微小RNA-29b在小鼠植入前胚胎中的表达及生物学功能

Expression and biological functions of microRNA-29b in mouse preimplantation embryos

目的研究微小RNA-29b(miR-29b)在植入前小鼠胚胎中的表达模式,探讨其在小鼠植入前胚胎发育中的生物学作用。方法采用实时定量PCR法检测小鼠植入前胚胎发育过程中miR-29b及其下游DNA甲基转移酶3a/3b(Dnmt3a/3b)的表达模式。采用显微注射商业化的miR-29b抑制剂的实验方法探讨其在早期胚胎中的功能。结果 miR-29b在小鼠二细胞阶段高表达,四细胞阶段开始表达下降,并在植入前维持较低的表达水平。Dnmt3a和Dnmt3b均在小鼠四细胞阶段表达量升高;其中,Dnmt3a在囊胚期表达降低并维持在较低的水平,而Dnmt3b在植入前均维持在较高的水平。miR-29b抑制剂抑制miR-29b功能后,小鼠胚胎从桑葚期到囊胚期发育明显阻滞。结论 miR-29b在小鼠植入前胚胎各阶段均有表达,呈现二细胞特异性高表达模式;Dnmt3a/3b在小鼠植入前胚胎中均有表达,且与miR-29b的表达呈现一定负相关性;miR-29b功能下调,引起植入前小鼠胚胎发育阻滞。

Objective To investigate the expression and biological functions of microRNA-29b （miR-29b） in the development of preimplantation embryo in mouse. Mehthods The quantitative RT-PCR method was used to detect the expression pattern of miR-29b and its downstream targets- DNA methyltransferase 3a/3b （Dnmt3a/3b） during mouse preimplantation embryonic development. Commercial miR-29b inhibitor microinjection was performed to study its function in preimplantation embryos. Results miR-29b showed a high expression at 2-cell embryo stage, then sharply decreased after 4-cell embryo stage, and subsequently kept a quite low level. The expressions of Dnrnt3a and Dnmt3b were increased distinctly in 4-cell embryo stage. The former was obviously reduced to a very low level at blastocyst, while the other one maintained a high level till blastocyst. The functional inhibition of miR-29b led to distinct developmental retardation in morula stage. Conclusion The miR-29b is expressed in all stages of preimplantation mouse embryos, which is the highest in 2-cell stage. Dnmt3a/3b is expressed in all stages of preimplantation mouse embryos and negatively correlated to miR-29b. Downregulation of nfiR-29b activity can cause the developmental retardation in mouse preimplantation embryos.