ALV-J对鸡骨髓源树突状细胞生物学功能影响的研究

发布时间：2018-06-25 11:08

本文选题：ALV-J + 免疫抑制　；参考：《华南农业大学》2015年博士论文

【摘要】：禽白血病是由反转录病毒科甲型反转录病毒属禽反转录病毒引起的一种禽类多种肿瘤性疾病的统称。此病自发现以后就在世界各国鸡群中广泛流行,主要通过垂直传播并引发肿瘤产生,导致鸡死亡和消瘦、生长发育不良,同时引起机体抵抗力下降和免疫抑制,是危害养禽业的主要疾病之一。由于对禽白血病的研究热点往往都集中在病毒的流行病学,致病性,致瘤和分子结构与功能方面,对于其导致免疫抑制的机制研究的并不十分清楚。本研究通过体外诱导培养免疫细胞分化和发育成熟的技术,研究和分析了ALV-J在树突状细胞分化、发育和成熟的过程中所造成的影响。由于树突状细胞的制作和分化过程研究的比较清楚,与其在抗原递呈和抗肿瘤方面的特殊地位,我们选择了建立体外培养并诱导分化的树突状细胞培养方法,并以此为基础,研究ALV-J对其分化成熟的影响。为了研究病毒是否能顺利感染分化中不同状态的树突状细胞,根据细胞培养和病毒培养所需的时间,在细胞分化培养的第1 d、2 d、4 d、6 d分别接种ALV-J。结果发现,在第1d、2d、4d病毒可以成功感染细胞,第六天接毒的样品,病毒感染失败。在实验过程中发现在接毒后的5-6天内,细胞的生长,分化情况并无明显差异。在接毒6天以后,细胞分化并接近成熟的时期,细胞数量开始减少,并且出现了形态变化。后经细胞凋亡实验发现,ALV-J可以导致细胞在分化的后期接近成熟时,细胞开始过早的凋亡。对部分TLR和细胞因子的mRNA表达水平的检测结果也显示,大部分检测的mRNA表达水平都出现了显著性下调。结果表明,ALV-J的感染抑制了树突状细胞的成熟,并且造成了树突状细胞的凋亡,并影响到了细胞的正常生物学功能。为了研究ALV-J对树突状细胞生物学功能影响的机制,进一步分析了树突状细胞的miRNA表达水平。发现了122种差异表达的miRNA。其中已知的miRNA81种,未知新发现的miRNA41种。在已知的81种差异表达的miRNA中,发现了4种与细胞免疫功能,ALV致病性有关的miRNA,分别是gga-mir-221、gga-mir-211、gga-mir-204和gga-mir-6651,这些miRNA共同调控着细胞内源性刺激应答、抗原递呈和加工、细胞凋亡和慢性骨髓白血病相关的细胞功能和通路。经miRNA表达水平的分析,这些细胞功能和代谢通路是呈激活状态的。说明,病毒感染没有直接对细胞的抗原递呈功能产生影响。同时,发现这些miRNA与慢性骨髓白血病有关,可能与ALV-J导致骨髓瘤型白血病有一定的关系。对显著性差异的细胞功能和代谢通路的分析结果发现多集中于细胞的生理代谢和能量物质代谢方面。ALV-J的感染使得这些功能的调控基因发生紊乱。细胞在分化的过程中,需要多种基因的调控,也需要营养和能量的供给。而ALV-J感染会影响到调控基因的表达,进一步影响细胞正常的生理和新陈代谢功能,从而导致细胞分化失败,活性降低和凋亡等。本研究发现ALV-J会严重影响树突状分化成熟。而这种情况可能设计大部分甚至所有免疫细胞的分化和成熟,进而导致免疫抑制。本研究首次从免疫细胞分化的角度去分析研究ALV-J导致免疫抑制的机制,对于ALV-J导致免疫抑制机制的研究,提供了一个新的研究方向,将有助于进一步全面的研究ALV-J导致免疫抑制的原因和机制。
[Abstract]:Avian leukosis is a common name for a variety of fowl diseases caused by the retrovirus of the family antiretroviral family. This disease has been widely popular among chickens in the world since its discovery, mainly through vertical transmission and initiation of cancer, resulting in death and emaciation of chickens, growth and development, and the cause of the body. The decline of resistance and immunosuppression is one of the major diseases that harm the poultry industry. The research focus on avian leukemia is often concentrated on the epidemiology, pathogenicity, tumorigenicity, molecular structure and function of the virus, and the mechanism of its immunosuppression is not very clear. This study has been induced in vitro by induction of immunization. The effects of ALV-J on the differentiation, development and maturation of dendritic cells are studied and analyzed. The study of the process and differentiation of dendritic cells is clear. We choose to establish in vitro culture and induce differentiation in specific sites of antigen presentation and antitumor. In order to study whether the virus could successfully infect different dendritic cells in the differentiation of the virus, in order to investigate whether the virus could successfully infect the dendritic cells of different states of differentiation, the results were found in first D, 2 D, 4 D, 6 D, respectively, in cell differentiation and culture, in order to study the effect of ALV-J on the differentiation and maturation of the cells. 1D, 2D, 4D virus can infect cells successfully, and the virus infection failed in sixth days. During the experiment, there was no obvious difference in the growth and differentiation of cells within 5-6 days after being poisoned. After 6 days of poison, the cell differentiation was close to the mature period, the number of cells began to decrease, and the morphological changes occurred. The apoptosis test found that ALV-J could lead to premature apoptosis when the cells were close to maturity in the late stage of differentiation. The results of mRNA expression levels of partial TLR and cytokines also showed that most of the levels of mRNA expression were significantly downregulated. The results showed that the infection of ALV-J inhibited the formation of dendritic cells. Mature, and cause the apoptosis of dendritic cells, and affect the normal biological function of the cells. In order to study the mechanism of the effect of ALV-J on the biological function of dendritic cells, the miRNA expression level of dendritic cells was further analyzed. The known miRNA81 species in 122 differentially expressed miRNA. were found, and the newly discovered miRNA41 was unknown. Species. In 81 known differentially expressed miRNA, 4 miRNA related to cellular immune function, ALV pathogenicity, are gga-mir-221, gga-mir-211, gga-mir-204 and gga-mir-6651, respectively. These miRNA co regulate the cellular power of cell endogenous stimulus response, antigen presentation and addition, cell apoptosis and chronic myelogenous leukemia. Energy and pathway. Through the analysis of miRNA expression level, these cell functions and metabolic pathways are activated. It shows that the virus infection does not directly affect the antigen presenting function of the cells. At the same time, it is found that these miRNA are associated with chronic myelogenous leukemia and may have a certain relationship with ALV-J induced myeloma leukemia. The analysis of different cellular and metabolic pathways found that the.ALV-J infection in the physiological metabolism of cells and the metabolism of energy materials caused disorder of the regulatory genes of these functions. In the process of differentiation, the cells needed the regulation of many genes and the supply of nutrition and energy. And the ALV-J infection would affect the regulation. The expression of controlled genes further affects the normal physiological and metabolic functions of the cells, which leads to the failure of cell differentiation, reduction of activity and apoptosis. This study found that ALV-J can seriously affect the maturation of dendritic differentiation. This situation may design most of the differentiation and maturation of all immune cells, which may lead to immunosuppression. The study for the first time from the angle of immune cell differentiation to analyze the mechanism of ALV-J induced immunosuppression provides a new direction for the study of the mechanism of ALV-J induced immunosuppression, which will help to further study the causes and mechanisms of ALV-J to lead to immunosuppression.
【学位授予单位】：华南农业大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：S858.31

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