CHRDL1基因无义变异引起大角膜免疫反应  

Nonsense Mutation of CHRDL1 Gene Causes Response of Immune in Megalocornea

作　　者：史平玲 但汉东 王艳歌 王娇娇 张贝贝 魏圆梦 唐贺 李苗 宋宗明 Pingling Shi;Handong Dan;Yange Wang;Jiaojiao Wang;Beibei Zhang;Yuanmeng Wei;He Tang;Miao Li;Zongming Song(Zhengzhou University People's Hospital,Henan Provincial People's Hospital,Henan Eye Hospital,Zhengzhou 450003,China)

机构地区：[1]郑州大学人民医院,河南省人民医院,河南省立眼科医院,郑州450003

出　　处：《中华眼视光学与视觉科学杂志》2024年第6期450-459,共10页Chinese Journal Of Optometry Ophthalmology And Visual Science

基　　金：国家自然科学基金(82101162);河南省重大科技专项(221100310200);河南省医学科技攻关计划联合共建项目(LHGJ20230654)

摘　　要：目的:确定先天性大角膜一家系致病基因变异,并探讨该基因致病机制。方法:实验研究。于2022年7月采集一X连锁大角膜(MGC1)家系先证者1例和其他家系成员2例的外周静脉血标本。并详细采集患者病史后,对患者行最佳矫正视力(BCVA)、眼压、裂隙灯显微镜联合前置镜、眼前节分析(OrbscanⅡ)、角膜地形图和眼底彩色照相检查。应用包含439个基因的眼前节疾病检测Panel和包含22个基因的青光眼检测Panel进行靶向捕获富集及高通量测序,筛选与疾病表型相关的致病基因变异,并使用Sanger测序进行验证。利用RT-PCR和免疫蛋白印迹法检测CHRDL1基因mRNA和蛋白的表达;荧光显微镜观察绿色荧光蛋白的表达。通过转录组测序和生物信息学分析,检测CHRDL1无义变异对分子功能和信号通路的影响。数据均采用独立样本t检验进行分析。结果:先证者被诊断为MGC1,眼压最高达27 mmHg(1 mmHg=0.133 kPa);角膜中央厚度右眼为469μm,左眼为470μm,均减少;角膜直径右眼为13.95 mm,左眼为13.65 mm。基因检测结果显示,先证者CHRDL1基因存在c.36delC(p.Tyr12^(*))半合子无义变异。与pHBLV-CHRDL1 wt组比较,pHBLV-CHRDL1 Tyr12^(*)组CHRDL1蛋白表达显著下降(t=24.16,P<0.001),CHRDL1Tyr12^(*)变异能够激活免疫系统信号通路并促进炎症因子和趋化因子表达。结论:CHRDL1基因c.36delC(p.Tyr12^(*))无义变异为MGC1家系的变异位点,该变异激活的免疫信号通路可能在MGC1的发病机制中发挥重要作用。Objective:To determine the mutation of pathogenic gene in a family with congenital megalocornea and to explore the pathogenic mechanism of gene.Methods:This was an experimental study.In July 2022,peripheral venous blood samples were collected from 1 probands of X-linked macrocornea(MGC1)pedigree and 2 members of other pedigrees.And the medical history of the patients was collected.The best corrected visual acuity(BCVA)and intraocular pressure(IOP)of the family were examined.The features of the anterior segment and fundus were observed via slit lamp biomicroscope and slit lamp lens.The anterior segment analysis and corneal topography of proband were carried out.And the fundus color photography was obtained as well.The targeted capture was carried out using anterior segment detection panel containing 439 genes and the glaucoma detection panel with 22 genes.The high-throughput sequencing was performed to screen the pathogenic mutations,and mutation sites were verified by Sanger sequencing.The expression of CHRDL1 mRNA and protein were performed by RT-PCR and WB.The expression of green fluorescent protein(GFP)was observed by fluorescence microscope.To evaluate the molecular functional modifications and the signal pathway alteration after CHRDL1 nonsense mutation transfected,the transcriptome sequencing and the bioinformatics analysis were carried out.Data were analyzed by independent t test.Results:The proband was diagnosed as MGC1.The highest IOP was 27 mmHg(1 mmHg=0.133 kPa).The central corneal thicknesses were 469μm and 470μm in the right and left eye,respectively.The corneal diameters were 13.95 mm and 13.65 mm in the right and left eye,respectively.The hemizygous nonsense mutation of c.36delC(p.Tyr12^(*))in CHRDL1 gene in this family was identified as pathogenic.The expression of CHRDL1 protein was significantly reduced(t=24.16,P<0.001),the immune system signaling pathways were significant up-regulated,and the expression of inflammatory and chemokines were increased in CHRDL1 mutation HCE-2.Conclusions:The nov

关 键 词：大角膜 青光眼 CHRDL1基因 免疫反应 

分 类 号：R772.2[医药卫生—眼科]