背景:代谢稳态与细胞命运决定和胚胎发育的早期损害密切相关。雷帕霉素的蛋白激酶机制靶标(mTOR)是体内细胞代谢的关键调节剂。早期胚胎中mTOR信号的抑制导致种植后发育失败,然而,机制仍然知之甚少。
方法:分别在体内和体外用mTOR抑制剂处理妊娠小鼠和植入前小鼠胚胎,随后检查了胚泡的形成,植入,和植入后的发展。我们用免疫荧光染色,RNA-Seqsmart2和全基因组亚硫酸氢盐测序技术,以研究mTOR抑制剂对质量的影响,细胞命运决定,和发育中胚胎的分子改变。
结果:我们发现植入前mTOR抑制降低了胚泡形成率和植入能力,损害植入后胚胎发育。我们发现阻断mTOR信号传导会对8细胞胚胎向胚泡的转化产生负面影响,并导致胚泡质量的各种缺陷。这些包括滋养外胚层细胞分化受损的问题,以及细胞命运规范的破坏。mTOR抑制显著影响胚胎的转录和DNA甲基化。用mTOR抑制剂治疗增加溶酶体活化并破坏胚泡中肌动蛋白细胞骨架的组织和动力学。
结论:这些结果表明,mTOR在8细胞到胚泡的转化中起着至关重要的作用,并在早期胚胎发育过程中保护胚胎质量。
BACKGROUND: Metabolic homeostasis is closely related to early impairment of cell fate determination and embryo development. The protein kinase mechanistic target of rapamycin (mTOR) is a key regulator of cellular metabolism in the body. Inhibition of mTOR signaling in early embryo causes postimplantation development failure, yet the mechanisms are still poorly understood.
METHODS: Pregnancy mice and preimplantation mouse embryo were treated with mTOR inhibitor in vivo and in vitro respectively, and subsequently examined the blastocyst formation, implantation, and post-implantation development. We used immunofluorescence staining, RNA-Seq smart2, and genome-wide bisulfite sequencing technologies to investigate the impact of mTOR inhibitors on the quality, cell fate determination, and molecular alterations in developing embryos.
RESULTS: We showed mTOR suppression during preimplantation decreases the rate of blastocyst formation and the competency of implantation, impairs the post implantation embryonic development. We discovered that blocking mTOR signaling negatively affected the transformation of 8-cell embryos into blastocysts and caused various deficiencies in blastocyst quality. These included problems with compromised trophectoderm cell differentiation, as well as disruptions in cell fate specification. mTOR suppression significantly affected the transcription and DNA methylation of embryos. Treatment with mTOR inhibitors increase lysosomal activation and disrupts the organization and dynamics of the actin cytoskeleton in blastocysts.
CONCLUSIONS: These results demonstrate that mTOR plays a crucial role in 8-cell to blastocyst transition and safeguards embryo quality during early embryo development.