腺病毒表达Ribozyme在细胞内对HBV的免疫作用

了解利用腺病毒腺体表达针对乙型肝炎病毒（HBV）的核酶在细胞内对HBV表达的抑制作用。将针对HBV的多位点核酶克隆入腺病毒载体中，利用包装细胞包装出含核酶及突变核酶的重组腺病毒，将腺病毒感染2．2．15细胞（含HBV），检测感染细胞中核酶及突变核酶的表达，利用ELISA，打点杂交及激光共聚光共聚焦等方法，检测核酶对细胞内乙型肝炎病毒表达的抑制作用。结果显示核酶及突变核酶的确被克隆入腺病毒载体，并经过包装后可形成包装腺病毒，感染2．2．15细胞后可表达出目的核酶，并发挥核酶的剪切作用，其对HBV表达的抑制率最高可达87．1％。证实包装腺病毒表达的多位点核酶对HBV的复制有明确的抑制作用，支持核酶的抗病毒效果，有希望成为HBV治疗的潜在方法之一。

INHIBITION OF HBV BY RIBOZYME IN ADENOVIRUS EXPRESSION SYSTEM

To investigate the inhibition of HBV expression by multi-target ribozyme-expressing adenovirus vector in vivo, the ribozyme and muta-ribozyme were cloned into adenovirus vector pAd-CMV. By using the adenovirus expression system, the two plasmids were co-transfected into 293 cell line with adenovirus gene recombinant plasmids pJM17. The recombinant adenovirus were detected by plaque assay. Then the recombinant adenovirus infected the 2.2.15 cells. Using ELISA, dot-bolt hybridization and image analysis system, the inhibition of HBV expression was detected. The ribozyme and muta-ribozyme were cloned into the adenovirus vector pAd-CMV. The recombinant adenovirus could infect 2.2.15 cells and express the ribozyme. The ribozyme could decrease the expression of HBeAg by 87.1%. All these results strongly indicate that ribozyme can inhibit HBV expression in the cell, which represents a potential gene therapy for HBV infection.