【作者】 金鑫；

【导师】 张卯年； 刘铁城； 袁慧军；

【作者基本信息】 中国人民解放军军医进修学院 ， 眼科学， 2008， 博士

【摘要】 目的:1.建立视网膜色素变性遗传资源收集保存的标准化操作程序。2.分析3个RP家系RP-GC-001、RP-DX-002、RP-PG-003临床表型及遗传学特点,寻找致病基因突变。3.分析2个Usher综合征家系USH-001、USH-002的临床表型特征及遗传学特点,筛选可能的已知致病基因位点。方法:1.对RP患者进行鉴定和临床分型,分析其遗传方式。收集、保存RP遗传资源,包括患者及家属的知情同意、填写RP调查表、抽取外周静脉血制备基因组DNA的、提取基因组DNA、保存及鉴定基因组DNA。建立RP遗传资源库,包括纸质档案管理系统和电子档案管理系统。2.分析RP-GC-001、RP-DX-002、RP-PG-003家系患者的临床表型特征、系谱特征,利用PCR扩增和直接测序的方法并对RHO、RDS、ROM1、NRL、CRX等5个已知致病基因的15个外显子进行筛查。3.分析USH-001、USH-002家系患者的临床表型特征、系谱特征,利用连锁分析的原理和方法,对12个已知致病基因位点周围的微卫星标记进行扫描,筛选可能的致病基因位点。结果:1.建立了规范、科学、合理,并且符合国际、国内遗传资源采集研究原则要求的视网膜色素变性遗传资源收集保存标准化操作系统。2.RP-GC-001、RP-PG-002、RP-DX-003表型具有共性:夜盲、进行性视野缩小、视网膜骨细胞状色素沉着、视乳头呈蜡黄色萎缩,表型垂直连续传递,男女均可发病。同时又具个性特征:RP-DX-002家系中患者视网膜骨细胞样色素沉着累及黄斑区和视乳头旁,多个患者伴有虹膜局限性前粘连、玻璃体烟灰样混浊。RP-PG-003家系患者在50岁左右出现急性闭角型青光眼。RP-GC-001家系中发现了RHO基因外显子2编码区403C＞T Arg135Trp R135W突变。RP-PG-002、RP-DX-003家系的RHO、RDS、ROM1、NRL、CRX基因未发现致病突变3.USH-001、USH-002家系均表现为儿童期出现的双耳听力下降,为非渐进性、中-重度感音神经性耳聋,以高频听力损失为主,前庭功能正常。10-13岁出现夜盲症状,随年龄增长夜盲逐渐加重,并出现视野逐渐缩窄至管状视野,中心视力下降不明显,临床表现符合USH2型。表型未出现连续遗传的现象,男女均有发病,患者的子女表型正常。USH-001家系中所有患者的D11S902、D17S785微卫星标记的Allele值完全一致,USH-002家系所有患者的微卫星标记D1S425、D9S1776的Allele值完全一致。结论:1.视网膜色素变性遗传资源收集保存标准化操作系统的建立,确保了本研究的规范性、保证了遗传资源搜集工作良好的延续性以及我们后续基因定位克隆、分子流行病学研究、基因筛查工作结果的真实可靠。2.RP-GC-001、RP-PG-002、RP-DX-003为常染色体显性遗传RP家系。RHO基因外显子2编码区403C＞T Arg135Trp R135W突变为RP-GC-001家系的致病突变。RHO、RDS、ROM1、NRL和CRX不是RP-PG-002、RP-DX-003家系的致病基因。3.USH-001、USH-002家系为USH2型家系,遗传方式为常染色体隐性遗传。USH1C、USH1G可能为USH-001家系的致病基因:USH2A、USH2D可能为USH-002家系的致病基因。更多还原

【Abstract】 Objective:1. To establish the standard system to collect and preserve the genetic resources of retinitis pigmentosa.2. To analyze the clinical and genetic features of the three retinitis pigmentosa families: RP-GC-001 RP-DX-002 RP-PG-003, and to find pathogenic genes for the families.3. To analyze the clinical features of the two Usher Syndrome families: USH-001 USH-002, and to find possible relevant USH gene map locus.Methods:1. RP patients were identified and classified, and their genetic patterns were analyzed. Collecting and preserving RP genetic resources including signing the informed consent, filling out RP questionnaire, extracting genomic DNA from peripheral blood, identification and preservation genomic DNA. Establishing RP genetic resources system including paper-based records management system and electronic records management system.2. Analysis of the clinical and pedigree features of RP-GC-001 RP-DX-002 RP-PG-003 families. And detecting the 15 exons of the five known pathogenic genes: RHO, RDS, ROM1, NRL and CRX by PCR and direct sequencing.3. Analysis of the clinical and pedigree features of USH-001 USH-002 families. And screening the short tandem repeat markers around the 12 known USH gene map locus to choose the possible relevant genes by use of the principles and methods of linkage analysis.Results:1. The system of collecting and preserving genetic resources of RP was standard scientific and reasonable. And it was in line with the international and domestic requirements and principles of genetic resources collection.2. The common phenotype of the families RP-GC-001 P-PG-002 and RP-DX-003 were night blindness, narrowing of vision, bone cell-like retinal pigmentation, shrink of papilla of optic nerve. The phenotype vertically transmitted and appeared in both male and female patients. However, RP-DX-002 family had some characteristics: bone cell-like retinal pigmentation involved in macular and environment of papilla of optic nerve, many patients were with iris adhesion and soot-like vitreous opacity. And patients in RP-PG-003 family would have acute angle-closure glaucoma in their fifties. RHO exon 2 coding region of 403 C>T Argl35Trp R135W mutation was found in the RP-GC-001 familiy, and no pathogenic mutations of RHO RDS ROM1 CRX and NRL were found in the RP-PG-002 and RP-DX-003 families.3. The phenotypes of the family USH-001 were similar to family USH-002: hearing loss in both ears in childhood, non-progressive sensorineural hearing loss, high-frequency hearing loss and normal vestibular function. Night blindness occurred in 10-13 years old and gradually got worse with age, visual field gradually narrowed to tubular vision, while the decline of visual acuity were not obvious. Their clinical manifestations were consistent with the diagnosis of USH2. The phenotype did not consecutively transmitte and appeared in both male and female patients. And patients in these two families all had normal children. In USH-001 family all the patients shared the same Allele of D11S902 and D17S785 short tandem repeat markers, and all the patients in USH-002 family shared the same Allele of D1S425and D9S1776 markers.Conclusion:1. Establishment of the system to preserve and collect RP genetic resources have ensured this study was standardized and had good continuity. It would make our follow-up results of positional cloning, molecular epidemiological study and genetic screening true and reliable. 2. The three RP families were autosomal dominant RP families. The mutation of RHO exon 2 coding region of 403 C>T Argl35Trp R135W was pathogenic mutation in RP-GC-001 family. RHO, RDS, ROM1, CRX and NRL were not the pathogenic genes for RP-PG-002 family and RP-DX-003 family.3. USH-001 and USH-002 were USH2 families and their genetic forms were autosomal recessive. USHIC and USH1G were likely pathogenic genes for USH-001 family, USH2A and USH2D were likely pathogenic genes for USH-002 family.更多还原