Bovine cumulus-oocyte disconnection in vitro

Summary Cumulus-oocyte complexes were obtained from cows by aspiration of small (1–6 mm in diameter) antral follicles after slaughter. Complexes with a compact multilayered cumulus investment were cultured and processed for transmission electron microscopy after different periods of culture including a 0 h control group. In 0 h control oocytes the cumulus cells had numerous projections which penetrated the zona pellucida and established gap junctions with the oolemma. A partial loss of these junctions was noticed as an early event of oocyte maturation occurring within the first 3 h of culture. A low frequency of gap junctions was maintained until 12–18 h of culture where the junctional contact was completely disrupted. This decrease in intercellular communication was parallelled by resumption of oocyte meiosis.     

Postnatal formation of the blood-testis barrier in the rat with special reference to the initiation of meiosis

Summary Postnatal formation of the Blood-Testis Barrier (BTB) in the rat was studied by either fixation in hypertonic fixative or employing lanthanum tracer. After 15 days of age, meiosis has reached different stages of spermatogenesis in differnt zones of the seminiferous cords. Only in those parts where germ cells are in the pachytene stage of meiosis do Sertoli cells form an effective barrier or tight compartment. Between 16 and 19 days of age, final formation of the BTB, which is to be found in the adult rat testis, occurs by zygotene and then leptotene stages successively entering the tight compartment. Thus, formation of a BTB by Sertoli cells does not occur synchronously along the length of the seminiferous cord but in accordance with the stage of meiosis of the associated germ cells.     

Meiotic differentiation during colony maturation in Saccharomyces cerevisiae

As yeast colonies ceased growth, cells at the edge of these colonies transited from the cell division cycle into meiosis at high efficiency. This transition occurred remarkably synchronously and only at late stages of colony maturation. The transition occurred on medium containing acetate or low concentrations of glucose, but not on medium containing high glucose. The repression by high glucose was overcome when IME1 was overexpressed from a plasmid. Experiments with different growth media imply that meiosis in colonies is triggered by changes in the nutrient environment as colonies mature. HAP2 is required to sporulate in any carbon source, whereas GRR1 is required for glucose repression of sporulation. CLN3 is required to repress meiosis in colonies but not in liquid cultures, indicating that the regulators that mediate the transition to meiosis in colonies are not identical to the regulators that mediate this transition in liquid cultures.     

女性生殖细胞发育过程的分子遗传学研究进展

女性生殖细胞的发育主要经历胚胎发育期和青春期后的卵泡发育期,其中有丝分裂-减数分裂转换、减数分裂阻滞和再激活是发育过程中的关键阶段。各发育阶段独具特征性分子事件,其顺序发生亦需严密的基因调控及与性腺体细胞的交互作用。近年来,单细胞转录组学研究揭示了生殖细胞及性腺体细胞的阶段特异性表达基因、信号传导通路及表观遗传学变化,为深入理解生殖细胞发育过程、阐明相关疾病的发病机制以及促进生殖遗传研究成果的临床转化提供了科学依据和理论基础。     

卵母细胞成熟障碍4例与同期卵母细胞大部分不成熟患者比较分析

目的分析4例体外受精-胚胎移植（IVF—ET）患者共8个辅助助孕周期,全部73个卵母细胞均处于GV、MetaphaseI期的可能原因。并与同期13例IVF—ET患者比较分析,比较组患者〉50％卵母细胞处于GV、MetaphaseI期。方法回顾分析本中心12年辅助助孕工作中出现的4例患者共8个周期所获卵母细胞均处于GV、MI的临床及实验室资料,并与同期13例超过50％卵母细胞处于GV、MI患者的临床、实验室资料进行比较。结果4例患者73个卵母细胞均处于GV或MI期,经体外成熟培养,24,48,72h仍无极体排出,停滞于GV、MI期。同期13例患者大部分卵母细胞不成熟,但体外培养后部分卵母细胞可进一步成熟,并可受精,获得妊娠。结论细胞和遗传机制引起卵母细胞成熟障碍,现有体外成熟培养方法尚无法促其成熟,目前赠卵是该类患者获得妊娠可供选择的助孕方法。但对于在控制性超促排卵中,出现大部分卵母细胞不成熟的患者可以通过延长促超排时间,增大hCG注射日卵泡直径,体外成熟培养等方法获得成熟的卵母细胞,获得妊娠。     

核质置换技术治疗线粒体疾病的研究进展

线粒体疾病是线粒体基因组发生基因突变所导致的一类遗传疾病,目前尚无治愈方法。近年核质置换技术的出现为线粒体疾病的治疗提供了新思路。现有的核质置换技术包括生发泡移植、纺锤体移植、极体移植和原核移植技术,其中生发泡移植涉及的体外成熟可影响卵母细胞的发育潜能;纺锤体移植最大的挑战是纺锤体组装对机械刺激敏感,缺乏核膜包绕,操作过程可能对纺锤体造成损伤;极体移植违背了自然选择过程,其远期影响尚不明了;原核移植操作方便、基础研究证据充分,具有短期内向临床转化的潜能,但使用已经受精的合子作为胞质供体面临的伦理争议较大。本文就4种核质置换技术治疗线粒体疾病的应用现状进行综述。     

The regulation of maturation promoting factor during prophase I arrest and meiotic entry in mammalian oocytes

Mammalian oocytes arrest at prophase of meiosis I at around birth and they remain arrested at this stage until puberty when the preovulatory surge of luteinizing hormone (LH) causes ovulation. Prophase I arrest in the immature oocyte results from the maintenance of low activity of maturation promoting factor (MPF), which consists of a catalytic subunit (CDK1) and regulatory subunit (cyclin B1). Phosphorylation-mediated inactivation of CDK1 and constant degradation of cyclin B1 keep MPF activity low during prophase I arrest. LH-mediated signaling manipulates a vast array of molecules to activate CDK1. Active CDK1 not only phosphorylates different meiotic phosphoproteins during the resumption of meiosis but also inhibits their rapid dephosphorylation by inhibiting the activities of CDK1 antagonizing protein phosphatases (PPs). In this way, CDK1 both phosphorylates its substrates and protects them from being dephosphorylated. Accumulating evidence suggests thatthe net MPF activity that drives the resumption of meiosis in oocytes depends on the activation status of CDK1 antagonizing PPs. This review aims to provide a summary of the current understanding of the signaling pathways involved in regulating MPF activity during prophase I arrest and reentry into meiosis of mammalian oocytes.     

人类卵母细胞减数分裂进程及形态学研究

目的对人类未成熟卵母细胞进行体外培养,观察卵母细胞直径和卵周隙随卵母细胞成熟所发生的形态学改变,并按照其染色体变化确定人类卵母细胞减数分裂进程。方法穿刺收集多囊卵巢患者未成熟卵母细胞进行体外培养成熟,分别于体外培养4、8、12、16、20、24和28h进行卵母细胞直径及最大卵周隙测量,然后行染色体形态学观察并进行分析。结果未成熟卵母细胞体外培养10h约有50％发生生发泡破裂（GVBD）恢复减数分裂;减数分裂Ⅰ（MⅠ）前中期由培养8h的31．37％上升到16h的45％,然后下降;MⅠ中期（Meta-MⅠ）在培养20h进入高峰,占到40．63％,MⅠ后、末期（Ana／tel-MⅠ）维持时问相对较短,24h后多数已进入减数分裂Ⅱ（MⅡ）成熟期,28h未成熟卵母细胞体外培养的成熟率为68．85％。另外发现随着卵母细胞的体外成熟,其直径不会发生明显改变,但卵周隙（PVS）却在逐步增加。结论按照常规的体外培养方法可大致确定人类未成熟卵母细胞的体外成熟进程,且认为卵周隙的扩大可作为卵母细胞成熟分期的另一标志。     

外源性化学物在卵母细胞成熟中的作用

系统综述了外源性化学物对卵母细胞减数分裂成熟的影响及其作用机制。有研究表明,外源性化学物中的双酚丙烷、对叔辛基苯酚、邻苯二甲酸酯类、有机氯类、金属类等都可干扰正常卵母细胞的成熟过程,但各种外源性化学物的作用机制不尽相同。     

Identification of meiotic anomalies with multiplex fluorescence in situ hybridization: Preliminary results

OBJECTIVE: To characterize meiotic anomalies in infertile men by multiplex fluorescence in situ hybridization (M-FISH) and to determine whether synaptic problems affect specific bivalents or whether anomalies are random. DESIGN: Analysis of meiotic preparations with standard techniques and M-FISH. SETTING: Assisted reproduction centers and Universitat Autònoma de Barcelona. PATIENT(S): Three fertile men undergoing vasectomy, four sterile patients with oligoasthenoteratozoospermia, and one patient with a Robertsonian translocation t(13;14). INTERVENTION(S): Unilateral testicular biopsy in controls and patients with oligoasthenoteratozoospermia and collection of a semen sample from the translocation carrier. MAIN OUTCOME MEASURE(S): Identification of bivalents in metaphase I and chromosomes in metaphase II and characterization of chromosome abnormalities. RESULT(S): All bivalents in metaphase I and all chromosomes in metaphase II could be identified. In controls and in one patient with oligoasthenoteratozoospermia, meiosis was normal. Other patients with oligoasthenoteratozoospermia showed different types of anomaly: desynapsis, breaks, precocious XY separation, or cryptic reorganizations. The Robertsonian translocation t(13;14) was easily identified. CONCLUSION(S): Results confirm the high incidence of synaptic errors in oligoasthenoteratozoospermia patients. Bivalents in metaphase I and chromosomes in metaphase II were individually identifiable. Nondisjunctional errors or small reorganizations overlooked in classic meiotic preparations were identified. Synaptic anomalies seem to affect meiotic bivalents at random.