The Expression Of ICAM-1 In Endothelium And The Expression Of MCP-1 And CD36 In Monocyte/macrophage Induced By Visfatin

Chapter One Expression of ICAM-1 induced by visfatin in human umbilical vein endothelial cellsBackground:Visfatin is the most recently identified adipocytokine(known previously as pre-B cell colony enhancing factor,PBEF)which appears to be preferentially produced by visceral adipose tissue.Visfatin plasma concentrations are increased in humans with abdominal obesity or type 2 diabetes mellitus,visfatin increased adipocytes and monocytes secrete cytokines such tumor necrosis factor alpha,interleukin-6 and interkin-1βin vitro.The human monocytes and adipocytes in vivo can secret visfatin also.Visfatin can stimulate the monocyte release intercellular adhesion molecule-1 and the endothelium release vascular endothelium growth factor.Clinical researchers found that visfatin expressed in carotid endarterectomy specimens and coronary artery plaques,and the symptomatic patients were marked high than asymptomatic patients.This indicates that visfatin is not only an cytokine but also interact with atherosclerosis.The intercellular adhesion molecule-1 on endothelium can mediate monocyte adhesion to the vascular endothelium and accelerate atherosclerosis.The effect of visfatin on endothelium expressing ICAM-1 hasn't been researched.Obejiective:The aim of this study was to evaluate the effect of visfatin or visfatin +ox-LDL or visfatin+high glucoseon on endothelium expression of ICAM-1.Method:HUVECs in the third to sixth passage were used in the present experiments. We evaluated the endothelium ICAM-1 in three different treatment with visfatin. 1.The ICMA-1 level of endothelium after different levels of visfatin(100-800ng/ml)treatment for 24 hours with or without ERK_1/2signal pathway inhibitor PD98059 pretreatment for 30 minutes.2.The ICAM-1 level of endothelium after ox-LDL(40ug/ml)and different levels of visfatin for 24 hours with or without ERK_1/2signal pathway inhibitor PD98059 pretreatment for 30 minutes.3.The ICAM-1 of endothelium after high glucose(30mM)and different levels of visfatin for 24 hours with or without ERK_1/2signal pathway inhibitor PD98059 pretreatment for 30 minutes.We use the RT-PCR method to detect the ICAM-1 mRNA levels and the western-blotting method to detect the ICAM-1 protein levels.And to elucidate the effects of visfatin or the visfatin+ox-LDL or the visfatin+high glucose on endothelium ICAM-1 expression in the nucleonic acid and protein level.Results:1.The endothelium stimulated with visfatin and resulted in a dose-dependent induction of ICAM-1,the level of ERK_1/2protein was increased also and the ERK_1/2inhibitor PD98059 can markedly inhibit the expression of ICAM-12.The endothelium stimulated with visfatin and ox-LDL and resulted in synergistic effect of induction of ICMA-1,the level of ERK_1/2protein was increased also and the ERK_1/2inhibitor PD98059 can markedly inhibit the expression of ICAM-1.3.The endothelium stimulated with visfatin and high glucose and resulted in synergistic effect of induction of ICMA-1,the level of ERK protein was increased also and the ERK_1/2inhibitor PD98059 can markedly inhibit the expression of ICAM-1.4.Visfatin promoted the endothelium ICAM-1 expression as well as the adhesion of monocyte and endothelium.Conclusion:The endothelium stimulated with visfatin can soon result in a dose-dependent ICAM-1 expression.The endothelium stimulated with visfatin and ox-LDL or visfatin and high glucose result in synergistic effect effect of promote ICAM-1 expression.The activation of ERK_1/2signaling pathway was crucial in endothelium expression of ICAM-1 induced by visfatin. Chapter Two Expression of Monocyte Chemoattractant Protein-1 Induced by visfatin in THP-1 cellsBackground:The increase of MCP-1 expression of Monocyte and the followed increase of migration of monocyte through endothelium luminal is an important accelerate factor during atherosclerosis formation.Visfatin is a new found adipocytokine recently,it was found increased in obesity and diabetes and hypertension,such diseases were intensively connected with atherosclerosis.The current research found that visfatin can promote ICAM-1 expression in monocyte and VEGF expression in endothelium.Visfatin was found markly increased in carotid plaques and coronary artery plaques,and the symptomatic patients were more than asymptomatic patients.This indicate that visfatin probably connected with the progression of atherosclerosis.The expression of MCP-1 in monocyte can accelerate the monocytes recruitment and migration.The impact of visfatin to the MCP-1 expression in monocyte doesn't been researched.Obejiective:The aim of this study was to evaluate the effect of visfatin or visfatin +ox-LDL or visfatin +high glucoseon on MCP-1 expression in THP-1 Monocyte cells and the migration of THP-1.Method:THP-1 cells in the third to sixth passage were used in the present experiments.We evaluated the cell MCP-1 in three different treatment with visfatin.1.The MCP-1 level in THP-1 cells after different levels of visfatin(100-800ng/ml)treatment for 24 hours with or without p38 MAPK signal pathway inhibitor SB202190 pretreatment for 30 minutes.2.The MCP-1 level in THP-1 cells after ox-LDL(40ug/ml)and different levels of visfatin treatment for 24 hours with or without p38 MAPK signal pathway inhibitor SB202190 pretreatment for 30 minutes.3.The MCP-1 in THP-1 cells after high glucose(30mM)and different levels of visfatin treatment for 24 hours with or without p38 MAPK signal pathway inhibitor SB202190 pretreatment for 30 minutes.We use the RT-PCR method to detect the MCP-1 mRNA levels and the western-blotting method to detect the MCP-1 protein levels so as to elucidate the effects of visfatin or the visfatin+ox-LDL or the visfatin+high glucose on THP-1 MCP-1 expression in the nucleonic acid and protein level.Results:1.The THP-1 monocytes stimulated with visfatin and resulted in a dose-dependent induction of MCP-1,the level of P38 protein was increased also and the P38 inhibitor SB202190 can markedly inhibit the expression of MCP-12.The THP-1 monocytes stimulated with visfatin and ox-LDL and resulted in synergistic effect of induction of MCP-1,the level of P38 protein was increased also and the P38 inhibitor SB202190 can markedly inhibit the expression of MCP-1.3.The THP-1 monocytes stimulated with visfatin and high glucose and resulted in synergistic effect of induction of MCP-1,the level of P38 protein was increased also and the P38 inhibitor SB202190 can markedly inhibit the expression of MCP-1.Conclusion:The monocyte stimulated with visfatin can soon result in a dose-dependent MCP-1 expression.The monocyte stimulated with visfatin and ox-LDL or visfatin and high glucose result in synergistic effect effect of MCP-1 expression. The activation of p38MAPK signaling pathway was crucial in monocyte expression of MCP-1 induced by visfatin. Chapter Three Expression of CD36 Induced by visfatin in THP-1 Macrophage cellsBackground:During the atherosclerosis plaque formation,the increase of CD36 expression in Macrophage and the followed increase of uptake of oxidized LDL and Advanced Glycation End products(AGEs)and then macrophage turned into foam cell accelerate the plaque formation.THE activation of PPARγsignaling pathway can increase the CD36 expression.Visfatin is a new found adipokine recently,it was found marldy increased in carotid plaques and coronary artery plaques,and the symptomatic patients were more than asymptomatic patients. This indicate that visfatin probably connected with the progression of atherosclerosis.The impact of visfatin to the CD36 expression in macrophage doesn't been researched.Obejiective:The aim of this study was to evaluate the effect of visfatin or visfatin+ox-LDL or visfatin+high glucose to CD36 expression in THP-1 Macrophage cells.Method:THP-1 cells in the exponential phase of growth were treated by phorbol and converted to macrophages,then the macrophages was used for experiments.We measured the THP-1 CD36 and PPARγin three different treatment with visfatin. 1.The CD36 and PPARγexpression in THP-1 after different levels of visfatin(100-800ng/ml)treatment for 24 hours with or without PPARγsignal pathway inhibitor GW9662 pretreatment for 30 minutes.2.The CD36 and PPARγexpression in THP-1 after ox-LDL(40ug/ml)and different levels of visfatin treatment for 24 hours with or without PPARγsignal pathway inhibitor GW9662 pretreatment for 30 minutes.3.The CD36 and PPARγexpression in THP-1 after high glucose(30mM)and different levels of visfatin treatment for 24 hours with or without PPARγsignal pathway inhibitor GW9662 pretreatment for 30 minutes.We use the RT-PCR method to detect the CD36 and PPARγmRNA and the western-blotting method to detect the CD36 protein so as to elucidate the effects of visfatin or the visfatin+ox-LDL or the visfatin+high glucose on THP-1 CD36 expression in the nucleonic acid and protein form.Results:1.Visfatin increased THP-1 macrophage CD36 and PPARγexpression in a dose dependent manner and the PPARγsignaling pathway inhibitor GW9662 can almost completely inhibit the CD36 expression.2.The THP-1 macrophage stimulated by visfatin+ox-LDL and resulted in synergistic effect of induction of CD36 and PPARγexpression and cholesterol accumulation increased,PPARγinhibitor GW9662 can almost completely inhibit the effect of visfatin+ox-LDL.3.The THP-1 macrophage stimulated by visfatin+high glucose and resulted in synergistic effect of induction of CD36 and PPARγexpression,PPARγinhibitor GW9662 can almost completely inhibit the effect of visfatin+high glucose.ConclusionThe macrophage stimulated with visfatin can soon result in a dose-dependent CD36 expression.The macrophage stimulated with visfatin and ox-LDL or visfatin and high glucose result in synergistic effect effect of CD36 expression. The activation of PPAR7 signaling pathway was crucial in macrophage expression of CD36 induced by visfatin.