OBJECTIVES: Redox status influences replication of some viruses but its effect on porcine circovirus type 2 (PCV2), the primary causative agent of the emerging swine disease post-weaning multisystemic wasting syndrome is not known. The interaction of PCV2 replication with intracellular redox status in PK15 cells was examined in this study. METHODS: Intracellular glutathione (GSH) was measured spectrophotometrically by reaction with 5, 5'-dithiobis (2-nitrobenzoic acid). Total superoxide dismutase activity (SOD) was assayed by inhibition of oxyamine oxidation by the xanthine oxidase system. Malondialdehyde (MDA) was assayed spectrophotometrically using the thiobarbituric acid reaction. Both quantification of PCV2 DNA by real-time polymerase chain reaction and indirect immunofluorescence of PCV2-infected cells were used to evaluate the replication of PCV2. RESULTS: Both GSH and SOD decreased significantly at 48 hours after PCV2 infection, whereas MDA concentration increased significantly after 48 hour post-infection. Furthermore, PCV2 replication in PK15 cells was significantly impaired after the elevation of intracellular GSH through treatment with the antioxidant N-acetyl-l-cysteine (NAC), a precursor in GSH synthesis. In contrast, PCV2 replication in PK15 cells was enhanced after reduction of GSH levels through H2O2-mediated oxidation. In addition, NAC treatment blocked the increase of virus replication induced by H2O2. CONCLUSIONS: This study suggests that PCV2 infection induces oxidative stress and that intracellular redox status influences PCV2 replication in PK15 cells.
OBJECTIVES: Redox status influences replication of some viruses but its effect on porcine circovirus type 2 (PCV2), the primary causative agent of the emerging swine disease post-weaning multisystemic wasting syndrome is not known. The interaction of PCV2 replication with intracellular redox status in PK15 cells was examined in this study. METHODS: Intracellular glutathione (GSH) was measured spectrophotometrically by reaction with 5, 5'-dithiobis (2-nitrobenzoic acid). Total superoxide dismutase activity (SOD) was assayed by inhibition of oxyamine oxidation by the xanthine oxidase system. Malondialdehyde (MDA) was assayed spectrophotometrically using the thiobarbituric acid reaction. Both quantification of PCV2 DNA by real-time polymerase chain reaction and indirect immunofluorescence of PCV2-infected cells were used to evaluate the replication of PCV2. RESULTS: Both GSH and SOD decreased significantly at 48 hours after PCV2 infection, whereas MDA concentration increased significantly after 48 hour post-infection. Furthermore, PCV2 replication in PK15 cells was significantly impaired after the elevation of intracellular GSH through treatment with the antioxidant N-acetyl-l-cysteine (NAC), a precursor in GSH synthesis. In contrast, PCV2 replication in PK15 cells was enhanced after reduction of GSH levels through H2O2-mediated oxidation. In addition, NAC treatment blocked the increase of virus replication induced by H2O2. CONCLUSIONS: This study suggests that PCV2 infection induces oxidative stress and that intracellular redox status influences PCV2 replication in PK15 cells.
Authors: Ishtiaq Qadri; Mieko Iwahashi; Juan M Capasso; Matthew W Hopken; Sonia Flores; Jerome Schaack; Francis R Simon Journal: Biochem J Date: 2004-03-15 Impact factor: 3.857