In Vitro Antiviral Activity of Sakuranetin against Human Rhinovirus 3

Abstract

Objectives: Rhinoviruses (RVs) cause common cold and are associated with exacerbation of chronic inflammatory respiratory diseases. Until now, no clinically effective antiviral chemotherapeutic agents to treat diseases caused by human rhinoviruses (HRVs) have been reported. We assessed the anti-HRV3 activity of sakuranetin isolated from Sorbus commixta Hedl. in human epithelioid carcinoma cervix (HeLa) cells, to evaluate its anti-rhinoviral potential in the clinical setting.

Methods: Antiviral activity and cytotoxicity as well as the effect of sakuranetin on HRV3-induced cytopathic effects (CPEs) were evaluated using the sulforhodamine B (SRB) method using CPE reduction. The morphology of HRV3-infected cells was studied using a light microscope.

Results: Sakuranetin actively inhibited HRV3 replication and exhibited antiviral activity of more than 67% without cytotoxicity in HeLa cells, at 100 μg/mL. Ribavirin showed anti-HRV3 activity similar to that of sakuranetin. Treatment of HRV-infected HeLa cells with sakuranetin visibly reduced CPEs.

Conclusion: The inhibition of HRV production by sakuranetin is mainly due to its general antioxidant activity through inhibition of viral adsorption. Therefore, the antiviral activity of sakuranetin should be further investigated to elucidate its mode of action and prevent HRV3-mediated diseases in pathological conditions.

Keywords: Sorbus commixta; antiviral; human rhinovirus; sakuranetin.

Figure 1

Structure of sakuranetin isolated from Sorbus commixta.

Figure 2

Antiviral activity of sakuranetin against human rhinovirus (HRV) 3 in human epithelioid carcinoma cervix (HeLa) cells. Antiviral activity of the three extracts against HRV3 in HeLa cells is shown; (A) Antiviral activity of sakuranetin against HRV3 in HeLa cells. (B) Cytotoxicity of sakuranetin in HeLa cells. The diluted virus suspension, containing 30 mM MgCl₂, 1% fetal bovine serum, and 50% tissue culture infective dose of the virus, was added to HeLa cells to produce the appropriate cytopathic effect (CPE) within 48 hours after infection. The antiviral activity and cytotoxicity of sakuranetin were investigated through the sulforhodamine B (SRB) assay with CPE reduction. Results are presented as the mean percentage values obtained from three independent experiments performed in triplicate ± standard deviation.

Figure 3

The effect of sakuranetin on human rhinovirus (HRV) 3-induced cytopathic effect (CPE). The effects of sakuranetin on HRV3-induced CPE are shown. Culture medium in 96-well tissue culture plates was aspirated and the cells were washed with phosphate buffered saline. Thereafter, 0.09 mL of the diluted virus suspension, containing 30 mM MgCl₂, 1% fetal bovine serum, and 50% tissue culture infective dose of the virus and 0.01 mL of medium were added to human epithelioid carcinoma cervix (HeLa) cells to produce the appropriate CPE within 48 hours after infection, and then sakuranetin or ribavirin (100 mg/mL) was added. After incubation at 32°C and 5% CO₂ for 2 days, the cells stained by SRB, and cellular morphology was studied using photographs taken under a light microscope (×400). (A) Non-infected cells; (B) HRV3-infected cells without sakuranetin or ribavirin treatment; (C) non-infected cells treated with sakuranetin; (D) virus-infected cells treated with sakuranetin; (E) non-infected cells with treated ribavirin; (F) virus-infected cells treated with ribavirin.