Attenuation of nitric oxide bioavailability in porcine aortic endothelial cells by classical swine fever virus

Abstract

Classical swine fever (CSF) causes severe disease in pigs, characterized by hemorrhage, fever, and leucopenia. A primary target of the virus is endothelial cells, where a pro-inflammatory and pro-coagulant response occurs with downregulation of gap junctional communication; these changes establish a basis for haemostatic imbalance. The aim of this study was to gain an understanding of the effect of classical swine fever virus (CSFV) on endothelial nitric oxide synthase (eNOS) expression and nitric oxide (NO) bioavailability. Porcine aortic endothelial cells (PAECs) were infected with CSFV at different multiplicity of infection (M.O.I.) for 48 h. Downregulation of the transcription and translation levels of eNOS was detected by semi-quantitative RT-PCR, immunoconfocal microscopy, and western blotting. This was accompanied by a reduction in NO bioavailability and attenuation of angiogenesis. Without influence from the progeny virus titer, the decrease in eNOS protein was reversed by an ERK inhibitor (PD98059) and two PI3/Akt inhibitors (LY294002 and wortmannin). In addition, we found that the transcription factors AP1, Sp1, and GATA1/2 may be involved in the downregulation of eNOS promoter activity. In conclusion, infection of PAECs with CSFV attenuated the expression of eNOS and reduced NO bioavailability through activation of the ERK and PI3/Akt pathways.

Classical swine fever virus induces oxidative stress in swine umbilical vein endothelial cells

“…Our current observation shows that CSFV infection induces ROS in a time dependent manner. In lines with our finding, the down-regulation of NO production and attenuation of the expression of NO synthase in the CSFV infected porcine macrophages and aortic endothelial cells have been reported [ 15 ]. The concentrations of ROS reduce bioactive NO through chemical inactivation, forming toxic peroxynitrite, which in turn can uncouple endothelial NO synthase to form a dysfunctional superoxide-generating enzyme that contributes further to oxidative stress [ 27 , 28 ].…”

“…The CSFV replication did not cause direct damage to the vascular endothelial cells; the cell morphology was normal and no CPE or plaque formation was induced in the infected cells [ 19 ]. However, the physiological function of vascular endothelial cells (VECs) was remarkably changed post infection [ 14 , 15 , 20 ]. It was found that vascular endothelial cells infected with CSFV induce tissue factor expression and inhibit apoptosis and interferon synthesis, which may lead to the establishment of long-term infection [ 14 ].…”

“…One intriguing possibility is that CSFV through its effect on oxidative stress plays a pathophysiological role in vascular dysfunction. Interestingly, the bioavailability of nitric oxide (NO), a dominant factor responsible for regulating vascular function, was found to be reduced in CSFV infected vascular endothelial cells [ 15 ]. And it was also reported to be decreased when oxidative stress and the aberrant expression of ROS occurred [ 16 ].…”