大肠杆菌侵袭基因缺失突变株与人脑微血管内皮细胞的相互作用

Interaction of E. coli Invasion Gene Deletion Mutants with Human Brain Microvascular Endothelial Cell

ibeA、ibeB、ibeC是与大肠杆菌侵袭人脑微血管内皮细胞(HBMEC)密切相关的基因,但迄今各基因的功能并不清楚。应用侵袭分析和免疫荧光技术分析了各基因的缺失突变型及野生型大肠杆菌对HBMEC的侵袭、细胞骨架与细胞间紧密连接的影响。结果显示:野生型菌大肠杆菌侵袭率为3.46%,而ibeA、ibeB、ibeC缺失突变株分别为0.54%、0.82%和0.73%;ibeA缺失突变型与野生型大肠杆菌作用相似,可引起HBMEC的细胞骨架蛋白分布改变,在细胞膜处呈明显的聚集,而ibeB和ibeC缺失突变株并未引起细胞骨架的明显改变;野生型和ibeA缺失突变型大肠杆菌可引起紧密连接结构的明显改变,而ibeB和ibeC缺失突变株对紧密连接结构的影响不明显。这些观察到的结果提示:ibeB和ibeC基因产物可能在调节细胞骨架和影响细胞紧密连接中起重要作用,而ibeA基因产物在其中的作用较小。

IbeA, ibeB and ibeC are important genes associated with E.coli K1 invasion to human brain microvascular endothelial cell (HBMEC), but the role of each gene in invasion process is not clear. The main purpose of this study is to make a preliminary conclusion of the function of each gene. We had HBMEC interacted with the invasion gene (ibeA, ibeB and ibeC) deletion mutants of E.coli, meanwhile the HBMEC interacted with wild type E. coli is as positive control .The invasion rate of each mutant , HBMEC cytoskeleton and tight junction changes were studied. In the invasion test, each mutant of E.coli showed remarkable low invasion rate to HBMEC in contrast to that of wild E.coli. HBMEC interacted with ibeA deletion mutant showed distinct change in cytoskeleton just same as the positive control, while, HBMEC interacted with ibeB , ibeC deletion mutants respectively showed relatively small in cytoskeletal change in contrast to negative control HBMEC. In tight junction test, HBMEC interacted with ibeA deletion mutant showed the same result as positive control HBMEC in which tight junction is heavily disturbed, HBMEC interacted with ibeB, ibeC deletion mutants respectively showed relatively small tight junction disturbance in contrast to negative control HBMEC . Taken together, we can propose that the IbeA not play any role in cytosk- eleton and tight junction regulation; IbeB and IbeC , on the contrary, play an important role in both processes .