Allergen inhalation increases mRNA expression in induced sputum from mild atopic asthmatics

Abstract

Rationale Sputum is increasingly used as a non-invasive method for the assessment of airway inflammation. We investigated whether it is possible to use cells derived from sputum for gene expression analysis by quantitative PCR. Methods Mild atopic asthmatics with allergen-induced dual airway responses were selected. Subjects were required to have stable asthma with FEV1 >70%. Subjects were exposed to allergen inhalation and induced-sputum was collected before and 7 and 24 hrs after allergen exposure. Total number of sputum cells were counted and processed into RNA and subsequently converted into cDNA. The mRNA expression of genes involved in Th2-inflammation was measured by quantitative PCR analysis. All values were normalized to the expression of house-keeping gene ubiquitin. Results Allergen inhalation induced upregulation of the Th2 genes IL-4, IL-5, and IL-13. The highest increase in expression was observed 7 hrs after allergen exposure and values returned to baseline levels within 24 hrs. Increased expression of CCR3 mRNA after allergen exposure was observed, indicative of increased numbers of eosinophils in these samples. In addition, mRNA expression of the Th2-attracting chemokines TARC and MDC was upregulated. TARC upregulation was maximum at 7 hrs post allergen, and MDC mRNA expression peaked at 24 hrs. Repeated allergen exposure in the same patients demonstrated a consistent level of increased Th2-gene expression. Conclusions The ability to evaluate the expression of a wide variety of genes in sputum samples opens up a new and promising way to assess airway inflammation in allergic asthma patients.