红花组织培养中细胞分化的超微结构研究

红花子叶脱分化及愈伤组织形成的超微结构研究揭示：处于脱分化过程中的细胞代谢极其活跃。贮藏的脂类在诱导的初期即被利用,伴随细胞脱分化的同时,叶绿体了同样经历一个脱分化的过程,内膜肿胀解体,同时质体分裂或出芽增殖。转移到分化培养基后,叶绿体细胞逐渐恢复,分化成熟,处于脱分化的细胞以及愈伤组织细胞中,常可以观察到处于质膜与细胞之间的所谓壁旁体的囊泡结构,对壁旁体可能的功能以及叶绿体脱分化的原因进行了讨论     

植物离体培养中器官发生调控机制的研究进展

本文就离体培养的植物组织对生长调节物质的吸收和代谢,外源生长调节物质对内源激素水平的影响,内源激素对细胞脱分化和再分化的调控,生长素和细胞分裂素基因与器官发生的关系,与器官发生有关的基因和特异蛋白等问题的研究进展进行了评述,并对下一步研究提出了自己的看法。     

植物离体培养中器官发生调控机制的研究进展

本文就离体培养的植物组织对生长调节物质的吸收和代谢，外源生长调节物质对内源激素水平的影响，内源激素对细胞脱分化和再分化的调控，生长素和细胞分裂素基因与器官发生的关系，与器官发生有关的基因和特异蛋白等问题的研究进展进行了评述，并对下一步研究提出了自己的看法。     

环境pH值对籼粳稻幼苗呼吸的影响

籼稻和粳稻无论在田间栽培条件下或外植体离体培养条件下,它们彼此间在某些生长习性上都存在明显的差异。但是迄今却很少见到有关籼、粳稻差异的生理学基础的研究报道。鉴于呼吸是代谢的中心环节,以及环境pH值能够明显影响植物离体培养物的脱分化启动和再分化潜力,我们进行了     

‘SK4—316’胡萝卜体胚的诱导和培养

以'SK4-316'胡萝卜无菌苗的下胚轴为外植体,研究不同培养基配方和培养条件对愈伤组织诱导、体细胞胚间接发生及其同步化培养的影响,以及不同脱分化时间、脱分化培养基及外植体续存时间对体细胞胚直接发生的诱导及其培养的影响.结果表明:含3%蔗糖、0.8%琼脂的1/2MS + 2,4-D 2.5 mg/L + 6-BA(或KT)0.5 mg/L + CH 300 mg/L是诱导愈伤组织的良好培养基;1/2MS + 2,4-D 1.25 mg/L + KT 0.25 mg/L + 6-BA 0.25 mg/L(含3%蔗糖)适于愈伤组织分化并诱导体胚发生,0.02% ABA对体胚的诱导有促进作用,0.06% ABA或15% PEG能促进体胚成熟;外植体在MS + 2,4-D 1.0 mg/L固体培养基上脱分化培养48 h,再转入MS + CH 300 g/L液体培养基中可诱导体胚直接发生,但随着外植体续存于诱导培养基中时间的延长,体胚发生变异的几率也渐增.     

扫描电镜下的器官发生——植物胚状体和不定芽的分化(封二说明)

在离体培养条件下,植物组织的器官分化基本可分为不定芽发生及胚状体(体细胞胚)发生这两大主要类型。它们既可以从外值体(即接种材料)的表面直接发生,也可以由外植体经脱分化后形成愈伤组织,由愈伤组织再分化而产生。 封二,图1、2,为刺五加(Acanthopanaxsenticosus)的胚接种在MS 2,4-D0.5ppm培养基上1—2个月后,由子叶及胚轴直接分化出大量胚状体的情况。胚状体的分化是不同步的,     

花椰菜(Brassica oleracea var botrytis)下胚轴培养过程中过氧化物酶活性及同工酶谱的变化

花椰菜下胚轴外植体在MS+6BA 5 ppm的培养基上能分化出芽,在MS+2,4-D2ppm的培养基上能脱分化而形成愈伤组织。用3种不同的酚类物质(咖啡酸、阿魏酸、愈创木酚及联苯胺)作氢供体发现分化过程中的过氧化物酶活性高于脱分化过程,其中以咖啡酸作氢供体显示的活性最高,阿魏酸及愈创木酚次之,而联苯胺最小。用聚丙烯酰胺凝胶电泳分离阴极向及阳极向过氧化物酶同工酶,在分化及脱分化培养过程中均不断出现新的酶带,前者有13条,后者为11条,两者的差别主要在阴极向酶带,在分化过程中多了两条酶带(C_1和C_3),同时C_2带活性也比脱分化的高。阳极向酶带也有差别,A_2和A_2两条酶带在分化过程中逐渐加强,但是在脱分化过程中却逐渐消失。反映了两个过程生理上的差别。     

烟草表皮细胞薄层培养系统中多种组织器官发生的研究

利用烟草表皮细胞薄层培养系统,研究了离体培养下表皮毛和气孔的发生,发现斜向分裂和不均等分裂是与脱分化细胞再分化相关的分裂方式,并观察了愈伤组织气孔的适应性分化现象,此外,在茎表皮细胞薄层上成功地发生了不定根,利用花茎表皮细胞薄层也在其愈伤组织上发生了花芽。     

烟草叶片外植体脱分化与再分化过程中FtsZ蛋白的表达（简报）

在植物叶肉细胞的脱分化、再分化过程中伴随着叶绿体与质体相互转化的过程。已高度分化的叶肉细胞脱分化为分生状态细胞时．其中的原质体主要由叶绿体出芽生殖产生．偶尔可以看到某些叶绿体分裂或分裂与出芽同时出现的情况。此外,叶绿体在出芽产生原质体的同时自身逐渐被巨大的淀粉粒所充满．从而转变为淀粉体。     

大麦直接游离小孢子培养中的脱分化启动和胚胎发生

采用DNA特异荧光染料,在共聚焦激光扫描显微镜下,对直接游离的大麦小孢子,在预处理过程中的脱分化启动,以及培养过程中的胚胎发生进行了较详尽的细胞学特征的观察,并在主要发育途径上与低温预处理后的小孢子进行了比较.研究结果表明:(1)小孢子的脱分化启动在预处理的12h内就已开始.完成脱分化启动的小孢子在细胞学上的主要特征为:细胞体积明显增大;核与核仁体积也显著增加,核仁极其明显,且高度浓缩,核/质比例高.(2)不同的预处理方法,都是促使离体小孢子完成脱分化启动,从而激发胚胎发生过程.(3)预处理方法可通过改变离体小孢子第1次有丝分裂的方式,而形成不同的主要发育途径.经低温预处理以形成A途径为主,直接游离以形成B途径为主.     

Ultrastructure of Early Secondary Embryogenesis by Multicellular and Unicellular Pathways in Cork Oak (Quercus suber L.)

Early cellular events during secondary embryogenesis were studiedin a cork oak recurrent embryogenic system in which embryosarise either in a multicellular budding pathway from a compactmass of proliferation or from isolated single cells in friablecallus. The compact mass of proliferation originated from theepidermal cells at the hypocotyl whose growth and convolutionwas characterized by a decrease in the nucleus/cytoplasm ratioand a marked increase in storage products. The transition fromthe compact mass to meristematic primordia occurred at the peripheryand was accompanied by cell dedifferentiation and a drasticreduction of storage products. Meristematic primordia evolvedto globular embryos by the organization of a protodermis andtwo internal centres. Microscope analysis of friable callusshowed an hypothetical sequence from single cells to aggregatesof a few cells, meristematic cell clusters and globular embryos.Single cells showed typical features of embryogenic cells suchas rich cytoplasm and a large number of starch grains and lipidbodies. A progressive cell dedifferentiation and a drastic reductionof storage products was observed when aggregates of a few cellsand meristematic cell clusters were compared. Progressive bipolarizationin large meristematic cell clusters initiated globular embryoformation. The comparison of both embryogenic pathways at theultrastructural level showed that subcellular changes followa similar sequential pattern, especially with regard to thestorage products. The possible role of plastid extrusions andmultivesicular bodies in the changing pattern of starch metabolismduring embryogenesis is discussed. Copyright 2001 Annals ofBotany Company Quercus suber L, cork oak, somatic embryogenesis, multicellular budding, friable callus, ultrastructural studies     

杜仲次生木质部细胞分化和脱分化过程中ATPase的超微细胞化学定位

采用磷酸铅沉淀技术,对杜仲（Ｅｕｃｏｍｍｉａ ｕｌｍｏｉｄｅｓ Ｏｌｉｖ．）次生木质部细胞分和脱化过程进行了ＡＴＰａｓｅ的超微细胞化学定位。随着分化过程中细胞程序性死亡（ｐｒｏｇｒａｍｍｅｄ ｃｅｌｌ ｄｅａｔｈ,ＰＣＤ）程度的加深,ＡＴＰａｓｅ在细胞核上的分布由少变多,而在各种细胞器上的分布由有到无,并且随着细胞质的解体,ＡＴＰａｓｅ在细胞壁内侧和纹孔处的分布也由少到多,说明它们的变化是由核基因     

杜仲次生木质部细胞分化和脱分化过程中ATPase的超微细胞化学定位

The ultracytochemical localization of ATPase in the secondary xylem cells during their differentiation and dedifferentiation in the girdled Eucommia ulmoides Oliv. was carried out using a lead phosphate precipitation technique. Throughout the differentiation, which is a typical programmed cell death (PCD) process, ATPase deposits increased in the nucleus but decreased and progressively disappeared in the cell organelles. At the same time, the distribution of ATPase increased in the inner face of the cell wall and pits with cytoplasmic degeneration. The results demonstrated that the PCD was an energy dependent active process and was controlled by nuclear genes. On the other hand, the distribution of ATPase in the intercellular spaces increased with the formation of the new cambium resulted from the dedifferentiation of the secondary xylem cells after girdling. However, ATPase was not found in the nucleus of the dividing cells, suggesting that nutrients were transported through protoplast during differentiation, and through both protoplast and apoplast during dedifferentiation. Thus, the energy required in cell division was provided mainly by intercellular spaces. These findings indicate that the dynamic distribution of ATPase reflected which cell component was actively taking part in the cell metabolism at various stages of the plant development, and its distribution was associated with the physiological state of the cell. Based on the characteristic distributions of ATPase, the critical stage of cell differentiation and the relationship between the critical stage and dedifferentiation were discussed.     

竹节海棠叶外植体脱分化启动的细胞学观察

竹节海棠叶外植体接种于MS+6-BA1ppm+NAA0.1ppm培养基上。外植体脱分化启动过程中,表皮及叶肉细胞主要以劈裂的无丝分裂方式进行分裂:最初核延伸为纺锤形,核仁大而明显,使整个核的轮廓呈“眼”状;随着核的中部出现裂缝,核断开成为两部分,稍后,由原来的母细胞形成两个子细胞。由于核分裂前向细胞中央移动的距离及断裂时断裂面的不同,从而造成细胞团内细胞大小悬殊及分裂面严重混乱的不等分裂现象。文中对栅栏组织细胞脱分化启动后重复进行无丝分裂形成梯状细胞团的现象也进行了讨论。     

Changes in the phospholipid molecular species composition of soybean hypocotyl and cotyledon after dedifferentiation

The effect of dedifferentiation on the molecular species composition of soybean phospholipids was studied by using hypocotyl, cotyledon and the suspension culture cells established from those organs. Three major phospholipids (phosphatidylcholine, phosphatidylethanolamine and phosphatidylinositol) and phosphatidylmonomethylethanolamine were composed of twelve molecular species. Major species were 1-palmitoyl-2-linoleoyl, 1-obeoyl-2-linoleoyl, 1-palmitoyl-2-linolenoyl and 1-linoleoyl-2-linoleoyl species. Different proportions of the molecular species were found among the three major phospholipids, but phosphatidylmonomethylethanolamine was composed of the same proportions of the molecular species as those of phosphatidylethanolamine. After dedifferentiation, the 1-palmitoyl-2-linoleoyl species increased in the cell established from hypocotyl. In the cells established from cotyledon, the 1-palmitoyl-2-linolenoyl species increased dramatically. In both cells, the 1-palmitoyl-2-linolenoyl species increased in response to increase in the 2,4-dichlorophenoxyacetic acid concentrations and the progress of cell growth.     

Dedifferentiation derived cells exhibit phenotypic and functional characteristics of epidermal stem cells

Differentiated epidermal cells can dedifferentiate into stem cells or stem cell‐like cells in vivo. In this study, we report the isolation and characterization of dedifferentiation‐derived cells. Epidermal sheets eliminated of basal stem cells were transplanted onto the skin wounds in 47 nude athymic (BALB/c‐nu/nu) mice. After 5 days, cells negative for CK10 but positive for CK19 and β₁‐integrin emerged at the wound‐neighbouring side of the epidermal sheets. Furthermore, the percentages of CK19 and β₁‐integrin⁺ cells detected by flow cytometric analysis were increased after grafting (P < 0.01) and CK10⁺ cells in grafted sheets decreased (P < 0.01). Then we isolated these cells on the basis of rapid adhesion to type IV collagen and found that there were 4.56% adhering cells (dedifferentiation‐derived cells) in the grafting group within 10 min. The in vitro phenotypic assays showed that the expressions of CK19, β₁‐integrin, Oct4 and Nanog in dedifferentiation‐derived cells were remarkably higher than those in the control group (differentiated epidermal cells) (P < 0.01). In addition, the results of the functional investigation of dedifferentiation‐derived cells demonstrated: (1) the numbers of colonies consisting of 5–10 cells and greater than 10 cells were increased 5.9‐fold and 6.7‐fold, respectively, as compared with that in the control (P < 0.01); (2) more cells were in S phase and G2/M phase of the cell cycle (proliferation index values were 21.02% in control group, 45.08% in group of dedifferentiation); (3) the total days of culture (28 days versus 130 days), the passage number of cells (3 passages versus 20 passages) and assumptive total cell output (1 × 10⁵ cells versus 1 × 10¹² cells) were all significantly increased and (4) dedifferentiation‐derived cells, as well as epidermal stem cells, were capable of regenerating a skin equivalent, but differentiated epidermal cells could not. These results suggested that the characteristics of dedifferentiation‐derived cells cultured in vitro were similar to epidermal stem cells. This study may also offer a new approach to yield epidermal stem cells for wound repair and regeneration.     

Microvesicles Derived from Human Umbilical Cord Mesenchymal Stem Cells Facilitate Tubular Epithelial Cell Dedifferentiation and Growth via Hepatocyte Growth Factor Induction

During acute kidney injury (AKI), tubular cell dedifferentiation initiates cell regeneration; hepatocyte growth factor (HGF) is involved in modulating cell dedifferentiation. Mesenchymal stem cell (MSC)-derived microvesicles (MVs) deliver RNA into injured tubular cells and alter their gene expression, thus regenerating these cells. We boldly speculated that MVs might induce HGF synthesis via RNA transfer, thereby facilitating tubular cell dedifferentiation and regeneration. In a rat model of unilateral AKI, the administration of MVs promoted kidney recovery. One of the mechanisms of action is the acceleration of tubular cell dedifferentiation and growth. Both in vivo and in vitro, rat HGF expression in damaged rat tubular cells was greatly enhanced by MV treatment. In addition, human HGF mRNA present in MVs was delivered into rat tubular cells and translated into the HGF protein as another mechanism of HGF induction. RNase treatment abrogated all MV effects. In the in vitro experimental setting, the conditioned medium of MV-treated injured tubular cells, which contains a higher concentration of HGF, strongly stimulated cell dedifferentiation and growth, as well as Erk1/2 signaling activation. Intriguingly, these effects were completely abrogated by either c-Met inhibitor or MEK inhibitor, suggesting that HGF induction is a crucial contributor to the acceleration of cell dedifferentiation and growth. All these findings indicate that MV-induced HGF synthesis in damaged tubular cells via RNA transfer facilitates cell dedifferentiation and growth, which are important regenerative mechanisms.     

MULTIPLE FORMS OF β -GALACTOSIDASE AND THE CHANGES IN ITS PATTERN DURING DEVELOPMENT OF DICTYOSTELIUM DISCOIDEUM

Four forms of β-galactosidase (EC 3.2.1.23) were separated from cell extracts of Dictyostelium discoideum by polyacrylamide gel electrophoresis. Changes in the pattern of multiple forms of this enzyme during differentiation and dedifferentiation were studied. The band closest to the anode (Band 1) existed throughout development, but the other three bands appeared and disappeared at certain stages. One of those bands was specific for the vegetative stage (Band 3), and another for the morphogenetic stages (Band 2). The last one predominantly appeared during dedifferentiation of disaggregated slug cells (Band 4), although it was slightly detected during culmination. During the process of dedifferentiation, the increase in activity of Band 4 coincided with the decrease in Band 2. After the completion of dedifferentiation, Band 4 also disappeared, only Band 1 remaining. These multiple forms were not only electrophoretically separable but different in their sensitivities to various inhibitors. All forms of the enzyme were localized in subcellular particles, probably in lysosomes and phagosomes, and the relative activities in these two fractions varied during development.     

Early events following amputation of adult newt limbs given regeneration inhibitory doses of X irradiation

X irradiation, (2000 R) prevented regeneration but had no apparent effect on morphological dedifferentiation. DNA synthesis and mitotic activity increased significantly in irradiated limbs after amputation but not as much as in regenerating unirradiated controls. Even though fewer mitotic cells were seen in irradiated limbs, those that occurred were distributed normally throughout the stump. The results indicate that morphological dedifferentiation and entry into the cell cycle are dissociable events and that X rays act by interfering with the latter. They also show that some cells in irradiated amputated limbs not only synthesize DNA but also divide.