Effect of interleukin-1β, tumour necrosis factor-α and interferon-γ on the induction of cyclo-oxygenase-2 in cultured human airway smooth muscle cells

Abstract

1. Increased levels of several pro-inflammatory cytokines including interleukin-1β (IL-1β) and tumour necrosis factor-α (TNFα) have been found in bronchoalveolar lavage fluid from symptomatic asthmatic patients. IL-1β, TNFα and interferon-γ (IFNγ) are known to stimulate a number of cells to produce inflammatory mediators such as prostaglandins. Although airway smooth muscle (ASM) is known to be a rich source of prostaglandins, the regulation of cyclo-oxygenase (COX) isoforms and prostanoid production by proinflammatory cytokines have not been studied in human airway smooth muscle.

2. We studied the effects of IL-1β, TNFα and IFNγ on the induction of two isoforms of cyclo-oxygenase and its relation to prostaglandin E₂ (PGE₂) release and COX activity (reflected by PGE₂ synthesis from exogenous arachidonic acid) in human cultured airway smooth muscle cells.

3. IL-1β, but not TNFα or IFNγ, caused a time- and concentration-dependent enhancement in PGE₂ and other prostanoid (6-keto-PGF_1α, PGF_2α, thromboxane B₂ (TXB₂) and PGD₂) production, with PGE₂ and 6-keto-PGF_1α as the principal products. This stimulation was accompanied by a corresponding increase in COX activity.

4. COX-2 protein measured by Western blot analysis was not detectable in untreated cells, but was increased in a time- and concentration-dependent manner by IL-1β, but not TNFα or IFNγ. In contrast, no variation in the expression of COX-1 protein was observed.

5. Pretreatment with the conventional non-steroidal anti-inflammatory drugs (NSAIDs), indomethacin and ibuprofen, and the selective COX-2 inhibitors, NS-398 and nimesulide, completely blocked IL-1β-induced PGE₂ release and COX activity. The glucocorticosteroid dexamethasone and protein synthesis inhibitors, cycloheximide and actinomycin D, not only markedly inhibited IL-1β-stimulated PGE₂ release and COX activity but also suppressed IL-1β-induced COX-2 induction.

6. This study demonstrates that human cultured ASM cells release prostanoids in response to IL-1β stimulation and that the response is mostly mediated by the induction of COX-2 rather than COX-1 isoenzyme, implying that airway smooth muscle may be an important source of prostaglandins in human airways and that COX-2 may play an important role in the regulation of the inflammatory process in asthma.

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