Abnormalities in the content of nucleic acids of peripheral blood mononuclear cells from patients with systemic lupus erythematosus

We have completed 59 cytofluorographic studies of DNA/RNA content in acridine orange-stained peripheral blood mononuclear cells from 44 patients with systemic lupus erythematosus, most of whom received no medications. Most such cells were in resting phases of the cell cycle, particularly those from patients with inactive disease. Nine patients with systemic lupus erythematosus had increased percentages of these cells in the synthesis and postsynthesis phases of the cell cycle; the B lymphocyte had a greatest proportions of activated cells. In 11 patients, we found that cells, particularly T lymphocytes, had increased RNA content without a proportional increase in DNA. This DNA block occurred primarily in patients with serum antibodies to DNA and it could be reproduced in normal mitogen-stimulated mononuclear cells incubated in heat-inactivated sera from patients with systemic lupus erythematosus whose own cells showed abnormalities of DNA/RNA content or in purified native DNA antibody. The DNA blocking potential of the DNA antibody was dependent on its Fc portion and on the presence of Fc receptors on T cells. Thus, saturation of Fc receptors by pretreatment with aggregated IgG or incubation with the whole antibody in the cold prevented the DNA block, indicating that it was an active process.     

神经颗粒素在系统性红斑狼疮患者外周血单个核细胞中的表达

目的 探讨神经颗粒素在系统性红斑狼疮(SLE)患者和正常人外周血单个核细胞(PBMCs)的表达状态.方法 对比分析SLE PBMCs基因表达系列分析(SAGE)文库的高表达标签,并采用反转录聚合酶链反应(RT-PCR)法检测与正常人群有明显差异表达的神经颗粒素在SLE患者PBMCs的mRNA表达水平.结果 对比分析SLE PBMCs和正常人各类淋巴细胞SAGE文库显示:神经颗粒素标签仅异位高表达于SLE患者,而几乎不表达于正常人各类淋巴细胞.RT-PCR检测证实:活动组SLE患者神经颗粒素mRNA表达水平较正常对照明显增加(P＜O.001),而缓解组SLE患者其表达水平仅稍有增加(P＞O.05).结论 神经颗粒素作为一种前凋亡因子,高表达于SLE PBMCs,可能介导或参与SLE异常的免疫调节反应.     

Testosterone suppresses anti-DNA antibody production in peripheral blood mononuclear cells from patients with systemic lupus erythematosus

Objective. To study the in vitro effect of testosterone on anti-DNA antibody production in peripheral blood mononuclear cells (PBMC) from patients with systemic lupus erythematosus (SLE) in order to elucidate its regulatory role in SLE. Methods. PBMC from SLE patients were cultured with testosterone. IgG anti-double-stranded DNA (anti-dsDNA) antibody, total IgG, and cytokine activity in the supernatants were measured by enzyme-linked immunosorbent assay. Results. Testosterone suppressed both IgG anti-dsDNA antibody and total IgG production in PBMC from SLE patients. Antibody production in B cells was also suppressed by testosterone, although the magnitude of its effect on B cells was lower than that on PBMC. Interleukin-6 (IL-6) partially restored the testosterone-induced decrease in antibody levels in PBMC. Testosterone reduced IL-6 production in monocytes. Conclusion. These results suggest that testosterone may directly suppress anti-DNA antibody production in PBMC from SLE patients by inhibiting B cell hyperactivity and, indirectly, by down-regulating IL-6 production in monocytes. These results support the therapeutic effects of testosterone on SLE.     

Estrogen receptor expression by peripheral blood mononuclear cells of patients with systemic lupus erythematosus

To investigate the influence of sex hormones on the development of systemic lupus erythematosus (SLE), we examined the estrogen receptor (ER) expression by peripheral blood mononuclear cells (PBMC) in patients with SLE using the real-time quantitative polymerase chain reaction (TaqMan) method. The expression of messenger RNA (mRNA) for ER alpha (ERa) was increased and expression of ER beta (ERb) mRNA was decreased in PBMC from SLE patients compared with PBMC from normal controls. These findings may be useful for elucidation of the pathophysiology of 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。     

JAK2 and PTPRC mRNA expression in peripheral blood mononuclear cells from patients with systemic lupus erythematosus

Clinical Rheumatology - In this study, we aimed to explore the expression levels of JAK2 and PTPRC in peripheral blood mononuclear cells (PBMCs) from SLE patients and controls, detect the effects...     

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。     

A proteomic study of peripheral blood mononuclear cells in systemic lupus erythematosus

Our objective was to analyze the changes in the protein expression profiles of peripheral blood mononuclear cells (PBMCs) in patients with systemic lupus erythematosus (SLE). Peripheral blood was obtained from patients with SLE and healthy controls. 2-D gel electrophoresis was performed, and gels were silver-stained. Differentially expressed protein spots were detected, some of which were identified by MALDI-TOF spectrometry. Match rates of 71% +/- 4% and 72% +/- 4% were gotten for control and patient gels, respectively. 791 +/- 17 spots were detected for control gels and 781 +/- 17 for patient gels. Eleven protein spots were up-regulated, and 9 protein spots were down-regulated in patients with SLE. Five differentially expressed proteins were identified as immunoglobulin J chain, apolipoprotein A-IV precursor, calprotectin L1H and zinc finger protein subfamily 1A (all up-regulated) and glutathione S-transferase (down-regulated), some of which had previously been shown to play a potential role in the pathogenesis of SLE. We conclude there are significant changes in the 2-D maps of PBMCs in patients with SLE and applying this proteomic approach may be a useful way to gain novel insights into SLE.     

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。     

系统性红斑狼疮患者外周血单个核细胞蛋白质组学研究

目的 分析系统性红斑狼疮(SLE)患者外周血单个核细胞(PBMCs)蛋白质表达谱的变化.方法 分别抽取SLE患者及健康对照外周血,分离单个核细胞,抽提总蛋白,进行双向电泳、电泳胶银染显色,用ImageMaster 2D Platinum 5.0软件对获得的蛋白图谱进行分析,寻找差异表达的蛋白质.利用基质辅助激光解析电离飞行时间质谱鉴定差异蛋白点.结果 对照组凝胶蛋白点匹配率为(71±4)%,患者组匹配率为(72±4)%.对照组凝胶共检出蛋白点(791±17)个,患者组检出(781±17)个.有11个蛋白点在SLE患者组表达上调,9个表达下调,质谱分析共鉴定5个蛋白.以前的研究显示,我们鉴定的部分蛋白在SLE的发病机制中起潜在的作用.结论 SLE患者外周血单个核细胞蛋白质表达发生了明显改变,为我们从淋巴细胞蛋白质谱变化的整体角度上阐明SLE发生的分子机制及免疫调控通路奠定了基础.     

系统性红斑狼疮患者外周血单个核细胞CD226基因表达的研究

目的 研究CD226基因在系统性红斑狼疮(SLE)患者与健康人外周血单个核细胞(PBMCs)的表达及其与CD226-Gly307Ser多态性、疾病活动度的相关性.方法 应用实时荧光定量聚合酶链反应检测SLE患者组90例及健康对照组30名PBMCs CD226 mRNA表达水平,同时应用单因素方差分析CD226基因表达量在3种基因之间是否存在差异,并且探讨其与临床指标的相关性以及CD226-Gly307Ser多态性的3种基因型进行Pearson相关性分析.结果 SLE组与健康对照组相比,CD226基因表达水平明显降低(P＜0.01);SLE组3种基因型之间的CD226 mRNA表达量差异无统计学意义(6.8±1.1与26.5±6.7,P＞0.05);红细胞沉降率、尿蛋白定量(24 h)、抗核抗体滴度、SLE疾病活动指数(SLEDAI)及血清补体C3水平与CD226基因表达均没有相关性.结论 在中国湖北汉族人群中,CD226-Gly307Ser多态性与SLE发病相关;T危险等位基因未影响CD226的mRNA表达水平,CD226分子在自身免疫性疾病中作用需进一步探讨.

Abstract：

Objective To investigate the expression level of CD226 mRNA in the peripheral blood mononuclear cells (PBMCs) of patients with systemic lupus erythematosus (SLE) and explore the relation between the gene expression and disease activity, and the relation between the gene expression and Gly307Ser polymorphism of CD226 was also examined. Methods CD226 gene was measured with real-time polymerase chain reaction (qRT- PCR) in PBMCs. The expression levels of CD226 gene in PBMCs were compared between 90 SLE patients and 30 healthy individuals. One-way ANOVA and pearson correlation were used for statistical analysis. Results The expression level of CD226 in the PBMCs of SLE patients (6.8±1.1) was significantly decreased compared to healthy individuals (26.5±6.7) (P＜0.01), while there was no association between mRNA level and genotype (P＞0.05). No correlation between ESR, CRP, ANA, SLEDAI scores, C3 and the expression level of CD226 gene was discovered. Conclusion In Hubei Chinese Han population, CD226-Gly307Ser locus is associated with the development of SLE, while T allele does not impact the expression of CD226 gene, thus the role of CD226 gene in autoimmune diseases should be explored in the future.     

Interleukin-2 activated cells from peripheral blood of patients with systemic lupus erythematosus

The effect of recombinant interleukin-2 (IL-2) on proliferative and cytotoxic response of peripheral blood mononuclear cells (PBMC) was investigated in a group of 21 patients with systemic lupus erythematosus (SLE). There was a significant decrease in thymidine uptake of peripheral blood mononuclear cells when compared with controls. However, IL-2 induced cytotoxicity was not diminished and minimal frequencies of lymphokine-activated killer (LAK) cells precursors remained in the range of control group. These data provide an evidence about the dissection between proliferative and cytotoxic response of PBMCs to exogenous interleukin-2 in patients with SLE in vitro. The factors contributing to this effect and the clinical significance of these findings remains to be answered.     

Peptides from antibodies to DNA elicit cytokine release from peripheral blood mononuclear cells of patients with systemic lupus erythematosus: relation of cytokine pattern to disease duration

Peptides from VH regions of antibodies to DNA drive immune responses in systemic lupus erythematosus (SLE). We studied peptide-induced cytokine release by peripheral blood mononuclear cells (PBMC) of patients, the influence of peptide concentration, disease characteristics and HLA-D haplotypes. Cells secreting cytokines (IFNgamma, IL-2, IL-4 and IL-10) were measured by ELISPOT in PBMC from 31 patients with SLE and 20 matched healthy controls in response to seven peptides (A-G) from the CDR1/FR2 to CDR2/FR3 VH regions of human anti-DNA MAbs. Disease activity was assessed by SELENA-SLEDAI. HLA-DR and -DQ alleles were determined by molecular typing techniques. PBMC from significantly higher proportions of SLE patients than controls responded to VH peptides by generating IFNgamma and IL-10. Type of cytokines released in response to at least one peptide (D) depended on antigen concentration. Cytokine release was not associated with clinical features of SLE except for disease duration. A shift occurred from IFNgamma, IL-4 and IL-10 production in early disease to IL-4 and IL-10 in late disease (suggesting increasing TH2-like responses over time). Three peptides (B, D, G) were more stimulatory in the SLE patients than controls. Although none of the peptides was restricted by any particular MHC class II allele, among responders there was increased prevalence of HLA- DQB1*0201 and/or DRB1*0301, alleles known to predispose to SLE. Thus, responses to some VH peptides are more frequent in SLE and vary with disease duration. Increased responses in individuals with HLA class II genotypes that predispose to SLE suggest that peptide presentation by those molecules permits brisker peripheral blood cell responses to autoantibody peptides, thus increasing risk for disease.     

In vitro response to influenza immunisation by peripheral blood mononuclear cells from patients with systemic lupus erythematosus and other autoimmune diseases.

Reduced in vitro anti-influenza antibody response by peripheral blood mononuclear cells (PBMs) after vaccination was confirmed in a group of 28 patients with systemic lupus erythematosus (SLE), and also in 16 patients with some other autoimmune syndromes. This group of patients with SLE had higher serum anti-DNA binding, but there was no evidence of increased autoantibody production after vaccination, nor any clinical or laboratory evidence of flares in disease activity that are sometimes seen to follow intercurrent infection. Although a reduced in vitro antibody response may, to some extent, reflect redistribution of antibody producing cells, there appears to be more generalised impairment of the immune response in these patients, which cannot be accounted for by steroid/immunosuppressive therapy.     

MX homologous protein in mononuclear cells from patients with systemic lupus erythematosus

The 76-kd human interferon (IFN)-induced MX protein is the homolog to the murine protein, which is necessary and sufficient to provide adequate resistance to influenza virus infection in murine cells and in mice. Fifty-one patients with systemic lupus erythematosus (SLE) were screened for the presence of the MX homolog in mononuclear cells and for IFN and anti-IFN antibodies in serum. In 47 of 51 patients, significant levels of the MX homolog were found, while only 15 of 51 patients had measurable alpha-IFN in their serum. The IFN activity found in these sera was characterized as a partially acid-labile alpha-IFN, by means of acid-stability cross-reactivity on heterologous cells, trypsin sensitivity, and neutralization by homologous or heterologous antisera. Four of the patients had no detectable MX homolog in their leukocytes; 3 of these 4 possessed an anti-alpha-IFN antibody that was able to neutralize both a natural alpha-IFN preparation and the acid-labile IFN in SLE sera. Also, acid-labile alpha-IFN-containing SLE sera induced the MX homolog in vitro in mononuclear cells from healthy donors. These observations suggest that endogenously produced alpha-IFN is responsible for the observed induction of the MX homolog in SLE and that the IFN system is activated in more than 90% of SLE patients.     

Glucocorticosteroid dependent decrease in the activity of calcineurin in the peripheral blood mononuclear cells of patients with systemic lupus erythematosus

OBJECTIVE: To compare the activity of calcineurin in the peripheral blood mononuclear cells (PBMC) of 32 patients with systemic lupus erythematosus (SLE) and 35 healthy controls. METHODS: The activity of calcineurin was assayed in the supernatants of sonicated mononuclear cells. RESULTS: There was no significant difference in the calcineurin activity of patients with SLE not taking glucocorticosteroids (GCS) compared with the healthy controls. On the other hand, the activity of calcineurin was reduced in patients with SLE taking GCS, correlating negatively with the dose of GCS. The stimulation of PBMC by phorbol ester and calcium ionophore decreased the calcineurin activity both in patients with SLE and in healthy controls. GCS could also reduce calcineurin activity in the mononuclear cells of healthy subjects in vitro. CONCLUSIONS: In patients with SLE the decrease in the calcineurin activity of PBMC depended on the dose of GCS used for treatment, and it was not a disease specific alteration. The higher the dose of GCS, the greater the inhibition of calcineurin activity. The reduction of calcineurin activity is a new element in the immunosuppressive effects of GCS during the treatment of patients with SLE.     

系统性红斑狼疮外周血单个核细胞生长激素受体和mRNA表达的研究

目的　探讨SLE患者外周血单个核细胞 (PBMC)的生长激素受体 (GHR)的表达与系统性红斑狼疮 (SLE)的关系。方法　采用放射性受体配基结合试验 (RLBA)及反转录 聚合酶链反应(RT PCR)研究外周血淋巴细胞GHR变化。结果　SLE活动期患者PBMC的GHR的特异性结合率 (SB) (3 4± 2 0 ) %、总结合率 (TB) (13 8± 5 7) %和静止期患者SB (1 3± 1 2 ) % ,TB (11 4±4 6 ) %均高于正常对照组SB (1 0± 1 0 ) % ,TB (8 3± 0 9) % (P <0 0 1) ;SLE活动期患者PBMC的mRNA的表达水平 (0 77± 0 6 6 )高于静止期患者 (0 4 5± 0 2 8) (P <0 0 5 )和正常对照组PBMC的mRNA的表达 (0 31± 0 2 2 ) (P <0 0 1)。结论　SLE患者PBMC的GHR高表达与SLE的病情活动性相关。