【作者】 柳长明；

【导师】 刘铜军；

【作者基本信息】 吉林大学 ， 外科学， 2009， 博士

【摘要】 结肠癌是胃肠道中常见的恶性肿瘤,以41-65岁发病率高。在我国近20年来,尤其在大城市发病率明显上升,且结肠癌多于直肠癌的趋势,结肠癌病因虽未明确,但其相关的高危因素逐渐被认识,如过多的动物脂肪及动物蛋白饮食,缺乏新鲜蔬菜及维生素食品;缺乏适度的体力活动。遗传易感性在结直肠癌的发病中也具有重要地位,结肠腺瘤,溃疡性结肠炎以及结肠血吸虫病肉芽肿,与结肠癌的发生有较密切的关系。中国人直肠癌与西方人比较有三个流行病学特点:①直肠癌比结肠癌发生率高,约1.5:1;②低位直肠癌所占比例高,约占直肠癌的60%-75%;③青年人(<30)直肠癌比例高,约10%-15%。癌的发生发展是一个多步骤、多阶段及多基因参与的细胞遗传性疾病[141]。随着分子生物学技术快速发展,进一步研究大肠癌发病机理、生物学诊断和治疗,成为目前研究的热点。高脂肪饮食与特定器官包括乳腺、胰腺、结肠、前列腺等发病率及肿瘤生长相关联。也证实花生四烯酸、多不饱和脂肪酸代谢是通过环氧合酶(cox)途径或脂氧合酶(lox)途径,产生一系列促炎性反应物质类花生酸类物质,包括前列腺素、血栓烷素、白细胞三烯等.这些物质在人类癌症发生、发展中发挥重要生物效应。环氧合酶在人许多恶性肿瘤发生、发展起到重要作用,它的抑制剂相应被呈现出。脂氧合酶和其代谢产物在人类癌细胞组织中明显增多,这一过度表达对于肿瘤细胞增殖、抵抗吞噬作用和血管发生更具有理论及实践意义。本实验通过RT-PCR、Western-blot和免疫组织化学法不同方法对结直肠癌患者切除标本进行5-lox和cox-2测定及5-lox抑制剂对5-loxmRNA与bcl-2mRNA和baxmRNA表达的影响,在国内尚未见报道。结果表明5-loxmRNA和cox-2mRNA在结直肠癌组织中表达增高,与结直肠癌临床分期、肿瘤大小、肿瘤分化程度有关;5-lox蛋白和cox-2蛋白表达癌肿组织明显高于癌旁组织(western-blot),5-lox和cox-2表达水平与微血管密度相关联;5-loxmRNA表达与bcl-2mRNA表达相一致,而与baxmRNA表达呈负相关;5-lox抑制剂MK886对5-loxmRNA、bcl-2mRNA表达有抑制作用,表明对环氧合酶和脂氧合酶的深入研究具有临床实用性和先进性,为今后结直肠癌诊断和治疗提供了很好理论和实践意义。更多还原

【Abstract】 Objective: To study expressions of 5-Lipoxygenase and Cyclooxygenase-2 in human colorectal cancer, and approach potential mechanism of 5-lox in the promotion and progression of human colorectal cancer, it provides a good theoretical and practical significance For the future diagnosis and treatment of colorectal cancer.Background:Evidence from epidemiological and animal studies suggests that a high-fat consumption is associated with an increased incidence and growth of tumors at several specific organ sites, it has been demonstrated that the metabolism of arachidonic acid, a polyunsaturated fatty acid by either the Cyclooxygenase (cox) pathway or the Lipoxygenase (lox) pathway, generates a host of pro-inflammatory substances called eicosanoids including prostaglandins, thromboxanes, leukotrienes, etc. that act as significance effect of biological in the promotion and progression of many types of human neoplasms. it suggests that the deep investigation ofcyclooxygenase and lipoxygenase possess clinical utility and advanced ,for the future diagnosis and treatment of colorectal cancer provides a good theoretical and practical significance.Part 1 Expressions of 5-Lipoxygenase and Cyclooxygenase-2 in human colorectal tumor and adjacent normal mucosaMethod:1、To investigate the expressions of 5-loxmRNA and cox-2mRNA in paired 39 colorectal cancer samples and their relationship by RT-PCR. 2、The relative quantity of protein 5-lox and cox-2 in colorectal tumor and adjacent normal mucosa samples was determined by western-blot.3、Expressions of 5-Lipoxygenase and Cyclooxygenase-2 in human colorectal cancer samples is related with microvessel density by immunohistochemical staining methods.Result:There were 33 cases of 5-loxmRNA amplified bands of 504bp in39 cases of colorectal cancer tissues, positive expression ratio was 84.6% (33/39); there were 30 cases cox-2mRNA amplified bands of 540bp in 39 cases of colorectal cancer tissues, positive expression ratio was 76.9% (30/39). Expressions of 5-loxmRNA and cox-2mRNA in colorectal cancer tissue were remarkably elevated,and correlated with clinical stage, tumor size,differentiation degree. Expressions of both were not coordinated. Expressions of 5-lox and cox-2 were remarkably elevated in human colorectal tumor tissue in comparison with normal colon mucosa. And the over expression level of 5-lox and cox-2 was positively correlated with microvessel density.Part 2 Expressions of 5-loxmRNA is correlated with expression of bcl-2mRNA and bax mRNA in paired 39 colorectal tumor and adjacent normal mucosa samples and the effect of 5-lox MK886 inhibitor on expression of 5-loxmRNA、bcl-2mRNA and baxmRNAMethod:1、Expressions of 5-loxmRNA、bcl-2mRNA and baxmRNA in paired 39 colorectal tumor and adjacent normal mucosa samples by RT-PCR.2、Expressions of 5-lox mRNA、bcl-2mRNA and baxmRNA compared with before and after MK886 in HT29 colon cancer cell lines.Result:Expressions of 5-loxmRNA、bcl-2mRNA and baxmRNA are separately 33/39, 29/39, 10/39 and 14/39, 10/39, 26/39in paired 39 colorectal tumor and adjacent normal mucosa samples.Expressions of 5-loxmRNA and bcl-2mRNA were more elevated than expression of baxmRNA in human colorectal tumor tissue, Expression of baxmRNA were more higher than expressions of 5-loxmRNA and bcl-2mRNA in human adjacent normal mucosa tissue, Expressions of 5-loxmRNA and bcl-2mRNA were lower by after Mk886 in comparison with befor Mk886 in HT29 colon cancer cell lines. Expressions of baxmRNA were higher by after Mk886 in comparison with before Mk886 in HT29 colon cancer cell lines. Expressions of 5-lox mRNA, bcl-2mRNA are gradually reduced with augmentation of drug concentration, Expression of baxmRNA is gradually increased With augmentation of drug concentration.Conclusion:1、Expressions of 5-loxmRNA and cox-2mRNA were simultaneously up- regulated in human colorectal cancer tissue, Expressions of both were not coordinated.2、Protein expressions of 5-lox and cox-2 were remarkably elevated in human colorectal tumor tissue in comparison with normal colon mucosa.3、The over expressions level of 5-lox and cox-2 was positively correlated with microvessel density.4、5-lox may play a role of promoter in the promotion and development of colorectal cancer by influencing the balance of between cell proliferation and apoptosis.更多还原