一种大量制备小鼠不同分化阶段成红细胞的简单方法

一种大量制备小鼠不同分化阶段成红细胞的简单方法张世馥，王鑫，刘彤，薛社普（中国医学科学院基础医学研究所细胞生物室，北京１００００５）在细胞分化、基因表达调控和某些药物诱导分化原理的研究中，哺乳类红细胞一向被人们作为一种理想的分化模型，它们在分化、发育...     

OP9-DL1细胞共培养系统在T细胞体外定向诱导分化中的应用

T细胞是一类在机体免疫应答中起核心作用的免疫活性细胞。T细胞的分化发育指来自骨髓的淋巴样祖细胞(lymph-oid pregnitors)进入胸腺增殖、分化,并从胸腺皮质迁移至髓质,成为成熟T细胞的过程。长期以来,T细胞发育的体内外研究一直依赖于胸腺。但人或鼠的胸腺组织来源有限且取材困难,加大了T细胞发育的研究难度。近年来,随着T细胞发育过程中一些关键分子的发现及其研究的不断深入,一种可高效诱导造血干/祖细胞在体外向T细胞分化的简单模型骨髓基质细胞OP9-DL1体外培养系统得以建立,这种系统的出现大大简化了T细胞分化发育的研究,本文将就其在T细胞体外定向诱导分化中的应用作一综述。     

髓质区单阳性T细胞发育研究进展和挑战

胸腺T细胞发育分为CIM／CD8双阴性、双阳性和单阳性三个主要阶段，相对于双阴性和双阳性阶段，单阳性阶段细胞发育一直未受到应有的关注。但近年来的研究提示，T细胞发育中的许多重要事件都发生于胸腺髓质区，即单阳性阶段。全面、系统地研究单阳性细胞发育必将深化人们对功能成熟、阴性选择、Treg和NKT细胞定向分化等重要问题的认识。本文在回顾该领域研究进展的同时，着重讨论发育过程中的阴性选择和NKT细胞定向分化问题，包括发育阻滞对其产生的影响。此外，     

线粒体功能在B细胞发育分化及活化中的作用

B细胞从骨髓起始发育，到外周器官成熟，是介导体液免疫的重要细胞。在抗原的刺激下，B细胞分化为浆细胞产生特异性抗体或分化为记忆B细胞。线粒体是细胞的能量产生及代谢中心，负责为细胞提供充足的能量。当线粒体功能受损时，会影响其能量供应，从而改变细胞的命运；而细胞的不同状态也会影响线粒体的功能。近年研究显示，线粒体功能在B细胞的发育和活化中起着重要作用，以适应细胞整个周期中遇到的表型和环境变化。本文主要综述了线粒体功能在B细胞发育、分化及活化过程中的作用，以及在B细胞的不同阶段、不同状态下发生的改变，为探究线粒体相关疾病的机制研究提供新思路，为B细胞相关疾病的治疗提供新的理论依据。     

癌基因研究进展及某些研究技术

第一讲 概述 肿瘤是许多癌细胞的聚集态,它们以失去控制的方式生长。人和其他生物一样,是由上百万细胞组织起来的有机体,这些细胞彼此之间存在着紧密的联系和控制。在生物学中一个最有兴趣的领域是研究细胞如何控制生长和分化。在采精卵发育为成体的过程中,这些细胞要经过生长、分化、专门化、停止生长、彼此互通信息等一系列复杂过程,而在肿瘤中这些     

心肌分化的调控

骨髓间质干细胞被认为是生物体内的自体修复库。了解生理情况下心肌细胞分化的调控过程是进行骨髓间质干细胞心肌分化研究的基础。根据对动物胚胎发育的研究，心脏的发育、心肌细胞的分化过程是多种基因和多种因子，通过多种途径，在不同时间点上进行调控的复杂过程。本文对与心肌细胞分化相关的基因、蛋白质、酶和生长因子等在心脏发育、心肌分化中的作用作一综述，并探讨了心脏不同部分心肌细胞形成过程中有关基因表达的区域和时相变化，为进一步研究骨髓间质干细胞向心肌分化的调控机制奠定基础。     

胸腺微环境及其作用

胸腺是Ｔ淋巴细胞分化发育的场所，胸腺微环境由胸腺基质细胞、细胞外基质和细胞因子等构成。在胸腺基质细胞中，上皮细胞最多，分布最广。上皮细胞的分化、成熟和增殖依赖于胸腺内发育阶段的各Ｔ细胞亚群。位于胸腺内不同区域的基质细胞在Ｔ细胞发育的不同阶段起重要作用。血胸屏障是胸腺微环境中一个特殊结构，前Ｔ细胞可通过粘附分子，在趋化因子等作用下穿过血胸屏障进入胸腺，在胸腺内完成其分化发育过程。     

B淋巴细胞发育分化中的特征性表面分子

B淋巴细胞是体内唯一能够产生抗体的细胞,介导体液免疫.B淋巴细胞发育分化是一个十分有序的过程,大体分为五个阶段,祖B细胞、前B细胞、未成熟B细胞、过渡B细胞和成熟B细胞.每个阶段均有特征性的表面标志,其表面分子的表达呈规律性改变.各发育分化阶段特征性表面分子表达异常可引起某些疾病,虽然目前国外对B淋巴细胞发育分化情况有较多研究,但对于B淋巴细胞发育分化过程中的一些问题及对某些疾病的影响与相关机制仍不明确,需进一步研究.     

小鼠诱导型多潜能干细胞定向分化为功能性肝细胞

目的:探讨在体外诱导型多潜能干细胞(IPS)能否遵循正常肝脏发育途径定向分化为具备良好生理功能的肝细胞。方法:根据正常肝脏体内发育的规律,序贯应用肝脏发育所需的生长因子,设计特色培养基,诱导IPS细胞遵循正常发育途径定向高效分化为肝细胞:首先将IPS细胞定向分化为前层确定性内胚层细胞(ADE),再进一步分化为肝细胞。结果:在本研究中IPS细胞在体外可循正常发育通路定向诱导为肝细胞:先分化为前层内胚层细胞,再继续分化为肝细胞。分化的各时段实时定量RT-PCR检测显示:在mRNA表达的水平,IPS细胞经历了肝细胞在体内从胚胎干细胞的发育过程,内胚层细胞特异性RNA如CXCR4、Sox17和FOXA2在分化的早期明显升高,而早期肝细胞特异性RNA如HNF4和AFP在分化的中期开始升高,肝细胞成熟特异性RNA如白蛋白在分化的晚期才达高峰,同时IPS细胞的未分化标记RNA如OCT4在早期就明显下调。同mRNA表达一致,在分化的第20d,大部分的肝脏细胞表达甲胎蛋白和白蛋白(阳性率80%)。更为重要的是IPS来源肝细胞在体外具有典型成熟肝细胞的功能,能够摄取和释放靛青绿(ICG)并具有药物代谢酶细胞色素P450酶的活性。结论:IPS细胞在体外能够遵循正常肝脏发育通路,序贯表达肝脏发育各阶段的特色基因和蛋白,能够高效分化为有功能的肝细胞,本研究可能为临床终末期肝病进行细胞治疗提供肝细胞。     

蛋白聚糖Agrin与神经系统

神经系统的发育是一个极其复杂的过程，它包括形态的形成以及结构之间联系的建立。在这一过程中，细胞与细胞间的相互作用是一个关键因素。这种细胞间相互作用包括分化诱导作用与分化抑制作用，细胞外一些物质如细胞因子、粘附分子、激素及蛋白聚糖等都可介导细胞之间的相互作用，其中蛋白聚糖在发育中对于促进细胞间相互联系起着重要的作用。蛋白聚糖分布广泛，并与神经系统的发育具有密切关系。     

微循环灌流状态的在体研究

本文介绍采用国内外先进的显微高速动态分析装置,定性和定量地检测了毛细血管前括约肌的舒缩运动;细胞流变特性;细胞与微血管的相互作用和动态耦合关系。提示:微循环灌流状态取决于流质与流场的动态耦合关系,且具有复杂性和多变性。而所谓的“海涛式灌注”是不确切的。     

高压氧对快速减压动物脑血流量和血细胞流变性的效用

目的 :探讨高压氧对快速减压脱险时应激性损伤的治疗效用。方法 :SD大鼠 30只 ,随机分为快速减压组、高压氧暴露组和正常对照组。用LDF 3激光血流仪测定了动物大脑局部组织血流量 ,用OPTON显微镜及录像系统观测了动物快速减压应激损伤及其高压氧暴露后血细胞流变性的变化。结果 :动物快速减压应激损伤时大脑皮质血流量明显下降 ,一般行为状态较差 ,红细胞出现畸形 ,白细胞、血小板激活 ,血栓形成 ,显示动物发生了减压应激损伤。然而 ,此时给损伤动物用高压氧暴露治疗 ,大脑血流量比对照组明显增加 (P <0 .0 1) ;畸形红细胞程度和发生率下降 (P<0 .0 1) ,白细胞与血小板激活现象减轻 ,血栓凝块消散。表明高压氧对快速减压应激损伤治疗中改善大脑皮质血流量和血细胞流变性有重要作用。结论 :(1)动物大脑皮质血流量下降及血液流变性异常是减压应激损伤重要特征之一 ;(2 )高压氧可通过改善大脑组织血液灌注和血细胞流变性 ,达到对动物减压应激损伤的治疗效用     

Changes of blood flow structure in precapillary microvessels during significant slow-down of flow

The experiments were carried out with a special "biological model" in which the rabbits' red blood cell suspension possessing low hematocrit circulated in frogs' mesenterial microvessels. Red blood cell behaviour was investigated in microvessels of 19-45 microns in diameter under conditions of arbitrarily changed flow velocity in mesenterial microvessels. Automatic frame-to-frame analysis of cinematographic films with the texture analysis system (Ernst Leitz, FRG) showed that the velocity fluctuations of individual red blood cells and their radial displacements increased significantly, while their velocity profile became blunt, during slowing-down of flow from 0.7 to 0.2 mm/s. Thus the normal blood flow structure in microvessels becomes disordered under ischemic conditions entailing disturbance of blood rheological properties and creating additionally increased resistance in the vessels.     

Possible treatment of some genetic deficiency diseases — A hypothesis

A conceptual hypothesis for the possibility of treatment of genetic deficiency diseases utilizing genetic engineering techniques is presented. It is proposed that the gene responsible for the synthesis of a protein which is missing in the patient may be inserted into the patient's stem cells using already established techniques of gene splicing and delivery. The so modified stem cells, if put back into the patient's system (e.g. bone marrow), by virtue of their ability of self multiplication are likely to start synthesizing and continue to produce missing protein. Since stem cells are also capable of differentiating into various blood cells, the nucleated blood cells are also likely to begin production of the protein whose gene was inserted into the stem cells. In this way a blood protein synthesized by any organ (e.g. liver) in normal persons may be synthesized by stem cells or their differentiated forms in the patient resulting in correction of the deficiency. Certain genetically deficient animals may be used to prove this hypothesis.     

在纤维蛋白凝块上诱导脐血间充质干细胞向膀胱尿路上皮细胞的分化◆

背景：在纤维蛋白凝块上诱导脐血间充质干细胞分化为膀胱尿路上皮细胞,尚无明确方案。 目的：探讨脐血来源间充质干细胞在纤维蛋白凝块上分化为膀胱尿路上皮细胞的可能性。 方法：实验组将脐血间充质干细胞接种于人工制备的脐血来源的纤维蛋白凝块上与胎儿膀胱尿路上皮细胞共培养,对照组将脐血间充质干细胞接种于人工制备的脐血来源的纤维蛋白凝胶上仅用DMEM/F1培养基培养。 结果与结论：实验组的脐血间充质干细胞逐渐伸展呈多角形,细胞扁平变大,透射电子显微镜下具有膀胱尿路上皮细胞的超微结构,抗广谱角蛋白(AE1/AE3)免疫组织化学染色阳性。对照组的脐血间充质干细胞呈长梭形,抗广谱角蛋白(CK AE1/AE3)免疫组织化学染色阴性。提示在纤维蛋白凝块上诱导脐血间充质干细胞有分化为膀胱尿路上皮细胞的能力。      

Effect of therapeutic plasmapheresis on rheological properties of blood in patients with ischemic heart disease]

Patients with angina pectoris, functional classes III-IV, were examined for changes in the rheological blood properties (red blood cell aggregation ratio, fluidity limit, apparent viscosity, electrophoretic mobility of red blood cells, fibrinogen, total plasma protein and ADP, induced platelet aggregation) under the effect of continuous plasmapheresis (PA). PA was established to produce a selective effect on the rheological blood properties depending on the initial level of its disorders. The most remarkable clinical effect was produced by PA in patients with initially high characteristics of hemorheology. The first PA session ameliorated hemorheology at the expense of the plasma component, reduced high platelet aggregation whereas the repeated sessions largely affected red blood cell function.     

Subclinical ischaemic episodes during the steady state of sickle cell anaemia.

AIMS: To determine the clinical, haematological, biochemical and rheological changes that occur in the asymptomatic steady state of sickle cell anaemia. METHODS: Patient self-assessment visual analogue scores (for wellbeing and tiredness), the blood concentration of acute phase proteins (C-reactive protein, orosomucoid, and fibrinogen), and blood rheology (percentage of dense cells and the number of sickled cells that occluded pores 5 microns in diameter) were studied longitudinally on 10 occasions in each of 20 outpatients with sickle cell anaemia. RESULTS: Patients in the steady state showed fluctuation in visual analogue scores, in concentration of acute phase proteins, and in rheological parameters consistent with minor episodes of tissue injury. Significantly more variation in acute phase proteins occurred in the steady state of 14 of the 20 patients who developed one or more vaso-occlusive crises during the 16 month study period. Rheological fluctuation in the steady state simulated rheological change during crisis, namely a transient rise and then fall in the number of dense and poorly filterable cells. CONCLUSIONS: The term "steady state" is a misnomer, being characterised by biochemical and rheological fluctuation consistent with minor episodes of microvascular occlusion that are insufficient to cause the overt tissue infarction of painful crisis.     

VHL-deficient vasculogenesis in hemangioblastoma

Hemangioblasts are capable of differentiation into vascular structures and blood. Patients with von Hippel–Lindau (VHL) disease develop hemangioblastomas which are composed of VHL-deficient tumor cells with protracted hemangioblastic differentiation potential. In a subset of these tumors, hemangioblastic differentiation is characterized by different stages of red blood cell formation. It has remained controversial, however, whether VHL-deficient hemangioblastic cells are similarly capable of differentiating into vascular cells and functioning vascular structures in vivo.     

Altered filtrability of white blood cells after myocardial infarction

Abnormal white blood cell rheological behaviour has been implicated as a cause of blood flow disturbances under conditions of ischaemia and reduced perfusion pressure. Accordingly, we have tested the mechanical properties of white cells following myocardial infarction by measuring the rate at which suspension of these cells cause plugging of Nuclepore filters. The number of clogging particles in a standard white cell suspension increased by the third day after infarction but subsequently decreased to the control levels. Since white cells can cause blockage of narrow blood vessels, it is assumed that such changes in cellular properties may influence the eventual extent of infarction.     

The osteogenic differentiation potentials of umbilical cord blood hematopoietic stem cells

Under specific culture conditions, umbilical cord blood derived mesenchymal stem cells (MSCs) can differentiate into osteogenic, adipogenic, and chondrogenic lineages. The purpose of the current study was to assess the differentiation potential of osteogenic umbilical cord blood derived hematopoietic stem cells (HSCs) and to develop an appropriate osteogenic differentiation medium for in vitro differentiation of umbilical cord blood derived HSCs. The study was conducted on 20 cord blood samples. The cells were cultured in osteogenic differentiating medium for 3?weeks. The HSCs differentiated into osteoblasts, which expressed osteoblast-associated genes (osteocalcin and bone sialoprotein), which were detected by RT-PCR. They showed alkaline phosphatase activity and a positive Alizarin red-S (AR-S) stain (calcium phosphate deposition). Umbilical cord blood is a rich source of hematopoietic stem cells that can be differentiated into osteoblasts; thus, it can be used for therapeutic strategies in the context of regenerative therapy.