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

Abstract

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.

Keywords: 3,3',5,5'-tetramethylbenzidine; 50% cytotoxic concentration; ATCC; CC(50); CVB4; DMEM; DMSO; Dulbecco's Modified Eagle medium; ELISA; FCS; HEp-2; HK-2; IC(50); IFN; Kidney injury; PBS; Respiratory tract infection; SI; She-Gan-Ma-Huang-Tang; Sheganmahuang decoction; Sheganmahuang decoction (Chinese); TCM; TMB; Yakammaoto; Ye-Gan-Ma-Huang-Tang; Ye-Gan-Ma-Huang-Tang (Taiwan); dimethylsulfoxide; enzyme-linked immunosorbent assay; fetal calf serum; human coxsackievirus B4; human kidney 2 cell; human larynx epidermoid carcinoma cell; interferon; minimal concentration required to inhibit 50% plaque formation; pfu; phosphate-buffered saline; plaque forming unit; selectivity index; the American Type Culture Collection; traditional Chinese medicine.