CHEMICAL INDUCTION OF INTERLEUKIN-8, A PROINFLAMMATORY CHEMOKINE, IN HUMAN EPIDERMAL KERATINOCYTE CULTURES AND ITS RELATION TO CYTOGENETIC TOXICITY

Tumor promoters, proinflammatory cytokines, endotoxins, and protein synthesis inhibitors can modulate cell cycle kinetics of various cell types, stimulate production of reactive oxygen species, and induce keratinocytes to produce interleukin-8 (IL-8) a potent chemotactant for polymorphonuclear neutrophils and T lymphocytes. The aim of this study was to determine whether perturbations of cytogenetic responses correlated with the induction of IL-8 expression Cultures of primary human keratinocytes were grown in serum-free medium with 5 mu mol/L bromodeoxyuridine to label DNA and exposed either to phorbol-13-myristate-12-acetate (PMA) (0.0001-100 ng/ml), cycloheximide (CHX) (0.01-50 mu g) lipopolysaccharide (0.1-100 mu g/ml), tumor necrosis factor-a (TNF alpha) (3.13-50 ng/ml), or interleukin-1 alpha (IL-1 alpha) (1-182 pg/ml). Metaphase chromosome preparations were stained by a fluorescence-plus-Giemsa technique to differentiate sister chromatids. For IL-8 production, keratinocytes were grown to 70% confluency and then exposed to chemicals for 24 h. Immunoreactive IL-8 was quantitated from the supernatants by ELISA. With the exception of benzo(a)pyrene used as a positive control, none of the agents induced sister chromatid exchanges. However, PMA and TNF alpha induced IL-8 production that coincided with significant cell cycle inhibition IL-1 alpha had no effect on cytogenetic endpoints, yet stimulated a 6.3-fold increase in IL-8. CHX inhibited cell cycle progression and mitotic activity at concentrations that were 200 times lower than required for IL-8 induction; however, puromycin (0.31-10 mu g/ml), another protein synthesis inhibitor, did not induce IL-8. At all concentrations tested, TNF alpha reduced the mitotic index by similar to 45%, slowed cell cycle progression by similar to 3.5 h, and induced a flat, albeit large, IL-8 response at concentrations greater than or equal to 12.5 ng/ml. These agent-specific response patterns suggest that induction of IL-8 production is not always the inevitable result of cell cycle perturbations or genetic damage.