| Objective: In this study,the model of high-fat diet induced insulin resistance(IR)in obese Wistar rats were used.The electro-acupuncture(EA)and caloric restriction(CR)were adopted as an intervention method to observe the hypothalamic leptin receptor(OB-Rb)and downstream orexin peptide agouti-related protein(AGRP).Taking the leptin resistance(LR)as a breakthrough point to investigate the different effect of the EA and CR on obese rats with IR,so as to provide experimental evidence for clinical application of EA combined with CR in preventing and curing IR,obesity and related diseases.Methods: 100 male Wistar rats were selected and fed with normal food,after 1 week adaptive feeding,the rats were randomly divided into normal diet group(n=20)and high-fat diet group(n=80)by using random number table.8 weeks later,5 rats in normal diet group and 20 rats in high-fat diet group were randomly picked up to detect the GIR by eurythmics hyperinsulinemia clamp technique.In the meantime,the average body weight and anal length were measured to calculate the Lee 's index,it was used to assess the obesity model of insulin resistance.After the IR obesity model rats were successfully bulid,the rats in the high-fat diet group were randomly divided into the model group(M group),the caloric restriction group(CR group),the electro-acupuncture group(E group),and the combined group(E+CR group),10 rats in each group.And 10 rats in the normal diet group were chosen as the normal group(N group).After allocation,the N group was given nothing but ordinary food;the other four groups were continue fed with high-fat diet,but no treatment was done in the M group;the quantity of feeding and remainder of the M group were recorded every day to calculate the average food intake,on the basis of this,the rats in the CR group were given 30% of the dietary restriction;the E group were given electro-acupuncture treatment,and the E+CR group adopted double intervention by using caloric restriction and electro-acupuncture.Acupoints are ST36,ST40,CV4 and CV12,the depth of acupuncture is 3-5mm and the frequency of electro-acupuncture instrument is 2Hz/1m A,three times a week(10min/once),for 8 weeks.The weight of rats was weighed and recorded every two weeks,before intervention(0 week)and 2,4,6,and 8 weeks after the intervention.After 6 weeks of intervention,the detection of IPGTT and IPITT were performed.After 8 weeks of intervention,the Elisa and biochemical methods were used for testing serum TG,TC,FFA,serum insulin,serum leptin,and serum resistin;the HE staining was adopted to observe the morphological changes of white adipose tissue(WAT);the expression of the protein OB-Rb and AGRP in the hypothalamus was detected by Western blotting,and the expression of OB-Rb and AGRP m RNA were detected by real-time fluorescence quantitative technique.Result1.The influence of EA and CR on body weight of obese rats with IR After 8-week intervention,the body weight of rats in the three intervention groups were significantly lower than those in the M group(P<0.01).Among them,the weight loss was most obvious in the E+CR group;and the EA and CR methods could also slow the growth of weight2.The effects of EA and CR on TC,TG,FFA and insulin sensitivity n obese rats with IR In the lipid metabolism: compared with the M group,the contents of TC,TG and FFA in the three intervention groups were significantly decreased(P<0.05),and the regulation of lipid metabolism in the E group and the E+CR group was the most obvious(P<0.01).In the level of serum insulin,the E group,E+CR group were significantly lower than that in the M group(P < 0.01),and CR group was likewise lower than that of the M group(P<0.05).In the insulin sensitivity: about the IPGTT experiment,the blood glucose levels of the three intervention groups were lower than those of the M group,and the IPGTT-AUC of three groups were significantly reduced than the M group(P<0.01),Among them,the degree of reduction in E+CR group was more obvious than CR group(P<0.05).About the IPITT experiment,the glucose levels in the three intervention groups were lower than those in the M group,and the IPITT-AUC of three groups were also significantly decreased than M group(P<0.01).and compared with the CR group,the E+CR group was decreased(P<0.05).3.The effect of EA and CR on the levels of serum leptin,resistin and the morphology of white adipose tissue in obese rats with IR In the comparison of adipokines: compared with the M group,the level of Leptin in the E group and the CR group were both decreased(P<0.05),and the CR group was significantly lower(P<0.01)than M group;In contrast of M group,the content of Resistin in the CR group decreased(P<0.05),while the content of EA group and E+CR group decreased significantly(P<0.01).In the form of white fat cells,the number of adipocytes in the CR group increased more than that in the M group(P<0.05),and the number of cells in the EA group,E+CR group were both increased significantly than M group(P<0.01).Compared with the M group,most of the adipocyte diameters of the three intervention groups were reduced(P<0.05),and the E+CR group was the most obvious(P < 0.01).There was no statistical difference between the three intervention groups.(P>0.05).4.The effect of EA and CR on expression of OB-Rb,AGRP protein and mRNA in hypothalamus of IR obesity rats In terms of protein expression: compared with the M group,after the EA or CR intervention,the expression of OB-Rb protein in the hypothalamus increased significantly,and the expression of AGRP protein decreased significantly(P<0.01),and the effect of the combined intervention was superior to single EA or CR intervention.(P<0.05).In terms of m RNA expression,compared with the M group,AGRP m RNA expression were significantly reduced in both the EA group and the E+CR group,OB-Rb m RNA expression in this 2 groups were significantly increased(P<0.01);compared with the M group,the expression of OB-Rb m RNA in the CR group was also increased(P<0.05),the expression of AGRP m RNA was significantly decreased(P<0.01).There was no statistical difference between the three intervention groups.Conclusion:1.EA combined with CR can reduce the body weight of IR obese rats,slow down the growth trend of body weight,lower the levels of blood lipids TC,TG,and FFA,enhance the insulin sensitivity of IR obese rats and improve the IR status.2.EA combined with CR can reduce the levels of serum leptin and resistin in IR obese rats,alleviate the status of LR and IR.At the same time,EA combined with CR can increase the number of white adipocytes and reduce the size of the cells,suggesting that EA combined with CR can effectively promote lipid metabolism and reduce lipid accumulation in adipocytes.3.EA combined with CR can up-regulate the expression of OB-Rb protein and m RNA in hypothalamus of IR obese rats,thus down-regulating the expression of AGRP m RNA and inhibiting the synthesis of AGRP protein.This indicates that EA combined with CR may be through up-regulating the expression of OB-Rb,so that leptin in the serum could pass through the blood-brain barrier and fully bind to OB-Rb,lowering the level of serum leptin,improving the status of LR to exert leptin function,inhibit AGRP expression,regulate diet,control body weight,improve peripheral insulin sensitivity,and improve IR obesity. |
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