小肝癌的临床病理及预后相关分析多灶与单灶肝细胞癌外周血白细胞基因表达谱的差异研究

发布时间：2018-02-20 18:48

本文关键词： 肝癌酒精 ALT 预后肝细胞癌多灶肿瘤微环境　出处：《北京协和医学院》2017年博士论文　论文类型：学位论文

【摘要】：该博士学位论文由临床研究及基础研究工作两部分组成。第一部分98例小肝癌的临床病理及预后相关分析[目的]分析小肝癌患者的临床病理特征及影响预后的相关因素。[方法]回顾性分析2009年1月至2013年12月行外科手术R0切除的98例小肝癌患者的临床病理资料,并进行完整随访。采用Kaplan-Meier法计算生存率,Logrank法进行生存曲线比较,多因素分析采用Cox比例风险模型,检验水准α=0.05。[结果]98例患者全部获得随访,98例患者术后1、3、5年生存率分别为99.0%、91.7%和76.3%,中位总生存时间(OS)为52个月。98例患者术后1、3、5年无进展生存率分别为86.7%、66.2%和55.0%,中位无病生存时间(DFS)为43.5个月。单因素分析显示,卫星结节、肝被膜侵犯和术后复发时间与OS有关(均P0.05),长期大量饮酒、卫星结节和肝被膜侵犯与PFS有关(均P0.05)。多因素分析显示,长期大量饮酒是影响小肝癌无病生存时间的独立因素(P=0.003),术后复发时间和肝被膜侵犯是影响小肝癌总生存时间的独立因素(均P0.05)。分组分析影响长期大量饮酒患者及非长期大量饮酒患者的预后因素,对于无长期大量饮酒患者,术前ALT是无病生存时间及总生存时间的显著性不良因素。对于有长期大量饮酒患者,单因素分析显示卫星灶对总生存时间及无病生存时间的影响有统计学差异(P0.05)。多因素分析结果显示:术后复发时间是影响总生存时间的独立危险因素(P0.05)。[结论]对于小肝癌的手术治疗,在保证肿瘤R0切除的前提下,尽量减少正常肝的切除范围对患者有利。小肝癌术后复发时间≤2年提示预后不良,长期大量饮酒能够促进小肝癌术后复发。对于无长期大量饮酒的患者,术前ALT升高能够影响小肝癌术后复发。第二部分多灶与单灶肝细胞肝癌外周血白细胞基因表达谱的差异研究[目的]多灶与单灶肝细胞癌是具有不同临床表现和分子生物学特征的异质性疾病。探索多灶与单灶病变的外周血单个核细胞发生的不同分子事件,分析多灶与单灶肝细胞癌肿瘤微环境的差异。[方法]本实验收集同一治疗组治疗的15例单灶肝癌与9例多灶肝癌患者外周血,采用Agilent 4*44k基因芯片平台,检测外周血单个核细胞的mRNA表达谱,通过生物信息学分析,获得差异表达基因。[结果]通过表达谱芯片分析,在多灶与单灶肝癌外周血单个核细胞中发现592个差异表达基因。主成分分析显示,多灶性肝癌患者与单发性肝癌患者外周血白细胞基因表达谱有明显差异。GO富集分析结果显示,536个上调的差异表达基因涉及的生物学过程包括:细胞因子介导的免疫应答、细胞间的信号传导、损伤修复反应,p38MAPK信号通路的正性调控,免疫系统过程调节以及神经系统和突触的形成等。56个表达下调的差异基因涉及的生物学过程主要包括DNA的转录过程与调控、核糖体的生物合成以及胶原纤维的合成过程等。Pathway分析结果显示,上调的差异表达基因主要与以下通路相关:酪氨酸代谢过程,神经活性配体-受体相互作用,细胞的紧密连接过程,血清素突触以及矿物质的吸收等信号通路相关。样本GSVA分析显示:多灶与单灶肝癌外周血白细胞对比,筛选出有显著性差异的基因集(p0.05)共339个,主要涉及:各种免疫细胞的功能调控(CD4T细胞,树突状细胞,CD8T细胞等),肝癌干细胞的功能,肿瘤微环境的细胞因子功能(TGF-β,IL-1,IL-6,IFN,HIF等),MAPK信号通路的调节,脂类代谢稳态与脂类合成的调节等。[结论]多灶与单灶肝癌患者外周血白细胞基因表达谱存在明显差异,多灶肝癌患者白细胞TGF-β信号通路、脂类合成过程以及HIF等促进肿瘤进展及转移的基因较单灶肝癌患者的表达明显上调,而p38/MAPK信号通路在多灶患者基因表达谱中下调。多灶肝癌较单灶肝癌在肿瘤微环境中存在更多促转移及进展的上调基因。
[Abstract]:This doctoral dissertation by clinical research and basic research work is composed of two parts. The first part of the 98 cases of small hepatocellular carcinoma clinical pathological and prognostic analysis to analyze prognostic factors. Methods: the clinical and pathological features and effects of small liver cancer patients were retrospectively analyzed from January 2009 to December 2013 underwent surgical resection of 98 cases of small hepatocellular carcinoma R0 the clinicopathological data of patients, and complete follow-up. The survival rate was calculated by Kaplan-Meier method, Logrank method to compare the survival curves, multivariate analysis using Cox proportional hazard model, a =0.05.[level test results]98 patients were followed up, 98 cases of patients with postoperative 1,3,5 year survival rates were 99%, 91.7% and 76.3%. The median survival time was 52 months (OS) of.98 patients after 1,3,5 years of progression free survival rates were 86.7%, 66.2% and 55%, the median disease-free survival time (DFS) for 43.5 months for single. Analysis showed that the satellite nodules, liver capsule invasion and recurrence time associated with OS (P0.05), a large number of long-term drinking, satellite nodules and liver capsule invasion associated with PFS (P0.05). Multivariate analysis showed that a large number of long-term drinking are independent factors affecting small liver disease free survival (P=0.003), postoperative recurrence time and liver membrane invasion were independent factors affecting the survival of small hepatocellular carcinoma (P0.05). Group analysis of long-term heavy drinking and non drinking a lot of patients with long-term prognostic factors for patients with long-term drinking, for patients, preoperative ALT is significant adverse factors of disease free survival time and total the survival time for patients with long-term drinking, the single factor analysis showed that the satellite focus on overall survival time and disease-free survival time was statistically different (P0.05). The results of multivariate analysis showed that postoperative recurrence time is the total effect Independent risk factors for the survival time (P0.05). Conclusion] for surgical treatment of small hepatocellular carcinoma, tumor resection under the premise of ensuring R0, try to reduce the resection range of normal liver is beneficial in patients with small hepatocellular carcinoma. Postoperative recurrence time is less than 2 years of poor prognosis, long-term drinking can promote the postoperative recurrence of small hepatocellular carcinoma. For a large number of long-term drinking in patients with elevated preoperative ALT can influence the recurrence of small hepatocellular carcinoma after operation. The second part multifocal hepatocellular carcinoma and single lesion of peripheral white blood cell differential gene expression study [Objective] multifocal spectrum and single lesion of hepatocellular carcinoma is a heterogeneous disease with different clinical manifestations and characteristics of molecular biology the different molecular events. Explore multifocal and single focal lesion of the peripheral blood mononuclear cells of the tumor and the difference analysis of multifocal single focal hepatocellular carcinoma microenvironment. Methods: We collected 15 cases with single lesion treatment in treatment group 9 cases of multifocal hepatocellular carcinoma and peripheral blood of patients with hepatocellular carcinoma, using Agilent 4*44k microarray platform, detection of peripheral blood mononuclear cells mRNA expression, bioinformatics analysis of differentially expressed genes. The results obtained by microarray expression analysis, in multifocal and single focal liver cancer in peripheral blood mononuclear cells found 592 differentially expressed genes. Principal component analysis showed that the multifocal hepatocellular carcinoma patients with solitary white blood cell gene expression profile of peripheral blood of patients with hepatocellular carcinoma were significantly different.GO enrichment analysis showed that the genes involved in the biological process including 536 up-regulated expression of immune response mediated by cytokines, cellular signal transduction the injury repair response, positive regulation of p38MAPK signaling pathway, immune system regulation and nervous system formation and synaptic.56 expression of genes down regulated biological process differences include The process of transcription and regulation of DNA, and ribosome biosynthesis of collagen synthesis process of.Pathway analysis showed that the upregulation of differentially expressed genes mainly with the following pathway: tyrosine metabolism, neuroactive ligand receptor interaction, is closely connected to the process of cell, serotonin synapses and mineral absorption and sample GSVA related signal pathway. Analysis showed that: compared to white cells in the peripheral blood of multifocal and single focal liver cancer, screening significant gene sets (P0.05) a total of 339, mainly involves: the functional regulation of various immune cells (CD4T cells, dendritic cells, CD8T cells), liver cancer stem cell function, cytokine function of tumor the environment (IL-1, IL-6, TGF- beta, IFN, HIF), MAPK pathway, lipid metabolism homeostasis and lipid synthesis regulation. Conclusion] multifocal hepatocellular carcinoma patients with single lesion of peripheral white blood cells There are obvious differences in gene expression spectrum, multifocal white blood cells in patients with hepatocellular carcinoma TGF- beta signaling pathway, lipid synthesis process and promote HIF expression was significantly increased in tumor progression and metastasis genes than a single lesion in patients with hepatocellular carcinoma, and p38/MAPK signaling pathway in patients with multifocal gene expression. Lower multifocal hepatocellular carcinoma than single focal hepatocellular carcinoma was up-regulated more progress and promoting gene transfer in the tumor microenvironment.

【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】：2017
【分类号】：R735.7

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