Expression of telomerase activity in 4NQO induced carcinoma in rat tongue

4NQO诱导大鼠舌癌端粒酶活性的表达

• 批准号: 11672027
• 负责人: SHIGERU Ueno
• 金额: $ 1.47万
• 依托单位: Osaka Dental University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 1999
• 资助国家: 日本
• 起止时间: 1999 至 2000
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-11672027/
• 关键词: telomerase; chemically induced carcinoma; 4-nitroquinoline 1-oxide; stretch PCR method

Correlation between telomerase activity and apoptosis was examined in chemically induced tongue carcinomas using 5-week-old male Sprague-Dowley rats. Animals were given 50 ppm 4-nitroquinoline 1-oxide solution as drinking water for 4, 8, 12, 16, 20, and 24 weeks and sacrificed. Tongue tissue was prepared to investigate apoptotic cells and p53 positive cells by TUNEL method and immunohistochemical method, respectively. Telomerase activity was also examined by stretch PCR method. Namely, telomerase containing solution was extracted from each group of rats, and stretch reaction by telomerase was performed at 30℃. After amplification by PCR with rTaq, samples were applied to an acrylamide gel. Electrophoresis was carried out at 100V under a constant temperature. The gel was stained with SYBRgreen and observed. Epithelial tissue of the tongue showed hyperplastic change in 8 weeks, dysplastic change in 12 weeks, and carcinogenesis in 16 weeks after the beginning of 4NQO administration. p53 overexpression increased in 4 weeks and was kept high level through experimental period. Apoptotic cells also increased till 16 weeks, but decreased thereafter. Telomerase activity was observed in all animals including 4-week group. As a result, telomerase activity began to appear coincident with p53 overexpression proceeding the initiation of histological changes. So it is suggested that apoptosis and immortalization of the tissue should be equally involved in the early stage of chemically induced carcinoma.

用5周龄雄性SD大鼠研究了化学诱导的舌癌中端粒酶活性与细胞凋亡的相关性。给予50ppm的4-硝基喹啉1-氧化物溶液作为饮用水，分别饮用4、8、12、16、20和24周后处死动物。取舌组织，分别用TUNEL法和免疫组织化学方法检测凋亡细胞和P53阳性细胞。用延伸聚合酶链式反应方法检测端粒酶活性。即每组大鼠抽提含端粒酶的溶液，在30℃进行端粒酶拉伸反应。经rTaq聚合酶链式反应扩增后，将样品加到丙烯酰胺凝胶上。在100V恒温下进行电泳法。用SYBRgreen对凝胶进行染色观察。舌上皮组织在4NQO给药后8周出现增生性改变，12周出现异常改变，16周出现癌变。P53过表达在4周后开始增加，并在整个实验期间保持较高水平。细胞凋亡率在16周前也有所增加，但之后逐渐减少。包括4周组在内的所有动物均观察到端粒酶活性。结果，端粒酶活性开始与P53的过度表达一致，从而启动了组织学改变。因此，在化学诱癌的早期阶段，应同时考虑组织的凋亡和永生化。