Molecular live cell bioimaging in stem cell research

Functional heterogeneity within stem and progenitor cells has been shown to influence cell fate decisions. Similarly, intracellular signaling activated by external stimuli is highly heterogeneous and its spatiotemporal activity is linked to future cell behavior. To quantify these heterogeneous states and link them to future cell fates, it is important to observe cell populations continuously with single cell resolution. Live cell imaging in combination with fluorescent biosensors for signaling activity serves as a powerful tool to study cellular and molecular heterogeneity and the long-term biological effects of signaling. Here, we describe these methodologies, their advantages over classical approaches, and we illustrate how they could be applied to improve our understanding of the importance of heterogeneous cellular and molecular responses to external signaling cues.     

Molecular imaging of embryonic stem cell misbehavior and suicide gene ablation

Numerous studies have demonstrated the potential use of stem cells for the repair and regeneration of injured tissues. However, tracking transplanted stem cell fate and function in vivo remains problematic. To address these issues, murine embryonic stem (ES) cells were stably transduced with self-inactivating lentiviral vectors carrying either a triple fusion (TF) or double fusion (DF) reporter gene construct. The TF consisted of monomeric red fluorescence protein (mrfp), firefly luciferase (Fluc), and herpes simplex virus truncated thymidine kinase (HSV-ttk) reporter genes. The DF consisted of enhanced green fluorescence protein (egfp) and Fluc reporter genes but lacked HSV-ttk. Stably transduced ES-TF or ES-DF cells were selected by fluorescence activated cell sorting based on either mrfp (TF) or egfp (DF) expression. Afterwards, cells were injected subcutaneously into the right (ES-TF cells) and left (ES-DF cells) shoulders of adult female nude mice. Cell survival was tracked noninvasively by bioluminescence and positron emission tomography imaging of Fluc and HSV-ttk reporter genes, respectively. Imaging signals progressively increased from day 2 to day 14, consistent with ES cell survival and proliferation in vivo. However, teratoma formation occurred in all nude mice after 5 weeks. Administration of ganciclovir (GCV), targeting the HSV-ttk gene, resulted in selective ablation of teratomas arising from the ES-TF cells but not ES-DF cells. These data demonstrate the novel use of multimodality imaging techniques to (1) monitor transplanted ES cell survival and proliferation in vivo and (2) assess the efficacy of suicide gene therapy as a backup safety measure against teratoma formation.     

Controversies over stem cell research

Much interest and effort has focused on the therapeutic potential of stem cell technology to treat presently intractable diseases. However, this scientific promise has been accompanied by important issues, including ethical hurdles, political policies and dilemmas concerning cell-source selection (embryonic versus adult stem cells). Although the contribution of stem cells to medical research seems enormous, many countries now face complex ethical and regulatory questions, which could represent a significant limitation to medical progress and could take years to overcome.     

International stem cell research considerations

Legislative bodies in the international arena and in individual countries are actively engaged in developing policies regarding the establishment, distribution and use of human embryonic stem cells. Present and anticipated policies concerning research on human adult and embryonic stem cells of possible medical importance reflect the wide spectrum of popular views that range from complete rejection to enthusiastic support. Since the public debate concerning the use of human gametes or embryos for research purposes is not likely to abate anytime soon, all the more urgent becomes the quest for alternative approaches toward generating stem cells that are not embryonic and yet are pluripotent.     

干细胞研究伦理

干细胞具有＂分化＂和＂脱分化＂的特点和潜能,干细胞研究有着良好的医学前景,许多人类严重疾病的治疗有了新的希望。伴随着干细胞研究的开展和深入,出现了诸多伦理问题的争论。拟就干细胞研究的希望和现实、伦理争论的主要观点及干细胞研究伦理准则的构建,作一简要介绍,并就加强干细胞管理提出建议。     

肿瘤干细胞研究进展

目前,癌症是导致人类死亡的主要因素之一。尽管在癌症治疗方面取得了巨大进展,但是,其较高的复发率还是会导致死亡。连续治疗失败的一个可能原因是,残留的恶性细胞有类似干细胞的分化潜能,这样就能再次形成肿瘤和造成病灶转移。肿瘤干细胞(cancer stem cell,CSC)假说认为,肿瘤组织中存在具有自我跟新能力,无限增殖和肿瘤形成能力的一小部分肿瘤细胞,近年来,随着在血液肿瘤和实体瘤中相继发现CSC存在的相关证据,对CSC的生物学特性的认识不断深入,对肿瘤的复发、病灶转移、耐药性形成也有了新的观点和研究方向,目前的研究主要集中在其分离鉴定阶段,本文就近年来该方面的研究进展作一综述。     

Perspectives on human stem cell research

Human stem cell research draws not only scientists' but the public's attention. Human stem cell research is considered to be able to identify the mechanism of human development and change the paradigm of medical practices. However, there are heated ethical and legal debates about human stem cell research. The core issue is that of human dignity and human life. Some prefer human adult stem cell research or iPS cell research, others hES cell research. We do not need to exclude any type of stem cell research because each has its own merits and issues, and they can facilitate the scientific revolution when working together. J. Cell. Physiol. 220: 535–537, 2009. © 2009 Wiley‐Liss, Inc.     

肿瘤干细胞的认知历程与研发热点

肿瘤组织中存在一小群能够自我更新、增殖和分化,对肿瘤的发生、发展、复发、转移起决定作用的细胞,即肿瘤干细胞（cancer stem cells,CSCs）。在传统理论方法已不能攻克癌症的情况下,肿瘤干细胞理论为我们重新认识肿瘤的起源和本质提供了新的方向和视角。从20世纪50年代至今,随着生物技术的发展,肿瘤干细胞理论经历了从设想到验证的漫长历程。但该理论自提出之日起便受到来自各方面不同观点的质疑。当今针对肿瘤干细胞癌症治疗主要集中在靶向问题上。因此,寻找特异的肿瘤干细胞标志物,探索肿瘤干细胞与周围微环境间的复杂关系以及发现调控其功能的关键信号通路成为当前研究的热点。     

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.     

干细胞的研究进展及临床应用前景

干细胞是一类具有自我复制和分化潜能的影响,它包括胚胎干细胞和成体干细胞。干细胞的发育受多种内在机制和微环境因素的影响。目前人类胚胎干细胞已可在体外培养。最新研究发现,成体干细胞可以横向分化为其他类型的细胞,为干细胞的广泛应用提供了基础。     

Critical appraisal of the side population assay in stem cell and cancer stem cell research

The "Side Population" (SP) discrimination assay is a flow cytometry method used to detect stem cells based on the dye efflux properties of ABC transporters. We discuss the SP assay and its applications in stem cell?biology, with an emphasis on the technical challenges related to sample preparation, data acquisition, analysis, and interpretation. We highlight the value of multicolor phenotyping, the impact of DNA ploidy, and the importance of distinguishing graft versus host cells for an appropriate SP discrimination. To improve the consistency and reliability of data between laboratories, we propose a set of recommendations for SP assay data reporting.     

The impact of cell culture on stem cell research

Induced pluripotent stem cell research has broadened possibilities for regenerative medicine and captured the world's attention in a way that science rarely does. However, clinical applications utilizing cultured stem cells have existed for >30 years and can assist benchers and bedsiders in identifying and expediting promising avenues for future therapies.     

Molecular mass spectrometry imaging in biomedical and life science research

This review describes the current state of mass spectrometry imaging (MSI) in life sciences. A brief overview of mass spectrometry principles is presented followed by a thorough introduction to the MSI workflows, principles and areas of application. Three major desorption-ionization techniques used in MSI, namely, secondary ion mass spectrometry (SIMS), matrix-assisted laser desorption ionization (MALDI), and desorption electrospray ionization (DESI) are described, and biomedical and life science imaging applications of each ionization technique are reviewed. A separate section is devoted to data handling and current challenges and future perspectives are briefly discussed at the end.