Background Eosinophil cationic protein is a clinical asthma biomarker that would be released into blood, especially gathered in bronchia. The signal peptide of eosinophil cationic protein (ECPsp) plays an important role in translocating ECP to the extracellular space. We previously reported that ECPsp inhibits microbial growth and regulates the expression of mammalian genes encoding tumor growth factor-a (TGF-a) and epidermal growth factor receptor (EGFR). Results In the present study, we first generated a DNA microarray dataset, which showed that ECPsp upregulated proinflammatory molecules, including chemokines, interferon-induced molecules, and Toll-like receptors. The levels of mRNAs encoding CCL5, CXCL10, CXCL11, CXCL16, STAT1, and STAT2 were increased in the presence of ECPsp by 2.07-, 4.21-, 7.52-, 2.6-, 3.58-, and 1.67-fold, respectively. We then generated a functional linkage network by integrating the microarray dataset with the pathway database of Kyoto Encyclopedia of Genes and Genomes. This revealed that STAT1[/2], an important transcriptional factor that regulates cytokine expression and release, served as a hub to connect the pathways of cytokine stimulation (TGF-a and EGFR pathways) and inflammatory responses. Furthermore, integrating TGF-a and EGFR with the functional linkage network indicated that STAT1 served as a hub that connects two functional clusters, including (1) cell proliferation and survival, and (2) inflammation. Finally, we found that conditioned medium in which cells that express ECPsp had been cultured could chemoattract macrophages. Therefore, we hypothesize that ECPsp may regulate the migration of macrophages in vivo. Conclusion The increased expression and release of various cytokines triggered by ECPsp may attract macrophages to bronchia to purge damaged cells. Our approach, involving experimental and computational systems biology, predicts pathways and potential biological functions for further characterization of this novel function of ECPsp under inflammatory conditions.
Overall design: The control group of this study is Beas-2B cells treated with pEGFP-C1, and the experiment group is the same cell line treated with pEGFPN1-ECPsp. Each group was conducted by two biological repeats and two technical repeats were done in DNA microarray analyses. On the microarray chip, 29,187 probes correspond to the annotated genes in the RefSeq v38 and Ensembl v56 databases. Furthermore, 1,088 control probes are also included for monitoring the sample quality and the hybridization process.
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