大建中汤调控小胶质细胞自噬治疗腹泻型肠易激综合征内脏痛和抑郁症共病的作用机制

武静; 田维毅; 蔡琨; 李尧锋; 杨莎莎

doi:10.3969/j.issn.1006-2157.2023.02.012

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大建中汤调控小胶质细胞自噬治疗腹泻型肠易激综合征内脏痛和抑郁症共病的作用机制

实验研究 | 更新时间：2023-03-07

- 大建中汤调控小胶质细胞自噬治疗腹泻型肠易激综合征内脏痛和抑郁症共病的作用机制
- Mechanism of Dajianzhong Decoction in treating visceral pain and depression associated with diarrhea irritable bowel syndrome through regulating microglial autophagy
- 北京中医药大学学报 2023年46卷第2期页码：215-223
- 作者机构：
  
  1.贵州中医药大学　贵阳　550025
  2.贵州中医药大学第一附属医院
- 作者简介：
  
  武静，男，硕士，副教授
  #杨莎莎，女，博士，副主任医师，硕士生导师，主要研究方向：中医药抗免疫机制研究，E-mail：yangyuansha88@163.com
- 基金信息：
  
  国家自然科学基金项目(82260855;81960821);贵州省基础研究计划（自然科学类）项目(黔科合基础-ZK〔2022〕一般495)
- DOI：10.3969/j.issn.1006-2157.2023.02.012
  中图分类号： R285.5
- 收稿：2022-05-30，
  
  网络首发：2022-12-27，
  
  纸质出版：2023-02-28
- 稿件说明：
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武静, 田维毅, 蔡琨, 等. 大建中汤调控小胶质细胞自噬治疗腹泻型肠易激综合征内脏痛和抑郁症共病的作用机制[J]. 北京中医药大学学报, 2023,46(2):215-223.

WU Jing, TIAN Weiyi, CAI Kun, et al. Mechanism of Dajianzhong Decoction in treating visceral pain and depression associated with diarrhea irritable bowel syndrome through regulating microglial autophagy[J]. Journal of Beijing University of Traditional Chinese Medicine, 2023, 46(2): 215-223. DOI: 10.3969/j.issn.1006-2157.2023.02.012.
武静, 田维毅, 蔡琨, 等. 大建中汤调控小胶质细胞自噬治疗腹泻型肠易激综合征内脏痛和抑郁症共病的作用机制[J]. 北京中医药大学学报, 2023,46(2):215-223. DOI： 10.3969/j.issn.1006-2157.2023.02.012.

WU Jing, TIAN Weiyi, CAI Kun, et al. Mechanism of Dajianzhong Decoction in treating visceral pain and depression associated with diarrhea irritable bowel syndrome through regulating microglial autophagy[J]. Journal of Beijing University of Traditional Chinese Medicine, 2023, 46(2): 215-223. DOI: 10.3969/j.issn.1006-2157.2023.02.012. DOI：

摘要

目的

探讨大建中汤治疗腹泻型肠易激综合征内脏痛和抑郁症共病的作用机制。

方法

采用慢性不可预知性刺激加葡聚糖硫酸钠方法建立腹泻型肠易激综合征内脏痛和抑郁症共病大鼠模型。大鼠随意分为正常对照组、模型组、大建中汤组[10.8 g/(kg·d)]、氟西汀组[0.01 g/(kg·d)]。治疗14 d后，采用强迫游泳评估各组大鼠的抑郁状况，腹壁撤退反应(AWR)评估各组大鼠肠道痛觉敏感性，酶联免疫吸附法检测各组大鼠前扣带回皮质(ACC)中白细胞介素(IL)－ 1β、肿瘤坏死因子－α(TNF-α)、IL-10、IL-4表达；蛋白质印迹法检测ACC区髓鞘脂蛋白(PLP)、髓鞘少突胶质细胞糖蛋白(MOG)、髓鞘碱性蛋白(MBP)及微管相关蛋白1轻链3－Ⅱ(LC3-Ⅱ)、p62、Beclin-1，以及小胶质细胞相关蛋白精氨酸酶1(Arg1)和诱导型一氧化氮合酶(iNOS)蛋白表达；免疫荧光双标法观察ACC区自噬相关蛋白LC3-Ⅱ和MG离子钙结合衔接分子1(Iba1)共表达情况。

结果

与正常组大鼠相比，共病大鼠强迫游泳时间减少(

0.01)，不动时间增加(

0.01)，在20、40、60 mmHg(1 mmHg≈0.133 kPa)3个压力下AWR分数上升(

0.01)，前扣带回皮质PLP、MOG、MBP蛋白表达下降(

0.01)，iNOS蛋白表达升高(

0.01)，促炎因子TNF-α、IL-1β升高(

0.01)，抗炎因子IL-10、IL-4水平有增加趋势，LC3－Ⅱ、Beclin-1蛋白表达下降(

0.01)，p62蛋白表达升高(

0.01)，LC3-Ⅱ、Iba1阳性细胞降低(

0.01)；与模型组大鼠比较，大建中汤组不动时间及各压力下AWR评分降低(

0.01)，ACC中PLP、MOG、MBP蛋白表达升高(

0.01)，Arg1蛋白表达升高(

0.01)、iNOS蛋白表达下降(

0.01)，TNF-α、IL-1β下降(

0.01)，IL-10、IL-4升高(

0.01)，LC3-Ⅱ、Beclin-1蛋白表达升高(

0.01)，p62蛋白表达降低(

0.01)，LC3-Ⅱ、Iba1阳性细胞增加(

0.01)。上述各指标，氟西汀组与大建中汤组差异均无统计学意义。

结论

通过促进自噬以调控小胶质细胞清除髓鞘碎片，是大建中汤治疗腹泻型肠易激综合征内脏痛和抑郁症共病的可能作用机制。

Abstract

Objective

To explore the mechanism of

Dajianzhong

Decoction in treating visceral pain and depression associated with diarrhea irritable bowel syndrome.

Methods

Chronic unpredictable stimulation and dextran sodium sulfate were used to establish a rat model with visceral pain and depression associated with diarrhea irritable bowel syndrome. The rats were randomly divided into normal group

model group

Dajianzhong

Decoction group [10.8 g/(kg·d)]

and fluoxetine group [0.01 g/(kg·d)]

. After 14 days of treatment

the depression level of the rats in each group was evaluated by forced swimming

the intestinal pain sensitivity of the rats in each group was evaluated by abdominal wall withdrawal reaction (AWR)

the interleukin (IL)- 1β

tumor necrosis factor-α (TNF-α)

IL-10

and IL-4 expression in the anterior cingulate cortex (ACC) of the rats in each group were detected by enzyme linked immunosorbent assay. Western blotting was used to detect the expression of myelin sheath lipoprotein (PLP)

myelin oligodendrocyte glycoprotein (MOG)

myelin basic protein (MBP)

microtubule associated protein 1 light chain 3-Ⅱ(LC3-Ⅱ)

p62

Beclin-1

microglial cell (MG)-associated protein Arg1

and inducible nitric oxide synthase (iNOS). The coexpression of autophagy related protein LC3-Ⅱ and MG calcium binding adaptor molecule 1 (Iba1) in the ACC region was observed by the immunofluorescence double labeling method .

Results

Compared with the normal rats

the model rats showed shorter forced swimming times (

0.01)

and increased immobility times (

0.01). The AWR fraction increased under pressures of 20

and 60 mmHg. The protein levels of PLP

MOG

and MBP in the anterior cingulate cortex decreased (

0.01)

and iNOS protein increased (

0.01). The proinflammatory factors TNF-α and IL-1β increased significantly (

0.01). We observed an increasing trend in the levels of the anti-inflammatory factors IL-10 and IL-4; LC3-Ⅱ and Beclin-1 protein decreased significantly (

0.01); p62 protein increased (

0.01); and the number of cells that were positive for LC3-Ⅱ and Iba1 decreased (

0.01). Compared with the model group

the immobility time and AWR scores under various pressures in the

Dajianzhong

Decoction group decreased (

0.01). In the

Dajianzhong

Decoction group

the protein levels of PLP

MOG

MBP in the anterior cingulate cortex increased (

0.01); Arg1 protein increased (

0.01); iNOS protein decreased (

0.01); TNF-α and IL-1β decreased (

0.01); IL-10 and IL-4 increased (

0.01); LC3-Ⅱ and Beclin-1 protein increased (

0.01); p62 protein decreased (

0.01); and LC3-Ⅱ

Iba1 positive cells increased significantly (

0.01). There was no significant difference in the above indices between the fluoxetine group and the

Dajianzhong

Decoction group.

Conclusion

The mechanism of

Dajianzhong

Decoction in treating visceral pain and depression associated with diarrhea irritable bowel syndrome may involve promoting autophagy to regulate microglia to clear myelin sheath fragments.

关键词

Keywords

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