Mechanism Of Lipid-induced White Adipose Tissue Insulin Resistance

Aims:Adipose tissue metabolic dysfunction is associated with metabolic diseases including obesity,type 2 diabetes mellitus and fatty liver diseases.One facet of adipose dysfunction is white adipose tissue(WAT)insulin resistance which is partly characterized by the inability of insulin to suppress lipolysis in obesity,resulting in higher rates of fatty acid flux and ectopic lipid accumulation.To date,numerous studies regarding molecular mechanisms of adipose tissue insulin resistance more focused on hypoxia,inflammatory infiltration,abnormal adipokine secretion,etc.However,mechanism of early adipose tissue insulin resistance is not fully understood.DAG-PKC pathway plays an important role in mediating hepatic and muscular insulin resistance.Furthermore,recent study has showed that PKC? activation is associated with adipose tissue insulin resistance.Nevertheless,the role of DAG-PKC pathway in mediating adipose tissue insulin resistance still need further study.The aims of the study are to assess the role of DAG-PKC pathway in mediating early white adipose tissue insulin resistance,and also to examine if phosphorylation of INSR Thr1150 play a role in mediating white adipose tissue insulin resistance,which provides a new therapeutic strategy to improve adipose tissue insulin resistance.Methods:(1)This study established 7-day high fat diet-induced adipose tissue insulin resistance rat model.We assessed adipose tissue insulin sensitivity by using hyperinsulinemic-euglycemic clamp combined with infusion of[1,1,2,3,3-d5]glycerol and[U-13C16]potassium palmitate,and evaluated basal and insulin stimulated lipolysis rates including glycerol turnover,palmitate turnover,fatty acid turnover by gas chromatography-mass spectrometer,insulin signaling pathway and downstream signaling with regards to lipolytic pathway by western blot(2)To further investigate the role of DAG-PKC pathway in meditating adipose tissue insulin resistance,we further assessed distribution of DAG stereoisomers in adipose tissue subcellular fractionations and PKCs activation based on the early adipose tissue insulin resistance rat model.In addition,we measured adipose tissue hypoxia and inflammatory infiltration related gene expression.(3)To further investigate the role of phosphorylation of insulin receptor Thr1150 in meditating adipose tissue insulin resistance,we assessed adipose tissue insulin sensitivity by using hyperinsulinemic-euglycemic clamp.In addition,we measured basal and insulin stimulated lipolysis rates by gas chromatography-mass spectrometer and insulin signaling pathway related to lipolytic machinery in WT and InsrT1150A mice subjected to 7-day high fat diet.Results:(1)Rats subjected to 7-day high fat diet(vs.RC)displayed impaired insulin suppression of lipolysis approximately 20%as reflected by decreased insulin's ability to suppress plasma non-esterified fatty acid,glycerol turnover,palmitate turnover,fatty acid turnover.In addition,7-day high fat diet rats showed impaired insulin signaling pathway as reflected by decreased pINSR/INSR,pAkt/Akt2;activation of cAMP/PKA mediated phosphorylation of PLIN?HSL and ATGL increased in rats fed 7-day high fat diet.(2)Rats subjected to 7-day high fat diet displayed specific distribution of DAG stereoisomers in adipose tissue subcellular fractionations which only sn-1,2 DAG increased in adipose tissue membrane fraction without alterations in sn-1,3 DAG and sn-2,3 DAG in all five subcellular compartments.Simultaneously,only PKC?translocation,rather than other PKC isoforms,increased in rats fed 7-day high fat diet.In addition,7-day high fat feeding has no effect on adipose tissue gene expression related to inflammatory infiltration such as TNFa?IL-6?F4/80?MCP-1,and hypoxia mediator HIF-1?.(3)InsrT1150A mice subjected to 7-day high fat diet exhibited increased insulin suppression of lipolysis as reflected by increased insulin's ability to suppress plasma non-esterified fatty acid,glycerol turnover,palmitate turnover,fatty acid turnover.Furthermore,InsrT1150A mice restored insulin signaling pathway as reflected by increased pINSR/INSR,pAkt/Akt2.Besides,cAMP/PKA mediated phosphorylation of PLIN?HSL and ATGL decreased in InsrT1150A mice compared to WT mice.Conclusions:(1)7-day high fat diet induces white adipose tissue insulin resistance in rats which is mainly characterized by impaired insulin suppression of lipolysis,defective insulin signaling pathway at the level of insulin receptor and impaired downstream signaling related to lipolytic pathway(2)Specific increase of sn 1,2-DAG in membrane fraction associated with activation of PKCB in white adipose tissue may play a role in mediating early adipose tissue insulin resistance independent of adipose tissue hypoxia and inflammatory infiltration.(3)InsrT1150A mice are protected from 7-day high fat diet-induced white adipose tissue insulin resistance as reflected by increased insulin suppression of lipolysis and improved insulin signaling pathway at the level of insulin(4)sn 1,2-DAG-PKC?-INSR Thr1150 pathway plays a major role in mediating lipid-induced white adipose tissue insulin resistance.