Effects of Yiqi Kaimi Formula on intestinal motility， protein expression of MLCK and p‑MLC in mice with slow transit constipation

MENG Lingyun; XIAO Changfang; ZHANG Jiayuan; WU Jingwen; YAO Yibo

doi:10.16305/j.1007-1334.2025.z20240606004

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Effects of Yiqi Kaimi Formula on intestinal motility， protein expression of MLCK and p‑MLC in mice with slow transit constipation

更新时间：2025-02-28

- Effects of Yiqi Kaimi Formula on intestinal motility， protein expression of MLCK and p‑MLC in mice with slow transit constipation
- Shanghai Journal of Traditional Chinese Medicine Vol. 59, Issue 2, Pages: 64-70(2025)
- 作者机构：
  
  上海中医药大学附属龙华医院肛肠科（上海 200032）
- 作者简介：
- 基金信息：
- DOI：10.16305/j.1007-1334.2025.z20240606004
  CLC：
- Received：06 June 2024，
  
  Published：10 February 2025
- 稿件说明：
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孟令昀,肖长芳,张嘉员,等.益气开秘方对慢传输型便秘小鼠肠道动力及MLCK、p‑MLC蛋白表达的影响[J].上海中医药杂志,2025,59(2):64-70.

MENG Lingyun,XIAO Changfang,ZHANG Jiayuan,et al.Effects of Yiqi Kaimi Formula on intestinal motility， protein expression of MLCK and p‑MLC in mice with slow transit constipation[J].Shanghai Journal of Traditional Chinese Medicine,2025,59(2):64-70.
孟令昀,肖长芳,张嘉员,等.益气开秘方对慢传输型便秘小鼠肠道动力及MLCK、p‑MLC蛋白表达的影响[J].上海中医药杂志,2025,59(2):64-70. DOI： 10.16305/j.1007-1334.2025.z20240606004.

MENG Lingyun,XIAO Changfang,ZHANG Jiayuan,et al.Effects of Yiqi Kaimi Formula on intestinal motility， protein expression of MLCK and p‑MLC in mice with slow transit constipation[J].Shanghai Journal of Traditional Chinese Medicine,2025,59(2):64-70. DOI： 10.16305/j.1007-1334.2025.z20240606004.

摘要

目的

建立稳定的慢传输型便秘小鼠模型并完善模型评价，探究益气开秘方治疗慢传输型便秘的作用机制。

方法

将健康的28只雄性C57BL/6J小鼠，随机分为正常组、模型组、莫沙必利组和益气开秘方组，每组7只。模型组、莫沙必利组、益气开秘方组小鼠采用灌胃洛哌丁胺［10 mg/（kg·d）］联合限水法进行造模，造模药物每天灌胃2次，持续给药7 d。期间，实验第3天第2次造模给药后，莫沙必利组［5 mg/（kg·d）］及益气开秘方组［6 g/（kg·d）］小鼠灌胃给予相应药物进行干预，每天1次，连续干预5 d；正常组和模型组小鼠灌胃给予0.9%氯化钠溶液，连续干预5 d。检测各组小鼠一般情况、粪便含水率、4 h内粪便排泄粒数和粪便排泄次数，并通过墨迹实验检测肠道推进率；通过吲哚箐绿（ICG）荧光染色实验检测首粒含荧光粪便排泄时间及粪便总荧光强度；通过苏木精-伊红（HE）染色实验检测结肠平滑肌厚度；通过Western blot法和免疫荧光法检测肌球蛋白轻链激酶（MLCK）、磷酸化肌球蛋白轻链（p-MLC）表达水平。

结果

与正常组相比，模型组小鼠体质量明显降低，粪便含水率降低，4 h内粪便粒数减少，4 h内粪便排泄次数减少，肠道推进率降低，首粒含荧光粪便排泄时间延长，粪便荧光总强度减弱，结肠平滑肌厚度变薄，MLCK和p

⁃

MLC蛋白表达水平下调，差异均有统计学意义（

0.05）。与模型组相比，益气开秘方组小鼠体质量明显增加，粪便含水率增加，4 h内粪便粒数增加，4 h内粪便排泄次数增加，肠道推进率增加，首粒含荧光粪便排泄时间缩短，粪便荧光总强度增强，结肠平滑肌厚度增厚，MLCK和p-MLC蛋白表达水平上调，差异均有统计学意义（

0.05）。

结论

益气开秘方能够改善慢传输型便秘小鼠肠道动力，其作用机制可能是通过上调MLCK蛋白表达及MLC磷酸化程度，从而改善慢传输型便秘小鼠便秘症状。

Abstract

Objective

To establish a stable slow transit constipation mouse model and improve the model evaluation，and explore action mechanism of Yiqi Kaimi Formula in treating slow transit constipation.

Methods

Twenty-eight healthy male C57BL/6J mice were randomly divided into normal group， model group， mosapride group， and Yiqi Kaimi Formula group， with 7 mice in each group. The mice in the model group， mosapride group and Yiqi Kaimi Formula group were treated with loperamide ［10 mg/（kg·d）］ by gavage twice a day for 7 d combined with water restriction method to make a model. During this period， on the third day of the experiment， after the second dose of modeling drug was administered， the mice in the mosapride group ［5 mg/（kg·d）］ and the Yiqi Kaimi Formula group ［6 g/（kg·d）］ were treated with corresponding drugs once a day for 5 consecutive days； the mice in the normal group and model group were orally administered with sodium chloride solution （9 g/L） for 5 consecutive days. Detect the general condition， fecal moisture content， number of fecal particles excreted and frequency of fecal excretion within 4 h of mice in each group. Ink blotting experiment were used to detect intestinal propulsion rate. Indocyanine green （ICG） fluorescence staining experiment were used to detect the first fluorescent fecal excretion time and total fecal fluorescence intensity. Hematoxylin-eosin （HE） staining experiment were used to detect the thickness of colon smooth muscle. Western blot and immunofluorescence were used to detect the expression levels of myosin light chain kinase （MLCK） and myosin light chain protein phosphorylation （p‑MLC）.

Results

Compared with the normal group， the body mass of mice in the model group was significantly reduced， the fecal moisture content was decreased， the number of fecal particles excreted within 4 h was decreased， the frequency of fecal excretion within 4 h was decreased， the intestinal propulsion rate was decreased， the first fluorescent fecal excretion time was prolonged， total fecal fluorescence intensity was weakened， the thickness of colon smooth muscle became thinner， and the protein expression levels of MLCK and p-MLC were down-regulated. The differences were statistically significant （

0.05）. Compared with the model group， the body mass of mice in the model group was significantly increased， the fecal moisture content was increased， the number of fecal particles excrete

d within 4 h was increased， the frequency of fecal excretion within 4 h was increased， the intestinal propulsion rate was increased， the first fluorescent fecal excretion time was shortened， total fecal fluorescence intensity was enhanced， the thickness of colon smooth muscle became thicker， and the protein expression levels of MLCK and p-MLC were up-regulated. The differences were statistically significant （

0.05）.

Conclusions

Yiqi Kaimi Formula can improve intestinal motility in mice with slow transit constipation. Its mechanism may be to up-regulate the expression of MLCK protein and the phosphorylation of MLC， so as to improve the symptoms of slow transit constipation in mice.

关键词

Keywords

references

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