Rationale Chronic obstructive pulmonary disease (COPD) is an irreversible inflammatory lung disease and is currently the fourth greatest burden of disease of worldwide. However a key issue is that patients show a lack of response to corticosteroid treatment. Corticosteroid insensitivity is mainly caused by oxidative stress which directly stimulates inflammatory transcription factors and reduces the activity of co-adaptor proteins essential for the inhibitory actions of corticosteroids. AMP-activated kinase (AMPK) is a serine/threonine protein kinase that regulates cellular energy homeostasis and anti-oxidant defences, and has recently been labelled as an anti-ageing molecule. We hypothesised that activation of AMPK using Quercetin reverses corticosteroid resistance caused by cigarette smoke extract (CSE) in a monocytic cell line.
Methods Human monocytic cell line, U937s, were initially incubated with Quercetin (20µM) for 24 hours and then exposed to CSE for 2 hours. Cells were then treated with dexamethasone (1x10-11 to 1x10-6M) for 45 minutes and stimulated with TNF-a (10ng/ml) for 24 hours. Supernatants were collected and CXCL-8 was measured using ELISA. Corticosteroid resistance was calculated as the ability of dexamethasone to inhibit 25% of TNF-a-induced CXCL-8 (IC25). Activation of AMPK by Quercetin was measured using the levels of the phosphorylated AMPK by Western Blot. Nuclear factor erythroid related factor 2 (Nrf-2) levels and glucocorticoid receptor (GR) nuclear translocation were also assessed using Western Blot.
Results CSE induced corticosteroid resistance in U937s (IC25 = 30nM vs IC25 = 5nM). Interestingly Quercetin restored corticosteroid sensitivity by approximately 3 fold (IC25 = 11nM) compared to CSE. Quercetin increased levels of activated AMPK and also up-regulated the expression of Nrf-2. However, Quercetin was unable to restore GR nuclear translocation.
Conclusions Quercetin was found to be a potential novel therapy for restoration of corticosteroid sensitivity in COPD. Although the mechanism of action remains to be elucidated, Nrf-2 and AMPK activations which increase anti-oxidant levels and prevents oxidative damage could be a key the mechanism of action Activation of AMPK could therefore be a potential novel mechanism for the restoration of corticosteroid sensitivity and Quercetin could be used as an add-on treatment to corticosteroids in COPD.