遗传 ›› 2018, Vol. 40 ›› Issue (9): 758-766.doi: 10.16288/j.yczz.18-053

• 研究报告 • 上一篇    下一篇

Lnc-RAP3对小鼠3T3-L1前脂肪细胞分化的影响

李欢1(),冯晋川2,李贵林1,王讯1,李明洲1,刘海峰1()   

  1. 1. 四川农业大学,畜禽遗传资源发掘与创新利用四川省重点实验室,成都 611130
    2. 四川农业大学动物科技学院,成都 611130
  • 收稿日期:2018-03-01 修回日期:2018-05-07 出版日期:2018-09-20 发布日期:2018-08-01
  • 作者简介:作者简介: 李欢,硕士研究生,专业方向：分子遗传与动物育种。E-mail: lihuan0461@163.com
  • 基金资助:
    国家自然科学基金项目(31522055);国家自然科学基金项目(31772576);四川省青年科技创新研究团队项目(2015TD0012)

The effect of lnc-RAP3 on 3T3-L1 preadipocyte differentiation in mouse

Li Huan1(),Feng Jinchuan2,Li Guilin1,Wang Xun1,Li Mingzhou1,Liu Haifeng1()   

  1. 1. Farm Animal Genetic Resources Exploration and Innovation Key Laboratory of Sichuan Province, Sichuan Agricultural University, Chengdu 611130, China
    2. College of Animal Science and Technology, Sichuan Agricultural University, Chengdu 611130, China
  • Received:2018-03-01 Revised:2018-05-07 Online:2018-09-20 Published:2018-08-01
  • Supported by:
    Supported by the National Natural Science Foundation of China(31522055);Supported by the National Natural Science Foundation of China(31772576);the Program for Innovative Research Team of Sichuan Province(2015TD0012)

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摘要：

长链非编码RNA (long non-coding RNA, lncRNA)是一类长度大于200nt、没有长开放阅读框架但往往具有mRNA结构特征的RNA,可以在转录及转录后水平参与基因的表达调控。近年来,有研究证实lncRNA对脂肪生成具有重要作用。Lnc-RAP3位于小鼠(Mus musculus)17号染色体,其表达量在小鼠脂肪细胞分化前后呈现显著差异,但其具体的生物学功能尚不清楚。为探讨lnc-RAP3在小鼠3T3-L1前脂肪细胞成脂分化中的作用,本文首先构建了lnc-RAP3的真核表达载体pcDNA3.1-RAP3,利用脂质体将pcDNA3.1-RAP3和人工合成的lnc-RAP3的siRNAs分别转染3T3-L1前脂肪细胞,并对转染后的细胞进行诱导分化,并通过油红O染色、qRT-PCR检测成脂分化相关基因表达等方法比较过表达和敲降lnc-RAP3对3T3-L1前脂肪细胞成脂分化的影响。结果显示,过表达lnc-RAP3后,细胞内脂滴聚集显著减少(P<0.05),在诱导分化第0 d、2 d和4 d时C/EBPα、Glut4、PPARγ、LPL和FAS的表达水平均呈显著(P<0.05)或极显著(P<0.01)下降;敲降lnc-RAP3后,细胞内脂滴聚集显著增多(P<0.05),同时在诱导分化第0 d、2 d时PPARγ、LPL、C/EBPα、FAS和Glut4的表达水平呈显著(P<0.05)或极显著(P<0.01)升高。本研究结果表明,lnc-RAP3可能通过影响成脂分化相关基因的表达来抑制3T3-L1前脂肪细胞的成脂分化。

关键词: lnc-RAP3, 3T3-L1前脂肪细胞, 成脂分化

Abstract:

The long non-coding RNAs (lncRNAs) are a type of RNAs with more than 200nt in length and without any long open reading frame, but often have mRNA structural features. They can regulate the expression of target genes in different manners at the transcriptional and post-transcriptional levels. In recent years, various studies demonstrated that lncRNAs play crucial roles in adipogenesis. The long non-coding RNA, lnc-RAP3, located on the mouse chromosome 17, possesses a significantly differential expression pattern during mouse adipocyte differentiation; but its specific biological function (s) remains unclear. To investigate the effect of lnc-RAP3 on adipogenesis in the mouse 3T3-L1 preadipocytes, we first constructed a eukaryotic expression vector pcDNA3.1-RAP3. pcDNA3.1-RAP3 and synthetic RAP3-siRNAs were transfected individually into 3T3-L1 preadipocytes by Lipofectamine TM 2000, thereby over-expressing and knocking-down lnc-RAP3 expression, respectively. The transfected preadipocytes were induced to undergo adipogenic differentiation. Oil Red O staining and qRT-PCR were used to detect the effects of lnc-RAP3 overexpression/knockdown on 3T3-L1 preadipocyte differentiation. The results showed that overexpression of lnc-RAP3 led to a notable decrease in lipid accumulation (P<0.05) and remarkably reduced the mRNA expression levels of C/EBPα, Glut4, PPARγ, LPL and FAS on day 0, day 2, and day 4 post differentiation (P<0.05, P<0.01). In contrast, quantitative analysis of Oil Red O on day 4 of differentiation revealed that inhibition of lnc-RAP3 increased the formation of neutral lipid droplets (P<0.05). In addition, silencing lnc-RAP3 also significantly increased the mRNA expression of PPARγ, LPL, C/EBPα, FAS and Glut4 on day 0 and day 2 post differentiation (P<0.05, P<0.01). Our study suggests that lnc-RAP3 might suppress 3T3-L1 preadipocyte differentiation by affecting the expression of the genes involved in adipogenic differentiation.

Key words: lnc-RAP3, 3T3-L1 preadipocytes, adipogenic differentiation