Rab18通过PPARγ下调PLIN2以减少巨噬细胞脂质蓄积

目的探讨Rab18是否通过过氧化物酶体增殖物激活受体γ（PPARγ）下调脂滴包被蛋白2（PLIN2）以减少巨噬细胞脂质蓄积。方法用氧化型低密度脂蛋白（ox-LDL）处理THP-1巨噬细胞,通过Westernblot法检测ox-LDL作用不同时间（0h、6h、12h和24h）后细胞内Rab18、PPARγ及PLIN2的表达情况,并使用油红O染色法观察细胞内脂质蓄积的变化。制备表达野生型、高活型和低活型Rab18的巨噬细胞,分别加入ox-LDL处理,用Westernblot及细胞免疫荧光染色法检测细胞内PPARγ和PLIN2的表达量,并采用油红O染色法观察细胞内脂质蓄积情况。分别用PPARγ激动剂GW1929和抑制剂T0070907预处理表达高活型Rab18的巨噬细胞,再用ox-LDL孵育该细胞,Westernblot及免疫荧光染色法分别检测巨噬细胞内PPARγ和PLIN2的表达量及PPARγ的核转位情况,同时采用油红O染色法观察细胞内脂质蓄积情况。结果ox-LDL处理24h后,THP-1巨噬细胞Rab18、PPARγ和PLIN2的表达水平显著增加,脂质蓄积增加（P<0.05）。ox-LDL处理后,表达野生型和高活型Rab18的巨噬细胞中PPARγ及PLIN2表达下调,脂质蓄积减少（P<0.05）,而表达低活型Rab18的巨噬细胞上述指标则无明显变化。Rab18能抑制PPARγ的激活,并抑制其核转位。表达高活型Rab18的巨噬细胞中PLIN2表达下调能被PPARγ激动剂GW1929逆转,并使脂质蓄积增多。PPARγ抑制剂处理后,表达高活型Rab18的巨噬细胞中PLIN2表达及脂质蓄积进一步减少。结论Rab18通过抑制PPARγ的激活下调PLIN2表达进而抑制巨噬细胞脂质蓄积。     

Cornus kousa F.Buerger ex Miquel increases glucose uptake through activation of peroxisome proliferator-activated receptor γ and insulin sensitization

Aim of the study

Cornus kousa F.Buerger ex Miquel, an oriental medicinal plant, has been traditionally used for the treatment of hyperglycemia, but its molecular mechanism remains unknown. The goal of this study was to investigate the peroxisome proliferator-activated receptor γ (PPARγ) ligand-binding activity of Cornus kousa and to determine the effects of Cornus kousa on insulin sensitization in 3T3-L1 cells for the treatment of type 2 diabetes.

Materials and methods

PPARγ luciferase transactivation assay was used to evaluate the PPARγ ligand-binding activity of Cornus kousa leaf extract. Western blot analysis, oil Red O staining, and glucose uptake assay were performed to evaluate PPARγ agonistic activity and insulin sensitizing effects of Cornus kousa leaf extract (CKE) in 3T3-L1 cells.

Results

CKE increased PPARγ ligand-binding activity in a dose-dependent manner. In addition, CKE enhanced adipogenesis and the expression of PPARγ target proteins, including glucose transporter 4 (GLUT4) and adiponectin, as well as proteins involved in adipogenesis, including PPARγ and CCAAT/enhancer binding protein α (C/EBPα) in 3T3-L1 adipocytes. Furthermore, CKE led to significant induction of glucose uptake and stimulated insulin signaling, but not to activation of AMP-activated protein kinase (AMPK) signaling. The enhanced glucose uptake by CKE were abolished by treatment with bisphenol a diglycidyl ether (BADGE), a PPARγ antagonist, or LY294002, an inhibitor of phosphoinositide 3-kinase (PI3K), but not by compound C, an AMPK inhibitor.

Conclusion

Consistent with the high PPARγ ligand-binding activity, CKE increased glucose uptake through PPARγ activation and insulin signaling. These results suggest that CKE could have pharmacological effects for the treatment of hyperglycemia and type 2 diabetes.     

肥胖大鼠海马内TNF-α、胰岛素降解酶、PPARγ和Aβ的表达及其作用

目的:检测肥胖大鼠海马内肿瘤坏死因子α(tumornecrosisfactorα,TNF-α)、胰岛素降解酶(insulin-degradingenzyme,IDE)、过氧化物酶体增殖物激活受体γ(peroxisomeproliferator-activatedreceptorγ,PPARγ)和淀粉样β肽(amyloidβ-peptide,Aβ)水平,探讨TNF-α对肥胖大鼠海马内IDE和Aβ的影响,观察TNF-α、Aβ与PPARγ的关系。方法:建立肥胖(obesity,OB)大鼠模型;葡萄糖氧化酶法检测大鼠血糖水平,放射免疫法测血浆胰岛素水平;实时荧光定量PCR检测大鼠海马内TNF-α、淀粉样β蛋白前体(amyloidβ-proteinprecursor,APP)、PPARγ及IDE的mRNA水平;蛋白免疫印记技术及免疫组织化学染色法检测大鼠海马内TNF-α、Aβ42(由APP降解产生)、PPARγ及IDE的蛋白表达。结果:OB组大鼠血糖较正常组无明显差别,胰岛素明显升高并存在胰岛素抵抗(P<0.05);实时荧光定量PCR结果显示肥胖大鼠海马内TNF-α和APPmRNA水平较正常组升高(P<0.01),而PPARγ和IDEmRNA表达减少(P<0.01);蛋白免疫印记技术及免疫组织化学法检测与实时荧光定量PCR结果一致。结论:肥胖时大鼠海马内存在炎性改变,TNF-α表达升高,IDE和PPARγ表达减少,Aβ沉积增加。IDE作为Aβ的重要降解酶,其表达减少是Aβ沉积的关键因素;PPARγ作为调控IDE转录的核转录因子,在本实验肥胖大鼠海马内表达下降,提示IDE生成减少可能与PPARγ作用下降有关。     

阿托伐他汀对自发性高血压大鼠心肌组织PPARs表达的影响

目的：探讨阿托伐他汀对自发性高血压大鼠心肌组织PPARs（peroxisomeproliferator-activatedreceptors,PPARs）表达的影响及其对心肌肥厚的逆转作用与可能机制。方法:自发性高血压大鼠分为阿托伐他汀灌胃治疗组（SHR-A,30mg·kg-1·d-1）及模型组（SHR）,治疗8周,同周龄Wistar-Kyoto鼠为正常血压对照组。治疗前及治疗后2、4、8周测量大鼠尾动脉血压。治疗后测血浆血脂水平,以心脏组织病理分析判断心肌肥厚,Westernblotting检测心肌组织PPARα、PPARγ的表达水平。结果:经过8周治疗,SHR-A组及SHR组血压及血脂水平无明显差异（P＞0.05）。SHR-A组左室重量指数低于SHR组（P＜0.01）。在SHR-A组,PPARα及PPARγ表达高于SHR组（P＜0.01）。结论:阿托伐他汀显著改善自发性高血压大鼠心肌组织PPARs表达,有效逆转左室肥厚,可能与其降压及降脂作用无关。     

PPAR γ与纤维化疾病关系研究进展

【摘要】 纤维化疾病是临床上常见的疾病,至今仍是一个治疗难题。近年来的研究发现,过氧化物酶体增殖物激活受体(PPARs)中PPAR γ亚型的活化或激动剂在器官纤维化的发生过程中有调节作用,可减缓纤维化进程。本文就PPAR γ与肝、肾、肺、心肌、眼及皮肤等器官纤维化发病关系的研究现状与进展做一综述。     

罗格列酮通过抑制RANKL及p-ERK活化抑制类风湿关节炎破骨细胞的分化及功能

目的:探讨罗格列酮(RSG)对类风湿关节炎(RA)成纤维样滑膜细胞(FLS)介导的破骨细胞(Oc)分化及功能的影响及其可能机制。方法:活动期RA患者滑膜体外分离培养FLS,与健康人外周血单核细胞(MNC)共培养,不同浓度RSG(0、5、10和15μmol/L)干预,抗酒石酸酸性磷酸酶(TRAP)染色鉴定Oc并计数;甲苯胺蓝染色和图像分析系统计算骨吸收陷窝面积;Real-timePCR检测共培养体系RANKL和OPG的mRNA表达,Westernblot检测RANKL、OPG、p-ERK、p-p38和p-JNK的蛋白含量。结果:与不加RSG组比较,15μmol/LRSG干预后Oc的数量明显减少(P<0.01),骨吸收陷窝面积也减少(P<0.05);共培养体系RANKL的mRNA及蛋白表达明显降低,OPG的mRNA及蛋白表达明显升高(P<0.01);p-ERK的蛋白含量明显降低(P<0.05),p-p38及p-JNK的蛋白含量则不受影响。结论:RSG通过抑制RANKL及p-ERK活化影响RA关节微环境中组织细胞与免疫细胞的相互作用,从而抑制Oc分化及骨吸收功能。     

消脂护肝胶囊对非酒精性脂肪性肝病大鼠病理及PPARα基因的影响

目的研究消脂护肝胶囊对非酒精性脂肪性肝病（NAFLD）大鼠模型肝组织过氧化物酶体增殖物激活受体α（PPARα）基因表达的影响。方法将32只SD大鼠随机分为4组,每组各8只,空白组予普通饲料,模型组、观察组、对照组以高脂饲料联合四环素腹腔注射建立NAFLD模型,并于造模1周后观察组给予消脂护肝胶囊,对照组予东宝肝泰片治疗,第7周处死大鼠,观察肝组织PPARαmRNA表达情况以及肝脏病理;丙氨酸氨基转移酶（ALT）、天门冬氨酸氨基转移酶（AST）、总胆固醇（TC）、甘油三酯（TG）水平,肝匀浆游离脂肪酸（FFA）、丙二醛（MDA）、超氧化物歧化酶（SOD）含量的变化。结果观察组PPARαmRNA表达增加,血清ALT、AST、TC、TG水平及肝匀浆FFA、MDA含量降低,SOD增加,肝脂肪变性程度减轻,效果优于对照组。结论消脂护肝胶囊能够激活PPARαmRNA表达,同时可阻止脂质过氧化反应,从而发挥防治NAFLD的作用。     

Peroxisomes and Hepatotoxicity

Peroxisomes are ubiquitous organelles of eukaryotic cells and are present in significant amounts in hepatic liver cells. Peroxisomal enzymes contribute to several metabolic pathways including fatty acid, purine and amino acid catabolism or bile acid synthesis. The peroxisomal oxidative reactions produce hydrogen peroxide, mostly degraded by catalase which prevents oxidative stress. Moreover, peroxisomes are involved in arylderivative drug detoxification through its epoxide hydrolase activity.In rodents the exposure of cells to xenobiotic compounds such as fibrates, phthalates/adipates and chlorophenoxyacetic acid derivatives, which are used as hypolipaemic drugs, plasticizers and pesticides respectively, lead to a liver mass increase and to a high peroxisome proliferation. This latter event is due to a strong genetic activation triggered by the PPAR (peroxisome proliferator activated nuclear receptor).Human contrasts with rodent since there is no, or little, effect of the above cited compounds. In contrast, the defect of single or multiple peroxisomal functions caused by genetic disorders lead to an increase of very long chain fatty acid level, which is toxic, especially for brain and kidney. The liver response to xenobiotics of the peroxisome proliferator class may be modulated by auxiliary compounds such as hormones (e.g. thyroid hormone) or nutriments (e.g. retinoids).Originally presented at the Second European Comparative Clinical Pathology Conference, Dijon.     

A case of pseudo-Zellweger syndrome with a possible bifunctional enzyme deficiency but detectable enzyme protein: Comparison of two cases of Zellweger syndrome

Three infants with peroxisomal disorders were investigated clinicobiochemically and neuroradiologically. Two had classical Zellweger syndrome, and cranial CT scans showed typical disproportionate enlargement of the occipital horns of the lateral ventricles (colpocephaly) with marked hypodensity of the white matter. In one female infant, although the clinical findings were similar to those in Zellweger syndrome, some findings, such as elevated transaminase levels, liver fibrosis, the absence of renal cortical cysts and colpocephaly, were negative or milder. Biochemical analyses revealed increased very long-chain fatty acids, dicar☐ylic aciduria and impaired β-oxidation of lignoceric acid. However, peroxisomes were abundantly present in hepatocytes and cultured fibroblasts, and all peroxisomal β-oxidation enzyme proteins were detected on immunoblot analysis. A cell fusion study suggested that the enzyme responsible for this case of ‘pseudo-Zellweger syndrome’ is bifunctional.     

The Hansenula polymorpha per6 mutant is affected in two adjacent genes which encode dihydroxyacetone kinase and a novel protein, Pak1p, involved in peroxisome integrity

The Hansenula polymorpha per6-210 mutant is impaired in respect of growth on methanol (Mut^–) and is characterized by aberrant peroxisome formation. The functionally complementing DNA fragment contains two open reading frames. The first encodes dihydroxyacetone kinase (DAK), a cytosolic enzyme essential for formaldehyde assimilation; the second ORF codes for a novel protein (Pak1p). We have demonstrated that per6-210 cells lack DAK activity, causing the Mut^– phenotype, and have strongly reduced levels of Pak1p, resulting in peroxisomal defects. Sequence analysis revealed that per6-210 contains a mutation in the 3′ end of the DAK coding region, which overlaps with the promoter region of PAK1. Possibly this mutation also negatively affects PAK1 expression. Received: 25 February / 12 May 1998