Model Of Sjögren's Syndrome Research Articles

Total glucosides of paeony alleviates experimental Sjögren's syndrome through inhibiting NLRP3 inflammasome activation of submandibular gland cells.

The mechanisms by which total glucosides of paeony (TGP) mitigates Sjögren's syndrome (SS) remains elusive. In the present study, we aim to explore the relationship between the therapeutic effects of TGP in the treatment of SS and NLRP3 inflammasome activation in submandibular gland (SG) cells. Female non-obese diabetic (NOD) mice were selected as the model of SS. The mice were divided into PBS and TGP treatment group. For treatment, TGP (400mg·kg-1) was administered intragastrically every day for 4 weeks. The SS-like symptoms and pathological changes of the SG of mice were compared between the PBS and TGP group. The activation of NLRP3 inflammasome in SG was detected by RT-qPCR, immunohistochemistry and western blot. The SG cells stimulated by lipopolysaccharide (LPS) and adenosine triphosphate (ATP) for activation of NLRP3 inflammasome were treated with or without TGP. Then, NLRP3 inflammasome activation was assessed. The IL-1β and IL-18 in homogenate of SG, serum and supernatant were detected by ELISA. Compared with balb/c mice, NOD mice showed SS-like symptoms and lymphocyte infiltration in SG, and the expression of NLRP3 inflammasome in SG was significantly increased. The SS-like symptoms were alleviated, and lymphocyte infiltration in SG was reduced, and the level of NLRP3 inflammasome in SG mice was decreased after TGP treatment. TGP also significantly inhibit the activation of NLRP3 inflammasome of SG cells in vitro. Collectively, our results indicated that TGP alleviates SS through inhibition of the activation of NLRP3 inflammasome of SG. These findings clarified the mechanism underlying the therapeutic effects of TGP on SS, and provided new evidence for the further application of TGP in the treatment of SS.

Rapamycin can alleviate the submandibular gland pathology of Sjögren's syndrome by limiting the activation of cGAS-STING signaling pathway.

Sjögren's Syndrome (SS) is also known as autoimmune exocrine gland disease. Previous studies have confirmed that adaptive immunity plays an important role in the development of this disease. But less is known about the role of the innate immune system. To identify the core pathways, and local infiltrated immune cells in the local immune microenvironment of SS. We verified the activation of these core genes and core signaling pathways in SS model mice by in vivo experiment and transcriptome sequencing. Finally, we identified 6 core genes EPSTI1, IFI44L, MX1, CXCL10, IFIT3, and IFI44. All the 6 genes had good diagnostic value. Based on multi-omics sequencing results and experimental studies, we found that cGAS-STING signaling pathway is most relevant to the pathogenesis of SS. By in vivo experiments, we verified that autophagy is the key brake to limit the activation of cGAS-STING signaling pathway. Maladaptive activation of autophagy and cGAS-STING signaling pathway are central contributors to the SG pathogenesis of pSS patient. Regulating autophagy by rapamycin may be a possible treatment for Sjögren's syndrome in the future.

Extracellular vesicles of iPS cells highly capable of producing HGF and TGF-β1 can attenuate Sjögren's syndrome via innate immunity regulation

Previous studies have demonstrated that extracellular vesicles (EVs) from dental pulp stem cells (DPSCs), which release abundant hepatocyte growth factor (HGF) and transforming growth factor-β1 (TGF-β1), contribute to the pathogenesis of Sjögren's syndrome (SS). However, depending on the condition of DPSCs, this effect is often not achieved. In this study, we established induced pluripotent stem (iPS) cells highly capable of releasing HGF and TGF-β1 and iPS cells barely capable of releasing them, and administered each EV to SS model mice to see if there was a difference in therapeutic effect. EVs were collected from each iPS cell and their characteristics and shapes were examined. When they were administered to SS model mice, the EVs from iPS cells with higher concentrations of HGF and TGF-β1 showed significantly reduced inflammatory cell infiltration in salivary gland tissues, increased saliva volume, and decreased anti-SS-A and anti-SS-B antibodies. A comprehensive search of microRNA arrays for differences among those EVs revealed that EVs from iPS cells with higher concentrations of HGF and TGF-β1 contained more of the let-7 family. Thereafter, we examined the expression of toll-like receptors (TLRs), which are said to be regulated by the let-7 family, by qPCR, and found decreased TLR4 expression. Focusing on MAPK, a downstream signaling pathway, we examined cytokine concentrations in mouse macrophage culture supernatants and Western blotting of murine splenic tissues and found higher concentrations of anti-inflammatory cytokines in the EVs-treated group and decreased TLR4, NF-κB and phosphorylation (p)-p-38 MAPK expression by Western blotting. Alternatively, p-Smad2/3 was upregulated in the EVs-treated group. Our findings suggest that the let-7 family in EVs may suppress the expression of TLR4 and NF-κB, which may be involved in the suppression of MAPK-mediated pro-inflammatory cytokine production.

Inhibition of CD4 + T cells by fanchinoline via miR506-3p/NFATc1 in Sjögren's syndrome.

The hyperproliferation and hyperactivation of CD4 + T cells in salivary gland tissues are hallmarks of Sjögren's syndrome (SS). Fangchinoline (Fan) is extracted from the root of Stephania tetrandra Moore, which is used for treating rheumatic diseases in many studies. This study aimed to identify the mechanism underlying the inhibition of CD4 + T cells by Fan in the SS model NOD/ShiLtj mice. In vivo, Fan alleviated the dry mouth and lymphocyte infiltration in the salivary gland tissues of the NOD/ShiLtj mice and inhibited the number of CD4 + T cells in the infiltrating focus. In vitro, Fan's inhibitory effect on the proliferation of mouse primary CD4 + T cells was verified by CFSE and EdU tests. Furthermore, qRT-PCR and WB analysis confirmed that Fan could inhibit the expression of NFATc1 (Nuclear factor of activated T-cells, cytoplasmic 1) by upregulating miR-506-3p. Dual luciferase reporter gene assay suggested that miR-506-3p interacted with NFATc1. CFSE and EdU tests showed that Fan could inhibit the proliferation of CD4 + T cells through miR-506-3p/NFATc1. The key role of NFATc1 in the activation of CD4 + T cells and the high expression of NFATc1 in samples from SS patients suggested that NFATc1 might become a therapeutic target for SS. In vivo, 11R-VIVIT (NFATc1 inhibitor) alleviated SS-like symptoms. This study not only explained the new mechanism of Fan inhibiting proliferation of CD4 + T cells and alleviating SS-like symptoms but also provided NFATc1 as a potential target for the subsequent research and treatment of SS.

Coupling annealed silver nanoparticles with a porous silicon Bragg mirror SERS substrate and machine learning for rapid non-invasive disease diagnosis.

Ag2O-Ag-porous silicon Bragg mirror (PSB) composite SERS substrates were successfully synthesized by using a combination of electrochemical and thermochemical methods. Test results showed that the SERS signal increased and decreased as the annealing temperature used for the substrate increased, where the most intense SERS signal was obtained using a substrate annealed at 300°C. Stability test results showed substantial enhancement of the SERS signal intensity of the Ag2O-Ag-PSB composite one month after preparation compared with that of conventional Ag-PSB. We conclude that Ag2O nanoshells play an essential role in SERS signal enhancement. Ag2O prevents natural oxidation of Ag nanoparticles (AgNPs) and has a solid localized surface plasmon resonance (LSPR). SERS signal enhancement was tested using this substrate for serum from patients with Sjögren's syndrome (SS) and Diabetic nephropathy (DN), as well as from healthy controls (HC). SERS feature extraction was performed using principal component analysis (PCA). The extracted features were analyzed by a support vector machine (SVM) algorithm. Finally, a rapid screening model for SS and HC, as well as DN and HC, was developed and used to perform controlled experiments. The results showed that the diagnostic accuracy, sensitivity and selectivity for SERS technology combined with machine learning algorithms reached 90.7%, 93.4% and 86.7% for SS/HC and 89.3%, 95.6% and 80% for DN/HC, respectively. The results of this study show that the composite substrate has excellent potential to be developed into a commercially available SERS chip for medical testing.

Depression-like behaviors in mouse model of Sjögren's syndrome: A role of gut–microbiota–brain axis

Depression is a common psychiatric symptom in patients with Sjögren's syndrome (SS). Prevalence of depression in SS patients is significantly higher than that in the general population. Increasing evidence suggests a crucial role of gut–microbiota–brain axis in depression. In this study, we investigated whether non-obese diabetic (NOD) mice, a widely used animal model of SS, exhibit depression-like phenotypes and abnormal composition of gut microbiota. Eleven-week-old NOD mice spontaneously exhibited SS-related symptoms without pancreatic destruction. NOD mice displayed depression-like behaviors, decreased expression of synaptic proteins in the prefrontal cortex (PFC), and abnormal composition of gut microbiota. Interestingly, SS-related proinflammatory factors in the submandibular gland (SMG) and autoantibodies (anti-SSA and anti-SSB) in the plasma were correlated with the expression of synaptic proteins in the PFC or depression-like behaviors. In addition, there were correlations between the relative abundance of microbiota and SS-related symptoms (or depression-related phenotypes). These data suggest that SS-related symptoms and abnormal composition of gut microbiota may play a role in depression-like phenotypes in NOD mice through gut–microbiota–brain axis.

Early Dry Eye Disease Onset in a NOD.H-2h4 Mouse Model of Sjögren's Syndrome.

PurposeTo develop a mouse model of human dry eye disease (DED) for investigation of sex differences in autoimmune-associated dry eye pathology.MethodsOcular surface disease was assessed by quantifying corneal epithelial damage with lissamine green stain in the NOD.H-2h4,IFNγ−/−,CD28−/− (NOD.H-2h4 DKO) mouse model of Sjögren's syndrome (SS). Lacrimal gland function was assessed by tear volume quantification with phenol red thread and lacrimal gland inflammation (i.e., dacryoadenitis) was assessed by quantification of immune cell foci, flow cytometric analysis of immune cell composition, and expression of proinflammatory markers.ResultsThe NOD.H-2h4 DKO mouse model of SS exhibits greater age-dependent increases in corneal damage than in NOD.H-2h4 parental mice and demonstrates an earlier disease onset in females compared to males. The severity of ocular surface disease correlates with loss of goblet cell density, increased conjunctivitis, and dacryoadenitis that is more pronounced in NOD.H-2h4 DKO than NOD.H-2h4 mice. B cells dominate lacrimal infiltrates in 16-week-old NOD.H-2h4 and NOD.H-2h4 DKO mice, but T helper cells and macrophages are also present. Lacrimal gland expression of proinflammatory genes, including the P2X7 and P2Y2 purinergic receptors, is greater in NOD.H-2h4 DKO than NOD.H-2h4 mice and correlates with dacryoadenitis.ConclusionsOur results demonstrate for the first time that autoimmune dry eye disease occurs in both sexes of NOD.H-2h4 DKO and NOD.H-2h4 mice, with earlier onset in female NOD.H-2h4 DKO mice when compared to males of the same strain. This study demonstrates that both NOD.H-2h4 and NOD.H-2h4 DKO mice are novel models that closely resemble SS-related and sex-dependent DED.

Salivary and Lacrimal Gland Alterations of the Epidermal Fatty Acid-Binding Protein (E-FABP) in Non-Obese Diabetic Mice.

The purpose of this study was to investigate the changes in E-FABP in the salivary and lacrimal glands of the Sjögren syndrome (SS) model non-obese diabetic mice (NOD). Cotton thread and ocular vital staining tests were performed on 10-week NOD male mice (n = 24) and age- and sex-matched wild-type (WT) mice (n = 25). Tear and saliva samples were collected at sacrifice for E-FABP ELISA assays. Salivary and lacrimal gland specimens underwent immunohistochemistry stainings for E-FABP. Real-time RT-PCR was also performed for the quantification of mRNA expression levels in the salivary and lacrimal glands. Corneal vital staining scores in the NOD mice were significantly higher compared with those for the wild-type mice (p = 0.0001). The mean tear E-FABP level showed a significantly lower concentration in the NOD mice (p = 0.001). The mean saliva E-FABP level also showed a significantly lower concentration in the NOD mice (p = 0.04). Immunohistochemistry revealed intense E-FABP staining in the LG acinar epithelium and less intense staining in the acinar epitheliae of the SGs in the NOD mice compared to the WT mice. Real-time RT-PCR for the mRNA expression of E-FABP showed a significantly decreased expression in the SG and a significant increase in the LG of the NOD mice compared to the WT mice. In conclusion, the E-FABP showed marked alterations in the tear film, saliva, lacrimal, and salivary glands of the NOD mouse, which may help explain the ocular surface changes in relation to the dry eye disease in this SS model mouse and keratoconjunctivitis sicca in SS patients.

Supra-lacrimal protein-based carriers for cyclosporine A reduce Th17-mediated autoimmunity in murine model of Sjögren's syndrome

Sjögren's syndrome (SS) is a multifactorial autoimmune disease with principal symptoms including inflammation and loss of function of lacrimal glands (LG) and salivary glands. While glandular infiltrates includes both B- and T-cells, CD4+ T cells are strongly implicated. Utilizing the male non-obese diabetic (NOD) mouse model of SS, this work: 1) identifies clinically-relevant elevations in cytokines (IL-17A, IL-2) in LG-derived CD4+ T cells; and 2) explores tissue-specific immunosuppression of SS using a novel protein-based drug carrier to concentrate cyclosporine A (CsA) directly in the LG. As a potent immunosuppressant, topical ophthalmic CsA is approved for dry eye disorders; however, it cannot effectively resolve inflammation due to limited accumulation in the LG. Systemic CsA has dose-limiting side effects that also limit its ability to block LG inflammation. Using elastin-like polypeptides (ELPs) fused genetically to cyclophilin, the intracellular cognate receptor of CsA, this manuscript reports a sustained-release formulation of CsA that maintains therapeutic drug concentrations in the LG and extends intervals between doses. This formulation blocked both in vitro Th17 cell differentiation and IL-17A secretion. In vivo treatment significantly decreased the abundance of Th17.1 cells, a helper cell population sharing phenotypes of both Th17 and Th1, in the LG of diseased NOD mice. Treatment with even a single dose of the sustained-release formulation was effective enough to improve basal levels of tear production. Thus, this sustained-release formulation suppressed local LG inflammation driven through IL-17 dependent pathways, while improving ocular surface function.

Short-chain fatty acid butyrate induces IL-10-producing B cells by regulating circadian-clock-related genes to ameliorate Sjögren's syndrome.

Sjögren's syndrome (SS) is an autoimmune disease caused by inflammation of the exocrine gland. The pathological hallmark of SS is the infiltration of lymphocytes into the salivary glands. Increased infiltration of T and B cells into salivary glands exacerbates symptoms of SS. Several recent studies have identified the role of gut microbiota in SS. Butyrate, one of the metabolites of the gut microbiota, regulates T cells; however, its effects on B cells and SS remain unknown. This study determined the therapeutic effect of butyrate on regulating B cells in SS. Various concentrations of butyrate were intraperitoneally injected three times per week in NOD/ShiLtJ (NOD) mice, the prototype animal model for SS, and observed for more than 10 weeks. Whole salivary flow rate and the histopathology of salivary glands were investigated. Human submandibular gland (HSG) cells and B cells in mouse spleen were used to confirm the anti-inflammatory and immunomodulatory effects of butyrate. Butyrate increased salivary flow rate in NOD mice and reduced inflammation of salivary gland tissues. It also regulated cell death and the expression of circadian-clock-related genes in HSG cells. Butyrate induced B cell regulation by increasing IL-10-producing B (B10) cells and decreasing IL-17-producing B cells, through the circadian clock genes RAR-related orphan receptor alpha and nuclear receptor subfamily 1 group D member 1. The findings of this study imply that butyrate may ameliorate SS via reciprocal regulation of IL-10- and IL-17-producing B cells.

Triptolide reduces salivary gland damage in a non-obese diabetic mice model of Sjögren's syndrome via JAK/STAT and NF-κB signaling pathways.

Triptolide (TP) has anti-inflammatory and immunosuppressive effects. However, the effect of triptolide on Sjögren's syndrome (SS) is rarely reported. In this paper, we studied the effects of triptolide on non-obese diabetes mice model of SS. In this study, salivary flow rate was measured every two weeks, and autoantibodies levels in the serum were detected. Salivary gland index and spleen index were detected, pathological changes of salivary gland were detected by hematoxylin-eosin staining, inflammatory factors were detected by enzyme linked immunosorbent assay, lymphocytes were detected by flow cytometry, proliferation of T cells and B cells were detected, and related proteins were detected by Western blot. Triptolide increased salivary flow rate and salivary gland index, and decreased spleen gland index. Moreover, triptolide reduced the infiltration of lymphocytes to salivary glands, decreased the level of autoantibodies in serum, and reduced the inflammatory factors in salivary glands and IFN-γ induced salivary gland epithelial cells. Further, triptolide inhibited activator of JAK/STAT pathway and NF-κB pathway. In conclusion, triptolide could inhibit the infiltration of lymphocytes and the expression of inflammatory factors through JAK/STAT pathway and NF-κB pathway. Thus, triptolide may be used as a potential drug to treat SS.

The NZB/WF1 mouse model for Sjögren's syndrome: A historical perspective and lessons learned.

Sjögren's syndrome (SS) is a chronic rheumatic autoimmune disorder affecting multiple organ systems. The clinical findings in SS patients show considerable heterogeneity and overlap with other autoimmune diseases. In addition, the autoimmune response in SS initiates several years before the appearance of clinical symptoms. Thus, understanding the pathogenic mechanisms involved in the disease process have been a challenge. Several animal model systems of SS-like disease have been developed to overcome these issues. The New Zealand Black (NZB) x New Zealand White (NZW) F1 (NZB/WF1) mouse represents the first spontaneous mouse model of SS. In this review, we provide a historical perspective and detailed description of this mouse model focusing on exocrine gland histopathology, autoantibody populations, and glandular dysfunction. Considering that NZB/WF1 mice also develop a systemic lupus erythematosus (SLE)-like disease, this mouse model mimics the clinical presentation of polyautoimmunity seen in a sizable subset of SS patients. It is plausible that such patients will require distinct therapeutic interventions necessary to treat both SLE and SS. Therefore, the NZB/WF1 mouse is a powerful tool to decipher pathogenic mechanisms involved in SS related polyautoimmunity and develop appropriate therapeutic strategies.

Small RNA Deep Sequencing Identifies a Unique miRNA Signature Released in Serum Exosomes in a Mouse Model of Sjögren's Syndrome.

Sjögren's Syndrome (SS) is an autoimmune disease characterized by lymphocytic infiltration and loss of function of moisture-producing exocrine glands as well as systemic inflammation. SS diagnosis is cumbersome, subjective and complicated by manifestation of symptoms that overlap with those of other rheumatic and ocular diseases. Definitive diagnosis averages 4–5 years and this delay may lead to irreversible tissue damage. Thus, there is an urgent need for diagnostic biomarkers for earlier detection of SS. Extracellular vesicles called exosomes carry functional small non-coding RNAs which play a critical role in maintaining cellular homeostasis via transcriptional and translational regulation of mRNA. Alterations in levels of specific exosomal miRNAs may be predictive of disease status. Here, we have assessed serum exosomal RNA using next generation sequencing in a discovery cohort of the NOD mouse, a model of early-intermediate SS, to identify dysregulated miRNAs that may be indicative of SS. We found five miRNAs upregulated in serum exosomes of NOD mice with an adjusted p < 0.05—miRNA-127-3p, miRNA-409-3p, miRNA-410-3p, miRNA-541-5p, and miRNA-540-5p. miRNAs 127-3p and 541-5p were also statistically significantly upregulated in a validation cohort of NOD mice. Pathway analysis and existing literature indicates that differential expression of these miRNAs may dysregulate pathways involved in inflammation. Future studies will apply these findings in a human cohort to understand how they are correlated with manifestations of SS as well as understanding their functional role in systemic autoimmunity specific to SS.

Mechanism of cAMP-PKA Signaling Pathway Mediated by Shaoyao Gancao Decoction () on Regulation of Aquaporin 5 and Muscarinic Receptor 3 Levels in Sjögren's Syndrome.

To investigate the mechanism of cAMP-PKA signaling pathway mediated by Chinese medicine formula Shaoyao Gancao Decoction (, SGD) on the regulation of aquaporin 5 (AQP5) and muscarinic receptor 3 (M3R) levels in Sjögren's syndrome (SS). Of the 30 mice, 5 were randomly selected as control, and others were used for creating SS model. After successful modeling, mice were randomly divided into 5 groups (n=5 per group) and intragastrically administered with saline (8 mL/kg), pilocarpine (1.4 mg/kg), or low, medium and high doses SGD (0.14, 0.21, 0.35 g/kg Radix paeoniae with 0.01 g/kg Radix glycyrrhizae, respectively) for 6 weeks. Human labial gland acinar cells were treated with pilocarpine or varying doses of SGD with saline as the placebo. Hematoxylin and eosin staining was used to observe the histopathological changes of the submandibular glands of mice. The serum levels of anti-SS antigen A (SS-A), anti-SS antigen B (SS-B), M3R, and α-fodrin in submandibular glands of mice were measured by enzyme-linked immunosorbent assay. Immunofluorescence staining was used to observe the spatial localization of AQP5 and M3R in acinar cells. Reverse transcriptase polymerase chain reaction and Western blot were used to detect the expressions of PKA, cAMP, Epac1, AQP5, M3R, nuclear factor kappa-B (NF-κB), and tumor necrosis factor (TNF)-α in submandibular gland tissues and cells of each group. Compared to normal mice, body weight, 5-min salivary secretion, 30-min secretion of tears and breakup time of tear film of model mice decreased at 1-6 weeks after immunization (all P<0.05), whereas water intake increased (all P<0.05). In the model group, glands of the submandibular glands showed atrophy, accompanied by acini of different sizes, decreased numbers and loose arrangement, with catheter dilatation and different degrees of lymphocyte infiltration. Conditions of mice in SGD groups were improved. The positive expression of AQP5 and M3R were higher in the acinar cells treated with all doses SGD compared to the normal group; serum levels of SS-A, SS-B, and α-fodrin were lower, and that of M3R was higher in all doses SGD treated animals than the model or pilocarpine treated ones (all P<0.05). Compared to the model and pilocarpine groups, the mRNA and protein levels of NF-κB and TNF-α were lower in mice or cells treated with medium or high-dose SGD (all P<0.05), while those of PKA, Epac1, AQP5 and M3R were higher (all P<0.05). SGD can improve symptoms of SS by regulating the cAMP-PKA signaling pathway and increasing AQP5 and M3R levels.

Achaete-Scute Homologue 2–Regulated Follicular Helper T Cells Promote Autoimmunity in a Murine Model for Sjögren Syndrome

Follicular helper T (Tfh) cells contribute to various immune responses as well as to the pathogenesis of several immune diseases. However, the precise mechanism underlying the onset or development of autoimmunity via Tfh cells remains unclear. Herein, the detailed relationship between autoimmune disease and Tfh cells was analyzed using a murine model for Sjögren syndrome (SS) wherein the mice underwent neonatal thymectomy. Germinal center (GC) development was promoted in this SS model along with an increase of Tfh cells and GC B cells. The severity of the autoimmune lesions was correlated with the number of Tfh cells detected in the spleen of the SS model mice. In addition, treatment with an anti-CD20 monoclonal antibody effectively suppressed the autoimmune lesions with a reduction of Tfh cells and GC B cells. Comprehensive gene analysis revealed that several genes associated with Tfh cell differentiation, including achaete-scute homologue 2 (Ascl2), were up-regulated in peripheral CD25- CD4+ T cells in SS model mice compared with those in control mice. Moreover, an experiment using CD4CreBcl6fl/fl mice that received neonatal thymectomy treatment demonstrated that Ascl2 contributes to the Tfh cell differentiation associated with autoimmunity during the early stages, independent of Bcl6. In conclusion, our results indicate that abnormal Tfh cell differentiation via Ascl2 regulation might contribute to the pathogenesis of autoimmunity.

Spontaneous Model of Sjögren's Syndrome in NOD Mice.

The non-obese diabetic (NOD) mouse model is the most widely described and validated method for investigating human primary Sjögren's syndrome (SS) and represents a useful model for translational studies. However, the systemic disease manifestation in NOD mice is sensitive to the housing environment, as stress modulates the immune system, so it is essential to confirm that readouts are robust, reproducible, and sensitive to known clinical treatments. This protocol describes the establishment of the spontaneous NOD model of SS and underscores the necessity of model validation to ensure that the housing environment is compatible. © 2019 by John Wiley & Sons, Inc.

Stem cells from exfoliated deciduous teeth alleviate hyposalivation caused by Sjögren syndrome.

To evaluate the effect of stem cells from exfoliated deciduous teeth on the hyposalivation caused by Sjögren syndrome (SS) and investigate the mechanism. Stem cells were injected into the tail veins of non-obese diabetic mice, the animal model of SS. The saliva flow was measured after pilocarpine intraperitoneal injection. Apoptosis and autophagy were evaluated by TUNEL and Western blot. Lymphocyte proportions were detected by flow cytometer. Fluid secretion was decreased in 21-week-old mice. Stem cell treatment increased fluid secretion, alleviated inflammation in the submandibular glands and reduced inflammatory cytokine levels in the serum, submandibular glands and saliva. Stem cells decreased the apoptotic cell number and the expressions of ATG5 and Beclin-1 in the submandibular glands. Stem cells have no effect on other organs. Furthermore, the infused stem cells migrated to the spleen and liver, not the submandibular gland. Stem cells directed T cells towards Treg cells and suppressed Th1 and Tfh cells in spleen lymphocytes. Stem cells from exfoliated deciduous teeth alleviate the hyposalivation caused by SS via decreasing the inflammatory cytokines, regulating the inflammatory microenvironment and decreasing the apoptosis and autophagy. The stem cells regulated in T-cell differentiation are involved in the immunomodulatory effects.

Ablation of the Chaperone Protein ERdj5 Results in a Sjögren's Syndrome-Like Phenotype in Mice, Consistent With an Upregulated Unfolded Protein Response in Human Patients.

Objective: Sjögren's syndrome (SS) is a chronic autoimmune disorder that affects mainly the exocrine glands. Endoplasmic reticulum (ER) stress proteins have been suggested to participate in autoimmune and inflammatory responses, either acting as autoantigens, or by modulating factors of inflammation. The chaperone protein ERdj5 is an ER-resident disulfide reductase, required for the translocation of misfolded proteins during ER-associated protein degradation. In this study we investigated the role of ERdj5 in the salivary glands (SGs), in association with inflammation and autoimmunity.Methods: In situ expression of ERdj5 and XBP1 activation were studied immunohistochemically in minor SG tissues from primary SS patients and non-SS sicca-complaining controls. We used the mouse model of ERdj5 ablation and characterized its features: Histopathological, serological (antinuclear antibodies and cytokine levels), and functional (saliva flow rate).Results: ERdj5 was highly expressed in the minor SGs of SS patients, with stain intensity correlated to inflammatory lesion severity and anti-SSA/Ro positivity. Moreover, SS patients demonstrated higher XBP1 activation within the SGs. Remarkably, ablation of ERdj5 in mice conveyed many of the cardinal features of SS, like spontaneous inflammation in SGs with infiltrating T and B lymphocytes, distinct cytokine signature, excessive cell death, reduced saliva flow, and production of anti-SSA/Ro and anti-SSB/La autoantibodies. Notably, these features were more pronounced in female mice.Conclusions: Our findings suggest a critical connection between the function of the ER chaperone protein ERdj5 and autoimmune inflammatory responses in the SGs and provide evidence for a new, potent animal model of SS.

Tissue-Resident Memory CD8+ T Cells Acting as Mediatorsof Salivary Gland Damage in a Murine Model of Sjögren's Syndrome.

Although a role for CD4+ T cells in the pathogenesis of Sjögren's syndrome (SS) has been documented, the pathogenic significance of CD8+ T cells is unclear. The aim of this study was to investigate the role of CD8+ T cells in the development of SS. Flow cytometry and immunofluorescence analyses were utilized to detect T cell infiltration within the labial salivary glands of patients with primary SS. In parallel, p40-/- CD25-/- mice were used as a murine model of SS. In addition, mice with genetic knockout of CD4, CD8a, or interferon-γ (IFNγ) were crossed with p40-/- CD25-/- mice to study the pathogenic significance of specific lineage subpopulations, including functional salivary gland tests as well as histopathologic and serologic data. A CD8+ T cell-specific depletion antibody was used in this murine SS model to evaluate its potential as a therapeutic strategy. CD8+ T cells with a tissue-resident memory phenotype outnumbered CD4+ T cells in the labial salivary glands of patients with SS, and were primarily colocalized with salivary duct epithelial cells and acinar cells. Furthermore, infiltrating CD8+ T cells with a CD69+CD103+/- tissue-resident phenotype and with a significant elevation of IFNγ production were dominant in the submandibular glands of mice in this murine SS model. CD8a knockout abrogated the development of SS in these mice. Knockout of IFNγ decreased CD8+ T cell infiltration and gland destruction. More importantly, depletion of CD8+ T cells fully protected mice against the pathologic manifestations of SS, even after the onset of disease. These data reveal the pathogenic significance of CD8+ T cells in the development and progression of SS in the salivary glands. Treatment directed against CD8+ T cells may be a rational therapy for the management of SS in human subjects.

Reduced Corneal Innervation in the CD25 Null Model of Sjögren Syndrome.

Decreased corneal innervation is frequent in patients with Sjögren Syndrome (SS). To investigate the density and morphology of the intraepithelial corneal nerves (ICNs), corneal sensitivity, epithelial cell proliferation, and changes in mRNA expression of genes that are involved in autophagy and axon targeting and extension were assessed using the IL-2 receptor alpha chain (CD25 null) model of SS. ICN density and thickness in male and female wt and CD25 null corneas were assessed at 4, 6, 8, and 10/11 wk of age. Cell proliferation was assessed using ki67. Mechanical corneal sensitivity was measured. Quantitative PCR was performed to quantify expression of beclin 1, LC3, Lamp-1, Lamp-2, CXCL-1, BDNF, NTN1, DCC, Unc5b1, Efna4, Efna5, Rgma, and p21 in corneal epithelial mRNA. A significant reduction in corneal axon density and mechanical sensitivity were observed, which negatively correlate with epithelial cell proliferation. CD25 null mice have increased expression of genes regulating autophagy (beclin-1, LC3, LAMP-1, LAMP-2, CXCL1, and BDNF) and no change was observed in genes that were related to axonal targeting and extension. Decreased anatomic corneal innervation in the CD25 null SS model is accompanied by reduced corneal sensitivity, increased corneal epithelial cell proliferation, and increased expression of genes regulating phagocytosis and autophagy.