Exendin-4对脂肪细胞分化及相关基因表达的影响
摘要
目的:探讨Exendin-4对3T3-L1前脂肪细胞分化及GLUT-4、PPAR-γ、HSL mRNA表达的影响。
方法:3T3-L1前脂肪细胞诱导分化不同时期分别用Exendin-4等干预,应用倒置相差显微镜观察3T3-L1前脂肪细胞诱导分化前后的细胞形态变化;油红O染色后观察脂滴数量;Q-PCR分别检测GLUT-4、PPAR-γ、HSL mRNA表达量;应用酶法测定脂肪细胞的甘油三酯(TG)含量。
结果:诱导分化成熟的脂肪细胞经油红O染色可见细胞质内大片脂滴呈亮红色,而未分化细胞则不被油红O染色。
在脂肪细胞分化第0d干预,Exendin-4可促进脂肪细胞分化,较空白组增加细胞内TG的含量(P<0.01),上调GLUT-4、PPAR-γ、HSL mRNA的表达(P<0.01),TNF-α组较空白组均能显著下调GLUT-4、HSL mRNA的表达(P<0.01);与Troglitazone组比较,EX-4组PPAR-γmRNA的表达降低(P<0.05),EX-4+Trogli组GLUT-4、HSL mRNA的表达显著增加(P<0.01),EX-4+Trogli组PPAR-γmRNA的表达增加(P<0.05)。
在脂肪细胞分化6d干预,Exendin-4可促进脂肪细胞分化,较空白组增加细胞内TG的含量(P<0.01),上调GLUT-4、HSL、PPAR-γmRNA的表达(P<0.01);TNF-α组较空白组能显著下调GLUT-4、HSL mRNA的表达(P<0.01),下调PPAR-γmRNA的表达(P<0.05);与Troglitazone组比较,EX-4组GLUT-4 mRNA的表达降低(P<0.05),HSL mRNA的表达显著降低(P<0.01),EX-4+Trogli组HSL、PPAR-γmRNA的表达增加(P<0.05)。
脂肪细胞分化第12d干预,Trogli组、EX-4组+Trogli组较空白组能上调GLUT-4 mRNA的表达(P<0.05);TNF-α组较空白组能下调HSL mRNA的表达(P<0.01);与Troglitazone组比较,EX-4组GLUT-4、HSL mRNA的表达减少(P<0.05),EX-4组+Trogli组HSL mRNA的表达增加(P<0.05),TG含量也增加(P<0.05)。
结论:Exendin-4促进脂肪细胞分化并上调糖脂代谢相关基因GLUT-4、PPAR-γ、HSL mRNA表达,可能为Exendin-4抗糖尿病的部分作用新机制。
Objective: To investigate the effects of Exendin-4 on differentiation of 3T3-L1 preadipocytes and mRNA expressions of GLUT-4、PPAR-γand HSL.
Methods: 3T3-L1 preadipocytes of different differentiation stages were treated by Exendin-4. Morphological changes of 3T3-L1 preadipocytes and cell differentiation were measured by inverted phase contrast mocroscope. The number of lipid droplets was observed by Oil red O staining. The mRNA expressions of GLUT-4、PPAR-γand HSL were detected by quantitative real-time polymerase chain reaction(RT-PCR).The triglyceride content of 3T3-L1 preadipocytes was quantitated with an acetyl acetone–based colorimetric kit.
Results: Induced differentiation of mature adipocytes showed large lipid droplets of red within the cytoplasm by oil red O staining, undifferentiated cells are not stained by oil red O.
Comparing with control group, Exendin-4 promoted preadipocyte differentiation, increased TG content (P<0.01) and the mRNA expression of GLUT-4, HSL and PPAR-γ(P<0.01) when treated with Exendin-4 at day 0.Compared with the control group,TNF-αsignificantly reduced mRNA expressions of GLUT-4 and HSL (P<0.01). Compared with troglitazone group, the expressions of PPAR-γmRNA reduced in EX-4 group (P<0.05), the expressions of GLUT-4 and HSL mRNA significantly increased (P<0.01) and the expressions of PPAR-γmRNA increased (P<0.05) in Ex-4 combined with troglitazone group.
Comparing with control group, Exendin-4 promoted preadipocyte differentiation, increased TG content (P<0.01) and the mRNA expressions of GLUT-4, HSL andPPAR-γ(P<0.01) when treated with Exendin-4 at day 6. Compared the control group, TNF-αgroup significantly reduced GLUT-4 and HSL mRNA expressions (P<0.01) and reduced PPAR-γmRNA expression (P<0.05); Compared with Troglitazone group , the expressions of GLUT-4 mRNA reduced in Ex-4 group (P<0.05) and the expression of HSL mRNA significantly reduced (P<0.01). Compared with Troglitazone group, the expressions of HSL and PPAR-γmRNA increased in Ex-4 combined with troglitazone group (P <0.05).
Compared with control group, the expressions of GLUT-4 mRNA increased in Ex-4 combined with troglitazone group and Trogli group(P<0.05). Compared with control group, the expressions of HSL mRNA reduced in TNF-αgroup (P<0.01) when treated with Exendin-4 at day. Compared with Troglitazone group, the expressions of GLUT-4 and HSL mRNA reduced in Ex-4 group (P<0.05), Compared with Troglitazone group, increased TG content (P<0.01) and the mRNA expression of HSL mRNA increased in Ex-4 combined with troglitazone group (P<0.05) and TG content also increased (P<0.05).
Conlusion: Exendin-4 promoted adipocyte differentiation,increased glucose and lipid metabolism related genes mRNA expression, which may be some new mechanisms of Exendin-4 anti-diabetic.
方法:3T3-L1前脂肪细胞诱导分化不同时期分别用Exendin-4等干预,应用倒置相差显微镜观察3T3-L1前脂肪细胞诱导分化前后的细胞形态变化;油红O染色后观察脂滴数量;Q-PCR分别检测GLUT-4、PPAR-γ、HSL mRNA表达量;应用酶法测定脂肪细胞的甘油三酯(TG)含量。
结果:诱导分化成熟的脂肪细胞经油红O染色可见细胞质内大片脂滴呈亮红色,而未分化细胞则不被油红O染色。
在脂肪细胞分化第0d干预,Exendin-4可促进脂肪细胞分化,较空白组增加细胞内TG的含量(P<0.01),上调GLUT-4、PPAR-γ、HSL mRNA的表达(P<0.01),TNF-α组较空白组均能显著下调GLUT-4、HSL mRNA的表达(P<0.01);与Troglitazone组比较,EX-4组PPAR-γmRNA的表达降低(P<0.05),EX-4+Trogli组GLUT-4、HSL mRNA的表达显著增加(P<0.01),EX-4+Trogli组PPAR-γmRNA的表达增加(P<0.05)。
在脂肪细胞分化6d干预,Exendin-4可促进脂肪细胞分化,较空白组增加细胞内TG的含量(P<0.01),上调GLUT-4、HSL、PPAR-γmRNA的表达(P<0.01);TNF-α组较空白组能显著下调GLUT-4、HSL mRNA的表达(P<0.01),下调PPAR-γmRNA的表达(P<0.05);与Troglitazone组比较,EX-4组GLUT-4 mRNA的表达降低(P<0.05),HSL mRNA的表达显著降低(P<0.01),EX-4+Trogli组HSL、PPAR-γmRNA的表达增加(P<0.05)。
脂肪细胞分化第12d干预,Trogli组、EX-4组+Trogli组较空白组能上调GLUT-4 mRNA的表达(P<0.05);TNF-α组较空白组能下调HSL mRNA的表达(P<0.01);与Troglitazone组比较,EX-4组GLUT-4、HSL mRNA的表达减少(P<0.05),EX-4组+Trogli组HSL mRNA的表达增加(P<0.05),TG含量也增加(P<0.05)。
结论:Exendin-4促进脂肪细胞分化并上调糖脂代谢相关基因GLUT-4、PPAR-γ、HSL mRNA表达,可能为Exendin-4抗糖尿病的部分作用新机制。
Objective: To investigate the effects of Exendin-4 on differentiation of 3T3-L1 preadipocytes and mRNA expressions of GLUT-4、PPAR-γand HSL.
Methods: 3T3-L1 preadipocytes of different differentiation stages were treated by Exendin-4. Morphological changes of 3T3-L1 preadipocytes and cell differentiation were measured by inverted phase contrast mocroscope. The number of lipid droplets was observed by Oil red O staining. The mRNA expressions of GLUT-4、PPAR-γand HSL were detected by quantitative real-time polymerase chain reaction(RT-PCR).The triglyceride content of 3T3-L1 preadipocytes was quantitated with an acetyl acetone–based colorimetric kit.
Results: Induced differentiation of mature adipocytes showed large lipid droplets of red within the cytoplasm by oil red O staining, undifferentiated cells are not stained by oil red O.
Comparing with control group, Exendin-4 promoted preadipocyte differentiation, increased TG content (P<0.01) and the mRNA expression of GLUT-4, HSL and PPAR-γ(P<0.01) when treated with Exendin-4 at day 0.Compared with the control group,TNF-αsignificantly reduced mRNA expressions of GLUT-4 and HSL (P<0.01). Compared with troglitazone group, the expressions of PPAR-γmRNA reduced in EX-4 group (P<0.05), the expressions of GLUT-4 and HSL mRNA significantly increased (P<0.01) and the expressions of PPAR-γmRNA increased (P<0.05) in Ex-4 combined with troglitazone group.
Comparing with control group, Exendin-4 promoted preadipocyte differentiation, increased TG content (P<0.01) and the mRNA expressions of GLUT-4, HSL andPPAR-γ(P<0.01) when treated with Exendin-4 at day 6. Compared the control group, TNF-αgroup significantly reduced GLUT-4 and HSL mRNA expressions (P<0.01) and reduced PPAR-γmRNA expression (P<0.05); Compared with Troglitazone group , the expressions of GLUT-4 mRNA reduced in Ex-4 group (P<0.05) and the expression of HSL mRNA significantly reduced (P<0.01). Compared with Troglitazone group, the expressions of HSL and PPAR-γmRNA increased in Ex-4 combined with troglitazone group (P <0.05).
Compared with control group, the expressions of GLUT-4 mRNA increased in Ex-4 combined with troglitazone group and Trogli group(P<0.05). Compared with control group, the expressions of HSL mRNA reduced in TNF-αgroup (P<0.01) when treated with Exendin-4 at day. Compared with Troglitazone group, the expressions of GLUT-4 and HSL mRNA reduced in Ex-4 group (P<0.05), Compared with Troglitazone group, increased TG content (P<0.01) and the mRNA expression of HSL mRNA increased in Ex-4 combined with troglitazone group (P<0.05) and TG content also increased (P<0.05).
Conlusion: Exendin-4 promoted adipocyte differentiation,increased glucose and lipid metabolism related genes mRNA expression, which may be some new mechanisms of Exendin-4 anti-diabetic.
引文
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[2].Nielsen LL,Baron AD.Pharmacology of exenatide (synthetic exendin-4) for the treatment of type 2 diabetes.Curr Opinion Investig Drugs 2003;4:401–405.
[3].Gedulin BR,Nikoulina SE,Smith PA,et al.Exenatide (exendin-4) improves insulin sensitivity and beta-cell mass in insulin-resistant obese fa/fa Zucker rats independent of glycemia and body weight. Endocrinology, 2005;146:2069–2076.
[4].Fineman MS,Shen LZ,Kristin Taylor T,et al.Effectiveness of progressive dose- escalation of exenatide(exendin-4) in reducing dose-limiting side effects in subjects with type 2 diabetes.Diabetes Metab Res Rev 200420:411-417.
[5].Kolterman D,Kim DD,Shen L,et al. Pharmacokinetics, pharmacolynamics,and safety of exenatide in patients with type 2 diabetes mellitus.[J].Am J Health Pharm, 2005;62:173-81.
[6].杨涛,杨永年.Exenatide与β细胞功能.国外医学内分泌分册,2005,25 (1):18-22.
[7].Otto TC,LaneMD. Adipose development: from stem cell to adipocyte.Crit Rev Biochem Mol Biol,2005;40(4):229-242.
[8].Kershaw EE,Hamm JK,Verhagen LA,et al.Adipose triglyceride lipase: function, regulation by insulin,and comparison with adiponutrin. Diabetes, 2006;55: 148-157.
[9].Pannacciulli N,Le DS, Salbe AD, et al. Postprandial glucagon like peptide -1(GLP-1) response is positively associated with changes in neuronal activity of brain areas implicated in satiety and food intake regulation in humans. Neuro image,2007;35(2):511-7.
[10].McDermott LC,Freel JA,West AP,et al.Zn-alpha2-glycoprotein,an MHC class I related glycoprotein regulator of adipose tissues: modification or abrogation of ligand binding by site directed mutagenesis.Biochemistry,2006;45:2035-2041.
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