| Part 1 Expression and effect of monocyte/macrophage chemoattractant protein in adipose tissue of obese infantile ratsObjective:to investigate the expression of monocyte macrophage and chemokine in adipose tissue of obese infantile rats and the correlation with obesity and insulin resistanceMethods: 20 weanling female SD rats were randomly divided into 2groups: control group(n=10)with based feed,obesity group with high fat diet,for 6 weeks.Measured the weight of rats;determined serum fasting insulin by chemiluminescence immunoassay,fasting blood glucose by dry chemical method;calculate insulin resistance index by HOMA model;determined serum total cholesterol by automatic biochemical analyzer;determined the serum monocyte chemoattractant protein-1(MCP-1)and macrophage inflammatory protein-1?(MIP-1?)byenzyme linked immunosorbent assay(ELISA);immunohistochemical(IHC)staining was used to detect the expression of MCP-1,MIP-1? and F4/80 protein in adipose tissue;to detect MCP-1,MIP-1?and NF-k B mRNA expression level in adipose tissue by real time fluorescencequantitative polymerase chain reaction(qPCR).Result: The body weight,fasting blood glucose,fasting insulin,HOMA-IR and serum total cholesterol in the obese group were significantly higher than those in the control group(P<0.05).The serum levels of MCP-1 and MIP-1? in obese group were higher than those in control group(P<0.05).Abdominal adipose tissue HE staining showed that the structure of the control group was more uniform than that of control group.The adipocytes arranged in the obese group were disordered,the size was different,the adipocytes were over inflated,and many of the newborned adipocytes were coexisted.The expression of MCP-1,MIP-1? and F4/80 in adipose tissue of obese group was higher than that of control group(P<0.05),and the mRNA expression of MCP-1,MIP-1? and NF-k B were higher than those of control group(P<0.05).The expression of MCP-1,MIP-1? and F4/80 protein were positively correlated with body weight,serum insulin,HOMA-IR and serum cholesterol;the expression level of MCP-1and MIP-1? was negatively correlated with InSI;only the expression of MCP-1is positively associated with blood glucose.Conclusion:In the adipose tissue of obese rats induced by high fat diet,the expression ofmacrophage infiltration and monocyte chemotactic factor MCP-1and MIP-1?enhance,and relatewith the metabolism of glucose and lipid and insulin resistance,suggesting that mononucytes/macrophages in adipose tissue and the secretion of chemokines may play a role in the pathophysiological process of obesity and its complications.Part 2 Effects of vitamin D on insulin resistance and expression of monocyte/macrophage chemotactic protein in obese infantile ratObjective: to study the effects of vitamin D on glucose and lipid metabolism and insulin resistance in obese young rats,and to explore the relationship between the expression of monocyte/macrophage chemoattractant protein and macrophage infiltration in adipose tissue.Methods:30 weanling female SD rats were randomly divided into 3 groups(n=10): control group were fed basic diet,obesity group and VD intervention group with high fat diet;VD group:intragastric administration with VD5ug/(kg.d),the control group and the obese model group were administrated vegetable oil instead,totally for 6 weeks.Measured body weight and body length,calculated Lee's Index,weighed intra-abdominal fat pad;determined serum fasting insulin by chemiluminescence immunoassay,fasting blood glucose by dry chemical method;calculate insulin resistance index by HOMA model;determined serum lipid metabolism by automatic biochemical analyzer;determined the serum monocyte chemoattractant protein-1(MCP-1)and macrophage inflammatory protein-1?(MIP-1?)byenzyme linked immunosorbent assay(ELISA);observed the structure of adipose tissue by hematoxylin eosin(HE)staining;immunohistochemical(IHC)staining was used to detect the expression of MCP-1,MIP-1? and F4/80 protein in adipose tissue;detectedthe expression of MCP-1,MIP-1?and NF-k B m RNA in adipose tissue by real time fluorescence quantitative polymerase chain reaction(q PCR);observed and calculated thenumber and size of mitochondria in skeletal muscle by transmission electron microscope.Result:VD intervention group weight,fasting blood glucose,fasting insulin,HOMA-IR and abdominal fat coefficient,blood lipids were lower than the obese model group(P<0.05).The levels of serum MCP-1 and MIP-1?in the VD intervention group were lower than those in the obese group(P<0.05).HE showed that the cells of VD group were more orderly and uniform in size.IHC showed that the expression of MCP-1,MIP-1?and F4/80 in adipose tissue of VD group were significantly lower than those in obese group(P<0.05).QPCR showed that the m RNA expression of MCP-1,MIP-1?and NF-k B in adipose tissue of VD group was higher than that of obese group(P<0.05).Transmission electron microscopy showed that the skeletal muscle mitochondria of rats with obesity were swelling,the number was reduced,and a large number of glycogen granules were deposited.The skeletal muscle mitochondria of VD group were more complete,the number of them was increased,and the abnormal deposition of glycogen granules decreased.VD group compared with the obese group,the area of mitochondria in skeletal muscle was increased(P<0.05).Conclusion:Vitamin D early intervention can ameliorate glucose and lipid metabolism and insulin resistance in obese infantile rats,the specific mechanism is not clear,vitamin D may inhibit adipose tissue macrophage infiltration,reduced expression of MCP-1 and MIP-1? in adipose tissue and stability of skeletal muscle mitochondrial function,reduced inflammatory state in adipose tissue of obese subject. |
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