Luyts Katrien, Napierska Dorota, Dinsdale David, Klein Sebastian G, Serchi Tommaso, Hoet Peter H M
Department of Public Health and Primary Care, Occupational and Environmental Toxicology, KULeuven, Leuven, Belgium.
Toxicol In Vitro. 2015 Feb;29(1):234-41. doi: 10.1016/j.tiv.2014.10.024.
We developed a coculture model of the lung–blood barrier using human bronchial epithelial cells(16HBE14o-), monocytes (THP-1) and human lung microvascular endothelial cells (HLMVEC) in which several parameters can be assessed simultaneously. The epithelial and endothelial cells were grown on opposite sides of a microporous membrane. Electron and confocal microscopic pictures show the presence of the cells in their appropriate compartment and both cell types do not show evidence of growing through the pores. Out of three endothelial cell types (EAhy.926, HUVEC and HLMVEC), the last was chosen as the most appropriate cell type, best resembling the pulmonary endothelium and allowing the expression of functional tight junctions in the 16HBE14o- monolayer with sufficiently high transepithelial electrical resistance (TEER) values. Finally, monocytes were added to the apical compartment. PMA-activated macrophages significantly affected barrier integrity (73% TEER reduction compared to control after 24 h) and disrupted the epithelial tight junctions as shown by redistribution of ZO-1 labeling. Alternatively, monocytes could be activated using lipopolysaccharide, at a sub-toxic level int he apical compartment and only induced a small, though significant, reduction in TEER.This coculture system is a representative model of the lung–blood barrier with barrier integrity as the main toxicity endpoint.
我们使用人支气管上皮细胞（16HBE14o-）、单核细胞（THP-1）和人肺微血管内皮细胞（HLMVEC）建立了肺-血屏障的共培养模型，该模型可以同时评估多个参数。上皮细胞和内皮细胞生长在微孔膜的两侧。电子显微镜和共聚焦显微镜照片显示细胞在各自合适的区域内，两种细胞类型均未显示出穿过孔隙生长的迹象。在三种内皮细胞类型（EAhy.926、HUVEC和HLMVEC）中，最后一种被选为最合适的细胞类型，它最类似于肺内皮细胞，并能在16HBE14o-单层中表达功能性紧密连接，且具有足够高的跨上皮电阻（TEER）值。最后，将单核细胞添加到顶侧腔室。如ZO-1标记的重新分布所示，经佛波酯（PMA）激活的巨噬细胞显著影响屏障完整性（24小时后与对照组相比TEER降低73%）并破坏上皮紧密连接。或者，可以使用脂多糖在顶侧腔室以亚毒性水平激活单核细胞，其仅诱导TEER出现虽小但显著的降低。这种共培养系统是以屏障完整性作为主要毒性终点的肺-血屏障的代表性模型。
