【作者】 侯振宇；

【导师】 周伟平； 吴孟超；

【作者基本信息】 第二军医大学 ， 外科学， 2008， 硕士

【摘要】 XAGE-1是Brinkmann和Vasmatzis等人于1999年利用同源漫步(homologouswalking)的生物信息学方法在人类EST库中搜索GAGE-PAGE的近缘基因时发现的一类新基因,定位于Xp11.21-Xp11.22,限制性地表达于健康睾丸组织及多种肿瘤中,是典型的CT-X抗原。XAGE-1的表达剪接体有4种,最主要的是XAGE-1b。临床研究已获得了较为详细的XAGE-1b在肿瘤中的表达谱,发现在肺癌、黑色素瘤、前列腺癌等癌症组织中有较高的XAGE-1b检出率,并且逐步揭示出XAGE-1b表达与病理进程和肿瘤亚型间的关联。本研究通过检测XAGE-1b在原发性肝癌(PHC)人群、肝炎肝硬化人群、良性肝病人群、健康人群中的表达,了解其是否可作为PHC诊断及预后判断的指标。目的:评价XAGE-1b作为PHC辅助诊断指标的可行性。方法:采用Real-time RT-PCR的方法定量检测125例PHC、23例肝炎肝硬化、34例良性肝病、41例健康志愿者血液中XAGE-1b mRNA表达,探讨其表达与PHC患者性别、年龄、肿瘤大小、病理类型、临床分期、AFP的关系,比较其在不同人群中表达差异,并用ROC曲线比较XAGE-1b mRNA、AFP、CEA、CA199诊断PHC的价值;进一步检测18例PHC癌组织、癌旁组织XAGE-1bmRNA表达水平并比较其表达差异,采用免疫组化S-P法对100例PHC癌组织和癌旁组织进行染色,研究XAGE-1b蛋白表达。结果:125例PHC患者血液XAGE-1b mRNA检出率为80.8%。其中男女患者检出率分别为80.58%,81.82%(P＞0.05),各年龄组(20—39岁、40—59岁、60—79岁)检出率分别为90.0%,77.9%,82.1%(P＞0.05)。血液XAGE-1b mRNA的表达水平在大肝癌组(肿瘤直径＞5cm)和小肝癌组(肿瘤直径≤5cm),肝细胞癌组和胆管细胞癌组,临床Ⅰ、Ⅱ、Ⅲ、Ⅳ期组,AFP阳性组和阴性组差异无统计学意义(P＞0.05);但肝细胞癌患者血液XAGE-1b mRNA的阳性率明显高于AFP~*(P＜0.01),尤其对于AFP阴性肝细胞癌其检出率为82.35%,XAGE-1bmRNA与AFP不具有相关性(95%可信区间为[-0.21,0.10])。血液XAGE-1bmRNA在PHC患者中的表达水平高于肝炎肝硬化、良性肝病、健康人群(P＜0.01),在肝炎肝硬化患者中的表达水平高于良性肝病和健康人群(P＜0.05),而在良性肝病和健康人群的表达差异无统计学意义(P＞0.05);ROC曲线评价XAGE-1b mRNA、AFP、CEA、CA199对PHC的诊断价值,其AUC值分别为0.879、0.759、0.648、0.511,XAGE-1b mRNA具有较高的诊断准确性,其次为AFP,而CEA和CA199诊断价值较低。选取的18例PHC癌组织XAGE-1b mRNA表达水平高于癌旁组织和血液(P＜0.01),而癌旁组织表达水平高于血液(P＜0.01)。免疫组化研究100例PHC患者XAGE-1b蛋白水平表达,在1例Ⅳ期患者中,转移癌组织的表达强度明显高于原位肿瘤,其他均未见表达。结论:XAGE-1b mRNA的表达与PHC患者性别、年龄、肿瘤大小、病理类型、临床分期、AFP无关,且对良性肝病、肝炎肝硬化、健康人群具有鉴别意义。XAGE-1b mRNA可以预测血液中肿瘤细胞的微转移并作为一种肿瘤标志物辅助诊断PHC,尤其对AFP阴性的肝细胞癌更显示出诊断价值,对PHC预后判断及治疗方案的选择可能有重要意义。更多还原

【Abstract】 XAGE-1 is a new gene discovered by Brinkmann,Vasmatzis and others in 1999 by means of homologous walking,a kind of bioinformatic method.When they searched human expressed sequence tags library near MAGE-PAGE,it was found and located in Xp11.21-Xp11.22.Further study shows that it is typical CT-X antigen expressed in testis and many kinds of tumor restrictively,and it expresses four spliceosomes among which XAGE-1b is main expression product.Clinical research has get detailed expression spectra of XAGE-1b.It is expressed with high percentage in lung cancer,melanoma,prostatic cancer and so on.Furthermore, relationship is gradually being revealed between XAGE-1b expression and clinical factors such as patho-proceeding and tumor hypotype.This study focuses on XAGE-1b expression in healthy individuals and patients with primary hepatic carcinoma(PHC),benign hepatic disease and liver cirrhosis caused by hepatitis,so as to investigate XAGE-1b as a marker of PHC for diagnosis and prognosis judgement.Objective:To value the feasibility of XAGE-1b as auxiliary diagnosis for PHC.Methods:Peripheral blood from 125 patients with PHC,34 patients with benign hepatic disease,23 patients with liver cirrhosis caused by hepatitis and 41 healthy individuals was collected,and XAGE-1b mRNA was detected by quantitative real-time reverse transcription polymerase chain reaction(RT-PCR).Relationship between XAGE-1b mRNA expression and clinical factors such as sex,age,tumor size, pathologic type,clinical stage and AFP was investigated in PHC patients and expression diversity in healthy individuals and different patients was compared.ROC curve was used to analyse the results of XAGE-1b mRNA,AFP,CEA and CA199 to compare their diagnostic value for PHC.Furthermore,XAGE-1b mRNA was detected in tumor tissue and their adjacent non-tumor tissue from 18 PHC patients to compare expression diversity and 100 cases of tumor tissue and their adjacent non-tumor tissue were stained by S-P immunohistochemistry to investigate XAGE-1b protein. Results:XAGE-1b mRNA was detectable in blood from 80.8%of PHC patients. Among them,80.58%of male patients and 81.82%of female patients were detectable (P＞0.05),and detectable rate of different age group(20-39,40-59,60-79 years old group) was 90.0%,77.9%and 82.1%respectively(P＞0.05).Levels of XAGE-1b mRNA in blood were not obviously different between big hepatic carcinoma(tumor diameter＞5cm) and small hepatic carcinoma(tumor diameter≤5cm),hepatocellular carcinoma and cholangiocellular carcinoma,clinical stageⅠ、Ⅱ、ⅢandⅣ,patients with AFP positive and the AFP negative(P＞0.05),but the positive rate of XAGE-1b mRNA in blood from patients with hepatocellular carcinoma was obviously higher than that of AFP(P＜0.01).Especially,82.35%of PHC patients with AFP negative was found XAGE-1b mRNA in blood and there was no dependablity between XAGE-1b mRNA and AFP(95%confidence interval from -0.21 to 0.10.Levels of XAGE-1b mRNA in blood from PHC patients were higher than from other patients and healthy individuals(P＜0.01),and from liver cirrhosis patients were higher than from patients with benign hepatic disease and healthy individuals(P＜0.05),yet from patients with benign hepatic disease and healthy individuals were similar(P＞0.05).ROC curve judged the diagnostic value of XAGE-1b mRNA,AFP,CEA and CA199 for PHC,the AUC was 0.879,0.759,0.648 and 0.511 respectively.XAGE-1b mRNA was more accuracy than others and AFP was the second,but CEA and Ca199 had little diagnostic value.Levels of XAGE-1b mRNA in tumor tissue from 18 PHC patients were higher than in their adjacent non-tumor tissue and peripheral blood(P＜0.01),and in adjacent non-tumor tissue were higher than in peripheral blood(P＜0.01).Protein of XAGE-1b was detected in tumor tissue and adjacent non-tumor tissue from 100 PHC patients by immunohistochemistry,and it was more increased in distant metastasis than in primary tumor,which was observed in one case,but other cases were negative.Conclusion:The expression of XAGE-1b mRNA in blood from PHC patients doesn’t associate with sex,age,tumor size,pathological category,clinical stage and AFP.Through XAGE-1b mRNA,PHC patients can be discriminated from healthy individuals and patients with benign hepatic disease or liver cirrhosis caused by hepatitis.It can predict tumor cell micrometastasis in blood and contribute to diagnose PHC as a tumor marker,especially has diagnostic value for hepatocellular carcinoma with AFP negative.It may have considerable significance for prognostic judgment and treatment selection for PHC.更多还原