Sugiyama Masaya, Tanaka Yasuhito, Kato Takanobu, Orito Etsuro, Ito Kiyoaki, Acharya Subrat K, Gish Robert G, Kramvis Anna, Shimada Takashi, Izumi Namiki, Kaito Masahiko, Miyakawa Yuzo, Mizokami Masashi
Department of Clinical Molecular Informative Medicine, Nagoya City University Graduate School of Medical Sciences, Nagoya, Japan.
Hepatology. 2006 Oct;44(4):915-24. doi: 10.1002/hep.21345.
Various genotypes of the hepatitis B virus (HBV) induce liver disease of distinct severity, but the underlying virological differences are not well defined. Huh7 cells were transfected with plasmids carrying 1.24-fold the HBV genome of different genotypes/subgenotypes (2 strains each for Aa/A1, Ae/A2, Ba/B2 and D; 3 each for Bj/B1 and C). HBV DNA levels in cell lysates, determined by Southern hybridization, were the highest for C followed by Bj/Ba and D/Ae (P < .01), and the lowest for Aa (P < .01), whereas in culture media, they were the highest for Bj, distantly followed by Ba/C/D and further by Ae/Aa (P < .01). The intracellular expression of core protein was more than 3-fold lower for Ae/Aa than the others. Hepatitis B e antigen (HBeAg) was excreted in a trend similar to that of HBV DNA with smaller differences. Secretion of hepatitis B surface antigen (HBsAg) was most abundant for Ae followed by Aa, Ba, Bj/C and remotely by D, which was consistent with mRNA levels. Cellular stress determined by the reporter assay for Grp78 promoter was higher for C and Ba than the other genotypes/subgenotypes (P < .01). Severe combined immunodeficiency mice transgenic for urokinase-type plasminogen activator (uPA/SCID), with the liver replaced for human hepatocytes, were inoculated with virions passed in mouse and recovered from culture supernatants. HBV DNA levels in their sera were higher for C than Ae by 2 logs during 4-7 weeks after inoculation. In conclusion, virological differences among HBV genotypes were demonstrated both in vitro and in vivo. These differences may influence HBV infections with distinct genotypes in clinical and epidemiological settings.
不同基因型的乙型肝炎病毒（HBV）可引发严重程度各异的肝脏疾病，但其潜在的病毒学差异尚未明确界定。用携带不同基因型/亚基因型HBV基因组1.24倍的质粒转染Huh7细胞（Aa/A1、Ae/A2、Ba/B2和D各2株；Bj/B1和C各3株）。通过Southern杂交测定细胞裂解物中的HBV DNA水平，C型最高，其次是Bj/Ba和D/Ae（P < 0.01），Aa型最低（P < 0.01）；而在培养基中，Bj型最高，Ba/C/D型次之，Ae/Aa型最低（P < 0.01）。Ae/Aa型核心蛋白的细胞内表达比其他类型低3倍以上。乙型肝炎e抗原（HBeAg）的分泌趋势与HBV DNA相似，但差异较小。乙型肝炎表面抗原（HBsAg）的分泌以Ae型最为丰富，其次是Aa型、Ba型、Bj/C型，D型最少，这与mRNA水平一致。通过Grp78启动子报告基因检测确定的细胞应激，C型和Ba型高于其他基因型/亚基因型（P < 0.01）。将表达尿激酶型纤溶酶原激活剂的严重联合免疫缺陷小鼠（uPA/SCID）的肝脏替换为人肝细胞，接种从小鼠体内传代并从培养上清液中回收的病毒粒子。接种后4至7周，其血清中C型的HBV DNA水平比Ae型高2个对数。总之，在体外和体内均证实了HBV基因型之间的病毒学差异。这些差异可能在临床和流行病学环境中影响不同基因型的HBV感染。
