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This chapter covers gene expression analysis by microarray to study and characterize stem cells. In a case-study scenario, we describe basic bioinformatic methodologies used to answer common questions in microarray experiments involving one or more stem cell populations. Service providers or departmental core labs usually carry out sample preparation, hybridization, and scanning of microarrays. Therefore, in this chapter, we focus on the state-of-the-art data analysis that avoids common pitfalls and introduces the reader to important controls that yield robust biologically relevant results. We describe evaluation of differentially expressed genes, clustering methods, gene-set enrichment analysis, and gene network discovery methods that can be used to formulate meaningful biological insights as well as suggest new wet lab experiments.

Neural stem cell (NSC) transplantation represents an unexplored approach for treating neurodegenerative disorders associated with cognitive decline such as Alzheimer disease (AD). Here, we used aged triple transgenic mice (3xTg-AD) that express pathogenic forms of amyloid precursor protein, presenilin, and tau to investigate the effect of neural stem cell transplantation on AD-related neuropathology and cognitive dysfunction. Interestingly, despite widespread and established Ass plaque and neurofibrillary tangle pathology, hippocampal neural stem cell transplantation rescues the spatial learning and memory deficits in aged 3xTg-AD mice. Remarkably, cognitive function is improved without altering Ass or tau pathology. Instead, the mechanism underlying the improved cognition involves a robust enhancement of hippocampal synaptic density, mediated by brain-derived neurotrophic factor (BDNF). Gain-of-function studies show that recombinant BDNF mimics the beneficial effects of NSC transplantation. Furthermore, loss-of-function studies show that depletion of NSC-derived BDNF fails to improve cognition or restore hippocampal synaptic density. Taken together, our findings demonstrate that neural stem cells can ameliorate complex behavioral deficits associated with widespread Alzheimer disease pathology via BDNF.

Stem cells are defined as self-renewing cell populations that can differentiate into multiple distinct cell types. However, hundreds of different human cell lines from embryonic, fetal and adult sources have been called stem cells, even though they range from pluripotent cells-typified by embryonic stem cells, which are capable of virtually unlimited proliferation and differentiation-to adult stem cell lines, which can generate a far more limited repertoire of differentiated cell types. The rapid increase in reports of new sources of stem cells and their anticipated value to regenerative medicine has highlighted the need for a general, reproducible method for classification of these cells. We report here the creation and analysis of a database of global gene expression profiles (which we call the 'stem cell matrix') that enables the classification of cultured human stem cells in the context of a wide variety of pluripotent, multipotent and differentiated cell types. Using an unsupervised clustering method to categorize a collection of approximately 150 cell samples, we discovered that pluripotent stem cell lines group together, whereas other cell types, including brain-derived neural stem cell lines, are very diverse. Using further bioinformatic analysis we uncovered a protein-protein network (PluriNet) that is shared by the pluripotent cells (embryonic stem cells, embryonal carcinomas and induced pluripotent cells). Analysis of published data showed that the PluriNet seems to be a common characteristic of pluripotent cells, including mouse embryonic stem and induced pluripotent cells and human oocytes. Our results offer a new strategy for classifying stem cells and support the idea that pluripotency and self-renewal are under tight control by specific molecular networks.

Neural stem cells are a self-renewing population that generates the neurons and glia of the developing brain. They can be isolated, proliferated, genetically manipulated and differentiated in vitro and reintroduced into a developing, adult or pathologically altered CNS. Neural stem cells have been considered for use in cell replacement therapies in various neurodegenerative diseases, and an unexpected and potentially valuable characteristic of these cells has recently been revealed--they are highly migratory and seem to be attracted to areas of brain pathology such as ischaemic and neoplastic lesions. Here, we speculate on the ways in which neural stem cells might be exploited as delivery vehicles for gene therapy in the CNS.

The introduction of stem cell-associated molecular factors into human patient-derived cells allows for their reprogramming in the laboratory environment. As a result, human induced pluripotent stem cells (hiPSC) can now be reprogrammed epigenetically without disruption of their overall genomic integrity. For patients with neurodegenerative diseases characterized by progressive loss of functional neurons, the ability to reprogram any individual's cells and drive their differentiation toward susceptible neuronal subtypes holds great promise. Apart from applications in regenerative medicine and cell replacement-based therapy, hiPSCs are increasingly used in preclinical research for establishing disease models and screening for drug toxicities. The rapid developments in this field prompted us to review recent progress toward the applications of stem cell technologies for movement disorders. We introduce reprogramming strategies and explain the critical steps in the differentiation of hiPSCs to clinical relevant subtypes of cells in the context of movement disorders. We summarize and discuss recent discoveries in this field, which, based on the rapidly expanding basic science literature as well as upcoming trends in personalized medicine, will strongly influence the future therapeutic options available to practitioners working with patients suffering from such disorders. © 2014 International Parkinson and Movement Disorder Society [ABSTRACT FROM AUTHOR]

It is widely accepted that the (now reversed) Bush administration's decision to restrict federal funding for human embryonic stem cell (hESC) research to a few "eligible" hESC lines is responsible for the sustained preferential use of a small subset of hESC lines (principally the H1 and H9 lines) in basic and preclinical research. Yet, international hESC usage patterns, in both permissive and restrictive political environments, do not correlate with a specific type of stem cell policy. Here we conducted a descriptive analysis of hESC line usage and compared the ability of policy-driven processes and collaborative processes inherent to biomedical research to recapitulate global hESC usage patterns. We find that current global hESC usage can be modelled as a cumulative advantage process, independent of restrictive or permissive policy influence, suggesting a primarily innovation-driven (rather than policy-driven) mechanism underlying human pluripotent stem cell usage in preclinical research. [ABSTRACT FROM AUTHOR]

The article discusses the challenges PluriTest and other global assays have to address to fulfill their potential for industrial, diagnostic and therapeutic applications. It focuses on the conceptual basis that underlies the unprecedented opportunity to build and use predictive, data-driven models in biology today. It suggests that these challenges can be solved within a framework of unbiased genome-wide datasets taken from nucleotide-based high-content measurements.

Pluripotent stem cells (PSCs) are defined by their potential to generate all cell types of an organism. The standard assay for pluripotency of mouse PSCs is cell transmission through the germline, but for human PSCs researchers depend on indirect methods such as differentiation into teratomas in immunodeficient mice. Here we report PluriTest, a robust open-access bioinformatic assay of pluripotency in human cells based on their gene expression profiles. [ABSTRACT FROM AUTHOR]

The article calls for the standardization of teratoma assays used to define human pluripotent cell lines. It indicates that germline transmission is the standard for proving pluripotency for mouse cells which demonstrated the ability to make all cell types. Researchers are suggested to explore a systematic reporting system for teratoma experiments and data. It claims that standards help them in the evaluation and the comparison of results across different reprogramming strategies and protocols.

Abstract: Human pluripotent stem cells have the unique properties of being able to proliferate indefinitely in their undifferentiated state and to differentiate into any somatic cell type. These cells are thus posited to be extremely useful for furthering our understanding of both normal and abnormal human development, providing a human cell preparation that can be used to screen for new reagents or therapeutic agents, and generating large numbers of differentiated cells that can be used for transplantation purposes. Critical among the applications for the latter are diseases and injuries of the nervous system, medical approaches to which have been, to date, primarily palliative in nature. Differentiation of human pluripotent stem cells into cells of the neural lineage, therefore, has become a central focus of a number of laboratories. This has resulted in the description in the literature of several dozen methods for neural cell differentiation from human pluripotent stem cells. Among these are methods for the generation of such divergent neural cells as dopaminergic neurons, retinal neurons, ventral motoneurons, and oligodendroglial progenitors. In this review, we attempt to fully describe most of these methods, breaking them down into five basic subdivisions: (1) starting material, (2) induction of loss of pluripotency, (3) neural induction, (4) neural maintenance and expansion, and (5) neuronal/glial differentiation. We also show data supporting the concept that undifferentiated human pluripotent stem cells appear to have an innate neural differentiation potential. In addition, we evaluate data comparing and contrasting neural stem cells differentiated from human pluripotent stem cells with those derived directly from the human brain. [Copyright &y& Elsevier]