非细胞体系小鼠卵母细胞游离生发泡减数分裂的启动

目的 :建立有效的哺乳动物卵母细胞生发泡的分离方法 ,并使之在构建的非细胞体系中发生减数分裂的重新启动。　方法 :采用直接取核法获取卵母细胞生发泡 ,用同步化的HeLa细胞裂解液构建非细胞体系 ,将游离的生发泡培养在裂解液中 ,并加入荧光染料Hoechest 33342 ,观察染色质的变化情况。　结果 :M期裂解液可以诱导生发泡的染色质出现凝集现象 ,而G2 期早期的裂解液无此现象发生。　结论 :用直接取核法可成功分离出游离的生发泡 ,且它们在体外培养条件下可以和体内一样发生染色质凝集现象     

Effects of body temperature on the expression and localization of meiosis-related proteins STRA8 and SCP3 in boar testes

Body temperature could lead to interruption of spermatogenesis, but the molecular mechanism was still unclear. Cryptorchidism was defined as the failure of testes to enter the scrotum, which exposed the testes to body temperature. Meiosis was a unique feature of germ cell development. Whether cryptorchidism damage the initiation of meiosis in boars had not been reported. The aim of this study was to determine whether spermatogonia in the cryptorchid testes entered into meiosis by detecting meiosis-related markers stimulated by retinoic acid gene 8 (STRA8) and synaptonemal complex protein 3 (SCP3). Three boars with spontaneous unilateral abdominal cryptorchidism were used. The testis located in the abdomen was cryptorchidism group, the scrotal testis of the same animal was used as control. HE results showed that only Sertoli cells, and a few spermatogonia remained in the seminiferous tubules, and no spermatids were seen compared with the control. Immunohistochemistry results showed that in both control and cryptorchidism group, STRA8 was mainly expressed in the nucleus of spermatogonia and spermatocytes. In control group, SCP3 was expressed in the nucleus of spermatocytes. In cryptorchidism group, SCP3 immunopositive cells were also observed. qRT-PCR and Western Blot results showed that the mRNA and protein levels of STRA8 and SCP3 were significantly decreased in cryptorchid boars. The expression of STRA8 and SCP3 in cryptorchidism suggested that spermatogonia could still enter meiosis in cryptorchid boars.     

Delivery of a chromosomally normal child from an oocyte with reciprocal aneuploid polar bodies

Purpose

To demonstrate that a euploid embryo derived from an oocyte with reciprocal aneuploid polar bodies is capable of producing a chromosomally normal child.

Methods

A case report of maternal MI error compensation where single nucleotide polymorphism (SNP) microarray based comprehensive chromosome screening (CCS) was performed on the 1st and 2nd polar body, the resulting embryo, and newborn DNA.

Results

CCS performed after embryo transfer identified a chromosomally normal embryo that resulted from an oocyte with reciprocal aneuploid polar bodies. The first polar body was found to be missing a single chromatid derived from chromosome 21 and the second polar body possessed an extra chromatid derived from chromosome 21. Compensation of the maternal meiotic error was verified by CCS analysis of a trophectoderm biopsy from the resulting blastocyst which was euploid for all 23 pairs of chromosomes. DNA fingerprinting and CCS of the resulting newborn confirmed a chromosomally normal child, demonstrating the developmental potential of an oocyte with reciprocal aneuploid polar bodies.

Conclusions

This is the first case report demonstrating the reproductive potential of a chromosomally normal embryo derived from an oocyte which had undergone meiosis I error. Systematic investigation into the frequency of meiosis I error compensation and the negative predictive value of polar body aneuploidy screening for reproductive potential should be conducted in order to confirm clinical relevance.     

Sterile UGA nonsense mutants of fission yeast

Summary Eight sterile mutants, which regain their fertility upon reactivation of an inactivated UGA suppressor allele of the serine tRNA gene sup3, are shown to carry UGA nonsense alleles of two established ste genes, ste1 (one mutant) and ste6 (two mutants), and of two novel genes, ste9 (four mutants) and ste10 (one leaky mutant of ras1 ^-/ste5-like cell morphology). The mutant alleles of ste1 and ste9 lead to a defect in both conjugation and meiosis, whereas those of ste6 and ste10 affect mating only. Two of the four genes map to chromosome I, ste1 in the left arm 6 cM distal of ura1, and ste9 in the right arm 3 cM distal of ade2. The ste10 and ste6 genes are located in the right arms of chromosomes II and III, respectively, the former 4 cM distal of trp1 and the latter 1 cM proximal or distal of trp3.     

The phosphodiesterase 3 inhibitor ORG 9935 inhibits oocyte maturation in the naturally selected dominant follicle in rhesus macaques

BACKGROUND: The study was conducted to determine whether the phosphodiesterase (PDE) 3 inhibitor ORG 9935 prevents the resumption of meiosis in primate oocytes during natural menstrual cycles. STUDY DESIGN: Regularly cycling adult female macaques (n=8) were followed during the follicular phase and then started on a 2-day treatment regimen of human recombinant gonadotropins to control the timing of ovulation. Monkeys received no further treatment (controls) or ORG 9935. Oocytes were recovered by laparoscopic follicle aspiration 27 h after an ovulatory stimulus, cultured in vitro in the absence of inhibitor and inseminated. The primary outcome was the meiotic stage of the oocyte. RESULTS: In six ORG 9935 cycles, five of the recovered oocytes were germinal vesicle (GV)-intact, and one exhibited GV breakdown (GVBD). In contrast, all three oocytes that recovered during control cycles were GVBD (p<.05). None of the ORG 9935-treated oocytes underwent fertilization compared with 2/3 (67%) from controls. CONCLUSIONS: These results demonstrate that ORG 9935 blocks resumption of meiosis in the naturally selected dominant follicle in primates and suggest that PDE3 inhibitors have potential clinical use as contraceptives in women.     

The phosphodiesterase 3 inhibitor ORG 9935 inhibits oocyte maturation during gonadotropin-stimulated ovarian cycles in rhesus macaques

To determine whether phosphodiesterase (PDE) 3 inhibitors prevent the resumption of meiosis by primate oocytes in vivo, rhesus macaques were stimulated to develop multiple preovulatory follicles by administering human recombinant gonadotropins, and follicles were aspirated 34 h after an ovulatory stimulus (human chorionic gonadotropin [hCG]). Monkeys received no further treatment (controls) or the PDE3 inhibitor ORG 9935 (a) exclusively in the periovulatory interval beginning 6-12 h prior to receiving hCG at 200 mg/kg every 12 h orally (PER200) or a 200 mg/kg oral loading dose followed by 50 mg/kg sc every 6 h (PER50) or (b) throughout the ovarian stimulation protocol with daily increases until a dose of 200 mg/kg bid was administered onward from the eighth day of ovarian stimulation (EXT200). The primary outcome was the number of oocytes that had resumed meiosis (germinal vesicle breakdown [GVBD]) at collection. At initial aspiration, 85% of oocytes recovered from control animals (n = 4) had progressed to GVBD compared with 53% (p<.01), 23% (p<.01), and 13% (p<.01) recovered from animals in the PER200 (n = 2), PER50 (n = 1) and EXT200 (n = 3) groups, respectively. Although spontaneous maturation of oocytes was observed during follow-up culture in the absence of ORG 9935, none of the oocytes in the PER50 or EXT200 underwent normal fertilization in vitro. These results demonstrate that the PDE3 inhibitor ORG 9935 blocks oocyte maturation during gonadotropin-stimulated ovarian cycles in rhesus macaques and suggest that PDE3 inhibitors have potential clinical use as contraceptives in women.     

人类精子染色体非整倍体研究进展

人类最常见的染色体异常是非整倍体。由于在胚胎发育的早期阶段多数的非整倍体胚胎会停止发育,因此对胚胎的非整倍体率的精确评估多通过对配子的直接研究来进行。大多数非整倍体胚胎的产生,是由于父源或母源性减数分裂中偶然的染色体不分离所导致。本文综述了正常人群的精子非整倍体发生率以及不同种类患者的精子非整倍体率,包括不育患者、体细胞核型异常患者,以及暴露于某些有害的环境或生活方式中的个体;并对精子非整倍体率升高的临床影响进行讨论。     

围生期低剂量双酚A对子代雄鼠生精功能的影响

目的: 研究围生期低剂量双酚A（BPA）对子代雄鼠睾丸生精功能的影响。方法: 妊娠母鼠在妊娠第10天到分娩后第7天,一组每天皮下注射2 μg/kg BPA（其雄性子代鼠为BPA染毒组）,另一组妊娠母鼠每日皮下注射等体积的溶剂橄榄油（其雄性子代鼠为对照组）。子代雄鼠分别在出生后18、21、24 d处死,观察睾丸组织学结构、检测睾丸内cyclin A1、cyclin B、c-jun和c-fos基因的表达,测定血浆卵泡刺激素（FSH）和黄体生成激素（LH）的水平,检测下丘脑视前区（POA）组织促性腺激素释放激素（GnRH）和弓状核（ARC）组织kiss1的mRNA表达。结果: 24日龄BPA组子代雄鼠的睾丸生精小管直径、圆形精子细胞数量以及圆形精子细胞管腔率较对照组增加（P<0.05）。21日龄BPA组子代雄鼠睾丸内cyclin A1、c-jun和c-fos的基因表达高于对照组（均P<0.01）。BPA组子代雄鼠出生后21、24 d的血浆FSH和LH的水平均高于对照组（均P<0.05）。然而2组的血浆睾酮（T）和雌二醇（E₂）水平差异无统计学意义（P>0.05）。同时,21日龄BPA组子代雄鼠的下丘脑POA-GnRH和ARC-kiss1的mRNA表达水平高于对照组（GnRH：P<0.05;kiss1：P<0.01）。结论: 围生期低剂量BPA长时间暴 露除直接作用于睾丸影响精子发生外,还通过激活ARC-kisspeptin神经元,提前启动生殖内分泌轴,使睾丸精子发生提前。     

Thymineless recombination in Saccharomyces cerevisiae is independent of the ability to undergo meiosis

Summary Thymine nucleotide starvation is recombinagenic in Saccharomyces cerevisiae and induces formation of the nuclear dense body, a structure characteristic of yeast cells in meiosis. Conceivably, thymineless recombination in yeast, presumed to be mitotic, might be meiotic in nature. We have tested this hypothesis and have found that thymineless recombination can be induced in strains incapable of meiotic exchange.     

Non-invasive method to assess genotoxicity of nocodazole interfering with spindle formation in mammalian oocytes

Trisomies due to nondisjunction in oogenesis are still a major cause of genetic diseases in humans. In this study, we analysed spindle morphology of in vitro matured nocodazole-exposed mouse oocytes by novel non-invasive Polscope-microscopy, and compared images to those obtained by anti-tubulin immunofluorescence of fixed oocytes. Polscope revealed a reduction in the numbers of oocytes expressing a birefringent spindle, and alterations in spindle morphology at concentrations of nocodazole below those inducing detectable aberrations in immunofluorescence. Hyperploidy increased significantly at a concentration of 40 nM nocodazole in mouse metaphase II oocytes, similar to thresholds inducing nondisjunction in cultured human lymphocytes. In conclusion, Polscope represents a novel highly sensitive, non-invasive method to identify chemicals inducing severe spindle aberrations that predispose mammalian oocytes to nondisjunction. Polscope may provide information on the functionality of the spindle in experimental studies but is also compatible with clinical trials in human assisted reproduction due to its non-invasive nature.