Cyclosporin-sensitive expression of cytokine mRNA in mouse macrophages responding to bacteria

Abstract

Characteristics of the cytokine response in resident mouse macrophages to certain Gram-positive and Gram-negative bacteria have been investigated by monitoring the expression of mRNA encoding interleukin-1α and -β ( IL- 1α β ) and tumor necrosis factor-α (TNF-α). Expression of these cytokine mRNAs occurred within 30–60 min. Both the flavonoid quercetin and phloretin inhibited the expression of IL- 1α β as well as TNF-α mRNA, with quercetin being more potent than phloretin and TNF-α expression somewhat more sensitive than that of IL- 1α β . Expression of all three cytokine mRNAs was also inhibited by prostaglandin E 2, with an IC 50 of > 1 μM, but not by the phosphodiesterase inhibitor pentoxifylline, although lipopolysaccharide-induced expression of TNF-α mRNA was inhibited. Down-regulation of phorbol ester-sensitive isoforms of protein kinase C had virtually no effect on the cytokine response to bacteria, and treatment of resting macrophages with phorbol ester did not cause expression of any of the cytokine mRNAs investigated. Among protein phosphatase inhibitors, cyclosporin A caused extensive inhibition of bacteria-induced expression of both IL- 1α β and TNF-α mRNA, while okadaic acid in itself caused selective induction of TNF-α, but not IL- 1α β mRNA, with a sharp peak at 0.3 μM concentration. At higher concentrations of okadaic acid, at which protein/phosphatase 2B/calcineurin would also be inhibited, the induction was completely reversed. This suggests that critical phosphorylation events, counteracted by one or more okadaic acid-sensitive protein phosphatase(s), and a dephosphorylation event carried out by a cyclosporin-sensitive protein phosphatase are both necessary for transcriptional activation of the TNF-α gene.