反义adipophilin寡核苷酸降低ACAT活性(英)

已经发现,随着细胞内胆固醇酯的积聚,adipophilin的表达明显增加.在此基础上,为了寻找adipophilin在细胞内胆固醇代谢的作用点.小鼠腹腔巨噬细胞与80 mg/L OxLDL或80 mg/L OxLDL加1 mmol/L adipophilin反义寡核苷酸共孵育,在不同的时间点取样,使用免疫荧光染色,流式细胞仪分析和细胞内胆固醇测定.结果显示,72 h后,反义寡核苷酸处理组细胞内胆固醇酯显著下降到（19.9±1.9）mg/g,油红O染色显示,该组细胞质内红色脂滴明显减少.96 h的观察期间内,两组细胞adipophilin蛋白的表达量都增加.从12 h开始,未用反义寡核苷酸处理组adipophilin蛋白的表达量开始大于反义寡核苷酸处理组.在96 h时间点,统计学处理,未用反义寡核苷酸处理组明显高于反义寡核苷酸处理组,差别有显著性.使用Ox-r［CL-³H］LDL观察两组细胞摄入OxLDL的量,实验发现,两组细胞摄入Ox-r［CL-³H］LDL的量逐渐增加.但是,反义寡核苷酸处理组摄入Ox-r［CL-³H］LDL的量从24 h后明显低于未用反义寡核苷酸处理组,在48 h和96 h时间点,统计学处理,两组差别有显著性.观察ACAT的活性发现,ACAT相对活性从6 h到48 h表现为增加,但从48 h到96 h,ACAT相对活性趋于稳定.在48 h时间点,两组比较差别有显著性,反义寡核苷酸处理组的活性明显低于未用反义寡核苷酸处理组.相关分析发现,ACAT的活性与adipophilin蛋白的表达量有一定的联系,但不是直线相关.结果表明,adipophilin表达的高低与细胞摄入外源性脂蛋白,与ACAT的活性有一定的联系.提示adipophilin与脂滴的代谢功能密切相关,且ACAT有可能是其潜在的位点.     

The effects of (22S,23S)- and (22R,23R)-3β-hydroxy-22,23-oxido-5α-ergost-8(14)-en-15-ones on biosynthesis of cholesteryl esters and activity of acyl-CoA:Cholesterol acyl transferase in Hep G2 cells

Novel synthetic oxysterols (22S,23S)-3β-hydroxy-22,23-oxido-5α-ergost-8(14)-en-15-one (I) and (22R,23R)-3β-hydroxy-22,23-oxido-5α-ergost-8(14)-en-15-one (II) influenced biosynthesis of cholesteryl esters from [¹⁴C]acetate (85% and 180% of control at 5 μM concentration) in the human hepatoma Hep G2 cell line. Ketosterol (I) increased the level of cholesteryl ester biosynthesis from [¹⁴C]oleate in Hep G2 cells in a dose dependent manner, whereas the level of cholesteryl esters biosynthesis in the presence of ketosterol (II) reached the maximal value (269±20% of control) at 1 μM concentration of this compound. In a cell free system ketosterol (I) increased the rate of ACAT-dependent cholesterol acylation similar to 25-hydroxycholesterol, however, ketosterol (II)), efficiently stimulated an initial rate of ACAT-catalyzed cholesterol esterification, followed by rapid inactivation of this enzyme.     

Acyl-CoA: cholesterol acyltransferase-2 gene polymorphisms and their association with plasma lipids and coronary artery disease risks

Acyl-CoA: cholesterol acyltransferase-2 (ACAT2), an intracellular cholesterol esterification enzyme found only in the intestine and liver, has been demonstrated to be associated with hypercholesterolemia and atherosclerosis in mice. To explore the possible impact of ACAT2 gene variants on CAD susceptibility and plasma lipid levels, three polymorphisms, 41A>G (Glu>Gly), 734C>T (Thr>Ile), and IVS4-57_58 ins48 bp (D/I), were genotyped in 809 CAD patients (CAD+) and 1,304 controls (CAD−) from three distinct Singaporean ethnic groups (1,228 Chinese, 367 Malays and 518 Indians). The 734T allele frequency was significantly lower in CAD+ (0.20) than CAD− (0.26) in Chinese (P=0.003) and I allele of D/I was significantly higher in CAD+ (0.17) than CAD− (0.10) in Indians (P=0.011). The 41G allele was significantly more frequent among normolipidemic (0.19) than dyslipidemic (0.13) individuals in Chinese (P=0.008). In normolipidemic females, 734C>T was associated with apoA1, apoB and lipoprotein (a) in Indians, and with apoA1 in Malays, whereas 41A>G is associated with total cholesterol in Indians. The 734C>T polymorphism was in almost complete linkage disequilibrium (LD) with the IVS4-57_58 ins48 bp and in very strong LD with 41A>G in all the three ethnic groups. In the normolipidemic females, the AG/CT had much higher apoB than AA/CC in Indians. We found that the three ACAT2 polymorphisms studied are associated with CAD risk and plasma lipid levels but their effects are not consistent across genders and ethnic groups.     

Hypocholesterolemic effect of rice protein is due to regulating hepatic cholesterol metabolism in adult rats

Aging is one of major risk factors for developing hypercholesterolemia. To elucidate the cholesterol-lowering mechanism exerted by rice protein (RP), the effects on hepatic cholesterol outputs and cholesterol metabolism related enzymes were investigated in adult rats, which were fed by casein (CAS) and RP without cholesterol in diets. After 2 weeks of feeding, the significant cholesterol-lowering effect was observed in adult rats fed by RP compared to CAS. The hepatic total- and VLDL-cholesterol secretions into circulation were significantly depressed in RP group, whereas biliary outputs of bile acids and cholesterol were effectively stimulated by RP-feeding, causing an increase in fecal sterol excretion compared to CAS. As a result, the apparent cholesterol absorption was significantly inhibited by RP. RP-feeding significantly increased the activity and gene expression of cholesterol 7α-hydroxylase, whereas acyl-CoA:cholesterol acyltransferase-2 activity and gene expression were significantly decreased by RP as compared with CAS. Neither activity nor gene expression of 3-hydroxy-3-methylglutaryl coenzyme A reductase of RP did differ from CAS in the liver. The present study demonstrates that rice protein can prevent hypercholesterolemia through modifying hepatic cholesterol metabolism under cholesterol-free dietary condition. The findings suggest that hypocholesterolemic action induced by rice protein is attributed in part to the inhibition of cholesterol absorption during the adult period.     

New functions of mitochondria associated membranes in cellular signaling

In all eukaryotic cells, the endoplasmic reticulum (ER) and the mitochondria establish a tight interplay, which is structurally and functionally modulated through a proteinaceous tether formed at specific subdomains of the ER membrane, designated mitochondria-associated membranes or MAMs. The tethering function of the MAMs allows the regulation of lipid synthesis and rapid transmission of calcium (Ca^2 +) signals between the ER and mitochondria, which is crucial to shape intracellular Ca^2 + signaling and regulate mitochondrial bioenergetics. Research on the molecular characterization and function of MAMs has boomed in the last few years and the list of signaling and structural proteins dynamically associated with the ER–mitochondria contact sites in physiological and pathological conditions, is rapidly increasing along with the realization of an unprecedented complexity underlying the functional role of MAMs. Besides their established role as a signaling hub for Ca^2 + and lipid transfer between ER and mitochondria, MAMs have been recently shown to regulate mitochondrial shape and motility, energy metabolism and redox status and to be central to the modulation of various key processes like ER stress, autophagy and inflammasome signaling. In this review we will discuss some emerging cell-autonomous and cell non-autonomous roles of the MAMs in mammalian cells and their relevance for important human diseases. This article is part of a Special Issue entitled: Calcium signaling in health and disease. Guest Editors: Geert Bultynck, Jacques Haiech, Claus W. Heizmann, Joachim Krebs, and Marc Moreau.     

人细胞内胆固醇酯化酶ACAT

酰基辅酶A:胆固醇酰基转移酶(ACAT)是细胞内唯一催化游离胆固醇与长链脂肪酸形成胆固醇酯的酶,在维持细胞内胆固醇、脂肪酸等脂质代谢平衡中起着极重要的作用。人类一些重要疾病如心脑血管病的动脉粥样硬化、神经系统的阿尔茨海默症、消化吸收系统的胆囊病变等都与ACAT紧密关联。本文主要从人ACAT的表达、功能效应、抑制剂应用前景等三方面介绍有关的研究进展。     

ACAT inhibition and amyloid beta reduction

Alzheimer's disease (AD) is a devastating neurodegenerative disorder. Accumulation and deposition of the beta-amyloid (Aβ) peptide generated from its larger amyloid precursor protein (APP) is one of the pathophysiological hallmarks of AD. Intracellular cholesterol was shown to regulate Aβ production. Recent genetic and biochemical studies indicate that not only the amount, but also the distribution of intracellular cholesterol is critical to regulate Aβ generation. Acyl-coenzyme A: cholesterol acyl-transferase (ACAT) is a family of enzymes that regulates the cellular distribution of cholesterol by converting membrane cholesterol into hydrophobic cholesteryl esters for cholesterol storage and transport. Using pharmacological inhibitors and transgenic animal models, we and others have identified ACAT1 as a potential therapeutic target to lower Aβ generation and accumulation. Here we discuss data focusing on ACAT inhibition as an effective strategy for the prevention and treatment of AD.