原发性闭角性青光眼致病基因的鉴定

Primary angle-closure glaucoma is one of the major causes of blindness among Chinese. Early prevention measures and effective therapy are not available until now. Genetic factors play a major role in the pathogenesis of PACG. But rare PACG related genes have been identified so far. In this study, we have collected 14 PACG families and 220 sporadic cases. Whole-genome Linkage Analysis was performed in a large family to target a number of positive sites. And Whole-Genome Sequencing was used in two confirmed affected members from the same family. Then the sequencing data were filtered by dbSNP131 and Genome Project. As a result, 523 SNPs and 53 insert/deletions were detected. We plan to combine the Whole-genome Linkage data with Whole-Genome Sequencing to screen and identify PACG genes and mutations, and verify those mutations in other families and sporadic cases. Further evidence of genes-pathogenesis links will come from preliminary functional study by experiment in vitro. Better understanding of the disease will improve the treatment and outcome of this condition.

原发性闭角性青光眼（PACG）是我国主要致盲眼病之一，目前缺乏有效的早期预防和治疗措施。遗传因素在PACG发病中起着重要作用，至今尚未鉴定出公认的PACG致病基因。我们收集了14个原发性闭角性青光眼家系和220例散发病例，对其中一个大家系进行了全基因组遗传连锁分析，获得了多个阳性候选位点；同时，对该家系的两个病人进行了全基因组测序分析，获得的所有核苷酸改变经dbSNP131和1000 Genome Project数据库过滤后有523个单核苷酸变异和53个插入缺失变异（indel）。本研究将结合全基因组遗传连锁分析和全基因组测序分析结果，鉴定该家系的致病基因及其突变；并在其他家系和散发病例中进行验证，初步研究鉴定的PACG基因的体外功能和致病机制，为PACG的预防和治疗奠定基础。

青光眼是一种具有很强遗传异质性的综合神经退行性疾病，其特点是造成视力逐渐缺失，从前房的解剖结构分类为开角型和闭角型。本课题已经收集160个先天性青光眼家系，1200例青光眼样本和3000例对照标本。为了寻找在编码区的新的青光眼致病基因，我们对其中100个家系和300例散发样本及1000例对照进行了全基因外显子组测序。我们对外显子组测序结果（1）首先采用多基因的分析模式进行了变异位点的关联分析，筛选到47个在编码区的非同义突变位点（p<10e-8），正在进行扩大样本验证工作。（2）还通过单点突变位点分析发现49个单点突变位点进行了sequenom扩大样本验证，下一步要进行再扩大样本验证、数据解释和功能研究工作。（3）最后，通过单基因的分析模式进行了变异位点分析，发现 TIMP4基因的编码区的4个变化位点exon2:c.211_212del:p.71_71del，exon3:c.C284T:p.T95M，exon5:c.T502G:p.C168G，exon5:c.C485G:p.T162S。在人眼睛冰冻切片中染色发现TIMP4基因在视网膜神经节细胞中表达，说明该基因可能在青光眼中起一定的作用。目前已经构建该基因的表达质粒，下一步将对该基因进行更深一步的功能研究。