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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 therapeutic effects of leflunomide on salivary gland secretion hypofunction in NOD mice with Sjogren′s syndrome.Methods:NOD mice were randomly divided into four groups: prevention group, prevention control group, treatment group and treatment control group. Salivary flow rate was measured after pilocarpine stimulation. Changes in the average number and area of infiltrating lesions were compared after hematoxylin and eosin (HE) staining. The percentages of CD3 + T, CD4 + T, CD8 + T, CD44 + CD62L -CD4 + T, CD19 + B and CD138 + B cells in submandibular gland and spleen were detected by flow cytometry. The levels of TNF-α, IL-17A and IL-6 in serum were detected by CBA method. Results:The salivary flow rate ( t=-5.81, P<0.001; t=-3.61, P<0.05), the number of infiltrating lesions( t=3.95, P<0.01; t=4.94, P<0.001)and the average area of infiltrating lesions( t=3.18, P<0.05; t=2.35, P<0.05)were significantly ameliorated in the prevention and treatment groups. Moreover, CD3 + CD4 + T cells( t=2.39, P<0.05; t=3.82, P<0.01)and CD44 + CD62L -CD4 + T cells( t=3.53, P<0.05; t=3.36, P<0.05)in the submandibular gland were significantly decreased. CD3 + T( t=6.08, P<0.001; t=2.76, P<0.05), CD3 + CD4 + T ( t=3.73, P<0.05; t=2.39, P<0.05), CD19 + B ( t=5.88, P<0.001; t=4.23, P<0.01) and CD138 + B cells ( t=4.30, P<0.001; t=4.46, P<0.01) in the spleen were also significantly reduced. Serum IL-17A ( t=4.15, P<0.01; t=3.36, P<0.01) in the two groups and TNF-α ( t=4.56, P<0.001) in the prevention group were down-regulated, but no significant difference was observed in IL-6 level. Conclusions:This study suggested that leflunomide could prevent and improve salivary gland hypofunction and inhibit immune activation in NOD mice, providing reference for evaluating leflunomide in the treatment of Sjogren′s syndrome.
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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 improve the comprehensive understanding and treatment level of the disease by analyzing the clinical characteristics of rheumatoid arthritis (RA) caused by pauciarticular arthritis.Methods The method was retrospective analysis and summary of clinical and laboratory data of 198 cases of patients with RA,in which 98 cases of pauciarticular arthritis belonged to the observation group and 100 cases of polyarticular arthritis belonged to the control group.Results Male patients in observation group were obviously more than those in the control group (t =2.456,P =0.015).Courses of disease was obviously shorter than those in the control group (t =-2.450,P =0.018).The number of involved joints was obviously less than those in the control group (t =-6.316,P ＜0.001).The incidence of morning.stiffness was significantly less than those in the control group (t =-3.884,P ＜ 0.001).Rating scores of disease activity score 28 were significantly lower than those in the control group (t =-8.694,P ＜ 0.001).And positive rate of anti-cyslic citrullinated peptide antibody is significantly higher than those in the control group (t =-2.299,P =0.022).Thyrotrophin levels were significantly higher than those in the control group (t =3.809,P ＜ 0.001).There was no significant differences in age,erythrocyte sedimentation rate and C reactive protein,immunoglobulin and complement level,free triiodothyronine,free thyroxine,blood lipids,blood system and kidney involvement between the two groups (all P ＞ 0.05).Conclusions The conclusion turns out to be that there are no typical clinical manifestations showing that rheumatoid arthrifts is caused by pauciarticular arthritis.Maybe it is the early performance of disease.It is worth attention that concurrent subclinical hypothyroidism turns out to be more.
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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 investigate the plasma vitamin D3 level and its association with B cell subsets of patients with primary Sj(o)gren's syndrome (pSS).The role of vitamin D3 levels in the pathogenesis of pSS was explored.Methods The expression of plasma vitamin D3 levels of 55 patients with pSS and 32 controls were analyzed by ELISA.Frequencies of peripheral blood CD19+CD27-na(i)ve B cells,CD19+CD27+ memory B cells and CD19+CD27high plasma cells were analyzed by flow cytometry in 34 pSS patients without therapy and 22 controls.The relationship between the vitamin D3 levels and B cell subsets,SSDAI,tear flow rate,saliva flow rate,rheumatoid factor,immunoglobulin was analyzed in pSS patients.Non-parametric test,t test,one-way ANOVA,x2test,Pearman's and Spearman's correlation analysis were used for statistical analysis.Results ① There was significant difference in the levels of plasma 1,25 (OH)2D3 between the pSS patients group and normal control group,1,25 (OH)2D3 was significantly lower in pSS patients than that in the normal control group [24.17(22.20,28.41) pg/ml and 41.25(23.38,62.18) pg/ml,P＜0.05],and that was also obviously lower in the active group [22.64(20.74,24.90) pg/ml] than that in the normal control group (P＜0.05),and that was also obviously lower in the active disease group than that in the inactive disease group [25.39 (23.16,33.09) pg/ml,P＜0.05],but there was no difference between the inactive group and the normal control group (P＞0.05).② The percentage of peripheral blood of CD19+CD27high plasma cells and CD19+CD27+ memory B cells in CD19+ cells was reduced in patients in the pSS group compared with the control group [(0.89±0.30)％ and (1.72±0.43)％,(24±8)％ and (34±5)％; P＜0.05],and that was also significantly lower in the active group [(1.03±0.59) ％ ; (26± 10)％] and inactive group [(1.00±0.16)％,(26± 3)％] than that in the normal controls (P＜0.05).However,there was no difference between the active group and the inactive group (P＞0.05),but the frequency of peripheral blood of CD19+ CD27-naive B cells in CD19+ B cells was increased in patients with pSS compared with normal control group [(75.4±7.5)％ and (63.9±5.2)％,P＜0.05],and that was also significantly higher in the active group [(73.4±9.7)％] and inactive group [(73.3±2.9)％] than that in the normal control (P＜0.05),there was no difference between the active group and the inactive group.③ Significant negative correlation was observed between 1,25 (OH)2D3 and the percentage of peripheral blood CD19+CD27+ memory B cells in CD19+ cells as well as immunoglobulin G(r=-0.627,P=0.039; r=-0.657,P＜0.01) level.Conclusions These results demonstrate that abnormality of vitamin D levels may play an important role in the pathogenesis of pSS.
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Objective To study the pathophysiology of ankylosing spondylitis (AS)-related osteoporosis by investigating the relation between bone mineral density (BMD) and bone turnover markers.Methods Fortysix AS patients were included in this study,including 35 male and 11 female patients.Twenty-five gender and age matched healthy subjects were served as the healthy control group.Informed consents were obtained from all subjects.BMD of lumbar spine (L2-4),fermoral neck and radius were tested using dual-energy X-ray absorption method.Data about BASDAi,ESR,Ig was collected.Bone formation markers including procollagen type Ⅰ N-terminal peptide (PINP) and osteocalcin (OC) were included for analysis,the formal was measured by radioimmunoassay and the later was measured by immunoradiometric assay and bone resorption marker.β-Crosslaps was measured by electrochemiluminescence immunoassay.Independent-samples t test was used to compare normal distributed data between the two groups.Analysis of variance was used to compare the data between the three groups.For those data which were not normally distributed,Rank sum test was performed.Correlations were determined by Pearson or Spearman's ranking.Results ① 48％ of AS patients had bone loss,while the percentage in the healthy control group was 20％ (x2=5.32,P=0.021).BMD of lumbar spine,fermoral neck and radius of the AS patients were lower than the controls (t=-3.73,-3.68,-5.24; P＜0.05).② Bone density (BMD) of lumbar spine in patients at the late stage was higher than patients at early disease stage (t=2.26,P=0.034),however,the BMD in the femoral neck was not.③ The procollagen (PINP),OC and β-CrossLaps were not significantly different in patients at different stage of diseases (P＞0.05).④BMD of lumbar spine was negatively correlated with the β-CrossLaps and ESR(r=-0.325,P=0.047; r=-0.314,P=0.046),The BMD in fermoral neck was negatively correlated with β-CrossLaps and OC (r=-0.387,P=0.024; r=-0.371,P=0.034),but they were not correlated with disease duration and BADSAI.Conclusion Osteoporosis is common in AS.Bone turnover markers including bone formation and bone resorption are increased in AS.β-CrossLaps is likely to predict bone loss in AS.
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Objective To explore the probable function of peptidyl arginine deiminase 4 (PAD4) in rheumatoid arthritis(RA).Methods Real-time quantitative polymerase chain reaction (PCR) was used to determine the expression of PAD4 mRNA in peripheral blood mononucleated cells (PBMCs) from 60 RA patients and 40 healthy individuals.Asymmetric di-methylation of histone H3R17,symmetric di-methylation and mono-methylation of H4R3 were semi-quantified by Western blotting in 12 patients with osteoarthritis (OA),26 patients with RA and 10 healthy controls.Results PAD4 mRNA in RA was significantly higher than that in healthy controls [34.6 (16.7,70.8) vs 20.6 (11.1,51.8),P ＜ 0.05].The level of histone H3R17 asymmetric di-methylation in RA was significantly higher than that of OA or control groups(71.34 ±25.65 vs 37.18 ± 18.62 vs 50.67 ± 13.99,P ＜0.01),which was positively related to Tender joint count and Swollen joint count in 28 joints (r =0.418,P =0.034 ; r =0.402,P =0.042).The level of histone H4R3 symmetric di-methylation was similar in RA,OA and control groups (75.02 ± 20.35 vs 57.92 ± 22.77 vs 68.37 ± 17.57,P ＞ 0.05).The level of histone H4R3 mono-methylation in RA patients was significantly lower than that of OA patients and healthy individuals (11.24 ±7.81 vs 32.77 ±30.77 vs 51.20 ±47.14,P ＜ 0.05).The level of histone H4R3 mono-methylation in RA patients was negatively correlated to PAD4 (r =-0.643,P ＜ 0.01).The level of histone H3R17 asymmetric di-methylation and H4R3 symmetric di-methylation was not associated with PAD4 level in RA group (r =-0.185,P =0.377; r =0.198,P =0.344).Conclusions The level of histone H3R17 asymmetric di-methylation is significantly higher and the level of histone H4R3 mono-methylation is significantly lower in RA patients comparing with OA and control groups.Abnormality of histone methylation may be one of the mechanisms for the development of RA.PAD4 probably plays an important role in rheumatoid arthritis by influencing histone methylation.
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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 effect of total glucosides of paeony(TGP) on hepatic dysfunction caused by Methotrexate (MTX) and Leflunomide (LEF) in patients with moderate or severe rheumatoid arthritis (RA).Methods From July 2010 to February 2011,204 cases with definite-diagnosed rheumatoid arthritis were included in three hospitals in Anhui province.All these patients suffered from moderate or severe rheumatoid arthritis and they were divided randomly into two groups,the therapeutic groups were treated with TGP combined with MTX and LEF and the control group were treated without TGP.The incidence of hepatic dysfunction was observed.Statistical anylysis was carried out by using t test and x2 test.Results One hundred and ninty-four patients had completed observation for at least 12 weeks.In the therapeutic group,only 10 of 105 patients had hepatic dysfunction compared to 31 of 89 patients in the control group.The incidence of hepatic dysfunction of the therapeutic group was significantly lower than the control group.After 12 weeks treatment with TGP,the effective respohse rates of the therapeutic group was 81.9％,which was higher than 74.6％ in the control group.The good response rate in the therapeutic group was 21.6％,which was better than 11.7％ in the control group.Although the difference between the two groups was not statistically significant,we could observed the tendency of increasing in effectiveness and the good response rate in the therapeutic group when compared with the control group.Conclusion TGP can decrease the incidence of hepatic dysfunction during the treatment with MTX and LEF,and it may improve the therapeutic effecacy of patients with RA.
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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 investigate the changes of plasmacytoid dendritic cells (pDC) in peripheral blood of patients with systemic lupus erythematosus(SLE) and its roles in the pathogenesis of SLE.Methods The level of pDC and the expressions of CD32,CD40,CD86,CD62L and CXCR4 were analyzed by flow cytometry.The concentrations of IFN-α in serum were detected by ELISA assay.Results The levels of circulating pDC were significantly decreased in SLE patients compared with healthy controls.Moreover,the pDC levels in active SLE patients were lower than those in inactive SLE patients,and compared with the primary group,the pDC levels were increased in the treatment group.The levels of pDC showed a significant decrease in SLE patients with arthritis,proteinuria or leucopenia in comparison with patients without those manifestations,showing a negative correlation with proteinuria.The expressions of cell surface molecules including CD32,CD86,CD62L and CXCR4 on pDCs were significantly increased in SLE patients compared with healthy controls,and the levels of pDC were negatively correlated with the expressions of CD32 and CXCR4 in patients with SLE.The concentrations of IFN-α in serum of patients with SLE were significantly higher than those in healthy controls,and the levels of pDC were positively correlated with the concentrations of IFN-α in patients with SLE.Conclusion The level of circulating pDC in patients with SLE was remarkably reduced,but the expressions of molecules involved in cell activation and migration were upregulated,accompanied by enhanced IFN-α production,which might promote the onset and progression of SLE.
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Objective To identify interleukin 17 (IL-17) and B cell activating factor (BAFF) that could influence B cell biology by detecting the expression of BAFF in the serum and labial salivary glands from primary Sj(o)gren's syndrome (pSS) patients and to test the apoptosis rates of B cells cultured with Th17 cells which were transfected with IL-17-siRNA,BAFF-siRNA.Methods A total of 40 patients with pSS who were referred to the Department of Rheumatology and Immunology at Anhui Provincial Hospital from June 2011 to June 2012 were enrolled into this study.The expression of BAFF on salivary gland and serum from pSS patients and healthy controls were detected by ELISA and immunohistochemical examination (22 patients with pSS).Flow cytometry was used to detect B cell's apoptosis,BAFF and IL-17 interfered with amplified Th17 cells,and co-cultured with B cells.Immunoblot was used to detect supernatant antibody in 5 patients with pSS.Independent samples t test was used for statistical analysis.Results In all pSS specimens,infiltrating inflammatory cells expressed BAFF,so did some ductual cells,but acinar cells did not express these markers.There was no expression of BAFF in the controls.BAFF-positive cell numbers in the labial salivary glands of pSS patients with focal infiltrating lymphocytes were more than that with non-focal infiltrating lymphocytes (888±372 vs 164±161,t=5.94,P＜0.05),and the percentage of BAFF-positive lymphocytes over the total infiltrating lymphocytes in the salivary glands of pSS patients with focal infiltrating lymphocytes [(0.18 ±0.08) ％] was higher than those with non-focal infiltrating lymphocytes [(0.09 ±0.07) ％] (t =3.03,P＜0.05).The level of soluble BAFF in patients with pSS [(6.0±2.8) ng/ml] was significantly higher than the controls [(3.8±1.7) ng/ml,t=3.26,P＜0.05].BAFF or IL-17 transfected group,B cell apoptosis rate [(24± 5)％,(23±5)％] were significantly higher than the non-transfected group [(7±4)％],t=4.6,4.4; P＜0.05].And there was no significant difference when compared with cultured B cells (P＞0.05).Compared with the controls,no antibody could be detected in the supernatants.Conclusion BAFF may be involved in the process of local inflammatory damage of the pSS,it may have a synergistic effect with IL-17 on abnormal B cell function.