鱼类卵子发育能力的进一步研究

作者中的二人[Tung,and Tung.(童第周与叶毓芬)1944;Tung,Chang,andTung.(童第周、张致一与叶毓芬)1945]曾以分割的方法,研究金鱼(Carassiusauratus)卵子的发育能力。他们将2至8细胞时期的金鱼卵子,在去膜后,用玻璃针,按第一次或第二次的分裂平面,将整个卵子分割为二半。每半含有半数分裂球,一半卵黄,和一半未流入分裂球的细胞质。分割后,每半都能继续发育,但产生下列6种不同的情形:(1)二半都发育为完整的,一半大小的胚胎;(2)一个为完整的胚胎,另一个为不完整的胚胎;(3)一个为完整的胚胎,另一人为囊胚;     

小鼠四倍体早期胚胎基因组甲基化模式

利用小鼠抗5-甲基胞嘧啶(5MeC)单克隆抗体检测了体外培养小鼠四倍体早期胚胎的基因组甲基化模式。结果表明: 利用电融合方法制备的小鼠四倍体胚胎在体外培养体系中经历细胞质融合、细胞核融合及细胞继续分裂发育直到囊胚期的过程, 在细胞质融合的时候胚胎卵裂球同体内体外培养二倍体胚胎一样, 呈现高度甲基化状态; 在细胞核开始融合的时候, 甲基化水平急速下降, 在细胞核完全融合的时候甲基化水平达到最低点; 随着胚胎继续分裂, 胚胎甲基化水平逐渐增加, 在桑葚胚期甲基化水平最高; 但是囊胚期四倍体胚胎内细胞团同滋养层细胞甲基化荧光信号没有差别, 这与体内体外培养二倍体囊胚内细胞团细胞甲基化荧光强度高于滋养层细胞甲基化荧光强度不同。因此, 小鼠体外培养四倍体胚胎的甲基化模式是不正常的, 这可能是四倍体小鼠难以发育到妊娠足月的原因之一。这是对小鼠四倍体早期胚胎基因组甲基化模式的首次报道。     

"生存还是毁灭",发育还是阻滞--小鼠早胚体外发育2-细胞阻滞产生机制初析

在体外培养条件下,小鼠受精卵往往经一次分裂后就停滞在２－细胞,不能完成到达囊胚的后续发育过程,称作２－细胞阻滞。氧自由基伤害、培养液成分不平衡等外界因素都能引起阻滞。小鼠的２－细胞阻滞现象受细胞质内母型物质制约,具有品系依赖性。在阻滞品系小鼠胚胎的细胞质内可能缺乏某些重要的蛋白因子,在无外源信号的培养体系内不能继续分裂。发生阻滞的胚胎细胞内ＭＰＦ前体物虽然储备充足,但因无法去磷酸化激活而最终导致发     

卵母细胞成熟过程中线粒体的变化

卵子成熟是一个复杂的过程,细胞核成熟和细胞质成熟必须和谐的统一在一起,才能保证卵子的正常受精和进一步的发育。作为细胞质内最重要的细胞器,线粒体的分布在卵子成熟过程中出现了显著变化。同时其产生的ATP是卵子、受精卵以及胚胎主要的能量来源。因此,对卵子成熟过程中线粒体的分布和功能变化的研究,有利于进一步了解生殖生理,并为解决辅助生育技术中所面临的难题提供新的思路。     

线粒体与卵母细胞发育

卵子发育、成熟是一个复杂的过程, 细胞核成熟和细胞质成熟过程必须同步化, 才能保证卵子的正常受精和进一步的发育。作为细胞质内最重要的细胞器, 线粒体在卵子成熟过程中的分布的变化、氧化磷酸化产生ATP的能力以及线粒体ＤＮＡ的含量和拷贝数或转录水平对卵母细胞发育成熟有着重要的影响。因此, 对卵子成熟过程中线粒体的分布和功能状况及线粒体DNA的研究, 有利于进一步了解生殖生理, 并为解决辅助生殖技术中及克隆胚胎技术所面临的困难提供新的思路。     

外源基因转染导致山羊体细胞过快衰老与端粒缩短

在对山羊体细胞进行外源基因转染过程中,无论电击法或脂质体法所得到的细胞克隆都有细胞过快衰老的现象。山羊体细胞转基因后出现细胞体积增大、细胞核膨大并逐步分裂成多核、细胞质空泡化和吐核等衰老的表型特征。转基因后衰老细胞的染色体核型正常,但经细胞染色体端粒长度的Southern检测发现,转基因衰老细胞比原代胎儿成纤维细胞染色体端粒长度减少了2.56 kb,超出了正常传代40代的细胞的衰老速度,但转基因衰老细胞仍能支持核移植克隆胚胎的早期发育。     

小麦雄配子体的超微结构 Ⅱ.精细胞的形成和发育

1.通过小孢子有丝分裂而形成的生殖细胞,在发育过程中有一系列的包括位移和变形的变化。最后生殖细胞变为纺锤体,准备另一次有丝分裂。在此时期,生殖细胞是裸露的,它只有自己的质膜和被营养细胞的质膜包围。生殖细胞的大部分为明显的椭圆形的核所占据,具高度凝集的染色质。在生殖细胞薄层的细胞质中,除核糖体外,所有的细胞器比营养细胞质中的明显的少,而且小。微管也能在纺锤形的生殖细胞的细胞质中看到,它们的排列方向是和细胞的长轴平行的。在生殖细胞中没有造粉质体。2.当生殖细胞已移位并和营养细胞再次紧密靠近后,它开始分裂。由生殖细胞分裂所形成的精细胞及其发育包括下列主要的变化:精细胞的形状由圆球形变为椭圆形,最后变为具尾延长的细胞。与此同时,细胞质的分布逐渐集中到精细胞的一端,形成细胞质的延伸,构成所谓的精细胞的尾部。有更多的细胞器,特别是线粒体集中在尾部。从生殖细胞分裂刚形成的精细胞是裸露的,所包围的质膜是不连续的。除精细胞的质膜之外,为营养细胞的质膜所包围,在后来的发育时期此双质膜变为连续的,并在一个极短的时期有壁物质——胼胝质沉积在质膜之间的空间。但在此时期之后细胞壁变为不连续的,壁物质明显降低。对生殖细胞分裂前的位置及精细胞发育中形态变化的意义进行了讨论。     

山羊(Capra hircus)重构胚与再重构胚早期发育的研究

选择山羊8细胞期到桑椹胚期的正常胚胎或重构胚的卵裂球,或选择胚泡期胚胎的内细胞团细胞作为供核,并以LRH注射后26—28h的去核成熟卵球作为受体,制备重构胚或再重构胚,琼脂糖包埋,植入山羊输卵管内,体内培养4—6天发育结果表明:不同发育时期(8细胞期至胚泡期)的正常胚胎细胞核或重构胚细胞核中,至少有部分细胞核均保留着发育的全能性,这些细胞核在母系基因表达产物的调控下,实现重新编程,启动并完成正常发育的全过程。     

斑马鱼Z-OTU蛋白在卵子发生和胚胎发育早期的表达(英文)

斑马鱼z-otu基因编码的蛋白可能具有DUBs活性,它包含OTU和TUDOR结构域,属于OTU相关蛋白酶家族的成员。该研究将原核表达的融合蛋白(OTU结构域和GST)纯化后免疫新西兰兔,获得多克隆抗体anti-Z-OTU,并利用该抗体对Z-OTU蛋白质在斑马鱼卵子发生和早期胚胎发育过程中的表达进行了分析。根据原位和整体免疫组织化学检测结果并结合以前的研究结论,分析并比较了z-otu基因的mRNA和蛋白质的分布,发现在卵子发生和早期胚胎发育过程中,z-otu基因的mRNA和蛋白质表达模式存在明显差异:mRNA仅在卵子发生早期表达,卵母细胞受精后才重新开始表达,而其蛋白在卵子发生过程中均表达;在卵子发生过程中,mRNA分布于细胞质中,而蛋白质先分布于细胞核中,然后向细胞质迁移,接着又向卵母细胞生发泡(germinal vesicle,GV)集中。推测Z-OTU蛋白类似于其他具有去泛素化酶活性的OTU相关蛋白酶,对于卵母细胞减数分裂过程中生发泡破裂、生发泡迁移及维持胚胎的分裂是必需的。     

剑尾鱼卵子发生的的组织学观察

应用光学显微镜对卵胎生硬骨鱼类剑尾鱼(Xiphophorus helleri)卵巢的组织结构进行了观察。结果显示,剑尾鱼卵子的发育过程可划分为6个时相。Ⅰ时相的卵母细胞呈原始分化状态,细胞外具一层细胞质膜。Ⅱ时相卵母细胞外不仅具有质膜,而且还包绕一层滤泡细胞。Ⅲ时相和Ⅳ时相的卵母细胞分化明显,胞质内开始积累脂滴和卵黄颗粒。Ⅴ时相为成熟卵子,卵子的卵膜极薄,胞质内含有丰富的脂滴和卵黄。Ⅵ时相卵母细胞进入退化期,滤泡细胞从卵周向中央突入,卵黄被完全吸收,滤泡细胞自身也变得肥大。结果表明,剑尾鱼卵巢中的卵母细胞的发育是不同步的。     

Effect of protein phosphatase inhibitors on cleavage furrow formation in newt eggs: Inhibition of normal furrow formation and concomitant induction of furrow-like dents

The effects of three protein phosphatase inhibitors, okadaic acid, calyculin A and tautomycin, on the formation of cleavage furrows and the induction of furrow-like dents in the egg of the newt, Cynops pyrrhogaster , were examined. Solutions of the individual compound were injected into the animal hemisphere of one of the two presumptive blastomere regions of the embryo during the first cleavage. Injection of a solution containing any of the chemicals often disturbed the formation of a normal furrow in the injected blastomere at second cleavage. Injection with okadaic acid or calyculin A often induced furrow-like dents on the surface of the injected blastomere at the same time as second cleavage in control embryos, while that with tautomycin usually did not induce them. In an injected blastomere, formation of dents started in the animal half and moved towards the vegetal half as the furrow in its counterpart blastomere extended from the animal half towards the vegetal. Dents gradually became slightly deeper and formed cytoplasmic projections that later degenerated, leaving a surface scar. Cytological observations on blastomeres injected with calyculin A revealed that nuclear division occurred normally.     

Dual blastomere analysis improves reliability of preimplantation trembler mouse diagnosis

Dual blastomere biopsy and independent blastomere analysis dramatically improved preimplantation diagnostic reliability as confirmed by testing the remaining biopsied eight-cell mouse embryo. The autosomal dominant trembler mouse point mutation was selected as a model for human preimplantation diagnosis because: (1) single cell assay failure is predicted to be the highest when testing autosomal dominant mutations; (2) point mutations represent the most common of all mutation categories and the most demanding mutation to assay reliably; and (3) the trembler mouse point mutation in peripheral myelin protein 22 (Pmp22) is a model of human Charcot-Marie-Tooth type 1A disease. Mathematical models predict our experimental results assuming amplification of 80% of each target allele as well as trembler sperm DNA contamination in 1 of 44 normal biopsied single blastomeres. Single blastomere analysis correctly predicted the genotype in only 84% of embryos that would have been implanted as normal. In contrast, when independent tests of both biopsied blastomeres agreed, test results were confirmed in 20 of 21 (95.2%) of the remaining six-cell biopsied embryos designated as normal. Thus, biopsied six-cell embryo confirmation demonstrated that dual biopsied blastomere analysis improved test reliability remarkably. Received: 20 November 1996 / Accepted: 4 April 1997     

The Determinant for Archenteron Formation in Starfish: Co-Culture of an Animal Egg Fragment-Derived Cell Cluster and a Selected Blastomere

A method of detecting blastomeres that carrying the determinant for archenteron formation was established, based on the reported localization of the determinant in the vegetal cytoplasm (17, 24). The essence of the method was to co-culture a selected blastomere with an animal egg fragment-derived cell cluster, so as to generate one joined embryo. The presence of the determinant in the blastomere was assessed by the formation of the archenteron in the developed joined embryos. The vegetal blastomeres that carried the determinant sometimes induced animal egg fragment-derived cells to form part of the archenteron.     

Early development and axis specification in the sea anemone Nematostella vectensis

We investigated the early development of the sea anemone Nematostella vectensis, an emerging model system of the Cnidaria. Early cleavage stages are characterized by substantial variability from embryo to embryo, yet invariably lead to the formation of a coeloblastula. The coeloblastula undergoes a series of unusual broad invaginations-evaginations which can be blocked by cell cycle inhibitors suggesting a causal link of the invagination cycles to the synchronized cell divisions. Blastula invagination cycles stop as cell divisions become asynchronous. Marking experiments show a clear correspondence of the animal-vegetal axis of the egg to the oral-aboral axis of the embryo. The animal pole gives rise to the concave side of the blastula and later to the blastopore of the gastrula, and hence the oral pole of the future polyp. Asymmetric distribution of granules in the unfertilized egg suggest an animal-vegetal asymmetry in the egg in addition to the localized position of the pronucleus. To determine whether this asymmetry reflects asymmetrically distributed determinants along the animal-vegetal axis, we carried out blastomere isolations and embryonic divisions at various stages. Our data strongly indicate that normal primary polyps develop only if cellular material from the animal hemisphere is included, whereas the vegetal hemisphere alone is incapable to differentiate an oral pole. Molecular marker analysis suggests that also the correct patterning of the aboral pole depends on signals from the oral half. This suggests that in Nematostella embryos the animal hemisphere contains organizing activity to form a normal polyp.     

Cellular interactions in early C. elegans embryos

In normal development both the anterior and posterior blastomeres in a 2-cell C. elegans embryo produce some descendants that become muscles. We show that cellular interactions appear to be necessary in order for the anterior blastomere to produce these muscles. The anterior blastomere does not produce any muscle descendants after either the posterior blastomere or one of the daughters of the posterior blastomere is removed from the egg. Moreover, we demonstrate that a daughter of the anterior blastomere that normally does not produce muscles appears capable of generating muscles when interchanged with its sister, a cell that normally does produce muscles. Embryos develop normally after these blastomeres are interchanged, suggesting that cellular interactions play a major role in determining the fates of some cells in early embryogenesis.     

Early patterning of the mouse embryo--contributions of sperm and egg

The first cleavage of the fertilised mouse egg divides the zygote into two cells that have a tendency to follow distinguishable fates. One divides first and contributes its progeny predominantly to the embryonic part of the blastocyst, while the other, later dividing cell, contributes mainly to the abembryonic part. We have previously observed that both the plane of this first cleavage and the subsequent order of blastomere division tend to correlate with the position of the fertilisation cone that forms after sperm entry. But does sperm entry contribute to assigning the distinguishable fates to the first two blastomeres or is their fate an intrinsic property of the egg itself? To answer this question we examined the distribution of the progeny of early blastomeres in embryos never penetrated by sperm - parthenogenetic embryos. In contrast to fertilised eggs, we found there is no tendency for the first two parthenogenetic blastomeres to follow different fates. This outcome is independent of whether parthenogenetic eggs are haploid or diploid. Also unlike fertilised eggs, the first 2-cell blastomere to divide in parthenogenetic embryo does not necessarily contribute more cells to the blastocyst. However, even when descendants of the first dividing blastomere do predominate, they show no strong predisposition to occupy the embryonic part. Thus blastomere fate does not appear to be decided by differential cell division alone. Finally, when the cortical cytoplasm at the site of sperm entry is removed, the first cleavage plane no longer tends to divide the embryo into embryonic and abembryonic parts. Together these results indicate that in normal development fertilisation contributes to setting up embryonic patterning, alongside the role of the egg.     

Changes in the Pattern of Intercellular Junctions during Early Embryogenesis of the Starfish, Asterias amurensis

Changes in the cellular adhesion pattern during the early embryogenesis of a starfish Asterias amurensis were examined using carboxyfluorescein (CF) dye as a probe. CF that was injected into one of the blastomeres at the 2- or 4-cell stage was in all cases restricted to the progeny cells of the CF-labelled blastomere. With the advancement of gastrulation, however, the injected dye was distributed not only to the progeny of the labelled blastomere, but also to cells that originated from non-injected blastomeres. At the beginning of mesenchyme cell release, the injected dye spread uniformly to most cells comprising the embryo. When one of the blastomeres situated in the vegetal hemisphere of an 8-cell embryo was labelled, the resulting embryo showed more intense fluorescence in the cells surrounding the archenteron than in the ectodermal layer, suggesting that the cells in ectodermal layer became associated more intimately or earlier than those surrounding the archenteron. Likewise, in double embryos formed by combining two denuded eggs, in which one egg had been labelled with CF, dye spread was observed when the ectodermal layer began to expand. The intercellular spread of CF dye in starfish embryo suggests that there is a dramatic change in the cellular adhesion pattern during the course of gastrulation.     

Topological determination of early morphogenesis in Metazoa

This paper presents a topological interpretation of some developmental events through the use of well-known concepts and theorems of combinatorial geometry. The organization of early embryo using a simulation of cleavage considering only blastomere contacts is examined. Each blastomere is modeled as a topological cell and whole embryo—as cell packing. The egg cleavage results in a pattern of cellular contacts on the surface of each blastomere and whole embryo, a discrete morphogenetic field. We find topological distinctions between different types of early egg cleavage and suggest a topological classification of cleavage. Blastulation and gastrulation may be related to an inevitable emergence of discrete curvature that directs development in three-dimensional space. The relationship between local and global orders in metazoan development, i.e., between local morphogenetic processes and integral developmental patterns, is established. Thus, this methodology reveals a topological imperative: a certain set of topological rules that constrains and directs biological morphogenesis.     

Regional specification during embryogenesis in the inarticulate brachiopod Discinisca.

The process of embryogenesis is described for the inarticulate brachiopod Discinisca strigata of the family Discinidae. A fate map has been constructed for the early embryo. The animal half of the egg forms the dorsal ectoderm of the apical and mantle lobes. The vegetal half forms mesoderm and endoderm and is the site of gastrulation; it also forms the ectoderm of the ventral regions of the apical and mantle lobes of the larva. The plane of the first cleavage goes through the animal-vegetal axis of the egg along the future plane of bilateral symmetry of the larva. The timing of regional specification in these embryos was examined by isolating animal, vegetal, or lateral regions at different times from the 2-cell stage through gastrulation. Animal halves isolated at the 8-cell and blastula stages formed an epithelial vesicle and did not gastrulate. When these halves were isolated from blastulae they formed the cell types typical of apical and mantle lobes. Vegetal halves isolated at all stages gastrulated and formed a more or less normal larva; the only defect these larvae had was the lack of an apical tuft, which normally forms from cells at the animal pole of the embryo. When lateral isolates were created at all developmental stages, these halves gastrulated. Cuts which separated presumptive anterior and posterior regions generated isolates at the 4-cell and blastula stages that formed essentially normal larvae; however, at the midgastrula stage these halves formed primarily anterior or posterior structures indicating that regional specification had taken place along the anterior-posterior axis. The plane of the first cleavage, which predicts the plane of bilateral symmetry, can be shifted by either changing the cleavage pattern that generates the bilateral 16-cell blastomere configuration or by isolating embryo halves prior to, or during, the 16-cell stage. These results indicate that while the plane of the first cleavage predicts the axis of bilateral symmetry, the axis is not established until the fourth cleavage. The development of Discinisca is compared to development in the inarticulate brachiopod Glottidia of the family Lingulidae and to Phoronis in the phylum Phoronida.