Open AccessFeature PaperReview
Assays to Monitor Autophagy in Saccharomyces cerevisiae
Cells 2017, 6(3), 23; doi:10.3390/cells6030023 -
Abstract
Autophagy is an intracellular process responsible for the degradation and recycling of cytoplasmic components. It selectively removes harmful cellular material and enables the cell to survive starvation by mobilizing nutrients via the bulk degradation of cytoplasmic components. While research over the last decades
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Autophagy is an intracellular process responsible for the degradation and recycling of cytoplasmic components. It selectively removes harmful cellular material and enables the cell to survive starvation by mobilizing nutrients via the bulk degradation of cytoplasmic components. While research over the last decades has led to the discovery of the key factors involved in autophagy, the pathway is not yet completely understood. The first studies of autophagy on a molecular level were conducted in the yeast Saccharomyces cerevisiae. Building up on these studies, many homologs have been found in higher eukaryotes. Yeast remains a highly relevant model organism for studying autophagy, with a wide range of established methods to elucidate the molecular details of the autophagy pathway. In this review, we provide an overview of methods to study both selective and bulk autophagy, including intermediate steps in the yeast Saccharomyces cerevisiae. We compare different assays, discuss their advantages and limitations and list potential applications. Full article
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Open AccessFeature PaperReview
Exploring Autophagy in Drosophila
Cells 2017, 6(3), 22; doi:10.3390/cells6030022 -
Abstract
Autophagy is a catabolic process in eukaryotic cells promoting bulk or selective degradation of cellular components within lysosomes. In recent decades, several model systems were utilized to dissect the molecular machinery of autophagy and to identify the impact of this cellular “self-eating” process
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Autophagy is a catabolic process in eukaryotic cells promoting bulk or selective degradation of cellular components within lysosomes. In recent decades, several model systems were utilized to dissect the molecular machinery of autophagy and to identify the impact of this cellular “self-eating” process on various physiological and pathological processes. Here we briefly discuss the advantages and limitations of using the fruit fly Drosophila melanogaster, a popular model in cell and developmental biology, to apprehend the main pathway of autophagy in a complete animal. Full article
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Open AccessFeature PaperReview
Studying Autophagy in Zebrafish
Cells 2017, 6(3), 21; doi:10.3390/cells6030021 -
Abstract
Autophagy is an evolutionarily conserved catabolic process which allows lysosomal degradation of complex cytoplasmic components into basic biomolecules that are recycled for further cellular use. Autophagy is critical for cellular homeostasis and for degradation of misfolded proteins and damaged organelles as well as
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Autophagy is an evolutionarily conserved catabolic process which allows lysosomal degradation of complex cytoplasmic components into basic biomolecules that are recycled for further cellular use. Autophagy is critical for cellular homeostasis and for degradation of misfolded proteins and damaged organelles as well as intracellular pathogens. The role of autophagy in protection against age-related diseases and a plethora of other diseases is now coming to light; assisted by several divergent eukaryotic model systems ranging from yeast to mice. We here give an overview of different methods used to analyse autophagy in zebrafish—a relatively new model for studying autophagy—and briefly discuss what has been done so far and possible future directions. Full article
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Open AccessFeature PaperReview
Assays to Monitor Autophagy Progression in Cell Cultures
Cells 2017, 6(3), 20; doi:10.3390/cells6030020 -
Abstract
The vast number of implications of autophagy in multiple areas of life sciences and medicine has attracted the interest of numerous scientists that aim to unveil the role of this process in specific physiological and pathological contexts. Cell cultures are one of the
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The vast number of implications of autophagy in multiple areas of life sciences and medicine has attracted the interest of numerous scientists that aim to unveil the role of this process in specific physiological and pathological contexts. Cell cultures are one of the most frequently used experimental setup for the investigation of autophagy. As a result, it is essential to assess this highly regulated molecular pathway with efficient and reliable methods. Each method has its own advantages and disadvantages. Here, we present a review summarizing the most established assays used to monitor autophagy induction and progression in cell cultures, in order to guide researchers in the selection of the most optimal solution for their experimental setup and design. Full article
Open AccessFeature PaperReview
Methods to Monitor and Quantify Autophagy in the Social Amoeba Dictyostelium discoideum
Cells 2017, 6(3), 18; doi:10.3390/cells6030018 -
Abstract
Autophagy is a eukaryotic catabolic pathway that degrades and recycles cellular components to maintain homeostasis. It can target protein aggregates, superfluous biomolecular complexes, dysfunctional and damaged organelles, as well as pathogenic intracellular microbes. Autophagy is a dynamic process in which the different stages
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Autophagy is a eukaryotic catabolic pathway that degrades and recycles cellular components to maintain homeostasis. It can target protein aggregates, superfluous biomolecular complexes, dysfunctional and damaged organelles, as well as pathogenic intracellular microbes. Autophagy is a dynamic process in which the different stages from initiation to final degradation of cargo are finely regulated. Therefore, the study of this process requires the use of a palette of techniques, which are continuously evolving and whose interpretation is not trivial. Here, we present the social amoeba Dictyostelium discoideum as a relevant model to study autophagy. Several methods have been developed based on the tracking and observation of autophagosomes by microscopy, analysis of changes in expression of autophagy genes and proteins, and examination of the autophagic flux with various techniques. In this review, we discuss the pros and cons of the currently available techniques to assess autophagy in this organism. Full article
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Open AccessFeature PaperReview
Vascular Mechanobiology: Towards Control of In Situ Regeneration
Cells 2017, 6(3), 19; doi:10.3390/cells6030019 -
Abstract
The paradigm of regenerative medicine has recently shifted from in vitro to in situ tissue engineering: implanting a cell-free, biodegradable, off-the-shelf available scaffold and inducing the development of functional tissue by utilizing the regenerative potential of the body itself. This approach offers a
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The paradigm of regenerative medicine has recently shifted from in vitro to in situ tissue engineering: implanting a cell-free, biodegradable, off-the-shelf available scaffold and inducing the development of functional tissue by utilizing the regenerative potential of the body itself. This approach offers a prospect of not only alleviating the clinical demand for autologous vessels but also circumventing the current challenges with synthetic grafts. In order to move towards a hypothesis-driven engineering approach, we review three crucial aspects that need to be taken into account when regenerating vessels: (1) the structure-function relation for attaining mechanical homeostasis of vascular tissues, (2) the environmental cues governing cell function, and (3) the available experimental platforms to test instructive scaffolds for in situ tissue engineering. The understanding of cellular responses to environmental cues leads to the development of computational models to predict tissue formation and maturation, which are validated using experimental platforms recapitulating the (patho)physiological micro-environment. With the current advances, a progressive shift is anticipated towards a rational and effective approach of building instructive scaffolds for in situ vascular tissue regeneration. Full article
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Open AccessFeature PaperReview
Standard Immunohistochemical Assays to Assess Autophagy in Mammalian Tissue
Cells 2017, 6(3), 17; doi:10.3390/cells6030017 -
Abstract
Autophagy is a highly conserved lysosomal degradation pathway with major impact on diverse human pathologies. Despite the development of different methodologies to detect autophagy both in vitro and in vivo, monitoring autophagy in tissue via immunohistochemical techniques is hampered due to the lack
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Autophagy is a highly conserved lysosomal degradation pathway with major impact on diverse human pathologies. Despite the development of different methodologies to detect autophagy both in vitro and in vivo, monitoring autophagy in tissue via immunohistochemical techniques is hampered due to the lack of biomarkers. Immunohistochemical detection of a punctate pattern of ATG8/MAP1LC3 proteins is currently the most frequently used approach to detect autophagy in situ, but it depends on a highly sensitive detection method and is prone to misinterpretation. Moreover, reliable MAP1LC3 immunohistochemical staining requires correct tissue processing and high-quality, isoform-specific antibodies. Immunohistochemical analysis of other autophagy-related protein targets such as SQSTM1, ubiquitin, ATG5 or lysosomal proteins is not recommended as marker for autophagic activity in tissue for multiple reasons including aspecific labeling of cellular structures and a lack of differential protein expression during autophagy initiation. To better understand the role of autophagy in human disease, novel biomarkers for visualization of the autophagic process with standard histology techniques are urgently needed. Full article
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Open AccessFeature PaperReview
Assessing Autophagy in Mouse Models and Patients with Systemic Autoimmune Diseases
Cells 2017, 6(3), 16; doi:10.3390/cells6030016 -
Abstract
Autophagy is a tightly regulated mechanism that allows cells to renew themselves through the lysosomal degradation of proteins, which are misfolded or produced in excess, and of damaged organelles. In the context of immunity, recent research has specially attempted to clarify its roles
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Autophagy is a tightly regulated mechanism that allows cells to renew themselves through the lysosomal degradation of proteins, which are misfolded or produced in excess, and of damaged organelles. In the context of immunity, recent research has specially attempted to clarify its roles in infection, inflammation and autoimmunity. Autophagy has emerged as a spotlight in several molecular pathways and trafficking events that participate to innate and adaptive immunity. Deregulation of autophagy has been associated to several autoimmune diseases, in particular to systemic lupus erythematosus. Nowadays, however, experimental data on the implication of autophagy in animal models of autoimmunity or patients remain limited. In our investigations, we use Murphy Roths Large (MRL)/lymphoproliferation (lpr) lupus-prone mice as a mouse model for lupus and secondary Sjögren’s syndrome, and, herein, we describe methods applied routinely to analyze different autophagic pathways in different lymphoid organs and tissues (spleen, lymph nodes, salivary glands). We also depict some techniques used to analyze autophagy in lupus patient’s blood samples. These methods can be adapted to the analysis of autophagy in other mouse models of autoinflammatory diseases. The understanding of autophagy implication in autoimmune diseases could prove to be very useful for developing novel immunomodulatory strategies. Our attention should be focused on the fact that autophagy processes are interconnected and that distinct pathways can be independently hyper-activated or downregulated in distinct organs and tissues of the same individual. Full article
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Open AccessFeature PaperReview
Telomere Biology—Insights into an Intriguing Phenomenon
Cells 2017, 6(2), 15; doi:10.3390/cells6020015 -
Abstract
Bacteria and viruses possess circular DNA, whereas eukaryotes with typically very large DNA molecules have had to evolve into linear chromosomes to circumvent the problem of supercoiling circular DNA of that size. Consequently, such organisms possess telomeres to cap chromosome ends. Telomeres are
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Bacteria and viruses possess circular DNA, whereas eukaryotes with typically very large DNA molecules have had to evolve into linear chromosomes to circumvent the problem of supercoiling circular DNA of that size. Consequently, such organisms possess telomeres to cap chromosome ends. Telomeres are essentially tandem repeats of any DNA sequence that are present at the ends of chromosomes. Their biology has been an enigmatic one, involving various molecules interacting dynamically in an evolutionarily well-trimmed fashion. Telomeres range from canonical hexameric repeats in most eukaryotes to unimaginably random retrotransposons, which attach to chromosome ends and reverse-transcribe to DNA in some plants and insects. Telomeres invariably associate with specialised protein complexes that envelop it, also regulating access of the ends to legitimate enzymes involved in telomere metabolism. They also transcribe into repetitive RNA which also seems to be playing significant roles in telomere maintenance. Telomeres thus form the intersection of DNA, protein, and RNA molecules acting in concert to maintain chromosome integrity. Telomere biology is emerging to appear ever more complex than previously envisaged, with the continual discovery of more molecules and interplays at the telomeres. This review also includes a section dedicated to the history of telomere biology, and intends to target the scientific audience new to the field by rendering an understanding of the phenomenon of chromosome end protection at large, with more emphasis on the biology of human telomeres. The review provides an update on the field and mentions the questions that need to be addressed. Full article
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Open AccessReview
Methods for Measuring Autophagy in Mice
Cells 2017, 6(2), 14; doi:10.3390/cells6020014 -
Abstract
Autophagy is a dynamic intracellular process that mediates the degradation of damaged cytoplasmic components by the lysosome. This process plays important roles in maintaining normal cellular homeostasis and energy balance. Measuring autophagy activity is critical and although the determination of autophagic flux in
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Autophagy is a dynamic intracellular process that mediates the degradation of damaged cytoplasmic components by the lysosome. This process plays important roles in maintaining normal cellular homeostasis and energy balance. Measuring autophagy activity is critical and although the determination of autophagic flux in isolated cells is well documented, there is a need to have reliable and quantitative assays to evaluate autophagy in whole organisms. Because mouse models have been precious in establishing the functional significance of autophagy under physiological or pathological conditions, we present in this chapter a compendium of the current available methods to measure autophagy in mice, and discuss their advantages and limitations. Full article
Open AccessFeature PaperReview
Activation of the EGF Receptor by Ligand Binding and Oncogenic Mutations: The “Rotation Model”
Cells 2017, 6(2), 13; doi:10.3390/cells6020013 -
Abstract
The epidermal growth factor receptor (EGFR) plays vital roles in cellular processes including cell proliferation, survival, motility, and differentiation. The dysregulated activation of the receptor is often implicated in human cancers. EGFR is synthesized as a single-pass transmembrane protein, which consists of an
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The epidermal growth factor receptor (EGFR) plays vital roles in cellular processes including cell proliferation, survival, motility, and differentiation. The dysregulated activation of the receptor is often implicated in human cancers. EGFR is synthesized as a single-pass transmembrane protein, which consists of an extracellular ligand-binding domain and an intracellular kinase domain separated by a single transmembrane domain. The receptor is activated by a variety of polypeptide ligands such as epidermal growth factor and transforming growth factor α. It has long been thought that EGFR is activated by ligand-induced dimerization of the receptor monomer, which brings intracellular kinase domains into close proximity for trans-autophosphorylation. An increasing number of diverse studies, however, demonstrate that EGFR is present as a pre-formed, yet inactive, dimer prior to ligand binding. Furthermore, recent progress in structural studies has provided insight into conformational changes during the activation of a pre-formed EGFR dimer. Upon ligand binding to the extracellular domain of EGFR, its transmembrane domains rotate or twist parallel to the plane of the cell membrane, resulting in the reorientation of the intracellular kinase domain dimer from a symmetric inactive configuration to an asymmetric active form (the “rotation model”). This model is also able to explain how oncogenic mutations activate the receptor in the absence of the ligand, without assuming that the mutations induce receptor dimerization. In this review, we discuss the mechanisms underlying the ligand-induced activation of the preformed EGFR dimer, as well as how oncogenic mutations constitutively activate the receptor dimer, based on the rotation model. Full article
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Open AccessReview
Major Tumor Suppressor and Oncogenic Non-Coding RNAs: Clinical Relevance in Lung Cancer
Cells 2017, 6(2), 12; doi:10.3390/cells6020012 -
Abstract
Lung cancer is the leading cause of cancer deaths worldwide, yet there remains a lack of specific and sensitive tools for early diagnosis and targeted therapies. High-throughput sequencing techniques revealed that non-coding RNAs (ncRNAs), e.g., microRNAs and long ncRNAs (lncRNAs), represent more than
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Lung cancer is the leading cause of cancer deaths worldwide, yet there remains a lack of specific and sensitive tools for early diagnosis and targeted therapies. High-throughput sequencing techniques revealed that non-coding RNAs (ncRNAs), e.g., microRNAs and long ncRNAs (lncRNAs), represent more than 80% of the transcribed human genome. Emerging evidence suggests that microRNAs and lncRNAs regulate target genes and play an important role in biological processes and signaling pathways in malignancies, including lung cancer. In lung cancer, several tumor suppressor/oncogenic microRNAs and lncRNAs function as biomarkers for metastasis and prognosis, and thus may serve as therapeutic tools. In this review, recent work on microRNAs and lncRNAs is introduced and briefly summarized with a focus on potential biological and therapeutic applications. Full article
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Open AccessFeature PaperReview
Taking a Bad Turn: Compromised DNA Damage Response in Leukemia
Cells 2017, 6(2), 11; doi:10.3390/cells6020011 -
Abstract
Genomic integrity is of outmost importance for the survival at the cellular and the organismal level and key to human health. To ensure the integrity of their DNA, cells have evolved maintenance programs collectively known as the DNA damage response. Particularly challenging for
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Genomic integrity is of outmost importance for the survival at the cellular and the organismal level and key to human health. To ensure the integrity of their DNA, cells have evolved maintenance programs collectively known as the DNA damage response. Particularly challenging for genome integrity are DNA double-strand breaks (DSB) and defects in their repair are often associated with human disease, including leukemia. Defective DSB repair may not only be disease-causing, but further contribute to poor treatment outcome and poor prognosis in leukemia. Here, we review current insight into altered DSB repair mechanisms identified in leukemia. While DSB repair is somewhat compromised in all leukemic subtypes, certain key players of DSB repair are particularly targeted: DNA-dependent protein kinase (DNA-PK) and Ku70/80 in the non-homologous end-joining pathway, as well as Rad51 and breast cancer 1/2 (BRCA1/2), key players in homologous recombination. Defects in leukemia-related DSB repair may not only arise from dysfunctional repair components, but also indirectly from mutations in key regulators of gene expression and/or chromatin structure, such as p53, the Kirsten ras oncogene (K-RAS), and isocitrate dehydrogenase 1 and 2 (IDH1/2). A detailed understanding of the basis for defective DNA damage response (DDR) mechanisms for each leukemia subtype may allow to further develop new treatment methods to improve treatment outcome and prognosis for patients. Full article
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Open AccessArticle
Distinct Fiber Type Signature in Mouse Muscles Expressing a Mutant Lamin A Responsible for Congenital Muscular Dystrophy in a Patient
Cells 2017, 6(2), 10; doi:10.3390/cells6020010 -
Abstract
Specific mutations in LMNA, which encodes nuclear intermediate filament proteins lamins A/C, affect skeletal muscle tissues. Early-onset LMNA myopathies reveal different alterations of muscle fibers, including fiber type disproportion or prominent dystrophic and/or inflammatory changes. Recently, we identified the p.R388P LMNA mutation
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Specific mutations in LMNA, which encodes nuclear intermediate filament proteins lamins A/C, affect skeletal muscle tissues. Early-onset LMNA myopathies reveal different alterations of muscle fibers, including fiber type disproportion or prominent dystrophic and/or inflammatory changes. Recently, we identified the p.R388P LMNA mutation as responsible for congenital muscular dystrophy (L-CMD) and lipodystrophy. Here, we asked whether viral-mediated expression of mutant lamin A in murine skeletal muscles would be a pertinent model to reveal specific muscle alterations. We found that the total amount and size of muscle fibers as well as the extent of either inflammation or muscle regeneration were similar to wildtype or mutant lamin A. In contrast, the amount of fast oxidative muscle fibers containing myosin heavy chain IIA was lower upon expression of mutant lamin A, in correlation with lower expression of genes encoding transcription factors MEF2C and MyoD. These data validate this in vivo model for highlighting distinct muscle phenotypes associated with different lamin contexts. Additionally, the data suggest that alteration of muscle fiber type identity may contribute to the mechanisms underlying physiopathology of L-CMD related to R388P mutant lamin A. Full article
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Open AccessArticle
Microinjection of Antibodies Targeting the Lamin A/C Histone-Binding Site Blocks Mitotic Entry and Reveals Separate Chromatin Interactions with HP1, CenpB and PML
Cells 2017, 6(2), 9; doi:10.3390/cells6020009 -
Abstract
Lamins form a scaffold lining the nucleus that binds chromatin and contributes to spatial genome organization; however, due to the many other functions of lamins, studies knocking out or altering the lamin polymer cannot clearly distinguish between direct and indirect effects. To overcome
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Lamins form a scaffold lining the nucleus that binds chromatin and contributes to spatial genome organization; however, due to the many other functions of lamins, studies knocking out or altering the lamin polymer cannot clearly distinguish between direct and indirect effects. To overcome this obstacle, we specifically targeted the mapped histone-binding site of A/C lamins by microinjecting antibodies specific to this region predicting that this would make the genome more mobile. No increase in chromatin mobility was observed; however, interestingly, injected cells failed to go through mitosis, while control antibody-injected cells did. This effect was not due to crosslinking of the lamin polymer, as Fab fragments also blocked mitosis. The lack of genome mobility suggested other lamin-chromatin interactions. To determine what these might be, mini-lamin A constructs were expressed with or without the histone-binding site that assembled into independent intranuclear structures. HP1, CenpB and PML proteins accumulated at these structures for both constructs, indicating that other sites supporting chromatin interactions exist on lamin A. Together, these results indicate that lamin A-chromatin interactions are highly redundant and more diverse than generally acknowledged and highlight the importance of trying to experimentally separate their individual functions. Full article
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Open AccessArticle
KDM2 Family Members are Regulated by HIF-1 in Hypoxia
Cells 2017, 6(1), 8; doi:10.3390/cells6010008 -
Abstract
Hypoxia is not only a developmental cue but also a stress and pathological stimulus in many human diseases. The response to hypoxia at the cellular level relies on the activity of the transcription factor family, hypoxia inducible factor (HIF). HIF-1 is responsible for
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Hypoxia is not only a developmental cue but also a stress and pathological stimulus in many human diseases. The response to hypoxia at the cellular level relies on the activity of the transcription factor family, hypoxia inducible factor (HIF). HIF-1 is responsible for the acute response and transactivates a variety of genes involved in cellular metabolism, cell death, and cell growth. Here, we show that hypoxia results in increased mRNA levels for human lysine (K)-specific demethylase 2 (KDM2) family members, KDM2A and KDM2B, and also for Drosophila melanogaster KDM2, a histone and protein demethylase. In human cells, KDM2 family member’s mRNA levels are regulated by HIF-1 but not HIF-2 in hypoxia. Interestingly, only KDM2A protein levels are significantly induced in a HIF-1-dependent manner, while KDM2B protein changes in a cell type-dependent manner. Importantly, we demonstrate that in human cells, KDM2A regulation by hypoxia and HIF-1 occurs at the level of promoter, with HIF-1 binding to the KDM2A promoter being required for RNA polymerase II recruitment. Taken together, these results demonstrate that KDM2 is a novel HIF target that can help coordinate the cellular response to hypoxia. In addition, these results might explain why KDM2 levels are often deregulated in human cancers. Full article
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Open AccessReview
Application of Induced Pluripotent Stem Cell Technology to the Study of Hematological Diseases
Cells 2017, 6(1), 7; doi:10.3390/cells6010007 -
Abstract
The burst of reprogramming technology in recent years has revolutionized the field of stem cell biology, offering new opportunities for personalized, regenerative therapies. The direct reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) has provided an invaluable tool to study and
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The burst of reprogramming technology in recent years has revolutionized the field of stem cell biology, offering new opportunities for personalized, regenerative therapies. The direct reprogramming of somatic cells to induced pluripotent stem cells (iPSCs) has provided an invaluable tool to study and model a wide range of human diseases. Here, we review the transforming potential of such a strategy in research and in therapies applicable to the hematology field. Full article
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Open AccessReview
Invariant Chain Complexes and Clusters as Platforms for MIF Signaling
Cells 2017, 6(1), 6; doi:10.3390/cells6010006 -
Abstract
Invariant chain (Ii/CD74) has been identified as a surface receptor for migration inhibitory factor (MIF). Most cells that express Ii also synthesize major histocompatibility complex class II (MHC II) molecules, which depend on Ii as a chaperone and a targeting factor. The assembly
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Invariant chain (Ii/CD74) has been identified as a surface receptor for migration inhibitory factor (MIF). Most cells that express Ii also synthesize major histocompatibility complex class II (MHC II) molecules, which depend on Ii as a chaperone and a targeting factor. The assembly of nonameric complexes consisting of one Ii trimer and three MHC II molecules (each of which is a heterodimer) has been regarded as a prerequisite for efficient delivery to the cell surface. Due to rapid endocytosis, however, only low levels of Ii-MHC II complexes are displayed on the cell surface of professional antigen presenting cells and very little free Ii trimers. The association of Ii and MHC II has been reported to block the interaction with MIF, thus questioning the role of surface Ii as a receptor for MIF on MHC II-expressing cells. Recent work offers a potential solution to this conundrum: Many Ii-complexes at the cell surface appear to be under-saturated with MHC II, leaving unoccupied Ii subunits as potential binding sites for MIF. Some of this work also sheds light on novel aspects of signal transduction by Ii-bound MIF in B-lymphocytes: membrane raft association of Ii-MHC II complexes enables MIF to target Ii-MHC II to antigen-clustered B-cell-receptors (BCR) and to foster BCR-driven signaling and intracellular trafficking. Full article
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Open AccessReview
May I Cut in? Gene Editing Approaches in Human Induced Pluripotent Stem Cells
Cells 2017, 6(1), 5; doi:10.3390/cells6010005 -
Abstract
In the decade since Yamanaka and colleagues described methods to reprogram somatic cells into a pluripotent state, human induced pluripotent stem cells (hiPSCs) have demonstrated tremendous promise in numerous disease modeling, drug discovery, and regenerative medicine applications. More recently, the development and refinement
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In the decade since Yamanaka and colleagues described methods to reprogram somatic cells into a pluripotent state, human induced pluripotent stem cells (hiPSCs) have demonstrated tremendous promise in numerous disease modeling, drug discovery, and regenerative medicine applications. More recently, the development and refinement of advanced gene transduction and editing technologies have further accelerated the potential of hiPSCs. In this review, we discuss the various gene editing technologies that are being implemented with hiPSCs. Specifically, we describe the emergence of technologies including zinc-finger nuclease (ZFN), transcription activator-like effector nuclease (TALEN), and clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9 that can be used to edit the genome at precise locations, and discuss the strengths and weaknesses of each of these technologies. In addition, we present the current applications of these technologies in elucidating the mechanisms of human development and disease, developing novel and effective therapeutic molecules, and engineering cell-based therapies. Finally, we discuss the emerging technological advances in targeted gene editing methods. Full article
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Open AccessReview
Stem Cell Therapies in Retinal Disorders
Cells 2017, 6(1), 4; doi:10.3390/cells6010004 -
Abstract
Stem cell therapy has long been considered a promising mode of treatment for retinal conditions. While human embryonic stem cells (ESCs) have provided the precedent for regenerative medicine, the development of induced pluripotent stem cells (iPSCs) revolutionized this field. iPSCs allow for the
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Stem cell therapy has long been considered a promising mode of treatment for retinal conditions. While human embryonic stem cells (ESCs) have provided the precedent for regenerative medicine, the development of induced pluripotent stem cells (iPSCs) revolutionized this field. iPSCs allow for the development of many types of retinal cells, including those of the retinal pigment epithelium, photoreceptors, and ganglion cells, and can model polygenic diseases such as age-related macular degeneration. Cellular programming and reprogramming technology is especially useful in retinal diseases, as it allows for the study of living cells that have genetic variants that are specific to patients’ diseases. Since iPSCs are a self-renewing resource, scientists can experiment with an unlimited number of pluripotent cells to perfect the process of targeted differentiation, transplantation, and more, for personalized medicine. Challenges in the use of stem cells are present from the scientific, ethical, and political realms. These include transplant complications leading to anatomically incorrect placement, concern for tumorigenesis, and incomplete targeting of differentiation leading to contamination by different types of cells. Despite these limitations, human ESCs and iPSCs specific to individual patients can revolutionize the study of retinal disease and may be effective therapies for conditions currently considered incurable. Full article