广西百色地区血红蛋白Westmead的实验及临床研究
发布时间：2018-07-13 11:45
【摘要】：目的:(1)评价利用聚丙烯酰胺凝胶肽链电泳(PAGE)检测异常血红蛋白Westmead(HbWS)区带的准确性,从检测蛋白质肽链层面提供筛查和诊断HbWS的新方法。(2)探讨百色地区人群中HbWS的基因型和血液学表型特点及其与其他不同基因型的地中海贫血(地贫)的血液学表型差异。方法:(1)实验研究:收集2015年-2016年、籍贯系百色地区、在我院进行地贫基因检查、年龄在2-60岁之间的病例,以基因检测作为诊断HbWS的金标准、同时应用PAGE方法检测HbWS区带即具有Gγ位置区带浓染而Aγ位置无浓染(HbF值正常)为HbWS区带阳性,通过四格表计算PAGE阳性诊断HbWS的敏感性、特异性等各项指标。(2)临床研究:收集2010年-2016年、来源于百色地区、门诊确诊为HbWS的不同基因型携带者(患者)为研究对象,根据纳入标准及排除标准从中筛选出HbWS各类型基因突变的携带者(患者)作为观察组,筛选出其他相应各类地贫基因型携带者(患者)分成若干对照组(亚组);所有样本均进行自动血细胞分析仪、血红蛋白自动分析仪进行血液学分析;采用Gap-PCR检测缺失型α-地贫和反向斑点印迹(RDB)法检测非缺失型α-地贫和β-地贫。应用SPSS 19.0统计软件对各组数据进行处理。结果:(1)入组病例600例,取得的样本经PAGE实验诊断HbWS区带阳性130例、阴性470例;同时按金标准方法共检出含HbWS基因突变的126例,无HbWS基因突变的474例。PAGE诊断HbWS的诊断符合率为98.00%、敏感度96.83%、特异度98.31%、阳性预测值0.9385、阴性预测值0.9915、阳性似然比57.3、阴性似然比0.0249。应用ROC曲线分析PAGE诊断HbWS的曲线下面积AUC为0.942,95%置信区间为(0.893,0.991)。(2)收集确诊为HbWS的不同基因型组合携带者(患者)共292例,基因类型组合分为八大种类,其中hbws杂合子(αwsα/αα)174例(占59.59%)、hbws复合轻型β-地贫43例(占14.73%)、hbws复合其它静止型α-地贫的双重杂合子30例(10.27%)、hbh-ws病28例(占9.59%)、含hbws的轻型α-地贫复合β-地贫8例(占2.74%)、hbh-ws病复合轻型β-地贫4例(占1.37%)、hbws复合重型β-地贫3例(占1.03%)和hbws纯合子2例(占0.68%)。(3)117例hbws杂合子组的hb(129.68±21.11g/l)、mcv(86.00±5.36fl)、mch(27.76±2.16pg)在正常范围内,其均值低于正常人组但高于αcsα/αα组、αqsα/αα组、αα/-α~(3.7)组和αα/-α~(4.2)组(该四个组mch均低于正常参考值);rdw-cv(13.10±1.10%)与其他静止型α-地贫比较无差异但高于正常人组。(4)hbws复合其它静止型α-地贫的双重杂合子的hb在正常范围(128.60±15.67g/l),高于hbws复合轻型β-地贫组和轻型β-地贫组;mcv(80.27±4.70fl)在临界范围、mch(25.29±1.52pg)低于正常参考值范围,但高于hbws复合轻型β-地贫组;rdw-cv(13.38±1.27%)低于hbws复合轻型β-地贫组、轻型β-地贫组和标准型的α-地贫。(5)hbws复合轻型β-地贫组的hb(112.59±14.03g/l),mcv(64.48±5.53fl)、mch(19.72±1.81pg)低于正常范围,表现为小细胞低色素性轻度贫血与轻型β-地贫组的hb、mcv和mch差异无统计学意义,rdw-cv(15.67±1.34%)低于轻型β-地贫组。(6)hbh-ws病的hb(104.67±27.03g/l)、mcv(67.76±6.10fl)、mch(20.64±1.82pg)与-α~(4.2)/--sea组、-α~(3.7)/--sea组一样表现为小细胞低色素性轻度贫血但hb水平比后两组高(p0.05);αwsα/--sea组的mcv低于αcsα/--sea组(p0.05),mch减低程度最轻,rdw-cv增加程度最轻(p0.01)。结论:(1)page方法hbws区带阳性诊断hbws的特异性、敏感性和诊断符合率高,准确性接近基因检测方法,可作为hbws新的筛查诊断方法供临床应用参考;此外,page方法还可检测ζ链诊断成年人标准型α-地贫。(2)百色地区hbws的基因突变类型主要以αwsα/αα为主,αwsα/αα的血液学表型基本正常,筛查时容易漏诊;hbws复合其它静止型α-地贫的双重杂合子,表现为小细胞低色素;hbws复合轻型β-地贫组表现为小细胞低色素性轻度贫血;hbh-ws病也表现为小细胞低色素性轻度贫血,但贫血程度比-α~(4.2)/--SEA组、-α~(3.7)/--SEA组轻。(3)HbWS不同基因型血液学表型各有其特点,但是贫血程度都要比同型的地贫较轻,研究结果对该地区地贫的遗传咨询和产前诊断有指导意义。
[Abstract]:Objective: (1) to evaluate the accuracy of detection of abnormal hemoglobin Westmead (HbWS) zone by polyacrylamide gel electrophoresis (PAGE), and to provide a new method for screening and diagnosing HbWS from the detection of protein peptide chain. (2) to explore the genotype and hematological phenotypes of HbWS in the population of Baise and the Mediterranean with other different genotypes. Haematological phenotypic difference of anemia (ground poor). Methods: (1) experimental study: in 2015 -2016 years, the native place of Baise region, in our hospital in the poor gene examination, age between 2-60 years of age, gene detection as the gold standard for the diagnosis of HbWS, and the use of PAGE method to detect the HbWS zone with G gamma location with concentration and A gamma position no Strong staining (HbF value) was positive in HbWS area, and the sensitivity and specificity of PAGE positive diagnosis of HbWS were calculated by four grid tables. (2) clinical study: collect the different genotype carriers (patients) from the Baise area in 2010, from the clinic diagnosed as HbWS, and select the Hb according to the inclusion criteria and the exclusion criteria. The carriers (patients) of different types of gene mutations of WS were divided into several control groups (subgroups), and all the samples were carried out by automatic blood cell analyzer, hemoglobin automatic analyzer for hematological analysis, and Gap-PCR detection of deficient alpha and reverse dot print. The RDB method was used to detect the non missing alpha ground poverty and beta poverty. The data were processed with SPSS 19 statistical software. Results: (1) 600 cases were enrolled in the group, 130 cases of HbWS zone positive and 470 cases were diagnosed by PAGE test, 126 cases of HbWS gene mutation were detected by the gold standard method, and 474 cases of.PAG without HbWS gene mutation were found. The diagnostic coincidence rate of E diagnosis HbWS was 98%, sensitivity 96.83%, specificity 98.31%, positive predictive value 0.9385, negative predictive value 0.9915, positive likelihood ratio 57.3, negative likelihood ratio 0.0249. application ROC curve analysis PAGE diagnosis HbWS curve area AUC was 0.942,95% confidence interval (0.893,0.991). (2) collect confirmed HbWS different genotypes group A total of 292 patients (patients) were divided into eight types, including 174 hbws heterozygote (alpha WS alpha / ALA) (59.59%), 43 hbws complex beta poor ground poor (14.73%), 30 double heterozygotes (10.27%), hbh-ws disease 28 (9.59%), and 8 cases (2), hbh-ws disease, and 8 cases (2) (8 cases) with hbws. .74%), 4 cases of hbh-ws disease combined with light beta poor (1.37%), 3 cases of hbws complex beta poor (1.03%) and 2 cases of hbws homozygote (0.68%). (3) the HB (129.68 + 21.11g/l), MCV (86 + 5.36fl), MCH (27.76 + 2.16pg) in the hbws heterozygote group were in the normal range, which was lower than the normal group but higher than the alpha alpha / alpha alpha group, alpha alpha A A / A / alpha / A / - Alpha ~ (3.7) and alpha / - alpha ~ (4.2) groups (the four groups of MCH are lower than normal reference values); rdw-cv (13.10 + 1.10%) is not different from other stationary alpha - poor, but higher than the normal human group. (4) the Hb of the double heterozygote of hbws compound other static alpha poor is in the normal range (128.60 + 15.67g/l), higher than the hbws composite light beta poor group and light beta - The poor group; MCV (80.27 + 4.70fl) in the critical range, MCH (25.29 + 1.52pg) lower than the normal reference range, but higher than the hbws composite light beta poor group; rdw-cv (13.38 + 1.27%) is lower than the hbws composite light beta poor group, the light beta poor group and the standard type alpha poor. (5) hbws composite light beta poor group HB (112.59 + 14.03g/l), MCV (64.48 + 5.53fl). MCH (19.72 + 1.81pg) was lower than normal range. There was no significant difference between small cell low pigment anemia and light beta poor group Hb, MCV and MCH, rdw-cv (15.67 + 1.34%) was lower than light beta poor group. (6) HB (104.67 + 27.03g/l), MCV (67.76 + 6.10fl), MCH (20.64 + 4.2), - alpha ~ (3.7) group, - alpha ~ (3.7) group. The same expression was small cell low pigmentary anemia, but the Hb level was higher than that of the two groups (P0.05); the MCV in the group of alpha WS alpha /--sea was lower than the alpha CS alpha /--sea group (P0.05), the degree of MCH decreased is the lightest, the rdw-cv increase was the lightest (P0.01). Conclusion: (1) the specificity of the positive diagnosis of the page method region is high, the sensitivity and diagnostic coincidence rate are high and the accuracy is close to the gene. The detection method can be used as a new hbws screening diagnostic method for clinical application. In addition, the page method can also detect the zeta chain for the diagnosis of adult standard alpha poverty. (2) the gene mutation types of hbws in Baise area are mainly alpha WS alpha / alpha alpha, and the hematological phenotype of alpha ws a / ala is basically normal, and it is easy to leak in screening; hbws compound other static alpha. The dual heterozygotes in the poor area were small cells and low pigments, and the hbws composite light beta poor group showed small cell low pigmented anemia; hbh-ws's disease also showed small cell low pigmented anemia, but the degree of anemia was less than - alpha ~ (4.2) /--SEA, - (3.7) /--SEA group light. (3) HbWS different genotypes have their characteristics, but they have their own characteristics. The degree of anemia is lighter than that of the same type of thalassemia. The research results are instructive to the genetic counseling and prenatal diagnosis of the land poverty in this area.
【学位授予单位】：右江民族医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R556.61
本文编号：2119257
[Abstract]:Objective: (1) to evaluate the accuracy of detection of abnormal hemoglobin Westmead (HbWS) zone by polyacrylamide gel electrophoresis (PAGE), and to provide a new method for screening and diagnosing HbWS from the detection of protein peptide chain. (2) to explore the genotype and hematological phenotypes of HbWS in the population of Baise and the Mediterranean with other different genotypes. Haematological phenotypic difference of anemia (ground poor). Methods: (1) experimental study: in 2015 -2016 years, the native place of Baise region, in our hospital in the poor gene examination, age between 2-60 years of age, gene detection as the gold standard for the diagnosis of HbWS, and the use of PAGE method to detect the HbWS zone with G gamma location with concentration and A gamma position no Strong staining (HbF value) was positive in HbWS area, and the sensitivity and specificity of PAGE positive diagnosis of HbWS were calculated by four grid tables. (2) clinical study: collect the different genotype carriers (patients) from the Baise area in 2010, from the clinic diagnosed as HbWS, and select the Hb according to the inclusion criteria and the exclusion criteria. The carriers (patients) of different types of gene mutations of WS were divided into several control groups (subgroups), and all the samples were carried out by automatic blood cell analyzer, hemoglobin automatic analyzer for hematological analysis, and Gap-PCR detection of deficient alpha and reverse dot print. The RDB method was used to detect the non missing alpha ground poverty and beta poverty. The data were processed with SPSS 19 statistical software. Results: (1) 600 cases were enrolled in the group, 130 cases of HbWS zone positive and 470 cases were diagnosed by PAGE test, 126 cases of HbWS gene mutation were detected by the gold standard method, and 474 cases of.PAG without HbWS gene mutation were found. The diagnostic coincidence rate of E diagnosis HbWS was 98%, sensitivity 96.83%, specificity 98.31%, positive predictive value 0.9385, negative predictive value 0.9915, positive likelihood ratio 57.3, negative likelihood ratio 0.0249. application ROC curve analysis PAGE diagnosis HbWS curve area AUC was 0.942,95% confidence interval (0.893,0.991). (2) collect confirmed HbWS different genotypes group A total of 292 patients (patients) were divided into eight types, including 174 hbws heterozygote (alpha WS alpha / ALA) (59.59%), 43 hbws complex beta poor ground poor (14.73%), 30 double heterozygotes (10.27%), hbh-ws disease 28 (9.59%), and 8 cases (2), hbh-ws disease, and 8 cases (2) (8 cases) with hbws. .74%), 4 cases of hbh-ws disease combined with light beta poor (1.37%), 3 cases of hbws complex beta poor (1.03%) and 2 cases of hbws homozygote (0.68%). (3) the HB (129.68 + 21.11g/l), MCV (86 + 5.36fl), MCH (27.76 + 2.16pg) in the hbws heterozygote group were in the normal range, which was lower than the normal group but higher than the alpha alpha / alpha alpha group, alpha alpha A A / A / alpha / A / - Alpha ~ (3.7) and alpha / - alpha ~ (4.2) groups (the four groups of MCH are lower than normal reference values); rdw-cv (13.10 + 1.10%) is not different from other stationary alpha - poor, but higher than the normal human group. (4) the Hb of the double heterozygote of hbws compound other static alpha poor is in the normal range (128.60 + 15.67g/l), higher than the hbws composite light beta poor group and light beta - The poor group; MCV (80.27 + 4.70fl) in the critical range, MCH (25.29 + 1.52pg) lower than the normal reference range, but higher than the hbws composite light beta poor group; rdw-cv (13.38 + 1.27%) is lower than the hbws composite light beta poor group, the light beta poor group and the standard type alpha poor. (5) hbws composite light beta poor group HB (112.59 + 14.03g/l), MCV (64.48 + 5.53fl). MCH (19.72 + 1.81pg) was lower than normal range. There was no significant difference between small cell low pigment anemia and light beta poor group Hb, MCV and MCH, rdw-cv (15.67 + 1.34%) was lower than light beta poor group. (6) HB (104.67 + 27.03g/l), MCV (67.76 + 6.10fl), MCH (20.64 + 4.2), - alpha ~ (3.7) group, - alpha ~ (3.7) group. The same expression was small cell low pigmentary anemia, but the Hb level was higher than that of the two groups (P0.05); the MCV in the group of alpha WS alpha /--sea was lower than the alpha CS alpha /--sea group (P0.05), the degree of MCH decreased is the lightest, the rdw-cv increase was the lightest (P0.01). Conclusion: (1) the specificity of the positive diagnosis of the page method region is high, the sensitivity and diagnostic coincidence rate are high and the accuracy is close to the gene. The detection method can be used as a new hbws screening diagnostic method for clinical application. In addition, the page method can also detect the zeta chain for the diagnosis of adult standard alpha poverty. (2) the gene mutation types of hbws in Baise area are mainly alpha WS alpha / alpha alpha, and the hematological phenotype of alpha ws a / ala is basically normal, and it is easy to leak in screening; hbws compound other static alpha. The dual heterozygotes in the poor area were small cells and low pigments, and the hbws composite light beta poor group showed small cell low pigmented anemia; hbh-ws's disease also showed small cell low pigmented anemia, but the degree of anemia was less than - alpha ~ (4.2) /--SEA, - (3.7) /--SEA group light. (3) HbWS different genotypes have their characteristics, but they have their own characteristics. The degree of anemia is lighter than that of the same type of thalassemia. The research results are instructive to the genetic counseling and prenatal diagnosis of the land poverty in this area.
【学位授予单位】：右江民族医学院
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R556.61
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