C-C基序趋化因子受体3介导的细胞外信号调节激酶1/2和p38丝裂原活化蛋白激酶信号传导：嗜酸性粒细胞趋化因子-2和嗜酸性粒细胞趋化因子-3诱导人气道上皮黏蛋白5AC的潜在靶点
C-C Motif Chemokine Receptor 3-Mediated Extracellular Signal-Regulated Kinase 1/2 and p38 Mitogen-Activated Protein Kinase Signaling: Promising Targets for Human Airway Epithelial Mucin 5AC Induction by Eotaxin-2 and Eotaxin-3.
作者信息
Jo Sooyeon, Na Hyung Gyun, Choi Yoon Seok, Bae Chang Hoon, Song Si-Youn, Kim Yong-Dae
机构信息
Department of Otorhinolaryngology-Head and Neck Surgery, College of Medicine, Yeungnam University, Daegu, Republic of Korea,
Department of Otorhinolaryngology-Head and Neck Surgery, College of Medicine, Yeungnam University, Daegu, Republic of Korea.
出版信息
Int Arch Allergy Immunol. 2023;184(9):893-902. doi: 10.1159/000531911. Epub 2023 Aug 8.
INTRODUCTION
Eotaxin-2 and -3 of the C-C chemokine subfamily function as potent chemoattractant factors for eosinophil recruitment and various immune responses in allergic and inflammatory airway diseases. Mucin 5AC (MUC5AC), a major gel-forming secretory mucin, is overexpressed in airway inflammation. However, the association between mucin secretion and eotaxin-2/3 expression in the upper and lower airway epithelial cells has not been fully elucidated. Therefore, in this study, we investigated the effects of eotaxin-2/3 on MUC5AC expression and its potential signaling mediators.
METHODS
We analyzed the effects of eotaxin-2 and -3 on NCI-H292 human airway epithelial cells and primary human nasal epithelial cells (HNEpCs) via reverse transcription-polymerase chain reaction, enzyme-linked immunosorbent assay, and western blotting. Along with immunoblot analyses with specific inhibitors and small interfering RNA (siRNA), we explored the signaling pathway involved in MUC5AC expression following eotaxin-2/3 treatment.
RESULTS
In HCI-H292 cells, eotaxin-2/3 activated the mRNA expression and protein production of MUC5AC. A specific inhibitor of C-C motif chemokine receptor 3 (CCR3), SB328437, suppressed eotaxin-2/3-induced MUC5AC expression at both the mRNA and protein levels. Eotaxin-2/3 induced the phosphorylation of extracellular signal-regulated kinase (ERK)-1/2 and p38, whereas pretreatment with a CCR3 inhibitor significantly attenuated this effect. Induction of MUC5AC expression with eotaxin-2/3 was decreased by U0126 and SB203580, specific inhibitors of ERK1/2 and p38 mitogen-activated protein kinase (MAPK), respectively. In addition, cell transfection with ERK1/2 and p38 siRNAs inhibited eotaxin-2/3-induced MUC5AC expression. Moreover, specific inhibitors (SB328437, U0126, and SB203580) attenuated eotaxin-2/3-induced MUC5AC expression in HNEpCs.
CONCLUSION
Our results imply that CCR3-mediated ERK1/2 and p38 MAPK are involved in the signal transduction of eotaxin-2/3-induced MUC5AC overexpression.
引言
C-C趋化因子亚家族的嗜酸性粒细胞趋化因子-2和-3在过敏性和炎症性气道疾病中作为嗜酸性粒细胞募集和各种免疫反应的有效趋化因子发挥作用。粘蛋白5AC（MUC5AC）是一种主要的凝胶形成分泌性粘蛋白，在气道炎症中过度表达。然而，上下气道上皮细胞中粘蛋白分泌与嗜酸性粒细胞趋化因子-2/3表达之间的关联尚未完全阐明。因此，在本研究中，我们研究了嗜酸性粒细胞趋化因子-2/3对MUC5AC表达的影响及其潜在的信号传导介质。
方法
我们通过逆转录-聚合酶链反应、酶联免疫吸附测定和蛋白质印迹分析了嗜酸性粒细胞趋化因子-2和-3对NCI-H292人气道上皮细胞和原代人鼻上皮细胞（HNEpC）的影响。除了用特异性抑制剂和小干扰RNA（siRNA）进行免疫印迹分析外，我们还探索了嗜酸性粒细胞趋化因子-2/3处理后参与MUC5AC表达的信号通路。
结果
在HCI-H292细胞中，嗜酸性粒细胞趋化因子-2/3激活了MUC5AC的mRNA表达和蛋白质产生。C-C基序趋化因子受体3（CCR3）的特异性抑制剂SB328437在mRNA和蛋白质水平上均抑制了嗜酸性粒细胞趋化因子-2/3诱导的MUC5AC表达。嗜酸性粒细胞趋化因子-2/3诱导细胞外信号调节激酶（ERK）-1/2和p38磷酸化，而用CCR3抑制剂预处理可显著减弱这种作用。分别用ERK1/2和p38丝裂原活化蛋白激酶（MAPK）的特异性抑制剂U0126和SB203580处理后，嗜酸性粒细胞趋化因子-2/3诱导的MUC5AC表达降低。此外，用ERK1/2和p38 siRNA转染细胞可抑制嗜酸性粒细胞趋化因子-2/3诱导的MUC5AC表达。此外，特异性抑制剂（SB328437、U0126和SB203580）减弱了嗜酸性粒细胞趋化因子-2/3诱导的HNEpC中MUC5AC的表达。
结论
我们的结果表明，CCR3介导的ERK1/2和p38 MAPK参与了嗜酸性粒细胞趋化因子-2/3诱导的MUC5AC过表达的信号转导。
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