Leukocyte attack in a 3D human coronary in-vitro model

Objective The importance of peripheral blood leukocytes for the development of early atherosclerosis and restenosis has confronted cardiologists with classical hematologic issues. Three-dimensional human coronary in-vitro uni ts of leukocyte attack (3DLA-units) open the field for exact studies of leu kocyte attack and its subsequent effects on human medial coronary smooth mu scle cells (HCMSMC). Methods Central part of 3DLA-units are polycarbonate membranes with a pore size of 5 mum that correspond to the internal elastic membrane. Human coron ary endothelial cells (HCAEC) were cultured on one side of the membranes, H CMSMC on the other side. Before leukocyte attack expression of adhesion mol ecules was up-regulated by tumour necrosis factor-alpha (TNF-alpha). Leukoc yte attack was mimicked by selective adding of human monocytes (MC), respec tively human CD4(+)-lymphocytes (CD4(+)-LC) to the HCAEC side of the 3DLA-u nits. Three-dimensional leukocyte attack units were fixed and stained after a period of 30 min, 1, 2, 3, 4, 6, and 24 h. Cell divisions of HCMSMC were analysed by measuring the uptake of bromodeoxyuridine (BrdU). Results Monocytes were able to adhere to the endothelial surface, pass thro ugh the filter-pores, and penetrate the HCMSMC side of the 3DLA-units. Huma n CD4(+)-lymphocytes (CD4(+)-LC) only attached to the HCAEC side, and no ch emotaxis to the HCMSMC side was detected. Proliferation of HCMSMC was incre ased 2.9-fold (P < 0.001) after selective MC-attack and 3.5-fold after sele ctive MC-attack and TNF-<alpha> stimulus. No significant increase was found after selective CD4(+)-LC attack, a significant increase (2.1-fold; P< 0.0 01) was seen after selective CD4(+)-LC attack and TNF-<alpha> stimulus. Conclusions Within the given limitations of the model the study emphasizes a predominance of MC in comparison to CD4(+)-LC in the process of adhesion, chemotaxis, and triggered reactive proliferation of co-cultured HCMSMC wit hin the first 24 h after leukocyte attack. 3DLA-units offer an elegant meth od to study directly the effects of intravascular and intramural treatment strategies. Coron Artery Dis 12:401-411 (C) 2001 Lippincott Williams & Wilk ins.