Implication of budding uninhibited by benomyl gene in spontaneous abortion embryos with numerical chromosomal abnormality / 中华检验医学杂志

ABSTRACT

Objective To explore the mechanism of hBUB1 gene in the developing of spontaneous abortion embryos with numerical chromosomal abnormalitv.Methods Quantitative real-time RT-PCR and Western blot were used to determine the mRNA and protein level of hsMAD2 gene both in spontaneous abortion embryos with numerical chromosomal abnormality(experimental group)and with numerical chromosomal normality(control group).Recombinant shRNA plasmids targeting hBUB1 gene was constructed to inhibit the expression of endogenous hBUB1 genes in embryonic cells.Interference efficiency was demonstrated by fluorescent quantitative PCR and Western blot.The inhibitory rate of cell proliferation was measured by MTT assay and cells-cycle was assessed by flow cytometry.Resuns Western blot analysis showed that protein level of hBUB1 in the experimental group was decreased signifieandv(the rate of positivity and strong positivity were 8%and 93.5%,respectively,P＜0.05)compared with the control group.The expression of hBUB1 gene in embryonic cells was significantly and specially inhibited by shRNA plasmids (the mRNA level before and after treatment witll RNAi were 0.196±0.067 and 0.042±0.006,respectively,P＜0.05).The inhibitory rate of cell proliferation was increased to 62%from 4%at 48 h after transfection.The rate of G2/M phase cells was decreased after transfection with efficient shRNA(control group40.2%and 41.3%,test group21.3%).Conclusions Down-regulation of hBUB1 gene leads to the inhibition of cell proliferation and the arrest of cell-cycle.It also probably plays an important role in the development of spontaneous abortion embryos with numerical chromosomal abnormality.The clinical relevance warrant further investigation.