• Xiao-Ying Dong, Wen-Jun Liu, Ming-Qiu Zhao, Jia-Ying Wang, Jing-Jing Pei, Yong-Wen Luo, Chun-Mei Ju, and Jin-Ding Chen.
• College of Veterinary Medicine, South China Agricultural University, No,483 Wu Shan Road, Tian He District, Guangzhou 510642, China. jdchen@scau.edu.cn.
• Virol J. 2013 Sep 13; 10: 286.

BackgroundClassical swine fever (CSF) caused by CSF virus (CSFV) is a highly contagious disease of pigs. The RNA helicases retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA-5) are differentially involved in the detection of various RNA viruses. In present study, we investigated the roles of RIG-I and MDA-5 in eliciting antiviral and inflammatory responses to CSFV shimen strain in Porcine alveolar macrophages (PAMs).MethodsCSFV Shimen strain was used as challenge virus in this study and PAMs were cultured in vitro. Interferon regulatory factor (IRF)-3 and nuclear factor-kappa B (NF-κB) translocation was detected using immunofluorescent staining; RIG-I, MDA5, interferon promoter-stimulating factor 1 (IPS-1), IRF-3 and NF-κB expression was measured by Western Blotting; Interferon beta (IFN-β), IFN-α, interleukin-1beta (IL-1β), IL-6 and tumor necrosis factor (TNF-α) expression was tested by Enzyme-linked immunosorbent assays (ELISA) and shRNA-mediated knockdown of MDA5 or RIG-I was performed.ResultsThe findings suggested that the initial response to CSFV infection resulted in the higher expression of RIG-I and MDA5 leading to the activation of IPS-1, IRF-3 and NF-κB in a dose-dependent manner. Evaluation of IFN-α, IFN-β, IL-1β, IL-6 or TNF-α expressed by PAMs showed significant differences between infected and uninfected cells. CSFV infected cells induced to express high levels of IFN-α, IFN-β, IL-1β, IL-6 and TNF-α in a dose-dependent way within 24 h post-infection (hpi). At the same time, CSFV improved the nuclear translocation of IRF-3 and NF-κB. We also directly compared and assessed the roles of RIG-I and MDA5 in triggering innate immune actions during CSFV infection through shRNA-mediated knockdown of MDA5 or RIG-I. We found that, compared to the control, the production of IFN-α, IFN-β, IL-1β, IL-6 and TNF-α in response to CSFV infection was heavily reduced in RIG-I knockdown cells while it was moderately decreased in MDA5 knockdown cells. PAMs derived from knockdown of both RIG-I and MDA5 almost failed to produce IFNs and inflammatory cytokines.ConclusionsIt indicates that CSFV can be recognized by both RIG-I and MDA5 to initiate the RIG-I signaling pathway to trigger innate defenses against infection.

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