Study on the grade of malignancy and postoperative recurrence in hepatocellular carcinoma with special reference to molecular biological analysis

利用分子生物学方法，我们检测了13q， 16q和17p染色体上的杂合性缺失（LOH），这是人类肝细胞癌（HCC）中经常检测到的。我们还检测了与肿瘤转移相关的抗转移基因nm23H1、粘附分子、细胞粘附调节剂（CAR）和E-cadherin （EC）的基因表达。最后，我们分析了这些因素与临床病理特征和DNA倍体模式的关系。我们的研究结果如下：随着hcc分期的进展，LOH的频率，尤其是16q染色体的频率以及nm23H1、CAR、EC基因的表达减少均增加。CAR和EC基因也存在于16q染色体上，在16q染色体LOH的肿瘤中，其表达减少的频率更高。在含16q染色体的2个及以上LOH， nm23H1、CAR、EC表达减少的肿瘤中，门静脉侵犯、肝内转移、非整倍体、肝内多发复发、肝外转移的频率较高。我们通过分析多结节性HCC的肿瘤LOH模式与复发性HCC切除标本中原发性和复发性肿瘤LOH模式的差异，证实了HCC的多中心发生。这些研究表明，分子生物学分析对于评估肝癌的恶性程度和诊断多中心发生是非常有用的。采用微卫星标记LOH的聚合酶链反应（PCR）方法和基因表达的逆转录酶PCR （RT-PCR）方法，我们可以分析极少量的样本，例如针头活检获得的样本，从而尝试提前评估预后。由于这些基因改变发生在HCC的晚期，因此研究早期的基因改变是很重要的。因此，我们计划在早期研究LOH，以在这些染色体上发现新的抑癌基因，并与遗传性非息肉性结直肠癌中检测到的异常DNA修复基因的关系，这些基因导致HCC的发生和发展。少

Using the molecular biological methods, we examined the loss of heterozygosity (LOH) on chromosomes 13q, 16q and 17p, which are frequently detected in human hepatocellular carcinoma (HCC). We also examined the gene expression of antimetastatic gene, nm23H1, adhesion molecules, cell adhesion regulator (CAR) and E-cadherin (EC) which are associated with cancer metastasis. Finally, we analyzed the relationships between these factors and clinicopathological features and DNA ploidy pattern. Our results were as follows ;1.The frequency of LOH,especially 16q chromosome and reduced expression of nm23H1, CAR,EC gene were increased along with the progression of the Stage of HCC.2.CAR and EC genes also existed on 16q chromosome and their frequency of reduced expression was higher in those tumors with LOH of 16q chromosome.3.In tumors with 2 and more LOH,including 16q chromosome, and reduced expression of nm23H1, CAR,EC,the frequency of portal vein invasion, intrahepatic metasstasis, aneuploid, mu … More ltiple intrahepatic recurrence, and extrahepatic metastasis were high.4.We were able to confirm the multicentric occurence of HCC by analysing the differences of LOH pattern of tumors of multinodular HCC and primary and recurrent tumors in the resected specimens of recurrent HCC.Thsese investigations demonstrated that molecular biological analysis was very useful to evaluate the grade of malignancy and diagnose muticentric occurence in HCC.With polymerase chain reaction (PCR) method with microsatellite marker for LOH and reverse transcriptase PCR (RT-PCR) method for gene expression, we can analyze a very small amount of sample, such as those obtained by needle biopsy, and thus, try to evaluate the prognosis preoeratively. As these genetic alteration occures in the late stage of HCC,it is important to investigate the genetic alteration in the early stage. Thus we plan to study the LOH in the early stage to identify new tumor suppressor genes on these chromosomes, and the relationship with abnormal DNA repair genes, detected in those who had hereditary nonpolyposis colorectal cancer, which causes carcinogenesis and progression of HCC. Less