过氧化物酶体增殖物激活受体α的研究进展

过氧化物酶体增殖物激活受体（Peroxisome proliferator activated receptors,PPARs）作为核受体超家族的一员,其作用广泛,可调节脂肪细胞因子表达、抑制炎症因子、改善胰岛素抵抗等。PPARs有三种亚型,分别是：PPARα、PPARβ／δ和PPARγ。其中PPARα是PPARs最主要的亚型,主要分布在肝脏中。PPARα由不饱和脂肪酸或贝特类降脂药物等配体活化后形成异二聚体,调控靶基因的表达,发挥生物学功能。PPARα参与调节肝脏脂质吸收、脂肪酸氧化、酮体生成、胆固醇代谢等脂代谢过程,以及糖代谢、炎症反应和细胞增殖等,与脂肪性肝病、肝脏炎症反应、乙肝病毒复制和肝癌等肝脏疾病密切相关。本文对PPARα的结构、作用机制、生物学功能及其与肝脏疾病的关系进行综述。PPARα作为肝脏疾病一个新的治疗靶点,阐明其与肝脏疾病发生机制之间的关系,有助于为肝脏疾病的治疗提供新的途径。     

外源性脂肪酸对罗非鱼脂肪细胞增殖与分化的影响

为了探究脂肪酸对罗非鱼(Oreochromis niloticus)脂肪细胞增殖和分化的影响, 在体外培养罗非鱼前脂肪细胞, 并在其增殖和分化过程中分别添加100 μmol/L的棕榈酸(Palmitic Acid, PA)、油酸(Oleic Acid, OA), 亚油酸(Linoleic Acid, LA)和α-亚麻酸(α-Linolenic Acid, LNA)进行处理。使用SRB (Sulforhodamine B)染色法和油红O染色法检测外源性脂肪酸对脂肪细胞增殖和分化的影响, Real-time qPCR检测增殖分化过程中基因表达情况。结果显示, 在培养8d时, 外源添加的不饱和脂肪酸可以促进罗非鱼前脂肪细胞增殖, 并且增殖过程中增殖相关基因(c-fos和c-myc)、脂解相关基因(ATGL)和脂合成相关基因(PPARγ和CD36)的表达与对照组相比均显著提高(P<0.05)。此外, 外源脂肪酸的加入可以抑制脂肪细胞的分化。棕榈酸的加入使得脂肪细胞中产生的脂滴面积较少, 数量较多; 分化过程中细胞的β氧化相关基因(CPT-1a)与对照组相比显著上调, 而脂解相关基因(ATGL)则显著下调。外源性不饱和脂肪酸可以促进罗非鱼前脂肪增殖, 而饱和脂肪酸主要抑制细胞分化。在增殖过程中, 过量的脂肪酸先通过脂合成储存在胞内, 再借助脂解等途径进行代谢, 从而帮助细胞适应环境中高浓度的脂肪酸。而在分化过程中, 添加外源脂肪酸, 可能通过抑制脂肪细胞内的脂合成和脂解的发生, 同时促进β氧化等方式来抑制脂肪细胞分化。     

Involvement of PPARα and PPARγ in apoptosis and proliferation of human hepatocarcinoma HepG2 cells

Peroxisome proliferator‐activated receptors (PPARs) mediate the effects of various ligands, known as peroxisome proliferators, a heterogeneous class of compounds including industrial chemicals, pharmaceuticals, and biomolecules such as fatty acids and eicosanoids. Among peroxisome proliferators, fibrate derivatives are considered specific ligands for PPARα, whereas eicosanoids, such as PGJ2, for PPARγ. The study aimed to clarify the relation between PPARs and apoptosis or proliferation on the same type of cells, using clofibrate as specific ligand of PPARα and PGJ2 as specific ligand of PPARγ. The cells used were human hepatocarcinoma HepG2 cells. The results showed that PPARα protein content increased in HepG2 cells treated with clofibrate, causing apoptosis in a time‐ and concentration‐dependent way, as evidenced by the citofluorimetric assay and determination of BAD, myc and protein phosphatase 2A protein content. It also emerged that PPARγ increased in the same cells when treated with a specific ligand of this PPAR; in this case the increase of PPARγ did not cause an increase of apoptosis, but a time‐ and concentration‐dependent inhibition of cell proliferation, evidenced by decreased cell numbers and increased number of cells in the G0/G1 phase of the cycle. It may be concluded that PPARα is chiefly related to apoptosis and PPARγ to cell proliferation. Copyright © 2010 John Wiley & Sons, Ltd.     

Docosahexaenoic acid suppresses the activity of peroxisome proliferator-activated receptors in a colon tumor cell line

Fatty acids are generally considered as agonists for peroxisome proliferator-activated receptors (PPARs). Fatty acids have been shown to bind to and transactivate PPARs; it is not known whether fatty acids act as generalized agonists for PPARs in different cell types, and thus, stimulate the expression of PPAR-regulated target genes. Here, we investigated the potency of unsaturated fatty acids on transactivation of PPRE, DNA-binding activity of PPARs, and the expression of a PPAR-regulated gene product, CD36. Docosahexaenoic acid (DHA) suppressed the basal and PPAR agonist-induced transactivation of PPRE, and DNA binding of PPARs in colon tumor cells (HCT116). The suppression of PPAR transactivation by DHA leads to reduced expression of CD36 in HCT116 cells and human monocytic cells (THP-1) as determined by promoter reporter gene assay and flow cytometric analysis. Our results demonstrate that DHA and other unsaturated fatty acids act as antagonists instead of agonists for transactivation of PPRE and PPAR-regulated gene expression in the cell lines tested. These results suggest that PPAR-mediated gene expression and cellular responses can be dynamically modulated by different types of dietary fatty acids consumed.     

The influence of peroxisome proliferator-activated receptor γ(1) during differentiation of mouse embryonic stem cells to neural cells

Peroxisome proliferator activated receptor γ, belongs to PPARs, which exerts various metabolic functions including differentiation process. To testify the importance of PPARγ in neural differentiation of mouse embryonic stem cells (mESCs), its expression level was assessed. Data revealed an elevation in expression level of PPARγ when neural precursors (NPs) are formed upon retinoic acid treatment. Thus, involvement of PPARγ in two stages of neural differentiation of mESCs, during and post-NPs formation was examined by application of its agonist and antagonist. Our results indicated that PPARγ inactivation via treatment with GW9662 during NPs formation, reduced expression of neural precursor and neural (neuronal and astrocytes) markers. However, PPARγ inactivation by antagonist treatment post-NPs formation stage only decreased the expression of mature astrocyte marker (Gfap) suggesting that inactivation of PPARγ by antagonist decreased astrocyte differentiation. Here, we have demonstrated the stage dependent role of PPARγ modulation on neural differentiation of mESCs by retinoic acid treatment for the first time.     

Aluminum affects membrane physical properties in human neuroblastoma (IMR-32) cells both before and after differentiation

The capacity of Al(3+) to induce changes in the physical properties of plasma membrane from human neuroblastoma cells (IMR-32) was investigated, and the magnitude of the changes was compared with that obtained after cell differentiation to a neuronal phenotype. Similarly to our previous results in liposomes, Al(3+) (10 to 100 microM) caused a significant loss of membrane fluidity, being the differentiated cells more affected than the nondifferentiated cells. Al(3+) also increased the relative content of lipids in gel phase and promoted lipid rearrangement through lateral phase separation, with the magnitude of this effect being similar in nondifferentiated and differentiated cells. Since membrane physical properties depend on bilayer composition, we characterized the content of proteins, phospholipids, cholesterol, and fatty acids in the IMR-32 cells before and after differentiation. Differentiated cells had a significantly higher content of unsaturated fatty acids, creating an environment that favors Al(3+)-mediated effects on the bilayer fluidity. The neurotoxic effects of Al(3+) may be, at least in part, due to alterations of neuronal membrane physical properties, with potential consequences on the normal functioning of membrane-related cellular processes.     

Activation of autophagy and PPARγ protect colon cancer cells against apoptosis induced by interactive effects of butyrate and DHA in a cell type-dependent manner: The role of cell differentiation

The short-chain and n-3 polyunsaturated fatty acids exhibit anticancer properties, and they may mutually interact within the colon. However, the molecular mechanisms of their action in colon cancer cells are still not fully understood. Our study focused on the mechanisms responsible for the diverse effects of sodium butyrate (NaBt), in particular when interacting with docosahexaenoic acid (DHA), in distinct colon cancer cell types, in which NaBt either induces cell differentiation or activates programmed cell death involving mitochondrial pathway. NaBt activated autophagy both in HT-29 cells, which are sensitive to induction of differentiation, and in nondifferentiating HCT-116 cells. However, autophagy supported cell survival only in HT-29 cells. Combination of NaBt with DHA-promoted cell death, especially in HCT-116 cells and after longer time intervals. The inhibition of autophagy both attenuated differentiation and enhanced apoptosis in HT-29 cells treated with NaBt and DHA, but it had no effect in HCT-116 cells. NaBt, especially in combination with DHA, activated PPARγ in both cell types. PPARγ silencing decreased differentiation and increased apoptosis only in HT-29 cells, therefore we verified the role of caspases in apoptosis, differentiation and also PPARγ activity using a pan-caspase inhibitor. In summary, our data suggest that diverse responses of colon cancer cells to fatty acids may rely on their sensitivity to differentiation, which may in turn depend on distinct engagement of autophagy, caspases and PPARγ. These results contribute to understanding of mechanisms underlying differential effects of NaBt, when interacting with other dietary fatty acids, in colon cancer cells.     

The metabolic syndrome and the hepatic fatty acid drainage hypothesis

Much data indicates that lowering of plasma triglyceride levels by hypolipidemic agents is caused by a shift in the liver metabolism towards activation of peroxisome proliferator activated receptor (PPAR)alpha-regulated fatty acid catabolism in mitochondria. Feeding rats with lipid lowering agents leads to hypolipidemia, possibly by increased channeling of fatty acids to mitochondrial fatty acid oxidation at the expense of triglyceride synthesis. Our hypothesis is that increased hepatic fatty acid oxidation and ketogenesis drain fatty acids from blood and extrahepatic tissues and that this contributes significantly to the beneficial effects on fat mass accumulation and improved peripheral insulin sensitivity. To investigate this theory we employ modified fatty acids that change the plasma profile from atherogenic to cardioprotective. One of these novel agents, tetradecylthioacetic acid (TTA), is of particular interest due to its beneficial effects on lipid transport and utilization. These hypolipidemic effects are associated with increased fatty acid oxidation and altered energy state parameters of the liver. Experiments in PPAR alpha-null mice have demonstrated that the effects hypolipidemic of TTA cannot be explained by altered PPAR alpha regulation alone. TTA also activates the other PPARs (e.g., PPAR delta) and this might compensate for deficiency of PPAR alpha. Altogether, TTA-mediated clearance of blood triglycerides may result from a lowered level of apo C-III, with a subsequently induction of hepatic lipoprotein lipase activity and (re)uptake of fatty acids from very low density lipoprotein (VLDL). This is associated with an increased hepatic capacity for fatty acid oxidation, causing drainage of fatty acids from the blood stream. This can ultimately be linked to hypolipidemia, anti-adiposity, and improved insulin sensitivity.     

Activation of protein kinase A and exchange protein directly activated by cAMP promotes adipocyte differentiation of human mesenchymal stem cells

Human mesenchymal stem cells are primary multipotent cells capable of differentiating into several cell types including adipocytes when cultured under defined in vitro conditions. In the present study we investigated the role of cAMP signaling and its downstream effectors, protein kinase A (PKA) and exchange protein directly activated by cAMP (Epac) in adipocyte conversion of human mesenchymal stem cells derived from adipose tissue (hMADS). We show that cAMP signaling involving the simultaneous activation of both PKA- and Epac-dependent signaling is critical for this process even in the presence of the strong adipogenic inducers insulin, dexamethasone, and rosiglitazone, thereby clearly distinguishing the hMADS cells from murine preadipocytes cell lines, where rosiglitazone together with dexamethasone and insulin strongly promotes adipocyte differentiation. We further show that prostaglandin I(2) (PGI(2)) may fully substitute for the cAMP-elevating agent isobutylmethylxanthine (IBMX). Moreover, selective activation of Epac-dependent signaling promoted adipocyte differentiation when the Rho-associated kinase (ROCK) was inhibited. Unlike the case for murine preadipocytes cell lines, long-chain fatty acids, like arachidonic acid, did not promote adipocyte differentiation of hMADS cells in the absence of a PPARγ agonist. However, prolonged treatment with the synthetic PPARδ agonist L165041 promoted adipocyte differentiation of hMADS cells in the presence of IBMX. Taken together our results emphasize the need for cAMP signaling in concert with treatment with a PPARγ or PPARδ agonist to secure efficient adipocyte differentiation of human hMADS mesenchymal stem cells.     

Peroxisome proliferator-activated receptor agonist regulation of glial activation: relevance to CNS inflammatory disorders

Peroxisome proliferator-activated receptors (PPARs) play key roles in lipid metabolism and inflammation. Recent studies indicated that PPARs are also capable of modulating immune responses. Microglia and astrocytes are cells resident to the central nervous system (CNS) that function to protect against environmental insults including pathogens. However, following CNS inflammation, reactive gliosis occurs which is characterized by astrocyte hypertrophy and increased glial proliferation. Under such conditions, glia can become chronically activated and may contribute to the neuropathology associated with a variety of neuroinflammatory disorders including multiple sclerosis (MS), Alzheimer's disease (AD), Parkinson's disease (PD), amyotrophic lateral sclerosis (ALS), and stroke. A review of the role of PPAR agonists in modulating glial cell activation is presented. Included is a discussion of the molecular mechanisms of action of these PPAR agonists and the potential utility of these agents for the treatment of neuroinflammatory disorders.     

Fatty acid transduction of nitric oxide signaling: multiple nitrated unsaturated fatty acid derivatives exist in human blood and urine and serve as endogenous peroxisome proliferator-activated receptor ligands

Mass spectrometric analysis of human plasma and urine revealed abundant nitrated derivatives of all principal unsaturated fatty acids. Nitrated palmitoleic, oleic, linoleic, linolenic, arachidonic and eicosapentaenoic acids were detected in concert with their nitrohydroxy derivatives. Two nitroalkene derivatives of the most prevalent fatty acid, oleic acid, were synthesized (9- and 10-nitro-9-cis-octadecenoic acid; OA-NO2), structurally characterized and determined to be identical to OA-NO2 found in plasma, red cells, and urine of healthy humans. These regioisomers of OA-NO2 were quantified in clinical samples using 13C isotope dilution. Plasma free and esterified OA-NO2 concentrations were 619 +/- 52 and 302 +/- 369 nm, respectively, and packed red blood cell free and esterified OA-NO2 was 59 +/- 11 and 155 +/- 65 nm. The OA-NO2 concentration of blood is approximately 50% greater than that of nitrated linoleic acid, with the combined free and esterified blood levels of these two fatty acid derivatives exceeding 1 microm. OA-NO2 is a potent ligand for peroxisome proliferator activated receptors at physiological concentrations. CV-1 cells co-transfected with the luciferase gene under peroxisome proliferator-activated receptor (PPAR) response element regulation, in concert with PPARgamma, PPARalpha, or PPARdelta expression plasmids, showed dose-dependent activation of all PPARs by OA-NO2. PPARgamma showed the greatest response, with significant activation at 100 nm, while PPARalpha and PPARdelta were activated at approximately 300 nm OA-NO2. OA-NO2 also induced PPAR gamma-dependent adipogenesis and deoxyglucose uptake in 3T3-L1 preadipocytes at a potency exceeding nitrolinoleic acid and rivaling synthetic thiazo-lidinediones. These data reveal that nitrated fatty acids comprise a class of nitric oxide-derived, receptor-dependent, cell signaling mediators that act within physiological concentration ranges.     

Lipid-binding proteins modulate ligand-dependent trans-activation by peroxisome proliferator-activated receptors and localize to the nucleus as well as the cytoplasm

Peroxisome proliferator-activated receptors (PPARs) are activated by a variety of fatty acids, eicosanoids, and hypolipidemic and insulin-sensitizing drugs. Many of these compounds bind avidly to members of a family of small lipid-binding proteins, the fatty acid-binding proteins (FABPs). Fatty acids are activated to CoA esters, which bind with high affinity to the acyl-CoA-binding protein (ACBP). Thus, the availability of known and potential PPAR ligands may be regulated by lipid-binding proteins. In this report we show by transient transfection of CV-1 cells that coexpression of ACBP and adipocyte lipid-binding protein (ALBP) exerts a ligand- and PPAR subtype-specific attenuation of PPAR-mediated trans-activation, suggesting that lipid-binding proteins, when expressed at high levels, may function as negative regulators of PPAR activation by certain ligands. Expression of ACBP, ALBP, and keratinocyte lipid-binding protein (KLBP) is induced during adipocyte differentiation, a process during which PPARgamma plays a prominent role. We present evidence that endogenous ACBP, ALBP, and KLBP not only localize to the cytoplasm but also exhibit a prominent nuclear localization in 3T3-L1 adipocytes. In addition, forced expression of ACBP, ALBP, and KLBP in CV-1 cells resulted in a substantial accumulation of all three proteins in the nucleus. These results suggest that lipid-binding proteins, contrary to the general assumption, may exert their action in the nucleus as well as in the cytoplasm.