Inhibition of hepatitis B virus replication by shRNAs in stably HBV expressed HEPG2 2.2.15 cell lines

Abstract

In this study, the effect of RNAi on HBV replication was observed in a cell culture model, HepG2 2.2.15 cell line, which supports human HBV ayw replication and expression. Aim of the study was to investigate effects of shRNAs (small hairpin RNAs) targeting hepatitis B virus mRNAs on the viral replication in HepG2 2.2.15 cells. We selected three target HBV mRNA regions with different putative secondary structures to test whether the secondary structure of RNA may affect the inhibition efficacy on the target HBV RNA. Three HBV-specific siRNAs (small interfering RNA) were designed targeting X (1689-1708), Core (2229-2248) and S (765-784 nt) transcripts. HepG2 2.2.15 cells were transfected with shRNA expressing plasmids, P765, P2229 and P1689 targeting S, core and X region, respectively or a mock plasmid targeting lacZ gene. The culture media was collected throughout six days after transfection and analyzed by real-time PCR. Viral DNA production was suppressed for 7 days. The HBV DNA levels were decreased by 73, 72 and 79% with P765, P2229 and P1689 vectors, respectively. In conclusion, the shRNAs designed for X, core and S regions, specifically and significantly suppressed HBV DNA. siRNAs potentially may be used in treatment of hepatitis B.

High load hepatitis B virus replication inhibits hepatocellular carcinoma cell metastasis through regulation of epithelial–mesenchymal transition

“…13,14 A number of studies have verified that specific siRNAs targeting four open reading frames (ORFs) of HBV are efficient at inhibiting the expression of viral antigens and the replication of HBV DNA. [15][16][17][18] Fu et al designed and evaluated 23 siRNAs targeted throughout all four ORFs of HBV. The results showed that most of them were effective, but the siRNAs targeting different ORFs could lead to different efficacy in inhibiting HBV expression and replication.…”

Efficient inhibition of human cytomegalovirus UL122 gene expression in cell by small interfering RNAs

“…Until now, two approaches have been developed to deliver extra siRNA into cells for the preselection of effective siRNAs, namely transient delivery of chemically synthesized siRNA and expression of siRNA intracellularly from vectors [27]. Vector-based RNAi has particular advantages, considering the likelihood of RNAse contamination, relatively short half-life and the great costs associated with chemically synthesized siRNA [28]. In addition, transgenic vector-based RNAi could function as an alternative method of gene silencing in vivo [29].…”

Development of sheep kidney cells with increased resistance to different subgenotypes of BVDV-1 by RNA interference

“…Specific gene silencing also can be triggered in mammalian cells by using synthetic short interfering RNA (siRNA), and plasmid or virus-mediated short hairpin RNA (shRNA). These RNAi strategies have been used successfully for suppressing the replication of human and animal viruses, including hepatitis B virus (HBV) (Kayhan et al, 2007), human immunodeficiency virus type (HIV) (Coburn and Cullen, 2002), porcine reproductive and respiratory syndrome virus (PRRSV) (Luo et al, 2013) and foot-and-mouth disease virus (FMDV) (De los Santos et al, 2005;Lv et al, 2009;Xu et al, 2012). Furthermore, we and others reported that transgenic animals expressing shRNA against virus genes showed a significant resistance to viral challenge (Pengyan et al, 2010;Li et al, 2014;Daniel-Carlier et al, 2013;Du et al, 2014).…”