Fig 1: Analysis of VIP and VIP receptors VPAC-1, VPAC-2, and CRTH2 receptors on human blood eosinophils and in vitro-generated mast cells and their migration in response to VIP. Anti-VIP, anti-VAPC1, anti-VACP2, and anti-CRTH2 antibodies were used to assess VIP and VIP-associated receptor (VAPC1, VACP2, and CRTH2) expression in human eosinophils with flow cytometry analysis (a–d). Further, the VIP and VIP receptors (VAPC1, VACP2, and CRTH2) were also analyzed in in vitro-generated mast cells from human bone marrow using flow cytometry (e–h). Furthermore, the analysis of the chemoattractant assay in response to dose-dependent VIP, PDG-2, and eotaxins-3 (i). Anti-CRTH2 antibody inhibited VIP-induced eosinophil migration in vitro but had no effect on eotaxin-2-induced eosinophil migration. These results demonstrate that the interaction between CRTH2 and VIP is essential for eosinophil motility (j(i)). Anti-CRTH2 antibody inhibits VIP–induced mast cell migration in vitro (j(ii)). Data are expressed as mean ± SD, n = 3 experiments. Migration data are presented as an average of three separate experiments. Data involving two groups were analyzed with a Mann–Whitney test; for more than two groups, Kruskal–Wallis test was used. Significance was set at p < 0.05 and p < 0.01, NS—no significant respectively.
Fig 2: Transcript analysis of VIP and correlation of VIP-associated receptors in esophageal biopsies of human EoE. ELISA analysis indicated that expression levels of VIP protein and mRNA (a,b) and VIP receptors VAPC1 and CRTH2 were higher in EoE and dysphagia biopsies compared to controls (c,d). A significant statistical transcript correlation between VIP and CRTH2 (p < 0.0001, r = 0.71) (e), as well as between CRTH2 and eosinophils (p < 0.005, r = 0.4), was observed in EoE patients (f). Significance was calculated using Mann–Whitney test for two groups, and Kruskal–Wallis for more than two groups, and correlation (r2) values were calculated using the Mann–Whitney test with the Bonferroni correction. Data are expressed as mean ± SD, n = 8–12. Each data point represents one individual patient.
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