Influence of anti-inflammatory drugs on adhesion of neutrophils to endothelial cells cultured on microcarriers: a novel in vitro system as an alternative to animal experimentation.

Pharmacological control of inflammation by steroidal (SAIDs) and nonsteroidal (NSAIDs) antiinflammatory drugs is of substantial clinical importance. To reduce the number of animals used in pharmacological and toxicological evaluation of these drugs we developed a novel assay to determine adhesion of bovine neutrophils (PMN) to bovine aortic endothelial cells (BAEC) cultured on microcarriers in a flow-through system. Pretreatment of BAEC with thrombin (10(-7)-10(-4) M) led to a dose-dependent increase of PMN-adhesion (10(-6)-10(-4) M:P < 0.05); platelet-activating factor (10(-9) M) and 1:200 diluted zymosan-activated serum (ZAS) had similar effects (P < 0.001). Pretreatment of PMN with SAIDs (50.9 and 509 microM dexamethasone, 12.2 and 24.4 microM flumethasone) did inhibit adhesion to ZAS-treated BAEC dose-dependently. Pretreatment of PMN with NSAIDs had a less consistent influence on adhesion to ZAS-stimulated BAEC. While phenylbutazone (0.33 and 3.3 mM), diclofenac (0.392 and 0.574 mM), indomethacine (0.436 and 0.872 mM), and acetylsalicylic acid (3.47 and 16.94 mM) induced dose-dependent inhibition of PMN-adhesion to ZAS-treated BAEC, piroxicam (0.377 and 0.754 mM) inhibited PMN-adhesion strongly (P < 0.001) but not dose-dependently, and ketoprofene (0.614 and 1.228 mM) had no effect on PMN-adhesion. The method presented here is efficient for evaluating the pharmacological modulation of PMN interaction with endothelial cells, and useful for studying further aspects of endothelial cell biology.