【作者】 李娟；

【导师】 李南方；

【作者基本信息】 石河子大学 ， 内科学， 2015， 硕士

【摘要】 目的:通过对Fabry病患者的GLA基因突变类型及临床表现的分析,探讨Fabry病基因型与临床表型之间的关系。方法:收集一个已确诊Fabry病的患者及其家庭成员的临床资料,包括一般资料、血生化指标、心电图、心脏超声、头颅核磁等资料,并在知情同意的情况下收集研究对象的清晨空腹肘静脉血样置于-20℃冰箱。用干血片法测定肝素抗凝血浆α-半乳糖苷酶A活性,利用聚合酶链反应扩增GLA基因并对其进行纯化测序。另外选取13名居住于同一地区的健康志愿者作为对照组人群,对他们的GLA基因突变型及临床表型进行分析。结果:在该Fabry病家系中共有19名成员,8人存在相关临床症状,包括3名男性和5名女性,平均年龄(36.38±19.41)岁。其中男性患者症状较重,半合子男性具有Fabry病的典型临床表现如血管角质瘤、肢端疼痛以及心、脑、肾等损害,而女性患者症状相对较轻,女性杂合子仅表现为肢端疼痛等症状。在13名健康对照者中,男5例,女8例,平均年龄(46.85±18.52)岁。对Fabry病患者α-半乳糖苷酶A活性研究发现,8例Fabry病患者α-半乳糖苷酶A活性下降,平均值为(96.96±81.15)10-15mol/day/spot,健康对照者α-半乳糖苷酶A活性的平均值为(188.31±42.23)10-15mol/day/spot,两组之间存在统计学差异(P<0.05)。其中男性患者的α-半乳糖苷酶A活性(8.66±2.15)10-15mol/day/spot为健康对照者的4.6%,差异有统计学意义(P<0.05);女性患者的α-半乳糖苷酶A活性(149.93±46.56)10-15mol/day/spot则为健康对照者的79.62%,与健康对照组相比有所下降,差异无统计学意义(P>0.05),但与男性半合子患者相比差异有统计学意义(P<0.05)。在对GLA基因1~7号外显子及其邻近区域测序分析中发现该Fabry病家系先证者的GLA基因5号外显子内发生碱基缺失,其突变类型为g.10208-10209del AA。在家系成员中共检出7例GLA突变基因携带者,其中男性半合子2例,女性杂合子5例,突变类型均与先证者相同。在对照组人群中也发现了GLA基因突变,为1号外显子1390位点的错义突变(1390A>G),但患者无相应临床症状。结论:α-半乳糖苷酶A活性测定在Fabry病诊断中存在局限性,男性半合子患者酶活性下降更为明显。Fabry基因型和临床表型之间存在一定的变异性,即使相同基因型的Fabry病患者其临床表现也有所不同,同时GLA基因突变中可能存在不致病的情况。更多还原

【Abstract】 Objective: To explore the correlation between genotype and phenotype through analyzing the clinical manifestation and the gene mutation in the alpha-galactosidase A(GLA) in families with Fabry disease.Methods: In this study, the clinical data of a diagnosed Fabry disease patient and his family members were collected, including general information, blood biochemical index, electrocardiogram, echocardiography,skull magnetic resonance imaging, and so on. After inform consent, the early morning fasting elbow venous blood of subjects were also collected. α-galactosidase A(α-Gal A) activity was measured using dried blood spots on filter paper, the GLA gene was amplified using polymerase chain reaction(PCR), and the amplified products were purified and sequenced. In addition, thirteen healthy individuals living in the same region were examined as controls. The mutations of GLA gene and clinical manifestation of subjects were analyzed.Results: There were 19 people in this Fabry disease pedigrees, 8 people had clinical symptoms, three male and five female. The average age of them is(36.38±19.41) years. The symptoms of male patients more severe than female patients. In hemizygous male the symptoms include angiokeratoma, limb pain and involvement of cardiovascular, cerebrovascular, renal. In heterozygous female the symptoms just is limb pain, hypohidrosis, and so on. In the thirteen healthy controls, there were 5 male and 8 female. The average age of them is(46.85±18.52) years. The α-Gal A activity analysis revealed the α-Gal A activity in patients with Fabry disease was reduced, and the mean of it is(96.96±81.15)10-15mol/day/spot, and there were significant difference with healthy controls(188.31±42.23)10-15mol/day/spot(P<0.05). What’s more, theα-Gal A activity in males with Fabry disease was only 4.6% of that of healthy controls(8.66±2.15 vs188.31±42.23), the difference between them was significant(P<0.05). Whereas the α-Gal A activity in females with Fabry disease was 79.62% of healthy controls(149.93±46.56 vs 188.31±42.23), although there was a tendency to decline, the difference between them was not significant(P>0.05); when compared with hemizygous male patients, there was significant difference(P<0.05).When all seven exons of the GLA gene and the adjacent areas sequencing were analyzed, the proband was found a two nucleotide deletion in exon 5, and the genotype was g.10208-10209 del AA. There were seven people with the same mutation in these Fabry pedigree, including two hemizygote and five heterozygotes. At the same time, in the healthy controls also exist people with GLA gene mutation, 1390 A to G transition in exon 1(1390A>G), while this person with no symptoms.Conclusion: The α-Gal A activity measured with diagnostic limitations in Fabry disease, the enzymatic activity is significant reduced in male patients. Although patients with Fabry disease carrying the same gene phenotype may not present the same clinical manifestations, and there may be non-pathogenic GLA gene mutation.更多还原