PP255—IFN-gamma Interfere the Effect of Beta-2 Agonist on Tnf-Alpha Induced CXCL10 Through Creb Phosphorylation in HASM Cells

IntroductionCXCL10 is a potent mast cell chemoattractant responsible for the mast cell myositis characteristic of asthma. We have identified human airway smooth muscle (HASM) as a rich source of CXCL10 which is increased by TNF-alpha and IFN-gamma. We have previously shown that beta-agonists inhibit TNF-alpha induced CXCL10 release but the effect is lost when IFN-gamma is given concomitantly through poorly defined mechanisms. Here we defined the mechanism involved.Patients (or Materials) and MethodsHASM cells taken from 3 normal donors were cultured using standard techniques. ELISA was used for quantitative measurement of CXCL10 release. Cyclic AMP assay was used to measure the cAMP level. Western Blot was used to indicate CREB phosphorylation.ResultsWe found that salmeterol time dependently increased cAMP generation. TNF-alpha or IFN-gamma alone or in combination had similar effects on salmeterol time dependently induced cAMP generation suggesting that the inhibitory effect of IFN-gamma on beta-2 agonist signaling was not at the level of cAMP generation but rather was likely to be an effect on downstream signaling pathways. Next we looked at the phosphorylation of CREB by Western blot and found that salmeterol and caused phosphorylation of CREB but this was differentially affected by the cytokines. Whereas TNF-alpha alone was without effect but in combination with IFN-gamma markedly impaired the ability of the long-acting beta-agonist to phosphorylate CREB. This suggests a novel mechanism whereby IFN-gamma interferes with beta-agonist signaling by impairing phosphorylation of CREB. This may be important in reducing the responses to beta-agonists in refractory asthma.ConclusionIFN-gamma interferes with beta-2 agonist signaling by impairing phosphorylation of CREB. This may be important in reducing the responses to beta-agonists in refractory asthma.Disclosure of InterestNone declared. IntroductionCXCL10 is a potent mast cell chemoattractant responsible for the mast cell myositis characteristic of asthma. We have identified human airway smooth muscle (HASM) as a rich source of CXCL10 which is increased by TNF-alpha and IFN-gamma. We have previously shown that beta-agonists inhibit TNF-alpha induced CXCL10 release but the effect is lost when IFN-gamma is given concomitantly through poorly defined mechanisms. Here we defined the mechanism involved.