Recapitulative haematopoietic development of human pluripotent stem cells in the absence of exogenous haematopoietic cytokines

To improve the recapitulative quality of human pluripotent stem cell (hPSC) differentiation, we removed exogenous haematopoietic cytokines from the defined differentiation system. Here, we show that endogenous stimuli and VEGF are sufficient to induce robust hPSC-derived haematopoiesis, intensive generation of haematopoietic progenitors, maturation of blood cells and the emergence of definitive precursor cells including those that phenotypically identical to early human embryonic haematopoietic stem cells (HSCs). Moreover, the cytokine-free system produces significantly higher numbers of haematopoietic progenitors compared to the published protocols. The removal of cytokines revealed a broad developmental potential of the early blood cells, stabilized the hPSC-derived definitive precursors and led to spontaneous activation of inflammatory signalling. Our cytokine-free protocol is simple, efficient, reproducible and applicable for embryonic stem cells (ESCs) and induced PSCs. The spectrum of recapitulative features of the novel protocol makes the cytokine-free differentiation a preferred model for studying the early human haematopoietic development.     

维持胚胎干细胞多能性的分子机制

胚胎干细胞（ES细胞）来源于早期发育的胚胎,具有分化为任何细胞类型的多能性,因此具有巨大的基础研究及潜在的应用前景.目前认为ES细胞主要通过一些外源性信号分子的作用及某些重要的内源性转录因子的表达共同起作用来达到其维持多能性的目的.外源性信号分子LIF、BMP4以及Wnt等介导的信号传导通路与内源性转录因子Oct4、Nanog、Sox2、FoxD3等共同起作用来抑制那些促进ES细胞分化的基因表达和激活那些有助于维持ES细胞多能性维持的基因表达,进而形成一个相互调控和依存的基因调控网络共同维持ES细胞的多能性.     

精原干细胞自我更新和分化的调控

精原干细胞（spermatogonial stem cells,SSCs）是体内自然状态下惟一能将遗传信息传至子代的成体干细胞,它们能通过维持自我更新和分化的稳定从而保证雄性生命过程中精子发生的持续进行。了解SSCs自我更新和分化的调节机制有助于阐明精子发生机理,并为探究其他组织中成体干细胞增殖分化的调节机制提供依据。然而目前对于SSCs自我更新和分化的调控机制所知甚少。SSCs的更新与分化遵循特定模式,受以睾丸支持细胞为主要成分的微环境及各种内分泌因素如胶质细胞源神经营养因子（GDNF）、维生素、Ets转录因子ERM/Etv5等的调控。本文评述了SSCs更新与分化的模式以及上述因素对其更新、分化的调控,探讨了其中可能涉及的信号通路,以期为本领域及其他成体干细胞相关研究提供借鉴。     

干细胞研究的新途径:miRNAs

微小RNA(microRNAs,miRNAs)是一类内源性的非编码单链RNA,能够通过与靶mRNA特异性的碱基配对而导致靶mRNA降解或抑制其翻译,从而对基因进行转录后调控。干细胞的自我更新和多向分化过程依赖于广泛而多样的调控机制,miRNAs正是这些调控机制中非常重要的一类分子。研究发现,干细胞的自我更新功能需要多种miRNAs的参与来维持;干细胞的分化也是多种miRNAs参与调控的结果。miRNAs可以作为干细胞研究的一个新的切入点。     

干细胞命运决定的分子调控网络

来源于囊胚期胚胎内细胞团的胚胎干细胞具有独特的生物学特性,包括无限自我更新的能力以及分化为内胚层、中胚层和外胚层各种细胞的潜能.阐明胚胎干细胞全能性维持以及向各种特定细胞分化的分子机制,不仅有助于我们了解胚胎发育过程,而且将促进胚胎干细胞尽早应用于疾病治疗.本文主要就干细胞的一种命运决定过程,维持胚胎干细胞全能性或失去全能性开始分化,结合最新的研究进展讨论该过程中的分子调控网络,包括信号转导通路、表达调控网络以及表观遗传调控.     

Circulating stem cell vary with NYHA stage in heart failure patients

We have investigated the blood levels of sub-classes of stem cells (SCs) [mesenchymal stem cells (MSCs), haematopoietic stem cells (HSCs), endothelial progenitor cells/circulating endothelial cells (EPCs/CECs) and tissue-committed stem cells (TCSCs)] in heart failure (HF) patients at different stage of pathology and correlated it with plasmatic levels of proangiogenic cytokines. Peripheral blood level of SCs were analysed in 97 HF patients (24 in NYHA class I, 41 in class II, 17 in class III and 15 in class IV) and in 23 healthy controls. Plasmatic levels of PDGF-BB, bFGF, HGF, vascular endothelial growth factor (VEGF), SDF-1α, TNF-α and NTproBNP were also measured. Compared with healthy individuals, MSC, and in particular the sub-classes CD45⁻CD34⁻CD90⁺, CD45⁻CD34⁻CD105⁺ and CD45⁻CD34⁻CXCR4⁺ were significantly enhanced in NYHA class IV patients (16.8-, 6.4- and 2.7-fold, respectively). Level of CD45⁻CD34⁻CD90⁺CXCR4⁺cells progressively increased from class II to class IV (fold increases compared with controls: 8.5, 12 and 21.5, respectively). A significant involvement of CXCR4⁺ subpopulation of HSC (CD45⁺CD34⁺CD90⁺CXCR4⁺, 1.4 versus 13.3 cells/μl in controls and NYHA class III patients, respectively) and TCSC (CD45⁻CD34⁺CXCR4⁺, 1.5 cells/ μl in controls versus 12.4 and 28.6 cells/μl in NYHA classes II and IV, respectively) were also observed. All tested cytokines were enhanced in HF patients. In particular, for PDGF-BB and SDF-1α we studied specific ligand/receptors pairs. Interestingly, the first one positively correlated with TCSCs expressing PDGFR (r = 0.52, P = 0.001), whereas the second one correlated with TCSCs (r = 0.34, P = 0.005) and with MSCs CD90⁺ expressing CXCR4 (r = 0.39, P = 0.001). HF is characterized by the increase in the circulating levels of different MSC, HSC, EPC and TCSC subsets. Both the entity and kinetic of this process varied in distinct cell subsets. Specifically, differently from HSCs and EPCs/CECs, MSCs and TCSCs significantly increased with the progression of the disease, suggesting a possible distinct role of these cells in the pathophysiology of HF.     

Requirements for human haematopoietic stem/progenitor cells

‘Requirements for human haematopoietic stem/progenitor cells’ is the first set of guidelines on human haematopoietic stem/progenitor cells in China, jointly drafted and agreed upon by experts from the Chinese Society for Stem Cell Research. This standard specifies the technical requirements, inspection methods, inspection rules, instructions for usage, labelling requirements, packaging requirements, storage requirements and transportation requirements for human haematopoietic stem/progenitor cells, which is applicable to the quality control for human haematopoietic stem/progenitor cells. We hope that publication of these guidelines will promote institutional establishment, acceptance and execution of proper protocols, and accelerate the international standardization of human haematopoietic stem/progenitor cells for applications.     

Prion potency in stem cells biology

Prion protein (PrP) can be considered a pivotal molecule because it interacts with several partners to perform a diverse range of critical biological functions that might differ in embryonic and adult cells. In recent years, there have been major advances in elucidating the putative role of PrP in the basic biology of stem cells in many different systems. Here, we review the evidence indicating that PrP is a key molecule involved in driving different aspects of the potency of embryonic and tissue-specific stem cells in self-perpetuation and differentiation in many cell types. It has been shown that PrP is involved in stem cell self-renewal, controlling pluripotency gene expression, proliferation, and neural and cardiomyocyte differentiation. PrP also has essential roles in distinct processes that regulate tissue-specific stem cell biology in nervous and hematopoietic systems and during muscle regeneration. Results from our own investigations have shown that PrP is able to modulate self-renewal and proliferation in neural stem cells, processes that are enhanced by PrP interactions with stress inducible protein 1 (STI1). Thus, the available data reveal the influence of PrP in acting upon the maintenance of pluripotent status or the differentiation of stem cells from the early embryogenesis through adulthood.     

Embryonic and adult neural stem cell research in China

Neural stem cells(NSCs) are one specific type of multipotential stem cells that have the ability to proliferate for a long time and to differentiate into neural cells,including neurons,astrocytes and oligodendrocytes.These NSCs exist in both the embryonic and adult central nervous system(CNS) of all mammalian species.Progress has been made in the understanding of the developmental regulation of NSCs and their function in neurogenesis.This review discusses recent progress in this area,with emphasis on work d...     

Multicellular cuddling in a stem cell niche

Haematopoietic stem and progenitor cells (HSPCs) can self-renew and differentiate in any blood cell type throughout life and thereby sustain the entire blood system. To do so, HSPCs had been shown to seed, in a multi-step process, intermediate haematopoietic niches before colonizing the adult marrow. While HSPC birth had been thoroughly characterized in the past, both in mammals and in zebrafish, how perivascular niches could host HSPCs and sustain their expansion was poorly understood. In an article published in the last issue of Cell, Tamplin et al.¹ elegantly exploited the many advantages provided by the zebrafish embryo to describe how endothelium remodeling in the perivascular niche, referred to as “cuddling,” favors HSPCs colonization and expansion.     

Hair follicle stem cell cultures reveal self‐organizing plasticity of stem cells and their progeny

Understanding how complex tissues are formed, maintained, and regenerated through local growth, differentiation, and remodeling requires knowledge on how single‐cell behaviors are coordinated on the population level. The self‐renewing hair follicle, maintained by a distinct stem cell population, represents an excellent paradigm to address this question. A major obstacle in mechanistic understanding of hair follicle stem cell (HFSC) regulation has been the lack of a culture system that recapitulates HFSC behavior while allowing their precise monitoring and manipulation. Here, we establish an in vitro culture system based on a 3D extracellular matrix environment and defined soluble factors, which for the first time allows expansion and long‐term maintenance of murine multipotent HFSCs in the absence of heterologous cell types. Strikingly, this scheme promotes de novo generation of HFSCs from non‐HFSCs and vice versa in a dynamic self‐organizing process. This bidirectional interconversion of HFSCs and their progeny drives the system into a population equilibrium state. Our study uncovers regulatory dynamics by which phenotypic plasticity of cells drives population‐level homeostasis within a niche, and provides a discovery tool for studies on adult stem cell fate.     

Enforced expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo

Hoxa5 is preferentially expressed in haematopoietic stem cells (HSCs) and multipotent progenitor cells (MPPs), and is more highly expressed in expanding HSCs. To date, little is known regarding the role of Hoxa5 in HSCs and downstream progenitor cells in vivo. In this study, we show that increased expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo. Hoxa5 differentially modifies the cell cycle of HSCs and lineage committed progenitor cells, depending on the cellular context. Hoxa5 drives HSCs, but not MPPs, through the cell cycle and arrests erythroid progenitor cells in G0 phase. Although the HSC pool shrinks after overexpression of Hoxa5, HSCs sustain the abilities of self-renewal and multipotency. In vivo, Hoxa5 has two effects on erythropoiesis: it causes a predominance of mature erythroid lineage cells and the partial apoptosis of erythroid progenitors. RNA-seq indicates that multiple biological processes, including erythrocyte homeostasis, cell metabolism, and apoptosis, are modified by Hoxa5. The results of this study indicate that Hoxa5 is a key regulator of the HSC cell cycle, and the inappropriate expression of Hoxa5 in lineage-committed progenitor cells leads to aberrant erythropoiesis.     

Enforced expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo

Hoxa5 is preferentially expressed in haematopoietic stem cells (HSCs) and multipotent progenitor cells (MPPs), and is more highly expressed in expanding HSCs. To date, little is known regarding the role of Hoxa5 in HSCs and downstream progenitor cells in vivo. In this study, we show that increased expression of Hoxa5 in haematopoietic stem cells leads to aberrant erythropoiesis in vivo. Hoxa5 differentially modifies the cell cycle of HSCs and lineage committed progenitor cells, depending on the cellular context. Hoxa5 drives HSCs, but not MPPs, through the cell cycle and arrests erythroid progenitor cells in G0 phase. Although the HSC pool shrinks after overexpression of Hoxa5, HSCs sustain the abilities of self-renewal and multipotency. In vivo, Hoxa5 has two effects on erythropoiesis: it causes a predominance of mature erythroid lineage cells and the partial apoptosis of erythroid progenitors. RNA-seq indicates that multiple biological processes, including erythrocyte homeostasis, cell metabolism, and apoptosis, are modified by Hoxa5. The results of this study indicate that Hoxa5 is a key regulator of the HSC cell cycle, and the inappropriate expression of Hoxa5 in lineage-committed progenitor cells leads to aberrant erythropoiesis.     

Phosphorylation state of the histone variant H2A.X controls human stem and progenitor cell fate decisions

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胚胎干细胞

胚胎干细胞具有自我复制并分化为人体各种功能细胞的潜能。胚胎干细胞具有的独特生物学特性使其被广泛应用于生物学研究的各个领域,特别是发育学。同时,它潜在的医学应用也成为世界范围内的研究热点。但是,由于人胚胎干细胞的来源为植入前的早期胚胎,人胚胎干细胞自诞生之日起便倍受争议。本文将从胚胎干细胞的来源、特性、鉴定标准、增殖机理、应用前景以及研究本身涉及的伦理学争论给予概述。     

Niche-mediated control of human embryonic stem cell self-renewal and differentiation

Complexity in the spatial organization of human embryonic stem cell (hESC) cultures creates heterogeneous microenvironments (niches) that influence hESC fate. This study demonstrates that the rate and trajectory of hESC differentiation can be controlled by engineering hESC niche properties. Niche size and composition regulate the balance between differentiation-inducing and -inhibiting factors. Mechanistically, a niche size-dependent spatial gradient of Smad1 signaling is generated as a result of antagonistic interactions between hESCs and hESC-derived extra-embryonic endoderm (ExE). These interactions are mediated by the localized secretion of bone morphogenetic protein-2 (BMP2) by ExE and its antagonist, growth differentiation factor-3 (GDF3) by hESCs. Micropatterning of hESCs treated with small interfering (si) RNA against GDF3, BMP2 and Smad1, as well treatments with a Rho-associated kinase (ROCK) inhibitor demonstrate that independent control of Smad1 activation can rescue the colony size-dependent differentiation of hESCs. Our results illustrate, for the first time, a role for Smad1 in the integration of spatial information and in the niche-size-dependent control of hESC self-renewal and differentiation.     

Therapy targets in glioblastoma and cancer stem cells: lessons from haematopoietic neoplasms

Despite intense efforts to identify cancer‐initiating cells in malignant brain tumours, markers linked to the function of these cells have only very recently begun to be uncovered. The notion of cancer stem cell gained prominence, several molecules and signalling pathways becoming relevant for diagnosis and treatment. Whether a substantial fraction or only a tiny minority of cells in a tumor can initiate and perpetuate cancer, is still debated. The paradigm of cancer‐initiating stem cells has initially been developed with respect to blood cancers where chronic conditions such as myeloproliferative neoplasms are due to mutations acquired in a haematopoietic stem cell (HSC), which maintains the normal hierarchy to neoplastic haematopoiesis. In contrast, acute leukaemia transformation of such blood neoplasms appears to derive not only from HSCs but also from committed progenitors that cannot differentiate. This review will focus on putative novel therapy targets represented by markers described to define cancer stem/initiating cells in malignant gliomas, which have been called ‘leukaemia of the brain’, given their rapid migration and evolution. Parallels are drawn with other cancers, especially haematopoietic, given the similar rampant proliferation and treatment resistance of glioblastoma multiforme and secondary acute leukaemias. Genes associated with the malignant conditions and especially expressed in glioma cancer stem cells are intensively searched. Although many such molecules might only coincidentally be expressed in cancer‐initiating cells, some may function in the oncogenic process, and those would be the prime candidates for diagnostic and targeted therapy. For the latter, combination therapies are likely to be envisaged, given the robust and plastic signalling networks supporting malignant proliferation.