PI3K-Akt信号通路在肝母细胞瘤的作用及肝母细胞瘤预后危险因素的评估

发布时间：2018-05-10 04:39

本文选题：PI3K-Akt通路 + 差异表达基因　；参考：《北京协和医学院》2015年博士论文

【摘要】：第一部分：PI3K-Akt信号通路在肝母细胞瘤的作用研究背景肝母细胞瘤(hepatoblastoma, HB)是儿童最常见的肝脏恶性肿瘤。近20年来由于肝肿瘤切除手术和化疗的优化结合,术前术后化疗方案的个体化措施的发展,肝母细胞瘤患儿的预后有了很大的改善。PI3K-Akt通路是一个经典的抗凋亡、促存活的信号转导途径,在肿瘤发生、增殖、侵袭、转移中发挥重要作用。研究目的本研究通过分析PI3K, AKT在肝母细胞瘤细胞系及其肿瘤和癌旁组织的表达情况,了解PI3K-Akt通路在肝母细胞瘤中的重要意义,为后续开发通过抑制该通路治疗肝母细胞瘤的靶向药物提供理论依据。研究方法我们通过差异基因表达筛查发现PI3K-Akt通路的相关因子包括PI3K,Akt,PKC,PKB及GSK-3等在肿瘤组织中特异性的高表达。利用MTT法测定LY294002作用前后肝母细胞瘤细胞系HuH-6和HepG2的增殖；通过Western blot分析PI3K通路抑制剂LY294002对肝母细胞瘤细胞系中PI3K、Akt表达的影响；RT-PCR检测PI3KmRNA在肿瘤和癌旁组织的表达差异；Western blot检测PI3K、p-Akt(Ser473)以及总的Akt蛋白在肿瘤和癌旁组织的差异。结果本研究中,我们发现HuH-6和HepG2细胞对不同浓度的PI3K抑制剂LY294002处理具有剂量依懒性增殖抑制；同时,随着PI3K通路抑制剂LY294002浓度的增加,PI3K、p-Akt(Ser473)蛋白在HuH-6和HepG2细胞中的表达逐渐下降,且各组之间的差异存在统计学差异(p0.001)；PI3K mRNA在肿瘤组织中的表达显著高于癌旁组织；PI3K和p-Akt(Ser473)蛋白在肿瘤组织的表达显著高于癌旁组织。而总的Akt和β-actin在两组病人中无显著差别。结论PI3K-Akt通路的激活可致使肝母细胞瘤肿瘤细胞增殖,相关基因在肿瘤组织的表达显著高于癌旁组织,提示PI3K-Akt通路在肝母细胞瘤的发生发展中起到了重要的作用。第二部分肝母细胞瘤预后危险因素的评估研究背景肝母细胞瘤(hepatoblastoma, HB)是儿童最常见的肝脏恶性肿瘤,化疗敏感的患儿术后具有较高的总体生存率(3年生存率在75%以上)。尽管如此,肝母细胞瘤患儿的预后仍存在较大差异,因为多个危险因素与其预后密切相关。研究目的本研究的目的是确定与亚洲人群肝母细胞瘤患儿预后相关的危险因素,并分析影响患儿无事件生存(Event-Free Survival, EFS)的各个危险因素之间的相互关系。研究方法回顾性分析首都儿科研究所附属儿童医院2001年1月至2014年9月行肝肿瘤切除术的肝母细胞瘤患儿共176例,采用Kaplan-Meier生存曲线分析和Cox比例风险模型评估与肝母细胞瘤相关的危险因素。结果所有患儿(176例)的中位无事件生存时间为80.4个月(95% CI：71.6-89.2)及5年无事件生存率(EFS)54.6%,五年总体生存率(OS)66.7%。肝母细胞瘤患儿预后与AFP水平、肿瘤单发多发、PRETEXT分期、有无转移、病理分型有关。多因素分析结果显示：AFP水平(HR：2.567,P=0.014),肿瘤是否多发(HR：2.187,P=0.028),肿瘤是否转移(HR：1.912,P=0.028)以及PRETEXT IV期(HR：2.254,P=0.005)均是影响肝母细胞瘤患儿EFS的独立为危险因素。不同的病理分型分组相互影响,分别引入多因素Cox模型中进行分析提示,完全胎儿型(HR：2.752,P=0.021)是影响肝母细胞瘤患儿EFS的独立为危险因素。结论肝母细胞瘤患儿预后与AFP水平、肿瘤单发多发、PRETEXT分期、有无转移、病理分型有关。肿瘤的病理分型与其他各因素之间存在着相互作用,不同危险因素的存在进一步影响病理分型对于预后的预测。
[Abstract]:The first part: the role of PI3K-Akt signaling pathway in hepatblastoma, background hepatoblastoma (HB) is the most common liver malignant tumor in children. In the last 20 years, the combination of hepatoma resection and chemotherapy, the development of preoperative and postoperative chemotherapy regimens, and the prognosis of the children with hepatblastoma A great improvement in the.PI3K-Akt pathway is a classic anti apoptotic and survival signal transduction pathway that plays an important role in tumor genesis, proliferation, invasion and metastasis. The purpose of this study was to analyze the expression of PI3K, AKT in the hepatblastoma cell lines and their tumor and paracancerous groups, and to understand the PI3K-Akt pathway in the hepatocytes. The important significance of the tumor is to provide a theoretical basis for subsequent development of targeted drugs for the treatment of hepatblastoma by inhibiting this pathway. We found that the related factors of PI3K-Akt pathway include PI3K, Akt, PKC, PKB and GSK-3 in specific high expression in tumor tissues by differential gene expression screening. The determination of LY294002 by MTT method The proliferation of hepatblastoma cell lines HuH-6 and HepG2, and the effect of LY294002 on the expression of PI3K and Akt in the hepatblastoma cell lines by Western blot; RT-PCR detected by RT-PCR to detect the difference in the expression of PI3KmRNA in the tumor and adjacent tissues; Western blot, and the total protein in tumor and cancer In this study, we found that HuH-6 and HepG2 cells have a dose dependent inhibition of laziness in different concentrations of PI3K inhibitor LY294002; meanwhile, the expression of PI3K, p-Akt (Ser473) protein in HuH-6 and HepG2 cells gradually decreases with the increase of LY294002 concentration in the PI3K pathway, and the difference between each group is poor. The difference was statistically significant (p0.001), and the expression of PI3K mRNA in the tumor tissues was significantly higher than that in the para cancerous tissue, and the expression of PI3K and p-Akt (Ser473) protein in the tumor tissues was significantly higher than that in the paracancerous tissues. The total Akt and beta -actin were not significantly different in the two groups. Conclusion the activation of PI3K-Akt path can lead to the proliferation of hepatblastoma tumor cells. The expression of related genes in tumor tissues is significantly higher than that of paracancerous tissue, suggesting that PI3K-Akt pathway plays an important role in the development of hepatblastoma. Second the evaluation of prognostic risk factors in partial hepatblastoma (hepatoblastoma, HB) is the most common malignant tumor of the liver in children, chemotherapy sensitivity A higher overall survival rate (3 year survival rate is above 75%) after surgery. However, there are still significant differences in the prognosis of children with hepatblastoma, because multiple risk factors are closely related to their prognosis. The purpose of this study is to determine the risk factors associated with the prognosis of children with Asians with hepatblastoma, and the purpose of this study is to determine the risk factors for the prognosis of the children with hepatblastoma. A retrospective analysis of 176 children with hepatoblastoma in children's Hospital Affiliated to the capital Pediatrics Institute from January 2001 to September 2014, using Kaplan-Meier survival curve analysis and Cox proportions risk, was analyzed in a retrospective analysis of the relationship between the various risk factors affecting Event-Free Survival (EFS). The model assessed the risk factors associated with hepatblastoma. Results the median survival time of all children (176 cases) was 80.4 months (95% CI:71.6-89.2) and 5 years of event free survival (EFS) 54.6%. The prognosis of the five year total survival rate (OS) 66.7%. hepatblastoma was associated with the AFP level, the multiple tumors, the PRETEXT staging, the metastasis, and the disease. The results of multi factor analysis showed that AFP level (HR:2.567, P=0.014), tumor multiple (HR:2.187, P=0.028), tumor metastasis (HR:1.912, P=0.028), and PRETEXT IV phase (HR:2.254, P=0.005) were all risk factors affecting the independence of EFS in children with hepatblastoma. Different pathological classification groups influenced each other, respectively. In the multi factor Cox model, it is suggested that complete fetal type (HR:2.752, P=0.021) is an independent risk factor affecting EFS in children with hepatblastoma. Conclusion the prognosis of the children with hepatblastoma is related to the level of AFP, the multiple tumors, the PRETEXT staging, the metastasis and the pathological classification. The pathological classification of the tumor is associated with the other factors. The presence of interaction and different risk factors further affect the prognosis of pathological classification.

【学位授予单位】：北京协和医学院
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R735.7

【共引文献】