Search results for: nuclear-reprogramming-and-stem-cells

Nuclear Reprogramming and Stem Cells

Author : Justin Ainscough
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Research into the field of stem cell biology has developed exponentially over recent years, and is beginning to offer significant promise for unravelling the molecular basis of a multitude of disease states. Importantly, in addition to offering the opportunity to delve deeply into the mechanisms that drive disease aetiology the research is realistically opening the doors for development of targeted and personalized therapeutic applications that many considered, until recently, to be nothing more that a far fetched dream. This volume provides a timely glimpse into the methods that have been developed to instigate, and the mechanisms that have been identified to drive, the process of nuclear reprogramming, chronicling how the field has developed over the last 50-60 years. Since the early 1950s a small number of notable experiments have provided significant impetus to the field, primarily the demonstration of reprogramming ability, first by the complex cytoplasmic milieu that constitutes the amphibian egg, then that of the mammalian egg, and finally that of the mammalian embryonic stem cell. Most recently, the demonstration that a limited pool of defined molecules is capable of reprogramming a multitude of cell types has provided massive impetus and facilitated transition towards realistic therapeutic application. We have therefore reproduced some of the key articles that elegantly document these dramatic stages of development of the field in an inclusive appendix to the book, for the benefit of readers keen to investigate the history of how the field of stem cell biology has evolved. Owing to the ever broadening nature of this field, and the incredible rate at which it is evolving, the main content of this volume focuses on areas that have shown significant movement in recent years, are most likely to translate into personalized therapeutic application, and thus provide greatest potential for significant impact on human health in the not too distant future. We recognize that research into many other disease states and cell types are all equally worthy of discussion. We would therefore like to acknowledge those researchers involved whose work we have not been able to include in this volume. Nuclear Reprogramming and Stem Cells will serve as a valuable resource for all researchers in the field of stem cell biology, including those just setting out on their career path as well as those already established in the field.

Nuclear Reprogramming

Author : Steve Pells
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This volume includes various methods of nuclear reprogramming, including nuclear transfer in several different species, both amphibian and mammalian, fusion achieved both by chemical treatment and by electrically shocking cells, and quantitative fusion and reprogramming in vitro treatment of cells with cell extracts. Also described are the different methods of monitoring nuclear programming after nuclear transfer, activation of telomerase as an ES-specific marker, observation of structural changes in the nucleus by both light and electron microscopy, and verification of stem cells' surface marker expression and differentiation potential.

Stem Cells

Author : Gregory R. Bock
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Understanding stem cells at the molecular level is essential to understanding their behaviour in a physiological context. This volume in our acclaimed Novartis Foundation series features animated discussion from the world’s experts in this topic on the important ethical issues that are raised by research on stem cells. They review the various regulatory regimes, which apply in different countries – a key factor in determining where future stem cell research is carried out. Potential clinical applications covered in the book include the production of cardiomyocytes to replace damaged heart tissue, the production of insulin-producing cells for patients with diabetes, and the generation of neurons for the treatment of patients with Parkinson’s disease or spinal cord injury. Particular attention is paid to the factors that maintain stem cells in a pluripotent state or which drive them to create differentiated and lineage-committed cells in vitro and in vivo. Nuclear reprogramming, the process by which a nucleus acquires developmental potential, is covered here as well. It is relevant to stem cell research generally, and also to research on the cloning of animals by nuclear transfer. This book is an essential purchase for all those engaged in stem cell research, whether in the laboratory, the clinic or the regulatory authorities. From the reviews: "...this book provides: a comprehensive overview of current issues in stem cell research, with contributions from leading figures..." —BRITISH SOCIETY OF CELL BIOLOGY

Assessment of Nuclear Reprogramming Activity in Mammalian Embryonic Stem Cells

Author : S. Sullivan
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Cell Reprogramming

Author : Paul J. Verma
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This volume provides an understanding of the factors involved in nuclear reprogramming, which is essential for the success of reprogramming. The book is aimed at reprogramming differentiated cells and germ line transmission of pluripotent stem cells and features chapters that deal with reprogramming-related issues such as analysis of mitochondrial DNA in reprogrammed cells and the isolation of reprogramming intermediates; alternative methods for nuclear transfer; the production of germ-line chimeras from embryonic stem cells and induced pluripotent stem cells; and neonatal care and management of somatic cell nuclear transfer derived offspring. Written in the highly successful Methods in Molecular Biology series format, chapters include introductions to their respective topics, lists of the necessary materials and reagents, step-by-step, readily reproducible laboratory protocols and tips on troubleshooting and avoiding known pitfalls. Authoritative and cutting-edge, Cell Reprogramming: Methods and Protocols

Nuclear Transfer Protocols

Author : Paul J. Verma
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"Nuclear Transfer Protocols: Cell Reprogramming and Transgenesis" is a comprehensive review of nuclear transfer technology in vertebrates, aimed at reprogramming differentiated nuclei and effecting targeted gene transfer. The emphasis here is on providing readily reproducible techniques for the generation of cloned embryos and animals in a number of key research and commercially important vertebrates. Additional chapters provide alternative cutting-edge methods for nuclear transfer, such as zona-free nuclear transfer and post-fusion enucleation nuclear transfer.

Stem Cells Differentiation and Nuclear Reprogramming

Author : Mathias Pawlak
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Nuclear Reprogramming and Culture Requirement for Somatic Cell Nuclear Transfer in Cattle and Mice

Author : Li-Ying Sung
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Secondly, our nuclear transfer experiments demonstrated that premature chromosome condensation (PCC) is not an indispensable prerequisite for the competent reprogramming of a differentiated somatic genome in cattle. The direct exposure of the donor nucleus to the MII cytoplast, presumably containing high levels of MPF, for a relatively short period of time, is sufficient to trigger a cascade of nuclear reprogramming and developmental events. Higher efficiency of blastocyst development was obtained by immediate activation. Similar in vivo developmental potentials of NT embryos derived from substantially varied protocols (IA vs. DA), and the birth of live clones from both treatments, suggest that cattle may represent a unique species with a greater plasticity available for mechanical, biochemical, and physiological manipulation.

Nuclear Reprogramming

Author : Steve Pells
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A wide-ranging collection of readily reproducible methods for performing nuclear reprogramming by nuclear transfer in several different species, by fusion through both chemical treatment and electrically shocking cells, and by in vivo treatment of cells with cell extracts. Several methods of monitoring nuclear reprogramming are also presented, including the use of transgenic markers, activation of telomerase as an ES-specific marker, light and electron microscopic observation of structural changes in the nucleus, and verification of surface marker expression and the differentiation potential of stem cells. Biochemical methods are provided for the examination of chromatin protein modifications, nucleosomal footprinting, transcription factor binding, and the study of DNA methylation changes both at the specific locus level and at the level of the whole nucleus.

Regulatory Networks in Stem Cells

Author : Vinagolu K. Rajasekhar
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Stem cells appear to be fundamental cellular units associated with the origin of multicellular organisms and have evolved to function in safeguarding the cellular homeostasis in organ t- sues. The characteristics of stem cells that distinguish them from other cells have been the fascinating subjects of stem cell research. The important properties of stem cells, such as ma- tenance of quiescence, self-renewal capacity, and differentiation potential, have propelled this exciting ?eld and presently form a common theme of research in developmental biology and medicine. The derivation of pluripotent embryonic stem cells, the prospective identi?cation of multipotent adult stem cells, and, more recently, the induced pluripotent stem cells (popularly called iPS) are important milestones in the arena of stem cell biology. Complex networks of transcription factors, different signaling molecules, and the interaction of genetic and epi- netic events constantly modulate stem cell behavior to evoke programming and reprogramming processes in normal tissue homeostasis during development. In any given cellular scenario, the regulatory networks can pose considerable complexity and yet exert an orderly control of stem cell differentiation during normal development. An aberration in these ?nely tuned processes during development usually results in a spectrum of diseases such as cancers and neurological disorders. Thisunderscorestheimminentneedforamorecompleteunderstandingofmolecular mechanisms underlying the regulatory circuitries required for stem cell maintenance. Overthepast3–5years,adiversegroupofbenchandphysicianscientistshaveprospectively enhanced our knowledge of stem cell biology. These studies are unveiling many unrecognized or previously unknown fundamentals of developmental biology.

Study of Nuclear Reprogramming

Author : Chih-Jen Lin
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Control and Regulation of Stem Cells

Author : Bruce Stillman
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Based on presentations at the 73rd annual Cold Spring Harbor Symposium on Quantitative Biology, this volume reviews the latest advances in stem cell biology. Topics include nuclear reprogramming, the stem cell niche, self-renewal and differentiation, and the role of stem cells in cancer.

STEM CELLS REGENERATIVE MEDI

Author : Walter C. Low
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Stem cells have the ability to differentiate into cells that are found throughout the body. This fundamental property of stem cells suggests that they can potentially be used to replace degenerative cells within the body, and regenerate the functional capacity of organ systems that have deteriorated because of disease or aging. This authoritative textbook provides an overview of the latest advances in the field of stem cell biology, spanning topics that include nuclear reprogramming, somatic cell cloning, and determinants of cell fate; embryonic stem cells for hematopoietic and pancreatic repair; adult stem cells for cardiovascular, neural, renal, and hepatic repair; and manufacturing of stem cells for clinical use.

Reprogramming of Mesenchymal Stem Cells and Adult Fibroblasts Following Nuclear Transfer in Rabbits

Author : Ru Hao
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Stem Cell Therapy for Diabetes

Author : Shimon Efrat
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Stem Cell Therapy for Diabetes, one of the latest installments of the Stem Cell Biology and Regenerative Medicine series, reviews the three main approaches for generation of sufficient numbers of insulin-producing cells for restoration of an adequate beta-cell mass: beta-cell expansion, stem-cell differentiation, and nuclear reprogramming. Adeptly collecting the research of the leading scientists in the field, Stem Cell Therapy for Diabetes compares the merits of employing autologous versus banked allogeneic cell sources for generation of surrogate beta cells, and addresses tissue engineering and ways for cell protection from recurring autoimmunity and graft rejection. Stem Cell Therapy for Diabetes provides essential reading for those especially interested in tracking the progress in applying of one of the most exciting new developments in bio-medicine towards a cure for diabetes.

Is this Cell a Human Being

Author : Antoine Suarez
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The central question of this book is whether or not particular cell entities of human origin ought to be considered human beings. The answer is crucial for making moral decisions for or against research and experimentation. Experts in the field discuss the production of embryonic-like pluripotent stem cells by altered nuclear transfer, parthenogenesis and reprogramming of adult somatic cells. They thoroughly analyse the biological and moral status of different cell entities, such as human stem cells, embryos and human-animal hybrid embryos, and make a decisive step towards establishing final criteria for what constitutes a human being. The topic is challenging in nature and of broad interest to all those concerned with current bioethical thought on embryonic human life and its implications for society.

Stem Cells Regenerative Medicine

Author : Krishnarao Appasani
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Defined as, “The science about the development of an embryo from the fertilization of the ovum to the fetus stage,” embryology has been a mainstay at universities throughout the world for many years. Throughout the last century, embryology became overshadowed by experimental-based genetics and cell biology, transforming the field into developmental biology, which replaced embryology in Biology departments in many universities. Major contributions in this young century in the fields of molecular biology, biochemistry and genomics were integrated with both embryology and developmental biology to provide an understanding of the molecular portrait of a “development cell.” That new integrated approach is known as stem-cell biology; it is an understanding of the embryology and development together at the molecular level using engineering, imaging and cell culture principles, and it is at the heart of this seminal book. Stem Cells and Regenerative Medicine: From Molecular Embryology to Tissue Engineering is completely devoted to the basic developmental, cellular and molecular biological aspects of stem cells as well as their clinical applications in tissue engineering and regenerative medicine. It focuses on the basic biology of embryonic and cancer cells plus their key involvement in self-renewal, muscle repair, epigenetic processes, and therapeutic applications. In addition, it covers other key relevant topics such as nuclear reprogramming induced pluripotency and stem cell culture techniques using novel biomaterials. A thorough introduction to stem-cell biology, this reference is aimed at graduate students, post-docs, and professors as well as executives and scientists in biotech and pharmaceutical companies.

Induced Pluripotency and Reprogramming by Defined Factors

Author : Bryce W. Carey
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The process by which the totipotent zygote undergoes development into an adult organism using a single genome is the foundation for epigenetics and cellular differentiation. Somatic cell nuclear transfer experiments (SCNT) provided unequivocal proof of nuclear equivalency between adult somatic cells. However the molecular mechanisms of somatic cell reprogramming have remained largely a mystery. Recent advancements in epigenetic reprogramming by defined factors provide new opportunities to explore factors that regulate induction of pluripotency in somatic cells. Nuclear reprogramming by SCNT occurs in an 'indirect' manner by unidentified components within oocyte cytoplasm and requires the destruction of embryos. The introduction of induced pluripotent stem cells (iPS cells) and 'direct' reprogramming methods created a tractable system to both study of the process in vitro and potentially derive personalized pluripotent stem cells free of the practical and ethical concerns surrounding embryonic stem (ES) cells and SCNT. Herein we study mouse somatic cell reprogramming by defined factors and develop novel tools to compare the induced pluripotent state to the gold standard of pluripotency, ES cells. First, we designed reprogramming vectors that minimize the number of viruses required to generate iPS cells, yielding pluripotent cells with minimal genomic alterations from reprogramming factors. This allowed the creation of transgenic "reprogrammable mouse" strains after gene targeting in ES cells, providing a renewable source of somatic cells that can be induced to pluripotency by addition of a drug. In addition we can easily introduce or mate these strains to study unique genetic variants during reprogramming. Third, we study factors that influence the induced pluripotent state, specifically how to generate pluripotent cells with all properties of embryonic stem cells including derivation of "all iPS cell mice" by tetraploid complementation assays. In contrast to previous reports, we find the majority ( - 80%) of iPS cell lines derived from adult somatic cells of varying organs contain the developmental potential of ES cells. This outcome correlated with high expression of Oct4 and Klf4 and low expression of Sox2 and c-Myc during reprogramming. In addition we report that adult mice derived from iPS cells are healthy and do not develop tumors. Together these results suggest in vitro reprogramming to pluripotency by defined factors holds great promise for regenerative medicine.

Manipulating Somatic Cells to Remove Barriers in Induced Pluripotent Stem Cell Reprogramming

Author : Julia Chung
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Development leads unidirectionally towards a more restricted cell fate that is usually stable. However, it has been proven that developmental systems are reversible by the success of animal cloning of a differentiated somatic genome through somatic cell nuclear transfer (SCNT). Recently, reprogramming of somatic cells to a pluripotent embryonic stem cell (ESC)-like state by introducing defined transcripton factor has been achieved, resulting in the generation of induced pluripotent stem cells (iPSCs), which resemble ESCs. iPSC reprogramming is of great medical interest, as it has the potential to generate a source of patient-specific cells. However, the dangerous delivery method, low efficiency, and slow kinetics of the reprogramming process have hampered progress with this technology.

Reprogramming Cellular Fate Using Defined Factors

Author : Ruth Kjelgaard Foreman
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Embryonic stem (ES) cells have a vast therapeutic potential given their pluripotency, or the ability to differentiate into tissues from all three germ layers. One of the ultimate goals of regenerative medicine is to isolate pluripotent stem cells from patients. Nuclear reprogramming offers the possibility of creating patient-specific cell lines, thus abrogating the need for immunosuppressants following cell transplantation therapy. It was recently reported that the forced expression of four transcription factors, Oct4, Sox2, c-Myc and Klf4 can induce a pluripotent state in somatic cells, without the need for embryo destruction. The work presented here aims to characterize reprogramming using defined factors and provide insight into the mechanisms governing this process. It also seeks to identify transient cues to induce reprogramming in somatic cells, alleviating the need for virally transduced transcription factors that hinder its eventual clinical use.