Olodaterol exerts anti-inflammatory effects on COPD airway epithelial cells

Abstract

Background: Airway inflammation is a key feature of chronic obstructive pulmonary disease (COPD) and inhaled corticosteroids (ICS) remain the main treatment for airway inflammation. Studies have noted the increased efficacy of ICS and long-acting beta 2 agonist (LABA) combination therapy in controlling exacerbations and improving airway inflammation than either monotherapy. Further studies have suggested that LABAs may have inherent anti-inflammatory potential, but this has not been well-studied.

Objective: We hypothesize that the LABA olodaterol can inhibit airway inflammation resulting from exposure to respiratory syncytial virus (RSV) via its binding receptor, the β2-adrenergic receptor.

Methods: Human bronchial epithelial brushing from patients with and without COPD were cultured into air-liquid interface (ALI) cultures and treated with or without olodaterol and RSV infection to examine the effect on markers of inflammation including interleukin-8 (IL-8) and mucus secretion. The cell line NCI-H292 was utilized for gene silencing of the β2-adrenergic receptor via siRNA as well as receptor blocking via ICI 118,551 and butaxamine.

Results: At baseline, COPD-ALIs produced greater amounts of IL-8 than control ALIs. Olodaterol reduced RSV-mediated IL-8 secretion in both COPD and control ALIs and also significantly reduced Muc5AC staining in COPD-ALIs infected with RSV. A non-significant reduction was seen in control ALIs. Gene silencing of the β2-adrenergic receptor in NCI-H292 negated the ability of olodaterol to inhibit IL-8 secretion from both RSV infection and lipopolysaccharide stimulus, as did blocking of the receptor with ICI 118,551 and butaxamine.

Conclusions: Olodaterol exhibits inherent anti-inflammatory properties on the airway epithelium, in addition to its bronchodilation properties, that is mediated through the β2-adrenergic receptor and independent of ICS usage.

Keywords: Airway epithelium; Airway inflammation; Beta 2-adrenergic receptor; COPD; Interleukin-8; Long-acting beta agonist; Olodaterol.

Fig. 1

Basal concentrations of IL-8 are higher in ALI cells of COPD versus control subjects. At baseline, COPD-ALIs produced four times greater concentrations of IL-8 than control ALIs (**p < 0.01). n = 3. Data was analyzed via two-tailed unpaired Student t-test

Fig. 2

The effect of olodaterol on Muc5AC expression with and without RSV infection. Mucus production was quantified by Muc5AC staining to visualize mucins. a Representative image of Muc5AC staining. b No significant change in Muc5AC expression with or without olodaterol and RSV treatment in control-ALIs. n = 4. c RSV infection significantly increased Muc5AC expression compared to the control (**p < 0.01) in COPD-ALIs. Additionally, olodaterol (OLO + RSV) significantly decreased RSV-mediated Muc5AC expression (**p < 0.01). Olodaterol concentration used was 10 µM. n = 3. Data was analyzed via one-way ANOVA with Bonferroni correction

Fig. 3

The effect of olodaterol on IL-8 secretion in ALI-cultures. a In control ALIs, olodaterol (OLO)-treated cultures demonstrated a significant reduction in RSV-induced IL-8 secretion compared to RSV alone (*p < 0.05). n = 4. b COPD-ALI cultures showed a significant decrease in RSV-mediated IL-8 secretion when pre-treated with olodaterol (OLO + RSV) compared to RSV infection alone (**p < 0.01). Olodaterol concentration used was 10 µM. n = 4. Data was analyzed via one-way ANOVA with Bonferroni correction

Fig. 4

The effect of olodaterol on RSV infections. RSV staining was used to quantify RSV particles. a Representative image of RSV staining. b Control ALIs showed significantly decreased RSV staining with olodaterol treatment (OLO + RSV) compared to RSV-only infection (*p < 0.05). n = 3. c COPD-ALIs also showed significantly decreased RSV staining with olodaterol treatment (**p < 0.01). Olodaterol concentration used was 10 µM. n = 3. Data was analyzed via two-tailed unpaired Student t-test

Fig. 5

Olodaterol dose response in NCI-H292 on LPS-mediated IL-8 secretion. NCI-H292 cells were treated with 2 µg/mL of LPS and increasing log doses of olodaterol (0.01, 0.1, 1 and 10 µM). Olodaterol concentrations of 0.01 µM, 0.1 µM, 1 µM and 10 µM all significantly inhibited IL-8 secretion compared to the LPS-only treatment (***p < 0.001, ****p < 0.0001, ****p < 0.0001 and ****p < 0.0001, respectively). n = 6. Data was analyzed via one-way ANOVA with Bonferroni correction

Fig. 6

The effect of β2AR gene silencing on IL-8 secretion induced by RSV and LPS. a In non-transfected and scrambled siRNA-transfected cells, olodaterol inhibited RSV-mediated IL-8 secretion (OLO + RSV, **p < 0.01 and ****p < 0.0001 respectively) but not in β2AR siRNA-transfected cells. b Olodaterol attenuated LPS-mediated IL-8 secretion in non-transfected (OLO + LPS, ***p < 0.001) and scrambled siRNA-transfected cells (OLO + LPS, ***p < 0.001) but not in β2AR siRNA-transfected cells. n = 3. In both (a) and (b), β2AR siRNA negated olodaterol-mediated reduction of LPS-induced IL-8 secretion. Olodaterol concentration used was 10 µM. n = 3. Data was analyzed via two-way ANOVA with Bonferroni correction

Fig. 7

The effect of the β2AR antagonists ICI 118,551 and butaxamine on olodaterol inhibition of IL-8 secretion. NCI-H292 cells were treated with 2 µg/mL LPS with and without 0.01 µM olodaterol. To antagonize β2AR, cells were pre-treated with 30 nM of ICI 118,551 or 100 nM of butaxamine for 1 h, followed by addition of 0.01 µM olodaterol for 8 h and 2 µg/mL LPS overnight for a total of 24 h. a Olodaterol significantly inhibited LPS-mediated IL-8 secretion (**p < 0.01) but not in the presence of 30 nM of ICI 118,551. b Olodaterol also inhibited LPS-mediated IL-8 secretion (*p < 0.05) but not in the presence of 100 nM of butaxamine. n = 6. Data was analyzed via one-way ANOVA with Bonferroni correction