HMGA1 Difference In 2-cell Embryo Blastomeres Sister Mouse Influence The Selection Of The Embryonic Cell Lineages

The differentiation of inner cell mass and trophoblast is considered to be one of the key events in the early embryonic development of mammals.The nourishing ectoderm will develop into the extraembryonic structure of the fetus,while the inner cell mass will develop into the embryo and part of the extraembryonic structure.According to the pedigree tracking experiment,the differentiation of this cell lineage can be traced back to the 2-cell stage.HMGA1 is a member of the non-histone high mobility family protein family.As a structural transcription factor,it binds to the A-T rich DNA sequences and participates in biological processes such as enhancer formation,chromatin remodeling and transcriptional regulation.HMGA1 also plays a role in many cell processes,including cell growth and differentiation.Early studies have shown that there is little or no expression of HMGA1 in normal mouse tissues,but the expression level is very high in the early preimplantation embryo development.So it is inferred that HMGA1 may play an important role in the early preimplantation embryo development.According to the results of sequencing analysis,it was found that HMGA1 was differentially expressed in mouse embryonic 2-cell stage.More studies have shown that in human embryonic stem cells,HMGA1 regulates the expression of pluripotent genes Sox2,Oct4 and Nanog,in which Sox2 is an important transcription factor in the inner cell mass.This lays a theoretical foundation for studying whether the differential expression of HMGA1 in sister blastomeres of mouse 2-cell embryos regulates the distribution of mouse embryonic cell lineages.Firstly,fluorescence quantitative PCR(q PCR)and immunofluorescence staining were used to detect the expression and localization of HMGA1 in mouse preimplantation embryos.The results showed that the expression of(m RNA)Hmga1 in MII and zygotic stage was significantly higher than that in 2-cell stage,and slightly increased in morula and blastocyst stage.HMGA1 protein was evenly distributed in MII oocytes,located in the nucleus from zygote to blastocyst stage after fertilization,remained relatively stable before 8-cell,and then decreased.Then immunofluorescence staining,in situ hybridization and q PCR,were used to detect the distribution of HMGA1 in mouse blastomeres at 2-cell and 4-cell stage at RNA and protein levels.The results showed that there were differences in the distribution of HMGA1 in the blastomeres of 2-cell stage and 4-cell stage.EGFP was used as a marker to further study whether HMGA1 regulates the differentiation of mouse embryonic cell lineage.After injection of Hmga1 si RNA and(m RNA)EGFP,and injection of control interference fragments and(m RNA)EGFP,blastocysts were collected for immunofluorescence staining,and the location relationship between EGFP and inner cell mass and trophoblast cells in the three groups was counted.The statistical results showed that interfering with the expression of HMGA1 in single blastomeres reduced its contribution to the differentiation of inner cell mass.This shows that the differential distribution of HMGA1 in the blastomeres of mouse 2-cell embryos affects the selection of mouse embryonic cell lineages.To explore whether HMGA1 regulates the distribution of mouse embryonic cell lineages by affecting the expression of pluripotent genes.The Hmga1 si RNA were transfected into F9 cells at the cellular level.q PCR results showed that the Oct4 and Sox2 in the interference group were significantly lower than those in the control group.In order to further study whether HMGA1 affects the expression of pluripotent genes in embryos.By injecting Hmga1 si RNA,the c DNA of 4-cell and 8-cell embryos after zygote injection was obtained.The results of q PCR showed that Sox2 decreased significantly after interfering with Hmga1 at 4-cell and 8-cell stage.Immunofluorescence staining also showed that the SOX2 of the interference group was significantly lower than that of the control group.To sum up,the asymmetric distribution of HMGA1 in sister blastomeres of mouse 2-cell and 4-cell embryos was detected at the level of m RNA and protein,and it was verified that HMGA1 could affect the selection of cell lineage of mouse early embryos,which laid an important theoretical foundation for exploring the mechanism of cell pedigree selection of early mouse embryos before implantation.