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Interleukin-4-induced posttranscriptional gene regulation of CCL26 by the RNA-binding protein HuR in primary human nasal polyp-derived epithelial cells.
International Forum of Allergy & Rhinology 2018 November 25
BACKGROUND: Much attention on the pathophysiology of nasal polyp (NP) has focused on eosinophils. Interleukin (IL)-4 and eotaxin-3 (C-C motif chemokine ligand 26, or CCL26) levels have been reported to be increased in eosinophilic nasal polyps. The aim of this study was to characterize CCL26 posttranscriptional regulation by the RNA-binding protein HuR in primary human nasal polyp-derived epithelial cells (hNPDECs) challenged with IL-4.
METHODS: A prospective, observational study was conducted. Nasal polyp tissues were obtained from eosinophilic (n = 12) and non-eosinophilic (n = 10) NP patients, and inferior turbinate (IT) tissues were taken from control subjects (n = 9) and cultured into hNPDECs. Expression of HuR and CCL26 were measured by immunohistochemistry, Western blot analysis, enzyme-linked immunoassay, and real-time polymerase chain reaction (PCR). The nucleocytoplasmic shuttling of HuR in hNPDECs was detected by immunofluorescence. Posttranscriptional regulation of CCL26 by HuR was tested by ribonucleoprotein immunoprecipitation assay (RIP) and dual-luciferase reporter assay. CCL26 mRNA stabilization was measured by quatititative PCR after treatment with actinomycin D. Student's t test and one-way analysis of variance were used.
RESULTS: Immunohistochemical data show that both HuR and CCL26 were highly expressed in NP tissues, especially eosinophilic NP tissues (p < 0.05). IL-4 stimulation increased CCL26 mRNA stability, and overexpression and knockdown of HuR affected CCL26 expression. Immunofluorescence data indicate that IL-4 altered the subcellular distribution of HuR. The RIP and dual-luciferase reporter assay results supply strong evidence for HuR binding to CCL26.
CONCLUSION: Our results provide strong support for the hypothesis that IL-4-induced expression of CCL26 in hNPDECs relies partly on CCL26 mRNA stabilization mediated by the interaction of HuR with CCL26 3'UTR.
METHODS: A prospective, observational study was conducted. Nasal polyp tissues were obtained from eosinophilic (n = 12) and non-eosinophilic (n = 10) NP patients, and inferior turbinate (IT) tissues were taken from control subjects (n = 9) and cultured into hNPDECs. Expression of HuR and CCL26 were measured by immunohistochemistry, Western blot analysis, enzyme-linked immunoassay, and real-time polymerase chain reaction (PCR). The nucleocytoplasmic shuttling of HuR in hNPDECs was detected by immunofluorescence. Posttranscriptional regulation of CCL26 by HuR was tested by ribonucleoprotein immunoprecipitation assay (RIP) and dual-luciferase reporter assay. CCL26 mRNA stabilization was measured by quatititative PCR after treatment with actinomycin D. Student's t test and one-way analysis of variance were used.
RESULTS: Immunohistochemical data show that both HuR and CCL26 were highly expressed in NP tissues, especially eosinophilic NP tissues (p < 0.05). IL-4 stimulation increased CCL26 mRNA stability, and overexpression and knockdown of HuR affected CCL26 expression. Immunofluorescence data indicate that IL-4 altered the subcellular distribution of HuR. The RIP and dual-luciferase reporter assay results supply strong evidence for HuR binding to CCL26.
CONCLUSION: Our results provide strong support for the hypothesis that IL-4-induced expression of CCL26 in hNPDECs relies partly on CCL26 mRNA stabilization mediated by the interaction of HuR with CCL26 3'UTR.
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