5种核碱丙氨酸的合成及其抑制大鼠肝癌细胞RH35增殖的作用机制

doi:10.16098/j.issn.0529-1356.2020.02.008

解剖学报 ›› 2020, Vol. 51 ›› Issue (2): 195-205.doi: 10.16098/j.issn.0529-1356.2020.02.008

5种核碱丙氨酸的合成及其抑制大鼠肝癌细胞RH35增殖的作用机制

徐存拴¹ 宋亚萍¹ 徐凯¹ 杨晖¹ 丁一¹ 毕晶晶² 赵茜怡^2* 徐霆^2*

1. 河南师范大学生命科学学院河南省科技部共建细胞分化调控国家重点实验室培育基地，河南新乡 453007；2. 河南师范大学化学化工学院，河南新乡 453007

收稿日期:2019-01-03 修回日期:2019-04-23 出版日期:2020-04-06 发布日期:2020-04-06
通讯作者: 赵茜怡;徐霆 E-mail:xtsheep@163.com
基金资助:
细胞分化调控国家重点实验室培育基地研究项目

Synthesis of five types of nucleobase-alanines and their mechanisms of inhibiting proliferation of rat hepatoma cells RH35

XU Cun-shuan¹ SONG Ya-ping¹ XU Kai¹ YANG Hui¹ DING Yi¹ BI Jing-jing² ZHAO Qian-yi^2* XU Ting^2*

1.State Key Laboratory Cultivation Base for Cell Differentiation Regulation, School of Life Science, He’nan Normal University, He’nan Xinxiang 453007, China; 2.School of Chemistry and Chemical Engineering, He’nan Normal University, He’nan Xinxiang 453007, China

Received:2019-01-03 Revised:2019-04-23 Online:2020-04-06 Published:2020-04-06
Contact: ZHAO Qian-yi;XU Ting E-mail:xtsheep@163.com

摘要/Abstract

摘要：

目的合成5种核碱丙氨酸，探讨其细胞毒性及机制。方法用丝氨酸-内酯亲核开环方法合成 5种核碱丙氨酸，用MTT法检测它们对大鼠正常肝细胞BRL-3A和大鼠肝癌细胞RH35活性的影响，流式细胞术检测它们对上述BLR-3A和RH35细胞周期进程和凋亡的影响，用Real-time PCR检测它们对上述两种细胞增殖/凋亡相关基因表达的影响。结果合成了1-尿嘧啶-L-丙氨酸、1-胸腺嘧啶-L-丙氨酸、1-胞嘧啶-L-丙氨酸、9-腺嘌呤-L-丙氨酸和9-鸟嘌呤-L-丙氨酸等5种核碱丙氨酸。其中，1-胞嘧啶-L-丙氨酸和9-鸟嘌呤-L-丙氨酸对RH35的半数抑制浓度（IC₅₀）值显著低于对BRL-3A的 IC₅₀ 值，它们促进RH35凋亡的作用显著强于对BRL-3A作用，它们促进RH35凋亡相关基因CASP3、CASP9和Bax的表达上调幅度，抑制细胞增殖相关基因CCNA2和PCAN的表达下调幅度和抑制抗凋亡相关基因Bcl-2的表达下调幅度均显著高于对BRL-3A作用。结论 1-胞嘧啶-L-丙氨酸和9-鸟嘌呤-L-丙氨酸通过显著抑制RH35增殖相关基因表达，促进凋亡相关基因表达而抑制其活性和促进其凋亡。

关键词: 核碱丙氨酸, 细胞毒性, 细胞增殖动力学, 流式细胞术, 大鼠

Abstract:

Objective Five types of nucleobase-alanines were synthesized to study their cytotoxicity and mechanism. Methods Five types of nucleobase-alanines were synthesized by the method of nucleophilic ring-opening of serine-lactone. MTT assay was used to detect their effects on the viability of rat normal hepatocytes BRL-3A and rat hepatoma cells RH35. Flow cytometry was used to detect their effects of cell cycle and apoptosis on BLR-3A and RH35. Real-time PCR was performed to detect their effects of proliferation/apoptosis-related gene expression on these two cells. Results Five types of nucleobase-alanines were synthesized, including 1-uracil-L-alanine, 1-thymine-L-alanine, 1-cytosine-L-alanine, 9-adenine-L-alanine and 9-guanine-L-alanine. Among them, 1-cytosine-L-alanine and 9-guanine-L-alanine had significant lower half inhibition concentration(IC₅₀) values for RH35 than for BRL-3A, and their effects of promoting RH35 apoptosis were significantly stronger than that of BRL-3A, the amplitude of their promoting RH35 up-regulation apoptosis-related genes CASP3, CASP9 and Bax, the down-regulation of cell proliferation-related genes CCNA2 and PCAN, as well as the down-regulation of anti-apoptosis-related gene Bcl-2 were significantly higher than those of BRL-3A. Conclusion 1-cytosine-L-alanine and 9-guanine-L-alanine showed significant effects of inhibiting the viability and promoting apoptosis by up-regulating the expression of proliferation-related genes and down-regulating the expression of apoptosis-related genes more potently.

Key words: Nucleobase-alanine, , Cytotoxicity, , Cell proliferation kinetics, , Flow cytometry, , Rat

中图分类号:

Q291

徐存拴宋亚萍徐凯杨晖丁一毕晶晶赵茜怡徐霆. 5种核碱丙氨酸的合成及其抑制大鼠肝癌细胞RH35增殖的作用机制[J]. 解剖学报, 2020, 51(2): 195-205.

XU Cun-shuan SONG Ya-ping XU Kai YANG Hui DING Yi BI Jing-jing ZHAO Qian-yi XU Ting . Synthesis of five types of nucleobase-alanines and their mechanisms of inhibiting proliferation of rat hepatoma cells RH35[J]. Acta Anatomica Sinica, 2020, 51(2): 195-205.

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5种核碱丙氨酸的合成及其抑制大鼠肝癌细胞RH35增殖的作用机制

Synthesis of five types of nucleobase-alanines and their mechanisms of inhibiting proliferation of rat hepatoma cells RH35

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