Prognostic Value Evaluation of HLA-DRB1*07:01, *10, *12, *13:01 Alleles for Alloimmunization in Transfusion-Dependent Thalassemia

BackgroundTransfusion is a lifesaving treatment for lots of patients. However, in chronic blood recipients such as thalassemia patients, there are high concerns about alloantibody production that affects the quality of their life. Therefore, research on risk factors of alloimmunization has been started and followed. This study aimed at the determination of correlation probability between some HLADRB1 alleles and alloimmunization in Iranian thalassemia patients.Materials and methodsThe present study was conducted on 60 alloimmunized and 60 non-alloimmunized transfusion-dependent thalassemia patients. Antibody screening and identification tests were carried out using the tube method, and HLA-DRB1 genotyping was done using a single specific primer-polymerase chain reaction (PCRSSP).ResultsThe results of antibody screening showed that the most prevalent alloantibodies were Anti-K (46.7 %), and followed by Anti-E (32.5 %), Anti-C (15.8 %), Anti-D (13.3 %), Anti-e (8 %), and Anti-c (5.8 %), respectively. DRB1*07:01 was detected more in non-responder patients (28.3 %). However, data analysis showed that there is no significant relationship between DRB1*07:01, *10, *13:01 frequency among responder and non-responder groups (p = 0·195, 0.648, 0.402, respectively). There was not any significant correlation between HLA-DRB1*10 and *13:01 allele and alloimmunization. There was a significant association between HLA-DRB1*12 and alloimmunization (p < 0·05, OR = 2.071, CI: 1.716–2.501).ConclusionThe findings of this study represented that there is a significant relationship between HLADRB1*12 and Kell and E alloantibodies. Although more developed studies on other HLA alleles are demanded, these findings can be valuable in determining important alloimmunization risk factors to better management of transfusion complications.     

HLA-DRB1＊15与162例儿童急性淋巴细胞白血病发病的关系

为了研究HLA—DRB1＊15与儿童急性淋巴细胞白血病（ALL）的关系,用特异性引物,对162例15岁以下儿童ALL患者进行HLA—DRB1＊15检测,对HLA—DRB5＊进行PCR扩增,并与1000份健康脐血做显著性比较,计算相对危险度。结果表明：儿童ALL患者DRB1＊15的抗原频率为40．12％,等位基因频率为22．62％;对照组抗原频率为30．8％,等位基因频率为16．81％。两组比较,差别有显著的统计学意义（χ^2=5．560,P=0．018,RR=1．506）。结论：儿童ALL患者HLA—DRB1＊15的抗原频率和等位基因频率均显著高于正常对照,此结果对儿童ALL的发病机制研究有重要意义。     

新等位基因HLA-DRB1＊1462的核苷酸序列分析

本研究验证一个新的HLA等位基因DRB1＊1462的核苷酸序列。应用MR-SSO和MR-SSP基因分型技术对HLA分型,对怀疑是HLA新等位基因进行DNA测序。结果发现,该序列与已知的所有HLA-DRB1等位基因序列不一致,与HLA—DRB1＊140101的序列差异仅表现在第2外显子区域中256位碱基发生了G-〉A的替换,导致相应的氨基酸由丙氨酸变为苏氨酸。结论：该基因是HLA-DRB1位点的一个新等位基因,已经被WHOHLA命名委员会于2006年1月30日正式命名为HLA-DRB1女1462。     

Foetal and neonatal alloimmune thrombocytopenia – The role of the HLA-DRB3*01:01 allele for HPA-1a-immunisation and foetal/neonatal outcome

Foetal and neonatal alloimmune thrombocytopenia (FNAIT) is the platelet counterpart of haemolytic disease of the foetus and newborn. Among Caucasians, around 80 % of FNAIT cases and some of the most severe cases, are caused by alloantibodies against the human platelet antigen 1a (HPA-1a). For around 3 decades it has been known that almost all HPA-1a-immunised women are HLA-DRB3*01:01 positive. The HLA molecule encoded by the HLA-DRA/DRB3*01:01 genes seems to be of crucial importance for initiating the immune response against HPA-1a. The HLA-DRB3*01:01 carrier status is not only important as a risk factor for immunisation, but does also have a significant impact on foetal/neonatal outcome. The possible role of HLA-DRB3*01:01 typing as tool for risk stratification is discussed.     

Genotyping analysis of the MNS blood group system of thalassemia patients with alloantibodies in Iran

BackgroundSerological methods are unreliable for accurate determination of blood group antigens in multi-transfused thalassemia patients. The MNS blood group system has five high-frequency antigens. Many studies demonstrated that some antibodies including anti-S, anti-s, and anti-U may cause acute and delayed transfusion reactions and hemolytic disease of the fetus and newborn. This study aimed to determine the genotype of the MNS blood group in thalassemia patients with alloantibodies by molecular methods.Material and methodsIn this study, 104 blood samples from thalassemia patients were collected. The blood group phenotype for M, N, S and s antigens was determined by the tube hemagglutination method. MNS blood group genotyping was performed using PCR-SSP and DNA Sequencing methods.ResultsAll patients were genotyped with a total of 6 pairs of primers. Discrepancies between genotype and phenotype were observed in 22 patients with S/s alleles and 2 patients with M/N alleles, however, there was full accordance between the results of SSP-PCR and DNA sequencing. The frequency of MNS blood group alleles was determined as follows: 25 % MNSs, 23 % MNss, 21 % MMSs, 9% MMSS, 9% MMss, 8% NNss, 2%MNSS, and NNSS, NNSs, MM genotypes at 1% each.ConclusionIn conclusion, molecular genotyping is more reliable than serological methods in multiple transfusion patients and can lead to a more compatible blood unit for transfusion in these patients.     

四川骨髓库汉族人群HLAD-RB1~*14等位基因分布

目的研究四川骨髓库中四川籍汉族人群HLAD-RB1*14等位基因分布。方法在四川骨髓库8934名四川汉族造血干细胞捐献志愿者中随机筛选107例中低分辨为DRB1*14的样本,以PCR-SBT分型技术检测DRB1位点的第2外显子以鉴定HLAD-RB1*14等位基因。结果共检出6种DRB1*14等位基因,包括DRB1*140101/1454(56.0%),DRB1*1403(1.8%),DRB1*1404(11.0%),DRB1*140501(27.5%),DRB1*140701(1.8%)和DRB1*1425(1.8%);其中最主要的DRB1*140101/1454频率与美国亚裔/太平洋岛民接近,而与高加索人、非洲裔美国人、西班牙裔及韩国人和中国北方汉族中的分布有显著性差异;DRB1*140501和DRB1*1404则与中国北方汉族分布类似;本研究未检到的等位基因在四川汉族DRB1*14阳性个体中的频率低于3%。结论四川汉族人群HLA-DRB1*14等位基因呈现多样性并存在自身特点。     

Unique clinical and pathological features in HLA-DRB1*0401-restricted MBP 111-129-specific humanized TCR transgenic mice

Amino acid residues 111-129 represent an immunodominant epitope of myelin basic protein (MBP) in humans with human leukocyte antigen (HLA)-DRB1*0401 allele(s). The MBP 111-129-specific T cell clone MS2-3C8 was repeatedly isolated from a patient with multiple sclerosis (MS), suggesting an involvement of MS2-3C8 T cells in the pathogenesis. To address the pathogenic potential of the MS2-3C8 T cell clone, we generated transgenic (Tg) mice expressing its T cell receptor and restriction element, HLA-DRB1*0401, to examine the pathogenic characteristics of MS2-3C8 Tg T cells by adoptive transfer into HLA-DRB1*0401 Tg mice. In addition to the ascending paralysis typical of experimental autoimmune encephalomyelitis, mice displayed dysphagia due to restriction in jaw and tongue movements and abnormal gait. In accordance with the clinical phenotype, infiltrates of MS2-3C8 Tg T cells and inflammatory lesions were predominantly located in the brainstem and the cranial nerve roots in addition to the spinal cord and spinal nerve roots. Together, these data suggest a pathogenic role of MBP-specific T cells in inflammatory demyelination within the brainstem and cranial nerve roots during the progression of MS. This notion may help to explain the clinical and pathological heterogeneity of MS.     

Assessment of lung function by spirometry in transfusion-dependent thalassemia patients in a tertiary care center in Sultanate of Oman

Introductionβ-thalassemia’s are hereditary chronic hemolytic diseases, the mainstay of treatment of thalassemia major is regular blood transfusion and iron chelation. They cause many complications, one of the recognized complications related to respiratory system is pulmonary hypertension. Respiratory functions in those patients are not well studied in most of the world. The studies done to assess respiratory function are inconsistent, some found a predominantly restrictive pattern, others found obstructive pattern, and few found normal spirometry. The aim of this study was to assess the spirometric patterns in asymptomatic Omani patients with transfusion-dependent thalassemia using spirometry studies.MethodsTransfusion-dependent thalassemia patients who are registered at Sultan Qaboos University Hospital who are > 15 years old and able to perform spirometry test were selected for the study after they signed the informed consent. All the patients were free of any respiratory disease. Spirometry was performed in all patients in the sitting position and FVC, FEV1, FEV1/FVC were obtained.ResultsTotal number of thalassemia patients enrolled in the study was 37. 32 patients are suffering from thalassemia major and 5 are suffering from thalassemia intermedia. The mean age of our patients was 29.95 years. We found that 37.8 % of the patients showed normal spirometry. Most patients had abnormal spirometry (62.1 %). Of these, 35.1 % showed a restrictive pattern while 27 % showed obstructive pattern.ConclusionSpirometry assessment of the lung function in thalassemia patients who are receiving regular transfusion showed that majority had abnormal spirometry results despite being asymptomatic from a respiratory point of view.     

消化性溃疡与HLA-DRB1基因位点相关性的研究

目的 探讨人类HLA-DRB1等位基因与消化性溃疡的病因与发病机制的相关性.方法 采用聚合酶链反应-序列特异性寡核苷酸探针(PCR-SSO)杂交的方法,对101例消化性溃疡和305例正常对照无偿献血者的血液标本进行HLA-DRB1等位基因的检测.结果 消化性溃疡组的HLA-DRBl*09基因频率明显低于正常对照组(7.43%vs 12.95%,RR=0.539159).结论 HLA-DRB1*09基因对消化性溃疡可能具有重要的抵抗作用.     

HLA-DRB1基因全长序列分子克隆及测序方法的建立

目的建立可靠的HLA-DRB1等位基因全长序列的分子克隆和测序方法。方法设计、合成HLA-DRB1基因全长序列PCR引物和探索PCR反应体系,采用长距离PCR技术,对10份经PCR产物直接测序、基因型已知的标本,扩增HLA-DRB1基因5'-UTR区、6个外显子、5个内含子和3'-UTR区,全长约11 kb。PCR产物纯化后作长片段分子克隆,筛选阳性克隆,提取质粒DNA,采用自行设计的测序引物测序。结果 PCR扩增获得了特异性目的片段,克隆测序后获得了全部10种等位基因的全长序列。将HLA-DRB1等位基因全长序列划分为9个亚区,群体遗传学分析发现第2外显子的平均核苷酸变异率(π)8.653%,高于其他外显子,并且非同义突变率(dn)9.029%大于同义突变率(ds)7.846%。第5内含子的平均核苷酸变异率高达13.690%,DRB1*09∶01∶02和DRB1*07∶01∶01∶02这2个等位基因第5内含子的序列与其他等位基因序列差别较大。结论采用HLA-DRB1基因全长序列分子克隆及测序方法获得了HLA-DRB1基因各亚区多态性分布特点等基础资料。     

The long-term efficacy in blood transfusions,hematologic parameter changes,and complications after splenectomy in patients with transfusion-dependent thalassemia

BackgroundA splenectomy can reduce transfusion requirements in patients with thalassemia. However, the role of a splenectomy remains controversial because its efficacy has not yet been fully determined and there are concerns over potential complications. The purpose of this study was to assess the efficacy, potential changes in hematologic parameters, and any complications associated with splenectomy.MethodsMedical records of 50 patients with transfusion-dependent thalassemia (TDT) who had undergone a splenectomy, along with those of 20 control subjects with intact spleens, were retrospectively reviewed.ResultsThe primary outcomes indicate the efficacy of a splenectomy in reducing red cell transfusions. Fifty TDT post-splenectomy patients were included in this study, of which 28 (56%) were female. The median age of all patients was 20.5 (18–28 years of age). Twenty-seven patients (54%) transformed from TDT to non-transfusion-dependent thalassemia (NTDT) after the splenectomy; 100% with Hb H disease, 58.3% with beta-thalassemia/Hb E disease, and 23.5% with homozygous beta-thalassemia. According to multivariable logistic regression analysis, Hb H disease (adjusted OR 55.23, 95% CI 1.35–22.8.10) and receiving a splenectomy at > ten years of age (adjusted OR 25.36, 95% CI 1.62–396.47) were associated with higher responses. The prevalence of pulmonary hypertension and thromboembolic events were similar between the splenectomy patients and non-splenectomy patients.ConclusionSplenectomy reduced transfusion requirements in TDT patients. The predictive factors as a response to a splenectomy included Hb H disease amongthose receiving a splenectomy at > ten years of age.     

HLA新等位基因A~*9217家系调查分析

背景:目前所发现的HLA等位基因是在人类进化过程中,为了适应某种外界环境过程中逐渐产生的,新近产生而不曾被人们发现的为新等位基因,这些新的等位基因产生诱因千变万化,在诱发因素消除之后这些突变是否持续存在,尚需进一步观察.目的:采用DNA测序技术确认聚合酶链反应-序列特异性寡核苷酸分型检测中的可疑结果,确认新的HLA等位基因,分析新等位基因HLA-A~*9217(WHO注册号:WHS10004629)的遗传方式.设计、时间及地点:以DNA为观察对象的开放性试验.其中常规初检聚合酶链反应-寡核苷酸探针分型反向杂交荧光微珠法于2007-11在河南省红十字血液中心中华骨髓库河南分库人类白细胞抗原组织配型实验室完成,测序实验于2008-02在戴诺生物技术(北京)有限公司人类白细胞抗原实验室完成.材料:中华骨髓库志愿捐献者(编号:371xxxxx)及其直系3代亲属血样,受试者共9人在实验室逐个登记,采集静脉血5 mL,EDTA抗凝.方法:用聚合酶链反应-序列特异性寡核苷酸流式荧光微珠HLA分型方法、SBT测序分型方法,对A~*9217携带者家庭成员进行中低分辨,DNA测序高分辨检测分析,血清学方法检测ABO,MN,Rh血型系统,分析新基因的发现及遗传方式.主要观察指标:HLA新等位基因A~*9217携带者A位点外显子3序列对比.结果:通过受检者HLA分型结果和红细胞系统ABO,MN,Rh血型分析,先证者A~*9217应来自父方,但父方并没有检出A~*9217,而是检出了与之序列相近的A~*020301,与A~*9217在处显子3有3处碱基改变391 T>G,414 C>G,418 T >G,A~*9217携带者的2个孩子均检出A~*9217.结论:新基因A~*9217先证者父方产生突变所致,该突变能正常遗传给后代,该基因是否能稳定遗传下去有待进一步观察研究.     

DNA序列分析鉴定HLA-DRB1~*新等位基因DRB1~*1449

目的鉴定HLADRB1位点新等位基因。方法应用基因克隆和DNA测序技术对1例受检样本HLADRB1基因第2外显子(Exon2)作核苷酸序列分析,与已知等位基因序列比对并作血清学分型。结果基因组DNA和DNA克隆的测序结果一致;DRB1Exon2的核苷酸序列与已知等位基因序列均不相同,与同源性最高的HLADRB11432相比,存在4处碱基的改变;以Exon2的第1位碱基为记数起点,核苷酸71位C→>G,196位G→>A,244位T→>G和245位上G→>T,引起相应编码氨基酸28位上天门冬氨酸(Asp)→>谷氨酸(Glu),70位上精氨酸(Arg)→>谷氨酸盐(Gln),86位上缬氨酸(Val)→>氨基乙酸(Gly)。血清学分型结果表明新等位基因血清学特异性为DR14。结论被检标本HLADRB位点存在新等位基因,2005年2月WHOHLA命名委员会正式将其命名为HLADRB11449。     

HLA-DRB1基因与湘西土家、汉族人群不明原因反复流产的相关性研究

目的 探讨HLA-DRB1基因多态性与湘西土家族、汉族人群不明原因反复流产(URSA)的遗传关联性.方法 采用聚合酶链反应-序列特异性引物(PCR-SSP)技术,对76例湘西土家族、68例汉族人群URSA患者和82例湘西土家族、75例汉族人群健康对照者的HLA-DRB1等位基因进行分析.结果 ①土家族、汉族URSA组的DRB1*04基因频率显著高于土家族及汉族对照组(RR>1,PC<0.01);DRB1*12的基因频率显著低于对照组(RR<1,PC<0.01).②土家族URSA组的DRB1*07基因频率明显高于汉族URSA组(18.08%比5.28%,PC<0.01).结论 HLA-DRB1*04可能是湘西土家族、汉族人群不明原因反复流产的易感基因,HLA-DRB1*12可能为保护基因.     

轻型地中海贫血者的血液学实验参数研究

目的探讨轻型地中海贫血的血液学检测参数变化及临床诊断意义。方法应用全自动血细胞分析仪检测红细胞各参数,用醋酸纤维薄膜电泳检测血红蛋白有无异常区带,一管法测定红细胞脆性,层析柱法测HbA2,抗碱法测HbF,Wright氏染色后镜检红细胞形态。结果对77例轻型地中海贫血、32例缺铁性贫血与27例健康人的检测结果显示,轻型地中海贫血者红细胞MCV及MCH等参数、红细胞脆性试验、HbA2及HbF与对照组有显著性意义(P<0.01)。结论上述相关血液学检测指标是筛查轻型地中海贫血的可靠指标,对Hb正常,RBC数高的疑似病人,应建议做地中海贫血基因检测,以进一步诊断。     

HLA-DRB1 differences in allelic distribution between familial and sporadic multiple sclerosis in a Hellenic cohort

Objective: Familial Multiple Sclerosis (fMS) is reported to have distinct clinical and imaging characteristics in comparison to the sporadic disease (sMS). Nevertheless, the genetic/immunogenetic profile of fMS has never been investigated in depth, so far. In this study, we examined differences of HLA-DRB1 allelic frequencies between 57 fMS and 141 sMS Hellenic patients, with reference to 246 previously genotyped healthy controls (HCs).

Patients and Methods: All patients underwent medical interview and DRB1 genotyping, using a low-resolution SSOP technique. Statistical analyses were performed using SPSS v.21.0 software, with significance set at 0.05, and p value corrected according to the Benjamini–Yekutieli method.

Results: 29 fMS cases had at least one 1^st degree relative affected (fMS 1^st), while the rest had at least one 2^nd or 3^rd degree relative affected (fMS 2^nd/3^rd). Parent-of-origin effects were observed, with the prevalence of maternal inheritance. Frequency of DRB1*15 was significantly increased in fMS and sMS, in comparison to HCs (p = 0.002 and <0.001, respectively). After fMS stratification, this result was mainly attributed to the fMS 2^nd/3^rd subgroup. DRB1*11 frequency was significantly decreased only in sMS (p < 0.001) with fMS approximating HCs’ frequency, especially for the fMS 1^st subgroup. Heterozygosity was favored over homozygosity in all groups.

Conclusion: We propose possible HLA-DRB1 allelic distribution differences between fMS and sMS, which become more apparent as proximity of affected relative/-es in fMS increases, supporting a rather degraded role of DRB1 alleles in fMS HLA/immunogenetics and indicating the concomitant implication of other HLA and non-HLA polymorphisms.     

Decreased cortisol production in male type 1 diabetic patients

BACKGROUND: It is unclear whether cortisol production and the 11betaHSD-mediated cortisol to cortisone interconversion are different between type 1 diabetic patients and healthy subjects. MATERIALS AND METHODS: Fourteen male, nonobese, normotensive type 1 diabetic patients without severe complications (HbA1c < 8.5%) were studied twice during a daily sodium intake of 50 and 200 mmol, and were then compared with 14 individually matched healthy subjects. Cortisol production was assessed by the sum of urinary cortisol metabolite excretion. Urinary ratios of (tetrahydrocortisol + allo-tetrahydrocortisol)/tetrahydro-cortisone [(THF + allo-THF)/THE] and of free cortisol/free cortisone [UFF/UFE] were determined as parameters of 11betaHSD activity. RESULTS: Sum of urinary cortisol metabolite excretion during low- and high-salt diet was 7.4 +/- 2.5 vs. 7.7 +/- 2.3 nmol min-1 m-2 (NS) in diabetic patients and 9.7 +/- 2.1 vs. 11.2 +/- 4.1 nmol min-1 m-2 (NS) in healthy subjects, respectively (P < 0.05 vs. healthy subjects at both diets). The allo-THF excretion and allo-THF/THF ratios were lower in the diabetic than in the healthy males during both diets (P < 0.05). Urinary (THF + alloTHF)/THE and UFF/UFE were similar in both groups and remained unchanged after salt loading. CONCLUSIONS: The sum of urinary cortisol metabolite excretion as a measure of cortisol production is lower in nonobese, normotensive type 1 diabetic males with adequate glycaemic control and without severe complications, irrespective of sodium intake. We suggest that this is at least in part as result of diminished 5alpha reductase activity, resulting in a decreased cortisol metabolic clearance. In type 1 diabetic and in healthy males, the 11betaHSD setpoint is not affected by physiological variations in sodium intake.