六味地黄丸入血成分增效自杀基因杀伤肝癌细胞的缝隙连接机制  

Gap Junction Mechanisms of Liuwei Dihuang Pills Constituents Acting on Blood for Enhancing Effect of Suicide Gene Therapy on Killing Hepatoma Cells

作　　者：刘娟[1] 周瑶[2] 张广献[3] 谭宇蕙[3] 肖建勇[3] 吴映雅[3] 曾玲[1] 杜标炎[4] 

机构地区：[1]广州中医药大学 [2]湖南中医药高等专科学校形态教研室,湖南株洲412012 [3]广州中医药大学生化教研室,广东广州510006 [4]广州中医药大学病理教研室,广东广州510006

出　　处：《广州中医药大学学报》2014年第1期103-108,170,共7页Journal of Guangzhou University of Traditional Chinese Medicine

基　　金：国家自然科学基金资助项目(编号:81072906)

摘　　要：【目的】探讨六味地黄丸主要入血成分处理的淋巴上清对大鼠肝癌细胞CBRH7919缝隙连接功能的影响,以期阐明其增强自杀基因疗法杀伤肝癌细胞效应的作用机制。【方法】取大鼠脾淋巴细胞,加入六味地黄丸的5种主要入血成分,体外培养后取含药淋巴上清,采用流式细胞术结合荧光示踪法分析其对细胞缝隙连接通讯功能(GJIC)的影响;利用Western blot法检测含药淋巴上清对大鼠肝癌CBRH7919细胞Cx43蛋白表达的影响;利用GJ阻滞剂甘草次酸(GA)观察阻滞GJIC后,含药淋巴细胞上清联合tk/GCV系统对细胞杀伤作用的影响。【结果】流式细胞术结合荧光示踪法结果表明,占培养液15%(体积分数)的含药淋巴上清能促进大鼠肝癌CBRH7919细胞的GJIC功能;Western blot法结果显示,含药淋巴细胞上清能提高大鼠肝癌CBRH7919细胞Cx43蛋白的表达水平;GA阻断GJIC功能后,含药淋巴细胞上清联合tk/GCV系统对细胞杀伤作用降低。【结论】六味地黄丸入血成分处理的淋巴上清可通过缝隙连接机制发挥对自杀基因杀伤大鼠肝癌细胞的增效作用。Objective To investigate the effects of lymphatic supernatant fluid activated by Liuwei Dihuang Pills d^-1） , and high-, middle- and low-dose KD groups （in the dosage of 18.3, 9.2, 4.6 g·kg^-1·d^-1, respectively） . UC rat model was established with trinitro-benzene-sulfonic acid and were treated with the corresponding medicine according to the experimental design. El, UBC5 and E3RSIKB mRNA expression levels in colonic mucosa were tested by reverse transcription-polymerase chain reaction （RT-PCR）, and El, UBC5 and E3RSIKB protein contents were detected by enzyme-linked immunosorbent assay （ELISA） . Results The relative contents of E1 and UBC5 mRNA and protein in the colon of the model group had no statistically difference from those of the normal group （P〉0.05）, and the relative contents in the colon of high-, middle- and low-dose KD groups either did not differ from those of the model group （P〉0.05, except for UBC5 protein expression in high-dose KD group） .The relative contents of E3RSIKB mRNA and protein in the model group were significantly higher than those of the normal group （P〈0.05） . The relative content of E3RSIKB mRNA expression in high- and middle-dose KD group and the relative contents of E3RSIKB protein expression in high- and low-dose KD groups were signifieantly lower than those of the model group （P〈0.05 or P〈0.01） Conclusion The therapeutie meehanism of KD for UC may be related with the obstruction of ubiquitin degradation through inhibiting E3RSIKB expression, thus to decrease the activation of NF-KB and relieve the inflammation.

关 键 词：肝癌 中药疗法 六味地黄丸 药理学 入血成分 淋巴上清 自杀基因 缝隙连接 肝癌细胞 病理学 细胞培养 

分 类 号：R285.5[医药卫生—中药学]