长链非编码RNA U90926在调节3T3-L1前脂肪细胞分化中的作用及可能机制
The role and possible mechanism of lncRNA U90926 in modulating 3T3-L1 preadipocyte differentiation.
作者信息
Chen J, Liu Y, Lu S, Yin L, Zong C, Cui S, Qin D, Yang Y, Guan Q, Li X, Wang X
机构信息
Department of Cell Biology, Shandong University School of Medicine, Jinan, Shandong, China.
Department of Endocrinology, Qingdao Municipal Hospital, Qingdao, Shandong, China.
出版信息
Int J Obes (Lond). 2017 Feb;41(2):299-308. doi: 10.1038/ijo.2016.189. Epub 2016 Oct 26.
BACKGROUND
Obesity is a risk factor for metabolic diseases, while preadipocyte differentiation or adipogenesis is closely related to obesity occurrence. Long noncoding RNAs (lncRNAs) are a unique class of transcripts in regulation of a variety of biological processes. Using cDNA microarray, we found lncRNA U90926 is negatively correlated with 3T3-L1 preadipocyte differentiation.
OBJECTIVE
The aim of this study was to explore the role of lncRNA U90926 (lnc-U90926) in adipogenesis and the underlying mechanisms.
METHODS
Quantitative real-time PCR (qPCR) was performed to determine lnc-U90926 expression in 3T3-L1 preadipocytes, differentiated adipocytes, and in adipose tissues form mice. RNA fluorescent in situ hybridization (FISH) was performed to determine the localization of lnc-U90926 in 3T3-L1 preadipocytes. The effects of lnc-U90926 on 3T3-L1 adipogenesis were analyzed with lentivirus-mediated gain- and loss-of-function experiments. Lipid accumulation was evaluated by oil red O staining; several adipogenesis makers were analyzed by qPCR and western blotting. Dual luciferase assay was applied to explore the transactivation of target genes modulated by lnc-U90926. All measurements were performed at least for three times.
RESULTS
Lnc-U90926 expression decreased along the differentiation of 3T3-L1 preadipocytes. In mice, lnc-U90926 is predominantly expressed in adipose tissue. Obese mice have lower lnc-U90926 expression in subcutaneous and visceral adipose tissue than non-obese mice. FISH results showed that lnc-U90926 was mainly located in the cytoplasm. Overexpression lnc-U90926 attenuated 3T3-L1 adipocyte differentiation as evidenced by its ability to inhibit lipid accumulation, to decrease the mRNA levels of peroxisome proliferator-activated receptor gamma 2 (PPARγ2), fatty acid binding protein 4 (FABP4) and adiponectin (AdipoQ) as well as to reduce the protein levels of PPARγ and FABP4 (P<0.05). Knockdown of lnc-U90926 showed opposite effects, which increased mRNA expression of PPARγ2, FABP4, CCAAT/enhancer-binding proteinα (C/EBPα) and AdipoQ.
CONCLUSION
Lnc-U90926 attenuates 3T3-L1 adipocyte differentiation via inhibiting the transactivation of PPARγ2 or PPARγ.
背景
肥胖是代谢性疾病的一个风险因素，而前脂肪细胞分化或脂肪生成与肥胖的发生密切相关。长链非编码RNA（lncRNA）是一类独特的转录本，参与调控多种生物学过程。通过cDNA微阵列，我们发现lncRNA U90926与3T3-L1前脂肪细胞分化呈负相关。
目的
本研究旨在探讨lncRNA U90926（lnc-U90926）在脂肪生成中的作用及其潜在机制。
方法
采用定量实时PCR（qPCR）检测lnc-U90926在3T3-L1前脂肪细胞、分化的脂肪细胞以及小鼠脂肪组织中的表达。采用RNA荧光原位杂交（FISH）检测lnc-U90926在3T3-L1前脂肪细胞中的定位。通过慢病毒介导的功能获得和功能缺失实验分析lnc-U90926对3T3-L1脂肪生成的影响。用油红O染色评估脂质积累；通过qPCR和蛋白质印迹分析几种脂肪生成标志物。应用双荧光素酶报告基因检测法探讨lnc-U90926对靶基因的反式激活作用。所有测量至少进行3次。
结果
lnc-U90926的表达随着3T3-L1前脂肪细胞的分化而降低。在小鼠中，lnc-U90926主要在脂肪组织中表达。肥胖小鼠皮下和内脏脂肪组织中lnc-U90926的表达低于非肥胖小鼠。FISH结果显示lnc-U90926主要位于细胞质中。lnc-U90926过表达减弱了3T3-L1脂肪细胞的分化，这表现为其抑制脂质积累的能力、降低过氧化物酶体增殖物激活受体γ2（PPARγ2）、脂肪酸结合蛋白4（FABP4）和脂联素（AdipoQ）的mRNA水平以及降低PPARγ和FABP4的蛋白质水平（P<0.05）。lnc-U90926敲低则表现出相反的效果，增加了PPARγ2、FABP4、CCAAT/增强子结合蛋白α（C/EBPα）和AdipoQ的mRNA表达。
结论
lnc-U90926通过抑制PPARγ2或PPARγ的反式激活作用减弱3T-L1脂肪细胞分化。
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