Tumor cell/endothelial cell tight contact upregulates endothelial adhesion molecule expression mediated by NFkappaB: differential role of the shear stress.

Cancer metastasis is a multistep process involving cell-cell interactions, but little is known about the adhesive interactions and signaling events during extravasation of tumor cells (TCs). In this study, cell adhesion molecule (CAM) expression was investigated using an in vitro assay, in which TCs were seeded onto an endothelial cell (ECs) monolayer and cocultured during 5 h. Flow cytometry, confocal microscopy as well as western blot analysis indicated that endothelial ICAM-1 (Inter Cellular Adhesion Molecule-1), VCAM-1 (Vascular Adhesion Molecule-1) and E-selectin were up-regulated after TC-EC coculture, whereas no change was observed for CAMs expression in tumor cells. This increased CAMs expression required tight contact between TCs and ECs. Incubation of ECs with the pyrrolidine-dithiocarbamate NFkappaB inhibitor prior to coculture, fully prevented coculture-induced expression of endothelial CAMs. Using specific blocking antibodies we showed an implication of ICAM-1 and VCAM-1 for TCs extravasation and VCAM-1 for adhesion. Moreover, fluid flow experiments revealed that high shear stress totally abolished coculture-induced as well as TNFalpha-induced CAMs over-expression. This study suggests that TCs could act as a potent inflammatory stimulus on ECs by inducing CAMs expression via NFkappaB activation, and that this action can be modulated by shear stress.

Morphological analysis of tumor cell/endothelial cell interactions under shear flow.

In the process of hematogenous cancer metastasis, tumor cells (TCs) must shed into the blood stream, survive in the blood circulation, migrate through the vascular endothelium (extravasation) and proliferate in the target organs. However, the precise mechanisms by which TCs penetrate the endothelial cell (EC) junctions remain one of the least understood aspects of TC extravasation. This question has generally been addressed under static conditions, despite the important role of flow induced mechanical stress on the circulating cell-endothelium interactions. Moreover, flow studies were generally focused on transient or firm adhesion steps of TC-EC interactions and did not consider TCs spreading or extravasation. In this paper, we used a parallel-plate flow chamber to investigate TC-EC interactions under flow conditions. An EC monolayer was cultured on the lower plate of the flow chamber to model the endothelial barrier. Circulating TCs were introduced into the flow channel under a well-defined flow field and TC cell shape changes on the EC monolayer were followed in vitro with live phase contrast and fluorescence microscopy. Two spreading patterns were observed: radial spreading which corresponds to TC extravasation, and axial spreading where TCs formed a mosaic TC-EC monolayer. By investigating the changes in area and minor/major aspect ratio, we have established a simple quantitative basis for comparing spreading modes under various shear stresses. Contrary to radial spreading, the extent of axial spreading was increased by shear stress.

A new flow chamber for the study of shear stress and transmural pressure upon cells adhering to a porous biomaterial.

Biomaterials used in some biomedical devices are porous and exposed to normal and tangential flow of biofluids. To examine the influence of flow induced forces on the morphology and the biochemical responses of cells adhering to such biomaterials, a Hele-Shaw cell with a porous bottom wall was designed and characterized experimentally. Theoretical predictions for the flow in the chamber are provided and allow to quantify the shear stress and/or transmural pressure exerted on cells. It is thus possible to follow up continuously the shape changes of cells that are adherent on a permeable membrane used in bioreactors.