艾灸对腹泻型肠易激综合征大鼠结肠TLR4/MyD88/NF-κB信号通路的影响

[Effect of moxibustion on TLR4/MyD88/NF-κB signaling pathway in colon of diarrhea-predo-minant irritable bowel syndrome rats].

作者信息

Chu Hao-Ran, Wang Yu, Tong Li, Wu Sheng-Bing, Wu Li-Bin, Li Nan, Cheng Hong-Liang

机构信息

The Second Affiliated Hospital of Anhui University of Chinese Medicine, Hefei 230061, China; Institute of Clinical Acupuncture and Moxibustion, Anhui Academy of Chinese Medicine, Hefei 230038.

Graduate School of Anhui University of Chinese Medicine, Hefei 230038.

出版信息

Zhen Ci Yan Jiu. 2020 Aug 25;45(8):633-9. doi: 10.13702/j.1000-0607.190950.

DOI:10.13702/j.1000-0607.190950

PMID:32869573

Abstract

OBJECTIVE

To observe the effect of moxibustion on serum inflammatory factors, Toll-like receptor 4 (TLR4), myeloid differentiation factor 88 (MyD88) and nuclear factor Kappa B (NF-κB) in colon tissue of rats with diarrhea-predominant irritable bowel syndrome (IBS-D)，so as to explore the mechanism of moxibustion in the treatment of IBS-D.

METHODS

Healthy male SD rats were randomly divided into blank group, model group and moxibustion group, with 8 rats in each group. The IBS-D model was established by chronic restraint combined with gavage of Senna-leaf solution. The rats of the moxibustion group was treated with moxibustion at "Tianshu"(ST25)and "Shangjuxu"(ST37)for 30 min， once a day for 7 d. After the intervention, the rate of loose stools and the minimum threshold volume of abdominal withdrawal reflex (AWR) induced by colorectal distension were observed. The contents of serum inflammatory factors IL-1β, TNF-α and IL-6 were detected by ELISA. The pathological changes of rat colon were observed by H.E. staining. The average optical density of TLR4 and NF-κB (p65) in colon tissue was detected by immunohistochemistry. The relative expressions of TLR4, MyD88 and NF-κB (p65) mRNAs in colon tissues were detected by real-time fluorescence quantitative PCR, and the expression levels of TLR4, MyD88 and NF-κB (p65) proteins in colon tissues were detected by Western blot.

RESULTS

There was a mild degree of inflammation in colon in rats of the model group. Compared with the blank group, the minimum volume threshold of AWR decreased significantly (< 0.01), and the rate of loose stools, the contents of IL-1β, IL-6 and TNF-α in serum as well as the expressions of TLR4, MyD88, NF-κB (p65) mRNAs and protein in colon tissues were significantly increased in the model group (<0.01). Compared with the model group, the inflammation was alleviated in colonic mucosa, the minimum volume threshold of AWR increased significantly (<0.01), and rate of loose stools decreased significantly (<0.01) in the moxibustion group, the contents of IL-1β, TNF-α and IL-6 in serum and the expression levels of TLR4, MyD88, NF-κB (p65) mRNAs and proteins in colon tissue decreased significantly in the moxibustion group (<0.01).

CONCLUSION

Moxibustion ST25 and ST37 can improve diarrhea symptom and visceral hypersensitivity in IBS-D rats, which may be related with its effects in inhibiting TLR4/MyD88/NF-κB signaling pathway and reducing the expression of inflammatory factors.

摘要

目的

观察艾灸对腹泻型肠易激综合征（IBS-D）大鼠血清炎症因子、Toll样受体4（TLR4）、髓样分化因子88（MyD88）及结肠组织中核因子κB（NF-κB）的影响，以探讨艾灸治疗IBS-D的作用机制。

方法

将健康雄性SD大鼠随机分为空白组、模型组和艾灸组，每组8只。采用慢性束缚联合番泻叶溶液灌胃法建立IBS-D模型。艾灸组大鼠取“天枢”（ST25）、“上巨虚”（ST37）穴艾灸30 min，每日1次，连续7 d。干预后，观察大鼠稀便率及结直肠扩张诱发的腹部退缩反射（AWR）最小阈值容积。采用酶联免疫吸附测定（ELISA）法检测血清炎症因子白细胞介素-1β（IL-1β）、肿瘤坏死因子-α（TNF-α）和IL-6含量。采用苏木精-伊红（H.E.）染色观察大鼠结肠组织病理变化。采用免疫组织化学法检测结肠组织中TLR4和NF-κB（p65）的平均光密度。采用实时荧光定量聚合酶链反应（PCR）检测结肠组织中TLR4、MyD88和NF-κB（p65）mRNA的相对表达量，采用蛋白质免疫印迹法检测结肠组织中TLR4、MyD88和NF-κB（p65）蛋白的表达水平。

结果

模型组大鼠结肠有轻度炎症。与空白组比较，模型组大鼠AWR最小容积阈值显著降低（<0.01），稀便率、血清中IL-1β、IL-６、TNF-α含量及结肠组织中TLR4、MyD88、NF-κB（p65）mRNA和蛋白表达均显著升高（<0.01）。与模型组比较，艾灸组大鼠结肠黏膜炎症减轻，AWR最小容积阈值显著升高（<0.01），稀便率显著降低（<0.01），血清中IL-1β, TNF-α和IL-6含量及结肠组织中TLR4、MyD88、NF-κB（p65）mRNA和蛋白表达均显著降低（<0.01）。