Novel antiviral activity of baicalein against dengue virus

Abstract

Background: Dengue is a serious arboviral disease currently with no effective antiviral therapy or approved vaccine available. Therefore, finding the effective compound against dengue virus (DENV) replication is very important. Among the natural compounds, bioflavonoids derived mainly from plants are of interest because of their biological and medicinal benefits.

Methods: In the present study, antiviral activity of a bioflavonoid, baicalein, was evaluated against different stages of dengue virus type 2 (DENV-2) replication in Vero cells using focus forming unit reduction assay and quantitative RT-PCR.

Results: Baicalein inhibited DENV-2 replication in Vero cells with IC50= 6.46 μg/mL and SI= 17.8 when added after adsorption to the cells. The IC50 against DENV-2 was 5.39 μg/mL and SI= 21.3 when cells were treated 5 hours before virus infection and continuously up to 4 days post infection. Baicalein exhibited direct virucidal effect against DENV-2 with IC 50= 1.55 μg/mL and showed anti-adsorption effect with IC50 = 7.14 μg/mL.

Conclusions: Findings presented here suggest that baicalein exerts potent antiviral activity against DENV. Baicalein possesses direct virucidal activity against DENV besides its effects against dengue virus adsorption and intracellular replication of DENV-2. Baicalein, hence, should be considered for in vivo evaluation in the development of an effective antiviral compound against DENV.

Figure 1

Cytotoxicity of baicalein on Vero cells. MTT assay was used to evaluate the cytotoxicity of the flavonoid. All experiments were conducted in triplicates.

Figure 2

DENV-2 foci in Vero cells treated with or without baicalein. prophylactic treatment with 12.5 μg/ml of baicalein (A) or without baicalein (B); during adsorption with 12.5 μg/ml of baicalein (C) or without baicalein (D); after virus adsorption with 12.5 μg/ml of baicalein (E) or without baicalein (F); direct virucidal activity with 12.5 μg/ml of baicalein (G) or without baicalein (H). Foci were detected using dengue hyperimmune sera after 4 days of incubation.

Figure 3

Prophylactic effects of baicalein against DENV-2 replication. Foci forming unit reduction assay was used to evaluate the prophylactic activity of the baicalein (A). and the DENV-2 RNA copies were quantified using qRT-PCR (B). All experiments were conducted in triplicates. The percentages of foci reduction (%RF) and RNA copy number reduction were obtained by comparing against the controls maintained in parallel.

Figure 4

Effects of baicalein against DENV-2 host cell adsorption. Foci forming unit reduction assay was used to determine the anti-adsorption activity of baicalein against dengue virus (A) the respective DENV-2 RNA copies were quantified using qRT-PCR (B). All experiments were conducted in triplicates. The percentages of foci reduction (%RF) and RNA copy number reduction were obtained by comparing against the controls maintained in parallel.

Figure 5

Antiviral activity of baicalein against DENV-2 intracellular replication. Foci forming unit reduction assay was used to evaluate the anti-dengue activity of baicalein after virus adsorption to the Vero cells (A) and the respective DENV-2 RNA copies were quantified using qRT-PCR (B). All experiments were conducted in triplicates. The percentages of foci reduction (%RF) and RNA copy number reduction were obtained by comparing against the controls maintained in parallel.

Figure 6

Anti-dengue effect of continuous treatment with baicalein. Foci forming unit reduction assay was used to evaluate the in vitro anti-dengue virus activities of the baicalein (A). The respective DENV-2 RNA copy number was quantified using qRT-PCR (B). The percentages of foci reduction (%RF) and RNA copy number reduction were obtained by comparing against untreated controls maintained in parallel.

Figure 7

Direct virucidal activity of baicalein against DENV-2. Foci forming unit reduction assay was used to determine the direct anti-dengue virus activity of the baicalein. Baicalein was added directly to virus inoculum prior to infecting cells (A) and the respective DENV-2 RNA copies were quantified using qRT-PCR (B). All experiments were conducted in triplicates. The percentages of foci reduction (%RF) and RNA copy number reduction were obtained by comparing against the controls maintained in parallel.

Figure 8

Chemical structure of flavone (A) and baicalein (B).