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目的:利用细胞角蛋白19(K19)启动子构建胃黏膜上皮细胞中JC病毒(John Cunningham virus,JCV)T抗原表达转基因鼠,试图建立胃黏膜自发肿瘤动物模型。方法:利用限制性内切酶NdeⅠ,酶切K19-JCV T抗原表达质粒,电泳分离带有K19启动子的T抗原核酸序列后,显微注射入C57BL/6J30只小鼠受精卵中。PCR检测转基因阳性鼠,组织学观察主要脏器的形态学表现及免疫组化检测T抗原表达。结果:显微注射K19-JCV T抗原(KT)DNA片段后成功获得11种转基因阳性鼠,其中2个月龄KT7的各脏器组织学检测显示,脑、肝、肾、食管、前胃、胃、小肠、大肠、胰腺、肺、心脏和脾组织未见异常形态学改变,JCV T抗原特异表达于胃黏膜上皮细胞和支气管上皮细胞的细胞核中,16个月龄的KT7转基因鼠13.3%(2/15)出现肺肿瘤和腺癌的病理学改变,T抗原定位于癌细胞细胞核中。结论:JCV T抗原直接诱发肺肿瘤动物模型的建立为JCV T抗原嵌入所致上皮肿瘤提供了直接的实验依据,为JCV所致上皮肿瘤发生和演进分子机制奠定基础,同时为抗肿瘤药物筛选及基因治疗监控提供了良好的动物模型。
OBJECTIVE: To construct a transgenic mouse expressing T antigen of JC virus (JCV) in gastric epithelial cells by using cytokeratin 19 (K19) promoter in an attempt to establish a spontaneous gastric tumor animal model. Methods: The restriction endonuclease NdeⅠ was used to digest the K19-JCV T antigen expression plasmid. The T antigen with K19 promoter was isolated by electrophoresis and then microinjected into the fertilized eggs of C57BL / 6J30 mice. The positive transgenic mice were detected by PCR, the morphological features of major organs were observed by histology and the expression of T antigen was detected by immunohistochemistry. RESULTS: Eleven transgenic mice were successfully obtained after KT-KT DNA fragment was injected microinjected. The histological examination of KT7 at 2 months showed that the brain, liver, kidney, esophagus, anterior stomach, No abnormal morphological changes were observed in the stomach, small intestine, large intestine, pancreas, lung, heart and spleen tissues. JCV T antigen was specifically expressed in the nucleus of gastric mucosal epithelial cells and bronchial epithelial cells, 13.3% of KT7 transgenic mice at 16 months of age 2/15) pathological changes of lung tumors and adenocarcinomas, T antigen located in the nucleus of cancer cells. Conclusion: The establishment of an animal model of lung tumor induced by JCV T antigen provides a direct experimental basis for JCV T antigen-embedded epithelial tumor, which lays a foundation for the molecular mechanism of JCV-induced epithelial tumorigenesis and progression. Meanwhile, Gene therapy monitoring provides a good animal model.