小鼠不同脂肪组织中Gata1的表达特点及其在白色脂肪棕色化中的作用研究
发布时间：2018-10-18 08:11
【摘要】：机体内脂肪主要有两类:与脂肪堆积有关的白色脂肪组织(White adipose tissue,WAT)和脂肪代谢有关的棕色脂肪组织(Brown adipose tissue,BAT)。前者通过甘油三酯的形式储存能量,与肥胖症,糖尿病和血管类疾病的发生密切相关,后者则通过氧化磷酸化代谢脂肪,负责能量的提供,可有效抗肥胖。有研究证实,在适当的刺激下,白色脂肪可以被转化为棕色脂肪,即白色脂肪棕色化。因此阐明白色脂肪棕色化的具体分子机制对于人类的健康具有重要意义。与脂肪发生有关的因子有很多,其中血管内皮生长因子(Vascular endothelial growth factor,VEGF-A)近年来被发现在脂肪组织中大量表达,并且与脂肪组织发育及能量代谢密切相关。我们建立了四环素-Vegfa可逆调节小鼠模型,并成功介导白色脂肪棕色化。在此进程中,除WAT和BAT关键调控因子Leptin,Prdm16,Ucp1,Cidea,Bmp4和Bmp7发生显著改变之外,Gata1也发生了显著性上调。过往对GATA1的报道主要集中于其在红系分化中的调控作用,在脂肪分化中的研究较少,然而其是否也在脂肪分化与能量代谢中发挥了重要调节作用正是我们本研究的主要内容。在本论文中,我们对小鼠不同脂肪组织间Gata1的表达特点及其在白色脂肪棕色化进程中的作用及相关机制进行了研究。首先我们对小鼠体内WAT和BAT进行了分离并分别提取RNA,Real-time PCR结果显示Gata1在BAT中的表达量要远远高于其在WAT中的表达,这与同时检测的棕色脂肪调控因子Prdm16、Ucp1和Cidea等的表达特点是一致的。继而利用构建好的四环素-Vegfa可逆调节小鼠模型,我们发现在白色脂肪棕色化的进程中Gata1发生了明显的上调,并且其表达与Vegfa呈现显著性的负相关。继而我们利用pc DNA3.1(-)载体及慢病毒载体pco EGFP-puro分别构建了适用于瞬时转染和慢病毒侵染的Gata1过表达质粒,并在不同细胞系中进行了瞬时转染和慢病毒侵染实验。Real-time PCR检测结果显示,不论在3T3-L1细胞系还是原代脂肪前体细胞中,外源过表达Gata1后均能够上调相同的棕色脂肪调控因子的表达,如Bmp4,Pgc1-α和Cidea;但是并不影响Bmp7、Prdm16和Ucp1的表达。我们的实验结果揭示在脂肪分化进程中,Gata1的表达与棕色脂肪发生及白色脂肪棕色化密切相关;其作用的发挥可能是通过上调Bmp4,Pgc1-α和Cidea等棕色脂肪调控因子的表达而实现的。GATA1在白色脂肪棕色化中的作用及其机制的阐明既补充了GATA1已有的生物学功能,为能量代谢研究提供了新的线索,同时也为预防和控制肥胖提供了一个潜在靶点。
[Abstract]:There are two main types of fat in the body: (White adipose tissue,WAT (white adipose tissue associated with fat accumulation) and (Brown adipose tissue,BAT (brown adipose tissue related to fat metabolism). The former is closely related to obesity, diabetes and vascular diseases by storing energy in the form of triglycerides, while the latter can effectively resist obesity by oxidizing phosphorylated metabolized fat, which is responsible for energy supply. Studies have shown that white fat can be turned into brown fat, or white fat, brown with appropriate stimulation. Therefore, it is important for human health to clarify the specific molecular mechanism of white fat browning. There are many factors related to adipogenesis, among which vascular endothelial growth factor (Vascular endothelial growth factor,VEGF-A) has been found to be highly expressed in adipose tissue in recent years, and is closely related to adipose tissue development and energy metabolism. We established a mouse model of reversible regulation of tetracycline-Vegfa and successfully mediated white fat browning. In addition to the significant changes in WAT and BAT key regulatory factors, Leptin,Prdm16,Ucp1,Cidea,Bmp4 and Bmp7, Gata1 also increased significantly during this process. Previous reports on GATA1 mainly focus on its regulatory role in erythroid differentiation, but less on adipose differentiation. However, whether GATA1 also plays an important regulatory role in adipodifferentiation and energy metabolism is the main content of this study. In this paper, we studied the expression of Gata1 in different adipose tissues of mice and its role in the brown process of white fat and its related mechanisms. First, we isolated WAT and BAT from mice and extracted RNA,Real-time PCR respectively. The results showed that the expression of Gata1 in BAT was much higher than that in WAT. This was consistent with the expression of brown fat regulatory factors Prdm16,Ucp1 and Cidea. Then we established a mouse model of reversible regulation of tetracycline-Vegfa. We found that Gata1 was significantly up-regulated in the course of brown white fat and its expression was negatively correlated with Vegfa. Then we constructed Gata1 overexpression plasmids suitable for transient transfection and lentivirus infection using pc DNA3.1 (-) vector and lentivirus vector pco EGFP-puro, and carried out transient transfection and lentivirus infection experiments in different cell lines. In both 3T3-L1 cell lines and primary adipose progenitor cells, exogenous overexpression of Gata1 could up-regulate the expression of the same brown fat regulatory factors, such as Bmp4,Pgc1- 伪 and Cidea;, but did not affect the expression of Bmp7,Prdm16 and Ucp1. Our results showed that the expression of Gata1 was closely related to brown adipogenesis and white fat browning during adipose differentiation. The role of GATA1 in the brown fat browning may be achieved by upregulating the expression of brown fat regulatory factors such as Bmp4,Pgc1- 伪 and Cidea. The role of GATA1 in the browning of white fat and its mechanism may complement the existing biological functions of GATA1. It provides a new clue for energy metabolism research and a potential target for the prevention and control of obesity.
【学位授予单位】：东北师范大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R589.2
本文编号：2278524
[Abstract]:There are two main types of fat in the body: (White adipose tissue,WAT (white adipose tissue associated with fat accumulation) and (Brown adipose tissue,BAT (brown adipose tissue related to fat metabolism). The former is closely related to obesity, diabetes and vascular diseases by storing energy in the form of triglycerides, while the latter can effectively resist obesity by oxidizing phosphorylated metabolized fat, which is responsible for energy supply. Studies have shown that white fat can be turned into brown fat, or white fat, brown with appropriate stimulation. Therefore, it is important for human health to clarify the specific molecular mechanism of white fat browning. There are many factors related to adipogenesis, among which vascular endothelial growth factor (Vascular endothelial growth factor,VEGF-A) has been found to be highly expressed in adipose tissue in recent years, and is closely related to adipose tissue development and energy metabolism. We established a mouse model of reversible regulation of tetracycline-Vegfa and successfully mediated white fat browning. In addition to the significant changes in WAT and BAT key regulatory factors, Leptin,Prdm16,Ucp1,Cidea,Bmp4 and Bmp7, Gata1 also increased significantly during this process. Previous reports on GATA1 mainly focus on its regulatory role in erythroid differentiation, but less on adipose differentiation. However, whether GATA1 also plays an important regulatory role in adipodifferentiation and energy metabolism is the main content of this study. In this paper, we studied the expression of Gata1 in different adipose tissues of mice and its role in the brown process of white fat and its related mechanisms. First, we isolated WAT and BAT from mice and extracted RNA,Real-time PCR respectively. The results showed that the expression of Gata1 in BAT was much higher than that in WAT. This was consistent with the expression of brown fat regulatory factors Prdm16,Ucp1 and Cidea. Then we established a mouse model of reversible regulation of tetracycline-Vegfa. We found that Gata1 was significantly up-regulated in the course of brown white fat and its expression was negatively correlated with Vegfa. Then we constructed Gata1 overexpression plasmids suitable for transient transfection and lentivirus infection using pc DNA3.1 (-) vector and lentivirus vector pco EGFP-puro, and carried out transient transfection and lentivirus infection experiments in different cell lines. In both 3T3-L1 cell lines and primary adipose progenitor cells, exogenous overexpression of Gata1 could up-regulate the expression of the same brown fat regulatory factors, such as Bmp4,Pgc1- 伪 and Cidea;, but did not affect the expression of Bmp7,Prdm16 and Ucp1. Our results showed that the expression of Gata1 was closely related to brown adipogenesis and white fat browning during adipose differentiation. The role of GATA1 in the brown fat browning may be achieved by upregulating the expression of brown fat regulatory factors such as Bmp4,Pgc1- 伪 and Cidea. The role of GATA1 in the browning of white fat and its mechanism may complement the existing biological functions of GATA1. It provides a new clue for energy metabolism research and a potential target for the prevention and control of obesity.
【学位授予单位】：东北师范大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：R589.2
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