巨噬细胞移动抑制因子在牛胚胎早期发育中的表达模式及作用初探

The Expression Pattern and Function of Macrophage Migration Inhibitory Factor(MIF) in Early Bovine Embryo Development

巨噬细胞移动抑制因子(macrophage migration inhibitory factor,MIF)在生殖过程中发挥重要作用,但其在早期胚胎发育中的作用尚未阐明。本研究通过RT-PCR、Western-blot和显微注射的方法对MIF在牛胚胎早期发育中的表达模式和功能进行初探。研究结果显示:MIF基因和MIF蛋白在牛体外受精(in vitro fertilization,IVF)胚胎和体细胞核移植(somatic cell nuclear transfer,SCNT)胚胎早期发育过程中持续性表达,MIF基因表达量较低,在SCNT胚胎2细胞、4细胞、8细胞和囊胚期的表达显著高于IVF胚胎,但MIF蛋白在SCNT胚胎中的表达量显著低于IVF胚胎;干扰MIF的表达后,胚胎8细胞期的阻滞率(51.2%vs.29.5%,P<0.05)显著升高,囊胚率(17.3%vs.36.5%,P<0.05)和囊胚细胞数(85.6±9.3 vs.110.2±10.5,P<0.05)显著降低,2细胞期和4细胞期胚胎的还原型谷胱甘肽(reduced glutathione,GSH)水平也显著降低,提高MIF干扰组胚胎GSH水平可以显著改善胚胎的囊胚率和囊胚细胞数(16.9%vs.32.9%,P<0.05;85.4±8.6 vs.102.2±10.4,P<0.05)。研究结果初步表明,MIF蛋白在牛胚胎早期发育中发挥重要作用,可能通过调控胚胎GSH水平影响胚胎早期发育率及囊胚质量。

Macrophage migration inhibitory factor(MIF) plays an important role in the process of reproduction, but its functions in early embryonic development have not been clarified yet. The objective of this study was to explore the expression pattern and function of MIF in the early development of bovine embryos by RT-PCR, Western-blot and microinjection. The results showed that MIF mRNA and protein were consistently expressed at a low level during the early development of in vitro fertilized(IVF) bovine embryos. The expression of MIF mRNA was significantly higher in somatic cell nuclear transfer(SCNT) embryos than in their IVF counterparts at the 2-cell, 4-cell, 8-cell and blastocyst stages, but the expression of MIF protein was significantly lower in SCNT embryos. After the expression of MIF protein was inhibited by antibody injection,more embryos were developmentally arrested before the 8-cell stage(51.2% vs. 29.5%, P<0.05). The blastocyst rate(17.3% vs. 36.5%, P<0.05) and number(85.6±9.3 vs. 110.2±10.5, P<0.05) were significantly decreased,and the levels of reduced glutathione(GSH) in 2-and 4-cell stage embryos were also decreased significantly.Increasing the GSH level of embryos in the MIF interference group could significantly improve the blastocyst rate and the total cell number of blastocysts(16.9% vs. 32.9%, P<0.05;85.4±8.6 vs. 102.2±10.4, P<0.05).These results preliminarily indicated that MIF may play an important role in the early development of IVF bovine embryos, and affect the early embryonic development rate and blastocyst quality by regulating embryonic GSH levels.