HCG在小鼠卵母细胞体外受精胚胎发育过程中的作用  

Effect of HCG in culture medium on development of mouse embryos in vitro

作　　者：邢雅欣 解其贵[2] 杨志勇 刘玉兵 王玲[2] 千日成 XING Ya-xin;XIE Qi-gui;YANG Zhi-yong;LIU Yu-bing;WANG Ling;CHIAN Ri-cheng(School of Medicine,Nantong University,Nantong 226000;Center for Reproductive Medicine,Shanghai Tenth People ＇ s Hospital,Shanghai 200072)

机构地区：[1]南通大学医学院,南通226000 [2]上海市第十人民医院生殖医学中心,上海200072

出　　处：《生殖医学杂志》2018年第10期1000-1005,共6页Journal of Reproductive Medicine

基　　金：国家重点研发计划资助(2017YFC1002003;2017YFC1001601)

摘　　要：目的探讨HCG在小鼠卵母细胞体外受精胚胎发育过程中的作用。方法选择性成熟4~6周ICR雌性小鼠进行促排卵,体外受精并培养至96h。在受精液及胚胎培养液中分别加入HCG浓度为0U/L(对照组)和1 500U/L(HCG组),对两组的体外受精率、2-细胞分裂率及囊胚形成率进行比较;用实时定量PCR法(qPCR)比较两组4个囊胚质量相关基因mRNA的表达水平,包括多能性相关基因八聚体结合蛋白4(Oct4)、分化相关基因尾端型同源盒2(Cdx2)、氧化应激相关基因锰超氧化物歧化酶(MnSOD)、侵袭相关基因基质金属蛋白酶9(Mmp9);采用免疫荧光法,对两组囊胚进行Cdx2蛋白标记,并用DAPI染色料标记所有细胞的细胞核,在激光共聚焦显微镜下对细胞总数、内细胞团(ICM)、外滋养层(TE)细胞数目进行计数并比较。结果HCG组的受精率(95.6%)、2-细胞率(96.6%)以及囊胚形成率(83.9%)与对照组(分别为95.8%,96.0%,81.6%)相比较,差异无统计学意义(P>0.05);HCG组相关基因Cdx2、MnSOD mRNA的表达水平显著高于对照组(P<0.05);与对照组相比,HCG组Mmp9基因mRNA表达水平显著上调(P<0.01);两组的Oct4基因mRNA表达水平无显著性差异(P>0.05);HCG组囊胚细胞总数(61.7±16.3)显著高于对照组(52.3±9.1)(P<0.05),HCG组ICM细胞数(15.4±3.8)及TE细胞数(46.2±13.1)多于对照组[分别为(13.2±3.9)和(39.1±7.8)],但差异无统计学意义(P>0.05);两组ICM/TE值相比较,差异也无统计学意义(P>0.05)。结论胚胎培养液中加入HCG增加囊胚细胞总数及某些质量相关基因的表达,提高体外发育潜能,HCG具有潜力成为优化植入前胚胎培养体系的有益成分。Objective： To study the effect of HCG in culture medium on the development of mouse embryos in Methods： The ICR female mice（4 6 weeks of age）were super ovulated. Oocytes were fertilized and embryos were cultured for 96 hours. The fertilization and embryonic development media were supplemented with HCG with 0 U/L（control group）and 1500 U/L（HCG group）respectively. The in vitro fertilization rate,2 cell division rate and blastocyst formation rate were compared between the two groups. Real time quantitative PCR（qPCR）was used to compare the expression levels of four blastocyst quality re mRNAs,including pluripotency related gene octamer binding protein 4（Oct4）, differentiation re ated gene ated gene tail -type homeobox 2（Cdx2）, oxidative stress related gene manganese superoxide dismutase（MnSOD）, and invasion related gene matrix metalloproteinase 9（Mmp 9）. The blastocysts were labeled with Cdx2 protein and the nucleus of all cells was labeled with DAPI staining by immunofluorescence. The total cell numbers of blastocyst,inner cell mess （ICM） and trophectoderm（TE）were counted by laser confocal microscope respectively. Results： The fertilization rate（95.6~）,2 cell rate（96.6~）and blastocyst formation rate（83.9~）in the HCG group were not significantly different from those in the control group（95.8~, 9G. 0~, 81. 6%, respectively） （P 〈 0.05）. The expression levels of Cd：c2 and MnSOD mRNA in HCG group were significantly higher than those in control group（P〈0.05）. Compared with the control group, the mRNA expression level of Mmp 9 gene in HCG group was significantly up regulated（P〈0.01）. There was no significant difference in the mRNA expression level of Oct4 gene between the two groups（P〉0.05）. The total cell number of blastocyst in HCG group （61.7 ± 16.3）was significantly more than control group （52.3±9.1） （P〈0.05）. The number of ICM cells（15.4±3.8）and TE cells（46.2±13.1）in the HCG group were higher than those i

关 键 词：人绒毛膜促性腺激素 小鼠胚胎 体外发育 

分 类 号：R714.8[医药卫生—妇产科学]