Effects of Pin1 antagonist on mouse preimplantation embryo development in vitro

Abstract

Abstract: Objective To elucidate the further function of Pin1 in mouse preimplantation embryos, We observed the distribution of Pin1 in mouse preimplantation embryos and the effect of Pin1 antagonist (Juglone) on the development of mouse preimplantation embryo in vitro. Methods The immunofluorescence technique was used in the present study in order to localize the distribution of pin1 protein in the preimplanration mouse embryo. The 1-cell embryos of KM mice were cultured in KSOM culture medium supplemented various concentrations pin1 inhibitor (Juglone), respectively. The different development potential was observed under an inverted microscope. The mRNA expression levels of Sox2, Oct4, Klf4 and c-Myc were detected by Real-time PCR. Results Pin1 protein was distributed in both cytoplasm and nucleus of mouse preimplantation embryos. The immunofluorescent intensities in nuclei were higher than those in the complasma. The 1-cell embryos treated with 10μmol/L and 25μmol/L Juglone for 93h (from post hCG 27h to 120h) in vitro could hardly develop to 4-cell embryos (P<0.001). The 1-cell embryos treated with 25μmol/L Juglone for 18h (from post hCG 27h to 45h) in vitro were arrested at 2-cell stage (P<0.001). In 25μmol/L Juglone group, the Sox2 mRNA levels were significantly low compared with KSOM group (P<0.05), while the mRNA levels of Oct4, c-Myc and klf4 were no significant differences compared with KSOM group (P>0.05). Conclusion Pin1 is mainly distributed in the nucleus of mouse preimplantation embryos, the results suggest Pin1 can participate in gene transcription. The development rate of mouse preimplantatiom embryos from 2-cell to 4-cell is markedly decreased and the mRNA expression levels of stem cell factor (Sox2) are down-regulated by Juglone treatment, which indicates Pin1 may play a important role in the early development of mouse preimplantation embryos by regulating zygotic genome activation.

Key words: Pin1, Juglone, 2-cell block, Real-time PCR, Immunofluorescence staining

References

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