串联序列amiRNA稳定转染后对cccDNA的影响(1)
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[摘要] 目的：观察优化设计的串联序列amiRNA(artificial microRNA)质粒表达载体长期干扰乙型肝炎病毒(HBV)后，对HBV复制源泉cccDNA的影响。方法：将前期构建的针对HBV RNA干扰效率高的单一序列及串联序列amiRNA质粒表达载体，瞬时转染入HepG2.2.15细胞，筛选出稳定转染细胞株，观察其对cccDNA的影响。结果：稳定转染单一序列amiRNA-HBV-4组相对于阴性对照质粒，对HBV cccDNA的抑制率为93.78%；串联序列amiRNA-HBV3-4组为99.65%，二者比较，差异有高度统计学意义(P<0.01)。结论：构建的针对HBV的新型amiRNA质粒表达载体可较明显地抑制HBV复制的根源cccDNA，串联序列组效果更佳，为进一步进行体内实验奠定了基础。
[关键词] 乙型肝炎病毒(HBV)；amiRNA；串联序列质粒表达载体；稳定转染；cccDNA
[中图分类号] R512 [文献标识码] A [文章编号] 1673-7210(2011)11(a)-020-04
Influence of the tandem sequence expression plasmids of amiRNA against HBV cccDNA
PU Chunwen1,2, WANG Lifen3*, ZHANG Yong2, LIU Wei1, ZHANG Ying1, CONG Shurong1
1.The Sixth People′s Hospital of Dalian City, Liaoning Province, Dalian 116033, China; 2.Dalian Medical University, Liaoning Province, Dalian 116033, China; 3.The Second Affiliated Hospital of Dalian Medical University, Liaoning Province, Dalian 116033, China
[Abstract] Objective: To observe the effect of cccDNA after the long-term inhibition of tandem sequence amiRNA plasmids expression vector against HBV. Methods: The earlier constructed amiRNA plasmids of singular and tandem sequences against HBV were transfected into HepG2.2.15 cells transiently. Then the cells which stably expressed miRNA were screened. Last cccDNA were measured to observe the long-term inhibition efficiency. Results: HBV cccDNA remarkably decreased in amiRNA-HBV-4 group by 93.78% compared with the negative control plasmid group; amiRNA-HBV3-4 group by 99.65%, there was a significant difference between them (P<0.01). Conclusion: The tandem sequence plasmid expression vector amiRNA can remarkably inhibit replication of HBV cccDNA, and the tandem sequence group is better. It pave the way for this plasmid expressing carrier to further study in vivo.
[Key words] HBV; amiRNA; Tandem sequence plasmids expression vector; Stable transfection; cccDN
乙型肝炎是由乙型肝炎病毒感染导致的传染病，目前已成为全球重要的公共卫生问题[1]。近几年抗病毒治疗的临床研究证实，只要血清中HBV病毒量持续阴性，患者的病情即会有根本性好转 ......
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