上海交通大学学报（医学版）

上海交通大学学报（医学版）

• 论著（基础研究） • 上一篇    下一篇

脾酪氨酸激酶对氧化应激诱导的气道上皮细胞黏液高分泌的影响

冉丹华1,陈灵修1，黄华萍2,韩忠2,周向东1，2   

  1. 1.重庆医科大学附属第二医院呼吸内科， 重庆 400010； 2.海南医学院附属医院， 海口 570102
  • 出版日期:2016-02-28 发布日期:2016-03-29
  • 通讯作者: 周向东, 电子信箱: zxd999@263.net。
  • 作者简介:冉丹华（1990—）， 女， 硕士生; 电子信箱: rdh900831@163.com。
  • 基金资助:

    国家自然科学基金（81370111， 31171346）

Effects of spleen tyrosine kinase on oxidative stress-induced mucous hypersecretion of airway epithelial cells

RAN Dan-hua1, CHEN Ling-xiu1, HUANG Hua-ping2, HAN Zhong2, ZHOU Xiang-dong1,2   

  1. 1.Department of Respiratory Medicine， The Second Affiliated Hospital， Chongqing Medical University, Chongqing 400010， China; 2.Affiliated Hospital of Hainan Medical University, Haikou 570102, China
  • Online:2016-02-28 Published:2016-03-29
  • Supported by:

    National Natural Science Foundation of China, 81370111, 31171346

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摘要：

目的 研究脾酪氨酸激酶（Syk）在氧化应激诱导的气道黏液高分泌中的作用。方法 体外培养人气道上皮细胞（16HBE），用H2O2刺激细胞构建黏液高分泌模型，以Syk siRNA、丝裂原活化蛋白激酶(MAPK)信号通路中的细胞外信号调节激酶（ERK1/2）特异性抑制剂U0126及P38特异性抑制剂SB203580为干预因素，将细胞随机分为对照组、H2O2刺激组、H2O2+Syk siRNA组、H2O2+阴性siRNA组、H2O2+U0126组及H2O2+SB203580组。四甲基偶氮唑盐法（MTT）测试细胞活力；RT-PCR和ELISA法分别检测黏蛋白(MUC)5AC的转录及分泌水平；Western blotting法检测Syk、磷酸化ERK1/2 (p-ERK1/2)和磷酸化P38(p-P38)蛋白表达水平；激光共聚焦技术检测MUC5AC蛋白的含量和分布。结果 H2O2刺激后Syk、p-ERK1/2、p-P38、MUC5AC表达水平显著高于对照组（均P=0.000），而利用Syk siRNA、SB203580和U0126处理后，H2O2导致的上述效应明显得到抑制（均P=0.000）。结论 Syk通过ERK1/2、P38 MAPK信号通路，参与氧化应激诱导的气道黏液高分泌。

关键词: 脾酪氨酸激酶, 信号通路, 黏蛋白5AC

Abstract:

Objective To explore the effects of spleen tyrosine kinase (Syk) on oxidative stress-induced airway mucous hypersecretion. Methods Human airway epithelial cells 16HBE were cultured in vitro. The mucous hypersecretion model was established by stimulating cells by H2O2. Intervention factors were Syk siRNA and extracellular signal regulated kinase (ERK1/2) specific inhibitor U0126 and P38 specific inhibitor SB203580 of mitogen-activated protein kinase (MAPK) signaling pathway. The cells were randomly divided into the control group, H2O2 group, H2O2+Syk siRNA group, H2O2+negative siRNA group, H2O2+U0126 group, and H2O2+SB203580 group. The cell viability was detected by MTT assay. Transcription and protein expression of mucin (MUC) 5AC were detected by RT-PCR and ELISA respectively. The protein expression of Syk, phosphorylated ERK1/2 (p-ERK1/2), and phosphorylated P38 (p-P38) were detected by Western blotting. The content and distribution of MUC5AC protein were detected by confocal laser technology. Results Compared with the control group, expressions of Syk, p-ERK1/2, p-P38, and MUC5AC significantly increased after being stimulated by H2O2 (P=0.000). Above effects of H2O2 were significantly inhibited after being treated by Syk siRNA, U0126, and SB203580 (P=0.000). Conclusion Syk involves the oxidative stress induced airway mucous hypersecretion via ERK1/2 and P38 MAPK signaling pathways.

Key words: spleen tyrosine kinase, signaling pathway, MUC5AC