弓形虫感染对小鼠、猪脾肝基因表达的影响

发布时间：2018-01-15 06:33

本文关键词：弓形虫感染对小鼠、猪脾肝基因表达的影响　出处：《中国农业科学院》2016年博士论文　论文类型：学位论文

【摘要】：弓形虫是一种寄生于真核细胞内的顶复门原虫。它能通过其自身蛋白改变宿主细胞信号通路状态、调节宿主基因的表达量,进而影响宿主的生命活动进程。本课题通过RNA-seq和iTRAQ技术、生物信息学分析、定量PCR验证等技术,对弓形虫感染后小鼠和猪的脾肝转录组、miRNA和蛋白组进行了研究。通过对小鼠感染弓形虫后脾脏的转录组进行分析,发现弓形虫感染能引起脾脏的免疫相关通路或组分上调。免疫细胞在脾脏中的含量可能会上升,因为趋化这些免疫细胞移动的趋化因子在脾脏中表达上调。此外,脾脏细胞的细胞器组分在弓形虫感染过程中发生改变,包括线粒体和内质网组分。通过检测小鼠脾脏感染弓形虫后miRNA的表达量,我们发现在小鼠脾脏差异表达的miRNA中,既有靶向转录因子的miRNA,也有靶向凋亡通路的miRNA。这提示弓形虫可以通过改变靶向转录因子的miRNA表达量来间接地调节宿主基因的表达。同时,弓形虫也可以通过改变靶向细胞凋亡相关基因的miRNA表达来调控宿主细胞凋亡进程;靶向凋亡相关基因的结果则取决于促进凋亡和抑制凋亡的miRNA之间的平衡。通过对小鼠感染弓形虫后的肝脏蛋白组进行检测和分析,我们发现小鼠肝脏表达上调的蛋白集中在免疫、感染以及蛋白折叠相关通路,而下调的蛋白则集中在代谢通路。对小鼠肝脏转录组分析结果表明,iTRAQ检测的结果准确、可靠。此外,通过分析转录组数据我们还发现,肝脏胆汁合成和分泌通路在弓形虫感染中被显著下调。通过转录因子富集,我们发现下调的基因有很大一部分是由PPAR转录因子控制的。PPAR信号通路与肝脏能量、脂类等代谢有密切关系;因此弓形虫可能通过劫持肝细胞的PPAR通路来引起肝脏代谢功能下调。由于猪的生理特点比小鼠更加接近于人,故本课题还对弓形虫感染猪后肝脏和脾脏的转录组变化进行了检测和分析。通过对肝脏、脾脏差异表达转录本的表达模式进行聚类,发现猪肝脏和脾脏的代谢进程都在弓形虫感染中下调,尤其是在感染后第6天的肝脏。肝脏中的炎症反应总体上要强于脾脏,尤其是感染后第18天猪肝脏出现了结缔组织增生及炎症细胞浸润的病理变化;猪肝脏在感染后第6天没有发现明显的病理变化,但小鼠肝脏在感染后第6天却发现了明显的病理变化。这可能是由于猪和小鼠之间的免疫系统和生理系统差异以及对弓形虫不同的易感性造成的。此外,我们还发现猪肝脏蛋白折叠和蛋白质量控制相关基因的表达受到弓形虫感染的影响,这与小鼠模型的研究结果一致。综上所述,本课题首次对弓形虫感染后小鼠脾脏的转录组、miRNA,小鼠肝脏的蛋白组、转录组以及猪脾脏和肝脏的转录组进行了研究和分析,描述了小鼠和猪感染弓形虫后脾脏和肝脏的基因表达变化。同时,我们的研究结果也证明了弓形虫感染对宿主基因表达的影响与感染进程以及宿主属性有关。本课题研究结果将为进一步深入研究弓形虫感染对宿主基因的调节、脾脏和肝脏功能的改变奠定基础。
[Abstract]:Toxoplasma gondii is a parasite in eukaryotic cells in apicomplexan protozoa. It can change the host cell signaling pathways through its protein expression regulation of host genes, thereby affecting the host life activity process. In this paper by RNA-seq and iTRAQ technology, bioinformatics analysis, quantitative PCR verification technology. The spleen and liver transcriptome of mouse and porcine Toxoplasma infection, studied miRNA and protein group. Through the analysis of mice infected with Toxoplasma gondii transcriptome of spleen, found that Toxoplasma infection can induce immune related pathways or group of spleen points raised. Content of immune cells in the spleen may rise, because chemotaxis of these immune cells move chemokines in spleen expression. In addition, the spleen cell organelles change in Toxoplasma gondii infection process, including mitochondria and endoplasmic reticulum by detecting components. The expression of measuring spleen of mice infected with Toxoplasma gondii miRNA, we found that the expression of miRNA in spleen of mice in both differences, miRNA targeting transcription factor expression, also targeting the apoptosis miRNA. suggesting that Toxoplasma can be changed by targeting transcription factor miRNA expression to indirectly regulate host genes. At the same time, Toxoplasma can shift targeting apoptosis related gene expression of miRNA in the regulation of host cell apoptosis process; targeting apoptosis related genes depends on the results between promote and inhibit apoptosis of miRNA balance. Through testing and Analysis on the liver protein group of mice infected with Toxoplasma gondii, we found that the mice the protein concentration in the liver increased expression of immunity, infection and protein folding pathway, the downregulated proteins are concentrated in metabolic pathways. That of mouse liver transcriptome analysis, iT RAQ detection result is accurate and reliable. In addition, through the analysis of transcriptome data we also found that the synthesis and secretion pathway was significantly downregulated in Toxoplasma gondii infection in the liver bile. Through transcription factor enrichment, we found that the down regulated genes are a large part of the.PPAR signaling pathway and liver energy controlled by the PPAR transcription factor, a closely related to lipid metabolism; therefore, Toxoplasma possibly via the PPAR pathway in liver cells to hijack cause liver metabolic function down. Because of the physiological characteristics of pigs closer than in mice, so this research on swine Toxoplasma infection after liver and spleen transcriptome changes were detected and analyzed. The liver was the expression pattern of clustering transcripts expression differences, found that the metabolic process of pig liver and spleen are downregulated in Toxoplasma infection, especially in sixth days after infection of the liver in the liver inflammation. In response to general than the spleen, especially after infection appeared pathological changes of connective tissue hyperplasia and infiltration of inflammatory cells on the eighteenth day of pig liver; pig liver sixth days found no obvious pathological changes in the liver of mice after infection, but in sixth days after infection and found obvious pathological changes. This may be due to the difference between the immune system of pigs and mice and physiological systems and different susceptibility to Toxoplasma gondii. In addition, we also found that the pig liver protein folding and protein quality control related gene expression affected by Toxoplasma gondii infection, the mice model research and the consistency of the results. To sum up, this is the first study on the transcriptome of mouse spleen the Toxoplasma gondii infection after miRNA protein in mouse liver, transcriptome and transcriptome of porcine spleen and liver were studied and analyzed, described mice and pigs infected with bow shaped worm The expression changes of spleen and liver genes. At the same time, our results also show that the related infection process and host attributes affect the expression of host gene of Toxoplasma gondii infection. The results of this study will further study on Toxoplasma infection in regulation of host genes, lay the foundation for the spleen and liver function change.

【学位授予单位】：中国农业科学院
【学位级别】：博士
【学位授予年份】：2016
【分类号】：S855.9

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