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IFN-CSP inhibiting hepatitis B virus in HepG2.2.15 cells involves JAK-STAT signal pathway.

Lu X, Wang J, Jin X, Huang Y, Zeng W, Zhu J - Biomed Res Int (2015)

Bottom Line: This targeting would target the IFNα2b specifically to the liver, thus reducing the adverse events.In the present study, we further investigated the anti-HBV effects and molecular mechanisms of recombinant IFN-CSP in HepG2.2.15 cell line.IFN-CSP could be a good substitute for IFNα2b for anti-HBV treatment.

View Article: PubMed Central - PubMed

Affiliation: School of Basic Courses, Guangdong Pharmaceutical University, Guangzhou Higher Education Mega Center, 280 Wai Huan Dong Road, Guangzhou 510006, China ; Guangdong Provincial Key Laboratory of Pharmaceutical Bioactive Substances, Guangzhou Higher Education Mega Center, 280 Wai Huan Dong Road, Guangzhou 510006, China.

ABSTRACT
Frequent and high-dose administration of interferon to patients with viral hepatitis results in various side effects. In our previous study, a novel liver-targeting interferon (IFN-CSP) combining Plasmodium region I peptide with IFNα2b was successfully designed and expressed in the Escherichia coli expression systems. This targeting would target the IFNα2b specifically to the liver, thus reducing the adverse events. In the present study, we further investigated the anti-HBV effects and molecular mechanisms of recombinant IFN-CSP in HepG2.2.15 cell line. Hepatitis B surface antigen (HBsAg) and HBe antigen (HBeAg) in the culture supernatants were analyzed by enzyme-linked immunosorbent assay (ELISA). HBV-DNA was measured by real-time quantitative PCR. HBV core protein was assayed by immunofluorescent and western blot analysis. The expressions of signal transducers and transactivator 1 (STAT1), STAT2, IFN regulatory factor 9 (IRF-9), and 2'-5'-oligoadenylate synthetase 1 (OAS1) were investigated by the reverse transcription PCR and western blot analysis. Results indicate IFN-CSP efficiently inhibited HBsAg and HBeAg secretion, HBV-DNA replication, and HBV core protein expression in HepG2.2.15 cells. The anti-HBV mechanisms involve activation of JAK-STAT signaling and increase of the anti-HBV protein OAS expression. IFN-CSP could be a good substitute for IFNα2b for anti-HBV treatment.

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Related in: MedlinePlus

Analysis of HBV core protein in HepG2.2.15 cells treated with IFN-CSP for 9 days. (a) HepG2.2.15 cells stained by immunofluorescent staining with anti-HBcAg antibody and scanned by confocal microscope. Representative photographs are presented. Green stained with HBV core protein. Blue nuclear stained with DAPI. Calibration bar = 30 μm for all images. (b) The HBV core protein was also measured by western blot analysis. (c) Optical densities of the core protein were analyzed using Image-Pro Plus (IPP) software. The data are the mean ± SEM (n = 3). *P < 0.05 versus untreated controls, **P < 0.01 versus untreated controls, and ##P < 0.01 versus IFN α2b controls.
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fig2: Analysis of HBV core protein in HepG2.2.15 cells treated with IFN-CSP for 9 days. (a) HepG2.2.15 cells stained by immunofluorescent staining with anti-HBcAg antibody and scanned by confocal microscope. Representative photographs are presented. Green stained with HBV core protein. Blue nuclear stained with DAPI. Calibration bar = 30 μm for all images. (b) The HBV core protein was also measured by western blot analysis. (c) Optical densities of the core protein were analyzed using Image-Pro Plus (IPP) software. The data are the mean ± SEM (n = 3). *P < 0.05 versus untreated controls, **P < 0.01 versus untreated controls, and ##P < 0.01 versus IFN α2b controls.

Mentions: The expression level of HBV core protein in HepG2.2.15 cells was assayed by immunofluorescent staining and western blot analysis. The photographs scanned by confocal microscope (Figure 2(a)) showed that the signal of HBV core protein is obviously decreased in the cytoplasm of HepG2.2.15 cells treated with IFN-CSP in a dose-dependent manner when compared with the control cells. IFNα2b control also suppress the expression of HBV core protein in HepG2.2.15 cells, but the suppress degree less than the same dose of IFN-CSP. The result of western blot (Figures 2(b) and 2(c)) was consistent with the immunofluorescent staining.


IFN-CSP inhibiting hepatitis B virus in HepG2.2.15 cells involves JAK-STAT signal pathway.

Lu X, Wang J, Jin X, Huang Y, Zeng W, Zhu J - Biomed Res Int (2015)

Analysis of HBV core protein in HepG2.2.15 cells treated with IFN-CSP for 9 days. (a) HepG2.2.15 cells stained by immunofluorescent staining with anti-HBcAg antibody and scanned by confocal microscope. Representative photographs are presented. Green stained with HBV core protein. Blue nuclear stained with DAPI. Calibration bar = 30 μm for all images. (b) The HBV core protein was also measured by western blot analysis. (c) Optical densities of the core protein were analyzed using Image-Pro Plus (IPP) software. The data are the mean ± SEM (n = 3). *P < 0.05 versus untreated controls, **P < 0.01 versus untreated controls, and ##P < 0.01 versus IFN α2b controls.
© Copyright Policy - open-access
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC4378597&req=5

fig2: Analysis of HBV core protein in HepG2.2.15 cells treated with IFN-CSP for 9 days. (a) HepG2.2.15 cells stained by immunofluorescent staining with anti-HBcAg antibody and scanned by confocal microscope. Representative photographs are presented. Green stained with HBV core protein. Blue nuclear stained with DAPI. Calibration bar = 30 μm for all images. (b) The HBV core protein was also measured by western blot analysis. (c) Optical densities of the core protein were analyzed using Image-Pro Plus (IPP) software. The data are the mean ± SEM (n = 3). *P < 0.05 versus untreated controls, **P < 0.01 versus untreated controls, and ##P < 0.01 versus IFN α2b controls.
Mentions: The expression level of HBV core protein in HepG2.2.15 cells was assayed by immunofluorescent staining and western blot analysis. The photographs scanned by confocal microscope (Figure 2(a)) showed that the signal of HBV core protein is obviously decreased in the cytoplasm of HepG2.2.15 cells treated with IFN-CSP in a dose-dependent manner when compared with the control cells. IFNα2b control also suppress the expression of HBV core protein in HepG2.2.15 cells, but the suppress degree less than the same dose of IFN-CSP. The result of western blot (Figures 2(b) and 2(c)) was consistent with the immunofluorescent staining.

Bottom Line: This targeting would target the IFNα2b specifically to the liver, thus reducing the adverse events.In the present study, we further investigated the anti-HBV effects and molecular mechanisms of recombinant IFN-CSP in HepG2.2.15 cell line.IFN-CSP could be a good substitute for IFNα2b for anti-HBV treatment.

View Article: PubMed Central - PubMed

Affiliation: School of Basic Courses, Guangdong Pharmaceutical University, Guangzhou Higher Education Mega Center, 280 Wai Huan Dong Road, Guangzhou 510006, China ; Guangdong Provincial Key Laboratory of Pharmaceutical Bioactive Substances, Guangzhou Higher Education Mega Center, 280 Wai Huan Dong Road, Guangzhou 510006, China.

ABSTRACT
Frequent and high-dose administration of interferon to patients with viral hepatitis results in various side effects. In our previous study, a novel liver-targeting interferon (IFN-CSP) combining Plasmodium region I peptide with IFNα2b was successfully designed and expressed in the Escherichia coli expression systems. This targeting would target the IFNα2b specifically to the liver, thus reducing the adverse events. In the present study, we further investigated the anti-HBV effects and molecular mechanisms of recombinant IFN-CSP in HepG2.2.15 cell line. Hepatitis B surface antigen (HBsAg) and HBe antigen (HBeAg) in the culture supernatants were analyzed by enzyme-linked immunosorbent assay (ELISA). HBV-DNA was measured by real-time quantitative PCR. HBV core protein was assayed by immunofluorescent and western blot analysis. The expressions of signal transducers and transactivator 1 (STAT1), STAT2, IFN regulatory factor 9 (IRF-9), and 2'-5'-oligoadenylate synthetase 1 (OAS1) were investigated by the reverse transcription PCR and western blot analysis. Results indicate IFN-CSP efficiently inhibited HBsAg and HBeAg secretion, HBV-DNA replication, and HBV core protein expression in HepG2.2.15 cells. The anti-HBV mechanisms involve activation of JAK-STAT signaling and increase of the anti-HBV protein OAS expression. IFN-CSP could be a good substitute for IFNα2b for anti-HBV treatment.

Show MeSH
Related in: MedlinePlus