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Ultrastructure of human mature oocytes after vitrification
M.A. Khalili,M. Maione,M.G. Palmerini,S. Bianchi
European Journal of Histochemistry , 2012, DOI: 10.4081/ejh.2012.e38
Abstract: Since the introduction of human assisted reproduction, oocyte cryopreservation has been regarded as an attractive option to capitalize the reproductive potential of surplus oocytes and preserve female fertility. However, for two decades the endeavor to store oocytes has been limited by the not yet optimized methodologies, with the consequence of poor clinical outcome or of uncertain reproducibility. Vitrification has been developed as the promising technology of cryopreservation even if slow freezing remains a suitable choice. Nevertheless, the insufficiency of clinical and correlated multidisciplinary data is still stirring controversy on the impact of this technique on oocyte integrity. Morphological studies may actually provide a great insight in this debate. Phase contrast microscopy and other light microscopy techniques, including cytochemistry, provided substantial morphofunctional data on cryopreserved oocyte, but are unable to unraveling fine structural changes. The ultrastructural damage is one of the most adverse events associated with cryopreservation, as an effect of cryo-protectant toxicity, ice crystal formation and osmotic stress. Surprisingly, transmission electron microscopy has attracted only limited attention in the field of cryopreservation. In this review, the subcellular structure of human mature oocytes following vitrification is discussed at the light of most relevant ultrastructural studies.
Cryopreservation of Human Unfertilized and Fertilized Oocytes
Vanderzwalmen P,Zech NH,Nagy ZP,Stecher A
Journal für Reproduktionsmedizin und Endokrinologie , 2013,
Abstract: Cryopreservation of human fertilized oocytes and embryos are nowadays well established in IVF practice with a wide range of clinical applications. However, freezing of unfertilized MII oocytes turned out to be one of the greatest challenges in the field of human reproductive cryobiology since the protocols remained ineffective for over 25 years. Only in the last 10 years the efficiency and safety of oocyte cryopreservation tremendously improved with the realization that zona hardening occurs in the process of cryopreservation and the introduction of vitrification. The possibility to cryopreserve unfertilized oocytes introduces new applications in the field of cryopreservation. We have now the opportunity to preserve fertility for cancer patients or to freeze oocytes for IVF patients when now sperm is available at the day of pick-up. This chapter will be dedicated to the vitrification of unfertilized MII oocytes and fertilized oocytes (pronuclear or zygote stage). An overview on the different indications for both stages of development will be described in the first part of this chapter. The second part will be about the technical and practical description of the vitrification process using hermetically closed and thus aseptic systems such as the Vitrisafe as carrier device. Finally, an overview on the clinical aspect, including our experience, will be given.
基于微量rna高通量测序技术的牦牛mii期卵母细胞转录组研究
兰道亮,熊显荣,林宝山,陈亚冰,胡敏,苏小珊,李键
牲畜兽医学报 , 2014,
Abstract: ?旨在现有高通量测序的基础上,建立一种针对牦牛卵母细胞等微量样本的rna高通量测序方法,为进一步研究牦牛卵母细胞的转录组学提供基础。以微量mii期牦牛卵母细胞rna(10ng)作为研究样本,应用smartcdnapcr扩增技术对样本进行富集并构建测序文库,再应用改进的rna高通量测序技术对其进行高通量测序分析。经illumina-solexa深度测序后,得到了一个包含47619254条测序序列,4gb大小数据量的微量mii期牦牛卵母细胞测序文库。质量控制(qualitycontrol,qc)及基本结果分析表明测序文库质量良好。基因组比对分析显示,共有15626个牦牛基因被映射。此外,还获得了7348个新的转录本。go分类注释结果显示,共有13795个比对上的基因参与了生物过程、细胞组分及分子功能3大主要类别。进一步富集分析显示,分别有333134及113个go类别在上述3大类中得到富集。kegg通路分析结果显示,共有13496个比对上的基因参与了258条通路,其中101条通路得到有效富集。本研究以牦牛卵母细胞为样本成功建立了一种起始rna量可低至10ng的微量转录组测序方法。该研究结果为进一步研究牦牛基因组提供了基础,同时也为研究牦牛mii期卵母细胞的分子机理提供了新的视角。
3种冷冻方法对猪mii期卵母细胞冷冻后微丝分布和脂肪颗粒变化的影响
吴彩凤,戴建军,张树山,李维杰,徐利,张德福
牲畜兽医学报 , 2014,
Abstract: ?本研究旨在探讨3种冷冻方法(clv法、ops法和straw法)对猪mii期卵母细胞玻璃化冷冻后微丝、脂肪颗粒分布及其超微结构的影响。结果显示:(1)clv(cryoloopvitrification)法冷冻速率最高,达42639℃?min-1;(2)clv法微丝正常分布率(47.06%)最高,显著高于ops(openpulledstraw)法的38.04%和straw法的30.95%(p<0.05)。(3)clv法和ops法的脂肪颗粒正常分布率之间没有显著差异(p>0.05),但两者均显著高于straw法(p<0.05);(4)解冻后卵母细胞超微结构观察结果显示,大的脂肪颗粒含量明显减少;大部分脂肪颗粒冷冻后呈均质化状态,只有很少的异质化脂肪颗粒存在。试验结果表明,冷冻过程中卵母细胞脂肪颗粒破坏严重,增加冷冻速率能降低对微丝的损伤,从而减少对脂肪颗粒异常分布的影响。
3种冷冻方法对猪mii期卵母细胞线粒体分布和损伤的影响
戴建军,吴彩凤,张廷宇,张树山,徐利,张德福
牲畜兽医学报 , 2012,
Abstract: ?本研究旨在探讨3种冷冻方法对猪mii期卵母细胞玻璃化冷冻后线粒体的分布和超微结构变化的影响。通过透射电镜、rhodamine-123(r-123)荧光染色和体外发育观察,结果显示,(1)无论是fda-dapi复染存活率,还是孤雌激活卵裂率,clv(cryoloopvitrification)法(72.00%,7.22%)效果最好,ops(openpulledstraw)法(60.00%,4.85%)次之,straw法(42.22%,0%)最低;(2)clv法经r123染色后,卵母细胞线粒体的正常分布率(52.24%)要比其它2组要高(ops,48.65%;straw,37.68%),但三者之间没有显著差异(p>0.05);(3)透射电镜超微结构表明,冻后卵母细胞线粒体变得粗糙和模糊,有的线粒体嵴减少甚至消失。结果表明,冷冻过程中卵母细胞线粒体的分布和形态损伤严重,clv法可提高冷冻速率,增强冻后卵母细胞的发育能力,降低冷冻造成的卵母细胞线粒体异常分布比例。
一种支持组播的snmpv3改进模型
程春玲,张登银,崔国亮,隋宗见?
计算机科学 , 2012,
Abstract: 随着三网融合的发展,业务类型逐渐增多,设备数量急剧增加。典型的基于单播模式的snmi〕网管系统在数据采集过程中存在大量的重复数据发送,造成带宽开销大、网管系统效率不高。提出一种支持组播的snmpv3改进模型,它引入了组播组的概念和组代理实体。模型扩展了mii3库的定义,改进了snmp引擎,使其能识别和处理组播消息;给出了管理站和代理端的模型结构和主要模块;随后阐述了基于改进模型的组管理流程和基于组播轮询的数据采集的实现流程。系统测试结果表明,基于改进模型所开发的原型系统采用组播轮询的方式进行数据采集,可以减少数据的重复发送,减少网络流量,提高数据查询和数据采集的效率。
Comparación de la calidad embrionaria entre fertilización in vitro (FIV) y cultivo intravaginal de ovocitos (INVO) en el Centro Colombiano de Fertilidad y Esterilidad - CECOLFES, Bogotá, Colombia Comparing embryo quality between in vitro fertilization (IVF) and intravaginal ovocyte culture (INVO) in the Colombian Centre for Fertility and Sterility - CECOLFES, Bogotá, Colombia
Doris Elena Navarro-Carbonell,Carlos Pulido-Torres,ángela María Saa-Madri?án,Elkin Lucena-Quevedo
Revista Colombiana de Obstetricia y Ginecología , 2012,
Abstract: Objetivo: comparar la calidad embrionaria y describir las tasas de implantación, embarazo y aborto en las técnicas de fertilización in vitro (FIV) y el cultivo intravaginal de ovocitos. Materiales y métodos: cohortes históricas de pacientes con tratamiento de fertilización in vitro y el cultivo intravaginal de ovocitos en el Centro Colombiano de Fertilidad y Esterilidad (Cecolfes) durante el a o 2010. Se incluyeron 137 pacientes aspiradas dentro de los cuatro grupos de estudio: Grupo 1, FIV/Incubadora; Grupo 2, FIV/INVO; Grupo 3, ICSI/INVO, y Grupo 4, ICSI/Incubadora. Se midió el peso de la paciente, el número de ovocitos recuperados y óvulos maduros (MII), la tasa de implantación y la tasa de embarazo y aborto en cada uno de los grupos. Se realizó análisis mediante la prueba de Kruskal-Wallis; la calidad embrionaria fue evaluada con un análisis de covarianza multivariado (MANOVA). Resultados: se observó diferencia significativa en la calidad embrionaria entre las dos técnicas FIV e INVO (p = 0,0388). En la técnica INVO se presentaron mayores tasas de división embrionaria (μ = 7,35/INVO frente a 6,64/Incubadora) y menor fragmentación (μ = 4,67/INVO frente a 4,59/ Incubadora). En cuanto a la tasa de implantación, embarazo y aborto se obtuvieron más altos porcentajes en los grupos INVO. Conclusión: la técnica INVO se asoció a mejor calidad embrionaria. Las tasas de implantación, embarazo y bajas tasas de aborto son semejantes a las descritas en la técnica FIV. Objective: Comparing embryo quality and describing implantation, pregnancy and abortion rates regarding in vitro fertilization (IVF) and intravaginal oocyte culture (INVO) techniques. Materials and methods: The study involved historical cohorts of patients undergoing IVF and INVO treatment in the Colombian Fertility and Sterility Centre (Centro Colombiano de Fertilidad y Esterilidad – Cecolfes) during 2010. It involved 137 aspirated patients, covering four study groups: Group 1 IVF/incubator, Group 2 IVF/INVO, Group 3 ICSI/INVO and Group 4 ICSI/incubator. The patients’ weight, the number of ovocytes retrieved, mature ovules (M2), implantation rate, pregnancy and abortion rates were measured in each group. The Kruskal-Wallis test was used for statistical analysis; embryo quality was evaluated by multivariate covariance analysis (MANOVA). Results: A significant difference was observed regarding embryo quality between IVF and INVO (p = 0.0388), the INVO technique having higher embryo cleavage rates (μ = 7.35/INVO cf 6.64/ incubator) and lower embryo fragmentation (μ = 4.67/INVO cf 4.59/incubator). INVO
Improved Model of SNMPv3 Supporting Multicast
一种支持组播的SNMPv3改进模型

程春玲,张登银,崔国亮,隋宗见
计算机科学 , 2012,
Abstract: With the development of triple play, the type of business increases and the number of devices increases dra- matically. Some problems, such as the data repeat sending, large broadband overhead and low efficiency, are more serious in typical SNMP network management system based on unicast mock. An improved SNMPv3 model supporting multi- cast was proposed and the concepts of multicast group and the entity of group proxy were introduced. In the improved SNMPv3 modcl,the definition of MII3 was extended and the SNMP engine was improved to identify and process the multicast messages. The improved model and main modules on management stations and proxies were given respective- 1y. Then the implementation flows of group management and data collection based on improved model were described. hhe test results for the prototype system show that the data collection based on multicast polling can save the resource of senders, reduce network traffic and improve the data query and data collection efficiency greatly.
Goat MII Ooplasts Support Preimplantation Development of Embryos Cloned from Other Species
山羊M II期卵母细胞胞质支持异种间克隆胚的着床前发育

Xujun Xu,Guohui Liu,Jianquan Chen,Juan Chen,Hongying Sh,Youbing Wu,Aimin Zhang,Guoxiang Cheng,
徐旭俊
,刘国辉,陈建泉,陈娟,沙红英,吴友兵,张爱民,成国祥

生物工程学报 , 2008,
Abstract: The preimplantation development competences of somatic cell nuclear transfer (SCNT) embryos reconstructed with enuleated goat (Capra hircus) Metaphase II (MII) oocytes matured in vivo and whole cells derived from adult fibroblasts of several mammalian species (goat, boer goat, bovine, tahr, panda) and human patient were evaluated. Results obtained from our experiments revealed that these reconstructed SCNT embryos could complete preimplantation development to form blastocysts. The fusion rate and blastocyst rate of intra-species SCNT embryos (Capra hircus as control) was 78.67 (557/708); 56.29% (264/469), that of sub-species or inter-species SCNT embryos were: boer goat 78.18% (541/692); 33.90% (40/118), bovine 70.53% (146/207); 22.52% (25/111), tahr 53.51% (61/114); 5.26% (3/570), panda 79.82% (1159/1452); 8.35% (75/898) and human 68.76% (317/461); 5.41% (16/296), respectively. It is concluded that (1) there are no relationships between fusion rate and relativeness of the recipient cytoplasm to nucleus donor cells, (2) cytoplast of the goat MII oocyte can support the preimplantation development of SCNT embryos reconstructed with nucleus from other species, (3) the blastocyst rate of close relative inter-species SCNT embryos is higher than that of distant relative inter-species SCNT embryos.
Goat MII Ooplasts Support Preimplantation Development of Embryos Cloned from Other Species
山羊M II期卵母细胞胞质支持异种间克隆胚的着床前发育

Xujun Xu,Guohui Liu,Jianquan Chen,Juan Chen,Hongying Sh,Youbing Wu,Aimin Zhang,Guoxiang Cheng,
徐旭俊
,刘国辉,陈建泉,陈娟,沙红英,吴友兵,张爱民,成国祥

微生物学报 , 2008,
Abstract: The preimplantation development competences of somatic cell nuclear transfer (SCNT) embryos reconstructed with enuleated goat (Capra hircus) Metaphase II (MII) oocytes matured in vivo and whole cells derived from adult fibroblasts of several mammalian species (goat, boer goat, bovine, tahr, panda) and human patient were evaluated. Results obtained from our experiments revealed that these reconstructed SCNT embryos could complete preimplantation development to form blastocysts. The fusion rate and blastocyst rate of intra-species SCNT embryos (Capra hircus as control) was 78.67 (557/708); 56.29% (264/469), that of sub-species or inter-species SCNT embryos were: boer goat 78.18% (541/692); 33.90% (40/118), bovine 70.53% (146/207); 22.52% (25/111), tahr 53.51% (61/114); 5.26% (3/570), panda 79.82% (1159/1452); 8.35% (75/898) and human 68.76% (317/461); 5.41% (16/296), respectively. It is concluded that (1) there are no relationships between fusion rate and relativeness of the recipient cytoplasm to nucleus donor cells, (2) cytoplast of the goat MII oocyte can support the preimplantation development of SCNT embryos reconstructed with nucleus from other species, (3) the blastocyst rate of close relative inter-species SCNT embryos is higher than that of distant relative inter-species SCNT embryos.
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