Comparison of cytokine mRNA expression in the bronchoalveolar lavage fluid of horses with inflammatory airway disease and bronchoalveolar lavage mastocytosis or neutrophilia using REST software analysis

Abstract

Objectives: The pathophysiology of inflammatory airway disease (IAD) is unknown, but in some cases involves the accumulation of mast cells, neutrophils, or both in the bronchoalveolar lavage fluid (BALF). The objective of this study was to characterize cytokine gene expression in the BALF cells of horses with IAD, including a comparison of cytokine gene expression between IAD horses with increased BALF mast cells (IAD-Mast) or neutrophils (IAD-Neutro).

Methods: The mRNA expression of IL-4, IFN-γ, IL-17, IL-8, IL-1β, IL-5, IL-6, IL-10, IL-12p35, and eotaxin-2 was studied by quantitative polymerase chain reaction (QPCR) with efficiency correction in BALF samples of 17 horses with IAD (IAD-total), also subcategorized as 8 IAD-Mast and 9 IAD-Neutro, and 10 controls. Glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was used as a reference gene. Relative expression software tool (REST) analysis provided ratios of expression, statistical analysis, and confidence intervals for the results.

Results: Compared with the control group, IL-5, IL-1β, IL-6, IL-8, and IL-10 mRNA expression was upregulated 3.5-, 3.4-, 2.8-, 2.2-, and 1.9-fold, respectively, in the IAD-total group. The IAD-Neutro group showed increased expression of IL-17, IL-8, and IL-5 (4.7-, 2.5-, and 2.9-fold, respectively) and a decreased expression of IL-4 (3.4-fold) compared with the IAD-Mast group.

Conclusion: Cytokines from the Th2 family plays a key role in IAD and a different pathophysiology may be involved in mast cell versus neutrophil BALF accumulation in IAD horses.