摘要
脂肪细胞中贮存的甘油三酯的水解是人和动物体内精细调节的动态平衡过程，与维持机体能量的动态平衡和代谢健康密切相关。Perilipin A是成熟脂肪细胞中脂滴膜上表达量较多的一种结构和功能蛋白，是多条脂解通路的终末靶点之一，对脂肪细胞的脂肪分解起着关键的调控作用。激素和细胞因子是影响脂肪细胞脂肪分解的重要因素，其中慢性高剂量胰岛素和白细胞介素(Interleukin，IL)-6均能够显著刺激人类及啮齿类动物脂肪细胞的脂肪分解，但在此过程中perilipinA的调控作用及其分子机制目前尚不清楚。猪的体脂沉积模式与啮齿类动物存在显著差异，却与人类更为接近，故可作为研究人类肥胖及相关疾病的理想动物模型。然而慢性高剂量胰岛素和IL-6对猪脂肪细胞脂肪分解的影响及其分子机制目前尚未见报导。
本研究以1—7日龄仔猪为试验动物，采用胶原酶消化法获得原代猪前体脂肪细胞并诱导分化。用不同浓度的胰岛素和IL-6处理分化的猪脂肪细胞适当的时间，通过测定培养液中甘油和游离脂肪酸的释放量来检测脂解率；采用形态学观察检测脂肪细胞中甘油三酯积聚量的变化；Western Blot检测perilipin A的蛋白表达；RT-PCR检测Perilipin A、PPARγ2 (Peroxisome proliferator-activated receptor-gamma2)、HSL (hormone sensitivelipase)、ATGL (adipose triglyceride lipase)、TNF-α(tumor necrosis factor-α)、IL-6、PGC-1α(peroxisome proliferators activated receptor gamma coactivator-1α)、CPT-1(carnitinepalmitoyl-transferase-1)和UCP2(uncoupling protein-2)等基因的mRNA表达；利用PKA(protein kinase A)、PKC(protein kinase C)脂解通路的抑制剂干预这两种因素刺激的脂肪分解，探讨所涉及的脂解通路及perilipinA在这一过程中的表达变化；利用罗格列酮(rosiglitazone)干预IL-6刺激的脂肪分解，研究perilipin A、PPARγ2、IL-6、PGC-1α、FAS(fatty acid synthase)等基因在这一过程中的表达变化，最后综合分析perilipin A调控慢性高剂量胰岛素和IL-6刺激的猪脂肪细胞脂肪分解的分子机制，取得以下主要结果：
1．慢性高剂量胰岛素能够以时间和剂量依赖性的方式促进猪脂肪细胞的脂肪分解，同降低脂肪细胞对异丙肾上腺素刺激的脂解脂解应答能力；在这一过程中perilipin A和PPARγ的mRNA和蛋白表达水平显著下降，TNFα和IL-6的mRNA表达水平显著上调，而ATGL和HSL的mRNA表达水平无显著变化。
2．PKA抑制剂H89和ERK(extracellular signal-related kinase)抑制剂PD98059、U0126均能够在一定程度上显著抑制慢性高剂量胰岛素刺激的脂肪分解；联合使用高剂量的PKA和ERK抑制剂几乎完全抑制了胰岛素刺激的脂肪分解；并且PD98059能够抑制胰岛素对perilipin A mRNA表达水平的下调作用，而H89却未表现出这种作用。
3．IL-6受体系统基因IL-6R和gpl30在猪脂肪细胞中表达；慢性高剂量IL-6显著促进猪脂肪细胞的脂肪分解，同时使perilipin A和PPARγ2的mRNA和蛋白表达水平显著降低，但HSL的蛋白表达并未发生显著变化；另外脂肪酸氧化相关基因PGC-1α、CPT-1和UCP2的mRNA表达也显著升高。
4．IL-6(100 ng／mL)不改变脂肪细胞内cAMP水平，PKA抑制剂H89对IL-6刺激的脂肪分解也没有显著影响；IL-6(100 ng／mL)显著促进脂肪细胞中ERKl／2的磷酸化，ERK抑制剂PD98059能够显著抑制IL-6刺激的脂肪分解，同时阻滞IL-6对perilipin AmRNA表达水平的下调作用。
5．罗格列酮能够显著抑制IL-6刺激的脂肪分解，并增强IL-6预处理的脂肪细胞对异丙肾上腺素刺激的脂解应答；在这一过程中，perilipin A、PPARγ2、FAS和PGC-1α的基因表达显著上调，但IL-6的mRNA表达水平显著下调。
综上所述，慢性高剂量胰岛素显著刺激猪脂肪细胞的脂肪分解，同时使perilipin A和PPARγ的mRNA和蛋白表达水平显著下调，并使TNFα和IL-6的mRNA表达水平显著上调。cAMP／PKA通路和MAPK／ERK通路同时参与了慢性高剂量胰岛素刺激的脂肪分解，但仅MAPK／ERK通路介导了perilipin A表达水平的下降。慢性高剂量IL-6通过抑制perilipin A、促进PGC-1α等基因的表达直接刺激猪脂肪细胞的脂肪分解。这一脂解过程主要由MAPK／ERK信号通路介导，而PKA介导的脂解通路并未参与。罗格列酮通过上调perilipinA和PGC-1α、下调IL-6的表达抑制IL-6刺激的脂肪分解。
The enzymatic hydrolysis of stored neutral lipid in adipocytes is an exquisitely regulatedprocess that maintains whole body energy homeostasis and is vital to metabolic health.Perilipin A is a important structural and functional protein,which locates on the surface oflipid droplets in adipocytes and acts as one of terminal targets among several lipolyticpathway,so it plays a key regulatory role during adipocytes lipolysis.Hormones andcytokines are important factors affecting lipolysis.However,up to now,the regulatorymechanisms of perilipin A during chronic high insulin and IL-6 stimulate lipolysis remainsunclear.Moreover,Porcine has more abundant body fat than rodents,and has many similarphysiological characters to human.Thus,porcine is regarded as an important animal modelfor researching human obesity and metabolic diseases.While at present,there are no reportsabout perilipin A regulating porcine lipolysis,and the effect of chronic high insulin and IL-6level on porcine adipocytes lipolysis also remains unclear.
In this study,porcine preadipocytes were isolated from 1-7 days old piglets and culturedto induce differentiation.Then the differentiated porcine adipocytes were incubated withdifferent insulin and IL-6 concentrations for various times(24h or 48h).After that,theconcentration of glycerol and free fatty acids(FFAs)in the media was measured as anindicator of the lipolysis and FFA consumption.Lipid accumulation morphology wasvisualized by light microscopy.Further,gene expression ofperilipin A,PPARγ2,HSL,ATGL,TNFα,IL-6,PGC-1α,CPT-1 and UCP2 were determined with semi-quantitative RT-PCR andperilipin A protein expression was detected by western blot.Meanwhile,PKA and ERKinhibitors were used to determine the involved lipolytic pathway,and rosiglitazone was usedto analyze the regulatory role of perilipin A in IL-6 treatment lipolysis.The results are as
follows:
1.Chronic high insulin dose significantly promoted lipolysis in porcine adipocytes.Theexpression of perilipin A protein were significantly reduced as well as the mRNA expressionof perilipin A and PPARγ2,however,the mRNA expression of TNFαand IL-6 wassignificantly increased.
2.The inhibitors of both PKA(H89)and ERK(PD98059)could repress the lipolysisstimulated by chronic high insulin concentration partially,and the combination of them couldrepress the lipolysis completely.Moreover,PD98059 could inhibit the down regulation ofperilipin A induced by insulin,but H89 did not present this kind of role.
3.IL-6 receptor system was expressed in porcine adipocytes.Chronic high IL-6 dosesignificantly promoted lipolysis in porcine adipocytes.Furthermore,the mRNA expression ofperilipin A and PPARγ2 were significantly reduced,but the expression of PGC-1α,CPT-1 andUCP2 mRNA was significantly increased.
4.The inhibitor of PKA(H89)didn't affect the lipolysis stimulated by IL-6(100ng/mL).However,the inhibitor of ERK(PD98059)had significant inhibitory role in this kind oflipolysis,and the down regulation ofperilipin A induced by IL-6 was significantly repressed.
5.Rosiglitazone significantly repressed the lipolysis stimulated by IL-6 in porcine adipocytes,and lipolytic response of the adipocytes to isoprenaline was enhanced.Meanwhile,rosiglitazone significantly increased the mRNA expression of perilipin A,PPARγ2,FAS andPGC-1α的mRNA,but reduced the expression oflL-6 mRNA.
Above all,Chronic high insulin dose significantly promoted lipolysis in porcineadipocytes,and decreased the expression of perilipin A protein and mRNA,this process couldbe associated with the up regulation of IL-6 and TNFα.Both PKA and ERK pathwaymediated the lipolysis stimulated by chronic high insulin dose,but the down regulation ofperilipin A expression was only involved in ERK pathway,rather than PKA pathway,so weinfered that the means of PKA lipolytic pathway during insulin treatment lipolysis wasinvolved in the phosphorylation of perilipin A and activation of HSL and ATGL.Chronichigh IL-6 dose directly stimulated lipolysis in porcine adipocytes by repressing perilipin Aand promoting gene expression of PGC-1α,CPT-1 and UCP2.This process was onlymediated by ERK pathway.Rosiglitazone repressed the lipolysis stimulated by IL-6 inporcine adipocytes by increasing perilipin A expression,which maybe related to the upregulation of PPARγ2 and FAS as well as the down regulation of IL-6.
引文
[1]Koubi HE,Desplanches D,Gabrielle C,et al.Exercise endurance and fuel utilization:a reevaluation of the effects of fasting[J].J.Appl.Physiol.,1991,70(3):1337-1343.
[2]Klein S,Young VR,Blackburn GL,et al.The impact of body composition on the regulation of lipolysis during short-term fasting[J].J.Am.Coll.Nutr.,1988,7(1):77-84.
[3]Boss O,Hagen T,Lowell BB.Uncoupling proteins 2 and 3:potential regulators of mitochondrial energy metabolism[J].Diabetes,2000,49(2):143-156.
[4]Jazet IM,Pijl H,Meinders AE.Adipose tissue as an endocrine organ:impact on insulin resistance[J]. Neth.J.Med.,2003,61(6):194-212.
[5]Kern PA,Ranganathan S,Li C.Adipose tissue tumor necrosis factor and interleukin-6 expression in human obesity and insulin resistance[J].Am J.Physiol.Endocrinol.Metab.,2001,280:E745-E751.
[6]Stouthard JM,Romijn JA,Vander Poll T.Endocrinologic and metabolic effects of interleukin-6 in humans[J].Am.J.Physiol.,1995,268(5 Pt 1):E813-E819.
[7]Bastard JP,Jardel C,Bruckert E.Elevated levels of interleukin-6 are reduced in serum and subcutaneous adipose tissue of obese women after weight loss[J].J.Clin.Endocrinol.Metab.,2000, 85:3338-3342.
[8]Miyoshi H,Souza SC,Zhang HH,et al.Perilipin promotes hormone-sensitive lipase-mediated adipocyte lipolysis via phosphorylation-dependent and-independent mechanisms[J].J.Biol.Chem., 2006,281(23):15837-15844.
[9]Miyoshi H,2nd Perfield JW,Souza SC,et al.Control of adipose triglyceride lipase action by serine 517 of perilipin A globally regulates protein kinase A-stimulated lipolysis in adipocytes[J].J.Biol. Chem.,2007,282(2):996-1002.
[10]Subramanian V,Rothenberg A,Gomez C,et al.Perilipin A Mediates the Reversible Binding of CGI-58 to Lipid Droplets in 3T3-L1 Adipocytes[J].J.Biol.Chem.,2004,279(40):42062-42071.
[11]Lass A,Zimmermann R,Haemmerle G,et al.Adipose triglyceride lipase-mediated lipolysis of cellular fat stores is activated by CGI-58 and defective in Chanarin-Dorfman Syndrome[J].Cell Metab.,2006, 3(5):309-319.
[12]Chung S,Brown JM,Provo JN,et al.Conjugated Linoleic Acid Promotes Human Adipocyte Insulin Resistance through NFκB-dependent Cytokine Production[J].The Journal of Biological Chemistry, 2005,280(46):38445-38456.
[13]Pages G,Lenormand P,L'Allemain G,et al.Mitogen-activated protein kinases p42mapk and p44mapk are required for fibroblast proliferation[J].Proc Natl Acad Sci USA,1993,90:8319-8323.
[14]Bost F,Caron L,Marchetti I,et al.Retinoic acid activation of the ERK pathway is required for embryonic stem cell commitment into the adipocyte lineage[J].Biochem J,2002,361:621-627.
[15]Kanda Y,Nishio E,Watanabe Y.Differential regulation of Na+/H+exchange and DNA synthesis in vascular smooth muscle cells[J].Eur J Pharmacol.,1999,371:69-74.
[16]Greenberg AS,Shen W J,Muliro K,et al.Stimulation of lipolysis and hormone-sensitive lipase via the extracellular signal-regulated kinase pathway [J]. J Biol Chem, 2001, 276: 45456-45461.
[17] Das UN. Is obesity an inflammatory condition?[J]. Nutrition, 2001,17 (11-12): 953-966.
[18] Wellen KE, Hotamisligil GS. Inflammation, stress, and diabetes[J]. J. Clin. Invest., 2005, 115: 1111-1119.
[19] Spiegelman BM. PPAR-gamma: adipogenic regulator and thiazolidinedione receptor [J]. Diabetes, 1998,47:507-514.
[20] Hauner H. The mode of action of thiazolidinediones [J]. Diabetes Metab. Res. Rev., 2002, 18: S10-15.
[21] Mohanty P, Aljada A, Ghanim H, et al. Evidence for a potent antiinfiammatory effect of rosiglitazone [J]. J Clin Endocrinol Metab, 2004, 89: 2728-35.
[22] Beckman M. Cell biology. Great balls of fat [J]. Science, 2006, 311 (5765): 1232-1234.
[23] Langin D. Control of fatty acid and glycerol release in adipose tissue lipolysis [J]. C R Biol, 2006, 329(8): 598-607; discussion 653-655.
[24] Langin D. Adipose tissue lipolysis as a metabolic pathway to define pharmacological strategies against obesity and the metabolic syndrome [J]. Pharmacol Res, 2006, 53 (6): 482-491.
[25] Boden G. Role of fatty acids in the pathogenesis of insulin resistance and NIDDM [J]. Diabetes, 1997, 46(1): 3-10.
[26] Frayn KN. Adipose tissue and the insulin resistance syndrome [J]. Proc Nutr Soc, 2001, 60 (3): 375-380.
[27] Langin D, Dicker A, Tavernier G, et al. Adipocyte lipases and defect of lipolysis in human obesity [J]. Diabetes, 2005, 54 (11): 3190-3197.
[28] Holm C, Kirchgessner TG, Svenson KL, et al. Hormone-sensitive lipase: sequence, expression, and chromosomal localization to 19 cent-q13.3 [J]. Science, 1988, 241(4872): 1503-1506.
[29] Holm C. Molecular mechanisms regulating hormone-sensitive lipase and lipolysis [J]. Biochem Soc Trans, 2003, 31 (Pt 6): 1120-1124.
[30] Osuga J, Ishibashi S, Oka T, et al. Targeted disruption of hormone-sensitive lipase results in male sterility and adipocyte hypertrophy, but not in obesity [J]. Proc Natl Acad Sci U S A, 2000, 97 (2): 787-792.
[31] Haemmerle G, Zimmermann R, Strauss JG, et al. Hormone-sensitive lipase deficiency in mice changes the plasma lipid profile by affecting the tissue-specific expression pattern of lipoprotein lipase in adipose tissue and muscle [J]. J Biol Chem, 2002, 277 (15): 12946-52.
[32] Yeaman SJ. Hormone-sensitive lipase-a multipurpose enzyme in lipid metabolism [J]. Biochim BiophysActa, 1990, 1052(1): 128-132.
[33] Kraemer FB, Shen WJ. Hormone-sensitive lipase: control of intracellular tri-(di-) acylglycerol and cholesteryl ester hydrolysis [J]. J Lipid Res, 2002, 43 (10): 1585-1594.
[34] Greenberg AS, Shen WJ, Muliro K, et al. Stimulation of lipolysis and hormone-sensitive lipase via the extracellular signal-regulated kinase pathway [J]. J Biol Chem, 2001, 276 (48): 45456-45461.
[35] Smih F, Rouet P, Lucas S, et al. Transcriptional regulation of adipocyte hormone-sensitive lipase by glucose [J]. Diabetes, 2002, 51 (2): 293-300.
[36] Berraondo B, Martinez JA. Free fatty acids are involved in the inverse relationship between hormone-sensitive lipase (HSL) activity and expression in adipose tissue after high-fat feeding or beta3-adrenergic stimulation [J]. Obes Res, 2000, 8 (3): 255-261.
[37] Zimmermann R, Strauss JG, Haemmerle G, et al. Fat mobilization in adipose tissue is promoted by adipose triglyceride lipase [J]. Science, 2004,306 (5700): 1383-1386.
[38] Jenkins CM, Mancuso DJ, Yan W, et al. Identification, cloning, expression, and purification of three novel human calcium-independent phospholipase A2 family members possessing triacylglycerol lipase and acylglycerol transacylase activities [J]. J Biol Chem, 2004,279 (47): 48968-48975.
[39] Villena J, Roy S, Sarkadi-Nagy E, et al. Desnutrin, an adipocyte gene encoding a novel patatin domain-containing protein, is induced by fasting and glucocorticoids: ectopic expression of desnutrin increases triglyceride hydrolysis [J]. J Biol Chem, 2004,279 (45): 47066-47075.
[40] Ali YB, Carriere F, Verger R, et al. Continuous monitoring of cholesterol oleate hydrolysis by hormone-sensitive lipase and other cholesterol esterases [J]. J Lipid Res, 2005,46 (5): 994-1000.
[41] Smirnova E, Goldberg EB, Makarova KS, et al. ATGL has a key role in lipid droplet/adiposome degradation in mammalian cells [J]. EMBO Rep, 2006, 7(1): 106-113.
[42] Haemmerle G, Lass A, Zimmermann R, et al. Defective lipolysis and altered energy metabolism in mice lacking adipose triglyceride lipase [J]. Science, 2006, 312 (5774): 734-737.
[43] Zechner R, Strauss JG, Haemmerle G, et al. Lipolysis: pathway under construction [J]. Curr Opin Lipidol, 2005, 16 (3): 333-340.
[44] Lehner R, Verger R. Purification and characterization of a porcine liver microsomal triacylglycerol hydrolase [J]. Biochemistry, 1997, 36 (7): 1861-1868.
[45] Lehner R, Cui Z, Vance DE. Subcellullar localization, developmental expression and characterization of a liver triacylglycerol hydrolase [J]. Biochem J, 1999, 338 (Pt 3): 761-768.
[46] Satoh T, Hosokawa M. The mammalian carboxylesterases: from molecules to functions [J]. Annu Rev Pharmacol Toxicol, 1998, 38: 257-288.
[47] Dolinsky VW, Sipione S, Lehner R, et al. The cloning and expression of a murine triacylglycerol hydrolase cDNA and the structure of its corresponding gene [J]. Biochim Biophys Acta, 2001, 1532 (3): 162-172.
[48] Matsushima M, Inoue H, Ichinose M, et al. The nucleotide and deduced amino acid sequences of porcine liver proline- beta-naphthylamidase. Evidence for the identity with carboxylesterase [J]. FEBS Lett, 1991, 293 (1-2): 37-41.
[49] Soni KG, Lehner R, Metalnikov P, et al. Carboxylesterase 3 (EC 3.1.1.1) is a major adipocyte lipase [J]. The Journal of biological chemistry, 2004, 279 (39): 40683-40689.
[50] Okazaki H, Osuga J, Tamura Y, et al. Lipolysis in the absence of hormone-sensitive lipase: evidence for a common mechanism regulating distinct Upases [J]. Diabetes, 2002, 51 (12): 3368-3375.
[51] Holm C. Molecular mechanisms regulating hormone-sensitive lipase and lipolysis [J]. Biochemical SocietyTransactions, 2003, 31: 1120-1124.
[52] Carey GB. Mechanisms regulating adipocyte lipolysis [J]. Advances in Experimental Medicine and Biology, 1998, 76: 157-170.
[53] Langin D, Lucas S, Lafontan M. Millennium fat-cell lipolysis reveals unsuspected novel tracks [J]. Hormone and Metabolic Research, 2000, 32: 443-452.
[54] Large V, Peroni O, Letexier D et al. Metabolism of lipids in human white adipocyte [J]. Diabetes and Metabolism, 2004, 30: 294-309.
[55] Bjornholm M, He AR, Attersand A et al. Absence of functional insulin receptor substrate-3 (IRS-3) gene in humans [J]. Diabetologia, 2002, 45: 1697-1702.
[56] Degerman E, Resjo S, Landstrom TR, et al. Methods to study phosphorylation and activation of the hormone-sensitive adipocyte phosphodiesterase type 3B in rat adipocytes [J]. Methods in Molecular Biology, 2001, 155: 167-180.
[57] Zhang Jin, Hupfeld CJ, Taylor SS, et al. Insulin disrupts p-adrenergic signalling to protein kinase A in adipocytes [J]. Nature, 2005,437 (22): 569-573.
[58] Hupfeld CJ, Dalle S, Olefsky JM. P-Arrestin 1 down-regulation after insulin treatment is associated with supersensitization of β2 adrenergic receptor Gas signalling in 3T3-L1 adipocytes [J]. Proc. Natl Acad. Sci. USA, 2003, 100: 161-166.
[59] Getty-Kaushik L, Richard AT., Corkey BE. Glucose-dependent insulin modulation of oscillatory lipolysis in perifused rat adipocytes [J]. Obesity Research, 2005,13 (12): 2058-2065.
[60] Pessin JE, Saltiel AR. Signaling pathways in insulin action: molecular targets of insulin resistance [J]. J. Clin. Invest., 2000, 106: 165-169.
[61] Hutley L, Prins JB. Fat as endocrine organ: relationship to the metabolic syndrome. Am J Med Sci, 2005, 330 (6): 280-289.
[62] 周丽斌, 陈明道. 脂肪因子和代谢综合征. 中国医学科学院学报, 2006,28 (6): 840-884.
[63] Ryden M, Arvidsson E, Blomqvist L, et al. Targets for TNF-alpha-induced lipolysis in human adipocytes[J]. Biochem Biophys Res Commun, 2004, 318 (1): 168-175.
[64] Langin D, Arner P. Importance of TNFalpha and neutral Upases in human adipose tissue lipolysis [J]. Trends Endocrinol Metab, 2006, 17 (8): 314-320.
[65] Grunfeld C, Feingold KR. The metabolic effects of tumor necrosis factor and other cytokines [J]. Biotherapy, 1991, 3 (2): 143-158.
[66] Hotamisligil GS, Spiegelman BM. Tumor necrosis factor alpha: a key component of the obesity-diabetes link [J]. Diabetes, 1994, 43 (11): 1271-1278.
[67] Sethi JK, Hotamisligil GS. The role of TNF alpha in adipocyte metabolism [J]. Semin Cell Dev Biol, 1999, 10(1): 19-29.
[68] Margetic S, Gazzola C, Pegg G G, et al.Leptin: a review of its peripheral actions and interactions [J]. Int J Obes, 2002, 26: 1407-1433.
[69] Lissett C, Clayton PE, Shalet S.The acute leptin response to GH [J]. J Clin Endocrinol Metab, 2001, 86:4412-4415.
[70] Devos P, Lefebvre AM, Shrivo I, et al.Glucocorticoids induce the expression of the leptin gene through a non - classical mechanism of transcriptional activation [J]. Eur J Biochem, 1998, 253: 619 -626.
[71] Fruhbeck G, Gomez-Ambrosi J, Muruzabal FJ, et al.The adipocyte: a model for integration of endocrine and metabolic signaling in energy metabolism regulation [J]. Am J Physiol, 2001, 280: 827 -847.
[72] Fruhbeck G, Gomez-Ambrosi J, Salvador J. Leptin-induced lipolysis opposes the tonic inhibition of endogenous adenosine in white adipocytes [J]. FASEB J, 2001, 15: 333 -340.
[73] Lonnqvist F, Arner P, Nordfors L, et al. Overexpression of the obese (ob) gene in adipose tissue of human obese subjects[J]. Nat. Med., 1995, 1:950-953.
[74] Fruhbeck G, Aguado M, Martinez JA. In vitro lipolytic effect of leptin on mouse adipocytes: evidence for a possible autocrine/paracrine role of leptin[J].Biochem.Biophys.Res.Commun.,1997,240: 590-594.
[75]Siegrist-Kaiser CA,Pauli V,Juge-Aubry CE,et al.Direct effects of leptin on brown and white adipose tissue5[J].J.Clin.Invest.,1997,100:52858-52864.
[76]Fruhbeck G,Aguado M,Gomez-Ambrosi J,et al.Lipolytic effect of in vivo leptin administration on adipocytes of lean and ob/ob mice,but not db/db mice[J].Biochem.Biophys.Res.Commun.,1998, 250:99-102.
[77]Muller G,Ertl J,Gerl M,et al.Leptin impairs metabolic actions of insulin in isolated rat adipocytes[J]. J.Biol.Chem.,1997,272:10585-10593.
[78]Steensberg A.The role of IL-6 in exercise-induced immune changes and metabolism.Exerc Immunol Rev.2003,9:40-47.
[79]Petersen AMW,Pedersen BK.The anti-inflammatory effect of exercise.J Appl Physiol.,2005,98: 1154-1162.
[80]Steensberg A,van Hall G,Osada T.Production ofinterleukin-6 in contracting human skeletal muscles can account for the exercise-induced increase in plasma interleukin-6[J].J.Physiol.,2000,529: 237-242.
[81]Fruhbeck G,Gomez-Ambrosi J,Muruzabal FJ.The adipocyte:a model for integration of endocrine and metabolic signaling in energy metabolism regulation[J].Am J Physiol,2001,280:E827-E847.
[82]van Hall G,Steensberg A,Sacchetti M,et al.Interleukin-6 stimulates lipolysis and fat oxidation in humans[J].J.Clin.Endocrinol.Metab.,2003,88(7):3005-3010.
[83]Lyngso D,Simonsen L,Bulow J.Metabolic effects of interleukin-6 in human splanchnic and adipose tissue[J].J.Physiol.,2002,543:379-386.
[84]Petersen EW,Carey AL,Sacchetti M,et al.Acute IL-6 treatment increases fatty acid turnover in elderly humans in vivo and in tissue culture in vitro[J].Am.J.Physiol.Endocrinol.Metab.,2005, 288:E155-E162.
[85]P(a|¨)th G,Bornstein SR,Gurniak M,et al.Human breast adipocytes express interleukin-6(IL-6)and its receptor system:increased IL-6 production by β-adrenergic activation and effects of IL-6 on adipocyte function[J].J.Clin.Endocrinol.Metab.,2001,86(5):2281-2288.
[86]Wallenius V,Wallenius K,Ahr(?)n B,et al.Interleukin-6 deficient mice develop mature onset obesity [J].Nat Med.2002,8:75-79.
[87]Kershaw EE,Flier JS.Adipose tissue as an endocrine organ[J].J Clin Endocrinol Meta,2004,89(6): 2548-2556.
[88]Bai Y,Zhang S,Kim K-S,et al.Obese Gene Expression Alters the Ability Of 30A5 Preadipocytes to Respond to Lipogenic Hormones[J].J.Biol.Chem.,1996,271:13939-13942.
[89]Hara K,Horikoshi M,Yamauchi T,et al.Measurement of the high-molecular weight form of adiponectin in plasma is useful for the prediction of insulin resistance and metabolic syndrome[J]. Diabetes Care,2006,29(6):1357-1362.
[90]Diez J J,Iglesias P.The role of the novel adipocyte derived hormone adiponectin in human disease[J]. Eur J Endocrinol,2003,148(3):293-300.
[91]周顺伍．动物生物化学(第三版)[M]．中国北京，中国农业出版社，2000年．
[92]Carmen GY,Victor SM.Signalling mechanisms regulating lipolysis[J].Cell Signal.2006,18: 401-408.
[93] Fain JN, Garcija-Sainz JA. Adrenergic regulation of adipocyte metabolism [J]. J. Lipid Res. 1983, 24:945-966.
[94] Langin D. Control of fatty acid and glycerol release in adipose tissue lipolysis [J]. C. R. Biol., 2006, 329: 598-607; discussion 653-55.
[95] Large V, Peroni O, Letexier D, et al. Metabolism of lipids in human white adipocyte [J]. Diabetes Metab., 2004, 30: 294-309.
[96] Londos C, Honnor RC, Dhillon GS. cAMP-dependent protein kinase and lipolysis in rat adipocytes.Ⅲ. Multiple modes of insulin regulation of lipolysis and regulation of insulin responses by adenylate cyclase regulators [J]. J. Biol. Chem., 1985, 260: 15139-15145.
[97] Olsson H, Belfrage P. The regulatory and basal phosphorylation sites of hormonesensitive lipase are dephosphorylated by protein phosphatase-1, 2A and 2C but not by protein phosphatase-2B [J]. Eur. J. Biochem., 1987, 168:399-405.
[98] Ragolia L, Begum N. Protein phosphatase-1 and insulin action [J]. Mol. Cell. Biochem., 1998, 182: 49-58.
[99] Stralfors P, Honnor RC. Insulin-induced dephosphorylation of hormone-sensitive lipase. Correlation with lipolysis and cAMP-dependent protein kinase activity [J]. Eur. J. Biochem., 1989, 182: 379-385.
[100] Zhang J, Hupfeld CJ, Taylor SS, et al. Insulin disrupts beta -adrenergic signaling to protein kinase A in adipocytes [J]. Nature, 2005, 437: 569-573.
[101] Schrager S. Dietary calcium intake and obesity [J]. J. Am. Board Fam. Pract., 2005, 18: 205-210.
[102] Melanson EL, Sharp TA, Schneider J, et al. Relation between calcium intake and fat oxidation in adult humans [J]. Int. J. Obes. Relat. Metab. Disord., 2003, 27: 196-203.
[103] Murray RK, GrannerDK, MayesPA, et al. Harper's Biochemistry [M]. Stamford, CT: Appleton & Lange, 2000, 927 pp.
[104] Shi H, Dirienzo D, Zemel MB. Effects of dietary calcium on adipocyte lipid metabolism and body weight regulation in energy-restricted aP2-agouti transgenic mice [J]. FASEB J., 2001, 15: 291-293
[105] Xue B, Greenberg AG, Kraemer FB, et al. Mechanism of intracellular calcium ([Ca2+]i) inhibition of lipolysis in human adipocytes [J]. FASEB J. 2001, 15: 2527-2529.
[106] Londos C, Cooper DM, Schlegel W, et al. Adenosine analogs inhibit adipocyte adenylate cyclase by a GTP-dependent process: basis for actions of adenosine and methylxanthines on cyclic AMP production and lipolysis [J]. Proc. Natl. Acad. Sci. USA, 1978, 75: 5362-5366.
[107] Borglum JD, Vassaux G, Richelsen B, et al. Changes in adenosine Al- and A2-receptor expression during adipose cell differentiation [J]. Mol. Cell.Endocrinol., 1996, 117: 17-25.
[108] Peers DG, Davies JI. Significance of the caffeine-like effect of various purines, pyrimidines and derivatives on adipose-tissue phosphodiesterase [J]. Biochem. J., 1971, 124: P8-9.
[109] Acheson KJ, Gremaud G, Meirim I, et al. Metabolic effects of caffeine in humans: lipid oxidation or futile cycling? [J]. Am. J. Clin. Nutr., 2004, 79: 40-46.
[110] Westerterp-Plantenga MS, Lejeune MP, Kovacs EM. Body weight loss and weight maintenance in relation to habitual caffeine intake and green tea supplementation [J]. Obes. Res., 2005, 13: 1195-1
[111] Siler SQ, Neese RA, Hellerstein MK. De novo lipogenesis, lipid kinetics, and whole-body lipid balances in humans after acute alcohol consumption [J]. Am. J. Clin. Nutr., 1999, 70: 928-936.
[112] Kang L, Nagy LE. Chronic ethanol feeding suppresses beta-adrenergic receptorstimulated lipolysis in adipocytes isolated from epididymal fat [J]. Endocrinology, 2006, 147:4330-4338.
[113] McCarty MF, Thomas CA. PTH excess may promote weight gain by impeding catecholamine -induced lipolysis-implications for the impact of calcium, vitamin D, and alcohol on body weight [J]. Med. Hypotheses, 2003,61: 535-542.
[114] Feve B, Emorine LJ, Lasnier F, et al. Atypical beta-adrenergic receptor in 3T3-F442A adipocytes. Pharmacological and molecular relationship with the human beta 3- adrenergic receptor [J]. J. Biol. Chem., 1991,266:203-209.
[115] Galitzky J, Carpene C, Bousquet-Melou A, et al. Differential activation of beta 1-, beta 2- and beta 3-adrenoceptors by catecholamines in white and brown adipocytes [J]. Fundam. Clin. Pharmacol., 1995,9:324-331.
[116] Van IL, Van Witzenburg VA., Vauquelin G. Multiple beta adrenergic receptor subclasses mediate the 1- isoproterenol-induced lipolytic response in rat adipocytes [J]. J. Pharmacol. Exp. Ther., 1992,262: 552.
[117] Belfrage P, Fredrikson G, Nilsson NO, et al. Regulation of adipose tissue lipolysis: phosphorylation of hormones sensitive lipase in intact rat adipocytes [J].FEBS Lett., 1980, 111 (1): 120-124.
[118] Kawamura M, Jensen DF, Wancewicz EV, et al. Hormone-sensitive lipase in differentiated 3T3-L1 cells and its activation by cyclic AMP-dependent protein kinase [J]. Proc Natl Acad Sci U S A, 1981, 78 (2): 732-736.
[119] Schimmel RJ, Buhlinger CA, Serio R. Activation of adenosine 3',5'-monophosphate-dependent protein kinase and its relationship to cyclic AMP and lipolysis in hamster adipose tissue [J]. J. Lipid Res., 1980,21(2): 250-256.
[120] Stralfors P, Belfrage P. Phosphorylation of hormone-sensitive lipase by cyclic AMP-dependent protein kinase [J]. J. Biol. Chem., 1983, 258 (24): 15146-15152.
[121] Pergola DG. The adipose tissue metabolism: role of testosterone and dehydroe-piandrosterone [J]. Int. J. Obes. Relat. Metab. Disord., 2000,24 (Suppl 2): S59-S63.
[122] Hadri K.E, Courtalon A, Gauthereau X, et al. Differential regulation by tumor necrosis factor-alpha of beta1-, beta2-, and beta3-adrenoreceptor gene expression in 3T3-F442A adipocytes [J]. J Biol Chem, 1997, 272 (39): 24514-24521.
[123] Carel JC, Stunff CL, Condamine L, et al. Resistance to the Lipolytic Action of Epinephrine: A New Feature of Protein Gs Deficiency [J] .The Journal of Clinical Endocrinology & Metabolism, 1999, 84 (11): 4127-4131.
[124] Chaudhry A, MacKenzie RG, Georgic LM, et al. Differential interaction of β1- and β3-adrenergic receptors with Gi in rat adipocytes [J] .Cellular Signalling, 1994,6 (4): 457-465.
[125] Feinstein DL, Reis DJ, Regunathan S. Inhibition of Astroglial Nitric Oxide Synthase Type 2 Expression by Idazoxan [J]. Mol Pharmacol 1999, 55 (2): 304-308.
[126] Soeder KJ, Snedden SK, Cao W, et al. The beta3-adrenergic receptor activates mitogen-activated protein kinase in adipocytes through a Gi-dependent mechanism [J]. J Biol Chem, 1999, 274 (17): 12017-12022.
[127] Lindquist JM , Fredriksson JM, Rehnmark S, et al. beta 3- and alpha 1-Adrenergic Erk1/2 Activation Is Src- but Not Gi-mediated in Brown Adipocytes [J]. J. Biol. Chem, 2000, 275 (30): 22670-22677.
[128] Lafontan M, Barbe P, Galitzky J, et al. Adrenergic regulation of adipocyte metabolism [J]. Hum. Reprod, 1997, 12 (Suppl 1): 6-20.
[129] Schwabe U, Ebert R, Erbler HC. Adenosine release from fat cells: effect on cyclic AMP levels and hormone actions [J]. Adv Cyclic Nucleotide Res., 1975, 5: 569-584.
[130] Gasic S, Tian B, Green A.Tumor necrosis factor alpha stimulates lipolysis in adipocytes by decreasing Gi protein concentrations [J]. J Biol Chem, 1999, 274 (10): 6770-6775.
[131] Manganiello VC, Murata T, Taira M, et al. Perspectives in biochemistry and biophysics. Diversity in cyclic nucleotide phosphodiesterase isoenzyme families [J]. Arch Biochem Biophys, 1995, 322 (1): 1-13.
[132] Degerman E, Belfrage P, Manganiello VC. Structure,localization,and regulation of cGMP-inhibited phosphodiesterase (PDE3) [J]. J Biol Chem, 1997, 272 (11): 6823-6826.
[133] Degerman E, Landstr TR, Wijkander J, et al. Phosphorylation and activation of hormone-sensitive adipocyte phosphodiesterase type 3B [J]. Methods., 1998, 14 (1): 43-53.
[134] Goswami A Rosenberg IN. Thyroid hormone modulation of epinephrine-induced lipolysis in rat adipocytes: a possible role of calcium [J]. Endocrinology, 1978, 103: 2223-2233.
[135] Van Inwegen RG, Robison GA, Thompson WJ. Cyclic nucleotide phosphodiesterases and thyroid hormones [J]. J. Biol. Chem, 1975, 250 (7): 2452-2456.
[136] Rahn Landstrom T, Mei J, Karlsson M, et al. Down-regulation of cyclic-nucleotide phosphodiesterase 3B in 3T3-L1 adipocytes induced by tumour necrosis factor alpha and cAMP [J]. Biochem J. 2000,346 (Pt 2): 337-343.
[137] Xue B, Greenberg AG, Kraemer FB, et al. Mechanism of intracellular calcium ([Ca2+]i) inhibition of lipolysis in human adipocytes [J]. FASEB J, 2001, 15 (13): 2527-2529.
[138] Jimenez M, Leger B, Canola K, et al. Beta(1)/beta(2)/beta(3)-adrenoceptor knockout mice are obese and cold-sensitive but have normal lipolytic responses to fasting [J]. FEBS Lett, 2002, 530 (1-3): 37-40.
[139] Flechtner-Mors M, Jenkinson CP, Alt A, et al. In Vivo 1-Adrenergic Lipolytic Activity in Subcutaneous Adipose Tissue of Obese Subjects [J]. J. Pharmacol. Exp. Ther, 2002, 301(1): 229-233.
[140] Arner P. Adrenergic receptor function in fat cells [J]. Am J Clin Nutr, 1992, 55 (1 Suppl): 228S-236S.
[141] Galvin-Parton PA, Chen X, Moxham CM, et al. Induction of Galpha q-specific Antisense RNA in Vivo Causes Increased Body Mass and Hyperadiposity [J]. J. Biol. Chem, 1997, 272 (7): 4335-4341.
[142] Sanchez-Margalet V, Gonzalez-Yanes C. Pancreastatin inhibits insulin action in rat adipocytes [J]. Am J Physiol Endocrinol Metab, 1998,275(6 Pt 1): El055-1060.
[143] Gonzalez-Yanes C, Sanchez-Margalet V. Pancreastatin modulates insulin signaling in rat adipocytes: mechanisms of cross-talk [J]. Diabetes, 2000,49 (8): 1288-1294.
[144] Naghshineh S, Noguchi M, Huang KP, et al. Activation of adipocyte adenylate cyclase by protein kinase C[J]. J. Biol. Chem, 1986, 261(31): 14534-14538.
[145]Fricke K,Heitland A,Maronde E.Cooperative activation of lipolysis by protein kinase A and protein kinase C pathways in 3T3-L1 adipocytes[J].Endocrinology,2004,145(11):4940-4947.
[146]Romanelli A,Werve GV.Activation of mitogen-,activated protein kinase in freshly isolated rat hepatocytes by both a.calcium-and a protein kinase C-dependent pathway[J].Metabolism,1997, 46(5):548-555.
[147]Lindquist JM.,Fredriksson JM,Rehnmark S,et al.Beta 3-and alphal-adrenergic Erk1/2 activation is Src-but not Gi-mediated in Brown adipocytes[J].J.Biol.Chem.,2000,275(30):22670-22677.
[148]Huang SH,Shen WJ,Yeo HL.Signaling pathway of magnolol-stimulated lipolysis in sterol ester-loaded 3T3-L1 preadipocytes[J].J Cell Biochem,2004,91(5):1021-1029.
[149]Franklin RA,Atherfold PA,McCubrey JA.Calcium-induced ERK activation in human T lymphocytes occurs via p56(Lck)and CaM-kinase[J].Mol Immunol.,2000,37(11):675-683.
[150]Soderling TR.The Ca-calmodulin-dependent protein kinase cascade[J].Trends Biochem.Sci,1999, 24(6):232-236.
[151]Sengen(?)s C,Berlan M,Glisezinski I,et al.Natriuretic peptides:a new lipolytic pathway in human adipocytes[J].FASEB J.,2000,14:1345-1351.
[152]Sengenes C,Zakaroff-Girard A,Moulin A,et al.Natriuretic peptide-dependent lipolysis in fat cells is a primate specificity[J].Am J Physiol Regul Integr Comp Physiol.,2002,283:R257-265.
[153]Egan J J,Greenberg AS,Chang MK,et al.Mechanism of hormone-stimulated lipolysis in adipocytes: Translocation of hormone-sensitive lipase to the lipid storage droplet[J].Proc.Natl.Acad.Sci.U.S. A.,1992,89:8537-8541.
[154]Smolenski A,Burkhardt AM,Eigenthaler,M,et al.Functional analysis of cGMP-dependent protein kinases I and II as mediators of NO/cGMP effects[J].Naunyn-Schmiedeberg's Arch.Pharmacol., 1998,358:134-139.
[155]Galitzky J,Sengenes C,Thalamas C,et al.The lipid-mobilizing effect of atrial natriuretic peptide is unrelated to sympathetic nervous system activation or obesity in young men[J].J Lipid Res.,2001, 42:5365-44.
[156]Moro C,Galitzky J,Sengenes C,et al.Functional and pharmacological characterization of the natriuretic peptide-dependent lipolytic pathway in human fat cells[J].J Pharmacol Exp Ther.,2004, 308:984-992.
[157]Sengenes C,Bouloumie A,Hauner H,et al.Involvement of a cGMP-dependent pathway in the natriuretic peptide-mediated hormone-sensitive lipase phosphorylation in human adipocytes[J].J Biol Chem.,2003,278:48617-48626.
[158]Moro C,Crampes F,Sengenes C,et al.Atrial natriuretic peptide contributes to physiological control of lipid mobilization in humans[J].FASEB J.,2004,18:908-910.
[159]Bolen B J,Brugge JS.Leukocyte protein tyrosine kinases:Potentail targets for drug discovery[J]. Annu Rev Immunol,1997,15:371-404.
[160]Pellegrini S,Dusanter-FI.The structure,regulation and function of the Janus kinase(JAKs)and the signal transducers and activators of transcriptions(STATs)[J].Eur J Biochem,1997,248:615-633.
[161]Ihle JN.STATs:Signal transducers and activators of transcription[J].Cell,1996,84:331-334.
[162]Bjorbaek C,Kahn BB.Leptin signaling in the central nervous system and the periphery[J].Recent Prog Horm Res.,2004,59:305-331.
[163] Myers MG. Leptin receptor signaling and the regulation of mammalian physiology[J]. Recent Prog Horm Res., 2004, 59: 287-304.
[164] Siegrist-Kaiser CA, Pauli V, Juge-Aubry CE, et al, Direct Effects of Leptin on Brown and White Adipose Tissue[J]. J. Clin. Invest, 1997, 100: 2858-2864.
[165] Wang MY, Lee Y, Unger RH. Novel Form of Lipolysis Induced by Leptin [J]. J. Biol. Chem., 1999, 274: 17541-17544.
[166] Zhou Y, Wang Z, Higa M, et al. Reversing Adipocyte Differentiation: Implications for Treatment of Obesity [J]. Proc Natl Acad Sci. 1999,96: 2391-2395.
[167] Bates SH, Stearns WH, Dundon TA, et al. STAT3 signalling is required for leptin regulation of energy balance but not reproduction [J]. Nature, 2003, 421 (6925): 856-859.
[168] Cernkovich ER, Deng J, Bond MC et al. Adipose-specific disruption of signal transducer and activator of transcription 3 increase bodyweight and adiposity [J]. Endocrinology, 2007, 78: 35-39.
[169] Bartz Rene, Zehmer Johnk, Liu Pingsheng. New face of lipid droplets [J]. Prog. Biochem. Biophys. 2005, 32 (5): 387-392.
[170] Londos C, Sztalryd C, Tansey JT, et al. Role of PAT proteins in lipid metabolism[J]. Biochimie, 2005, 87: 45-49.
[171] Wolins NE, Quaynor BK, Skinner JR, et al. S3-12, adipophilin, and TIP47 package lipid in adipocytes [J]. J BiolChem, 2005,280:19146-19155.
[172] Xu G, Sztalryd C, Londos C. Degradation of perilipin is mediated through ubiquitination -proteasome pathway [J]. Biochimca et Biophysica Acta, 2006, 1761: 83-90.
[173] Brasaemle DL, Barber T, Kimmel AR, et al. Post-translational regulation of perilipin expression [J]. J Biol Chem, 1997, 272: 9378-9387.
[174] Tansey JT , Huml AM, Vogt R, et al. Function studies on native and mutated forms of perilipins [J]. J Biol Chem, 2003, 278: 8401-8406.
[175] Beckman M. Great Balls of Fat [J]. 2006, 311: SCIENCE, 1232-1234.
[176] Sztalryd C, Xu G, Dorward H, et al. Perilip in A is essential for the translocation of hormone-sensitive lipase during lipolytic activation [J]. J Cell Biol, 2003, 161: 1093-1103.
[177] Sztalryd C, Xu G, Dorward H, et al. Perilipin A is essential for the translocation of hormone-sensitive lipase during lipolytic activation [J]. J Cell Biol, 2003,161: 1093-1103.
[178] Tansey JT, Sztalryd C, Hlavin EM., et al. The Central Role of Perilipin A in Lipid Metabolism and Adipocyte Lipolysis [J]. IUBMB Life, 2004, 56 (7): 379-385.
[179] Cohen AW, Razani B, SchubertW, et al. Role of caveolin-1 in the modulation of lipolysis and lipid droplet formation [J]. Diabetes, 2004, 53: 1261-1270.
[180] Cohen AW, Razani B, Schubert W, et al. Role of caveolin-1 in the modulation of lipolysis and lipid droplet formation [J]. Diabetes, 2004, 53: 1261-1270.
[181] Arimura N, Horiba T, Imagawa M, et al. The peroxisome proliferators activated receptor gamma regulates expression of the perilipin gene in adipocytes [J]. J Biol Chem, 2004, 279: 10070-10076.
[182] Mottagui TS, Ryden M, Lofgren P, et al. Evidence for an important role of perilipin in the regulation of human adipocyte lipolysis [J]. Diabetologia, 2003,46: 789-797.
[183] Kern PA, Di Gregorio G, Lu T, et al. Perilipin expression in human adipose tissue is elevated with obesity[J]. J Clin Endocrinol Metab, 2004, 89: 1352-1358.
[184]Zhang HH,Souza SC,Muliro KV,et al.Lipase-selective functional domains of Perilipin A differentially regulate constitutive and protein kinase A-stimulated lipolysis[J].J.Biol.Chem., 2003,278:51535-51542.
[185]Zhang HH.,Halbleib M,Ahmad F,et al.Tumor necrosis factor-α stimulates lipolysis in differentiated human adipocytes through activation of extracellular signal-related kinase and elevation of intracellular cAMP[J].Diabetes,2002,51:2929-2935.
[186]Gerrity RG,Natarajan R,Nadler JL,et al.Diabetes-Induced Accelerated Atherosclerosis in Swine[J]. Diabetes,2001,50:1654-1665.
[187]李影，杨公社，卢容华．猪前体脂肪细胞培养方法的优化[J]．细胞生物学杂志，2005，27：697-700．
[188]Lin YQ,Zhuang HL,Yang GS.Effects of RXRα gene silencing on the porcine adipocyte differentiation in vitro[J].Comp.Biochem.Physiol.,Part D,2007,2:207-214.
[189]Fryer HJ,Davis GE,Manthorpe M,et al.Lowry protein assay using an automatic microtiter plate spectrophotometer[J].Anal.Biochem.,1986,153(2):262-266.
[190]Elks M L,Manganiello V C.Antilipolytic action of insulin:role of cAMP phosphodiesterase activation[J].Endocrinology,1985,116:2119-2121.
[191]Manganiello VC,Smith C J,Degerman E,et al.Molecular mechanisms involved in the antilipolytic action of insulin:phosphorylation and activation of a particulate adipocyte cAMP phosphodiesterase [J].New York:Plenum Press,1991,239-248.
[192]Coppack SW,Frayn KN,Humphreys SM,et al.Effects of insulin on human adipose tissue metabolism in vivo[J].Clin Sci.,1986,77:663-670.
[193]Smith U.Influence of insulin and medium glucose concentration on cellular metabolism[J].J Clin Invest.,1974,53:91-98.
[194]李晓华，郑升，刘优萍，等．胰岛素的长期作用对人脂肪细胞代谢的影响[J]．上海第二医科大学学报，2005，25(3)：239-241。
[195]Hardy RW,Grover MW,Carroll JW.Insulin inhibition of lipolysis in rat adipocytes without a concomitant decrease in cAMP[J].Diabetes,2001,50:A270.
[196]Rosenstock M,Greenberg AS,Rudich A.Distinct long-term regulation of glycerol and non-esterified fatty acid release by insulin and TNF-alpha in 3T3-L1 adipocytes[J].Diabetologia,2001,44:55- 62.
[197]Kang ES,Betts D,Fain.JN,et al.Chronic exposure of rat fat cells to insulin enhances lipolysis and activation of partially purified hormone-sensitive l ipase[J].Diabetes,1993,42:1415-24.
[198]Schweiger M,Schreiber R,Haemmerle G,et al.Adipose triglyceride lipase and hormone-sensitive lipase are the major enzymes in adipose tissue triacyiglycerol catabolism[J].J.Biol.Chem.,2006, 281(52):40236-40241.
[199]Miyoshi H,Souza SC,Zhang HH,et al.Perilipin promotes hormone-sensitive lipase-mediated adipocyte lipolysis via phosphorylation-dependent and-independent mechanisms[J].J.Biol. Chem.,2006,281(23):15837-15844.
[200]Kralisch S,Klein J,Lossner U,et al.Isoproterenol,TNFalpha,and insulin downregulate adipose triglyceride lipase in 3T3-L1 adipocytes[J].Mol Cell Endocrinol,2005,240(12):43-49.
[201]彭永德，陈家伦．PPARs与脂肪细胞分化及脂质代谢[J]．《国外医学》内分泌学分册，1998，12(18)： 172-175.
[202]Tontonoz PE,Spiegelman BM.Stimulation of adipogenesis in fibroblasts by PPARγ2,a lipid activated transcription factor[J].Cell,1994,79:1147-1156.
[203]Arimura N,Horiba T,Imagawa M,et al.The peroxisome proliferator-activated receptor-γ regulates expression of the Perilipin gene in adipocytes[J].J.Biol.Chem.2004,279(11):10070-10076.
[204]卢建雄，杨公社，陈粉粉．胰岛素对原代培养猪脂肪细胞生脂和脂解相关基因转录表达及脂代谢的影响[J]．农业生物技术学报，2006，14(1)：11-16．
[205]DeFronzo RA,Bonadonna RC,Ferrannini E.Pathogenesis of NIDDM:a balanced overview[J]. Diabetes Care,1992,15:318-368.
[206]Yki-Jarvinen H.Role of insulin resistance in the pathogenesis ofNIDDM[J].Diabetologia,1995,38: 1378-1388.
[207]Pickup JC,Chusney GD,Thomas SM,et al.Plasma interleukin-6,tumour necrosis factor alpha and blood cytokine production in type 2 diabetes[J].Life Sci,2000,67:291-300.
[208]Rotter V,Nagaev I,Smith U.Interleukin-6(IL-6)induces insulin resistance in 3T3-L1 adipocytes and is,like il-8 and tumor necrosis factor-α,overexpressed in human fat cells from insulin-resistant subjects[J].J.Biol.Chem.2003,278(46):45777-45784.
[209]Chung S,Brown JM,Provo JN,et al.Conjugated linoleic acid promotes human adipocyte insulin resistance through NFκB-dependent cytokine production[J].J.Biol.Chem.,2005,280(46): 38445-38456.
[210]Fasshauer M,Klein J,Lossner U,et al.Interleukin(IL)-6 mRNA expression is stimulated by insulin, isoproterenol,tumour necrosis factor alpha,growth hormone,and IL-6 in 3T3-L1 adipocytes[J]. Horm Metab Res.,2003,35(3):147-52.
[211]Chang L and Karin M.Mammalian MAP kinase signalling cascades[J].Nature,2001,410:37-40.
[212]Ajuwon KM,Spurlock ME.Direct regulation of lipolysis by interleukin-15 in primary pig adipocytes [J].Am J Physiol Regul Integr Comp Physiol,2004,287:R608-R611.
[213]Aiessi DR,Cuenda A,Cohen P,et ai.PD98059 is a specific inhibitor of the activation of mitogen-activated protein kinase kinase in vitro and in vivo[J].J Biol Chem,1995,270: 27489-27494.
[214]Favata MF,Horiuchi KY,Manos EJ,et al.Identification of a novel inhibitor of mitogen-activated protein kinase kinase[J].J Biol Chem,1998,273:18623-18632.
[215]Brasaemle DL,Rubin B,Harten IA,et al.Perilipin A Increases Triacylglycerol Storage by Decreasing the Rate of Triacylglycerol Hydrolysis[J].The Journal Of Biological Chemistry,2000, 275(49):38486-38493.
[216]Juan CC,Chang CL,Lai YH,et al.Endothelin-1 induces lipolysis in 3T3-L1 adipocytes[J].Am J Physiol Endocrinol Metab,2005,288:E1146-E1152.
[217]Fricke K,Heitland A,Maronde E.Cooperative Activation of Lipolysis by Protein Kinase A and Protein Kinase C Pathways in 3T3-L1 Adipocytes[J].Endocrinology,2004,145(11):4940-4947.
[218]Landstrom R,Mie J,Karlsson M,et al.Downregulation of cyclic nucleotide phosphodiesterase 3B in 3T3-L1 adipocytes induced by tumor necrosis factor α and cAMP[J].Biochem J,2000,346: 337-343.
[219]Prusty D,Park BH,Davis KE,et al.Activation of MEK/ERK signaling promotes adipogenesis by enhancing peroxisome proliferator-activated receptor(PPAR)and C/EBPα gene expression during the differentiation of 3T3-L1 preadipocytes[J].J Biol Chem.,2002,277:46226-46232.
[220]Fujishiro M,Gotoh Y,Katagiri H,et al.Three mitogen-activated protein kinases inhibit insulin signaling by different mechanisms in 3T3-L1 adipocytes[J].Mol Endocrinol,2003,17:487-497.
[221]Ryden M,Dicker A,van Harmelen V,et al.Mapping of early signaling events in tumor necrosis factor α mediated lipolysis in human fat cells[J].J Biol Chem,2002,277:1085-1091.
[222]Daeipour M,Kumar G,Amaral MC,et al.Recombinant IL-6 activates p42 and p44 mitogenactivated protein kinases in the IL-6 responsive B cell line AF-10[J].J Immunol,1993,150: 4743-4753.
[223]Browning DD,Diehl WC,Hsu MH,et al.Autocrine regulation of interleukin-8 production in human monocytes[J].Am J Physiol Lung Cell Mol Physiol,2000,279:L1129-L1136.
[224]Marette A.Mediators of cytokine-induced insulin resistance in obesity and other inflammatory settings[J].Curr.Opin.Clin.Nutr.Metab.Care,2002,5:377-383.
[225]Pradhan AD,Manson JE,Rifai N,et al.C-reactive protein,interleukin 6,and risk of developing type 2 diabetes mellitus[J].JAMA,2001,286:327-334.
[226]Trujillo ME,Sullivan S,Harten I,et al.Interleukin-6 regulates human adipose tissue lipid metabolism and leptin production in vitro[J].J.Clin.Endocrinol.Metab.,2004,89(11): 5577-5582.
[227]Petersen AM,Pedersen BK.The anti-inflammatory effect of exercise:The role of IL-6 in mediating the anti-inflammatory effects of exercise[J].J.Physiol.Pharmacol,2006,57(Suppl.10):43-51.
[228]Xing Z,Gauldie J,Cox G,et al.IL-6 is an anti-inflammatory cytokine required for controlling local or systemic acute inflammatory responses[J].J.Clin.Invest,1998,101:311-320.
[229]Schindler R,Mancilla J,Endres S,et al.Correlations and interactions in the production of interleukin-6(IL-6),IL-1,and tumor necrosis factor(TNF)in human blood mononuclear cells:IL-6 suppresses IL-1 and TNF[J].Blood,1990,75:40-47.
[230]Tilg H,Trehu E,Atkins MB,et al.Interleukin-6(IL-6)as an anti-inflammatory cytokine:induction of circulating IL-1 receptor antagonist and soluble tumor necrosis factor receptor p55[J].Blood, 1994,83(1):113-118.
[231]Ruderman NB,Keller C,Richard A.M,et al.Interleukin-6 regulation of AMP-activated protein kinase,potential role in the systemic response to exercise and prevention of the metabolic syndrome [J].Diabetes,2006,55(Supplement 2):S47-S54.
[232]Li Y,Yang GS,Lu RH.Optimal culture method of the porcine preadipocyte[J].Chin.J.Cell Biol., 2005,27:697-700.
[233]Kershaw EE,Flier JS.Adipose tissue as an endocrine organ[J].The Journal of Clinical Endocrinology & Metabolism,2004,89(6):2548-2556.
[234]Ryd(?)n M,Arvidsson E,Blomqvist L,et al.Targets for TNF-a-induced lipolysis in human adipocytes [J].Biochemical and Biophysical Research Communications,2004,318:168-175.
[235]Wang MY,Lee Y,Unger RH.Novel form of lipolysis induced by leptin[J].The Journal of Biological Chemistry,1999,274(25):17541-17544.
[236]Tschritter O,Fritsche A,Thamer C.Plasma adiponectin concentration predict insulin sensitivity of both glucose and lipid metabolism[J].Diabetes,2003,52(2):239-243.
[237] Petersen AM, Pedersen BK. The anti-inflammatory effect of exercise [J]. J. Appl. Physiol., 2005, 98: 1154-1162.
[238] Fasshaue M, Klein J, Lossner U, et al. Interleukin (IL)-6 mRNA expression is stimulated by insulin, isoproterenol, tumour necrosis factor alpha, growth hormone, and IL-6 in 3T3-L1 adipocytes [J]. Horm. Metab. Res., 2003, 35 (3): 147-152.
[239] Wang MY, Lee Y, Unger RH. Novel form of lipolysis induced by leptin [J]. J. Biol. Chem., 1999, 274(25): 17541-17544.
[240] Wang SP, Laurin N, Himms-Hagen J, et al. The adipose tissue phenotype of hormone-sensitive lipase deficiency in mice [J]. Obes. Res., 2001, 9: 119-128.
[241] Zimmermann R, Strauss JG, Haemmerle G, et al. Fat mobilization in adipose tissue is promoted by adipose triglyceride lipase [J]. Science, 2004, 306 (5700): 1383-1386.
[242] Xu G, Sztalryd C, Londos C. Degradation of Perilipin is mediated through ubiquitination -proteasome pathway [J]. Biochim. Biophys. Acta, 2006,1761: 83-90.
[243] Lagathu C, Bastard JP, Auclair M, et al. Chronic interleukin-6 (IL-6) treatment increased IL-6 secretion and induced insulin resistance in adipocyte: prevention by rosiglitazone [J]. Biochem. Biophys. Res. Commun., 2003, 311: 372-379.
[244] Kim MS, Sweeney TR, Shigenaga JK, et al. Tumor necrosis factor and interleukin 1 decrease RXRalpha, PPARalpha, PPARgamma, LXRalpha, and the coactivators SRC-1, PGC-1 alpha, and PGC-1beta in liver cells [J]. Metabolism, 2007, 56 (2): 267-79.
[245] Watt MJ, Carey AL, Wolsk-Petersen E, et al. Hormone-sensitive lipase is reduced in the adipose tissue of patients with type 2 diabetes mellitus: influence of IL-6 infusion [J]. Diabetologia, 2005, 48: 105-112.
[246] St-Pierre J, Lin J, Krauss S, et al. Bioenergetic analysis of peroxisome proliferator-activated receptor coactivators 1α and 1β (PGC-1α and PGC-1β) in muscle cells [J]. J. Biol. Chem., 2003, 278: 26597-26603.
[247] Luo GF, Yu TY, Wen XH, et al. Alteration of mitochondrial oxidative capacity during porcine preadipocyte differentiation and in response to leptin [J]. Mol. Cell. Biochem., 2008, 307: 83-91.
[248] Nicholls DG, Locke RM. Thermogenic mechanisms in brown fat [J]. Physiol. Rev., 1984, 64: 1-64.
[249] Patti ME, Butte AJ, Crunkhorn S, et al. Coordinated reduction of genes of oxidative metabolism in humans with insulin resistance and diabetes: potential role of PGC1 and NRF1 [J]. Proc. Natl. Acad. Sci. U S A, 2003, 100: 8466-8471.
[250] Petersen KF, Befroy D, Dufour S, et al. Mitochondrial dysfunction in the elderly: possible role in insulin resistance [J]. Science, 2003, 300: 1140-1142.
[251] Puigserver P, Spiegelman BM. Peroxisome proliferator-activated receptor gamma coactivator 1 alpha (PGC-1 alpha): transcriptional coactivator and metabolic regulator [J]. Endocr. Rev., 2003, 24: 78-90.
[252] Bruce CR, Dyck DJ. Cytokine regulation of skeletal muscle fatty acid metabolism: effect of interleukin-6 and tumor necrosis factor-α [J]. Am. J. Physiol. Endocrinol. Metab., 2004, 287: E616-E621.
[253] F(?)ldt J, Wernstedt I, Fitzgerald SM, et al. Reduced exercise endurance in interleukin-6-deficient mice [J]. Endocrinology, 2004, 145: 2680-2686.
[254]Kanda Y,Nishio E,Watanabe Y.Differential regulation ofNa+/H+exchange and DNA synthesis in vascular smooth muscle cells[J].Eur J Pharmacol.,1999,371:69-74.
[255]Beckman M.Great Balls of Fat[J].Science,2006,311:1232-1234.
[256]Solini A,Santini E,Madec S,et al.Rosiglitazone increases matrix production and quenches inflammation:studies in human cells[J].Diabetes Metab Res Rev.,2008,24(3):197-204.
[257]Kolak M,Yki-J(a|¨)rvinen H,Kannisto K,et al.Effects of Chronic Rosiglitazone Therapy on Gene Expression in Human Adipose Tissue in Vivo in Patients with Type 2 Diabetes[J].The Journal of Clinical Endocrinology & Metabolism,2007,92(2):720-724.
[258]Celec P.Nuclear factor kappa B molecular biomedicine:the next generation[J].Biomed Pharmacother,2004,58:365-71.
[259]Suzawa M,Takada I,Yanagisawa J,et al.Cytokines suppress adipogenesis and PPAR-gamma function through the TAK1/ /NIK cascade[J].Nat Cell Biol.,2003,5:224-230.
[260]Ruan H,Hacohen N,Golub TR,et al.Tumor necrosis factor-alpha suppresses adipocyte-specific genes and activates expression of preadipocyte genes in 3T3-L1 adipocytes:nuclear factor-kappaB activation by TNFalpha is obligatory[J].Diabetes,2002,51:1319-1336.
[261]Festuccia WT,Laplante M,Berthiaume M,et al.PPARgamma agonism increases rat adipose tissue lipolysis,expression of glyceride lipases,and the response of lipolysis to hormonal control[J]. Diabetologia,2006,49(10):2427-36.
[262]Hung Y J,Lin SH,Pei D,et al.Rosiglitazone improves insulin sensitivity in nonobese subjects with impaired glucose tolerance:the role ofadiponectin and C-reactive protein[J].Metabolism,2006,55 (4):439-44.
[263]Sutinen J,Kannisto K,Korsheninnikova E,et al.Effects of rosiglitazone on gene expression in subcutaneous adipose tissue in highly active antiretroviral therapy-associated lipodystrophy[J].Am J Physiol Endocrinol Metab,2004,286:E941-E949.
