钩端螺旋体对各种宿主细胞及ECM分子的黏附和作用
发布时间：2018-08-18 11:41
【摘要】： 目的:了解问号钩端螺旋体(简称钩体)对宿主细胞及细胞外基质(ECM)分子的黏附作用及其差异。了解多种宿主细胞ECM分子的种类及其分布差异。 方法:建立问号钩体黄疸出血群赖型56601株对J774A.1、L929和Vero细胞感染模型,采用Fontana镀银染色法及显微镜检查法,了解问号钩体56601株黏附细胞ECM的效果。建立ELISAs,检测问号钩体56601株黏附ECM分子层粘连蛋白(LN)、纤维纤连蛋白(FN)、核心蛋白多糖(DEN)及胶原蛋白1、2、4(COL1,2,4)的作用,采用竞争性抑制试验对上述实验结果进行验证。采用免疫荧光染色法和激光共聚焦显微镜观察ECM分子FN、LN、DEN、COL1～4在THP-1、J774A.1、HEK293、NRK、COS-7、NRK-49F、L-02、IAR-20和HUVEC细胞中的分布。建立各种ELISAs定性验证以上结果并定量检测以上ECM分子在各细胞株的含量 结果问号钩体56601株能以一端或两端黏附于L929、J774A.1和Vero细胞的ECM部位。问号钩体56601株可与上述6种ECM分子发生黏附,其中对COL1、LN、COL4的黏附作用较强。问号钩体56601株分别与上述6种ECM分子预孵育后,对相应ECM分子的黏附作用明显下降。各细胞ECM分子种类和含量存在明显差异。人单核-巨噬细胞THP-1和小鼠单核-巨噬样细胞J774A.1仅含有FN。人肾上皮细胞HEK293含有FN和COL4,大鼠肾上皮细胞NRK含有FN、LN,其中FN含量较高。猴肾成纤维细胞COS-7含有LN、COL3和COL4,其中LN和COL4含量较高。大鼠肾成纤维细胞NRK-49F含有FN、LN、DEN、COL3和COL4,其中FN、DEN和COL4含量较高。人肝上皮细胞L-02含有FN、LN、COL3和COL4,其中FN和COL4含量较高,而小鼠肝上皮细胞IAR-20仅含有COL4。人静脉内皮细胞HUVEC含有FN、LN、COL3和COL4。 结论ECM是问号钩体黏附宿主细胞的主要部位。LN、FN、DEN、COL1、COL2、COL4均是问号钩体黏附的受体分子,但问号钩体对不同ECM分子的黏附效果有一定差异。不同的ECM分子分布在不同的问号钩体宿主细胞中,且含量存在明显差异。
[Abstract]:Aim: to investigate the adhesion of Leptospira interrogans (Leptospira interrogans) to host cells and extracellular matrix (ECM) molecules. To understand the species and distribution of ECM molecules in various host cells. Methods: the model of infection of J774A.1nL929 and Vero cells was established by 56601 strains of Leptospira interrogans with jaundice hemorrhage. The effect of ECM on the adhesion cells of 56601 strains of Leptospira interrogans was studied by Fontana silver staining and microscope examination. Elisas was established to detect the effects of ECM laminin (LN), fibronectin (FN), core proteoglycan (DEN) and collagen1 (1 FN 2) 4 (Coll 1) on the adhesion of Leptospira interrogans strain 56601. The results were verified by competitive inhibition test. Immunofluorescence staining and laser confocal microscopy were used to observe the distribution of ECM molecule ECM in THP-1 J774A.1 HEK293NRK-49FN-L-02IAR-20 and HUVEC cells. All kinds of ELISAs were established to qualitatively verify the above results and to quantitatively detect the content of the above ECM molecules in each cell line. The results showed that Leptospira 56601 could adhere to the ECM site of L929J774A.1 and Vero cells at one or both ends. Leptospira interrogans strain 56601 could bind to the 6 ECM molecules mentioned above, and the adhesion to COL1 LNNCOL4 was stronger. The adhesion of Leptospira interrogans strain 56601 to the corresponding ECM molecules decreased significantly after preincubation with the six ECM molecules mentioned above. There were significant differences in the types and contents of ECM molecules among different cells. Human monocyte-macrophage THP-1 and murine monocyte-macrophage J774A.1 contained only FNs. Human renal epithelial cells (HEK293) and rat renal epithelial cells (NRK) contained FN and COL4, and FN content was higher in NRK. The COS-7 of monkey renal fibroblasts contained LNCOL3 and COL4, and the contents of LN and COL4 were higher. The NRK-49F of rat renal fibroblasts contained FNNLNDEN3 and COL4, in which FNNNDEN and COL4 were higher. Human liver epithelial cells L-02 contained FNLNnncCOL3 and COL4, among which FN and COL4 were higher, while IAR-20 of mouse hepatocytes contained only COL4. Human vein endothelial cells (HUVEC) contain FNLNCol 3 and COL4. Conclusion ECM is the main part of Leptospira which adheres to host cells. All of them are receptor molecules of Leptospira question Leptospira, but the effect of Leptospira interrogans on the adhesion of different Leptospira molecules to different ECM molecules is different. Different ECM molecules were distributed in different host cells of Leptospira interrogans, and the content of Leptospira was significantly different.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R377
本文编号：2189360
[Abstract]:Aim: to investigate the adhesion of Leptospira interrogans (Leptospira interrogans) to host cells and extracellular matrix (ECM) molecules. To understand the species and distribution of ECM molecules in various host cells. Methods: the model of infection of J774A.1nL929 and Vero cells was established by 56601 strains of Leptospira interrogans with jaundice hemorrhage. The effect of ECM on the adhesion cells of 56601 strains of Leptospira interrogans was studied by Fontana silver staining and microscope examination. Elisas was established to detect the effects of ECM laminin (LN), fibronectin (FN), core proteoglycan (DEN) and collagen1 (1 FN 2) 4 (Coll 1) on the adhesion of Leptospira interrogans strain 56601. The results were verified by competitive inhibition test. Immunofluorescence staining and laser confocal microscopy were used to observe the distribution of ECM molecule ECM in THP-1 J774A.1 HEK293NRK-49FN-L-02IAR-20 and HUVEC cells. All kinds of ELISAs were established to qualitatively verify the above results and to quantitatively detect the content of the above ECM molecules in each cell line. The results showed that Leptospira 56601 could adhere to the ECM site of L929J774A.1 and Vero cells at one or both ends. Leptospira interrogans strain 56601 could bind to the 6 ECM molecules mentioned above, and the adhesion to COL1 LNNCOL4 was stronger. The adhesion of Leptospira interrogans strain 56601 to the corresponding ECM molecules decreased significantly after preincubation with the six ECM molecules mentioned above. There were significant differences in the types and contents of ECM molecules among different cells. Human monocyte-macrophage THP-1 and murine monocyte-macrophage J774A.1 contained only FNs. Human renal epithelial cells (HEK293) and rat renal epithelial cells (NRK) contained FN and COL4, and FN content was higher in NRK. The COS-7 of monkey renal fibroblasts contained LNCOL3 and COL4, and the contents of LN and COL4 were higher. The NRK-49F of rat renal fibroblasts contained FNNLNDEN3 and COL4, in which FNNNDEN and COL4 were higher. Human liver epithelial cells L-02 contained FNLNnncCOL3 and COL4, among which FN and COL4 were higher, while IAR-20 of mouse hepatocytes contained only COL4. Human vein endothelial cells (HUVEC) contain FNLNCol 3 and COL4. Conclusion ECM is the main part of Leptospira which adheres to host cells. All of them are receptor molecules of Leptospira question Leptospira, but the effect of Leptospira interrogans on the adhesion of different Leptospira molecules to different ECM molecules is different. Different ECM molecules were distributed in different host cells of Leptospira interrogans, and the content of Leptospira was significantly different.
【学位授予单位】：浙江大学
【学位级别】：硕士
【学位授予年份】：2010
【分类号】：R377
【参考文献】
相关期刊论文 前1条
1 任士飞,徐建生,董国雄,李俊宝;细菌黏附研究进展[J];中国预防兽医学报;2004年03期
，本文编号：2189360
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