Intrinsic transition of embryonic stem-cell differentiation into neural progenitors

The neural fate is generally considered to be the intrinsic direction of embryonic stem (ES) cell differentiation. However, little is known about the intracellular mechanism that leads undifferentiated cells to adopt the neural fate in the absence of extrinsic inductive signals. Here we show that the zinc-finger nuclear protein Zfp521 is essential and sufficient for driving the intrinsic neural differentiation of mouse ES cells. In the absence of the neural differentiation inhibitor BMP4, strong Zfp521 expression is intrinsically induced in differentiating ES cells. Forced expression of Zfp521 enables the neural conversion of ES cells even in the presence of BMP4. Conversely, in differentiation culture, Zfp521-depleted ES cells do not undergo neural conversion but tend to halt at the epiblast state. Zfp521 directly activates early neural genes by working with the co-activator p300. Thus, the transition of ES cell differentiation from the epiblast state into neuroectodermal progenitors specifically depends on the cell-intrinsic expression and activator function of Zfp521.     

Bone morphogenetic proteins inhibit the tumorigenic potential of human brain tumour-initiating cells

Transformed, oncogenic precursors, possessing both defining neural-stem-cell properties and the ability to initiate intracerebral tumours, have been identified in human brain cancers. Here we report that bone morphogenetic proteins (BMPs), amongst which BMP4 elicits the strongest effect, trigger a significant reduction in the stem-like, tumour-initiating precursors of human glioblastomas (GBMs). Transient in vitro exposure to BMP4 abolishes the capacity of transplanted GBM cells to establish intracerebral GBMs. Most importantly, in vivo delivery of BMP4 effectively blocks the tumour growth and associated mortality that occur in 100% of mice after intracerebral grafting of human GBM cells. We demonstrate that BMPs activate their cognate receptors (BMPRs) and trigger the Smad signalling cascade in cells isolated from human glioblastomas (GBMs). This is followed by a reduction in proliferation, and increased expression of markers of neural differentiation, with no effect on cell viability. The concomitant reduction in clonogenic ability, in the size of the CD133+ population and in the growth kinetics of GBM cells indicates that BMP4 reduces the tumour-initiating cell pool of GBMs. These findings show that the BMP-BMPR signalling system--which controls the activity of normal brain stem cells--may also act as a key inhibitory regulator of tumour-initiating, stem-like cells from GBMs and the results also identify BMP4 as a novel, non-cytotoxic therapeutic effector, which may be used to prevent growth and recurrence of GBMs in humans.     

Corridors of migrating neurons in the human brain and their decline during infancy

The subventricular zone of many adult non-human mammals generates large numbers of new neurons destined for the olfactory bulb. Along the walls of the lateral ventricles, immature neuronal progeny migrate in tangentially oriented chains that coalesce into a rostral migratory stream (RMS) connecting the subventricular zone to the olfactory bulb. The adult human subventricular zone, in contrast, contains a hypocellular gap layer separating the ependymal lining from a periventricular ribbon of astrocytes. Some of these subventricular zone astrocytes can function as neural stem cells in vitro, but their function in vivo remains controversial. An initial report found few subventricular zone proliferating cells and rare migrating immature neurons in the RMS of adult humans. In contrast, a subsequent study indicated robust proliferation and migration in the human subventricular zone and RMS. Here we find that the infant human subventricular zone and RMS contain an extensive corridor of migrating immature neurons before 18 months of age but, contrary to previous reports, this germinal activity subsides in older children and is nearly extinct by adulthood. Surprisingly, during this limited window of neurogenesis, not all new neurons in the human subventricular zone are destined for the olfactory bulb--we describe a major migratory pathway that targets the prefrontal cortex in humans. Together, these findings reveal robust streams of tangentially migrating immature neurons in human early postnatal subventricular zone and cortex. These pathways represent potential targets of neurological injuries affecting neonates.     

三七总皂苷对新生大鼠海马神经干细胞活性影响及促分化作用

目的研究三七总皂苷对海马神经干细胞活性的影响和分化作用。方法体外培养海马神经干细胞,分别接种于96孔板和12孔板,96孔板细胞按三七总皂苷不同浓度梯度和同一浓度的不同时间点进行干预,应用MTT法检测海马神经干细胞的OD值,观察三七总皂苷对海马神经干细胞活性的影响;12细胞孔板分为对照组和给药组,应用免疫荧光染色方法检测神经元新生特异抗原（Tuj-1）和胶质细胞新生抗原（Vimentin）的表达,以观察三七总皂苷对海马神经干细胞分化的影响。结果（1）一定浓度范围内三七总皂苷能增强海马神经干细胞活性;（2）三七总皂苷能促进海马神经干细胞向神经元和胶质细胞方向分化。结论三七总皂苷能增强海马神经干细胞的活性并能促进海马神经干细胞分化。     

碱性成纤维细胞生长因子对骨髓基质细胞的生物学效应

目的探讨碱性成纤维细胞生长因子(bFGF)对骨髓基质细胞(BMSC)粘附、增殖和分化等生物学效应的影响.方法利用体视学计数、MTT法及ALP试剂盒分别测定不同浓度bFGF诱导一定时间后BMSG的粘附特性、增殖和分化情况的变化.结果10n/mLbFGF明显促进BMSG的粘附,但是200ng/mLbFGF反而不利于细胞粘附;在细胞增殖和分化测定中,100ng/mLbFGF明显促进细胞增殖,但细胞碱性磷酸酶含量也最低.结论碱性成纤维细胞生长因子对骨髓基质细胞的生物效应是复杂和多方面的,可以作用于骨髓基质细胞的粘附、增殖和分化等多个环节,这种影响与碱性成纤维细胞生长因子的剂量有关.     

<Emphasis Type="Italic">In vitro</Emphasis> induction of mouse meningeal-derived ips cells into neural-like cells

Previous research has shown that mouse embryonic stem (ES) cells can be induced to form neural cells in adherent monocultures. In this study, pluripotent stem (iPS) C5 cells derived from meningeal membranes were converted successfully into neural-like cells using the same protocol generally used for ES cells. Meningeal-iPS C5 cells were induced to express neural markers Sox1, Sox3, Pax6, Nestin and Tuj1 and to reduce the expression of ES markers Oct4 and Nanog during neural differentiation, and can be differentiated into Pax6 and Nestin positive neural progenitors, and further into neuronal, astrocytic, and oligodendrocytic cells. In vitro differentiation of iPS cells into patient-specific neural cells could serve as a model to study mechanisms of genetic diseases and develop promising candidates for therapeutic applications in dysfunctional or aging neural tissues. Meningeal cells express a high level of the embryonic master regulator Sox2, allowing them to be reprogrammed into iPS cells more easily than other somatic cells.     

人骨髓间充质干细胞对小鼠神经干细胞增殖与分化培养的影响

通过在体外培养、鉴定人的骨髓间充质干细胞与小鼠神经干细胞,用骨髓间充质干细胞条件培养基分别在增殖与分化条件下对神经干细胞进行培养.发现,间充质干细胞条件培养基在增殖条件下能加快神经球内神经干细胞的迁移,使神经球解聚,对神经干细胞增殖没有影响;而间充质干细胞条件培养基在分化条件下,能增加神经干细胞向少突胶质细胞分化的能力,降低向星型胶质细胞的分化能力,对向神经元分化能力没有影响,间充质干细胞可能是通过促进神经干细胞迁移、分化而加快神经损伤的修复的.     

毛囊干细胞及Noggin 对其调控作用的研究进展

摘要: 毛囊干细胞( HFSC) 通常能够分化为角质细胞、皮脂腺细胞和短暂扩充祖细胞,因其快速增殖的能力,HFSC为细胞的治疗提供了构建干细胞来源的基础。本综述将系统描述HFSC 的定位、培养以及分子标志物,为鉴定、分离细胞提供有力的参考。同时针对HFSC 增殖和分化调控的关键基因Noggin,从基因发现、分子功能等进行全面阐述,并结合BMP 信号通路叙述Noggin 调控HFSC 研究的最新进展。     

神经干细胞的诱导分化

目的：探讨神经干细胞（NSCs）的诱导分化机制。方法：本文就Nscs的来源、分布、诱导分化的调控机制和因素以及其应用前景作一综述。结果：神经干细胞可从鼠、人的胚胎和成年中枢神经系统（CNS）成功分离并培养，具有自我更新和多分化潜能，经体外诱导或植入体内后均能分化为成熟神经元和神经胶质细胞。结论：神经干细胞的发现和研究的深入为神经发育研究及中枢神经功能重建提供了新的思路。     

Effect of Dy3+on osteogenic and adipogenic differentiation of mouse primary bone marrow stromal cells and adipocytic trans-differentiation of mouse primary osteoblasts

A series of experimental methods including 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) test, alkaline phosphatase (ALP) activity measurement, mineralized function, Oil Red O stain and measurement were employed to assess the effect of Dy³⁺ on the osteogenic and adipogenic differentiation of mouse primary bone marrow stromal cells (BMSCs) and the adipogenic trans-differentiation of mouse primary osteoblasts (OBs). The results showed that Dy³⁺ had no effect on BMSC proliferation at concentrations of 1×10⁻⁸ and 1×10⁻⁵ mol/L, but inhibited BMSC proliferation at other concentrations. Dy³⁺ had no effect on OB proliferation at concentrations of 1×10⁻¹⁰ and 1×10⁻⁹ mol/L, but inhibited OB proliferation at other concentrations. Dy³⁺ had no effect on the osteogenic differentiation of BMSCs at concentrations of 1×10⁻⁹ and 1×10⁻⁷ mol/L, and promoted osteogenic differentiation of BMSCs at other concentrations at the 7th day. The osteogenic differentiation of BMSCs was inhibited by Dy³⁺ at concentration of 1×10⁻⁵ mol/L at the 14th day, but promoted osteogenic differentiation of BMSCs at concentrations of 1×10⁻⁹, 1×10⁻⁸, 1×10⁻⁷ and 1×10⁻⁶ mol/L with the maximal effect at concentration of 10⁻⁶ mol/L. Dy³⁺ promoted mineralized function of BMSCs at any concentration. Dy³⁺ had no effect on adipogenic differentiation of BMSCs at concentration of 1×10⁻⁷ mol/L, but inhibited adipogenic differentiation of BMSCs at other concentrations. Dy³⁺ inhibited adipocytic trans-differentiation of OBs at any concentration, suggesting that Dy³⁺ had protective effect on bone and the protective effect on bone may be mediated by modulating differentiation of BMSCs away from the adipocyte and inhibiting adipocytic trans-differentiation of OBs which may promote differentiation and mineralization of OBs. These results may be valuable for better understanding the mechanism of the effect of Dy³⁺ on pathogenesis of osteoporosis. Supported by the Foundation for Key Program of Ministry of Education of China (Grant No. 208018)     

Differentiating germ cells can revert into functional stem cells in Drosophila melanogaster ovaries

Many tissues including blood, skin, gut and germ cells are continuously maintained by tissue stem cells. Under certain conditions, however, other organs can undergo repair using stem-cell-like progenitors generated by cell de-differentiation. Cell fates have been broadened experimentally, but mechanisms allowing de-differentiation to a stem cell state are poorly known. Germline stem cells begin to differentiate by forming interconnected germ cell cysts (cystocytes), and under certain conditions male mouse cystocytes have been postulated to revert into functional progenitors. Here we report that four- and eight-cell Drosophila germline cystocytes generated either in second instar larval ovaries or in adults over-producing the BMP4-like stem cell signal Decapentaplegic efficiently convert into single stem-like cells. These de-differentiated cells can develop into functional germline stem cells and support normal fertility. Our results show that cystocytes represent a relatively abundant source of regenerative precursors that might help replenish germ cells after depletion by genotoxic chemicals, radiation or normal ageing. More generally, Drosophila cystocytes now provide a system for studying de-differentiation and its potential as a source of functional stem cells.     

新生大鼠海马神经干细胞的体外培养及鉴定

的探讨新生大鼠海马神经干细胞（NSC）的体外培养和诱导分化的条件和特点。方法分离出生1d大鼠海马,在表皮生长因子、碱性成纤维生长因子和B27联合作用下使其稳定增殖,用5-溴脱氧尿苷（BMU）标记处于增殖状态的神经干细胞,应用免疫荧光染色方法行巢蛋白（Nestin）、5-溴脱氧尿苷（BrdU）、β-Ⅲ型微管蛋白（Tuj-1）、波形蛋白（Vimenfin）和Galc-C免疫荧光染色,对NSC的增殖及其分化的细胞进行鉴定。结果体外培养的NSC增殖成神经干细胞球并传代,鉴定为Nestin染色阳性细胞和5-溴脱氧尿苷（BrdU）标记染色阳性细胞,并可诱导分化为神经元细胞（Tuj-1染色阳性细胞）、神经胶质细胞（Vimentin染色阳性细胞）和少突胶质细胞（GMc-C染色阳性细胞）。结论采用无血清培养基中加入特定生长因子的培养技术,可培养出在体外稳定增殖并有多向分化潜能的新生大鼠海马神经干细胞。     

脉冲磁场对应激大鼠海马神经干细胞增殖的影响

建立大鼠束缚应激模型及脉冲磁场环境，观察应激大鼠在脉冲磁场环境中海马神经千细胞数及增殖变化．实验设对照、应激、磁场、应激磁场4个组，应用免疫组织化学法观察和计算各组海马巢蛋白（Nestin）和溴化脱氧核糖尿嘧啶（BrdU）的阳性细胞数．研究结果显示，Nestin在各组海马CA1和CA2区表达，BrdU在海马齿状回表达．实验各组Nestin和BrdU的阳性细胞比对照组明显增加，应激磁场组增加更显著，而磁场组和应激磁场组则无明显差异．这表明应激能引起大鼠海马神经千细胞内源性增殖，脉冲磁场能显著提高应激大鼠海马神经千细胞的增殖反应，提示在应激脑损伤的基础上，脉冲磁场作为外源性物理因素能刺激神经千细胞增殖，并可能修复受损神经元。     

一些神经营养因子对神经干细胞的增殖和分化的影响

胚胎和成年哺乳动物脑内存在能分化为神经元和神经胶质细胞的细胞干细胞,从成年脑和胚脑分离的神经干细胞能在体外分裂并进一步分化成神经元和胶质细胞,许多生长因子,如成纤维细胞生长因子和表皮生长因子等都参与了这一分裂、分化过程。对神经干细胞的增殖及分化产生一定的影响,但在不同的情况下,它们对增殖及分化的作用不同。     

马钱子苷对神经干细胞的增殖、存活和分化的调节作用

为探讨不同浓度的马钱子苷对神经干细胞的增殖、存活和分化的调节作用及其相关分子机制，本实验从成年小鼠大脑中分离培养了神经干细胞，用不同浓度的马钱子苷进行干预，观察马钱子苷对神经干细胞增殖、存活和分化的影响。结果显示：成年小鼠神经干细胞在含有中、高浓度马钱子苷的增殖培养基中培养5 d和7 d后，神经球数量和直径与对照相比显著增加（P<0.05）；中、高剂量的马钱子苷能够促进神经干细胞的有丝分裂，低剂量马钱子苷处理显著促进神经干细胞发生分化（P<0.01），并增加神经元和星形胶质细胞的数量及比例（P<0.01）；中、高剂量马钱子苷抑制神经干细胞分化（P<0.05），高剂量的马钱子苷使得神经元的数量减少（P<0.05）。研究结果表明，高浓度的马钱子苷能够促进神经干细胞存活，并通过促进神经干细胞有丝分裂来提高其增殖能力；低浓度的马钱子苷促进神经干细胞分化，有利于神经再生和少突胶质细胞再生。研究结果为神经干细胞治疗中枢神经系统疾病的研究奠定了理论和实验基础。     

重组人骨形态发生蛋白-2和-6对人骨肉瘤细胞株MG63和U2OS的作用

为研究重组人骨形态发生蛋白rhBMP-2和rhBMP-6对人骨肉瘤细胞株MG63和U20S的作用,分别用含rhBMP-2和rhBMP-6及重组绿色荧光蛋白的条件培养液干预人骨肉瘤细胞MG63和U2OS,利用台盼蓝拒染法、TUNEL法、吖啶橙/溴乙啶(AO/EB)荧光双染法、Transwell小窒和碱性磷酸酶活性测定法分别检测细胞增殖、凋亡、迁移以及成骨分化能力的变化.结果显示与未实施任何千预的空白对照组和用rGFP干预实验对照组相比.rhBMP-2和rhBMP-6的干预致两种骨肉瘤细胞株:细胞存活率均随时间逐渐降低;凋亡率呈时间依赖性增加;穿膜数明显减低;碱性磷酸酶(ALP)活性逐渐增加.以上差异均有统计学意义(P＜0.01).表明rhBMP-2和rhBMP-6可抑制人骨肉瘤细胞株MG63和U2OS的增殖和迁移、诱导其凋亡和向成骨细胞分化.     

Effects of genistein, daidzein and glycitein on the osteogenic and adipogenic differentiation of bone marrow stromal cells and on the adipogenic trans-differentiation of osteoblasts

The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), alkaline phosphatase (ALP) activity and oil red O assays were used to examine the effects of genistein, daidzein and glycitein on the osteogenic and adipogenic differentiation of primary mouse bone marrow stromal cells (MSCs) and the adipogenic trans-differentiation of primary mouse osteoblasts. The results indicated that daidzein, genistein and glycitein at concentrations from 1×10^-8 mol/L to 1×10^-5 mol/L promoted the proliferation of MSCs and osteoblasts; genistein, daidzein and glycitein promoted osteogenic differentiation and inhibited adipogenic differentiation of MSCs, and inhibited adipocytic transdifferentiation of osteoblasts at appropriate concentrations as 17β-estradiol. It suggests that genistein, daidzein and glycitein regulate a dual differentiational process of MSCs into the osteogenic and adipogenic lineages, and trans-differentiational process of primary osteoblasts into the adipocyte lineages, causing a lineage shift toward osteoblast. Protective effects of them on bone may be mediated by a reversal of adipogenesis which may promote the proliferation, differentiation and mineralization of osteoblasts, and make adipocytes secrete less cytokines which may promote osteoclast formation and activation. In addition, the results also indicated that genistein, daidzein and glycitein may be helpful in preventing the development of steroid induced osteonecrosis.     

全反式维甲酸对小鼠胚胎不同阶段神经干细胞诱导作用研究

目的:探讨全反式维甲酸对小鼠胚胎不同阶段神经干细胞(NSCs)的诱导分化情况.方法:分离孕12.5d及15.5d的胚胎小鼠脑皮质.取第3代NSCs,用含1μmol·L-1的全反式维甲酸在体外诱导小鼠胚胎不同阶段的NSCs.诱导5d后,通过神经元微管相关蛋白2(MAP2)免疫荧光染色和Westernblots检测NSCs分化为神经元的比例.结果:与对照组相比,全反式维甲酸可明显提高神经元分化的比例.E12.5干细胞和E15.5胚胎干细胞分化为神经元的比例分别为30%±1.47%和16.21%±1.36%.结论全反式维甲酸具有显著的促神经干细胞分化成神经元的效应,并对胚胎不同阶段神经干细胞的诱导作用有所不同.