LPS Exposure Exacerbates or Attenuates Ovalbumin-Induced Functional And Inflammatory Responses And Induces Corticosteroid Insensitivity In A Guinea-Pig Model of Asthma

A.P.P Lowe1, A.T Nials2, W.R Ford1, E.J Kidd1, K.J Broadley1. 1Cardiff University, Cardiff, UK, 2GSK, Stevenage, UK

Background: Exacerbations of asthma contribute to decreased responsiveness to inhaled steroids (Fahy et al, 1995; in't Veen et al, 1999). Both bacterial and environmental lipopolysaccaride (LPS) contribute to steroid insensitivity and asthma exacerbations in humans (Goleva et al, 2008; Bisgaard et al, 2010). This study examined the temporal effects of LPS exposure on allergen challenge responses and sensitivity to steroid in guinea-pigs.

Methods: Guinea-pigs (200-250g, male, Charles River) were sensitised with 150µg ovalbumin (Ova) and 100mg Al(OH)3. Airway responses to inhaled Ova (0.03%), in conscious guinea-pigs were determined by whole body plethysmography (Buxco Systems, USA). Changes in specific airways conductance (sGaw) were determined over 12hrs and at 24hrs. LPS (30μg/ml, 1h) was administered in two protocols 1) 72 and/or 24hrs pre-Ova, 2) 48hrs pre-Ova and co-administered with Ova. Fluticasone propionate (FP, 0.5 mg/ml daily) was administered by inhalation for 6 days, split into twice daily doses. Airways hyper-responsiveness (AHR) was investigated from the bronchoconstriction by inhaled histamine (0.3mM) 7 days pre- and 24hr post-Ova challenge. Inflammatory cell influx was determined by bronchoalveolar lavage at 24hr post-challenge.

Results: Animals challenged with Ova alone displayed peak early asthmatic responses (EAR, -70.9±7.2%) which were significantly attenuated by LPS exposure 24hrs pre-Ova (-34.4±4.6%, P<0.05). LPS exposure 48hrs pre- and co-administered with Ova prolonged the EAR (time to 50% of peak value (4.8±0.6h, compared to Ova alone, 2.3±0.7h, P<0.05). Late asthmatic responses (LAR) to Ova measured as the 6-12hr peak (-22.4±3.9%) were unchanged in all LPS treatment groups. Ova challenge of saline-treated animals displayed significant AHR, bronchoconstriction to histamine increasing from 0.7±4.0% to -21.6±4.6%, (P<0.01) change in sGaw. LPS exposure 24hr pre-Ova did not significantly alter the bronchoconstriction to histamine but did attenuate it when combined with a 72hr pre-Ova exposure (-4.4±2.3%; -5.7±2.9%; pre- and post-Ova respectively). LPS co-administered and given 48hr pre-Ova exacerbated AHR (7.3±2.7% pre-; -41.1±7.0% post-Ova, P<0.001). Ova alone in saline-treated animals produced eosinophilic inflammatory responses (8.9±0.4x106 cells/ml). Two LPS exposures, either co-administered and given 48hr pre- or 24 and 72hr pre-Ova significantly increased total cells (21.0±0.9x106/ml; 18.3±3.5x106/ml respectively, P<0.05), including neutrophils and macrophages. In all combined Ova and LPS protocols tested, FP failed to reduce LAR, AHR and total and differential cells, at a dose effective in reducing these parameters after Ova exposure alone.

Conclusions: LPS exposure exerts a temporal effect on the functional responses to allergen challenge. LPS 24hrs before allergen challenge diminishes functional responses, whereas co-administration with allergen augments these responses. LPS has no temporal effects on inflammatory responses and sensitivity to steroid, with both pre- and co-challenge rendering both functional and inflammatory responses steroid insensitive. The experimental combination of Ova and LPS co-administration might represent a useful preclinical model of asthma exacerbation and steroid-resistant airway inflammation.

Bisgaard H et al (2010) BMJ 341:c4978

Fahy JV, et al (1995) J Allergy Clin Immunol. 95:843-52.

Goleva et al, (2008) J.AllergyClin.Immunol 122:550–559

in't Veen et al (1999) Am J Respir Crit Care Med 1999;160:93–99