【作者】 杨丽；

【导师】 李博安；

【作者基本信息】 厦门大学 ， 生物学， 2019， 硕士

【摘要】 现有的常规非侵入性肿瘤标志物用于检测早期胃癌(EGC)时,表现出较低的敏感性和特异性。因此,本项目致力于筛选鉴定具有较高敏感性和稳定性的早期胃癌特异性外泌体长非编码RNA作为生物标志物。在本研究中,分离了来自5个健康个体和10个I期胃癌患者的血浆外泌体,以及来自4个人原代胃上皮细胞系和4个胃癌细胞系培养基上清的外泌体。然后,通过RNA-seq绘制外泌体RNA表达谱,筛选鉴定早期胃癌特异性外泌体长非编码RNA。与健康个体和人原代胃上皮细胞系相比,在I期胃癌患者和胃癌细胞系中,分别有79个和285个外泌体RNA的表达水平显著上调。对RNA测序结果进行组合分析,我们筛选到两个早期胃癌特异性外泌体长非编码RNAs:lncUEGC1和lncUEGC2。通过在早期胃癌血浆和胃癌细胞系进一步进行实时荧光定量PCR实验,证实了外泌体中lncUEGC1和lncUEGC2显著上调。此外,稳定性测试表明几乎所有的外泌体lncUEGCl都被包裹在外泌体里面,因此免受RNA酶降解。接着,通过绘制受试者工作特征曲线(AUC)对外泌体lncUEGC1的诊断准确性进行评估。lncUEGC1在区分早期胃癌患者和健康个体时,AUC值是0.8760;而在区分癌前病变期慢性萎缩性胃炎患者和健康个体时,AUC值是0.8406。癌胚抗原在区分早期胃癌患者和健康个体时,AUC值是0.6424;而在区分癌前病变期慢性萎缩性胃炎患者和健康个体时,AUC值是0.6123。由此可知,lncUEGC1诊断准确率高于癌胚抗原的诊断准确率。由于外泌体lncUEGC1对早期胃癌的诊断具有高灵敏度,高稳定性和非侵入性的特点。因此,外泌体lncUEGC1有望被开发作为早期胃癌诊断的生物标志物。更多还原

【Abstract】 Existing conventional tumor markers for non-invasive diagnosis of gastric cancer(GC)exhibit lower sensitivity and specificity when were used in the detection of early gastric cancer(EGC).Therefore,this proj ect is dedicated to screening and identifying specific exosomal 1ncRNA biomarkers for early gastric cancer,which have high sensitivity and stability for the diagnosis of EGC.In the present study,we isolated plasma exosomes from 5 healthy individuals and 10 stage I GC patients,as well as cell line culture supernatants exosomes from 4 human primary gastric epithelial cell lines(HPGECs)and 4 GC.Then,Exosomal RNA profiling was performed using RNA sequencing to screened and identify EGC-specific exosomal IncRNAs.Respectively,compared with healthy individuals and HPGECs,the expression levels of 79 and 285 exosomal RNAs were significantly up-regulated in stage I gastric cancer and gastric cancer cell lines.Through combinational analysis of the RNA sequencing results,we found two EGC-specific exosomal IncRNAs,lncUEGCl and lncUEGC2.Further real-time fluorescence quantitative PCR experiments in EGC plasma and gastric cancer cell lines(GCCs)confirmed that lncUEGC1 and lncUEGC2 were significantly up-regulated in exosomes.In addition,stability testing indicated that almost all exosomes lncUEGCl were encapsulated in the exosomes and thus protected from RNase degradation.Next,the diagnostic accuracy of the exosomal lncUEGC1 was evaluated.When lncUEGC1 distinguishes between early gastric cancer patients and healthy individuals,the AUC value is 0.8760.When lncUEGC1 differentiates patients with chronic atrophic gastritis and healthy individuals from precancerous lesions,the AUC value is 0.8406.However,the carcinoembryonic antigen has an AUC value of 0.6424 when distinguishing between early gastric cancer patients and healthy individuals.When carcinoembryonic antigen differentiates patients with chronic atrophic gastritis and healthy individuals from precancerous lesions,the AUC value is 0.6123.It can be seen that the diagnostic accuracy of IncUEGC1 is higher than carcinoembryonic antigen.The exosomal IncUEGC1 is highly sensitive,stable and non-invasive to the diagnosis of early gastric cancer.Therefore,it is expected to develop biomarkers for the diagnosis of early gastric cancer.更多还原