p38/MAPK信号通路在高渗透压破坏角膜上皮屏障功能中的作用

秦爽; 周文君; 李华; 宋胜仿

doi:10.16016/j.1000-5404.201707055

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p38/MAPK信号通路在高渗透压破坏角膜上皮屏障功能中的作用

基础医学 | 更新时间：2026-01-09

- p38/MAPK信号通路在高渗透压破坏角膜上皮屏障功能中的作用
- Role of p38/MAPK pathway in hyperosmosis-induced functional impairment of human corneal epithelial barrier in vitro
- 第三军医大学学报 2017年39卷第23期页码：2282-2288
- 作者机构：
  
  402160 重庆，重庆医科大学附属永川医院眼科
- 作者简介：
  
  [宋胜仿，E-mail: ssf868591@163.com SONG Shengfang, E-mail: ssf868591@163.com
- 基金信息：
  
  重庆市基础科学与前沿技术研究项目(CSTC2016jcyjA0249);重庆医科大学附属永川医院研究生创新基金(YJSCX201602)
- DOI：10.16016/j.1000-5404.201707055
  中图分类号： R322.91;R329.24;R363.22
- 收稿：2017-07-06，
  
  修回：2017-09-09，
  
  纸质出版：2017-12-15
- 稿件说明：
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秦爽, 周文君, 李华, 等. p38/MAPK信号通路在高渗透压破坏角膜上皮屏障功能中的作用[J]. 第三军医大学学报, 2017,39(23):2282-2288.

Shuang QIN, Wenjun ZHOU, Hua LI, et al. Role of p38/MAPK pathway in hyperosmosis-induced functional impairment of human corneal epithelial barrier in vitro[J]. Journal of Third Military Medical University, 2017, 39(23): 2282-2288.
秦爽, 周文君, 李华, 等. p38/MAPK信号通路在高渗透压破坏角膜上皮屏障功能中的作用[J]. 第三军医大学学报, 2017,39(23):2282-2288. DOI： 10.16016/j.1000-5404.201707055.

Shuang QIN, Wenjun ZHOU, Hua LI, et al. Role of p38/MAPK pathway in hyperosmosis-induced functional impairment of human corneal epithelial barrier in vitro[J]. Journal of Third Military Medical University, 2017, 39(23): 2282-2288. DOI： 10.16016/j.1000-5404.201707055.

摘要

目的

探讨高渗透压对角膜上皮屏障功能的影响及p38/MAPK信号通路在此过程中潜在的作用。

方法

体外培养人永生化角膜上皮细胞(human corneal epithelial cells，HCEC)，用不同浓度渗透压[302(正常渗透压)、350、500 mOsm/L]作用于HCEC细胞24 h，CCK-8法检测细胞增殖活性；免疫荧光检测细胞紧密连接蛋白ZO-1和Claudin-1的表达和分布；p38/MAPK阻断剂预处理HCEC细胞前后，Western blot分别检测紧密连接蛋白ZO-1、Claudin-1和p38/MAPK信号通路表达及其磷酸化水平；跨膜电阻仪检测上皮细胞跨膜电阻(transepithelial electrical resistance

TEER)的变化。

结果

高渗透压对HCEC细胞增殖活性没有影响；免疫荧光染色可见正常渗透压组ZO-1、Claudin-1沿着细胞膜呈线性分布，而350、500 mOsm/L高渗透压组ZO-1、Claudin-1荧光信号均减弱，且细胞间紧密连接结构破坏(

0.05)；在高渗透压下，HCEC细胞紧密连接蛋白ZO-1、Claudin-1表达量明显下降(

0.05)，p38/MAPK磷酸化水平增加(

0.05)，上皮细胞跨膜电阻(TEER)降低(

0.05)，而p38/MAPK阻断剂SB203580可抑制高渗透压引起的上述反应(

0.05)。

结论

高渗透压可以使人角膜上皮细胞屏障功能破坏，其机制为通过激活p38/MAPK信号通路导致紧密连接蛋白ZO-1、Claudin-1表达下降。

Abstract

Objective

To study the effect of hyperosmosis on corneal epithelial barrier function and explore the role of p38/MAPK pathway in this process.

Methods

Human corneal epithelial cells (HCECs) cultured in normal osmotic (302 mOsm/L) or in hyperosmotic (350 and 500 mOsm/L) conditions for 24 h were examined for cell proliferation using CCK-8 assay and for distribution of tight junction proteins zonula occludens 1 (ZO-1) and claudin-1 using immunofluorescence analysis. Western blotting was used to detect the expression of ZO-1

claudin-1

p38 and phosphorylation of p38 in the cells cultured in different osmotic conditions with or without prior treatment of 10 μg/L SP203580

a p38/MAPK specific inhibitor. The transepithelial electrical resistance (TEER) of the cells was measured using a transmembrane resistance meter.

Results

Hyperosmosis produced no obvious effect on HCEC cell proliferation. Immunofluorescence assay showed that in the cells cultured in normal osmotic condition

ZO-1 and claudin-1 emitted fluorescence in a linear pattern along the cell membrane

while the cells cultured at 350 or 500 mOsm/L exhibited a significantly lowered intensity of the fluorescence signals with obvious structural collapse of the tight junctions (

0.05). In hyperosmotic conditions

the cells expressed significantly decreased levels of tight junction proteins ZO-1 and claudin-1 (

0.05) with significantly increased phosphorylation of p38 (

0.05) and reduced TEER (

0.05). Treatment of the cells with the p38 inhibitor (SB203580) prior to hyperosmotic exposures obviously suppressed these responses to hyperosmosis (

0.05).

Conclusions

Hyperosmosis induces impairment of corneal epithelial barrier function by activating the p38/MAPK pathway to cause down-regulation of the tight junction proteins ZO-1 and claudin-1.

关键词

Keywords

references

Definition and Classfication Subcommittee members. The definition and classification of dry eye disease: report of the Definition and Classification Subcommittee of the International Dry Eye WorkShop (2007)[J]. Ocul Surf, 2007, 5(2): 75-92. DOI: 10.1016/S1542-0124(12)70081-2.

林泉, 刘伟民.干眼诊断的研究进展[J].中国临床新医学, 2016, 9(3): 269-274. DOI: 10.3969/j.issn.1674-3806.2016.03.28.

LIN Q, LIU W M. Research progress of diagnosis of dry-eye disease[J]. Chin J New Clin Med, 2016, 9(3): 269-274. DOI: 10.3969/j.issn.1674-3806.2016.03.28.

WARCOIN E, CLOUZEAU C, BRIGNOLE-BAUDOUIN F, et al . Hyperosmolarity: intracellular effects and implication in dry eye disease[J ] . J Fr Ophtalmol, 2016, 39(7): 641-651. DOI: 10.1016/j.jfo.2016.07.006.

李晶. 一种新的干眼病模型——高渗盐水点眼小鼠干眼病模型的建立与评估[D]. 重庆: 第三军医大学, 2010.

LI J. A new dry eye disease model:evaluation of A murine model of dry eye disease induced by topical hyperosmolar saline[D]. Chongqing: Third Military Medical University, 2010.

YAGHI A, ZAMAN A, DOLOVICH M B. The direct effect of hyperosmolar agents on ciliary beating of human bronchial epithelial cells[J]. J Aerosol Med Pulm Drug Deliv, 2012, 25(2): 88-95. DOI: 10.1089/jamp.2011.0914.

HUANG S M, WU C S, CHAO D, et al . High-glucose-cultivated peripheral blood mononuclear cells impaired keratinocyte function via reduced IL-22 expression: implications on impaired diabetic wound healing[J ] . Exp Dermatol, 2015, 24(8): 639-641. DOI:10.1111/exd.12733..

KATSUYAMA I, ARAKAWA T. A convenient rabbit model of ocular epithelium damage induced by osmotic dehydration[J]. J Ocul Pharmacol Ther, 2003, 19(3): 281-289. DOI:10.1089/108076803321908400.

LEONG Y Y, TONG L. Barrier function in the ocular surface: from conventional paradigms to new opportunities[J]. Ocul Surf, 2015, 13(2): 103-109. DOI: 10.1016/j.jtos. 2014.10.003.

李方辉, 薛少青, 修焕娟, 等. JNK/SAPK和p38信号转导通路在高渗透压介导的兔髓核细胞凋亡中的作用[J].复旦学报(医学版), 2012, 39(2): 128-134. DOI: 10.3969/j.issn.1672-8467.2012.02.004.

LI F H, XUE S Q, XIU H J, et al . The role of JNK/SAPK and p38 signal pathways in apoptosis of rabbit nucleus pulposus cells induced by high osmolality[J ] . Fudan Univ J Med Sci, 2012, 39(2): 128-134.DOI:10.3969/j.issn. 1672-8467.2012.02.004.

CHEN Y, LI M, LI B, et al . Effect of reactive oxygen species generation in rabbit corneal epithelial cells on inflammatory and apoptotic signaling pathways in the presence of high osmotic pressure[J ] . PLoS ONE, 2013, 8(8): e72900. DOI: 10.1371/journal.pone.0072900.

LUO L, LI D Q, CORRALES R M, et al . Hyperosmolar saline is a proinflammatory stress on the mouse ocular surface[J ] . Eye Contact Lens, 2005, 31(5): 186-193.

ACERA A, VECINO E, DURAN J A. Tear MMP-9 levels as a marker of ocular surface inflammation in conjunctivochalasis[J]. Invest Ophthalmol Vis Sci, 2013, 54(13): 8285-8291. DOI:10.1167/iovs.13-12235.

MAURIS J, WOODWARD A M, CAO Z, et al . Molecular basis for MMP9 induction and disruption of epithelial cell-cell contacts by galectin-3[J ] . J Cell Sci, 2014, 127(Pt 14): 3141-3148. DOI:10.1242/jcs.148510.

WHITTAMORE J M. Osmoregulation and epithelial water transport: lessons from the intestine of marine teleost fish[J]. J Comp Physiol B, 2012, 182(1): 1-39. DOI:10.1007/s00360-011-0601-3.

ZOLOTAREV V A, ANDREEVA IuV, KHROPYCHEVA R P. Effect of mild irritants on gastric outputs of bicarbonates and pepsinogen depends on the rate of acid secretion[J]. Ross Fiziol Zh Im I M Sechenova, 2012, 98(6): 744-756.

XUE Q, WANG X, WANG P, et al . Role of p38MAPK in apoptosis and autophagy responses to photodynamic therapy with Chlorin e6[J ] . Photodiagnosis Photodyn Ther, 2015, 12(1): 84-91. DOI: 10.1016/j.pdpdt.2014.12.001.

HALL-JACKSON C A, EYERS P A, COHEN P, et al . Paradoxical activation of Raf by a novel Raf inhibitor[J ] . Chem Biol, 1999, 6(8): 559-568. DOI: 10.1016/ S1074-5521(99)80088-X.

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