IP6对卵巢癌SKOV3细胞的侵袭转移能力及Wnt通路的影响

发布时间：2018-03-28 21:00

本文选题：卵巢癌　切入点：SKOV3　出处：《河北医科大学》2017年硕士论文

【摘要】：在妇科恶性肿瘤中,死亡率第一位的是卵巢癌,组织分型中以上皮性肿瘤占85%-90%,由于卵巢癌早期无特异性症状,且缺乏高敏感性及高特异性的筛查手段以及特效的治疗方法,而传统的化疗方法存在许多副反应以及化疗药物耐药的问题,使卵巢癌的生存率仍然很低。天然药物具有经济、高效、来源广、低毒副作用等优点。近年来,以肌醇六磷酸(IP6)为代表的植物提取的天然药物以其强大的抗肿瘤作用成为近来研究的热点。Wnt通路近来被广泛关注,其可调控动物组织分化、器官形成等生理过程,通路中的关键蛋白表达异常,可导致癌症的发生。其中β-连环蛋白(β-catenin)与T细胞因子4(TCF4)都是Wnt通路中的主要成员。有研究表明卵巢良性疾病的β-catenin低于卵巢恶性疾病,并且β-catenin的异常表达与卵巢癌的临床分期、病理分型、血行转移情况有关。TCF4与β-catenin结合可以促进基因转录,许多肿瘤的发生发展均与TCF4有关。上皮间质转化过程(epithelial mesenchymal transition,EMT)是上皮细胞向间质细胞转变的过程。EMT被认为是肿瘤转移的主要原因。Wnt经典通路与EMT发生关联密切,β-catenin、TCF4均与癌细胞EMT的发生以及肿瘤侵袭转移密切相关。目的:本研究旨在探讨IP6+肌醇对卵巢癌SKOV3细胞侵袭转移能力的影响及其作用机制,以及顺铂与IP6+肌醇联合应用对卵巢癌是否有协同作用。方法:1 Transwell肿瘤细胞侵袭实验,观察IP6+肌醇剂量变化对卵巢癌细胞侵袭转移能力的影响。2 Western blot法检测IP6+肌醇对β-catenin蛋白的影响。3 RealtimePCR技术检测Wnt通路中β-catenin mRNA,TCF4mRNA表达的影响。4统计学方法:采用SPSS21.0统计学软件处理实验数据。结果:1transwell侵袭实验结果显示:联合组细胞数为26.2±2.6,顺铂组细胞数为32.0±3.4,联合组、顺铂组与对照组82.8±5.7相比细胞数明显减少,差异显著(p0.05),ip6+肌醇高、中剂量组细胞数分别为62.6±2.3,73.2±3.0,与对照组相比,差异有统计学意义(p0.05)。ip6+肌醇低剂量组细胞数为80.2±3.3,与对照组相比,差异无统计学意义(p0.05),ip6+肌醇各浓度组中,随着ip6浓度增高细胞数量减少,差异有统计学意义(p0.05)。2westernblot检测β-catenin蛋白表达:ip6+肌醇高、中剂量组分别为0.886±0.262,0.987±0.707,与对照组1.291±0.431相比,差异有统计学意义(p0.05)。ip6+肌醇低剂量组为1.245±0.580,与对照组相比无明显差异(p0.05)。随着ip6+肌醇剂量的增高,各组蛋白表达量逐渐降低,联合组蛋白表达量为0.601±0.019,顺铂组为0.694±0.039,与对照组相比,差异有统计学意义(p0.05)。且联合组蛋白表达量低于顺铂组,差异有统计学意义(p0.05)。3rt-pcr检测β-cateninmrna的表达:对照组为1.000±0.000,ip6+肌醇低剂量0.984±0.103,两组无明显差异(p0.05),ip6中剂量组、高剂量组、顺铂组、联合组分别为0.906±0.010,0.786±0.104,0.570±0.123,0.488±0.126,与对照组相比,差异均有统计学意义(p0.05),联合用药组表达量低于顺铂组,差异有统计学意义(p0.05)。ip6各剂量组间,随着剂量的增加,表达逐渐减弱,差异有统计学意义(p0.05)。4rt-pcr检测tcf4mrna的表达:ip6+肌醇高剂量组表达量为0.793±0.341,中剂量组为0.853±0.148,低剂量组为0.985±0.133,与对照组1.000±0.000相比,ip6各剂量组表达均降低,但与低剂量组差异不明显(p0.05),与高、中剂量组相比,差异有统计学意义(p0.05)。随着ip6剂量的升高,mrna的表达逐渐减弱。结论:1ip6+肌醇能抑制卵巢癌的侵袭转移。2ip6+肌醇与顺铂具有协同抗癌作用。3ip6+肌醇阻碍卵巢癌侵袭转移的机制可能是影响wnt信号通路,抑制肿瘤的emt,最终抑制其侵袭转移。
[Abstract]:In gynecological malignant tumors, the first mortality is ovarian cancer, histological type of tumor of the skin above accounted for 85%-90%, due to early ovarian cancer specific symptoms, and the lack of means of screening Gao Min perceptual and high specificity and treatment effect, and the method of traditional chemotherapy has many side effects and chemotherapy the problem of drug resistance, the survival rate of ovarian cancer is still very low. The natural medicine is economic, efficient, wide source, low toxicity and other advantages. In recent years, with the inositol six phosphate (IP6) as the representative of the natural medicine plant extraction becomes a hot topic in recent research of.Wnt pathway has been extensively concerned with its anti-tumor effect strong, the regulation of animal tissue differentiation, organ formation and other physiological processes, the abnormal expression of key proteins in the pathway, there may lead to cancer. The beta catenin (beta -catenin) and T cell factor 4 (TCF4) is Key members of Wnt pathway. Studies have shown that ovarian benign disease beta -catenin was lower than that of ovarian malignant disease, and clinical abnormal expression in ovarian cancer and beta -catenin staging, pathological type, metastasis related.TCF4 and beta -catenin binding can promote gene transcription, occurrence and development of many tumors are associated with TCF4. Epithelial mesenchymal transition process (epithelial mesenchymal transition, EMT) is epithelial to mesenchymal transition.EMT is considered the main cause of tumor metastasis.Wnt classic pathway and EMT associated closely, beta -catenin, TCF4 and EMT cancer cells and tumor invasion and metastasis. Objective: This study aimed to investigate the effect of inositol IP6+ on invasion and metastasis of ovarian cancer SKOV3 cells and its mechanism, and the combination of cisplatin and IP6+ inositol on ovarian cancer have synergistic effect. Methods: 1 Tra Nswell tumor cell invasion assay, to observe the effect of.2 Western blot IP6+ inositol dose changes on invasion and metastasis of ovarian cancer cells to detect IP6+ inositol effect on beta -catenin protein.3 RealtimePCR detection of beta -catenin Wnt pathway mRNA, TCF4mRNA expression of.4 statistical methods: SPSS21.0 statistical software with experimental data. Results: 1transwell experimental results show that the combined group invasion cell number was 26.2 + 2.6, cisplatin group cells was 32 + 3.4, 82.8 + group, cisplatin group and control group compared to 5.7 cells significantly reduced the number of significant differences (P0.05), ip6+ high dose group of inositol, cell numbers were 62.6 + 2.3,73.2 + 3. Compared with the control group, the difference was statistically significant (P0.05).Ip6+ inositol low dose group of cells was 80.2 + 3.3, compared with the control group, the difference was not statistically significant (P0.05), ip6+ muscle alcohol each concentration group, with IP6 concentration The degree of increased number of cells decreased, the difference was statistically significant (P0.05) expression of beta -catenin protein.2westernblot: ip6+ inositol high, medium dose group were 0.886 + 0.262,0.987 + 0.707, 1.291 + 0.431 compared with the control group, the difference was statistically significant (P0.05).Ip6+ inositol low dose group was 1.245 + 0.580, compared with the control group no significant difference (P0.05). With the increase of ip6+ inositol dose, each protein expression decreased gradually, the combined group protein expression was 0.601 + 0.019, 0.694 + cisplatin group was 0.039, compared with the control group, the difference was statistically significant (P0.05). The expression of protein is lower than the combined group and cisplatin group, was statistically significant the difference (P0.05) to detect the expression of.3rt-pcr beta -cateninmrna: the control group is 1 + 0, 0.984 + 0.103 ip6+ inositol low dose, no significant difference between the two groups (P0.05), IP6 middle dose group, high dose group, cisplatin group and combination group were 0.906 + 0 .010,0.786 + 0.104,0.570 + 0.123,0.488 + 0.126, compared with the control group, the differences were statistically significant (P0.05), combination group expression was lower than that of cisplatin group, the difference was statistically significant (P0.05.Ip6) between the groups of different dose, with the dose increased, the expression decreased gradually, and there was statistically significant difference (P0.05) to detect the expression of.4rt-pcr tcf4mrna the high dose group of inositol: ip6+ expression level was 0.793 + 0.341, 0.853 + 0.148 in middle dose group, low dose group was 0.985 + 0.133, 1 + 0 compared with the control group, the expression of IP6 in each dose group were decreased, but with the low dose group was not significantly different (P0.05), and high dose. Compared with the group, the difference was statistically significant (P0.05). With the increase of IP6 dose, the expression of mRNA decreased. Conclusion: 1ip6+ inositol can inhibit ovarian cancer invasion and metastasis of.2ip6+ and cisplatin have synergistic anticancer effect of inositol inositol.3ip6+ hinder the invasion and metastasis of ovarian cancer machine It may affect the Wnt signaling pathway, inhibit the tumor's EMT, and eventually inhibit its invasion and metastasis.

【学位授予单位】：河北医科大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R737.31

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