title: Adjustment of sensitisation and challenge protocols restores functional and inflammatory responses to ovalbumin in guinea-pigs.
abstract: BACKGROUND
  Inhalation of antigen in atopic asthma induces early (EAR) and late asthmatic responses (LARs), inflammatory cell infiltration and airways hyperresponsiveness (AHR). Previously, we have established a protocol of sensitisation and subsequent ovalbumin (Ova) inhalation challenge in guinea-pigs which induced these 4 features (Smith & Broadley, 2007). However, the responses of guinea-pigs to Ova challenge have recently declined, producing no LAR or AHR and diminished EAR and cells. By making cumulative modifications to the protocol, we sought to restore these features.
  
  
  METHODS
  Guinea-pigs were sensitised with Ova (i.p. 100 or 150 μg) on days 1 and 5 or days 1, 4 and 7 and challenged with nebulised Ova (100 or 300 μg/ml, 1h) on day 15. Airway function was measured in conscious guinea-pigs by whole-body plethysmography to record specific airway conductance (sGaw). Airway responsiveness to aerosolized histamine (0.3mM) was determined before and 24h after Ova challenge. Bronchoalveolar lavage was performed for total and differential inflammatory cell counts. Lung sections were stained for counting of eosinophils.
  
  
  RESULTS
  Lack of AHR and LAR with the original protocol was confirmed. Increasing the Ova challenge concentration from 100 to 300 μg/ml restored AHR and eosinophils and increased the peak of the EAR. Increasing the number of sensitisation injections from 2 to 3 did not alter the responses. Increasing the Ova sensitisation concentration from 100 to 150 μg significantly increased total cells, particularly eosinophils. A LAR was revealed and lymphocytes and eosinophils increased when either the Al(OH)3 concentration was increased or the duration between the final sensitisation injection and Ova challenge was extended from 15 to 21 days.
  
  
  CONCLUSIONS
  This study has shown that declining allergic responses to Ova in guinea-pigs could be restored by increasing the sensitisation and challenge conditions. It has also demonstrated an important dissociation between EAR, LAR, AHR and inflammation.
authors.affiliation: Cardiff School of Pharmacy, Cardiff University, Redwood Building, King Edward VII Ave, Cardiff CF10 3NB, United Kingdom.
authors.forename: Alexander P P
authors.initials: AP
authors.lastname: Lowe
authors.affiliation: Cardiff School of Pharmacy, Cardiff University, Redwood Building, King Edward VII Ave, Cardiff CF10 3NB, United Kingdom. Electronic address: BroadleyKJ@cardiff.ac.uk.
authors.forename: Kenneth J
authors.initials: KJ
authors.lastname: Broadley
authors.affiliation: Discovery Biology, Respiratory Centre of Excellence for Drug Discovery, GlaxoSmithKline Medicines Research Centre, Gunnels Wood Road, SG1 2NY Stevenage, United Kingdom.
authors.forename: Anthony T
authors.initials: AT
authors.lastname: Nials
authors.affiliation: Cardiff School of Pharmacy, Cardiff University, Redwood Building, King Edward VII Ave, Cardiff CF10 3NB, United Kingdom.
authors.forename: William R
authors.initials: WR
authors.lastname: Ford
authors.affiliation: Cardiff School of Pharmacy, Cardiff University, Redwood Building, King Edward VII Ave, Cardiff CF10 3NB, United Kingdom.
authors.forename: Emma J
authors.initials: EJ
authors.lastname: Kidd
mesh_terms: D000280:Administration, Inhalation; D000485:Allergens; D000818:Animals; D001249:Asthma / Q000139:chemically induced*; D016535:Bronchial Hyperreactivity / Q000276:immunology; D006168:Guinea Pigs; D006632:Histamine / Q000633:toxicity; D007114:Immunization; D007249:Inflammation / Q000139:chemically induced*; D008297:Male; D010047:Ovalbumin / Q000008:administration & dosage / Q000633:toxicity*
chemical_list: D000485:Allergens; D006632:Histamine; D010047:Ovalbumin
keywords: Airways hyperresponsiveness; Allergic inflammation; Bronchoconstriction; Early asthmatic response; Late asthmatic response; Ovalbumin