小鼠视网膜神经节细胞在视神经损伤后的基因表达谱分析  

Analysis of gene expression profile of mouse retinal ganglion cells after optic nerve crush

作　　者：彭碧燕 梁嘉敏 韩可伊 王兵玉 甘子晖 罗丹妮 何丽婷 朱易 黄诗颖 郭晓萍 施维 张俊 孙俊铭 薛康宁 方茜 欧阳轶强 王春苗 张名媛 PENG Biyan;LIANG Jiamin;HAN Keyi;WANG Bingyu;GAN Zihui;LUO Danni;HE Liting;ZHU Yi;HUANG Shiying;GUO Xiaoping;SHI Wei;ZHANG Jun;SUN Junming;XUE Kangning;FANG Xi;OUYANGYiqiang;WANG Chunmiao;ZHANG Mingyuan(Laboratory Animal Center,Guangxi Medical University,Nanning 530021,China;School of Basic Medical Sciences,Guangxi Medical University,Nanning 530021,China;Life Sciences Institute,Guangxi Medical University,Nanning 530021,China;Guangxi Medical University Cancer Hospital,Nanning 530021,China;The First Affiliated Hospital of Guangxi Medical University,Nanning 530021,China)

机构地区：[1]广西医科大学实验动物中心,南宁530021 [2]广西医科大学基础医学院,南宁530021 [3]广西医科大学生命科学研究院,南宁530021 [4]广西医科大学附属肿瘤医院,南宁530021 [5]广西医科大学第一附属医院,南宁530021

出　　处：《广西医科大学学报》2024年第2期168-176,共9页Journal of Guangxi Medical University

基　　金：国家自然科学基金资助项目(No.32160137);广西自然科学基金项目资助(No.2021GXNSFAA220005);广西医科大学大学生创新创业项目资助(No.X202310598374)。

摘　　要：目的:揭示小鼠视神经损伤后视网膜神经节细胞(RGCs)凋亡的分子机制和潜在的治疗靶点。方法:构建视神经损伤(ONC)小鼠模型,对视网膜组织进行HE染色观察组织病理学改变,并使用实时荧光定量PCR(RT-qPCR)检测RGCs标志物—具有多重剪接的RNA结合蛋白(Rbpms)的mRNA表达水平;透射电镜(TEM)观察RGCs线粒体损伤情况;采用转录组测序分析ONC 7 d组小鼠的视网膜差异表达基因,并对差异表达基因进行Gene Ontology(GO)功能、Kyoto Encyclopedia of Genes and Genomes(KEGG)通路富集和韦恩分析,RT-qPCR检测差异基因mRNA表达水平。结果:与正常组相比,ONC 7 d组RGCs数量显著减少,细胞排列疏松不规则,RGCs标志物Rbpms的mRNA表达水平显著降低(P<0.001),ONC小鼠模型构建成功。透射电镜结果表明,ONC 7 d组RGCs的线粒体出现肿胀,嵴消失;转录组测序结果表明,ONC 7 d组与正常组比,存在562个差异表达基因,其中152个基因表达下调和410个基因表达上调。GO功能富集分析和KEGG通路富集分析表明ONC后差异表达基因主要富集于神经元损伤修复的多个重要信号通路;RT-qPCR结果显示,与正常组相比,与神经元损伤修复相关的基因Ecel1、Atf3、Sprr1a的表达均在损伤后第4天显著上调(均P<0.05),与测序结果一致。结论:ONC小鼠RGCs凋亡与线粒体损伤有关,而神经元损伤修复相关基因Atf3、Ecel1、Sprr1a在视神经损伤的早期参与了小鼠视神经损伤后RGCs的保护。Objective:To explore the molecular mechanism and potential therapeutic targets of retinal ganglion cells(RGCs)apoptosis after optic nerve crush in mice.Methods:An optic nerve crush(ONC)mouse model was constructed,the retinal tissue was stained with HE staining to observe the histamathological changes,and the mRNA expression level of RGCs marker,RNA binding proteins with multiple splicing(Rbpms)was detected by reverse transcription-quantitative PCR(RT-qPCR).The mitochondrial damage of RGCs was observed by transmission electron microscopy(TEM).Transcriptomic sequencing was used to analyze the retinal differentially expressed genes of ONC 7 d group mice,and the differentially expressed genes were analyzed by Gene Ontology(GO)function,Kyoto Encyclopedia of Genes and Genomes(KEGG)pathway enrichment and Wayne analysis.The mRNA expression levels of differentially expressed genes were detected by RT-qPCR.Results:Compared with the normal group,the number of RGCs in the ONC 7 d group was significantly reduced,the cell arrangement was loose and irregular,and the mRNA expression level of RGCs marker Rbpms was significantly decreased(P<0.001).The ONC mouse model was successfully constructed.The results of TEM showed that the mitochondria of RGCs in the ONC 7 d group were swollen and the ridge disappeared.Transcriptome sequencing results showed that there were 562 differentially expressed genes in the ONC 7 d group compared with the normal group,among which 152 genes were down-regulated and 410 genes were up-regulated.GO functional enrichment analysis and KEGG pathway enrichment analysis showed that the differentially expressed genes after ONC were mainly concentrated in several important signaling pathways involved in neuronal damage repair.The RTqPCR results showed that compared with the normal group,the expression of Ecel1,Atf3,and Sprr1a genes related to neuronal damage repair was significantly up-regulated on the 4th day after injury(all P<0.05),which was consistent with the sequencing results.Conclusion:The apoptosis of

关 键 词：视神经损伤 视网膜神经节细胞 青光眼 转录组分析 

分 类 号：R774.1[医药卫生—眼科]