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基于“治肠脑”原则观察电针对血管性痴呆大鼠肠道菌群及血清白细胞介素-1β和白细胞介素-18的影响

[Effect of electroacupuncture on gut microbiota and serum IL-1β and IL-18 in rats with vascular dementia based on principle of "curing brain disorders by treating intestines"].

作者信息

Chen Dan-Feng, Zhang Hong, Xie Ju-Ying, Deng Chang, Qiu Rong-Rong, Xu Ya-Yu, Xiong Dan, Tan Jie

机构信息

School of Acupuncture-Message and Rehabilitation, Hunan University of Chinese Medicine, Changsha 410208, China.

出版信息

Zhen Ci Yan Jiu. 2022 Mar 25;47(3):216-23. doi: 10.13702/j.1000-0607.20210766.

Abstract

OBJECTIVE

To observe the effect of electroacupuncture (EA) of "Baihui"(GV20), "Dazhui"(GV14), "Shenshu" (BL23)and "Zusanli"(ST36) on the intestinal flora and serum interleukin (IL)-1β and IL-18 contents in vascular dementia (VD) rats.

METHODS

SD rats were randomized into sham operation, VD model, GV20+GV14+BL23 (EA-basic acupoints), and EA-basic acupoints+ST36 and EA-basic acupoints+probiotics groups (=10 in each group). EA (10 Hz/50 Hz) was conducted for 30 min, once daily for 4 consecutive weeks. Rats of the EA-basic acupoints+probiotics received gavage of probiotics (2 mL/d containing 2.0×10 CFU of live bifidobacterium), once a day for 4 weeks, and those of the EA-basic acupoints and EA-basic acupoints+ST36 groups received gavage of the same dose of normal saline. The Morris water maze test was used to evalua-te the rats' lear-ning and memory ability before and after the treatment. The serum IL-1β and IL-18 levels were determined by ELISA, and the histopathological changes of the intestinal mucosa were observed by H.E. staining. The ultrastructural changes of hippocampal neurons were observed by using transmission electron microscopy and 16S rDNA sequencing technique was used to analyze the composition of intestinal microbiome.

RESULTS

Compared with the sham operation group, the escape latency, serum levels of IL-1β and IL-18, as well as the relative abundance of harmful bacteria (including Catabacter, obinsoniella and Desulfovibrio) in the intestine were significantly increased (<0.01). In comparison with the model group, the escape latency, serum levels of IL-1β and IL-18 in the three treatment groups, and the relative abundance of harmful bacteria (such as the Catabacter, Robinsoniella and Desulfovibrio) in the EA-basic acupoints+ST36 group were down-regulated obviously(<0.05,<0.01), and the relative abundance of Clostridiales-unclassified in both EA-basic acupoints+probiotics and EA-basic acupoints+ST36 groups was significantly up-regulated (<0.05). The effects of EA-basic acupoints+ST36 and EA-ba-sic acupoints+probiotics were significantly superior to that of EA-basic acupoints in down-regulating IL-18 content (<0.05). H.E. staining showed atrophy of the whole mucosal layer, loss of goblet cells, destruction of glands, infiltration of a large number of inflammatory cells, and transmission microscope displayed fuzziness of the nucleus membrane boundary, cystic dilation of the rough endoplasmic reticulum with unclear structure swelling of the mitochondria, and disordered arrangement or dissolution of the inner cristae in the model group, which was relatively milder in the EA-basic acupoints+ST36 and EA-basic acupoints+probiotics groups.

CONCLUSION

EA of GV20+GV14+BL23+ ST36 can improve the cognitive dysfunction of VD model rats, which may be related to its function in regulating the imbalance of intestinal microbiota, thereby inhibiting the peripheral inflammatory factor.

摘要

目的

观察电针“百会”(GV20)、“大椎”(GV14)、“肾俞”(BL23)和“足三里”(ST36)对血管性痴呆(VD)大鼠肠道菌群及血清白细胞介素(IL)-1β和IL-18含量的影响。

方法

将SD大鼠随机分为假手术组、VD模型组、GV20+GV14+BL23(电针基础穴位)组、电针基础穴位+ST36组和电针基础穴位+益生菌组(每组10只)。采用10Hz/50Hz电针刺激30min,每日1次,连续4周。电针基础穴位+益生菌组大鼠给予益生菌灌胃(2mL/d,含2.0×10CFU的活性双歧杆菌),每日1次,共4周,电针基础穴位组和电针基础穴位+ST36组大鼠给予相同剂量的生理盐水灌胃。采用Morris水迷宫试验评估大鼠治疗前后的学习记忆能力。采用ELISA法检测血清IL-1β和IL-18水平,采用苏木精-伊红(H.E.)染色观察肠黏膜组织病理学变化。采用透射电子显微镜观察海马神经元超微结构变化,采用16S rDNA测序技术分析肠道微生物群落组成。

结果

与假手术组比较,模型组大鼠逃避潜伏期、血清IL-1β和IL-18水平以及肠道有害菌(包括解脲棒杆菌、罗氏菌和脱硫弧菌)相对丰度显著升高(P<0.01)。与模型组比较,三个治疗组大鼠逃避潜伏期、血清IL-1β和IL-18水平以及电针基础穴位+ST36组肠道有害菌(如解脲棒杆菌、罗氏菌和脱硫弧菌)相对丰度明显下调(P<0.05,P<0.01),电针基础穴位+益生菌组和解脲棒杆菌电针基础穴位+ST36组未分类梭菌目相对丰度显著上调(P<0.05)。电针基础穴位+ST36组和电针基础穴位+益生菌组下调IL-18含量的作用明显优于电针基础穴位组(P<0.05)。H.E.染色显示模型组全层黏膜萎缩、杯状细胞丢失、腺体破坏、大量炎性细胞浸润,透射电镜显示模型组细胞核膜边界模糊、粗面内质网囊性扩张、结构不清、线粒体肿胀、线粒体内嵴排列紊乱或溶解,电针基础穴位+ST36组和电针基础穴位+益生菌组病变相对较轻。

结论

电针GV20+GV14+BL23+ST36可改善VD模型大鼠的认知功能障碍,其机制可能与调节肠道微生物群失衡、抑制外周炎性因子有关。

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