NRG1在肝细胞肝癌中杂合性丢失和蛋白表达的研究

目的：分析在肝细胞性肝癌中NRG1基因的两个多态性位点D8S278和D8S499杂合性丢失（loss ofheterozygosity,LOH）情况及其与NRG1蛋白表达水平之间的关系,探讨NRG1在肝细胞肝癌发生发展中的作用。方法：酚氯仿抽提经石蜡包埋的肝癌和癌旁正常组织DNA,进行PCR扩增,扩增出来的产物进行变性聚丙烯酰胺凝胶电泳并分析两位点杂合性丢失情况;用免疫组织化学方法检测肝癌组织和癌旁正常组织中NRG1的表达情况。结果：D8S278和D8S499的LOH率分别为32.43%（24/74）、37.86%（26/69）,总频率为48.65%（36/74）;NRG1在肝癌组织中的阳性表达率为67.57%（50/74）,显著低于在正常组织中的表达水平（97.30%,72/74）（P〈0.05）,这种降低与NRG1基因多态性位点的LOH有关;NRG1表达下调与肿瘤大小、Edmondson分级相关,而与年龄、性别、HBsAg、AFP、肝硬化和肝内外转移无明显相关。结论：NRG1基因可能是肝细胞肝癌的一个候选抑癌基因,在肝癌发生发展中起一定的作用;可为临床诊断、治疗及预后评估提供理论依据。

Study on the loss of heterozygosity and protein expression of NRG1 in hepatocellular carcinoma

Objective：To analyse the loss of heterozygosity（LOH）of two microsatellites D8S278 and D8S499 of NRG1 in hepatocellular carcinoma（HCC） and the relationship between LOH and NRG1 expression level in HCC tissues,and to investigate the role of NRG1 in the occurrence and development of HCC.Methods：DNA were extracted from paraffin-embedded hepatocellular carcinoma tissues and para-cancer normal tissues using phenol-chloroform.The products of PCR amplification were used to electrophorese in denaturing polyacrylamide gel,then we analyzed the results of LOH at the two microsatellites.Immunohistochemical method was used to detect the expression of NRG1 in 74 cases of HCC tissues and their adjacent tissues.Results：The percentages of LOH at D8S278 and D8S499 were 32.43%（24/74） and 37.86%（26/69） respectively and the total percentage was 48.65%（36/74）.The positive rate of NRG1 protein in HCC was 67.57%（50/74）,significantly lower than in normal tissues（97.30%,72/74）（P0.05）,and the level was inversely correlated with LOH.The down-regulation of NRG1 was associated with tumor size and Edmondson grade,but there was no association with age,gender,HBsAg,AFP,hepatic cirrhosis and intra-and extrahepatic metastasis.Conclusion：NRG1 may be a candidate tumor suppressor gene in HCC,playing an important role in tumorigenesis and development,it can provide theoretical basis for clinical diagnosis,treatment and evaluation of prognosis.