Activation Of Avian Innate Immunity By Baculovirus Of Antheraea Pernyi

Chickens are an economically important livestock animal, while avian diseases pose a significant threat to the poultry industry. Although vaccines are available for most poultry diseases, it often takes time to provide optimal protection. Exposure to viral infections during this period of time would still result in the animal acquiring the diseases. The innate immune system is the first line of defence and is responsible for the recognition of pathogens. Thus, there is a need to identify and characterize immunomodulators that offer rapid protection to avian by enhancing the innate immune response.In this study, baculovirus-induced immune response in avian was characterized, and the molecular mechanisms of baculovirus uptake were evaluated. In addition, the signal pathway of baculovirus-induced activation in avian macrophage-like cell line was further investigated1. Proinflammatory cytokines expression in budded baculovirus (BV) induced-chicken peripheral blood mononuclear cells (PBMC) and macrophage cell line (HD11) was measured by real-time PCR. Nitric oxide (NO) production in HD11 cells was measured by Griess assay. Degranulation of chicken heterophils was evaluated by determining the activity of releasedβ-glucuronidase. BV significantly increased the mRNA levels of IL-12, IL-6 and IFN-γ, up-regulated NO production, and inducted chicken heterophils degranulation. In addition, two-day old chicken were inoculated intranasally with BV, followed by i.n. administration of infectious bronchitis virus (IBV) 24 h later. The survival rates of the chickens were assessed over a period of 14 days. Neonatal chickens treated with 25μg BV were protected from a lethal challenge of IBV. These data indicated that baculovirus stimulated strong innate immune response in avian.2. Effects of inactivated-BV, BV DNA with or without liposomes and two endosomal inhibitors, monodansylcadaverine (MDC) and chloroquine on cytokine expression were examined to investigate the way in which BV may enter the cell. BV treated with UV, BEI or Triton X-100 exhibited diminished ability to activate an innate immune response. BV DNA was incapable of stimulating an immune response, but addition of liposomes to the DNA preparatiuon restored this activity. Inhibitors of clathrin-dependent endocytosis and endosomalmaturation abolished most BV-stimulated cytokine expression in HD11 cells. These data provided evidence that internalization of BV DNA via endocytosis mediated by the viral envelope glycoprotein, as well as endosomal maturation, which releases the viral genome into cellular compartments, was necessary for the induction of the innate immune response by BV.3. We investigated the expression of the ChTLR genes in resting and BV infected HD11 (chicken macrophage-like cell line). Expressions of ChTLR (ChTLR1-5, 7, 21) were detected in resting HD11 cells. After BV stimulation, ChTLR21 was the only upregulated ChTLR in HD11 cells, while all other TLR were downregulated. To further determine the role of ChTLR21 in BV-mediated immune response in HD11 cells, small interfering RNA was used to silence the ChTLR21 gene. The results indicated that expression of proinflammatory cytokines (IFN-γand IL-12) and production of nitric oxide (NO) were dependent on ChTLR21 expression.4. Activation of MAPK and NF-κB in BV-inducted HD11 cells was examined by Western blot and the signaling cascades responsible for the expression of proinflammatory cytokines (IFN-γand IL-12) and production of nitric oxide (NO) were evaluated by using highly selective pharmacological inhibitors. Western blot analysis revealed that ERK1/2, p38 MAPK and JNK were phosphorylated followed by activation of p65-NF-κB after HD11 cells were challenged with BV. P38 and NF-κB inhibition resulted in abrogation of BV-induced production of IL-12, IFN-γand NO, and JNK blockade had this effect only for cytokines expression. ERK inhibition had no impact. Taken together, these findings indicated that in BV-induced HD11 cells, the induction of cytokines was mediated by the p38 MAPK, JNK and NF-κB pathways, and NO production was mediated through the p38 MAPK/NF-κB pathways.