Effect of TNF-α on adipogenesis of SD rat bone marrow mesenchymal stem cells and the related molecular mechanism

Li Lin; Xie Youhong

doi:10.13406/j.cnki.cyxb.001040

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Effect of TNF-α on adipogenesis of SD rat bone marrow mesenchymal stem cells and the related molecular mechanism

更新时间：2024-01-17

- Effect of TNF-α on adipogenesis of SD rat bone marrow mesenchymal stem cells and the related molecular mechanism
- Journal of Chongqing Medical University Vol. 41, Issue 10, Pages: 1016-1021(2016)
- 作者机构：
  
  1. 重庆医科大学附属第一医院消化内科
  2. 重庆医科大学附属大学城医院消化中心
- 作者简介：
- 基金信息：
- DOI：10.13406/j.cnki.cyxb.001040
  CLC： R329.2
- Published：2016，
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[1]李琳,谢友红.TNF-α对SD大鼠骨髓间充质干细胞成脂分化的影响及其相关分子机制的探讨[J].重庆医科大学学报,2016,41(10):1016-1021.

Li Lin, Xie Youhong. Effect of TNF-α on adipogenesis of SD rat bone marrow mesenchymal stem cells and the related molecular mechanism. [J]. Journal of Chongqing Medical University 41(10):1016-1021(2016)
[1]李琳,谢友红.TNF-α对SD大鼠骨髓间充质干细胞成脂分化的影响及其相关分子机制的探讨[J].重庆医科大学学报,2016,41(10):1016-1021. DOI： 10.13406/j.cnki.cyxb.001040.

Li Lin, Xie Youhong. Effect of TNF-α on adipogenesis of SD rat bone marrow mesenchymal stem cells and the related molecular mechanism. [J]. Journal of Chongqing Medical University 41(10):1016-1021(2016) DOI： 10.13406/j.cnki.cyxb.001040.

摘要

目的:探讨炎症因子转化生长因子（transforming growth factor-α

TNF-α）对SD大鼠骨髓间充质干细胞（bone marrow mesenchymal stem cells

BMSCs）成脂分化的影响及相关的分子机制。方法:将生长良好的第3代（P3）SD大鼠BMSCs分为对照组（Con组）、成脂诱导组（Ad组）和10 ng/m L TNF-α干预成脂诱导组（Ad+TNF-α组）

每组各接种5块6孔板

培养14 d后

收集各组细胞的RNA和蛋白质。于第14天时间点处进行油红O染色和异丙醇萃取

酶标仪490 nm处检测吸光度（absorbance

A）值

观察各组成脂分化的程度。q RT-PCR检测各组基因Wnt10b、前体脂肪细胞因子-1（preadipocyte factor-1

Pref-1）、CCAAT区增强子结合蛋白-α（CCAAT/enhancer-binding protein-α

C/EBP-α）、过氧化物酶体增殖物激活受体-γ（peroxisome proliferator-activated receptor-γ

PPAR-γ）、脂肪酸结合蛋白-4（fatty acid binding protein-4

FABP-4）的m RNA表达水平。Western blot检测第14天3组基因β联蛋白（β-catenin）、PPAR-γ、C/EBP-α、FABP-4的蛋白表达水平。结果:TNF-α可以明显抑制SD大鼠BMSCs的成脂分化。在Ad+TNF-α组

油红O染色明显低于Ad组

异丙醇萃取后A值检测也明显低于Ad组（0.360±0.035 vs.0.770±0.025

P=0.000）。q RT-PCR结果显示

在Ad+TNF-α组

Wnt10b、Pref-1的m RNA水平（17.050±2.706

4.135±0.280）明显高于Con组（P=0.019

P=0.003）和Ad组（0.575±0.065

0.514±0.060）（P=0.020

P=0.006）

而C/EBP-α、PPAR-γ、FABP-4的m RNA水平（0.200±0.012

0.768±0.030

0.883±0.048）明显低于Ad组（2.965±0.455

2.330±0.211

3.847±0.171）（P=0.019

P=0.000

P=0.001）。Western blot结果显示

诱导14 d后

Ad+TNF-α组的C/EBP-α、PPAR-γ、FABP-4蛋白表达水平（0.586±0.013

0.356±0.008

0.118±0.002）明显低于Ad组（1.082±0.018

0.840±0.112

1.094±0.038）（均P=0.000）。在Ad+TNF-α组

磷酸化β-catenin（P-β-catenin）的蛋白水平（0.648±0.005）明显低于Ad组（3.376±0.211）（P=0.000）。结论:TNF-α抑制SD大鼠BMSCs成脂过程分化相的作用可能与Wnt/β-catenin信号通路活性的增加和Pref-1的高表达有关

由此BMSCs的成脂分化停留在了前体脂肪细胞阶段。

Abstract

Objective:To investigate the effect of transforming growth factor-α(TNF-α)on adipogenesis of SD rat bone marrow mesenchymal stem cells(BMSCs)and the related molecular mechanism. Methods:The well grown third-passage(P3)SD rat BMSCs were divided into three groups:control group

Ad group and Ad+TNF-α group. RNA and proteins of the cells in three groups were collected at day 14. Cells were stained with oil red O on day 14 to examine the lipid accumulation in each group. Optical density for oil red O was determined by isopropanol extraction and was measured in a plate reader at 490 nm. The m RNA expressions of Wnt10 b

peroxisome proliferator-activated receptor-γ(PPAR-γ)

CCAAT/enhancer-binding protein-α(C/EBP-α)

fatty acid binding protein-4(FABP-4)and preadipocyte factor-1(Pref-1)were detected by q RT-PCR. The protein expression levels of P-β-catenin

PPAR-γ

C/EBP-α

FABP-4 were detected by Western blot. Results:The absorbance(A)value measured at 490 nm was significantly lower in Ad+TNF-α group(0.360±0.035)than in Ad group(0.770±0.025)(P = 0.000). q RT-PCR analysis showed that m RNA expressions of Wnt10 b and Pref-1 were significantly higher in Ad+TNF-α group(17.050±2.706

4.135±0.280)than in Ad group(0. 575 ± 0. 065

0. 514 ± 0. 060)(P = 0. 02

P = 0. 006).While the m RNA expressions of C/EBP-α

PPAR-γ and FABP-4 were significantly lower in Ad +TNF-α group(0.200 ±0.012

0.768 ±0.030

0.883 ±0.048) than in Ad group(2.965 ±0.455

2.330 ±0.211

3.847 ±0.171)(P =0.019

P =0.000

P =0.001).Western blot showed the protein expressions of C/EBP-ɑ

PPAR-γ and FABP-4 were significantly lower in Ad+TNF-α group(0.586±0.013

0.356±0.008

0.118±0.002)than in Ad group(1.082±0.018

0.840±0.112

1.094±0.038)(P=0.000). The protein expression of P-β-catenin was significantly lower in Ad+TNF-α group(0.648±0.005)than in Ad group(3.376±0.211)(P=0.000). Conclusion :The fact that TNF- α inhibits the terminal differentiation of the adipogenesis of SD rat BMSCs and blocks the adipogenesis of SD BMSCs at preadipocyte stage may relate to the increased activity of Wnt/β-catenin signaling pathway and high expression of Pref-1.

关键词

骨髓间充质干细胞成脂分化Wnt/β-catenin信号通路前体脂肪细胞因子-1

Keywords

bone marrow mesenchymal stem cellsadipogenesisWnt/β-catenin signaling pathwaypreadipocyte factor-1

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