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Objective:To investigate the expression of B lymphocyte induced maturation protein 1 (Blimp-1) in peripheral blood CD4 + T cells, CD19 + B cells and labial glands of patients with primary Sj?gren′s syndrome (pSS) and the correlation of Blimp-1 with clinical features. Methods:Expression of PRDM1 at mRNA level in CD4 + T cells, CD19 + B cells and labial gland tissues were detected by RT-PCR. Immunohistochemistry (IHC) was used to observe the distribution of Blimp-1. Correlation of PRDM1 expression at mRNA level with clinical indicators was analyzed. Results:PRDM1 expression at mRNA level in CD4 + T and CD19 + B cells were significantly higher in pSS group than in healthy control (HC) group ( P<0.01). Based on European League Against Rheumatism Sj?gren′s Syndrome Disease Activity Index (ESSDAI), the patients were classified into two groups: active group (ESSDAI≥5) and inactive group (ESSDAI<5). PRDM1 expression at mRNA level in CD4 + T and CD19 + B cells were also higher in the active group than in inactive group ( P<0.05). Blimp-1 protein accumulated around the acinus and duct of labial gland and in the germinal center in pSS patients. PRDM1 expression at mRNA level in labial gland tissues of pSS patients was positively correlated with lymphocyte infiltration ( r=0.781, P<0.001). Conclusions:pSS displayed high expression of Blimp-1. Blimp-1 might affect pSS disease activity and have clinical significance in pSS treatment.
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Objective To investigate the change and significance of regulatory T cells (Tregs) in peripheral blood in patients with acute and chronic gout. Methods Flow cytometry was used to detect the ratio of Tregs in peripheral blood of healthy controls, patients with acute gout and patients with chronic gout. Enzyme linked immunosorbent assay (ELISA) was used to detect the levels of transforming growth factor (TGF)-βand interleukin (IL)-1βin plasma. Then, statistical analysis was conducted to analyze the changes and significance in different stages of gout, such as F test, Kruskal-Walls H test, q test and Pearson and Spearman correlation analysis. Results ① The percentage of CD4+CD25+Foxp+Treg/CD4+ T cells in peripheral blood was (1.22±0.27)％ in control group. While in patients with acute gout, it was (1.51±0.43)％, and (0.47±0.26)％in patients with chronic gout. There were statistical significant difference among the three groups ( F=101.39, P<0.05). The percentage of Tregs in acute gout group was significantly higher than that in control group and chronic gout group, while it was significantly lower in chronic gout group than in control group ( P<0.05). ②The concentration of TGF-β in plasma was (170 ±12) ng/L in control group, (214 ±77) ng/L in patients with acute gout and (179±21) ng/L in patients with chronic gout, the difference was statistically-significant (F=6.20, P<0.05). The concentration of TGF-β in plasma in acute gout group was significantly higher than the control group and chronic gout group, the difference was statistically significant (P<0.05), while the difference between chronic gout group and the control group was not statistically significant ( P>0.05). ③The con-centration of IL-1β in plasma in the control group was (4.8 ±1.3) ng/L, while that in patients with acute and chronic gout was (10.1±8.5) ng/L and (11.50±12.57) ng/L respectively, the difference between these three groupswas stati-sticallysignificant (P<0.05). The concentration of IL-1β in plasma in acute gout group and chronic gout group wassignificantly higher than that in the control group, the difference was statistical significant ( P<0.05). There was no significant difference between acute gout and chronic gout (P>0.05) patients. ④ The percentage of Tregs in peripheral blood of gout patients was negatively correlated with the duration of disease and the number of gout attacks within six months (duration of disease: r =-0.381, P <0.01 ; number of gout attacks: r=-0.518, P<0.01). But there wasno significant correlation to the concentration of TGF-β and IL-1β. Conclusion Tregsincreasesin acute gout and participate in the alleviation of gout inflammation, while the persistence of chronic gout may be related to the decrease of Tregs. Therefore, Tregs play an important regulatory role in the transformation of acute and chronic gout.
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Objective To investigate the relationship of skin-resident memory T cells (Trm cells) with the skin lesions in systemic lupus erythematosus (SLE) so as to deepen our understanding of the pathogenesis of skin lesions in SLE. Methods Peripheral blood and skin samples were collected from SLE patients and matched healthy volunteers. The percentages of effector memory T cells (Tem cells) and effcctor T cells (Teff cells) from peripheral blood were analyzed by flow cytometry. By using direct immunofluoresence, we detected the presence of immune complex,which deposited on the basement membrane of normal appearance skin from SLE patients or healthy individuals. Immunohistochemical staining was performed to detect the two characteristic surface markers of tissue-resident memory T lymphocytes,CD69 and CD103 and to analyze the expression of these T lymphocytes within skins from SLE patients or healthy volunteers. Data analysis was performed using t test. P<0.05 was considered statistically significant. Results As compared with control individuals,the proportions of CD4+Tem (12.6±3.4 vs 8.2±2.5,t=-3.15,P<0.05),CD4+Teff (2.5±1.5 vs 1.3± 0.8,t=-2.79,P<0.05)、CD8+Tem(15.3±3.6 vs 7.0±3.0,t=-6.22,P<0.05)and CD8+Teff cells(13.1±5.4 vs 3.7± 1.3,F=-7.36,P<0.05)in T cell subset were significantly increased. Also,the proportions of CD4+Tem(8.4±2.7 vs 5.8±2.0,t=-2.74,P<0.05),CD4+Teff (1.6±1.0 vs 0.8±0.5,t=-2.84,P<0.05),CD8+Tem (10.4±3.6 vs 5.3±2.4, t=-4.03, P<0.05) and CD8+Teff cells (8.2±4.1 vs 2.6±0.8, t=-6.15, P<0.05) in peripheral blood were increased as well. By direct immunofluorescence,we noticed the deposition of immunoglobulin IgA, IgM and complement C3 on the basement membrane zone of skins from SLE patients but not on heanlhy individuals. The amount of infiltrated lymphocytes in skin samples from SLE patients were significantly iecreased compared to that of healthy individuals,and the quantities of CD4, CD8 and CD103 positive T cells from SLE skin samples were all increased by various degrees than that of controls. Conclusion Skin-resident memory T cells may be involved in the pathogenesis of skin lesions in SLE.
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Objective To investigate the effect of long-term low dose prednisone administration on bone mineral density (BMD) in patients with inactive systemic lupus erythematosus (SLE).Methods A total of 118 inactive female SLE patients with long-term administration of low dose prednisone were recruited from the Department of Rheumatology and Immunology at An hui Provincial Hospital.All patients were given low dose prednisone for long-term (≤ 10 mg/d,more than half a year).According to prednisone doses,subjects were divided into two groups,namely group A (≤7.5 mg/d) and group B (7.5-10 mg/d).In addition,patients were also divided into four groups based on the duration of administration,including group Ⅰ ≤3 years,Ⅱ from 4-5 years,Ⅲ 6-10 years and Ⅳ ＞ 10 years.Twenty-nine healthy people were recruitedas normal controls.The BMD was measured by dual energy X-ray absorptiometry.The association of BMD with prednisone dose and duration was compared between different groups.Results The incidencc of osteopenia in all patients with SLE was 42.4％ (50/118),and the incidence of osteoporosis was 14.4％ (17/118).BMD of all bone sites in both group A and B were significantly lower than that in normal control group (P ＜ 0.05).Similarly,the BMD of all bone sites in group Ⅰ,Ⅱ,Ⅲ and Ⅳ were significantly decreased (P ＜ 0.05).What needed to be stressed was the BMD in group Ⅳ was lower than those in other three groups (P ＜ 0.05).Multiple logistic regression analysis showed that the cumulative prednisone dose was the risk factor for osteopenia,while taking calcium and alfacalcidol were protective factors.Conclusion Long-term use of low dose prednisone result in the decrease of BMD in patients with inactive SLE.The lumbar spine and femoral neck had more severe osteopenia.Long-term administration of prednisone,even less than 7.5 mg/d,can also cause osteopenia.Calcium and alfacalcidol were protective factors of BMD.
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Objective To investigate the significance of ultrasonography in the examination of rheumatoid arthritis(RA). Methods The wrist joints of activated RA and wrist, metacarpophalangeal, proximal interphalangeal joints of remission of RA were determined, and the imaging features were analyzed. Results Patients with activated RA were divided into two groups. The bone erosion and sheath lesions were lower in the group of duration less than 1 year than those in the group of duration over 1 year (P < 0.01). The positive rate of ultrasound was higher than that of X-ray in bone erosion (P < 0.05). To patients with the remission of RA, the positive rate of ultrasound was higher than that of the physical examination in synovitis ( P < 0 . 05 ) . Conclusions For bone erosion, ultrasound is better than X-ray for patients with early RA. For synovitis, the sensitivity of ultrasonography is higher than the physical examination in remission for RA patients.
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Objective To detect the levels of peptidylarginine deiminase 4 (PAD4) mRNA and neutrophil extracellular traps (NETs) of the peripheral blood neutrophils in rheumatoid arthritis (RA) patients and to study the association between PAD4 and NETs in RA.The anti-cyclic citrullinated peptide (anti CCP) antibodies,disease activity score (DAS28) score,erythrocyte sedimentation rate (ESR) were recorded.Its value in the pathogenesis of RA was explored.T test,rank-sum test,Pearson and Spearman correlative analysis were used for statical analysis.Methods The serum double-stranded DNA (dsDNA)/NETs in 43 RA patients and 20 healthy controls were detected by PicoGreen dsDNA Quantitation Kits (Invitrogen).Real-time quantitative polymerase chain reaction (RT-PCR) was used to determine the expression of PAD4 mRNA in neutrophils,and the relationship between dsDNA/NETs and their clinical indexes were analyzed.Results ① The level of peripheral blood neutrophils PAD4 mRNA in RA was significantly higher than healthy controls [1.493(0.831,2.607)] vs [0.631(0.358,1.489)],(Z=-2.07,P=0.039).The levels of serum PAD4 mRNA in RA treated with disease-modifying antirheumatic drugs (DMARDs) were lower than untreated with DMARDs but no significant difference was found (Z=-1.19,P=0.234).② The levels of serum dsDNA/NETs in RA patients were significantly higher than in healthy controls (t=3.4,P=0.001).③ The levels of serum dsDNA/NETs in RA were positively correlated with DAS28 score,ESR,CRP and PAD4 mRNA (r=0.36,P=0.036;r=0.345,P=0.042;r=0.36,P=0.043;r=0.42,P=0.017) but not with anti-CCP antibodies (r=0.277,P=0.154).Conclusion ① PAD4 may play an important role in the pathogenesis of RA.② NETs maybe associated with disease activity and play an important role in RA pathogenesis,inhibit NETs generation or improve the ability to clear NETs,perhaps can treat RA.③ PAD4 probably play an important role in rheumatoid arthritis by influencing the formation of the NETs.
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Objective To analyze the association of miR-326 mRNA expression level on regulatory T cells between clinical manifestations of patients with systemic lupus erythematosus (SLE), in order to inves-tigate the role of miR-326 on Treg cells and pathogenesis as well as its association with disease activity of SLE. Methods Twenty milimeter anti-coagulated peripheral blood was obtained from 52 SLE patients and 20 healthy controls. Treg were purified by MACS from peripheral blood, in which the quantity of miR-326 and Ets-1 mRNA were assessed by real-time polymerase chain reaction (PCR). Data were analyzed by using Mann-Whitney U test and Spearman correlation analysis. Results Compared with healthy controls [0.921(0.345, 1.879)], the expression of miR-326 mRNA level was significantly higher in Treg from SLE patients [2.927 (0.518, 8.662) (Z=-3.756, P<0.05). The difference between new-onset SLE patients [8.878(5.922, 12.466)] and recurrence group [3.512(0.582, 11.223)] with healthy controls was significant (Z=-2.135, Z=-4.928, P<0.05). The expression level of miR-326 in new-onset SLE patients was higher than inactive SLE patients (Z=-4.928, P<0.05). Significant difference of the expression level of miR-326 mRNA was found between new-onset SLE patients with serous cavity effusion [7.606(0.656, 16.795)] and new-onset SLE patients without it [1.840(0.576, 13.025)](Z=4.263, P<0.05). Our analysis showed that significant positive correlation was found between the expression of miR-326 mRNA in Treg with CRP (rs=0.481, P<0.05) and anti-C1q antibody (rs=0.729, P<0.05) from new-onset SLE patients. Conclusion The expression level of miR-326 is upregulated in Treg from SLE patients and is associated with disease active index, which suggests that miR-326 in Treg may participate the pathological process and disease activity in SLE.
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Objective To explore the role of serum neutrophil extracellular traps (NETs) in rheumatoid arthritis (RA).Methods The serum circulating free DNA (cf-DNA) / NETs in 45 RA patients and 18 healthy controls were detected by Pico Green dsDNA Quantitation Kits,and the association between serum NETs and ESR,DAS28 score,anti-CCP antibodies was analyzed.T test,rank sum test,Pearson or Spearman correlation analysis were ued for statistical analysis.Results ① The levels of serum dsDNA/NETs in RA patients [0.523 0(0.282 0,1.637 0) μg/ml] were significantly higher than those in healthy controls [0.410 5 (0.140 0,0.966 0)μg/ml] (Z=-2.419,P=0.016).② The level of serum dsDNA/NETs in RA was positively correlated with ESR and DAS28 score (r=0.357,P=0.016; r=0.325,P=0.029),but not with anti-CCP antibodies(r=0.146,P=0.434).③ The levels of serum dsDNA/NETs in RA treated by DMARDs were lower than those of untreated with DMARDs[0.516 5(0.282 0,1.637 0) μg/ml,0.523 0 (0.369 0,1.485 0) μg/ml,Z=-1.215,P=0.225],but the difference was not significant.Conclusion NETs maybe associated with disease activity and play an important role in RA pathogenesis.
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Objective To explore the etiopathogenisis of fungal infections induced by systemic lupus erythematosus (SLE). Methods One hundred and forty-seven SLE patients with fungal infections during 2004-2013 were assigned as experimental group and the same number of SLE patients without infections were randomly selected as control group. Clinical and laboratory documents of these patients were comparatively analyzed. Results The fungal infections in SLE patients affected oral mucosa , lower respiratory tract and skin , and the pathogenic bacteria were mainly candida albicans. The wide application of glucocorticoid, immunosuppressants and antibiotics pushed the rise of incidence of fungal infections in SLE patients significantly. Conclusions The patients′ use of glucocorticoid , immunosuppressants and antibiotics attributes to a higher risk of fungal infections , especially infections by candida albicans. Fungus infection may raise the level of C4.
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Objective To explore the significance of aryl hydrocarbon receptor (AhR) in patients with rheumatoid arthritis (RA) through detecting the levels of AhR and its response gene cytochrome P4501 A1 (CYP1 A1) in peripheral blood mononuclear cells (PBMCs).Methods Peripheral blood samples were collected from 35 patients with RA and 20 healthy subjects.The expression of AhR and CYP1A1 at mRNA level were detected by real-time PCR.The percentages of AhR-positive cells in PBMCs were detected by flow cytometry (FCM).The effects of leflunomide (LEF) on the expression of AhR and CYP1A1 were analyzed.The detailed clinical data of RA patients were recorded.The disease activity score (DAS) was calculated.Correlation analysis between AhR/CYP1A1 level and clinical data was conducted.Results (1) Both the expression of AhR at mRNA level and the percentage of AhR-positive cells in PBMCs from RA patients without LEF treatment were significantly higher than those from healthy subjects [(3.61±1.65) vs.(2.00±1.27),P=0.002; (34.21±11.30)％ vs.(18.83±7.32)％,P＜0.01].There were no statistically significant differences in the expression of AhR at mRNA level and the percentages of AhR-positive cells between patients with or without LEF treatment [(3.83 ± 1.62) vs.(3.61 ± 1.65),P =0.670 ; (36.69±10.61)％ vs.(34.21±11.30)％,P=0.462].(2) Non-LEF treatment group showed a higher relative expression of CYP1 A1 at mRNA level than that from control group [1.33 (0.08,7.86) vs.(0.62 ±0.29),z=-3.922,P＜0.01],but there was no statistical difference between LEF treatment group and non-LEF treatment group [(2.62±2.08) vs.1.33(0.08,7.86) z=-0.133,P=0.894].(3) Neither the expression of AhR and CYP1A1 at mRNA level nor the percentages of AhR-positive cells showed significant correlations with clinical data.Conclusion AhR was highly expressed in PBMCs from patients with RA,which might participate in the progression of rheumatoid arthritis.But the high expression of AhR did not reflect disease activity.Moreover,the treatment of LEF showed no significant influences on the expression of AhR and CYP1A1 in PBMCs from patients with RA.
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Objective To analyze the expression of purinergic receptor P2X ligand-gated ion channel 7 (P2X7R) on different cells and peripheral blood mononuclear cell (PBMC) and to investigate its correlation with inflammatory cytokines in patients with SLE.Methods Flow cytometry was used to detect surface expression of P2X7R on lymphocytes,CD4+ cells,and CD19+ cell in 29 SLE patients and 28 healthy human controls to compare the difference between the SLE patients and the controls in P2X7R expression.Enzyme linked immunosorbent assay (ELISA) was performed to detect P2X7R-related serum cytokines interleukin (IL)-1β,IL-6,tumor necrosis factor (TNF)-α level.T test,Wilcoxon rank sum test,Spearman's correlation analysis were used for statitical analysis.Results ① SLE patients had significantly higher expression of P2X7R on CD4+,CD8+ lymphocytes compared to controls [CD4+ cells∶ 2.21(3.55) vs 0.89(1.15),Z=-1.527,P=0.015； CD19+ cells∶ 11.53(20.01) vs 6.66 (6.27),Z=-2.091,P=0.037]； ② The levels of three cytokines in patients with SLE were significantly higher than those in control.The positive relationship between P2X7R expression in lymphocytes with the serum IL-6 level was found in SLE patients (r=0.449,P=0.015)；③ Patients with arthritis showed significantly higher expression of P2X7R on lym-phocytes compared to patients without arthritis (Z=-2.772,P=0.006).The expression of P2X7R on lymphocytes and CD19+ cell was significantly positively correlated with the SLEDAI score.Positive correlation with anti-β2GP Ⅰ in lymphocyteswas also found.Conclusion P2X7R may mediate the release of inflammatory cytokines involved in the pathogenesis of SLE,and may participate the development of arthritis,lupus nephritis and NPSLE in SLE patients.
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Objective To investigate the expression of micro RNA-146a (miR-146a),TNF receptorassociated factor 6 (TRAF6) gene and IL-1 receptor-associated kinase 1 (IRAK1) gene in the peripheral mononuclear cells (PBMCs) of patients with ankylosing spondylitis (AS) and their relationship with the disease activity.The role of miR-146a,TRAF6,IRAK1 in the pathogenesis of AS was explored.Methods Expression of miR-146a,TRAF-6 and IRAK-1 in peripheral blood mononuclear cells was studied using realtime polymerase chain reaction (qRT-PCR) in 45 AS patients and 22 healthy controls.The indicators of disease activity adopted in this study were Bath ankylosing spondylitis disease activity index (BASDAI),erythrocyte sedimentation rate (ESR),C-reactive protein (CRP) level,and immunoglobulin (Ig).The relationship was analyzed in AS patients between the relative expression levels miR-146a,TRAF6,IRAK1 and BASDAI,ESR,CRP,Ig concentration.Non-parametric test,t test,One-way ANOVA,Pearson's and Spearman's correlation analysis were used for statistical analysis.Results ①The relative expression level of miR-146a which was observed in PBMCs of AS patients was significantly higher than that in normal control group [1.46(0.39,4.79)and 0.81(0.17,1.90),P＜0.05].The expression of miR-146a was significantly higher in active AS patients group than that in inactive patients [2.93(0.95,7.95) and 0.54(0.28,1.69),P＜0.05],there was no difference between the treatment group and without treatment group [1.28(0.31,2.37) and 2.22(0.49,7.71),P＞0.05].② There was significant difference in the relative expression level of IRAK-1 between AS patients and the normal control group.IRAK1 was significantly higher in AS patients than that in normal control group (1.4±0.7,1.1±0.4,P＜0.05).However,there was not difference between active AS patients group and inactive patients group as well as treated group and untreated group (1.5±0.9,1.4±0.5; 1.6±0.7,1.3±0.7,P＞0.05).③ TRAF6 expression was obviously lower in AS patients than that in normal control group (1.3±0.6,1.7±0.8,P＜0.05),and that was also significantly lower in the untreated group and active group than that in the normal control group (1.1±0.7,1.7±0.8; 1.1±0.5,1.7±0.8,P＜0.05).④ Signi-ficant positive correlation was observed between the miR-146a level and BASDAI,as well as duration of morning stiffness (r=0.557,P=0.000; r=0.363,P=0.018).The expression level of IRAK1 was significantly negative correlated with IgM (r=-0.313,P=0.046).Conclusion ① miR-146a expression is up-regulated in patients with AS,and it may be a potential useful marker for disease activity in AS patients; ② The abnormal expression of IRAK1,TRAF6 in AS patients may play a role in the pathogenesis of AS.
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Objective To enhance the understanding of systemic lupus erythematosus (SLE) associated with intestinal pseudo-obstruction (IPO) and ureterohydronephrosis. Methods Ten cases of SLE with IPO and ureterohydronephrosis were analyzed retrospectively. Results IPO was the initial manifestation of SLE in three patients. Seven cases (7/9) had concurrent bilateral ureterohydronephrosis, 5 (5/9) had thickened bladder wall, and 6 cases (6/8) were associated with thickened gallbladder wall simultaneously. All cases were ANA positive and hypocomplementemia, 6 had positive anti-dsDNA, 5 had positive anti-SSA, 6 had positive anti-RNP, and 4 were ACA-IgG positive (4/7). IPO responded readily to high dose steroid therapy within one week in all patients. Conclusions The pathogenic mechanism of IPO is not fully understood. The coexistence of IPO, ureterohydronephrosis and thickened gallbladder wall probably suggests a visceral smooth muscle dysmotility. Early recognition of the syndrome is necessary for the initiation of the appropriate medical treatment and prevention of inappropriate surgical intervention.