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Inhibition of hepatitis C virus infection and expression in vitro and in vivo by recombinant adenovirus expressing short hairpin RNA.

ABSTRACT: BACKGROUND AND AIM:We have reported previously that synthetic small interfering RNA (siRNA) and DNA-based siRNA expression vectors efficiently and specifically suppress hepatitis C virus (HCV) replication in vitro. In this study, we investigated the effects of the siRNA targeting HCV-RNA in vivo. METHODS:We constructed recombinant retrovirus and adenovirus expressing short hairpin RNA (shRNA), and transfected into replicon-expressing cells in vitro and transgenic mice in vivo. RESULTS:Retroviral transduction of Huh7 cells to express shRNA and subsequent transfection of an HCV replicon into the cells showed that the cells had acquired resistance to HCV replication. Infection of cells expressing the HCV replicon with an adenovirus expressing shRNA resulted in efficient vector delivery and expression of shRNA, leading to suppression of the replicon in the cells by approximately 10(-3). Intravenous delivery of the adenovirus expressing shRNA into transgenic mice that can be induced to express HCV structural proteins by the Cre/loxP switching system resulted in specific suppression of virus protein synthesis in the liver. CONCLUSION:Taken together, our results support the feasibility of utilizing gene targeting therapy based on siRNA and/or shRNA expression to counteract HCV replication, which might prove valuable in the treatment of hepatitis C.

SUBMITTER: Sakamoto N

PROVIDER: S-EPMC7166320 | biostudies-literature | 2008 Sep

REPOSITORIES: biostudies-literature

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Inhibition of hepatitis C virus infection and expression in vitro and in vivo by recombinant adenovirus expressing short hairpin RNA.

Sakamoto Naoya N Tanabe Yoko Y Yokota Takanori T Satoh Kenichi K Sekine-Osajima Yuko Y Nakagawa Mina M Itsui Yasuhiro Y Tasaka Megumi M Sakurai Yuki Y Cheng-Hsin Chen C Yano Masahiko M Ohkoshi Shogo S Aoyagi Yutaka Y Maekawa Shinya S Enomoto Nobuyuki N Kohara Michinori M Watanabe Mamoru M

Journal of gastroenterology and hepatology 20070807 9

<h4>Background and aim</h4>We have reported previously that synthetic small interfering RNA (siRNA) and DNA-based siRNA expression vectors efficiently and specifically suppress hepatitis C virus (HCV) replication in vitro. In this study, we investigated the effects of the siRNA targeting HCV-RNA in vivo.<h4>Methods</h4>We constructed recombinant retrovirus and adenovirus expressing short hairpin RNA (shRNA), and transfected into replicon-expressing cells in vitro and transgenic mice in vivo.<h4> ...[more]

PMID: 17683479

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Project description:Down-regulation of the HIV-1 coreceptor CCR5 holds significant potential for long-term protection against HIV-1 in patients. Using the humanized bone marrow/liver/thymus (hu-BLT) mouse model which allows investigation of human hematopoietic stem/progenitor cell (HSPC) transplant and immune system reconstitution as well as HIV-1 infection, we previously demonstrated stable inhibition of CCR5 expression in systemic lymphoid tissues via transplantation of HSPCs genetically modified by lentiviral vector transduction to express short hairpin RNA (shRNA). However, CCR5 down-regulation will not be effective against existing CXCR4-tropic HIV-1 and emergence of resistant viral strains. As such, combination approaches targeting additional steps in the virus lifecycle are required. We screened a panel of previously published shRNAs targeting highly conserved regions and identified a potent shRNA targeting the R-region of the HIV-1 long terminal repeat (LTR). Here, we report that human CD4(+) T-cells derived from transplanted HSPC engineered to co-express shRNAs targeting CCR5 and HIV-1 LTR are resistant to CCR5- and CXCR4- tropic HIV-1-mediated depletion in vivo. Transduction with the combination vector suppressed CXCR4- and CCR5- tropic viral replication in cell lines and peripheral blood mononuclear cells in vitro. No obvious cytotoxicity or interferon response was observed. Transplantation of combination vector-transduced HSPC into hu-BLT mice resulted in efficient engraftment and subsequent stable gene marking and CCR5 down-regulation in human CD4(+) T-cells within peripheral blood and systemic lymphoid tissues, including gut-associated lymphoid tissue, a major site of robust viral replication, for over twelve weeks. CXCR4- and CCR5- tropic HIV-1 infection was effectively inhibited in hu-BLT mouse spleen-derived human CD4(+) T-cells ex vivo. Furthermore, levels of gene-marked CD4(+) T-cells in peripheral blood increased despite systemic infection with either CXCR4- or CCR5- tropic HIV-1 in vivo. These results demonstrate that transplantation of HSPCs engineered with our combination shRNA vector may be a potential therapy against HIV disease.

Dataset Information

Inhibition of hepatitis C virus infection and expression in vitro and in vivo by recombinant adenovirus expressing short hairpin RNA.

Publications

Inhibition of hepatitis C virus infection and expression in vitro and in vivo by recombinant adenovirus expressing short hairpin RNA.

Similar Datasets

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