Flos Farfarae Inhibits Enterovirus 71-Induced Cell Injury by Preventing Viral Replication and Structural Protein Expression.

Enterovirus 71 (EV71) infection can cause airway symptoms, brainstem encephalitis, neurogenic shock, and neurogenic pulmonary edema with high morbidity and mortality. There is no proven therapeutic modality. Flos Farfarae is the dried flower bud of Tussilago farfara L. that has been used to manage airway illnesses for thousands of years. It has neuro-protective activity and has been used to manage neuro-inflammatory diseases. However, it is unknown whether Flos Farfarae has activity against EV71-induced neuropathy. The current study used both human foreskin fibroblast (CCFS-1/KMC) and human rhabdomyosarcoma (RD) cell lines to test the hypothesis that a hot water extract of Flos Farfarae could effectively inhibit EV71 infection. The authenticity of Flos Farfarae was confirmed by HPLC-UV fingerprint. Through plaque reduction assays and flow cytometry, Flos Farfarae was found to inhibit EV71 infection ([Formula: see text]). Inhibition of viral replication and protein expression were further confirmed by reverse transcription polymerase chain reaction (RT-PCR) and quantitative RT-PCR (qRT-PCR), and western blot, respectively. The estimated IC[Formula: see text]s were 106.3[Formula: see text][Formula: see text]g/mL in CCFS-1/KMC, and 15.0[Formula: see text][Formula: see text]g/mL in RD cells. Therefore, Flos Farfarae could be beneficial to inhibit EV71 infection by preventing viral replication and structural protein expression.

Gan-Lu-Siao-Du-yin, a prescription of traditional Chinese medicine, inhibited enterovirus 71 replication, translation, and virus-induced cell apoptosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Gan-Lu-Siao-Du-yin (GLSDY) is a prescription of traditional Chinese medicine. GLSDY contains 11 ingredients and is commonly used for endemic diseases. Enterovirus 71 (EV71) is an endemic disease that can cause meningoencephalitis with mortality and neurologic sequelae without any effective management. It is unknown whether GLSDY is effective against EV71 infection. AIM OF THE STUDY: To test the hypothesis that GLSDY can protect cell from EV71-induced injury. MATERIALS AND METHODS: Effects of a hot water extract of GLSDY on EV71 were tested in human foreskin fibroblast cells (CCFS-1/KMC) and human rhabdomyosarcoma cells (RD cells) by plaque reduction assay and flow cytometry respectively. Inhibition of viral replication was further examined by reverse quantitative RT-PCR (qRT-PCR). Its effect on viral protein translation and virus-induced apoptosis were examined by western blot. RESULTS: GLSDY was dose-dependently effective against EV71 infection (p<0.0001) in both CCFS-1/KMC cells and RD cells. GLSDY was highly effective when supplemented after viral inoculation (P<0.0001) with an IC50 of 8.7µg/mL. GLSDY inhibited viral RNA replication (P<0.0001), formation of viral structural proteins (VP0, VP1, VP2 and VP3) and non-structural proteins (protease 2B and 3AB). Furthermore, 300µg/mL GLSDY is effective to inhibit virus-induced apoptosis possibly through direct inhibition of caspase-8 and indirectly by inhibition of Bax. CONCLUSIONS: GLSDY is cheap and readily available to manage EV71 infection by inhibiting viral replication, viral protein formations, and EV71-induced apoptosis.

Yakammaoto inhibits enterovirus 71 infection by reducing viral attachment, internalization, replication, and translation.

Enterovirus 71 (EV71) can cause central nervous system infections with mortality and neurologic sequelae. At present, there is no effective therapeutic modality for EV71 infection. The infection is more common in families with poor socioeconomic status. Therefore, finding a readily available, cost-effective therapeutic modality would be very helpful to these socioeconomically disadvantaged families. Yakammaoto is a cheap and readily available traditional prescription that is proven to have antiviral activity against coxsackievirus B4 (CVB4). CVB4 and EV71 are enteroviruses. In this study, we evaluated the antiviral activity of hot water extract of yakammaoto against EV71. The results of plaque reduction assay and flow cytometry demonstrated that yakammaoto dose dependently inhibited EV71 infection. In addition, reverse transcription-polymerase chain reaction (RT-PCR) and quantitative RT-PCR results showed that yakammaoto reduced viral replication. Western blotting analysis showed that yakammaoto can inhibit viral protein production. Thus, our results suggest that yakammaoto should be considered to manage EV71 infection in the future.

Yakammaoto inhibited human coxsackievirus B4 (CVB4)-induced airway and renal tubular injuries by preventing viral attachment, internalization, and replication.

ETHNOPHARMACOLOGICAL RELEVANCE: Yakammaoto is a prescription of traditional Chinese medicine (TCM) containing nine ingredients, including Ephedra sinica, Pinellia ternate, Zingiber officinale, Tussilago farfara, Aster tataricus, Ziziphus jujube, Belamcanda chinensis, Asarum sieboldii, and Schisandra chinensis. Yakammaoto has been used against flu-like symptoms for more than two thousand years in China and Japan. Coxsackievirus B4 (CVB4) causes not only flu-like symptoms but life-threatening diseases, such as pneumonia, acute kidney injury, and so forth with severe morbidity and mortality. There is no effective therapeutic modality against CVB4 infection. It is unknown whether yakammaoto is effective against CVB4 infection. We tested the hypothesis that yakammaoto can effectively inhibit CVB4-induced plaque formation in human airway and renal tubular cell lines by preventing viral attachment, internalization, and replication. MATERIALS AND METHODS: The fingerprint of yakammaoto was assessed by HPLC. Effects of yakammaoto on CVB4 infection were tested by plaque reduction assay, reverse transcription polymerase chain reaction (RT-PCR), and enzyme-linked immunosorbent assay (ELISA). RESULTS: Yakammaoto dose-dependently inhibited CVB4-induced plaque formation in HK-2, A549, and HEp-2 cells (p<0.0001). Yakammaoto was both effective when supplemented prior to and after viral inoculation (p<0.0001) by preventing viral attachment (p<0.0001), internalization (p<0.0001), and replication (p<0.0001). Yakammaoto could decrease NGAL secretion before cytolysis to protect against viral injury. CONCLUSIONS: Yakammaoto had antiviral activity against CVB4-induced cellular injuries in airway mucosa and renal tubular epithelia by preventing viral attachment, internalization, and replication. The current study provides a basic support of its potential use against CVB4-induced airway and concomitant renal injuries.

Water extract of Pueraria lobata Ohwi has anti-viral activity against human respiratory syncytial virus in human respiratory tract cell lines.

Human respiratory syncytial virus (HRSV) infects all age groups and causes bronchiolitis, pneumonia, and acute respiratory distress syndrome with a significant mortality rate. To date, only ribavirin has been used to manage HRSV infection. However, ribavirin is expensive with an only modest effect. Furthermore, ribavirin has several side effects, which means it has limited clinical benefit. Pueraria lobata Ohwi (P. lobata) is a common ingredient of Ge-Gen-Tang (Kakkon-to) and Sheng-Ma-Ge-Gen-Tang (Shoma-kakkon-to), which are prescriptions of Chinese traditional medicine proven to have antiviral activity against HRSV. Therefore, it was hypothesized that P. lobata might be effective against HRSV. To find a cost-effective therapeutic modality, both human upper (HEp-2) and lower (A549) respiratory tract cell lines were used to test the hypothesis that P. lobata could inhibit HRSV-induced plaque formation. Results showed that the water extract of P. lobata was effective (p < 0.0001) against HRSV-induced plaque formation. P. lobata was more effective when given prior to viral inoculation (p < 0.0001) by inhibiting viral attachment (p < 0.0001) and penetration (p < 0.0001). However, supplementation with P. lobata could not stimulate interferon secretion after HRSV infection. In conclusion, P. lobata has antiviral activity against HRSV-induced plaque formation in airway mucosa mainly by inhibiting viral attachment and internalization. Further identification of effective constituents could contribute to the prevention of HRSV infection.

Xiao-Qing-Long-Tang (Sho-seiryu-to) inhibited cytopathic effect of human respiratory syncytial virus in cell lines of human respiratory tract.

ETHNOPHARMACOLOGICAL RELEVANCE: Xiao-Qing-Long-Tang (XQLT, TJ-19, Sho-seiryu-to, so-cheong-ryong-tang) has been used against acute airway diseases for thousands of year in ancient China. Most of the acute airway illnesses are caused by virus. However, without activity against influenza virus, XQLT has been questioned to manage respiratory tract viral infection. Nevertheless, XQLT might be active against airway viruses other than influenza. Human respiratory syncytial virus (HRSV) is one of the most common respiratory viral pathogens without effective management. However, it is unknown whether XQLT has anti-HRSV activity. AIM OF THE STUDY: We tested the hypothesis that XQLT can effectively minimize HRSV-induced plaque formation in respiratory tract mucosal cell lines. MATERIALS AND METHODS: Anti-HRSV activity of a hot water extract of XQLT was examined by plaque reduction assay in both human upper (HEp-2) and low (A549) respiratory tract cell lines. Its effects on syncytial formation and viral fusion (F) protein were examined directly by microscopy and by western blot, respectively. Ability of XQLT to stimulate IFN-ß was evaluated by enzyme-linked immunosorbent assay (ELISA). RESULTS: Hot water extract of XQLT dose-dependently inhibited HRSV-induced plaque formation in both HEp-2 and A549 cells (P<0.0001), particularly when given before viral inoculation (p<0.0001). XQLT inhibited viral attachment (p<0.0001) and internalization (p<0.0001). 300µg/ml XQLT could decrease both the number and the size of HRSV-induced syncytium without clear effect on the production of viral F protein. XQLT could stimulate epithelial cells to secrete IFN-ß before and after viral inoculation to counteract viral infection (p<0.0001). CONCLUSIONS: XQLT is effective against HRSV infection on airway epithelia by preventing viral attachment, internalization, syncytial formation, and by stimulating interferon secretion.

Water extract of Cinnamomum cassia Blume inhibited human respiratory syncytial virus by preventing viral attachment, internalization, and syncytium formation.

ETHNOPHARMACOLOGICAL RELEVANCE: Cinnamomum cassia Blume is a popular traditional Chinese herbal medicine that has been used to manage respiratory tract disease, including common cold and chronic bronchitis for thousand years. Human respiratory syncytial virus (HRSV) is one of the leading causes of severe lower respiratory tract illness worldwide. No effective therapeutic modality against HRSV infection has been proved. It is unknown whether Cinnamomum cassia is effective against HRSV. AIM OF THE STUDY: This study tested the hypothesis that Cinnamomum cassia can effectively decrease HRSV-induced plaque formation and syncytium formation in respiratory mucosal cell lines. MATERIALS AND METHODS: Antiviral activity of the hot water extract of Cinnamomum cassia against HRSV was tested by plaque reduction assay in both human upper (HEp-2) and low (A549) respiratory tract cell lines. Its ability to inhibit the synthesis of viral fusion (F) protein was examined by Western blot assay. RESULTS: Cinnamomum cassia dose-dependently inhibited HRSV-induced plaque formation in both HEp-2 and A549 cell lines (p<0.0001). Cinnamomum cassia was more effective when given before viral infection (p<0.0001) mainly by inhibition of viral attachment (p<0.0001) and internalization (p<0.0001). Cinnamomum cassia could inhibit F protein production and syncytium formation to interfere with HRSV spreading. CONCLUSIONS: Cinnamomum cassia prevented airway epithelia from HRSV infection through inhibiting viral attachment, internalization and syncytium formation. Cinnamomum cassia could be a candidate to develop therapeutic modalities to manage HRSV infection in the future.

Fresh ginger (Zingiber officinale) has anti-viral activity against human respiratory syncytial virus in human respiratory tract cell lines.

ETHNOPHARMACOLOGICAL RELEVANCE: Ginger, Zingiber officinale Roscoe, is a common spice and also a widely used medicinal plant in ancient China. Ginger is an ingredient of Ge-Gen-Tang (Kakkon-to; GGT). GGT has been proved to have antiviral activity against human respiratory syncytial virus (HRSV). However, it is unknown whether ginger is effective against HRSV. AIM OF THE STUDY: To find a readily available agent to manage HRSV infection, the authors tested the hypothesis that ginger can effectively decrease HRSV-induced plaque formation in respiratory mucosal cell lines. MATERIALS AND METHODS: Effect of hot water extracts of fresh and dried gingers on HRSV was tested by plaque reduction assay in both human upper (HEp-2) and low (A549) respiratory tract cell lines. Ability of ginger to stimulate anti-viral cytokines was evaluated by enzyme-linked immunosorbent assay (ELISA). RESULTS: Fresh ginger dose-dependently inhibited HRSV-induced plaque formation in both HEp-2 and A549 cell lines (p<0.0001). In contrast, dried ginger didn't show any dose-dependent inhibition. 300 µg/ml fresh ginger could decrease the plaque counts to 19.7% (A549) and 27.0% (HEp-2) of that of the control group. Fresh ginger was more effective when given before viral inoculation (p<0.0001), particularly on A549 cells. 300 µg/ml fresh ginger could decrease the plaque formation to 12.9% when given before viral inoculation. Fresh ginger dose-dependently inhibited viral attachment (p<0.0001) and internalization (p<0.0001). Fresh ginger of high concentration could stimulate mucosal cells to secrete IFN-ß that possibly contributed to counteracting viral infection. CONCLUSIONS: Fresh, but not dried, ginger is effective against HRSV-induced plaque formation on airway epithelium by blocking viral attachment and internalization.