Antiviral Activity of Tannic Acid Modified Silver Nanoparticles: Potential to Activate Immune Response in Herpes Genitalis

Abstract

(1) Background: Tannic acid is a plant-derived polyphenol showing antiviral activity mainly because of an interference with the viral adsorption. In this work, we tested whether the modification of silver nanoparticles with tannic acid (TA-AgNPs) can provide a microbicide with additional adjuvant properties to treat genital herpes infection. (2) Methods: The mouse model of the vaginal herpes simplex virus 2 (HSV-2) infection was used to test immune responses after treatment of the primary infection with TA-AgNPs, and later, after a re-challenge with the virus. (3) Results: The mice treated intravaginally with TA-AgNPs showed better clinical scores and lower virus titers in the vaginal tissues soon after treatment. Following a re-challenge, the vaginal tissues treated with TA-AgNPs showed a significant increase in the percentages of IFN-gamma+ CD8+ T-cells, activated B cells, and plasma cells, while the spleens contained significantly higher percentages of IFN-gamma+ NK cells and effector-memory CD8+ T cells in comparison to NaCl-treated group. TA-AgNPs-treated animals also showed significantly better titers of anti-HSV-2 neutralization antibodies in sera; and (4) Conclusions: Our findings suggest that TA-AgNPs sized 33 nm can be an effective anti-viral microbicide to be applied upon the mucosal tissues with additional adjuvant properties enhancing an anti-HSV-2 immune response following secondary challenge.

Keywords: HSV-2; silver nanoparticle; tannic acid.

Figure 1

Scanning transmission electron microscopy (STEM) image of silver nanoparticles (AgNPs) (A), STEM size distribution histogram (B), dynamic light scattering (DLS) size distribution histogram (C), and the overall results of the AgNPs’ characterization (D).

Figure 2

(A) The ultrastructure of Herpes simplex virus-2 (HSV-2) with attached several AgNP particles and EDX spectra of silver and uranium aligned with observed objects, acquired using the line-scan mode; scanning-transmission electron microscope, scale bar = 100 nm. (B) The ultrastructure of HSV-2 covered with silver nanoparticles; transmission electron microscope, scale bar = 50 nm.

Figure 3

Tannic acid modified silver nanoparticles (TA-AgNPs) reduce genital HSV-2-infection in mice. C57BL6 mice were inoculated intravaginally with 10³ PFU of HSV-2, then treated with 5 µg/mouse of TA-AgNPs at 6, 24, and 48 h after infection. Disease development expressed as mean disease score (A); (n = 5–20). (B) The numbers of the HSV-2 DNA copies in the vaginal tissues obtained on day eight post-infection, and in the spinal cord obtained on day eight of infection were assessed by PCR. The means are expressed as mean ± standard error of the mean (SEM) for n = 20; * represents significant differences with p ≤ 0.05, while ** means p ≤ 0.01.

Figure 4

Treatment with TA-AgNPs helps to boost T and NK cells’ response after re-challenge. C57BL6 mice were infected intravaginally with HSV-2, then treated with 5 µg/mouse of TA-AgNPs 6, 24, and 48 h after infection. Thirty days later, the mice were re-challenged with the same virus dose. The cell suspensions from the vaginal tissues and spleens were prepared 10 days after re-challenge. The percentage of CD8+ T cells and CD8+/IFN-gamma+ T cells (A), and monocytes and dendritic cells (B) in the vaginal tissue, as well as the percentage of NK1.1/IFN-gamma+ cells and CD8+/IFN-gamma+ T cells (C), and the effector memory CD8+ T cells (D) were accessed by flow cytometry. The means are expressed as mean ± SEM for n = 20; * represents significant differences with p ≤ 0.05, while ** means p ≤ 0.01.

Figure 5

Treatment with TA-AgNPs helps to boost B cells’ response after re-challenge in the vaginal tissue. C57BL6 mice were infected intravaginally with HSV-2, and treated with 5 µg/mouse of TA-AgNPs 6, 24, and 48 h after infection. Thirty days later, the mice were re-challenged with the same virus dose. The cell suspensions from the vaginal tissues prepared 10 days after re-challenge with HSV-2 were stained for plasma B cells (CD138+/B220low+/IgD−) and activated B cells (CD27+/B220+/IgD+). The means are expressed as mean ± SEM for n = 20; * represents significant differences with p ≤ 0.05, while ** means p ≤ 0.01.

Figure 6

Intravaginal treatment with TA-AgNPs of HSV-2 infection induces better seroneutralization titers. The C57BL6 mice were infected intravaginally with HSV-2, and treated with 5 µg/mouse of TA-AgNPs, then re-challenged thirty days later with the same virus dose. Sera were taken at 10 days after re-challenge with HSV-2, and were subjected to seroneutralization tests. The means are expressed as mean ± SEM for n = 20; * represents significant differences with p ≤ 0.05.