Human endothelial cells cultured on microporous filters used for leukocytetransmigration studies form monolayers on both sides of the filter

A growing number of studies on the mechanism of leukocyte transendothelial migration use endothelial cells grown on microporous filters as an in vitro model of endothelium. Ultrastructural examination of such a model system p reviously demonstrated that human pulmonary artery endothelial cells (HPAEC ) formed confluent monolayers on Loth sides of the 3-mu m-pore filter (Mack arel et al., 1999). To determine whether this was a characteristic specific to pulmonary artery endothelial cells, the growth characteristics of a hum an pulmonary microvascular endothelial cell type (HMVEC-L) and the widely u sed human umbilical vein endothelial cells (HUVEC) on 3-mu m microporous fi lters were examined by transmission electron microscopy (TEM). Similar to H PAEC, HMVEC-L and HUVEC were also found to grow on both sides of the filter . All three endothelial cell types were capable of migrating through the 3 mu m pores of the filter to form a monolayer on the filter underside. The e ndothelial cells on the underside were orientated in an inverted position w ith the luminal surface facing away from the filter Such 'bilayer' formatio n was observed at a range of seeding densities and in different culture med ia. Despite the presence of a bilayer of endothelial cells, TEM demonstrate d that neutrophils migrated successfully across the cell-filter-cell system . Previous transmigration reports in which an in vitro model similar to our s was used have often assumed only one layer of endothelial cells. The obse rvations reported here indicate that while endothelial cells on microporons filters are useful models for examining leukocyte-endothelial interactions , they are not appropriate for studies examining endothelial cell 'sidednes s.'