Genetic analysis of autoimmune regulator haplotypes in alopecia areata

Alopecia areata is an immune-mediated disorder, occurring with the highest observed frequency in the rare recessive autoimmune polyendocrinopathy-candidiasis-ectodermal dystrophy (APECED) syndrome caused by mutations of the autoimmune regulator (AIRE) gene on chromosome 21q22.3. We have previously detected association between alopecia areata and a single nucleotide polymorphism (SNP) in the AIRE gene in patients without APECED, and we now report the findings of an extended examination of the association of alopecia areata with haplotype analysis including six SNPs in the AIRE gene: C-103T, C4144G, T5238C, G6528A, T7215C and T11787C. In Caucasian groups of 295 patients and 363 controls, we found strong association between the AIRE 7215C allele and AA [P = 3.8 x 10(-8), OR (95% CI): 2.69 (1.8-4.0)]. The previously reported association between AA and the AIRE 4144G allele was no longer significant on correction for multiple testing. The AIRE haplotypes CCTGCT and CGTGCC showed a highly significant association with AA [P = 6.05 x 10(-6), 9.47 (2.91-30.8) and P = 0.001, 3.51 (1.55-7.95), respectively]. To select the haplotypes most informative for analysis, we tagged the polymorphisms using SNPTag software. Employing AIRE C-103T, G6528A, T7215C and T11787C as tag SNPs, two haplotypes were associated with AA; AIRE CGCT and AIRE CGCC [P = 3.84 x 10(-7), 11.40 (3.53-36.9) and P = 3.94 x 10(-4), 2.13 (1.39-3.24) respectively]. The AIRE risk haplotypes identified in this study potentially account for a major component of the genetic risk of developing alopecia areata.     

Unravelling the genetic complexity of autoimmune thyroid disease: HLA, CTLA-4 and beyond

The autoimmune thyroid diseases (AITDs) including Graves' disease (GD) and autoimmune hypothyroidism (AIH) are the commonest of the autoimmune conditions affecting 2-5% of the western population. Twin studies have clearly demonstrated that AITDs are caused by a combination of both environmental and genetic factors. Association of the HLA class II region with AITD has been documented for over 20 years now, but the primary aetiological variant in this region remains unknown. More recently the CTLA-4 gene region has been identified as the second locus conferring susceptibility to AITD. In contrast to HLA, a polymorphism of the CTLA-4 gene, which encodes an important negative regulator of the immune system, has been identified as a candidate for a primary determinant for AITD. A large number of candidate gene and genome wide linkage studies have been involved in the search for the elusive 'third' locus. The thyroglobulin (Tg) gene in humans maps to chromosome 8q, which has been linked in family studies to AITD. A number of association studies in humans and the mouse model for AITD are beginning to implicate the Tg gene although convincing evidence for a primary causative role is still needed. The establishment of large DNA disease resources along with more detailed genetic maps and the development of faster, more effective, high throughput genotyping and sequencing methods, provides some sense of optimism that novel loci will be identified in the near future and the complex aetiology of AITD will be further unraveled.     

Genetic variants in IL4RA,IL6, and IL12B genes and susceptibility to hepatitis B and C virus infections among Iraqi patients

Hepatitis B and C viruses (HBV and HCV) are common causative pathogens of viral hepatitis. Progression of both infections is determined by virus- and host-related factors. Cytokines are important host genetic factors that may have a predisposing role in HBV and HCV infections. This case-control study evaluated the genetic association of IL4RA₊₁₉₀₂ (rs1801275), IL6₋₁₇₄ (rs1800795), IL6₋₅₉₇ (rs1800797), and IL12B₋₁₁₈₈ (rs3212227) variants with chronic HBV and HCV infections among Iraqi patients. A total of 220 viral hepatitis patients were enrolled in the study (113 HBV and 107 HCV), together with 141 healthy subjects. Sequence-specific primer polymerase chain reaction assay was the genotyping method. Results revealed that under a dominant genetic model, IL6₋₁₇₄ variant was significantly associated with HBV infection, whereas no association with the HCV risk was reported. However, the risk for both infections was markedly associated with IL6₋₅₉₇ variant under recessive, dominant, and codominant genetic models. Estimation of IL6₋₁₇₄-IL6₋₅₉₇ haplotypes depicted that G-A haplotype was significantly associated with an increased risk to develop HBV infection, whereas a significantly decreased risk was associated with G-G and C-G haplotypes. For HCV, G-G and C-A haplotypes were significantly associated with risk of HCV infection. IL4RA₊₁₉₀₂ and IL12B₋₁₁₈₈ variants showed no association with HBV or HCV risk. Analysis of variance revealed no significant association between genotypes of the four determined single-nucleotide polymorphisms and HBV or HCV viral load. In conclusion, the study supports the concept that IL6₋₅₉₇ variant is associated with susceptibility to HBV and HCV infections among Iraqis. The risk of HBV infection is further associated with IL6₋₁₇₄ variant.     

The non-synonymous C1858T substitution in the PTPN22 gene is associated with susceptibility to the severe forms of alopecia areata

Alopecia areata is an acquired hair loss disorder resulting from an immunologically- mediated attack on hair follicles and autoimmunity may play a part in its pathogenesis. The non-synonymous C1858T substitution in the PTPN22 gene, which encodes lymphoid protein tyrosine phosphatase, has been shown to be associated with susceptibility to autoimmune disorders. In this study, the objective was to ascertain whether or not the disease-associated 1858T (W620) allele was associated with alopecia areata. For this, the allelic distribution of the PTPN22 C1858T alleles was determined in 196 English patients with alopecia areata and 507 healthy subjects in a case control study using a restriction fragment length polymorphism-polymerase chain reaction (PCR-RFLP) genotyping method. The results indicated that the frequency of the 1858T allele did not differ significantly between the alopecia areata patient group and the control cohort: of 392 alopecia areata alleles, 41 (10.5%) encoded the W620 variant compared to 86 of 1014 (8.5%) control alleles. However, in patients with severe disease, 25/168 (14.9%) alleles were 1858T and this frequency differed from that in the control group (P = 0.0127; OR, 95% CI = 1.89, 1.17 - 3.05). These results suggest that the non-synonymous C1858T substitution in the PTPN22 gene may have an influence on the severity of alopecia areata and provide further evidence for autoimmunity as an aetiological factor in this disorder.     

PTPN22 G788A和A10281188G与广东地区人群RA易感性的关系研究

目的探讨蛋白酪氨酸磷酸酶非受体型22（PTPN22）基因多态性（rs33996649/G788A/R263Q和rs1310182/A10281188G）与广东地区汉族人群类风湿性关节炎（RA）易感性间的关系。方法选取广东地区人群中218例RA患者以及229例健康对照者进行病例对照研究,采用PCR-RFLP技术检测PTPN22G788A和A10281188G两个多态性位点的基因型,计算比较两组基因型和等位基因频率。结果 RA患者和健康对照者PTPN22在788位点均为G等位基因,未检测到A等位基因,没有发现单核苷酸多态性的存在;10281188位点G等位基因在病例组和对照组中的频率分别为12.4%和13.3%（P〉0.05）。结论广东地区人群PTPN22788位点不存在多态性,G788A和A10281188G基因位点与广东汉族人群RA的发病无相关性。     

广西地区壮族Graves病与CTLA4基因多态性相关性研究

目的分析CTLA4基因微卫星多态性与广西地区壮族Graves病（Graves’disease，GD）相关性。方法提取48例广西地区壮族GD患者和44名正常对照组的外周血白细胞基因组DNA，应用聚合酶链反应检测CTLA4第4外显子的3’非翻译区包含（AT）n[CTLA4（AT）n]重复序列的特异性等位基因。结果广西地区壮族人CTLA4微卫星多态性检出19种等位基因。壮族GD组106bp等位基因频率与正常对照组比较显著增高，差异有统计学意义（P〈0．05）。结论广西壮族GD患者与CTLA4基因多态性明显相关,CTA4（AT）n 106bp可能是广西壮族GD的易感等位基因。     

Identification of susceptibility SNPs in CTLA-4 and PTPN22 for scleritis in Han Chinese

The aim of this study was to determine the association between 13 single nucleotide polymorphisms (SNPs) in the cytotoxic T lymphocyte-associated antigen-4 (CTLA4) and protein tyrosine phosphatase non-receptor type 22 (PTPN22) genes with scleritis in a Chinese Han population. We recruited 432 scleritis patients and 710 healthy controls. Four tag SNPs of CTLA4 and nine tag SNPs of PTPN22 were selected using Haploview. Genotyping was performed with the Sequenom MassArray® iPLEX GOLD Assay. Genotype and allele frequency differences were analyzed by χ² test and Bonferroni correction. Haplotype analysis was performed to further evaluate the association of these two genes with scleritis. In this study, CTLA4/rs3087243 G allele frequency and GG genotype frequency were significantly increased in scleritis patients compared to healthy controls [corrected P-value (Pc) = 0·02, odds ratio (OR) = 1·475, 95% confidence interval (CI) = 1·175–1·851; Pc = 0·04, OR = 1·546, 95% CI = 1·190–2·008, respectively]. None of the tested SNPs in the PTPN22 gene showed an association with scleritis. Haplotype analysis revealed a lower frequency of a CTLA4 TCAA haplotype (order of SNPs: rs733618, rs5742909, rs231775, rs3087243) (Pc = 4·26 × 10^–3, OR = 0·618, 95% CI = 0·540–0·858) and a higher frequency of a PTPN22 TTATACGCG haplotype (order of SNPs: rs3789604, rs150426536, rs1746853, rs1217403, rs1217406, rs3789609, rs1217414, rs3789612, rs2488457) (Pc = 2·83 × 10^–4, OR = 1·457, 95% CI = 1·210-1·754) in scleritis patients when compared to healthy controls. In conclusion, our findings indicate that CTLA4 and PTPN22 might confer genetic susceptibility to scleritis in a Chinese Han population.     

云南人群中PTPN22和PADI4基因多态性与类风湿关节炎的关联研究

目的 探讨云南人群PTPN22和PADI4基因的7个SNP多态与类风湿关节炎易感性的相关性.方法 选取192例类风湿关节炎患者和288名正常人进行病例对照研究.分别用PCR-RFLP法检测PTPN22基因的rs33996649和1858位点、PADI4基因的rs11203366和rs874881位点,用焦磷酸测序法检测PADI4基因的rs1635579、rs2428736和rs2240340共7个SNP位点的基因型.结果 在7个SNP位点中,PADI4基因的rs2240340位点的等位基因频率和基因型频率在病例组和对照组间差异有统计学意义(P＜0.05).结论 在云南人群中PADI4基因的rs2240340多态性与类风湿关节炎的易感性存在相关性.     

Highly polymorphic promoter regions of HLA DQA1 and DQB1 genes do not help to further define disease susceptibility in insulin-dependent diabetes mellitus

HLA DQA1, HLA DQB1 genes confer susceptibility to insulin-dependent (type 1) diabetes mellitus (IDDM). Since variants of their upstream regulatory regions are linked to the exons, we investigated their promoter polymorphisms (QAP and QBP) by a combination of PCR-based typing protocols in 136 IDDM patients, 167 controls and 6 families with an IDDM proband to identify possible additional susceptibility markers. Of major interest for IDDM susceptibility are the promoter "splits" of HLA DQA1*0301 (QAP3.1 and QAP3.2) and HLA DQB1*0302 (QBP3.2 and QBP3.3). QAP 3.1 (96% in patients vs 98% in controls) and QBP3.2 (100% vs 99%) were found to be the most frequent promoter variants for HLA DQA1*0301 and DQB1*0302, respectively, whereas QAP3.2 and QBP3.3 were very rare. Furthermore the promoter "splits" were equally distributed on the respective exon alleles in all groups and cosegregated in families as expected. In conclusion, HLA DQ-mediated susceptibility and protection in IDDM is not restricted to the exon but extends to the promoter region without further defining the genetic risk.     

The 1858T PTPN22 gene variant contributes to a genetic risk of type 1 diabetes in a Ukrainian population

The 1858T variant of the protein tyrosine phosphatase gene, PTPN22, is associated with an increased risk of several autoimmune diseases. The aim of this study has been to investigate the possible association of 1858C-->T PTPN22 polymorphism and type 1 diabetes (T1D) in Caucasians from Ukraine. Overall, the distribution of 1858 PTPN22 genotypes differed significantly between the T1D patient group (n = 296) and the control group (n = 242) (P = 0.0036). When both groups were classified according to sex, the TT genotype and T allele showed a statistically significant higher frequency in T1D female patients (5.9 and 22.8%, respectively) in comparison with the female controls (0 and 11.9%) (P = 0.008 for both analyses). The patients with the TT genotype were significantly younger at the onset of T1D compared with those with genotypes TC and CC (P = 0.035 and 0.019, respectively). In our Ukrainian Caucasian cohort, we confirmed the association between T1D and the PTPN22,1858T allele.     

The association of PTPN22 (rs2476601) and IL2RA (rs11594656) polymorphisms with T1D in Egyptian children

BackgroundType 1 diabetes mellitus (T1D) is a T cell-mediated autoimmune disease characterized by the destruction of pancreatic β cells. PTPN22 and IL2RA polymorphisms have been found to be associated with several autoimmune diseases including T1D.AimsWe aimed to elucidate the role of PTPN22 and IL2RA polymorphisms in predisposition of T1D in Egyptian children.MethodsWe studied 150 children and adolescents with T1D and 165 healthy controls. The PTPN22 (rs2476601) and IL2RA (rs11594656) polymorphisms were genotyped using polymerase chain reaction.ResultsWe found that carriers of the T allele of PTPN22 were significantly more likely to develop T1D (OR = 2.2, 95% CI = 1.2–4, P = 0.01). Also, the carrier of TT genotype and T allele of IL2RA more likely to develop T1D (OR = 2.8, 1.4, respectively, P = 0.03). There was a statistically significant association between T allele of PTPN22 gene and females ⩽10 years old at the onset of diabetes (OR = 4, 95% CI = 1.2–13.4, P = 0.019).ConclusionThis study suggests a possible association between the T allele of PTPN22 gene and TT genotype of IL2RA with T1D in studied Egyptian children, especially, females with early onset diabetes who carried the 1858T allele.     

Associative role of HLA‐DRB1 SNP genotypes as risk factors for susceptibility and severity of rheumatoid arthritis: A North‐east Indian population‐based study

Rheumatoid arthritis (RA) is a complex, multifactorial, systemic autoimmune disease. Reports are suggestive of the role of HLA especially HLA‐DRB1 alterations in RA pathogenesis. Existing data involving different geographical populations on the role of alterations in specific locus of HLA‐DRB1 in RA susceptibility and severity are equivocal, with no data available from ethnically distinct North‐east Indian population, where RA cases are alarmingly increasing. This study aimed to evaluate the association of HLA‐DRB1 gene SNPs (rs13192471, rs660895 and rs6457617) with susceptibility and severity of RA in an ethnically distinct North‐east Indian population. Whole blood was collected from clinically characterized RA cases (satisfying the American College of Rheumatology 1987 criteria) (n = 123) and community‐based age and sex‐matched healthy controls (n = 156) with informed consent. The HLA‐DRB1 SNP analysis was performed for all the RA and control cases using ARMS‐PCR using case and control genomic DNA as template. Statistical analysis was performed by SPSSv13.0 software. The HLA‐DRB1 rs660895 showed both wild (AA) and heterozygote (AG) genotype but the heterozygote allele was found to be associated with reduced risk of RA compared to controls [OR = 0.531, p = .024]. The difference in distribution of rs6457617 polymorphism between RA and control cases was comparable [OR = 0.525, p = .079]. Significantly higher distribution of variant rs13192471 genotype was observed in RA cases (69.92%) compared to controls (46.75%) (p < .001) and was associated with increased risk of susceptibility to RA [OR = 2.576, p < .001] compared to controls, as well as progression to severity in RA cases [OR = 2.404, p = .048]. Combinatorially also, the presence of rs13192471 variant genotype was associated with increased risk of RA susceptibility [OR = 8.267, p = .026] and RA severity [OR = 3.647, p = .280]. Alterations in HLA‐DRB1 are associated with RA susceptibility. HLA‐DRB1 rs13192471 SNP plays a critical role in RA susceptibility and severity in North‐east Indian cases and has prognostic significance in RA.     

Contribution of HLA class II genes to susceptibility in achalasia

Abstract: Achalasia is a motor disorder of the esophagus resulting in functional obstruction. The cause of the lesion is unknown although genetic and immunologic factors have been suggested. An association with serological HLA epitopes has been previously reported. In this study, we have further examined this HLA class II association with susceptibility to achalasia by DNA based methods. Achalasia patients ( n =40) and healthy controls ( n = 275), all Caucasians and unrelated, were included in the analysis. The strongest associations were with HLA-DQAl*0101 and two HLA-DQαβ heterodimers having their a chain encoded by this allele. Moreover, relative risk was significantly higher in DQAl*0101 homozygotes as compared to heterozygotes and results suggested that DQB1*02 may have a protective role.     

HLA Class I in Egyptian patients with Behçet’s disease: new association with susceptibility,protection, presentation and severity of manifestations

Introduction: Behçet’s disease is an autoimmune disease with diverse clinical manifestations with vasculitis being the hallmark of the disease. The aim of this work is to study the genetic association between human leukocyte antigen (HLA) class-I molecules of Egyptians with Behçet’s disease and the disease susceptibility and clinical patterns.

Methods: Fifty-seven patients diagnosed with Behçet’s disease according to the 1990 International Study Group (ISG) criteria for Behçet’s disease coming from Egyptian origin up to the third grandfather were included in the study. Healthy controls were taken from HLA Class-I case control studies in Egyptian population yielding a pool of 221 healthy controls. HLA Class-I typing for patients was done using Reverse Sequence specific oligonucleotide probes (rSSO).

Results: Male patients represented 89% of the sample. Mean age of onset was 25.81 (± 6.7) years and mean disease duration was 9.47 (± 7.4) years. Behçet’s disease was associated with HLA-A*24 and HLA-B*42 (p = 0.001) and highly associated with HLA-A*68 and B*15 and B*51 (p < 0.001). While HLA A*03 and B*52 were protective for Behçet’s (p = 0.002 and 0.007). Interestingly, HLA-B*51 and HLA-A*68 (p = 0.005 and 0.023) were associated with the blinding eye disease. HLA-B*51 was protective from Neurological and vascular involvement (p = 0.005 and 0.032, respectively).

Conclusion: Behçet’s disease is associated with HLA Class-I A*24, A*68 and B*15, B*42 and B*51 in Egyptian patients while A*03 and B*52 were found to be protective. Interestingly, HLA B*51 and A*68 could be considered as poor prognostic factor for eye involvement.     

Genetic variant of IL28B rs12979860, as predictive marker of interferon‐based therapy in Pakistani population

Hepatitis C virus (HCV) genotypes and genetic variants of interleukin 28B (IL28B) are significantly associated with interferon plus ribavirin treatment of HCV infection. We investigated the distribution of HCV genotypes and single‐nucleotide polymorphisms (SNPs) of IL28B (rs12979860 and rs8099917) in Pakistani population. IL28B genotyping was performed by allele‐specific PCR and restriction fragment length polymorphism PCR in 140 chronic hepatitis C patients (CHC) and 120 healthy controls. HCV genotype 3 (HCVG3) was the most prevalent genotype, 71.4% (n = 100/140) and with the highest treatment response of 90% (n = 90/100). The overall treatment response of all the HCV genotypes was 82% (n = 115/140). The distribution of IL28B rs12979860CC genotype in treatment responder and non‐responder groups was 40.8% (n = 47/115) and 16% (n = 4/25) respectively. IL28B rs12979860CC genotype demonstrated a significant correlation (p = 0.019) with interferon‐based therapy of HCV infection. However, there was no observed association of IL28B rs8099917 polymorphism with treatment response in CHC patients (p = 0.264). In conclusion, HCV genotypes and IL28B rs12979860 are predictive markers for the efficiency of interferon plus ribavirin combinational therapy of HCV infection. We recommend the inclusion of testing for these markers in the clinical criteria for decision making for HCV therapy in Pakistani population.     

RET基因调控区遗传变异与先天性巨结肠患病风险研究

目的先天性巨结肠（HSCR）是一种复杂的先天性疾病,RET是其主要的易感性基因。本研究对RET非编码区单核甘酸多态性（SNP）-5G〉A（rs10900296）,-1A〉C（rs10900297）和intron1 C〉T（rs2435357）进行分型分析,评估RET基因调控区SNPs及单倍型与先天性巨结肠之间的相关性。方法选取115名病例组病人和139名对照组正常人群,应用聚合酶链反应（PCR）技术和直接测序的方法进行基因分型。回归模型中使用OR值和95%置信区间（CI）作为基因型危险性的评价指标。结果 -5G〉A,-1A〉C,intron1C〉T各基因型频率在病例和对照人群的分布具有显著差异。-5 AA（OR=6.26,95%CI=3.62-10.83）,-1 CC（OR=7.54,95%CI=2.06-27.66）和intron1 TT（OR=19.22,95% CI=7.54-48.99）基因型均能显著增加HSCR发病的风险。单倍型A-C-T（OR=6.28,95% CI=3.77-10.46）和双体型A-C-T/A-C-T（OR=13.62,95% CI=3.48-53.30）分析同样表明与HSCR发病风险存在较强的相关性,并呈现出一定的累积效应。结论 RET基因调控区的基因多态性可能与HSCR的发病易感性有关,支持RET通路的常见变异在HSCR的发展过程中起着重要的作用的假设。     

IRF5, PTPN22, CD28, IL2RA,KIF5A,BLK and TNFAIP3 genes polymorphisms and lupus susceptibility in a cohort from the Egypt Delta; relation to other ethnic groups

ObjectiveTo replicate a single nucleotide polymorphism (SNP) of known genes for lupus (IRF5 rs10488631, PTPN22 rs2476601, BLK rs2736340 and TNFAIP3 rs5029939) and other autoimmune diseases (CD28 rs1980422, IL2RA rs2104286 and KIF5A rs1678542) on a newly studied Egyptian cohort to investigate the genetic disparity with different studied ethnic groups in relation to lupus susceptibility.Methods170 Egyptian patients from Egypt Delta with SLE and 241 matched healthy controls were genotyped by Taqman real time PCR for the selected SNPs.ResultsThe results revealed significant association with IRF5 (p < 0.0001) and PTPN22 (p = 0.008) and insignificant association with KIF5A, CD28, IL2RA, BLK and TNFAIP3 genes.ConclusionsThis study may provide an additional evidence for the association between IRF5 and PTPN22 and lupus susceptibility and may exclude it for CD28, IL2RA, and KIF5A.     

广东汉族人类风湿关节炎易感性与HLA-DRB1基因相关性研究

目的探讨ＨＬＡ－ＤＲＢ１基因与类风湿关节炎（ＲＡ）相关性。方法采用ＰＣＲ－ＳＳＰ方法对４７例广东汉族人ＲＡ患者进行ＨＬＡ－ＤＲＢ１基因分型，并与相应人群健康者１０２例结果比较。结果ＨＬＡ－ＤＲ４基因在ＲＡ组显著增高（３５．１％，ＲＲ＝３．５５，Ｐ＜０．００５，ＥＦ＝０．２５２），ＤＲ１６在ＲＡ组也高于正常（ＲＲ＝２．５７，Ｐ＜０．０５）；而ＤＲ９基因在ＲＡ组显著减少（Ｐ＜０．００５）。３１例ＤＲ４＋患者患病年龄较早，病情较重（类风湿因子阳性率和Ⅱ期ＲＡ骨关节Ｘ线改变者显著高于ＤＲ４－患者，Ｐ值分别＜０．０５和０．０２５）。结论广东汉族人ＲＡ易感性与宿主ＤＲ４基因密切相关，ＨＬＡ－ＤＲ４可能是一个对判断病情和预后有价值的实验指标。