空肠弯曲菌致GBS动物模型的建立及模式动物体内与GBS相关基因的挖掘

发布时间：2018-06-19 07:51

本文选题：空肠弯曲菌 + GBS　；参考：《扬州大学》2012年硕士论文

【摘要】：空肠弯曲菌Campylobacter jejuni(CJ),是人类肠胃炎的最主要的致病菌,临床约90%的急性细菌肠炎均与该菌相关。在欧美等国,其致病率已超过沙门氏菌、志贺氏菌、O157大肠杆菌等。CJ除了引起肠炎之外它还是是吉兰-巴雷综合征Guillain-Barre syndrome (GBS)的主要诱因之一,有报道指出,在CJ感染之后人类GBS的发病率会提升100倍。GBS又称急性炎症性脱髓鞘性多发性神经病(AIDP),是神经系统由体液和细胞共同介导的单向性自身免疫性周围神经病,死亡率约5～10%。鉴于CJ极高的感染率以及后续的潜在威胁,已受到人们的密切关注。本研究利用巴马小型猪构建空肠弯曲菌诱导GBS的动物模型,挖掘GBS模式动物中与病发相关的基因。 1空肠弯曲菌致GBS动物模型的建立选取致人GBS源空肠弯曲菌株,采用3×1012cfu/ml、3×1011cfu/ml、3×1010cfu/ml、3×109cfu/ml等浓度对4组巴马小型猪灌胃攻毒,每头猪攻毒10ml,同时设立对照组。从第二周开始定期采集组织样及血液样本,通过免疫学检测及组织病理切片判定病变情况。免疫学检测发现,在16份攻毒组样品中有12份血清中GM1-IgG抗体为检测阳性,阳性检出率为81.25%。但是,并没有显著的攻毒浓度梯度差异。病理组织切片H/E染色显示,抗体阳性试验猪的大脑出现了噬神经元、小血管周围间隙增宽、神经中央染色溶解等现象；切片LFB染色显示,在发病模型动物的小脑白质和腰膨大出现了轻度脱髓鞘现象。结果表明空肠弯曲菌诱导GBS的浓度在3X109cfVml-3X1012cfu/ml均可,组织样本及血液采集在攻毒后4～5周较理想。 2空肠弯曲菌致GBS病发时宿主体内相关基因的研究根据病理组织切片和免疫学检测结果,选取典型发病样本和对照组样本的神经组织。通过猪全基因组表达谱芯片检测得到差异探针2156个(P0.05),其中上调差异探针896条,下调差异探针1260条,这些探针涉及到的基因共222个,其中上调基因114个,下调基因108个。基于Gene Ontology数据库对222个差异基因进行分析,得到显著基因功能(GO)130个(P0.05),其中上调基因显著性GO55个,下调基因显著性GO75个；基于KEGG数据库,分析所有差异基因参与的信号通路(Pathway),得到相关Pathway共127项,其中上调基因相关的显著性Pathway30项,下调基因相关的显著性Pathway37项；采用显著性GO内所含的基因与显著性Pathway所含的基因,对二者取交集,使用交集内的基因分别构建病变组和对照组的共表达网络,最终得到21个关键基因。其中HPRT1、PKC和PPARGC-1等处于网络图的核心位置,PPARGC-1、SUS2DD、AMPKA2等11个关键基因在病发时都表现出显著性被抑制的状态。
[Abstract]:Campylobacter jejunii CJ is the main pathogen of human gastroenteritis, and 90% of acute bacterial enteritis is associated with it. In Europe, America and other countries, the pathogenic rate of Guillain-Barre syndrome is higher than that of Salmonella. Besides causing enteritis, E. coli. CJ of Shigella spp. O157 is also one of the main causes of Guillain-Barre syndrome. After CJ infection, the incidence of GBS in human will increase 100 times. GBS is also called acute inflammatory demyelinating polyneuropathy (AIDP). It is a unidirectional autoimmune peripheral neuropathy mediated by body fluid and cells in the nervous system. The death rate is about 510g. In view of the extremely high infection rate of CJ and its potential threat, people have paid close attention to it. In this study, the animal model of GBS induced by Campylobacter jejuni was constructed from Bama miniature pig. 1 the establishment of Campylobacter jejuni animal model induced by Campylobacter jejuni caused by human Campylobacter jejuni. 3 脳 1012cfur / ml 3 脳 1011cfuml 3 脳 1010cfuml / ml 3 脳 1010cfuml / ml 3 脳 10 10cfuml / ml 3 脳 109cfu/ml were administered to 4 groups of small pigs. Each pig was treated with 10 ml of virus, and the control group was set up at the same time. Tissue samples and blood samples were collected regularly from the second week, and pathological changes were determined by immunological examination and histopathological section. Immunological examination showed that the GM1-IgG antibody was positive in 12 of the 16 samples of the group, and the positive rate was 81.25%. However, there was no significant difference in concentration gradient. Histopathological sections showed that phagocytic neurons were present in the brain of pigs with positive antibodies, the spaces around small vessels were widened, and central nerve staining was dissolving, and LFB staining showed that, Mild demyelination of cerebellar white matter and lumbar swelling was observed in the model animals. The results showed that the concentration of GBS induced by Campylobacter jejuni was at 3X109cfVml-3X1012cfur / ml. Tissue samples and blood samples were collected at 4 to 5 weeks after attack. 2 the study of genes related to GBS caused by Campylobacter jejuni according to the results of histopathological sections and immunological examination. The nerve tissue of the typical disease and the control group were selected. A total of 2156 differentially expressed probes were detected by porcine genomic expression microarray, of which 896 were up-regulated and 1260 were down-regulated. These probes involved 222 genes, including 114 up-regulated genes and 108 down-regulated genes. Based on the analysis of 222 differentially expressed genes based on Gene Ontology database, 130 significant gene functions (P0.05) were obtained, of which 55 genes were up-regulated and 75 genes were down-regulated, which were based on KEGG database. A total of 127 related Pathway genes were obtained by analyzing the signal pathways involved in all the differentially expressed genes, among which 30 were up-regulated and 37 were down-regulated, and the genes contained in significant go and those contained in significant Pathway were used. The gene in the intersection was used to construct the coexpression network of the diseased group and the control group. Finally, 21 key genes were obtained. Among them, 11 key genes such as HPRT1PKC and PPARGC-1, such as PPARGC-1, SUS2DDD, AMPKA2 and so on, showed significant inhibition at the onset of the disease.
【学位授予单位】：扬州大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R-332;R379.6

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