优化的串联 amiRNA 介导更强的乙型肝炎病毒感染抑制作用。

Optimized tandem amiRNA mediates stronger inhibitory effects on hepatitis B virus infection.

机构信息

Dalian Sixth People's Hospital, Dalian 116033, China.

出版信息

J Gastrointestin Liver Dis. 2011 Sep;20(3):271-8.

PMID:21961095

Abstract

BACKGROUND AND AIMS

RNA interference (RNAi) has emerged as a potential new approach against hepatitis B virus (HBV) infection but unfortunately it also selects resistant virus mutants. In this study we combined the advantages of artificial micro RNAs (amiRNAs) reported previously with the purpose of constructing a more practical amiRNA with high inhibition effects against HBV.

METHOD

Aiming at conserved sites, we constructed singular-sequence vectors amiRNA-HBV1, amiRNA-HBV2, amiRNA-HBV3 and amiRNA-HBV4. We chose the two sequences of high efficiency, then built the tandem-sequence vector amiRNA-HBV3-HBV4. These vectors were transfected into HepG2.2.15 transiently. The secreted HBV surface antigen (HBsAg) and HBV 'e' antigen (HBeAg) were measured with a chemiluminescent microparticle immunoassay, and intracellular and extracellular HBV DNA was quantified by real-time PCR.

RESULTS

Our results demonstrated that amiRNA-HBV1, amiRNA-HBV2, amiRNA-HBV3, and amiRNA-HBV4 achieved a maximum inhibition of HBV mRNA expression of 29.3%, 14.9%, 61.2%, and 75.6%, respectively, while the tandem amiRNA-HBV3-HBV4 vector led to an inhibition of 87.2%.

CONCLUSION

Taken together, our data suggest that vector-based multiple artificial microRNAs are a promising therapeutic approach for chronic HBV infection.

摘要

背景与目的

RNA 干扰（RNAi）已成为一种针对乙型肝炎病毒（HBV）感染的潜在新方法，但不幸的是，它也会选择耐药病毒突变体。在这项研究中，我们结合了先前报道的人工 microRNA（amiRNA）的优势，旨在构建一种更实用的 amiRNA，对 HBV 具有更高的抑制作用。

方法

针对保守位点，我们构建了单序列载体 amiRNA-HBV1、amiRNA-HBV2、amiRNA-HBV3 和 amiRNA-HBV4。我们选择了两个高效的序列，然后构建了串联序列载体 amiRNA-HBV3-HBV4。这些载体被瞬时转染到 HepG2.2.15 中。用化学发光微粒子免疫测定法测量分泌的 HBV 表面抗原（HBsAg）和 HBV 'e' 抗原（HBeAg），实时 PCR 定量细胞内和细胞外 HBV DNA。