Kops S K, Lowe D K, Bement W M, West A B
Department of Laboratory Medicine, Yale University School of Medicine, New Haven, CT 06520, USA.
Microbiol Immunol. 1996;40(11):799-811. doi: 10.1111/j.1348-0421.1996.tb01145.x.
This study characterizes the transmigration of enteroinvasive Salmonella typhi in vitro, using a human intestinal epithelial cell line as a model of small intestinal epithelium. C2BBe cells, a subclone of CACO-2 with a highly differentiated enterocytic phenotype, were grown to maturity on Transwell filters. S. typhi Ty2 and the vaccine strain, Ty21a, the S. typhi mutant X7344 and parent strain SB130, and S. typhimurium 5771 in logarithmic phase were introduced to the upper chamber of the filter units. Numbers of bacteria in the lower chamber, TER and permeability of the monolayer to mannitol were measured over time. Monolayers were examined by light, electron and confocal microscopy to determine the pathway of bacterial transmigration, and intracellular bacteria were estimated by gentamicin assay. Epithelial cell injury was quantified by light microscopy. S. typhi transmigrated earlier and in larger numbers than S. typhimurium, inducing marked changes in electrical resistance and permeability. Unlike S. typhimurium, S. typhi selected epithelial cells in small number and caused their death and extrusion from the monolayers leaving holes through which S. typhi transmigrated. Ty2 consistently transmigrated in larger numbers and with more injury to monolayers than Ty21a. S. typhi crosses the monolayers of C2BBe cells by a paracellular route in contrast to the transcellular pathway described for other Salmonellae. This may be related to the unique pathophysiology of S. typhi infection and the restricted host specificity of this pathogen. In these assays the vaccine strain, Ty21a, is slightly less invasive than its parent, though more invasive than S. typhimurium.
本研究以人肠道上皮细胞系作为小肠上皮模型，对肠侵袭性伤寒沙门氏菌的体外迁移特性进行了表征。C2BBe细胞是具有高度分化肠细胞表型的CACO-2亚克隆，在Transwell滤器上生长至成熟。将对数期的伤寒沙门氏菌Ty2和疫苗株Ty21a、伤寒沙门氏菌突变体X7344及其亲本菌株SB130，以及鼠伤寒沙门氏菌5771接种到滤器单元的上室。随时间测量下室中的细菌数量、跨上皮电阻（TER）和单层对甘露醇的通透性。通过光学显微镜、电子显微镜和共聚焦显微镜检查单层细胞，以确定细菌迁移途径，并通过庆大霉素测定法估算细胞内细菌数量。通过光学显微镜对上皮细胞损伤进行定量。伤寒沙门氏菌比鼠伤寒沙门氏菌更早且以更大数量迁移，导致电阻和通透性发生显著变化。与鼠伤寒沙门氏菌不同，伤寒沙门氏菌选择少量上皮细胞并导致其死亡和从单层中挤出，留下孔洞供伤寒沙门氏菌迁移。与Ty21a相比，Ty2始终以更大数量迁移且对单层细胞造成更多损伤。与其他沙门氏菌所描述的跨细胞途径相反，伤寒沙门氏菌通过细胞旁途径穿过C2BBe细胞单层。这可能与伤寒沙门氏菌感染的独特病理生理学以及该病原体有限的宿主特异性有关。在这些试验中，疫苗株Ty21a的侵袭性略低于其亲本，但比鼠伤寒沙门氏菌更具侵袭性。
