目的：研究磷脂酰肌醇激酶（PI3K）抑制剂LY294002对小鼠前脂肪细胞分化和CCAAT增强子结合蛋白α（C/EBPα）及过氧化物酶体增殖物激活受体γ（PPARγ）表达的影响，探讨胰岛素受体底物（IRSs）/PI3K信号通路在前脂肪细胞分化中的作用。方法：小鼠3T3-L1细胞分为实验组和对照组，实验组加入LY294002（25 μmol/L），对照组加入同体积DMSO。应用0.5 mmol/L 3-异丁基-1-甲基黄嘌呤(IBMX)、10-6 mol/L地塞米松和5 μg/mL胰岛素诱导两组前脂肪细胞分化，在培养的0 d、2 d、4 d、8 d分别收集细胞，实时PCR法及Western blot法检测细胞中C/EBPα和PPARγ表达水平。培养8 d油红O染色，观察细胞分化情况。结果：小鼠3T3-L1细胞基础状态下两组C/EBPα及PPARγ表达差异无统计学意义（P＞0.05）；诱导分化时实验组C/EBPα及PPARγ表达低于对照组（分别P＜0.05，P＜0.01）。油红O染色示实验组细胞无明显脂滴。结论：PI3K抑制剂LY294002能抑制小鼠前脂肪细胞分化及C/EBPα和PPARγ的表达，提示IRSs/PI3K信号通路可通过调节PPARγ和C/EBPα的表达在3T3-L1前脂肪细胞的分化中发挥重要作用。
Abstract
OBJECTIVE: This study examined the effects of PI3K inhibitor LY294002 on the differentiation of mouse preadipocytes and the expression of CCAAT enhancer binding protein α (C/EBPα) and peroxisome proliferation activated receptor γ (PPARγ), in order to study the possible roles of insulin receptor substrate (IRSs)/PI3K signal pathway in the differentiation of preadipocytes. METHODS: The mouse 3T3-L1 cells were cultured normally and divided into experimental and control groups. 3T3-L1 cells in the experimental group were treated with PI3K inhibitor LY294002 (25 μmol/L) and those in the control group were treated with DMSO culture medium. 3-isobutyl-1-methylxanthine (IBMX) (0.5 mmol/L), dexamethasone (10-6 mol/L) and insulin (5 μg/mL) were used to induce the differentiation of 3T3-L1 preadipocytes in both groups. Before culture, and 2, 4 and 8 days after culture, the cells were collected to detect the expression of C/EBPα and PPARγ by real-time PCR and Western blot assays. The lipid droplets of 3T3-L1 preadipocytes were observed by oil-red O staining. RESULTS: PI3K inhibitor LY294002 did not affect the expression of C/EBPα and PPARγ in un-induced 3T3-L1 preadipocytes (P>0.05), but decreased the expression of C/EBPα and PPARγ during the in vitro induced differentiation of 3T3-L1 preadipocytes compared with the control group (P<0.05 or 0.01). The lipid droplets count was greatly reduced by LY294002. CONCLUSIONS: PI3K inhibitor LY294002 can inhibit the differentiation of mouse 3T3-LI preadipocytes and the expression of C/EBPα and PPARγ in the differentiation of 3T3-LI preadipoeytes, suggesting that IRSs/PI3K signal pathway may play an important role in the differentiation of 3T3-L1 preadipocytes by regulating the expression of C/EBPα and PPARγ.
关键词
PI3K抑制剂 /
3T3-L1细胞 /
分化 /
C/EBPα /
PPARγ /
小鼠
Key words
PI3K inhibitor /
3T3-L1 cell /
Differentiation /
C/EBPα /
PPARγ /
Mice
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参考文献
［1］鲍鹏丽，刘戈力，杨菁岩，姜丽红，魏莹，穆青，等.体格测量指标对肥胖儿童胰岛素抵抗的预测意义［J］.实用儿科临床杂志，2010，25（7）：484-485.
［2］Achike FI, To NH, Wang H, Kwan CY. Obesity, metabolic syndrome, adipocytes and vascular function: A holistic viewpoint［J］. Clin Exp Pharmacol Physiol, 2011, 38(1): 1-10.
［3］Tseng YH, Butte AJ, Kokkotou E, Yechoor VK, Taniquchi CM, Kriauciunas KM, et al. Prediction of preadipocyte differentiation by gene expression reveals role of insulin receptor substrates and nectin［J］. Nat Cell Biol, 2005, 7(6): 601-611.
［4］Rosen ED, Hsu CH, Wang X, Sakai S, Freeman MW, Gonzalez FJ, et al. C/EBPalpha induces adipogenesis through PPARgamma: a unified pathway［J］. Genes Dev, 2002, 16(1): 22-26.
［5］Ros Pérez M, Medina-Gómez G. Obesity, adipogenesis and insulin resistance ［J］. Endocrinol Nutr, 2011, 58(7): 360-369.
［6］Hayashi T, Nozaki Y, Nishizuka M, Ikawa M, Osada S, Imagawa M. Factor for adipocyte differentiation 158 gene disruption prevents the body weight gain and insulin resistance induced by a high-fat diet［J］. Biol Pharm Bull, 2011, 34(8): 1257-1263.
［7］Rosen ED, MacDougald OA. Adipocyte differentiation from the inside out［J］. Nat Rev Mol Cell Biol, 2006, 7(12): 885-896.
［8］李瑶，辛颖．胰岛素受体底物在宫内发育迟缓大鼠胰腺组织中的表达［J］．中国当代儿科杂志，2010，12（12）：972-975.
［9］刘晓梅，焦伊胜，潘莉莉，卢岩，李书琴．宫内生长受限大鼠肝脏磷酸肌醇-3-激酶信号通路分子的变化［Ｊ］．中国当代儿科杂志，2009，11（3）：221-224.
［10］Taniguchi CM, Emanuelli B, Kahn CR. Critical nodes in signaling pathways: insights into insulin action［J］. Nat Rev Mol Cell Biol, 2006, 7(2): 85-96.
［11］Kohn AD, Summers SA, Birnbaum MJ, Roth RA. Expression of a constitutively active Akt Ser/Thr kinase in 3T3L1 adipocytes stimulates glucose uptake and glucose transporter 4 translocation［J］. J Biol Chem, 1996, 271(49): 31372-31378.
［12］Yun SJ, Kim EK, Tucker DF, Kim CD, Birnbaum MJ, Bae SS. Isoform-specific regulation of adipocyte differentiation by Akt/protein kinase Balpha［J］. Biochem Biophys Res Commun, 2008, 371(1): 138-143.
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