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关键词： spinal cord   neuronal culture   tissue culture inserts   amino acid release   NMDA  

摘要： In the present study, using tissue culture inserts (TCI) coupled with a primary spinal cord neuronal culture. we characterize a new perfusion system, which permits continuous perfusate collection from cultured neurons. Primary spinal cord neurons were isolated from the lumbar portion of E14 spinal cords of Sprague-Dawley rats, plated on TO and fed with DMEM/B27/10% FBS. At 1-4 weeks after isolation the. development of synapses and neurotransmitter phenotype in cultured neurons was verified using immunofluorescence. A time-dependent development of synapses (Syn) was seen with a dense Syn-positive network identified at 3-4 weeks after plating. A sub-population of plated neurons (35-40%) showed GABA immunoreactivity and expressed NMDARI receptor. To measure neurotransmitter release. a chamber accommodating TO was constructed permitting per-fusion of the insert across the membrane. To evoke amino acid release from cultured neurons, NMDA (10mmol/1) was added into the perfusion buffer. Stimulation with NMDA evoked a significant GABA (4050 +/- 950%) and glutamate release (130 +/- 42%) during first 10 min after exposure. In control non-stimulated cells no significant changes,were measured. These data show that by using TCI it is possible to maintain embryonic spinal cord neurons for an extended period and that this system may represent a simple tool to identify neurotransmitter and/or peptides associated with a specific population of cultured brain and/or spinal cord neurons. (C) 2004 Elsevier B.V. All rights reserved.

关键词： CHROMOSOMES   FIBROBLASTS   WERNER'S syndrome   POLYMERASE chain reaction  

摘要： To determine whether the mouse Werner syndrome homologue (Wrn) and the poly (ADP-ribose) polymerase-1 (PARP-1) enzymes act in concert to prevent specific chromosomal rearrangements, mice with a mutation in the helicase domain of the Wrn gene (wrn(Deltahel/Deltahel) mice) were crossed to PARP-1 null mice. Spectral karyotyping of the mouse metaphases was used in correlation with conventional G-banded karyotype analysis to precisely define the chromosomal aberrations in cells. Although there was no recurrent clonal chromosome aberration, PARP-1 null/Wrn(Deltahel/Deltahel) fibroblasts were distinguished by an increased frequency of chromatid breaks. Interestingly, multiradial structures were the only type of DNA rearrangement that was significantly higher in such PARP-1 null/Wrn(Deltahel/Deltahel) cells. These results indicate that Wrn and PARP-1 enzymes may be part of a protein complex involved in the processing of DNA breaks that can ultimately lead to multiradial structures when both enzymes are nonfunctional. Finally, regions of chromosomes known to be fragile sites in the mouse genome are not more prone to DNA rearrangements in the absence of both PARP-1 and functional Wrn proteins. Moreover, the low number of recurrent rearranged chromosome at any given site suggest a random mutagenesis process in PARP-1 null/Wrn(Deltahel/Deltahel) fibroblasts. (C) 2005 Elsevier Inc. All rights reserved.

关键词： 早期胚胎   发育   Nestin   CK19   PCNA   细胞骨架蛋白   免疫组织化学  

摘要： 目的:研究NestinCK19及PCNA在胚胎早期组织器官的分布。方法:对6-10w的胚胎组织采用免疫组化SABC法染色观察。结果:胚胎6w时肝脏内血细胞前体细胞Nestin阳性;7w时肝、椎间盘及皮肤有Nestin的表达,导管上皮组织开始出现CK19阳性反应。PCNA的表达出现在肝脏,9w时表达达到高峰。结论:NestinCK19及PCNA在胚胎早期发育过程中对肝、椎间盘、皮肤等器官组织的发育可能起着调控作用。

关键词： 组织学   胚胎学   实验教学   教学效果   医学教育  

摘要： 提高组织学与胚胎学实验教学效果,有助于学生更好地掌握相关基础理论,为今后学习其他医学课程奠定扎实的基础.就如何搞好组织学与胚胎学的实验教学进行探讨,旨在总结经验,有成效地提高教学质量.

关键词： 流产   微小病毒B19   聚合酶链反应  

摘要： 目的 :应用聚合酶链反应 (PCR)技术检测早、中期自然流产患者胚胎及胎盘等组织中的微小病毒B19,对比分析早期流产的原因。方法 :收集早期自然流产患者的胚胎绒毛组织和母亲血清 ,孕中期自然流产胎儿的胎盘、胎儿组织和母亲血清 ,足月分娩者的胎盘组织及母亲血清 ,应用PCR检测微小病毒B19DNA ,同时检测母血清抗体。结果 :6 1例早期流产胚胎绒毛中 ,3例 (5 % )B19病毒阳性 ;4 5例孕中期胚胎组织中 ,5例 (11% )检测出B19病毒 ;5 0例足月分娩胎盘组织中未检测出B19病毒。早期流产患者血清中特异性抗体IgG检出 31%、IgM阴性 ;检出孕中期母血清B19病毒特异性抗体IgG占 35 %、IgM占 11%。结论 :B19病毒是导致早期流产的原因之一 。

关键词： 病理学教学   临床医学   基础医学   病理改变   转归   组织胚胎学   医学生   学生学习   兴趣   激发  

摘要： 病理学是一门研究疾病的病因、发病机制、病理改变和转归的医学基础学科。它不仅需要以基础医学中的解剖学、组织胚胎学、生理学、生物化学、分子生物学等为其学习基础，同时又为临床医学学习提供必要的理论依据，是沟通基础医学和临床医学的桥梁，在医学院校的教学中占有重要地位。但是众所周知，病理学内容比较繁杂、抽象，不易理解记忆，尤其对于处于基础学习阶段的医学生而言，极易产生枯燥、乏味感，严重影响教学效果。针对这一问题，我们进行了一系列探索，总结出一套行之有效的教学方法，旨在从激发学生学习兴趣着手提高病理学教学质量。

关键词： 组织胚胎学   多媒体教学   教学效果   医学教育  

摘要： 多媒体教学具有传统教学无法比拟的优点,但对于多媒体教学中出现的一些问题不及时加以改进就达不到多媒体教学的目的,反而影响教学质量.针对这些问题,提出了一些改进措施.

关键词： alkaline phosphatase   chick embryos   digoxigenin   gene expression   histology   in situ hybridization   NBT/BCIP   plastic embedding   whole mounts  

摘要： We describe a novel method that allows reliable detection of in situ hybridization signals in thin sections of plastic embedded embryos. Sections from plastic embedded embryos are thinner and have superior histological quality compared to paraffin, gelatin, agarose embedded sections or cryosections;however, plastic resin traditionally has not been used as an embedding medium following,in situ hybridization because of loss of signal. When signal is detected with alkaline phosphatase and NBT/BCIP, the resulting colored precipitate is subject to fading when samples are exposed to organic compounds. The colored precipitate can be redeposited by repeating the NBT/BCIP reaction following plastic sectioning. This recolorization shows no loss of specificity, because signal is detected only where the anti-digoxigenin/alkaline phosphatase conjugated antibody is bound to the riboprobe. Strong signals can be detected without recolorization;however, weaker signals require the recolorization step. This novel method of re-depositing colored precipitate after processing and sectioning allows accurate determination of the location of gene expression and study of this expression in high quality histological sections of early chick embryos..

关键词： Helicoverpa armigera   cathepsin B-like proteinose   expression   embryonic development  

摘要： Cathepsin B-like proteinase from Helicoverpa armigera (HCB) was proposed as being involved in the degradation of yolk proteins during embryonic development. Recombinant HCB was expressed as a fusion protein with GST in Escherichia coli BL21 on the basis of its cDNA and purified to homogeneity. The fusion protein was cleaved with thrombin to generate a soluble protease with a mass of 37 kDa. A polyclonal antiserum against this recombinant protein, raised in the rabbit, recognized three isoforms of HCB in an ovary homogenate of this insect. Expression of this enzyme during embryonic development was studied using immunoblotting, immunohistochemistry and activity assay. It was found that HCB was expressed during embryonic development and that its proteolytic activity was detected from embryonic developmental eggs. The fact that HCB activity is observed in ovaries and developing eggs suggested that the enzyme had already been activated before embryonic development. Immunohistochemistry indicated that the enzyme was located in follicular cells, the sphere of yolk granules, and the fat bodies of female adult. These lines of evidence suggested strongly that HCB takes part in the degradation of yolk proteins during the development of embryo. (C) 2004 Wiley-Liss, Inc.

关键词： in vitro propagation   medicinal plant   Quassia amara   silver nitrate  

摘要： In vitro propagation of Quassia amara L. (Simaroubaceae) was attempted using mature and juvenile explants. Attempts to establish in, vitro culture using leaf and internode explants from a plant more than 15 yr old were unsuccessful due to severe phenolic exudation. Plant regeneration through direct and indirect somatic embryogenesis was established from cotyledon explants. Murashige and Skoog (MS) medium with 8.9 muM N-6-benzyladenine (BA) and 11.7 muM silver nitrate induced the highest number (mean of 32.4 embryos per cotyledon) of somatic embryos. Direct somatic embryogenesis as well as callus formation was observed on medium with BA (8.9-13.3 muM). Semi-mature pale green cotyledons were superior for the induction of somatic embryos. Embryos developed from the adaxial side as well as from the point of excision of the embryonic axis. More embryos were developed on the proximal end compared to mid and distal regions of the cotyledons. Subculture of callus (developed along with the somatic embryos on medium with BA alone) onto medium containing 8.9 muM BA and 11.7 muM silver nitrate produced a mean of 17.1 somatic embryos. Primary somatic embryos cultured on MS medium with 8.9 muM BA and 11.7 muM silver nitrate produced a mean of 9.4 secondary somatic embryos. Most of the embryos developed up to early cotyledonary stage. Reduced concentration of BA (2.2 or 4.4 muM) improved maturation and conversion of embryos to plantlets. Ninety percent of the embryos converted to plantlets. The optimized protocol facilitated recovery of 30 plantlets per cotyledon explant within 80 d. Plantlets transferred to small cups were subsequently transferred to field conditions with a survival rate of 90%.