Effect Of PPARγ On Cholesterol Efflux Of C57 Mice Peritoneal Macrophage With Inflammatory Situation

Atherosclerosis ( AS ) is the pathologic background of many cardiovascular diseases,it is induced by many factors and regulated by many genes. The foam cells resulted from cholesterol dysequilibrium and dysbolism in macrophages is the most important pathologic character. As one of predecessors of foam cells,the macrophage cholesterol counter transport(cholesterol efflux) is an important link to maintain the balance of cholesterol in cells and influence the formation of foam cells. This process is regulated by many signal iter and transcription factors. The peroxisome proliferators-activated receptor gamma(PPARγ) is a member of nuclear hormone superfamily, PPARγhave been implicated in such dirverse pathways as lipid metabolism and glucose homeostasis, promoting adipocyte and monocyte/macrophage differentiation. Many studies of recent years have indicated:PPARγand its ligands are important roles in cholesterol efflux of macrophage. It can promote cholesterol efflux of macrophage through PPARγ-LXRα-ABCA1 channel. In addition, PPARγcan downregulate expressed of pro-imflammation genes in macrophage by transaction and suppresses the pro-inflammatory responses obviously. Research on the anti-inflammatory effects of PPARγhas showed that its agonists obviously inhibit the secretion of pro-inflammatory mediators such as TNFα,IL-1,IL-6 and NO in activated monocytes and macrophages. Pretreatment of wild-type mice with PPARγligands can decrease the expression of pro-inflammatory cytokines, alleviate the injury of local and distant tissues, which plays the therapeutic effect on many inflammatory diseases, such as acute myocarditis, autoimmune encephalitis and multiple sclerosis. Now as well known that the atherosclerosis is the consequence injured by disorder of lipid metabolism and chronic imflammation. Thus , with multip factors(hypercholesterolemia,acute,chronic inflammation) of promoting AS existing in solo or coexist,Making clear how the cholesterol efflux of macrophage changes and the role that PPARγplaying in these situations is very significant. It will conduce treveal the cause of foam cells and expand new methods in preventing and curing atherosclerosis.In view of the behavior of PPARγon various key transcriptional factors,This research make the PPARγas the key point between regulated macrophage cholesterol efflux and suppressesed inflammatory response, observe its effection and how macrophage cholesterol efflux changes with inflammation in vivo and vitro. firstly constructing C57BN/6J mice model under different metabolic situation including hyperlipoidemia,acute inflammation and isolating culture its peritoneal macrophage,observing the expression of PPARγand IκB-α,identify the character of macrophage cholesterol efflux in every group;then pretreat the normol C57 mice peritoneal macrophage with PPARγligand ciglitazone and PPARγantisense oligonucleotide,observing the effection to cholesterol efflux after simulated with LPS(100ng/ml)in vitro.It just provide an evidence to elucidate the possible mechanism of PPARγon C57 mice peritoneal macrophage with inflammatory situation cholesterol efflux and the role of PPARγin maintaining the balance between the cholesterol efflux and anti-inflammatory.The main results are as follows:1. The level of mice serum lipids ( total cholesterol and total triglyceride) of high fat/high cholesterol diet(HFHC)group(4.17±0.68,3.47±0.59 )was higher significantly than that of normal diet group(1.71±0.33,1.21±0.29,P<0.05),There were no significantly difference between the two groups after stimulated by LPS. The level of mice with HFHC and normal diet blood plasma TNF-αstimulated by LPS(7.09±1.95,6.64±1.39)were higher significantly than the mice with normal diet(2.09±0.37,P<0.01),there were no difference in statistics between the mice with HFHC diet and with normal diet.2. It can be observed by oil red O stained that lipid droplet deposited in macrophage of HFHC diet. The results of immuno chemistry showed that no PPARγprotein expression in groups of HFHC diet and normal diet, the expression increased lightly after stimulated by LPS; There were no NF-κB expression in group of normal diet, but the expression increased significantly after stimulated by LPS.3. The results of Western-blotting showed that the expression of PPARγprotein in groups of HFHC diet and stimulated by LPS(1.97±0.31,3.19±1.21,3.27±0.88 )were higher significany than that of control group(1.00, P<0.05) ; The expression of IκB-αin HFHC diet group(0.88±0.19) was lower lightly than in control group(1.00, P>0.05),but the expression in groups stimulated by LPS(0.49±0.07,0.44±0.11) was all lower significantly than in control grouph(1.00 ,P<0.01).4.The results of RT-PCR showed that the expression of PPARγmRNA of HFHC diet group and stimulated by LPS(0.67±0.18,0.75±0.22) were higher significantly than that of control group(0.39±0.11, P<0.05),the expression of normal diet stimulated by LPS (0.47±0.13,P<0.05)was higher lightly than that of control group.5.The determination of cholesterol efflux showed that this function of macrophage with HFHC die(t37%)was more enhanced than that of control group(25%,P<0.05) but was inhibited in group stimulated by LPS(18%,14%, P<0.05).6. To normal peritoneal macrophage pretreat with ciglitazone and stimulated by LPS : the expression of PPARγprotein and mRNA (0.31±0.11,0.17±0.08)was higher than that of control group(1.00,0.42±0.12, P<0.05) but the expression of IκB-α(0.81±0.12) was depressed obviously (1.00,P<0.05),The cholesterol efflux was depressed but the amplitude(11.37%) was lower than that of control group(25.72%).7.To normol peritoneal macrophage pretreat with PPARγantisense oligonucleotide and stimulated by LPS:the expression of PPARγprotein and mRNA (0.31±0.11,0.17±0.08)was lower than that of control group(1.00,0.42±0.12,P<0.05)but the expression of IκB-α(0.29±0.08)was depressed obviously(1.00,P<0.01),The cholesterol efflux was depressed but the amplitude (47.82%)was higher than that of control group(25.72%).Conclusions:1.Stimulated with LPS can increase the expression of PPARγin C57 mice peritoneal macrophage.2. The cholesterol efflux of C57 mice peritoneal macrophage was depressed after stimulated by LPS.3.The PPARγligand ciglitazone can increase the cholesterol efflux of C57 mice peritoneal macrophage and weaken the inhibition stimulated by LPS;The PPARγantisense oligonucleotide can depress it and aggravate the inhibition.4. The anti-imflammation character of PPARγresult from suppress the activity of NF-κB signal iter. The inhibition from LPS to peritoneal macrophage cholesterol efflux may because of anti-imflammation character of PPARγ.