Background: Genetic studies have shown that variants in SPINK5 may be associated with atopic diseases and asthma. However, the functional role of SPINK5 protein in asthma has not been elucidated.
Objectives: To determine the effects of SPINK5 on asthma related physiological events such as apoptosis, mucus and cytokine production by epithelial cells.
Methods: A549 cells were transfected with SPINK5 expression vector and stimulated with increasing doses of hydrogen peroxide and neutrophil elastase (NE) for measurement of cell viability or apoptosis and analysis of mucus production. Cell viability was measured by MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5- diphenyl-tetrazolium bromide) assay and apoptosis by Annexin V/PI staining. Levels of IL-4, IL-6, IL-8, IL-12, IL-13, IFNγ, IL-1β and RANTES were determined by ELISA in cell culture supernatants. Mucus production was determined by RT-PCR of the MUC5AC gene and PAS staining in NE treated cells.
Results: Epithelial cells transfected with SPINK5 expression vector produced more IL-6, IL-8 and RANTES compared to non-transfected cells (p < 0.001, p = 0.003, p < 0.001, respectively). Even though cells transfected with SPINK5 vector displayed significantly higher cell death, we have not observed any clear effect of SPINK5 on apoptosis. PAS staining showed that SPINK5 slightly decreased the mucin production induced by neutrophil elastase in A549 cells. However, SPINK5 had no effect on MUC5AC transcription.
Conclusion: SPINK5 is an important molecule in asthma. Its role extends beyond its well known protease inhibitor properties.
Copyright © 2011 Elsevier Ltd. All rights reserved.