SiRNA pool targeting different sites of human hepatitis B surface antigen efficiently inhibits HBV infection

Yong Chen, Ram Mahato

Research output: Contribution to journalArticle

18 Citations (Scopus)

Abstract

The main objective was to determine whether a pool of small interfering RNAs (siRNAs) targeting different regions of hepatitis B virus surface antigen (HBsAg) efficiently inhibits hepatitis B virus (HBV) infection. siRNAs targeting different regions of HBsAg were transfected into HBV-producing HepG2.2.15 cells and at 72 h post-transfection, the culture medium was collected for ELISA to determine HBsAg, while total RNA was isolated from the cells for real-time PCR. Three siRNA sequences that efficiently inhibited HBV infection were converted into small hairpin RNAs (shRNAs) and then cloned into a single plasmid psiSTRIKE™ driven by a single U6 promoter. These shRNA expressing plasmids were tested for HBsAg gene silencing in HepG2.2.15 cells. A pool of siRNAs targeting HBsAg efficiently inhibited HBV replication and antigen expression when transfected into HepG2.2.15 cells, compared with the use of single siRNA. Similarly, the plasmid encoding three different shRNAs driven by a single U6 promoter was more effective in silencing HBsAg at DNA, mRNA and protein levels compared with the plasmid encoding single shRNA. No apoptotic change was observed in the cells when the plasmid was transfected at a dose of 0.5-2 μg/1 × 106 cells after complex formation with Lipofectamine LTX™. Furthermore, transfection with siRNA or shRNA did not increase interferon-γ (IFNs-γ) release, suggesting no induction of IFN response. In conclusion, a pool of chemically synthesised siRNAs as well as the shRNA expression plasmid encoding multiple shRNAs targeting different regions of HBsAg showed high gene silencing in HepG2.2.15 cells.

Original languageEnglish (US)
Pages (from-to)140-148
Number of pages9
JournalJournal of Drug Targeting
Volume16
Issue number2
DOIs
StatePublished - Feb 1 2008

Fingerprint

Virus Diseases
Hepatitis B Surface Antigens
Hepatitis B virus
Small Interfering RNA
Plasmids
Gene Silencing
Transfection
Hepatitis B Antigens
Virus Replication
Interferons
Culture Media
Real-Time Polymerase Chain Reaction
Enzyme-Linked Immunosorbent Assay

Keywords

  • HBV replication
  • HBsAg expression
  • Hepatitis B virus
  • ShRNA
  • SiRNA

ASJC Scopus subject areas

  • Pharmaceutical Science

Cite this

SiRNA pool targeting different sites of human hepatitis B surface antigen efficiently inhibits HBV infection. / Chen, Yong; Mahato, Ram.

In: Journal of Drug Targeting, Vol. 16, No. 2, 01.02.2008, p. 140-148.

Research output: Contribution to journalArticle

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