title: Enhanced production of human influenza virus in PBS-12SF cells with a reduced interferon response.
abstract: Influenza is one of the most important infectious diseases in humans. The best way to prevent severe illness caused by influenza infection is vaccination. Cell culture-derived influenza vaccines are being considered in addition to the widely used egg-based system in order to support the increasing seasonal demand and to be prepared in case of a pandemic. Cell culture based systems offer increased safety, capacity, and flexibility with reduced downstream processing relative to embryonated eggs. We have previously reported a chick embryo cell line, termed PBS-12SF, that supports replication of human and avian influenza A viruses to high titers (>10(7) PFU/ml) without the need for exogenous proteases or serum proteins. Viral infections in cells are limited by the Interferon (IFN) response typified by production of type I IFNs that bind to the IFNα/β receptor and activate an antiviral state. In this study, we investigated how neutralizing the interferon (IFN) response in PBS-12SF cells, via shRNA-mediated knock-down of IFNAR1 mRNA expression, affects influenza virus production. We were successful in knocking down ∼90% of IFNAR1 protein expression by this method, resulting in a significant decrease in the response to recombinant chIFNα stimulation in PBS-12SF cells as shown by a reduction in expression of interferon-responsive genes when compared to control cells. Additionally; IFNAR1-knock-down cells displayed enhanced viral HA production and released more virus into cell culture supernatants than parental PBS-12SF cells.
authors.affiliation: a Molecular Pathogenesis Laboratory; Department of Animal Science; Michigan State University ; East Lansing , MI USA.
authors.forename: Monica
authors.initials: M
authors.lastname: Carvajal-Yepes
authors.forename: Kelly R B
authors.initials: KR
authors.lastname: Sporer
authors.forename: Jenna L
authors.initials: JL
authors.lastname: Carter
authors.forename: Christopher J
authors.initials: CJ
authors.lastname: Colvin
authors.forename: Paul M
authors.initials: PM
authors.lastname: Coussens
mesh_terms: D000818:Animals; D002460:Cell Line; D002645:Chickens; D055785:Gene Knockdown Techniques; D007372:Interferons / Q000096:biosynthesis*; D009975:Orthomyxoviridae / Q000254:growth & development* / Q000276:immunology / Q000302:isolation & purification*; D034741:RNA, Small Interfering / Q000378:metabolism*; D053633:Receptor, Interferon alpha-beta / Q000037:antagonists & inhibitors*; D013678:Technology, Pharmaceutical / Q000379:methods*; D019562:Viral Load; D014776:Virus Cultivation / Q000379:methods*
chemical_list: D034741:RNA, Small Interfering; D053633:Receptor, Interferon alpha-beta; D007372:Interferons
keywords: H1N1; Influenza; Interferon; Vaccine production; shRNA