Cytokine-induced sequential migration of neutrophils through endothelium and epithelium.

OBJECTIVE AND DESIGN

To better understand the mechanisms by which cytokines induced neutrophils to migrate into the airways, we constructed a novel in vitro model system.

MATERIALS

Human umbilical vein endothelial cell (HUVE) monolayers were grown on top of permeable filters and human lung type II-like alveolar epithelial cell (A549) monolayers were grown on the undersurface of the filters.

METHODS

The sequential migration of human neutrophils through the endothelium (apical to basal movement) and subsequently through the epithelium (basal to apical movement) in response to IL-1 beta or TNF alpha located basally to the epithelium was measured.

RESULTS

We found that IL-1 beta and TNF alpha induced dose-responsive and time-dependent migration through the double monolayers-filter complex. The pattern of migration was similar, and the amount greater than or equal to that observed through either single monolayer/filter complex. Neutrophil migration through naked filters was generally less than that observed through the cellular barriers. The contribution of the monolayer orientation was also examined and found to favor the more physiologic directional migration of neutrophils through an endothelial and epithelial barrier, apical to basal and basal to apical, respectively. In contrast, FMLP-induced neutrophil migration was not dependent upon either the orientation or presence of the monolayer(s).

CONCLUSIONS

Thus, we have established an in vitro model system to examine cytokine-induced sequential migration of neutrophils through endothelium and the respiratory epithelium in a manner analogous to that occurring with an in vivo airway stimulus causing neutrophil-rich airway inflammatory responses.