A coculture model of the lung–blood barrier: The role of activated phagocytic cells

Abstract

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.

Dual and Triple Epithelial Coculture Model Systems with Donor-Derived Microbiota and THP-1 Macrophages To Mimic Host-Microbe Interactions in the Human Sinonasal Cavities

“…Coculture of Calu-3 cell layers with THP-1-derived macrophage-like cells are known to decrease epithelial resistance when seeded at concentrations of 2 ϫ 10 5 cells ml Ϫ1 , and yet this effect is not apparent when lower seeding densities (2 ϫ 10 4 cells ml Ϫ1 ) are applied (73). Similar drops in TEER from THP-1-derived macrophages were previously noted for coculture models of the lung-blood barrier (74). While phorbol-12-myristate-12-acetate (PMA), the differentiation agent for THP-1 cells, is known to induce an inflammatory phenotype in vitro (75), it is not the cause for lower barrier integrity in our setup since the epithelial cell layers were not exposed to PMA.…”

A microfluidic renal proximal tubule with active reabsorptive function

“…Very few models have been developed to incorporate microvascular tissue to mimic reabsorptive barriers in vitro [ 33 , 34 ] . Transwells provide either a non-contact orientation with a 1 mm distance between the two cell types, or cells on both sides of the Transwell insert membrane to establish an approximation of a close-contact orientation of the two cell types[ 35 , 36 ]. The second orientation provides a more realistic architecture, but the Transwells deprive cells of physiological cues [ 19 , 37 ], such as BM topography and FSS which are present in vivo and accommodated in our model.…”

The Effect of Immunosuppression on Airway Integrity

“…Human Bronchial Epithelial cells, 16HBE14o -(16HBE) (provided by Dr. Gruenert, University of California, San Fransisco, USA) and Human Lung microvascular endothelial cells (HLMVEC) were purchased from the European Collection of Cell Cultures (Salisbury, UK) and cultured as previously described 10 . 16HBE cells were used for experiments between passages 3-15, HLMVEC cells between passages 3-10.…”

“…To mimic the barrier between blood stream and epithelium of the lung (ie, to mimic drug delivery of immunosuppressive drugs to the lungs via the blood), a biculture of 16HBE cells and HLMVEC cells was set up as described in Luyts et al 10 ( Figure S1, SDC, http://links.lww.com/TP/B446). When the medium of both compartments was renewed at day 1, 4 and 7, the supernatant of both compartments was collected (200 µL).…”