We found that the numbers of mast cells in the airway epithelium were increased in some, but not all, of the 26 asthmatic subjects we studied. By classifying the asthmatic subjects as having TH
2-high or TH
2-low asthma based on expression of IL-13–responsive genes in their epithelial brushings, we discovered that IEMC numbers in the TH
2-high subgroup were significantly higher than those in the TH
2-low subgroup. In fact, the numbers of IEMCs in the TH
2-low subgroup were similar to those in the healthy nonasthmatic subjects. We also found that mast cell genes and TH
2 signature genes clustered together in unsupervised hierarchical clustering analyses of the microarray data from epithelial brushings from asthmatic and healthy subjects, lending further support to a link between IEMCs and TH
2 inflammation. Thus our findings here add increases in IEMC numbers to the list of pathologies characteristic of TH
2-high asthma. Other characteristics we have reported include airway and systemic eosinophilia, sub-epithelial fibrosis, and abnormal expression of gel-forming mucins.7
The levels of expression of tryptase
in epithelial brushings in the asthma subgroup correlated positively with lung function improvement after treatment with ICSs. This finding is consistent with the TH
2 subtype of asthma being corticosteroid sensitive,7
and it suggests that some of the clinical improvement from ICSs could be a result of decreased activation of IEMCs.
, a cell-surface receptor on mast cells, was also in this mast cell cluster. SCF is a key ligand for c-kit, and levels of SCF correlate with asthma disease severity.22
We found SCF
expression to be increased among subjects with TH
2-high asthma and also found SCF
expression to be increased at both the gene transcript and protein levels in cultured airway epithelial cells stimulated with IL-13. Interestingly, SCF protein was secreted basally by the epithelial cells, raising the possibility that basal secretion of this mast cell chemokine and growth factor by epithelial cells in response to IL-13 could be a mechanism for mast cell migration from the subepithelial matrix into the intraepithelial space.
We found IEMCs to have upregulation of tryptase and CPA3 but no upregulation of chymase. This pattern of protease expression is not consistent with the patterns observed for MC-T or MC-TC cells. MC-T cells predominantly express high levels of tryptase, but not chymase or carboxypeptidases, whereas MC-TC cells express all 3 proteases.3
To our knowledge, our data for a tryptase-high and CPA3-high subset of mast cells is the first evidence that IEMCs in subjects with mild-to-moderate asthma have a unique phenotype with a different program of gene expression from mast cells in other lung microenvironments. It has previously been shown that mast cells grown in coculture with epithelial cells downregulate chymase.21
We have shown here that CM from IL-13–stimulated epithelial cells downregulate chymase while maintaining CPA3 and tryptase expression in cord blood–derived cultured mast cells. These findings raise the possibility that the TH
2-activated epithelium alters the phenotype of mast cells once they accumulate in this location. Our data contrast with those for IEMCs in mouse intestine, which produce chymase that is important for worm expulsion.23
In addition, increased numbers of chymase-producing mast cells have been reported in the small airways of patients with severe asthma,24
suggesting that mast cell phenotypes in the airway vary by disease severity.
The expression of CPA3 in IEMCs suggests a potentially important role for this protease in asthma. Mast cell carboxypeptidase cleaves C-terminal aromatic and some aliphatic amino acids, and several potential natural substrates have been identified, including angiotensins, apolipoproteins, endothelin, and venoms.25–28
Murine mast cell CPA3 can convert angiotensin I to angiotensin II by removing C-terminal amino acids.29
Angiotensin II is proposed to play a major role in airway bronchoconstriction and enhanced airway hyperresponsiveness in asthmatic subjects.30
It is therefore possible that CPA3 plays a role in asthma through production of angiotensin II. Our finding of CPA3 as a major product of IEMCs should prompt further investigation into the potential biologic role of carboxypeptidases in asthma.
The mechanism for how mast cells accumulate in the epithelium is incompletely understood, although there are clues. For example, mast cells adhere avidly to tracheal epithelium through interactions between epithelial cells and cell-surface proteins on mast cells.31
Human lung mast cells express several integrins and immunoglobulin superfamily adhesion receptors, including CD18, very late antigen 4, and αEβ7.32–34
Normal epithelial cells express important counterligands for these receptors, including intercellular adhesion molecule 1 (ICAM-1), vascular cell adhesion molecule 1, and E-cadherin, respectively.35,36
Mast cell products have several important paracrine effects on airway epithelial cell function. First, tryptases can stimulate production of IL-8 and ICAM-1 by epithelial cells.37
ICAM-1 is the primary receptor for the major group rhinoviruses, an important cause of asthma exacerbations. Second, numerous mast cell products can act as mucin secretagogues, including prostaglandin D2
, chymase, and leukotriene C4
Finally, mast cells can produce TH
2 cytokines, including IL-4 and IL-13,41
which activate specific programs of gene expression by epithelial cells, leading to mucus hypersecretion, eosinophilic inflammation, and airway hyperresponsiveness.42,43
In summary, we conclude that increases in IEMC numbers occurs in subjects with TH2-high asthma but not in subjects with TH2-low asthma and that IEMCs have an unusual phenotype, showing upregulation of tryptases and CPA3 but no upregulation of chymase. These findings highlight an important pathologic abnormality in asthma that has implications for mechanisms of epithelial cell activation.
- IEMCs are a pathologic characteristic of TH2-high asthma, a distinct molecular phenotype of asthma.
- The levels of expression of mast cell genes in the airway epithelium correlate positively with lung function improvements with corticosteroid treatment.
- The mechanism of accumulation of mast cells in the airway epithelium in asthmatic subjects might involve IL-13–stimulated secretion of SCF by airway epithelial cells.