复合因素急性应激诱导肠道屏障功能障碍及其作用机制研究

林启运; 候鹏飞; 郎和东; 张钧; 陈卡; 易龙; 糜漫天

doi:10.16016/j.2097-0927.202309077

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复合因素急性应激诱导肠道屏障功能障碍及其作用机制研究

基础医学 | 更新时间：2026-01-09

- 复合因素急性应激诱导肠道屏障功能障碍及其作用机制研究
- Effect and mechanism of acute stress induced by composite factors on intestinal barrier dysfunction
- 陆军军医大学学报 2024年46卷第1期页码：73-83
- 作者机构：
  
  400038 重庆，陆军军医大学(第三军医大学)军事预防医学系军队营养与食品安全研究中心，重庆市医学营养研究中心，重庆市营养与健康重点实验室
- 作者简介：
  
  [易龙，E-mail：amu_yilong@tmmu.edu.cn YI Long, E-mail: amu_yilong@tmmu.edu.cn
  [糜漫天，E-mail：mantian_mi@tmmu.edu.cn MI Mantian, E-mail: mantian_mi@tmmu.edu.cn
- 基金信息：
- DOI：10.16016/j.2097-0927.202309077
  中图分类号： R339.5;R363;R574
- 收稿：2023-09-19，
  
  修回：2023-10-21，
  
  纸质出版：2024-01-15
- 稿件说明：
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林启运, 候鹏飞, 郎和东, 等. 复合因素急性应激诱导肠道屏障功能障碍及其作用机制研究[J]. 陆军军医大学学报, 2024,46(1):73-83.

Qiyun LIN, Pengfei HOU, Hedong LANG, et al. Effect and mechanism of acute stress induced by composite factors on intestinal barrier dysfunction[J]. Journal of Army Medical University, 2024, 46(1): 73-83.
林启运, 候鹏飞, 郎和东, 等. 复合因素急性应激诱导肠道屏障功能障碍及其作用机制研究[J]. 陆军军医大学学报, 2024,46(1):73-83. DOI： 10.16016/j.2097-0927.202309077.

Qiyun LIN, Pengfei HOU, Hedong LANG, et al. Effect and mechanism of acute stress induced by composite factors on intestinal barrier dysfunction[J]. Journal of Army Medical University, 2024, 46(1): 73-83. DOI： 10.16016/j.2097-0927.202309077.

摘要

目的

研究复合因素急性应激对肠道屏障功能的影响，并探索相关作用机制。

方法

20只7~8周龄的雄性C57BL/6小鼠(体质量20~22 g)，用随机数表法分为对照(CON)组和模型(MOD)组，

=10。MOD组小鼠于24 h内先后给予4 h休息、15 h的睡眠剥夺、3 h的噪音(120 dB)/闪光(2 000±500 lx)和2 h的强迫游泳运动(水温26±1 ℃、水深45 cm)，建立复合因素急性应激模型；CON组小鼠置于恒温恒湿、正常昼夜节律的无噪音环境中不做任何处理。MOD组小鼠强迫游泳运动结束后，将小鼠麻醉采血，颈椎脱臼处死并取材。通过免疫酶联吸附实验试剂盒检测血清肠损伤标志物，HE染色法检测结肠组织形态变化，PAS染色检测肠道杯状细胞数目，免疫荧光、免疫组化和RT-qPCR法检测结肠组织屏障功能和炎症因子相关蛋白及基因表达，16S rRNA技术分析小鼠肠道菌群变化。

结果

与CON组比较，MOD组小鼠血清肠道损伤标志物脂多糖(Lipopolysaccharide，LPS)、肠脂肪酸结合蛋白(Intestinal fatty acid binding protein，I-FABP)、连蛋白(Zonulin)明显升高(

0.05)；HE染色显示MOD组小鼠肠道组织损伤明显；免疫荧光染色和RT-qPCR结果显示机械屏障相关分子紧密连接蛋白-1(Zona occludens 1，ZO-1)、闭合蛋白(Occludin)表达显著降低(

0.05)，免疫组化和PAS染色提示化学屏障相关杯状细胞数目及黏蛋白2(mucin2，Muc-2)、抗菌肽再生胰岛衍生蛋白3γ(regenerating islet derived-3γ，Reg3γ)、再生胰岛衍生蛋白3β(regenerating islet derived-3β，Reg3β)显著降低(

0.05)；肠道菌群β多样性显著改变，有害菌拟杆菌门、脱硫弧菌门显著增加，有益菌瘤胃球菌科、丹毒丝菌(

norank_f_Erysipelotrichaceae

)属、孪生球菌(

Gemella

)属、丹毒丝菌(

Erysipelotrichaceae

)属显著减少(

0.05)；炎症相关肿瘤坏死因子α(tumor necrosis factor，TNF-α)、白细胞介素6(interleukin-6，IL-6)以及活性氧(reactive oxygen species，ROS)水平均显著增加(

0.05)。

结论

复合因素急性应激显著诱导小鼠肠道损伤和肠屏障功能障碍。

Abstract

Objective

To identify the effect of acute stress induced by composite factors on intestinal barrier dysfunction and explore the relevant mechanism.

Methods

Twenty male C57BL/6 mice (7-8 weeks old

body mass 20-22 g) were randomly divided into control group (CON group

=10) and model group (MOD group

=10). To e

stablish an acute stress model of complex factors

the mice of the MOD group were given rest of 4 h

sleep deprivation of 15 h

and then followed by 3 h of noise (120 dB)/flash (2 000±500 lx) and 2 h of forced swimming (water temperature 26±1 ℃

water depth 45 cm) within 24 h. While the mice in the CON group were placed in a noiseless environment with constant temperature and humidity and normal circadian rhythm but been given no any treatment. At the end of forced swimming

the mice from the MOD group were anesthetized for blood samples and then euthanized. Serum markers of intestinal injury were detected by immunoenzyme-linked adsorption assay kit

morphological changes in colonic tissues were observed by HE staining

protein and mRNA levels of colonic tissue barrier function related molecules and inflammatory factors were detected by immunofluorescence

immunohistochemistry and RT-qPCR

and intestinal flora changes were analyzed by 16S rRNA technique.

Results

Compared with the CON group

the serum markers of intestinal damage

lipopolysaccharide (LPS)

intestinal fatty acid binding protein (I-FABP) and Zonulin were significantly increased in the MOD group (

0.05). HE staining showed that intestinal tissue damage was obvious in the MOD group. The results of immunofluorescence staining and RT-qPCR showed that the levels of mechanical barrier related molecules Zona occludens 1 (ZO-1) and Occludin were obviously decreased (

0.05). Immunohistochemistry and PAS staining indicated the number of chemical barriers associated goblet cells

and expression of mucin2 (Muc-2)

antimicrobial peptide regenerating islet derived-3γ (Reg3γ) and regenerating islet derived-3β (Reg3β) were notably reduced (

0.05). Intestinal flora β diversity significantly changed

with harmful bacteria

Bacteroidetes

and

Desulfovibrio

significantly increased

while beneficial bacteria

Rumenococcus

mily

norank_f_Erysipelotrichaceae

Gemella genus

and

Erysipelotrichaceae genus

declined (

0.05). The levels of TNF-α

IL-6 and ROS were significantly enhanced (

0.05).

Conclusion

Acute stress of complex factors significantly induces intestinal injury and barrier dysfunction.

关键词

Keywords

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