伏立诺他增强肝癌细胞对5-氟尿嘧啶及奥沙利铂的化疗敏感性

发布时间：2018-03-21 04:15

本文选题：肝细胞癌　切入点：伏立诺他　出处：《华中科技大学》2015年博士论文　论文类型：学位论文

【摘要】：第一部分伏立诺他通过抑制胸苷酸合成酶表达增强肝癌细胞对5-氟尿嘧啶的化疗敏感性目的：研究伏立诺他和5-氟尿嘧啶在肝癌细胞中的联合作用,探讨两者联用的协同效应。方法：用伏立诺他和(或)5-氟尿嘧啶处理HepG2、SMMC7721及BEL7402细胞。进行细胞毒性实验、平板克隆形成实验、软琼脂克隆形成实验、细胞周期检测、细胞凋亡检测、普通PCR、蛋白免疫印迹、免疫组织化学及动物实验。结果：伏立诺他及5-氟尿嘧啶均能抑制HepG2、SMMC7721及BEL7402细胞的增殖。在这三种细胞系中，两者联合作用的联合指数均小于1,减量指数均大于1,表明两者具有协同效应。两者联用引起G0/G1期细胞周期阻滞和caspase相关通路的细胞凋亡。两者联用在体内体外均使成瘤能力下降。5-氟尿嘧啶能引起胸苷酸合成酶表达的上调,而无论其存在与否,伏立诺他均能抑制胸苷酸合成酶的表达。结论：伏立诺他及5-氟尿嘧啶在肝癌中联用具有协同效应。两者联用通过引起细胞周期阻滞和凋亡以及抑制胸苷酸合成酶表达在体内体外均抑制细胞增殖和成瘤能力。这两者的合用或许能在肝癌治疗中起到一定作用。第二部分伏立诺他通过抑制Notch通路增强肝癌细胞对奥沙利铂的化疗敏感性目的：探讨伏立诺他和奥沙利铂在肝癌细胞中的联合作用及两者联用的协同效应。方法：用伏立诺他和(或)奥沙利铂处理SMMC772、BEL7402及HepG2细胞。进行细胞活性实验、平板克隆形成实验、软琼脂克隆形成实验、细胞周期分析、细胞凋亡分析、蛋白免疫印迹、动物皮下成瘤实验及免疫组织化学。结果：伏立诺他及奥沙利铂均能抑制：SMMC7721、BEL7402及HepG2细胞的增殖。两者联合作用后,这三种细胞系中的联合指数均小于1,而减量指数均大于1,表明两者具有协同效应。两者联用引起G2/M期细胞周期阻滞和caspase相关通路的细胞凋亡。在体内体外实验中,两者联用均抑制成瘤能力。奥沙利铂激活Notch信号通路表达,而无论其存在与否,伏立诺他均能抑制Notch信号通路的表达,从而增加奥沙利铂的化疗敏感性。结论：体内体外实验证实伏立诺他及奥沙利铂在肝癌中协同抑制细胞增殖和成瘤能力。这是通过引起细胞周期阻滞和凋亡以及抑制]Notch通路表达来实现的。这两种药物的联用可能在今后的肝癌治疗中发挥作用。
[Abstract]:Part one: Volinotaxel enhances the chemosensitivity of hepatoma cells to 5-fluorouracil by inhibiting the expression of thymidine synthase. Aim: to study the synergistic effect of volenotta and 5-fluorouracil in hepatocellular carcinoma cells. Methods: HepG2SM-SMMC7721 and BEL7402 cells were treated with volenostata and (or 5-fluorouracil). Cytotoxicity test, plate clone formation test, soft Agar clone formation assay, cell cycle detection, apoptosis detection, ordinary PCR, Western blotting were performed. Immunohistochemistry and animal experiments. Results: both volenotta and 5-fluorouracil inhibited the proliferation of HepG2SMMC7721 and BEL7402 cells. The combined index of the combined action was less than 1, and the decrement index was greater than 1, which indicated that both of them had synergistic effect. Combined use of both could induce cell cycle arrest in G _ 0 / G _ 1 phase and apoptosis of caspase related pathway. In vitro and in vivo, both of them were used in vitro and in vivo. Decreased tumorigenesis. 5 fluorouracil could induce the up-regulation of thymidine synthase expression. Volenoate inhibited the expression of thymidine synthase, whether it existed or not. Conclusion: the combined use of volenotta and 5-fluorouracil in hepatocellular carcinoma has synergistic effect, which can inhibit cell proliferation and proliferation in vitro and in vivo by inducing cell cycle arrest and apoptosis and inhibiting thymidine synthase expression in vitro and in vivo. Tumor capacity. The combination of the two may play a role in the treatment of liver cancer. The second part of Volinoxat enhances the chemosensitivity of hepatoma cells to oxaliplatin by inhibiting the Notch pathway. Aim: to investigate the synergistic effect of volenotta and oxaliplatin in hepatocellular carcinoma cells. Methods: SMMC772BEL7402 and HepG2 cells were treated with volenostata and / or oxaliplatin. The cell activity, plate clone formation, soft Agar clone formation, cell cycle analysis, apoptosis analysis, Western blot analysis were performed. Animal subcutaneous tumorigenesis and immunohistochemistry. Results: both volenotta and oxaliplatin could inhibit the proliferation of HepG2 and BEL7402 cells. The combined index of these three cell lines is less than 1, and the decrement index is greater than 1, which indicates that both have synergistic effect. The combination of them can induce cell cycle arrest and apoptosis related to caspase pathway in G 2 / M phase. In vitro and in vivo, Oxaliplatin activates the expression of Notch signaling pathway, but volenoside inhibits the expression of Notch signaling pathway, thus increasing the chemosensitivity of oxaliplatin. Conclusion: in vitro and in vivo experiments have proved that volenotta and oxaliplatin co-inhibit cell proliferation and tumorigenesis in hepatocellular carcinoma by inducing cell cycle arrest and apoptosis and inhibiting the expression of] Notch pathway. Combination of these drugs may play a role in the treatment of liver cancer in the future.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R735.7

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