To investigate the effect of bromodomain and extra-terminal (BET) protein inhibitor JQ1 on expression of autoimmune-related genes in CD4+T cells from patients with systemic lupus erythematosus (SLE).
 Methods: Peripheral CD4+T cells were isolated by positive selection with CD4 microbeads. The percentage of CD4+T cells were detected by flow cytometry. CD4+T cells were treated by JQ1 at 100 nm/L for 6, 24, 48 h. The expression of T cell-related genes was measured by quantitative real-time PCR (qPCR). The secretion levels of cytokines in culture supernatant were measured by ELISA at 48 h.
 Results: The percentage of CD4+T cells isolated by CD4 microbeads is 97.2%. Compared with the control group, the mRNA expression levels of IFNG, IL-17F, IL-21, CXCR5 and FOXP3 were down-regulated at 6, 24 and 48 h (P<0.05), and IL-17A mRNA level was decreased at 6 and 24 h (P<0.01); while IL-4 mRNA level was up-regulated at 24, 48 h (P<0.01), and TGF-β1 mRNA level was up-regulated at 6 and 48 h (P<0.05) in SLE CD4+T cells treated with JQ1. The secretion levels of IFN-γ and IL-21 in JQ1-treated group were decreased significantly (P<0.05), while the secretion levels of IL-4 and TGF-β were up-regulated compared with control group (P<0.05).
 Conclusion: JQ1 can reverse the immune dysregulation and improve the immunity homeostasis in CD4+T cells from patients with SLE.
Azepines ; pharmacology ; CD4 Lymphocyte Count ; CD4-Positive T-Lymphocytes ; cytology ; drug effects ; metabolism ; Cytokines ; analysis ; metabolism ; Flow Cytometry ; Humans ; Interferon-gamma ; metabolism ; Lupus Erythematosus, Systemic ; immunology ; metabolism ; Proteins ; antagonists & inhibitors ; RNA, Messenger ; metabolism ; Time Factors ; Transforming Growth Factor beta1 ; Triazoles ; pharmacology

BackgroundImbalance of interferon-gamma (IFN-γ), interleukin (IL)-4, and IL-17 producing by T cells is confirmed to contribute to the pathogenesis of systemic lupus erythematosus (SLE). Autophagy is now emerging as a core player in the development and the function of the immune system. Therefore, we investigated the autophagic behavior in IFN-γ-, IL-4-, and IL-17-producing T cells from patients with SLE.

MethodsThirty patients with SLE and 25 healthy controls matched for gender and age were recruited between September 2016 and May 2017. The autophagic levels in IFN-γ T cells, IL-4 T cells, and IL-17 T cells from patients with newly diagnosed SLE and healthy controls were measured using flow cytometry. The plasma levels of IFN-γ were determined by enzyme-linked immunosorbent assay in SLE patients and healthy controls. Unpaired t-tests and the nonparametric Mann-Whitney U-test were used to compare data from patients with SLE and controls. Spearman's rank correlation coefficient was applied for calculation of the correlation between parallel variables in single samples.

ResultsOur results showed increased percentage of autophagy in IFN-γ T cells from patients with SLE and healthy controls ([8.07 ± 2.72]% vs. [3.76 ± 1.67]%, t = 5.184, P < 0.001), but not in IL-4 T cells or IL-17 T cells (P > 0.05) as compared to healthy donors. Moreover, the plasma levels of IFN-γ in SLE patients were significantly higher than those in healthy controls ([68.9 ± 29.1] pg/ml vs. [24.7 ± 17.6] pg/ml, t = 5.430, P < 0.001). Moreover, in SLE patients, the percentage of autophagy in IFN-γ T cells was positively correlated with the plasma levels of IFN-γ (r = 0.344, P = 0.046), as well as the disease activity of patients with SLE (r = 0.379, P = 0.039).

ConclusionThe results indicate that autophagy in IFN-γ T cells from SLE patients is activated, which might contribute to the persistence of T cells producing IFN-γ, such as Th1 cells, and consequently result in the high plasma levels of IFN-γ, and then enhance the disease activity of SLE.

Adult ; Autophagy ; China ; Female ; Humans ; Interferon-gamma ; metabolism ; Interleukin-17 ; metabolism ; Interleukin-4 ; metabolism ; Lupus Erythematosus, Systemic ; immunology ; Male ; Middle Aged ; Th1 Cells ; physiology

We describe an unusual case of systemic lupus erythematosus with pulmonary manifestations presenting as hypoglycemia due to anti-insulin receptor antibodies. A 38-year-old female suffered an episode of unconsciousness and was admitted to hospital where her blood glucose was found to be 18 mg/dL. During the hypoglycemic episode, her serum insulin level was inappropriately high (2,207.1 pmol/L; normal range, 18 to 173) and C-peptide level was elevated (1.7 nmol/L; normal range, 0.37 to 1.47). Further blood tests revealed the presence of antinuclear antibodies, anti-double-stranded DNA antibodies, and anti-Ro/SSA, anti-La/SSB, anti-ribonucleoprotein, and anti-insulin receptor antibodies. A computed tomography scan of the abdomen, aimed at tumor localization, such as an insulinoma, instead revealed ground-glass opacities in both lower lungs, and no abnormal finding in the abdomen. For a definitive diagnosis of the lung lesion, video-associated thoracoscopic surgery was performed and histopathological findings showed a pattern of fibrotic non-specific interstitial pneumonia.
Adult ; Autoantibodies/*blood ; *Autoimmunity ; Biological Markers/blood ; Blood Glucose/metabolism ; Female ; Humans ; Hypoglycemia/blood/*complications/immunology ; Insulin/blood ; *Insulin Resistance ; Lung Diseases, Interstitial/diagnosis/*etiology/immunology/surgery ; Lupus Erythematosus, Systemic/*complications/diagnosis/immunology ; Receptor, Insulin/*immunology ; Thoracic Surgery, Video-Assisted ; Tomography, X-Ray Computed ; Treatment Outcome

OBJECTIVETo observe the effects of Xuebijing injection on dendritic cells (DCs) and T lymphocytes, and the potential mechanisms of its therapeutic effect on systemic lupus erythematosus (SLE).

METHODSA widely used mouse model, SLE-prone BLLF1 mice aged 8-10 weeks, was employed. Mice were randomly divided into 4 groups: a normal group, a model group and two treatment groups treated with Xuebijing Injection with a dose of 6.4 mL/kg via intraperitoneal administration for SLE-prone BLLF1 mice aged 8 weeks (treatment A group) and 10 weeks (treatment B group). Renal tissue sections were stained with Masson's trichrome and periodic acid-silver methenamine. Histopathological changes in the kidney were evaluated by a light microscopy. The capacity of the DCs isolated from the spleen to stimulate the T cell proliferation in response to concanavalin A (Con A) was determined.

RESULTSCompared with the model group, levels of anti-dsDNA antibodies in the two treatment groups decreased remarkablly (P<0.01, P<0.05), and levels of serum creatinine and blood urea nitrogen increased (P<0.01, P<0.05). Pathological changes were found in the kidney in the model group. Histopathological abnormalities were alleviated in the two treatment groups. Treatment with Xuebijing injection also significantly upregulated the expression of CD80, CD86 and major histocompatibility class II by DCs compared with the model group (P<0.05). When splenic T lymphocytes from BLLF1 mice were co-cultured with DCs at ratios of 1:100, 1:150 and 1:200 for 3 and 5 days, the proliferation of T lymphocytes was suppressed compared with the normal group (P<0.05), but this was restored by Xuebijing Injection under the same conditions. In the model group, levels of tumor necrosis factor (TNF)-α in supernatants were significantly elevated compared with the normal group (P<0.01), interleukin-2 levels decreased (P<0.05), while these changes were significantly alleviated in the Xuebijing treatment groups.

CONCLUSIONSXuebijing Injection alleviated renal injury in SLE-prone BLLF-1 mice. The mechanism might be through influencing T cell polarization mediated by DCs, and Xuebijing Injection might be a potential drug that suppresses immune dysfunction in patients with SLE.

Animals ; Antibodies, Antinuclear ; blood ; Cell Differentiation ; drug effects ; Cell Proliferation ; drug effects ; Concanavalin A ; pharmacology ; Dendritic Cells ; drug effects ; immunology ; pathology ; Drugs, Chinese Herbal ; administration & dosage ; pharmacology ; therapeutic use ; Injections ; Interleukin-2 ; metabolism ; Kidney ; drug effects ; pathology ; physiopathology ; ultrastructure ; Kidney Function Tests ; Lupus Erythematosus, Systemic ; blood ; drug therapy ; immunology ; physiopathology ; Mice ; Phenotype ; T-Lymphocytes ; drug effects ; immunology ; pathology ; Tumor Necrosis Factor-alpha ; metabolism

Recently, subpopulations of regulatory T (Treg) cells, resting Treg (rTreg) and activated Treg (aTreg), have been discovered. The authors investigated the relationship between the change of Treg, aTreg and rTreg and autoimmune diseases. Treg cells and those subpopulations were analyzed by using the human regulatory T cell staining kit and CD45RA surface marker for 42 rheumatoid arthritis (RA), 13 systemic lupus sclerosis (SLE), 7 Behcet's disease (BD), and 22 healthy controls. The proportion of Treg cells was significantly lower in RA (3.8% +/- 1.0%) (P < 0.001) and BD (3.3% +/- 0.5%) (P < 0.01) compared to healthy controls (5.0% +/- 1.3%). The proportion of aTreg cells was also significantly lower in RA (0.4% +/- 0.2%) (P = 0.008) and BD (0.3% +/- 0.1%) (P = 0.013) compared to healthy controls (0.6% +/- 0.3%). The rTreg cells showed no significant differences. The ratio of aTreg to rTreg was lower in RA patients (0.4% +/- 0.2%) than that in healthy controls (0.7% +/- 0.4%) (P = 0.002). This study suggests that the decrement of aTreg not rTreg cells contributes the decrement of total Treg cells in peripheral blood of RA and BD autoimmune diseases. Detailed analysis of Treg subpopulations would be more informative than total Treg cells in investigating mechanism of autoimmune disease.
Adult ; Aged ; Antigens, CD4/metabolism ; Antigens, CD45/metabolism ; Arthritis, Rheumatoid/*immunology/metabolism ; Behcet Syndrome/*immunology/metabolism ; Female ; Forkhead Transcription Factors/metabolism ; Humans ; Interleukin-2 Receptor alpha Subunit/metabolism ; Leukocyte Count ; Lupus Erythematosus, Systemic/*immunology/metabolism ; Male ; Middle Aged ; T-Lymphocytes, Regulatory/*cytology/immunology/metabolism

BACKGROUNDPrevious studies indicate that CD43 plays a role in regulating the adhesion of lymphocytes, cell mutation and activation, however, little is known about its effect on systemic lupus erythematosus (SLE). This study was designed to explore the clinical significance of CD43 in SLE patients.

METHODSWe used microarray and real-time PCR to detect the mRNA and protein expression of magnetic bead sorted T cells and B cells from peripheral blood mononuclear cells (PBMCs) of SLE patients, and analyzed the relationship between CD43 and the clinical indexes.

RESULTSBoth microarray and real-time PCR results showed that CD43 mRNA was significantly decreased in PBMCs of SLE patients compared with healthy controls (P < 0.001). There were no significant differences between lupus nephritis and non-lupus nephritis patients, and neuropsychiatric and non-neuropsychiatric patients. CD43 mRNA expression was significantly reduced in T cells but not in B-cells in SLE patients compared to healthy controls (P < 0.01). Compared with healthy controls, the percentage of CD43(+) cells in the PBMCs of SLE was significantly decreased (P = 0.004), and the CD43 fluorescence intensity in CD3(+)/CD43(+) cells and CD19(+)/CD43(+) cells was also significantly weaker than in healthy controls (P = 0.039 and 0.003). There was no significant difference in the percentage of CD3(+)/CD43(+) cells, CD19(+)/CD43(+) cells between the two groups. The CD43 fluorescence intensity in CD3(+)/CD43(+) cells was inversely correlated with the levels of IgG and IgM (r = -0.8 and -0.6).

CONCLUSIONSCompared to healthy controls, both CD43 mRNA and protein expressions were reduced in T cells from patients with SLE, and were inversely correlated with IgG.

B-Lymphocytes ; immunology ; metabolism ; Humans ; Leukocytes, Mononuclear ; Leukosialin ; genetics ; metabolism ; Lupus Erythematosus, Systemic ; immunology ; metabolism ; Oligonucleotide Array Sequence Analysis ; Real-Time Polymerase Chain Reaction ; T-Lymphocytes ; immunology ; metabolism

OBJECTIVE: To investigate the effect of total glucosides of peony (TGP) on expression and DNA methylation status of ITGAL gene (CD11a) in CD4(+) T cells from patients with systemic lupus erythematosus (SLE). METHODS: CD4(+) T cells were isolated by positive selection using CD4 beads. CD4(+) T cells were treated by TGP at 0, 62.5, 312.5 and 1562.5 mg/L for 48 h. The MTT method was used to assess cell viability; mRNA expression level was measured by realtime-PCR; protein level of CD11a was measured by flow cytometric analysis; DNA methylation status was assayed by bisulfite sequencing. RESULTS: No significant change in cell viability was found in CD4(+) T cells among the different concentration groups (P>0.05). Compared with control, the mRNA and protein levels of ITGAL were down-regulated significantly in SLE CD4(+) T cells treated with TGP (1562.5 mg/L) (P< 0.01). Furthermore, the extent of DNA methylation of ITGAL promoter was increased in TGP (1562.5 mg/L) treated CD4(+) T cells compared with control group (P<0.01). CONCLUSION TGP can repress CD11a gene expression through enhancing DNA methylation of ITGAL promoter in CD4(+) T cells from patients with SLE. This observation represents a preliminary step in understanding the mechanism of TGP in SLE therapy.
CD11a Antigen ; genetics ; metabolism ; CD4-Positive T-Lymphocytes ; immunology ; metabolism ; DNA Methylation ; drug effects ; Down-Regulation ; drug effects ; Glucosides ; pharmacology ; Humans ; Lupus Erythematosus, Systemic ; genetics ; immunology ; Paeonia ; chemistry ; Promoter Regions, Genetic ; genetics ; RNA, Messenger ; genetics ; metabolism

BACKGROUNDActivation in vitro of natural killer T (NKT) cells in systemic lupus erythematosus (SLE) with α-galactosylceramide (α-GalCer) and dendritic cells (DC) may affect the immunoregulatory role of NKT cells. This study was designed to compare the number of NKT cells in patients with SLE to the number in healthy volunteers and measure the cytokines secreted from these NKT cells in vitro.

METHODSThree sets of culture conditions using (i) α-GalCer, (ii) DC, or (iii) both α-GalCer and DC (α-GalCer+DC) were adopted to expand NKT cells from peripheral blood mononuclear cells (PBMC) of patients with SLE and healthy volunteers. Flow cytometry was used to assess the levels of interleukin (IL)-4, IL-10, interferon (IFN)-γ and tumor necrosis factor (TNF)-α produced by the Vα24(+)Vβ11(+) NKT cells.

RESULTSAfter 14 days in culture, the total cell count and percentage of Vα24(+)Vβ11(+) NKT cells were increased under all conditions but were highest in the α-GalCer+DC group. The level of IL-4 and IL-10 secreted by Vα24(+)Vβ11(+) NKT cells from patients with active SLE was found to be higher than that of inactive patients and the control group (P < 0.05), while the levels of IFN-γ and TNF-α were lower than those found in the inactive and control groups (P < 0.05).

CONCLUSIONSVα24(+)Vβ11(+) NKT cells showed the greatest expansion in vitro with α-GalCer and DC. Th2-type cytokines from Vα24(+)Vβ11(+) NKT cells are the predominant type in patients with SLE, while Th1 cytokines predominate in the control group. This evolution of NKT cell function during the progression of the disease may have important implications in understanding the mechanism of SLE and for the development of possible therapies using NKT cell agonists.

Adolescent ; Adult ; Cell Proliferation ; drug effects ; Cells, Cultured ; Cytokines ; metabolism ; Dendritic Cells ; metabolism ; Female ; Flow Cytometry ; Galactosylceramides ; pharmacology ; Humans ; Interferon-gamma ; metabolism ; Interleukin-10 ; metabolism ; Interleukin-4 ; metabolism ; Lupus Erythematosus, Systemic ; immunology ; metabolism ; Male ; Middle Aged ; Natural Killer T-Cells ; cytology ; drug effects ; metabolism ; Receptors, Antigen, T-Cell ; metabolism ; Receptors, Antigen, T-Cell, alpha-beta ; metabolism ; Tumor Necrosis Factor-alpha ; metabolism ; Young Adult

Tissue and organ differentiation is tightly controlled to ensure proper development and function of the growing embryo as well as cells such as lymphocytes that differentiate throughout the adult stage. Therefore it is vital that the genes and the protein they encode that are involved in these processes function accurately. Hence, any mutation or error that occurs along the way can result in extensive damage, which is expressed in various ways in the embryo and can result in immune pathogenesis, including immunodeficiency and autoimmune diseases, when lymphocyte development is altered. A number of studies have been carried out to look at the genes regulating transcription in tissue differentiation, including the transcription factors Pbx1. This gene is of particular interest to us as we have identified that it is associated with systemic lupus erythematosus susceptibility (Cuda et al., in press). This perspective summarizes the known roles of Pbx1 in tissue differentiation as well as our recent findings associating genetic variations in Pbx1 to lupus susceptibility, and we will speculate on how this gene controls the maintenance of immune tolerance in T cells.
Animals ; Cell Differentiation ; Chromatin Immunoprecipitation ; DNA-Binding Proteins ; genetics ; immunology ; Genetic Loci ; immunology ; Genetic Predisposition to Disease ; Homeodomain Proteins ; genetics ; immunology ; Humans ; Immune Tolerance ; Lupus Erythematosus, Systemic ; genetics ; immunology ; Lymphocyte Activation ; Mice ; Mice, Transgenic ; Pre-B-Cell Leukemia Transcription Factor 1 ; Protein Structure, Tertiary ; Proto-Oncogene Proteins ; genetics ; immunology ; Signal Transduction ; T-Lymphocytes, Regulatory ; cytology ; immunology ; Transcription Factors ; genetics ; immunology ; Tretinoin ; metabolism

PURPOSE: To distinguish lupus flare-up from infection in systemic lupus erythematosus (SLE), we analyze the expression of circulating CD27(high) plasma cells in SLE patients with and without infection, in comparison to non-SLE patients with infection. MATERIALS AND METHODS: The percentage of circulating CD27(high) plasma cells was measured by flow cytometry in the following four groups: 36 SLE patients without infection, 23 SLE patients with infection, eight non-SLE patients with infection, and 26 healthy controls. RESULTS: The frequency of CD27(high) plasma cells had a correlation with the SLE disease activity index (SLEDAI) (r = 0.866, p < 0.05), level of anti-dsDNA (r = 0.886, p < 0.05), C3 (r = - 0.392, p < 0.05), and C4 (r = - 0.337, p < 0.05) in SLE patients without infection, but there was no correlation with disease activity in SLE patients with infection. Among three groups in particular-SLE without infection, SLE with infection, and non-SLE with infection-the percentages of CD27(high) plasma cells were elevated. The percentage of CD27(high) plasma cells was higher in SLE patients with infection, when compared to SLE patients without infection. CONCLUSION: The percentage of CD27(high) plasma cells is a biomarker for disease activity of SLE without infection, under correlation with SLEDAI, anti-dsDNA, and C3 and C4 level. However, when the SLE patients have an infection, the percentage of CD27(high) plasma cells is not an adequate biomarker for the survey of disease activity. The percentage of CD27(high) plasma cells may serve as a potential parameter to distinguish a lupus flare-up from infection.
Adult ; Antigens, CD27/*biosynthesis ; Bacterial Infections/complications ; Biological Markers/metabolism ; Case-Control Studies ; Female ; Flow Cytometry/methods ; Humans ; Lupus Erythematosus, Systemic/*blood/immunology ; Male ; Plasma Cells/cytology/*immunology ; Virus Diseases/complications