鉤端螺旋體對(duì)各種宿主細(xì)胞及ECM分子的黏附和作用
發(fā)布時(shí)間：2018-08-18 11:41
【摘要】： 目的:了解問(wèn)號(hào)鉤端螺旋體(簡(jiǎn)稱(chēng)鉤體)對(duì)宿主細(xì)胞及細(xì)胞外基質(zhì)(ECM)分子的黏附作用及其差異。了解多種宿主細(xì)胞ECM分子的種類(lèi)及其分布差異。 方法:建立問(wèn)號(hào)鉤體黃疸出血群賴(lài)型56601株對(duì)J774A.1、L929和Vero細(xì)胞感染模型,采用Fontana鍍銀染色法及顯微鏡檢查法,了解問(wèn)號(hào)鉤體56601株黏附細(xì)胞ECM的效果。建立ELISAs,檢測(cè)問(wèn)號(hào)鉤體56601株黏附ECM分子層粘連蛋白(LN)、纖維纖連蛋白(FN)、核心蛋白多糖(DEN)及膠原蛋白1、2、4(COL1,2,4)的作用,采用競(jìng)爭(zhēng)性抑制試驗(yàn)對(duì)上述實(shí)驗(yàn)結(jié)果進(jìn)行驗(yàn)證。采用免疫熒光染色法和激光共聚焦顯微鏡觀察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細(xì)胞中的分布。建立各種ELISAs定性驗(yàn)證以上結(jié)果并定量檢測(cè)以上ECM分子在各細(xì)胞株的含量 結(jié)果問(wèn)號(hào)鉤體56601株能以一端或兩端黏附于L929、J774A.1和Vero細(xì)胞的ECM部位。問(wèn)號(hào)鉤體56601株可與上述6種ECM分子發(fā)生黏附,其中對(duì)COL1、LN、COL4的黏附作用較強(qiáng)。問(wèn)號(hào)鉤體56601株分別與上述6種ECM分子預(yù)孵育后,對(duì)相應(yīng)ECM分子的黏附作用明顯下降。各細(xì)胞ECM分子種類(lèi)和含量存在明顯差異。人單核-巨噬細(xì)胞THP-1和小鼠單核-巨噬樣細(xì)胞J774A.1僅含有FN。人腎上皮細(xì)胞HEK293含有FN和COL4,大鼠腎上皮細(xì)胞NRK含有FN、LN,其中FN含量較高。猴腎成纖維細(xì)胞COS-7含有LN、COL3和COL4,其中LN和COL4含量較高。大鼠腎成纖維細(xì)胞NRK-49F含有FN、LN、DEN、COL3和COL4,其中FN、DEN和COL4含量較高。人肝上皮細(xì)胞L-02含有FN、LN、COL3和COL4,其中FN和COL4含量較高,而小鼠肝上皮細(xì)胞IAR-20僅含有COL4。人靜脈內(nèi)皮細(xì)胞HUVEC含有FN、LN、COL3和COL4。 結(jié)論ECM是問(wèn)號(hào)鉤體黏附宿主細(xì)胞的主要部位。LN、FN、DEN、COL1、COL2、COL4均是問(wèn)號(hào)鉤體黏附的受體分子,但問(wèn)號(hào)鉤體對(duì)不同ECM分子的黏附效果有一定差異。不同的ECM分子分布在不同的問(wèn)號(hào)鉤體宿主細(xì)胞中,且含量存在明顯差異。
[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.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R377
本文編號(hào)：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.
【學(xué)位授予單位】：浙江大學(xué)
【學(xué)位級(jí)別】：碩士
【學(xué)位授予年份】：2010
【分類(lèi)號(hào)】：R377
【參考文獻(xiàn)】
相關(guān)期刊論文 前1條
1 任士飛,徐建生,董國(guó)雄,李俊寶;細(xì)菌黏附研究進(jìn)展[J];中國(guó)預(yù)防獸醫(yī)學(xué)報(bào);2004年03期
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