Article Figures & Data
Figures
- FIG 1
BX795 attenuates HSV-2 infection. (A) VK2 cells were infected with HSV-2 333 at 0.1 MOI, and then transcript level of viral protein UL30 was measured 24 hpi with black representing DMSO-treated cells, red representing ACV-treated cells (50 μM), and gray representing BX795-treated cells (10 μM). (B) VK2 cells were infected with HSV-2 333 at 0.1 MOI, and then transcript level of immediate early viral protein ICP27 was measured 24 hpi. (C) VK2 cells were infected with HSV-2 333 at 0.1 MOI, and then transcript level of late viral protein gD was measured 24 hpi. (D) VK2 cells were infected with HSV-2 333 at 0.1 MOI and then treated after infection with DMSO, ACV, or BX795. Samples were taken 0, 12, 24, and 36 hpi. Whole-cell lysates were probed by immunoblotting with antibodies against viral proteins VP16 and gD. (E) VK2 cells were infected with HSV-2 333 GFP at 0.1 MOI, and then cells were treated after infection with DMSO, ACV (50 μM), or BX795 (10 μM). Fluorescent images were taken at 12, 24, and 36 hpi. (F) VK2 cells were infected with HSV-2 333 at 0.1 MOI, and then medium samples were taken every 12 hpi until 48 hpi. Viral plaques of virus shed from cells into medium with black representing DMSO and gray representing BX795. (G) VK2 cells were infected with HSV-2 333 at 0.1 MOI, and then medium samples were taken every 12 hpi until 48 hpi. Viral plaques of intracellular virus, with black representing DMSO and gray representing BX795(10 μM). (H) VK2 cells were infected with HSV-2 333 GFP at 0.1 MOI and then treated with 10 μM BX795, and then cells were collected and flow cytometry was performed measuring cell GFP florescence. (I) Quantification of panel H. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
- FIG 2
Mechanism of antiviral action of BX795. (A) VK2 cells were infected with HSV-2 333 at 0.1 MOI and then treated with either mock DMSO, BX795 (10 μM), or insulin (10 nM). The cells were stained with Hoechst (nuclear blue stain) and an antibody against p-AKT. Green represents p-AKT expression. (B) Quantification of panel A. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
- FIG 3
Efficacy of BX795 as a treatment for HSV-2 infection. (A) Representative immunofluorescence microscopy images of VK2 cells infected with HSV-2 333 GFP and treated with 0 μM to 80 μM BX795. Images were taken 24 hpi. (B) Viral plaques of virus shed from cells into medium after infection with HSV-2 333 at 0.1 MOI and subsequent treatment with increasing concentrations of BX795. Cells were treated 2 hpi and samples collected 24 hpi. (C) Viral plaques of intracellular virus after infection with HSV-2 333 at 0.1 MOI and subsequent treatment with increasing concentrations of BX795. Cells were treated 2 hpi and samples collected 24 hpi. (D) VK2 cells treated with 0 μM to 80 μM BX795 were collected 24 h after treatment. An MTT assay was preformed to check viability. (E) Representative immunofluorescence microscopy images of VK2 cells infected with HSV-2 333 GFP at 0.1 MOI and then treated with 10 μM BX795 at 2, 4, 6, and 12 hpi. Images were taken 24 hpi. (F) Viral plaques of virus shed from cells into medium after infection with HSV-2 333 at 0.1 MOI and then treated with 10 μM BX795 at 2, 4, 6, and 12 hpi. Samples were taken 24 hpi. (G) Viral plaques of virus shed from cells after infection with HSV-2 333 at 0.1 MOI and then treated with 10 μM BX795 at 2, 4, 6, and 12 hpi. Samples were taken 24 hpi.
- FIG 4
In vivo efficacy of BX795 as a treatment for HSV-2 infection. (A) Secreted virus titers assessed from the swabs of vaginas (n = 5 per treatment group) 2 days postinfection. (B) Secreted virus titers assessed from the swabs of vaginas (n = 5 per treatment group) 4 days postinfection. (C) Representative 10-μm sections of epithelium from mice as follows: noninfected, infected nontreated, infected low dose, infected high dose. Quantification of panel C. *, P < 0.05; **, P < 0.01; ***, P < 0.001; ****, P < 0.0001.
Tables
- TABLE 1
List of reagents and their respective suppliers used for this study
- TABLE 2
List of all the primer sequences used for quantitative reverse transcriptase-PCR amplification
Supplemental material
- Supplemental file 1 -
Fig. S1 and S2
PDF, 302K
- Supplemental file 1 -
