Involvement of mast cells in eosinophilic esophagitis

Abstract

Background: Eosinophilic esophagitis (EE) is an emerging disorder with poorly understood pathogenesis.

Objective: Whereas prior studies have primarily focused on the role of eosinophils in disease diagnosis and pathogenesis, this study investigates the involvement of mast cells.

Methods: Total and degranulated mast cell counts were correlated to microarray and RT-PCR data to generate transcriptome expression profiles related to mast cell number and degranulation in patients with EE and healthy control subjects.

Results: Esophageal mastocytosis and mast cell degranulation were readily apparent in patients with EE compared with control subjects (P < .01), as assessed by staining for total mast cells and the presence of extracellular mast cell tryptase (P < .01). Microarray analysis revealed that mast cell levels correlated with the dysregulation of 0.8% (301 genes) of the genome, which was partially distinct from the genes that correlated with tissue eosinophilia. The expression of transcripts for the mast cell proteases carboxypeptidase A3 and tryptase, but not chymase, correlated with mast cell levels and distinguished patients with EE from control subjects. Suprabasilar mast cell counts (P < .01) and degranulation (P < .01) were proportional with KIT ligand mRNA expression. Treatment of patients with EE with swallowed fluticasone propionate normalized levels of mast cells and the mast cell-related transcriptome in responder patients.

Conclusion: Herein we have identified local mastocytosis and mast cell degranulation in the esophagi of patients with EE; identified an esophageal mast cell-associated transcriptome that is significantly divergent from the eosinophil-associated transcriptome, with carboxypeptidase A3 mRNA levels serving as the best mast cell surrogate marker; and provided evidence for the involvement of KIT ligand in the pathogenesis of EE.

Figure 1. Mast cell counts in EE patient samples versus normal or chronic esophagitis samples

The maximum number of tryptase positive mast cells per high power field in EE samples is increased (mean ± SEM) relative to NL, and CE patients (A). The maximum number of degranulated mast cells per high field is increased in EE relative to NL patients (B). Immunohistochemistry with anti-tryptase antibody in normal and EE patients (40×) (C). High power field (400×) from an EE patient demonstrating an intact mast cell with a dashed arrow and degranulated mast cells with solid arrows (D).

Figure 1. Mast cell counts in EE patient samples versus normal or chronic esophagitis samples

The maximum number of tryptase positive mast cells per high power field in EE samples is increased (mean ± SEM) relative to NL, and CE patients (A). The maximum number of degranulated mast cells per high field is increased in EE relative to NL patients (B). Immunohistochemistry with anti-tryptase antibody in normal and EE patients (40×) (C). High power field (400×) from an EE patient demonstrating an intact mast cell with a dashed arrow and degranulated mast cells with solid arrows (D).

Figure 2. Relationship between mast cell counts and relative expression of CPA3 and tryptase

Mast cell counts were determined by anti-tryptase IHC and correlated against the relative expression CPA3 (r = 0.68, p < 0.01)(A) and tryptase (r = 0.75, p < 0.01)(B) as determined by real-time PCR.

Figure 3. Correlation of CPA3 expression with mast cell gene expression

Correlations between microarray expression of CPA3 and mast cell related genes including: KIT (r = 0.82, p < 0.01), KITLG (r = 0.82, p < 0.01), PGDS (r = 0.91, p < 0.01), CCL26 (r = 0.81, p < 0.01), CXCL6 (r = 0.85, p < 0.01), CXCL1 (r = 0.63, p < 0.01)(A–F).

Figure 3. Correlation of CPA3 expression with mast cell gene expression

Correlations between microarray expression of CPA3 and mast cell related genes including: KIT (r = 0.82, p < 0.01), KITLG (r = 0.82, p < 0.01), PGDS (r = 0.91, p < 0.01), CCL26 (r = 0.81, p < 0.01), CXCL6 (r = 0.85, p < 0.01), CXCL1 (r = 0.63, p < 0.01)(A–F).

Figure 4. Correlation of KITLG expression with intact and degranulated mast cell

The normalized expression of KITLG is plotted against the maximum number of intact mast cells (r = 0.87, p < 0.01)(A) or degranulated mast cells (r = 0.86, p < 0.01) (B).

Figure 5. Mast cell counts, gene expression, and mast cell transcriptome in patients treated with swallowed FP

Maximum mast cell counts from normal, CE, EE, and compared to EE patients whom responded to fluticasone propionate (EE FP Resp), or who did not respond to fluticasone propionate therapy (EE FP Non-Resp). Response to treatment with swallowed FP was defined as ≤ 1 eosinophil per HPF (A). CPA3 expression on microarray (B). The mast cell related-transcriptome under varying conditions listed above (C).

Figure 5. Mast cell counts, gene expression, and mast cell transcriptome in patients treated with swallowed FP

Maximum mast cell counts from normal, CE, EE, and compared to EE patients whom responded to fluticasone propionate (EE FP Resp), or who did not respond to fluticasone propionate therapy (EE FP Non-Resp). Response to treatment with swallowed FP was defined as ≤ 1 eosinophil per HPF (A). CPA3 expression on microarray (B). The mast cell related-transcriptome under varying conditions listed above (C).