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We have examined the link between the aggregation or spreading of cells adhering to substrata of differing biocompatibility and activation of the cyclic AMP (cAMP) pathway. We compared the rate at which the Mouse Swiss 3T3 fibroblasts attached to Cuprophan (CU), AN69 and a control plastic in the presence and absence of foetal calf serum (FCS). Serum had no effect on the kinetics of cell attachment to CU or AN69. Cells incubated in culture medium containing 10% FCS aggregated on CU, whereas they spread on AN69 and plastic. Aggregated cells contained significantly higher concentrations of cAMP than cells spreading, and aggregation was prevented by treatment with miconazole, an inhibitor of adenylyl cyclase. cAMP-dependent cell aggregation occurred on all three substrata in serum-free medium, suggesting that proteins adsorbed onto AN69 and plastic in the presence of serum helped protect the cells. Far less serum protein was adsorbed onto CU than onto AN69 or plastic, consistent with the similar increases in cAMP in cells attached to CU with or without serum.

Glutaraldehyde (GTA) is used to cross-link collagen-based biomaterials, but these materials are often cytotoxic. In order to overcome this problem, we have proposed the use of the acyl azide methods with either hydrazine or diphenylphosphoryl azide (DPPA) as reagents. In this paper we determine the cytocompatibility of acyl azide- and GTA-treated pericardium in vitro, by an organotypic chick aorta culture technique developed for the evaluation of the propensity of vascular cells (both endothelial and smooth muscle cells) to migrate and grow on the surface of biomaterials. We first examined pericardium stabilization as a function of GTA concentration and time, so that we could minimize residual GTA molecules in the material. Treatment for 72 h with 0.05% GTA was optimal for thermal stabilization of the pericardium with a denaturation temperature (Td) of 86.8 degrees C, providing similar results to treatment with 0.6% GTA for 4 h (Td = 85.1 degrees C). Pericardium treated in this way was, however, poorly cytocompatible with little vascular cell migration and growth when compared with tissues treated by the acyl azide methods. The best results were obtained with 0.5% DPPA; treated tissues showed a high level of cross-linking (Td = 82.4 degrees C) and three-fold increases in cell growth and migration over those in a non-toxic control.

The trypsin sensitivity of chick embryo cellular layers cultivated by an in vitro organ culture method toward different biomaterials has been analysed. Monolayer cells grown on to tested samples were enzymatically dissociated in 5 min - 1 h. Cumulative cell numbers, expressed as the percentage of the totally detached cell number, were plotted versus time, thus permitting the calculation of the function. The mathematical treatment by a computerized system of this function represents the trypsin sensitivity. This value modulated by the migration cell number is the static adhesion modulated index (SAMI). The trypsin sensitivity expressed according to this index allowed the establishment of an abacus wherein several zones, A, B, C and D, define cell adhesion behaviour on different biomaterials. Scanning electron microscopy analysis performed on dissociation steps showed the selective activity of trypsin on cells toward different substrata, revealing the role of an extracellular matrix and cytoskeleton in the adhesion behaviour.