肉鸡感染FAdV-4后肝脏组织损伤及炎症相关基因表达研究
发布时间：2018-12-16 19:57
【摘要】：禽腺病毒Ⅰ群血清4型(Fowl adenovirus serotype 4,FAdV-4)病毒感染可引起家禽心包积液综合征(Hydropericardium syndrome,HPS)。肉鸡感染FAdV-4后剖检病变主要表现为心包腔内充盈大量黄色液体、肝脏肿大和坏死。肝脏最典型的病理组织学变化是肝细胞核内出现嗜碱性包涵体。肝脏是FAdV-4主要的靶器官,然而目前关于FAdV-4对肉鸡肝脏炎症损伤机理的研究鲜有报道。本研究用FAdV-4人工感染3周龄白羽艾维茵肉鸡,HE染色观察病毒感染后肝脏的组织病变;采用real-time PCR检测不同感染时间点的肝脏病毒载量;应用real-time RT-PCR检测肝脏炎症相关基因在不同感染时间的表达变化。目的是揭示肉鸡感染FAdV-4致使肝脏炎症损伤的机理。试验结果如下:1.FAdV-4分离株人工感染能引起非免健康白羽艾维茵肉鸡发生心包积液综合征。用FAdV-4人工感染非免健康肉鸡,分别于1 dpi、3 dpi、4 dpi、5 dpi、7 dpi、9 dpi、11dpi、15 dpi颈部放血处死8只感染肉鸡,按无菌操作采取肝脏。FAdV-4感染肉鸡在4 dpi天出现死亡,5 dpi病鸡出现精神萎靡、食欲减退等。剖检病鸡可见心包腔内充满大量黄色液体和肝坏死。病鸡的肝脏HE染色可见大量炎性细胞浸润,肝细胞变性、坏死,肝细胞核内嗜碱性包涵体。2.建立了检测FAdV-4 Hexon基因real-time PCR的方法。参照GenBank中FAdV-4保守序列Hexon设计引物,建立了检测FAdV-4 Hexon基因的real-time PCR的方法。并从感染FAd V-4不同时间点肉鸡肝脏中均检测出该病毒,病毒载量在4 dpi达到峰值。3.采用real-time RT-PCR检测了FAdV-4感染后不同时间点肉鸡肝脏中主要炎症相关基因mRNA表达变化。IL-2基因在肝脏中极显著上调(P0.001),IL-6/IFN-β基因上调;IFN-α基因在感染初期下调,在5 dpi显著上调(P0.01);CXCLi1、CXCLi2和CCR5基因都表现上调,其中CXCLi2基因上调倍数最高(P0.001)。IL-4和IL-10基因均呈上调趋势,其中IL-4基因在第3 dpi,IL-10基因在4 dpi极显著上调(P0.001);TGF-β1和TGF-β2基因在1 dpi下调(P0.01),随后上调,其中TGF-β2基因在4 dpi显著上调(P0.001)。综上所述,FAdV-4感染白羽艾维茵肉鸡后能造成肝脏严重病理损伤,引起肝脏组织中炎症相关基因mRNA的表达发生明显变化。试验结果为揭示FAdV-4感染肉鸡肝脏组织炎症损伤机制提供了试验数据。
[Abstract]:Avian adenovirus group I serotype 4 (Fowl adenovirus serotype 4 (FAdV-4) virus infection may cause pericardial effusion syndrome (Hydropericardium syndrome,HPS) in poultry. The pathological changes after FAdV-4 infection in broilers were mainly yellow fluid in pericardial cavity, hepatomegaly and necrosis. The most typical histopathological changes of the liver are basophilic inclusion bodies in the nucleus of the liver. Liver is the main target organ of FAdV-4. However, there are few reports on the mechanism of liver inflammation injury caused by FAdV-4 in broilers. In this study, three weeks old white feather Avian broilers infected with FAdV-4 were used to observe the liver pathological changes by HE staining, and real-time PCR was used to detect the viral load of the liver at different time points. Real-time RT-PCR was used to detect the expression of inflammatory genes in liver at different infection time. The aim of this study was to elucidate the mechanism of liver inflammatory injury caused by FAdV-4 infection in broilers. The results were as follows: artificial infection of 1.FAdV-4 isolate could induce pericardial effusion syndrome in non-immune white feather Avian broilers. Non-immune broilers were artificially infected with FAdV-4, and 8 infected broilers were killed in 1 dpi,3 dpi,4 dpi,5 dpi,7 dpi,9 dpi,11dpi,15 dpi neck bloodletting. FAdV-4 infected broilers died on 4 dpi day, 5 dpi infected broilers suffered from mental retardation and anorexia. A large amount of yellow fluid and liver necrosis were found in the pericardial cavity of the diseased chicken. A large number of inflammatory cell infiltration, hepatocyte degeneration, necrosis and basophilic inclusion bodies were observed in liver HE staining of diseased chicken. A method for detecting real-time PCR of FAdV-4 Hexon gene was established. According to the FAdV-4 conserved sequence Hexon in GenBank, the primers were designed and a method for detecting real-time PCR of FAdV-4 Hexon gene was established. The virus was detected from the liver of broilers infected with FAd V-4 at different time points, and the viral load reached a peak value of 3. 3 at 4 dpi. Real-time RT-PCR was used to detect the changes of mRNA expression of the major inflammatory related genes in the liver of broilers at different time points after FAdV-4 infection. IL-2 gene was significantly up-regulated (P0.001) and IL-6/IFN- 尾 gene was up-regulated in the liver. IFN- 伪 gene was down-regulated at the early stage of infection and significantly up-regulated at 5 dpi (P0.01). Both CXCLi1,CXCLi2 and CCR5 genes were up-regulated, among which CXCLi2 gene had the highest up-regulation (P0.001). Both IL-4 and IL-10 genes were up-regulated, and IL-4 gene was significantly up-regulated at 4 dpi in the 3rd dpi,IL-10 gene (P0.001). TGF- 尾 1 and TGF- 尾 2 genes were down-regulated at 1 dpi (P0.01) and then up-regulated. TGF- 尾 2 gene was significantly up-regulated at 4 dpi (P0.001). To sum up, FAdV-4 infection with Avian broilers can cause serious pathological damage of liver and change the expression of inflammatory related gene mRNA in liver tissue. The results provide experimental data for revealing the mechanism of liver inflammation injury in broilers infected with FAdV-4.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S858.31
，
本文编号：2382967
[Abstract]:Avian adenovirus group I serotype 4 (Fowl adenovirus serotype 4 (FAdV-4) virus infection may cause pericardial effusion syndrome (Hydropericardium syndrome,HPS) in poultry. The pathological changes after FAdV-4 infection in broilers were mainly yellow fluid in pericardial cavity, hepatomegaly and necrosis. The most typical histopathological changes of the liver are basophilic inclusion bodies in the nucleus of the liver. Liver is the main target organ of FAdV-4. However, there are few reports on the mechanism of liver inflammation injury caused by FAdV-4 in broilers. In this study, three weeks old white feather Avian broilers infected with FAdV-4 were used to observe the liver pathological changes by HE staining, and real-time PCR was used to detect the viral load of the liver at different time points. Real-time RT-PCR was used to detect the expression of inflammatory genes in liver at different infection time. The aim of this study was to elucidate the mechanism of liver inflammatory injury caused by FAdV-4 infection in broilers. The results were as follows: artificial infection of 1.FAdV-4 isolate could induce pericardial effusion syndrome in non-immune white feather Avian broilers. Non-immune broilers were artificially infected with FAdV-4, and 8 infected broilers were killed in 1 dpi,3 dpi,4 dpi,5 dpi,7 dpi,9 dpi,11dpi,15 dpi neck bloodletting. FAdV-4 infected broilers died on 4 dpi day, 5 dpi infected broilers suffered from mental retardation and anorexia. A large amount of yellow fluid and liver necrosis were found in the pericardial cavity of the diseased chicken. A large number of inflammatory cell infiltration, hepatocyte degeneration, necrosis and basophilic inclusion bodies were observed in liver HE staining of diseased chicken. A method for detecting real-time PCR of FAdV-4 Hexon gene was established. According to the FAdV-4 conserved sequence Hexon in GenBank, the primers were designed and a method for detecting real-time PCR of FAdV-4 Hexon gene was established. The virus was detected from the liver of broilers infected with FAd V-4 at different time points, and the viral load reached a peak value of 3. 3 at 4 dpi. Real-time RT-PCR was used to detect the changes of mRNA expression of the major inflammatory related genes in the liver of broilers at different time points after FAdV-4 infection. IL-2 gene was significantly up-regulated (P0.001) and IL-6/IFN- 尾 gene was up-regulated in the liver. IFN- 伪 gene was down-regulated at the early stage of infection and significantly up-regulated at 5 dpi (P0.01). Both CXCLi1,CXCLi2 and CCR5 genes were up-regulated, among which CXCLi2 gene had the highest up-regulation (P0.001). Both IL-4 and IL-10 genes were up-regulated, and IL-4 gene was significantly up-regulated at 4 dpi in the 3rd dpi,IL-10 gene (P0.001). TGF- 尾 1 and TGF- 尾 2 genes were down-regulated at 1 dpi (P0.01) and then up-regulated. TGF- 尾 2 gene was significantly up-regulated at 4 dpi (P0.001). To sum up, FAdV-4 infection with Avian broilers can cause serious pathological damage of liver and change the expression of inflammatory related gene mRNA in liver tissue. The results provide experimental data for revealing the mechanism of liver inflammation injury in broilers infected with FAdV-4.
【学位授予单位】：西北农林科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S858.31
，
本文编号：2382967
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