SIRT4在人胃肠道肿瘤中的表达及其对人结肠癌细胞生物学行为的影响研究

发布时间：2018-10-24 10:15

【摘要】：背景:SIRT家族(SIRT1-7)是一个NAD+-依赖的去乙酰化酶、乙酰化酶以及ADP-核糖基转移酶家族,几乎所有SIRT家族成员都被认为在肿瘤形成中起到重要作用。最近有研究表明,SIRT4能通过调节谷氨酰胺代谢而具有抑癌基因的作用,但目前还没有直接在人体组织蛋白水平进行的SIRT4与人体肿瘤的相关性研究。方法:我们通过RT-PCR检测了16对人结肠癌和癌旁正常结肠组织标本中SIRT4的mRNA表达水平,然后进一步分析了TCGA数据库中包含223例样本的人结肠癌表达谱芯片数据中SIRT4的mRNA表达水平,接下来分别对包含89例人结肠癌组织和75例人胃癌组织的组织芯片进行免疫组化实验并分析SIRT4表达与人结肠癌和胃癌患者的临床病理参数之间的关系。我们通过慢病毒构建过表达SIRT4的人结肠癌细胞RKO和HT29的稳定株,分析了过表达SIRT4对人结肠癌RKO和HT29的增殖、克隆形成的影响,并进一步分析过表达SIRT4之后这些细胞对葡萄糖代谢抑制剂2-DG以及化疗药5-FU的敏感性的影响。我们还分析了过表达SIRT4对RKO和HT29细胞在谷氨酰胺缺乏培养基中存活率的影响。结果:SIRT4在结肠癌和胃癌组织中的表达水平显著低于癌旁正常组织,SIRT4表达降低提示结肠癌和胃癌更差的病理分化程度(分别为:p=0.01,p=0.002),同时SIRT4表达低的结肠癌患者比SIRT4表达高的结肠癌患者术后5年生存率更差(p=0.0306)。过表达SIRT4抑制人结肠癌细胞RKO和HT29的增殖,并增加它们对葡萄糖代谢抑制剂2-DG以及化疗药5-FU的敏感性,同时过表达SIRT4减少了RKO和HT29细胞在谷氨酰胺缺乏培养基中的存活率。结论:SIRT4在人结肠癌和胃癌中扮演了抑癌基因的角色,SIRT4与人结肠癌和胃癌的病理分化程度相关,与人结肠癌患者的术后5年生存率相关。SIRT4增加人结肠癌细胞对葡萄糖抑制剂2-DG和化疗药5-FU的敏感性。SIRT4对人结肠癌细胞的抑制作用不仅仅是通过抑制谷氨酰胺代谢引起的。SIRT4是人结肠癌和胃癌一个很有希望的的诊断标志物,是人结肠癌一个很好的治疗靶点。
[Abstract]:Background: SIRT family (SIRT1-7) is a NAD-dependent deacetylase, acetylase and ADP- ribonyltransferase family. Almost all members of SIRT family are considered to play an important role in tumor formation. Recent studies have shown that SIRT4 can inhibit oncogene by regulating glutamine metabolism, but there is no direct study on the relationship between SIRT4 and human tumor at human tissue protein level. Methods: we detected the mRNA expression of SIRT4 in 16 pairs of human colon cancer and adjacent normal colon tissues by RT-PCR, and further analyzed the mRNA expression level of SIRT4 in the TCGA database containing 223 samples of human colon cancer expression microarray. Then the tissue microarray containing 89 cases of human colon cancer and 75 cases of human gastric carcinoma were examined by immunohistochemistry and the relationship between the expression of SIRT4 and the clinicopathological parameters of patients with human colon cancer and gastric cancer was analyzed. We constructed human colon cancer cells RKO and HT29 stably by lentivirus and analyzed the effect of overexpression of SIRT4 on the proliferation and clone formation of RKO and HT29 in human colon cancer. The effects of overexpression of SIRT4 on the sensitivity of these cells to glucose metabolism inhibitor 2-DG and chemotherapeutic agent 5-FU were further analyzed. We also analyzed the effect of overexpression of SIRT4 on the survival rate of RKO and HT29 cells in glutamine deficient medium. Results: the expression level of SIRT4 in colon cancer and gastric cancer was significantly lower than that in adjacent normal tissues. The decrease of SIRT4 expression indicated the worse pathological differentiation of colon cancer and gastric cancer (p0. 01 p0. 002, respectively). Meanwhile, the expression of SIRT4 in colon cancer patients with low SIRT4 expression was higher than that in SIRT4 table. The 5-year survival rate of colon cancer patients was significantly lower than that of patients with colon cancer (p0. 0306). Overexpression of SIRT4 inhibited the proliferation of RKO and HT29 in human colon cancer cells and increased their sensitivity to glucose metabolism inhibitor 2-DG and chemotherapeutic drug 5-FU. Overexpression of SIRT4 reduced the survival rate of RKO and HT29 cells in glutamine deficient medium. Conclusion: SIRT4 plays an important role as a tumor suppressor gene in human colon cancer and gastric cancer. SIRT4 is related to the degree of pathological differentiation of human colon cancer and gastric cancer. SIRT4 increases the sensitivity of human colon cancer cells to glucose inhibitor 2-DG and chemotherapeutic agent 5-FU. The inhibitory effect of SIRT4 on human colon cancer cells is not only by inhibiting glutamine metabolism. SIRT4 is a promising diagnostic marker for colon and stomach cancer. It is a good therapeutic target for human colon cancer.
【学位授予单位】：上海交通大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R735

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