LEUKOCYTE MIGRATION - A NEW TRIPLE MIGRATION CHAMBER ASSAY ALLOWS INVESTIGATION OF VARIOUS CELL-INTERACTIONS SIMULTANEOUSLY

Examination of the interactions between various cells of the vascular wall and blood components are essential for understanding different pa thophysiological processes. Such investigations require appropriate te chniques. Several groups have attempted to establish different methods . In all blood vessels except capillaries, endothelial cells (EC) and smooth muscle cells (SMC) coexist and interact very closely. The curre nt study describes a new 3-dimensional triple chamber migration assay, studying leukocyte migration through human endothelial cell monolayer s (ECM) towards human SMC layers simultaneously. To test the new assay , SMC-layers were prestimulated with different concentrations of tumor necrosis factor alpha (TNF-alpha, 1 ng/ml, 10 ng/ml, 100 ng/ml) over 6 hours. Then, two microporous membranes, a collecting membrane and a third membrane with cultured ECM, were inserted. Freshly isolated peri pheral blood mononuclear cells (PBMNC) were seeded on the ECM and tran smigrated cells were measured after further 3 hours incubation. The mi gration against non stimulated SMC-layers was used as control. Prestim ulated SMC-layers led to a dose dependent increase of PBMNC migration into the subendothelial cell space. Antibodies against interleukin-1 r educed the PBMNC migration. In conclusion, this assay allows to study cell migration into the subendothelial space and interactions between different vascular cells. Moreover, this assay can also be used for st udies on other cell-cell interactions in man.