| Objectives Obesity is closely related to insulin resistance and lipid metabolism disorders.Insulin resistance is a key pathologic basement for the development of obesity as well as type 2 diabetes,and it is also considered as the beginning and center course of the nonalcoholic fatty liver disease.Early intervention with insulin resistance and obesity is of great importance in preventing the development of type 2 diabetes and nonalcoholic fatty liver.As Sirt1 / PPAR-α plays an important role in the regulation of lipid metabolism in liver,we aimed to observe the effect of electroacupuncture(EA)and calorie restriction(CR)on the model of obese rats with IR via its regulation on protein and m RNA expression of SIRT1/PPAR-α,insulin sensitivity,lipid metabolism and hepatic cell morphology.It was expected to provide experimental evidence for the application of EA combined with CR in the treatment of obesity with IR and related metabolic diseases.Methods The 90 SPF male Wistar rats of 8 weeks were randomly divided into normal group(n = 18)and modeling group(n = 72)by using random number table,the normal group was fed with standard diet,the modeling group was fed with high-fat diet.8 weeks later,3 rats in normal group and 12 in model group were randomly picked to participate in high insulin-regular glucose clamps,and it was used to evaluate the model rat of IR.In the meantime,the weight and length of all rats were measured,Lee ’s index was calculated,both of them were used to assess the model of obesity.All the indicators testing above were finished in one week.After the model rat of obesity with IR was gotten successfully,the rats in modeling group would be divided into the model group,EA+CR group,EA group,CR group randomly,each group consisting of 15 rats.At the tenth week of the experiment,each group was given different interventions.Normal group was given nothing but standard diet,the model group was given nothing but high-fat diet;EA group and EA+CR group were both fed with high-fat diet and EA treatment at acupoint ST36,ST40,CV4 and CV12.0.3×25mm needle and HANS LH202 H electroacupuncture instrument were used,setting parameters were 1m A,2Hz,continuous wave,10 minutes once time,3 times a week,the total course is 8 weeks;CR group and EA+CR group were both given high-fat diet and 30% calorie restriction.The weight and the length of all rats were measured and Lee’s index of all rats was calculated before and after the intervention for 2,4,6,8 weeks.After 6 weeks of intervention,intraperitoneal glucose tolerance test(IPGTT)and insulin tolerance test(IPITT)of all rats were tested.After 8 weeks of intervention,3 rats were randomly selected to test GIR with high insulin-regular glucose clamps.After 8 weeks of intervention,serum triglycerides(TG),total cholesterol(TC)and free fatty acids(FFA)levels were detected,the liver tissue of rats were taken into-80 ℃ refrigerators.The liver tissue of rats was observed with hematoxylin-eosin staining under the light microscope.The protein expression of SIRT1 and PPAR-αin hepatic tissue were detected by western blotting,and the m RNA expression of SIRT1 and PPAR-αin hepatic tissues were detected by Real-time PCR.Finally,all the experimental datas were analyzed statistically.Results1.The effect of high-fat diet in eight weeks on the body mass,Lee,s index and GIR of obese rats with IR.After 8 weeks of high-fat diet feeding,the rat weight and Lee’s index of modeling group were significantly higher than that of normal group(P<0.01),GIR of modeling group was significantly lower than that of normal group(P<0.01).2.The effects of 8 weeks of EA and CR on weight,Lee’s index and lipid metabolism of obese rats with IR.Compared with the weight of normal group,that of EA group and model group were increased(P<0.05),that of the other two groups were increased with no statistical difference(P>0.05).Compared with the weight of model group,that of CR group and EA group were decreased(P<0.05),and EA group was decreased with no statistical difference(P>0.05).Compared with the weight of EA group,the weight of EA+CR group and CR group were decreased with no statistical difference(P>0.05).There was no statistically significant difference between the weight of CR group and EA+CR group(P>0.05).Compared with Lee’s index of the normal group,that of model group was increased(P<0.05).Compared with Lee’s index of model group,that of EA+CR group and CR group and EA group were significantly decreased(P<0.05).There was no statistical difference between Lee’s index of other groups(P>0.05).The level of TC,TG,FFA of the model group were significantly increased compared with that of normal group(P<0.01).Compared with the level of TC,TG,FFA of model group,that of EA+CR group and EA group were both significantly reduced(P<0.01),that of CR group was decreased(P<0.05).Compared with the level of TC,TG,and FFA of EA group,that of EA+CR group were reduced(P<0.05).Compared with the level of TC,TG,FFA of CR group,that of EA+CR group were reduced(P<0.05).There was no statistical difference between the level of TC,TG,FFA of other groups(P<0.05).3.The effects of EA and CR on intraperitoneal glucose tolerance(IPGT),intraperitoneal insulin tolerance(IPIT)and systemic insulin sensitivity of obese rats with IR.The intraperitoneal glucose tolerance of rat was observed by IPGT test.IPGT of model group was significantly impaired compared with that of normal group(P<0.01);IPGT of EA+CR group,EA group and CR group were both significantly enhanced compared with that of model group(P<0.01);Compared with EA+CR group,IPGT of CR group was impaired(P<0.05).There was no statistical difference between IPGT of other groups(P>0.05).The intraperitoneal insulin tolerance of rat was observed by IPIT test.IPIT of model group was significantly impaired compared with that of normal group(P<0.01);IPIT of EA+CR group,EA group,and CR group were significantly enhanced compared with IPIT of model group(P<0.01).IPIT in CR group was impaired compared with that of EA+CR group(P<0.05).There was no statistical difference between IPIT of other groups(P>0.05).Systemic insulin sensitivity in rats was observed by GIR.GIR of model group,EA group,and CR group were significantly reduced compared with that of normal group(P<0.01).GIR of EA group,CR group and EA+CR group significantly enhanced compared with GIR of model group(P<0.01),there was no statistically significant difference between GIR of other groups(P>0.05).4.The effects of EA and CR on the morphology of hepatocytes in obese rats with IR.In model group,the hepatic cell cords and lobular structure were not clear,the hepatocytes were severely ballooning,and a large number of large vacuolar fat vacuoles and obvious lipid accumulation were observed,and the hepatic cell piece-meal necrosis was observed.There was no difference between the liver cell morphology of EA group and CR group.The pathological changes were slightly in EA group and CR group compared with that of model group,characterized by hepatocyte ballooning,small bubble type lipid drops,hepatocyte foci or punctate necrosis.Compared with EA group and CR group,pathological changes were more slightly in EA+CR group,hepatic cell cords and lobular structure were clear,fat cavitation and the accumulation of lipid droplets were reduced,punctate necrosis was fewer.In terms of the comparison of liver steatosis and pathological integrals,the pathological integrals of the model group were significantly larger than that of normal group(P<0.01).The pathological integrals of EA+CR group,EA group,and CR group were all larger than that of normal group(P<0.05).EA+CR group was significantly smaller than that of model group(P<0.01).The pathological integrals of EA group and CR group were smaller than that of model group(P<0.05).The pathological integrals of EA group and CR group were larger than that of EA+CR group(P<0.05).5.The effects of EA and CR on protein and m RNA expression levels of hepatic SIRT1,PPAR-αin obese rats with IR.The protein expression level of hepatic SIRT1 of the other four groups was decreased compared with that of normal group(P<0.05).Compared with the hepatic SIRT1 protein expression level of the model group,that of EA+CR group and CR group were increased(P<0.05),there was no significant difference between the other groups(P>0.05).The hepatic PPAR-αprotein expression of the other four groups was significantly decreased compared with that of normal group(P<0.01).Compared with the hepatic PPAR-αprotein expression of model group,that of EA+CR group and CR group were significantly increased(P<0.01),and that of EA group increased(P<0.05).Compared with EA+CR group,the hepatic PPAR-αprotein expression of EA group was significantly decreased(P<0.01).Compared with EA group,the hepatic PPAR-αprotein expression of CR group was significantly increased(P<0.01),there was no statistical difference between the other groups.Compared with the hepatic SIRT1 m RNA expression of the normal group,that of model group was significantly decreased(P<0.01)and that of EA group was decreased(P<0.05).Compared with the hepatic SIRT1 m RNA expression of model group,that of EA+CR group and CR group were both increased(P<0.05),and there was no statistical difference between the other groups.Compared with the hepatic PPAR-αm RNA expression of the normal group,that of model group was significantly decreased(P<0.01)and that of EA group was decreased(P<0.05).Compared with the hepatic PPAR-αm RNA expression of model group,that of CR group and EA+CR group were both increased(P<0.05).There was no statistical difference between the other groups(P>0.05).Conclusions1.EA+CR,EA,CR can reduce the weight gain of the obese rats with IR,and they also can reduce serum TC,TG,FFA levels,improve IPGT and IPIT,improve insulin sensitivity,reduce the pathological changes of hepatocytes in morphology.2.EA+CR has a obvious inhibitory effect on the weight growth of obese rats with IR,the reduced effect on serum TC,TG,FFA of EA+CR was much greater than that of EA or CR,the enhancement effect on IPIT and IPGT of EA+CR was greater than that of CR.EA+CR could reduce pathological morphological changes of hepatocytes,and the degree of reduction on steatosis was greater than EA or CR.EA+CR could raise SIRT1 and PPAR-αprotein and m RNA expression level of hepatic tissue.The effect of EA+CR on the upregulation of PPAR-αprotein level was greater than that of EA.3.CR can inhibit the body weight gain of obese rat with IR.CR can upregulate the protein and m RNA levels of PPAR-αin hepatic tissue,the upregulation of PPAR-α protein level of CR is greater than that of EA.CR could upregulate the protein and m RNA expression level of SIRT1 in hepatic tissue.4.In conclusion,in terms of regulating lipid metabolism and improving the pathologic changes of hepatocytes and improving insulin sensitivity,there may be a synergistic effect between EA and CR.There were an significant differences between the regulative effect of EA and CR on the protein and m RNA levels of SIRT1,PPAR-αin hepatic tissue.The regulative effect of EA+CR on glycolipid metabolism may be related to the increase of SIRT1,PPAR-αprotein and m RNA expression levels in hepatic tissue. |
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