Increased mast cell density and airway responses to allergic and non-allergic stimuli in a sheep model of chronic asthma

Abstract

Background: Increased mast cell (MC) density and changes in their distribution in airway tissues is thought to contribute significantly to the pathophysiology of asthma. However, the time sequence for these changes and how they impact small airway function in asthma is not fully understood. The aim of the current study was to characterise temporal changes in airway MC density and correlate these changes with functional airway responses in sheep chronically challenged with house dust mite (HDM) allergen.

Methodology/principal findings: MC density was examined on lung tissue from four spatially separate lung segments of allergic sheep which received weekly challenges with HDM allergen for 0, 8, 16 or 24 weeks. Lung tissue was collected from each segment 7 days following the final challenge. The density of tryptase-positive and chymase-positive MCs (MC(T) and MC(TC) respectively) was assessed by morphometric analysis of airway sections immunohistochemically stained with antibodies against MC tryptase and chymase. MC(T) and MC(TC) density was increased in small bronchi following 24 weeks of HDM challenges compared with controls (P<0.05). The MC(TC)/MC(T) ratio was significantly increased in HDM challenged sheep compared to controls (P<0.05). MC(T) and MC(TC) density was inversely correlated with allergen-induced increases in peripheral airway resistance after 24 weeks of allergen exposure (P<0.05). MC(T) density was also negatively correlated with airway responsiveness after 24 challenges (P<0.01).

Conclusions: MC(T) and MC(TC) density in the small airways correlates with better lung function in this sheep model of chronic asthma. Whether this finding indicates that under some conditions mast cells have protective activities in asthma, or that other explanations are to be considered requires further investigation.

Figure 1. Tryptase-positive mast cell (MC_T) density following challenge with house dust mite (HDM) allergen.

Bars show the number of MC_T per mm² in the airway wall of bronchi in four spatially separate lung segments from control and HDM challenged groups. The number in italics indicates the number of HDM challenges each lung segment received. Line shows the mean, boxes show 25^th and 75^th percentile, whiskers represent 5^th to 95^th percentile. n = 7, *P<0.05. R – right; L – left.

Figure 2. Airway sections stained with antibodies against mast cell tryptase and chymase.

(a) Tryptase-positive mast cells and (b) chymase-positive mast cells. The house dust mite (HDM) treated segment received 24 weekly challenges and the control segment was untreated. Negative control shows omission of the primary antibody. Arrows indicate positive cells.

Figure 3. Chymase-positive mast cell (MC_TC) density increases following chronic allergen challenge.

(a) MC_TC per mm² of airway wall (b) ratio of MC_TC and tryptase-positive mast cells (MC_T). The house dust mite (HDM) treated segment received 24 weekly challenges and the control segment was untreated. Line shows the mean, boxes show 25^th and 75^th percentile, whiskers represent 5^th to 95th percentile. n = 7, *P<0.05, **P<0.01 compared to controls.

Figure 4. Relationship between mast cell density and peak early bronchoconstriction response.

Correlation between the percentage increase in peripheral resistance (R_p) 30 minutes post-HDM challenge and (a) MC_T (r_s = −0.89, P<0.05) and (b) MC_TC (r_s = −0.82, P<0.05).

Figure 5. Relationship between mast cell density and airway responsiveness.

Correlation between the percent dose of methacholine required to increase R_p by 100% (PC₁₀₀[MCh]) from baseline and (a) MC_T (r_s = 0.92, P<0.01) and (b) MC_TC.

Figure 6. Airway remodeling and eosinophils following chronic allergen challenge.

(a) Airway smooth muscle (ASM) per mm basement membrane (BM) length (b) collagen area per mm BM and (c) eosinophils per mm² airway wall, in untreated control sheep and sheep challenged with house dust mite (HDM) allergen weekly for 24 weeks. Line shows the mean, boxes show 25th and 75th percentile, whiskers represent 5th to 95th percentile. n = 7, *P<0.05, ***P<0.001 compared to controls.