大蹼铃蟾皮肤共生菌分离和两栖类IL-1β加工过程的研究
文献类型：学位论文
| 作者 | 严超 |
| 学位类别 | 硕士 |
| 答辩日期 | 2013-05 |
| 授予单位 | 中国科学院研究生院 |
| 授予地点 | 北京 |
| 导师 | 张云 |
| 关键词 | 大蹼铃蟾 天然免疫 皮肤共生菌 白细胞介素-1β |
| 其他题名 | Separation of Commensal Bacteria from the Bombina Maxima Skin and Study of Amphibian IL-1β Processing |
| 学位专业 | 生物化学与分子生物学 |
| 中文摘要 | 两栖类是联系哺乳动物和鱼类的重要分类单元，其中非洲爪蟾长期以来一直是重要的模式动物。随着21世纪免疫学与生态学的蓬勃发展，两栖类在研究天然免疫或生态毒理相关问题时成为可资利用的研究模型。由于皮肤裸露且生存条件多变，传统上普遍认为两栖类拥有强大的天然免疫系统，其中尤以抗菌肽、吞噬细胞以及NK细胞三类为代表。但在面对某些致死性病原体感染时，令人困惑的是不同两栖类物种在易感性和抗性上有着极大的区别。而不同的皮肤共生菌组成与体内炎症激活方式（包括了IL-1β的产生与加工）上的差异可能是两栖类免疫功能同中有异的关键所在，而这两方面的研究将对扩充我们在天然免疫学上的视野大有助益。 本研究中首先采用标准I号营养琼脂培养从大蹼铃蟾皮肤中分离能够在固体培养基中定植生长的共生菌，共得到5个不同细菌的16SrRNA近全长序列，测序后使用RDP-II进行比对鉴定，结果显示这些细菌分属变形菌门、厚壁菌门和放线菌门中5个不同的菌属。 其次将大蹼铃蟾IL-1β的部分片段进行了重组表达并得到IL-1β的多克隆抗体。以具有条件致病性的大蹼铃蟾皮肤共生菌作为病原体进行体内腹腔感染和体外脾细胞刺激实验。通过Western blot发现了大蹼铃蟾IL-1β可能的前体以及成熟肽形式。最后基于生化分析与结构比对，结合目前对于非哺乳类IL-1β转化酶酶切位点的研究进展，预测了大蹼铃蟾Caspase-1加工IL-1β的酶切位点。 |
| 英文摘要 | Amphibian that links mammals and fish is an important taxon in evolution, and Xenopus laevis is a model organism for a long time. With the development of immunology and ecology in 21st Century, amphibian is the model of choice for autarcesiology and ecotoxicology. Due to the exposed skin and the complexity of living conditions, a common thought is that amphibian has a strong innate immune system, such as antibacterial peptides, phagocytes and NK cells. Some species of amphibians are very resistant to lethal pathogen infections, whereas others are more susceptible. And the factors that determine resistance or susceptibility are not well understood. Different compositions of commensal bacteria on the skin and different patterns of inflammation activation (including IL-1β processing) in vivo could explain the contradiction above.The research for these two aspects would contribute to expand our vision in innate immunity. Commensal bacteria were separate from frog skin of Bombina maxima and cultured by Standard I nutrient medium.Then five different bacterial near full-length 16SrRNA sequences were found from cultured bacteria. After sequencing we used RDP- II to identify them, the results showed that these bacteria belonged to five different genera in Proteobacteria, Firmicutes and Actinobacteria. A partial recombinant IL-1β protein was expressed and rabbit polyclonal antibodies against Bombina maxima IL-1β was obtained. Opportunistic pathogen from frog skin was used for intra-abdominal infection and stimulation of splenocytes in vitro. Through the Western blot analysis we found probably inactive pro-IL-1β protein and a putative mature IL-1β. Based on the biochemical analysis and comparation of protein sequence and structure, combined with research progress of non-mammalian IL-1β converting enzyme cut site nowadays, we finally localized the putative Caspase1 cleavage site in Bombina maxima IL-1β. |
| 语种 | 中文 |
| 公开日期 | 2013-06-09 |
| 源URL | [http://159.226.149.42:8088/handle/152453/7460]  |
| 专题 | 昆明动物研究所_动物活性蛋白多肽组学 |
| 推荐引用方式 GB/T 7714 | 严超. 大蹼铃蟾皮肤共生菌分离和两栖类IL-1β加工过程的研究[D]. 北京. 中国科学院研究生院. 2013. |
入库方式： OAI收割
来源：昆明动物研究所
浏览0
下载0
收藏0
其他版本
除非特别说明，本系统中所有内容都受版权保护，并保留所有权利。