Generation of functional endothelial-like cells from adult mouse germline-derived pluripotent stem cells

Functional endothelial cells and their progenitors are required for vascular development, adequate vascular function, vascular repair and for cell-based therapies of ischemic diseases. Currently, cell therapy is limited by the low abundance of patient-derived cells and by the functional impairment of autologous endothelial progenitor cells (EPCs). In the present study, murine germline-derived pluripotent stem (gPS) cells were evaluated as a potential source for functional endothelial-like cells.     

Epigenetic alteration of imprinted genes during neural differentiation of germline-derived pluripotent stem cells

Spermatogonial stem cells (SSCs), which are unipotent stem cells in the testes that give rise to sperm, can be converted into germline-derived pluripotent stem (gPS) by self-induction. The androgenetic imprinting pattern of SSCs is maintained even after their reprogramming into gPS cells. In this study, we used an in vitro neural differentiation model to investigate whether the imprinting patterns are maintained or altered during differentiation. The androgenetic patterns of H19, Snrpn, and Mest were maintained even after differentiation of gPS cells into NSCs (gPS-NSCs), whereas the fully unmethylated status of Ndn in SSCs was altered to somatic patterns in gPS cells and gPS-NSCs. Thus, our study demonstrates epigenetic alteration of genomic imprinting during the induction of pluripotency in SSCs and neural differentiation, suggesting that gPS-NSCs can be a useful model to study the roles of imprinted genes in brain development and human neurodevelopmental disorders.     

Coloring skin with pluripotent stem cells

Comment on: Nissan X, et al. Proc Natl Acad Sci USA 2011; 108:14861-6.     

In vitro organogenesis from pluripotent stem cells

Pluripotent stem cells (PSCs) have the ability to spontaneously generate structured tissues in vitro reminiscent of embryonic tissue development. Recently, complex organoids such as cortical tissues, cerebral brain organoids, optical cups, intestinal tissues, and liver buds have been generated from PSCs derived from healthy individuals and patients with genetic diseases, providing powerful tools to understand morphogenesis and disease pathology. This article highlights recent advances in the state-of-art generation of organoids from PSCs, possible signaling pathways and mechanisms involved in organogenesis, and the understanding of extracellular microenvironment. Challenges involved in the organoid generation such as increasing organoid size, enhancing the tissue complexity, and improving functional maturation are also discussed.     

In vitro organogenesis from pluripotent stem cells

Pluripotent stem cells (PSCs) have the ability to spontaneously generate structured tissues in vitro reminiscent of embryonic tissue development. Recently, complex organoids such as cortical tissues, cerebral brain organoids, optical cups, intestinal tissues, and liver buds have been generated from PSCs derived from healthy individuals and patients with genetic diseases, providing powerful tools to understand morphogenesis and disease pathology. This article highlights recent advances in the state-of-art generation of organoids from PSCs, possible signaling pathways and mechanisms involved in organogenesis, and the understanding of extracellular microenvironment. Challenges involved in the organoid generation such as increasing organoid size, enhancing the tissue complexity, and improving functional maturation are also discussed.     

Generating human intestinal tissue from pluripotent stem cells in vitro

Here we describe a protocol for generating 3D human intestinal tissues (called organoids) in vitro from human pluripotent stem cells (hPSCs). To generate intestinal organoids, pluripotent stem cells are first differentiated into FOXA2(+)SOX17(+) endoderm by treating the cells with activin A for 3 d. After endoderm induction, the pluripotent stem cells are patterned into CDX2(+) mid- and hindgut tissue using FGF4 and WNT3a. During this patterning step, 3D mid- or hindgut spheroids bud from the monolayer epithelium attached to the tissue culture dish. The 3D spheroids are further cultured in Matrigel along with prointestinal growth factors, and they proliferate and expand over 1-3 months to give rise to intestinal tissue, complete with intestinal mesenchyme and epithelium comprising all of the major intestinal cell types. To date, this is the only method for efficiently directing the differentiation of hPSCs into 3D human intestinal tissue in vitro.     

Avian pluripotent stem cells

Pluripotent embryonic stem cells are undifferentiated cells capable of proliferation and self-renewal and have the capacity to differentiate into all somatic cell types and the germ line. They provide an in vitro model of early embryonic differentiation and are a useful means for targeted manipulation of the genome. Pluripotent stem cells in the chick have been derived from stage X blastoderms and 5.5 day gonadal primordial germ cells (PGCs). Blastoderm-derived embryonic stem cells (ESCs) have the capacity for in vitro differentiation into embryoid bodies and derivatives of the three primary germ layers. When grafted onto the chorioallantoic membrane, the ESCs formed a variety of differentiated cell types and attempted to organize into complex structures. In addition, when injected into the unincubated stage X blastoderm, the ESCs can be found in numerous somatic tissues and the germ line. The potential give rise to somatic and germ line chimeras is highly dependent upon the culture conditions and decreases with passage. Likewise, PGC-derived embryonic germ cells (EGCs) can give rise to simple embryoid bodies and can undergo some differentiation in vitro. Interestingly, chicken EG cells contribute to somatic lineages when injected into the stage X blastoderm, but only germ line chimeras have resulted from EGCs injected into the vasculature of the stage 16 embryo. To date, no lines of transgenic chickens have been generated using ESCs or EGCs. Nevertheless, progress towards the culture of avian pluripotent stem cells has been significant. In the future, the answers to fundamental questions regarding segregation of the avian germ line and the molecular basis of pluripotency should foster the full use of avian pluripotent stem cells.     

体细胞诱导为多能干细胞的最新进展

2007年11-12月,Cell、Science和Nature发表一系列体外诱导人类体细胞转变为多能干细胞的论文。来自日本和美国的研究小组利用慢病毒载体分别将Oct-4、Sox2、C-Myc、Klf4和Oct-4、Sox2、Nanog、Lin28两套基因转入人成纤维细胞,均获得类似ES细胞的克隆。小鼠诱导性多能干细胞已初步用于镰刀细胞性贫血的基因治疗。短短一年半,诱导性多能干细胞的研究和关注度呈现了爆炸式成长;体细胞重编程、去分化、多能干细胞来源等一系列热点问题再次成为大众瞩目的中心。     

Induced pluripotent stem cells

Induced pluripotent stem cells (iPS) result from a reprogramming of somatic cells via transduction with viral vectors expressing the Oct4, Sox2, c-Myc, Klf4, Nanog, and Lin28 genes, which are essential for the establishment and maintenance of the pluripotent state. In properties, iPS are almost fully similar to embryonic stem cells (ESC). To date, iPS have been obtained from various differentiated cells of mice and humans. Along with ESC, iPSs are highly promising for research and medicine.     

MicroRNAs与多潜能干细胞的研究进展

microRNAs(miRNAs)是一类内源性非编码小RNA,通过调控基因表达来参与生命过程中的一系列重要进程。越来越多的证据表明,miRNAs参与了几乎所有生物代谢过程,其胚胎干细胞的自我更新与分化和在多能干细胞(iPSCs)中的诱导调节作用也日益受到关注。该文介绍了miRNAs的生成、检测方法以及miRNAs对胚胎干细胞(ESCs)及诱导多能性干细胞的调控作用,并对miRNAs的应用前景进行了展望。     

Epithelial stem cells

New molecular markers for epidermal stem cells have enabled their isolation both in vitro and from the epidermis lying between hair follicles. Micro-dissection experiments have localised a second population of stem cells within hair follicles. Epidermal stem cells have a patterned distribution in vivo. The patterning can be reconstituted in vitro, showing that it is generated by interactions between keratinocytes and that the differentiation of epidermal stem cells is regulated by signals from other keratinocytes. Recent evidence from transgenic mice suggests that stem cell behaviour in the gut may be regulated by similar cell-cell interactions in vivo. Candidate genes for mediating these interactions are the homologues of Drosophila cell fate patterning genes such as Notch and Wingless and the Cadherin family of cell-cell adhesion molecules. The roles of stem cells and of mutations of the Patched gene in epithelial carcinogenesis are discussed.     

Metabolism of pluripotent stem cells

Background

Recently, growing attention has been directed toward stem cell metabolism, with the key observation that metabolism not only fuels the proper functioning of stem cells but also regulates the fate of these cells. There seems to be a clear link between the self-renewal of pluripotent stem cells (PSCs), in which cells proliferate indefinitely without differentiation, and the activity of specific metabolic pathways. The unique metabolism in PSCs plays an important role in maintaining pluripotency by regulating signaling pathways and resetting the epigenome.

Objective

To review the most recent publications concerning the metabolism of pluripotent stem cells and the role of metabolism in PSC self-renewal and differentiation.

Methods

A systematic literature search related to the metabolism of PSCs was conducted in databases including Medline, Embase, and Web of Science. The search was performed without language restrictions on all papers published before May 2016. The following keywords were used: “metabolism” combined with either “embryonic stem cell” or “epiblast stem cell.”

Results

Hundreds of papers focusing specifically on the metabolism of pluripotent stem cells were uncovered and summarized.

Conclusion

Identifying the specific metabolic pathways involved in pluripotency maintenance is crucial for progress in the field of developmental biology and regenerative medicine. Additionally, better understanding of the metabolism in PSCs will facilitate the derivation and maintenance of authentic PSCs from species other than mouse, rat, and human.

     

Ovarian pluripotent/multipotent stem cells and in vitro oogenesis in mammals

There has been a long persisting dilemma about potential ovarian stem cells in adult mammalian ovaries, including human, and now there is steadily increasing experimental evidence on their existence. After some previous indirect evidence about the presence of stem cells in adult mouse ovaries, an important breakthrough was made by Zou and his co-workers who successfully established long-persisting pluripotent/multipotent ovarian stem cell lines in neonatal and adult mice, and were followed by some other important studies in mouse and human. Moreover, oocyte-like cells can be developed in vitro from pluripotent stem cells of different origins (embryonic stem cells, induced pluripotent stem cells, fetal skin stem cells, pancreatic stem cells). The aim of this article is to elucidate the fast growing new knowledge on the ovarian stem cells and potential in vitro oogenesis in mammals.     

Breathing chromatin in pluripotent stem cells

Only a few factors controlling stem cell pluripotency have been identified to date, and we do not yet fully understand how they act to maintain pluripotency and control differentiation. A report in this issue of Developmental Cell (Meshorer et al., 2006) describes a new trait of pluripotent cells: hyperdynamic or "breathing" chromatin. According to this report, hyperdynamic chromatin is both specific and functionally relevant for pluripotent cells.     

诱导多能干细胞研究进展

人类诱导多能干细胞（induced pluripotent stem cells,iPS细胞）的建立被公认为目前最重要的科技进展之一。iPS细胞在动物疾病模型上的成功治疗,病患特异性iPS细胞的研究及iPS细胞的定向分化研究将有可能使人们避开治疗性克隆的伦理和技术障碍,给人类疾病的干细胞治疗带来光明的前景。本文从iPS细胞的诱导策略和方法,来源细胞及筛选、重编程机制的研究现状、应用前景以及研究中存在的问题等方面对其作一综述和讨论。     

Mouse induced pluripotent stem cells

The recent discovery that it is possible to directly reprogramme somatic cells to an embryonic stem (ES) cell-like pluripotent state, by retroviral transduction of just four genes (Oct3/4, Sox2, c-Myc and Klf4), represents a major breakthrough in stem cell research. The reprogrammed cells, known as induced pluripotent stem (iPS) cells, possess many of the properties of ES cells, and represent one of the most promising sources of patient-specific cells for use in regenerative medicine. While the ultimate goal is the use of iPS cells in the treatment of human disease, much of the research to date has been carried out with murine cells, and improved mouse iPS cells have been shown to contribute to live chimeric mice that are germ-line competent. Very recently, it has been reported that iPS cells can be generated by three factors without c-Myc, and these cells give rise to chimeric mice with a reduced risk of tumour development.     

A molecular view on pluripotent stem cells

Pluripotent stem cells are undifferentiated cells that are capable of differentiating to all three embryonic germ layers and their differentiated derivatives. They are transiently found during embryogenesis, in preimplantation embryos and fetal gonads, or as established cell lines. These unique cell types are distinguished by their wide developmental potential and by their ability to be propagated in culture indefinitely, without loosing their undifferentiated phenotype. This short review intends to give a general overview on the pluripotent nature of embryo-derived stem cells with a focus on human embryonic stem cells.