关键词： 地中海贫血   实验筛查   进展  

摘要： 地中海贫血是由于珠蛋白肽链合成障碍而导致的一组遗传性溶血性疾病,是世界上最常见和发病率最高的遗传性溶血性贫血,也是危害最严重的血红蛋白病之一。由于该病至今尚无有效的治疗方法,预防是最有效的措施。因此,早期进行地贫筛查并对高危孕妇进行地贫基因诊断是预防中、重型地中海贫血患儿出生的最有效措施。目前用于地中海贫血筛查的技术有:血细胞分析、红细胞渗透脆性实验、血红蛋白电泳、毛细管电泳法、高效液相色谱技术(HPLC)等。

关键词： 造血干细胞移植   重型β地中海贫血   移植物抗宿主疾病   细胞因子  

摘要： 研究背景重型β地中海贫血(β-thalassemia major)又称珠蛋白生成障碍性贫血,为进行性溶血性贫血,以特殊地中海贫血面容、肝脾肿大及血红蛋白F明显升高为特征的一种血液系统疾病。重型β地中海贫血患者必需定期输血及应用去铁药物。异基因造血干细胞移植是目前根治地中海贫血的唯一方法。但是因为重型p地中海贫血患者因本身的骨髓富细胞特性及反复输血的致敏效应,使得骨髓移植相对于其他疾病而言具有更大的难度,GVHD是目前造成移植失败和移植相关死亡的重要因素。预处理、感染等引起胃肠道黏膜上皮细胞损伤,使细菌的降解产物如脂多糖进入血液,引发“细胞因子风暴”,是GVHD致靶器官各种病理损害的重要原因之一。相关研究显示细胞因子在GVHD的发生发展中起着至关重要的作用。IL-2和TNF-α是研究最多的细胞因子。Miyamoto等率先报道了移植后第3天sIL-2R水平可预测急性GVHD, Mathias等报道sIL-2R水平与急性GVHD的发病及严重程度有关。Choi等测量438名采用清髓预处理患者移植后第7天可溶性的TNFR1水平增加超过原来的2.5倍,与Ⅱ-Ⅵ度急性GVHD发生存在相关性,增加了移植相关死亡率。Berger等报道含有较高的TNF时则预示发生GVHD的风险率增高。众多研究提示IL-10升高可能降低GVHD的发生。尽管相关研究表明细胞因子与GVHD关系密切,但还有很多细胞因子与GVHD的关系尚不清楚,同时地中海贫血为非肿瘤性疾病,重型p地中海贫血患者发生GVHD可能存在其特殊性。因此,了解重型β地中海贫血患者造血干细胞移植后发生GVHD时细胞因子的变化,为临床防治重型β地中海贫血造血干细胞移植后GVHD,提高重型p地中海贫血患者的生存质量,具有重要意义。本研究采用液相芯片技术检测重型β地中海贫血患者造血干细胞移植后发生GVHD前后IL-6、IL-8、IL-12 TNF-α、MIF的变化,评估其是否可以作为GVHD发生及疗效的监测指标。研究对象与方法对象：选取南方医科大学附属南方医院儿科病房2011年7月～2015年3月52例接受造血干细胞移植治疗患儿,其中47例为重型地贫患儿,5例非地贫发生aGVHD为对照。重型地贫患儿均作了地贫基因检查,基因型为β地贫基因纯合子或双重杂合子,其中男27例,女20例,中位年龄6.28岁(年龄范围在2-14岁)。移植前47例重型β-地贫患儿都按国内地贫临床分度(根据年龄、肝大情况和铁蛋白水平分为三度),类似于Pesaro分度,其中4例为Ⅰ度,42例为Ⅱ度,1例为Ⅲ度。亲缘干细胞移植24例,非亲缘干细胞移植23例,骨髓干细胞移植2例,外周干细胞移植29例,脐血干细胞移植7例,骨髓+脐血干细胞移植4例,骨髓+外周干细胞移植1例,外周+脐血干细胞移植4例。41例为HLA配型全相合(6/6,8/8或10/10),3例患者符合HLA 8/10相合,1例患者符合HLA 7/8相合,2例半相合。16例地贫发生GVHD中,Ⅰ度2例,Ⅱ度4例,Ⅲ度8例,Ⅵ度2例。预处理方案：对大部分采用环磷酰胺/白消安为基础的清髓性方案：环磷酰胺(50～60mg/kg/d,-10d～-9d);白消安(2.8～4.4mg/kg/d,-8d～-6d);氟达拉滨(40mg/m2/d,-6d--2d);噻替哌(10mg/kg/d,-5d);加或不加抗T-淋巴细胞兔免疫球蛋白(5～10mg/kg/d,-3d～-1d)或兔抗人胸腺细胞免疫球蛋白(2.5mg/kg/d,-3d～-1d)。预防治疗GVHD:采用环孢霉素A、霉酚酸酯联合甲氨蝶呤常规预防GVHD, GVHD诊断按照西雅图诊断标准。GVHD的治疗：仅有局部皮疹的aGVHD,可不治疗或局部涂用可的松软膏控制。但如果有如皮疹面积迅速扩大,皮肤损害程度加重,出现发热、流感样症状,或怀疑有肠道或肝脏GVHD等情况时给予全身性治疗。急性GVHD的一线治疗：甲基泼尼松龙2mg/kg/d分次给予或地塞米松；二线治疗：霉酚酸酯,普乐可复,人T-淋巴细胞兔免疫球蛋白,CD25单抗,抗淋巴细胞球蛋白等。慢性GVHD一线治疗：甲基强的松龙和环孢素交替使用；二线治疗：硫唑嘌呤、普乐可复、霉酚酸酯、环磷酰胺、甲氨蝶呤治疗等。造血干细胞的采集与输注：在预处理完成后,次日行造血干细胞移植物输注,骨髓干细胞+脐血干细胞移植则在移植当天先将供者体内采集的骨髓造血干细胞直接从中心静脉尽快输入,次日上午输注脐血干细胞。输注的CD34+干细胞数的均值为7×106/kg。并发症的防治：(1) GVHD预防：环孢霉素A移植前-10天至-2天予1.5mg/kg/d静脉滴注,移植前-1至移植后+25天剂量增至3mg/kg/d静脉滴注,1月后静脉滴注改为口服,根据浓度调整环孢素A口服量,维持血药浓度200±50ng/ml,移植2月后

关键词： Iran   Xmn1 polymorphism   α-thalassemia   β-thalassemia intermedia   β-thalassemia major  

摘要： Background: Thalassemia syndromes are the most prevalent single gene disorders in Iran. This study aimed to evaluate the effect of different types of beta-globin gene mutations, co-inheritance of alpha-globin gene mutations and/or Xmn1 SNP on disease phenotype in a large cohort of Iranian patients. Subjects and Methods: In total, 433 patients were clinically classified into β-thalassemia major (TM) or intermedia (TI). Multiplex PCR, ARMS-PCR, RFLP-PCR and DNA sequencing were performed to identify both α-and β-globin gene mutations and Xmn1 polymorphism as well. All data were compared and analyzed by SPSS software in TM and TI groups, consequently. Results: A total of 39 different β-globin mutations were identified. Among them, the most common were IVS IInt1 (40.33%) followed by IVS Int5 (9.56%), C30 (7.22%) and Fr8-9(7%). All patients were subjected to evaluate common a-globin gene deletions. The patients inherited concomitant mutations of a- and β-globin, showed no clinical modifications compared with those who had only β-globin mutation. The TI patients showed a significant increase in frequency of both heterozygous and homozygous form of the Xmn1 polymorphism. It was also found that β⁰/β⁰ genotype patients, inherited the Xmn1 polymorphism required lesser blood transfusion. Conclusion: No significant differences were observed, on the severity of disease, between patient's inherited defective α- and β-globin genes and ones with just β-globin gene mutation. Taking the results of this research into account, Xmn1 polymorphism can be considered as an important genetic factor modulating the severity of disease. © 2015, Tehran University of Medical Sciences (TUMS). All rights reserved.

关键词： GENETICS of thalassemia   ANIMAL experimentation   BIOLOGICAL models   ELECTROPHORESIS   GENES   RESEARCH methodology   MICE   MUTATION (Biology)   POLYMERASE chain reaction   FINANCING of research   TISSUE culture   WESTERN immunoblotting   FLUORESCENCE in situ hybridization  

摘要： Mouse models that carry mutations causing thalassemia represent a suitable tool to test in vivo new mutation-specific therapeutic approaches. Transgenic mice carrying the beta-globin IVSI-6 mutation (the most frequent in Middle-Eastern regions and recurrent in Italy and Greece) are, at present, not available. We report the production and characterization of a transgenic mouse line (TG-beta-IVSI-6) carrying the IVSI-6 thalassemia point mutation within the human beta-globin gene. In the TG-beta-IVSI-6 mouse (a) the transgenic integration region is located in mouse chromosome 7;(b) the expression of the transgene is tissue specific;(c) as expected, normally spliced human beta-globin mRNA is produced, giving rise to beta-globin production and formation of a humanmouse tetrameric chimeric hemoglobin mu beta-globin 2 /hu beta-globin 2 and, more importantly, (d) the aberrant beta-globin-IVSI-6 RNAs are present in blood cells. The TG-beta-IVSI-6 mouse reproduces the molecular features of IVSI-6 beta-thalassemia and might be used as an in vivo model to characterize the effects of antisense oligodeoxynucleotides targeting the cryptic sites responsible for the generation of aberrantly spliced beta-globin RNA sequences, caused by the IVSI-6 mutation. These experiments are expected to be crucial for the development of a personalized therapy for beta-thalassemia.

关键词： Prenatal Screening   Gene Pool   beta-Thalassemia   Homozygote   Consanguinity   Mutation   Nepal   Awareness  

摘要： Mutation spectrum varies significantly in different parts and different ethnic groups of India. Social factors such as preference to marry within the community and among 1st degree relatives (consanguinity) play an important role in impeding the gene pool of the disease within the community and so in society by and large. The present paper discusses the role of consanguinity in profiling of beta thalassemia mutation, and thus the approach for prenatal screening and prevention based awareness programme. Clinically diagnosed 516 cases of beta thalassemia were screened at molecular level. A detailed clinical Proforma was recorded with the information of origin of the family, ethnicity, and consanguinity. The present study reports that subjects originating from Uttar Pradesh, Uttarakhand, Bihar, and Jharkhand have c.92+5G>C and c.124-127delTTCT mutation as the commonest mutation compared to the subjects hailing from Madhya Pradesh and Chhattisgarh and Nepal where sickle mutation was found more common. In 40 consanguineous unions more common and specific beta mutations with higher rate of homozygosity have been reported. This consanguinity-based data helps not only in deciding target oriented prenatal diagnostic strategies but also in objective based awareness programmes in prevention of thalassemia major birth. © 2015 Ravindra Kumar et al.

关键词： 地中海贫血   溶血性贫血   遗传筛查   产前诊断   贵州地区   基因突变类型  

摘要： 目的：本研究通过对患有地中海贫血的夫妻及其胎儿的血液学相关检查的分析了解贵州地区孕妇及其胎儿的地贫基因突变类型，并进一步了解本地区地中海贫血基因的分布及产前诊断状况，为我省地贫遗传学咨询提供参考资料。　　方法：收集2012年10月至2014年10月共2年时间在贵阳医学院附属医院及贵州省人民医院产前诊断中心行地中海贫血产前诊断的患者资料，通过检查平均红细胞容积（MCV）、血红蛋白（Hb）、平均红细胞血红蛋白（MCH）进行初步筛查，当患者的MCV＜80fl，Hb＜110g/L，MCH＜27pg时进行血红蛋白电泳检测，当血红蛋白异常区带或血红蛋白A2(HbA2)含量异常者，可以用RDB和PCR法行地中海贫血的基因分型，分析其基因类型和产前诊断结果。　　结果：1.在受检的82例患者中，共检出α-地中海贫血患者有26例，占31.7％，β地中海贫血患者有56例，占68.3%。26例α-地中海的贫患者中，检测出中国人最常见的缺失型α地中海贫血（--SEA、α3.7、α4.2）;56例β-地贫阳性患者的标本中，检测出中国人常见的7种基因突变类型，以CD17（A→T）、CD41-42（－TCTT）、IVS－2－654（C→T）最常见。2.双方基因诊断为均携带α地贫或携带有β地贫的夫妻有51对，采用抽脐血或羊水的方式进行产前基因诊断，产前诊断为轻型地贫的胎儿有33例，地贫阴性胎儿有10例，中重型地贫胎儿有8例。其中，产前诊断检测出患有重型β地中海贫血的胎儿有5例，1例胎儿为bart's水肿胎，2例胎儿患有血红蛋白 H病，均采取终止妊娠的措施。3.在产前筛查中平均红细胞血红蛋白（MCH）、血红蛋白（Hb）以及平均红细胞体积（MCV）在α、β两者之间的差异无统计学意义;血红蛋白A2在α和β之间的差异具有统计学意义。　　结论：1.在本研究中，地中海贫血患者的发生率有较为显著的差异，但其基因突变率有相似的趋势，其中，β-地贫基因携带者的检出率明显要高于α-地贫基因携带者;2.贵州地区是地贫疾病的高发地带，通过产前筛查可以避免重型及中间型地贫患儿的出生，为本地区人群产前基因诊断，生育指导以及提高人口素质具有及其重要的意义。3.羊膜腔、脐静脉穿刺法在地中海贫血检出率的差异无统计学意义。4.定量血红蛋白电泳和红细胞参数是地中海贫血筛查的两项重要指标，通过PCR-RDB、gap-PCR方法可以快速有效诊断母胎地贫基因型，以干预地贫出生缺陷。

关键词： Alloantibody   Thalassemia   Transfusion  

摘要： Background Multiple transfusions in thalassemia patients may lead to antibody production against blood group antigens and hemolytic transfusion reaction might occur. In this study, antibody screening test was performed by tube and gel methods to determine the prevalence and specificity of alloantibodies in thalassemia patients. Materials and Methods In this cross-sectional study, overall of 100 thalassemia patients from Yazd thalassemia clinic were recruited from July to September 2013. Two blood samples with volume of 6 ml were collected from each patient for standard tube and gel method antibody screening tests and a questionnaire consisting of demographic, health and blood transfusion status was completed. Results Out of 100 cases, 54 were female (54%) and 46 male (46%). The patients' age mean was 14.97 +/- 7.91 years with 2 to 33 years age range. Only 4% (n=4) had developed alloantibodies. (One patient developed dual alloantibody (Anti-C and Anti-D) and three patients developed single alloantibody (Anti-K)). Gel method detected 4 patients with alloantibody but in two patients not detected by the standard tube method. Conclusion The prevalence of RBC alloantibody production in this study was less than most previous studies. Anti-K was the most prevalent alloantibody in thalassemia patients in Yazd. It seems Rh and Kell blood group phenotyping in a newly diagnosed thalassemia patient and selection of matched blood for transfusion is very important.

关键词： 孕前检查   数据挖掘   阈值   地中海贫血   分类模型   叶酸  

摘要： 在中国,每一年婴儿诞生后的缺陷率和0～4岁期间出现的先天残疾率共计5%左右,这就表明我国每年新增先天残疾儿童100多万。为保证一个健康的孩子的出生,孕前检查显得尤为重要。本课题主要应用数据挖掘和数据统计方面的知识和工具,针对孕前检查相关的两项内容,地中海贫血筛查和人体叶酸含量指标进行研究。在研究地中海贫血筛查分类模型过程中,对于多属性值的分类问题,大多采用人为设定阈值确定特定的特征属性。这种方法有很大的主观随意性从而影响分类的准确度。本文针对此问题,提出了一种解决方法。首先应用巴氏距离算法对特征属性排序；然后应用遗传算法选取最优的支持向量机模型,通过遍历搜索的方式对特征属性逐一剔除分别得到分类准确度；最后选取分类准确度最高的阈值点。实验结果表明,本方法优于手动方法,用于地中海贫血筛查有良好效果,尤其对于大样本数据阈值处理效果比较明显。在地中海贫血筛查方面的研究,主要通过统计云南省人群在不同民族和不同地区间中地中海贫血发病率,了解地中海贫血和民族、地区之间的关系。在传统方法利用血常规检测数据和血红蛋白电泳检测数据基础上,通过增加了正常人群与患地中海贫血人群中的性别、年龄、籍贯和民族等特征属性,利用常规的六种地中海贫血分类模型进行测试,筛选出了最优的logistic回归和支持向量机模型对地中海贫血进行筛查。实验结果表明,云南省人群中地中海贫血的发病率与民族和地区没有显著的关联关系。但是通过六种分类模型,对比传统的地中海贫血筛查准确率和诊断比数比,筛查准确率和诊断比数比都有明显的提高。另外,在叶酸含量方面的研究,主要通过了解云南省育龄人群红细胞叶酸水平,建立云南省地区的红细胞叶酸参考范围。通过检测有不良孕产史育龄人群的红细胞叶酸与正常人群红细胞叶酸水平,了解叶酸缺乏与不良孕产史之间的关系。统计结果表明,云南地区普通育龄人群20-29岁女性红细胞叶酸参考水平为11.31～35.73 ng/ml,20～29岁男性为10.85～33.59 ng/ml,女性叶酸测定数值较男性为高；在云南省各个民族之间叶酸水平差异无统计学意义(P>0.05)。通过对正常人群组2639例与不良生育史246例患者的分析,正常人群组与脑部异常儿生育史患者的叶酸水平比较,差异有统计学意义(P<0.05),正常人群组与胎儿畸形、自然流产和胎停育、神经管缺陷引产史、唐氏综合征生育史、其他染色体异常生育史组间叶酸水平比较,无统计学差异(P>0.05)。利用自动寻找最优阈值算法,能够更好的得到特征属性的最佳阈值点,从而使得的地贫筛查得到最优分类模型,能够更好获得临床筛查结果。通过建立云南地区红细胞叶酸参考范围为云南地区育龄人群补充叶酸提供了理论依据,而增补叶酸对预防出生缺陷有一定意义,育龄人群应适时增补叶酸。

关键词： β地中海贫血   KLF1突变   群体遗传   基因型-表型   修饰基因  

摘要： 背景与目的p地中海贫血(β-thalassemia,简称p地贫)是一种常见的严重威胁人类健康的致死、致残的遗传性血液病,主要分布在全球的热带和亚热带地区,在我国广东和广西是发生率最高的两个省区,其基因携带率分别高达2.54%和6.43%。β地贫为常染色体单基因隐性遗传病,是由染色体11p15.3的β-globin基因突变导致p珠蛋白肽链合成减少或缺失而导致的一种疾病,β地贫携带者(轻型β地贫；β-thalassemia trait, TT)婚配的下一代有1/4的机会罹患有严重溶血性贫血症状的中、重型p地中海贫血(P-thalassemia intermedia, TI; β-thalassemia major,TM)。值得注意的是同是纯合子或双重杂合子的β地贫患者,其临床表型从中间型(Hb 60-100 g/L)到重型(Hb<60 g/L,严重者20-40 g/L)存在很大差异,研究表明β地贫的表型变化首先是由其致病基因的突变来决定的,即临床表现与β珠蛋白基因的突变基因型直接相关,β基因完全被抑制的β0突变纯合子(β0/β0)一般都表现为重型β地贫,而β基因部分被抑制的旷突变纯合子(β+/β+)则多表现为中间型β地贫,而双重杂合子(β0/β+)可能表现为中间型β地贫,也可能是重型β地贫。目前无论是在人类还是在小鼠模型的研究中均已有报道一些β-globin基因以外的基因对β地贫临床表型的严重性起着修饰作用,例如分子伴侣AHSP (Alpha Hemoglobin Stabilizing Protein), HRI基因(Heme-regulated eIF2alpha kinase)等等。但下列两种能减轻α/β链比例失衡程度的遗传修饰因素是最主要因素：(1)患者同时合并α地贫突变而减轻(反之,合并α-globin基因三联体或多拷贝而加重)；(2)患者同时合并引起胎儿血红蛋白(Fetal hemoglobin, Hb F; α2γ2)升高的遗传变异缓解疾病的严重性。我们课题组之前的研究证实AHSP基因变异不是中国人p地贫临床表型的主要遗传因素,此外,我们还系统研究了中国人中间型p地贫的分子缺陷原因,结果提示还存在其他一些未知的遗传修饰因素对β地贫的表型变化有贡献。EKLF/KLF1是Kruppel样转录因子家族(Kruppel-like transcription factors, KLFs)成员之一,其羧基端含三个串联的锌指结构域,通过特异性的识别结合保守的DNA序列5'-CCMCRCCCN-3'启动下游基因的转录从而调节红细胞一系列的生长发育过程。近几年研究证明,KLF1作为一个重要的调控因子参与调控了胎儿血红蛋白(Hb F)向成人血红蛋白(Adult hemoglobin A, Hb A; α2γ2)的转化过程；2010年的一项研究发现KLF1的显性突变(p.E325K)可以导致先天性红细胞生成异常性贫血四型(Congenital dyserythropoietic anemia, CDAⅣ)；2014年报道了KLF1双重突变可以导致先天性的慢性非球形红细胞溶血性贫血(Chronic nonspherocytic hemolytic anemia, CNSHA);但是有更多的研究显示KLFl单等位基因的突变仅仅导致一系列的良性表型,如In(Lu)稀有血型,Hb F和成人血红蛋白A2 (Adult hemoglobin A2,Hb A2; α2δ2)升高(这两点均为p地贫的重要特征,且Hb F升高有利于改善p地贫患者临床表型严重性)等。然而,目前没有任何研究展开基于人群遗传学的KLFl突变研究,亦没有关于KLF1突变与β-globin突变共遗传的表型研究。因此,本次研究首次通过分子筛查一个大样本中国人群来调查KLFl基因的突变率与突变谱,并通过表型研究来评估KLF1突变是否对p地贫具有遗传修饰作用。设计与方法一、研究人群、病人和表型检测本研究通过随机抽样对长期居住在中国南方地区的3918例非亲缘关系个体进行了突变分析,这个大人群队列包括：3839例连续采集的样本和79例表型导向型样本(Hb A2水平：3.3%-4.1%和/或Hb F水平大于1.5%)。KLF1人群突变频率通过调查3839例连续采集的样本得出,而KLF1突变谱由所有3918例筛查到的所有突变组成,所有KLF1突变的阳性样本均用来进行表型分析。3839例连续采集的样本(年龄1-50岁；1987例男性,1852例女性)由三个队列组成：队列A,1971例非地贫人群；队列B,946例β地贫携带者；队列C,922例p地贫中重型病人(根据回顾性的临床指标分为568例重型,354例中间型)。队列A从本课题组前期对广西地区进行地贫流行病学调查的样本中随机抽出,队列B和C样本收集于

摘要： Background. Nontransfusion dependent thalassemia (NTDT) is a milder form of thalassemia that does not require regular transfusion. It is associated with many complications, which differ from that found in transfusion-dependent thalassemia (TDT). Currently available information is mostly derived from beta-NTDT;consequently, more data is needed to describe complications found in the alpha-NTDT form of this disease. Methods. We retrospectively reviewed the medical records of NTDT patients from January 2012 to December 2013. Complications related to thalassemia were reviewed and compared. Results. One hundred patients included 60 females with a median age of 38 years. The majority (54 patients) had alpha-thalassemia. Overall, 83 patients had one or more complications. The three most common complications were cholelithiasis (35%), abnormal liver function (29%), and extramedullary hematopoiesis (EMH) (25%). EMH, cardiomyopathy, cholelithiasis, and pulmonary hypertension were more commonly seen in beta-thalassemia. Osteoporosis was the only complication that was more common in alpha-thalassemia. The risk factors significantly related to EMH were beta-thalassemia type and hemoglobin < 8 g/dL. The risk factors related to osteoporosis were female gender and age > 40 years. Iron overload (ferritin > 800 ng/mL) was the only risk factor for abnormal liver function. Conclusion. The prevalence of alpha-NTDT complications was lower and different from beta-thalassemia.