title: MDCK cell line with inducible allele B NS1 expression propagates delNS1 influenza virus to high titres.
abstract: Influenza A viruses lacking the gene encoding the non-structural NS1 protein (delNS1) have potential use as live attenuated vaccines. However, due to the lack of NS1, virus replication in cell culture is considerably reduced, prohibiting commercial vaccine production. We therefore established two stable MDCK cell lines that show inducible expression of the allele B NS1 protein. Upon induction, both cell lines expressed NS1 to about 1000-fold lower levels than influenza virus-infected cells. Nevertheless, expression of NS1 increased delNS1 virus titres to levels comparable to those obtained with an isogenic virus strain containing an intact NS1 gene. Recombinant NS1 expression increased the infectious virus titres 244 to 544-fold and inhibited virus induced apoptosis. However, NS1 expression resulted in only slightly, statistically not significant, reduced levels of interferon-β production. Thus, the low amount of recombinant NS1 is sufficient to restore delNS1 virus replication in MDCK cells, but it remains unclear whether this occurs in an interferon dependent manner. In contrast to previous findings, recombinant NS1 expression did not induce apoptosis, nor did it affect cell growth. These cell lines thus show potential to improve the yield of delNS1 virus for vaccine production.
authors.affiliation: Central Veterinary Institute of Wageningen UR (CVI), P.O. Box 65, 8200 AB Lelystad, the Netherlands. rutger.vanwielink@wur.nl
authors.forename: R
authors.initials: R
authors.lastname: van Wielink
authors.forename: M M
authors.initials: MM
authors.lastname: Harmsen
authors.forename: D E
authors.initials: DE
authors.lastname: Martens
authors.forename: B P H
authors.initials: BP
authors.lastname: Peeters
authors.forename: R H
authors.initials: RH
authors.lastname: Wijffels
authors.forename: R J M
authors.initials: RJ
authors.lastname: Moormann
mesh_terms: D000483:Alleles; D000818:Animals; D017209:Apoptosis / Q000235:genetics; D002460:Cell Line; D002522:Chlorocebus aethiops; D004285:Dogs; D053118:Influenza A Virus, H1N1 Subtype / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D053124:Influenza A Virus, H5N1 Subtype / Q000235:genetics / Q000276:immunology / Q000378:metabolism; D009980:Influenza A virus / Q000235:genetics* / Q000276:immunology / Q000378:metabolism*; D007252:Influenza Vaccines / Q000235:genetics / Q000378:metabolism; D016899:Interferon-beta / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics / Q000276:immunology; D014613:Vaccines, Attenuated / Q000235:genetics / Q000378:metabolism; D014709:Vero Cells; D019562:Viral Load; D017361:Viral Nonstructural Proteins / Q000096:biosynthesis* / Q000235:genetics* / Q000276:immunology; D014779:Virus Replication / Q000235:genetics*
chemical_list: C085820:INS1 protein, influenza virus; D007252:Influenza Vaccines; D011994:Recombinant Proteins; D014613:Vaccines, Attenuated; D017361:Viral Nonstructural Proteins; D016899:Interferon-beta