Kim Dae-Ghon
Division of GI and Hepatology, Department of Internal Medicine, Chonbuk National University Medical School and Hospital, Jeonbuk, Republic of Korea.
Methods Mol Biol. 2006;317:141-55. doi: 10.1385/1-59259-968-0:141.
HBx and MHBst products from hepatitis B virus-DNA (HBV-DNA), which become transcriptional transactivators of cellular and viral genes, are known to play causative roles in the development of hepatocellular carcinoma (HCC). However, the biomolecular mechanism(s) for their roles in hepatocarcinogenesis in vivo remain poorly understood. To identify authentic cellular genes involved in HBx and MHBst-transactivated carcinogenesis,we used mRNA differential display polymerase chain reaction (DD-PCR). We examined HBx and MHBs-positive or -negative HCC, which had chromosomally integrated HBV DNA, vs nontumor tissues, respectively, and differentially expressed genes in either type of HCC were identified and compared with each other. Using 240 different combinations of three one-base anchored oligo-dT primers and 80 arbitrary 13mers, 16 genes were differentially expressed in the HBx and MHBs-positive HCC including RoRNA hY1, glutamine synthetase, factor H homologue 3' end, voltage-dependent anionc hannel 3 (VDAC3), three ribosomal proteins, four mitochondrial genes, and four novel genes. Unexpectedly, upregulated genes in association with functional HBV proteins were different from those reportedly transactivated by HBV viral proteins in vitro. Ten genes were downregulated, including three novel genes. In contrast, 15 genes in HCC tissue negative for HBx and MHBs-expression were preferentially expressed including pancreatic secretory trypsin inhibitor (PSTI), H19, guanidine nucleotide-binding protein alpha-1 subunit (GNAZ), carbamyl phosphate synthetase I (CPS I), insulin-like growth factor (IGF)-II, and 10 ribosomal proteins genes. Eighteen genes were downregulated including acute phase genes, a novel gene, and particularly the retinoblastoma susceptibility gene. Only two genes (ribosomal protein P0 and L37a) were commonly upregulated in both types of HCC tissues. These results suggest that cellular genes involved in the viral protein-transactivation may generally differ from those not associated with transactivation in established HCC, and that the specific oncogenic coordination through the transactivation by viral proteins which works in experiments in vitro, may play only a potential role in hepatocarcinogenesis in vivo. In addition, the functional analyses of the eight novel genes identified in this study might be valuable to further understand the mechanism(s) of hepatocarcinogenesis.
乙型肝炎病毒DNA（HBV-DNA）产生的HBx和中分子乙肝表面抗原（MHBst）产物可成为细胞和病毒基因的转录激活因子，已知它们在肝细胞癌（HCC）的发生发展中起致病作用。然而，它们在体内肝癌发生过程中作用的生物分子机制仍知之甚少。为了鉴定参与HBx和MHBst激活致癌作用的真实细胞基因，我们使用了mRNA差异显示聚合酶链反应（DD-PCR）。我们分别检测了染色体整合有HBV DNA的HBx和MHBs阳性或阴性HCC与非肿瘤组织，并鉴定了两种HCC中差异表达的基因并进行相互比较。使用三种单碱基锚定寡聚dT引物和80种任意的13聚体的240种不同组合，在HBx和MHBs阳性HCC中有16个基因差异表达，包括RoRNA hY1、谷氨酰胺合成酶、因子H同源物3'端、电压依赖性阴离子通道3（VDAC3）、三种核糖体蛋白、四个线粒体基因和四个新基因。出乎意料的是，与功能性HBV蛋白相关的上调基因与据报道在体外被HBV病毒蛋白激活的基因不同。有10个基因下调，包括三个新基因。相比之下，在HBx和MHBs表达阴性的HCC组织中有15个基因优先表达，包括胰腺分泌性胰蛋白酶抑制剂（PSTI）、H19、鸟苷酸结合蛋白α-1亚基（GNAZ）、氨基甲酰磷酸合成酶I（CPS I）、胰岛素样生长因子（IGF）-II和10个核糖体蛋白基因。有18个基因下调，包括急性期基因、一个新基因，特别是视网膜母细胞瘤易感基因。在两种HCC组织中只有两个基因（核糖体蛋白P0和L37a）共同上调。这些结果表明，参与病毒蛋白激活的细胞基因通常可能与已建立的HCC中未与激活相关的基因不同，并且在体外实验中起作用的通过病毒蛋白激活的特定致癌协同作用可能仅在体内肝癌发生中起潜在作用。此外，本研究中鉴定的八个新基因的功能分析可能对进一步了解肝癌发生机制有价值。
