无脉络膜症的基因筛查与临床表型分析

发布时间：2018-05-07 22:14

本文选题：无脉络膜症 + 基因型　；参考：《北京协和医学院》2013年博士论文

【摘要】：目的：明确国人无脉络膜症(choroideremia, CHM)突变位点,分析CHM患者临床表型特征,探索基因型与临床表型的关系。方法：收集2009～2013年于北京协和医院眼科就诊并诊断为CHM的患者及其家族成员。1.临床研究：询问病史、家族史并绘制家系图,行眼科一般检查(视力、屈光度、眼前节、眼底)、色觉检查、眼底彩色照相、相干光断层扫描(optical coherence tomography,OCT)、眼底自发荧光成像、视网膜电图(electroretinogram, ERG)以及视野(visual field, VF)检查。2.分子遗传学研究：采集患者及家族成员静脉血5-6ml,同时收集同期正常人作为对照,提取DNA。针对,基因的15个外显子及外显子-内含子接合处进行聚合酶链式反应(polymerase chain reaction, PCR)扩增及直接测序。发现DNA序列改变后,进行单核苷酸多态性(single nucleotide polymorphism, SNP)分析,以除外已知正常多态位点,并查阅遗传学数据库及文献,明确是否为已报突变。若为新发突变,对100条染色体进行该位点的扩增、测序,以排除未知多态位点可能。结果：1.共收集13个家系资料,确定男性患者51例,女性携带者52例。其中24例男性患者及21例女性携带者接受临床检查。男性患者中,87.5%在儿童及青少年期出现夜盲,46.7%存在色觉异常,就诊时最佳矫正视力(best-corrected visual acuity, BCVA)介于光感～1.0。眼底表现：视网膜色素上皮(retinal pigment epithelium, RPE)层及脉络膜毛细血管层呈不同程度萎缩,可暴露脉络膜大血管及巩膜；黄斑区可正常或萎缩；2例患者存在大量片状棕黑色色素沉着。OCT:IS/OS层消失、RPE层不连续是主要改变,严重病例出现中心凹形态消失、神经上皮厚度变薄。自发荧光：大部分患者仅在黄斑区残留正常荧光。ERG：83.3%的患眼呈暗视反应熄灭型,58.3%的患眼呈明视反应熄灭型。女性携带者,均无夜盲主诉、无色觉异常,就诊时BCVA介于0.3-1.5。眼底表现：以斑驳样色素变动为特征,其中5例携带者特别表现为黄白色类似玻璃膜疣样色素变动。OCT.绝大多数呈正常表现,玻璃膜疣样色素变动区域在OCT上对应RPE层局限性中高信号物质堆积。自发荧光：呈斑驳样低荧光表现。2.10个参与分子遗传学研究的家系全部明确了突变位点：包括3种无义突变,5种剪接突变,1种小片段缺失突变,1种小插入缺失突变。70%为新发突变。结论：1.临床表型：男性患者多于儿童及青少年期以夜盲起病,视力水平随年龄下降,色觉异常与中心视力相关；RPE及脉络膜萎缩是普遍眼底表现,晚期可累及黄斑。女性携带者多无临床症状,视力水平随年龄下降不明显；斑驳样色素变动是普遍眼底表现。2.基因型-临床表型关系：相似突变位点及类型存在临床异质性,相似临床表型存在遗传异质性,仍需继续探索基因型-临床表型相关性。
[Abstract]:Objective: to identify the choroideremia (CHM) mutation site in Chinese patients without choroid disease, analyze the clinical phenotypic characteristics of CHM patients, and explore the relationship between genotype and clinical phenotype. Methods: the patients and their family members who were diagnosed with CHM in Peking Union Hospital from 2009 to 2013 were collected. Clinical studies: asking for medical history, family history and drawing pedigree map, general ophthalmology examination (visual acuity, diopter, anterior segment, fundus, color vision, fundus color photography, optical coherence tomographic CT imaging, fundus autofluorescence imaging), Electroretinogramgrams (ERG) and visual field, VF). Molecular genetics study: collecting 5-6 ml venous blood from patients and family members, and collecting the same period normal people as control, extract DNA. The 15 exons and exon-intron conjugates of the gene were amplified by polymerase chain reaction (PCR) and sequenced directly. After the DNA sequence was changed, single nucleotide polymorphism (SNPs) was analyzed to exclude the known normal polymorphic loci, and the genetic database and literature were consulted to determine whether the mutation was reported. For new mutations, 100 chromosomes were amplified and sequenced to rule out unknown polymorphic loci. The result is 1: 1. A total of 13 families were collected and 51 cases of male patients and 52 cases of female carriers were identified. Clinical examination was performed in 24 male patients and 21 female carriers. In male patients, 87.5% had night blindness in children and adolescents, 46.7% had color vision abnormality, and the best corrected visual acuity (visual acuity, BCVA) was between 1.0 and 1.0 at the time of treatment. Fundus manifestation: retinal pigment epithelium (RPE) layer and choroidal capillary layer were atrophied in varying degrees, which could expose choroidal large vessels and sclera. In 2 patients with normal macular area or atrophy, there were a large number of flake brown black pigmentation. The disappearance of OCTI / S / OS layer and the discontinuity of RPE layer were the main changes. In severe cases, the central fovea disappeared and the thickness of neuroepithelium became thinner. Autofluorescence: only 83.3% of the eyes with residual normal fluorescence in macular area showed dark vision reaction extinguishing type and 58.3% eyes showed obvious vision reaction extinguishing type. There was no nocturnal blindness and no abnormal color vision in female carriers. The BCVA was between 0.3 and 1.5 at the time of visit. Fundus manifestations: mottled pigmentation was characteristic, among which 5 carriers showed yellow and white vitreous verrucous pigment changes. Most of them showed normal appearance, and the changes of vitreous verrucous pigments on OCT corresponded to the accumulation of medium and high signal substances in RPE layer. Autofluorescence: mottled low fluorescence. 2.10 families involved in molecular genetic studies all identified mutation sites: including three nonsense mutations, five splicing mutations, one small fragment deletion mutation, one small insertion deletion mutation. 70% is a new mutation. Conclusion 1. Clinical phenotype: in male patients more than in children and adolescents the onset of nocturnal blindness the visual acuity level decreased with age color vision abnormalities and central vision related to RPE and choroidal atrophy are common fundus manifestations late macula can be involved. Most female carriers had no clinical symptoms, and the visual acuity level did not decrease significantly with age, and mottled pigmentation was the common fundus manifestation. 2. Genotypic and clinical phenotypic relationship: similar mutation sites and types have clinical heterogeneity, similar clinical phenotypes have genetic heterogeneity, it is necessary to continue to explore the relationship between genotype and clinical phenotypes.
【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2013
【分类号】：R773.4

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