Granulocyte macrophage colony stimulating factor is elevated in alveolar macrophages from sheep naturally infected with maedi-visna virus and stimulates maedi-visna virus replication in macrophagesin vitro

Abstract

SUMMARYInfection by maedi-visna virus, a lentivirus of sheep, leads to chronic inflammatory reactions of various tissues. In this report we have analysed the role of specific cytokines in the disease process. A significant increase in expression of interleukin-6, interleukin-10, granulocyte macrophage-colony stimulating factor (GM-CSF) and transforming growth factor-b1 mRNA was observed in alveolar macrophages isolated from the lungs of naturally infected animals when compared with lungs of seronegative controls. Levels of GM-CSF mRNA expression in alveolar macrophages correlated with the presence of lung lesions, but there was no correlation of interleukin-10, interleukin-6, tumour necrosis factor-a and transforming growth factor-b1 mRNA levels in alveolar macrophages from animals with pulmonary lesions. In vitro investigation showed that GM-CSF in the range 0·1-10 ng/ml induced a significant increase in viral p25 production after 7 days in acutely infected blood monocytederived macrophages. The production of p25 peaked between 7 and 14 days exposure to 10 ng/ml of GM-CSF. Quantitative polymerase chain reaction showed that the level of viral DNA in monocytederived macrophages was dose-dependent following GM-CSF treatment in the range 0·1-100 ng/ml after 7 days. Viral mRNA expression was also enhanced. These findings indicate a role for GM-CSF in the pathogenesis of lymphoid interstitial pneumonia in infected animals.

Immune Response to Individual Maedi-Visna VirusgagAntigens

“…The ability of MVV-infected accessory cells to present antigen creates an immunopathological feedback loop in MVV lesions. CD4 T cells responding to accessory cells presented antigen release cytokines, e.g., gamma interferon and granulocytemacrophage colony-stimulating factor, which further enhance monocyte to macrophage differentiation and virus expression (63), and these activated macrophages provide the cellular microenvironment for enhanced viral replication. …”

Role of Alveolar Macrophages in Respiratory Transmission of Visna/Maedi Virus

“…The purification of AMs was based on a procedure described previously (28,50) that relies on the preferential adherence of macrophage populations to tissue culture flasks. After pelleting and counting BAL cells, cells were resuspended in 7 ml RMPI 1640 medium (Invitrogen, Paisley, United Kingdom) containing 10% fetal calf serum (FCS), 2 mM L-glutamine, 50 g/ml penicillin, 50 g/ml streptomycin, 2 g/ml amphotericin B, and 0.05 M ␤-mercaptoethanol and were seeded into 25-cm 2 tissue culture flasks.…”

“…It is possible that AMs mediate a similar inflammatory response during early VMV infection in vivo, as suggested by the observed up-regulation of interleukin-8 (IL-8) expression by AMs within 48 h of in vitro infection (28). This may have implications for the enhancement of virus uptake, as a number of proinflammatory cytokines have been shown to enhance VMV replication in AMs and other macrophages in vitro (15,50), and generalized inflammation is likely to result in the recruitment of VMV target cells, namely, macrophages and dendritic cells (DCs) (7,36).…”

“…AMs also have been implicated as the main cell type involved in VMV-induced lung inflammation, primarily by the dysregulation of cytokine gene expression and subsequent recruitment of inflammatory cells (29,50). It is possible that AMs mediate a similar inflammatory response during early VMV infection in vivo, as suggested by the observed up-regulation of interleukin-8 (IL-8) expression by AMs within 48 h of in vitro infection (28).…”

Development and comparison of strain specific gag and pol real-time PCR assays for the detection of Visna/maedi virus

“…The British VMV strain EV1 was isolated from a sheep displaying symptoms of arthritis and pneumonia and its complete genome was cloned and sequenced ( Sargan et al, 1991). This strain has been and is currently used for in vivo and in vitro studies focusing on diverse topics such as the identification of viral promoter elements, or cytokines that control the expression and replication of the virus , Sutton et al, 1997and Zhang et al, 2002, the dissection of the initial steps of the infection ( Bird et al, 1993, Blacklaws et al, 1995and Ryan et al, 2000, the characterization of the immune response ( Bird et al, 1993 andSingh et al, 2006), the routes of viral entry and dissemination ( Blacklaws et al, 1995, McNeilly et al, 2007 and the development of immunization strategies ( Gonzalez et al, 2005, Niesalla et al, 2009and Reina et al, 2008. In some of these studies the presence of the EV1 genome has been investigated by conventional PCR ( Bird et al, 1993and Ryan et al, 2000 or by quantitative competitive PCR (QC PCR) ( Zhang et al, 2000).…”