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。     

IGF and FGF cooperatively establish the regulatory stem cell niche of pluripotent human cells in vitro

Distinctive properties of stem cells are not autonomously achieved, and recent evidence points to a level of external control from the microenvironment. Here, we demonstrate that self-renewal and pluripotent properties of human embryonic stem (ES) cells depend on a dynamic interplay between human ES cells and autologously derived human ES cell fibroblast-like cells (hdFs). Human ES cells and hdFs are uniquely defined by insulin-like growth factor (IGF)- and fibroblast growth factor (FGF)-dependence. IGF 1 receptor (IGF1R) expression was exclusive to the human ES cells, whereas FGF receptor 1 (FGFR1) expression was restricted to surrounding hdFs. Blocking the IGF-II/IGF1R pathway reduced survival and clonogenicity of human ES cells, whereas inhibition of the FGF pathway indirectly caused differentiation. IGF-II is expressed by hdFs in response to FGF, and alone was sufficient in maintaining human ES cell cultures. Our study demonstrates a direct role of the IGF-II/IGF1R axis on human ES cell physiology and establishes that hdFs produced by human ES cells themselves define the stem cell niche of pluripotent human stem cells.     

Mouse neural stem cells culturedin vitro and expressing an exogenous gene

Neural stem cells are the multipotential, self-renewing cells in central nerve system, and play an essential role in the development and differentiation of nerve system. Neural stem cells can be used to treat the nerve system diseases, especially, the transplantation of neural stem cells to rescue the degenerated neural cells has become a very promising therapeutic way. We successfully cultured neural stem cells isolated from the brains of embryonic micein vitro and determined their distribution in the E17 mice brains. The neural stem cells were transfected with adenoviral vector carrying GFP (green fluorescence protein) gene and then highly expressed the exogenous gene. It paves the way for gene therapy of degenerative nerve system diseases.     

Forced expression of theOct-4 gene influences differentiation of embryonic stem cells

By transfecting an Oct-4 expression plasmid into embryonic stem cells (ES cells), the ES-O cell line was constructed, which sustained the expression of Oct-4 gene when induced by retinoic acid. Forced expression of Oct-4 gene could not sustain the stem property of ES-O cells without the differentiation inhibiting factor LIF, but if LIF exists, forced expression of Oct-4 gene could enhance the ability to sustain the undifferentiation state and inhibit cell differentiation induced by retinoic acid. It was indicated that Oct-4 must cooperate with LIF to sustain the undifferentiation state of ES cells. During the cell differentiation, ES-O cells tend to differentiate into neural cells, suggesting that forced expression of Oct-4 gene may be in relation with the differentiation of neuroderm.     

Regulation of BMP4 on the proliferation and differentiation in SVZa neural stem cells

The neural stem cells in the anterior subventricular zone (SVZa) mainly generate the progenitors that will differentiate into neurons, and along a highly circumscribed migratory access Rostral migratory stream (RMS), they migrate to the olfactory bulbs (OB). To understand the effects of BMPs on SVZa neural stem cells, in this study BMP4 at various concentrations was used to induce SVZa neural stem cells, and the living cell labeling using BMP4 promotor conjugated with red fluorescence protein showed the expression of BMP4 dynamically. The results demonstrated that low BMP4 doses (1-5 ng/mL) promoted while high doses (10-100 ng/mL) inhibited the proliferation of SVZa neural stem cells, and BMP4 promotedneuron differentiation in the early stage (1-3 d), howeverm, it inhibited the neuron commitment after 4 d. Noggin, the antagonist of BMP4, blocked the physiological effects of BMP4. In OB, BMP4 is mainly to accelerate the progenitors to withdraw from the cell cycle and trigger the differentiation, and in RMS, it promotes the proliferation of committed progenitors and not differentiation, further in SVZa, BMP4 enhances astrocyte commitment.     

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。     

FGFR-related gene nou-darake restricts brain tissues to the head region of planarians

The study of planarian regeneration may help us to understand how we can rebuild organs and tissues after injury, disease or ageing. The robust regenerative abilities of planarians are based upon a population of totipotent stem cells (neoblasts), and among the organs regenerated by these animals is a well-organized central nervous system. In recent years, methodologies such as whole-mount in situ hybridizations and double-stranded RNA have been extended to planarians with the aim of unravelling the molecular basis of their regenerative capacities. Here we report the identification and characterization of nou-darake (ndk), a gene encoding a fibroblast growth factor receptor (FGFR)-like molecule specifically expressed in the head region of the planarian Dugesia japonica. Loss of function of ndk by RNA interference results in the induction of ectopic brain tissues throughout the body. This ectopic brain formation was suppressed by inhibition of two planarian FGFR homologues (FGFR1 and FGFR2). Additionally, ndk inhibits FGF signalling in Xenopus embryos. The data suggest that ndk may modulate FGF signalling in stem cells to restrict brain tissues to the head region of planarians.     

Attenuation of FGF signalling in mouse beta-cells leads to diabetes

Fibroblast growth factor (FGF) signalling has been implicated in patterning, proliferation and cell differentiation in many organs, including the developing pancreas. Here we show that the FGF receptors (FGFRs) 1 and 2, together with the ligands FGF1, FGF2, FGF4, FGF5, FGF7 and FGF10, are expressed in adult mouse beta-cells, indicating that FGF signalling may have a role in differentiated beta-cells. When we perturbed signalling by expressing dominant-negative forms of the receptors, FGFR1c and FGFR2b, in the pancreas, we found that that mice with attenuated FGFR1c signalling, but not those with reduced FGFR2b signalling, develop diabetes with age and exhibit a decreased number of beta-cells, impaired expression of glucose transporter 2 and increased proinsulin content in beta-cells owing to impaired expression of prohormone convertases 1/3 and 2. These defects are all characteristic of patients with type-2 diabetes. Mutations in the homeobox gene Ipf1/Pdx1 are linked to diabetes in both mouse and human. We also show that Ipf1/Pdx1 is required for the expression of FGFR1 signalling components in beta-cells, indicating that Ipf1/Pdx1 acts upstream of FGFR1 signalling in beta-cells to maintain proper glucose sensing, insulin processing and glucose homeostasis.     

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。     

A PPARγ-FGF1 axis is required for adaptive adipose remodelling and metabolic homeostasis

Although feast and famine cycles illustrate that remodelling of adipose tissue in response to fluctuations in nutrient availability is essential for maintaining metabolic homeostasis, the underlying mechanisms remain poorly understood. Here we identify fibroblast growth factor 1 (FGF1) as a critical transducer in this process in mice, and link its regulation to the nuclear receptor PPARγ (peroxisome proliferator activated receptor γ), which is the adipocyte master regulator and the target of the thiazolidinedione class of insulin sensitizing drugs. FGF1 is the prototype of the 22-member FGF family of proteins and has been implicated in a range of physiological processes, including development, wound healing and cardiovascular changes. Surprisingly, FGF1 knockout mice display no significant phenotype under standard laboratory conditions. We show that FGF1 is highly induced in adipose tissue in response to a high-fat diet and that mice lacking FGF1 develop an aggressive diabetic phenotype coupled to aberrant adipose expansion when challenged with a high-fat diet. Further analysis of adipose depots in FGF1-deficient mice revealed multiple histopathologies in the vasculature network, an accentuated inflammatory response, aberrant adipocyte size distribution and ectopic expression of pancreatic lipases. On withdrawal of the high-fat diet, this inflamed adipose tissue fails to properly resolve, resulting in extensive fat necrosis. In terms of mechanisms, we show that adipose induction of FGF1 in the fed state is regulated by PPARγ acting through an evolutionarily conserved promoter proximal PPAR response element within the FGF1 gene. The discovery of a phenotype for the FGF1 knockout mouse establishes the PPARγ–FGF1 axis as critical for maintaining metabolic homeostasis and insulin sensitization.     

Bmi-1 determines the proliferative capacity of normal and leukaemic stem cells

An emerging concept in the field of cancer biology is that a rare population of 'tumour stem cells' exists among the heterogeneous group of cells that constitute a tumour. This concept, best described with human leukaemia, indicates that stem cell function (whether normal or neoplastic) might be defined by a common set of critical genes. Here we show that the Polycomb group gene Bmi-1 has a key role in regulating the proliferative activity of normal stem and progenitor cells. Most importantly, we provide evidence that the proliferative potential of leukaemic stem and progenitor cells lacking Bmi-1 is compromised because they eventually undergo proliferation arrest and show signs of differentiation and apoptosis, leading to transplant failure of the leukaemia. Complementation studies showed that Bmi-1 completely rescues these proliferative defects. These studies therefore indicate that Bmi-1 has an essential role in regulating the proliferative activity of both normal and leukaemic stem cells.     

微重力环境下Notch信号对神经干细胞分化的影响

基于微重力条件下神经干细胞的分化情况,结合Notch信号通路对神经干细胞分化的影响,论述模拟微重力对神经干细胞分化产生影响的可能分子机制。     

Human cardiovascular progenitor cells develop from a KDR+ embryonic-stem-cell-derived population

The functional heart is comprised of distinct mesoderm-derived lineages including cardiomyocytes, endothelial cells and vascular smooth muscle cells. Studies in the mouse embryo and the mouse embryonic stem cell differentiation model have provided evidence indicating that these three lineages develop from a common Flk-1(+) (kinase insert domain protein receptor, also known as Kdr) cardiovascular progenitor that represents one of the earliest stages in mesoderm specification to the cardiovascular lineages. To determine whether a comparable progenitor is present during human cardiogenesis, we analysed the development of the cardiovascular lineages in human embryonic stem cell differentiation cultures. Here we show that after induction with combinations of activin A, bone morphogenetic protein 4 (BMP4), basic fibroblast growth factor (bFGF, also known as FGF2), vascular endothelial growth factor (VEGF, also known as VEGFA) and dickkopf homolog 1 (DKK1) in serum-free media, human embryonic-stem-cell-derived embryoid bodies generate a KDR(low)/C-KIT(CD117)(neg) population that displays cardiac, endothelial and vascular smooth muscle potential in vitro and, after transplantation, in vivo. When plated in monolayer cultures, these KDR(low)/C-KIT(neg) cells differentiate to generate populations consisting of greater than 50% contracting cardiomyocytes. Populations derived from the KDR(low)/C-KIT(neg) fraction give rise to colonies that contain all three lineages when plated in methylcellulose cultures. Results from limiting dilution studies and cell-mixing experiments support the interpretation that these colonies are clones, indicating that they develop from a cardiovascular colony-forming cell. Together, these findings identify a human cardiovascular progenitor that defines one of the earliest stages of human cardiac development.     

Cell fusion-independent differentiation of neural stem cells to the endothelial lineage

Somatic stem cells have been claimed to possess an unexpectedly broad differentiation potential (referred to here as plasticity) that could be induced by exposing stem cells to the extracellular developmental signals of other lineages in mixed-cell cultures. Recently, this and other experimental evidence supporting the existence of stem-cell plasticity have been refuted because stem cells have been shown to adopt the functional features of other lineages by means of cell-fusion-mediated acquisition of lineage-specific determinants (chromosomal DNA) rather than by signal-mediated differentiation. In this study we co-cultured mouse neural stem cells (NSCs), which are committed to become neurons and glial cells, with human endothelial cells, which form the lining of blood vessels. We show that in the presence of endothelial cells six per cent of the NSC population converted to cells that did not express neuronal or glial markers, but instead showed the stable expression of multiple endothelial markers and the capacity to form capillary networks. This was surprising because NSCs and endothelial cells are believed to develop from the ectoderm and mesoderm, respectively. Experiments in which endothelial cells were killed by fixation before co-culture with live NSCs (to prevent cell fusion) and karyotyping analyses, revealed that NSCs had differentiated into endothelial-like cells independently of cell fusion. We conclude that stem-cell plasticity is a true characteristic of NSCs and that the conversion of NSCs to unanticipated cell types can be accomplished without cell fusion.