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BACKGROUND: Autoimmune regulator gene (Aire) and Wnt signaling pathway play an important role in the maintenance and differentiation of mouse embryonic stem cell pluripotency. However, whether the Wnt signal and Aire are involved in the differentiation of embryonic stem cells to thymic epithelial progenitor cells remains poorly understood. OBJECTIVE: To investigate the relationship of the Wnt signaling pathway and Aire with the differentiation of embryonic stem cells. METHODS: A two-step differentiation method was used to induce mouse embryonic stem cells to differentiate into endoderm and then into thymic epithelial progenitor cells. Mouse embryonic stem cells were infected with Aire shRNA lentivirus, and monoclonal stable strains were screened by puromycin. Mouse embryonic stem cells were collected on days 0, 3 and 10 of the directed induction of differentiation after the induced differentiation by the two-step differentiation method. Cellular immunofluorescence, flow cytometry, western blot assay, and real-time qPCR were used to detect the expression changes of related genes and proteins. RESULTS AND CONCLUSION: (1) Immunofluorescence staining showed positive expression of SSEA1 and OCT4 on day 0 of targeted induction of differentiation. (2) Immunofluorescence staining showed double-positive expression of SOX17 and FOXA2 on day 3 of targeted induction of differentiation. (3) Flow cytometry results showed positive expression of EPCAM1, K5 and K8 on day 10 of targeted induction of differentiation. (4) Compared with undifferentiated mouse embryonic stem cells, the expressions of Wnt7a, β-catenin, and Gsk-3β proteins were elevated, and the expression level of Aire protein was decreased in induced differentiated thymic epithelial progenitor cells. (5) Compared with undifferentiated mouse embryonic stem cells, the expressions of Wnt7a, β-catenin, Gsk-3β and Aire mRNA were elevated in thymic epithelial progenitor cells. (6) Compared with normal cultured mouse embryonic stem cells and their ultimately differentiated thymic epithelial progenitor cells, the expression levels of Wnt7a, β-catenin and Gsk-3β proteins were reduced in mouse embryonic stem cells with knockdown of Aire genes and their final differentiated thymic epithelial progenitor cells. In conclusion, the Wnt signaling pathway and Aire are jointly involved in the process of targeted induction of differentiation of mouse embryonic stem cells into mouse thymic epithelial progenitor cells. [ABSTRACT FROM AUTHOR]

Background/Aim: Autoimmune regulator (AIRE) is a transcription factor that plays pivotal role in controlling autoimmunity. In the thymus, it supports the presentation of peripheral tissue antigens to developing T cells, where recognition of these self-antigens negatively selects the autoimmune naïve T-cells by central tolerance. Studies demonstrated that single-nucleotide polymorphisms (SNPs) in AIRE alter transcription and propagate clonal survival of autoimmune T cells, therefore increase susceptibility to autoimmune diseases. This study intended to identify SNPs in exon and intron sequences that determine AIRE transcription, where their genotypes are associated with rheumatoid arthritis (RA) risk and clinical parameters. Patients and Methods: After a thorough in silico research, we enrolled 100 patients with RA and 100 healthy controls to analyze the association of SNP rs870881(C>T) and rs1003854(T>C) in AIRE coding sequence with RA risk by using five different genetic models and selected clinical parameters. Multiplex quantitative polymerase chain reaction was used to determine allelic discrimination of SNPs. RA risk was assessed by odds ratios (ORs) and confidence intervals (CIs). Results: In a recessive model of rs878081, minor allele TT homozygotes were associated with RA (p=0.032, OR=5.44, 95%CI=1.16-25.52); in a recessive model of rs1003854, minor allele CC homozygotes were associated with RA (p=0.047, OR=4.84, 95%CI=1.02-23.02). Higher C-reactive protein (CRP) levels in patients with RA were significantly associated with minor allele homozygotes in recessive and codominant genetic models (p=0.029 and p=0.043, respectively) of rs1003854. Conclusion: Genotypes for minor alleles of rs878081 and rs1003854 might be involved in RA pathogenesis and risk prediction. [ABSTRACT FROM AUTHOR]

Autoimmune polyendocrine syndrome type-1 (APS1) is characterized by autoimmune manifestations affecting different organs from early childhood on. Immunological abnormalities, the resulting endocrinopathies, and their treatments may compromise bone health. For the first time in APS1, we analyzed transiliac bone biopsy samples by bone histomorphometry and quantitative backscattered electron imaging in three adult patients (female P1, 38 years; male P2, 47 years; male P3, 25 years). All had biallelic mutations in the autoimmune regulator gene and in addition to endocrinopathies, also significant bone fragility. Histomorphometry showed bone volume in the lower normal range for P1 (BV/TV, − 0.98 SD) and P3 (− 1.34 SD), mainly due to reduced trabecular thickness (TbTh, − 3.63 and − 2.87 SD). In P1, osteoid surface was low (OS/BS, − 0.96 SD); active osteoblasts and double labeling were seen only on cortical bone. P3 showed a largely increased bone turnover rate (BFR/BV, + 4.53 SD) and increased mineralization lag time (Mlt, + 3.40 SD). Increased osteoid surface (OS/BS, + 2.03 and + 4.71 SD for P2 and P3) together with a large proportion of lowly mineralized bone area (Trab CaLow, + 2.22 and + 9.81 SD for P2 and P3) and focal mineralization defects were consistent with abnormal mineralization. In all patients, the density and area of osteocyte lacunae in cortical and trabecular bone were similar to healthy adults. The bone tissue characteristics were variable and included decreased trabecular thickness, increased amount of osteoid, and abnormal mineralization which are likely to contribute to bone fragility in patients with APS1. [ABSTRACT FROM AUTHOR]

A genetic variant in the SAND domain of autoimmune regulator (AIRE), R247C, was identified in a patient with type I diabetes mellitus (T1DM) and his mother with rheumatoid arthritis. In vitro, the variant dominantly inhibited AIRE; however, typical features of Autoimmune Polyendocrinopathy Candidiasis and Ectodermal Dysplasia (APECED) were not seen in the subjects. Rather, early manifestation of autoimmunity appeared to be dependent on additional genetic factors. On a population level, diverse variants were identified in this region. Surprisingly, many likely pathogenic variants were seen disproportionately in Africans when compared to Europeans, reinforcing the importance of these variants in altering the immune repertoire. In light of these findings, we propose that R247C and other variants within the SAND-domain alter protein function in a dominant fashion and hold potential as drivers of autoimmunity.

Thymically-derived Foxp3+ regulatory T cells (Treg) critically control immunological tolerance. These cells are generated in the medulla through high affinity interactions with medullary thymic epithelial cells (mTEC) expressing the Autoimmune regulator (Aire). Recent advances have revealed that thymic Treg contain not only developing but also recirculating cells from the periphery. Although Aire is implicated in the generation of Foxp3+ Treg, its role in the biology of recirculating Treg remains elusive. Here, we show that Aire regulates the suppressive signature of recirculating Treg independently of the remodeling of the medullary 3D organization throughout life where Treg reside. Accordingly, the adoptive transfer of peripheral Foxp3+ Treg in AireKO recipients led to an impaired suppressive signature upon their entry into the thymus. Furthermore, recirculating Treg from AireKO mice failed to attenuate the severity of multiorgan autoimmunity, demonstrating that their suppressive function is altered. Using bone marrow chimeras, we reveal that mTEC-specific expression of Aire controls the suppressive signature of recirculating Treg. Finally, mature mTEC lacking Aire were inefficient in stimulating peripheral Treg both in polyclonal and antigen-specific co-culture assays. Overall, this study demonstrates that Aire confers to mTEC the ability to restimulate recirculating Treg, unravelling a novel function for this master regulator in Treg biology. [ABSTRACT FROM AUTHOR]

Atherosclerosis is a chronic, lipid-driven disease of medium sized arteries which causes myocardial infarction and stroke. Recently, an adaptive immune response against the plaque-associated autoantigen Apolipoprotein B100 (ApoB), the structural protein component of low-density lipoprotein, has been implicated in atherogenesis. In healthy individuals, CD4+ T cells responding to ApoB mainly comprised regulatory T cells, which confer immune tolerance and atheroprotection. Mice and patients with atherosclerosis harbor increased numbers of proatherogenic ApoB-reactive T-helper cell subsets. Given the lack of therapies targeting proatherogenic immunity, clarification of the underlying mechanisms is of high clinical relevance. T cells develop in the thymus, where strong autoreactive T cells are eliminated in the process of negative selection. Herein, we investigated whether the transcription factor autoimmune regulator (AIRE), which controls expression of numerous tissue-restricted self-antigens in the thymus, is involved in mediating tolerance to ApoB and whether Aire deficiency might contribute to atherogenesis. Mice deficient for Aire were crossbred to apolipoprotein E-deficient mice to obtain atherosclerosis-prone Aire−/−Apoe−/− mice, which were fed a regular chow diet (CD) or western-type diet (WD). CD4+ T cells responding to the ApoB peptide p6 were analyzed by flow cytometry. We demonstrate that Aire deficiency influences neither generation nor activation of ApoB-reactive T cells and has only minor and overall inconsistent impacts on their phenotype. Furthermore, we show that atherosclerotic plaque size is not affected in Aire−/−Apoe−/− compared to Aire+/+Apoe−/−, irrespective of diet and gender. In conclusion, our data suggests that AIRE is not involved in regulating thymic expression of ApoB or atherosclerosis. Alternative mechanisms how ApoB-reactive CD4 T cells are selected in the thymus will have to be investigated.

Coexistence of bullous pemphigoid (BP) and vitiligo vulgaris (VV) is very rare. We present a unique case of BP associated with VV in a 76-year-old Japanese man, in which BP eruption developed exclusively on preexisting VV regions. The patient was referred to us with a 3-month history of blistering eruption with severe pruritis on the right forearm and left lower limb. In addition, he had been suffering from a widespread depigmented eruption on the trunk and extremities for at least 20 years. Curiously, the blistering eruption developed exclusively on preexisting depigmented eruption. Histopathological and laboratory examinations identified the blistering eruption as BP. The depigmented eruption was diagnosed as VV. BP eruption responded quickly to oral prednisolone therapy, improving within 1 week, and the prednisolone dose was gradually tapered. Within 1 month, BP lesions almost completely resolved. In the present case, BP not only coexisted with VV, but also developed exclusively on preexisting vitiliginous regions. The present case strongly suggests that BP and VV are partly caused by common pathological mechanisms.

Senescence-related decline of thymus affects immune function in the elderly population and contributes to the prevalence of many relevant diseases like cancer, autoimmune diseases, and other chronic diseases. In this study, we investigated the therapeutic effects of curcumin, an agent that could counter aging, and explored its optimal intake and the alteration of autoimmune regulator (Aire) after curcumin treatment in the D-galactose (D-gal)-induced accelerated aging mice. ICR mice were intraperitoneally injected with D-gal for 8 weeks to establish the accelerated aging model and given curcumin with 50, 100, and 200 mg/kg body weight per day by gavage, respectively, for 6 weeks. It indicated that the D-gal-treated mice developed structural changes in the thymi compared with the control group without D-gal and curcumin treatment. As the supplements of curcumin, it resulted in a restoration of the normal thymic anatomy with an increase of proliferating cells and a reduction of apoptotic cells in the thymi of the D-gal-induced aging model mice. Curcumin administration could also expand the expression level of Aire from mRNA level and protein level. The current study demonstrated that curcumin could ameliorate senescence-related thymus involution via upregulating Aire expression, suggesting that curcumin can rejuvenate senescence-associated alterations of thymus induced by D-gal accumulation. [ABSTRACT FROM AUTHOR]

Autoimmune regulator+ (Aire) medullary thymic epithelial cells (mTECs) play a critical role in tolerance induction. Several studies demonstrated that Aire+mTECs differentiate further into Post‐Aire cells. Yet, the identification of terminal stages of mTEC maturation depends on unique fate‐mapping mouse models. Herein, we resolve this limitation by segmenting the mTEChi(MHCIIhiCD80hi) compartment into mTECA/hi (CD24−Sca1−), mTECB/hi (CD24+Sca1−), and mTECC/hi (CD24+Sca1+). While mTECA/hi included mostly Aire‐expressing cells, mTECB/hi contained Aire+ and Aire− cells and mTECC/hi were mainly composed of cells lacking Aire. The differential expression pattern of Aire led us to investigate the precursor‐product relationship between these subsets. Strikingly, transcriptomic analysis of mTECA/hi, mTECB/hi, and mTECC/hi sequentially mirrored the specific genetic program of Early‐, Late‐ and Post‐Aire mTECs. Corroborating their Post‐Aire nature, mTECC/hi downregulated the expression of tissue‐restricted antigens, acquired traits of differentiated keratinocytes, and were absent in Aire‐deficient mice. Collectively, our findings reveal a new and simple blueprint to survey late stages of mTEC differentiation. [ABSTRACT FROM AUTHOR]

As a transcription factor, autoimmune regulator (Aire) participates in thymic negative selection and maintains immune tolerance mainly by regulating the ectopic expression of tissue-restricted antigens (TRAs) in medullary thymic epithelial cells (mTECs). Aire is also expressed in dendritic cells (DCs). DCs are professional antigen-presenting cells (APCs) that affect the differentiation of T cells toward distinct subpopulations and participate in the immune response and tolerance, thereby playing an important role in maintaining homeostasis. To determine the role of Aire in maintaining immune tolerance by bone marrow-derived dendritic cells (BMDCs), in the present study we utilized Aire-knockout mice to examine the changes of maturation status and TRAs expression on BMDCs, additionally investigate the differentiation of CD4+ T cells. The results showed that expression of costimulatory molecule and major histocompatibility complex class II (MHC-II) molecule was increased and expression of various TRAs was decreased in BMDCs from Aire-knockout mice. Aire deficiency reduced the differentiation of naïve CD4+ T cells into type 2T helper (Th2) cells and regulatory T cells (Tregs) but enhanced the differentiation of naïve CD4+ T cells into Th1 cells, Th17 cells, and follicular helper T (Tfh) cells. The results demonstrate that Aire expressed by BMDCs plays an important role in the maintenance of homeostasis by regulating TRA expression and the differentiation of T cell subsets.

BACKGROUND: Autoimmune diseases are a class of diseases that cause a strong immune response to the continuous lack of self-tissue-specific antigens in the thymus. Hypothyroidism and unstable expression of tissue-specific antigens in the thymus can limit the therapeutic effect. The thymus is mainly composed of thymic epithelial cells, but the limited number of mature thymic epithelial cells and thymic epithelial progenitor cells in the thymus has greatly limited related research. OBJECTIVE: To detect the expression of autoimmune regulator (AIRE) when mouse embryonic stem cells were transformed into thymic epithelial progenitor cells. METHODS: A two-step differentiation method was used to induce the differentiation of mouse embryonic stem cells into endoderm and then into thymic epithelial progenitor cells. The cells were collected at 0, 3, and 13 days of induced differentiation. Immunofluorescence, flow cytometry, western blot and real-time PCR were used to detect the expression of cell-associated genes and proteins. RESULTS AND CONCLUSION: Positive expression of OCT4 and SSEA1 was detected by immunofluorescence at 0 day of induction. The double positive expression of SOX17 and FoxA2 was measured by immunofluorescence at 3 days of induction. The positive expression of EpCAM, K5 and K8 were analyzed by flow cytometry at 13 days of induction. During the directional differentiation of mouse embryonic stem cells, real-time PCR indicated that the expression of PAX1, PAX9, FOXN1 and PLET1 showed an increasing trend. The expression of AIRE gene increased significantly at 0, 3, and 13 days of induction. At the same time, the expression of INS2 gene and GAD67 gene also increased. Western blot assay showed that the expression of AIRE protein gradually decreased at 0, 3, and 13 days of induction; however, insulin protein and GAD67 protein were not detected. Overall findings indicate that mouse embryonic stem cells can successfully differentiate into thymic epithelial progenitor cells with highly expressed AIRE gene, which promotes the expression of INS2 and GAD67 genes, and provides an evaluation basis for cell transplantation in the treatment of autoimmune diseases. [ABSTRACT FROM AUTHOR]

Autoimmune polyendocrine syndrome type 1 (APS-1) is a rare inherited autoimmune disease, characterized by a classic triad, including chronic mucocutaneous candidiasis, primary adrenocortical insufficiency and hypoparathyroidism. The present study investigated phenotypes and pathogenic variants in a Chinese woman with non-classical APS-1. Disease-associated variants in a patient with APS-1 were identified via targeted next generation sequencing and the variant was confirmed via Sanger sequencing. Serum levels of calcium, phosphorus, parathyroid hormone (PTH), follicle-stimulating hormone (FSH), luteinizing hormone (LH), estradiol and urinary levels of calcium were measured. Blood count assays and bone marrow morphology were investigated. The patient was a 32-year-old woman who had suffered from typical carpopedal spasms since she was 7 years old. She developed syncope, primary amenorrhea, intermittent diarrhea and general fatigue in subsequent years. Hypocalcemia, hyperphosphatemia, low levels of PTH and estradiol, elevated levels of FSH and LH, and absence of erythroblasts were observed, which indicated hypoparathyroidism, primary ovarian insufficiency and pure red cell aplasia. A novel heterozygous missense variant (NM_000383.2: c.623G>T, NP_000374.1: p.Gly208Val) in exon 5 of autoimmune regulator and a reported variant (NM_000383.2: c.371C>T, NP_000374.1: p.Pro124Leu) in exon 3 were detected, of which the c.623G>T variant may be a pathogenic variation that induces APS-1. Under a regular follow-up and therapeutic adjustment of calcium, calcitriol, hormone replacement therapy and methylprednisolone, the endocrine function and clinical symptoms of the patient were notably improved. The results of the present study expand the known genetic and phenotypical spectra of APS-1. [ABSTRACT FROM AUTHOR]

Alopecia areata (AA) is a non-scarring hair loss of autoimmune etiology. The autoimmune regulator (AIRE) gene is believed to be an important driver in AA pathogenesis. Genetic variants can alter mRNA expression levels which may provoke an autoimmune response. A total of 337 males (97 AA patients and 240 controls) were enrolled in the current case–control study. On screening of the most frequent variants in the gene, rs2075876 (A/G) polymorphism in intron 5 was selected and genotyped using Real-Time PCR (polymerase chain reaction) technology. Additionally, circulatory AIRE expression levels were quantified by quantitative reverse-transcription PCR (qRT-PCR). Allelic discrimination analysis revealed GG genotype to be more frequent in patients (90.7% in AA compared to 32.5% in controls, p <.001). G variant conferred increased risk to alopecia under homozygote comparison (GG versus AA: OR = 16.1, 95%CI = 5.57–46.3), dominant model (GG+AG versus AA: OR = 7.24, 95%CI = 2.5–20.5), recessive model (GG versus AG+AA: OR = 20.3, 95%CI = 9.7–42.4), and allelic model (G versus A: OR = 11.6, 95%CI = 6.47–21.1). The expression levels of AIRE gene did not differ significantly between patients and controls and were not related to rs2075876 variant. In conclusion, the intronic variant (rs2075876) is suggested to be a potent susceptibility variant for AA development in the studied population. Abbreviations: AA: Alopecia areata; AIRE: Autoimmune Regulator; APECED: Autoimmune, Polyendocrinopathy Candidiasis Ectodermal Dystrophy; DLQI: Dermatology life quality index questionnaire; MIQE: Minimum information for publication of quantitative real-time PCR experiments; mTEC: Medullary thymic epithelial cells; PHD: Plant homeodomain; qRT-PCR: Quantitative reversetranscription-polymerase chain reaction; RA: Rheumatoid arthritis [ABSTRACT FROM AUTHOR]

Coexistence of bullous pemphigoid (BP) and vitiligo vulgaris (VV) is very rare. We present a unique case of BP associated with VV in a 76-year-old Japanese man, in which BP eruption developed exclusively on preexisting VV regions. The patient was referred to us with a 3-month history of blistering eruption with severe pruritis on the right forearm and left lower limb. In addition, he had been suffering from a widespread depigmented eruption on the trunk and extremities for at least 20 years. Curiously, the blistering eruption developed exclusively on preexisting depigmented eruption. Histopathological and laboratory examinations identified the blistering eruption as BP. The depigmented eruption was diagnosed as VV. BP eruption responded quickly to oral prednisolone therapy, improving within 1 week, and the prednisolone dose was gradually tapered. Within 1 month, BP lesions almost completely resolved. In the present case, BP not only coexisted with VV, but also developed exclusively on preexisting vitiliginous regions. The present case strongly suggests that BP and VV are partly caused by common pathological mechanisms. [ABSTRACT FROM AUTHOR]

Background/Aims: AIRE is known for its involvement in autoreactive T-cell deletion in thymic epithelium. Though extrathymic expression of AIRE is well documented, the functional relevance of AIRE in non-thymus tissues is emerging. AIRE is expressed in neonatal and adult testis, and has been implicated in sporadic germ cell apoptosis in developing testis. In this study we examined whether AIRE has any role in inducing apoptosis in cultured spermatogonial cells. Methods: We over-expressed AIRE or CARD domain of AIRE in GC1-spg cells and evaluated its impact on cell cycle using fluorescence activated cell sorting following Hoechst 33342 staining. Apoptosis was assayed using Annexin-V staining. Caspase-3 cleavage was assessed on western blots and caspase-3 expression was quantitated using realtime PCR. Results: We report that C18-4 cells which are derived from Type A spermatogonia expressed AIRE, while GC1-spg which is closer to Type B spermatogonia was negative for AIRE expression. Overexpression of AIRE or CARD domain of AIRE induced Caspase-3 expression in GC1-spg cells. Silencing of AIRE in C18-4 cells inhibited Caspase-3 expression. When overexpressed, AIRE and CARD brought about a very negligible increase in germ cell death and resulted in altered cell cycle pattern with a reduction in G1 phase. This was not associated with any increase in activation of Caspase-3. Conclusion: We conclude that the CARD domain of AIRE enhances caspase-3 expression through possible direct DNA binding and triggers non-apoptotic downstream signaling in cultured spermatogonial cells. [ABSTRACT FROM AUTHOR]

Autoimmune regulator (Aire), primarily expressed in medullary thymic epithelial cells (mTECs), maintains central immune tolerance through the clearance of self-reactive T cells. Aire can also be expressed in dendritic cells (DCs), and DCs can mediate T follicular helper (T FH) cell differentiation and self-reactive B cell activation through inducible costimulator molecule ligand (ICOSL) and interleukin 6 (IL-6), which can cause autoimmune diseases. To confirm whether Aire in DCs affects T FH cell differentiation and to determine the role of Aire in the maintenance of peripheral immune tolerance, this study observed the effects of Aire deficiency on T FH cells using Aire knockout mice. The results showed that Aire deficiency caused increased number of T FH cells, both in vivo and in vitro. Further studies showed that Aire deficiency promoted T FH differentiation through the upregulation of ICOSL and IL-6 in DCs. Thus Aire could suppress the expression of ICOSL and IL-6 to inhibit T FH cell differentiation. [ABSTRACT FROM AUTHOR]

Autoimmune regulator (AIRE) is a transcription factor that is expressed in medullary thymic epithelial cells. It plays an essential role in central tolerance by eliminating self-reactive T cells. Recently, extrathymic AIRE-expressing cells have been revealed, which are associated with peripheral tolerance. Moreover, AIRE expression has been demonstrated in skin tumors and breast cancer. However, the expression of AIRE in osteosarcoma is unknown. We used immunohistochemistry to investigate AIRE expression in biopsy samples from 43 patients with conventional osteosarcoma and statistically analyzed the association between AIRE expression and clinicopathological characteristics. High AIRE expression was detected in 25 patients (58.1%), and significantly associated with the presence of lung metastasis (P = 0.014) and an increased number of forkhead box P3-positive tumor-infiltrating lymphocytes (regulatory T cells) (P = 0.014). The overall survival rate for all osteosarcoma patients with high AIRE expression was significantly shorter than that for those with low AIRE expression (P = 0.046). In a subgroup analysis of American Joint Committee on Cancer stage II patients who underwent complete surgical resection and conventional chemotherapy, the overall survival and metastasis-free survival rates were significantly shorter for patients with high AIRE expression than for those with low AIRE expression (P = 0.019 and P < 0.01, respectively). High AIRE expression was confirmed to be an independent poor prognostic factor for both overall survival (hazard ratio: 3.841, P = 0.038) and metastasis-free survival (hazard ratio: 4.348, P = 0.022) in the multivariate analysis. The evaluation of AIRE expression may be useful for stratifying osteosarcoma patients for more effective clinical follow-up. [ABSTRACT FROM AUTHOR]

Problem: Dedicator of cytokinesis (DOCK 180) involved in cytoskeletal reorganization is primarily a cytosolic molecule. It is recently shown to be nuclear in HeLa cells but its nuclear function is not known. Method of study: The spatiotemporal distribution of DOCK180 in uterus was studied in uterine cytoplasmic and nuclear compartments during the “window of implantation.” The functional significance of nuclear DOCK180 was explored by homology modeling, co‐immunoprecipitation assays, and mass spectrometric analysis. Dock180's role in early pregnancy was ascertained by Dock 180 silencing and subsequent quantitative real‐time PCR and Western blotting analysis. Results: Our study shows a nuclear DOCK180 in the uterus during “window of implantation.” Estrogen and progesterone mediate expression and nuclear translocation of DOCK180. The nuclear function of DOCK180 is attributed to its ability to import autoimmune regulator (AIRE) into the nucleus. Silencing of Dock180 inhibited AIRE nuclear shuttling which influenced its downstream targets, thereby affecting decidualization with AIRE and HOXA‐10 as the major players as well as lack of implantation site formation due to impact on angiogenesis‐associated genes. Conclusion: DOCK180 has an indispensable role in pregnancy establishment as knocking down Dock180 abrogates pregnancy by a consolidated impact on decidualization and angiogenesis by regulating AIRE nuclear entry. [ABSTRACT FROM AUTHOR]

A genetic variant in the SAND domain of autoimmune regulator (AIRE), R247C, was identified in a patient with type I diabetes mellitus (T1DM) and his mother with rheumatoid arthritis. In vitro , the variant dominantly inhibited AIRE; however, typical features of Autoimmune Polyendocrinopathy Candidiasis and Ectodermal Dysplasia (APECED) were not seen in the subjects. Rather, early manifestation of autoimmunity appeared to be dependent on additional genetic factors. On a population level, diverse variants were identified in this region. Surprisingly, many likely pathogenic variants were seen disproportionately in Africans when compared to Europeans, reinforcing the importance of these variants in altering the immune repertoire. In light of these findings, we propose that R247C and other variants within the SAND-domain alter protein function in a dominant fashion and hold potential as drivers of autoimmunity. [ABSTRACT FROM AUTHOR]

As a transcription factor, autoimmune regulator (Aire) participates in thymic negative selection and maintains immune tolerance mainly by regulating the ectopic expression of tissue-restricted antigens (TRAs) in medullary thymic epithelial cells (mTECs). Aire is also expressed in dendritic cells (DCs). DCs are professional antigen-presenting cells (APCs) that affect the differentiation of T cells toward distinct subpopulations and participate in the immune response and tolerance, thereby playing an important role in maintaining homeostasis. To determine the role of Aire in maintaining immune tolerance by bone marrow-derived dendritic cells (BMDCs), in the present study we utilized Aire-knockout mice to examine the changes of maturation status and TRAs expression on BMDCs, additionally investigate the differentiation of CD4+T cells. The results showed that expression of costimulatory molecule and major histocompatibility complex class II (MHC-II) molecule was increased and expression of various TRAs was decreased in BMDCs from Aire-knockout mice. Aire deficiency reduced the differentiation of naïve CD4+T cells into type 2T helper (Th2) cells and regulatory T cells (Tregs) but enhanced the differentiation of naïve CD4+T cells into Th1 cells, Th17 cells, and follicular helper T (Tfh) cells. The results demonstrate that Aire expressed by BMDCs plays an important role in the maintenance of homeostasis by regulating TRA expression and the differentiation of T cell subsets. [ABSTRACT FROM PUBLISHER]

Autoimmune regulator (Aire) is one of the most crucial genes expressed in the thymus, where it orchestrates the promiscuous expression and presentation of tissue-specific antigens during thymocyte selection. The presence of Aire-expressing cells outside the thymus points toward its plausible extrathymic functions; however, the relative contribution of Aire-expressing cells of hematopoietic origin and their role in the modulation of autoimmune diseases are still obscure. Here, we report that non-obese diabetic mice with transgenic Aire expression under the control of the CD11c (integrin alpha X) promoter were significantly protected from autoimmune diabetes compared with their non-transgenic littermates. The protective effect of Aire transgene was mediated primarily by an increase in the “exhausted” populations of CD4+ and CD8+ T cells, both demonstrating poor expressions of interferon-γ and tumor necrosis factor-α. Both CD4+ and CD8+ effector T cells in transgenic mice displayed distinctive and differential expression of T-bet and Eomesodermin, respectively, in conjunction with high expression of programmed cell death protein-1 and other exhaustion-associated markers. Importantly, transgenic Aire expression did not result in any detectable changes in the population of Foxp3+ regulatory T (Treg) cells. Co-transfer experiments also demonstrated that Aire transgenic dendritic cells, as a “stand-alone” cell population, had the potential to suppress effector T cells in vivo without the support of Treg cells, but eventually failed to prevent the diabetogenesis in recipient mice. In conclusion, our study suggests that apart from its role in clonal deletion of autoreactive T cells or clonal diversion to Treg lineage, Aire can also contribute to tolerance by forcing effector T cells into a state of exhaustion with poor effector functions, thereby effectively containing autoimmune diseases.

High titer autoantibodies produced by B lymphocytes are clinically important features of many common autoimmune diseases. APECED patients with deficient autoimmune regulator (AIRE) gene collectively display a broad repertoire of high titer autoantibodies, including some which are pathognomonic for major autoimmune diseases. AIRE deficiency severely reduces thymic expression of gene-products ordinarily restricted to discrete peripheral tissues, and developing T cells reactive to those gene-products are not inactivated during their development. However, the extent of the autoantibody repertoire in APECED and its relation to thymic expression of self-antigens are unclear. We here undertook a broad protein array approach to assess autoantibody repertoire in APECED patients. Our results show that in addition to shared autoantigen reactivities, APECED patients display high inter-individual variation in their autoantigen profiles, which collectively are enriched in evolutionarily conserved, cytosolic and nuclear phosphoproteins. The APECED autoantigens have two major origins; proteins expressed in thymic medullary epithelial cells and proteins expressed in lymphoid cells. These findings support the hypothesis that specific protein properties strongly contribute to the etiology of B cell autoimmunity.

Autoimmune regulator (Aire) is one of the most crucial genes expressed in the thymus, where it orchestrates the promiscuous expression and presentation of tissue-specific antigens during thymocyte selection. The presence of Aire-expressing cells outside the thymus points toward its plausible extrathymic functions; however, the relative contribution of Aire-expressing cells of hematopoietic origin and their role in the modulation of autoimmune diseases are still obscure. Here, we report that non-obese diabetic mice with transgenic Aire expression under the control of the CD11c (integrin alpha X) promoter were significantly protected from autoimmune diabetes compared with their non-transgenic littermates. The protective effect of Aire transgene was mediated primarily by an increase in the “exhausted” populations of CD4+ and CD8+ T cells, both demonstrating poor expressions of interferon-γ and tumor necrosis factor-α. Both CD4+ and CD8+ effector T cells in transgenic mice displayed distinctive and differential expression of T-bet and Eomesodermin, respectively, in conjunction with high expression of programmed cell death protein-1 and other exhaustion-associated markers. Importantly, transgenic Aire expression did not result in any detectable changes in the population of Foxp3+ regulatory T (Treg) cells. Co-transfer experiments also demonstrated that Aire transgenic dendritic cells, as a “stand-alone” cell population, had the potential to suppress effector T cells in vivo without the support of Treg cells, but eventually failed to prevent the diabetogenesis in recipient mice. In conclusion, our study suggests that apart from its role in clonal deletion of autoreactive T cells or clonal diversion to Treg lineage, Aire can also contribute to tolerance by forcing effector T cells into a state of exhaustion with poor effector functions, thereby effectively containing autoimmune diseases. [ABSTRACT FROM AUTHOR]

High titer autoantibodies produced by B lymphocytes are clinically important features of many common autoimmune diseases. APECED patients with deficient autoimmune regulator (AIRE) gene collectively display a broad repertoire of high titer autoantibodies, including some which are pathognomonic for major autoimmune diseases. AIRE deficiency severely reduces thymic expression of gene-products ordinarily restricted to discrete peripheral tissues, and developing T cells reactive to those gene-products are not inactivated during their development. However, the extent of the autoantibody repertoire in APECED and its relation to thymic expression of self-antigens are unclear. We here undertook a broad protein array approach to assess autoantibody repertoire in APECED patients. Our results show that in addition to shared autoantigen reactivities, APECED patients display high inter-individual variation in their autoantigen profiles, which collectively are enriched in evolutionarily conserved, cytosolic and nuclear phosphoproteins. The APECED autoantigens have two major origins; proteins expressed in thymic medullary epithelial cells and proteins expressed in lymphoid cells. These findings support the hypothesis that specific protein properties strongly contribute to the etiology of B cell autoimmunity. [ABSTRACT FROM AUTHOR]

Next generation sequencing of T and B cell receptors is emerging as a valuable and effective method to diagnose and monitor hematopoietic malignancies. So far, this approach has not been fully explored in regard to autoimmune diseases. T cells develop in the thymus where they undergo positive and negative selection, and the autoimmune regulator (Aire) is central in the establishment of immunological tolerance. Loss of Aire leads to severe multiorgan autoimmune disease with infiltration of autoreactive T cells in affected organs. Here, we have utilized next generation sequencing technology to investigate the T cell receptor repertoire in autoimmunity induced by immunization of mice with a self-antigen, myeloperoxidase. By investigating the T cell receptor repertoire in peripheral blood, spleen and lumbar lymph nodes from naïve and immunized Aire −/− mice and wild type littermates, changes in the usage of V and J genes were evident. Our results identify TCR clonotypes which could be potential targets for immune therapy. Also, Aire −/− autoimmunity is driven by a variety of autoantigens where the autoimmune response is highly polyclonal, and access to the most adjacent immunologically active tissue is required to identify T cell receptor sequences that are potentially unique to the antigen in Aire−/− immunized mice. [ABSTRACT FROM AUTHOR]

Polymorphism in autoimmune regulator ( AIRE ) gene is associated with various autoimmune disorders. Abnormal AIRE expression is associated with the development of myasthenia gravis (MG). We investigated the association of polymorphism in AIRE gene and the clinical features and severity of MG. The frequencies of alleles and genotypes were compared between 480 MG patients and 487 healthy controls, as well as among subgroups of MG patients. The frequencies of rs3761389 G allele in MG group (OR = 1.213, CI 95% 1.014–1.451, p = 0.035) and in mild (Oosterhuis score 0–2) subgroup (OR = 1.393, CI 95% 1.110–1.751, p = 0.004) were significantly higher than those in the control group. There were significant differences in the frequencies of rs3761389 genotypes (OR = 1.20, CI 95% 1.00–1.43, p = 0.046, log-additive model) and mild subgroup (OR = 1.32, CI 95% 1.03–1.69, p = 0.0058, log-additive model) compared with the control group. A Logistic regression analysis did not identify rs3761389 genotype as an independent risk factor to predict the severity of MG. This study provides the necessary preliminary data on the association with rs3761389 in AIRE gene with the susceptibility of MG, but not with the severity of MG. [ABSTRACT FROM AUTHOR]

Autoimmune polyendocrinopathy with candidiasis and ectodermal dystrophy (APECED) is a rare disorder of immune dysregulation caused by mutations in the autoimmune regulator (AIRE) gene. Individuals affected with APECED develop a clinical syndrome characterized by ectodermal abnormalities, autoantibody production, and organspecific autoimmune manifestations. Inflammatory arthritis is usually not described as a part of the syndrome, and only sporadic cases are reported. We describe the case of a preschool-age girl who presented with hypoparathyroidism, hepatitis, interstitial pneumonitis, and chronic polyarthritis at 4 years of age and was found to have two compound heterozygous disease-associated mutations in the AIRE gene. We also conducted a literature review of the main characteristics of inflammatory arthritis in APECED patients. Our case and review demonstrate that (1) inflammatory arthritis, although rare, can be an early manifestation of APECED; (2) the diagnosis of APECED should be considered if mucocutaneous candidiasis, multiple organ-specific autoimmune manifestations, polyendocrinopathy, especially hypoparathyroidism or adrenal failure, or ectodermal dystrophy accompany joint symptoms; and (3) genotyping interpretation should take into account that mutations are found in the 14 exons of the gene, compound heterozygosity is common, and in some cases, only one or no mutated alleles are found. [ABSTRACT FROM AUTHOR]

Autoimmune regulator (Aire) mutations result in autoimmune polyendocrinopathy candidiasis ectodermal dystrophy (APECED), which manifests as multi-organ autoimmunity and chronic mucocutaneous candidiasis (CMC). Indendritic cells (DCs), pattern recognition receptors (PRR), such as Toll-like receptors (TLRs), are closely involved in the recognition of various pathogens, activating the intercellular signaling pathway, followed by the activation of transcription factors and the expression of downstream genes, which take part in mediating the immune response and maintaining immune tolerance. In this study, we found that Aire up-regulated TLR3 expression and modulated the downstream cytokine expression and nuclear factor-κB (NF-κB) of the TLR3 signaling pathway. [ABSTRACT FROM AUTHOR]