (CRC). In this observer-blinded, case-control study, we analyzed total circulating serum IgE, IgG1 and IgG4 levels in CRC (n = 38) patients with (n = 13, TxNxM1) or without (n = 25, TxNxM0) metastasis, and in healthy donors (n = 21). Primary cultures of circulating monocyte-derived macrophages from healthy controls and CRC patients were further evaluated in their responses to stimulation with IgG1 or IgG4. We found higher absolute serum levels of IgG4 in patients with CRC. IgG4 enabled polarization of macrophages derived from CRC patients and healthy controls into alternatively-activated tolerogenic M2b phenotypes. IgG4-stimulated M2 macrophages were characterized by lower surface CD206, CD163, CD14, and CD11b expression and higher CCL-1, IL-10, and IL-6 production. IgG4 was less potent that IgG1 in triggering antibody-dependent cell-mediated phagocytosis (ADCP) of cancer cells. Further, higher z-normalized IgG4/-IgE sera level ratios correlated with the presence of metastasis (<i>p</i> = .0247 and <i>p</i> = .0009, respectively) in CRC patients. High IgG4 in CRC synergizes with macrophages in shaping an immunosuppressive microenvironment and impairs anti-cancer effector cell functions. The shift of serum IgG4/IgE ratios toward enhanced tolerance induction in metastatic disease indicates a role for high IgG4 in disease progression and poor prognostic outcome.
Project description:IgG4-related disease (IgG4-RD) is a novel systemic disease entity characterized by elevated serum IgG4 and tissue infiltration of IgG4-positive plasma cells accompanied by severe fibrosis. Although recent studies demonstrated that innate immune cells including monocytes and macrophages might promote local fibrosis and IgG4 production, the pathological mechanism remains unclear. In this study, we sought to identify the disease-associated genes, especially innate immune molecules. Gene expression was analyzed by DNA microarray in submandibular glands (SMGs) from patients with IgG4-RD (n?=?5), chronic sialoadenitis (CS) (n?=?3), and controls (n?=?3). Differentially expressed genes (DEGs) were validated by real-time polymerase chain reaction (PCR) and immunohistochemical staining in IgG4-RD (n?=?18), CS (n?=?4), Sjögren syndrome (n?=?11), and controls (n?=?10). Gene expression patterns in the 3 groups were quite different from each other by the pvclust method and principal components analysis. In IgG4-RD, 1028 upregulated genes and 692 downregulated genes were identified as DEGs (P?<?0.05). Gene Ontology (GO) term analysis indicated that the upregulated DEGs in IgG4-RD encoded proteins involved in T/B cell activation and chemotaxis. PCR validated significantly higher expression of macrophage receptor with collagenous structure (MARCO), a pattern-recognition receptor, in IgG4-RD compared with the other groups (P?<?0.01). Immunohistochemical analysis confirmed that the expression pattern of MARCO was similar to that of the M2 macrophage marker CD163. MARCO was identified as a disease-associated molecule in IgG4-RD by DNA microarray. Moreover, M2 macrophages might contribute to the initiation of IgG4-RD via MARCO.
Project description:We previously suggested a relationship between ocular immunoglobulin (Ig)G4-related disease (IgG4-RD) and marginal zone lymphomas (MZLs). However, the cytokine background associated with these disorders and whether it differs between ocular adnexal MZLs with (IgG4-associated MZL) and without (IgG4-negative MZL) numerous IgG4(+) plasma cells are unknown. In this study, we identified the mRNA expression pattern of Th2 and regulatory T-cell (Treg) cytokines in IgG4-RD and in IgG4-associated MZL and IgG4-negative MZL using real-time polymerase chain reaction analysis. Ocular IgG4-RD and IgG4-associated MZL exhibited significantly higher expression ratios of interleukin (IL)-4/β-actin, IL-10/β-actin, IL-13/β-actin, transforming growth factor (TGF) β1/β-actin, and FOXP3/β-actin than did IgG4-negative MZL (p < 0.05). This finding further supports our prior observations that a significant subset of ocular MZLs arises in the setting of IgG4-RD. Furthermore, the presence of a different inflammatory background in IgG4-negative MZLs suggests that IgG4-associated MZLs may have a different pathogenesis.
Project description:Background:The exact etiology and pathogenesis of granulomatous lobular mastitis (GLM) are yet to be illuminated. This study aimed to investigate CD68, CD163-positive M2 macrophages, CD57-positive natural killer (NK) cells, and IgG4 in GLM lesion tissue to explore their correlation with the occurrence and clinical features of GLM. Methods:Surgical pathologic specimens of GLM were collected from patients admitted to Hunan Provincial People's Hospital between October, 2014 and October 2015. Based on the postoperative pathological diagnosis, the tissues were divided into 3 groups: the experimental group (GLM, n=36), control group 1 (plasma cell mastitis, PCM, n=17), and control group 2 (breast cystic hyperplasia, n=10). Immunohistochemical staining was carried out using Elivision super testing to detect CD68, CD163, CD57, and IgG4 expression in the pathological tissue samples. The relationship between clinical parameters, including age, reproductive condition, nipple retraction, and tumor size, and the expressions of CD68, CD163, CD57, and IgG4 was analyzed. Results:There was no obvious difference in the levels of CD68, CD163, and CD57 expression between the GLM group and the PCM group, although both groups had higher expression levels of expression than the breast cystic hyperplasia group (P<0.05). In the GLM group, the expression level of CD57 at 2 weeks-3 months was significantly higher than at ?2 weeks (P<0.05). The expression level of CD57 in PCM patients >2 years after lactation was significantly higher than in patients ?2 years after lactation (P<0.05). The level of IgG4 expression in GLM patients with nipple retraction was significantly higher than in those without nipple retraction (P<0.05). Conclusions:Inflammatory cells are closely linked to the occurrence of GLM and PCM. In our study, both the GLM and PCM groups had low expression of IgG4, but the expression level of IgG4 in GLM patients with inverted nipples was significantly higher than that in patients without inverted nipples. This suggests that there may be two different clinical subtypes of GLM. Furthermore, our research also found that NK cells can provide a basis for GLM clinical staging.
Project description:IgG4-related disease (IgG4-RD) is a systemic condition of unknown cause characterized by highly fibrotic lesions with dense lymphoplasmacytic infiltrates. CD4(+) T cells constitute the major inflammatory cell population in IgG4-RD lesions.We used an unbiased approach to characterize CD4(+) T-cell subsets in patients with IgG4-RD based on their clonal expansion and ability to infiltrate affected tissue sites.We used flow cytometry to identify CD4(+) effector/memory T cells in a cohort of 101 patients with IgG4-RD. These expanded cells were characterized by means of gene expression analysis and flow cytometry. Next-generation sequencing of the T-cell receptor ? chain gene was performed on CD4(+)SLAMF7(+) cytotoxic T lymphocytes (CTLs) and CD4(+)GATA3(+) TH2 cells in a subset of patients to identify their clonality. Tissue infiltration by specific T cells was examined by using quantitative multicolor imaging.CD4(+) effector/memory T cells with a cytolytic phenotype were expanded in patients with IgG4-RD. Next-generation sequencing revealed prominent clonal expansions of these CD4(+) CTLs but not CD4(+)GATA3(+) memory TH2 cells in patients with IgG4-RD. The dominant T cells infiltrating a range of inflamed IgG4-RD tissue sites were clonally expanded CD4(+) CTLs that expressed SLAMF7, granzyme A, IL-1?, and TGF-?1. Clinical remission induced by rituximab-mediated B-cell depletion was associated with a reduction in numbers of disease-associated CD4(+) CTLs.IgG4-RD is prominently linked to clonally expanded IL-1?- and TGF-?1-secreting CD4(+) CTLs in both peripheral blood and inflammatory tissue lesions. These active, terminally differentiated, cytokine-secreting effector CD4(+) T cells are now linked to a human disease characterized by chronic inflammation and fibrosis.
Project description:Background: Although there are multiple ways to manage immunoglobulin G4-related disease (IgG4-RD), including treatment with glucocorticoids, "steroid-sparing" immunosuppressive drugs, or biologic agents, few clinical trials on IgG4-RD have been conducted. This study aimed to compare the efficacy and safety of glucocorticoids (GCs) combined with cyclophosphamide (CYC) or mycophenolate mofetil (MMF) in IgG4-RD patients. This cohort study was registered at ClinicalTrials.gov (ID: NCT01670695). Methods: This retrospective study included 155 IgG4-RD patients who received GCs with CYC or MMF at the Department of Rheumatology at Peking Union Medical College Hospital between January 2012 and July 2018. Propensity score matching (PSM) was conducted to match two groups of patients based on their baseline clinical characteristics. Treatment response, relapse rate, and drug safety were analyzed. The treatment response was evaluated based on complete response (CR), partial response (PR), and no change (NC), and the cumulative relapse rate and adverse events in each treatment group were compared using Kaplan-Meier curves and log-rank test, respectively. Results: Of the 155 IgG4-RD patients, 90 were treated with GCs plus CYC (group I) and 65 with GCs plus MMF (group II). After propensity score-matched (PSM) analysis, 108 patients were selected (54 in each group), 49 of whom had "definite" IgG4-RD, 8 "probable" IgG4-RD, and 51 "possible" IgG4-RD. At the last follow-up, the total response in groups I and II was 98.15 and 96.3%, respectively, and within 12 months, the cumulative relapse rate in group II was significantly higher than that in group I (14.8 vs. 3.7%, P = 0.046). Recurrence occurred at the paranasal sinus, lacrimal glands, skin, lung, pancreas, and bile ducts, and the relapsed patients achieved remission after switching immunosuppressants or/and increasing the GC dose. Conclusions: In IgG4-RD patients with internal organ involvement, GCs plus CYC or MMF are both effective with similar effects in disease response, while GCs plus CYC reduced the relapse rate better than GCs plus MMF.
Project description:BACKGROUND:This study aimed to investigate the contribution of a proliferation-inducing ligand (APRIL), a member of the tumor necrosis factor (TNF) superfamily implicated in plasma cell survival, to the development of plasma cell-rich lesions in immunoglobulin G4-related disease (IgG4-RD). METHODS:We performed immunohistochemical staining for APRIL with Stalk-1 and Aprily-8 antibodies specifically recognizing APRIL-producing cells and secreted APRIL, respectively, in renal and submandibular lesions of IgG4-RD in comparison with those of Sjögren's syndrome and sialolithiasis. RESULTS:Numerous Stalk-1-positive APRIL-producing cells were detectable in lesions of IgG4-RD. These cells, identified as CD163-positive M2 macrophages, secreted APRIL that distributed close to and even on infiltrating plasma cells. In contrast, APRIL-producing cells and the secreted form of APRIL were rarely detectable in lesions of Sjögren's syndrome or sialolithiasis. Notably, APRIL expression decreased concomitantly with the level of plasma cell infiltration after successful glucocorticoid treatment. CONCLUSIONS:Abundant infiltration into tissue lesions of APRIL-producing M2 macrophages and retention of secreted APRIL in plasma-cell-rich areas support a role for APRIL in the pathogenesis of plasma cell-rich lesions in IgG4-RD.