系统性红斑狼疮患者外周血单个核细胞骨调素表达及临床意义

目的 探讨骨调素在系统性红斑狼疮（ＳＬＥ）患者中的表达及临床意义。方法 用流式细胞仪和Ｎｏｒｔｈｅｒｎ ｂｌｏｔ杂交方法分别检测ＳＬＥ患者外周血单个核细胞（ＰＢＭＣ）ＯＰＮ蛋白和ｍｔｎＡ的表达。结果 活动期和稳定期ＳＬＥ患者ＰＢＭＣ ＯＰＮ蛋白和ｍＲＮＡ的表达均高于正常人（Ｐ〈０．０１）,ＳＬＥ患者ＯＰＮ蛋白主要在淋巴细胞表达,而活动期患者ＰＢＭＣ ＯＰＮ蛋白及ｍＲＮＡ的表达高于稳定期患者（Ｐ〈０     

外周血单个核细胞巨噬细胞移动抑制因子表达及其与狼疮活动

目的 观察系统性红斑狼疮(SLE)患者外周血单个核细胞(PBMC)巨噬细胞移动抑制因子(MIF)mRNA表达及血MIF水平的变化,并分析其与SLE疾病活动性、血清免疫学指标的相关关系。方法 采用逆转录-PCR技术及ELISA方法分别测定34例活动期和21例静止期SLE患者的PBMC中MIF mRNA表达及血清MIF浓度,并以18例正常人作对照,观察SLE患者血MIF的改变及其与疾病活动性、血清免疫学指标的相关关系。结果 SLE患者PBMC中MIF mRNA表达及血MIF水平较正常人显著升高(P<0.01),且活动期患者增高更为明显,显著高于静止期患者(P<0.01)。SLE患者血MIF水平及PBMC中MIF mRNA表达与疾病活动指数、抗ds-DNA抗体、血补体C3、C4及IgG水平明显相关,但与抗核抗体无关。结论 SLE患者血MIF mRNA表达及血MIF水平显著增高,其变化可反映SLE患者疾病的活动性。     

淋巴细胞趋化因子受体mRNA在系统性红斑狼疮患者外周血单个核细胞的表达

目的探讨淋巴细胞趋化因子受体(XCR1)mRNA在系统性红斑狼疮(SLE)患者外周血单个核细胞(PBMC)中的表达,及其与疾病的相关性。方法收集93例确诊的SLE患者和30名健康对照组,收集PBMC,提取RNA。应用反转录-聚合酶链反应(RT-PCR)方法检测研究对象XCR1mRNA表达水平。结果!"XCR1mRNA在PBMC中表达水平,患者组(1.57±0.84)与正常对照组(0.19±0.14),两者差异无统计学意义(P>0.05)。活动期(2.09±1.38)与对照组比较,两者差异有统计学意义(t=2.392,P=0.02);活动期与非活动期(0.31±0.24)比较,差异有统计学意义(t=3.091,P=0.003);非活动期与对照组比较,差异无统计学意义(P>0.05)。"#SLE患者组XCR1mRNA水平与疾病活动度评分(SLEDAI)关系:SLE患者组PBMC中XCR1mRNA表达水平与SLEDAI作直线相关性分析,XCR1mRNA水平与SLEDAI(r=0.540,t=5.445,P=0.000)呈正相关。"$SLE患者PBMC中XCR1mRNA表达与临床表现及实验室检查相关性:SLE抗SSA抗体阳性患者(17/68,1.4±1.2)比阴性患者(51/68,3.5±4.9),其XCR1mRNA表达水平降低,差异有统计学意义(t=2.78,P=0.007)。结论XCR1mRNA表达水平在PBMC中活动期SLE患者比非活动期及对照组增高,与疾病的活动性(SLEDAI)正相关,非活动期与对照组差异无统计学意义。SLE抗SSA抗体阳性患者比阴性患者,其XCR1mRNA表达水平降低,两组均较健康对照组增高。提示XCR1参与疾病的发展,与疾病的活动性相关,XCR1可能参与抗SSA抗体的形成过程,造成组织损伤。     

系统性红斑狼疮患者外周血单个核细胞CD28 mRNA的表达

目的　探讨CD28在系统性红斑狼疮(SLE)患者外周血单个核细胞(PBMC)中的表达水平及其意义。方法　应用反转录-聚合酶链反应(RT-PCR)检测了34例活动期SLE患者和30名正常人PBMC中CD28mRNA的表达水平。结果　活动期SLE患者CD28的阳性表达率为20.6%,明显低于正常人对照组(70.0%),差异非常显著(P＜0.001);活动期SLE组CD28的平均表达水平(0.19±0.21)亦明显低于正常对照组(0.43±0.11),差异显著(P＜0.05)。结论　CD28的异常表达可能在SLE发病机制中起作用,CD28mRNA的低水平表达可能与外周血CD28＋T细胞凋亡增加或迁移到炎症部位有关。     

FOXO1和FOXO3a在系统性红斑狼疮患者外周血单个核细胞中的表达及其与疾病活动性的关系

目的 探讨转录因子FOXO1和FOXO3a在系统性红斑狼疮(SLE)患者外周血单个核细胞(PBMC)中的表达水平及其与SLE疾病活动性的关系.方法 选择SLE活动性、SLE非活动性患者和正常对照,运用反转录一聚合酶链反应(RT-PCR)和Western blot检测SLE患者和健康对照组PBMC中FOXO1和FOXO3a mRNA及蛋白的表达水平.结果 活动性SLE患者PBMC中FOXO1 mRNA和蛋白的表达水平明显低于SLE非活动性患者和正常对照(P<0.05),非活动性SLE患者FOXO1 mRNA和蛋白的表达水平明显低于正常对照(P<0.05).FOX03a mRNA和蛋白的表达水平在3组间差异无统计学意义.FOXO1 mRNA水平与狼疮活动指数(SLEDAI)呈显著负相关(r=-0.651,P<0.05).结论 SLE患者PBMC中FOXO1 mRNA和蛋白的表达水平显著低于健康对照,随着SLE患者病情的好转,FOXO1 mRNA和蛋白的表达水平逐渐增高.FOXO1与SLE疾病的活动性呈负相关.FOXO3a与SLE疾病的活动性无关.     

类风湿关节炎外周血单个核细胞和滑膜中人类软骨糖蛋白-39的研究

目的　通过观察人类软骨糖蛋白 39(HCgp 39)mRNA在类风湿关节炎 (RA)外周血单个核细胞 (PBMC)和滑膜中的表达水平 ,寻找反映RA早期骨破坏的敏感指标。方法　采集 31例RA、6例骨关节炎 (OA)、10例血清阴性脊柱关节病 (SpA)、5例系统性红斑狼疮 (SLE)以及 10例健康人新鲜抗凝静脉血 ,分离单个核细胞 ,并采集 7例RA、5例OA的滑膜 ,行半定量逆转录 (RT) PCR。结果　RA患者PBMC中HCgp 39mRNA行RT PCR与内对照tubulin共扩增后吸光度比值为0 86 90± 0 5 2 4 0 ,与OA(P =0 0 2 4 )、SpA(P =0 0 4 9)、SLE(P =0 0 4 3)以及健康对照 (P =0 0 33)相比差异均有显著性。而OA、SpA、SLE与健康对照之间差异无显著性。RA滑膜HCgp 39mRNA行RT PCR与tubulin共扩增后吸光度比值为 1 986 3± 1 9397,与OA相比差异有显著性 (P =0 0 4 )。在RA和OA中 ,同一患者的PBMC与滑膜HCgp 39mRNA表达差异无显著性。 结论　HCgp 39mRNA在RA患者PBMC和滑膜的表达明显高于其他炎症性关节病 ,提示HCgp 39可能是RA发病机制中重要的自身抗原 ,它的异常表达可能是造成T细胞介导自身免疫反应的机制之一。     

系统性红斑狼疮患者外周血单个核细胞环氧合酶同工酶mRNA的表达

目的　探讨原生型环氧合酶 (COX 1)和诱生型环氧合酶 (COX 2 )在系统性红斑狼疮(SLE)患者外周血单个核细胞 (PBMC)中的表达水平及其在SLE发病机制中的作用及其意义。方法　应用逆转录 聚合酶链反应 (RT PCR)检测了 34例活动期SLE患者和 30名正常人PBMC中COX 1和COX 2mRNA的表达水平。结果　活动期SLE患者COX 1和COX 2的阳性表达率与正常对照组相比差异无显著性 (P >0 0 5 ) ;活动期SLE患者PBMC中COX 2mRNA的平均表达水平 (0 6 6±0 2 7)明显高于正常对照组 (0 43± 0 16 ) ,差异有非常显著性 (P <0 0 1) ;活动期SLE组COX 1的平均表达水平与正常对照组处于同一表达水平 ,差异性无统计学意义 (P >0 0 5 )。结论　活动期SLE患者外周血单个核细胞中COX 2mRNA的表达增加 ,通过诱导前列腺素 (PGs)的产生参与SLE的发病过程。为特异性COX 2抑制剂应用于SLE的治疗提供了理论依据。     

In vitro apoptosis and expression of apoptosis-related molecules in lymphocytes from patients with systemic lupus erythematosus and other autoimmune diseases

Objective. To analyze factors related to apoptosis in systemic lupus erythematosus (SLE) peripheral blood mononuclear cells (PBMC) and to compare the findings in SLE PBMC with those in normal donor PBMC or PBMC from patients with other autoimmune diseases. Methods. PBMC from normal healthy donors or patients with SLE, mixed connective tissue disease (MCTD), rheumatoid arthritis (RA), or various vasculitides were isolated. The percentage of apoptosis after activation through different signaling pathways was quantified using propidium iodide staining. Protein expression of Fas/APO-1 or bcl-2, and messenger RNA (mRNA) expression of bcl-2, bcl-x_L, bax, bak, Fas/APO-1, Fas ligand (Fas-L), c-myc, mad, or max were determined. Results. We confirmed previous findings of increased numbers of apoptotic cells in SLE PBMC compared with normal donor cells after in vitro incubation. After activation of PBMC with CD28 monoclonal antibody plus phorbol myristate acetate (CD28 MAb/PMA), staphylococcal enterotoxin B (SEB), or phytohemagglutinin (PHA), the percentage of apoptotic cells was unchanged (SEB) or diminished (CD28 MAb/PMA, PHA) in SLE cells, and the difference between normal donor and SLE cells was less pronounced. On the mRNA level, expression of apoptosis-related gene products did not differ between SLE cells and normal donor cells. Expression of Fas/APO-1 protein was increased in freshly isolated SLE T lymphocytes compared with normal donor T lymphocytes, whereas bcl-2 protein was up-regulated after a 3-day culture period. Cellular activation further increased bcl-2 protein levels, eliminating differences between normal donors and SLE patients. In RA cells, the percentage of apoptosis was similar to that in normal donor PBMC, whereas results using cells from patients with other autoimmune diseases (MCTD, Wegener's granulomatosis, Takayasu arteritis, polyarteritis nodosa) were comparable with those found using SLE PBMC. Addition of growth factors such as interleukin-2 (IL-2), IL-4, or IL-15 to culture medium decreased the percentage of in vitro apoptosis in both normal donor and SLE cells. Conclusion. Based on these data, we conclude that accelerated in vitro apoptosis and increased Fas/APO-1 and bcl-2 protein expression in SLE are nonspecific for the disease, and might be explained at least in part by the increased in vivo activation levels of PBMC from patients with SLE, MCTD, or autoimmune vasculitides combined with in vitro incubation under “noninflammatory” conditions and growth factor withdrawal.     

SLE患者外周血CD4+ CD25+调节性T细胞及相关基因Foxp3的变化

目的研究系统性红斑狼疮（SLE）合并狼疮性肾炎患者经肾上腺糖皮质激素（简称激素）冲击治疗后，外周血CD4^＋CD25^＋调节性T细胞（regulatoryTcell，Treg）及相关基因Foxp3表达的变化，从而探讨CD4^＋CD25^＋Treg和Foxp3与SLE发病的相关性。方法采用流式细胞术检测SLE患者外周血单个核细胞（PBMC）中CD4^＋CD25^＋T、CD4^＋CD25highT细胞数量的变化，RTPCR检测PBMC中CD4^＋CD25^＋Treg功能相关基因Foxp3mRNA的表达。结果激素冲击治疗后的SLE患者CD4^＋CD25^＋T／CD4^＋T及CD4^＋CD25highT／CD4^＋T比值高于正常对照组；Foxp3mRNA水平表达与对照组差异不显著。结论CD4^＋CD25^＋Treg数量和功能的变化可能参与SLE的发病，激素可能通过提升CD4^＋CD25^＋Treg治疗SLE。     

PDCD5在系统性红斑狼疮患者外周血单个核细胞中表达的初步研究

目的研究程序性死亡因子5（PDCD5）在系统性红斑狼疮（SLE）患者外周血单个核细胞（PBMC）中的表达情况,探讨其可能作用机制。方法应用免疫细胞化学和蛋白免疫印迹法检测45例SLE活动期患者、22例稳定期患者和50例健康对照者PBMC中PDCD5蛋白的细胞定位和表达情况。结果SLE活动期患者PDCD5蛋白的阳性表达率高于正常对照组（P〈0.01）;活动期患者PDCD5蛋白的表达水平高于稳定期患者和正常对照,稳定期患者高于正常对照（P〈0.01）。SLE活动期患者PBMC中PDCD5的表达与SLE疾病活动指标SLEDAI、补体C3、抗ds-DNA抗体滴度具有相关性。结论PDCD5可能作为淋巴细胞凋亡的正性调控基因,在SLE发生和发展中具有重要意义。     

Dysregulated expression of interleukin-23 and interleukin-12 subunits in systemic lupus erythematosus patients

The aim of this study was to investigate the regulation of interleukin (IL)-12 and IL-23 expression in the autoimmune disease, systemic lupus erythematosus (SLE). mRNA from healthy subjects and SLE patients were prepared from peripheral blood mononuclear cells (PBMC) and quantitative real-time polymerase chain reaction was performed to quantify IL-23 specific subunit P19, IL-12 specific subunit P35, and their common subunit P40. IL-12 specific subunit P35 mRNA expression in untreated and treated SLE patients was significantly lower than healthy controls (P = 0.015 and 0.000, respectively). Compared with untreated SLE patients, treatment of SLE patients with corticosteroids or corticosteroids plus another immunosuppressor significantly suppressed P40 and P19 expression (P = 0.002 and 0.015, respectively). The mRNA levels of p19, p40, and p35 in active SLE patients (SLEDAT > 10) were significantly higher compared with those in the inactive SLE patients (SLEDAI ≤ 10) (P = 0.000, 0.000, and 0.017, respectively). These results suggest that deficiency of IL-12 and possibly upregulation of IL-23 may contribute to SLE pathogenesis and both cytokines may be therapeutic targets in SLE.     

白细胞介素-17在狼疮性肾炎外周血单个核细胞上的表达及意义

目的　探讨白细胞介素 - 17(IL - 17)在狼疮性肾炎患者 (LN)外周血单个核细胞 (PBMC)表达和分泌及其与LN疾病活动性的关系。方法　用酶联免疫吸附法 (ELISA)检测LN活动期患者血浆及PBMC体外自发培养或刺激培养的上清液中IL - 17蛋白水平 ,流式细胞仪免疫荧光分析法检测PBMC表达IL - 17的水平 ;用逆转录 -聚合酶联反应 (RT -PCR)法测定LN活动期患者PBMC体外自发培养和刺激培养后的IL - 17mRNA的表达水平。结果　LN活动期患者血浆检测不出IL - 17(<5ng/L) ;PBMC自发培养和PHA刺激培养下 ,LN活动期、LN静止期和正常对照 ,三组均检测不出IL - 17;而PBMC在CD3mAb +CD2 8mAb和PMA +ionomycin刺激培养下 ,较自发培养和PHA刺激培养 ,上述 3组IL - 17水平均明显升高 (P <0 .0 1) ,LN活动期PBMC表达、分泌IL - 17水平与SLE疾病活动指数 (SLEDAI)呈明显正相关。结论　LN活动期患者PBMC表达、分泌IL -17水平存在明显异常 ,并能反映LN疾病活动程度     

系统性红斑狼疮合并抗磷脂综合征患者外周血CD1d的表达

目的通过检测系统性红斑狼疮(SLE)合并抗磷脂综合征(APS)患者外周血单个核细胞(PBMC)CD1d表达,探讨CD1d分子和APS疾病和实验室指标的相关性。方法利用流式细胞分析方法对20例合并APS患者,30例SLE对照,23名正常对照PBMC CD1d表达进行分析,并与患者临床资料和实验结果进行相关分析。结果SLE合并APS组PBMC中CD1d表达高于正常对照组,和SLE对照组比较差异无统计学意义。SLE合并APS组单核细胞群中CD1d表达高于正常对照组,和SLE对照组比较差异无统计学意义。CD1d和SLE疾病活动指数(SLEDAI)评分显著相关,和补体明显呈负相关。结论CD1d表达异常与SLE发病相关,并可作为评价SLE活动的指标。     

甲状腺肿瘤组织中雌激素受体β1和β2的表达及其意义

采用RT-PCR检测10例甲状腺乳头状腺癌(PTC)和10例癌旁正常甲状腺组织雌激素受体(ER)β1、ERβ2以及ERα的表达,同时用免疫组织化学方法检测107例甲状腺癌、56例甲状腺腺瘤和10例正常甲状腺组织中ERβ1和ERβ2表达,并结合甲状腺癌ERα表达及其它主要临床病理参数进行分析.结果显示,ERα mRNA在PTC组织的表达明显高于癌旁正常组织,ERβ2 mRNA在PTC组织的表达明显低于癌旁正常组织(P＜0.05);正常甲状腺组织、甲状腺腺瘤和甲状腺癌中3种ER的表达差异均有统计学意义( P＜0.05);ERβ1和ERβ2在甲状腺癌中的表达与其组织类型有关;甲状腺组织中ERβ1和ERβ2的表达与ERα呈低、中度负相关(ERβ1:r=-0.206,P=0.042;ERβ2:r=-0.545,P＜0.01);ERβ2的表达还与淋巴结转移以及TNM分期有关.提示ERβ2的表达缺失可能导致甲状腺滤泡上皮细胞的增生和癌变以及肿瘤的演进,并且有一定的预后评估价值.     

紫外线暴露对系统性红斑狼疮患者DNA甲基化水平的影响

目的 研究紫外线照射对系统性红斑狼疮(SEE)患者DNA甲基化水平的影响,探讨其在SLE发病中的作用. 方法 提取45例SLE患者及20名健康对照的外周血单个核细胞(PBMC)进行培养,用不同剂量的311 nm窄谱中波紫外线(NB-UVB)照射培养的PBMC,高效毛细管电泳法(HPCE)检测基因组DNA甲基化水平,实时荧光定量聚合酶链反应(RT-PCR)法测定甲基化转移酶1(DNMT1) mRNA的表达.结果 SLE患者DNA甲基化水平低于健康对照,差异有统汁学意义(P=0.014);SLE患者经50,100mJ/cm2紫外线照射后DNA甲基化水平降低,差异有统计学意义(P＜0.01);SLE患者DNMT1 mRNA的表达与健康对照相比差异无统计学意义;紫外线照射后SLE患者DNMT1 mRNA的表达差异无统计学意义;SLE患者DNA甲基化水平与SLE疾病活动指数(SLEDAI)无相天性. 结论 紫外线暴露可改变SLE患者基因组DNA甲基化水平;紫外线可能通过改变DNA甲基化影响SLE.     

Dysfunction of TRIM21 in interferon signature of systemic lupus erythematosus

Abstract

Objectives: TRIM21 is an E3 ubiquitin ligase for interferon regulatory factors (IRFs) that are involved in innate and acquired immunity. Here, we evaluated the role of TRIM21 in the interferon (IFN) signature of systemic lupus erythematosus (SLE).

Methods: Twenty SLE patients and 24 healthy controls were enrolled in this study. We analyzed mRNA expression of TRIM21, type I IFN, and IFN-inducible genes in peripheral blood mononuclear cell (PBMC). The protein levels of IRFs were assessed by Western blotting in PBMCs cultured with or without MG-132.

Results: The expression of TRIM21 mRNA and protein was significantly higher in SLE PBMCs as compared to healthy controls. There was a correlation between TRIM21 mRNA expression and SLE activities. In contrast to a negative correlation between mRNA expression level of TRIM21 and those of type I IFNs in healthy controls, we found a positive correlation between them in anti-TRIM21 antibody-positive SLE patients. Neither positive nor negative correlation was observed in the autoantibody-negative SLE patients. Western-blotting analysis revealed impaired ubiquitin-dependent proteasomal degradation of IRFs in SLE PBMCs.

Conclusion: Our study showed ubiquitin-dependent proteasomal degradation of IRFs was impaired in anti-TRIM21 antibody-dependent and -independent fashions, leading to amplification of IFN signature in SLE.