[Electroacupuncture intervention improves lipid metabolism and promotes browning of white adipose tissue by activating AMPK/Sirt1 pathway and up-regulating Nrg4 content in middle-aged and aged obese rats].

OBJECTIVE: To investigate the mechanism of electroacupuncture （EA） in promoting the browning of white adipose tissue in middle-aged and aged obese rats induced by high fat by regulating AMP-activated protein kinase （AMPK） /silence-information regulatory factor 1 （Sirt1） pathway and neuregulin 4 （Nrg4）. METHODS: Twenty-four male SD rats were randomized into blank control, model and EA groups （n=8 per group）. The obesity model was established by feeding the rats with high-fat diet for 6 weeks. For the EA group, EA （2 Hz/15 Hz, 1.5 mA） was applied to "Guanyuan" （CV4） and bilateral "Shenshu" （BL23）, "Fenglong" （ST40） and "Tianshu" （ST25） for 20 min, once a day, 5 days a week for 6 weeks. Rats of the blank control and model groups were also restrained for 20 min. The body mass and food intake were measured every week, and the Lee's index, epididymal fat, perirenal fat and brown adipose tissue were weighed. The contents of serum total cholesterol （TC）, triglyceride （TG）, high density lipoprotein cholesterol （HDL-C）, low density lipoprotein cholesterol （LDL-C） and norepinephrine （NE） were determined by ELISA. H.E. staining was used to observe the morphological changes of white and brown adipose tissue. The mRNA expression levels of mitochondrial uncoupling protein 1 （UCP1）, peroxisome proliferator activated receptor γ co-activator 1α （PGC-1α）, PR-domain protein 16 （PRDM16）, peroxisome proliferator activated receptor γ （PPARγ） and Nrg4 in the adipose tissue were detected by quantitative real time PCR, and the protein expression levels of Nrg4, AMPKα, Sirt1 and interleukin-6 （IL-6） in the white and brown adipose tissue were detected by Western blot. RESULTS: Compared with the blank control group, the body mass, food intake, the Lee's index, epididymal fat and perirenal fat mass, and serum TG, TC and LDL-C contents and the expression level of IL-6 protein were significantly increased （P<0.01, P<0.05, P<0.001）, and the brown adipose mass, serum HDL-C and NE contents, the expression levels of UCP1, PGC-1α, PRDM16, PPARγ and Nrg4 mRNAs, and the protein expression levels of AMPKα, Sirt1 and Nrg4 proteins in both white and brown adipose tissues were significantly decreased in the model group （P<0.01, P<0.05）. After EA intervention, the increased levels of body mass, food intake, Lee's index, epididymal fat and perirenal fat mass, serum TG, TC and LDL-C contents, and the expression of IL-6 protein, and the decreased levels of brown adipose mass, serum HDL-C and NE contents, expression levels of UCP1, PGC-1α, PRDM16, PPARγ and Nrg4 mRNAs, and those of AMPKα, Sirt1 and Nrg4 proteins in both white and brown adipose tissues were apparently reversed（P<0.05, P<0.01, P<0.001）. H.E. staining showed an increase of the volume and content of intracellular vacuoles of both white and brown adipose tissues, disordered arrangement of cells with vague boundary in the model group, which was relatively milder including a decrease of volume and content of vacuoles of both white and brown adipose, neat arrangement of cells with clear boundary. CONCLUSION: EA intervention can improve lipid metabolism and promote white adipose tissue browning in middle-aged and aged obese rats, which is possibly associated with its functions in activating AMPK/Sirt1 signaling pathway and up-regulating the level of Nrg4.

[The anti-inflammatory effect of electroacupuncture in mice with spinal cord injury and molecular mechanism based on transcriptome sequencing technology].

OBJECTIVE: To observe the effect of electroacupuncture(EA) on neural function and spinal cord pathological morphology in spinal cord injury(SCI) mice and investigate the anti-inflammatory molecular mechanism of EA on SCI mice from the aspects of gene by using bioinformatics. METHODS: Seventy-two female C57BL/6 mice were randomized into sham operation, model and EA groups, with 24 mice in each group. The SCI model was established by clamping the spinal cord with a serrefine after laminectomy at the 1st lumbar vertebra(L1). EA(1.5 Hz/7.5 Hz, 1.0 mA) was applied to bilateral "Jiaji"(EX-B2) and "Zusanli"(ST36) for 10 min, once a day for 14 consecutive days. Basso Mouse Scale(BMS) score was used to assess the hindlimb locomotor function of mice. Histopathological changes of the injured area of the spinal cord were determined by HE staining. The spinal cord RNA was sequenced by using RNA-Seq technology. The bioinformatic analysis was then performed to detect the diffe-rential genes between groups, and the function classification and the involved pathways were enriched. The mRNA and protein expressions of differential genes were detected and verified by using qRT-PCR and Western blot. RESULTS: Compared with the sham operation group, BMS score of the model group was significantly decreased(P<0.05), while that of EA group was increased relevant to the model group (P<0.05). HE staining showed loose and disordered structure and arrangement, cavitation, more inflammatory infiltration, nucleus pycnosis, and neuronal necrosis in the model group, which was alleviated in the EA group. Compared with the sham operation group, 565 differential genes were detected in the model group, including 545 up-regulated and 20 down-regulated, while 41 were detected between the EA and the model group, including 2 up-regulated and 39 down-regulated in the EA group. Fifteen genes that were all up-regulated after modeling and down-regulated after EA intervention were detected by using Venn plot, which are Retn, Adipoq, Myh1, Actn2, Pck1, Klhl41, Fabp4, Hspb7, Myot, Ankrd2, Hrc, Cox6a2, Obscn, Col2a1, Mybpc1, and 3 inflammation-related genes(Fabp4, Adipoq and Pck1) were finally acquired. The 15 differential genes were annotated into main biological processes, cell composition and molecular function in the GO function classification analysis. The 15 differential genes were then enriched into different KEGG pathways, including the peroxisome proliferatorsactivated receptor (PPAR) signaling pathway, Adipocytokine signaling pathway. The mRNA and protein expressions of Fabp4, Adipoq and Pck1 in spinal cord detected by qRT-PCR and Western blot were significantly increased in the model group (P<0.001, P<0.01), while these were significantly decreased in the EA group relevant to the model group(P<0.001, P<0.01, P<0.05). CONCLUSION: EA can promote the repair of nerve function and improve inflammatory infiltration in SCI mice. The mechanism may be closely related to the down-regulation of inflammatory factors Fabp4, Adipoq and Pck1 expression, and the regulation of PPAR and Adipocytokine signaling pathways.

[Effect of mild moxibustion on vascular aging in senescence rats based on local MCP-1/MMP-2/TGF-ß1 pro-inflammatory signal loop].

OBJECTIVE: To observe the effect of mild moxibustion on monocyte chemotaxis protein 1 (MCP-1)/matrix metalloproteinase 2 (MMP-2)/transforming growth factor ß1 (TGF-ß1) pro-inflammatory signal loop in senile rats, so as to explore its mechanisms underlying improving vascular aging (VA). METHODS: Twenty-four male VA SD rats were randomized into senium (VA) control, medication and moxibustion groups, and other 8 young SD rats (aged 2 months) were used as the young control group. The VA model was established by intraperitoneal injection of D-galactose (300 mg·kg-1·d-1) once daily for 4 weeks, and verified by serum total testosterone (TT) and free testosterone (FT) levels. For rats of the moxibustion group, mild moxibustion was applied to bilateral "Shenshu"(BL23) and "Guanyuan"(CV4) for 20 min, once a day, 5 days a week for 8 weeks. Rats of the medication group were treated by intraperitoneal injection of testosterone propionate (7 mg·kg-1·[3 d]-1) once daily for 8 weeks except weekends, and rats of the senium control and young control groups treated by intraperitoneal injection of the same dose of normal saline, once daily for 8 weeks except weekends. The duration of exhausted swimming (DES) before and after the treatment was recorded. H.E. staining and Masson staining were used to observe histopathological changes and collagen fiber content of the thoracic aortic tissue, respectively. The contents of serum TT, FT and angiotensin 2 (Ang Ⅱ) were determined by ELISA. The immunoactivity of aortic MCP-1 was detected by immunohistochemistry, and the expression levels of aortic MCP-1, MMP-2 and TGF-ß1 were detected by Western blot. RESULTS: Compared with the young control group, the levels of DES, serum TT and FT contents were significantly decreased (P<0.01), while those of serum AngⅡ and collagen fiber contents, aortic MCP-1 immunoactivity and MMP-2, TGF-ß1 and MCP-1 protein expression considerably increased in the senium control group (P<0.01). After the interventions, the decreased levels of DES, serum TT and FT contents and the increased levels of serum AngⅡ, collagen fiber contents, aortic MCP-1 immunoactivity and MMP-2, TGF-ß1 and MCP-1 protein expression were reversed in both medication and moxibustion groups (P<0.01, P<0.05). The therapeutic effect of mild moxibustion was significantly superior to that of medication in down-regulating the aortic collagen fiber, serum AngⅡ contents and MCP-1 immunoactivity and protein expression (P<0.05). H.E. staining showed thickened endometrium and disordered arrangement of vascular smooth muscles of the aorta in the senium group, and thinner endometrium and regular and ordered arrangement of aortic vascular smooth muscles in both moxibustion and medication groups. CONCLUSION: Mild moxibustion may improve vascular aging in senescence rats, which is possibly by suppressing vascular MCP-1/MMP-2/TGF-ß1 pro-inflammatory signal loop.