系统性红斑狼疮患者16q12精细定位研究

目的预初研究提示16q12区与系统性红斑狼疮(SLE)存在关联,本研究旨在对该区域精细定位,以明确16号染色体与中国人SLE相关性。方法在16号染色体57.79~69.05cM(47218~52919kb)范围内选择8对微卫星标记(D16S540、D16S3044、D16S409、D16S517、D16S3136、D16S416、D16S419和D16S3034),扩增288个SLE患者家系DNA,产物经377DNA测序仪电泳,所得数据由Genescan软件收集进行基因分型,以ETDT及Genehunter进行连锁不平衡分析。结果D16S409及D16S517与SLE存在传递不平衡(P=0.0048和P<0.00001)。D16S517中,271bp等位基因优先传递给患病子代(P<0.00001),277bp等位基因则优先传递给正常子代(P<0.001)。结论研究结果证实人类SLE在16号染色体上存在疾病易感基因,中国人群16q12区(58.46cM,49Mb附近)与SLE发病相关联。     

HLA-DP does not contribute towards susceptibility to systemic lupus erythematosus.

OBJECTIVES--To determine whether HLA-DP genes are involved in determining susceptibility to systemic lupus erythematosus (SLE). METHODS--HLA-DPA1 and DPB1 genes were amplified by PCR of DNA samples from a panel of patients with SLE and normal controls. Amplified DNA was blotted on to nylon filters and probed with sequence-specific oligonucleotide (SSO) probes. RESULTS--No DPA1 or DPB1 allele was significantly associated with SLE, or with any immunological or clinical subset of SLE. Evidence was found for only limited linkage disequilibrium between HLA-DP and HLA-DQ/DR variants, and none between HLA-DP and the TAP2 gene. CONCLUSIONS--These data indicate that HLA-DP genes do not contribute towards determining susceptibility to SLE.     

TNF-308A and HLA-DR3 alleles contribute independently to susceptibility to systemic lupus erythematosus

OBJECTIVE: To evaluate the respective contributions of tumor necrosis factor (TNF) promoter polymorphisms and HLA-DR alleles to susceptibility to systemic lupus erythematosus (SLE). METHODS: TNF-238G/A and 308G/A promoter polymorphisms and HLA-DRB1 alleles were determined in 99 consecutive Caucasian SLE patients and 177 Caucasian controls. Standard and Mantel-Haenszel odds ratios were calculated to assess the magnitude of the susceptibility factors. The presence or absence of the SLE classification criteria was determined and correlated with the TNF promoter and HLA-DRB1 genotypes. RESULTS: The frequency of the TNF-308A/A and 308G/A genotypes was significantly higher in SLE patients (odds ratio 5.0). Conversely, TNF-238G/A and 238A/A genotypes were equally prevalent in SLE patients and controls. The HLA-DR3 specificity (DRBI*0301 allele) was significantly more prevalent in the SLE population (odds ratio 4.4). Stratification to correct for interdependence of the 2 loci confirmed the association of both TNF-308A and HLA-DR3 with SLE (Mantel-Haenszel odds ratio 3.2 and 2.4, respectively). No correlation was found between TNF promoter and HLA-DRB1 genotypes and any SLE classification criterion or disease manifestation. CONCLUSION: TNF-308A and HLA-DR3 alleles are independent susceptibility factors for SLE.     

Association of PDCD1 with susceptibility to systemic lupus erythematosus

Objective

The A allele of the PD1.3 single‐nucleotide polymorphism (SNP) on the programmed cell death gene PDCD1 was markedly more frequent in patients with systemic lupus erythematosus (SLE) than in unaffected controls in a recent study involving large sets of Swedish, European American, and Mexican families. This study sought to determine the role of PDCD1 in susceptibility to SLE in the Spanish population.

Methods

Seven PDCD1 SNPs were studied in 518 SLE patients and 800 healthy control subjects who had been recruited in 5 distant towns spanning continental Spain. Patients and controls were of Spanish ancestry. The diagnosis of SLE was in accordance with the American College of Rheumatology updated classification criteria.

Results

The A allele of the PD1.3 polymorphism was significantly less frequent in Spanish female patients with SLE than in Spanish female controls (9.0% versus 13.0%, odds ratio 0.67, 95% confidence interval 0.50–0.89). This difference was consistent across the 5 sets of samples grouped by town of recruitment. The other PDCD1 SNPs were not associated with SLE susceptibility. The haplotype structure of PDCD1 in the Spanish controls was different from that reported in other healthy control populations.

Conclusion

Our results confirm the association of PDCD1 with susceptibility to SLE, but the findings show a lack of involvement of the PD1.3 SNP, which is contrary to the role of the PD1.3 A allele observed previously. These contradictory results probably reflect population differences in the haplotype structure of the PDCD1 locus. More research focusing on new polymorphisms and identifying associations in other populations will be needed to clarify the role of PDCD1 in SLE susceptibility.

     

Genetic linkage and association of Fcgamma receptor IIIA (CD16A) on chromosome 1q23 with human systemic lupus erythematosus

OBJECTIVE: Although low-affinity alleles of human Fcgamma receptor types IIA and IIIA (FcgammaRIIA and FcgammaRIIIA, respectively) polymorphisms have been associated with systemic lupus erythematosus (SLE) in case-control studies, the relative contribution of these genes to SLE susceptibility has not been resolved. METHODS: We analyzed the distribution of alleles of FcgammaRIIA, FcgammaRIIIA, and FcgammaRIIIB in 126 multiplex-SLE pedigrees and FcgammaRIIA and FcgammaRIIIA in a case-control replication study, using allele-specific polymerase chain reaction and direct sequencing of genomic DNA. Statistical tests of association were performed to detect evidence of linkage between the single nucleotide polymorphisms and SLE. RESULTS: We found evidence for linkage at both the FcgammaRIIIA (single-point nonparametric linkage [NPL] 1.8, P = 0.038; multipoint NPL 2.7, P = 0.004) and the FcgammaRIIA (single-point NPL 2.0, P = 0.021; multipoint NPL 2.6, P = 0.006) loci, but not the FcgammaRIIIB locus. Family-based tests of association demonstrated increased transmission of the low-affinity F176 allele at the FcgammaRIIIA locus (odds ratio [OR] 2.18, P = 0.0005 by transmission disequilibrium test and P = 0.002, by pedigree disequilibrium test [PDT]), but little evidence of preferential transmission of alleles at FcgammaRIIA (P = 0.089 by PDT). Stratification by ethnicity showed preferential transmission of the associated FcgammaRIIIA allele both in families of African American ancestry and in those of European American ancestry. Despite significant linkage disequilibrium between these genes, 2- and 3-locus haplotype analysis of the extended Fcgamma receptor cluster did not reveal any significant association beyond that observed with FcgammaRIIIA alone. In a large case-control replication study of 438 patients with SLE and 219 controls, FcgammaRIIIA provided the strongest evidence of an FcgammaR-SLE association (additive model: V/V 176 versus V/F 176 OR 1.51, V/V 176 versus F/F 176 OR 1.98, P = 0.007). CONCLUSION: To our knowledge, these data are the first to demonstrate linkage and both family-based and case-control-based association of FcgammaRIIIA with SLE. These data provide genetic evidence supporting a role for the physiologically relevant single nucleotide polymorphism of the FcgammaRIIIA gene in the pathophysiology of this complex genetic disease.     

The C1q binding assay in systemic lupus erythematosus

To investigate the relationship of C1q binding assay (C1qBA) to disease activity in systemic lupus erythematosus (SLE), a retrospective study was carried out on 232 C1qBA performed in 33 patients with SLE. When initial values only were assessed (33 tests in 33 patients), there was no relationship between positive and negative C1qBA and abnormal renal function (P = 0.482, Fisher exact test). Of 87 tests performed during active renal disease, 34 (39%) were positive; of 48 tests during active non-renal disease, 25(52%) were positive; of 83 tests when the disease was inactive, 45(54%) were positive; and of 14 tests during episodes of infection, 10 (71%) were positive. Corresponding means for the C1qBA were as follows: renal 65.66, non-renal 78.02, in-active 56.04, infection 135.79 (no significant differences by Student's t-test). There was no significant relationship with the C1qBA when comparing active disease (renal and non-renal) with inactive disease (X² Yates = 1.875, P = 0.171). When renal function abnormalities were analyzed separately, the C1qBA values were independent of azotemia or proteinuria (X² Yates = 1.399, P = 0.237). Patients were seen with progressive renal disease who had consistently negative C1qBA, as well as patients with benign clinical courses despite elevated C1qBA. The discordance of the C1qBA results with disease activity in SLE highlights the limitations of immune complex (IC) determinations by a single technique, and stresses the importance of evaluating such tests in terms of both their specificity and sensitivity. These data further suggest that the relationship of IC to the disease process in SLE may be a complex one.     

Anti-chromatin and anti-C1q antibodies in systemic lupus erythematosus compared to other systemic autoimmune diseases

OBJECTIVE: To evaluate the prevalence, sensitivity, and specificity of anti-chromatin and anti-C1q antibodies in systemic lupus erythematosus (SLE) and lupus nephritis compared to small vessel vasculitis and other connective tissue diseases. To provide long-term follow-up data for anti-chromatin antibodies in lupus nephritis. METHODS: We determined the significance of anti-nuclear antibodies (ANA), anti- double-stranded DNA (anti-dsDNA), anti-chromatin, and anti-C1q antibodies, as well as complement factors C3 and C4, in relation to disease activity in SLE patients with (n = 47; long-term follow-up data for 33 patients) and without (n = 31) biopsy-confirmed lupus nephritis, microscopic polyangiitis (n = 37), Wegener's granulomatosis (n = 66), primary Sj?gren's syndrome (n = 17), limited scleroderma (CREST syndrome) (n = 6), and progressive systemic scleroderma (PSS) (n = 11). RESULTS: Anti-chromatin antibodies were more specific and sensitive than anti-C1q antibodies in distinguishing SLE patients from those with other systemic autoimmune diseases [anti-chromatin: sensitivity 64.1%, specificity 99.2%, odds ratio (OR) 219.6; anti-C1q: sensitivity 50%, specificity 72.6%, OR 2.65]. Anti-C1q antibodies were present in 75% of patients with Sj?gren's syndrome and 35.1% of patients with microscopic polyangiitis. Anti-chromatin antibodies could identify SLE in patients with positive ANA but negative anti-dsDNA antibodies. Persisting anti-chromatin antibodies indicated SLE disease activity, even if anti-dsDNA antibodies had become negative. In long-term follow-up, those SLE patients with negative anti-dsDNA antibodies but persisting ANA and anti-chromatin antibodies relapsed if immunosuppression had been tapered. Anti-chromatin antibodies correlated with the SLE disease activity index (SLEDAI) as a marker of disease activity. CONCLUSIONS: The measurement of anti-chromatin, but not anti-C1q, antibodies in patients with systemic autoimmune diseases increases diagnostic sensitivity and specificity for SLE and assists in treatment decisions in anti-dsDNA-negative patients.     

Major histocompatibility complex genes and susceptibility to systemic lupus erythematosus

Susceptibility to systemic lupus erythematosus is associated with major histocompatibility complex (MHC)--encoded genes. We have used nucleotide sequence analysis to better define the disease-associated MHC alleles. HLA-DR2, DQw1, and especially the rare allele DQ beta 1. AZH confer high relative risk (RR = 14) for lupus nephritis in a Caucasian population of patients. Pilot studies using historical controls suggest that these genes also confer a high risk in non-Caucasian ethnic groups (RR = 24-78). We have found that DR4 is significantly decreased in patients with lupus nephritis. Fifty percent of the patients with lupus nephritis had either the DQ beta 1.1, the DQ beta 1.AZH, or the DQ beta 1.9 alleles. These alleles share amino acid residues that have been predicted to be the contact points for antigen and the T cell receptor. These HLA alleles appear to have a direct role in the predisposition to lupus nephritis, whereas DR4 may have a "protective" effect.     

Association of HLA-DQB1 allelic sequence variation with susceptibility to systemic lupus erythematosus

Systemic lupus erythematosus (SLE) is an autoimmune disease in which polymorphisms within the human leukocyte antigen (HLA) region have been associated to its etiology. We conducted this study to compare the HLA-DQB1 allelic sequence variation among SLE patients and controls in the northeast of Iran. Genomic DNA of 40 SLE patients and 83 healthy controls were amplified by Polymerase Chain Reaction with Sequence-Specific Primers technique (PCR-SSP). Seven serological subclasses of the HLA DQB1 were detected. Allele distribution comparison showed in the SLE group a significant increase of HLA DQ6 (*0601-*0609) (p=0.006); whereas alleles HLA DQ7 (*0301-*0304) were significantly decreased (p=0.005). Combination of DQ5 (*0501-*0504)-DQ6 (*0601-*0609) was increased in patients. These results suggest that DQ6 is the dominant HLA DQB1 allele probably associated with genetic susceptibility to SLE in the northeast of Iran. The association supports the importance of ethnic background and indicates the importance of various genes that has been observed in different SLE populations.     

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-23和白细胞介素-12亚单位在系统性红斑狼疮中的表达研究

目的研究自身免疫性疾病系统性红斑狼疮(SLE)中白细胞介素(IL)-23和IL-12的调节作用。方法78例SLE患者[以SLE疾病活动评分(SLEDAI)进行活动性评分]和正常对照36名,分别抽提SLE患者和正常对照外周血单个核细胞(PBMC)的mRNA,并反转录成cDNA,用实时定量聚合酶链反应(PCR)的方法对IL-23特异的亚单位P19、IL-12特异的亚单位P35以及它们共有的亚单位P40作定量分析,数据采用看家基因进行标化。结果未经治疗、已经治疗的SLE患者的IL-12特异性的亚单位P35表达水平显著低于正常对照(P值分别<0.05,<0.01)。与未经治疗相比,用类固醇激素或类固醇类激素加免疫抑制剂治疗的SLE患者显著抑制P40和P19的表达(P值分别<0.01和<0.05)。活动性SLE患者(SLEDAI>10)的P19、P40、P35表达水平显著高于非活动性SLE患者(SLEDAI≤10)(P值分别<0.01,<0.01和<0.05)。结论IL-12的表达下降或IL-23的表达上调也许参与SLE的发病过程,这两种细胞因子有可能是SLE患者的治疗靶点。     

Anti-C1q antibodies in pregnant patients with systemic lupus erythematosus

OBJECTIVE: To study anti-C1q antibodies in pregnant patients with systemic lupus erythematosus (SLE) and to evaluate their prognostic significance for the occurrence of disease flares or pregnancy complications. METHODS: Twenty-one pregnancies in 19 SLE patients prospectively followed were analyzed. Disease activity was evaluated on the basis of the physician's intention to treat and a modified version of the ECLAM index. Anti-C1q and anti-dsDNA antibodies were detected in the sera by an ELISA assay. Antinuclear antibodies, anti-ENA antibodies, anticardiolipin antibodies and lupus anticoagulant were also performed. RESULTS: In all the patients the disease was inactive at the beginning of the pregnancy. Four flares of disease activity were observed in 4 pregnancies (19%) and obstetric complications were encountered in 7 pregnancies (43%). Anti-C1q antibodies were positive in 4 (19%) pregnancies and anti-dsDNA antibodies in 8 (38%). The presence of anti-phospholipid antibodies at the first assessment was correlated with the occurrence of obstetric complications (p<0.05). The presence of anti-C1q and anti-dsDNA antibodies at the first assessment had no prognostic significance for the occurrence of flares or obstetric complications during the course of pregnancy. Although the small number of patients studied did not allow for statistically significant analysis, flares appeared to be more likely to occur in patients presenting with anti-dsDNA or anti-C1q antibodies during pregnancy compared to patients with no changes in these antibody titers (43% vs 8% respectively). CONCLUSIONS: The presence of anti-C1q and anti-dsDNA antibodies does not seem to be prognostic for the occurrence of flares during pregnancy. Further studies are warranted to explore this possibility.     

在中国人群中鉴定发现PBX1基因为1q 23-24区域新的狼疮易感基因

目的用连锁不平衡(linkage disequilibrium)的方法对1号染色体狼疮易感区域进行进一步的精细定位,并由此定位新的系统性红斑狼疮(SLE)候选基因.方法用9对高密度的微卫星标记对1q23-24易感区域进行连锁不平衡分析;用延伸型传递不平衡试验(ETDT)、Gene Hunter和家系为基础的相关分析(FBAT)软件分析连锁不平衡的结果;用TaqMan荧光实时定量聚合酶链反应(PCR)检测候选基因mRNA的表达量;用等位基因分型PCR对候选基因内的4个SNP分型.结果①发现微卫星标记D1S2628 (P＝0.000 987,Pc＝0.001 2)和D1S2673 (P＝0.007 082,Pc＝0.010 4)的等位基因存在整体的显著传递不平衡);②发现D1S2628-119 bp (Pc＝0.001 2;传递∶不传递＝91∶45,P＝0.000 1)和D1S2673-103 bp (Pc＝0.010 4,传递∶不传递＝109∶67,P＝0.0016)等位基因从杂合子父母优势传递给受累后代;③在中国汉族红斑狼疮核心家系中单倍型D1S2628 (119 bp)-D1S2673 (103 bp) (传递∶不传递＝43∶18,P＝0.001 4)和单倍型D1S2628 (119 bp)-D1S2673 (107 bp) (传递∶不传递＝29∶12,P＝0.007 9)存在传递不平衡.在中国汉族SLE患者中PBX1的mRNA的表达量显著下降(P＜0.000 1).PBX1基因中的单倍型SNP1(G)-SNP2(C)优势传递给受累后代.结论在中国汉族人群的1q23区存在与微卫星标记D1S2628和D1S2673物理距离相近的SLE易感基因;PBX1基因表达较低,其SNP单倍型与SLE易感显著相关.这提示PBX1基因可能是一个新的候选基因.     

Genetics of susceptibility and severity in systemic lupus erythematosus

PURPOSE OF REVIEW: The genetic basis of systemic lupus erythematosus, a complex genetic trait, may provide important insights into autoimmune disease. Innovation in both practical and theoretical approaches will assist in accelerating the pace of discovery and our understanding of pathogenesis. RECENT FINDINGS: Significant progress has been made in the last year with respect to the refinement of genetic intervals to promising candidate genes involved in systemic lupus erythematosus pathogenesis and specific phenotype susceptibility. This review highlights these discoveries and suggests platforms that may affect the future of analysis of this complex disease. SUMMARY: Understanding the genetic basis for systemic lupus erythematosus disease and sub-phenotype susceptibility will have a substantial effect on the therapeutic interventions used to care for patients.     

Evidence for gene–gene epistatic interactions among susceptibility loci for systemic lupus erythematosus

Objective

Several confirmed genetic susceptibility loci for lupus have been described. To date, no clear evidence for genetic epistasis in lupus has been established. The aim of this study was to test for gene–gene interactions in a number of known lupus susceptibility loci.

Methods

Eighteen single‐nucleotide polymorphisms tagging independent and confirmed lupus susceptibility loci were genotyped in a set of 4,248 patients with lupus and 3,818 normal healthy control subjects of European descent. Epistasis was tested by a 2‐step approach using both parametric and nonparametric methods. The false discovery rate (FDR) method was used to correct for multiple testing.

Results

We detected and confirmed gene–gene interactions between the HLA region and CTLA4, IRF5, and ITGAM and between PDCD1 and IL21 in patients with lupus. The most significant interaction detected by parametric analysis was between rs3131379 in the HLA region and rs231775 in CTLA4 (interaction odds ratio 1.19, Z = 3.95, P = 7.8 × 10⁻⁵ [FDR ≤0.05], P for multifactor dimensionality reduction = 5.9 × 10⁻⁴⁵). Importantly, our data suggest that in patients with lupus, the presence of the HLA lupus risk alleles in rs1270942 and rs3131379 increases the odds of also carrying the lupus risk allele in IRF5 (rs2070197) by 17% and 16%, respectively (P = 0.0028 and P = 0.0047, respectively).

Conclusion

We provide evidence for gene–gene epistasis in systemic lupus erythematosus. These findings support a role for genetic interaction contributing to the complexity of lupus heritability.