Abstract
BackgroundThe association of eosinophils with inflammation and tissue remodeling is at least partially due to their release of toxic granule proteins and other mediators, including cytokines. Tissue remodeling and consequent functional defects are affected by activity of connective tissue fibroblasts. Exaggerated fibroblast activation, accumulation and change of phenotype may lead to fibrosis and loss of tissue function. So far, little information has been reported on how eosinophils affect inflammation and tissue remodeling via the activation of fibroblasts. We have recently shown that eosinophil activation with IL-3 led to a robust eosinophil degranulation on immunoglobin-G (IgG) coated plates. Thus, in the present study, we analyze the effects of IL-3-activated eosinophil degranulation products on primary human lung fibroblasts (HLF) using whole transcriptome sequencing.MethodsConditioned media was obtained from eosinophils that were pre-activated with IL-3 or IL-5 and subsequently cultured for 6 h on IgG to induce degranulation. This conditioned media was added on human lung fibroblasts (HLF) for 24 h and the cell lysates were then subjected to whole transcriptome sequencing to identify global changes in gene expression. Differentially expressed genes were analyzed using the Ingenuity Pathway Analysis (IPA), and validated by qPCR.ResultsIn HLF, the expression level of 300 genes was changed by conditioned media from IL-3-activated eosinophils compared to control fibroblast cultures. Among these 300 genes, the expression level of 35 genes coding for known proteins was upregulated by IL-3- versus IL-5-pre-activated eosinophils. Of the 35 upregulated genes, IPA identified C3, CH25H, CXCL1, CXCL8, CYP1A1, ICAM1, IL6 and UCN2 as having downstream functions on inflammation, tissue remodeling and lipid synthesis. This analysis combined with previous RNA sequencing analyses of eosinophils suggest IL-1ß, OSM and TNFSF12 as potential upstream regulators of fibroblasts.ConclusionsThis study has identified several novel pro-inflammatory and pro–remodeling mediators produced by fibroblasts in response to activated eosinophils. These findings may have significant implications on the role of eosinophil/fibroblast interactions in eosinophilic disorders.
Highlights
The association of eosinophils with inflammation and tissue remodeling is at least partially due to their release of toxic granule proteins and other mediators, including cytokines
Generation of conditioned media from degranulated eosinophils Eosinophils activated for 20 h with IL-3 or IL-5 were added on coated heat-aggregated (HA)-human serum IgG for 6 h (IL3IgG and IL-5 on HA-IgG (IL5IgG))
The morphology of the eosinophils pre-activated with IL-3 or IL-5 and seeded on coated human IgG (HA-IgG) is shown on Additional file 2: Figure E1, where pictures of cultures were taken at 2 h, 4 h and 6 h
Summary
The association of eosinophils with inflammation and tissue remodeling is at least partially due to their release of toxic granule proteins and other mediators, including cytokines. Evidence suggests that eosinophils contribute to asthma pathogenesis via the release of cytotoxic granule proteins and other mediators, including cytokines and lipids [5, 6]. The release of these mediators from eosinophils leads to tissue damage and recruitment/activation of other cell types, including fibroblasts. The structural changes of airway remodeling result in airflow limitation and are associated with increased asthma severity [7]. Airway remodeling includes epithelial changes, angiogenesis, and increased smooth muscle mass, and activation of fibroblasts to differentiate into pro-inflammatory-producing fibroblasts and extracellular matrix (ECM)-producing myofibroblasts [8]
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