Chemical induction of interleukin-8, a proinflammatory chemokine,in human epidermal keratinocyte cultures and its relation to cytogenetic toxicity |
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Authors: | J L Wilmer M I Luster |
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Institution: | (1) Environmental Immunology and Neurobiology Section, National Institute of Environmental Health Sciences, PO Box 12233, 27709 Research Triangle Park, North Carolina, USA |
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Abstract: | 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 mol/L bromodeoxyuridine to label DNA and exposed either to phorbol-13-myristate-12-acetate (PMA) (0.0001–100 ng/ml), cycloheximice (CHX) (0.01–50 g), lipopolysaccharide (0.1–100 g/ml), tumor necrosis factor- (TNF) (3.13–50 ng/ml), or interleukin-1 (IL-1) (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 induced IL-8 production that coincided with significant cell cycle inhibition. IL-1 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 g/ml), another protein synthesis inhibitor, did not induce IL-8. At all concentrations tested, TNF reduced the mitotic index by 45%, slowed cell cycle progression by 3.5 h, and induced a flat, albeit large, IL-8 response at concentrations 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.Abbreviations B(a)P
benzo(a)pyrene
- BrdU
5-bromo-2-deoxyuridine
- CHX
cycloheximide
- ICAM
intercellular adhesion molecules
- IL-1
interleukin-1
- IL-8
interleukin-8
- KGM
keratinocyte growth medium
- LPS
lipopolysaccharide
- PKC
protein kinase C
- PMA
phorbol-13-myristate-12-acetate
- PMN
polymorphonuclear neutrophil
- ROS
reactive oxygen species
- SCE
sister chromatid exchange
- TNF
tumor necrosis factor |
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Keywords: | interleukin-1" target="_blank">gif" alt="agr" align="BASELINE" BORDER="0"> tumor necrosis factor-" target="_blank">gif" alt="agr" align="BASELINE" BORDER="0"> phorbol-13-myristate-12-cetate cyclohexemide lipopolysaccharide puromycin benzo(a)pyrene inflammation sister chromatid exchanges |
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