锤头状核酶RCP在HepG 2215细胞中阻断HBV基因表达的初步研究

Blocking effects of hammerhead ribozyme RCP on HBV gene expression in HepG2215 cell line

目的：在细胞水平上观察核酶对ＨＢＶ基因表达的作用。方法：将在体外具有切割活性的核酶ＲＣＰ的基因构建在表达载体ｐＳＶＬ中，获得重组质粒ＲＣＰ／ｐＳＶＬ，运用脂质体介导的基因转染技术将重组核酶基因引入ＨｅｐＧ２２１５细胞，并以空载体ｐＳＶＬ转染组为对照，通过固相放射免疫测定方法检测细胞培养上清的ＨＢｓＡｇ及ＨＢｅＡｇ分泌量，以此观察核酶ＲＣＰ对ＨＢＶ基因表达的影响。结果：在暂时性表达系统中，针对Ｐ基因５＇－端２３６０位点的核酶ＲＣＰ对ＨｅｐＧ２２１５细胞中ＨＢｓＡｇ和ＨＢｅＡｇ分泌量的抑制率分别是２６．７％、２４．８％。结论：锤头状核酶ＲＣＰ在细胞水平上能一定程度抑制ＨＢＶ基因的表达。

Objective: To investigate the cleavage activities of ribozyme RCP in blocking HBV gene expression and replication in eukaryotic cells. Methods: The recombinant plasmid RCP/pSVL which can transiently express ribozyme RCP in eukaryotic cells was constructed and then intrduced into HepG2215 cell line with the technique of lipofectaminemediated gene transfer. The vector pSVL transfection group was employed to serve as the control. HBsAg and HBeAg from the culture medium of the cells were tested with solidphase radioimmunoassay (RIA). Results: The cpm obtained form the medium samples showed that the inhibition rates of ribozyme RCP for HBsAg and HBeAg were 26.7% and 24.8% respectively in this transient expression system. Conclusion: Hammerhead ribozyme RCP can inhibit HBV gene expression in eukaryotic cells on the cell level.