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肥胖已经成为全球性的公共卫生问题,严重影响人类的身体健康和生活品质。本文运用CRISPR/Cas9技术,构建了肥胖相关基因——瘦素受体(leptin receptor,lepr)和黑素皮质素受体4(melanocortin-4 receptor,mc4r)的斑马鱼突变体,并对其进行形态和功能分析。结果显示,lepr~(-/-)和mc4r~(-/-)斑马鱼在胚胎和幼鱼期没有明显异常,但在成年期,lepr~(-/-)和mc4r~(-/-)斑马鱼相比对照进食增多、体重增加、体脂含量升高,呈现肥胖表型。血糖测定结果显示,饱食喂养条件可以诱导lepr~(-/-)和mc4r~(-/-)成年斑马鱼出现糖耐量受损的现象。进一步地,运用real time RT-PCR对76个能量代谢相关基因在lepr~(-/-)和mc4r~(-/-)斑马鱼肝脏中转录表达水平进行检测,并与Lep~(ob/ob)小鼠肝脏c DNA microarray的数据比较,发现胰岛素/胰岛素样生长因子信号转导通路(insulin/IGF signaling pathway,ISS)相关的基因在lepr~(-/-)斑马鱼和Lep~(ob/ob)小鼠肝脏中表达变化具有较高的正相关性,提示肥胖调控网络在进化中维持了一定的功能保守性。
Obesity has become a global public health problem that seriously affects human health and quality of life. In this study, zebrafish mutants with leptin receptor (lepr) and melanocortin-4 receptor (mc4r) were constructed using CRISPR / Cas9 technology Morphological and functional analysis. The results showed that there was no obvious abnormalities in the embryonic and juvenile stages of lepr ~ (- / -) and mc4r ~ (- / - Compared with the control, zebrafish increased food intake, weight gain, body fat content, showing obesity phenotype. Blood glucose measurements showed that satiety feeding induced impaired glucose tolerance in lepr ~ (- / -) and mc4r ~ (- / -) adult zebrafish. Further, the transcriptional level of 76 energy metabolism related genes in lepr ~ (- / -) and mc4r ~ (- / -) zebrafish liver was detected by real time RT-PCR and correlated with Lep ~ ) Mice liver c DNA microarray data and found that insulin / IGF signaling pathway (insulin / IGF signaling pathway, ISS) related genes in lepr ~ (- / -) zebrafish and Lep ~ (ob / ob) mouse liver expression has a high positive correlation, suggesting that obesity regulatory networks in the evolution of the maintenance of a certain degree of functional conservation.