Introduction: Alveolar macrophages (AMs) are dominant airway cells and have been strongly implicated in the development and progression of asthmatic disease. Macrophage polarisation and function is influenced by exposure to the Th-2 cytokines IL-4 and IL-13; these cytokines are associated with allergic asthma and have been shown to be elevated following exposure to house dust mite (HDM) allergen.
Aim: Examine changes in surface marker expression and cytokine production by AMs from ‘resting’ naïve (pre-challenge AMs), and ‘resting’ but allergen-exposed lungs (post-challenge AMs).
Methods: Bronchoalveolar lavage (BAL) samples were collected from HDM-sensitised sheep (n=7) on 2 separate occasions; before (pre-challenge AMs), as well as 2 weeks after sheep had been given 4x HDM airway challenges at 3-week intervals (post-challenge AMs). BAL adherent cells (AMs) were cultured in the presence of innate stimulators (LPS, Poly I:C, or PAMCys3K) or the cytokines IL-4 or IL-13 for 24h; IL-6, TNF-α and IL-10 production in vitro was assessed by ELISA. In separate experiments, freshly collected AMs were stained with surface markers and analysed by flow cytometry.
Results: IL-6 levels were low or undetected in culture supernatants with no discernible differences comparing pre- and post-challenge AMs or responses to the different stimuli in vitro. Post-challenge AMs cultured in the presence of LPS showed increased TNF-α production (p<0.05) but decreased IL-10 production (p<0.05) compared to pre-challenge AMs. IL-10 secretion was also lower in post-challenge AMs following stimulation with the TLR-2 agonist PAMCys3K. Conversely, AM cultures containing IL-13 induced higher levels of IL-10 secretion in post- compared to pre-challenge AMs (p<0.05). There were no significant changes in MHC class II, CD1a, CD11a, CD11b, CD11c, CD44 or CD69 expression on pre- compared to post-challenge AMs.
Conclusions: No phenotypic changes were detected in resting AMs collected from naïve compared to allergen-challenged airways. However, an altered cytokine profile of AMs (pre- vs post-challenge) indicates key changes in the resident AM population following allergen exposure in the HDM sheep asthma model.