生长分化因子-9在大鼠卵巢中的表达

目的：探讨生长分化因子-9(GDF-9)在大鼠卵巢中的表达部位。方法：选择不同发育时期的大鼠卵巢,采用免疫组化和Western blot技术检测大鼠卵泡各发育阶段GDF-9蛋白表达情况及分布规律。结果：免疫组化方法显示,出生0～2d卵巢中未见GDF-9表达,出生3d卵巢中见部分原始卵泡卵母细胞GDF-9表达阳性,以后从初级卵泡到发育成熟的卵泡卵母细胞均明显可见GDF-9蛋白表达。Western blot技术进一步提示,出生0-2d卵巢中无GDF-9蛋白形成,第3d开始有GDF-9蛋白产生,以后在卵巢发育的各时期均有GDF-9蛋白分泌。结论：GDF-9在卵巢发育的早期即开始产生,以后在各阶段卵巢中持续表达,提示卵母细胞通过产生GDF-9等因子对卵泡的发育进行调节。     

短额负蝗卵子发生及卵母细胞凋亡的组织学观察

用组织学方法对短额负蝗卵子发生和卵母细胞凋亡进行了显微观察.根据卵母细胞的大小、形态、胞核的变化、卵黄物质的形成以及滤泡细胞的形态变化等特征,将短额负蝗卵子发生分为3个时期8个阶段,并描述了各个阶段卵母细胞和滤泡细胞的形态特征.卵子发生过程中卵母细胞凋亡现象主要发生在卵母细胞生长期,特征主要表现为滤泡细胞向卵母细胞质内分裂增殖,卵母细胞的胞质不均匀,出现许多空泡状物质.     

牛体外受精卵发育培养条件的研究

将采自屠宰母牛卵巢的未成熟卵子在含有促进性腺激素和胎牛血清的TCM199培养基内培养成熟后,使用以钙离子载体Ionophore A23187诱导获能的牛精子进行授精处理.在受精卵发育用培养基中添加了谷氨酰胺,并以此与无添对照进行了对比,探索了培养基内谷氨酰胺的存在与否对胚胎发育的影响.另外,在发育用培养基内还分别添加了胎牛血清(FCS),阉牛血清(NSS)以及发情母牛血清(OCS)以观察其培养效果,结果表明,谷氨酰胺对胚胎的发育有极大的促进作用,添加谷氨酰胺的处理组与对照组中发育为桑椹胚、囊胚的胚胎比例分别为23.4%和6.6%,具有极显著的整异.而在含有FCS、NSS和OCS的TCM199培养基内培养的卵子中分别有81.4%,70.7%和82.9%发育为二细胞期胚胎,将胚胎继续培养至授精后第8天时,三个处理组中发育为桑椹胚、囊胚的胚胎比例分别为30.0%,40.2%和47.1%,以OCS的培养效果为最佳.本研究的结果证明,谷氨酰胺和发情母牛血清对牛体外受精卵的体外发育具有良好的促进作用.     

鲤鱼卵母细胞早期发育的细胞学观察

处在配丝期的鲤鱼卵母细胞,划分细线、偶线、凝线、粗线和双线五个发育时相。本文重点阐明和分析了各时相卵母细胞的染色体行为,形态以及灯刷染色体的形成和弥散过程。     

Reconstruction of human embryos derived from somatic cells

Reconstruction of human nuclear transfer embryos is a necessary step of therapeutic cloning. In this study we injected somatic cell nuclei into M Ⅱ oocytes and activated reconstructed oocytes with calcium ionophore A23187 (CaA) and 6-dimethylaminopurine (6-DMAP). After oocyteactivation and 2PN formation, we removed the female PN.By using this method, we avoided the application of DNA fluorescent stain and ultraviolet light for oocyte enucleation,and over elimination of ooplasm was also mitigated. Some reconstructed embryos developed into the blastocyst stage in vitro.     

动物卵子膜凝集素受体及其在生殖中的功能

凝集素及其受体是一新研究热点 ,卵子膜凝集素受体参与受精过程 .本文从卵子膜凝集素受体的结构分布特点、积累和功能 (精卵识别和结合 ,促有丝分裂 )三方面概述了卵子膜凝集素受体的研究进展 .     

MAPK regulates cell cycle progression in pig oocytes and fertilized eggs

Oocytes collected from prepubertal gilt ovaries were matured in vitro (IVM), and fertilized in vitro (IVF) or electrically activated. Phosphorylation of mitogenactivated protein kinase (MAPK) was detected with SDS-PAGE and Western blotting, and translocation of ERK₂ was observed with immunofluorescent cytochemistry. We found that the quantity of MAPK kept unchanged during oocyte maturation. There was no phosphorylated MAPK in porcine oocytes at the germinal vesicle (GV) stage; a little MAPK was phosphorylated at 20 h of IVM; a high level phosphorylation was detected at 30 h, while MAPK phosphorylation decreased at 36 h; and then MAPK phosphorylation increased again to the peak level from 40 to 60 h. ERK2 translocated from the peripheral cytoplasm to inner cytoplasm and nuclear area during oocyte maturation. There was nearly no phosphorylated MAPK at 18 and 20 h of electrically activated oocytes, but phosphorylation increased at 22 h. There was no phosphorylated MAPK at 12 h of IVF, while phosphorylation resumed at 16 h. These results suggest that MAPK may play an important regulatory role in MI-MII transition, pronucleus formation and the initiation of the first mitosis in pig eggs.     

基于USB的卵母细胞电压钳同步数据采集和分析软件设计

介绍了基于USB接口的卵母细胞电压钳系统的数据采集分析系统,应用Delphi语言开发了系统的人机操作实验界面,探讨了电压钳的USB同步传输、数据采集和和显示的实现方法.在细胞模型测试中,电压钳放大器的跨膜电流分辨达到了0.01nA,噪声测试低于10pA.在非洲爪蟾卵母细胞表达Kv4.2钾通道实验中,记录跨膜电流波形清晰,数据分析可靠.与国外同类仪器相比:该系统操作简单,价格低,数据采集和分析方便,适用于卵母细胞表达实验.     

Effects of different nuclear transfer and activation methods on the development of mouse somatic cell cloned embryos

A group of adult somatic cell cloned mice were obtained by using cumulus cells as nuclei donor cells. To study the effect of different nuclear transfer (NT) and activation methods on the development of mouse cloned embryos, embryos were reconstructed using two traditional NT methods (electrofusion and direct injection) and four activation treatments (electric pulse, ethanol, SrCl2 and electric pulse combined with SrCl2). The data showed that the efficiency of reconstruction using the direct injection method is significantly higher (90.7%) than that of the electrofusion method (49.7%). Parthenogenetic embryos can develop to blastocyst stage with three activation conditions, including ethanol, electric pulse and SrCl2; however, the rates of development to blastocyst after ethanol and electric pulse acti-vation (52.4%, 54.2%) are significantly lower than after SrCl2 activation (76.9%). Treatment of embryos for 6 h with 10 mmol/L SrCl2 was found to be the best condition for activation of parthenogenetic as well as reconstructed embryos. By contrast, reconstructed embryos failed to develop to blastocyst stage after being activated by ethanol. The use of either injection or electrofusion for embryo reconstruction affected the pre-implantation development. However, after transfer in pseudopregnant mice, cloned mice were obtained from both methods.     

Lipid characterization of individual porcine oocytes by dual mode DESI-MS and data fusion

The development of sensitive measurements to analyze individual cells is of relevance to elucidate specialized roles or metabolic functions of each cell under physiological and pathological conditions. Lipids play multiple and critical roles in cellular functions and the application of analytical methods in the lipidomics area is of increasing interest. In this work, in vitro maturation of porcine oocytes was studied. Two independent sources of chemical information (represented by mass spectra in the positive and negative ion modes) from single oocytes (immature oocytes, 24-h and 44-h in vitro matured oocytes) were acquired by using desorption electrospray ionization-mass spectrometry (DESI-MS). Low and mid-level data fusion strategies are presented with the aim of better exploring the large amount of chemical information contained in the two mass spectrometric lipid profiles. Data were explored by principal component analysis (PCA) within the two multi-block approaches to include information on free fatty acids, phospholipids, cholesterol-related molecules, di- and triacylglycerols. After data fusion, clearer differences among immature and in vitro matured porcine oocytes were observed, which provide novel information regarding lipid metabolism throughout oocyte maturation. In particular, changes in TAG composition, as well as increase in fatty acid metabolism and membrane complexity were evidenced during the in vitro maturation process. This information can assist the improvement of in vitro embryo production for porcine species.