BCL6 Regulation Of Skeletal Muscle Glucose Metabolism And Its Mechanism

PART ONE THE RELATIONSHIP BETWEEN BCL6 AND SKELETAL MUSCLE GLUCOSE METABOLISMObjective Explore the relationship between BCL6 and skeletal muscle glucose metabolism.Methods The relationship between BCL6 and insulin resistance in skeletal muscle was explored through GEO data mining.Detection the expression changes of BCL6 in skeletal muscle of wild type mice with insulin resistance.The expression of BCL6 before and after the single extreme exercise was detected in mice fed with normal and high-fat diet.The effects of palmitic acid and insulin on the expression of BCL6 were detected in myotubes.The effects of inhibition and overexpression of BCL6 on the insulin signaling pathway would also be detected in myotubes.Results We found that BCL6 expression was significantly lower in skeletal muscle of ob/ob and db/db mice than in wild type mice by GEO data mining.This result was also proved in ob/ob,db/db mice and wild-type mice fed with high-fat diet using RT-PCR and western blot.There was also a significantly decreased in myotubes treated with palmitic acid.The effects of insulin on BCL6 expression was time and dose dependence and negatively correlated with time and concentration.The BCL6 expression was significantly reduced after exercise in mice fed with normal diet,while there was no difference before and after exercise in mice fed with high fat diet.Inhibition of BCL6 can significantly promote the phosphorylation of IR and AKT in vitro.Conclusion BCL6 expression was significantly reduced both in insulinresistant mice and cells.BCL6 may be associated with a significant decreased exercise performance in mice fed with high fat diet.Insulin treatment with different concentrations and times in myotobes can inhibit the BCL6 expression in skeletal muscle.Inhibition the BCL6 in cells can significantly affect insulin signaling pathways.These results suggested that BCL6 may be involved in the glucose metabolism of skeletal muscles.PART TWO EFFECTS OF BCL6 ON GLUCOSE METABOLISM IN SKELETAL MUSCLEObjective Observe the effects of knockout and overexpression BCL6 on glucose metabolism of skeletal muscle in vivo.Methods The changes of energy metabolism in whole body and skeletal muscle was dectected in conditional knockout of BCL6 mice fed with normal diet and high-fat diet.In addition,the effects of BCL6 on glucose metabolism in skeletal muscle were also observed by injection of BCL6 overexpression adenovirus.General energy metabolism indicators,including body weight,relative mass of adipose tissues and skeletal muscle,food intake,respiration quotient were being detected.Haematological indicators,including fasting blood glucose and insulin,postprandial blood glucose and blood lipid were detected.The glucose and insulin tolerance were tested on indicted time.Glucose uptake and glycogen content in skeletal muscle and exercise performance were also been detected.Results There was no significant difference in body weight of BCL6MKO mice fed with normal diet,while it was decreased significantly in BCL6 MKO mice fed with high fat diet compared to the control group(P < 0.05).The relative mass of subcutaneous and epididymis fat was less in highfat-fed BCL6 MKO mice than that of the control group(P < 0.01),while there was no significant difference among the gastrocnemius,soleus and quadriceps.Although there was no significant difference in food intake of BCL6 MKO mice fed with normal or high-fat diet,the high-fat-diet BCL6 MKO mice were reduced in respiration quotient caused by significantly decreased glucose oxidation and increased fatty acid oxidation(P < 0.01 or P < 0.05).BCL6 MKO mice were lower in the level of fasting blood glucose and insulin,postprandial blood glucose and blood lipid,improved insulin and glucose tolerance and HOMA-IR,increased glucose uptake,glycogen content and exercise performace than control mice fed with high fat diet(P < 0.05 or P < 0.01);while there was no significant difference in BCL6 MKO mice than control mice fed with normal diet.BCL6 overexpression in skeletal muscle reduces glucose uptake,glycogen content and exercise performace in mice fed with high fat diet.Conclusion Knockout BCL6 in skeletal muscle significantly improved insulin sensitivity,promoted skeletal muscle glucose uptake and glycogen storage and enhanced exercise performance in mice with high-fat diet.PART THREE THE MECHANISM OF BCL6 REGULATION THE GLUCOSE METABOLISM IN SKELETAL MUSCLEObjective Investigation the mechanism of BCL6 regulation the glucose metabolism in skeletal muscle.Methods Detection changes in insulin signaling pathways and key proteins of glucose metabolism after knockout or overexpression BCL6 in skeletal muscle.The target genes were screened by searching the websites of transcription factor and reviewing literatures.Detection the changes of BCL6 and target genes in mice fed with normal and high-fat diet,then observation the effect of knockout and overexpression BCL6 on the target genes.Detection of whether BCL6 affects the transcription of downstream target genes with double luciferase reporting experiment.BCL6 regulation the glucose metabolism in skeletal muscle was further verification in vitro.Results We found that the downstream targets of BCL6 may be the FOXO family genes by screening the website of the transcription factor and reviewing literatures.The FOXO1 expression was significantly increased in wild-type mice and increased more in BCL6 MKO mice,but significantly reduced in BCL6 MOE mice fed with high-fat diet(P<0.01).Inhibition of BCL6 promoted the transcription of FOXO1,while overexpression of BCL6 significantly inhibited of it in the double luciferase reporting experiment(P<0.01).The inhibited glucose uptake by overexpression of BCL6 was partially recovered by overexpression of FOXO1,conversely,the improved glucose uptake by inhibition of BCL6 was partially reversed by overexpression of FOXO1.While the effect of BCL6 on glycogen content can not be reversed by FOXO1 overexpression or inhibiton.Conclusion BCL6 may affect the glucose uptake of skeletal muscle partly by regulation the expression of FOXO1.In addition,BCL6 may affect the use of glucose and glycogen storage by directly or indirectly affected the transcription or activation of the key proteins of glucose metabolism,such as PDK4,GS,etc.