Influence of 8-bromo-cyclicAMP on interleukin -6 and -8 mRNA levels in A549 human lung epithelial cells exposed to organic dust: A time-kinetic study

Abstract

Exposure in a swine confinement building of previously unexposed subjects leads to an intense inflammatory reaction with increased number of inflammatory cells and mediators in the upper and lower respiratory tract. In vitro the organic dust induces cytokine release from respiratory epithelial cells. Whether the dust-induced release of IL-6 and IL-8 protein from A549 lung epithelial cells is a result of sustained mRNA expression during the 24 h exposure generally applied is unknown. Furthermore, it is not known if the previously demonstrated effects on basal and dust-induced IL-6 and IL-8 protein production by 8-bromo-cyclicAMP are time-dependent, since only cumulative effects are observed by measurement of cytokine release. In the present study reverse transcription- (RT-) PCR was applied to investigate expression of IL-6 and IL-8 mRNA in A549 cells exposed to organic dust in a time-kinetic manner. The dust increased IL-6 and IL-8 mRNA expression at all times tested (1 h–48 h). The IL-6 mRNA expression peaked at 1–1.5 h and was reduced with time, whereas the dust-induced IL-8 mRNA expression remained elevated. At 1–1.5 h, 8-bromo-cAMP stimulated basal and dust-induced IL-6 mRNA expression and attenuated dust-induced IL-8 mRNA expression by activation of protein kinase A- (PKA), as assessed with the PKA inhibitor H-89. On prolonged exposure (>3 h), the dust-induced IL-6 mRNA was PKA-dependently decreased, whereas at 17 h and longer the IL-8 mRNA expression induced by a dust-suspension with 8-bromo-cAMP was similar to, or enhanced, relative to the dust-induced IL-8 mRNA. Thus, 8-bromo-cAMP exerted opposite action on dust-induced IL-6 and IL-8 mRNA expression with time.