Study On Inflammatory Injury Of Chicken Embryo Liver Caused By FAdV-4

FAdV-4 is the agent of Ankara disease in poultry,which can cause HHS when it infects poultry.The main target organ for FAdV-4 invasion is the liver,which causes degeneration and necrosis of hepatocytes,and basophilic or intracytoplasmic inclusions.At present,researches on FAdV-4 mainly focus on the etiology and preventive immunology of FAdV-4,but few studies on the inflammatory injury of FAdV-4 to chicken embryo liver.In this study,FAdV-4 was inoculated with SPF chicken embryo at 11 days old,and the liver was dissected on the 7th day after infection.Pathological sections were prepared to observe the pathological changes of the liver after infection of FAdV-4 in chicken embryo.And through the qRT-PCR to detect inflammatory cytokines?IL-2?IL-4?IL-10?IFN-??and TLRs/NF-?B signaling pathway related cytokines?TLR2?TLR3?TLR4?p65?IL-1??IL-6?IL-8?IL-12??IL-12??mRNA expression of the quantity,in order to make clear FAdV-4 infection to chicken embryos after liver inflammation injury related changes in the level of gene expression,and explore the liver injury and inflammation related factor to express the relationship between.The test results show that:1.The FAdV-4 with virus titer of 10^-5-41 TCID₅₀/0.1 mL was inoculated into the allantoic cavity of 11-day-old SPF chicken embryo 0.2 mL per ege.The vital signs of the chicken embryo were observed every day in the eggs.Autopsy results showed that after infection with FAdV-4,the chicken embryo liver was swollen,with obtuse rounded edges,brittle texture,pale color,earthen yellow,and occasional stasis and bleeding on the surface.Microscopically,the cords of the liver are disorganized,hepatocytes are enlarged,vacuolar degeneration is present,and some are necrotic.Basophilic inclusions are seen in the liver cells.Erythrocytosis of central vein and hepatic sinus in the center of hepatic lobule;Kupffer cells also increased.2.qRT-PCR was used to detect the mRNA levels of inflammation-related cytokines in the liver of FAdV-4 infected chicken embryos.The results showed that the expression of IFN-?in chicken embryo liver was up-regulated after infection with FAdV-4.The expression of anti-inflammatory factor IL-4 was decreased,while the expression of IL-10 was up-regulated.The expression level of chemokine IL-2 decreased,and the difference was extremely significant compared with the control group?P<0.01?.This indicates that FAdV-4 infection of chicken embryo liver causes inflammation in the liver and changes the expression of related inflammatory cytokines.3.In order to further explore the role of nf-kappa B inflammatory signaling pathway in chicken embryos infected with FAdV-4,qRT-PCR was used to detect the mRNA expression level of the inflammatory signaling pathway related to FAdV-4 in the liver after infection with chicken embryos.The results showed that the expression levels of TLR3 and TLR4 in chicken embryo liver were up-regulated after FAdV-4 infection,and the differences were extremely significant compared with the control group?P<0.01?,and the expression levels of TLR2 were not significantly changed.The expression of NF-?B p65 was up-regulated.Expressions of cytokines-pro-inflammatory cytokines IL-1?and IL-6 were up-regulated at the downstream of the NF-?B signaling pathway.The expressions of cytokines--chemokine IL-8,IL-12?and IL-12?at the downstream of the NF-?B signaling pathway were up-regulated,and the difference was extremely significant compared with that of the control group?P<0.01?.It indicates that FAdV-4 can activate the expression of TLR4 and TLR3 in the liver cell of chicken embryo,mediate the activation of inflammatory signal pathway of NF-?B,promote the expression of pro-inflammatory factors IL-1?,IL-6,and chemokine IL-8,IL-12?and IL-12?,thereby increasing vascular permeability,vascular endothelial swelling,stasis and necrosis,and promoting the occurrence and development of inflammation.In summary,after chicken embryo was infected with FAdV-4,substantial hepatitis lesions occurred in the liver,with changes in the expression of a variety of inflammatory cytokines in the liver tissue,among which the inflammatory signal pathway of NF-?B was also activated,mediating the production of a large number of inflammatory cytokines,inducing the inflammatory response in the liver,leading to inflammatory injury in the chicken embryo liver.This study provides a theoretical basis for the development of symptomatic cytokine therapy.If the release of inflammatory cytokines can be reduced during the treatment,the injury of hepatitis caused by FAdV-4 can be effectively improved,so as to protect the liver and reduce the death caused by liver injury.