格列本脲对平滑肌细胞及肌源性泡沫细胞内ACAT转录水平的影响

目的:研究格列本脲(glibenclamide,优降糖)对酰基辅酶A[胆固醇酰基转移酶(ACAT)]mRNA水平的影响,探讨其对ACAT酶的调节水平。方法:在小鼠主动脉平滑肌细胞及肌源性泡沫细胞分别加入格列本脲,孵育24h,用半定量RT-PCR检测ACAT酶mRNA表达。结果:(1)平滑肌细胞转变为泡沫细胞后,ACATmRNA的表达均增加。(2)格列本脲抑制平滑肌源性泡沫细胞中ACAT酶的mRNA的表达。结论:格列本脲在转录水平抑制ACATmRNA的表达,抑制平滑肌细胞向泡沫细胞的转化,可作为防止动脉粥样硬化的有效药物。     

格列本脲对巨噬细胞载脂蛋白E分泌的影响

目的： 通过研究格列本脲对巨噬细胞载脂蛋白E(apo E)分泌的影响，评价格列本脲在2型糖尿病治疗中的应用价值。方法：培养THP 1单核细胞并诱导其分化成为THP 1巨噬细胞；通过负载胆固醇制备巨噬细胞源性泡沫细胞。将格列本脲与THP 1巨噬细胞和泡沫细胞一起孵育， 孵育结束时分别通过酶联免疫测定和Northern blot测定格列本脲对细胞apo E的产生、分泌以及apo E基因表达的影响。结果：格列本脲明显减少THP 1巨噬细胞的apo E分泌(P＜0.01)，对巨噬细胞源性泡沫细胞亦表现出同样作用，同时不改变巨噬细胞和泡沫细胞内apo E含量和apo E mRNA表达水平。 结论：格列本脲影响巨噬细胞apo E的净分泌，这可能不利于2型糖尿病患者血管壁的脂质代谢，尤其不宜于治疗已经存在血脂异常的糖尿病患者。     

PPARγ激动剂对RAW264.7巨噬细胞源性泡沫细胞胆固醇流出的影响

目的观察PPARγ激动剂吡格列酮对RAW264.7巨噬细胞源性泡沫细胞胆固醇流出调节蛋白三磷酸腺苷结合核转运蛋白G1(ABCG1)、肝X受体α(LXRα)表达的影响。方法 (1)体外培养RAW 264.7巨噬细胞,用50 mg/L的氧化型低密度脂蛋白胆固醇(ox-LDL)孵育48 h诱导成泡沫细胞,油红O染色并在光镜下鉴定泡沫细胞形态及变化。(2)以不同浓度吡格列酮(0、5、10、20、30μmol/L)作用泡沫细胞24 h后,酶法检测泡沫细胞内胆固醇酯的含量。用反转录-聚合酶链反应(RT-PCR)及免疫印迹法测定ABCG1、LXRα的mRNA及蛋白的表达。结果 PPARγ激动剂吡格列酮可显著减少泡沫细胞内胆固醇酯含量,并呈浓度依赖性增加RAW264.7巨噬细胞源性泡沫细胞ABCG1、LXRα的mRNA和蛋白的表达。结论 PPARγ激动剂吡格列酮减少泡沫细胞内胆固醇酯的含量可能是通过上ABCG1、LXRα的mRNA及蛋白表达来实现的。     

洛伐他汀、普罗布考对小鼠巨噬细胞源性泡沫细胞内胆固醇含量的影响

目的　观察不同剂量下洛伐他汀和普罗布考对巨噬细胞源性泡沫细胞中胆固醇含量的影响 ,借此进一步探讨药物在促进胆固醇外流中的作用。方法　离体培养小鼠腹膜巨噬细胞 ,利用氧化的低密度脂蛋白蓄积胆固醇形成泡沫细胞 ,给予不同剂量的洛伐他汀与普罗布考后 ,利用高效液相方法测定细胞内胆固醇的含量。结果　模型组胆固醇酯(CE)的含量和胆固醇酯 /总胆固醇 (CE/TC)比值均较正常组和血清对照组升高 ,且CE/TC >5 0 % ,提示泡沫细胞模型建立成功。洛伐他汀可明显减少泡沫细胞内TC、CE含量 ,降低CE/TC比值 ,且有一定剂量依赖性。普罗布考可明显减少泡沫细胞内TC、CE含量 ,但无剂量依赖性 ,对CE/TC比值亦无明显影响。结论　洛伐他汀可促进泡沫细胞内胆固醇外流、降低细胞内胆固醇含量 ,减轻泡沫细胞化。普罗布考可降低细胞内胆固醇含量但对泡沫细胞化的影响不明显。     

NG701通过减少CD36 mRNA表达抑制泡沫细胞形成

目的:观察NG701对泡沫细胞形成及CD36mRNA表达的影响。方法:利用大鼠腹腔提取的巨噬细胞与氧化低密度脂蛋白共培养得到巨噬细胞源性泡沫细胞,加入不同浓度的NG701,观察泡沫细胞内胆固醇含量的变化,并利用RT-PCR法对CD36mRNA表达情况进行检测。结果:NG701能够显著减少泡沫细胞内总胆固醇和胆固醇酯的含量,降低CE/TC的值。并剂量依赖性的降低CD36mRNA表达。结论:NG701能通过减少CD36表达抑制巨噬细胞源性泡沫细胞的形成。     

白细胞介素1β对美伐他汀处理的HepG2细胞胞内胆固醇含量的影响

目的观察白细胞介素1β（IL-1β）能否干扰美伐他汀降低HepG2细胞内胆固醇含量的作用。方法给予不同浓度的美伐他汀(0,1,10,50,100μmol·L^-1)单独处理或联合20 ng·mL^-1IL-1β处理HepG2细胞24 h。用酶学反应法检测3-羟基3-甲基戊二酸单酰辅酶A还原酶（HMG-CoA-R）的酶活性,用实时荧光定量聚合酶链式反应（Real-Time PCR）方法测定低密度脂蛋白胆固醇（LDL）受体中mRNA的表达水平。然后选取100μmol·L^-1美伐他汀单独处理或联合20 ng·mL^-1IL-1β处理HepG2细胞24 h,用酶学反应法测定细胞内胆固醇含量,用Real-Time PCR测定细胞内酰基辅酶A-胆固醇酰基转移酶（ACAT）酶表达。结果美伐他汀能显著抑制HepG2细胞的HMG-CoA还原酶活性。IL-1β能有效地削弱美伐他汀对酶活性的抑制作用,同时进一步上调LDL受体的mRNA表达。IL-1β还能促进美伐他汀处理的HepG2细胞中ACAT mRNA的表达,增加细胞内胆固醇酯的含量。结论 IL-1β能增强HepG2细胞的HMG-CoA还原酶活性,促进ACAT酶和LDL受体表达,导致细胞脂质摄入和酯化增加。     

罗格列酮对人单核巨噬细胞THP-1中胆固醇代谢的影响机制研究

目的:研究罗格列酮(RG)对氧化低密度脂蛋白(oxLDL)诱导的人单核巨噬细胞THP-1源性泡沫细胞中胆固醇代谢的影响机制。方法:将THP-1巨噬细胞分为空白对照组、oxLDL(100 mg.L-1)组和oxLDL(100 mg.L-1)+RG(10μmol.L-1)组,后2组先加oxLDL培养48 h,最后1组再加RG培养48 h,采用高效液相色谱法检测每组细胞内游离胆固醇(FC)和胆固醇酯(CE)的含量,逆转录-聚合酶链式反应(RT-PCR)、蛋白质印迹法分别检测每组细胞中三磷酸腺苷结合盒转运蛋白(ABC)A1、ABCG1 mRNA及蛋白的表达情况。结果:与空白对照组比较,oxLDL组FC、CE含量明显升高,ABCA1、ABCG1 mRNA及蛋白表达均明显降低(P<0.01);与oxLDL组比较,oxLDL+RG组FC、CE含量明显降低,ABCA1、ABCG1 mRNA及蛋白表达均明显升高(P<0.01)。结论:RG可能通过上调THP-1巨噬细胞ABCA1、ABCG1的表达,减少FC在细胞内的蓄积,促进细胞内胆固醇代谢,进而抑制动脉粥样硬化的形成。     

坎地沙坦对泡沫细胞胆固醇代谢的影响

目的观察坎地沙坦(CAN)对氧化低密度脂蛋白(ox-LDL)诱导急性单核白血病细胞(THP-1)源性泡沫细胞及对三磷腺苷结合盒转运子A1(ABCA1)表达水平的作用。方法 THP-1源性细胞加入160 nmol.L-1佛波酯,在RPMI-1640液中孵育48 h,将其诱导分化为巨噬细胞后随机分为3组,对照组加入50 mg.L-1ox-LDL培养液孵育48 h;血管紧张肽Ⅱ(AngⅡ)组加入1×10-6mol.L-1AngⅡ后再加入50 mg.L-1ox-LDL培养液孵育48 h;CAN组加入1×10-6mol.L-1CAN后再加入1×10-6mol.L-1AngⅡ孵育,然后以50 mg.L-1ox-LDL处理48 h。分别采用油红O染色观察细胞内脂滴变化,酶化学法测定总胆固醇和胆固醇酯含量,Western blotting法检测ABCA1蛋白表达水平。结果 AngⅡ作用后,细胞内脂滴明显增多,细胞内总胆固醇和胆固醇酯含量及胆固醇酯/总胆固醇比值明显升高,ABCA1蛋白表达水平降低(P<0.05)。CAN能减少泡沫细胞内脂滴形成,导致细胞内总胆固醇和胆固醇酯含量及胆固醇酯/总胆固醇比值明显降低,一定程度上调泡沫细胞转运蛋白ABCA1表达(P<0.05),促进细胞内胆固醇酯流出。结论 AngⅡ减少泡沫细胞中ABCA1的表达,CAN可通过阻断AngⅡ1型受体促进ABCA1的表达及胆固醇酯流出。     

脂多糖对B类I型清道夫受体表达的调节

目的观察脂多糖（LPS）对THP-1巨噬细胞源性泡沫细胞B类I型清道夫受体（sR-BI）表达和胆固醇流出的影响,并探讨核因子-κB（NF-κB）信号途径在此过程中的作用。方法THP-1巨噬细胞源性泡沫细胞以LPS单独或NF-κB抑制剂对甲苯磺酰-L-苯丙氨酸氯甲基甲酮（TPCK）预处理后再加入LPS处理。Western blot检测sR．BI及核内NF-κBp65蛋白质的表达,液体闪烁计数器检测细胞内胆固醇流出,高效液相色谱分析细胞内总胆固醇、游离胆固醇和胆固醇酯含量。结果LPS抑制sR-BI蛋白质的表达,而增加核内NF-K-κB65蛋白质的表达,LPS使泡沫细胞细胞内胆固醇流出减少,细胞总胆固醇、游离胆固醇与胆固醇酯增加。TPCK预处理后,LPS的这种作用被部分抑制。结论NF-κB信号途径介导LPS对sR—BI表达及细胞内胆固醇流出的抑制作用。     

阿托伐他汀对金黄地鼠糖尿病模型巨噬细胞胆固醇外流的影响

目的：研究不同剂量阿托伐他汀对糖尿病动物模型体内巨噬细胞胆固醇含量的影响，进一步探讨他汀类药物抑制动脉粥样硬化斑块进展的机制。方法：建立糖尿病金黄地鼠动物模型，体外培养腹腔巨噬细胞，在体外与不同剂量阿托伐他汀孵育，利用酶法结合高效液相色谱法测定细胞内胆固醇及胆固醇脂含量。RT—PCR方法检测细胞ABCA1 mRNA表达量的不同。结果：在apoA1存在时，阿托伐他汀可以显著促进巨噬细胞内胆固醇外流，可能与药物增强细胞内胆固醇酯水解酶活性、促进巨噬细胞AB—CA1的表达有关。结论：阿托伐他汀促进ABCA1介导的细胞内胆固醇外流。     

Potential Role of Acyl-Coenzyme A:Cholesterol Transferase (ACAT) Inhibitors as Hypolipidemic and Antiatherosclerosis Drugs

Acyl-coenzyme A:cholesterol transferase (ACAT) is an integral membrane protein localized in the endoplasmic reticulum. ACAT catalyzes the formation of cholesteryl esters from cholesterol and fatty acyl coenzyme A. The cholesteryl esters are stored as cytoplasmic lipid droplets inside the cell. This process is very important to the organism as high cholesterol levels have been associated with cardiovascular disease. In mammals, two ACAT genes have been identified, ACAT1 and ACAT2. ACAT1 is ubiquitous and is responsible for cholesteryl ester formation in brain, adrenal glands, macrophages, and kidneys. ACAT2 is expressed in the liver and intestine. The inhibition of ACAT activity has been associated with decreased plasma cholesterol levels by suppressing cholesterol absorption and by diminishing the assembly and secretion of apolipoprotein B-containing lipoproteins such as very low density lipoprotein (VLDL). ACAT inhibition also prevents the conversion of macrophages into foam cells in the arterial walls, a critical event in the development of atherosclerosis. This review paper will focus on the role of ACAT in cholesterol metabolism, in particular as a target to develop novel therapeutic agents to control hypercholesterolemia, atherosclerosis, and Alzheimer's disease.     

Glibenclamide inhibits cholesterol metabolism in macrophage

Sulfonylureas are generally used in the therapeutic treatment of non-insulin-dependent diabetes mellitus. Little is known, however, whether they also affect the lipid metabolism. Using glibenclamide (GB), a typical sulfonylurea, we have investigated its effects on the lipid metabolism in macrophages, J774 and phorbol ester-treated THP-1 cells. In the whole-cell assay system for cholesteryl ester (CE) accumulation that is induced by addition of chemically modified low-density lipoprotein (LDL), such as Ac-LDL and ox-LDL, GB effectively inhibited the CE accumulation of J774 cells in dose-dependent manners. The inhibition was resulted from increase in free cholesterol but not from change in total cholesterol amount. The results suggest that GB acts on acyl-CoA: cholesterol acyltransferase (ACAT) and inhibits its activity. To confirm the possibility, we then tested GB by another assay system using ACAT that was solubilized from the cells and reconstituted into the liposome composed of phosphatidyl choline- cholesterol. GB inhibition was not so much effective as those by CI-976 and NTE-122, known ACAT inhibitors, but the inhibition was complete in the presence of 100 microM GB. Using cell homogenates of PMA-stimulated THP-1 cells, GB also inhibited the ACAT activity to the level of undifferentiated THP-1 cells. These results indicate that GB acts as ACAT inhibitor but the chemical structure is quite different from the conventional ACAT inhibitors, suggesting it can be a seed to generate potential ACAT inhibitors which do not exhibit toxicity in adrenal gland.     

Effects of NTE-122, a novel acyl-CoA:cholesterol acyltransferase inhibitor, on cholesterol esterification and high-density lipoprotein-induced cholesterol efflux in macrophages

We investigated the effects of a novel acyl-CoA:cholesterol acyltransferase (ACAT) inhibitor, NTE-122 (trans-1,4-bis[[1-cyclohexyl-3-(4-dimethylamino phenyl)ureido]methyl]cyclohexane), on ACAT activities in macrophages originating from several species and high-density lipoprotein (HDL)-induced cholesterol efflux in phorbol 12-myristate 13-acetate (PMA)-treated THP-1 cells. NTE-122 inhibited cell-free ACAT activities in human PMA-treated THP-1 cells and mouse J774.1 cells with IC50 values of 0.88 and 360 nM, respectively. NTE-122 competively inhibited the ACAT activity in PMA-treated THP-1 cells. NTE-122 also inhibited cellular ACAT activities in PMA-treated THP-1 cells, rat peritoneal macrophages and J774.1 cells with IC50 values of 3.5, 84 and 6800 nM, respectively. Furthermore, NTE-122 prevented cholesterol accumulation in PMA-treated THP-1 cells incubated with acetylated low density lipoprotein, simultaneously with HDL, while it caused accumulation of a significant amount of free cholesterol in the absence and even in the presence of HDL. NTE-122 also enhanced HDL-induced cholesterol efflux from established foam cells converted from PMA-treated THP-1 cells. These results suggest that NTE-122, capable of inhibiting macrophage ACAT activity in humans more strongly than those in the other species, exhibits anti-atherogenic effects by preventing the foam cell formation and enhancing the foam cell regression in humans.     

Acyl coenzyme A:cholesterol acyltransferase inhibitors as hypolipidemic and antiatherosclerotic drugs

Acyl coenzyme A:cholesterol acyltransferase (ACAT) is the enzyme that catalyzes the conversion of intracellular cholesterol into cholesteryl esters. Two ACAT isoforms, termed ACAT1 and ACAT2, have been described. ACAT1 is ubiquitously found, with high expression levels in macrophages, adrenals, sebaceous glands and foam cells from human atherosclerotic lesions. In contrast, ACAT2 expression is restricted to the intestine and the liver of mice and non-human primates. The reaction catalyzed by ACAT is essential for intestinal cholesterol absorption, synthesis and secretion of apolipoprotein B (apoB)-containing lipoproteins, and intracellular storage of cholesterol. Therefore, ACAT inhibitors would theoretically reduce plasma cholesterol levels by blocking cholesterol absorption from the diet and by reducing hepatic VLDL synthesis. Moreover, ACAT inhibition could limit the accumulation of cholesteryl esters in the cytoplasm of macrophages, thus reducing the formation of foam cells. In view of these attractive possibilities, a great deal of molecules with ACAT inhibitory properties have been synthesized in the last 20 years. However, only a few of them have reached clinical studies, mainly due to unexpected side effects. On the other hand, most of the compounds assayed in humans have not shown substantial hypolipidemic efficacy. The present article focuses on the current knowledge of the pharmacology of ACAT inhibitors, and, specifically, on the different pharmacological approaches used to evaluate these compounds as hypolipidemic and antiatherosclerotic agents.     

Molecular target of decursins in the inhibition of lipid droplet accumulation in macrophages

During screening for inhibitors of lipid droplet accumulation in mouse peritoneal macrophages, two coumarins identified as decursin and decursinol angelate were isolated from the roots of Angelicae gigantis. The cellular molecular target of these inhibitors in macrophages was studied. Decursin and decursinol angelate inhibited cholesteryl ester (CE) synthesis with IC50 values of 9.7 and 10.1 microM, respectively, whereas they enhanced triacylglycerol (TG) synthesis. Lysosomal metabolism of cholesterol to CE was inhibited by the compounds, indicating that the site of inhibition is one of the steps between the exiting of cholesterol from the lysosomes and CE synthesis in the endoplasmic reticulum. Therefore, acyl-CoA:cholesterol acyltransferase (ACAT) activity in the microsomal fractions prepared from mouse macrophages was studied, and the results showed inhibition of this activity by decursin and decursinol angelate with IC50 values of 43 and 22 microM, respectively. Thus, it was concluded that the compounds inhibit macrophage ACAT activity to decrease CE synthesis, leading to a reduction of lipid droplets in macrophages.     

Molecular target of piperine in the inhibition of lipid droplet accumulation in macrophages

An alkaloid piperine isolated from the Piper Nigrum was found to inhibit lipid droplet accumulation in mouse macrophages, and especially inhibited cholesteryl ester (CE) synthesis (IC50: 25 microM). The metabolism of cholesterol from lysosome to lipid droplet was inhibited with a similar IC50 (18 microM), indicating that the site of inhibition is one of the steps between the lysosomes and the endoplasmic reticulum. Therefore, effects of piperine on acyl-CoA:cholesterol acyltransferase (ACAT) activity in the microsomes prepared from mouse macrophage and liver were studied, to show that the compounds inhibited the activity in both cases (IC50: 9.1, 7.0 microM, respectively). Furthermore, piperine was found to inhibit both ACAT1 and ACAT2 isozymes to a similar extent (IC50: 16, 18 microM, respectively) in cell-based assays using ACAT1- or ACAT2-expressing cells. Thus, it was suggested that piperine inhibited macrophage ACAT to decrease CE synthesis, leading to a reduction of lipid droplets.     

Glibenclamide inhibits accumulation of cholesteryl ester in THP-1 human macrophages

Glibenclamide is an adenosine triphosphate (ATP)-sensitive potassium channel inhibitor that is widely used in treating diabetes mellitus. However, the effects of this drug on cholesterol metabolism and atherogenesis are not well known. We investigated the effects of this agent on the cellular cholesterol metabolism in cultured human macrophages. The effect of glibenclamide was evaluated by the measurement of the cellular contents of total cholesterol, free cholesterol, and cholesteryl ester in the presence of low-density lipoprotein (LDL). The effect on the degradation and association of 125I-labeled LDL (125I-LDL) also were determined. Cholesterol efflux was measured in the absence and the presence of high-density lipoprotein (HDL). The secretion of apolipoprotein E also was determined. The synthesis and hydrolysis of cholesteryl ester were evaluated. Glibenclamide stimulated both synthesis and hydrolysis of cholesteryl ester, and inhibited the net accumulation of cholesteryl ester by LDL in a concentration-dependent manner and even decreased its content compared with time 0 control. This drug had no effect on the degradation or association of 125I-LDL. Glibenclamide promoted the HDL-independent cholesterol efflux by decreasing esterified cholesterol and increasing the release of free cholesterol and secretion of apolipoprotein E into the medium. The other potassium channel inhibitors or openers had no effect on the cellular cholesterol levels. These results suggest that glibenclamide inhibits the accumulation of cholesteryl ester in macrophages by enhancing the hydrolysis of cholesteryl ester as well as by increasing cholesterol efflux, and possibly, by increasing the secretion of apolipoprotein E. These effects appeared to be unrelated to an effect on the potassium channel. Inhibition of accumulation of cellular cholesterol by glibenclamide might be favorable for the prevention of atherosclerotic disease.     

Acyl-coenzyme A:cholesterol acyltransferase inhibitors for controlling hypercholesterolemia and atherosclerosis

Acyl-coenzyme A:cholesterol acyltransferase (ACAT; Sterol O-acyltransferase/SOAT) is an intracellular enzyme that catalyzes the formation of cholesteryl esters from cholesterol and fatty acyl-coenzyme A. ACAT inhibitors reduce plasma cholesterol levels by suppressing absorption of dietary cholesterol and by suppressing the assembly and secretion of apolipoprotein B-containing lipoproteins such as very low density lipoprotein in liver and chylomicron in intestine. Moreover, ACAT inhibitors prevent the conversion of macrophages into foam cells in the arterial walls. Thus, ACAT inhibitors are under investigation for controlling hypercholesterolemia and the development of atherosclerosis. Some potent ACAT inhibitors have been tested for their efficacy and safety in humans.     

Inhibition of cytosolic phospholipase A(2) suppresses production of cholesteryl ester through the reesterification of free cholesterol but not formation of foam cells in oxidized LDL-stimulated macrophages

Macrophage-derived foam cells are formed as a result of the accumulation of cholesteryl ester (CE) not only in cytoplasm where CE is produced by the reesterification of free cholesterol derived from oxidized low density lipoprotein (OxLDL) undergoing hydrolysis, but also in lysosomes where the remaining CE of OxLDL is deposited. We examined the possible involvement of cytosolic phospholipase A(2)s (cPLA(2)s) in the production of CE through the reesterification and in the formation of foam cells. In [(3)H]oleic acid-labeled human acute monocytic leukemia (THP-1) cell-derived macrophages (THP-M) and mouse peritoneal macrophages (MPM), which possessed at least cPLA(2)alpha and cPLA(2)gamma, stimulation with OxLDL induced the production of [(3)H]cholesteryl oleate ([(3)H]CE).The production was suppressed by an inhibitor of cPLA(2)s. However, the inhibitor tended to slightly decrease total intracellular levels of CE, and did not affect the formation of foam cells, as estimated by staining with Oil Red O. In cPLA(2)alpha-knockout MPM, OxLDL-induced increases in [(3)H]CE and total CE did not differ from those in wild-type MPM. Our results suggest that cPLA(2)s other than cPLA(2)alpha contribute to the supply of fatty acids, which are utilized for the production of CE through the reesterification, in OxLDL-stimulated macrophages. However, the formation of foam cells could not be inhibited only by the suppression of cPLA(2)-mediated CE production.     

Effects of troglitazone on intracellular cholesterol distribution and cholesterol-dependent cell functions in MA-10 Leydig tumor cells

Troglitazone treatment of MA-10 Leydig tumor cells resulted in cellular cholesteryl esters decreasing and cell free cholesterol increasing. This was not an effect unique to this chemical entity; rosiglitazone and pioglitazone caused these changes also. The excess free cholesterol was recovered largely in the cholesterol oxidase susceptible, plasma membrane cholesterol pool. This effect of troglitazone probably is not mediated by activation of peroxisome proliferator activated receptors since it immediately reversed with washing and did not occur at all in cells treated with the peroxisome proliferator activated receptor agonist, 15-deoxy Delta 12,14 prostaglandin J-2. Plasma membrane cholesterol esterification was inhibited by troglitazone in a dose-dependent manner. Plasma membrane cholesterol esterification was inhibited half-maximally by 14 microM troglitazone and by more than 90% by 40 microM troglitazone. This effect was not unique for MA-10 cells. Similar results were found using fibroblasts. Troglitazone was not simply inhibiting internalization of plasma membrane cholesterol. Dibutyryl-cAMP stimulation of troglitazone-treated cells resulted in more progesterone synthesis than in stimulated control cells; moreover, radioactive plasma membrane cholesterol was readily converted into progesterone in troglitazone-treated cells. Studies of LDL uptake in troglitazone-treated cells indicated that intracellular membranes were cholesterol replete. Troglitazone inhibited plasma membrane cholesterol esterification with kinetics similar to 58-035, a known inhibitor of the acyl coenzyme A: cholesterol acyltranserase (ACAT) enzyme. It is not likely an ACAT inhibitor since troglitazone did not block incorporation of exogenous free fatty acids into cholesteryl esters. Thus, it appears that troglitazone prevented presentation of free fatty acid to the ACAT enzyme.