Objective: To observe the effect of electroacupuncture （EA） on white adipose tissue （WAT） browning by regulating central glucagon-like peptide-1 （GLP-1）, so as to explore the possible central mechanisms of EA in improving obesity.
Methods: Thirty male Wistar rats were randomly divided into normal group, model group, EA group, HM3D group, and EA+HM4D group, with 6 rats in each group. The obesity rat model was obtained by feeding with high-fat diet for 8 weeks. Adeno-associated virus combined with DREADDs was injected into bilateral nucleus of solitary tract （NTS）, with rAAV-GLP-1+rAAV-4D applied to the EA+HM4D group, rAAV-GLP-1+rAAV-3D applied to the HM3D group, and rAAV-GLP-1+rAAV-GFP applied to other 3 groups. After modeling, rats in the EA and EA+HM4D groups received EA treatment at bilateral "Zusanli"（ST36）, "Fenglong"（ST40）, "Guanyuan"（CV4） and "Zhongwan"（CV12）, with successive waves （2 Hz, 1 mA） for 10 minutes, 3 times a week, for a total of 8 weeks. Body mass of rats in each group were measured before and 2, 4, 6, and 8 weeks after intervention. Abdominal and perirenal WAT mass was weighed, serum triglyceride （TG） and total cholesterol （TC） contents were detected by using automatic analyzer, and nonestesterified fatty acid （NEFA） content was detected by using colorimetric assay kit. The morphology of abdominal WAT lipid droplets was observed by HE staining. The mRNA expressions of GLP-1 in NTS, AMPK in ventromedial nucleus of hypothalamus（VMH）, UCP1 and PGC-1α in subcutaneous fat were detected by real-time PCR. The protein expression levels of GLP-1, AMPK, phosphorylated-AMPK, UCP1 and PGC-1α were detected by Western blot. The activation level of GLP-1 neurons in NTS was observed by immunofluorescence.
Results: Compared with the normal group, abdominal WAT lipid droplets were enlarged, body weight, serum TG, TC, NEFA contents, abdominal and perirenal WAT mass, mRNA and protein expression levels of AMPK were significantly increased（P<0.01, P<0.05）, while GLP-1 neurons activation level, mRNA and protein expression levels of GLP-1, UCP1 and PGC-1α, and AMPK protein phosphorylation were decreased （P<0.01） in the model group. After EA intervention, body weight at 6 and 8 weeks after intervention and other indexes mentioned above were all significantly reversed （P<0.01, P<0.05） in the EA group in comparison with those of the model group. Compared with the EA group, the HM3D group had reduced abdominal WAT lipid droplets size, decreased serum TG, TC, and NEFA contents, and protein expression level of AMPK（P<0.01, P<0.05）, with increased mRNA and protein expression levels of GLP-1, UCP1 and PGC-1α, and phosphorylation level of AMPK protein（P<0.01, P<0.05）, while the EA+HM4D group had enlarged abdominal WAT lipid droplets, increased body weight 6 and 8 weeks after intervention, abdominal and renal WAT mass, and NEFA content （P<0.01, P<0.05）, with decreased serum TG content, activation level of GLP-1 neurons in the NTS, mRNA and protein expression levels of GLP-1, UCP1 and PGC-1α （P<0.01, P<0.05）, as well as down-regulated phosphorylation of AMPK protein and mRNA （P<0.01, P<0.05）.
Conclusion: EA can effectively promote the browning of WAT, which may be related to the activation of GLP-1 neurons in the NTS, as well as the promotion of the phosphorylation of AMPK in the VMH and up-regulation of UCP1.
目的：观察电针通过调控中枢胰高血糖素样肽-1（GLP-1）促进白色脂肪（WAT）褐色化的效应，探讨电针改善肥胖的可能中枢机制。方法：30只Wistar雄性大鼠随机分为正常组、模型组、电针组、HM3D组、电针+HM4D组，每组6只。采用高脂饲料喂养8周制备肥胖大鼠模型。各组大鼠均予双侧孤束核（NTS）注射相应的腺相关病毒结合特定药物激活特定受体。选取双侧“足三里”“丰隆”和“关元”“中脘”给予电针治疗（2 Hz，1 mA，连续波），每次10 min，每周3 次，共 8 周。测定各组大鼠干预前及干预后第2、4、6、8周的体质量，WAT质量，血清甘油三酯（TG）、总胆固醇（TC）、游离脂肪酸（NEFA）含量；用HE染色观察腹部WAT脂滴形态；用实时荧光定量PCR法检测大鼠NTS中GLP-1，下丘脑腹内侧核（VMH）中AMP活化蛋白激酶（AMPK），皮下脂肪中解偶联蛋白1 （UCP1）、过氧化物酶体增殖物激活受体γ共激活因子-1α（PGC-1α）的mRNA水平；应用Western blot法检测NTS中GLP-1，VMH中AMPK、p-AMPK，皮下脂肪中UCP1、PGC-1α的蛋白表达；应用免疫荧光染色法观察各组大鼠NTS中GLP-1神经元的激活水平。结果：与正常组比较，模型组大鼠体质量及血清TG、TC、NEFA含量显著升高（P<0.01），NTS中GLP-1神经元的激活水平、mRNA与蛋白表达水平，皮下脂肪中UCP1、PGC-1α的mRNA与蛋白表达水平显著降低（P<0.01），腹部及肾周WAT质量增加（P<0.05），腹部WAT脂滴增大，VMH中AMPK的mRNA与蛋白表达水平显著升高（P<0.01），磷酸化水平显著降低（P<0.01）。与模型组比较，电针组大鼠在治疗第6周和第8周时的体质量显著降低（P<0.01），其余各指标均被逆转（P<0.01，P<0.05）。与电针组比较，HM3D组大鼠腹部WAT脂滴减小，血清TG、TC、NEFA含量，VMH中AMPK的蛋白表达水平显著降低（P<0.01，P<0.05），NTS中GLP-1 mRNA与蛋白表达水平，VMH中AMPK的磷酸化水平，皮下脂肪中UCP1、PGC-1α的mRNA与蛋白表达水平显著升高（P<0.01，P<0.05）；电针+HM4D组大鼠干预后第6、8周的体质量、腹部与肾周WAT总质量、血清NEFA含量升高（P<0.01，P<0.05），血清TG含量，NTS中GLP-1神经元的激活水平、mRNA与蛋白表达水平，皮下脂肪中UCP1、PGC-1α的mRNA与蛋白表达水平降低（P<0.01，P<0.05），腹部WAT脂滴变大，VMH中AMPK的蛋白表达水平升高（P<0.01），mRNA与蛋白磷酸化水平显著降低（P<0.05，P<0.01）。结论：电针能够有效促进肥胖大鼠WAT褐色化，其内在机制可能与激活NTS中的GLP-1神经元，进而促进VMH中AMPK磷酸化，上调皮下脂肪中UCP1的表达有关。.
Keywords: AMP-activated protein kinase; Browning of white adipose tissue; Electroacupuncture; Glucagon-like peptide-1; Lipid metabolism; Obesity.