【作者】 于少华；

【导师】 高鹏； 张慧；

【作者基本信息】 山东大学 ， 临床病理学（专业学位）， 2024， 硕士

【摘要】 目的:特殊肝细胞癌(Hepatocellular carcinoma,HCC)的临床病理特征尚未得到充分地揭示。我们的目的是研究特殊亚型HCC的临床病理特征及预后差异,分析不同临床病理参数对HCC预后的影响。为了加深对该肿瘤的认识,我们对特殊病例进行了全面的组织学、免疫组化及分子特征的分析。方法:根据纳入和排除标准,收集2020年1月1日—2022年月1日山东大学齐鲁医院行手术切除的所有原发性HCC样本,共485例。回顾患者所有可用组织切片,重新进行组织学分型。综合分析485例HCC的临床参数及病理特征,临床参数包括性别、年龄、病毒感染状态、饮酒史,是否存在背景肝组织的肝硬化及脂肪变性情况,病理特征包括不同亚型HCC的肿瘤大小、ES分级、MVI分级、卫星结节形成及TNM分期情况。评估血清肿瘤标志物AFP、CEA、CA125、CA199与不同亚型HCC的相关性。以肿瘤内中性粒细胞数目大于100/10HPF(×400)为评价标准,评估中性粒细胞浸润预后的关系。评估血清肿瘤标志物AFP、CEA、CA125、CA199,不同的临床病理特征及不同亚型与预后的关系。为进一步探究肉瘤样HCC的分子特征,我们对5例肉瘤样HCC进行显微分割,对肉瘤样区域和上皮样区域分别进行DNA靶向测序及全转录组测序。此外,我们采用全外显子测序检测了一例肝OGCT的分子遗传特征,并对已报道病例的临床病理特征进行全面的回顾。结果:1、本研究共纳入485例原发性HCC患者,患者年龄在26岁-84岁,中位年龄56岁,男女比例为4.6:1。80%以上患者伴有乙肝病毒感染及肝硬化。肿瘤TNM分期以Ⅰ期、Ⅱ期为主,组织学分级主要为Ⅱ-Ⅲ级。2、根据阅片结果,HCC病例被分类为粗梁型HCC 90例、脂肪性肝炎型HCC30例、硬化型HCC21例、富于淋巴细胞型HCC7例、嫌色细胞型HCC 7例及富于中性粒细胞型HCC 2例。透明细胞型以50%诊断标准共确定22例,以80%诊断标准确定7例。同时,未被纳入2019版WHO的肉瘤样型HCC共5例、泡沫细胞型HCC共2例。3、约1/3的HCC可以归类为不同的亚型。粗梁型HCC在临床及组织学方面均呈现出明显的预后不良的特征。脂肪性肝炎型HCC的肿瘤体积更小,血管侵犯更少,可能提示预后更佳。硬化型HCC更容易出现微血管及门静脉的侵犯。肉瘤样HCC未显示出独特的临床病理特征,但预后较普通型HCC更差。以50%及80%为标准诊断的透明细胞型HCC均未表现出明显的临床病理特征的相关性。4、实验室检查结果分析显示,粗梁型HCC与AFP升高具有一定关联性,脂肪性肝炎型HCC与AFP升高者明显少于普通型HCC。硬化型HCC患者AFP升高程度明显高于普通型HCC。5、生存分析显示,AFP升高、ES分级、MVI分级、卫星结节形成数量、原发肿瘤T分期、M分期、TNM分期及中性粒细胞浸润均为影响预后的因素。肝硬化患者与非肝硬化患者的预后无明显差异。多因素COX比例风险模型分析,结果显示原发性肿瘤T分期及TNM分期是影响预后的独立预测因子。6、对于目前WHO分类中未提及的罕见的肉瘤样HCC及肝脏OGCT,我们进行了高通量基因检测,结果显示:肉瘤样HCC的肉瘤样分化区与上皮样区之间的DNA突变特征无明显差异,RNA表达水平的调控导致了该亚型组织学形态的转换,而肝OGCT的分子改变与HCC存在显著差异,可能是一种独立于HCC的组织学类型。根据基因组测序结果分析,APC和RNF43基因及其负调控的Wnt信号通路可能在肝脏OGCT的发生发展中发挥重要作用。结论:1、一些HCC亚型表现出特定的临床病理特征,粗梁型HCC呈现出明显的预后不良的特征,脂肪性肝炎型HCC肿瘤体积更小,血管侵犯更少,可能提示预后更佳。硬化型HCC更容易出现微血管及门静脉的侵犯。在2019版WHO分类之外,肉瘤样型HCC也显示出更差的预后特征。2、AFP升高、ES分级、MVI分级、卫星结节形成数量、原发肿瘤T分期、M分期、TNM分期及中性粒细胞浸润均为影响预后的因素。其中,原发性肿瘤T分期及TNM分期是影响预后的独立预测因子。3、肉瘤样HCC中的肉瘤样分化区与上皮样区的DNA特征无明显差异。RNA表达水平的调控导致了该亚型组织学形态的转换,继发性的突变导致了该亚型的预后不良。4、肝OGCT是一类不同于HCC的肿瘤,APC和RNF43基因及其负调控的Wnt信号通路可能在肝脏OGCT的发生发展中发挥重要作用。更多还原

【Abstract】 Objective:There are significant differences in the risk factors and clinicopathological features of Hepatocellular carcinoma(HCC)in different regions.At the same time,the clinicopathological characteristics of different HCC subtypes have not been fully revealed.Our objective was to study the clinicopathological features and prognostic differences of HCC subtypes and analyze the influence of different clinicopathological parameters on HCC prognosis.In order to deepen the understanding of this tumor,we performed a comprehensive histological,immunohistochemical and molecular analysis of special cases.Methods:Per the specified criteria for inclusion and exclusion,every primary HCC specimen was subjected to surgical removal at Shandong University’s Qilu Hospital between January 1,2020,and January 1,2022,amounting to a total of 485 cases.Every accessible tissue sample from the patient underwent a thorough examination and subsequent tissue typing.A detailed analysis was conducted on the clinical and pathological characteristics of 485 HCC patients,encompassing their gender,age,viral infection status,history of alcohol intake,and any existing liver cirrhosis and steatosis.The pathological characteristics encompassed the size of the tumor,ES grade,MVI grade,formation of satellite nodules,and TNM stage in various HCC subtypes.Assessing the relationship between serum tumor indicators AFP,CEA,CA125,CA199,and various HCC subtypes.Neutrophil count in the tumor exceeded 100/10HPF(×400).An assessment was conducted on the interplay among serum tumor indicators AFP,CEA,CA125,CA199,various clinicopathological characteristics,and diverse subtypes and prognoses.To further explore the molecular characteristics of sarcomatoid HCC,we performed micro-segmentation of 5 cases of sarcomatoid HCC,and performed DNA targeted sequencing and full transcriptome sequencing of sarcomatoid and epithelioid regions respectively.Furthermore,whole genome sequencing identified the molecular genetic characteristics of a single hepatic OGCT case,and a thorough examination of the clinicopathological aspects of the cases was conducted.Result:1.This research encompassed 485 individuals suffering from primary HCC,aged between 26 and 84,with an average age of 56,and a gender ratio of 4.6 males to 1 female.Over 80%of the patients are afflicted with hepatitis B virus infection and cirrhosis.The tumor’s TNM phase predominantly consisted of stage I and II,while its histological classification was largely between grade II and III.2.The findings indicate that HCC instances were categorized into 90 macrotrabecular-massive HCC,30 steatohepatitis HCC,21 sclerosis HCC,7 lymphocytic HCC,7 chromophobe HCC,and 2 neutrophilic HCC.Out of the total,22 instances of clear cell types were pinpointed using 50%of the diagnostic standards,and 7 instances were recognized with 80%of the diagnostic criteria.Concurrently,the WHO 2019 edition excluded 5 instances of sarcomatoid HCC and 2 instances of foam cell HCC.3.The predominant organizational form of HCC is the common type,constituting roughly two-thirds.Approximately a third of HCC cases fall into various subtypes.Macrotrabecular-massive HCC exhibits a more aggressive nature and a poorer outlook compared to typical HCC,in both clinical and histological terms.The reduced size of steatohepatitis HCC tumors and lesser vascular penetration could indicate a more favorable outlook.Sclerosis HCC tends to be more susceptible to invasion into micro-vessels and portal veins.While sarcomatoid HCC lacks distinct clinicopathological characteristics,its outlook is more unfavorable compared to traditional HCC.No notable clinical-pathological link was found between 50%and 80%clear cell HCC.4.Laboratory analysis revealed a notable link between macrotrabecular-massive HCC and elevated AFP levels,with the rise in AFP in steatohepatitis HCC being considerably lower compared to regular HCC.Notably,sclerosis HCC patients exhibited a much greater AFP increase than in typical HCC patients.5.The prognosis was impacted by various factors including AFP levels,ES grades,MVI grades,satellite nodule formation count,primary tumor T stages,M stages,TNM stages,and neutrophil infiltration,as revealed by survival studies.The prognosis remained largely unchanged between cirrhosis patients and non-cirrhosis patients.Analysis using the multivariate COX proportional risk model revealed that T stage and TNM stage independently forecasted the prognosis.6.For rare sarcomatoid HCC and hepatic OGCT not currently mentioned in the WHO classification,we performed high-throughput genetic testing,and the results showed that:there was no significant difference in DNA mutation characteristics between sarcomatoid and epithelioid regions of sarcomatoid HCC,and the regulation of RNA expression level led to the histological transformation of this subtype,while the molecular changes of hepatic OGCT were significantly different from those of HCC,which may be an independent histological type of HCC.According to the analysis of genome sequencing results,APC and RNF43 genes and their negatively regulated Wnt signaling pathway may play an important role in the occurrence and development of liver OGCT.Conclusion:1.Certain subtypes of HCC display distinct clinicopathological characteristics,with macrotrabecular-massive HCC being notably more aggressive and carrying a poorer prognosis.A reduced tumor dimension and lesser vascular invasion in steatohepatitis HCC could indicate a more favorable outlook.Sclerosis HCC tends to be more susceptible to invasion into micro-vessels and portal veins.Beyond the scope of the 2019 WHO categorization,sarcomatoid HCC exhibited a poorer predictive outlook.2.Survival analysis showed that AFP elevation,ES grade,MVI grade,number of satellite nodules forming,primary tumor T stage,M stage,TNM stage,and neutrophil infiltration were all prognostic factors.Independently,the T stage and TNM stage in the primary tumor served as prognostic factors.3.There was no significant difference in DNA characteristics between sarcomatoid differentiation and epithelioid differentiation in sarcomatoid HCC.Regulation of RNA expression level leads to the transformation of histological morphology of this subtype,and secondary mutations lead to poor prognosis of this subtype.4.Hepatic OGCT represents a type of tumor that differs from HCC.The APC and RNF43 genes,along with their negatively controlled Wnt signaling route,could be crucial in the emergence and progression of hepatic OGCT.更多还原