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miRNA作为内源性非编码小RNA,主要在转录后水平调控基因表达。近年来越来越多的证据表明,miRNA通过调控脂肪细胞分化相关的转录因子和信号通路来影响脂肪细胞的分化。为探究miR-26a对小鼠(Mus musculus)3T3-L1前脂肪细胞分化的影响及具体的调节作用,本研究分别用miR-26a的agomir和antagomir对miR-26a进行过表达和敲除。结果显示,miR-26a的表达量在3T3-L1细胞分化过程中逐渐上升。过表达miR-26a极显著增加了脂肪合成相关基因——过氧化物酶体增殖物激活受体γ(peroxisome proliferator-activated receptor gamma,PPARγ)、脂肪酸合酶(fatty acid synthase,FAS)和脂蛋白脂酶(lipoprotein lipase,LPL)的mRNA水平(P<0.01),PPARγ和FAS的蛋白水平也显著升高(P<0.05)。同时,细胞内的脂质聚集也明显增加。相反,抑制miR-26a的表达量降低了3T3-L1细胞的分化。通过靶基因预测和双荧光素酶报告系统分析,证明miR-26a能直接结合到同源性磷酸酶-张力蛋白(phosphatase and tensin homolog,PTEN)mRNA的3’UTR,并且导致其miRNA和蛋白水平降低(P<0.05)。本研究的结果表明,miR-26a可能通过直接抑制PTEN来正向的调节3T3-L1细胞的脂肪分化,该结果为miRNA成为治疗肥胖相关疾病的药物靶分子提供了一定的理论依据。
As an endogenous non-coding small RNA, miRNA mainly regulates gene expression at the post-transcriptional level. In recent years, more and more evidences show that miRNAs affect adipocyte differentiation by regulating transcription factors and signaling pathways involved in adipocyte differentiation. In order to investigate the effect of miR-26a on the differentiation of 3T3-L1 preadipocytes in mice and to investigate the role of miR-26a in miR-26a pre-adipocyte differentiation, miR-26a was overexpressed and knocked out by agomir and antagomir. The results showed that the expression of miR-26a gradually increased during the differentiation of 3T3-L1 cells. Overexpression of miR-26a significantly increased the expression of adipogenic genes related to peroxisome proliferator-activated receptor gamma (PPARγ), fatty acid synthase (FAS) and lipoprotein The mRNA levels of lipoprotein lipase (LPL) (P <0.01) and the protein levels of PPARγ and FAS were also significantly increased (P <0.05). At the same time, intracellular lipid accumulation is also significantly increased. In contrast, inhibition of miR-26a expression reduced the differentiation of 3T3-L1 cells. MiR-26a was shown to directly bind to the 3’UTR of homologous phosphatase and tensin homolog (PTEN) mRNA and to induce its miRNA and protein levels through target gene prediction and dual luciferase reporter system analysis Decreased (P <0.05). The results of this study indicate that miR-26a may positively regulate adipose differentiation of 3T3-L1 cells through direct inhibition of PTEN, providing a theoretical basis for miRNAs to become drug target molecules for the treatment of obesity-related diseases.