喜树细胞悬浮培养中生理生化指标的测定

以喜树(Camptotheca acuminata Decsne.)幼嫩叶片为材料,诱导和筛选愈伤组织,进行细胞悬浮培养。初步研究了液体培养条件下光照对喜树细胞生长和生理生化特性的影响。结果显示：在光照条件下培养细胞生长周期30d,在黑暗条件下培养细胞生长周期为27d,细胞的增殖曲线呈“S”形;在两种不同光照条件下,其培养液pH值先下降后回升;培养细胞中可溶性蛋白质含量及过氧化物酶活性均出现两个峰值,但出现的时间不同。     

水分胁迫对小麦幼苗及悬浮培养细胞中诱导蛋白表达的影响

丰抗8号小麦幼苗及成熟胚诱导的悬浮培养细胞在水分胁迫（－1．0MPa PEG6000）下,可溶性蛋白含量与蛋白组分变化有差异,幼苗可溶性蛋白含量高于对照,并随生长的延长呈降低趋势;悬浮培养细胞可溶性蛋白含量低于对照,且略有上升;复水后均可恢复对照水平,SDS－PAGE电泳及薄层扫描分析结果表明,幼苗受水分胁迫诱导,出现44．2kD蛋白亚基,该蛋白亚基含量可随胁迫时间延长上升,复水后消失,在正常条件下悬浮培养细胞中含有44．2kD蛋白亚基表达,轻度胁迫处理时,该蛋白亚基含量上升,对悬浮培养细胞进行水分胁迫,该蛋白则表现下降趋势,复水后又可上升。     

绞股蓝细胞培养及其某些生理生化特性

从绞股蓝幼叶诱导并筛选出了胚性愈伤组织,其悬浮培养细胞生长曲线呈S形,生长周期中培养液pH值先降低后回升,可溶性糖含量避渐降低,培养细胞可溶性蛋白质含量及过氧化物酶活性出现两个高峰,过氧化物酶谱带数目无变化。培养细胞能合成皂苷,静止期皂苷含量最高。     

遮荫对半夏叶片光合色素与保护酶活性的影响

对半夏进行不同程度遮荫处理,研究了其生长过程中叶组织的光合色素含量、叶绿素a/b值、保护酶体系(SOD、POD、CAT)活性和丙二醛(MDA)含量的变化及其生理特性.结果表明:不同处理的半夏在生长过程中,光合色素的含量呈先升后降的趋势;与全光照相比,遮荫处理的叶绿素含量较高,叶绿素a/b值较低,类胡萝卜素含量在生长发育中早期较低而后期较高;SOD、POD、CAT活性呈由低到高再降低的趋势,MDA含量持续升高;遮荫处理的SOD、POD活性和MDA含量均低于全光照处理,CAT活性则表现为先高于后低于全光照处理.适度的遮荫可以提高半夏叶绿素的含量,降低膜系统受到的伤害,改善半夏的倒苗状况,为半夏叶片的生长提供较好的环境.     

不同培养条件对半夏悬浮培养细胞生长及总生物碱形成的影响

研究了半夏悬浮培养过程中细胞鲜重的变化和MS培养基组分中碳源、钙盐、Fe盐及肌醇变化对半夏悬浮培养细胞生长及总生物碱合成的影响。结果表明 ,半夏细胞的生长曲线呈“S”型 ,细胞的最佳收获时间为 2 1d。在两种碳源中 ,葡萄糖比蔗糖利于细胞的生长和总生物碱的合成 ,最适浓度为 2 0g L。最适合细胞生长和总生物碱形成的钙盐浓度为 1 8mmol L、Fe盐浓度为 0 0 6mmol L和肌醇浓度为 10 0mg L。     

银杏液体悬浮细胞培养产生黄酮的研究

探讨了光照与黑暗、培养时间、细胞聚集体大小、前体物对银杏液体悬浮细胞系产生黄酮的影响。试验结果表明:光照条件下培养14 d黄酮含量最高,3～4 mm粒径的细胞聚集体是黄酮合成的适宜细胞聚集体,前体物L-苯丙氨酸和乙酸钠对黄酮合成无明显的促进作用,且对细胞生长有抑制作用。     

营养胁迫诱导的油菜子叶衰老过程中IAA氧化酶和细胞分裂素氧化酶活性的变化

以离体油菜子叶为材料,研究了营养胁迫诱导的子叶衰老过程中吲哚乙酸氧化酶(IAA氧化酶)和细胞分裂素氧化酶活性的变化。在光照条件下,离体子叶在不含任何无机元素的0．8％的琼脂中培养9d后,出现明显的衰老迹象(叶绿素含量下降,丙二醛含量上升),15d时完全死亡。在营养胁迫诱导的衰老过程中,IAA氧化酶和细胞分裂素氧化酶的活性表现出相似的变化趋势,在诱导处理1d时,两种酶的活性均比处理前有明显下降,之后又随着衰老进程逐渐上升。IAA氧化酶活性在诱导处理11d时达到高峰,超出处理前30％以上;比对照高出1倍以上;而细胞分裂素氧化酶活性在诱导处理13d时达到高峰,比对照高出3倍以上,也超过了处理前的水平。衰老过程中IAA氢化酶和细朐分裂素氧化酶活性的上升可能是导致内源激素含量下降的重要原因。     

黑暗和光照对丹参培养细胞生长和生理生化特性的影响

以丹参(Salvia miltiorrhiza Bunge)幼嫩叶片为外植体,在Ms 2,4-D0.5mg/L 6BA1．5mg/L培养基上诱导形成愈伤组织,愈伤组织置于不同条件下培养。探讨丹参细胞培养过程中蛋白质、酶活性的变化与细胞生长周期之间的关系。结果显示黑暗和光照下,培养细胞的生长周期为27天;可溶性蛋白含量和超氧化物歧化酶(SOD)活性均出现两个明显峰值,而过氧化物酶(POD)活性变化较复杂。丹参细胞生长呈“S”形周期性变化,其可溶性蛋白、酶活性也呈相应的周期性变化。     

中国仓鼠卵巢细胞的悬浮适应及代谢特征

目的：通过悬浮适应,使中国仓鼠卵巢细胞（CHO细胞）获得悬浮生长的特性,并可在悬浮培养条件下较快地生长。方法：将CHO细胞以3×10^5／mL接种于100mL的三角瓶内,培养时加入1％小牛血清、1g/LPIuronic F-68、25μg／mL硫酸葡聚糖,培养体积35mL,摇床转速90r／min,每24h离心换液,当细胞增殖为2×10^6／mL时传代。结果：经过悬浮适应,细胞的平均比生长速率由适应最初的0．27／d提高为适应后的0．48／d,最大总细胞密度由适应初期的2．5×10^6／mL提高为适应后的6．3×10^6／mL,目的蛋白活性也由适应前的2781U／mL提高为适应后的8878U／mL,适应后细胞的葡萄糖平均比消耗率为1．42μmol／（10^6细胞·d）,低于适应前的2．16μmol／（10^6细胞·d）。结论：贴壁生长的CHO细胞经过悬浮适应,不仅可以在悬浮培养条件下快速生长,而且细胞对葡萄糖的利用率也得到提高。     

干旱胁迫对菜苔叶片保护酶活性和膜脂过氧化的影响

以3个不同耐旱性的菜苔（Brassica parachinensis L.H.Bailey）品种为试材,研究了干旱胁迫对叶片保护酶活性和膜脂过氧化的影响及其与抗旱性的关系。干旱胁迫条件下,菜苔叶片的电解质外渗率和MDA含量呈上升趋势,叶绿素含量、抗坏血酸含量和SOD活性呈下降趋势,CAT活性表现为先上升后下降。耐旱品种比不耐旱品种具有较高的叶绿素含量和抗坏血酸含量,具有较低的电解质外渗率和MDA含量;耐旱品种的SOD活性比不耐旱品种下降幅度小。轻度干旱胁迫下,耐旱品种的CAT活性上升幅度比不耐旱品种高;重度干旱胁迫下耐旱品种的CAT活性下降程度比不耐旱品种低。耐旱品种的POD活性在干旱条件下先上升而后降低,不耐品种的POD活性处于下降趋势。干旱6d后,耐旱品种的SOD、CAT和POD活性显著高于不耐旱品种。     

微流控芯片技术在细胞生物学研究中的应用进展

20世纪90年代以来,微流控芯片技术得到了快速发展。由于具有小型化、集成化、高通量、低消耗、分析快速等特点,微流控芯片作为一种新型的生物学研究平台,能够提供传统方法不具备的精细和可控制的细胞研究条件,在细胞生物学研究领域中得到了广泛关注。该文主要介绍其在细胞培养、分选、裂解、计数、凋亡检测、迁移、单细胞捕获、细胞间作用等方面的研究进展。     

The role of glutamine, serum and energy factors in growth of enterocyte-like cell lines

Background: Glutamine is routinely added to most cell cultures. Glutamine has been found to be the preferential nutrient to the rapidly replicating intestinal mucosa, but whether this is a metabolic effect or due to other properties of this amino acid is not determined. To study the importance of glutamine on the growth of two enterocyte-like cell lines, the effects of depriving the media or supplementing it with glutamine were assessed in media with different serum and energy supplements. Methods: CaCo-2 and HT-29 cells were grown in serum-free medium, with fetal bovine or synthetic serum, and with or without glucose or galactose. The glutamine content was varied between 0 and 4 mM. All growth assays were performed in triplicate by counting in a hemocytometer. Results: Both cell lines were dependent of serum factors for growth, but displayed distinct requirements on glutamine supplementation. Glutamine was an obligate supplement with dose-dependent correlation to growth (r=0.87, p<0.01) for CaCo-2 cells cultured in synthetic, but not in fetal bovine serum. In HT-29 cells, the correlation between glutamine and growth was significant (r=0,68, p<0,05) only in fetal bovine serum in the absence of galactose. Conclusion: This study shows that glutamine has different growth stimulating effects on two enterocyte-like cell lines studied. This could reflect different modes of action of glutamine on proliferation and differentiation in an enterocyte cell population.     

Besides Purkinje cells and granule neurons: an appraisal of the cell biology of the interneurons of the cerebellar cortex

Ever since the groundbreaking work of Ramon y Cajal, the cerebellar cortex has been recognized as one of the most regularly structured and wired parts of the brain formed by a rather limited set of distinct cells. Its rather protracted course of development, which persists well into postnatal life, the availability of multiple natural mutants, and, more recently, the availability of distinct molecular genetic tools to identify and manipulate discrete cell types have suggested the cerebellar cortex as an excellent model to understand the formation and working of the central nervous system. However, the formulation of a unifying model of cerebellar function has so far proven to be a most cantankerous problem, not least because our understanding of the internal cerebellar cortical circuitry is clearly spotty. Recent research has highlighted the fact that cerebellar cortical interneurons are a quite more diverse and heterogeneous class of cells than generally appreciated, and have provided novel insights into the mechanisms that underpin the development and histogenetic integration of these cells. Here, we provide a short overview of cerebellar cortical interneuron diversity, and we summarize some recent results that are hoped to provide a primer on current understanding of cerebellar biology.     

哺乳动物体细胞核移植中供体细胞的研究进展

在哺乳动物体细胞核移植中,供体细胞是影响其效率的主要因素之一。供体细胞的类型、细胞周期、细胞的培养代数、冷藏与冷冻处理,以及供体动物的性别、年龄等都可能影响核移植胚胎的发育。根据现有资料,简要综述了在哺乳动物体细胞核移植中有关供体细胞的研究进展。     

微囊化K562细胞生长周期及代谢特性的研究

以K562细胞为模型,分别进行微囊化和游离培养,运用流式细胞术考察两种培养体系下细胞周期和生长代谢变化;建立数学模型,模拟了两种培养体系下细胞的生长活性和代谢特性。实验发现：微囊化培养过程中的K562细胞处于DNA合成期（S期）的百分含量显著高于游离培养,并且细胞保持较高的增殖活性。模型计算表明,所建模型动力学参数能够很好地描述微囊化和游离两种培养体系下细胞的代谢情况;对细胞活性的理论计算表明,微囊化的细胞具有较高的增殖和代谢活性,同时细胞能够较长时间保持此活性;模型参数表明,两种培养体系下,葡萄糖对细胞生长的影响无显著差别 (k^Free_L≈k^APA_L),乳酸对游离培养细胞的生长具有明显抑制作用,但对微囊化培养细胞抑制作用较小(kFreeL>≈kAPAL)。     

Ultrastructural evidence for two-cell and three-cell neural pathways in the tentacle epidermis of the sea anemone Aiptasia pallida.

Sensory and ganglion cells in the tentacle epidermis of the sea anemone Aiptasia pallida were traced in serial transmission electron micrographs to their synaptic contacts on other cells. Sensory cell synapses were found on spirocytes, muscle cells, and ganglion cells. Ganglion cells, in turn, synapsed on sensory cells, spirocytes, muscle cells, and other neurons and formed en passant axo-axonal synapses. Axonal synapses on nematocytes and gland cells were not traced to their cells of origin, i.e., identified sensory or ganglion cells. Direct synaptic contacts of sensory cells with spirocytes and sensory cells with muscle cells suggest a local two-cell pathway for spirocyst discharge and muscle cell contraction, whereas interjection of a ganglion cell between the sensory and effector cells creates a local three-cell pathway. The network of ganglion cells and their processes allows for a through-conduction system that is interconnected by chemical synapses. Although the sea anemone nervous system is more complex than that of Hydra, it has similar two-cell and three-cell effector pathways that may function in local responses to tentacle contact with food.     

Of mice, frogs and flies: generation of membrane asymmetries in early development

Embryonic development begins with cleavage of the fertilized egg. Cleavage comprises two major processes: cytokinesis and formation of a polarized epithelial cell layer. The focus of this review is comparison of the generation of membrane polarity during embryonic cleavage in three different developmental model systems. In mammalian embryos, as exemplified by analysis of the mouse, generation of distinct membrane domains is uncoupled from cleavage divisions and is initiated in a specific developmental phase, called compaction. In Xenopus laevis embryos, generation of polarized blastomeres occurs simultaneously with cytokinesis. The origin of specific membrane domains of X. laevis polar blastomeres, however, can be traced back to oogenesis. Finally, in Drosophila melanogaster, generation of polarized cells occurs at cellularization. The relevance of cell adhesion, cell junctions and cytocortical scaffolds will be discussed for each of the model systems. Despite enormous morphologic differences, the three models share many common features; in particular, many important molecular interactions are conserved.     

Mammalian testis: a target of in vivo electroporation

Mammalian spermatogenesis consists of three biologically significant processes: stem cell self-renewal and differentiation, meiosis, and haploid cell morphogenesis. Understanding the molecular mechanisms behind these processes might provide clues to the puzzle of species preservation and evolution, and to treatments for male infertility. However, few useful in vitro systems exist to investigate these processes at present. To elucidate these mechanisms, in vivo electroporation of the testis might be a convenient option. Since DNA solution can be injected into the seminiferous tubule via the rete testis, similar to germ cell transplantation, it is easy to transfect expression vectors into various differentiated germ cells and supporting Sertoli cells with adequate electric shock. Unfortunately, it is difficult to create transgenic animals using this method because of its low efficiency. However, gain- and loss-of-function assays, promoter assays, and tagged-protein behavior assays can be conducted with this technique, as in in vitro culture systems.     

Towards the development of a minimal cell model by generalization of a model of Escherichia coli: use of dimensionless rate parameters.

A model of a minimal cell would be a valuable tool in identifying the organizing principles that relate the static sequence information of the genome to the dynamic functioning of the living cell. Our approach for developing a minimal cell model is to first generalize an existing model of Escherichia coli by expressing reaction rates as ratios to a set of reference parameters. This generalized model is a prototype minimal cell model that will be developed by adding detail to explicitly include each chemical species. We tested the concept of a generalized model by testing the effect of scaling all enzyme-catalyzed reactions in the E. coli model. The scaling has little effect on cellular function for a wide range of kinetic ratios, where the kinetic ratio is defined as the rate of all enzyme-catalyzed reactions in a given model relative to those in the E. coli model.     

牛脂肪间充质干细胞的分离、培养与鉴定

为了给组织工程提供种子细胞,对牛间充质干细胞(Adipose-derived stem cells,ADSCs)进行体外分离培养。首先应用胶原酶消化法分离牛ADSCs,进行体外培养、连续传代,并观察细胞的形态变化,通过细胞计数绘制生长曲线,细胞压片进行染色体分析,采用细胞免疫荧光化学方法检测细胞表面标记,利用成骨分化和成脂分化检测其分化能力。结果显示牛ADSCs体外培养时细胞形态呈成纤维细胞样,增殖稳定;Vimentin、CD49d、CD13表达呈阳性,CD34表达呈阴性;成骨诱导条件下的细胞碱性磷酸酶活性高,茜素红染色呈阳性;成脂诱导条件下细胞周围脂滴明显,油红-O染色呈阳性。结果证明牛ADSCs体外生长稳定、增殖速度快、定向分化能力强,简易的体外分离培养及诱导方法为其在组织工程中的应用奠定了基础。