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电针治疗对肥胖大鼠脂肪组织中SIRT1/NF-κB信号通路的影响

[Effect of electroacupuncture on SIRT1/NF-κB signaling pathway in adipose tissue of obese rats].

作者信息

Huang Qi, Chen Rui, Peng Miao, Li Lun, Li Tong, Liang Feng-Xia, Xu Fen

机构信息

Department of Rehabilitation, Wuhan Center Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology, Wuhan 430014, Hubei Province, China; College of Acupuncture and Orthopedics, Hubei University of CM, Wuhan 430061.

Department of Integrative Chinese and Western Medicine, Union Hospital Affiliated to Tongji Medical College of Huazhong University of Science and Technology.

出版信息

Zhongguo Zhen Jiu. 2020 Feb 12;40(2):185-91. doi: 10.13703/j.0255-2930.20190324-00054.

Abstract

OBJECTIVE

To explore the effect of electroacupuncture (EA) on insulin sensitivity, adipose tissue inflammatory reaction and silent information regulation factor 1(SIRT1)/nuclear factor kappa B (NF-κB) signaling pathway in obese rats.

METHODS

A total of 100 SPF-grade Wistar male rats were collected. Thirteen rats of them were selected randomly as the normal group and fed with common forage, and the rest rats were fed with high-fat forage. Eight weeks later, 39 rats that met the obesity criteria were randomized into a model group, an EA group and a sham-EA group, 13 rats in each one. In each group, 3 rats were collected randomly and the hyperinsulinemic-euglycemic clamp was exerted to record glucose infusion rate (GIR) so as to determine insulin sensitivity. Afterwards, in the EA group, EA was applied to "Zusanli" (ST 36), "Fenglong" (ST 40), "Zhongwan" (CV 12) and "Guanyuan" (CV 4), stimulated with continuous wave, 2 Hz in frequency, 1 mA in current intensity, for 15 min. The treatment was given once every 2 days, 3 times a week, for 8 weeks totally. In the sham-EA group, the needles were inserted shallowly at the sites, 5 mm lateral to each of the acupoints stimulated in the EA group, and the electrodes were attached to the needle handles, but without electric stimulation exerted. The rest management was the same as the EA group. Before and after intervention, the body mass and the insulin sensitivity were measured. After intervention, the white adipose tissue was collected from the kidney in the rats. Western blot was adopted to detect the relative protein expressions of SIRT1, interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α) and acetylated NF-κB (Ac-NFκB). The real-time fluorescence quantitative PCR was used to detect the mRNA expressions of SIRT1, IL-6 and TNF-α. The immunofluorescence double labeling method was applied to detect the co-expression of SIRT1 and Ac-NFκB in adipose tissue.

RESULTS

After fed with high-fat forage for 8 weeks, the body mass was significantly increased and GIR decreased in the rats of the model group as compared with the normal group (<0.01), suggesting that the model of obese rat with insulin resistance was successfully established. After 8-week intervention, compared with the model group, the body mass was reduced and GIR increased in the rats of the EA group (<0.01). The differences were not significant statistically in comparison between the sham-EA group and the model group (>0.05). Compared with the normal group, in the model group, the protein and mRNA expressions of SIRT1 in adipose tissue were decreased, and the protein expression of Ac-NFκB increased, the protein and mRNA expressions of IL-6 and TNF-α increased (<0.05, <0.01). Compared with the model group, in the EA group, the protein and mRNA expressions of SIRT1 in adipose tissue were increased significantly, the protein expression of Ac-NFκB decreased, and the protein and mRNA expressions of IL-6 and TNF-α significantly decreased (<0.05, <0.01). There was no significant difference in each index between the sham-EA group and the model group (>0.05). The results of immunofluorescence double labeling showed that SIRT1 and Ac-NFκB were co-expressed in adipose tissue.

CONCLUSION

Electroacupuncture significantly reduces the body mass, inflammatory reaction conditions of adipose tissue and improves insulin sensitivity in obese rats. Regarding the potential mechanism, after the activation of SIRT1/NF-κB signaling pathway by electroacupuncture, and down-regulates the transcription of downstream inflammatory factors.

摘要

目的

探讨电针(EA)对肥胖大鼠胰岛素敏感性、脂肪组织炎症反应及沉默信息调节因子1(SIRT1)/核因子κB(NF-κB)信号通路的影响。

方法

选取100只SPF级雄性Wistar大鼠,随机选取13只作为正常组,给予普通饲料喂养,其余大鼠给予高脂饲料喂养。8周后,将符合肥胖标准的39只大鼠随机分为模型组、电针组和假电针组,每组13只。每组随机选取3只大鼠,进行高胰岛素-正葡萄糖钳夹试验,记录葡萄糖输注速率(GIR)以测定胰岛素敏感性。之后,电针组在“足三里”(ST 36)、“丰隆”(ST 40)、“中脘”(CV 12)和“关元”(CV 4)穴位施加电针,连续波刺激,频率2 Hz,电流强度1 mA,刺激15 min。每2天治疗1次,每周3次,共8周。假电针组在与电针组刺激穴位外侧5 mm处浅刺进针,将电极连接于针柄,但不施加电刺激。其余处理同电针组。干预前后测量大鼠体重及胰岛素敏感性。干预后,取大鼠肾周白色脂肪组织。采用蛋白质印迹法检测SIRT1、白细胞介素-6(IL-6)、肿瘤坏死因子-α(TNF-α)及乙酰化NF-κB(Ac-NFκB)的相对蛋白表达。采用实时荧光定量PCR检测SIRT1、IL-6及TNF-α的mRNA表达。采用免疫荧光双标法检测脂肪组织中SIRT1与Ac-NFκB的共表达情况。

结果

高脂饲料喂养8周后,模型组大鼠体重显著增加,GIR降低,与正常组比较差异有统计学意义(<0.01),提示成功建立胰岛素抵抗肥胖大鼠模型。干预8周后,与模型组比较,电针组大鼠体重降低,GIR升高(<0.01)。假电针组与模型组比较差异无统计学意义(>0.05)。与正常组比较,模型组脂肪组织中SIRT1蛋白及mRNA表达降低,Ac-NFκB蛋白表达升高,IL-6及TNF-α蛋白和mRNA表达升高(<0.05,<0.01)。与模型组比较,电针组脂肪组织中SIRT1蛋白及mRNA表达显著升高,Ac-NFκB蛋白表达降低,IL-6及TNF-α蛋白和mRNA表达显著降低(<0.05,<0.01)。假电针组与模型组各指标比较差异无统计学意义(>0.05)。免疫荧光双标结果显示,SIRT1与Ac-NFκB在脂肪组织中共表达。

结论

电针可显著降低肥胖大鼠体重、脂肪组织炎症反应程度,提高胰岛素敏感性。其潜在机制可能是电针激活SIRT1/NF-κB信号通路后,下调下游炎症因子的转录。

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