Fabry's Disease: Detection of Heterozygotes Using Blastoid Lymphocytes StimuIated by PhytohemaggIutinin

Cultured lymphocytes with phytohemagglutinin (PHA) were used for samples of α-galactosidase (α-gal) assay. In control subjects α-gal activity of cultured lymphocytes with PHA increased compared to that of uncultured or cultured lymphocytes without lectin. While the carrier females of Fabry's disease showed a slight decrease of α-gal activity in cultured lymphocytes with PHA. Therefore, heterozygotes could be definitely distinguished from controls. Cultured lymphocytes with PHA were thought to be available for the detection of heterozygotes.     

Plasma transfusion combined with chelating therapy alleviates fulminant Wilson's disease with a single Arg778Leu heterozygote mutation

Wilson's disease (WD), resulting from homozygote and compound heterozygote mutations in ATB7B, is an autosomal recessive disease. WD associated acute liver failure (ALF) is fatal, and a revised Wilson's disease prognostic index (RWPI) >11 is a reliable indication of liver transplantation (LT) or artificial liver support (ALS). We described a WD patient who initially presented with ALF and severe hemolytic anemia. A single heterozygote c.2333G>T mutation (p. Arg778Leu, R778L) in ATP7B was screened by whole exome sequencing and validated by Sanger sequencing. Rapid diagnostic criteria (ALP/TBIL <4 and AST/ALT >2.2) are suitable for early diagnosis. Although the RWPI amounted to 15, the patient recovered after intermittent plasma transfusion and subsequent chelating therapy without LT or ALS. In conclusion, WD patients with a single R778L heterozygote mutation can present with ALF as the initial clinical manifestation, and intermittent plasma transfusion combined with chelating therapy may alleviate fulminant WD without LT or ALS.     

Relationship between loss of heterozygosity of deleted in colorectal carcinoma gene microsatellites and prognosis of colorectal adenocarcinoma

AIM: To investigate the relationship between the loss of heterozygosity (LOH) of microsatellites on the deleted in colorectal carcinoma (DCC) gene and prognosis of colorectal adenocarcinoma.METHODS: A retrospective study of 58 colorectal adenocarcinoma cases with follow-up data and paired control normal mucosal tissues from 1983 to 1985 from files from the West China University of Medical Sciences Department of Pathology was carried out by PCR microsatellite analysis. Sixteen, 35, and seven cases had well-, moderately, and poorly differentiated tumors, respectively; 11, 30, and 17 cases were staged as Dukes’ A, B, and C, respectively.RESULTS: LOH of DCC microsatellites was detected in 18 cases (31.0%). The 5-year survival rate between LOH-positive and LOH-negative patients was 44.4% and 77.5%, respectively (P < 0.05). The results suggest that LOH of DCC microsatellites correlate with prognosis but not with differentiation (P > 0.05) and Dukes’ stage (P > 0.05) in colorectal adenocarcinoma.CONCLUSION: LOH of DCC microsatellites may be a marker of malignancy. Combined with the traditional prognostic indicators, LOH can predict prognosis of colorectal adenocarcinoma.     

斑状角膜营养不良一家系发现新的复合杂合突变

目的 明确一个中国斑状角膜营养不良(MCD)家系中致病基因及其突变的类型及组织病理学改变.方法 对行穿透性角膜移植术后获得的先证者角膜进行光镜和透射电子显微镜检查.收集此家系11个成员的外周血,提取DNA;合成特异性引物,利用聚合酶链式反应(PCR)扩增CHST6基因的编码区;PCR产物进行直接测序和限制性内切酶消化分析.选取100个无关健康个体作为对照.结果 组织特殊染色显示胶性铁染色阳性.透射电子显微镜下观察,可见滑面内质网扩张,胞质空泡化,有细颗粒物质.CHST6基因编码区进行突变检测发现,此家系3个患者CHST6基因第3个外显子存在一个复合杂合突变(Q298X和Y358H),其中第892位碱基C→T,导致第298位的谷氨酰胺变成终止密码子;第1072位碱基 T→C,导致第358位的酪氨酸被组氨酸取代.6个表型正常的个体均为杂合子,而在100个无血缘关系的健康个体中没有检测到突变.结论 确定了两个新的突变导致了该家系CHST6基因功能的丧失,引起MCD.该复合杂合突变在国内外为首次报道.     

血友病A携带者检测与产前诊断

目的 建立简便、快速的血友病A携带者检测与产前诊断体系。方法 用PCR方法直接检测FⅧ内含子22倒位或对FⅧ基因内的BclⅠ位点、内含子13和22中STR和FⅧ基因外的DXS52（ST14）位点的多态性进行遗传连锁分析。结果 应用内含子22例位的直接诊断率为47．6％;Bcl Ⅰ位点的可诊断率为27．8％;内含子13和22中STR的可诊断率分别为28．6％及29．4％;DCS52的可诊断率为81．3％;综合应用直接诊断和间接遗传连锁分析,对21个家系进行检测,可诊断率为94．7％。结论 血友病A的携带者检测与产前诊断可先进行内含子22倒位的检测,若结果为阳性即可作出诊断;若内含子22例位的检测结果为阴性,则可利用FⅧ基因内、外的多个位点多态性结果进行遗传连锁分析,以作出最终的诊断。     

应用细菌重组系统构建小鼠Flt3配体的重组腺病毒及其在肝癌细胞中的表达

目的 构建携带小鼠Flt3配体（murine flt3 ligand,mFL)的重组腺病毒载体并检测它的感染后的小鼠肝癌细胞中的表达。为肝癌基因治疗提供实验基础。方法 PCR扩增含有mFL的质粒,应用高效细菌内同源重组系统构建携带mFL的重组腺病毒,感染小鼠肝癌细胞株Hepal-6,一步法提取细胞总RNA,RT-PCR检测mFL的表达。结果 构建了携带mFL的重组腺病毒,并在感染后的Hepal-6细胞中检测到了mFL的mRNA。结论 应用该细菌重组系统成功地构建了携带mFL的重组腺病毒载体。为下一步的基因治疗研究奠定了基础。     

汉族肥胖儿童黑皮质素4受体基因筛查

目的探讨肥胖儿童黑皮质素4受体(MC4R)的突变频率及其与临床生化指标改变的关系。 方法浙江大学医学院附属儿童医院等单位于2004—2005年,对200例肥胖及100例正常体重儿童,利用PCR及基因测序技术进行MC4R基因筛查,同时对200例肥胖儿童进行生化检测和口服葡萄糖耐量试验。 结果(1)200例肥胖儿童中检出杂合子错义突变、无义突变3例;100例对照组中未检出突变;Val103Ile基因多态性在两组中分别有6例、2例。肥胖组中发现了3个新的杂合子突变位点Val166Ile、Cys277Stop、Arg310Lys;肥胖组和对照组中同时检测到新的杂合子变异Leu23Arg。(2)MC4R突变组和非突变组的肥胖儿童BMI、ALT、AST、TG、CHO、WBISI比较均无统计学意义(P>0.05)。 结论(1)首次在汉族儿童人群中发现了MC4R基因3个新的杂合子突变位点(Val166Ile、Cys277Stop、Arg310Lys)。(2)Leu23Arg可能是汉族人群MC4R的基因多态性。     

Mild dopa-responsive dystonia in heterozygous tyrosine hydroxylase mutation carrier: Evidence of symptomatic enzyme deficiency?

We present a case of mild, adult-onset dopa-responsive dystonia (DRD) with a heterozygous mutation in the tyrosine hydroxylase (TH) gene. We propose that this genetic state may have led to partial enzyme deficiency. Future studies should attempt to identify and characterize the phenotype of other patients with single TH variants.     

单细胞四重巢式荧光聚合酶链反应进行血友病A基因诊断和性别鉴定的研究

目的 建立快速有效检测血友病A基因诊断和性别鉴定的单细胞聚合酶链反应（PCR）方法。方法 联合6个与凝血因子Ⅷ（FⅧ）紧密连锁的STR位点（DXS15、DXS9901、G6PD、DXS1073、DXS1108和F8Civs13）通过多重荧光PCR进行正常人群多态性分析和血友病A家系基因诊断，从中选取杂合度和诊断率较高的STR位点联合性别位点牙釉蛋白基因（Amelogenin gene）建立单淋巴细胞多重巢式荧光PCR的检测方法。结果 选择了3个杂合度和诊断率较高的STR位点（F8Civs13、DXS1073和DXS15）联合性别位点建立了单细胞四重巢式荧光PCR的检测方法。Amelogenin、DXS15、F8Civs13和DXS1073在男性的单细胞扩增成功率分别为94．3％、91．4％、100％和100％，在女性为100％、97．1％、97．1％和97．1％。该女性的DXS1073和DXSl5位点呈杂合子，均未发现等位基因脱失现象。扩增皆未发现假阳性和假阴性的情况。结论 建立的单细胞四重巢式荧光PCR方法快速有效，有望应用于临床血友病A非创伤性产前基因诊断。     

DHCR7基因复合杂合变异致新生儿Smith-Lemli-Opitz综合征1例

本文报告了1例 DHCR7基因复合杂合变异导致的Smith-Lemli-Opitz综合征（Smith-Lemli-Opitz syndrome,SLOS）患儿。患儿生后即起病,以“多发畸形、喂养困难”为主诉从当地医院转诊至湖南师范大学第一附属医院（湖南省人民医院）。入院时体格检查见全身硬肿、头皮缺损、阴茎短...