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Sample records for adult human stem

  1. Expansion of Multipotent Stem Cells from the Adult Human Brain

    PubMed Central

    Murrell, Wayne; Palmero, Emily; Bianco, John; Stangeland, Biljana; Joel, Mrinal; Paulson, Linda; Thiede, Bernd; Grieg, Zanina; Ramsnes, Ingunn; Skjellegrind, Håvard K.; Nygård, Ståle; Brandal, Petter; Sandberg, Cecilie; Vik-Mo, Einar; Palmero, Sheryl; Langmoen, Iver A.

    2013-01-01

    The discovery of stem cells in the adult human brain has revealed new possible scenarios for treatment of the sick or injured brain. Both clinical use of and preclinical research on human adult neural stem cells have, however, been seriously hampered by the fact that it has been impossible to passage these cells more than a very few times and with little expansion of cell numbers. Having explored a number of alternative culturing conditions we here present an efficient method for the establishment and propagation of human brain stem cells from whatever brain tissue samples we have tried. We describe virtually unlimited expansion of an authentic stem cell phenotype. Pluripotency proteins Sox2 and Oct4 are expressed without artificial induction. For the first time multipotency of adult human brain-derived stem cells is demonstrated beyond tissue boundaries. We characterize these cells in detail in vitro including microarray and proteomic approaches. Whilst clarification of these cells’ behavior is ongoing, results so far portend well for the future repair of tissues by transplantation of an adult patient’s own-derived stem cells. PMID:23967194

  2. [Multipotency of adult stem cells derived from human amnion].

    PubMed

    Shi, Mingxia; Li, Weijia; Li, Bingzong; Li, Jing; Zhao, Chunhua

    2009-05-01

    Adult stem cells are drawing more and more attention due to the potential application in degenerative medicine without posing any moral problem. There is growing evidence showing that the human amnion contains various types of adult stem cell. Since amniotic tissue is readily available, it has the potential to be an important source of regenerative medicine material. In this study we tried to find multipotent adult stem cells in human amnion. We isolated stem cells from amniotic mesenchymal cells by limiting dilution assay. Similar to bone marrow derived mesenchymal stem cells, these cells displayed a fibroblast like appearance. They were positive for CD105, CD29, CD44, negative for haematopoietic (GlyA, CD31, CD34, CD45) and epithelial cell (pan-CK) markers. These stem cells had the potential to differentiate not only into osteogenic, adipogenic and endothelial lineages, but also hepatocyte-like cells and neural cells at the single-cell level depending on the culture conditions. They had the capacity for self-renewal and multilineage differentiation even after being expanded for more than 30 population doublings in vitro. So they may be an ideal stem cell source for inherited or degenerative diseases treatment.

  3. Neural stem cells in the adult human brain

    PubMed Central

    Gonzalez-Perez, Oscar

    2012-01-01

    For decades, it was believed that the adult brain was a quiescent organ unable to produce new neurons. At the beginning of the1960's, this dogma was challenged by a small group of neuroscientists. To date, it is well-known that new neurons are generated in the adult brain throughout life. Adult neurogenesis is primary confined to the subventricular zone (SVZ) of the forebrain and the subgranular zone of the dentate gyrus within the hippocampus. In both the human and the rodent brain, the primary progenitor of adult SVZ is a subpopulation of astrocytes that have stem-cell-like features. The human SVZ possesses a peculiar cell composition and displays important organizational differences when compared to the SVZ of other mammals. Some evidence suggests that the human SVZ may be not only an endogenous source of neural precursor cells for brain repair, but also a source of brain tumors. In this review, we described the cytoarchitecture and cellular composition of the SVZ in the adult human brain. We also discussed some clinical implications of SVZ, such as: stem-cell-based therapies against neurodegenerative diseases and its potential as a source of malignant cells. Understanding the biology of human SVZ and its neural progenitors is one of the crucial steps to develop novel therapies against neurological diseases in humans. PMID:23181200

  4. Brain stem auditory evoked responses in human infants and adults

    NASA Technical Reports Server (NTRS)

    Hecox, K.; Galambos, R.

    1974-01-01

    Brain stem evoked potentials were recorded by conventional scalp electrodes in infants (3 weeks to 3 years of age) and adults. The latency of one of the major response components (wave V) is shown to be a function both of click intensity and the age of the subject; this latency at a given signal strength shortens postnatally to reach the adult value (about 6 msec) by 12 to 18 months of age. The demonstrated reliability and limited variability of these brain stem electrophysiological responses provide the basis for an optimistic estimate of their usefulness as an objective method for assessing hearing in infants and adults.

  5. Transcriptional profiling of adult neural stem-like cells from the human brain.

    PubMed

    Sandberg, Cecilie Jonsgar; Vik-Mo, Einar O; Behnan, Jinan; Helseth, Eirik; Langmoen, Iver A

    2014-01-01

    There is a great potential for the development of new cell replacement strategies based on adult human neural stem-like cells. However, little is known about the hierarchy of cells and the unique molecular properties of stem- and progenitor cells of the nervous system. Stem cells from the adult human brain can be propagated and expanded in vitro as free floating neurospheres that are capable of self-renewal and differentiation into all three cell types of the central nervous system. Here we report the first global gene expression study of adult human neural stem-like cells originating from five human subventricular zone biopsies (mean age 42, range 33-60). Compared to adult human brain tissue, we identified 1,189 genes that were significantly up- and down-regulated in adult human neural stem-like cells (1% false discovery rate). We found that adult human neural stem-like cells express stem cell markers and have reduced levels of markers that are typical of the mature cells in the nervous system. We report that the genes being highly expressed in adult human neural stem-like cells are associated with developmental processes and the extracellular region of the cell. The calcium signaling pathway and neuroactive ligand-receptor interactions are enriched among the most differentially regulated genes between adult human neural stem-like cells and adult human brain tissue. We confirmed the expression of 10 of the most up-regulated genes in adult human neural stem-like cells in an additional sample set that included adult human neural stem-like cells (n = 6), foetal human neural stem cells (n = 1) and human brain tissues (n = 12). The NGFR, SLITRK6 and KCNS3 receptors were further investigated by immunofluorescence and shown to be heterogeneously expressed in spheres. These receptors could potentially serve as new markers for the identification and characterisation of neural stem- and progenitor cells or as targets for manipulation of cellular fate.

  6. Transcriptional Profiling of Adult Neural Stem-Like Cells from the Human Brain

    PubMed Central

    Sandberg, Cecilie Jonsgar; Vik-Mo, Einar O.; Behnan, Jinan; Helseth, Eirik; Langmoen, Iver A.

    2014-01-01

    There is a great potential for the development of new cell replacement strategies based on adult human neural stem-like cells. However, little is known about the hierarchy of cells and the unique molecular properties of stem- and progenitor cells of the nervous system. Stem cells from the adult human brain can be propagated and expanded in vitro as free floating neurospheres that are capable of self-renewal and differentiation into all three cell types of the central nervous system. Here we report the first global gene expression study of adult human neural stem-like cells originating from five human subventricular zone biopsies (mean age 42, range 33–60). Compared to adult human brain tissue, we identified 1,189 genes that were significantly up- and down-regulated in adult human neural stem-like cells (1% false discovery rate). We found that adult human neural stem-like cells express stem cell markers and have reduced levels of markers that are typical of the mature cells in the nervous system. We report that the genes being highly expressed in adult human neural stem-like cells are associated with developmental processes and the extracellular region of the cell. The calcium signaling pathway and neuroactive ligand-receptor interactions are enriched among the most differentially regulated genes between adult human neural stem-like cells and adult human brain tissue. We confirmed the expression of 10 of the most up-regulated genes in adult human neural stem-like cells in an additional sample set that included adult human neural stem-like cells (n = 6), foetal human neural stem cells (n = 1) and human brain tissues (n = 12). The NGFR, SLITRK6 and KCNS3 receptors were further investigated by immunofluorescence and shown to be heterogeneously expressed in spheres. These receptors could potentially serve as new markers for the identification and characterisation of neural stem- and progenitor cells or as targets for manipulation of cellular fate. PMID

  7. Adult somatic stem cells in the human parasite, Schistosoma mansoni

    PubMed Central

    Collins, James J.; Wang, Bo; Lambrus, Bramwell G.; Tharp, Marla; Iyer, Harini; Newmark, Phillip A.

    2013-01-01

    Summary Schistosomiasis is among the most prevalent human parasitic diseases, affecting more than 200 million people worldwide1. The etiological agents of this disease are trematode flatworms (Schistosoma) that live and lay eggs within the vasculature of the host. These eggs lodge in host tissues, causing inflammatory responses that are the primary cause of morbidity. Because these parasites can live and reproduce within human hosts for decades2, elucidating the mechanisms that promote their longevity is of fundamental importance. Although adult pluripotent stem cells, called neoblasts, drive long-term homeostatic tissue maintenance in long-lived free-living flatworms3,4 (e.g., planarians), and neoblast-like cells have been described in some parasitic tapeworms5, little is known about whether similar cell types exist in any trematode species. Here, we describe a population of neoblast-like cells in the trematode Schistosoma mansoni. These cells resemble planarian neoblasts morphologically and share their ability to proliferate and differentiate into derivatives of multiple germ layers. Capitalizing on available genomic resources6,7 and RNAseq-based gene expression profiling, we find that these schistosome neoblast-like cells express a fibroblast growth factor receptor ortholog. Using RNA interference we demonstrate that this gene is required for the maintenance of these neoblast-like cells. Our observations suggest that adaptation of developmental strategies shared by free-living ancestors to modern-day schistosomes likely contributed to the success of these animals as long-lived obligate parasites. We expect that future studies deciphering the function of these neoblast-like cells will have important implications for understanding the biology of these devastating parasites. PMID:23426263

  8. Human oocytes reprogram adult somatic nuclei of a type 1 diabetic to diploid pluripotent stem cells.

    PubMed

    Yamada, Mitsutoshi; Johannesson, Bjarki; Sagi, Ido; Burnett, Lisa Cole; Kort, Daniel H; Prosser, Robert W; Paull, Daniel; Nestor, Michael W; Freeby, Matthew; Greenberg, Ellen; Goland, Robin S; Leibel, Rudolph L; Solomon, Susan L; Benvenisty, Nissim; Sauer, Mark V; Egli, Dieter

    2014-06-26

    The transfer of somatic cell nuclei into oocytes can give rise to pluripotent stem cells that are consistently equivalent to embryonic stem cells, holding promise for autologous cell replacement therapy. Although methods to induce pluripotent stem cells from somatic cells by transcription factors are widely used in basic research, numerous differences between induced pluripotent stem cells and embryonic stem cells have been reported, potentially affecting their clinical use. Because of the therapeutic potential of diploid embryonic stem-cell lines derived from adult cells of diseased human subjects, we have systematically investigated the parameters affecting efficiency of blastocyst development and stem-cell derivation. Here we show that improvements to the oocyte activation protocol, including the use of both kinase and translation inhibitors, and cell culture in the presence of histone deacetylase inhibitors, promote development to the blastocyst stage. Developmental efficiency varied between oocyte donors, and was inversely related to the number of days of hormonal stimulation required for oocyte maturation, whereas the daily dose of gonadotropin or the total number of metaphase II oocytes retrieved did not affect developmental outcome. Because the use of concentrated Sendai virus for cell fusion induced an increase in intracellular calcium concentration, causing premature oocyte activation, we used diluted Sendai virus in calcium-free medium. Using this modified nuclear transfer protocol, we derived diploid pluripotent stem-cell lines from somatic cells of a newborn and, for the first time, an adult, a female with type 1 diabetes.

  9. Adult stem cells: simply a tool for regenerative medicine or an additional piece in the puzzle of human aging?

    PubMed

    Tollervey, James R; Lunyak, Victoria V

    2011-12-15

    Adult stem cells have taken center stage in current research related to regenerative medicine and pharmacogenomic studies seeking new therapeutic interventions. As we learn more about these cells, it is becoming apparent that the next big leap in our understanding of adult stem cell biology and adult stem cell aging will depend on the integration of approaches from various disciplines. Major advances and technological breakthroughs at the crossroad of fields such as biomaterials, genomics, epigenomics, and proteomics will enable the design of better tools to model human diseases, and warrant safe usage of adult stem cells in the clinic.

  10. Human germ cell differentiation from fetal- and adult-derived induced pluripotent stem cells

    PubMed Central

    Panula, Sarita; Medrano, Jose V.; Kee, Kehkooi; Bergström, Rosita; Nguyen, Ha Nam; Byers, Blake; Wilson, Kitchener D.; Wu, Joseph C.; Simon, Carlos; Hovatta, Outi; Reijo Pera, Renee A.

    2011-01-01

    Historically, our understanding of molecular genetic aspects of human germ cell development has been limited, at least in part due to inaccessibility of early stages of human development to experimentation. However, the derivation of pluripotent stem cells may provide the necessary human genetic system to study germ cell development. In this study, we compared the potential of human induced pluripotent stem cells (iPSCs), derived from adult and fetal somatic cells to form primordial and meiotic germ cells, relative to human embryonic stem cells. We found that ∼5% of human iPSCs differentiated to primordial germ cells (PGCs) following induction with bone morphogenetic proteins. Furthermore, we observed that PGCs expressed green fluorescent protein from a germ cell-specific reporter and were enriched for the expression of endogenous germ cell-specific proteins and mRNAs. In response to the overexpression of intrinsic regulators, we also observed that iPSCs formed meiotic cells with extensive synaptonemal complexes and post-meiotic haploid cells with a similar pattern of ACROSIN staining as observed in human spermatids. These results indicate that human iPSCs derived from reprogramming of adult somatic cells can form germline cells. This system may provide a useful model for molecular genetic studies of human germline formation and pathology and a novel platform for clinical studies and potential therapeutical applications. PMID:21131292

  11. Germline stem cells and neo-oogenesis in the adult human ovary.

    PubMed

    Liu, Yifei; Wu, Chao; Lyu, Qifeng; Yang, Dongzi; Albertini, David F; Keefe, David L; Liu, Lin

    2007-06-01

    It remains unclear whether neo-oogenesis occurs in postnatal ovaries of mammals, based on studies in mice. We thought to test whether adult human ovaries contain germline stem cells (GSCs) and undergo neo-oogenesis. Rather than using genetic manipulation which is unethical in humans, we took the approach of analyzing the expression of meiotic marker genes and genes for germ cell proliferation, which are required for neo-oogenesis, in adult human ovaries covering an age range from 28 to 53 years old, compared to testis and fetal ovaries served as positive controls. We show that active meiosis, neo-oogenesis and GSCs are unlikely to exist in normal, adult, human ovaries. No early meiotic-specific or oogenesis-associated mRNAs for SPO11, PRDM9, SCP1, TERT and NOBOX were detectable in adult human ovaries using RT-PCR, compared to fetal ovary and adult testis controls. These findings are further corroborated by the absence of early meiocytes and proliferating germ cells in adult human ovarian cortex probed with markers for meiosis (SCP3), oogonium (OCT3/4, c-KIT), and cell cycle progression (Ki-67, PCNA), in contrast to fetal ovary controls. If postnatal oogenesis is confirmed in mice, then this species would represent an exception to the rule that neo-oogenesis does not occur in adults.

  12. A mystery unraveled: nontumorigenic pluripotent stem cells in human adult tissues

    PubMed Central

    Simerman, Ariel A; Perone, Marcelo J; Gimeno, María L; Dumesic, Daniel A; Chazenbalk, Gregorio D

    2014-01-01

    Introduction: Embryonic stem cells and induced pluripotent stem cells have emerged as the gold standard of pluripotent stem cells and the class of stem cell with the highest potential for contribution to regenerative and therapeutic application; however, their translational use is often impeded by teratoma formation, commonly associated with pluripotency. We discuss a population of nontumorigenic pluripotent stem cells, termed Multilineage Differentiating Stress Enduring (Muse) cells, which offer an innovative and exciting avenue of exploration for the potential treatment of various human diseases. Areas covered: This review discusses the origin of Muse cells, describes in detail their various unique characteristics, and considers future avenues of their application and investigation with respect to what is currently known of adult pluripotent stem cells in scientific literature. We begin by defining cell potency, then discuss both mesenchymal and various reported populations of pluripotent stem cells, and finally delve into Muse cells and the characteristics that set them apart from their contemporaries. Expert opinion: Muse cells derived from adipose tissue (Muse-AT) are efficiently, routinely and painlessly isolated from human lipoaspirate material, exhibit tripoblastic differentiation both spontaneously and under media-specific induction, and do not form teratomas. We describe qualities specific to Muse-AT cells and their potential impact on the field of regenerative medicine and cell therapy. PMID:24745973

  13. Adult Human Nasal Mesenchymal-Like Stem Cells Restore Cochlear Spiral Ganglion Neurons After Experimental Lesion

    PubMed Central

    Bas, Esperanza; Van De Water, Thomas R.; Lumbreras, Vicente; Rajguru, Suhrud; Goss, Garrett; Hare, Joshua M.

    2014-01-01

    A loss of sensory hair cells or spiral ganglion neurons from the inner ear causes deafness, affecting millions of people. Currently, there is no effective therapy to repair the inner ear sensory structures in humans. Cochlear implantation can restore input, but only if auditory neurons remain intact. Efforts to develop stem cell-based treatments for deafness have demonstrated progress, most notably utilizing embryonic-derived cells. In an effort to bypass limitations of embryonic or induced pluripotent stem cells that may impede the translation to clinical applications, we sought to utilize an alternative cell source. Here, we show that adult human mesenchymal-like stem cells (MSCs) obtained from nasal tissue can repair spiral ganglion loss in experimentally lesioned cochlear cultures from neonatal rats. Stem cells engraft into gentamicin-lesioned organotypic cultures and orchestrate the restoration of the spiral ganglion neuronal population, involving both direct neuronal differentiation and secondary effects on endogenous cells. As a physiologic assay, nasal MSC-derived cells engrafted into lesioned spiral ganglia demonstrate responses to infrared laser stimulus that are consistent with those typical of excitable cells. The addition of a pharmacologic activator of the canonical Wnt/β-catenin pathway concurrent with stem cell treatment promoted robust neuronal differentiation. The availability of an effective adult autologous cell source for inner ear tissue repair should contribute to efforts to translate cell-based strategies to the clinic. PMID:24172073

  14. Alternative Sources of Adult Stem Cells: Human Amniotic Membrane

    NASA Astrophysics Data System (ADS)

    Wolbank, Susanne; van Griensven, Martijn; Grillari-Voglauer, Regina; Peterbauer-Scherb, Anja

    Human amniotic membrane is a highly promising cell source for tissue engineering. The cells thereof, human amniotic epithelial cells (hAEC) and human amniotic mesenchymal stromal cells (hAMSC), may be immunoprivileged, they represent an early developmental status, and their application is ethically uncontroversial. Cell banking strategies may use freshly isolated cells or involve in vitro expansion to increase cell numbers. Therefore, we have thoroughly characterized the effect of in vitro cultivation on both phenotype and differentiation potential of hAEC. Moreover, we present different strategies to improve expansion including replacement of animal-derived supplements by human platelet products or the introduction of the catalytic subunit of human telomerase to extend the in vitro lifespan of amniotic cells. Characterization of the resulting cultures includes phenotype, growth characteristics, and differentiation potential, as well as immunogenic and immunomodulatory properties.

  15. Micropatterning control of tubular commitment in human adult renal stem cells.

    PubMed

    Sciancalepore, Anna G; Portone, Alberto; Moffa, Maria; Persano, Luana; De Luca, Maria; Paiano, Aurora; Sallustio, Fabio; Schena, Francesco P; Bucci, Cecilia; Pisignano, Dario

    2016-07-01

    The treatment of renal injury by autologous, patient-specific adult stem cells is still an unmet need. Unsolved issues remain the spatial integration of stem cells into damaged areas of the organ, the commitment in the required cell type and the development of improved bioengineered devices. In this respect, biomaterials and architectures have to be specialized to control stem cell differentiation. Here, we perform an extensive study on micropatterned extracellular matrix proteins, which constitute a simple and non-invasive approach to drive the differentiation of adult renal progenitor/stem cells (ARPCs) from human donors. ARPCs are interfaced with fibronectin (FN) micropatterns, in the absence of exogenous chemicals or cellular reprogramming. We obtain the differentiation towards tubular cells of ARPCs cultured in basal medium conditions, the tubular commitment thus being specifically induced by micropatterned substrates. We characterize the stability of the tubular differentiation as well as the induction of a polarized phenotype in micropatterned ARPCs. Thus, the developed cues, driving the functional commitment of ARPCs, offer a route to recreate the microenvironment of the stem cell niche in vitro, that may serve, in perspective, for the development of ARPC-based bioengineered devices. PMID:27105437

  16. Apple ethanol extract promotes proliferation of human adult stem cells, which involves the regenerative potential of stem cells.

    PubMed

    Lee, Jienny; Shin, Moon Sam; Kim, Mi Ok; Jang, Sunghee; Oh, Sae Woong; Kang, Mingyeong; Jung, Kwangseon; Park, Yong Seek; Lee, Jongsung

    2016-09-01

    Tissue regeneration using adult stem cells (ASCs) has significant potential as a novel treatment for many degenerative diseases. Previous studies have established that age negatively affects the proliferation status and differentiation potential of ASCs, suggesting a possible limitation in their potential therapeutic use. Therefore, we hypothesized that apple extract might exert beneficial effects on ASCs. The specific objectives were to investigate the proliferative effect of apple ethanol extract on human adipose tissue-derived mesenchymal stem cells (ADSCs) and human cord blood-derived mesenchymal stem cells (CB-MSCs), and identify the possible molecular mechanisms. Apple extract promoted proliferation of ADSCs and CB-MSCs as determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and Click-iT 5-ethynyl-2'-deoxyuridine flow cytometry assays. In addition, phosphorylation of p44/42 MAPK (ERK), mammalian target of rapamycin (mTOR), p70 S6 kinase (p70S6K), S6 ribosomal protein (S6RP), eukaryotic initiation factor (eIF) 4B and eIF4E was induced stepwise in ADSCs. Furthermore, apple extract significantly induced the production of vascular endothelial growth factor and interleukin-6 in both ADSCs and CB-MSCs. Similarly, apple extract-induced phosphorylation of the mTOR/p70S6K/S6RP/eIF4B/eIF4E pathway was blocked by pretreatment with PD98059, a specific ERK inhibitor. These results indicate that apple extract-induced proliferation of ADSCs under serum-free conditions is mediated by ERK-dependent cytokine production. Moreover, the beneficial effect of apple extract on proliferation of ASCs may overcome the limitation in therapeutic use of stem cells in tissue regeneration and maintenance of stem cell homeostasis. PMID:27632912

  17. Isolation and Characterization of Human Adult Epithelial Stem Cells from the Periodontal Ligament.

    PubMed

    Athanassiou-Papaefthymiou, M; Papagerakis, P; Papagerakis, S

    2015-11-01

    We report a novel method for the isolation of adult human epithelial stem cells (hEpiSCs) from the epithelial component of the periodontal ligament-the human epithelial cell rests of Malassez (hERM). hEpiSC-rich integrin-α6(+ve) hERM cells derived by fluorometry can be clonally expanded, can grow organoids, and express the markers of pluripotency (OCT4, NANOG, SOX2), polycomb protein RING1B, and the hEpiSC supermarker LGR5. They maintain the growth profile of their originating hERM in vitro. Subcutaneous cotransplantation with mesenchymal stem cells from the dental pulp on poly-l-lactic acid scaffolds in nude mice gave rise to perfect heterotopic ossicles in vivo with ultrastructure of dentin, enamel, cementum, and bone. These remarkable fully mineralized ossicles underscore the importance of epithelial-mesenchymal crosstalk in tissue regeneration using human progenitor stem cells, which may have already committed to lineage despite maintaining hallmarks of pluripotency. In addition, we report the clonal expansion and isolation of human LGR5(+ve) cells from the hERM in xeno-free culture conditions. The genetic profile of LGR5(+ve) cells includes both markers of pluripotency and genes important for secretory epithelial and dental epithelial cell differentiation, giving us a first insight into periodontal ligament-derived hEpiSCs. PMID:26392003

  18. Long-term culture of genome-stable bipotent stem cells from adult human liver.

    PubMed

    Huch, Meritxell; Gehart, Helmuth; van Boxtel, Ruben; Hamer, Karien; Blokzijl, Francis; Verstegen, Monique M A; Ellis, Ewa; van Wenum, Martien; Fuchs, Sabine A; de Ligt, Joep; van de Wetering, Marc; Sasaki, Nobuo; Boers, Susanne J; Kemperman, Hans; de Jonge, Jeroen; Ijzermans, Jan N M; Nieuwenhuis, Edward E S; Hoekstra, Ruurdtje; Strom, Stephen; Vries, Robert R G; van der Laan, Luc J W; Cuppen, Edwin; Clevers, Hans

    2015-01-15

    Despite the enormous replication potential of the human liver, there are currently no culture systems available that sustain hepatocyte replication and/or function in vitro. We have shown previously that single mouse Lgr5+ liver stem cells can be expanded as epithelial organoids in vitro and can be differentiated into functional hepatocytes in vitro and in vivo. We now describe conditions allowing long-term expansion of adult bile duct-derived bipotent progenitor cells from human liver. The expanded cells are highly stable at the chromosome and structural level, while single base changes occur at very low rates. The cells can readily be converted into functional hepatocytes in vitro and upon transplantation in vivo. Organoids from α1-antitrypsin deficiency and Alagille syndrome patients mirror the in vivo pathology. Clonal long-term expansion of primary adult liver stem cells opens up experimental avenues for disease modeling, toxicology studies, regenerative medicine, and gene therapy. PMID:25533785

  19. Long-Term Culture of Genome-Stable Bipotent Stem Cells from Adult Human Liver

    PubMed Central

    Huch, Meritxell; Gehart, Helmuth; van Boxtel, Ruben; Hamer, Karien; Blokzijl, Francis; Verstegen, Monique M.A.; Ellis, Ewa; van Wenum, Martien; Fuchs, Sabine A.; de Ligt, Joep; van de Wetering, Marc; Sasaki, Nobuo; Boers, Susanne J.; Kemperman, Hans; de Jonge, Jeroen; Ijzermans, Jan N.M.; Nieuwenhuis, Edward E.S.; Hoekstra, Ruurdtje; Strom, Stephen; Vries, Robert R.G.; van der Laan, Luc J.W.; Cuppen, Edwin; Clevers, Hans

    2015-01-01

    Summary Despite the enormous replication potential of the human liver, there are currently no culture systems available that sustain hepatocyte replication and/or function in vitro. We have shown previously that single mouse Lgr5+ liver stem cells can be expanded as epithelial organoids in vitro and can be differentiated into functional hepatocytes in vitro and in vivo. We now describe conditions allowing long-term expansion of adult bile duct-derived bipotent progenitor cells from human liver. The expanded cells are highly stable at the chromosome and structural level, while single base changes occur at very low rates. The cells can readily be converted into functional hepatocytes in vitro and upon transplantation in vivo. Organoids from α1-antitrypsin deficiency and Alagille syndrome patients mirror the in vivo pathology. Clonal long-term expansion of primary adult liver stem cells opens up experimental avenues for disease modeling, toxicology studies, regenerative medicine, and gene therapy. PMID:25533785

  20. Human adult stem cells as the target cells for the initiation of carcinogenesis and for the generation of "cancer stem cells".

    PubMed

    Trosko, James E

    2008-11-01

    The inference to stem cells has been found in ancient myths and the concept of stem cells has existed in the fields of plant biology, developmental biology and embryology for decades. In the field of cancer research, the stem cell theory was one of the earliest hypotheses on the origin of a cancer from a single cell. However, an opposing hypothesis had it that an adult differentiated somatic cell could "de-differentiate" to become a cancer cell. Only within the last decade, via the "cloning" of Dolly, the sheep, did the field of stem cell biology really trigger an exciting revolution in biological research. The isolation of human embryonic stem cells has created a true revolution in the life sciences that has led to the hope that these human stem cells could lead to (a) basic science understanding of gene regulation during differentiation and development; (b) stem cell therapy; (c) gene therapy via stem cells; (d) the use of stem cells for drug discovery; (e) screening for toxic effects of chemicals; and (f) understand the aging and diseases of aging processes. PMID:24855504

  1. Induction of Stem Cell Gene Expression in Adult Human Fibroblasts without Transgenes

    PubMed Central

    Ambady, Sakthikumar; Holmes, William F.; Vilner, Lucy; Kole, Denis; Kashpur, Olga; Huntress, Victoria; Vojtic, Ina; Whitton, Holly; Dominko, Tanja

    2009-01-01

    Abstract Reprogramming of differentiated somatic cells into induced pluripotent stem (iPS) cells has potential for derivation of patient-specific cells for therapy as well as for development of models with which to study disease progression. Derivation of iPS cells from human somatic cells has been achieved by viral transduction of human fibroblasts with early developmental genes. Because forced expression of these genes by viral transduction results in transgene integration with unknown and unpredictable potential mutagenic effects, identification of cell culture conditions that can induce endogenous expression of these genes is desirable. Here we show that primary adult human fibroblasts have basal expression of mRNA for OCT4, SOX2, and NANOG. However, translation of these messages into detectable proteins and their subcellular localization depends on cell culture conditions. Manipulation of oxygen concentration and FGF2 supplementation can modulate expression of some pluripotency related genes at the transcriptional, translational, and cellular localization level. Changing cell culture condition parameters led to expression of REX1, potentiation of expression of LIN28, translation of OCT4, SOX2, and NANOG, and translocation of these transcription factors to the cell nucleus. We also show that culture conditions affect the in vitro lifespan of dermal fibroblasts, nearly doubling the number of population doublings before the cells reach replicative senescence. Our results suggest that it is possible to induce and manipulate endogenous expression of stem cell genes in somatic cells without genetic manipulation, but this short-term induction may not be sufficient for acquisition of true pluripotency. Further investigation of the factors involved in inducing this response could lead to discovery of defined culture conditions capable of altering cell fate in vitro. This would alleviate the need for forced expression by transgenesis, thus eliminating the risk of

  2. Clonal analysis of the differentiation potential of human adipose-derived adult stem cells.

    PubMed

    Guilak, Farshid; Lott, Kristen E; Awad, Hani A; Cao, Qiongfang; Hicok, Kevin C; Fermor, Beverley; Gimble, Jeffrey M

    2006-01-01

    Pools of human adipose-derived adult stem (hADAS) cells can exhibit multiple differentiated phenotypes under appropriate in vitro culture conditions. Because adipose tissue is abundant and easily accessible, hADAS cells offer a promising source of cells for tissue engineering and other cell-based therapies. However, it is unclear whether individual hADAS cells can give rise to multiple differentiated phenotypes or whether each phenotype arises from a subset of committed progenitor cells that exists within a heterogeneous population. The goal of this study was to test the hypothesis that single hADAS are multipotent at a clonal level. hADAS cells were isolated from liposuction waste, and ring cloning was performed to select cells derived from a single progenitor cell. Forty-five clones were expanded through four passages and then induced for adipogenesis, osteogenesis, chondrogenesis, and neurogenesis using lineage-specific differentiation media. Quantitative differentiation criteria for each lineage were determined using histological and biochemical analyses. Eighty one percent of the hADAS cell clones differentiated into at least one of the lineages. In addition, 52% of the hADAS cell clones differentiated into two or more of the lineages. More clones expressed phenotypes of osteoblasts (48%), chondrocytes (43%), and neuron-like cells (52%) than of adipocytes (12%), possibly due to the loss of adipogenic ability after repeated subcultures. The findings are consistent with the hypothesis that hADAS cells are a type of multipotent adult stem cell and not solely a mixed population of unipotent progenitor cells. However, it is important to exercise caution in interpreting these results until they are validated using functional in vivo assays.

  3. In vitro generation of pancreatic endocrine cells from human adult fibroblast-like limbal stem cells.

    PubMed

    Criscimanna, Angela; Zito, Giovanni; Taddeo, Annalisa; Richiusa, Pierina; Pitrone, Maria; Morreale, Daniele; Lodato, Gaetano; Pizzolanti, Giuseppe; Citarrella, Roberto; Galluzzo, Aldo; Giordano, Carla

    2012-01-01

    Stem cells might provide unlimited supply of transplantable cells for β-cell replacement therapy in diabetes. The human limbus is a highly specialized region hosting a well-recognized population of epithelial stem cells, which sustain the continuous renewal of the cornea, and the recently identified stromal fibroblast-like stem cells (f-LSCs), with apparent broader plasticity. However, the lack of specific molecular markers for the identification of the multipotent limbal subpopulation has so far limited the investigation of their differentiation potential. In this study we show that the human limbus contains uncommitted cells that could be potentially harnessed for the treatment of diabetes. Fourteen limbal biopsies were obtained from patients undergoing surgery for ocular diseases not involving the conjunctiva or corneal surface. We identified a subpopulation of f-LSCs characterized by robust proliferative capacity, expressing several pluripotent stem cell markers and exhibiting self-renewal ability. We then demonstrated the potential of f-LSCs to differentiate in vitro into functional insulin-secreting cells by developing a four-step differentiation protocol that efficiently directed f-LSCs towards the pancreatic endocrine cell fate. The expression of specific endodermal, pancreatic, islet, and β-cell markers, as well as functional properties of f-LSC-derived insulin-producing cells, were evaluated during differentiation. With our stage-specific approach, up to 77% of f-LSCs eventually differentiated into cells expressing insulin (also assessed as C-peptide) and exhibited phenotypic features of mature β-cells, such as expression of critical transcription factors and presence of secretory granules. Although insulin content was about 160-fold lower than what observed in adult islets, differentiated cells processed ∼98% of their proinsulin content, similar to mature β-cells. Moreover, they responded in vitro in a regulated manner to multiple secretory stimuli

  4. Human Adult Dental Pulp Stem Cells Enhance Poststroke Functional Recovery Through Non-Neural Replacement Mechanisms

    PubMed Central

    Leong, Wai Khay; Henshall, Tanya L.; Arthur, Agnes; Kremer, Karlea L.; Lewis, Martin D.; Helps, Stephen C.; Field, John; Hamilton-Bruce, Monica A.; Warming, Scott; Manavis, Jim; Vink, Robert; Gronthos, Stan

    2012-01-01

    Human adult dental pulp stem cells (DPSCs), derived from third molar teeth, are multipotent and have the capacity to differentiate into neurons under inductive conditions both in vitro and following transplantation into the avian embryo. In this study, we demonstrate that the intracerebral transplantation of human DPSCs 24 hours following focal cerebral ischemia in a rodent model resulted in significant improvement in forelimb sensorimotor function at 4 weeks post-treatment. At this time, 2.3 ± 0.7% of engrafted cells had survived in the poststroke brain and demonstrated targeted migration toward the stroke lesion. In the peri-infarct striatum, transplanted DPSCs differentiated into astrocytes in preference to neurons. Our data suggest that the dominant mechanism of action underlying DPSC treatment that resulted in enhanced functional recovery is unlikely to be due to neural replacement. Functional improvement is more likely to be mediated through DPSC-dependent paracrine effects. This study provides preclinical evidence for the future use of human DPSCs in cell therapy to improve outcome in stroke patients. PMID:23197777

  5. Pluripotency of adult stem cells derived from human and rat pancreas

    NASA Astrophysics Data System (ADS)

    Kruse, C.; Birth, M.; Rohwedel, J.; Assmuth, K.; Goepel, A.; Wedel, T.

    Adult stem cells are undifferentiated cells found within fully developed tissues or organs of an adult individuum. Until recently, these cells have been considered to bear less self-renewal ability and differentiation potency compared to embryonic stem cells. In recent studies an undifferentiated cell type was found in primary cultures of isolated acini from exocrine pancreas termed pancreatic stellate cells. Here we show that pancreatic stellate-like cells have the capacity of extended self-renewal and are able to differentiate spontaneously into cell types of all three germ layers expressing markers for smooth muscle cells, neurons, glial cells, epithelial cells, chondrocytes and secretory cells (insulin, amylase). Differentiation and subsequent formation of three-dimensional cellular aggregates (organoid bodies) were induced by merely culturing pancreatic stellate-like cells in hanging drops. These cells were developed into stable, long-term, in vitro cultures of both primary undifferentiated cell lines as well as organoid cultures. Thus, evidence is given that cell lineages of endodermal, mesodermal, and ectodermal origin arise spontaneously from a single adult undifferentiated cell type. Based on the present findings it is assumed that pancreatic stellate-like cells are a new class of lineage uncommitted pluripotent adult stem cells with a remarkable self-renewal ability and differentiation potency. The data emphasize the versatility of adult stem cells and may lead to a reappraisal of their use for the treatment of inherited disorders or acquired degenerative diseases.

  6. Tissue-specific mutation accumulation in human adult stem cells during life

    NASA Astrophysics Data System (ADS)

    Blokzijl, Francis; de Ligt, Joep; Jager, Myrthe; Sasselli, Valentina; Roerink, Sophie; Sasaki, Nobuo; Huch, Meritxell; Boymans, Sander; Kuijk, Ewart; Prins, Pjotr; Nijman, Isaac J.; Martincorena, Inigo; Mokry, Michal; Wiegerinck, Caroline L.; Middendorp, Sabine; Sato, Toshiro; Schwank, Gerald; Nieuwenhuis, Edward E. S.; Verstegen, Monique M. A.; van der Laan, Luc J. W.; de Jonge, Jeroen; Ijzermans, Jan N. M.; Vries, Robert G.; van de Wetering, Marc; Stratton, Michael R.; Clevers, Hans; Cuppen, Edwin; van Boxtel, Ruben

    2016-10-01

    The gradual accumulation of genetic mutations in human adult stem cells (ASCs) during life is associated with various age-related diseases, including cancer. Extreme variation in cancer risk across tissues was recently proposed to depend on the lifetime number of ASC divisions, owing to unavoidable random mutations that arise during DNA replication. However, the rates and patterns of mutations in normal ASCs remain unknown. Here we determine genome-wide mutation patterns in ASCs of the small intestine, colon and liver of human donors with ages ranging from 3 to 87 years by sequencing clonal organoid cultures derived from primary multipotent cells. Our results show that mutations accumulate steadily over time in all of the assessed tissue types, at a rate of approximately 40 novel mutations per year, despite the large variation in cancer incidence among these tissues. Liver ASCs, however, have different mutation spectra compared to those of the colon and small intestine. Mutational signature analysis reveals that this difference can be attributed to spontaneous deamination of methylated cytosine residues in the colon and small intestine, probably reflecting their high ASC division rate. In liver, a signature with an as-yet-unknown underlying mechanism is predominant. Mutation spectra of driver genes in cancer show high similarity to the tissue-specific ASC mutation spectra, suggesting that intrinsic mutational processes in ASCs can initiate tumorigenesis. Notably, the inter-individual variation in mutation rate and spectra are low, suggesting tissue-specific activity of common mutational processes throughout life.

  7. Adult human neural stem cell therapeutics: Current developmental status and prospect

    PubMed Central

    Nam, Hyun; Lee, Kee-Hang; Nam, Do-Hyun; Joo, Kyeung Min

    2015-01-01

    Over the past two decades, regenerative therapies using stem cell technologies have been developed for various neurological diseases. Although stem cell therapy is an attractive option to reverse neural tissue damage and to recover neurological deficits, it is still under development so as not to show significant treatment effects in clinical settings. In this review, we discuss the scientific and clinical basics of adult neural stem cells (aNSCs), and their current developmental status as cell therapeutics for neurological disease. Compared with other types of stem cells, aNSCs have clinical advantages, such as limited proliferation, inborn differentiation potential into functional neural cells, and no ethical issues. In spite of the merits of aNSCs, difficulties in the isolation from the normal brain, and in the in vitro expansion, have blocked preclinical and clinical study using aNSCs. However, several groups have recently developed novel techniques to isolate and expand aNSCs from normal adult brains, and showed successful applications of aNSCs to neurological diseases. With new technologies for aNSCs and their clinical strengths, previous hurdles in stem cell therapies for neurological diseases could be overcome, to realize clinically efficacious regenerative stem cell therapeutics. PMID:25621112

  8. Adult human neural stem cell therapeutics: Current developmental status and prospect.

    PubMed

    Nam, Hyun; Lee, Kee-Hang; Nam, Do-Hyun; Joo, Kyeung Min

    2015-01-26

    Over the past two decades, regenerative therapies using stem cell technologies have been developed for various neurological diseases. Although stem cell therapy is an attractive option to reverse neural tissue damage and to recover neurological deficits, it is still under development so as not to show significant treatment effects in clinical settings. In this review, we discuss the scientific and clinical basics of adult neural stem cells (aNSCs), and their current developmental status as cell therapeutics for neurological disease. Compared with other types of stem cells, aNSCs have clinical advantages, such as limited proliferation, inborn differentiation potential into functional neural cells, and no ethical issues. In spite of the merits of aNSCs, difficulties in the isolation from the normal brain, and in the in vitro expansion, have blocked preclinical and clinical study using aNSCs. However, several groups have recently developed novel techniques to isolate and expand aNSCs from normal adult brains, and showed successful applications of aNSCs to neurological diseases. With new technologies for aNSCs and their clinical strengths, previous hurdles in stem cell therapies for neurological diseases could be overcome, to realize clinically efficacious regenerative stem cell therapeutics.

  9. Immunomodulatory effect of human adipose tissue-derived adult stem cells: comparison with bone marrow mesenchymal stem cells.

    PubMed

    Puissant, Bénédicte; Barreau, Corinne; Bourin, Philippe; Clavel, Cyril; Corre, Jill; Bousquet, Christine; Taureau, Christine; Cousin, Béatrice; Abbal, Michel; Laharrague, Patrick; Penicaud, Luc; Casteilla, Louis; Blancher, Antoine

    2005-04-01

    Like mesenchymal stem cells from bone marrow (BM-MSCs), adipose tissue-derived adult stem cells (ADAS cells) can differentiate into several lineages and present therapeutical potential for repairing damaged tissues. The use of allogenic stem cells can enlarge their therapeutical interest, provided that the grafted cells could be tolerated. We investigate here, for the first time, the immunosuppressive properties of ADAS cells compared with the well-characterized immunosuppressive properties of BM-MSCs. ADAS cells did not provoke in vitro alloreactivity of incompatible lymphocytes and, moreover, suppressed mixed lymphocyte reaction (MLR) and lymphocyte proliferative response to mitogens. The impairment of inhibition when ADAS cells and BM-MSCs were separated from lymphocytes by a permeable membrane suggests that cell contact is required for a full inhibitory effect. Hepatocyte growth factor is secreted by both stem cells but, similar to interleukin-10 and transforming growth factor-beta (TGF-beta), the levels of which were undetectable in supernatants of MLR inhibited by ADAS cells or BM-MSCs, it did not seem implicated in the stem cell suppressive effect. These findings support that ADAS cells share immunosuppressive properties with BM-MSCs. Therefore, ADAS cell-based reconstructive therapy could employ allogenic cells and because of their immunosuppressive properties, ADAS cells could be an alternative source to BM-MSCs to treat allogenic conflicts.

  10. Encapsulation of adult human mesenchymal stem cells within collagen-agarose microenvironments.

    PubMed

    Batorsky, Anna; Liao, Jiehong; Lund, Amanda W; Plopper, George E; Stegemann, Jan P

    2005-11-20

    Reliable control over the process of cell differentiation is a major challenge in moving stem cell-based therapies forward. The composition of the extracellular matrix (ECM) is known to play an important role in modulating differentiation. We have developed a system to encapsulate adult human mesenchymal stem cells (hMSC) within spherical three-dimensional (3D) microenvironments consisting of a defined mixture of collagen Type I and agarose polymers. These protein-based beads were produced by emulsification of liquid hMSC-matrix suspensions in a silicone fluid phase and subsequent gelation to form hydrogel beads, which were collected by centrifugation and placed in culture. Bead size and size distribution could be varied by changing the encapsulation parameters (impeller speed and blade separation), and beads in the range of 30-150 microns in diameter were reliably produced. Collagen concentrations up to 40% (wt/wt) could be incorporated into the bead matrix. Visible light and fluorescence microscopy confirmed that the collagen matrix was uniformly distributed throughout the beads. Cell viability post-encapsulation was in the range of 75-90% for all bead formulations (similar to control slab gels) and remained at this level for 8 days in culture. Fluorescent staining of the actin cytoskeleton revealed that hMSC spreading increased with increasing collagen concentration. This system of producing 3D microenvironments of defined matrix composition therefore offers a way to control cell-matrix interactions and thereby guide hMSC differentiation. The bead format allows the use of small amounts of matrix proteins, and such beads can potentially be used as a cell delivery vehicle in tissue repair applications.

  11. The simplest method for in vitro β-cell production from human adult stem cells.

    PubMed

    Bhandari, Dilli Ram; Seo, Kwang-Won; Sun, Bo; Seo, Min-Soo; Kim, Hyung-Sik; Seo, Yoo-Jin; Marcin, Jurga; Forraz, Nicolas; Roy, Helene Le; Larry, Denner; Colin, McGuckin; Kang, Kyung-Sun

    2011-10-01

    Diabetes mellitus is a challenging autoimmune disease. Biomedical researchers are currently exploring efficient and effective ways to solve this challenge. The potential of stem cell therapies for treating diabetes represents one of the major focuses of current research on diabetes treatment. Here, we have attempted to differentiate adult stem cells from umbilical cord blood-derived mesenchymal cells (UCB-MSC), Wharton's jelly-derived mesenchymal stem cells (WJ-MSC) and amniotic epithelial stem cells (AE-SC) into insulin-producing cells. The serum-free protocol developed in this study resulted in the differentiation of cells into definitive endoderm, pancreatic foregut, pancreatic endoderm and, finally, pancreatic endocrine cells, which expressed the marker genes SOX17, PDX1, NGN3, NKX6.1, INS, GCG, and PPY, respectively. Detection of the expression of the gap junction-related gene connexin-36 (CX36) using RT-PCR provided conclusive evidence for insulin-producing cell differentiation. In addition to this RT-PCR result, insulin and C-peptide protein were detected by immunohistochemistry and ELISA. Glucose stimulation test results showed that significantly greater amounts of C-peptide and insulin were released from differentiated cells than from undifferentiated cells. In conclusion, the methods investigated in this study can be considered an effective and efficient means of obtaining insulin-producing cells from adult stem cells within a week.

  12. Intrastriatal transplantation of adult human neural crest-derived stem cells improves functional outcome in parkinsonian rats.

    PubMed

    Müller, Janine; Ossig, Christiana; Greiner, Johannes F W; Hauser, Stefan; Fauser, Mareike; Widera, Darius; Kaltschmidt, Christian; Storch, Alexander; Kaltschmidt, Barbara

    2015-01-01

    Parkinson's disease (PD) is considered the second most frequent and one of the most severe neurodegenerative diseases, with dysfunctions of the motor system and with nonmotor symptoms such as depression and dementia. Compensation for the progressive loss of dopaminergic (DA) neurons during PD using current pharmacological treatment strategies is limited and remains challenging. Pluripotent stem cell-based regenerative medicine may offer a promising therapeutic alternative, although the medical application of human embryonic tissue and pluripotent stem cells is still a matter of ethical and practical debate. Addressing these challenges, the present study investigated the potential of adult human neural crest-derived stem cells derived from the inferior turbinate (ITSCs) transplanted into a parkinsonian rat model. Emphasizing their capability to give rise to nervous tissue, ITSCs isolated from the adult human nose efficiently differentiated into functional mature neurons in vitro. Additional successful dopaminergic differentiation of ITSCs was subsequently followed by their transplantation into a unilaterally lesioned 6-hydroxydopamine rat PD model. Transplantation of predifferentiated or undifferentiated ITSCs led to robust restoration of rotational behavior, accompanied by significant recovery of DA neurons within the substantia nigra. ITSCs were further shown to migrate extensively in loose streams primarily toward the posterior direction as far as to the midbrain region, at which point they were able to differentiate into DA neurons within the locus ceruleus. We demonstrate, for the first time, that adult human ITSCs are capable of functionally recovering a PD rat model.

  13. Advances in homology directed genetic engineering of human pluripotent and adult stem cells.

    PubMed

    Ramamoorthi, Kalpith; Curtis, Donald; Asuri, Prashanth

    2013-10-26

    The ability to introduce precise genomic modifications in human cells has profound implications for both basic and applied research in stem cells, ranging from identification of genes regulating stem cell self-renewal and multilineage differentiation to therapeutic gene correction and creation of in vitro models of human diseases. However, the overall efficiency of this process is challenged by several factors including inefficient gene delivery into stem cells and low rates of homology directed site-specific targeting. Recent studies report the development of novel techniques to improve gene targeting efficiencies in human stem cells; these methods include molecular engineering of viral vectors to efficiently deliver episomal genetic sequences that can participate in homology directed targeting, as well as the design of synthetic proteins that can introduce double-stranded breaks in DNA to initiate such recombination events. This review focuses on the potential of these new technologies to precisely alter the human stem cell genome and also highlights the possibilities offered by the combination of these complementary strategies.

  14. Adult Stem Cells and Diabetes Therapy

    PubMed Central

    Ilgun, Handenur; Kim, Joseph William; Luo, LuGuang

    2016-01-01

    The World Health Organization estimates that diabetes will be the fourth most prevalent disease by 2050. Developing a new therapy for diabetes is a challenge for researchers and clinicians in field. Many medications are being used for treatment of diabetes however with no conclusive and effective results therefore alternative therapies are required. Stem cell therapy is a promising tool for diabetes therapy, and it has involved embryonic stem cells, adult stem cells, and pluripotent stem cells. In this review, we focus on adult stem cells, especial human bone marrow stem cells (BM) for diabetes therapy, its history, and current development. We discuss prospects for future diabetes therapy such as induced pluripotent stem cells which have popularity in stem cell research area. PMID:27123495

  15. Human embryonic and fetal mesenchymal stem cells differentiate toward three different cardiac lineages in contrast to their adult counterparts.

    PubMed

    Ramkisoensing, Arti A; Pijnappels, Daniël A; Askar, Saïd F A; Passier, Robert; Swildens, Jim; Goumans, Marie José; Schutte, Cindy I; de Vries, Antoine A F; Scherjon, Sicco; Mummery, Christine L; Schalij, Martin J; Atsma, Douwe E

    2011-01-01

    Mesenchymal stem cells (MSCs) show unexplained differences in differentiation potential. In this study, differentiation of human (h) MSCs derived from embryonic, fetal and adult sources toward cardiomyocytes, endothelial and smooth muscle cells was investigated. Labeled hMSCs derived from embryonic stem cells (hESC-MSCs), fetal umbilical cord, bone marrow, amniotic membrane and adult bone marrow and adipose tissue were co-cultured with neonatal rat cardiomyocytes (nrCMCs) or cardiac fibroblasts (nrCFBs) for 10 days, and also cultured under angiogenic conditions. Cardiomyogenesis was assessed by human-specific immunocytological analysis, whole-cell current-clamp recordings, human-specific qRT-PCR and optical mapping. After co-culture with nrCMCs, significantly more hESC-MSCs than fetal hMSCs stained positive for α-actinin, whereas adult hMSCs stained negative. Furthermore, functional cardiomyogenic differentiation, based on action potential recordings, was shown to occur, but not in adult hMSCs. Of all sources, hESC-MSCs expressed most cardiac-specific genes. hESC-MSCs and fetal hMSCs contained significantly higher basal levels of connexin43 than adult hMSCs and co-culture with nrCMCs increased expression. After co-culture with nrCFBs, hESC-MSCs and fetal hMSCs did not express α-actinin and connexin43 expression was decreased. Conduction velocity (CV) in co-cultures of nrCMCs and hESC-MSCs was significantly higher than in co-cultures with fetal or adult hMSCs. In angiogenesis bioassays, only hESC-MSCs and fetal hMSCs were able to form capillary-like structures, which stained for smooth muscle and endothelial cell markers.Human embryonic and fetal MSCs differentiate toward three different cardiac lineages, in contrast to adult MSCs. Cardiomyogenesis is determined by stimuli from the cellular microenvironment, where connexin43 may play an important role.

  16. Inhibition of activated pericentromeric SINE/Alu repeat transcription in senescent human adult stem cells reinstates self-renewal

    PubMed Central

    Hostikka, Sirkka Liisa; Atallah, Michelle; Blackwell, Benjamin; Lee, Elbert; Cook, Peter J; Pasaniuc, Bogdan; Shariat, Goli; Halperin, Eran; Dobke, Marek; Rosenfeld, Michael G

    2011-01-01

    Cellular aging is linked to deficiencies in efficient repair of DNA double strand breaks and authentic genome maintenance at the chromatin level. Aging poses a significant threat to adult stem cell function by triggering persistent DNA damage and ultimately cellular senescence. Senescence is often considered to be an irreversible process. Moreover, critical genomic regions engaged in persistent DNA damage accumulation are unknown. Here we report that 65% of naturally occurring repairable DNA damage in self-renewing adult stem cells occurs within transposable elements. Upregulation of Alu retrotransposon transcription upon ex vivo aging causes nuclear cytotoxicity associated with the formation of persistent DNA damage foci and loss of efficient DNA repair in pericentric chromatin. This occurs due to a failure to recruit of condensin I and cohesin complexes. Our results demonstrate that the cytotoxicity of induced Alu repeats is functionally relevant for the human adult stem cell aging. Stable suppression of Alu transcription can reverse the senescent phenotype, reinstating the cells' self-renewing properties and increasing their plasticity by altering so-called “master” pluripotency regulators. PMID:21862875

  17. Adult Stem and Progenitor Cells

    NASA Astrophysics Data System (ADS)

    Geraerts, Martine; Verfaillie, Catherine M.

    The discovery of adult stem cells in most adult tissues is the basis of a number of clinical studies that are carried out, with therapeutic use of hematopoietic stem cells as a prime example. Intense scientific debate is still ongoing as to whether adult stem cells may have a greater plasticity than previously thought. Although cells with some features of embryonic stem cells that, among others, express Oct4, Nanog and SSEA1 are isolated from fresh tissue, it is not clear if the greater differentiation potential is acquired during cell culture. Moreover, adult more pluripotent cells do not have all pluripotent characteristics typical for embryonic stem cells. Recently, some elegant studies were published in which adult cells could be completely reprogrammed to embryonic stem cell-like cells by overexpression of some key transcription factors for pluripotency (Oct4, Sox2, Klf4 and c-Myc). It will be interesting for the future to investigate the exact mechanisms underlying this reprogramming and whether similar transcription factor pathways are present and/or can be activated in adult more pluripotent stem cells.

  18. Perspectives on human stem cell research.

    PubMed

    Jung, Kyu Won

    2009-09-01

    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.

  19. Adult stem-like cells in kidney

    PubMed Central

    Hishikawa, Keiichi; Takase, Osamu; Yoshikawa, Masahiro; Tsujimura, Taro; Nangaku, Masaomi; Takato, Tsuyoshi

    2015-01-01

    Human pluripotent cells are promising for treatment for kidney diseases, but the protocols for derivation of kidney cell types are still controversial. Kidney tissue regeneration is well confirmed in several lower vertebrates such as fish, and the repair of nephrons after tubular damages is commonly observed after renal injury. Even in adult mammal kidney, renal progenitor cell or system is reportedly presents suggesting that adult stem-like cells in kidney can be practical clinical targets for kidney diseases. However, it is still unclear if kidney stem cells or stem-like cells exist or not. In general, stemness is defined by several factors such as self-renewal capacity, multi-lineage potency and characteristic gene expression profiles. The definite use of stemness may be obstacle to understand kidney regeneration, and here we describe the recent broad findings of kidney regeneration and the cells that contribute regeneration. PMID:25815133

  20. Adult stem-like cells in kidney.

    PubMed

    Hishikawa, Keiichi; Takase, Osamu; Yoshikawa, Masahiro; Tsujimura, Taro; Nangaku, Masaomi; Takato, Tsuyoshi

    2015-03-26

    Human pluripotent cells are promising for treatment for kidney diseases, but the protocols for derivation of kidney cell types are still controversial. Kidney tissue regeneration is well confirmed in several lower vertebrates such as fish, and the repair of nephrons after tubular damages is commonly observed after renal injury. Even in adult mammal kidney, renal progenitor cell or system is reportedly presents suggesting that adult stem-like cells in kidney can be practical clinical targets for kidney diseases. However, it is still unclear if kidney stem cells or stem-like cells exist or not. In general, stemness is defined by several factors such as self-renewal capacity, multi-lineage potency and characteristic gene expression profiles. The definite use of stemness may be obstacle to understand kidney regeneration, and here we describe the recent broad findings of kidney regeneration and the cells that contribute regeneration. PMID:25815133

  1. Evidence for tissue-resident mesenchymal stem cells in human adult lung from studies of transplanted allografts.

    PubMed

    Lama, Vibha N; Smith, Lisa; Badri, Linda; Flint, Andrew; Andrei, Adin-Cristian; Murray, Susan; Wang, Zhuo; Liao, Hui; Toews, Galen B; Krebsbach, Paul H; Peters-Golden, Marc; Pinsky, David J; Martinez, Fernando J; Thannickal, Victor J

    2007-04-01

    The origin and turnover of connective tissue cells in adult human organs, including the lung, are not well understood. Here, studies of cells derived from human lung allografts demonstrate the presence of a multipotent mesenchymal cell population, which is locally resident in the human adult lung and has extended life span in vivo. Examination of plastic-adherent cell populations in bronchoalveolar lavage samples obtained from 76 human lung transplant recipients revealed clonal proliferation of fibroblast-like cells in 62% (106 of 172) of samples. Immunophenotyping of these isolated cells demonstrated expression of vimentin and prolyl-4-hydroxylase, indicating a mesenchymal phenotype. Multiparametric flow cytometric analyses revealed expression of cell-surface proteins, CD73, CD90, and CD105, commonly found on mesenchymal stem cells (MSCs). Hematopoietic lineage markers CD14, CD34, and CD45 were absent. Multipotency of these cells was demonstrated by their capacity to differentiate into adipocytes, chondrocytes, and osteocytes. Cytogenetic analysis of cells from 7 sex-mismatched lung transplant recipients harvested up to 11 years after transplant revealed that 97.2% +/- 2.1% expressed the sex genotype of the donor. The presence of MSCs of donor sex identity in lung allografts even years after transplantation provides what we believe to be the first evidence for connective tissue cell progenitors that reside locally within a postnatal, nonhematopoietic organ.

  2. Sphingosine-1-phosphate mediates proliferation maintaining the multipotency of human adult bone marrow and adipose tissue-derived stem cells.

    PubMed

    He, Xiaoli; H'ng, Shiau-Chen; Leong, David T; Hutmacher, Dietmar W; Melendez, Alirio J

    2010-08-01

    High renewal and maintenance of multipotency of human adult stem cells (hSCs), are a prerequisite for experimental analysis as well as for potential clinical usages. The most widely used strategy for hSC culture and proliferation is using serum. However, serum is poorly defined and has a considerable degree of inter-batch variation, which makes it difficult for large-scale mesenchymal stem cells (MSCs) expansion in homogeneous culture conditions. Moreover, it is often observed that cells grown in serum-containing media spontaneously differentiate into unknown and/or undesired phenotypes. Another way of maintaining hSC development is using cytokines and/or tissue-specific growth factors; this is a very expensive approach and can lead to early unwanted differentiation. In order to circumvent these issues, we investigated the role of sphingosine-1-phosphate (S1P), in the growth and multipotency maintenance of human bone marrow and adipose tissue-derived MSCs. We show that S1P induces growth, and in combination with reduced serum, or with the growth factors FGF and platelet-derived growth factor-AB, S1P has an enhancing effect on growth. We also show that the MSCs cultured in S1P-supplemented media are able to maintain their differentiation potential for at least as long as that for cells grown in the usual serum-containing media. This is shown by the ability of cells grown in S1P-containing media to be able to undergo osteogenic as well as adipogenic differentiation. This is of interest, since S1P is a relatively inexpensive natural product, which can be obtained in homogeneous high-purity batches: this will minimize costs and potentially reduce the unwanted side effects observed with serum. Taken together, S1P is able to induce proliferation while maintaining the multipotency of different human stem cells, suggesting a potential for S1P in developing serum-free or serum-reduced defined medium for adult stem cell cultures.

  3. Three-Dimensional Adult Cardiac Extracellular Matrix Promotes Maturation of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

    PubMed

    Fong, Ashley H; Romero-López, Mónica; Heylman, Christopher M; Keating, Mark; Tran, David; Sobrino, Agua; Tran, Anh Q; Pham, Hiep H; Fimbres, Cristhian; Gershon, Paul D; Botvinick, Elliot L; George, Steven C; Hughes, Christopher C W

    2016-08-01

    Pluripotent stem cell-derived cardiomyocytes (CMs) have great potential in the development of new therapies for cardiovascular disease. In particular, human induced pluripotent stem cells (iPSCs) may prove especially advantageous due to their pluripotency, their self-renewal potential, and their ability to create patient-specific cell lines. Unfortunately, pluripotent stem cell-derived CMs are immature, with characteristics more closely resembling fetal CMs than adult CMs, and this immaturity has limited their use in drug screening and cell-based therapies. Extracellular matrix (ECM) influences cellular behavior and maturation, as does the geometry of the environment-two-dimensional (2D) versus three-dimensional (3D). We therefore tested the hypothesis that native cardiac ECM and 3D cultures might enhance the maturation of iPSC-derived CMs in vitro. We demonstrate that maturation of iPSC-derived CMs was enhanced when cells were seeded into a 3D cardiac ECM scaffold, compared with 2D culture. 3D cardiac ECM promoted increased expression of calcium-handling genes, Junctin, CaV1.2, NCX1, HCN4, SERCA2a, Triadin, and CASQ2. Consistent with this, we find that iPSC-derived CMs in 3D adult cardiac ECM show increased calcium signaling (amplitude) and kinetics (maximum upstroke and downstroke) compared with cells in 2D. Cells in 3D culture were also more responsive to caffeine, likely reflecting an increased availability of calcium in the sarcoplasmic reticulum. Taken together, these studies provide novel strategies for maturing iPSC-derived CMs that may have applications in drug screening and transplantation therapies to treat heart disease. PMID:27392582

  4. Biological character of human adipose-derived adult stem cells and influence of donor age on cell replication in culture.

    PubMed

    Lei, Lei; Liao, WeiMing; Sheng, PuYi; Fu, Ming; He, AiShan; Huang, Gang

    2007-06-01

    To investigate the biological character of human adipose-derived adult stem cells (hADAS cells) when cultured in vitro and the relationship between hADAS cell's replication activity and the donor's age factor, and to assess the stem cells as a new source for tissue engineering. hADAS cells are isolated from human adipose tissue of different age groups (from adolescents to olds: <20 years old, 21-40 years old, 41-60 years old and >61 years old groups). The protein markers (CD29, CD34, CD44, CD45, CD49d, HLA-DR, CD106) of hADAS cells were detected by flow cytometry (FCM) to identify the stem cell, and the cell cycle was examined for P20 hADAS cells to evaluate the safety of the subculture in vitro. The generative activity of hADAS cells in different age groups was also examined by MTT method. The formula "TD = t x log2/logNt - logN0" was used to get the time doubling (TD) of the cells. The results showed that the cells kept heredity stabilization by chromosome analysis for at least 20 passages. The TD of these cells increased progressively by ageing, and the TD of the <20 years old group was lower than that of the >61 years old group (statistical analysis of variance (ANOVA), P=0.002, P<0.05). These findings suggested that a higher level of hADAS cells replication activity was found in the younger donators, and they represent novel and valuable seed cells for studies of tissue engineering.

  5. Recapitulation of endochondral bone formation using human adult mesenchymal stem cells as a paradigm for developmental engineering.

    PubMed

    Scotti, Celeste; Tonnarelli, Beatrice; Papadimitropoulos, Adam; Scherberich, Arnaud; Schaeren, Stefan; Schauerte, Alexandra; Lopez-Rios, Javier; Zeller, Rolf; Barbero, Andrea; Martin, Ivan

    2010-04-20

    Mesenchymal stem/stromal cells (MSC) are typically used to generate bone tissue by a process resembling intramembranous ossification, i.e., by direct osteoblastic differentiation. However, most bones develop by endochondral ossification, i.e., via remodeling of hypertrophic cartilaginous templates. To date, endochondral bone formation has not been reproduced using human, clinically compliant cell sources. Here, we aimed at engineering tissues from bone marrow-derived, adult human MSC with an intrinsic capacity to undergo endochondral ossification. By analogy to embryonic limb development, we hypothesized that successful execution of the endochondral program depends on the initial formation of hypertrophic cartilaginous templates. Human MSC, subcutaneously implanted into nude mice at various stages of chondrogenic differentiation, formed bone trabeculae only when they had developed in vitro hypertrophic tissue structures. Advanced maturation in vitro resulted in accelerated formation of larger bony tissues. The underlying morphogenetic process was structurally and molecularly similar to the temporal and spatial progression of limb bone development in embryos. In particular, Indian hedgehog signaling was activated at early stages and required for the in vitro formation of hypertrophic cartilage. Subsequent development of a bony collar in vivo was followed by vascularization, osteoclastic resorption of the cartilage template, and appearance of hematopoietic foci. This study reveals the capacity of human MSC to generate bone tissue via an endochondral program and provides a valid model to study mechanisms governing bone development. Most importantly, this process could generate advanced grafts for bone regeneration by invoking a "developmental engineering" paradigm.

  6. Effects of FGF-2 on human adipose tissue derived adult stem cells morphology and chondrogenesis enhancement in Transwell culture

    SciTech Connect

    Kabiri, Azadeh; Esfandiari, Ebrahim; Hashemibeni, Batool; Kazemi, Mohammad; Mardani, Mohammad; Esmaeili, Abolghasem

    2012-07-27

    Highlights: Black-Right-Pointing-Pointer We investigated effects of FGF-2 on hADSCs. Black-Right-Pointing-Pointer We examine changes in the level of gene expressions of SOX-9, aggrecan and collagen type II and type X. Black-Right-Pointing-Pointer FGF-2 induces chondrogenesis in hADSCs, which Bullet Increasing information will decrease quality if hospital costs are very different. Black-Right-Pointing-Pointer The result of this study may be beneficial in cartilage tissue engineering. -- Abstract: Injured cartilage is difficult to repair due to its poor vascularisation. Cell based therapies may serve as tools to more effectively regenerate defective cartilage. Both adult mesenchymal stem cells (MSCs) and human adipose derived stem cells (hADSCs) are regarded as potential stem cell sources able to generate functional cartilage for cell transplantation. Growth factors, in particular the TGF-b superfamily, influence many processes during cartilage formation, including cell proliferation, extracellular matrix synthesis, maintenance of the differentiated phenotype, and induction of MSCs towards chondrogenesis. In the current study, we investigated the effects of FGF-2 on hADSC morphology and chondrogenesis in Transwell culture. hADSCs were obtained from patients undergoing elective surgery, and then cultured in expansion medium alone or in the presence of FGF-2 (10 ng/ml). mRNA expression levels of SOX-9, aggrecan and collagen type II and type X were quantified by real-time polymerase chain reaction. The morphology, doubling time, trypsinization time and chondrogenesis of hADSCs were also studied. Expression levels of SOX-9, collagen type II, and aggrecan were all significantly increased in hADSCs expanded in presence of FGF-2. Furthermore FGF-2 induced a slender morphology, whereas doubling time and trypsinization time decreased. Our results suggest that FGF-2 induces hADSCs chondrogenesis in Transwell culture, which may be beneficial in cartilage tissue engineering.

  7. The development and validation of a LIPUS system with preliminary observations of ultrasonic effects on human adult stem cells.

    PubMed

    Marvel, Skylar; Okrasinski, Stan; Bernacki, Susan H; Loboa, Elizabeth; Dayton, Paul A

    2010-09-01

    To study the potential effects of low-intensity pulsed ultrasound (LIPUS) on cell response in vitro, the ability to alter LIPUS parameters is required. However, commercial LIPUS systems have very little control over parameter selection. In this study, a custom LIPUS system was designed and validated by exploring the effects of using different pulse repetition frequency (PRF) parameters on human adipose derived adult stem cells (hASCs) and bone marrow derived mesenchymal stem cells (hMSCs), two common stem cell sources for creating bone constructs in vitro. Changing the PRF was found to affect cellular response to LIPUS stimulation for both cell types. Proliferation of LIPUS-stimulated cells was found to decrease for hASCs by d 7 for all three groups compared with unstimulated control cells (P = 0.008, 0.011, 0.014 for 1 Hz, 100 Hz and 1 kHz PRF, respectively) and for hMSCs by d 14 (donor 1: P = 0.0005, 0.0002, 0.0003; donor 2: P = 0.0003, 0.0002, 0.0001; for PRFs of 1 Hz, 100 Hz, and 1 kHz, respectively). Additionally, LIPUS was shown to strongly accelerate osteogenic differentiation of hASCs based on amount of calcium accretion normalized by total DNA (P = 0.003, 0.001, 0.003, and 0.032 between control/100 Hz, control/1 kHz, 1 Hz/1 kHz, and 100 Hz/1 kHz pulse repetition frequencies, respectively). These findings promote the study of using LIPUS to induce osteogenic differentiation and further encourage the exploration of LIPUS parameter optimization. The custom LIPUS system was successfully designed to allow extreme parameter variation, specifically PRF, and encourages further studies. PMID:20875987

  8. Isolation of small SSEA-4-positive putative stem cells from the ovarian surface epithelium of adult human ovaries by two different methods.

    PubMed

    Virant-Klun, Irma; Skutella, Thomas; Hren, Matjaz; Gruden, Kristina; Cvjeticanin, Branko; Vogler, Andrej; Sinkovec, Jasna

    2013-01-01

    The adult ovarian surface epithelium has already been proposed as a source of stem cells and germinal cells in the literature, therefore it has been termed the "germinal epithelium". At present more studies have confirmed the presence of stem cells expressing markers of pluripotency in adult mammalian ovaries, including humans. The aim of this study was to isolate a population of stem cells, based on the expression of pluripotency-related stage-specific embryonic antigen-4 (SSEA-4) from adult human ovarian surface epithelium by two different methods: magnetic-activated cell sorting and fluorescence-activated cell sorting. Both methods made it possible to isolate a similar, relatively homogenous population of small, SSEA-4-positive cells with diameters of up to 4  μm from the suspension of cells retrieved by brushing of the ovarian cortex biopsies in reproductive-age and postmenopausal women and in women with premature ovarian failure. The immunocytochemistry and genetic analyses revealed that these small cells--putative stem cells--expressed some primordial germ cell and pluripotency-related markers and might be related to the in vitro development of oocyte-like cells expressing some oocyte-specific transcription factors in the presence of donated follicular fluid with substances important for oocyte growth and development. The stemness of these cells needs to be further researched.

  9. Translational research of adult stem cell therapy.

    PubMed

    Suzuki, Gen

    2015-11-26

    Congestive heart failure (CHF) secondary to chronic coronary artery disease is a major cause of morbidity and mortality world-wide. Its prevalence is increasing despite advances in medical and device therapies. Cell based therapies generating new cardiomyocytes and vessels have emerged as a promising treatment to reverse functional deterioration and prevent the progression to CHF. Functional efficacy of progenitor cells isolated from the bone marrow and the heart have been evaluated in preclinical large animal models. Furthermore, several clinical trials using autologous and allogeneic stem cells and progenitor cells have demonstrated their safety in humans yet their clinical relevance is inconclusive. This review will discuss the clinical therapeutic applications of three specific adult stem cells that have shown particularly promising regenerative effects in preclinical studies, bone marrow derived mesenchymal stem cell, heart derived cardiosphere-derived cell and cardiac stem cell. We will also discuss future therapeutic approaches.

  10. Dominant-Negative Effects of Adult-Onset Huntingtin Mutations Alter the Division of Human Embryonic Stem Cells-Derived Neural Cells

    PubMed Central

    Lopes, Carla; Aubert, Sophie; Bourgois-Rocha, Fany; Barnat, Monia; Rego, Ana Cristina; Déglon, Nicole

    2016-01-01

    Mutations of the huntingtin protein (HTT) gene underlie both adult-onset and juvenile forms of Huntington’s disease (HD). HTT modulates mitotic spindle orientation and cell fate in mouse cortical progenitors from the ventricular zone. Using human embryonic stem cells (hESC) characterized as carrying mutations associated with adult-onset disease during pre-implantation genetic diagnosis, we investigated the influence of human HTT and of an adult-onset HD mutation on mitotic spindle orientation in human neural stem cells (NSCs) derived from hESCs. The RNAi-mediated silencing of both HTT alleles in neural stem cells derived from hESCs disrupted spindle orientation and led to the mislocalization of dynein, the p150Glued subunit of dynactin and the large nuclear mitotic apparatus (NuMA) protein. We also investigated the effect of the adult-onset HD mutation on the role of HTT during spindle orientation in NSCs derived from HD-hESCs. By combining SNP-targeting allele-specific silencing and gain-of-function approaches, we showed that a 46-glutamine expansion in human HTT was sufficient for a dominant-negative effect on spindle orientation and changes in the distribution within the spindle pole and the cell cortex of dynein, p150Glued and NuMA in neural cells. Thus, neural derivatives of disease-specific human pluripotent stem cells constitute a relevant biological resource for exploring the impact of adult-onset HD mutations of the HTT gene on the division of neural progenitors, with potential applications in HD drug discovery targeting HTT-dynein-p150Glued complex interactions. PMID:26863614

  11. Dominant-Negative Effects of Adult-Onset Huntingtin Mutations Alter the Division of Human Embryonic Stem Cells-Derived Neural Cells.

    PubMed

    Lopes, Carla; Aubert, Sophie; Bourgois-Rocha, Fany; Barnat, Monia; Rego, Ana Cristina; Déglon, Nicole; Perrier, Anselme L; Humbert, Sandrine

    2016-01-01

    Mutations of the huntingtin protein (HTT) gene underlie both adult-onset and juvenile forms of Huntington's disease (HD). HTT modulates mitotic spindle orientation and cell fate in mouse cortical progenitors from the ventricular zone. Using human embryonic stem cells (hESC) characterized as carrying mutations associated with adult-onset disease during pre-implantation genetic diagnosis, we investigated the influence of human HTT and of an adult-onset HD mutation on mitotic spindle orientation in human neural stem cells (NSCs) derived from hESCs. The RNAi-mediated silencing of both HTT alleles in neural stem cells derived from hESCs disrupted spindle orientation and led to the mislocalization of dynein, the p150Glued subunit of dynactin and the large nuclear mitotic apparatus (NuMA) protein. We also investigated the effect of the adult-onset HD mutation on the role of HTT during spindle orientation in NSCs derived from HD-hESCs. By combining SNP-targeting allele-specific silencing and gain-of-function approaches, we showed that a 46-glutamine expansion in human HTT was sufficient for a dominant-negative effect on spindle orientation and changes in the distribution within the spindle pole and the cell cortex of dynein, p150Glued and NuMA in neural cells. Thus, neural derivatives of disease-specific human pluripotent stem cells constitute a relevant biological resource for exploring the impact of adult-onset HD mutations of the HTT gene on the division of neural progenitors, with potential applications in HD drug discovery targeting HTT-dynein-p150Glued complex interactions.

  12. Dental Stem Cell in Tooth Development and Advances of Adult Dental Stem Cell in Regenerative Therapies.

    PubMed

    Tan, Jiali; Xu, Xin; Lin, Jiong; Fan, Li; Zheng, Yuting; Kuang, Wei

    2015-01-01

    Stem cell-based therapies are considered as a promising treatment for many clinical usage such as tooth regeneration, bone repairation, spinal cord injury, and so on. However, the ideal stem cell for stem cell-based therapy still remains to be elucidated. In the past decades, several types of stem cells have been isolated from teeth, including dental pulp stem cells (DPSCs), stem cells from human exfoliated deciduous teeth (SHED), periodontal ligament stem cells (PDLSCs), dental follicle progenitor stem cells (DFPCs) and stem cells from apical papilla (SCAP), which may be a good source for stem cell-based therapy in certain disease, especially when they origin from neural crest is considered. In this review, the specific characteristics and advantages of the adult dental stem cell population will be summarized and the molecular mechanisms of the differentiation of dental stem cell during tooth development will be also discussed.

  13. Markers of epidermal stem cell subpopulations in adult mammalian skin.

    PubMed

    Kretzschmar, Kai; Watt, Fiona M

    2014-10-01

    The epidermis is the outermost layer of mammalian skin and comprises a multilayered epithelium, the interfollicular epidermis, with associated hair follicles, sebaceous glands, and eccrine sweat glands. As in other epithelia, adult stem cells within the epidermis maintain tissue homeostasis and contribute to repair of tissue damage. The bulge of hair follicles, where DNA-label-retaining cells reside, was traditionally regarded as the sole epidermal stem cell compartment. However, in recent years multiple stem cell populations have been identified. In this review, we discuss the different stem cell compartments of adult murine and human epidermis, the markers that they express, and the assays that are used to characterize epidermal stem cell properties.

  14. Expression of Genes Related to Germ Cell Lineage and Pluripotency in Single Cells and Colonies of Human Adult Germ Stem Cells

    PubMed Central

    Conrad, Sabine; Azizi, Hossein; Hatami, Maryam; Kubista, Mikael; Bonin, Michael; Hennenlotter, Jörg; Sievert, Karl-Dietrich; Skutella, Thomas

    2016-01-01

    The aim of this study was to elucidate the molecular status of single human adult germ stem cells (haGSCs) and haGSC colonies, which spontaneously developed from the CD49f MACS and matrix- (collagen−/laminin+ binding-) selected fraction of enriched spermatogonia. Single-cell transcriptional profiling by Fluidigm BioMark system of a long-term cultured haGSCs cluster in comparison to human embryonic stem cells (hESCs) and human fibroblasts (hFibs) revealed that haGSCs showed a characteristic germ- and pluripotency-associated gene expression profile with some similarities to hESCs and with a significant distinction from somatic hFibs. Genome-wide comparisons with microarray analysis confirmed that different haGSC colonies exhibited gene expression heterogeneity with more or less pluripotency. The results of this study confirm that haGSCs are adult stem cells with a specific molecular gene expression profile in vitro, related but not identical to true pluripotent stem cells. Under ES-cell conditions haGSC colonies could be selected and maintained in a partial pluripotent state at the molecular level, which may be related to their cell plasticity and potential to differentiate into cells of all germ layers. PMID:26649052

  15. Potential of embryonic and adult stem cells in vitro.

    PubMed

    Czyz, Jaroslaw; Wiese, Cornelia; Rolletschek, Alexandra; Blyszczuk, Przemyslaw; Cross, Michael; Wobus, Anna M

    2003-01-01

    Recent developments in the field of stem cell research indicate their enormous potential as a source of tissue for regenerative therapies. The success of such applications will depend on the precise properties and potentials of stem cells isolated either from embryonic, fetal or adult tissues. Embryonic stem cells established from the inner cell mass of early mouse embryos are characterized by nearly unlimited proliferation, and the capacity to differentiate into derivatives of essentially all lineages. The recent isolation and culture of human embryonic stem cell lines presents new opportunities for reconstructive medicine. However, important problems remain; first, the derivation of human embryonic stem cells from in vitro fertilized blastocysts creates ethical problems, and second, the current techniques for the directed differentiation into somatic cell populations yield impure products with tumorigenic potential. Recent studies have also suggested an unexpectedly wide developmental potential of adult tissue-specific stem cells. Here too, many questions remain concerning the nature and status of adult stem cells both in vivo and in vitro and their proliferation and differentiation/transdifferentiation capacity. This review focuses on those issues of embryonic and adult stem cell biology most relevant to their in vitro propagation and differentiation. Questions and problems related to the use of human embryonic and adult stem cells in tissue regeneration and transplantation are discussed.

  16. Insulin-Producing Cells From Adult Human Bone Marrow Mesenchymal Stem Cells Control Streptozotocin-Induced Diabetes In Nude Mice

    PubMed Central

    Gabr, Mahmoud M.; Zakaria, Mahmoud M.; Refaie, Ayman F.; Ismail, Amani M.; Abou-El-Mahasen, Mona A.; Ashamallah, Sylvia A.; Khater, Sherry M.; El-Halawani, Sawsan M.; Ibrahim, Rana Y.; Uin, Gan Shu; Kloc, Malgorzata; Calne, Roy Y.; Ghoneim, Mohamed A.

    2013-01-01

    Harvesting, expansion and directed differentiation of human bone marrow-derived mesenchymal stem cells (BM-MSCs) could provide an autologous source of surrogate β-cells that would alleviate the limitations of availability and/or allogenic rejection following pancreatic or islet transplantation. Bone marrow cells were obtained from three adult type 2 diabetic volunteers and 3 non-diabetic donors. After 3 days in culture, adherent MSCs were expanded for 2 passages. At passage 3, differentiation was carried out in a 3-staged procedure. Cells were cultured in a glucose-rich medium containing several activation and growth factors. Cells were evaluated in-vitro by flow cytometry, immunolabelling, Rt-PCR and human insulin and c-peptide release in responses to increasing glucose concentrations. One thousand cell-clusters were inserted under the renal capsule of diabetic nude mice followed by monitoring of their diabetic status. At the end of differentiation, ~5–10% of cells were immunofluorescent for insulin, c-peptide or glucagon; insulin and c-peptide were co-expressed. Nanogold immunolabelling for electron microscopy demonstrated the presence of c-peptide in the rough endoplasmic reticulum. Insulin-producing cells (IPCs) expressed transcription factors and genes of pancreatic hormones similar to those expressed by pancreatic islets. There was a stepwise increase in human insulin and c-peptide release by IPCs in response to increasing glucose concentrations. Transplantation of IPCs into nude diabetic mice resulted in control of their diabetic status for 3 months. The sera of IPC-transplanted mice contained human insulin and c-peptide but negligible levels of mouse insulin. When the IPCs-bearing kidneys were removed, rapid return of diabetic state was noted. BM-MSCs from diabetic and non-diabetic human subjects could be differentiated without genetic manipulation to form IPCs which, when transplanted, could maintain euglycaemia in diabetic mice for 3 months

  17. Low Proliferation and Differentiation Capacities of Adult Hippocampal Stem Cells Correlate with Memory Dysfunction in Humans

    ERIC Educational Resources Information Center

    Coras, Roland; Siebzehnrubl, Florian A.; Pauli, Elisabeth; Huttner, Hagen B.; Njunting, Marleisje; Kobow, Katja; Villmann, Carmen; Hahnen, Eric; Neuhuber, Winfried; Weigel, Daniel; Buchfelder, Michael; Stefan, Hermann; Beck, Heinz; Steindler, Dennis A.; Blumcke, Ingmar

    2010-01-01

    The hippocampal dentate gyrus maintains its capacity to generate new neurons throughout life. In animal models, hippocampal neurogenesis is increased by cognitive tasks, and experimental ablation of neurogenesis disrupts specific modalities of learning and memory. In humans, the impact of neurogenesis on cognition remains unclear. Here, we…

  18. Genetic expression and functional characterization of the RUNX2 gene in human adult bone marrow mesenchymal stem cells.

    PubMed

    Zhang, J Y; Li, L C

    2015-01-01

    Past studies have revealed the critical role of runt-related transcription factor 2 (RUNX2) in the proliferation and differentiation of mesenchymal stem cells (MSCs). This study therefore aimed to investigate the expression profile of the RUNX2 gene in human bone marrow MSCs and its biological characteristics. Bone marrow MSCs were separated from 12 patients who had received hip joint replacement surgery. After purification and culture, the MSCs were subjected to the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay, flow cytometry, the alkaline phosphatase assay, reverse transcription polymerase chain reaction, and RUNX2 protein quantification. The cell growth curve, staining images, and information on the membrane antigens and the levels of RUNX2 mRNA and protein were obtained based on the results. The growth curve showed, after a 2-day lag period, cultured MSCs entered into the log phase between d3 (Day 3) and d6, when they reached a plateau. Flow cytometry data suggested 94.38% of MSCs were CD90-positive, while only 3.99 and 1.71% of total cells were positive for CD35 and CD45, respectively. With the elongated induction period, cultured MSCs were polygonal in shape. After a 14-day induction, cell fusion occurred in the center of the cell nodule accompanied by the disappearance of cellular structure to form the calcium nodule, which was stained red. There was also a statistically significant increase in the level of RUNX2 protein at d7 and d14. An osteogenic medium is required for the differentiation of adult MSCs, which is also under RUNX2 regulation. These findings are potentially valuable for clinical practice. PMID:26782468

  19. Comparative analysis of paracrine factor expression in human adult mesenchymal stem cells derived from bone marrow, adipose, and dermal tissue.

    PubMed

    Hsiao, Sarah Tzu-Feng; Asgari, Azar; Lokmic, Zerina; Sinclair, Rodney; Dusting, Gregory James; Lim, Shiang Yong; Dilley, Rodney James

    2012-08-10

    Human adult mesenchymal stem cells (MSCs) support the engineering of functional tissue constructs by secreting angiogenic and cytoprotective factors, which act in a paracrine fashion to influence cell survival and vascularization. MSCs have been isolated from many different tissue sources, but little is known about how paracrine factor secretion varies between different MSC populations. We evaluated paracrine factor expression patterns in MSCs isolated from adipose tissue (ASCs), bone marrow (BMSCs), and dermal tissues [dermal sheath cells (DSCs) and dermal papilla cells (DPCs)]. Specifically, mRNA expression analysis identified insulin-like growth factor-1 (IGF-1), vascular endothelial growth factor-D (VEGF-D), and interleukin-8 (IL-8) to be expressed at higher levels in ASCs compared with other MSC populations whereas VEGF-A, angiogenin, basic fibroblast growth factor (bFGF), and nerve growth factor (NGF) were expressed at comparable levels among the MSC populations examined. Analysis of conditioned media (CM) protein confirmed the comparable level of angiogenin and VEGF-A secretion in all MSC populations and showed that DSCs and DPCs produced significantly higher concentrations of leptin. Functional assays examining in vitro angiogenic paracrine activity showed that incubation of endothelial cells in ASC(CM) resulted in increased tubulogenic efficiency compared with that observed in DPC(CM). Using neutralizing antibodies we concluded that VEGF-A and VEGF-D were 2 of the major growth factors secreted by ASCs that supported endothelial tubulogenesis. The variation in paracrine factors of different MSC populations contributes to different levels of angiogenic activity and ASCs maybe preferred over other MSC populations for augmenting therapeutic approaches dependent upon angiogenesis.

  20. Generalized Potential of Adult Neural Stem Cells

    NASA Astrophysics Data System (ADS)

    Clarke, Diana L.; Johansson, Clas B.; Wilbertz, Johannes; Veress, Biborka; Nilsson, Erik; Karlström, Helena; Lendahl, Urban; Frisén, Jonas

    2000-06-01

    The differentiation potential of stem cells in tissues of the adult has been thought to be limited to cell lineages present in the organ from which they were derived, but there is evidence that some stem cells may have a broader differentiation repertoire. We show here that neural stem cells from the adult mouse brain can contribute to the formation of chimeric chick and mouse embryos and give rise to cells of all germ layers. This demonstrates that an adult neural stem cell has a very broad developmental capacity and may potentially be used to generate a variety of cell types for transplantation in different diseases.

  1. Adult stem cells and tissue repair.

    PubMed

    Körbling, M; Estrov, Z; Champlin, R

    2003-08-01

    Recently, adult stem cells originating from bone marrow or peripheral blood have been suggested to contribute to repair and genesis of cells specific for liver, cardiac and skeletal muscle, gut, and brain tissue. The mechanism involved has been termed transdifferentiation, although other explanations including cell fusion have been postulated. Using adult stem cells to generate or repair solid organ tissue obviates the immunologic, ethical, and teratogenic issues that accompany embryonic stem cells.

  2. Ovarian adult stem cells: hope or pitfall?

    PubMed Central

    2014-01-01

    For many years, ovarian biology has been based on the dogma that oocytes reserve in female mammals included a finite number, established before or at birth and it is determined by the number and quality of primordial follicles developed during the neonatal period. The restricted supply of oocytes in adult female mammals has been disputed in recent years by supporters of postnatal neo-oogenesis. Recent experimental data showed that ovarian surface epithelium and cortical tissue from both mouse and human were proved to contain very low proportion of cells able to propagate themselves, but also to generate immature oocytes in vitro or in vivo, when transplanted into immunodeficient mice ovaries. By mentioning several landmarks of ovarian stem cell reserve and addressing the exciting perspective of translation into clinical practice as treatment for infertility pathologies, the purpose of this article is to review the knowledge about adult mammalian ovarian stem cells, a topic that, since the first approach quickly attracted the attention of both the scientific media and patients. PMID:25018783

  3. Markers of Epidermal Stem Cell Subpopulations in Adult Mammalian Skin

    PubMed Central

    Kretzschmar, Kai; Watt, Fiona M.

    2014-01-01

    The epidermis is the outermost layer of mammalian skin and comprises a multilayered epithelium, the interfollicular epidermis, with associated hair follicles, sebaceous glands, and eccrine sweat glands. As in other epithelia, adult stem cells within the epidermis maintain tissue homeostasis and contribute to repair of tissue damage. The bulge of hair follicles, where DNA-label-retaining cells reside, was traditionally regarded as the sole epidermal stem cell compartment. However, in recent years multiple stem cell populations have been identified. In this review, we discuss the different stem cell compartments of adult murine and human epidermis, the markers that they express, and the assays that are used to characterize epidermal stem cell properties. PMID:24993676

  4. Human induced pluripotent stem cell‐derived versus adult cardiomyocytes: an in silico electrophysiological study on effects of ionic current block

    PubMed Central

    Paci, M; Hyttinen, J; Rodriguez, B

    2015-01-01

    Background and Purpose Two new technologies are likely to revolutionize cardiac safety and drug development: in vitro experiments on human‐induced pluripotent stem cell‐derived cardiomyocytes (hiPSC‐CMs) and in silico human adult ventricular cardiomyocyte (hAdultV‐CM) models. Their combination was recently proposed as a potential replacement for the present hERG‐based QT study for pharmacological safety assessments. Here, we systematically compared in silico the effects of selective ionic current block on hiPSC‐CM and hAdultV‐CM action potentials (APs), to identify similarities/differences and to illustrate the potential of computational models as supportive tools for evaluating new in vitro technologies. Experimental Approach In silico AP models of ventricular‐like and atrial‐like hiPSC‐CMs and hAdultV‐CM were used to simulate the main effects of four degrees of block of the main cardiac transmembrane currents. Key Results Qualitatively, hiPSC‐CM and hAdultV‐CM APs showed similar responses to current block, consistent with results from experiments. However, quantitatively, hiPSC‐CMs were more sensitive to block of (i) L‐type Ca2+ currents due to the overexpression of the Na+/Ca2+ exchanger (leading to shorter APs) and (ii) the inward rectifier K+ current due to reduced repolarization reserve (inducing diastolic potential depolarization and repolarization failure). Conclusions and Implications In silico hiPSC‐CMs and hAdultV‐CMs exhibit a similar response to selective current blocks. However, overall hiPSC‐CMs show greater sensitivity to block, which may facilitate in vitro identification of drug‐induced effects. Extrapolation of drug effects from hiPSC‐CM to hAdultV‐CM and pro‐arrhythmic risk assessment can be facilitated by in silico predictions using biophysically‐based computational models. PMID:26276951

  5. In situ characterization of the mTORC1 during adipogenesis of human adult stem cells on chip.

    PubMed

    Wu, Xuanye; Schneider, Nils; Platen, Alina; Mitra, Indranil; Blazek, Matthias; Zengerle, Roland; Schüle, Roland; Meier, Matthias

    2016-07-19

    Mammalian target of rapamycin (mTOR) is a central kinase integrating nutrient, energy, and metabolite signals. The kinase forms two distinct complexes: mTORC1 and mTORC2. mTORC1 plays an essential but undefined regulatory function for regeneration of adipose tissue. Analysis of mTOR in general is hampered by the complexity of regulatory mechanisms, including protein interactions and/or phosphorylation, in an ever-changing cellular microenvironment. Here, we developed a microfluidic large-scale integration chip platform for culturing and differentiating human adipose-derived stem cells (hASCs) in 128 separated microchambers under standardized nutrient conditions over 3 wk. The progression of the stem cell differentiation was measured by determining the lipid accumulation rates in hASC cultures. For in situ protein analytics, we developed a multiplex in situ proximity ligation assay (mPLA) that can detect mTOR in its two complexes selectively in single cells and implemented it on the same chip. With this combined technology, it was possible to reveal that the mTORC1 is regulated in its abundance, phosphorylation state, and localization in coordination with lysosomes during adipogenesis. High-content image analysis and parameterization of the in situ PLA signals in over 1 million cells cultured on four individual chips showed that mTORC1 and lysosomes are temporally and spatially coordinated but not in its composition during adipogenesis. PMID:27382182

  6. In situ characterization of the mTORC1 during adipogenesis of human adult stem cells on chip.

    PubMed

    Wu, Xuanye; Schneider, Nils; Platen, Alina; Mitra, Indranil; Blazek, Matthias; Zengerle, Roland; Schüle, Roland; Meier, Matthias

    2016-07-19

    Mammalian target of rapamycin (mTOR) is a central kinase integrating nutrient, energy, and metabolite signals. The kinase forms two distinct complexes: mTORC1 and mTORC2. mTORC1 plays an essential but undefined regulatory function for regeneration of adipose tissue. Analysis of mTOR in general is hampered by the complexity of regulatory mechanisms, including protein interactions and/or phosphorylation, in an ever-changing cellular microenvironment. Here, we developed a microfluidic large-scale integration chip platform for culturing and differentiating human adipose-derived stem cells (hASCs) in 128 separated microchambers under standardized nutrient conditions over 3 wk. The progression of the stem cell differentiation was measured by determining the lipid accumulation rates in hASC cultures. For in situ protein analytics, we developed a multiplex in situ proximity ligation assay (mPLA) that can detect mTOR in its two complexes selectively in single cells and implemented it on the same chip. With this combined technology, it was possible to reveal that the mTORC1 is regulated in its abundance, phosphorylation state, and localization in coordination with lysosomes during adipogenesis. High-content image analysis and parameterization of the in situ PLA signals in over 1 million cells cultured on four individual chips showed that mTORC1 and lysosomes are temporally and spatially coordinated but not in its composition during adipogenesis.

  7. Invariant natural killer T cells generated from human adult hematopoietic stem-progenitor cells are poly-functional.

    PubMed

    Sun, Wenji; Wang, Yi; East, James E; Kimball, Amy S; Tkaczuk, Katherine; Kesmodel, Susan; Strome, Scott E; Webb, Tonya J

    2015-03-01

    Invariant natural killer T (iNKT) cells constitute an important subset of T cells that can both directly and indirectly mediate anti-tumor immunity. However, cancer patients have a reduction in both iNKT cell number and function, and these deficits limit the potential clinical application of iNKT cells for cancer therapy. To overcome the problem of limited iNKT cell numbers, we investigated whether iNKT cells can be generated in vitro from bone marrow-derived adult hematopoietic stem-progenitor cells (HSPC). Our data demonstrate that co-culture of HSPC with OP9-DL1 stromal cells, results in a functional CD3(+) T cell population. These T cells can be further differentiated into iNKT cells by secondary culture with CD1d-Ig-based artificial antigen-presenting cells (aAPC). Importantly, these in vitro-generated iNKT cells are functional, as demonstrated by their ability to proliferate and secrete IFN-γ and GM-CSF following stimulation. PMID:25569376

  8. Invariant natural killer T cells generated from human adult hematopoietic stem-progenitor cells are poly-functional

    PubMed Central

    Sun, Wenji; Wang, Yi; East, James E.; Kimball, Amy S.; Tkaczuk, Katherine; Kesmodel, Susan; Strome, Scott E.; Webb, Tonya J.

    2014-01-01

    Invariant natural killer T (iNKT) cells constitute an important subset of T cells that can both directly and indirectly mediate anti-tumor immunity. However, cancer patients have a reduction in both iNKT cell number and function, and these deficits limit the potential clinical application of iNKT cells for cancer therapy. To overcome the problem of limited iNKT cell numbers, we investigated whether iNKT cells can be generated in vitro from bone marrow-derived adult hematopoietic stem-progenitor cells (HSPC). Our data demonstrate that co-culture of HSPC with OP9-DL1 stromal cells, results in a functional CD3+ T cell population. These T cells can be further differentiated into iNKT cells by secondary culture with CD1d-Ig-based artificial antigen-presenting cells (aAPC). Importantly, these in vitro-generated iNKT cells are functional, as demonstrated by their ability to proliferate and secrete IFN-γ and GM-CSF following stimulation. PMID:25569376

  9. Adult Stem Cells and Diseases of Aging

    PubMed Central

    Boyette, Lisa B.; Tuan, Rocky S.

    2014-01-01

    Preservation of adult stem cells pools is critical for maintaining tissue homeostasis into old age. Exhaustion of adult stem cell pools as a result of deranged metabolic signaling, premature senescence as a response to oncogenic insults to the somatic genome, and other causes contribute to tissue degeneration with age. Both progeria, an extreme example of early-onset aging, and heritable longevity have provided avenues to study regulation of the aging program and its impact on adult stem cell compartments. In this review, we discuss recent findings concerning the effects of aging on stem cells, contributions of stem cells to age-related pathologies, examples of signaling pathways at work in these processes, and lessons about cellular aging gleaned from the development and refinement of cellular reprogramming technologies. We highlight emerging therapeutic approaches to manipulation of key signaling pathways corrupting or exhausting adult stem cells, as well as other approaches targeted at maintaining robust stem cell pools to extend not only lifespan but healthspan. PMID:24757526

  10. Adult Stem Cell Responses to Nanostimuli

    PubMed Central

    Tsimbouri, Penelope M.

    2015-01-01

    Adult or mesenchymal stem cells (MSCs) have been found in different tissues in the body, residing in stem cell microenvironments called “stem cell niches”. They play different roles but their main activity is to maintain tissue homeostasis and repair throughout the lifetime of an organism. Their ability to differentiate into different cell types makes them an ideal tool to study tissue development and to use them in cell-based therapies. This differentiation process is subject to both internal and external forces at the nanoscale level and this response of stem cells to nanostimuli is the focus of this review. PMID:26193326

  11. Nop2 is expressed during proliferation of neural stem cells and in adult mouse and human brain.

    PubMed

    Kosi, Nina; Alić, Ivan; Kolačević, Matea; Vrsaljko, Nina; Jovanov Milošević, Nataša; Sobol, Margarita; Philimonenko, Anatoly; Hozák, Pavel; Gajović, Srećko; Pochet, Roland; Mitrečić, Dinko

    2015-02-01

    The nucleolar protein 2 gene encodes a protein specific for the nucleolus. It is assumed that it plays a role in the synthesis of ribosomes and regulation of the cell cycle. Due to its link to cell proliferation, higher expression of Nop2 indicates a worse tumor prognosis. In this work we used Nop2(gt1gaj) gene trap mouse strain. While lethality of homozygous animals suggested a vital role of this gene, heterozygous animals allowed the detection of expression of Nop2 in various tissues, including mouse brain. Histochemistry, immunohistochemistry and immunoelectron microscopy techniques, applied to a mature mouse brain, human brain and on mouse neural stem cells revealed expression of Nop2 in differentiating cells, including astrocytes, as well as in mature neurons. Nop2 was detected in various regions of mouse and human brain, mostly in large pyramidal neurons. In the human, Nop2 was strongly expressed in supragranular and infragranular layers of the somatosensory cortex and in layer III of the cingulate cortex. Also, Nop2 was detected in CA1 and the subiculum of the hippocampus. Subcellular analyses revealed predominant location of Nop2 within the dense fibrillar component of the nucleolus. To test if Nop2 expression correlates to cell proliferation occurring during tissue regeneration, we induced strokes in mice by middle cerebral artery occlusion. Two weeks after stroke, the number of Nop2/nestin double positive cells in the region affected by ischemia and the periventricular zone substantially increased. Our findings suggest a newly discovered role of Nop2 in both mature neurons and in cells possibly involved in the regeneration of nervous tissue.

  12. Comparative Analysis of AhR-Mediated TCDD-Elicited Gene Expression in Human Liver Adult Stem Cells

    PubMed Central

    Kim, Suntae; Dere, Edward; Burgoon, Lyle D.; Chang, Chia-Cheng; Zacharewski, Timothy R.

    2009-01-01

    Time course and dose-response studies were conducted in HL1-1 cells, a human liver cell line with stem cell–like characteristics, to assess the differential gene expression elicited by 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) compared with other established models. Cells were treated with 0.001, 0.01, 0.1, 1, 10, or 100nM TCDD or dimethyl sulfoxide vehicle control for 12 h for the dose-response study, or with 10nM TCDD or vehicle for 1, 2, 4, 8, 12, 24, or 48 h for the time course study. Elicited changes were monitored using a human cDNA microarray with 6995 represented genes. Empirical Bayes analysis identified 144 genes differentially expressed at one or more time points following treatment. Most genes exhibited dose-dependent responses including CYP1A1, CYP1B1, ALDH1A3, and SLC7A5 genes. Comparative analysis of HL1-1 differential gene expression to human HepG2 data identified 74 genes with comparable temporal expression profiles including 12 putative primary responses. HL1-1–specific changes were related to lipid metabolism and immune responses, consistent with effects elicited in vivo. Furthermore, comparative analysis of HL1-1 cells with mouse Hepa1c1c7 hepatoma cell lines and C57BL/6 hepatic tissue identified 18 and 32 commonly regulated orthologous genes, respectively, with functions associated with signal transduction, transcriptional regulation, metabolism and transport. Although some common pathways are affected, the results suggest that TCDD elicits species- and model-specific gene expression profiles. PMID:19684285

  13. Emerging restorative treatments for Parkinson's disease: manipulation and inducement of dopaminergic neurons from adult stem cells.

    PubMed

    Zhao, Junpeng; Xu, Qunyuan

    2011-06-01

    Parkinson's disease (PD) is a common neurodegenerative disease, characterized by a selective loss of midbrain Dopaminergic (DA) neurons. To address this problem, various types of stem cells that have potential to differentiate into DA neurons are being investigated as cellular therapies for PD, including cells derived from embryonic or adult donor tissue, and embryonic stem cells. These cell sources, however, have raised certain questions with regard to ethical and rejection issues. Recent progress in adult stems has further proved that the cells derived from adult tissue could be expanded and differentiated into DA precursor cells in vitro, and cell therapy with adult stem cells could produce a clear improvement for PD models. Using adult stem cells for clinic application may not only overcome the ethical problem inherent in using human fetal tissue or embryonic stem cells, but also open the possibility for autologous transplantation. The patient-specific adult stem cell is therefore a potential and prospective candidate for PD treatment.

  14. Human fetal cardiac progenitors: The role of stem cells and progenitors in the fetal and adult heart.

    PubMed

    Bulatovic, Ivana; Månsson-Broberg, Agneta; Sylvén, Christer; Grinnemo, Karl-Henrik

    2016-02-01

    The human fetal heart is formed early during embryogenesis as a result of cell migrations, differentiation, and formative blood flow. It begins to beat around gestation day 22. Progenitor cells are derived from mesoderm (endocardium and myocardium), proepicardium (epicardium and coronary vessels), and neural crest (heart valves, outflow tract septation, and parasympathetic innervation). A variety of molecular disturbances in the factors regulating the specification and differentiation of these cells can cause congenital heart disease. This review explores the contribution of different cardiac progenitors to the embryonic heart development; the pathways and transcription factors guiding their expansion, migration, and functional differentiation; and the endogenous regenerative capacity of the adult heart including the plasticity of cardiomyocytes. Unfolding these mechanisms will become the basis for understanding the dynamics of specific congenital heart disease as well as a means to develop therapy for fetal as well as postnatal cardiac defects and heart failure.

  15. The significance of the host inflammatory response on the therapeutic efficacy of cell therapies utilising human adult stem cells

    SciTech Connect

    Navarro, Melba; Pu, Fanrong; Hunt, John A.

    2012-02-15

    Controlling the fate of implanted hMSCs is one of the major drawbacks to be overcome to realize tissue engineering strategies. In particular, the effect of the inflammatory environment on hMSCs behaviour is poorly understood. Studying and mimicking the inflammatory process in vitro is a very complex and challenging task that involves multiple variables. This research addressed the questions using in vitro co-cultures of primary derived hMSCs together with human peripheral blood mononucleated cells (PBMCs); the latter are key agents in the inflammatory process. This work explored the in vitro phenotypic changes of hMSCs in co-culture direct contact with monocytes and lymphocytes isolated from blood using both basal and osteogenic medium. Our findings indicated that hMSCs maintained their undifferentiated phenotype and pluripotency despite the contact with PBMCs. Moreover, hMSCs demonstrated increased proliferation and were able to differentiate specifically down the osteogenic lineage pathway. Providing significant crucial evidence to support the hypothesis that inflammation and host defence mechanisms could be utilised rather than avoided and combated to provide for the successful therapeutic application of stem cell therapies.

  16. Ectopically hTERT expressing adult human mesenchymal stem cells are less radiosensitive than their telomerase negative counterpart

    SciTech Connect

    Serakinci, Nedime . E-mail: nserakinci@health.sdu.dk; Christensen, Rikke; Graakjaer, Jesper; Cairney, Claire J.; Keith, W. Nicol; Alsner, Jan; Saretzki, Gabriele; Kolvraa, Steen

    2007-03-10

    During the past several years increasing evidence indicating that the proliferation capacity of mammalian cells is highly radiosensitive, regardless of the species and the tissue of origin of the cells, has accumulated. It has also been shown that normal bone marrow cells of mice have a similar radiosensitivity to other mammalian cells so far tested. In this study, we investigated the genetic effects of ionizing radiation (2.5-15 Gy) on normal human mesenchymal stem cells and their telomerised counterpart hMSC-telo1. We evaluated overall genomic integrity, DNA damage/repair by applying a fluorescence-detected alkaline DNA unwinding assay together with Western blot analyses for phosphorylated H2AX and Q-FISH was applied for investigation of telomeric damage. Our results indicate that hMSC and TERT-immortalized hMSCs can cope with relatively high doses of {gamma}-rays and that overall DNA repair is similar in the two cell lines. The telomeres were extensively destroyed after irradiation in both cell types suggesting that telomere caps are especially sensitive to radiation. The TERT-immortalized hMSCs showed higher stability at telomeric regions than primary hMSCs indicating that cells with long telomeres and high telomerase activity have the advantage of re-establishing the telomeric caps.

  17. Synthesis of calcium phosphate-zirconia scaffold and human endometrial adult stem cells for bone tissue engineering.

    PubMed

    Alizadeh, Aliakbar; Moztarzadeh, Fathollah; Ostad, Seyed Naser; Azami, Mahmoud; Geramizadeh, Bita; Hatam, Gholamreza; Bizari, Davood; Tavangar, Seyed Mohammad; Vasei, Mohammad; Ai, Jafar

    2016-01-01

    To address the hypothesis that using a zirconia (ZrO2)/ β-tricalcium phosphate (β-TCP) composite might improve both the mechanical properties and cellular compatibility of the porous material, we fabricated ZrO2/β-TCP composite scaffolds with different ZrO2/β-TCP ratios, and evaluated their physical and mechanical characteristics, also the effect of three-dimensional (3D) culture (ZrO2/β-TCP scaffold) on the behavior of human endometrial stem cells. Results showed the porosity of a ZrO2/β-TCP scaffold can be adjusted from 65% to 84%, and the compressive strength of the scaffold increased from 4.95 to 6.25 MPa when the ZrO2 content increased from 30 to 50 wt%. The cell adhesion and proliferation in the ZrO2/β-TCP scaffold was greatly improved when ZrO2 decreased. Moreover, in vitro study showed that an osteoblasts-loaded ZrO2/β-TCP scaffold provided a suitable 3D environment for osteoblast survival and enhanced bone regeneration. We thus showed that a porous ZrO2/β-TCP composite scaffold has excellent mechanical properties, and cellular/tissue compatibility, and would be a promising substrate to achieve both bone reconstruction and regeneration needed during in vivo study for treatment of large bone defects.

  18. A facile method to establish human induced pluripotent stem cells from adult blood cells under feeder-free and xeno-free culture conditions: a clinically compliant approach.

    PubMed

    Chou, Bin-Kuan; Gu, Haihui; Gao, Yongxing; Dowey, Sarah N; Wang, Ying; Shi, Jun; Li, Yanxin; Ye, Zhaohui; Cheng, Tao; Cheng, Linzhao

    2015-04-01

    Reprogramming human adult blood mononuclear cells (MNCs) cells by transient plasmid expression is becoming increasingly popular as an attractive method for generating induced pluripotent stem (iPS) cells without the genomic alteration caused by genome-inserting vectors. However, its efficiency is relatively low with adult MNCs compared with cord blood MNCs and other fetal cells and is highly variable among different adult individuals. We report highly efficient iPS cell derivation under clinically compliant conditions via three major improvements. First, we revised a combination of three EBNA1/OriP episomal vectors expressing five transgenes, which increased reprogramming efficiency by ≥10-50-fold from our previous vectors. Second, human recombinant vitronectin proteins were used as cell culture substrates, alleviating the need for feeder cells or animal-sourced proteins. Finally, we eliminated the previously critical step of manually picking individual iPS cell clones by pooling newly emerged iPS cell colonies. Pooled cultures were then purified based on the presence of the TRA-1-60 pluripotency surface antigen, resulting in the ability to rapidly expand iPS cells for subsequent applications. These new improvements permit a consistent and reliable method to generate human iPS cells with minimal clonal variations from blood MNCs, including previously difficult samples such as those from patients with paroxysmal nocturnal hemoglobinuria. In addition, this method of efficiently generating iPS cells under feeder-free and xeno-free conditions allows for the establishment of clinically compliant iPS cell lines for future therapeutic applications.

  19. A facile method to establish human induced pluripotent stem cells from adult blood cells under feeder-free and xeno-free culture conditions: a clinically compliant approach.

    PubMed

    Chou, Bin-Kuan; Gu, Haihui; Gao, Yongxing; Dowey, Sarah N; Wang, Ying; Shi, Jun; Li, Yanxin; Ye, Zhaohui; Cheng, Tao; Cheng, Linzhao

    2015-04-01

    Reprogramming human adult blood mononuclear cells (MNCs) cells by transient plasmid expression is becoming increasingly popular as an attractive method for generating induced pluripotent stem (iPS) cells without the genomic alteration caused by genome-inserting vectors. However, its efficiency is relatively low with adult MNCs compared with cord blood MNCs and other fetal cells and is highly variable among different adult individuals. We report highly efficient iPS cell derivation under clinically compliant conditions via three major improvements. First, we revised a combination of three EBNA1/OriP episomal vectors expressing five transgenes, which increased reprogramming efficiency by ≥10-50-fold from our previous vectors. Second, human recombinant vitronectin proteins were used as cell culture substrates, alleviating the need for feeder cells or animal-sourced proteins. Finally, we eliminated the previously critical step of manually picking individual iPS cell clones by pooling newly emerged iPS cell colonies. Pooled cultures were then purified based on the presence of the TRA-1-60 pluripotency surface antigen, resulting in the ability to rapidly expand iPS cells for subsequent applications. These new improvements permit a consistent and reliable method to generate human iPS cells with minimal clonal variations from blood MNCs, including previously difficult samples such as those from patients with paroxysmal nocturnal hemoglobinuria. In addition, this method of efficiently generating iPS cells under feeder-free and xeno-free conditions allows for the establishment of clinically compliant iPS cell lines for future therapeutic applications. PMID:25742692

  20. PGE2 maintains self-renewal of human adult stem cells via EP2-mediated autocrine signaling and its production is regulated by cell-to-cell contact

    PubMed Central

    Lee, Byung-Chul; Kim, Hyung-Sik; Shin, Tae-Hoon; Kang, Insung; Lee, Jin Young; Kim, Jae-Jun; Kang, Hyun Kyoung; Seo, Yoojin; Lee, Seunghee; Yu, Kyung-Rok; Choi, Soon Won; Kang, Kyung-Sun

    2016-01-01

    Mesenchymal stem cells (MSCs) possess unique immunomodulatory abilities. Many studies have elucidated the clinical efficacy and underlying mechanisms of MSCs in immune disorders. Although immunoregulatory factors, such as Prostaglandin E2 (PGE2), and their mechanisms of action on immune cells have been revealed, their effects on MSCs and regulation of their production by the culture environment are less clear. Therefore, we investigated the autocrine effect of PGE2 on human adult stem cells from cord blood or adipose tissue, and the regulation of its production by cell-to-cell contact, followed by the determination of its immunomodulatory properties. MSCs were treated with specific inhibitors to suppress PGE2 secretion, and proliferation was assessed. PGE2 exerted an autocrine regulatory function in MSCs by triggering E-Prostanoid (EP) 2 receptor. Inhibiting PGE2 production led to growth arrest, whereas addition of MSC-derived PGE2 restored proliferation. The level of PGE2 production from an equivalent number of MSCs was down-regulated via gap junctional intercellular communication. This cell contact-mediated decrease in PGE2 secretion down-regulated the suppressive effect of MSCs on immune cells. In conclusion, PGE2 produced by MSCs contributes to maintenance of self-renewal capacity through EP2 in an autocrine manner, and PGE2 secretion is down-regulated by cell-to-cell contact, attenuating its immunomodulatory potency. PMID:27230257

  1. Stem Cell Basics

    MedlinePlus

    ... stem cells? What are the potential uses of human stem cells and the obstacles that must be overcome before ... two kinds of stem cells from animals and humans: embryonic stem cells and non-embryonic "somatic" or "adult" stem cells . ...

  2. Isolation, characterization, and differentiation of human multipotent dermal stem cells.

    PubMed

    Li, Ling; Fukunaga-Kalabis, Mizuho; Herlyn, Meenhard

    2013-01-01

    Skin, as the body's largest organ, has been extensively used to study adult stem cells. Most previous skin-related studies have focused on stem cells isolated from hair follicles and from keratinocytes. Here we present a protocol to isolate multipotent neural crest stem-like dermis-derived stem cells (termed dermal stem cells or DSCs) from human neonatal foreskins. DSCs grow like neural spheres in human embryonic stem cell medium and gain the ability to self-renew and differentiate into several cell lineages including melanocytes, neuronal cells, Schwann cells, smooth muscle cells, adipocytes, and chondrocytes. These cells express neural crest stem cell markers (NGFRp75 and nestin) as well as an embryonic stem cell marker (OCT4).

  3. Embryonic and adult stem cell therapy.

    PubMed

    Brignier, Anne C; Gewirtz, Alan M

    2010-02-01

    There are many types of stem cells. All share the characteristics of being able to self-renew and to give rise to differentiated progeny. Over the last decades, great excitement has been generated by the prospect of being able to exploit these properties for the repair, improvement, and/or replacement of damaged organs. However, many hurdles, both scientific and ethical, remain in the path of using human embryonic stem cells for tissue-engineering purposes. In this report we review current strategies for isolating, enriching, and, most recently, inducing the development of human pluripotent stem cells. In so doing, we discuss the scientific and ethical issues associated with this endeavor. Finally, progress in the use of stem cells as therapies for type 1 diabetes mellitus, congestive heart failure, and various neurologic and immunohematologic disorders, and as vehicles for the delivery of gene therapy, is briefly discussed. PMID:20061008

  4. Wnt signaling in adult intestinal stem cells and cancer.

    PubMed

    Krausova, Michaela; Korinek, Vladimir

    2014-03-01

    Signaling initiated by secreted glycoproteins of the Wnt family regulates many aspects of embryonic development and it is involved in homeostasis of adult tissues. In the gastrointestinal (GI) tract the Wnt pathway maintains the self-renewal capacity of epithelial stem cells. The stem cell attributes are conferred by mutual interactions of the stem cell with its local microenvironment, the stem cell niche. The niche ensures that the threshold of Wnt signaling in the stem cell is kept in physiological range. In addition, the Wnt pathway involves various feedback loops that balance the opposing processes of cell proliferation and differentiation. Today, we have compelling evidence that mutations causing aberrant activation of the Wnt pathway promote expansion of undifferentiated progenitors and lead to cancer. The review summarizes recent advances in characterization of adult epithelial stem cells in the gut. We mainly focus on discoveries related to molecular mechanisms regulating the output of the Wnt pathway. Moreover, we present novel experimental approaches utilized to investigate the epithelial cell signaling circuitry in vivo and in vitro. Pivotal aspects of tissue homeostasis are often deduced from studies of tumor cells; therefore, we also discuss some latest results gleaned from the deep genome sequencing studies of human carcinomas of the colon and rectum. PMID:24308963

  5. Adult Stem Cell Therapy for Stroke: Challenges and Progress

    PubMed Central

    Bang, Oh Young; Kim, Eun Hee; Cha, Jae Min; Moon, Gyeong Joon

    2016-01-01

    Stroke is one of the leading causes of death and physical disability among adults. It has been 15 years since clinical trials of stem cell therapy in patients with stroke have been conducted using adult stem cells like mesenchymal stem cells and bone marrow mononuclear cells. Results of randomized controlled trials showed that adult stem cell therapy was safe but its efficacy was modest, underscoring the need for new stem cell therapy strategies. The primary limitations of current stem cell therapies include (a) the limited source of engraftable stem cells, (b) the presence of optimal time window for stem cell therapies, (c) inherited limitation of stem cells in terms of growth, trophic support, and differentiation potential, and (d) possible transplanted cell-mediated adverse effects, such as tumor formation. Here, we discuss recent advances that overcome these hurdles in adult stem cell therapy for stroke. PMID:27733032

  6. Neural Crest As the Source of Adult Stem Cells

    PubMed Central

    Pierret, Chris; Spears, Kathleen; Maruniak, Joel A.; Kirk, Mark D.

    2012-01-01

    Recent studies suggest that adult stem cells can cross germ layer boundaries. For example, bone marrow-derived stem cells appear to differentiate into neurons and glial cells, as well as other types of cells. How can stem cells from bone marrow, pancreas, skin, or fat become neurons and glia; in other words, what molecular and cellular events direct mesodermal cells to a neural fate? Transdifferentiation, dediffereniation, and fusion of donor adult stem cells with fully differentiated host cells have been proposed to explain the plasticity of adult stem cells. Here we review the origin of select adult stem cell populations and propose a unifying hypothesis to explain adult stem cell plasticity. In addition, we outline specific experiments to test our hypothesis. We propose that peripheral, tissue-derived, or adult stem cells are all progeny of the neural crest. PMID:16646675

  7. DNA damage response in adult stem cells.

    PubMed

    Insinga, Alessandra; Cicalese, Angelo; Pelicci, Pier Giuseppe

    2014-04-01

    This review discusses the processes of DNA-damage-response and DNA-damage repair in stem and progenitor cells of several tissues. The long life-span of stem cells suggests that they may respond differently to DNA damage than their downstream progeny and, indeed, studies have begun to elucidate the unique stem cell response mechanisms to DNA damage. Because the DNA damage responses in stem cells and progenitor cells are distinctly different, stem and progenitor cells should be considered as two different entities from this point of view. Hematopoietic and mammary stem cells display a unique DNA-damage response, which involves active inhibition of apoptosis, entry into the cell-cycle, symmetric division, partial DNA repair and maintenance of self-renewal. Each of these biological events depends on the up-regulation of the cell-cycle inhibitor p21. Moreover, inhibition of apoptosis and symmetric stem cell division are the consequence of the down-regulation of the tumor suppressor p53, as a direct result of p21 up-regulation. A deeper understanding of these processes is required before these findings can be translated into human anti-aging and anti-cancer therapies. One needs to clarify and dissect the pathways that control p21 regulation in normal and cancer stem cells and define (a) how p21 blocks p53 functions in stem cells and (b) how p21 promotes DNA repair in stem cells. Is this effect dependent on p21s ability to inhibit p53? Such molecular knowledge may pave the way to methods for maintaining short-term tissue reconstitution while retaining long-term cellular and genomic integrity.

  8. Immunological control of adult neural stem cells

    PubMed Central

    Gonzalez-Perez, Oscar; Quiñones-Hinojosa, Alfredo; Garcia-Verdugo, Jose Manuel

    2010-01-01

    Adult neurogenesis occurs only in discrete regions of adult central nervous system: the subventricular zone and the subgranular zone. These areas are populated by adult neural stem cells (aNSC) that are regulated by a number of molecules and signaling pathways, which control their cell fate choices, survival and proliferation rates. For a long time, it was believed that the immune system did not exert any control on neural proliferative niches. However, it has been observed that many pathological and inflammatory conditions significantly affect NSC niches. Even more, increasing evidence indicates that chemokines and cytokines play an important role in regulating proliferation, cell fate choices, migration and survival of NSCs under physiological conditions. Hence, the immune system is emerging is an important regulator of neurogenic niches in the adult brain, which may have clinical relevance in several brain diseases. PMID:20861925

  9. Adult neural stem cells stake their ground

    PubMed Central

    Lim, Daniel A.; Alvarez-Buylla, Arturo

    2014-01-01

    The birth of new neurons in the walls of the adult brain lateral ventricles has captured the attention of many neuroscientists for over two decades, yielding key insights into the identity and regulation of neural stem cells (NSCs). In the adult ventricular-subventricular zone (V-SVZ), NSCs are a specialized form of astrocyte that generates several types of neurons for the olfactory bulb. Here we discuss recent findings regarding the unique organization of the V-SVZ NSCs niche, the multiple regulatory controls of neuronal production, the distinct regional identities of adult NSCs, and the epigenetic mechanisms that maintain adult neurogenesis. Understanding how V-SVZ NSCs establish and maintain lifelong neurogenesis continues to provide surprising insights into the cellular and molecular regulation of neural development. PMID:25223700

  10. Adult Human Biliary Tree Stem Cells Differentiate to β-Pancreatic Islet Cells by Treatment with a Recombinant Human Pdx1 Peptide

    PubMed Central

    Scafetta, Gaia; Renzi, Anastasia; De Canio, Michele; Sicilia, Francesca; Nevi, Lorenzo; Casa, Domenico; Panetta, Rocco; Berloco, Pasquale Bartolomeo; Reid, Lola M.; Federici, Giorgio; Gaudio, Eugenio; Maroder, Marella; Alvaro, Domenico

    2015-01-01

    Generation of β-pancreatic cells represents a major goal in research. The aim of this study was to explore a protein-based strategy to induce differentiation of human biliary tree stem cells (hBTSCs) towards β-pancreatic cells. A plasmid containing the sequence of the human pancreatic and duodenal homeobox 1 (PDX1) has been expressed in E. coli. Epithelial-Cell-Adhesion-Molecule positive hBTSCs or mature human hepatocyte cell line, HepG2, were grown in medium to which Pdx1 peptide was added. Differentiation toward pancreatic islet cells were evaluated by the expression of the β-cell transcription factors, Pdx1 and musculoapo-neurotic fibrosarcoma oncogene homolog A, and of the pancreatic hormones, insulin, glucagon, and somatostatin, investigated by real time polymerase chain reaction, western blot, light microscopy and immunofluorescence. C-peptide secretion in response to high glucose was also measured. Results indicated how purified Pdx1 protein corresponding to the primary structure of the human Pdx1 by mass spectroscopy was efficiently produced in bacteria, and transduced into hBTSCs. Pdx1 exposure triggered the expression of both intermediate and mature stage β-cell differentiation markers only in hBTSCs but not in HepG2 cell line. Furthermore, hBTSCs exposed to Pdx1 showed up-regulation of insulin, glucagon and somatostatin genes and formation of 3-dimensional islet-like structures intensely positive for insulin and glucagon. Finally, Pdx1-induced islet-like structures exhibited glucose-regulated C-peptide secretion. In conclusion, the human Pdx1 is highly effective in triggering hBTSC differentiation toward functional β-pancreatic cells. PMID:26252949

  11. Adult Human Biliary Tree Stem Cells Differentiate to β-Pancreatic Islet Cells by Treatment with a Recombinant Human Pdx1 Peptide.

    PubMed

    Cardinale, Vincenzo; Puca, Rosa; Carpino, Guido; Scafetta, Gaia; Renzi, Anastasia; De Canio, Michele; Sicilia, Francesca; Nevi, Lorenzo; Casa, Domenico; Panetta, Rocco; Berloco, Pasquale Bartolomeo; Reid, Lola M; Federici, Giorgio; Gaudio, Eugenio; Maroder, Marella; Alvaro, Domenico

    2015-01-01

    Generation of β-pancreatic cells represents a major goal in research. The aim of this study was to explore a protein-based strategy to induce differentiation of human biliary tree stem cells (hBTSCs) towards β-pancreatic cells. A plasmid containing the sequence of the human pancreatic and duodenal homeobox 1 (PDX1) has been expressed in E. coli. Epithelial-Cell-Adhesion-Molecule positive hBTSCs or mature human hepatocyte cell line, HepG2, were grown in medium to which Pdx1 peptide was added. Differentiation toward pancreatic islet cells were evaluated by the expression of the β-cell transcription factors, Pdx1 and musculoapo-neurotic fibrosarcoma oncogene homolog A, and of the pancreatic hormones, insulin, glucagon, and somatostatin, investigated by real time polymerase chain reaction, western blot, light microscopy and immunofluorescence. C-peptide secretion in response to high glucose was also measured. Results indicated how purified Pdx1 protein corresponding to the primary structure of the human Pdx1 by mass spectroscopy was efficiently produced in bacteria, and transduced into hBTSCs. Pdx1 exposure triggered the expression of both intermediate and mature stage β-cell differentiation markers only in hBTSCs but not in HepG2 cell line. Furthermore, hBTSCs exposed to Pdx1 showed up-regulation of insulin, glucagon and somatostatin genes and formation of 3-dimensional islet-like structures intensely positive for insulin and glucagon. Finally, Pdx1-induced islet-like structures exhibited glucose-regulated C-peptide secretion. In conclusion, the human Pdx1 is highly effective in triggering hBTSC differentiation toward functional β-pancreatic cells. PMID:26252949

  12. In vitro generation of mechanically functional cartilage grafts based on adult human stem cells and 3D-woven poly(ε-caprolactone) scaffolds

    PubMed Central

    Valonen, P.K.; Moutos, F.T.; Kusanagi, A.; Moretti, M.; Diekman, B.O.; Welter, J.F.; Caplan, A.I.; Guilak, F.

    2009-01-01

    Three-dimensionally woven poly(ε-caprolactone)(PCL) scaffolds were combined with adult human mesenchymal stem cells (hMSC) to engineer mechanically functional cartilage constructs in vitro. The specific objectives were to: (i) produce PCL scaffolds with cartilage-like mechanical properties, (ii) demonstrate that hMSCs formed cartilage after 21-days of culture on PCL scaffolds, and (iii) study effects of scaffold structure (loosely vs. tightly woven), culture vessel (static dish vs. oscillating bioreactor), and medium composition (chondrogenic additives with or without serum). Aggregate moduli of 21-day constructs approached normal articular cartilage for tightly woven PCL cultured in bioreactors, were lower for tightly woven PCL cultured statically, and lowest for loosely woven PCL cultured statically (p<0.05). Construct DNA, total collagen, and glyocosaminoglycans (GAG) increased in a manner dependent on time, culture vessel, and medium composition. Chondrogenesis was verified histologically by rounded cells within a hyaline-like matrix that immunostained for collagen type II but not type I. Bioreactors yielded constructs with higher collagen content (p<0.05) and more homogenous matrix than static controls. Chondrogenic additives yielded constructs with higher GAG (p<0.05) and earlier expression of collagen II mRNA if serum was not present in medium. These results show feasibility of functional cartilage tissue engineering from hMSC and 3D woven PCL scaffolds. PMID:20034665

  13. Bilateral transplantation of allogenic adult human bone marrow-derived mesenchymal stem cells into the subventricular zone of Parkinson's disease: a pilot clinical study.

    PubMed

    Venkataramana, N K; Pal, Rakhi; Rao, Shailesh A V; Naik, Arun L; Jan, Majahar; Nair, Rahul; Sanjeev, C C; Kamble, Ravindra B; Murthy, D P; Chaitanya, Krishna

    2012-01-01

    The progress of PD and its related disorders cannot be prevented with the medications available. In this study, we recruited 8 PD and 4 PD plus patients between 5 to 15 years after diagnosis. All patients received BM-MSCs bilaterally into the SVZ and were followed up for 12 months. PD patients after therapy reported a mean improvement of 17.92% during "on" and 31.21% during "off" period on the UPDRS scoring system. None of the patients increased their medication during the follow-up period. Subjectively, the patients reported clarity in speech, reduction in tremors, rigidity, and freezing attacks. The results correlated with the duration of the disease. Those patients transplanted in the early stages of the disease (less than 5 years) showed more improvement and no further disease progression than the later stages (11-15 years). However, the PD plus patients did not show any change in their clinical status after stem cell transplantation. This study demonstrates the safety of adult allogenic human BM-MSCs transplanted into the SVZ of the brain and its efficacy in early-stage PD patients. PMID:22550521

  14. Differentiation of embryonic and adult stem cells into insulin producing cells.

    PubMed

    Zulewski, H

    2008-03-01

    Replacement of insulin producing cells represents an almost ideal treatment for patients with diabetes mellitus type 1. Transplantation of pancreatic islets of Langerhans is successful in experienced centers. The wider application of this therapy, however, is limited by the lack of donor organs. Insulin producing cells generated from stem cells represent an attractive alternative. Stem cells with the potential to differentiate into insulin producing cells include embryonic stem cells (ESC) as well as adult stem cells from various tissues including the pancreas, liver, bone marrow and adipose tissue. The use of human ESC is hampered by ethical concerns but research with human ESC may help us to decipher important steps in the differentiation process in vitro since almost all information available on pancreas development are based on animal studies. The present review summarizes the current knowledge on the development of insulin producing cells from embryonic and adult stem cells with special emphasis on pancreatic, hepatic and human mesenchymal stem cells. PMID:18427390

  15. Embryonic Stem Cell Patents and Human Dignity

    PubMed Central

    Resnik, David B.

    2009-01-01

    This article examines the assertion that human embryonic stem cells patents are immoral because they violate human dignity. After analyzing the concept of human dignity and its role in bioethics debates, this article argues that patents on human embryos or totipotent embryonic stem cells violate human dignity, but that patents on pluripotent or multipotent stem cells do not. Since patents on pluripotent or multipotent stem cells may still threaten human dignity by encouraging people to treat embryos as property, patent agencies should carefully monitor and control these patents to ensure that patents are not inadvertently awarded on embryos or totipotent stem cells. PMID:17922198

  16. The effects of Ca2SiO4-Ca3(PO4)2 ceramics on adult human mesenchymal stem cell viability, adhesion, proliferation, differentiation and function.

    PubMed

    De Aza, Piedad N; García-Bernal, David; Cragnolini, Francesca; Velasquez, Pablo; Meseguer-Olmo, Luis

    2013-10-01

    Bioceramic samples with osteogenic properties, suitable for use in the regeneration of hard tissue, were synthesized. The materials consisting of α-tricalcium phosphate (αTCP) and also αTCP doped with either 1.5 wt.% or 3.0 wt.% of dicalcium silicate (C2S) in the system Dicalcium Silicate-Tricalcium Phosphate (C2S-TCP) were obtained by solid state reaction. All materials were composed of a single phase, αTCP in the case of a pure material, or solid solution of C2S in αTCP (αTCPss) for the doped αTCP. Viability, proliferation and in vitro osteoinductive capacity were investigated by seeding, adult mesenchymal stem cells of human origin (ahMSCs) which were CD73(+), CD90(+), CD105(+), CD34(-) and CD45(-) onto the 3 substrates for 30 days. Results show a non-cytotoxic effect after applying an indirect apoptosis test (Annexin V/7-AAD staining), so ahMSCs adhered, spread, proliferated and produced extracellular matrix (Heparan-sulfate proteoglycan (HS) and osteopontin (OP)) on all the ceramics studied. Finally, the cells lost the cluster differentiation marker expression CD73, CD90 y CD105 characteristic of ahMSCs and they showed an osteoblastic phenotype (Alkaline phosphatase activity (ALP), Osteocalcin production (OC), Collagen type I expression (Col-I), and production of mineralization nodules on the extracellular matrix). These observations were more evident in the αTCP ceramic doped with 1.5 wt.% C2S, indicating osteoblastic differentiation as a result of the increased concentration of solid solution of C2S in αTCP (αTCPss). Overall, these results suggest that the ceramics studied are cytocompatible and they are able to induce osteoblastic differentiation of undifferentiated ahMSCs.

  17. Isolation, identification and differentiation of human embryonic cartilage stem cells.

    PubMed

    Fu, Changhao; Yan, Zi; Xu, Hao; Zhang, Chen; Zhang, Qi; Wei, Anhui; Yang, Xi; Wang, Yi

    2015-07-01

    We isolated human embryonic cartilage stem cells (hECSCs), a novel stem cell population, from the articular cartilage of eight-week-old human embryos. These stem cells demonstrated a marker expression pattern and differentiation potential intermediate to those of human embryonic stem cells (hESCs) and human adult stem cells (hASCs). hECSCs expressed markers associated with both hESCs (OCT4, NANOG, SOX2, SSEA-3 and SSEA-4) and human adult stem cells (hASCs) (CD29, CD44, CD90, CD73 and CD10). These cells also differentiated into adipocytes, osteoblasts, chondrocytes, neurons and islet-like cells under specific inducing conditions. We identified N(6), 2'-O-dibutyryl cyclic adenosine 3':5'-monophosphate (Bt2cAMP) as an inducer of chondrogenic differentiation in hECSCs. Similar results using N(6), 2'-O-dibutyryl cyclic adenosine 3':5'-monophosphate (Bt2cAMP) were obtained for two other types of human embryonic tissue-derived stem cells, human embryonic hepatic stem cells (hEHSCs) and human embryonic amniotic fluid stem cells (hEASCs), both of which exhibited a marker expression pattern similar to that of hECSCs. The isolation of hECSCs and the discovery that N(6), 2'-O-dibutyryl cyclic adenosine 3':5'-monophosphate (Bt2cAMP) induces chondrogenic differentiation in different stem cell populations might aid the development of strategies in tissue engineering and cartilage repair.

  18. Spatial control of adult stem cell fate using nanotopographic cues

    PubMed Central

    An, Steven S.; Afzal, Junaid; Lee, Suengwon; Kwak, Moonkyu; Suh, Kahp-Yang; Levchenko, Andre

    2015-01-01

    Adult stem cells hold great promise as a source of diverse terminally differentiated cell types for tissue engineering applications. However, due to the complexity of chemical and mechanical cues specifying differentiation outcomes, development of arbitrarily complex geometric and structural arrangements of cells, adopting multiple fates from the same initial stem cell population, has been difficult. Here, we show that the topography of the cell adhesion substratum can be an instructive cue to adult stem cells and topographical variations can strongly bias the differentiation outcome of the cells towards adipocyte or osteocyte fates. Switches in cell fate decision from adipogenic to osteogenic lineages were accompanied by changes in cytoskeletal stiffness, spanning a considerable range in the cell softness/rigidity spectrum. Our findings suggest that human mesenchymal stem cells (hMSC) can respond to the varying density of nanotopographical cues by regulating their internal cytoskeletal network and use these mechanical changes to guide them toward making cell fate decisions. We used this finding to design a complex two-dimensional pattern of co-localized cells preferentially adopting two alternative fates, thus paving the road for designing and building more complex tissue constructs with diverse biomedical applications. PMID:24388388

  19. Human stem cell ethics: beyond the embryo.

    PubMed

    Sugarman, Jeremy

    2008-06-01

    Human embryonic stem cell research has elicited powerful debates about the morality of destroying human embryos. However, there are important ethical issues related to stem cell research that are unrelated to embryo destruction. These include particular issues involving different types of cells used, the procurement of such cells, in vivo use of stem cells, intellectual property, and conflicts of interest.

  20. Patterning as a signature of human epidermal stem cell regulation

    PubMed Central

    Klein, Allon M.; Nikolaidou-Neokosmidou, Varvara; Doupé, David P.; Jones, Philip H.; Simons, Benjamin D.

    2011-01-01

    Understanding how stem cells are regulated in adult tissues is a major challenge in cell biology. In the basal layer of human epidermis, clusters of almost quiescent stem cells are interspersed with proliferating and differentiating cells. Previous studies have shown that the proliferating cells follow a pattern of balanced stochastic cell fate. This behaviour enables them to maintain homeostasis, while stem cells remain confined to their quiescent clusters. Intriguingly, these clusters reappear spontaneously in culture, suggesting that they may play a functional role in stem cell auto-regulation. We propose a model of pattern formation that explains how clustering could regulate stem cell activity in homeostatic tissue through contact inhibition and stem cell aggregation. PMID:21632613

  1. Comparison of human fetal liver, umbilical cord blood, and adult blood hematopoietic stem cell engraftment in NOD-scid/gammac-/-, Balb/c-Rag1-/-gammac-/-, and C.B-17-scid/bg immunodeficient mice.

    PubMed

    Lepus, Christin M; Gibson, Thomas F; Gerber, Scott A; Kawikova, Ivana; Szczepanik, Marian; Hossain, Jaber; Ablamunits, Vitaly; Kirkiles-Smith, Nancy; Herold, Kevan C; Donis, Ruben O; Bothwell, Alfred L; Pober, Jordan S; Harding, Martha J

    2009-10-01

    Immunodeficient mice bearing components of a human immune system present a novel approach for studying human immune responses. We investigated the number, phenotype, developmental kinetics, and function of developing human immune cells following transfer of CD34(+) hematopoietic stem cell (HSC) preparations originating from second trimester human fetal liver (HFL), umbilical cord blood (UCB), or granulocyte colony-stimulating factor-mobilized adult blood (G-CSF-AB) delivered via intrahepatic injection into sublethally irradiated neonatal NOD-scid/gammac(-/-), Balb/c-Rag1(-/-)gammac(-/-), and C.B-17-scid/bg mice. HFL and UCB HSC provided the greatest number and breadth of developing cells. NOD-scid/gammac(-/-) and Balb/c-Rag1(-/-)gammac(-/-) harbored human B and dendritic cells as well as human platelets in peripheral blood, whereas NOD-scid/gammac(-/-) mice harbored higher levels of human T cells. NOD-scid/gammac(-/-) mice engrafted with HFL CD34(+) HSC demonstrated human immunological competence evidenced by white pulp expansion and increases in total human immunoglobulin following immunization with T-dependent antigens and delayed-type hypersensitivity-infiltrating leukocytes in response to antigenic challenge. In conclusion, we describe an encouraging base system for studying human hematopoietic lineage development and function utilizing human HFL or UCB HSC-engrafted NOD-scid/gammac(-/-) mice that is well suited for future studies toward the development of a fully competent humanized mouse model.

  2. Adipose-derived adult stem cells: available technologies for potential clinical regenerative applications in dentistry.

    PubMed

    Catalano, Enrico; Cochis, Andrea; Varoni, Elena; Rimondini, Lia; Carrassi, Antonio; Azzimonti, Barbara

    2013-01-01

    Tissue homeostasis depends closely on the activity and welfare of adult stem cells. These cells represent a promising tool for biomedical research since they can aid in treatment and promote the regeneration of damaged organs in many human disorders. Adult stem cells indefinitely preserve their ability to self-renew and differentiate into various phenotypes; this capacity could be promoted in vitro by particular culture conditions (differentiation media) or spontaneously induced in vivo by exploiting the biochemical and mechanical properties of the tissue in which the stem cells are implanted. Among the different sources of adult stem cells, adipose tissue is an attractive possibility thanks to its ready availability and the standard extraction techniques at our disposal today. This review discusses the isolation, characterization, and differentiation of human adipose-derived adult stem cells, as well as regeneration strategies, therapeutic uses, and adverse effects of their delivery. In particular, since oral disorders (e.g., trauma, erosion, and chronic periodontitis) often cause the loss of dental tissue along with functional, phonetic, and aesthetic impairment, this review focuses on the application of human adipose-derived adult stem cells, alone or in combination with biomaterials, in treating oral diseases.

  3. Stem cell differentiation and human liver disease

    PubMed Central

    Zhou, Wen-Li; Medine, Claire N; Zhu, Liang; Hay, David C

    2012-01-01

    Human stem cells are scalable cell populations capable of cellular differentiation. This makes them a very attractive in vitro cellular resource and in theory provides unlimited amounts of primary cells. Such an approach has the potential to improve our understanding of human biology and treating disease. In the future it may be possible to deploy novel stem cell-based approaches to treat human liver diseases. In recent years, efficient hepatic differentiation from human stem cells has been achieved by several research groups including our own. In this review we provide an overview of the field and discuss the future potential and limitations of stem cell technology. PMID:22563188

  4. [Progress in treating diabetes mellitus with adult stem cells].

    PubMed

    Zhang, Lixin; Teng, Chunbo; An, Tiezhu

    2008-02-01

    Diabetes mellitus is a metabolic diseases, mainly including type 1 and type 2 diabetes. Treatment for type 1 and part of type 2 often involves regular insulin injection. However, this treatment neither precisely controls the blood sugar levels, nor prevents the diabetes complications. Transplantation of islets of Langerhans offers an attractive strategy for diabetes therapies, but its wide application has been limited by donor shortage and immunological rejection after transplantation. Stem cells with strong proliferation capacity and multipotential may be potential cell sources in diabetes therapies. For this, adult stem cells are interesting because of absence of teratoma formation and ethnical problems. Adult pancreatic stem cells (PSCs) really exist and could produce insulin-secreting cells both under the condition of pancreatic injury and in vitro culture, but lack of effective markers to enrich PSCs hampers the studies of exploring the expanding and differentiating conditions in vitro. Some other adult stem cells, such as hepatic stem cells, marrow stem cells or intestine stem cells, were also suggested to transdifferentiate into insulin-producing cells under special culture conditions in vitro or by genetic modifications. Moreover, transplanting these adult stem cells-derived insulin-secreting cells into the diabetic mouse could cure diabetes. Thus, adult stem cells would supply the abundant beta-cell sources for cell replacement therapy of diabetes. PMID:18464596

  5. [Progress in treating diabetes mellitus with adult stem cells].

    PubMed

    Zhang, Lixin; Teng, Chunbo; An, Tiezhu

    2008-02-01

    Diabetes mellitus is a metabolic diseases, mainly including type 1 and type 2 diabetes. Treatment for type 1 and part of type 2 often involves regular insulin injection. However, this treatment neither precisely controls the blood sugar levels, nor prevents the diabetes complications. Transplantation of islets of Langerhans offers an attractive strategy for diabetes therapies, but its wide application has been limited by donor shortage and immunological rejection after transplantation. Stem cells with strong proliferation capacity and multipotential may be potential cell sources in diabetes therapies. For this, adult stem cells are interesting because of absence of teratoma formation and ethnical problems. Adult pancreatic stem cells (PSCs) really exist and could produce insulin-secreting cells both under the condition of pancreatic injury and in vitro culture, but lack of effective markers to enrich PSCs hampers the studies of exploring the expanding and differentiating conditions in vitro. Some other adult stem cells, such as hepatic stem cells, marrow stem cells or intestine stem cells, were also suggested to transdifferentiate into insulin-producing cells under special culture conditions in vitro or by genetic modifications. Moreover, transplanting these adult stem cells-derived insulin-secreting cells into the diabetic mouse could cure diabetes. Thus, adult stem cells would supply the abundant beta-cell sources for cell replacement therapy of diabetes.

  6. Human stem cells and articular cartilage regeneration.

    PubMed

    Inui, Atsuyuki; Iwakura, Takashi; Reddi, A Hari

    2012-11-05

    The regeneration of articular cartilage damaged due to trauma and posttraumatic osteoarthritis is an unmet medical need. Current approaches to regeneration and tissue engineering of articular cartilage include the use of chondrocytes, stem cells, scaffolds and signals, including morphogens and growth factors. Stem cells, as a source of cells for articular cartilage regeneration, are a critical factor for articular cartilage regeneration. This is because articular cartilage tissue has a low cell turnover and does not heal spontaneously. Adult stem cells have been isolated from various tissues, such as bone marrow, adipose, synovial tissue, muscle and periosteum. Signals of the transforming growth factor beta superfamily play critical roles in chondrogenesis. However, adult stem cells derived from various tissues tend to differ in their chondrogenic potential. Pluripotent stem cells have unlimited proliferative capacity compared to adult stem cells. Chondrogenesis from embryonic stem (ES) cells has been studied for more than a decade. However, establishment of ES cells requires embryos and leads to ethical issues for clinical applications. Induced pluripotent stem (iPS) cells are generated by cellular reprogramming of adult cells by transcription factors. Although iPS cells have chondrogenic potential, optimization, generation and differentiation toward articular chondrocytes are currently under intense investigation.

  7. Mitochondrial respiration regulates adipogenic differentiation of human mesenchymal stem cells.

    PubMed

    Zhang, Yanmin; Marsboom, Glenn; Toth, Peter T; Rehman, Jalees

    2013-01-01

    Human mesenchymal stem cells (MSCs) are adult multipotent stem cells which can be isolated from bone marrow, adipose tissue as well as other tissues and have the capacity to differentiate into a variety of mesenchymal cell types such as adipocytes, osteoblasts and chondrocytes. Differentiation of stem cells into mature cell types is guided by growth factors and hormones, but recent studies suggest that metabolic shifts occur during differentiation and can modulate the differentiation process. We therefore investigated mitochondrial biogenesis, mitochondrial respiration and the mitochondrial membrane potential during adipogenic differentiation of human MSCs. In addition, we inhibited mitochondrial function to assess its effects on adipogenic differentiation. Our data show that mitochondrial biogenesis and oxygen consumption increase markedly during adipogenic differentiation, and that reducing mitochondrial respiration by hypoxia or by inhibition of the mitochondrial electron transport chain significantly suppresses adipogenic differentiation. Furthermore, we used a novel approach to suppress mitochondrial activity using a specific siRNA-based knockdown of the mitochondrial transcription factor A (TFAM), which also resulted in an inhibition of adipogenic differentiation. Taken together, our data demonstrates that increased mitochondrial activity is a prerequisite for MSC differentiation into adipocytes. These findings suggest that metabolic modulation of adult stem cells can maintain stem cell pluripotency or direct adult stem cell differentiation.

  8. Direct reprogramming of human neural stem cells by OCT4.

    PubMed

    Kim, Jeong Beom; Greber, Boris; Araúzo-Bravo, Marcos J; Meyer, Johann; Park, Kook In; Zaehres, Holm; Schöler, Hans R

    2009-10-01

    Induced pluripotent stem (iPS) cells have been generated from mouse and human somatic cells by ectopic expression of four transcription factors (OCT4 (also called POU5F1), SOX2, c-Myc and KLF4). We previously reported that Oct4 alone is sufficient to reprogram directly adult mouse neural stem cells to iPS cells. Here we report the generation of one-factor human iPS cells from human fetal neural stem cells (one-factor (1F) human NiPS cells) by ectopic expression of OCT4 alone. One-factor human NiPS cells resemble human embryonic stem cells in global gene expression profiles, epigenetic status, as well as pluripotency in vitro and in vivo. These findings demonstrate that the transcription factor OCT4 is sufficient to reprogram human neural stem cells to pluripotency. One-factor iPS cell generation will advance the field further towards understanding reprogramming and generating patient-specific pluripotent stem cells. PMID:19718018

  9. Therapeutics from Adult Stem Cells and the Hype Curve.

    PubMed

    Maguire, Greg

    2016-05-12

    The Gartner curve for regenerative and stem cell therapeutics is currently climbing out of the "trough of disillusionment" and into the "slope of enlightenment". Understanding that the early years of stem cell therapy relied on the model of embryonic stem cells (ESCs), and then moved into a period of the overhype of induced pluripotent stem cells (iPSCs), instead of using the model of 40 years of success, i.e. adult stem cells used in bone marrow transplants, the field of stem cell therapy has languished for years, trying to move beyond the early and poorly understood success of bone marrow transplants. Recent studies in the lab and clinic show that adult stem cells of various types, and the molecules that they release, avoid the issues associated with ESCs and iPSCs and lead to better therapeutic outcomes and into the slope of enlightenment. PMID:27190588

  10. Therapeutics from Adult Stem Cells and the Hype Curve.

    PubMed

    Maguire, Greg

    2016-05-12

    The Gartner curve for regenerative and stem cell therapeutics is currently climbing out of the "trough of disillusionment" and into the "slope of enlightenment". Understanding that the early years of stem cell therapy relied on the model of embryonic stem cells (ESCs), and then moved into a period of the overhype of induced pluripotent stem cells (iPSCs), instead of using the model of 40 years of success, i.e. adult stem cells used in bone marrow transplants, the field of stem cell therapy has languished for years, trying to move beyond the early and poorly understood success of bone marrow transplants. Recent studies in the lab and clinic show that adult stem cells of various types, and the molecules that they release, avoid the issues associated with ESCs and iPSCs and lead to better therapeutic outcomes and into the slope of enlightenment.

  11. Developing human-nonhuman chimeras in human stem cell research: ethical issues and boundaries.

    PubMed

    Karpowicz, Phillip; Cohen, Cynthia B; van der Kooy, Derek

    2005-06-01

    The transplantation of adult human neural stem cells into prenatal non-humans offers an avenue for studying human neural cell development without direct use of human embryos. However, such experiments raise significant ethical concerns about mixing human and nonhuman materials in ways that could result in the development of human-nonhuman chimeras. This paper examines four arguments against such research, the moral taboo, species integrity, "unnaturalness," and human dignity arguments, and finds the last plausible. It argues that the transfer of human brain or retinal stem cells to nonhuman embryos would not result in the development of human-nonhuman chimeras that denigrate human dignity, provided such stem cells are dissociated. The article provides guidelines that set ethical boundaries for conducting such research that are consonant with the requirements of human dignity.

  12. Purification of oogonial stem cells from adult mouse and human ovaries: an assessment of the literature and a view toward the future.

    PubMed

    Woods, Dori C; White, Yvonne A R; Tilly, Jonathan L

    2013-01-01

    Contemporary claims that mitotically active female germ line or oogonial stem cells (OSCs) exist and support oogenesis during postnatal life in mammals have been debated in the field of reproductive biology since March 2004, when a mouse study posed the first serious challenge to the dogma of a fixed pool of oocytes being endowed at birth in more than 50 years. Other studies have since been put forth that further question the validity of this dogma, including the isolation of OSCs from neonatal and adult mouse ovaries by 4 independent groups using multiple strategies. Two of these groups also reported that isolated mouse OSCs, once transplanted back into ovaries of adult female mice, differentiate into fully functional eggs that ovulate, fertilize, and produce healthy embryos and offspring. Arguably, one of the most significant advances in this emerging field was provided by a new research study published this year, which reported the successful isolation and functional characterization of OSCs from ovaries of reproductive age women. Two commentaries on this latest work, one cautiously supportive and one highly skeptical, were published soon afterward. This article evaluates the current literature regarding postnatal oogenesis in mammals and discusses important next steps for future work on OSC biology and function.

  13. A Comparison of Culture Characteristics between Human Amniotic Mesenchymal Stem Cells and Dental Stem Cells.

    PubMed

    Yusoff, Nurul Hidayat; Alshehadat, Saaid Ayesh; Azlina, Ahmad; Kannan, Thirumulu Ponnuraj; Hamid, Suzina Sheikh Abdul

    2015-04-01

    In the past decade, the field of stem cell biology is of major interest among researchers due to its broad therapeutic potential. Stem cells are a class of undifferentiated cells that are able to differentiate into specialised cell types. Stem cells can be classified into two main types: adult stem cells (adult tissues) and embryonic stem cells (embryos formed during the blastocyst phase of embryological development). This review will discuss two types of adult mesenchymal stem cells, dental stem cells and amniotic stem cells, with respect to their differentiation lineages, passage numbers and animal model studies. Amniotic stem cells have a greater number of differentiation lineages than dental stem cells. On the contrary, dental stem cells showed the highest number of passages compared to amniotic stem cells. For tissue regeneration based on animal studies, amniotic stem cells showed the shortest time to regenerate in comparison with dental stem cells.

  14. Cancer Genes Hypermethylated in Human Embryonic Stem Cells

    PubMed Central

    Calvanese, Vincenzo; Horrillo, Angelica; Hmadcha, Abdelkrim; Suarez-Álvarez, Beatriz; Fernandez, Agustín F.; Lara, Ester; Casado, Sara; Menendez, Pablo; Bueno, Clara; Garcia-Castro, Javier; Rubio, Ruth; Lapunzina, Pablo; Alaminos, Miguel; Borghese, Lodovica; Terstegge, Stefanie; Harrison, Neil J.; Moore, Harry D.; Brüstle, Oliver; Lopez-Larrea, Carlos; Andrews, Peter W.; Soria, Bernat; Esteller, Manel; Fraga, Mario F.

    2008-01-01

    Developmental genes are silenced in embryonic stem cells by a bivalent histone-based chromatin mark. It has been proposed that this mark also confers a predisposition to aberrant DNA promoter hypermethylation of tumor suppressor genes (TSGs) in cancer. We report here that silencing of a significant proportion of these TSGs in human embryonic and adult stem cells is associated with promoter DNA hypermethylation. Our results indicate a role for DNA methylation in the control of gene expression in human stem cells and suggest that, for genes repressed by promoter hypermethylation in stem cells in vivo, the aberrant process in cancer could be understood as a defect in establishing an unmethylated promoter during differentiation, rather than as an anomalous process of de novo hypermethylation. PMID:18820729

  15. Ciprofloxacin Improves the Stemness of Human Dermal Papilla Cells.

    PubMed

    Kiratipaiboon, Chayanin; Tengamnuay, Parkpoom; Chanvorachote, Pithi

    2016-01-01

    Improvement in the expansion method of adult stem cells may augment their use in regenerative therapy. Using human dermal papilla cell line as well as primary dermal papilla cells as model systems, the present study demonstrated that ciprofloxacin treatment could prevent the loss of stemness during culture. Clonogenicity and stem cell markers of dermal papilla cells were shown to gradually decrease in the culture in a time-dependent manner. Treatment of the cells with nontoxic concentrations of ciprofloxacin could maintain both stem cell morphology and clonogenicity, as well as all stem cells markers. We found that ciprofloxacin exerted its effect through ATP-dependent tyrosine kinase/glycogen synthase kinase3β dependent mechanism which in turn upregulated β-catenin. Besides, ciprofloxacin was shown to induce epithelial-mesenchymal transition in DPCs as the transcription factors ZEB1 and Snail were significantly increased. Furthermore, the self-renewal proteins of Wnt/β-catenin pathway, namely, Nanog and Oct-4 were significantly upregulated in the ciprofloxacin-treated cells. The effects of ciprofloxacin in preserving stem cell features were confirmed in the primary dermal papilla cells directly obtained from human hair follicles. Together, these results revealed a novel application of ciprofloxacin for stem cell maintenance and provided the underlying mechanisms that are responsible for the stemness in dermal papilla cells.

  16. Adult Human Neurogenesis: From Microscopy to Magnetic Resonance Imaging

    PubMed Central

    Sierra, Amanda; Encinas, Juan M.; Maletic-Savatic, Mirjana

    2011-01-01

    Neural stem cells reside in well-defined areas of the adult human brain and are capable of generating new neurons throughout the life span. In rodents, it is well established that the new born neurons are involved in olfaction as well as in certain forms of memory and learning. In humans, the functional relevance of adult human neurogenesis is being investigated, in particular its implication in the etiopathology of a variety of brain disorders. Adult neurogenesis in the human brain was discovered by utilizing methodologies directly imported from the rodent research, such as immunohistological detection of proliferation and cell-type specific biomarkers in postmortem or biopsy tissue. However, in the vast majority of cases, these methods do not support longitudinal studies; thus, the capacity of the putative stem cells to form new neurons under different disease conditions cannot be tested. More recently, new technologies have been specifically developed for the detection and quantification of neural stem cells in the living human brain. These technologies rely on the use of magnetic resonance imaging, available in hospitals worldwide. Although they require further validation in rodents and primates, these new methods hold the potential to test the contribution of adult human neurogenesis to brain function in both health and disease. This review reports on the current knowledge on adult human neurogenesis. We first review the different methods available to assess human neurogenesis, both ex vivo and in vivo and then appraise the changes of adult neurogenesis in human diseases. PMID:21519376

  17. Cell motion predicts human epidermal stemness

    PubMed Central

    Toki, Fujio; Tate, Sota; Imai, Matome; Matsushita, Natsuki; Shiraishi, Ken; Sayama, Koji; Toki, Hiroshi; Higashiyama, Shigeki

    2015-01-01

    Image-based identification of cultured stem cells and noninvasive evaluation of their proliferative capacity advance cell therapy and stem cell research. Here we demonstrate that human keratinocyte stem cells can be identified in situ by analyzing cell motion during their cultivation. Modeling experiments suggested that the clonal type of cultured human clonogenic keratinocytes can be efficiently determined by analysis of early cell movement. Image analysis experiments demonstrated that keratinocyte stem cells indeed display a unique rotational movement that can be identified as early as the two-cell stage colony. We also demonstrate that α6 integrin is required for both rotational and collective cell motion. Our experiments provide, for the first time, strong evidence that cell motion and epidermal stemness are linked. We conclude that early identification of human keratinocyte stem cells by image analysis of cell movement is a valid parameter for quality control of cultured keratinocytes for transplantation. PMID:25897083

  18. Novel in vitro culture condition improves the stemness of human dermal stem/progenitor cells.

    PubMed

    Shim, Joong Hyun; Lee, Tae Ryong; Shin, Dong Wook

    2013-12-01

    Cell therapy using adult stem cells has emerged as a potentially new approach for the treatment of various diseases. Therefore, it is an essential procedure to maintain the stemness of adult stem cells for clinical treatment. We previously reported that human dermal stem/progenitor cells (hDSPCs) can be enriched using collagen type IV. However, hDSPCs gradually lose their stem cell properties as in vitro passages continue. In the present study, we developed optimized in vitro culture condition to improve the stemness of these hDSPCs. To evaluate whether the stemness of hDSPCs is well sustained in various culture conditions, we measured the expression levels of SOX2, NANOG, and S100B, which are well-known representative dermal progenitor markers. We observed that hDSPCs grown in three-dimensional (3D) culture condition had higher expression levels of those markers compared with hDSPCs grown in two-dimensional (2D) culture condition. Under the 3D culture condition, we further demonstrated that a high glucose (4.5 g/L) concentration enhanced the expression levels of the dermal progenitor markers, whereas O(2) concentration did not affect. We also found that skin-derived precursor (SKP) culture medium was the most effective, among various culture media, in increasing the dermal progenitor marker expression. We finally demonstrated that this optimized culture condition enhanced the expression level of human telomerase reverse transcriptase (hTERT), the proliferation, and the multipotency of hDSPCs, an important characteristic of stem cells. Taken together, these results suggested that this novel in vitro culture condition improves the stemness of hDSPCs.

  19. Strategies to Optimize Adult Stem Cell Therapy for Tissue Regeneration

    PubMed Central

    Liu, Shan; Zhou, Jingli; Zhang, Xuan; Liu, Yang; Chen, Jin; Hu, Bo; Song, Jinlin; Zhang, Yuanyuan

    2016-01-01

    Stem cell therapy aims to replace damaged or aged cells with healthy functioning cells in congenital defects, tissue injuries, autoimmune disorders, and neurogenic degenerative diseases. Among various types of stem cells, adult stem cells (i.e., tissue-specific stem cells) commit to becoming the functional cells from their tissue of origin. These cells are the most commonly used in cell-based therapy since they do not confer risk of teratomas, do not require fetal stem cell maneuvers and thus are free of ethical concerns, and they confer low immunogenicity (even if allogenous). The goal of this review is to summarize the current state of the art and advances in using stem cell therapy for tissue repair in solid organs. Here we address key factors in cell preparation, such as the source of adult stem cells, optimal cell types for implantation (universal mesenchymal stem cells vs. tissue-specific stem cells, or induced vs. non-induced stem cells), early or late passages of stem cells, stem cells with endogenous or exogenous growth factors, preconditioning of stem cells (hypoxia, growth factors, or conditioned medium), using various controlled release systems to deliver growth factors with hydrogels or microspheres to provide apposite interactions of stem cells and their niche. We also review several approaches of cell delivery that affect the outcomes of cell therapy, including the appropriate routes of cell administration (systemic, intravenous, or intraperitoneal vs. local administration), timing for cell therapy (immediate vs. a few days after injury), single injection of a large number of cells vs. multiple smaller injections, a single site for injection vs. multiple sites and use of rodents vs. larger animal models. Future directions of stem cell-based therapies are also discussed to guide potential clinical applications. PMID:27338364

  20. Strategies to Optimize Adult Stem Cell Therapy for Tissue Regeneration.

    PubMed

    Liu, Shan; Zhou, Jingli; Zhang, Xuan; Liu, Yang; Chen, Jin; Hu, Bo; Song, Jinlin; Zhang, Yuanyuan

    2016-06-21

    Stem cell therapy aims to replace damaged or aged cells with healthy functioning cells in congenital defects, tissue injuries, autoimmune disorders, and neurogenic degenerative diseases. Among various types of stem cells, adult stem cells (i.e., tissue-specific stem cells) commit to becoming the functional cells from their tissue of origin. These cells are the most commonly used in cell-based therapy since they do not confer risk of teratomas, do not require fetal stem cell maneuvers and thus are free of ethical concerns, and they confer low immunogenicity (even if allogenous). The goal of this review is to summarize the current state of the art and advances in using stem cell therapy for tissue repair in solid organs. Here we address key factors in cell preparation, such as the source of adult stem cells, optimal cell types for implantation (universal mesenchymal stem cells vs. tissue-specific stem cells, or induced vs. non-induced stem cells), early or late passages of stem cells, stem cells with endogenous or exogenous growth factors, preconditioning of stem cells (hypoxia, growth factors, or conditioned medium), using various controlled release systems to deliver growth factors with hydrogels or microspheres to provide apposite interactions of stem cells and their niche. We also review several approaches of cell delivery that affect the outcomes of cell therapy, including the appropriate routes of cell administration (systemic, intravenous, or intraperitoneal vs. local administration), timing for cell therapy (immediate vs. a few days after injury), single injection of a large number of cells vs. multiple smaller injections, a single site for injection vs. multiple sites and use of rodents vs. larger animal models. Future directions of stem cell-based therapies are also discussed to guide potential clinical applications.

  1. sb203580 preconditioning recharges matrix-expanded human adult stem cells for chondrogenesis in an inflammatory environment – a feasible approach for autologous stem cell based osteoarthritic cartilage repair

    PubMed Central

    Zhang, Ying; Pizzute, Tyler; Li, Jingting; He, Fan; Pei, Ming

    2015-01-01

    Autologous stem cells are a promising cell source for cartilage regeneration; however, cell replicative senescence and joint posttraumatic inflammation provide challenges to bring this treatment modality to fruition. In this study, we hypothesized that preconditioning with p38 MAPK inhibitor (sb203580) would recharge decellularized extracellular matrix (dECM) expanded human synovium-derived stem cell (hSDSC) chondrogenesis in an inflammatory environment. We found that preconditioning with sb203580 greatly enhanced dECM expanded hSDSC proliferation and chondrogenic potential while supplementation with sb203580 in an induction medium dramatically retarded hSDSC chondrogenic differentiation, even for dECM expanded cells. We also found that sb203580 preconditioning enhanced matrix-expanded hSDSC chondrogenic capacity even in an interleukin-1 (IL-1) induced inflammatory environment. Non-detectable expression of HLA-DR in the hSDSCs grown on allogeneic dECM indicates the feasibility of commercial preparation of these dECMs from healthy, young donors for patients who need autologous transplantation. Our study indicated that p38 MAPK inhibitor has a distinctive priming effect on dECM mediated stem cell cartilage regeneration. Combined rejuvenation with sb203580 and dECM expansion can precondition hSDSCs’ resurfacing capacity for osteoarthritic patients with cartilage defects. PMID:26122165

  2. Human Adipose Stem Cells: From Bench to Bedside.

    PubMed

    De Francesco, Francesco; Ricci, Giulia; D'Andrea, Francesco; Nicoletti, Giovanni Francesco; Ferraro, Giuseppe Andrea

    2015-12-01

    Stem cell-based therapies for repair and regeneration of different tissues are becoming more important in the treatment of several diseases. Adult stem cells currently symbolize the most available source of cell progenitors for tissue engineering and repair and can be harvested using minimally invasive procedures. Moreover, mesenchymal stem cells (MSCs), the most widely used stem cells in stem cell-based therapies, are multipotent progenitors, with capability to differentiate into cartilage, bone, connective, muscle, and adipose tissue. So far, bone marrow has been regarded as the main source of MSCs. To date, human adult adipose tissue may be the best suitable alternative source of MSCs. Adipose stem cells (ASCs) can be largely extracted from subcutaneous human adult adipose tissue. A large number of studies show that adipose tissue contains a biologically and clinically interesting heterogeneous cell population called stromal vascular fraction (SVF). The SVF may be employed directly or cultured for selection and expansion of an adherent population, so called adipose-derived stem cells (ASCs). In recent years, literature based on data related to SVF cells and ASCs has augmented considerably: These studies have demonstrated the efficacy and safety of SVF cells and ASCs in vivo in animal models. On the basis of these observations, in several countries, various clinical trials involving SVF cells and ASCs have been permitted. This review aims at summarizing data regarding either ASCs cellular biology or ASCs-based clinical trials and at discussing the possible future clinical translation of ASCs and their potentiality in cell-based tissue engineering.

  3. [Therapeutic use of stem cells. II. Adult stem cells].

    PubMed

    Uzan, Georges

    2004-09-30

    Many degenerative diseases are not curable by means of classical medicine. The long term objective of cell therapy is to treat the patients with their own stem cells that could be either purified from the diseased organ or from "reservoirs" of stem cells such as that constituted by the bone marrow. The existence of stem cells in the organs or reservoirs is now established in vitro and in some cases, in animal models. Numbers of technical problems linked to the scarcity of these cells still delay the clinical use of purified stem cells. However, clinical protocols using heterogeneous cell populations have already started to treat a growing number of diseases. In some case, autologous cells can be used, as it is the case for bone marrow transplantation in blood diseases. Mesenchymal cells, also purified from the bone marrow are currently used in orthopaedic diseases. Because these cells reveal a broad differentiation potential, active research programs explore their possible use for treatment of other diseases. Bone marrow also contains vascular stem cells that could be active in reappearing defective vessels responsible for ischaemic diseases. Indeed, clinical trials in which bone marrow cells are injected in the cardiac muscle of patients with myocardial infarction or in the leg muscle (gastrocnemius) of patients with hind limb ischaemia have already started. Artificial skin prepared from skin biopsies is used for the reconstitution of the derma of severely burned patients. Clinical trials have also started, using allogenic cells. The patients must be treated by immunosuppressive drugs. Neurodegenerative diseases such as Parkinson have been successfully treated by intra-cerebral injection of foetal neurones. Pancreatic islets implanted in the liver have shown to re-establish a normal glycaemia in diabetic patients. However, all these clinical trials use differentiated cells or at least progenitors which display differentiation potential and lifetime much more

  4. Generation of human melanocytes from induced pluripotent stem cells.

    PubMed

    Ohta, Shigeki; Imaizumi, Yoichi; Akamatsu, Wado; Okano, Hideyuki; Kawakami, Yutaka

    2013-01-01

    The discovery of human induced pluripotent stem cells (iPSCs) has provided a model system for studying early events during human development. Developmentally melanocytes originate from migratory neural crest cells that emerge from the neural plate during embryogenesis after a complex process of differentiation, proliferation, and migration out of the neural tube along defined pathways. In the adult, human melanocytes are located in the basal layer of the epidermis, hair follicles, uvea, inner ear, and meninges. In the epidermis, melanocytes produce melanin pigment that gives color to the skin as well as providing protection from ultraviolet light damage. In addition, melanocytes transfer melanin pigment to hair matrix keratinocytes during each hair cycle to maintain hair pigmentation. Characterization of mouse melanocyte stem cells (MELSCs) is more complete than for humans. MELSCs are located in the bulge region of hair follicles, where hair follicle stem cells (HFSCs) also reside. Recently, it has been demonstrated that HFSCs provide a functional nice for MELSCs. According to current cancer stem cell theory, melanomas are considered to evolve from MELSCs, although the exact mechanism remains to be elucidated fully. In humans, importantly, the lack of more specific markers of MELSCs, current understanding of the molecular regulations of melanocyte development remains incomplete. Recently, the generation of melanocytes from iPSCs has lead to some clarification of human melanocyte development in vitro. Utilization of iPSC-derived melanocytes may prove invaluable in further study of human melanocytic development and novel therapies for patients suffering with pigmentation disorders and melanoma.

  5. Human mesenchymal stem cells - current trends and future prospective

    PubMed Central

    Ullah, Imran; Subbarao, Raghavendra Baregundi; Rho, Gyu Jin

    2015-01-01

    Stem cells are cells specialized cell, capable of renewing themselves through cell division and can differentiate into multi-lineage cells. These cells are categorized as embryonic stem cells (ESCs), induced pluripotent stem cells (iPSCs) and adult stem cells. Mesenchymal stem cells (MSCs) are adult stem cells which can be isolated from human and animal sources. Human MSCs (hMSCs) are the non-haematopoietic, multipotent stem cells with the capacity to differentiate into mesodermal lineage such as osteocytes, adipocytes and chondrocytes as well ectodermal (neurocytes) and endodermal lineages (hepatocytes). MSCs express cell surface markers like cluster of differentiation (CD)29, CD44, CD73, CD90, CD105 and lack the expression of CD14, CD34, CD45 and HLA (human leucocyte antigen)-DR. hMSCs for the first time were reported in the bone marrow and till now they have been isolated from various tissues, including adipose tissue, amniotic fluid, endometrium, dental tissues, umbilical cord and Wharton's jelly which harbours potential MSCs. hMSCs have been cultured long-term in specific media without any severe abnormalities. Furthermore, MSCs have immunomodulatory features, secrete cytokines and immune-receptors which regulate the microenvironment in the host tissue. Multilineage potential, immunomodulation and secretion of anti-inflammatory molecules makes MSCs an effective tool in the treatment of chronic diseases. In the present review, we have highlighted recent research findings in the area of hMSCs sources, expression of cell surface markers, long-term in vitro culturing, in vitro differentiation potential, immunomodulatory features, its homing capacity, banking and cryopreservation, its application in the treatment of chronic diseases and its use in clinical trials. PMID:25797907

  6. Human skeletal muscle-derived stem cells retain stem cell properties after expansion in myosphere culture

    SciTech Connect

    Wei, Yan; Li, Yuan; Chen, Chao; Stoelzel, Katharina; Kaufmann, Andreas M.

    2011-04-15

    Human skeletal muscle contains an accessible adult stem-cell compartment in which differentiated myofibers are maintained and replaced by a self-renewing stem cell pool. Previously, studies using mouse models have established a critical role for resident stem cells in skeletal muscle, but little is known about this paradigm in human muscle. Here, we report the reproducible isolation of a population of cells from human skeletal muscle that is able to proliferate for extended periods of time as floating clusters of rounded cells, termed 'myospheres' or myosphere-derived progenitor cells (MDPCs). The phenotypic characteristics and functional properties of these cells were determined using reverse transcription-polymerase chain reaction (RT-PCR), flow cytometry and immunocytochemistry. Our results showed that these cells are clonogenic, express skeletal progenitor cell markers Pax7, ALDH1, Myod, and Desmin and the stem cell markers Nanog, Sox2, and Oct3/4 significantly elevated over controls. They could be maintained proliferatively active in vitro for more than 20 weeks and passaged at least 18 times, despite an average donor-age of 63 years. Individual clones (4.2%) derived from single cells were successfully expanded showing clonogenic potential and sustained proliferation of a subpopulation in the myospheres. Myosphere-derived cells were capable of spontaneous differentiation into myotubes in differentiation media and into other mesodermal cell lineages in induction media. We demonstrate here that direct culture and expansion of stem cells from human skeletal muscle is straightforward and reproducible with the appropriate technique. These cells may provide a viable resource of adult stem cells for future therapies of disease affecting skeletal muscle or mesenchymal lineage derived cell types.

  7. Intestinal stem cells in the adult Drosophila midgut

    SciTech Connect

    Jiang, Huaqi; Edgar, Bruce A.

    2011-11-15

    Drosophila has long been an excellent model organism for studying stem cell biology. Notably, studies of Drosophila's germline stem cells have been instrumental in developing the stem cell niche concept. The recent discovery of somatic stem cells in adult Drosophila, particularly the intestinal stem cells (ISCs) of the midgut, has established Drosophila as an exciting model to study stem cell-mediated adult tissue homeostasis and regeneration. Here, we review the major signaling pathways that regulate the self-renewal, proliferation and differentiation of Drosophila ISCs, discussing how this regulation maintains midgut homeostasis and mediates regeneration of the intestinal epithelium after injury. -- Highlights: Black-Right-Pointing-Pointer The homeostasis and regeneration of adult fly midguts are mediated by ISCs. Black-Right-Pointing-Pointer Damaged enterocytes induce the proliferation of intestinal stem cells (ISC). Black-Right-Pointing-Pointer EGFR and Jak/Stat signalings mediate compensatory ISC proliferation. Black-Right-Pointing-Pointer Notch signaling regulates ISC self-renewal and differentiation.

  8. Potential antitumor therapeutic strategies of human amniotic membrane and amniotic fluid-derived stem cells.

    PubMed

    Kang, N-H; Hwang, K-A; Kim, S U; Kim, Y-B; Hyun, S-H; Jeung, E-B; Choi, K-C

    2012-08-01

    As stem cells are capable of self-renewal and can generate differentiated progenies for organ development, they are considered as potential source for regenerative medicine and tissue replacement after injury or disease. Along with this capacity, stem cells have the therapeutic potential for treating human diseases including cancers. According to the origins, stem cells are broadly classified into two types: embryonic stem cells (ESCs) and adult stem cells. In terms of differentiation potential, ESCs are pluripotent and adult stem cells are multipotent. Amnion, which is a membranous sac that contains the fetus and amniotic fluid and functions in protecting the developing embryo during gestation, is another stem cell source. Amnion-derived stem cells are classified as human amniotic membrane-derived epithelial stem cells, human amniotic membrane-derived mesenchymal stem cells and human amniotic fluid-derived stem cells. They are in an intermediate stage between pluripotent ESCs and lineage-restricted adult stem cells, non-tumorigenic, and contribute to low immunogenicity and anti-inflammation. Furthermore, they are easily available and do not cause any controversial issues in their recovery and applications. Not only are amnion-derived stem cells applicable in regenerative medicine, they have anticancer capacity. In non-engineered stem cells transplantation strategies, amnion-derived stem cells effectively target the tumor and suppressed the tumor growth by expressing cytotoxic cytokines. Additionally, they also have a potential as novel delivery vehicles transferring therapeutic genes to the cancer formation sites in gene-directed enzyme/prodrug combination therapy. Owing to their own advantageous properties, amnion-derived stem cells are emerging as a new candidate in anticancer therapy.

  9. Patenting human genes and stem cells.

    PubMed

    Martin-Rendon, Enca; Blake, Derek J

    2007-01-01

    Cell lines and genetically modified single cell organisms have been considered patentable subjects for the last two decades. However, despite the technical patentability of genes and stem cell lines, social and legal controversy concerning their 'ownership' has surrounded stem cell research in recent years. Some granted patents on stem cells with extremely broad claims are casting a shadow over the commercialization of these cells as therapeutics. However, in spite of those early patents, the number of patent applications related to stem cells is growing exponentially. Both embryonic and adult stem cells have the ability to differentiate into several cell lineages in an organism as a result of specific genetic programs that direct their commitment and cell fate. Genes that control the pluripotency of stem cells have been recently identified and the genetic manipulation of these cells is becoming more efficient with the advance of new technologies. This review summarizes some of the recent published patents on pluripotency genes, gene transfer into stem cells and genetic reprogramming and takes the hematopoietic and embryonic stem cell as model systems.

  10. Arts & Humanities in Adult Education.

    ERIC Educational Resources Information Center

    Word's Worth: A Quarterly Newsletter of the Lifelong Learning Network, 1998

    1998-01-01

    This issue of a quarterly newsletter on lifelong learning focuses on the theme of the arts and humanities in adult literacy education. The following articles are included: (1) "In Defense of a Practical Education" (Earl Shorris); (2) "From the Program Director" (Elizabeth Bryant McCrary); (3) "Vermont Council on the Humanities: Book Discussion…

  11. 3 CFR - Guidelines for Human Stem Cell Research

    Code of Federal Regulations, 2010 CFR

    2010-01-01

    ... 3 The President 1 2010-01-01 2010-01-01 false Guidelines for Human Stem Cell Research Presidential Documents Other Presidential Documents Memorandum of July 30, 2009 Guidelines for Human Stem Cell Research..., scientifically worthy human stem cell research, including human embryonic stem cell research, to the...

  12. A novel view of the adult bone marrow stem cell hierarchy and stem cell trafficking.

    PubMed

    Ratajczak, M Z

    2015-04-01

    This review presents a novel view and working hypothesis about the hierarchy within the adult bone marrow stem cell compartment and the still-intriguing question of whether adult bone marrow contains primitive stem cells from early embryonic development, such as cells derived from the epiblast, migrating primordial germ cells or yolk sac-derived hemangioblasts. It also presents a novel view of the mechanisms that govern stem cell mobilization and homing, with special emphasis on the role of the complement cascade as a trigger for egress of hematopoietic stem cells from bone marrow into blood as well as the emerging role of novel homing factors and priming mechanisms that support stromal-derived factor 1-mediated homing of hematopoietic stem/progenitor cells after transplantation.

  13. A novel view of the adult bone marrow stem cell hierarchy and stem cell trafficking

    PubMed Central

    Ratajczak, M Z

    2015-01-01

    This review presents a novel view and working hypothesis about the hierarchy within the adult bone marrow stem cell compartment and the still-intriguing question of whether adult bone marrow contains primitive stem cells from early embryonic development, such as cells derived from the epiblast, migrating primordial germ cells or yolk sac-derived hemangioblasts. It also presents a novel view of the mechanisms that govern stem cell mobilization and homing, with special emphasis on the role of the complement cascade as a trigger for egress of hematopoietic stem cells from bone marrow into blood as well as the emerging role of novel homing factors and priming mechanisms that support stromal-derived factor 1-mediated homing of hematopoietic stem/progenitor cells after transplantation. PMID:25486871

  14. Adult mesenchymal stem cells for tissue engineering versus regenerative medicine.

    PubMed

    Caplan, Arnold I

    2007-11-01

    Adult mesenchymal stem cells (MSCs) can be isolated from bone marrow or marrow aspirates and because they are culture-dish adherent, they can be expanded in culture while maintaining their multipotency. The MSCs have been used in preclinical models for tissue engineering of bone, cartilage, muscle, marrow stroma, tendon, fat, and other connective tissues. These tissue-engineered materials show considerable promise for use in rebuilding damaged or diseased mesenchymal tissues. Unanticipated is the realization that the MSCs secrete a large spectrum of bioactive molecules. These molecules are immunosuppressive, especially for T-cells and, thus, allogeneic MSCs can be considered for therapeutic use. In this context, the secreted bioactive molecules provide a regenerative microenvironment for a variety of injured adult tissues to limit the area of damage and to mount a self-regulated regenerative response. This regenerative microenvironment is referred to as trophic activity and, therefore, MSCs appear to be valuable mediators for tissue repair and regeneration. The natural titers of MSCs that are drawn to sites of tissue injury can be augmented by allogeneic MSCs delivered via the bloodstream. Indeed, human clinical trials are now under way to use allogeneic MSCs for treatment of myocardial infarcts, graft-versus-host disease, Crohn's Disease, cartilage and meniscus repair, stroke, and spinal cord injury. This review summarizes the biological basis for the in vivo functioning of MSCs through development and aging. PMID:17620285

  15. Human adipose stem cells: current clinical applications.

    PubMed

    Gir, Phanette; Oni, Georgette; Brown, Spencer A; Mojallal, Ali; Rohrich, Rod J

    2012-06-01

    Adipose-derived stem cells are multipotent cells that can easily be extracted from adipose tissue, are capable of expansion in vitro, and have the capacity to differentiate into multiple cell lineages, which have the potential for use in regenerative medicine. However, several issues need to be studied to determine safe human use. For example, there are questions related to isolation and purification of adipose-derived stem cells, their effect on tumor growth, and the enforcement of U.S. Food and Drug Administration regulations. Numerous studies have been published, with the interest in the potential for regenerative medicine continually growing. Several clinical trials using human adipose stem cell therapy are currently being performed around the world, and there has been a rapid evolution and expansion of their number. The purpose of this article was to review the current published basic science evidence and ongoing clinical trials involving the use of adipose-derived stem cells in plastic surgery and in regenerative medicine in general. The results of the studies and clinical trials using adipose-derived stem cells reported in this review seem to be promising not only in plastic surgery but also in a wide variety of other specialties. Nevertheless, those reported showed disparity in the way adipose-derived stem cells were used. Further basic science experimental studies with standardized protocols and larger randomized trials need to be performed to ensure safety and efficacy of adipose-derived stem cells use in accordance with U.S. Food and Drug Administration guidelines.

  16. Centre for human development, stem cells & regeneration.

    PubMed

    Oreffo, Richard O C

    2014-01-01

    The Centre for Human Development, Stem Cells and Regeneration (CHDSCR) was founded in 2004 as a cross-disciplinary research and translational program within the Faculty of Medicine at the University of Southampton. The Centre undertakes fundamental research into early development and stem cells together with applied translational research for patient benefit. The Centre has vibrant and thriving multidisciplinary research programs that harness the translational strength of the Faculty together with an innovative Stem Cell PhD program, outstanding clinical infrastructure and enterprise to deliver on this vision.

  17. Human Stem Cells for Craniomaxillofacial Reconstruction

    PubMed Central

    Kirkpatrick, William Niall Alexander; Cameron, Malcolm Gregor

    2014-01-01

    Human stem cell research represents an exceptional opportunity for regenerative medicine and the surgical reconstruction of the craniomaxillofacial complex. The correct architecture and function of the vastly diverse tissues of this important anatomical region are critical for life supportive processes, the delivery of senses, social interaction, and aesthetics. Craniomaxillofacial tissue loss is commonly associated with inflammatory responses of the surrounding tissue, significant scarring, disfigurement, and psychological sequelae as an inevitable consequence. The in vitro production of fully functional cells for skin, muscle, cartilage, bone, and neurovascular tissue formation from human stem cells, may one day provide novel materials for the reconstructive surgeon operating on patients with both hard and soft tissue deficit due to cancer, congenital disease, or trauma. However, the clinical translation of human stem cell technology, including the application of human pluripotent stem cells (hPSCs) in novel regenerative therapies, faces several hurdles that must be solved to permit safe and effective use in patients. The basic biology of hPSCs remains to be fully elucidated and concerns of tumorigenicity need to be addressed, prior to the development of cell transplantation treatments. Furthermore, functional comparison of in vitro generated tissue to their in vivo counterparts will be necessary for confirmation of maturity and suitability for application in reconstructive surgery. Here, we provide an overview of human stem cells in disease modeling, drug screening, and therapeutics, while also discussing the application of regenerative medicine for craniomaxillofacial tissue deficit and surgical reconstruction. PMID:24564584

  18. Adult stem cells and their ability to differentiate.

    PubMed

    Tarnowski, Maciej; Sieron, Aleksander L

    2006-08-01

    This is a review of the current status of knowledge on adult stem cells as well as the criteria and evidence for their potential to transform into different cell types and cell lineages. Reports on stem cell sources, focusing on tissues from adult subjects, were also investigated. Numerous reports have been published on the search for early markers of both stem cells and the precursors of various cell lineages. The question is still open about the characteristics of the primary stem cell. The existing proofs and hypotheses have not yielded final solutions to this problem. From a practical point of view it is also crucial to find a minimal set of markers determining the phenotypes of the precursor cells of a particular cell lineage. Several lines of evidence seem to bring closer the day when we will be able to detect the right stem cell niche and successfully isolate precursor cells that are needed for the treatment of a particular disorder. Recent reports on cases of cancer in patients subjected to stem cell therapy are yet another controversial issue looked into in this review, although the pros and cons emerging from the results of published studies still do not provide satisfying evidence to fully understand this issue.

  19. Endothelial juxtaposition of distinct adult stem cells activates angiogenesis signaling molecules in endothelial cells.

    PubMed

    Mohammadi, Elham; Nassiri, Seyed Mahdi; Rahbarghazi, Reza; Siavashi, Vahid; Araghi, Atefeh

    2015-12-01

    Efficacy of therapeutic angiogenesis needs a comprehensive understanding of endothelial cell (EC) function and biological factors and cells that interplay with ECs. Stem cells are considered the key components of pro- and anti-angiogenic milieu in a wide variety of physiopathological states, and interactions of EC-stem cells have been the subject of controversy in recent years. In this study, the potential effects of three tissue-specific adult stem cells, namely rat marrow-derived mesenchymal stem cells (rBMSCs), rat adipose-derived stem cells (rADSCs) and rat muscle-derived satellite cells (rSCs), on the endothelial activation of key angiogenic signaling molecules, including VEGF, Ang-2, VEGFR-2, Tie-2, and Tie2-pho, were investigated. Human umbilical vein endothelial cells (HUVECs) and rat lung microvascular endothelial cells (RLMECs) were cocultured with the stem cells or incubated with the stem cell-derived conditioned media on Matrigel. Following HUVEC-stem cell coculture, CD31-positive ECs were flow sorted and subjected to western blotting to analyze potential changes in the expression of the pro-angiogenic signaling molecules. Elongation and co-alignment of the stem cells were seen along the EC tubes in the EC-stem cell cocultures on Matrigel, with cell-to-cell dye communication in the EC-rBMSC cocultures. Moreover, rBMSCs and rADSCs significantly improved endothelial tubulogenesis in both juxtacrine and paracrine manners. These two latter stem cells dynamically up-regulated VEGF, Ang-2, VREGR-2, and Tie-2 but down-regulated Tie2-pho and the Tie2-pho/Tie-2 ratio in HUVECs. Induction of pro-angiogenic signaling in ECs by marrow- and adipose-derived MSCs further indicates the significance of stem cell milieu in angiogenesis dynamics. PMID:26068799

  20. Vascular Potential of Human Pluripotent Stem Cells

    PubMed Central

    Iacobas, Ionela; Vats, Archana; Hirschi, Karen K.

    2010-01-01

    Cardiovascular disease is the number one cause of death and disability in the US. Understanding the biological activity of stem and progenitor cells, and their ability to contribute to the repair, regeneration and remodeling of the heart and blood vessels affected by pathologic processes is an essential part of the paradigm in enabling us to achieve a reduction in related deaths. Both human embryonic stem (ES) cells and induced pluripotent stem (iPS) cells are promising sources of cells for clinical cardiovascular therapies. Additional in vitro studies are needed, however, to understand their relative phenotypes and molecular regulation toward cardiovascular cell fates. Further studies in translational animal models are also needed to gain insights into the potential and function of both human ES- and iPS-derived cardiovascular cells, and enable translation from experimental and pre-clinical studies to human trials. PMID:20453170

  1. Arsenic, stem cells, and the developmental basis of adult cancer.

    PubMed

    Tokar, Erik J; Qu, Wei; Waalkes, Michael P

    2011-03-01

    That chemical insults or nutritive changes during in utero and/or postnatal life can emerge as diseases much later in life are now being accepted as a recurring phenomenon. In this regard, inorganic arsenic is a multisite human carcinogen found at high levels in the drinking water of millions of people, although it has been difficult until recently to produce tumors in rodents with this metalloid. A mouse transplacental model has been developed where maternal exposure to inorganic arsenic either acts as a complete carcinogen or enhances carcinogenic response to other agents given subsequently in the offspring, producing tumors during adulthood. Similarly, human data now have emerged showing that arsenic exposure during the in utero period and/or in early life is associated with cancer in adulthood. The mouse arsenic transplacental model produces tumors or enhances response to other agents in multiple strains and tissues, including sites concordant with human targets of arsenic carcinogenesis. It is now believed that cancer often is a stem cell (SC)-based disease, and there is no reason to think cancer induced by developmental chemical exposure is any different. Indeed, arsenic impacts human SC population dynamics in vitro by blocking exit into differentiation pathways and whereby creating more key targets for transformation. In fact, during in vitro malignant transformation, arsenic causes a remarkable survival selection of SCs, creating a marked overabundance of cancer SCs (CSCs) compared with other carcinogens once a cancer phenotype is obtained. In addition, skin cancers produced following in utero arsenic exposure in mice are highly enriched in CSCs. Thus, arsenic impacts key, long-lived SC populations as critical targets to cause or facilitate later oncogenic events in adulthood as a possible mechanism of developmental basis of adult disease.

  2. Arsenic, Stem Cells, and the Developmental Basis of Adult Cancer

    PubMed Central

    Tokar, Erik J.; Qu, Wei; Waalkes, Michael P.

    2011-01-01

    That chemical insults or nutritive changes during in utero and/or postnatal life can emerge as diseases much later in life are now being accepted as a recurring phenomenon. In this regard, inorganic arsenic is a multisite human carcinogen found at high levels in the drinking water of millions of people, although it has been difficult until recently to produce tumors in rodents with this metalloid. A mouse transplacental model has been developed where maternal exposure to inorganic arsenic either acts as a complete carcinogen or enhances carcinogenic response to other agents given subsequently in the offspring, producing tumors during adulthood. Similarly, human data now have emerged showing that arsenic exposure during the in utero period and/or in early life is associated with cancer in adulthood. The mouse arsenic transplacental model produces tumors or enhances response to other agents in multiple strains and tissues, including sites concordant with human targets of arsenic carcinogenesis. It is now believed that cancer often is a stem cell (SC)–based disease, and there is no reason to think cancer induced by developmental chemical exposure is any different. Indeed, arsenic impacts human SC population dynamics in vitro by blocking exit into differentiation pathways and whereby creating more key targets for transformation. In fact, during in vitro malignant transformation, arsenic causes a remarkable survival selection of SCs, creating a marked overabundance of cancer SCs (CSCs) compared with other carcinogens once a cancer phenotype is obtained. In addition, skin cancers produced following in utero arsenic exposure in mice are highly enriched in CSCs. Thus, arsenic impacts key, long-lived SC populations as critical targets to cause or facilitate later oncogenic events in adulthood as a possible mechanism of developmental basis of adult disease. PMID:21071725

  3. Adult stem cells therapy for urine incontinence in women.

    PubMed

    Stangel-Wójcikiewicz, Klaudia; Majka, Marcin; Basta, Antoni; Stec, Małgorzata; Pabian, Wojciech; Piwowar, Monika; Chancellor, Michael B

    2010-05-01

    The past few years brought high development in obtaining and culturing autologous adult stem cells. In this paper we review publications of experimental investigations and clinical trials of the muscle-derived cells and the application in the treatment of stress urinary incontinence among women. Mesenchymal stem cells (MSCs) can be obtained from bone marrow but it is associated with a painful biopsy procedure. Collection of muscle-derived stem cells (MDSCs) is less harmful because the skeletal muscle biopsy is performed with a small caliber needle in local anesthesia. The stem-based therapy could be the next step in the treatment of urinary incontinence. There are still many elements of therapy such as effectiveness or long-term side effects which need to be researched.

  4. Characterization of human pluripotent stem cells.

    PubMed

    Gokhale, Paul J; Andrews, Peter W

    2013-12-18

    Human pluripotent stem cells (PSCs), whether embryonic stem cells or induced PSCs, offer enormous opportunities for regenerative medicine and other biomedical applications once we have developed the ability to harness their capacity for extensive differentiation. Central to this is our ability to identify and characterize such PSCs, but this is fraught with potential difficulties that arise from a tension between functional definitions of pluripotency and the more convenient use of 'markers', a problem exacerbated by ethical issues, our lack of knowledge of early human embryonic development, and differences from the mouse paradigm.

  5. Towards Personalized Regenerative Cell Therapy: Mesenchymal Stem Cells Derived from Human Induced Pluripotent Stem Cells.

    PubMed

    Lin, Lin; Bolund, Lars; Luo, Yonglun

    2016-01-01

    Mesenchymal stem cells (MSCs) are adult stem cells with the capacity of self-renewal and multilineage differentiation, and can be isolated from several adult tissues. However, isolating MSCs from adult tissues for cell therapy is hampered by the invasive procedure, the rarity of the cells and their attenuated proliferation capacity when cultivated and expanded in vitro. Human MSCs derived from induced pluripotent stem cells (iPSC-MSCs) have now evolved as a promising alternative cell source for MSCs and regenerative medicine. Several groups, including ours, have reported successful derivation of functional iPSC-MSCs and applied these cells in MSC-based therapeutic testing. Still, the current experience and understanding of iPSC-MSCs with respect to production methods, safety and efficacy are primitive. In this review, we highlight the methodological progress in iPSC-MSC research, describing the importance of choosing the right sources of iPSCs, iPSC reprogramming methods, iPSC culture systems, embryoid body intermediates, pathway inhibitors, basal medium, serum, growth factors and culture surface coating. We also highlight some progress in the application of iPSC-MSCs in direct cell therapy, tissue engineering and gene therapy.

  6. Epigenetic regulation of human adipose-derived stem cells differentiation.

    PubMed

    Daniunaite, Kristina; Serenaite, Inga; Misgirdaite, Roberta; Gordevicius, Juozas; Unguryte, Ausra; Fleury-Cappellesso, Sandrine; Bernotiene, Eiva; Jarmalaite, Sonata

    2015-12-01

    Adult stem cells have more restricted differentiation potential than embryonic stem cells (ESCs), but upon appropriate stimulation can differentiate into cells of different germ layers. Epigenetic factors, including DNA modifications, take a significant part in regulation of pluripotency and differentiation of ESCs. Less is known about the epigenetic regulation of these processes in adult stem cells. Gene expression profile and location of DNA modifications in adipose-derived stem cells (ADSCs) and their osteogenically differentiated lineages were analyzed using Agilent microarrays. Methylation-specific PCR and restriction-based quantitative PCR were applied for 5-methylcytosine (5mC) and 5-hydroxymethylcytosine (5hmC) detection in selected loci. The level of DNA modifications in the POU5F1 locus was quantified with deep sequencing. Expression levels of selected genes were assayed by real-time PCR. ADSCs differentiation into osteogenic lineages involved marked changes in both 5mC and 5hmC profiles, but 5hmC changes were more abundant. 5mC losses and 5hmC gains were the main events observed during ADSCs differentiation, and were accompanied by increased expression of TET1 (P = 0.009). In ADSCs, POU5F1 was better expressed than NANOG or SOX2 (P ≤ 0.001). Both 5mC and 5hmC marks were present in the POU5F1 locus, but only hydroxymethylation of specific cytosine showed significant effect on the gene expression. In summary, the data of our study suggest significant involvement of changes in 5hmC profile during the differentiation of human adult stem cells.

  7. Telocytes and putative stem cells in ageing human heart.

    PubMed

    Popescu, Laurentiu M; Curici, Antoanela; Wang, Enshi; Zhang, Hao; Hu, Shengshou; Gherghiceanu, Mihaela

    2015-01-01

    Tradition considers that mammalian heart consists of about 70% non-myocytes (interstitial cells) and 30% cardiomyocytes (CMs). Anyway, the presence of telocytes (TCs) has been overlooked, since they were described in 2010 (visit www.telocytes.com). Also, the number of cardiac stem cells (CSCs) has not accurately estimated in humans during ageing. We used electron microscopy to identify and estimate the number of cells in human atrial myocardium (appendages). Three age-related groups were studied: newborns (17 days-1 year), children (6-17 years) and adults (34-60 years). Morphometry was performed on low-magnification electron microscope images using computer-assisted technology. We found that interstitial area gradually increases with age from 31.3 ± 4.9% in newborns to 41 ± 5.2% in adults. Also, the number of blood capillaries (per mm(2) ) increased with several hundreds in children and adults versus newborns. CMs are the most numerous cells, representing 76% in newborns, 88% in children and 86% in adults. Images of CMs mitoses were seen in the 17-day newborns. Interestingly, no lipofuscin granules were found in CMs of human newborns and children. The percentage of cells that occupy interstitium were (depending on age): endothelial cells 52-62%; vascular smooth muscle cells and pericytes 22-28%, Schwann cells with nerve endings 6-7%, fibroblasts 3-10%, macrophages 1-8%, TCs about 1% and stem cells less than 1%. We cannot confirm the popular belief that cardiac fibroblasts are the most prevalent cell type in the heart and account for about 20% of myocardial volume. Numerically, TCs represent a small fraction of human cardiac interstitial cells, but because of their extensive telopodes, they achieve a 3D network that, for instance, supports CSCs. The myocardial (very) low capability to regenerate may be explained by the number of CSCs, which decreases fivefold by age (from 0.5% to 0.1% in newborns versus adults).

  8. FDA regulation of adult stem cell therapies as used in sports medicine.

    PubMed

    Chirba, Mary Ann; Sweetapple, Berkley; Hannon, Charles P; Anderson, John A

    2015-02-01

    In sports medicine, adult stem cells are the subject of great interest. Several uses of stem cells are under investigation including cartilage repair, meniscal regeneration, anterior cruciate ligament reconstruction, and tendinopathy. Extensive clinical and basic science research is warranted as stem cell therapies become increasingly common in clinical practice. In the United States, the Food and Drug Administration (FDA) is responsible for regulating the use of stem cells through its "Human Cells, Tissues, and Cellular and Tissue-Based Products" regulations. This report provides a brief overview of FDA regulation of adult stem cells. Several common clinical case scenarios are then presented that highlight how stem cells are currently being used in sports medicine and how current FDA regulations are likely to affect the physicians who use them. In the process, it explains how a variety of factors in sourcing and handling these cells, particularly the extent of cell manipulation, will affect what a physician can and cannot do without first obtaining the FDA's express approval.

  9. Successful isolation of viable adipose-derived stem cells from human adipose tissue subject to long-term cryopreservation: positive implications for adult stem cell-based therapeutics in patients of advanced age.

    PubMed

    Devitt, Sean M; Carter, Cynthia M; Dierov, Raia; Weiss, Scott; Gersch, Robert P; Percec, Ivona

    2015-01-01

    We examined cell isolation, viability, and growth in adipose-derived stem cells harvested from whole adipose tissue subject to different cryopreservation lengths (2-1159 days) from patients of varying ages (26-62 years). Subcutaneous abdominal adipose tissue was excised during abdominoplasties and was cryopreserved. The viability and number of adipose-derived stem cells isolated were measured after initial isolation and after 9, 18, and 28 days of growth. Data were analyzed with respect to cryopreservation duration and patient age. Significantly more viable cells were initially isolated from tissue cryopreserved <1 year than from tissue cryopreserved >2 years, irrespective of patient age. However, this difference did not persist with continued growth and there were no significant differences in cell viability or growth at subsequent time points with respect to cryopreservation duration or patient age. Mesenchymal stem cell markers were maintained in all cohorts tested throughout the duration of the study. Consequently, longer cryopreservation negatively impacts initial live adipose-derived stem cell isolation; however, this effect is neutralized with continued cell growth. Patient age does not significantly impact stem cell isolation, viability, or growth. Cryopreservation of adipose tissue is an effective long-term banking method for isolation of adipose-derived stem cells in patients of varying ages.

  10. Successful Isolation of Viable Adipose-Derived Stem Cells from Human Adipose Tissue Subject to Long-Term Cryopreservation: Positive Implications for Adult Stem Cell-Based Therapeutics in Patients of Advanced Age

    PubMed Central

    Devitt, Sean M.; Carter, Cynthia M.; Dierov, Raia; Weiss, Scott; Percec, Ivona

    2015-01-01

    We examined cell isolation, viability, and growth in adipose-derived stem cells harvested from whole adipose tissue subject to different cryopreservation lengths (2–1159 days) from patients of varying ages (26–62 years). Subcutaneous abdominal adipose tissue was excised during abdominoplasties and was cryopreserved. The viability and number of adipose-derived stem cells isolated were measured after initial isolation and after 9, 18, and 28 days of growth. Data were analyzed with respect to cryopreservation duration and patient age. Significantly more viable cells were initially isolated from tissue cryopreserved <1 year than from tissue cryopreserved >2 years, irrespective of patient age. However, this difference did not persist with continued growth and there were no significant differences in cell viability or growth at subsequent time points with respect to cryopreservation duration or patient age. Mesenchymal stem cell markers were maintained in all cohorts tested throughout the duration of the study. Consequently, longer cryopreservation negatively impacts initial live adipose-derived stem cell isolation; however, this effect is neutralized with continued cell growth. Patient age does not significantly impact stem cell isolation, viability, or growth. Cryopreservation of adipose tissue is an effective long-term banking method for isolation of adipose-derived stem cells in patients of varying ages. PMID:25945096

  11. EMPOWERING ADULT STEM CELLS FOR MYOCARDIAL REGENERATION

    PubMed Central

    Mohsin, Sadia; Siddiqi, Sailay; Collins, Brett; Sussman, Mark A.

    2012-01-01

    Treatment strategies for heart failure remain a high priority for ongoing research due to the profound unmet need in clinical disease coupled with lack of significant translational progress. The underlying issue is the same whether the cause is acute damage, chronic stress from disease, or aging: progressive loss of functional cardiomyocytes and diminished hemodynamic output. To stave off cardiomyocyte losses, a number of strategic approaches have been embraced in recent years involving both molecular and cellular approaches to augment myocardial structure and performance. Resultant excitement surrounding regenerative medicine in the heart has been tempered by realizations that reparative processes in the heart are insufficient to restore damaged myocardium to normal functional capacity and that cellular cardiomyoplasty is hampered by poor survival, proliferation, engraftment and differentiation of the donated population. To overcome these limitations, a combination of molecular and cellular approaches needs to be adopted involving use of genetic engineering to enhance resistance to cell death and increase regenerative capacity. This review will highlight biological properties of approached to potentiate stem cell-mediated regeneration to promote enhanced myocardial regeneration, persistence of donated cells, and long lasting tissue repair. Optimizing cell delivery and harnessing the power of survival signaling cascades for ex vivo genetic modification of stem cells prior to reintroduction into the patient will be critical to enhance the efficacy of cellular cardiomyoplasty. Once this goal is achieved, then cell-based therapy has great promise for treatment of heart failure to combat the loss of cardiac structure and function associated with acute damage, chronic disease or aging. PMID:22158649

  12. Haploidentical Stem Cell Transplantation in Adult Haematological Malignancies

    PubMed Central

    Parmesar, Kevon; Raj, Kavita

    2016-01-01

    Haematopoietic stem cell transplantation is a well-established treatment option for both hematological malignancies and nonmalignant conditions such as aplastic anemia and haemoglobinopathies. For those patients lacking a suitable matched sibling or matched unrelated donor, haploidentical donors are an alternative expedient donor pool. Historically, haploidentical transplantation led to high rates of graft rejection and GVHD. Strategies to circumvent these issues include T cell depletion and management of complications thereof or T replete transplants with GVHD prophylaxis. This review is an overview of these strategies and contemporaneous outcomes for hematological malignancies in adult haploidentical stem cell transplant recipients. PMID:27313619

  13. Techniques of Human Embryonic Stem Cell and Induced Pluripotent Stem Cell Derivation.

    PubMed

    Lewandowski, Jarosław; Kurpisz, Maciej

    2016-10-01

    Developing procedures for the derivation of human pluripotent stem cells (PSCs) gave rise to novel pathways into regenerative medicine research. For many years, stem cells have attracted attention as a potentially unlimited cell source for cellular therapy in neurodegenerative disorders, cardiovascular diseases, and spinal cord injuries, for example. In these studies, adult stem cells were insufficient; therefore, many attempts were made to obtain PSCs by other means. This review discusses key issues concerning the techniques of pluripotent cell acquisition. Technical and ethical issues hindered the medical use of somatic cell nuclear transfer and embryonic stem cells. Therefore, induced PSCs (iPSCs) emerged as a powerful technique with great potential for clinical applications, patient-specific disease modelling and pharmaceutical studies. The replacement of viral vectors or the administration of analogous proteins or chemical compounds during cell reprogramming are modifications designed to reduce tumorigenesis risk and to augment the procedure efficiency. Intensified analysis of new PSC lines revealed other barriers to overcome, such as epigenetic memory, disparity between human and mouse pluripotency, and variable response to differentiation of some iPSC lines. Thus, multidimensional verification must be conducted to fulfil strict clinical-grade requirements. Nevertheless, the first clinical trials in patients with spinal cord injury and macular dystrophy were recently carried out with differentiated iPSCs, encouraging alternative strategies for potential autologous cellular therapies.

  14. HLA Engineering of Human Pluripotent Stem Cells

    PubMed Central

    Riolobos, Laura; Hirata, Roli K; Turtle, Cameron J; Wang, Pei-Rong; Gornalusse, German G; Zavajlevski, Maja; Riddell, Stanley R; Russell, David W

    2013-01-01

    The clinical use of human pluripotent stem cells and their derivatives is limited by the rejection of transplanted cells due to differences in their human leukocyte antigen (HLA) genes. This has led to the proposed use of histocompatible, patient-specific stem cells; however, the preparation of many different stem cell lines for clinical use is a daunting task. Here, we develop two distinct genetic engineering approaches that address this problem. First, we use a combination of gene targeting and mitotic recombination to derive HLA-homozygous embryonic stem cell (ESC) subclones from an HLA-heterozygous parental line. A small bank of HLA-homozygous stem cells with common haplotypes would match a significant proportion of the population. Second, we derive HLA class I–negative cells by targeted disruption of both alleles of the Beta-2 Microglobulin (B2M) gene in ESCs. Mixed leukocyte reactions and peptide-specific HLA-restricted CD8+ T cell responses were reduced in class I–negative cells that had undergone differentiation in embryoid bodies. These B2M−/− ESCs could act as universal donor cells in applications where the transplanted cells do not express HLA class II genes. Both approaches used adeno-associated virus (AAV) vectors for efficient gene targeting in the absence of potentially genotoxic nucleases, and produced pluripotent, transgene-free cell lines. PMID:23629003

  15. Human-Mouse Chimerism Validates Human Stem Cell Pluripotency.

    PubMed

    Mascetti, Victoria L; Pedersen, Roger A

    2016-01-01

    Pluripotent stem cells are defined by their capacity to differentiate into all three tissue layers that comprise the body. Chimera formation, generated by stem cell transplantation to the embryo, is a stringent assessment of stem cell pluripotency. However, the ability of human pluripotent stem cells (hPSCs) to form embryonic chimeras remains in question. Here we show using a stage-matching approach that human induced pluripotent stem cells (hiPSCs) and human embryonic stem cells (hESCs) have the capacity to participate in normal mouse development when transplanted into gastrula-stage embryos, providing in vivo functional validation of hPSC pluripotency. hiPSCs and hESCs form interspecies chimeras with high efficiency, colonize the embryo in a manner predicted from classical developmental fate mapping, and differentiate into each of the three primary tissue layers. This faithful recapitulation of tissue-specific fate post-transplantation underscores the functional potential of hPSCs and provides evidence that human-mouse interspecies developmental competency can occur.

  16. Fetal programming of adult Leydig cell function by androgenic effects on stem/progenitor cells

    PubMed Central

    Kilcoyne, Karen R.; Smith, Lee B.; Atanassova, Nina; Macpherson, Sheila; McKinnell, Chris; van den Driesche, Sander; Jobling, Matthew S.; Chambers, Thomas J. G.; De Gendt, Karel; Verhoeven, Guido; O’Hara, Laura; Platts, Sophie; Renato de Franca, Luiz; Lara, Nathália L. M.; Anderson, Richard A.; Sharpe, Richard M.

    2014-01-01

    Fetal growth plays a role in programming of adult cardiometabolic disorders, which in men, are associated with lowered testosterone levels. Fetal growth and fetal androgen exposure can also predetermine testosterone levels in men, although how is unknown, because the adult Leydig cells (ALCs) that produce testosterone do not differentiate until puberty. To explain this conundrum, we hypothesized that stem cells for ALCs must be present in the fetal testis and might be susceptible to programming by fetal androgen exposure during masculinization. To address this hypothesis, we used ALC ablation/regeneration to identify that, in rats, ALCs derive from stem/progenitor cells that express chicken ovalbumin upstream promoter transcription factor II. These stem cells are abundant in the fetal testis of humans and rodents, and lineage tracing in mice shows that they develop into ALCs. The stem cells also express androgen receptors (ARs). Reduction in fetal androgen action through AR KO in mice or dibutyl phthalate (DBP) -induced reduction in intratesticular testosterone in rats reduced ALC stem cell number by ∼40% at birth to adulthood and induced compensated ALC failure (low/normal testosterone and elevated luteinizing hormone). In DBP-exposed males, this failure was probably explained by reduced testicular steroidogenic acute regulatory protein expression, which is associated with increased histone methylation (H3K27me3) in the proximal promoter. Accordingly, ALCs and ALC stem cells immunoexpressed increased H3K27me3, a change that was also evident in ALC stem cells in fetal testes. These studies highlight how a key component of male reproductive development can fundamentally reprogram adult hormone production (through an epigenetic change), which might affect lifetime disease risk. PMID:24753613

  17. Biology of the adult enteric neural stem cell.

    PubMed

    Estrada-Mondaca, Sandino; Carreón-Rodríguez, Alfonso; Belkind-Gerson, Jaime

    2007-01-01

    An increasing body of evidence has accumulated in recent years supporting the existence of neural stem cells in the adult gut. There are at least three groups that have obtained them using different methodologies and have described them in vitro. There is a growing amount of knowledge on their biology, but many questions are yet unanswered. Among these questions is whether these cells are part of a permanent undifferentiated pool or are recruited in a regular basis; in addition, the factors and genes involved in their survival, proliferation, migration, and differentiation are largely unknown. Finally, with between 10 and 20% of adults suffering from diseases involving the enteric nervous system, most notably irritable bowel syndrome and gastroesophageal reflux, what is the possible role of enteric nervous stem cells in health and disease?

  18. Stem Cell-Mediated Regeneration of the Adult Brain

    PubMed Central

    Jessberger, Sebastian

    2016-01-01

    Acute or chronic injury of the adult mammalian brain is often associated with persistent functional deficits as its potential for regeneration and capacity to rebuild lost neural structures is limited. However, the discovery that neural stem cells (NSCs) persist throughout life in discrete regions of the brain, novel approaches to induce the formation of neuronal and glial cells, and recently developed strategies to generate tissue for exogenous cell replacement strategies opened novel perspectives how to regenerate the adult brain. Here, we will review recently developed approaches for brain repair and discuss future perspectives that may eventually allow for developing novel treatment strategies in acute and chronic brain injury. PMID:27781019

  19. Use of genome editing tools in human stem cell-based disease modeling and precision medicine.

    PubMed

    Yuda, Wei; Shuang, Li; Gaigai, Liu; Yongxian, Zhang; Qiurong, Ding

    2015-10-01

    Precision medicine emerges as a new approach that takes into account individual variability. The successful conduct of precision medicine requires the use of precise disease models. Human pluripotent stem cells (hPSCs), as well as adult stem cells, can be differentiated into a variety of human somatic cell types that can be used for research and drug screening. The development of genome editing technology over the past few years, especially the CRISPR/Cas system, has made it feasible to precisely and efficiently edit the genetic background. Therefore, disease modeling by using a combination of human stem cells and genome editing technology has offered a new platform to generate " personalized " disease models, which allow the study of the contribution of individual genetic variabilities to disease progression and the development of precise treatments. In this review, recent advances in the use of genome editing in human stem cells and the generation of stem cell models for rare diseases and cancers are discussed.

  20. Hematopoietic stem cells, progenitor cells and leukemic stem cells in adult myeloproliferative neoplasms.

    PubMed

    Ng, Ashley P

    2013-05-01

    The understanding of myeloproliferative neoplasms has changed dramatically since Dameshek proposed his classification over 50 years ago. Our knowledge of the types of cells which constitute the hematopoietic system and of how they are regulated has also appreciated significantly over this time. This review relates what is currently known about the acquired genetic mutations associated with adult myeloproliferative neoplasms to how they lead to the hematopoietic perturbations of myeloproliferative disease. There is a particular focus on how stem and progenitor cell compartments are affected by BCR-ABL1 and JAK2V617F mutations, and the particular issue of resistance of leukemic stem cells to conventional and targeted therapies. PMID:23013358

  1. Enriched retinal ganglion cells derived from human embryonic stem cells.

    PubMed

    Gill, Katherine P; Hung, Sandy S C; Sharov, Alexei; Lo, Camden Y; Needham, Karina; Lidgerwood, Grace E; Jackson, Stacey; Crombie, Duncan E; Nayagam, Bryony A; Cook, Anthony L; Hewitt, Alex W; Pébay, Alice; Wong, Raymond C B

    2016-01-01

    Optic neuropathies are characterised by a loss of retinal ganglion cells (RGCs) that lead to vision impairment. Development of cell therapy requires a better understanding of the signals that direct stem cells into RGCs. Human embryonic stem cells (hESCs) represent an unlimited cellular source for generation of human RGCs in vitro. In this study, we present a 45-day protocol that utilises magnetic activated cell sorting to generate enriched population of RGCs via stepwise retinal differentiation using hESCs. We performed an extensive characterization of these stem cell-derived RGCs by examining the gene and protein expressions of a panel of neural/RGC markers. Furthermore, whole transcriptome analysis demonstrated similarity of the hESC-derived RGCs to human adult RGCs. The enriched hESC-RGCs possess long axons, functional electrophysiological profiles and axonal transport of mitochondria, suggestive of maturity. In summary, this RGC differentiation protocol can generate an enriched population of functional RGCs from hESCs, allowing future studies on disease modeling of optic neuropathies and development of cell therapies. PMID:27506453

  2. Enriched retinal ganglion cells derived from human embryonic stem cells

    PubMed Central

    Gill, Katherine P.; Hung, Sandy S. C.; Sharov, Alexei; Lo, Camden Y.; Needham, Karina; Lidgerwood, Grace E.; Jackson, Stacey; Crombie, Duncan E.; Nayagam, Bryony A.; Cook, Anthony L.; Hewitt, Alex W.; Pébay, Alice; Wong, Raymond C. B.

    2016-01-01

    Optic neuropathies are characterised by a loss of retinal ganglion cells (RGCs) that lead to vision impairment. Development of cell therapy requires a better understanding of the signals that direct stem cells into RGCs. Human embryonic stem cells (hESCs) represent an unlimited cellular source for generation of human RGCs in vitro. In this study, we present a 45-day protocol that utilises magnetic activated cell sorting to generate enriched population of RGCs via stepwise retinal differentiation using hESCs. We performed an extensive characterization of these stem cell-derived RGCs by examining the gene and protein expressions of a panel of neural/RGC markers. Furthermore, whole transcriptome analysis demonstrated similarity of the hESC-derived RGCs to human adult RGCs. The enriched hESC-RGCs possess long axons, functional electrophysiological profiles and axonal transport of mitochondria, suggestive of maturity. In summary, this RGC differentiation protocol can generate an enriched population of functional RGCs from hESCs, allowing future studies on disease modeling of optic neuropathies and development of cell therapies. PMID:27506453

  3. GLIOMA STEM CELL HOMING TO THE HUMAN SUBVENTRICULAR ZONE

    PubMed Central

    Sanai, Nader; Kusne, Yael; Su, Ning; Hemdan, Sandy; Mirzadeh, Zaman; Shi, Fu-Dong

    2014-01-01

    BACKGROUND: (blind field) METHODS: Using intraoperatively-derived human tissue, we employ a combination of cell culture, FACS-sorting, and immunohistochemistry techniques to characterize human glioma and GSC homing to the SVZ, as well as identify new, targetable cytokine pathways relevant to this niche. RESULTS: Our analysis identifies glioma cells homing to human SVZ and co-opting the gap layer for tangential migration. Interestingly, we find that human SVZ-invasive glioma cells are 50-fold more likely to function as glioma stem cells. Organotypic slice culture and co-culture assays also identify chemotactic effects, associated integrins, and related extracellular matrix proteins of 3 SVZ-enriched ligands (EGF, PDGF-BB, and SDF1) known to drive pro-migratory glioma pathways. CONCLUSIONS: Despite its quiescence, the adult human SVZ retains the signaling machinery to support cell migration and these mechanisms may be co-opted by glioma stem cells during subependymal spread in humans. SECONDARY CATEGORY: n/a.

  4. Regenerative medicine using adult neural stem cells: the potential for diabetes therapy and other pharmaceutical applications.

    PubMed

    Kuwabara, Tomoko; Asashima, Makoto

    2012-06-01

    Neural stem cells (NSCs), which are responsible for continuous neurogenesis during the adult stage, are present in human adults. The typical neurogenic regions are the hippocampus and the subventricular zone; recent studies have revealed that NSCs also exist in the olfactory bulb. Olfactory bulb-derived neural stem cells (OB NSCs) have the potential to be used in therapeutic applications and can be easily harvested without harm to the patient. Through the combined influence of extrinsic cues and innate programming, adult neurogenesis is a finely regulated process occurring in a specialized cellular environment, a niche. Understanding the regulatory mechanisms of adult NSCs and their cellular niche is not only important to understand the physiological roles of neurogenesis in adulthood, but also to provide the knowledge necessary for developing new therapeutic applications using adult NSCs in other organs with similar regulatory environments. Diabetes is a devastating disease affecting more than 200 million people worldwide. Numerous diabetic patients suffer increased symptom severity after the onset, involving complications such as retinopathy and nephropathy. Therefore, the development of treatments for fundamental diabetes is important. The utilization of autologous cells from patients with diabetes may address challenges regarding the compatibility of donor tissues as well as provide the means to naturally and safely restore function, reducing future risks while also providing a long-term cure. Here, we review recent findings regarding the use of adult OB NSCs as a potential diabetes cure, and discuss the potential of OB NSC-based pharmaceutical applications for neuronal diseases and mental disorders.

  5. Multipotent stem cells in the Malpighian tubules of adult Drosophila melanogaster

    PubMed Central

    Singh, Shree Ram; Hou, Steven X.

    2009-01-01

    Summary Excretion is an essential process of an organism's removal of the waste products of metabolism to maintain a constant chemical composition of the body fluids despite changes in the external environment. Excretion is performed by the kidneys in vertebrates and by Malpighian tubules (MTs) in Drosophila. The kidney serves as an excellent model organ to investigate the cellular and molecular mechanisms underlying organogenesis. Mammals and Drosophila share common principles of renal development. Tissue homeostasis, which is accomplished through self-renewal or differentiation of stem cells, is critical for the maintenance of adult tissues throughout the lifetime of an animal. Growing evidence suggests that stem cell self-renewal and differentiation is controlled by both intrinsic and extrinsic factors. Deregulation of stem cell behavior results in cancer formation, tissue degeneration, and premature aging. The mammalian kidney has a low rate of cellular turnover but has a great capacity for tissue regeneration following an ischemic injury. However, there is an ongoing controversy about the source of regenerating cells in the adult kidney that repopulate injured renal tissues. Recently, we identified multipotent stem cells in the MTs of adult Drosophila and found that these stem cells are able to proliferate and differentiate in several types of cells in MTs. Furthermore, we demonstrated that an autocrine JAK-STAT (Janus kinase–signal transducers and activators of transcription) signaling regulates stem cell self-renewal or differentiation of renal stem cells. The Drosophila MTs provide an excellent in vivo system for studying the renal stem cells at cellular and molecular levels. Understanding the molecular mechanisms governing stem cell self-renewal or differentiation in vivo is not only crucial to using stem cells for future regenerative medicine and gene therapy, but it also will increase our understanding of the mechanisms underlying cancer formation

  6. Adult stem cell and mesenchymal progenitor theories of aging

    PubMed Central

    Fukada, So-ichiro; Ma, Yuran; Uezumi, Akiyoshi

    2014-01-01

    Advances in medical science and technology allow people live longer lives, which results in age-related problems. Humans cannot avoid the various aged-related alterations of aging; in other words, humans cannot remain young at molecular and cellular levels. In 1956, Harman proposed the “free radical theory of aging” to explain the molecular mechanisms of aging. Telomere length, and accumulation of DNA or mitochondrial damage are also considered to be mechanisms of aging. On the other hand, stem cells are essential for maintaining tissue homeostasis by replacing parenchymal cells; therefore, the stem cell theory of aging is also used to explain the progress of aging. Importantly, the stem cell theory of aging is likely related to other theories. In addition, recent studies have started to reveal the essential roles of tissue-resident mesenchymal progenitors/stem cells/stromal cells in maintaining tissue homeostasis, and some evidence of their fundamental roles in the progression of aging has been presented. In this review, we discuss how stem cell and other theories connect to explain the progress of aging. In addition, we consider the mesenchymal progenitor theory of aging to describing the process of aging. PMID:25364718

  7. Adult Palatum as a Novel Source of Neural Crest-Related Stem Cells

    PubMed Central

    Widera, Darius; Zander, Christin; Heidbreder, Meike; Kasperek, Yvonne; Noll, Thomas; Seitz, Oliver; Saldamli, Belma; Sudhoff, Holger; Sader, Robert; Kaltschmidt, Christian; Kaltschmidt, Barbara

    2009-01-01

    Somatic neural and neural crest stem cells are promising sources for cellular therapy of several neurodegenerative diseases. However, because of practical considerations such as inadequate accessibility of the source material, the application of neural crest stem cells is strictly limited. The secondary palate is a highly regenerative and heavily innervated tissue, which develops embryonically under direct contribution of neural crest cells. Here, we describe for the first time the presence of nestin-positive neural crest-related stem cells within Meissner corpuscles and Merkel cell-neurite complexes located in the hard palate of adult Wistar rats. After isolation, palatal neural crest-related stem cells (pNC-SCs) were cultivated in the presence of epidermal growth factor and fibroblast growth factor under serum-free conditions, resulting in large amounts of neurospheres. We used immunocytochemical techniques and reverse transcriptase-polymerase chain reaction to assess the expression profile of pNC-SCs. In addition to the expression of neural crest stem cell markers such as Nestin, Sox2, and p75, we detected the expression of Klf4, Oct4, and c-Myc. pNC-SCs differentiated efficiently into neuronal and glial cells. Finally, we investigated the potential expression of stemness markers within the human palate. We identified expression of stem cell markers nestin and CD133 and the transcription factors needed for reprogramming of somatic cells into pluripotent cells: Sox2, Oct4, Klf4, and c-Myc. These data show that cells isolated from palatal rugae form neurospheres, are highly plastic, and express neural crest stem cell markers. In addition, pNC-SCs may have the ability to differentiate into functional neurons and glial cells, serving as a starting point for therapeutic studies. Stem Cells 2009;27:1899–1910 PMID:19544446

  8. Isolation and Characterization of Pluripotent Human Spermatogonial Stem Cell-Derived Cells

    PubMed Central

    Kossack, Nina; Meneses, Juanito; Shefi, Shai; Nguyen, Ha Nam; Chavez, Shawn; Nicholas, Cory; Gromoll, Joerg; Turek, Paul J; Reijo-Pera, Renee A

    2009-01-01

    Several reports have documented the derivation of pluripotent cells (multipotent germline stem cells) from spermatogonial stem cells obtained from the adult mouse testis. These spermatogonia-derived stem cells express embryonic stem cell markers and differentiate to the three primary germ layers, as well as the germline. Data indicate that derivation may involve reprogramming of endogenous spermatogonia in culture. Here, we report the derivation of human multipotent germline stem cells (hMGSCs) from a testis biopsy. The cells express distinct markers of pluripotency, form embryoid bodies that contain derivatives of all three germ layers, maintain a normal XY karyotype, are hypomethylated at the H19 locus, and express high levels of telomerase. Teratoma assays indicate the presence of human cells 8 weeks post-transplantation but limited teratoma formation. Thus, these data suggest the potential to derive pluripotent cells from human testis biopsies but indicate a need for novel strategies to optimize hMGSC culture conditions and reprogramming. PMID:18927477

  9. Directed Differentiation of Human-Induced Pluripotent Stem Cells to Mesenchymal Stem Cells.

    PubMed

    Lian, Qizhou; Zhang, Yuelin; Liang, Xiaoting; Gao, Fei; Tse, Hung-Fat

    2016-01-01

    Multipotent stromal cells, also known as mesenchymal stem cells (MSCs), possess great potential to generate a wide range of cell types including endothelial cells, smooth muscle cells, bone, cartilage, and lipid cells. This protocol describes in detail how to perform highly efficient, lineage-specific differentiation of human-induced pluripotent stem cells (iPSCs) with an MSCs fate. The approach uses a clinically compliant protocol with chemically defined media, feeder-free conditions, and a CD105 positive and CD24 negative selection to achieve a single cell-based MSCs derivation from differentiating human pluripotent cells in approximately 20 days. Cells generated with this protocol express typical MSCs surface markers and undergo adipogenesis, osteogenesis, and chondrogenesis similar to adult bone marrow-derived MSCs (BM-MSCs). Nonetheless, compared with adult BM-MSCs, iPSC-MSCs display a higher proliferative capacity, up to 120 passages, without obvious loss of self-renewal potential and constitutively express MSCs surface antigens. MSCs generated with this protocol have numerous applications, including expansion to large scale cell numbers for tissue engineering and the development of cellular therapeutics. This approach has been used to rescue limb ischemia, allergic disorders, and cigarette smoke-induced lung damage and to model mesenchymal and vascular disorders of Hutchinson-Gilford progeria syndrome (HGPS). PMID:27236679

  10. Evolutionary dynamics of adult stem cells: Comparison of random and immortal-strand segregation mechanisms

    NASA Astrophysics Data System (ADS)

    Tannenbaum, Emmanuel; Sherley, James L.; Shakhnovich, Eugene I.

    2005-04-01

    This paper develops a point-mutation model describing the evolutionary dynamics of a population of adult stem cells. Such a model may prove useful for quantitative studies of tissue aging and the emergence of cancer. We consider two modes of chromosome segregation: (1) random segregation, where the daughter chromosomes of a given parent chromosome segregate randomly into the stem cell and its differentiating sister cell and (2) “immortal DNA strand” co-segregation, for which the stem cell retains the daughter chromosomes with the oldest parent strands. Immortal strand co-segregation is a mechanism, originally proposed by [Cairns Nature (London) 255, 197 (1975)], by which stem cells preserve the integrity of their genomes. For random segregation, we develop an ordered strand pair formulation of the dynamics, analogous to the ordered strand pair formalism developed for quasispecies dynamics involving semiconservative replication with imperfect lesion repair (in this context, lesion repair is taken to mean repair of postreplication base-pair mismatches). Interestingly, a similar formulation is possible with immortal strand co-segregation, despite the fact that this segregation mechanism is age dependent. From our model we are able to mathematically show that, when lesion repair is imperfect, then immortal strand co-segregation leads to better preservation of the stem cell lineage than random chromosome segregation. Furthermore, our model allows us to estimate the optimal lesion repair efficiency for preserving an adult stem cell population for a given period of time. For human stem cells, we obtain that mispaired bases still present after replication and cell division should be left untouched, to avoid potentially fixing a mutation in both DNA strands.

  11. Recent advances in bone regeneration using adult stem cells.

    PubMed

    Zigdon-Giladi, Hadar; Rudich, Utai; Michaeli Geller, Gal; Evron, Ayelet

    2015-04-26

    Bone is a highly vascularized tissue reliant on the close spatial and temporal association between blood vessels and bone cells. Therefore, cells that participate in vasculogenesis and osteogenesis play a pivotal role in bone formation during prenatal and postnatal periods. Nevertheless, spontaneous healing of bone fracture is occasionally impaired due to insufficient blood and cellular supply to the site of injury. In these cases, bone regeneration process is interrupted, which might result in delayed union or even nonunion of the fracture. Nonunion fracture is difficult to treat and have a high financial impact. In the last decade, numerous technological advancements in bone tissue engineering and cell-therapy opened new horizon in the field of bone regeneration. This review starts with presentation of the biological processes involved in bone development, bone remodeling, fracture healing process and the microenvironment at bone healing sites. Then, we discuss the rationale for using adult stem cells and listed the characteristics of the available cells for bone regeneration. The mechanism of action and epigenetic regulations for osteogenic differentiation are also described. Finally, we review the literature for translational and clinical trials that investigated the use of adult stem cells (mesenchymal stem cells, endothelial progenitor cells and CD34(+) blood progenitors) for bone regeneration.

  12. Insights from a Chimpanzee Adipose Stromal Cell Population: Opportunities for Adult Stem Cells to Expand Primate Functional Genomics

    PubMed Central

    Pfefferle, Lisa W.; Wray, Gregory A.

    2013-01-01

    Comparisons between humans and chimpanzees are essential for understanding traits unique to each species. However, linking important phenotypic differences to underlying molecular changes is often challenging. The ability to generate, differentiate, and profile adult stem cells provides a powerful but underutilized opportunity to investigate the molecular basis for trait differences between species within specific cell types and in a controlled environment. Here, we characterize adipose stromal cells (ASCs) from Clint, the chimpanzee whose genome was first sequenced. Using imaging and RNA-Seq, we compare the chimpanzee ASCs with three comparable human cell lines. Consistent with previous studies on ASCs in humans, the chimpanzee cells have fibroblast-like morphology and express genes encoding components of the extracellular matrix at high levels. Differentially expressed genes are enriched for distinct functional classes between species: immunity and protein processing are higher in chimpanzees, whereas cell cycle and DNA processing are higher in humans. Although hesitant to draw definitive conclusions from these data given the limited sample size, we wish to stress the opportunities that adult stem cells offer for studying primate evolution. In particular, adult stem cells provide a powerful means to investigate the profound disease susceptibilities unique to humans and a promising tool for conservation efforts with nonhuman primates. By allowing for experimental perturbations in relevant cell types, adult stem cells promise to complement classic comparative primate genomics based on in vivo sampling. PMID:24092797

  13. Clinical Trials of Adult Stem Cell Therapy in Patients with Ischemic Stroke.

    PubMed

    Bang, Oh Young

    2016-01-01

    Stem cell therapy is considered a potential regenerative strategy for patients with neurologic deficits. Studies involving animal models of ischemic stroke have shown that stem cells transplanted into the brain can lead to functional improvement. With current advances in the understanding regarding the effects of introducing stem cells and their mechanisms of action, several clinical trials of stem cell therapy have been conducted in patients with stroke since 2005, including studies using mesenchymal stem cells, bone marrow mononuclear cells, and neural stem/progenitor cells. In addition, several clinical trials of the use of adult stem cells to treat ischemic stroke are ongoing. This review presents the status of our understanding of adult stem cells and results from clinical trials, and introduces ongoing clinical studies of adult stem cell therapy in the field of stroke.

  14. Pluripotent stem cell transcription factors during human odontogenesis.

    PubMed

    da Cunha, Juliana Malta; da Costa-Neves, Adriana; Kerkis, Irina; da Silva, Marcelo Cavenaghi Pereira

    2013-09-01

    Stem cells are capable of generating various cell lines and can be obtained from adult or embryonic tissues for clinical therapies. Stem cells from deciduous dental pulp are among those that are easily obtainable from adult tissues and have been widely studied because of their ability to differentiate into a variety of cell lines in the presence of various chemical mediators. We have analyze the expression of several proteins related to the differentiation and proliferative potential of cell populations that compose the tooth germ of human fetuses. We evaluate 20 human fetuses of both genders. After being paraffin-embedded, cap and bell stages of tooth germ development were subjected to immunohistochemistry for the following markers: Oct-4, Nanog, Stat-3 and Sox-2. The studied antibodies showed nuclear or cytoplasmic immunnostaining within various anatomical structures and with various degrees of expression, indicating the action of these proteins during tooth development. We conclude that the interrelationship between these transcription factors is complex and associated with self-renewal and cell differentiation. Our results suggest that the expression of Oct-4, Nanog, Sox-2 and Stat-3 are related to differentiation in ameloblasts and odontoblasts.

  15. Challenges and strategies for generating therapeutic patient-specific hemangioblasts and hematopoietic stem cells from human pluripotent stem cells

    PubMed Central

    PETERS, ANN; BURRIDGE, PAUL W.; PRYZHKOVA, MARINA V.; LEVINE, MICHAL A.; PARK, TEA-SOON; ROXBURY, CHRISTOPHER; YUAN, XUAN; PÉAULT, BRUNO; ZAMBIDIS, ELIAS T.

    2012-01-01

    Recent characterization of hemangioblasts differentiated from human embryonic stem cells (hESC) has further confirmed evidence from murine, zebrafish and avian experimental systems that hematopoietic and endothelial lineages arise from a common progenitor. Such progenitors may provide a valuable resource for delineating the initial developmental steps of human hemato-endotheliogenesis, which is a process normally difficult to study due to the very limited accessibility of early human embryonic/fetal tissues. Moreover, efficient hemangioblast and hematopoietic stem cell (HSC) generation from patient-specific pluripotent stem cells has enormous potential for regenerative medicine, since it could lead to strategies for treating a multitude of hematologic and vascular disorders. However, significant scientific challenges remain in achieving these goals, and the generation of transplantable hemangioblasts and HSC derived from hESC currently remains elusive. Our previous work has suggested that the failure to derive engraftable HSC from hESC is due to the fact that current methodologies for differentiating hESC produce hematopoietic progenitors developmentally similar to those found in the human yolk sac, and are therefore too immature to provide adult-type hematopoietic reconstitution. Herein, we outline the nature of this challenge and propose targeted strategies for generating engraftable human pluripotent stem cell-derived HSC from primitive hemangioblasts using a developmental approach. We also focus on methods by which reprogrammed somatic cells could be used to derive autologous pluripotent stem cells, which in turn could provide unlimited sources of patient-specific hemangioblasts and HSC. PMID:20563986

  16. Adult stem cells applied to tissue engineering and regenerative medicine.

    PubMed

    Cuenca-López, M D; Zamora-Navas, P; García-Herrera, J M; Godino, M; López-Puertas, J M; Guerado, E; Becerra, J; Andrades, J A

    2008-01-01

    Regeneration takes place in the body at a moment or another throughout life. Bone, cartilage, and tendons (the key components of the structure and articulation in the body) have a limited capacity for self-repair and, after traumatic injury or disease, the regenerative power of adult tissue is often insufficient. When organs or tissues are irreparably damaged, they may be replaced by an artificial device or by a donor organ. However, the number of available donor organs is considerably limited. Generation of tissue-engineered replacement organs by extracting stem cells from the patient, growing them and modifying them in clinical conditions after re-introduction in the body represents an ideal source for corrective treatment. Mesenchymal stem cells (MSCs) are the multipotential progenitors that give rise to skeletal cells, vascular smooth muscle cells, muscle (skeletal and cardiac muscle), adipocytes (fat tissue) and hematopoietic (blood)-supportive stromal cells. MSCs are found in multiple connective tissues, in adult bone marrow, skeletal muscles and fat pads. The wide representation in adult tissues may be related to the existence of a circulating blood pool or that MSCs are associated to the vascular system.

  17. The decision on the "optimal" human pluripotent stem cell.

    PubMed

    Rosner, Margit; Schipany, Katharina; Hengstschläger, Markus

    2014-05-01

    Because of recent advances, the array of human pluripotent stem cells now contains embryonic stem cells, derived from "surplus" in vitro fertilization embryos or from cloned embryos; induced pluripotent stem cells; and amniotic fluid stem cells. Here, we compare these stem cell types regarding ethical and legal concerns, cultivation conditions, genomic stability, tumor developing potentials, and applicability for disease modeling and human therapy. This overview highlights that in the future appropriate methodological management must include a decision on the "optimal" stem cell to use before the specific application.

  18. Switching roles: the functional plasticity of adult tissue stem cells

    PubMed Central

    Wabik, Agnieszka; Jones, Philip H

    2015-01-01

    Adult organisms have to adapt to survive, and the same is true for their tissues. Rates and types of cell production must be rapidly and reversibly adjusted to meet tissue demands in response to both local and systemic challenges. Recent work reveals how stem cell (SC) populations meet these requirements by switching between functional states tuned to homoeostasis or regeneration. This plasticity extends to differentiating cells, which are capable of reverting to SCs after injury. The concept of the niche, the micro-environment that sustains and regulates stem cells, is broadening, with a new appreciation of the role of physical factors and hormonal signals. Here, we review different functions of SCs, the cellular mechanisms that underlie them and the signals that bias the fate of SCs as they switch between roles. PMID:25812989

  19. Human amniotic fluid stem cell preconditioning improves their regenerative potential.

    PubMed

    Rota, Cinzia; Imberti, Barbara; Pozzobon, Michela; Piccoli, Martina; De Coppi, Paolo; Atala, Anthony; Gagliardini, Elena; Xinaris, Christodoulos; Benedetti, Valentina; Fabricio, Aline S C; Squarcina, Elisa; Abbate, Mauro; Benigni, Ariela; Remuzzi, Giuseppe; Morigi, Marina

    2012-07-20

    Human amniotic fluid stem (hAFS) cells, a novel class of broadly multipotent stem cells that share characteristics of both embryonic and adult stem cells, have been regarded as promising candidate for cell therapy. Taking advantage by the well-established murine model of acute kidney injury (AKI), we studied the proregenerative effect of hAFS cells in immunodeficient mice injected with the nephrotoxic drug cisplatin. Infusion of hAFS cells in cisplatin mice improved renal function and limited tubular damage, although not to control level, and prolonged animal survival. Human AFS cells engrafted injured kidney predominantly in peritubular region without acquiring tubular epithelial markers. Human AFS cells exerted antiapoptotic effect, activated Akt, and stimulated proliferation of tubular cells possibly via local release of factors, including interleukin-6, vascular endothelial growth factor, and stromal cell-derived factor-1, which we documented in vitro to be produced by hAFS cells. The therapeutic potential of hAFS cells was enhanced by cell pretreatment with glial cell line-derived neurotrophic factor (GDNF), which markedly ameliorated renal function and tubular injury by increasing stem cell homing to the tubulointerstitial compartment. By in vitro studies, GDNF increased hAFS cell production of growth factors, motility, and expression of receptors involved in cell homing and survival. These findings indicate that hAFS cells can promote functional recovery and contribute to renal regeneration in AKI mice via local production of mitogenic and prosurvival factors. The effects of hAFS cells can be remarkably enhanced by GDNF preconditioning.

  20. Human Amniotic Fluid Stem Cell Preconditioning Improves Their Regenerative Potential

    PubMed Central

    Rota, Cinzia; Imberti, Barbara; Pozzobon, Michela; Piccoli, Martina; De Coppi, Paolo; Atala, Anthony; Gagliardini, Elena; Xinaris, Christodoulos; Benedetti, Valentina; Fabricio, Aline S.C.; Squarcina, Elisa; Abbate, Mauro; Benigni, Ariela; Remuzzi, Giuseppe

    2012-01-01

    Human amniotic fluid stem (hAFS) cells, a novel class of broadly multipotent stem cells that share characteristics of both embryonic and adult stem cells, have been regarded as promising candidate for cell therapy. Taking advantage by the well-established murine model of acute kidney injury (AKI), we studied the proregenerative effect of hAFS cells in immunodeficient mice injected with the nephrotoxic drug cisplatin. Infusion of hAFS cells in cisplatin mice improved renal function and limited tubular damage, although not to control level, and prolonged animal survival. Human AFS cells engrafted injured kidney predominantly in peritubular region without acquiring tubular epithelial markers. Human AFS cells exerted antiapoptotic effect, activated Akt, and stimulated proliferation of tubular cells possibly via local release of factors, including interleukin-6, vascular endothelial growth factor, and stromal cell–derived factor-1, which we documented in vitro to be produced by hAFS cells. The therapeutic potential of hAFS cells was enhanced by cell pretreatment with glial cell line–derived neurotrophic factor (GDNF), which markedly ameliorated renal function and tubular injury by increasing stem cell homing to the tubulointerstitial compartment. By in vitro studies, GDNF increased hAFS cell production of growth factors, motility, and expression of receptors involved in cell homing and survival. These findings indicate that hAFS cells can promote functional recovery and contribute to renal regeneration in AKI mice via local production of mitogenic and prosurvival factors. The effects of hAFS cells can be remarkably enhanced by GDNF preconditioning. PMID:22066606

  1. Adult stem cell plasticity: will engineered tissues be rejected?

    PubMed Central

    Fang, Te-Chao; Alison, Malcolm R; Wright, Nicholas A; Poulsom, Richard

    2004-01-01

    The dogma that adult tissue-specific stem cells remain committed to supporting only their own tissue has been challenged; a new hypothesis, that adult stem cells demonstrate plasticity in their repertoires, is being tested. This is important because it seems possible that haematopoietic stem cells, for example, could be exploited to generate and perhaps deliver cell-based therapies deep within existing nonhaematopoietic organs. Much of the evidence for plasticity derives from histological studies of tissues from patients or animals that have received grafts of cells or whole organs, from a donor bearing (or lacking) a definitive marker. Detection in the recipient of appropriately differentiated cells bearing the donor marker is indicative of a switch in phenotype of a stem cell or a member of a transit amplifying population or of a differentiated cell. In this review, we discuss evidence for these changes occurring but do not consider the molecular basis of cell commitment. In general, the extent of engraftment is low but may be increased if tissues are damaged. In model systems of liver regeneration, the repeated application of a selection pressure increases levels of engraftment considerably; how this occurs is unclear. Cell fusion plays a part in regeneration and remodelling of the liver, skeletal muscle and even regions of the brain. Genetic disease may be amenable to some forms of cell therapy, yet immune rejection will present challenges. Graft-vs.-host disease will continue to present problems, although this may be avoided if the cells were derived from the recipient or they were tolerized. Despite great expectations for cellular therapies, there are indications that attempts to replace missing proteins could be confounded simply by the development of specific immunity that rejects the new phenotype. PMID:15255965

  2. Stem cells on the brain: modeling neurodevelopmental and neurodegenerative diseases using human induced pluripotent stem cells.

    PubMed

    Srikanth, Priya; Young-Pearse, Tracy L

    2014-01-01

    Seven years have passed since the initial report of the generation of induced pluripotent stem cells (iPSCs) from adult human somatic cells, and in the intervening time the field of neuroscience has developed numerous disease models using this technology. Here, we review progress in the field and describe both the advantages and potential pitfalls of modeling neurodegenerative and neurodevelopmental diseases using this technology. We include tables with information on neural differentiation protocols and studies that developed human iPSC lines to model neurological diseases. We also discuss how one can: investigate effects of genetic mutations with iPSCs, examine cell fate-specific phenotypes, best determine the specificity of a phenotype, and bring in vivo relevance to this in vitro technique.

  3. Robust G2 pausing of adult stem cells in Hydra.

    PubMed

    Buzgariu, Wanda; Crescenzi, Marco; Galliot, Brigitte

    2014-01-01

    Hydra is a freshwater hydrozoan polyp that constantly renews its two tissue layers thanks to three distinct stem cell populations that cannot replace each other, epithelial ectodermal, epithelial endodermal, and multipotent interstitial. These adult stem cells, located in the central body column, exhibit different cycling paces, slow for the epithelial, fast for the interstitial. To monitor the changes in cell cycling in Hydra, we established a fast and efficient flow cytometry procedure, which we validated by confirming previous findings, as the Nocodazole-induced reversible arrest of cell cycling in G2/M, and the mitogenic signal provided by feeding. Then to dissect the cycling and differentiation behaviors of the interstitial stem cells, we used the AEP_cnnos1 and AEP_Icy1 transgenic lines that constitutively express GFP in this lineage. For the epithelial lineages we used the sf-1 strain that rapidly eliminates the fast cycling cells upon heat-shock and progressively becomes epithelial. This study evidences similar cycling patterns for the interstitial and epithelial stem cells, which all alternate between the G2 and S-phases traversing a minimal G1-phase. We also found interstitial progenitors with a shorter G2 that pause in G1/G0. At the animal extremities, most cells no longer cycle, the epithelial cells terminally differentiate in G2 and the interstitial progenitors in G1/G0. At the apical pole ~80% cells are post-mitotic differentiated cells, reflecting the higher density of neurons and nematocytes in this region. We discuss how the robust G2 pausing of stem cells, maintained over weeks of starvation, may contribute to regeneration.

  4. Derivation of Neural Precursor Cells from Human Embryonic Stem Cells for DNA Methylomic Analysis.

    PubMed

    Roubal, Ivan; Park, Sun Joo; Kim, Yong

    2016-01-01

    Embryonic stem cells are self-renewing pluripotent cells with competency to differentiate into all three-germ lineages. Many studies have demonstrated the importance of genetic and epigenetic molecular mechanisms in the maintenance of self-renewal and pluripotency. Stem cells are under unique molecular and cellular regulations different from somatic cells. Proper regulation should be ensured to maintain their unique self-renewal and undifferentiated characteristics. Understanding key mechanisms in stem cell biology will be important for the successful application of stem cells for regenerative therapeutic medicine. More importantly practical use of stem cells will require our knowledge on how to properly direct and differentiate stem cells into the necessary type of cells. Embryonic stem cells and adult stem cells have been used as study models to unveil molecular and cellular mechanisms in various signaling pathways. They are especially beneficial to developmental studies where in vivo molecular/cellular study models are not available. We have derived neural stem cells from human embryonic stem cells as a model to study the effect of teratogen in neural development. We have tested commercial neural differentiation system and successfully derived neural precursor cells exhibiting key molecular features of neural stem cells, which will be useful for experimental application.

  5. Isolation, culture and analysis of adult subependymal neural stem cells.

    PubMed

    Belenguer, Germán; Domingo-Muelas, Ana; Ferrón, Sacri R; Morante-Redolat, José Manuel; Fariñas, Isabel

    2016-01-01

    Individual cells dissected from the subependymal neurogenic niche of the adult mouse brain proliferate in medium containing basic fibroblast growth factor (bFGF) and/or epidermal growth factor (EGF) as mitogens, to produce multipotent clonal aggregates called neurospheres. These cultures constitute a powerful tool for the study of neural stem cells (NSCs) provided that they allow the analysis of their features and potential capacity in a controlled environment that can be modulated and monitored more accurately than in vivo. Clonogenic and population analyses under mitogen addition or withdrawal allow the quantification of the self-renewing and multilineage potency of these cells and the identification of the mechanisms involved in these properties. Here, we describe a set of procedures developed and/or modified by our group including several experimental options that can be used either independently or in combination for the ex vivo assessment of cell properties of NSCs obtained from the adult subependymal niche. PMID:27016251

  6. The molecular nature of very small embryonic-like stem cells in adult tissues.

    PubMed

    Kim, YongHwan; Jeong, Jaeho; Kang, Hyunsook; Lim, Jisun; Heo, Jinbeom; Ratajczak, Janina; Ratajczak, Mariusz Z; Shin, Dong-Myung

    2014-11-01

    Pluripotent stem cells (PSCs) have been considered as the most important cells in regenerative medicine as they are able to differentiate into all types of cells in the human body. PSCs have been established from several sources of embryo tissue or by reprogramming of terminally differentiated adult tissue by transduction of so-called Yamanaka factors (Oct4, Sox2, Klf4, and cMyc). Interestingly, accumulating evidence has demonstrated the residence of PSCs in adult tissue and with the ability to differentiate into multiple types of tissue-committed stem cells (TCSCs). We also recently demonstrated that a population of pluripotent Oct4(+) SSEA-1(+)Sca-1(+)Lin(-)CD45(-) very small embryonic-like stem cells (VSELs) resides in the adult murine bone marrow (BM) and in other murine tissue. These very small (∼3-6 μm) cells express pluripotent markers such as Oct4, Nanog, and SSEA-1. VSELs could be specified into several tissue-residing TCSCs in response to tissue/organ injury, and thus suggesting that these cells have a physiological role in the rejuvenation of a pool of TCSCs under steady-state conditions. In this review article, we discuss the molecular nature of the rare population of VSELs which have a crucial role in regulating the pluripotency, proliferation, differentiation, and aging of these cells. PMID:25473442

  7. Deciding on human embryonic stem cell research.

    PubMed

    Burgin, Eileen

    2009-03-01

    This paper examines the influences that congressional staff people viewed as important in shaping legislators' voting decisions on the human embryonic stem (ES) cell research bill in the 109th Congress, the first legislation vetoed by President George W. Bush. The analysis illuminates factors that impact congressional decision making on a salient issue with a strong moral component. Constituent concerns, ideology, and a desire to make good public policy all centrally affected members' choices; however, moral overtones permeated considerations relevant to the human ES cell research question. In addition, at least three influences that directly reflect or relate to members' moral claims - religious convictions, personal connections to potential beneficiaries of human ES cell research, and moral pressure from outside interests - were important also. The analysis draws on data gathered from interviews with congressional aides.

  8. Telocytes and putative stem cells in ageing human heart

    PubMed Central

    Popescu, Laurentiu M; Curici, Antoanela; Wang, Enshi; Zhang, Hao; Hu, Shengshou; Gherghiceanu, Mihaela

    2015-01-01

    Tradition considers that mammalian heart consists of about 70% non-myocytes (interstitial cells) and 30% cardiomyocytes (CMs). Anyway, the presence of telocytes (TCs) has been overlooked, since they were described in 2010 (visit http://www.telocytes.com). Also, the number of cardiac stem cells (CSCs) has not accurately estimated in humans during ageing. We used electron microscopy to identify and estimate the number of cells in human atrial myocardium (appendages). Three age-related groups were studied: newborns (17 days–1 year), children (6–17 years) and adults (34–60 years). Morphometry was performed on low-magnification electron microscope images using computer-assisted technology. We found that interstitial area gradually increases with age from 31.3 ± 4.9% in newborns to 41 ± 5.2% in adults. Also, the number of blood capillaries (per mm2) increased with several hundreds in children and adults versus newborns. CMs are the most numerous cells, representing 76% in newborns, 88% in children and 86% in adults. Images of CMs mitoses were seen in the 17-day newborns. Interestingly, no lipofuscin granules were found in CMs of human newborns and children. The percentage of cells that occupy interstitium were (depending on age): endothelial cells 52–62%; vascular smooth muscle cells and pericytes 22–28%, Schwann cells with nerve endings 6–7%, fibroblasts 3–10%, macrophages 1–8%, TCs about 1% and stem cells less than 1%. We cannot confirm the popular belief that cardiac fibroblasts are the most prevalent cell type in the heart and account for about 20% of myocardial volume. Numerically, TCs represent a small fraction of human cardiac interstitial cells, but because of their extensive telopodes, they achieve a 3D network that, for instance, supports CSCs. The myocardial (very) low capability to regenerate may be explained by the number of CSCs, which decreases fivefold by age (from 0.5% to 0.1% in newborns versus adults). PMID:25545142

  9. Genome editing in human stem cells.

    PubMed

    Byrne, Susan M; Mali, Prashant; Church, George M

    2014-01-01

    The use of custom-engineered sequence-specific nucleases (including CRISPR/Cas9, ZFN, and TALEN) allows genetic changes in human cells to be easily made with much greater efficiency and precision than before. Engineered double-stranded DNA breaks can efficiently disrupt genes, or, with the right donor vector, engineer point mutations and gene insertions. However, a number of design considerations should be taken into account to ensure maximum gene targeting efficiency and specificity. This is especially true when engineering human embryonic stem or induced pluripotent stem cells (iPSCs), which are more difficult to transfect and less resilient to DNA damage than immortalized tumor cell lines. Here, we describe a protocol for easily engineering genetic changes in human iPSCs, through which we typically achieve targeting efficiencies between 1% and 10% without any subsequent selection steps. Since this protocol only uses the simple transient transfection of plasmids and/or single-stranded oligonucleotides, most labs will easily be able to perform it. We also describe strategies for identifying, cloning, and genotyping successfully edited cells, and how to design the optimal sgRNA target sites and donor vectors. Finally, we discuss alternative methods for gene editing including viral delivery vectors, Cas9 nickases, and orthogonal Cas9 systems.

  10. Pluripotent states of human embryonic stem cells.

    PubMed

    Chen, Yifei; Lai, Dongmei

    2015-02-01

    Since human embryonic stem cells (hESCs) were first isolated and successfully cultured in vitro, the pluripotent potential of hESCs has been underestimated. The pluripotency of mouse embryonic stem cells (mESCs) can be categorized as naïve and primed, depending on their corresponding in vivo developing phases. mESC morphology differs at distinct pluripotent states, which differ in signaling dependence, gene expression, epigenetic features, and developmental potential. hESCs resemble mouse stem cells at primed pluripotency, and consequently are believed to correspond to a later developmental stage in vivo than mESCs. Nevertheless, recent studies indicate that a naïve state of pluripotency may exist in hESCs, and the pluripotency of hESCs also can be enhanced by genetic modification or optimized culture systems. These findings provide novel insight into the properties and differentiation potential of hESCs. Here, we review the recent advances in characterization of ESC states and investigate the mechanisms regulating hESC pluripotency. PMID:25393391

  11. Stem Cell Models of Human Brain Development.

    PubMed

    Kelava, Iva; Lancaster, Madeline A

    2016-06-01

    Recent breakthroughs in pluripotent stem cell technologies have enabled a new class of in vitro systems for functional modeling of human brain development. These advances, in combination with improvements in neural differentiation methods, allow the generation of in vitro systems that reproduce many in vivo features of the brain with remarkable similarity. Here, we describe advances in the development of these methods, focusing on neural rosette and organoid approaches, and compare their relative capabilities and limitations. We also discuss current technical hurdles for recreating the cell-type complexity and spatial architecture of the brain in culture and offer potential solutions.

  12. Maturing human pluripotent stem cell-derived cardiomyocytes in human engineered cardiac tissues.

    PubMed

    Feric, Nicole T; Radisic, Milica

    2016-01-15

    Engineering functional human cardiac tissue that mimics the native adult morphological and functional phenotype has been a long held objective. In the last 5 years, the field of cardiac tissue engineering has transitioned from cardiac tissues derived from various animal species to the production of the first generation of human engineered cardiac tissues (hECTs), due to recent advances in human stem cell biology. Despite this progress, the hECTs generated to date remain immature relative to the native adult myocardium. In this review, we focus on the maturation challenge in the context of hECTs, the present state of the art, and future perspectives in terms of regenerative medicine, drug discovery, preclinical safety testing and pathophysiological studies.

  13. Adult stem cells for cardiac repair: a choice between skeletal myoblasts and bone marrow stem cells.

    PubMed

    Ye, Lei; Haider, Husnain Kh; Sim, Eugene K W

    2006-01-01

    The real promise of a stem cell-based approach for cardiac regeneration and repair lies in the promotion of myogenesis and angiogenesis at the site of the cell graft to achieve both structural and functional benefits. Despite all of the progress and promise in this field, many unanswered questions remain; the answers to these questions will provide the much-needed breakthrough to harness the real benefits of cell therapy for the heart in the clinical perspective. One of the major issues is the choice of donor cell type for transplantation. Multiple cell types with varying potentials have been assessed for their ability to repopulate the infarcted myocardium; however, only the adult stem cells, that is, skeletal myoblasts (SkM) and bone marrow-derived stem cells (BMC), have been translated from the laboratory bench to clinical use. Which of these two cell types will provide the best option for clinical application in heart cell therapy remains arguable. With results pouring in from the long-term follow-ups of previously conducted phase I clinical studies, and with the onset of phase II clinical trials involving larger population of patients, transplantation of stem cells as a sole therapy without an adjunct conventional revascularization procedure will provide a deeper insight into the effectiveness of this approach. The present article discusses the pros and cons of using SkM and BMC individually or in combination for cardiac repair, and critically analyzes the progress made with each cell type.

  14. Using Human Induced Pluripotent Stem Cells to Model Skeletal Diseases.

    PubMed

    Barruet, Emilie; Hsiao, Edward C

    2016-01-01

    Musculoskeletal disorders affecting the bones and joints are major health problems among children and adults. Major challenges such as the genetic origins or poor diagnostics of severe skeletal disease hinder our understanding of human skeletal diseases. The recent advent of human induced pluripotent stem cells (human iPS cells) provides an unparalleled opportunity to create human-specific models of human skeletal diseases. iPS cells have the ability to self-renew, allowing us to obtain large amounts of starting material, and have the potential to differentiate into any cell types in the body. In addition, they can carry one or more mutations responsible for the disease of interest or be genetically corrected to create isogenic controls. Our work has focused on modeling rare musculoskeletal disorders including fibrodysplasia ossificans progressive (FOP), a congenital disease of increased heterotopic ossification. In this review, we will discuss our experiences and protocols differentiating human iPS cells toward the osteogenic lineage and their application to model skeletal diseases. A number of critical challenges and exciting new approaches are also discussed, which will allow the skeletal biology field to harness the potential of human iPS cells as a critical model system for understanding diseases of abnormal skeletal formation and bone regeneration.

  15. Human Naive Embryonic Stem Cells: How Full Is the Glass?

    PubMed

    Wang, Yixuan; Gao, Shaorong

    2016-03-01

    Human naive embryonic stem cells in the ground state of pluripotency provide a new opportunity to study human developmental biology and potential clinical applications. Two studies now report related work in human naive stem cell derivation and DNA methylation analysis, with one reporting some differences from oocyte and blastocyst profiles. PMID:26942847

  16. Adult stem cells in the small intestine are intrinsically programmed with their location-specific function.

    PubMed

    Middendorp, Sabine; Schneeberger, Kerstin; Wiegerinck, Caroline L; Mokry, Michal; Akkerman, Ronald D L; van Wijngaarden, Simone; Clevers, Hans; Nieuwenhuis, Edward E S

    2014-05-01

    Differentiation and specialization of epithelial cells in the small intestine are regulated in two ways. First, there is differentiation along the crypt-villus axis of the intestinal stem cells into absorptive enterocytes, Paneth, goblet, tuft, enteroendocrine, or M cells, which is mainly regulated by WNT. Second, there is specialization along the cephalocaudal axis with different absorptive and digestive functions in duodenum, jejunum, and ileum that is controlled by several transcription factors such as GATA4. However, so far it is unknown whether location-specific functional properties are intrinsically programmed within stem cells or if continuous signaling from mesenchymal cells is necessary to maintain the location-specific identity of the small intestine. Using the pure epithelial organoid technique, we show that region-specific gene expression profiles are conserved throughout long-term cultures of both mouse and human intestinal stem cells and correlated with differential Gata4 expression. Furthermore, the human organoid culture system demonstrates that Gata4-regulated gene expression is only allowed in absence of WNT signaling. These data show that location-specific function is intrinsically programmed in the adult stem cells of the small intestine and that their differentiation fate is independent of location-specific extracellular signals. In light of the potential future clinical application of small intestine-derived organoids, our data imply that it is important to generate GATA4-positive and GATA4-negative cultures to regenerate all essential functions of the small intestine.

  17. Human embryonic stem cell research: implications from an ethical and legal standpoint.

    PubMed

    Trepagnier, D M

    2000-12-01

    The purpose of this paper is to discuss the ethical and legal implications of one of the newest and most controversial medical breakthroughs. Stem cell research has been performed on mice for many years, but human embryonic stem cells are believed by scientists to be the basis for possible treatments and/or cures to many diseases affecting millions of people around the world. In order to perform research on human embryonic stem cells, numerous ethical issues must be addressed. Guidelines and protocols can be established in order to allow scientists to pursue new medical advances while maintaining the highest ethical standards in the use of human embryos. An alternative to using embryos is adult stem cells which have recently proven to be more versatile than previously believed. Opposing views will always be encountered when facing new science technologies. Where should the ethical line be drawn?

  18. [Human pluripotent stem cell and neural differentiation].

    PubMed

    Wataya, Takafumi; Muguruma, Keiko; Sasai, Yoshiki

    2008-10-01

    Recovery of lost brain function is an important issue in medical studies because neurons of the central nervous system (CNS) have poor potential for regeneration. Since few CNS diseases can be treated completely by medicines, regenerative therapy by using stem cells should be studied as a new type of therapeutic intervention. The efficacy of cell replacement therapy in Parkinson's disease has been well investigated. Several studies on fetal tissue transplantation have revealed that quantity and purity of transplanted cells are necessary for recovery of symptoms. SFEB (Serum-free floating culture of embryoid body-like aggregates) method is capable of inducing multi-potential CNS progenitors that can be steered to differentiate into region-specific tissues. On the basis of the existing knowledge of embryology, we have succeeded in the generating of various types of neurons such as telencephalic, cerebeller (Purkinje and granule cells), retinal (photoreceptor cells) and hypothalamic neurons. Application of this culture method to human ES (hES) cells is necessary for clinical purpose: however, poor survival of hES cells in SFEB culture might limit the possibility of using these cells for future medical applications. We found that a selective Rho-associated kinase (ROCK) inhibitor, Y-27632, markedly diminished the dissociation-induced apoptosis of hES cells and enabled the cells to form aggregates in SFEB culture. For both mouse and human ES cells, SFEB culture is a favorable method that can generate large amounts of region-specific neurons. However, stem cell-based therapy continues to face several obstacles. It is important that researchers in the basic sciences and clinical medicine should discuss these problems together to overcome both scientific and ethical issues related to stem cells.

  19. Nanotechnology & human stem cells: Applications in cardiogenesis and neurogenesis

    NASA Astrophysics Data System (ADS)

    Tomov, Martin L.

    Human stem cell research holds an unprecedented promise to revolutionize the way we approach medicine and healthcare in general, moving us from a position of mostly addressing the symptoms to a state where treatments can focus on removing the underlying causes of a condition. Stem cell research can shed light into normal developmental pathways, as we are beginning to replicate them in a petri dish and can also be used to model diseases and abnormal conditions. Direct applications can range from finding cures for single or multigene diseases to demonstrating that we can replace these genes with a normal copy. We can even begin to model lifelong conditions such as aging by iPSC technology by relying on fetal, young, adult, and centenarian populations to provide insights into the process. We have also begun to understand the microenvironment in which specific cell populations reside. Being able to replicate the chemical, physical mechanical, and spatial needs of those cells, research groups are successfully generating full organs using cadaver scaffolds of heart and kidney, and there is promising research to reach the same success with other organs, such as the liver, and pancreas. Advances in those areas open an enormous potential to study organs, organoids, organ valves, tubes or other functional elements such as beating cardiomyocytes in vitro. There is also the need to evaluate the whole genome of induced and differentiated cells, with its myriad of interacting pathways. Bioinformatics can help our understanding of embryogenesis, organ differentiation and function. It can also help optimize our stem cell and bio-scaffold tools to advance closer to functional organs and tissues. Such a combination approach will also include pluripotency evaluation and multi-lineage differentiation, as well as platforms that may assist in cell therapies: 3D structures, micro-ribbons, directed patterning to name a few. There is now a clearer path forward with stem cell research than

  20. Future research and therapeutic applications of human stem cells: general, regulatory, and bioethical aspects.

    PubMed

    Liras, Antonio

    2010-01-01

    There is much to be investigated about the specific characteristics of stem cells and about the efficacy and safety of the new drugs based on this type of cells, both embryonic as adult stem cells, for several therapeutic indications (cardiovascular and ischemic diseases, diabetes, hematopoietic diseases, liver diseases). Along with recent progress in transference of nuclei from human somatic cells, as well as iPSC technology, has allowed availability of lineages of all three germ layers genetically identical to those of the donor patient, which permits safe transplantation of organ-tissue-specific adult stem cells with no immune rejection. The main objective is the need for expansion of stem cell characteristics to maximize stem cell efficacy (i.e. the proper selection of a stem cell) and the efficacy (maximum effect) and safety of stem cell derived drugs. Other considerations to take into account in cell therapy will be the suitability of infrastructure and technical staff, biomaterials, production costs, biobanks, biosecurity, and the biotechnological industry. The general objectives in the area of stem cell research in the next few years, are related to identification of therapeutic targets and potential therapeutic tests, studies of cell differentiation and physiological mechanisms, culture conditions of pluripotent stem cells and efficacy and safety tests for stem cell-based drugs or procedures to be performed in both animal and human models in the corresponding clinical trials. A regulatory framework will be required to ensure patient accessibility to products and governmental assistance for their regulation and control. Bioethical aspects will be required related to the scientific and therapeutic relevance and cost of cryopreservation over time, but specially with respect to embryos which may ultimately be used for scientific uses of research as source of embryonic stem cells, in which case the bioethical conflict may be further aggravated. PMID:21143967

  1. Future research and therapeutic applications of human stem cells: general, regulatory, and bioethical aspects

    PubMed Central

    2010-01-01

    There is much to be investigated about the specific characteristics of stem cells and about the efficacy and safety of the new drugs based on this type of cells, both embryonic as adult stem cells, for several therapeutic indications (cardiovascular and ischemic diseases, diabetes, hematopoietic diseases, liver diseases). Along with recent progress in transference of nuclei from human somatic cells, as well as iPSC technology, has allowed availability of lineages of all three germ layers genetically identical to those of the donor patient, which permits safe transplantation of organ-tissue-specific adult stem cells with no immune rejection. The main objective is the need for expansion of stem cell characteristics to maximize stem cell efficacy (i.e. the proper selection of a stem cell) and the efficacy (maximum effect) and safety of stem cell derived drugs. Other considerations to take into account in cell therapy will be the suitability of infrastructure and technical staff, biomaterials, production costs, biobanks, biosecurity, and the biotechnological industry. The general objectives in the area of stem cell research in the next few years, are related to identification of therapeutic targets and potential therapeutic tests, studies of cell differentiation and physiological mechanisms, culture conditions of pluripotent stem cells and efficacy and safety tests for stem cell-based drugs or procedures to be performed in both animal and human models in the corresponding clinical trials. A regulatory framework will be required to ensure patient accessibility to products and governmental assistance for their regulation and control. Bioethical aspects will be required related to the scientific and therapeutic relevance and cost of cryopreservation over time, but specially with respect to embryos which may ultimately be used for scientific uses of research as source of embryonic stem cells, in which case the bioethical conflict may be further aggravated. PMID:21143967

  2. Future research and therapeutic applications of human stem cells: general, regulatory, and bioethical aspects.

    PubMed

    Liras, Antonio

    2010-12-10

    There is much to be investigated about the specific characteristics of stem cells and about the efficacy and safety of the new drugs based on this type of cells, both embryonic as adult stem cells, for several therapeutic indications (cardiovascular and ischemic diseases, diabetes, hematopoietic diseases, liver diseases). Along with recent progress in transference of nuclei from human somatic cells, as well as iPSC technology, has allowed availability of lineages of all three germ layers genetically identical to those of the donor patient, which permits safe transplantation of organ-tissue-specific adult stem cells with no immune rejection. The main objective is the need for expansion of stem cell characteristics to maximize stem cell efficacy (i.e. the proper selection of a stem cell) and the efficacy (maximum effect) and safety of stem cell derived drugs. Other considerations to take into account in cell therapy will be the suitability of infrastructure and technical staff, biomaterials, production costs, biobanks, biosecurity, and the biotechnological industry. The general objectives in the area of stem cell research in the next few years, are related to identification of therapeutic targets and potential therapeutic tests, studies of cell differentiation and physiological mechanisms, culture conditions of pluripotent stem cells and efficacy and safety tests for stem cell-based drugs or procedures to be performed in both animal and human models in the corresponding clinical trials. A regulatory framework will be required to ensure patient accessibility to products and governmental assistance for their regulation and control. Bioethical aspects will be required related to the scientific and therapeutic relevance and cost of cryopreservation over time, but specially with respect to embryos which may ultimately be used for scientific uses of research as source of embryonic stem cells, in which case the bioethical conflict may be further aggravated.

  3. Genetic Manipulation of Human Embryonic Stem Cells.

    PubMed

    Eiges, Rachel

    2016-01-01

    One of the great advantages of embryonic stem (ES) cells over other cell types is their accessibility to genetic manipulation. They can easily undergo genetic modifications while remaining pluripotent, and can be selectively propagated, allowing the clonal expansion of genetically altered cells in culture. Since the first isolation of ES cells in mice, many effective techniques have been developed for gene delivery and manipulation of ES cells. These include transfection, electroporation, and infection protocols, as well as different approaches for inserting, deleting, or changing the expression of genes. These methods proved to be extremely useful in mouse ES cells, for monitoring and directing differentiation, discovering unknown genes, and studying their function, and are now being extensively implemented in human ES cells (HESCs). This chapter describes the different approaches and methodologies that have been applied for the genetic manipulation of HESCs and their applications. Detailed protocols for generating clones of genetically modified HESCs by transfection, electroporation, and infection will be described, with special emphasis on the important technical details that are required for this purpose. All protocols are equally effective in human-induced pluripotent stem (iPS) cells.

  4. Mesenchymal stem cells instruct oligodendrogenic fate decision on adult neural stem cells.

    PubMed

    Rivera, Francisco J; Couillard-Despres, Sebastien; Pedre, Xiomara; Ploetz, Sonja; Caioni, Massimiliano; Lois, Carlos; Bogdahn, Ulrich; Aigner, Ludwig

    2006-10-01

    Adult stem cells reside in different tissues and organs of the adult organism. Among these cells are MSCs that are located in the adult bone marrow and NSCs that exist in the adult central nervous system (CNS). In transplantation experiments, MSCs demonstrated neuroprotective and neuroregenerative effects that were associated with functional improvements. The underlying mechanisms are largely unidentified. Here, we reveal that the interactions between adult MSCs and NSCs, mediated by soluble factors, induce oligodendrogenic fate decision in NSCs at the expense of astrogenesis. This was demonstrated (a) by an increase in the percentage of cells expressing the oligodendrocyte markers GalC and myelin basic protein, (b) by a reduction in the percentage of glial fibrillary acidic protein (GFAP)-expressing cells, and (c) by the expression pattern of cell fate determinants specific for oligodendrogenic differentiation. Thus, it involved enhanced expression of the oligodendrogenic transcription factors Olig1, Olig2, and Nkx2.2 and diminished expression of Id2, an inhibitor of oligodendrogenic differentiation. Results of (a) 5-bromo-2'-deoxyuridine pulse-labeling of cells, (b) cell fate analysis, and (c) cell death/survival analysis suggested an inductive mechanism and excluded a selection process. A candidate factor screen excluded a number of growth factors, cytokines, and neurotrophins that have previously been shown to influence neurogenesis and neural differentiation from the oligodendrogenic activity derived from the MSCs. This work might have major implications for the development of future transplantation strategies for the treatment of degenerative diseases in the CNS. PMID:16763198

  5. Hedgehog signaling activation induces stem cell proliferation and hormone release in the adult pituitary gland

    PubMed Central

    Pyczek, Joanna; Buslei, Rolf; Schult, David; Hölsken, Annett; Buchfelder, Michael; Heß, Ina; Hahn, Heidi; Uhmann, Anja

    2016-01-01

    Hedgehog (HH) signaling is known to be essential during the embryonal development of the pituitary gland but the knowledge about its role in the adult pituitary and in associated tumors is sparse. In this report we investigated the effect of excess Hh signaling activation in murine pituitary explants and analyzed the HH signaling status of human adenopituitary lobes and a large cohort of pituitary adenomas. Our data show that excess Hh signaling led to increased proliferation of Sox2+ and Sox9+ adult pituitary stem cells and to elevated expression levels of adrenocorticotropic hormone (Acth), growth hormone (Gh) and prolactin (Prl) in the adult gland. Inhibition of the pathway by cyclopamine reversed these effects indicating that active Hh signaling positively regulates proliferative processes of adult pituitary stem cells and hormone production in the anterior pituitary. Since hormone producing cells of the adenohypophysis as well as ACTH-, GH- and PRL-immunopositive adenomas express SHH and its target GLI1, we furthermore propose that excess HH signaling is involved in the development/maintenance of hormone-producing pituitary adenomas. These findings advance the understanding of physiological hormone regulation and may open new treatment options for pituitary tumors. PMID:27109116

  6. Human Wharton's jelly stem cells have unique transcriptome profiles compared to human embryonic stem cells and other mesenchymal stem cells.

    PubMed

    Fong, Chui-Yee; Chak, Li-Ling; Biswas, Arijit; Tan, Jee-Hian; Gauthaman, Kalamegam; Chan, Woon-Khiong; Bongso, Ariff

    2011-03-01

    The human umbilical cord that originates from the embryo is an extra-embryonic membrane and the Wharton's jelly within it is a rich source of stem cells (hWJSCs). It is not definitely known whether these cells behave as human embryonic stem cells (hESCs), human mesenchymal stem cells (hMSC) or both. They have the unique properties of high proliferation rates, wide multipotency, hypoimmunogenicity, do not induce teratomas and have anticancer properties. These advantages are important considerations for their use in cell based therapies and treatment of cancers. In a search for properties that confer these advantages we compared a detailed transcriptome profiling of hWJSCs using DNA microarrays with that of a panel of known hESCs, hMSCs and stromal cells. hWJSCs expressed low levels of the pluripotent embryonic stem cell markers including POUF1, NANOG, SOX2 and LIN28, thus explaining why they do not produce teratomas. Several cytokines were significantly upregulated in hWJSCs including IL12A which is associated with the induction of apoptosis, thus explaining their anticancer properties. When GO Biological Process analysis was compared between the various stem cell types, hWJSCs showed an increased expression of genes associated with the immune system, chemotaxis and cell death. The ability to modulate immune responses makes hWJSCs an important compatible stem cell source for transplantation therapy in allogeneic settings without immunorejection. The data in the present study which is the first detailed report on hWJSC transcriptomes provide a foundation for future functional studies where the exact mechanisms of these unique properties of hWJSCs can be confirmed.

  7. Stirred bioreactors for the expansion of adult pancreatic stem cells.

    PubMed

    Serra, Margarida; Brito, Catarina; Leite, Sofia B; Gorjup, Erwin; von Briesen, Hagen; Carrondo, Manuel J T; Alves, Paula M

    2009-01-01

    Adult pluripotent stem cells are a cellular resource representing unprecedented potential for cell therapy and tissue engineering. Complementary to this promise, there is a need for efficient bioprocesses for their large scale expansion and/or differentiation. With this goal in mind, our work focused on the development of three-dimensional (3-D) culture systems for controlled expansion of adult pancreatic stem cells (PSCs). For this purpose, two different culturing strategies were evaluated, using spinner vessels: cell aggregated cultures versus microcarrier technology. The use of microcarrier supports (Cytodex 1 and Cytodex 3) rendered expanded cell populations which retained their self-renewal ability, cell marker, and the potential to differentiate into adipocytes. This strategy surmounted the drawbacks of aggregates in culture which were demonstrably unfeasible as cells clumped together did not proliferate and lost PSC marker expression. Furthermore, the results obtained showed that although both microcarriers tested here were suitable for sustaining cell expansion, Cytodex 3 provided a better substrate for the promotion of cell adherence and growth. For the latter approach, the potential of bioreactor technology was combined with the efficient Cytodex 3 strategy under controlled environmental conditions (pH-7.2, pO2-30% and temperature-37 degrees C); cell growth was more efficient, as shown by faster doubling time, higher growth rate and higher fold increase in cell concentration, when compared to spinner cultures. This study describes a robust bioprocess for the controlled expansion of adult PSC, representing an efficient starting point for the development of novel technologies for cell therapy.

  8. Primary Bioassay of Human Myeloma Stem Cells

    PubMed Central

    Hamburger, Anne; Salmon, Sydney E.

    1977-01-01

    The ability to clone primary tumors in soft agar has proven useful in the study of the kinetics and biological properties of tumor stem cells. We report the development of an in vitro assay which permits formation of colonies of human monoclonal plasma cells in soft agar. Colony growth has been observed from bone marrow aspirates from 75% of the 70 patients with multiple myeloma or related monoclonal disorders studied. Growth was induced with either 0.02 ml of human type O erythrocytes or 0.25 ml of medium conditioned by the adherent spleen cells of mineral oil-primed BALB/c mice. 5-500 colonies appeared after 2-3 wk in culture yielding a plating efficiency of 0.001-0.1%. The number of myeloma colonies was proportional to the number of cells plated between concentrations of 105-106 and back-extrapolated through zero, suggesting that colonies were clones derived from single myeloma stem cells. Morphological, histochemical, and functional criteria showed the colonies to consist of immature plasmablasts and mature plasma cells. 60-80% of cells picked from colonies contained intracytoplasmic monoclonal immunoglobulin. Colony growth was most easily achieved from the bone marrow cells of untreated patients or those in relapse. Only 50% of bone marrow samples from patients in remission were successfully cultured. Tritiated thymidine suicide studies provided evidence that for most myeloma patients, a very high proportion of myeloma colony-forming cells was actively in transit through the cell cycle. Velocity sedimentation at 1 g showed myeloma stem cells sedimented in a broad band with a peak at 13 mm/h. Antibody to granulocyte colony-stimulating factor did not reduce the number or size of the colonies. Increased numbers of myeloma colonies were seen when the marrow was depleted of colony-stimulating factor elaborating adherent cells before plating. This bioassay should prove useful in studying the in vitro biological behavior of certain bone marrow-derived (B

  9. Axonal control of the adult neural stem cell niche.

    PubMed

    Tong, Cheuk Ka; Chen, Jiadong; Cebrián-Silla, Arantxa; Mirzadeh, Zaman; Obernier, Kirsten; Guinto, Cristina D; Tecott, Laurence H; García-Verdugo, Jose Manuel; Kriegstein, Arnold; Alvarez-Buylla, Arturo

    2014-04-01

    The ventricular-subventricular zone (V-SVZ) is an extensive germinal niche containing neural stem cells (NSCs) in the walls of the lateral ventricles of the adult brain. How the adult brain's neural activity influences the behavior of adult NSCs remains largely unknown. We show that serotonergic (5HT) axons originating from a small group of neurons in the raphe form an extensive plexus on most of the ventricular walls. Electron microscopy revealed intimate contacts between 5HT axons and NSCs (B1) or ependymal cells (E1) and these cells were labeled by a transsynaptic viral tracer injected into the raphe. B1 cells express the 5HT receptors 2C and 5A. Electrophysiology showed that activation of these receptors in B1 cells induced small inward currents. Intraventricular infusion of 5HT2C agonist or antagonist increased or decreased V-SVZ proliferation, respectively. These results indicate that supraependymal 5HT axons directly interact with NSCs to regulate neurogenesis via 5HT2C. PMID:24561083

  10. Axonal Control of the Adult Neural Stem Cell Niche

    PubMed Central

    Tong, Cheuk Ka; Chen, Jiadong; Cebrián-Silla, Arantxa; Mirzadeh, Zaman; Obernier, Kirsten; Guinto, Cristina D.; Tecott, Laurence H.; García-Verdugo, Jose Manuel; Kriegstein, Arnold; Alvarez-Buylla, Arturo

    2014-01-01

    SUMMARY The ventricular-subventricular zone (V-SVZ) is an extensive germinal niche containing neural stem cells (NSC) in the walls of the lateral ventricles of the adult brain. How the adult brain’s neural activity influences the behavior of adult NSCs remains largely unknown. We show that serotonergic (5HT) axons originating from a small group of neurons in the raphe form an extensive plexus on most of the ventricular walls. Electron microscopy revealed intimate contacts between 5HT axons and NSCs (B1) or ependymal cells (E1) and these cells were labeled by a transsynaptic viral tracer injected into the raphe. B1 cells express the 5HT receptors 2C and 5A. Electrophysiology showed that activation of these receptors in B1 cells induced small inward currents. Intraventricular infusion of 5HT2C agonist or antagonist increased or decreased V-SVZ proliferation, respectively. These results indicate that supraependymal 5HT axons directly interact with NSCs to regulate neurogenesis via 5HT2C. PMID:24561083

  11. Live Imaging of Adult Neural Stem Cells in Rodents

    PubMed Central

    Ortega, Felipe; Costa, Marcos R.

    2016-01-01

    The generation of cells of the neural lineage within the brain is not restricted to early development. New neurons, oligodendrocytes, and astrocytes are produced in the adult brain throughout the entire murine life. However, despite the extensive research performed in the field of adult neurogenesis during the past years, fundamental questions regarding the cell biology of adult neural stem cells (aNSCs) remain to be uncovered. For instance, it is crucial to elucidate whether a single aNSC is capable of differentiating into all three different macroglial cell types in vivo or these distinct progenies constitute entirely separate lineages. Similarly, the cell cycle length, the time and mode of division (symmetric vs. asymmetric) that these cells undergo within their lineage progression are interesting questions under current investigation. In this sense, live imaging constitutes a valuable ally in the search of reliable answers to the previous questions. In spite of the current limitations of technology new approaches are being developed and outstanding amount of knowledge is being piled up providing interesting insights in the behavior of aNSCs. Here, we will review the state of the art of live imaging as well as the alternative models that currently offer new answers to critical questions. PMID:27013941

  12. LPS-Stimulated Human Skin-Derived Stem Cells Enhance Neo-Vascularization during Dermal Regeneration.

    PubMed

    Kisch, Tobias; Weber, Caroline; Rapoport, Daniel H; Kruse, Charli; Schumann, Sandra; Stang, Felix H; Siemers, Frank; Matthießen, Anna E

    2015-01-01

    High numbers of adult stem cells are still required to improve the formation of new vessels in scaffolds to accelerate dermal regeneration. Recent data indicate a benefit for vascularization capacity by stimulating stem cells with lipopolysaccharide (LPS). In this study, stem cells derived from human skin (SDSC) were activated with LPS and seeded in a commercially available dermal substitute to examine vascularization in vivo. Besides, in vitro assays were performed to evaluate angiogenic factor release and tube formation ability. Results showed that LPS-activated SDSC significantly enhanced vascularization of the scaffolds, compared to unstimulated stem cells in vivo. Further, in vitro assays confirmed higher secretion rates of proangiogenic as well as proinflammatoric factors in the presence of LPS-activated SDSC. Our results suggest that combining activated stem cells and a dermal substitute is a promising option to enhance vascularization in scaffold-mediated dermal regeneration.

  13. Regulation of neural stem cell in the human SVZ by trophic and morphogenic factors

    PubMed Central

    Alvarez-Palazuelos, Lucia E.; Robles-Cervantes, Martha S.; Castillo-Velazquez, Gabriel; Rivas-Souza, Mario; Guzman-Muniz, Jorge; Moy-Lopez, Norma; Gonzalez-Castaneda, Rocio E.; Luquin, Sonia; Gonzalez-Perez, Oscar

    2011-01-01

    The subventricular zone (SVZ), lining the lateral ventricular system, is the largest germinal region in mammals. In there, neural stem cells express markers related to astoglial lineage that give rise to new neurons and oligodendrocytes in vivo. In the adult human brain, in vitro evidence has also shown that astrocytic cells isolated from the SVZ can generate new neurons and oligodendrocytes. These proliferative cells are strongly controlled by a number of signals and molecules that modulate, activate or repress the cell division, renewal, proliferation and fate of neural stem cells. In this review, we summarize the cellular composition of the adult human SVZ (hSVZ) and discuss the increasing evidence showing that some trophic modulators strongly control the function of neural stem cells in the SVZ. PMID:22053150

  14. Biological impact of human embryonic stem cells.

    PubMed

    Martín, Miguel; Menéndez, Pablo

    2012-01-01

    Research on human embryonic stem cells (hESCs) and induced pluripotent (iPS) stem cells is currently a field of great potential in biomedicine. These cells represent a highly valuable tool for developmental biology studies, disease models, and drug screening and toxicity. The ultimate goal of hESCs and iPS cell research is the treatment of diseases or disorders for which there is currently no treatment or existing therapies are only partially effective. Despite the disproportionate short-term hopes generated, which are putting too much pressure on scientists, the international scientific community is making rapid progress in understanding hESCs and iPS cells. Nonetheless, great efforts have to be made to provide an answer to still quite basic questions concerning their biology. Moreover, translation to clinical applications in cell replacement therapy requires prior solution to ethical barriers. The recent development of iPS cells has provided a strong alternative to overcome ethical issues concerning hESCs. However, an in-depth characterization of their genetic and epigenetic features, as well as their differentiation potential still remains to be undertaken. This chapter will describe, precisely, what the critical issues are, where scientific and ethical barriers stand, and how we are to overcome them. Only then, we shall finally discover whether hESCs and iPS cells will allow building reproducible disease models, and whether they really are a safe tool, with great potential for regenerative medicine.

  15. In search of human hematopoietic stem cell identity.

    PubMed

    Ivanovs, Andrejs; Medvinsky, Alexander

    2015-01-01

    Better insight into hematopoietic stem cell (HSC) development in the human embryo and fetus is crucial for translational research. In this issue of Cell Stem Cell, Prashad et al. (2014) describe a novel surface marker for human fetal liver HSCs, glycosylphosphatidylinositol-anchored protein GPI-80, that is functionally required for their self-renewal.

  16. Isolation and Culture of Neural Crest Stem Cells from Human Hair Follicles.

    PubMed

    Yang, Ruifeng; Xu, Xiaowei

    2016-01-01

    The hair follicle undergoes lifelong cycling and growth. Previous studies have been focused on epithelial stem cells in the hair follicles. Neural crest stem cells (NCSCs) are pluripotent cells that can persist in adult tissues. We have previously demonstrated that human NCSCs can be isolated from hair follicles. Here, we present a protocol to isolate NCSCs from human hair follicles based on their specific surface-marker expression of CD271/HNK1 or CD271/CD49D (alpha4 integrin). NCSCs can be expanded in the culture as neural spheres or attached cells.

  17. Human Salivary Gland Stem Cells Functionally Restore Radiation Damaged Salivary Glands.

    PubMed

    Pringle, Sarah; Maimets, Martti; van der Zwaag, Marianne; Stokman, Monique A; van Gosliga, Djoke; Zwart, Erik; Witjes, Max J H; de Haan, Gerald; van Os, Ronald; Coppes, Rob P

    2016-03-01

    Adult stem cells are often touted as therapeutic agents in the regenerative medicine field, however data detailing both the engraftment and functional capabilities of solid tissue derived human adult epithelial stem cells is scarce. Here we show the isolation of adult human salivary gland (SG) stem/progenitor cells and demonstrate at the single cell level in vitro self-renewal and differentiation into multilineage organoids. We also show in vivo functionality, long-term engraftment, and functional restoration in a xenotransplantation model. Indeed, transplanted human salisphere-derived cells restored saliva production and greatly improved the regenerative potential of irradiated SGs. Further selection for c-Kit expression enriched for cells with enhanced regenerative potencies. Interestingly, interaction of transplanted cells with the recipient SG may also be involved in functional recovery. Thus, we show for the first time that salispheres cultured from human SGs contain stem/progenitor cells capable of self-renewal and differentiation and rescue of saliva production. Our study underpins the therapeutic promise of salisphere cell therapy for the treatment of xerostomia. PMID:26887347

  18. [Generation of new nerve cells in the adult human brain].

    PubMed

    Poulsen, Frantz Rom; Meyer, Morten; Rasmussen, Jens Zimmer

    2003-03-31

    Generation of new nerve cells (neurogenesis) is normally considered to be limited to the fetal and early postnatal period. Thus, damaged nerve cells are not expected to be replaced by generation of new cells. The brain is, however, more plastic than previously assumed. This also includes neurogenesis in the adult human brain. In particular two brain regions show continuous division of neural stem and progenitor cells generating neurons and glial cells, namely the subgranular zone of the dentate gyrus and the subventricular zones of the lateral ventricles. From the latter region newly generated neuroblasts (immature nerve cells) migrate toward the olfactory bulb where they differentiate into neurons. In the dentate gyrus the newly generated neurons become functionally integrated in the granule cell layer, where they are believed to be of importance to learning and memory. It is at present not known whether neurogenesis in the adult human brain can be manipulated for specific repair after brain damage.

  19. Regulation of seminiferous tubule-associated stem Leydig cells in adult rat testes.

    PubMed

    Li, Xiaoheng; Wang, Zhao; Jiang, Zhenming; Guo, Jingjing; Zhang, Yuxi; Li, Chenhao; Chung, Jinyong; Folmer, Janet; Liu, June; Lian, Qingquan; Ge, Renshan; Zirkin, Barry R; Chen, Haolin

    2016-03-01

    Testicular Leydig cells are the primary source of testosterone in males. Adult Leydig cells have been shown to arise from stem cells present in the neonatal testis. Once established, adult Leydig cells turn over only slowly during adult life, but when these cells are eliminated experimentally from the adult testis, new Leydig cells rapidly reappear. As in the neonatal testis, stem cells in the adult testis are presumed to be the source of the new Leydig cells. As yet, the mechanisms involved in regulating the proliferation and differentiation of these stem cells remain unknown. We developed a unique in vitro system of cultured seminiferous tubules to assess the ability of factors from the seminiferous tubules to regulate the proliferation of the tubule-associated stem cells, and their subsequent entry into the Leydig cell lineage. The proliferation of the stem Leydig cells was stimulated by paracrine factors including Desert hedgehog (DHH), basic fibroblast growth factor (FGF2), platelet-derived growth factor (PDGF), and activin. Suppression of proliferation occurred with transforming growth factor β (TGF-β). The differentiation of the stem cells was regulated positively by DHH, lithium- induced signaling, and activin, and negatively by TGF-β, PDGFBB, and FGF2. DHH functioned as a commitment factor, inducing the transition of stem cells to the progenitor stage and thus into the Leydig cell lineage. Additionally, CD90 (Thy1) was found to be a unique stem cell surface marker that was used to obtain purified stem cells by flow cytometry.

  20. Regulation of seminiferous tubule-associated stem Leydig cells in adult rat testes

    PubMed Central

    Li, Xiaoheng; Wang, Zhao; Jiang, Zhenming; Guo, Jingjing; Zhang, Yuxi; Li, Chenhao; Chung, Jinyong; Folmer, Janet; Liu, June; Lian, Qingquan; Ge, Renshan; Zirkin, Barry R.; Chen, Haolin

    2016-01-01

    Testicular Leydig cells are the primary source of testosterone in males. Adult Leydig cells have been shown to arise from stem cells present in the neonatal testis. Once established, adult Leydig cells turn over only slowly during adult life, but when these cells are eliminated experimentally from the adult testis, new Leydig cells rapidly reappear. As in the neonatal testis, stem cells in the adult testis are presumed to be the source of the new Leydig cells. As yet, the mechanisms involved in regulating the proliferation and differentiation of these stem cells remain unknown. We developed a unique in vitro system of cultured seminiferous tubules to assess the ability of factors from the seminiferous tubules to regulate the proliferation of the tubule-associated stem cells, and their subsequent entry into the Leydig cell lineage. The proliferation of the stem Leydig cells was stimulated by paracrine factors including Desert hedgehog (DHH), basic fibroblast growth factor (FGF2), platelet-derived growth factor (PDGF), and activin. Suppression of proliferation occurred with transforming growth factor β (TGF-β). The differentiation of the stem cells was regulated positively by DHH, lithium- induced signaling, and activin, and negatively by TGF-β, PDGFBB, and FGF2. DHH functioned as a commitment factor, inducing the transition of stem cells to the progenitor stage and thus into the Leydig cell lineage. Additionally, CD90 (Thy1) was found to be a unique stem cell surface marker that was used to obtain purified stem cells by flow cytometry. PMID:26929346

  1. Lifespan Extension and Sustained Expression of Stem Cell Phenotype of Human Breast Epithelial Stem Cells in a Medium with Antioxidants

    PubMed Central

    Wang, Kai-Hung; Kao, An-Pei; Chang, Chia-Cheng; Lin, Ta-Chin

    2016-01-01

    We have previously reported the isolation and culture of a human breast epithelial cell type with stem cell characteristics (Type I HBEC) from reduction mammoplasty using the MSU-1 medium. Subsequently, we have developed several different normal human adult stem cell types from different tissues using the K-NAC medium. In this study, we determined whether this low calcium K-NAC medium with antioxidants (N-acetyl-L-cysteine and L-ascorbic acid-2-phosphate) is a better medium to grow human breast epithelial cells. The results clearly show that the K-NAC medium is a superior medium for prolonged growth (cumulative population doubling levels ranged from 30 to 40) of normal breast epithelial cells that expressed stem cell phenotypes. The characteristics of these mammary stem cells include deficiency in gap junctional intercellular communication, expression of Oct-4, and the ability to differentiate into basal epithelial cells and to form organoid showing mammary ductal and terminal end bud-like structures. Thus, this new method of growing Type I HBECs will be very useful in future studies of mammary development, breast carcinogenesis, chemoprevention, and cancer therapy. PMID:27807451

  2. Multipotent (adult) and pluripotent stem cells for heart regeneration: what are the pros and cons?

    PubMed

    Liao, Song-Yan; Tse, Hung-Fat

    2013-12-24

    Heart failure after myocardial infarction is the leading cause of mortality and morbidity worldwide. Existing medical and interventional therapies can only reduce the loss of cardiomyocytes during myocardial infarction but are unable to replenish the permanent loss of cardiomyocytes after the insult, which contributes to progressive pathological left ventricular remodeling and progressive heart failure. As a result, cell-based therapies using multipotent (adult) stem cells and pluripotent stem cells (embryonic stem cells or induced pluripotent stem cells) have been explored as potential therapeutic approaches to restore cardiac function in heart failure. Nevertheless, the optimal cell type with the best therapeutic efficacy and safety for heart regeneration is still unknown. In this review, the potential pros and cons of different types of multipotent (adult) stem cells and pluripotent stem cells that have been investigated in preclinical and clinical studies are reviewed, and the future perspective of stem cell-based therapy for heart regeneration is discussed.

  3. Nucleosome Organization in Human Embryonic Stem Cells.

    PubMed

    Yazdi, Puya G; Pedersen, Brian A; Taylor, Jared F; Khattab, Omar S; Chen, Yu-Han; Chen, Yumay; Jacobsen, Steven E; Wang, Ping H

    2015-01-01

    The fundamental repeating unit of eukaryotic chromatin is the nucleosome. Besides being involved in packaging DNA, nucleosome organization plays an important role in transcriptional regulation and cellular identity. Currently, there is much debate about the major determinants of the nucleosome architecture of a genome and its significance with little being known about its role in stem cells. To address these questions, we performed ultra-deep sequencing of nucleosomal DNA in two human embryonic stem cell lines and integrated our data with numerous epigenomic maps. Our analyses have revealed that the genome is a determinant of nucleosome organization with transcriptionally inactive regions characterized by a "ground state" of nucleosome profiles driven by underlying DNA sequences. DNA sequence preferences are associated with heterogeneous chromatin organization around transcription start sites. Transcription, histone modifications, and DNA methylation alter this "ground state" by having distinct effects on both nucleosome positioning and occupancy. As the transcriptional rate increases, nucleosomes become better positioned. Exons transcribed and included in the final spliced mRNA have distinct nucleosome profiles in comparison to exons not included at exon-exon junctions. Genes marked by the active modification H3K4m3 are characterized by lower nucleosome occupancy before the transcription start site compared to genes marked by the inactive modification H3K27m3, while bivalent domains, genes associated with both marks, lie exactly in the middle. Combinatorial patterns of epigenetic marks (chromatin states) are associated with unique nucleosome profiles. Nucleosome organization varies around transcription factor binding in enhancers versus promoters. DNA methylation is associated with increasing nucleosome occupancy and different types of methylations have distinct location preferences within the nucleosome core particle. Finally, computational analysis of nucleosome

  4. Mitochondria in human pluripotent stem cell apoptosis.

    PubMed

    TeSlaa, Tara; Setoguchi, Kiyoko; Teitell, Michael A

    2016-04-01

    Human pluripotent stem cells (hPSCs) have great potential in regenerative medicine because they can differentiate into any cell type in the body. Genome integrity is vital for human development and for high fidelity passage of genetic information across generations through the germ line. To ensure genome stability, hPSCs maintain a lower rate of mutation than somatic cells and undergo rapid apoptosis in response to DNA damage and additional cell stresses. Furthermore, cellular metabolism and the cell cycle are also differentially regulated between cells in pluripotent and differentiated states and can aid in protecting hPSCs against DNA damage and damaged cell propagation. Despite these safeguards, clinical use of hPSC derivatives could be compromised by tumorigenic potential and possible malignant transformation from failed to differentiate cells. Since hPSCs and mature cells differentially respond to cell stress, it may be possible to specifically target undifferentiated cells for rapid apoptosis in mixed cell populations to enable safer use of hPSC-differentiated cells in patients.

  5. Mitochondria in human pluripotent stem cell apoptosis.

    PubMed

    TeSlaa, Tara; Setoguchi, Kiyoko; Teitell, Michael A

    2016-04-01

    Human pluripotent stem cells (hPSCs) have great potential in regenerative medicine because they can differentiate into any cell type in the body. Genome integrity is vital for human development and for high fidelity passage of genetic information across generations through the germ line. To ensure genome stability, hPSCs maintain a lower rate of mutation than somatic cells and undergo rapid apoptosis in response to DNA damage and additional cell stresses. Furthermore, cellular metabolism and the cell cycle are also differentially regulated between cells in pluripotent and differentiated states and can aid in protecting hPSCs against DNA damage and damaged cell propagation. Despite these safeguards, clinical use of hPSC derivatives could be compromised by tumorigenic potential and possible malignant transformation from failed to differentiate cells. Since hPSCs and mature cells differentially respond to cell stress, it may be possible to specifically target undifferentiated cells for rapid apoptosis in mixed cell populations to enable safer use of hPSC-differentiated cells in patients. PMID:26828436

  6. [Research with human embryo stem cells. Foundations and judicial limits].

    PubMed

    Eser, Albin; Koch, Hans-Georg

    2004-01-01

    Research with human embryos, and particularly, the use for scientific purposes of human embryonic stem cells has given raise to different sort of problems at the international level. One of the most strict regulation in this field, is this lecture Professors Albin Eser and Hans-Georg Koch analyse the german legal framework in relation with the use of embryos and human embryonic stem cells for scientific purposes.

  7. Neoplastic human embryonic stem cells as a model of radiation resistance of human cancer stem cells

    PubMed Central

    Dingwall, Steve; Lee, Jung Bok; Guezguez, Borhane; Fiebig, Aline; McNicol, Jamie; Boreham, Douglas; Collins, Tony J.; Bhatia, Mick

    2015-01-01

    Studies have implicated that a small sub-population of cells within a tumour, termed cancer stem cells (CSCs), have an enhanced capacity for tumour formation in multiple cancers and may be responsible for recurrence of the disease after treatment, including radiation. Although comparisons have been made between CSCs and bulk-tumour, the more important comparison with respect to therapy is between tumour-sustaining CSC versus normal stem cells that maintain the healthy tissue. However, the absence of normal known counterparts for many CSCs has made it difficult to compare the radiation responses of CSCs with the normal stem cells required for post-radiotherapy tissue regeneration and the maintenance of tissue homeostasis. Here we demonstrate that transformed human embryonic stem cells (t-hESCs), showing features of neoplastic progression produce tumours resistant to radiation relative to their normal counterpart upon injection into immune compromised mice. We reveal that t-hESCs have a reduced capacity for radiation induced cell death via apoptosis and exhibit altered cell cycle arrest relative to hESCs in vitro. t-hESCs have an increased expression of BclXL in comparison to their normal counterparts and re-sensitization of t-hESCs to radiation upon addition of BH3-only mimetic ABT737, suggesting that overexpression of BclXL underpins t-hESC radiation insensitivity. Using this novel discovery platform to investigate radiation resistance in human CSCs, our study indicates that chemotherapy targeting Bcl2-family members may prove to be an adjuvant to radiotherapy capable of targeting CSCs. PMID:26082437

  8. Neoplastic human embryonic stem cells as a model of radiation resistance of human cancer stem cells.

    PubMed

    Dingwall, Steve; Lee, Jung Bok; Guezguez, Borhane; Fiebig, Aline; McNicol, Jamie; Boreham, Douglas; Collins, Tony J; Bhatia, Mick

    2015-09-01

    Studies have implicated that a small sub-population of cells within a tumour, termed cancer stem cells (CSCs), have an enhanced capacity for tumour formation in multiple cancers and may be responsible for recurrence of the disease after treatment, including radiation. Although comparisons have been made between CSCs and bulk-tumour, the more important comparison with respect to therapy is between tumour-sustaining CSC versus normal stem cells that maintain the healthy tissue. However, the absence of normal known counterparts for many CSCs has made it difficult to compare the radiation responses of CSCs with the normal stem cells required for post-radiotherapy tissue regeneration and the maintenance of tissue homeostasis. Here we demonstrate that transformed human embryonic stem cells (t-hESCs), showing features of neoplastic progression produce tumours resistant to radiation relative to their normal counterpart upon injection into immune compromised mice. We reveal that t-hESCs have a reduced capacity for radiation induced cell death via apoptosis and exhibit altered cell cycle arrest relative to hESCs in vitro. t-hESCs have an increased expression of BclXL in comparison to their normal counterparts and re-sensitization of t-hESCs to radiation upon addition of BH3-only mimetic ABT737, suggesting that overexpression of BclXL underpins t-hESC radiation insensitivity. Using this novel discovery platform to investigate radiation resistance in human CSCs, our study indicates that chemotherapy targeting Bcl2-family members may prove to be an adjuvant to radiotherapy capable of targeting CSCs.

  9. Cerebellar stem cells do not produce neurons and astrocytes in adult mouse

    SciTech Connect

    Su, Xin; Guan, Wuqiang; Yu, Yong-Chun; Fu, Yinghui

    2014-07-18

    Highlights: • No new neurons and astrocytes are generated in adult mouse cerebellum. • Very few mash1{sup +} or nestin{sup +} stem cells exist, and most of them are quiescent. • Cell proliferation rate is diversified among cerebellar regions and decreases over time. - Abstract: Although previous studies implied that cerebellar stem cells exist in some adult mammals, little is known about whether these stem cells can produce new neurons and astrocytes. In this study by bromodeoxyuridine (BrdU) intraperitoneal (i.p.) injection, we found that there are abundant BrdU{sup +} cells in adult mouse cerebellum, and their quantity and density decreases significantly over time. We also found cell proliferation rate is diversified in different cerebellar regions. Among these BrdU{sup +} cells, very few are mash1{sup +} or nestin{sup +} stem cells, and the vast majority of cerebellar stem cells are quiescent. Data obtained by in vivo retrovirus injection indicate that stem cells do not produce neurons and astrocytes in adult mouse cerebellum. Instead, some cells labeled by retrovirus are Iba1{sup +} microglia. These results indicate that very few stem cells exist in adult mouse cerebellum, and none of these stem cells contribute to neurogenesis and astrogenesis under physiological condition.

  10. Generation and characterization of LIF-dependent canine induced pluripotent stem cells from adult dermal fibroblasts.

    PubMed

    Whitworth, Deanne J; Ovchinnikov, Dmitry A; Wolvetang, Ernst J

    2012-08-10

    Dogs provide a more clinically relevant model of human disease than rodents, particularly with respect to hereditary diseases. Thus, the availability of canine stem cells will greatly facilitate the use of the dog in the development of stem cell-based gene therapies and regenerative medicine. In this study we describe the production of canine induced pluripotent stem cells (ciPSCs) from adult dermal fibroblasts. These cells have a morphology resembling previously described canine embryonic stem cells, a normal karyotype, and express pluripotency markers including alkaline phosphatase, Nanog, Oct4, Telomerase, SSEA1, SSEA4, TRA1-60, TRA1-81, and Rex1. Furthermore, the inactive X chromosome is reactivated indicating a ground-state pluripotency. In culture they readily form embryoid bodies, which in turn give rise to cell types from all 3 embryonic germ layers, as indicated by expression of the definitive endoderm markers Cxcr4 and α-fetoprotein, mesoderm markers Collagen IIA and Gata2, and ectoderm markers βIII-tubulin, Enolase, and Nestin. Of particular significance is the observation that these ciPSCs are dependent only on leukemia inhibitory factor (LIF), making them similar to mouse and canine embryonic stem cells, but strikingly unlike the ciPSCs recently described in two other studies, which were dependent on both basic fibroblast growth factor and LIF in order to maintain their pluripotency. Thus, our ciPSCs closely resemble mouse ESCs derived from the inner cell mass of preimplantation embryos, while the previously described ciPSCs appear to be more representative of cells from the epiblast of mouse postimplantation embryos.

  11. Reverse engineering human neurodegenerative disease using pluripotent stem cell technology.

    PubMed

    Liu, Ying; Deng, Wenbin

    2016-05-01

    With the technology of reprogramming somatic cells by introducing defined transcription factors that enables the generation of "induced pluripotent stem cells (iPSCs)" with pluripotency comparable to that of embryonic stem cells (ESCs), it has become possible to use this technology to produce various cells and tissues that have been difficult to obtain from living bodies. This advancement is bringing forth rapid progress in iPSC-based disease modeling, drug screening, and regenerative medicine. More and more studies have demonstrated that phenotypes of adult-onset neurodegenerative disorders could be rather faithfully recapitulated in iPSC-derived neural cell cultures. Moreover, despite the adult-onset nature of the diseases, pathogenic phenotypes and cellular abnormalities often exist in early developmental stages, providing new "windows of opportunity" for understanding mechanisms underlying neurodegenerative disorders and for discovering new medicines. The cell reprogramming technology enables a reverse engineering approach for modeling the cellular degenerative phenotypes of a wide range of human disorders. An excellent example is the study of the human neurodegenerative disease amyotrophic lateral sclerosis (ALS) using iPSCs. ALS is a progressive neurodegenerative disease characterized by the loss of upper and lower motor neurons (MNs), culminating in muscle wasting and death from respiratory failure. The iPSC approach provides innovative cell culture platforms to serve as ALS patient-derived model systems. Researchers have converted iPSCs derived from ALS patients into MNs and various types of glial cells, all of which are involved in ALS, to study the disease. The iPSC technology could be used to determine the role of specific genetic factors to track down what's wrong in the neurodegenerative disease process in the "disease-in-a-dish" model. Meanwhile, parallel experiments of targeting the same specific genes in human ESCs could also be performed to control

  12. The role of CD44 in fetal and adult hematopoietic stem cell regulation.

    PubMed

    Cao, Huimin; Heazlewood, Shen Y; Williams, Brenda; Cardozo, Daniela; Nigro, Julie; Oteiza, Ana; Nilsson, Susan K

    2016-01-01

    Throughout development, hematopoietic stem cells migrate to specific microenvironments, where their fate is, in part, extrinsically controlled. CD44 standard as a member of the cell adhesion molecule family is extensively expressed within adult bone marrow and has been previously reported to play important roles in adult hematopoietic regulation via CD44 standard-ligand interactions. In this manuscript, CD44 expression and function are further assessed and characterized on both fetal and adult hematopoietic stem cells. Using a CD44(-/-) mouse model, conserved functional roles of CD44 are revealed throughout development. CD44 is critical in the maintenance of hematopoietic stem and progenitor pools, as well as in hematopoietic stem cell migration. CD44 expression on hematopoietic stem cells as well as other hematopoietic cells within the bone marrow microenvironment is important in the homing and lodgment of adult hematopoietic stem cells isolated from the bone/bone marrow interface. CD44 is also involved in fetal hematopoietic stem cell migration out of the liver, via a process involving stromal cell-derived factor-1α. The absence of CD44 in neonatal bone marrow has no impact on the size of the long-term reconstituting hematopoietic stem cell pool, but results in an enhanced long-term engraftment potential of hematopoietic stem cells.

  13. Human amniotic fluid stem cells support undifferentiated propagation and pluripotency of human embryonic stem cell without b-FGF in a density dependent manner.

    PubMed

    Ma, Xiaorong; Li, Huanqi; Xin, Shujia; Ma, Yueting; Ouyang, Tianxiang

    2014-01-01

    Human embryonic stem cells (hESCs) are pluripotent cells which can give rise to almost all adult cell lineages. Culture system of hESCs is complex, requiring exogenous b-FGF and feeder cell layer. Human mesenchymal stem cells (MSCs) not only synthesize soluble cytokines or factors such as b-FGF, but also provide other mechanism which might play positive role on sustaining hESCs propagation and pluripotency. Human amniotic fluid stem (AFS) cells, which share characteristics of both embryonic and adult stem cells, have been regarded as promising cells for regenerative medicine. Taking advantage by AFS cells, we studied the ability of AFS cells in supporting undifferentiated propagation and pluripotency of Chinese population derived X-01 hESCs. Human AF-type amniotic fluid stem cells (hAF-AFSCs) transcribed genes including Activin A, TGF-β1, Noggin and b-FGF, which involved in maintaining pluripotency and self-renewal of hESCs. Compared to mouse embryonic fibroblasts (MEFs), hAF-AFSCs secreted higher concentration of b-FGF which was important in hESCs culture (P < 0.05). The hESCs were propagated more than 30 passages on hAF-AFSCs layer with exogenous b-FGF supplementation, keeping undifferentiated status. While exogenous b-FGF was obviated, propagation of hESCs with undifferentiated status was dependent on density of hAF-AFSC feeder layer. Lower density of hAF-AFSCs resulted in rapid decline in undifferentiated clone number, while higher ones hindered the growth of colonies. The most appropriate hAF-AFSCs feeder density to maintain the X-01 hESC line without exogenous b-FGF was 15-20×10(4)/well. To the best of our knowledge, this is the first study demonstrating that hAF-AFSCs could support undifferentiated propagation and pluripotency of Chinese population derived hESCs without exogenous b-FGF supplementation.

  14. Immunological properties of embryonic and adult stem cells

    PubMed Central

    Bifari, Francesco; Pacelli, Luciano; Krampera, Mauro

    2010-01-01

    The possibility of treating degenerative diseases by stem cell-based approaches is a promising therapeutical option. Among major concerns for the clinical application of stem cells, some derive from the possibility that stem cells may be rejected by the immune system as a consequence of histoincompatibility and that stem cells themselves may interfere with the normal functions of host immune response. Therefore, the immunogenicity and the immunomodulatory properties of stem cells must be carefully addressed. Although these properties are common features of different stem cell types, some peculiarities can be recognized and characterized for their proper clinical use. PMID:21607122

  15. Developing defined culture systems for human pluripotent stem cells

    PubMed Central

    Valamehr, Bahram; Tsutsui, Hideaki; Ho, Chih-Ming; Wu, Hong

    2013-01-01

    Human pluripotent stem cells hold promising potential in many therapeutics applications including regenerative medicine and drug discovery. Over the past three decades, embryonic stem cell research has illustrated that embryonic stem cells possess two important and distinct properties: the ability to continuously self-renew and the ability to differentiate into all specialized cell types. In this article, we will discuss the continuing evolution of human pluripotent stem cell culture by examining requirements needed for the maintenance of self-renewal in vitro. We will also elaborate on the future direction of the field toward generating a robust and completely defined culture system, which has brought forth collaborations amongst biologists and engineers. As human pluripotent stem cell research progresses towards identifying solutions for debilitating diseases, it will be critical to establish a defined, reproducible and scalable culture system to meet the requirements of these clinical applications. PMID:21916597

  16. Generation of induced pluripotent stem cells from human blood.

    PubMed

    Loh, Yuin-Han; Agarwal, Suneet; Park, In-Hyun; Urbach, Achia; Huo, Hongguang; Heffner, Garrett C; Kim, Kitai; Miller, Justine D; Ng, Kitwa; Daley, George Q

    2009-05-28

    Human dermal fibroblasts obtained by skin biopsy can be reprogrammed directly to pluripotency by the ectopic expression of defined transcription factors. Here, we describe the derivation of induced pluripotent stem cells from CD34+ mobilized human peripheral blood cells using retroviral transduction of OCT4/SOX2/KLF4/MYC. Blood-derived human induced pluripotent stem cells are indistinguishable from human embryonic stem cells with respect to morphology, expression of surface antigens, and pluripotency-associated transcription factors, DNA methylation status at pluripotent cell-specific genes, and the capacity to differentiate in vitro and in teratomas. The ability to reprogram cells from human blood will allow the generation of patient-specific stem cells for diseases in which the disease-causing somatic mutations are restricted to cells of the hematopoietic lineage. PMID:19299331

  17. Ex vivo targeting of p21Cip1/Waf1 permits relative expansion of human hematopoietic stem cells.

    PubMed

    Stier, Sebastian; Cheng, Tao; Forkert, Randolf; Lutz, Christoph; Dombkowski, David M; Zhang, Jie Lin; Scadden, David T

    2003-08-15

    Relative quiescence is a defining characteristic of hematopoietic stem cells. Reasoning that inhibitory tone dominates control of stem cell cycling, we previously showed that mice engineered to be deficient in the cyclin-dependent kinase inhibitor, p21Cip1/Waf1 (p21), have an increased stem cell pool under homeostatic conditions. Since p21 was necessary to maintain stem cell quiescence and its absence sufficient to permit increased murine stem cell cycling, we tested whether reduction of p21 alone in human adult-derived stem cells could affect stem cell proliferation. We demonstrate here that interrupting p21 expression ex vivo resulted in expanded stem cell number and in vivo stem cell function compared with control, manipulated cells. Further, we demonstrate full multilineage reconstitution capability in cells where p21 expression was knocked down. Therefore, lifting the brake on cell proliferation by altering cell cycle checkpoints provides an alternative paradigm for increasing hematopoietic stem cell numbers. This approach may be useful for relative ex vivo human stem cell expansion. PMID:12702511

  18. Adult bone marrow: which stem cells for cellular therapy protocols in neurodegenerative disorders?

    PubMed

    Wislet-Gendebien, Sabine; Laudet, Emerence; Neirinckx, Virginie; Rogister, Bernard

    2012-01-01

    The generation of neuronal cells from stem cells obtained from adult bone marrow is of significant clinical interest in order to design new cell therapy protocols for several neurological disorders. The recent identification in adult bone marrow of stem cells derived from the neural crests (NCSCs) might explain the neuronal phenotypic plasticity shown by bone marrow cells. However, little information is available about the nature of these cells compared to mesenchymal stem cells (MSCs). In this paper, we will review all information available concerning NCSC from adult tissues and their possible use in regenerative medicine. Moreover, as multiple recent studies showed the beneficial effect of bone marrow stromal cells in neurodegenerative diseases, we will discuss which stem cells isolated from adult bone marrow should be more suitable for cell replacement therapy.

  19. High-efficiency immunomagnetic isolation of solid tissue-originated integrin-expressing adult stem cells.

    PubMed

    Palmon, Aaron; David, Ran; Neumann, Yoav; Stiubea-Cohen, Raluca; Krief, Guy; Aframian, Doron J

    2012-02-01

    Isolation of highly pure specific cell types is crucial for successful adult stem cell-based therapy. As the number of such cells in adult tissue is low, an extremely efficient method is needed for their isolation. Here, we describe cell-separation methodologies based on magnetic-affinity cell sorting (MACS) MicroBeads with monoclonal antibodies against specific membrane proteins conjugated to superparamagnetic particles. Cells labeled with MACS MicroBeads are retained in a magnetic field within a MACS column placed in a MACS separator, allowing fast and efficient separation. Both positively labeled and non-labeled fractions can be used directly for downstream applications as the separated cell fractions remain viable with no functional impairment. As immunomagnetic separation depends on the interaction between a cell's membrane and the magnetically labeled antibody, separation of specific cells originating from solid tissues is more complex and demands a cell-dissociating pretreatment. In this paper, we detail the use of immunomagnetic separation for the purpose of regenerating damaged salivary gland (SG) function in animal and human models of irradiated head and neck cancer. Each year 500,000 new cases of head and neck cancer occur worldwide. Most of these patients lose SG function following irradiation therapy. SGs contain integrin α6β1-expressing epithelial stem cells. We hypothesized that these cells can be isolated, multiplied in culture and auto-implanted into the irradiated SGs to regenerate damaged SG function.

  20. Generation of functional hepatocytes from human spermatogonial stem cells

    PubMed Central

    Chen, Zheng; Sun, Min; Yuan, Qingqing; Niu, Minghui; Yao, Chencheng; Hou, Jingmei; Wang, Hong; Wen, Liping; Liu, Yun; Li, Zheng; He, Zuping

    2016-01-01

    To generate functional human hepatocytes from stem cells and/or extra-hepatic tissues could provide an important source of cells for treating liver diseases. Spermatogonial stem cells (SSCs) have an unlimited plasticity since they can dedifferentiate and transdifferentiate to other cell lineages. However, generation of mature and functional hepatocytes from human SSCs has not yet been achieved. Here we have for the first time reported direct transdifferentiation of human SSCs to mature and functional hepatocytes by three-step induction using the defined condition medium. Human SSCs were first transdifferentiated to hepatic stem cells, as evidenced by their morphology and biopotential nature of co-expressing hepatocyte and cholangiocyte markers but not hallmarks for embryonic stem cells. Hepatic stem cells were further induced to differentiate into mature hepatocytes identified by their morphological traits and strong expression of CK8, CK18, ALB, AAT, TF, TAT, and cytochrome enzymes rather than CK7 or CK19. Significantly, mature hepatocytes derived from human SSCs assumed functional attributes of human hepatocytes, because they could produce albumin, remove ammonia, and uptake and release indocyanine green. Moreover, expression of β-CATENIN, HNF4A, FOXA1 and GATA4 was upregulated during the transdifferentiation of human SSCs to mature hepatocytes. Collectively, human SSCs could directly transdifferentiate to mature and functional hepatocytes. This study could offer an invaluable source of human hepatocytes for curing liver disorders and drug toxicology screening and provide novel insights into mechanisms underlying human liver regeneration. PMID:26840458

  1. Adult stem cells in neural repair: Current options, limitations and perspectives.

    PubMed

    Mariano, Eric Domingos; Teixeira, Manoel Jacobsen; Marie, Suely Kazue Nagahashi; Lepski, Guilherme

    2015-03-26

    Stem cells represent a promising step for the future of regenerative medicine. As they are able to differentiate into any cell type, tissue or organ, these cells are great candidates for treatments against the worst diseases that defy doctors and researchers around the world. Stem cells can be divided into three main groups: (1) embryonic stem cells; (2) fetal stem cells; and (3) adult stem cells. In terms of their capacity for proliferation, stem cells are also classified as totipotent, pluripotent or multipotent. Adult stem cells, also known as somatic cells, are found in various regions of the adult organism, such as bone marrow, skin, eyes, viscera and brain. They can differentiate into unipotent cells of the residing tissue, generally for the purpose of repair. These cells represent an excellent choice in regenerative medicine, every patient can be a donor of adult stem cells to provide a more customized and efficient therapy against various diseases, in other words, they allow the opportunity of autologous transplantation. But in order to start clinical trials and achieve great results, we need to understand how these cells interact with the host tissue, how they can manipulate or be manipulated by the microenvironment where they will be transplanted and for how long they can maintain their multipotent state to provide a full regeneration.

  2. Epistatic adult plant resistance in wheat to stem rust cosegregates with Sr12 seedling resistance

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Wheat adult plant resistance (APR) to stem rust is desirable. Researchers have characterized the inheritance of APR in cultivar Thatcher as complex. In order to identify the loci providing APR in Thatcher, we evaluated 160 RILs derived from Thatcher/McNeal for stem rust reaction in the field in Keny...

  3. Lin-28 promotes symmetric stem cell division and drives adaptive growth in the adult Drosophila intestine.

    PubMed

    Chen, Ching-Huan; Luhur, Arthur; Sokol, Nicholas

    2015-10-15

    Stem cells switch between asymmetric and symmetric division to expand in number as tissues grow during development and in response to environmental changes. The stem cell intrinsic proteins controlling this switch are largely unknown, but one candidate is the Lin-28 pluripotency factor. A conserved RNA-binding protein that is downregulated in most animals as they develop from embryos to adults, Lin-28 persists in populations of adult stem cells. Its function in these cells has not been previously characterized. Here, we report that Lin-28 is highly enriched in adult intestinal stem cells in the Drosophila intestine. lin-28 null mutants are homozygous viable but display defects in this population of cells, which fail to undergo a characteristic food-triggered expansion in number and have reduced rates of symmetric division as well as reduced insulin signaling. Immunoprecipitation of Lin-28-bound mRNAs identified Insulin-like Receptor (InR), forced expression of which completely rescues lin-28-associated defects in intestinal stem cell number and division pattern. Furthermore, this stem cell activity of lin-28 is independent of one well-known lin-28 target, the microRNA let-7, which has limited expression in the intestinal epithelium. These results identify Lin-28 as a stem cell intrinsic factor that boosts insulin signaling in intestinal progenitor cells and promotes their symmetric division in response to nutrients, defining a mechanism through which Lin-28 controls the adult stem cell division patterns that underlie tissue homeostasis and regeneration.

  4. Lin-28 promotes symmetric stem cell division and drives adaptive growth in the adult Drosophila intestine

    PubMed Central

    Chen, Ching-Huan; Luhur, Arthur; Sokol, Nicholas

    2015-01-01

    Stem cells switch between asymmetric and symmetric division to expand in number as tissues grow during development and in response to environmental changes. The stem cell intrinsic proteins controlling this switch are largely unknown, but one candidate is the Lin-28 pluripotency factor. A conserved RNA-binding protein that is downregulated in most animals as they develop from embryos to adults, Lin-28 persists in populations of adult stem cells. Its function in these cells has not been previously characterized. Here, we report that Lin-28 is highly enriched in adult intestinal stem cells in the Drosophila intestine. lin-28 null mutants are homozygous viable but display defects in this population of cells, which fail to undergo a characteristic food-triggered expansion in number and have reduced rates of symmetric division as well as reduced insulin signaling. Immunoprecipitation of Lin-28-bound mRNAs identified Insulin-like Receptor (InR), forced expression of which completely rescues lin-28-associated defects in intestinal stem cell number and division pattern. Furthermore, this stem cell activity of lin-28 is independent of one well-known lin-28 target, the microRNA let-7, which has limited expression in the intestinal epithelium. These results identify Lin-28 as a stem cell intrinsic factor that boosts insulin signaling in intestinal progenitor cells and promotes their symmetric division in response to nutrients, defining a mechanism through which Lin-28 controls the adult stem cell division patterns that underlie tissue homeostasis and regeneration. PMID:26487778

  5. Aggregation of Human Eyelid Adipose-derived Stem Cells by Human Body Fluids.

    PubMed

    Song, Yeonhwa; Yun, Sujin; Yang, Hye Jin; Yoon, A Young; Kim, Haekwon

    2012-12-01

    Fetal bovine serum (FBS) is the most frequently used serum for the cultivation of mammalian cells. However, since animal-derived materials might not be appropriate due to safety issues, allogeneic human serum (HS) has been used to replace FBS, particularly for the culture of human cells. While there has been a debate about the advantages of HS, its precise effect on human adult stem cells have not been clarified. The present study aimed to investigate the effect of HS on the human eyelid adipose stem cells (HEACs) in vitro. When HEACs were cultivated in a medium containing 10% HS, many cells moved into several spots and aggregated there. The phenomenon was observed as early as 9 days following 10% HS treatment, and 12 days following 5% HS plus 5% FBS treatment. However, the aggregation was never observed when the same cells were cultivated with 10% FBS or bovine serum albumin. To examine whether cell density might affect the aggregation, cells were seeded with different densities on 12-well dish. Until the beginning of aggregation, cells seeded at low densities exhibited the longest culture period of 16 days whereas cells seeded at high densities showed the shortest period of 9 days to form aggregation. The number of cells was 15.1±0.2×10(4) as the least for the low density group, and 29.3±2.8×10(4) as the greatest for the high density group. When human cord blood serum or normal bovine serum was examined for the same effect on HEACs, interestingly, cord blood serum induced the aggregation of cells whereas bovine serum treatment has never induced. When cells were cultivated with 10% HS for 9 days, they were obtained and analyzed by RT-PCR. Compared to FBS-cultivated HEACs, HS-cultivated HEACs did not express VIM, and less expressed GATA4, PALLD. On the other hand, HS-cultivated HEACs expressed MAP2 more than FBS-cultivated HEACs. In conclusion, human adult stem cells could move and form aggregates by the treatment with human body fluids.

  6. Aggregation of Human Eyelid Adipose-derived Stem Cells by Human Body Fluids

    PubMed Central

    Song, Yeonhwa; Yun, Sujin; Yang, Hye Jin; Yoon, A Young; Kim, Haekwon

    2012-01-01

    Fetal bovine serum (FBS) is the most frequently used serum for the cultivation of mammalian cells. However, since animal-derived materials might not be appropriate due to safety issues, allogeneic human serum (HS) has been used to replace FBS, particularly for the culture of human cells. While there has been a debate about the advantages of HS, its precise effect on human adult stem cells have not been clarified. The present study aimed to investigate the effect of HS on the human eyelid adipose stem cells (HEACs) in vitro. When HEACs were cultivated in a medium containing 10% HS, many cells moved into several spots and aggregated there. The phenomenon was observed as early as 9 days following 10% HS treatment, and 12 days following 5% HS plus 5% FBS treatment. However, the aggregation was never observed when the same cells were cultivated with 10% FBS or bovine serum albumin. To examine whether cell density might affect the aggregation, cells were seeded with different densities on 12-well dish. Until the beginning of aggregation, cells seeded at low densities exhibited the longest culture period of 16 days whereas cells seeded at high densities showed the shortest period of 9 days to form aggregation. The number of cells was 15.1±0.2×104 as the least for the low density group, and 29.3±2.8×104 as the greatest for the high density group. When human cord blood serum or normal bovine serum was examined for the same effect on HEACs, interestingly, cord blood serum induced the aggregation of cells whereas bovine serum treatment has never induced. When cells were cultivated with 10% HS for 9 days, they were obtained and analyzed by RT-PCR. Compared to FBS-cultivated HEACs, HS-cultivated HEACs did not express VIM, and less expressed GATA4, PALLD. On the other hand, HS-cultivated HEACs expressed MAP2 more than FBS-cultivated HEACs. In conclusion, human adult stem cells could move and form aggregates by the treatment with human body fluids. PMID:25949109

  7. Testicular Niche Required for Human Spermatogonial Stem Cell Expansion

    PubMed Central

    Smith, James F.; Yango, Pamela; Altman, Eran; Choudhry, Shweta; Poelzl, Andrea; Zamah, Alberuni M.; Rosen, Mitchell; Klatsky, Peter C.

    2014-01-01

    Prepubertal boys treated with high-dose chemotherapy do not have an established means of fertility preservation because no established in vitro technique exists to expand and mature purified spermatogonial stem cells (SSCs) to functional sperm in humans. In this study, we define and characterize the unique testicular cellular niche required for SSC expansion using testicular tissues from men with normal spermatogenesis. Highly purified SSCs and testicular somatic cells were isolated by fluorescence-activated cell sorting using SSEA-4 and THY1 as markers of SSCs and somatic cells. Cells were cultured on various established niches to assess their role in SSC expansion in a defined somatic cellular niche. Of all the niches examined, cells in the SSEA-4 population exclusively bound to adult testicular stromal cells, established colonies, and expanded. Further characterization of these testicular stromal cells revealed distinct mesenchymal markers and the ability to undergo differentiation along the mesenchymal lineage, supporting a testicular multipotent stromal cell origin. In vitro human SSC expansion requires a unique niche provided exclusively by testicular multipotent stromal cells with mesenchymal properties. These findings provide an important foundation for developing methods of inducing SSC growth and maturation in prepubertal testicular tissue, essential to enabling fertility preservation for these boys. PMID:25038247

  8. Characterizing the radioresponse of pluripotent and multipotent human stem cells.

    PubMed

    Lan, Mary L; Acharya, Munjal M; Tran, Katherine K; Bahari-Kashani, Jessica; Patel, Neal H; Strnadel, Jan; Giedzinski, Erich; Limoli, Charles L

    2012-01-01

    The potential capability of stem cells to restore functionality to diseased or aged tissues has prompted a surge of research, but much work remains to elucidate the response of these cells to genotoxic agents. To more fully understand the impact of irradiation on different stem cell types, the present study has analyzed the radioresponse of human pluripotent and multipotent stem cells. Human embryonic stem (ES) cells, human induced pluripotent (iPS) cells, and iPS-derived human neural stem cells (iPS-hNSCs) cells were irradiated and analyzed for cell survival parameters, differentiation, DNA damage and repair and oxidative stress at various times after exposure. While irradiation led to dose-dependent reductions in survival, the fraction of surviving cells exhibited dose-dependent increases in metabolic activity. Irradiation did not preclude germ layer commitment of ES cells, but did promote neuronal differentiation. ES cells subjected to irradiation exhibited early apoptosis and inhibition of cell cycle progression, but otherwise showed normal repair of DNA double-strand breaks. Cells surviving irradiation also showed acute and persistent increases in reactive oxygen and nitrogen species that were significant at nearly all post-irradiation times analyzed. We suggest that stem cells alter their redox homeostasis to adapt to adverse conditions and that radiation-induced oxidative stress plays a role in regulating the function and fate of stem cells within tissues compromised by radiation injury.

  9. Glycomics of human embryonic stem cells and human induced pluripotent stem cells.

    PubMed

    Furukawa, Jun-Ichi; Okada, Kazue; Shinohara, Yasuro

    2016-10-01

    Most cells are coated by a dense glycocalyx composed of glycoconjugates such as glycosphingolipids, glycoproteins, and proteoglycans. The overall glycomic profile is believed to be crucial for the diverse roles of glycans, which are mediated by specific interactions that regulate cell-cell adhesion, the immune response, microbial pathogenesis, and other cellular events. Many cell surface markers were discovered and identified as glycoconjugates such as stage-specific embryonic antigen, Tra-1-60/81 and various other cell surface molecules (e.g., cluster of differentiation). Recent progress in the development of analytical methodologies and strategies has begun to clarify the cellular glycomics of various cells including human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs). The glycomic profiles of these cells are highly cell type-specific and reflect cellular alterations, such as development, differentiation and cancerous change. In this mini review, we briefly summarize the glycosylation spectra specific to hESCs and hiPSCs, which cover glycans of all major glycoconjugates (i.e., glycosphingolipids, N- and O-glycans of glycoproteins, and glycosaminoglycans) and free oligosaccharides.

  10. The therapeutic potential of human olfactory-derived stem cells.

    PubMed

    Marshall, C T; Lu, C; Winstead, W; Zhang, X; Xiao, M; Harding, G; Klueber, K M; Roisen, F J

    2006-06-01

    Stem cells from fetal and adult central nervous system have been isolated and characterized, providing populations for potential replacement therapy for traumatic injury repair and neurodegenerative diseases. The regenerative capacity of the olfactory system has attracted scientific interest. Studies focusing on animal and human olfactory bulb ensheathing cells (OECs) have heightened the expectations that OECs can enhance axonal regeneration and repair demyelinating diseases. Harvest of OECs from the olfactory bulb requires highly invasive surgery, which is a major obstacle. In contrast, olfactory epithelium (OE) has a unique regenerative capacity and is readily accessible from its location in the nasal cavity, allowing for harvest without lasting damage to the donor. Adult OE contains progenitors responsible for the normal life-long continuous replacement of neurons and supporting cells. Culture techniques have been established for human OE that generate populations of mitotically active neural progenitors that form neurospheres (Roisen et al., 2001; Winstead et al., 2005). The potential application of this technology includes autologous transplantation where minimal donor material can be isolated, expanded ex vivo, and lineage restricted to a desired phenotype prior to/or after re-implantation. Furthermore, these strategies circumvent the ethical issues that arise with embryonic or fetal tissues. The long term goal is to develop procedures through which a victim of a spinal cord injury or neurodegenerative condition would serve as a source of progenitors for his/her own regenerative grafts, avoiding the need for immunosuppression and ethical controversy. In addition, these cells can provide populations for pharmacological and/or diagnostic evaluation.

  11. Human Dermal Stem/Progenitor Cell-Derived Conditioned Medium Improves Senescent Human Dermal Fibroblasts.

    PubMed

    Jung, Ji-Yong; Shim, Joong Hyun; Choi, Hyun; Lee, Tae Ryong; Shin, Dong Wook

    2015-08-13

    Adult skin stem cells are recognized as potential therapeutics to rejuvenate aged skin. We previously demonstrated that human dermal stem/progenitor cells (hDSPCs) with multipotent capacity could be enriched from human dermal fibroblasts using collagen type IV. However, the effects of hDSPCs on cellular senescence remain to be elucidated. In the present study, we investigated whether conditioned medium (CM) collected from hDSPC cultures (hDSPC-CM) exhibits beneficial effects on senescent fibroblasts. We found that hDSPC-CM promoted proliferation and decreased the expression level of senescence-associated β-galactosidase in senescent fibroblasts. In addition, p53 phosphorylation and p21 expression were significantly reduced in senescent fibroblasts treated with hDSPC-CM. hDSPC-CM restored the expression levels of collagen type I, collagen type III, and tissue inhibitor of metalloproteinase, and antagonized the increase of matrix metalloproteinase 1 expression. Finally, we demonstrated that hDSPC-CM significantly reduced reactive oxygen species levels by specifically up-regulating the expression level of superoxide dismutase 2. Taken together, these data suggest that hDSPC-CM can be applied as a potential therapeutic agent for improving human aged skin.

  12. The architectural organization of human stem cell cycle regulatory machinery.

    PubMed

    Stein, Gary S; Stein, Janet L; van J Wijnen, Andre; Lian, Jane B; Montecino, Martin; Medina, Ricardo; Kapinas, Kristie; Ghule, Prachi; Grandy, Rodrigo; Zaidi, Sayyed K; Becker, Klaus A

    2012-01-01

    Two striking features of human embryonic stem cells that support biological activity are an abbreviated cell cycle and reduced complexity to nuclear organization. The potential implications for rapid proliferation of human embryonic stem cells within the context of sustaining pluripotency, suppressing phenotypic gene expression and linkage to simplicity in the architectural compartmentalization of regulatory machinery in nuclear microenvironments is explored. Characterization of the molecular and architectural commitment steps that license human embryonic stem cells to initiate histone gene expression is providing understanding of the principal regulatory mechanisms that control the G1/S phase transition in primitive pluripotent cells. From both fundamental regulatory and clinical perspectives, further understanding of the pluripotent cell cycle in relation to compartmentalization of regulatory machinery in nuclear microenvironments is relevant to applications of stem cells for regenerative medicine and new dimensions to therapy where traditional drug discovery strategies have been minimally effective.

  13. Cryopreservation of Human Stem Cells for Clinical Application: A Review

    PubMed Central

    Hunt, Charles J.

    2011-01-01

    Summary Stem cells have been used in a clinical setting for many years. Haematopoietic stem cells have been used for the treatment of both haematological and non-haematological disease; while more recently mesenchymal stem cells derived from bone marrow have been the subject of both laboratory and early clinical studies. Whilst these cells show both multipotency and expansion potential, they nonetheless do not form stable cell lines in culture which is likely to limit the breadth of their application in the field of regenerative medicine. Human embryonic stem cells are pluripotent cells, capable of forming stable cell lines which retain the capacity to differentiate into cells from all three germ layers. This makes them of special significance in both regenerative medicine and toxicology. Induced pluripotent stem (iPS) cells may also provide a similar breadth of utility without some of the confounding ethical issues surrounding embryonic stem cells. An essential pre-requisite to the commercial and clinical application of stem cells are suitable cryopreservation protocols for long-term storage. Whilst effective methods for cryopreservation and storage have been developed for haematopoietic and mesenchymal stem cells, embryonic cells and iPS cells have proved more refractory. This paper reviews the current state of cryopreservation as it pertains to stem cells and in particular the embryonic and iPS cell. PMID:21566712

  14. Comparison of Human Fetal Liver, Umbilical Cord Blood, and Adult Blood Hematopoietic Stem Cell Engraftment in NOD-scid/γc−/−, Balb/c-Rag1−/−γc−/−, and C.B-17-scid/bg Immunodeficient Mice

    PubMed Central

    Lepus, Christin M.; Gibson, Thomas F.; Gerber, Scott A.; Kawikova, Ivana; Szczepanik, Marian; Hossain, Jaber; Ablamunits, Vitaly; Kirkiles-Smith, Nancy; Herold, Kevan C.; Donis, Ruben O.; Bothwell, Alfred L.; Pober, Jordan S.; Harding, Martha J.

    2010-01-01

    Immunodeficient mice bearing components of a human immune system present a novel approach for studying human immune responses. We investigated the number, phenotype, developmental kinetics and function of developing human immune cells following transfer of CD34+ hematopoietic stem cell (HSC) preparations, originating from second trimester human fetal liver (HFL), umbilical cord blood (UCB), or granulocyte colony-stimulating factor-mobilized adult blood (G-CSF-AB) delivered via intrahepatic injection into sublethally irradiated neonatal NOD-scid/γc−/−, Balb/c-Rag1−/−γc−/−, and C.B-17-scid/bg mice. HFL and UCB HSC provided the greatest number and breadth of developing cells. NOD-scid/γc−/− and Balb/c-Rag1−/−γc−/− harbored human B and dendritic cells as well as human platelets in peripheral blood, whereas NOD-scid/γc−/− mice harbored higher levels of human T cells. NOD-scid/γc−/− mice engrafted with HFL CD34+ HSC demonstrated human immunological competence evidenced by white pulp expansion and increases in total human immunoglobulin following immunization with T-dependent antigens, and delayed type hypersensitivity-infiltrating leukocytes in response to antigenic challenge. In conclusion, we describe an encouraging base system for studying human hematopoietic lineage development and function utilizing human HFL or UCB HSC-engrafted NOD-scid/γc−/− mice that is well suited for future studies toward the development of a fully competent humanized mouse model. PMID:19524633

  15. Somatic stem cells express Piwi and Vasa genes in an adult ctenophore: ancient association of "germline genes" with stemness.

    PubMed

    Alié, Alexandre; Leclère, Lucas; Jager, Muriel; Dayraud, Cyrielle; Chang, Patrick; Le Guyader, Hervé; Quéinnec, Eric; Manuel, Michaël

    2011-02-01

    Stem cells are essential for animal development and adult tissue homeostasis, and the quest for an ancestral gene fingerprint of stemness is a major challenge for evolutionary developmental biology. Recent studies have indicated that a series of genes, including the transposon silencer Piwi and the translational activator Vasa, specifically involved in germline determination and maintenance in classical bilaterian models (e.g., vertebrates, fly, nematode), are more generally expressed in adult multipotent stem cells in other animals like flatworms and hydras. Since the progeny of these multipotent stem cells includes both somatic and germinal derivatives, it remains unclear whether Vasa, Piwi, and associated genes like Bruno and PL10 were ancestrally linked to stemness, or to germinal potential. We have investigated the expression of Vasa, two Piwi paralogues, Bruno and PL10 in Pleurobrachia pileus, a member of the early-diverging phylum Ctenophora, the probable sister group of cnidarians. These genes were all expressed in the male and female germlines, and with the exception of one of the Piwi paralogues, they showed similar expression patterns within somatic territories (tentacle root, comb rows, aboral sensory complex). Cytological observations and EdU DNA-labelling and long-term retention experiments revealed concentrations of stem cells closely matching these gene expression areas. These stem cell pools are spatially restricted, and each specialised in the production of particular types of somatic cells. These data unveil important aspects of cell renewal within the ctenophore body and suggest that Piwi, Vasa, Bruno, and PL10 belong to a gene network ancestrally acting in two distinct contexts: (i) the germline and (ii) stem cells, whatever the nature of their progeny.

  16. Engineering physiologically stiff and stratified human cartilage by fusing condensed mesenchymal stem cells.

    PubMed

    Bhumiratana, Sarindr; Vunjak-Novakovic, Gordana

    2015-08-01

    For a long time, clinically sized and mechanically functional cartilage could be engineered from young animal chondrocytes, but not from adult human mesenchymal stem cells that are of primary clinical interest. The approaches developed for primary chondrocytes were not successful when used with human mesenchymal cells. The method discussed here was designed to employ a mechanism similar to pre-cartilaginous condensation and fusion of mesenchymal stem cells at a precisely defined time. The formation of cartilage was initiated by press-molding the mesenchymal bodies onto the surface of a bone substrate. By image-guided fabrication of the bone substrate and the molds, the osteochondral constructs were engineered in anatomically precise shapes and sizes. After 5 weeks of cultivation, the cartilage layer assumed physiologically stratified histomorphology, and contained lubricin at the surface, proteoglycans and type II collagen in the bulk phase, collagen type X at the interface with the bone substrate, and collagen type I within the bone phase. For the first time, the Young's modulus and the friction coefficient of human cartilage engineered from mesenchymal stem cells reached physiological levels for adult human cartilage. We propose that this method can be effective for generating human osteochondral tissue constructs.

  17. [The three-dimensional culture of adult mesenchymal stem cells for intervertebral disc tissue engineering].

    PubMed

    Feng, Ganjun; Liu, Hao; Deng, Li; Chen, Xiaohe; Zhao, Xianfeng; Liang, Tao; Li, Xiuqiong

    2009-12-01

    Intervertebral disc (IVD) degeneration is one of the major causes of low back pain. As current clinical treatments are aimed at restoring biomechanical function and providing symptomatic relief, the methods focused on biological repair have aroused interest and several tissue engineering approaches using different cell types have been proposed. Owing to the unsuitable nature of degenerate cells for tissue engineering, attention has been given to the use of mesenchymal stem cells (MSCs). In this connection, we have made a study on the characteristics of MSCs derived from adult bone marrow and on the feasibility of constructing IVD tissue-engineering cell under a Three-Dimensional Pellet Culture System. The human bone marrow MSCs were isolated and purified with density gradient solution and attachment-independent culture system. MSCs isolated using this method are a homogeneous population as indicated by morphology and other criteria. They have the capacity for self-renewal and proliferation, and the multilineage potential to differentiate.

  18. Progress in stem cell therapy for major human neurological disorders.

    PubMed

    Martínez-Morales, P L; Revilla, A; Ocaña, I; González, C; Sainz, P; McGuire, D; Liste, I

    2013-10-01

    Human neurological disorders such as Alzheimer's disease (AD), Parkinson's disease, stroke or spinal cord injury are caused by the loss of neurons and glial cells in the brain or spinal cord in the Central Nervous System (CNS). Stem cell technology has become an attractive option to investigate and treat these diseases. Several types of neurons and glial cells have successfully been generated from stem cells, which in some cases, have ameliorated some dysfunctions both in animal models of neurological disorders and in patients at clinical level. Stem cell-based therapies can be beneficial by acting through several mechanisms such as cell replacement, modulation of inflammation and trophic actions. Here we review recent and current remarkable clinical studies involving stem cell-based therapy for AD and stroke and provide an overview of the different types of stem cells available nowadays, their main properties and how they are developing as a possible therapy for neurological disorders.

  19. Ethical questions concerning research on human embryos, embryonic stem cells and chimeras.

    PubMed

    Bobbert, Monika

    2006-12-01

    Research using human embryos and embryonic stem cells is viewed as important for various reasons. Apart from questions concerning legal regulations, numerous ethical objections are raised pertaining to the use of surplus embryos from reproductive medicine as well as the creation of embryos and stem cells through cloning. In the hopes of avoiding ethical problems, alternatives have been proposed including the extraction of egg cells from "dead" embryos derived from in vitro fertilization procedures, the extraction of pluripotent stem cells from blastocysts, technologies such as "altered nuclear transfer" (ANT) and "oocyte-assisted reprogramming" (ANT-OAR) as well as parthenogenesis. Initial ethical assessments show that certain questions pertaining to such strategies have remained unanswered. Furthermore, with the help of new or more differentiated biotechnological procedures, it is possible to create chimeras and hybrids in which human and non-human cells are combined. Human-animal chimeras, in which gametes or embryonic tissue have been mixed with embryonic or adult stem cells, demonstrate a different "quality" and "degree of penetration" from those produced in previous experiments. Not only does this have consequences regarding questions of patentability, this situation also raises fundamental questions concerning the human being's self image, the concept of person, identity and species and the moral rights and duties that are connected with such concepts. There is a need for legal regulation, on the national as well as the international level.

  20. Direct Reprogramming of Human Amniotic Fluid Stem Cells by OCT4 and Application in Repairing of Cerebral Ischemia Damage

    PubMed Central

    Qin, Mingde; Chen, Ruihua; Li, Hong; Liang, Hansi; Xue, Qun; Li, Fang; Chen, Ying; Zhang, Xueguang

    2016-01-01

    Amniotic fluid stem cells (AFSCs) are a type of fetal stem cell whose stemness encompasses both embryonic and adult stem cells, suggesting that they may be easily and efficiently reprogrammed into induced pluripotent stem cells (iPSCs). To further simplify the reprogramming process, the creation of AFSC-derived iPSCs using a single factor is desirable. Here we report the generation of one-factor human AFSC-iPSCs (AiPSCs) from human AFSCs by ectopic expression of the transcription factor OCT4. Just like human embryonic stem cells, AiPSCs exhibited similar epigenetic status, global gene expression profiles, teratoma formation and in vitro & in vivo pluripotency. Our results indicate that the OCT4 is necessary and sufficient to directly reprogram human AFSCs into pluripotent AiPSCs. Moreover, reflecting the similar memory characteristics of AFSCs and neural stem cells, we show that AiPSC membrane-derived vesicles (MVs) repair cerebral ischemia damage. We anticipate that the successful generation of one-factor AiPSCs will facilitate the creation of patient-specific pluripotent stem cells without the need for transgenic expression of oncogenes. Moreover, MVs from tissue-specific AiPSCs have potential in tissue repair, representing a novel application of iPSCs. PMID:27019637

  1. Derivation of human embryonic stem cells in defined conditions.

    PubMed

    Ludwig, Tenneille E; Levenstein, Mark E; Jones, Jeffrey M; Berggren, W Travis; Mitchen, Erika R; Frane, Jennifer L; Crandall, Leann J; Daigh, Christine A; Conard, Kevin R; Piekarczyk, Marian S; Llanas, Rachel A; Thomson, James A

    2006-02-01

    We have previously reported that high concentrations of basic fibroblast growth factor (bFGF) support feeder-independent growth of human embryonic stem (ES) cells, but those conditions included poorly defined serum and matrix components. Here we report feeder-independent human ES cell culture that includes protein components solely derived from recombinant sources or purified from human material. We describe the derivation of two new human ES cell lines in these defined culture conditions.

  2. Derivation of three new human embryonic stem cell lines.

    PubMed

    Bradley, Cara K; Chami, Omar; Peura, Teija T; Bosman, Alexis; Dumevska, Biljana; Schmidt, Uli; Stojanov, Tomas

    2010-04-01

    Human embryonic stem cells are pluripotent cells capable of extensive self-renewal and differentiation to all cells of the embryo proper. Here, we describe the derivation and characterization of three Sydney IVF human embryonic stem cell lines not already reported elsewhere, designated SIVF001, SIVF002, and SIVF014. The cell lines display typical compact colony morphology of embryonic stem cells, have stable growth rates over more than 40 passages and are cytogenetically normal. Furthermore, the cell lines express pluripotency markers including Nanog, Oct4, SSEA3 and Tra-1-81, and are capable of generating teratoma cells derived from each of the three germ layers in immunodeficient mice. These experiments show that the cell lines constitute pluripotent stem cell lines. PMID:20198447

  3. Stem Cells

    MedlinePlus

    Stem cells are cells with the potential to develop into many different types of cells in the body. They serve as a repair ... body. There are two main types of stem cells: embryonic stem cells and adult stem cells. Stem ...

  4. The ethics of patenting human embryonic stem cells.

    PubMed

    Chapman, Audrey R

    2009-09-01

    Just as human embryonic stem cell research has generated controversy about the uses of human embryos for research and therapeutic applications, human embryonic stem cell patents raise fundamental ethical issues. The United States Patent and Trademark Office has granted foundational patents, including a composition of matter (or product) patent to the Wisconsin Alumni Research Foundation (WARF), the University of Wisconsin-Madison's intellectual property office. In contrast, the European Patent Office rejected the same WARF patent application for ethical reasons. This article assesses the appropriateness of these patents placing the discussion in the context of the deontological and consequentialist ethical issues related to human embryonic stem cell patenting. It advocates for a patent system that explicitly takes ethical factors into account and explores options for new types of intellectual property arrangements consistent with ethical concerns.

  5. Hurdles to clinical translation of human induced pluripotent stem cells.

    PubMed

    Neofytou, Evgenios; O'Brien, Connor Galen; Couture, Larry A; Wu, Joseph C

    2015-07-01

    Human pluripotent stem cells are known to have the capacity to renew indefinitely, being intrinsically able to differentiate into many different cell types. These characteristics have generated tremendous enthusiasm about the potential applications of these cells in regenerative medicine. However, major challenges remain with the development and testing of novel experimental stem cell therapeutics in the field. In this Review, we focus on the nature of the preclinical challenges and discuss potential solutions that could help overcome them. Furthermore, we discuss the use of allogeneic versus autologous stem cell products, including a review of their respective advantages and disadvantages, major clinical requirements, quality standards, time lines, and costs of clinical grade development.

  6. Vascular potential of human pluripotent stem cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Cardiovascular disease is the number one cause of death and disability in the US. Understanding the biological activity of stem and progenitor cells, and their ability to contribute to the repair, regeneration and remodeling of the heart and blood vessels affected by pathological processes is an ess...

  7. [Ethical aspects of human embryonic stem cell use and commercial umbilical cord blood stem cell banking. Ethical reflections on the occasion of the regulation of the European Council and Parliament on advanced therapy medicinal products].

    PubMed

    Virt, G

    2010-01-01

    The regulation of the European Council and Parliament on advanced therapy medicinal products also includes therapies with human embryonic stem cells. The use of these stem cells is controversially and heavily discussed. Contrary to the use of adult stem cells, medical and ethical problems concerning the use of human embryonic stem cells persists, because this use is based on the destruction of human life at the very beginning. The regulation foresees, therefore, subsidiarity within the European Member States. Although there are no ethical problems in principle with the use of stem cells from the umbilical cord blood, there are social ethical doubts with the banking of these stem cells for autologous use without any currently foreseeable medical advantage by commercial blood banks. Also in this case subsidiarity is valid.

  8. Cryopreservation of human pluripotent stem cells: a general protocol.

    PubMed

    Miyazaki, Takamichi; Suemori, Hirofumi

    2015-01-01

    Cryopreservation is an essential technique to preserve stem cells, semipermanently sustaining their potentials. There are two main approaches of cryopreservation for human pluripotent stem cells (hPSCs). The first is the vitrification, which involves instantaneous freeze and thaw of hPSCs. The second is the conventional slow-cooling method and a rapid thaw. Both cryopreservation protocols have been standardized and optimized to yield high survivability of hPSCs.

  9. Derivation of a Homozygous Human Androgenetic Embryonic Stem Cell Line.

    PubMed

    Ding, Chenhui; Huang, Sunxing; Qi, Quan; Fu, Rui; Zhu, Wanwan; Cai, Bing; Hong, Pingping; Liu, Zhengxin; Gu, Tiantian; Zeng, Yanhong; Wang, Jing; Xu, Yanwen; Zhao, Xiaoyang; Zhou, Qi; Zhou, Canquan

    2015-10-01

    Human embryonic stem cells (hESCs) have long been considered as a promising source for cell replacement therapy. However, one major obstacle for the use of these cells is immune compatibility. Histocompatible human parthenogenetic ESCs have been reported as a new method for generating human leukocyte antigen (HLA)-matched hESCs. To further investigate the possibility of obtaining histocompatible stem cells from uniparental embryos, we tried to produce androgenetic haploid human embryos by injecting a single spermatozoon into enucleated human oocyte, and establish human androgenetic embryonic stem (hAGES) cell lines from androgenetic embryos. In the present study, a diploid hAGES cell line has been established, which exhibits typical features of human ESCs, including the expression of pluripotency markers, having differentiation potential in vitro and in vivo, and stable propagation in an undifferentiated state (>P40). Bisulfite sequencing of the H19, Snrpn, Meg3, and Kv imprinting control regions suggested that hAGES cells maintained to a certain extent a sperm methylation pattern. Genome-wide single nucleotide polymorphism, short tandem repeat, and HLA analyses revealed that the hAGES cell genome was highly homozygous. These results suggest that hAGES cells from spermatozoon could serve as a useful tool for studying the mechanisms underlying genomic imprinting in humans. It might also be used as a potential resource for cell replacement therapy as parthenogenetic stem cells.

  10. Cartilage tissue engineering identifies abnormal human induced pluripotent stem cells.

    PubMed

    Yamashita, Akihiro; Liu, Shiying; Woltjen, Knut; Thomas, Bradley; Meng, Guoliang; Hotta, Akitsu; Takahashi, Kazutoshi; Ellis, James; Yamanaka, Shinya; Rancourt, Derrick E

    2013-01-01

    Safety is the foremost issue in all human cell therapies, but human induced pluripotent stem cells (iPSCs) currently lack a useful safety indicator. Studies in chimeric mice have demonstrated that certain lines of iPSCs are tumorigenic; however a similar screen has not been developed for human iPSCs. Here, we show that in vitro cartilage tissue engineering is an excellent tool for screening human iPSC lines for tumorigenic potential. Although all human embryonic stem cells (ESCs) and most iPSC lines tested formed cartilage safely, certain human iPSCs displayed a pro-oncogenic state, as indicated by the presence of secretory tumors during cartilage differentiation in vitro. We observed five abnormal iPSC clones amoungst 21 lines derived from five different reprogramming methods using three cellular origins. We conclude that in vitro cartilage tissue engineering is a useful approach to identify abnormal human iPSC lines.

  11. Advances in culture and manipulation of human pluripotent stem cells.

    PubMed

    Qian, X; Villa-Diaz, L G; Krebsbach, P H

    2013-11-01

    Recent advances in the understanding of pluripotent stem cell biology and emerging technologies to reprogram somatic cells to a stem cell-like state are helping bring stem cell therapies for a range of human disorders closer to clinical reality. Human pluripotent stem cells (hPSCs) have become a promising resource for regenerative medicine and research into early development because these cells are able to self-renew indefinitely and are capable of differentiation into specialized cell types of all 3 germ layers and trophoectoderm. Human PSCs include embryonic stem cells (hESCs) derived from the inner cell mass of blastocyst-stage embryos and induced pluripotent stem cells (hiPSCs) generated via the reprogramming of somatic cells by the overexpression of key transcription factors. The application of hiPSCs and the finding that somatic cells can be directly reprogrammed into different cell types will likely have a significant impact on regenerative medicine. However, a major limitation for successful therapeutic application of hPSCs and their derivatives is the potential xenogeneic contamination and instability of current culture conditions. This review summarizes recent advances in hPSC culture and methods to induce controlled lineage differentiation through regulation of cell-signaling pathways and manipulation of gene expression as well as new trends in direct reprogramming of somatic cells.

  12. Recent developments in StemBase: a tool to study gene expression in human and murine stem cells

    PubMed Central

    Sandie, Reatha; Palidwor, Gareth A; Huska, Matthew R; Porter, Christopher J; Krzyzanowski, Paul M; Muro, Enrique M; Perez-Iratxeta, Carolina; Andrade-Navarro, Miguel A

    2009-01-01

    Background Currently one of the largest online repositories for human and mouse stem cell gene expression data, StemBase was first designed as a simple web-interface to DNA microarray data generated by the Canadian Stem Cell Network to facilitate the discovery of gene functions relevant to stem cell control and differentiation. Findings Since its creation, StemBase has grown in both size and scope into a system with analysis tools that examine either the whole database at once, or slices of data, based on tissue type, cell type or gene of interest. As of September 1, 2008, StemBase contains gene expression data (microarray and Serial Analysis of Gene Expression) from 210 stem cell samples in 60 different experiments. Conclusion StemBase can be used to study gene expression in human and murine stem cells and is available at . PMID:19284540

  13. Use of human pluripotent stem cells to study and treat retinopathies

    PubMed Central

    Ben M’Barek, Karim; Regent, Florian; Monville, Christelle

    2015-01-01

    Human cell types affected by retinal diseases (such as age-related macular degeneration or retinitis pimentosa) are limited in cell number and of reduced accessibility. As a consequence, their isolation for in vitro studies of disease mechanisms or for drug screening efforts is fastidious. Human pluripotent stem cells (hPSCs), either of embryonic origin or through reprogramming of adult somatic cells, represent a new promising way to generate models of human retinopathies, explore the physiopathological mechanisms and develop novel therapeutic strategies. Disease-specific human embryonic stem cells were the first source of material to be used to study certain disease states. The recent demonstration that human somatic cells, such as fibroblasts or blood cells, can be genetically converted to induce pluripotent stem cells together with the continuous improvement of methods to differentiate these cells into disease-affected cellular subtypes opens new perspectives to model and understand a large number of human pathologies, including retinopathies. This review focuses on the added value of hPSCs for the disease modeling of human retinopathies and the study of their molecular pathological mechanisms. We also discuss the recent use of these cells for establishing the validation studies for therapeutic intervention and for the screening of large compound libraries to identify candidate drugs. PMID:25914766

  14. Stiffening of Human Mesenchymal Stem Cell Spheroid Microenvironments Induced by Incorporation of Gelatin Microparticles

    PubMed Central

    Baraniak, Priya R.; Cooke, Marissa T.; Saeed, Rabbia; Kinney, Melissa A.; Fridley, Krista M.; McDevitt, Todd C.

    2012-01-01

    Culturing multipotent adult mesenchymal stem cells as 3D aggregates augments their differentiation potential and paracrine activity. One caveat of stem cell spheroids, though, can be the limited diffusional transport barriers posed by the inherent 3D structure of the multicellular aggregates. In order to circumvent such limitations, polymeric microparticles have been incorporated into stem cell aggregates as a means to locally control the biochemical and physical properties of the 3D microenvironment. However, the introduction of biomaterials to the 3D stem cell microenvironment could alter the mechanical forces sensed by cells within aggregates, which in turn could impact various cell behaviors and overall spheroid mechanics. Therefore, the objective of this study was to determine the acute effects of biomaterial incorporation within mesenchymal stem cell spheroids on aggregate structure and mechanical properties. The results of this study demonstrate that although gelatin microparticle incorporation results in similar multi-cellular organization within human mesenchymal stem cell spheroids, the introduction of gelatin materials significantly impacts spheroid mechanical properties. The marked differences in spheroid mechanics induced by microparticle incorporation may hold major implications for in vitro directed differentiation strategies and offer a novel route to engineer the mechanical properties of tissue constructs ex vivo. PMID:22658155

  15. Regulation of intestinal stem cell proliferation by human methyl-CpG-binding protein-2 in Drosophila.

    PubMed

    Lee, Shin-Hae; Kim, In-Joo; Kim, Joong-Gook; Park, Joung-Sun; Kim, Young-Shin; Yamaguchi, Masamitsu; Kim, Cheol-Min; Yoo, Mi-Ae

    2011-01-01

    Recent studies have suggested the involvement of epigenetic factors such as methyl-CpG-binding protein-2 (MeCP2) in tumorigenesis. In addition, cancer may represent a stem cell-based disease, suggesting that understanding of stem cell regulation could provide valuable insights into the mechanisms of tumorigenesis. However, the function of epigenetic factors in stem cell regulation in adult tissues remains poorly understood. In the present study, we investigated the role of human MeCP2 (hMeCP2), a bridge factor linked to DNA modification and histone modification, in stem cell proliferation using adult Drosophila midgut, which appears to be an excellent model system to study stem cell biology. Results show that enterocyte (EC)-specific expression of hMeCP2 in adult midgut using an exogenous GAL4/UAS expression system induced intestinal stem cell (ISC) proliferation marked by staining with anti-phospho-histone H3 antibody and BrdU incorporation assays. In addition, hMeCP2 expression in ECs activated extracellular stress-response kinase signals in ISCs. Furthermore, expression of hMeCP2 modulated the distribution of heterochromatin protein-1 in ECs. Our data suggests the hypothesis that the expression of hMeCP2 in differentiated ECs stimulates ISC proliferation, implying a role of MeCP2 as a stem cell regulator.

  16. Wildtype adult stem cells, unlike tumor cells, are resistant to cellular damages in Drosophila.

    PubMed

    Ma, Meifang; Zhao, Hang; Zhao, Hanfei; Binari, Richard; Perrimon, Norbert; Li, Zhouhua

    2016-03-15

    Adult stem cells or residential progenitor cells are critical to maintain the structure and function of adult tissues (homeostasis) throughout the lifetime of an individual. Mis-regulation of stem cell proliferation and differentiation often leads to diseases including cancer, however, how wildtype adult stem cells and cancer cells respond to cellular damages remains unclear. We find that in the adult Drosophila midgut, intestinal stem cells (ISCs), unlike tumor intestinal cells, are resistant to various cellular damages. Tumor intestinal cells, unlike wildtype ISCs, are easily eliminated by apoptosis. Further, their proliferation is inhibited upon autophagy induction, and autophagy-mediated tumor inhibition is independent of caspase-dependent apoptosis. Interestingly, inhibition of tumorigenesis by autophagy is likely through the sequestration and degradation of mitochondria, as compromising mitochondria activity in these tumor models mimics the induction of autophagy and increasing the production of mitochondria alleviates the tumor-suppression capacity of autophagy. Together, these data demonstrate that wildtype adult stem cells and tumor cells show dramatic differences in sensitivity to cellular damages, thus providing potential therapeutic implications targeting tumorigenesis. PMID:26845534

  17. A planarian p53 homolog regulates proliferation and self-renewal in adult stem cell lineages

    PubMed Central

    Pearson, Bret J.; Alvarado, Alejandro Sánchez

    2010-01-01

    The functions of adult stem cells and tumor suppressor genes are known to intersect. However, when and how tumor suppressors function in the lineages produced by adult stem cells is unknown. With a large population of stem cells that can be manipulated and studied in vivo, the freshwater planarian is an ideal system with which to investigate these questions. Here, we focus on the tumor suppressor p53, homologs of which have no known role in stem cell biology in any invertebrate examined thus far. Planaria have a single p53 family member, Smed-p53, which is predominantly expressed in newly made stem cell progeny. When Smed-p53 is targeted by RNAi, the stem cell population increases at the expense of progeny, resulting in hyper-proliferation. However, ultimately the stem cell population fails to self-renew. Our results suggest that prior to the vertebrates, an ancestral p53-like molecule already had functions in stem cell proliferation control and self-renewal. PMID:20040488

  18. Monitoring the Bystander Killing Effect of Human Multipotent Stem Cells for Treatment of Malignant Brain Tumors

    PubMed Central

    Leten, Cindy; Trekker, Jesse; Struys, Tom; Roobrouck, Valerie D.; Dresselaers, Tom; Vande Velde, Greetje; Lambrichts, Ivo; Verfaillie, Catherine M.; Himmelreich, Uwe

    2016-01-01

    Tumor infiltrating stem cells have been suggested as a vehicle for the delivery of a suicide gene towards otherwise difficult to treat tumors like glioma. We have used herpes simplex virus thymidine kinase expressing human multipotent adult progenitor cells in two brain tumor models (hU87 and Hs683) in immune-compromised mice. In order to determine the best time point for the administration of the codrug ganciclovir, the stem cell distribution and viability were monitored in vivo using bioluminescence (BLI) and magnetic resonance imaging (MRI). Treatment was assessed by in vivo BLI and MRI of the tumors. We were able to show that suicide gene therapy using HSV-tk expressing stem cells can be followed in vivo by MRI and BLI. This has the advantage that (1) outliers can be detected earlier, (2) GCV treatment can be initiated based on stem cell distribution rather than on empirical time points, and (3) a more thorough follow-up can be provided prior to and after treatment of these animals. In contrast to rodent stem cell and tumor models, treatment success was limited in our model using human cell lines. This was most likely due to the lack of immune components in the immune-compromised rodents. PMID:26880961

  19. Monitoring the Bystander Killing Effect of Human Multipotent Stem Cells for Treatment of Malignant Brain Tumors.

    PubMed

    Leten, Cindy; Trekker, Jesse; Struys, Tom; Roobrouck, Valerie D; Dresselaers, Tom; Vande Velde, Greetje; Lambrichts, Ivo; Verfaillie, Catherine M; Himmelreich, Uwe

    2016-01-01

    Tumor infiltrating stem cells have been suggested as a vehicle for the delivery of a suicide gene towards otherwise difficult to treat tumors like glioma. We have used herpes simplex virus thymidine kinase expressing human multipotent adult progenitor cells in two brain tumor models (hU87 and Hs683) in immune-compromised mice. In order to determine the best time point for the administration of the codrug ganciclovir, the stem cell distribution and viability were monitored in vivo using bioluminescence (BLI) and magnetic resonance imaging (MRI). Treatment was assessed by in vivo BLI and MRI of the tumors. We were able to show that suicide gene therapy using HSV-tk expressing stem cells can be followed in vivo by MRI and BLI. This has the advantage that (1) outliers can be detected earlier, (2) GCV treatment can be initiated based on stem cell distribution rather than on empirical time points, and (3) a more thorough follow-up can be provided prior to and after treatment of these animals. In contrast to rodent stem cell and tumor models, treatment success was limited in our model using human cell lines. This was most likely due to the lack of immune components in the immune-compromised rodents. PMID:26880961

  20. Angiogenic properties of adult human thymus fat.

    PubMed

    Salas, Julián; Montiel, Mercedes; Jiménez, Eugenio; Valenzuela, Miguel; Valderrama, José Francisco; Castillo, Rafael; González, Sergio; El Bekay, Rajaa

    2009-11-01

    The endogenous proangiogenic properties of adipose tissue are well recognized. Although the adult human thymus has long been known to degenerate into fat tissue, it has never been considered as a potential source of angiogenic factors. We have investigated the expression of diverse angiogenic factors, including vascular endothelial growth factor A and B, angiopoietin 1, and tyrosine-protein kinase receptor-2 (an angiopoietin receptor), and then analyzed their physiological role on endothelial cell migration and proliferation, two relevant events in angiogenesis. The detection of the gene and protein expression of the various proteins has been performed by immunohistochemistry, Western blotting, and quantitative real-time polymerase chain reaction. We show, for the first time, that adult thymus fat produces a variety of angiogenic factors and induces the proliferation and migration of human umbilical cord endothelial cells. Based on these findings, we suggest that this fat has a potential angiogenic function that might affect thymic function and ongoing adipogenesis within the thymus.

  1. Development of Scalable Culture Systems for Human Embryonic Stem Cells

    PubMed Central

    Azarin, Samira M.; Palecek, Sean P.

    2009-01-01

    The use of human pluripotent stem cells, including embryonic and induced pluripotent stem cells, in therapeutic applications will require the development of robust, scalable culture technologies for undifferentiated cells. Advances made in large-scale cultures of other mammalian cells will facilitate expansion of undifferentiated human embryonic stem cells (hESCs), but challenges specific to hESCs will also have to be addressed, including development of defined, humanized culture media and substrates, monitoring spontaneous differentiation and heterogeneity in the cultures, and maintaining karyotypic integrity in the cells. This review will describe our current understanding of environmental factors that regulate hESC self-renewal and efforts to provide these cues in various scalable bioreactor culture systems. PMID:20161686

  2. Chimeric animal models in human stem cell biology.

    PubMed

    Glover, Joel C; Boulland, Jean-Luc; Halasi, Gabor; Kasumacic, Nedim

    2009-01-01

    The clinical use of stem cells for regenerative medicine is critically dependent on preclinical studies in animal models. In this review we examine some of the key issues and challenges in the use of animal models to study human stem cell biology-experimental standardization, body size, immunological barriers, cell survival factors, fusion of host and donor cells, and in vivo imaging and tracking. We focus particular attention on the various imaging modalities that can be used to track cells in living animals, comparing their strengths and weaknesses and describing technical developments that are likely to lead to new opportunities for the dynamic assessment of stem cell behavior in vivo. We then provide an overview of some of the most commonly used animal models, their advantages and disadvantages, and examples of their use for xenotypic transplantation of human stem cells, with separate reviews of models involving rodents, ungulates, nonhuman primates, and the chicken embryo. As the use of human somatic, embryonic, and induced pluripotent stem cells increases, so too will the range of applications for these animal models. It is likely that increasingly sophisticated uses of human/animal chimeric models will be developed through advances in genetic manipulation, cell delivery, and in vivo imaging.

  3. Stem Cells: A Renaissance in Human Biology Research.

    PubMed

    Wu, Jun; Izpisua Belmonte, Juan Carlos

    2016-06-16

    The understanding of human biology and how it relates to that of other species represents an ancient quest. Limited access to human material, particularly during early development, has restricted researchers to only scratching the surface of this inherently challenging subject. Recent technological innovations, such as single cell "omics" and human stem cell derivation, have now greatly accelerated our ability to gain insights into uniquely human biology. The opportunities afforded to delve molecularly into scarce material and to model human embryogenesis and pathophysiological processes are leading to new insights of human development and are changing our understanding of disease and choice of therapy options.

  4. Stem Cells: A Renaissance in Human Biology Research.

    PubMed

    Wu, Jun; Izpisua Belmonte, Juan Carlos

    2016-06-16

    The understanding of human biology and how it relates to that of other species represents an ancient quest. Limited access to human material, particularly during early development, has restricted researchers to only scratching the surface of this inherently challenging subject. Recent technological innovations, such as single cell "omics" and human stem cell derivation, have now greatly accelerated our ability to gain insights into uniquely human biology. The opportunities afforded to delve molecularly into scarce material and to model human embryogenesis and pathophysiological processes are leading to new insights of human development and are changing our understanding of disease and choice of therapy options. PMID:27315475

  5. An insulin-like peptide regulates size and adult stem cells in planarians.

    PubMed

    Miller, Claire M; Newmark, Phillip A

    2012-01-01

    Animal growth depends on nutritional intake during development. In many animals, nutritional status is uncoupled from moderation of adult stature after adult size is achieved. However, some long-lived animals continue to regulate adult size and fertility in a nutrition-dependent manner. For example, the regenerating flatworm Schmidtea mediterranea becomes smaller, or degrows, during periods of starvation. These animals provide an opportunity to readily observe adult stem cell population dynamics in response to nutritional cues. We explored the role of insulin signaling in S. mediterranea. We disrupted insulin signaling via RNA interference and showed that animals, despite eating, degrew similarly to starved animals. Utilizing in situ hybridization and immunofluorescence, we assessed cellular changes in proliferative populations including the planarian adult stem cell population (neoblasts) and the germline. Both impaired insulin signaling and nutritional deprivation correlated with decreased neoblast proliferation. Additionally, insulin signaling played a role in supporting spermatogenesis that was distinct from the effects of starvation. In sum, we have demonstrated that insulin signaling is responsible for regulation of adult animal size and tissue homeostasis in an organism with plastic adult size. Importantly, insulin signaling continued to affect stem cell and germline populations in a mature organism. Furthermore, we have shown that adult organisms can differentially regulate specific cell populations as a result of environmental challenges.

  6. Fetal and adult liver stem cells for liver regeneration and tissue engineering.

    PubMed

    Fiegel, H C; Lange, Claudia; Kneser, U; Lambrecht, W; Zander, A R; Rogiers, X; Kluth, D

    2006-01-01

    For the development of innovative cell-based liver directed therapies, e.g. liver tissue engineering, the use of stem cells might be very attractive to overcome the limitation of donor liver tissue. Liver specific differentiation of embryonic, fetal or adult stem cells is currently under investigation. Different types of fetal liver (stem) cells during development were identified, and their advantageous growth potential and bipotential differentiation capacity were shown. However, ethical and legal issues have to be addressed before using fetal cells. Use of adult stem cells is clinically established, e.g. transplantation of hematopoietic stem cells. Other bone marrow derived liver stem cells might be mesenchymal stem cells (MSC). However, the transdifferentiation potential is still in question due to the observation of cellular fusion in several in vivo experiments. In vitro experiments revealed a crucial role of the environment (e.g. growth factors and extracellular matrix) for specific differentiation of stem cells. Co-cultured liver cells also seemed to be important for hepatic gene expression of MSC. For successful liver cell transplantation, a novel approach of tissue engineering by orthotopic transplantation of gel-immobilized cells could be promising, providing optimal environment for the injected cells. Moreover, an orthotopic tissue engineering approach using bipotential stem cells could lead to a repopulation of the recipients liver with healthy liver and biliary cells, thus providing both hepatic functions and biliary excretion. Future studies have to investigate, which stem cell and environmental conditions would be most suitable for the use of stem cells for liver regeneration or tissue engineering approaches.

  7. Detection, characterization, and spontaneous differentiation in vitro of very small embryonic-like putative stem cells in adult mammalian ovary.

    PubMed

    Parte, Seema; Bhartiya, Deepa; Telang, Jyoti; Daithankar, Vinita; Salvi, Vinita; Zaveri, Kusum; Hinduja, Indira

    2011-08-01

    The present study was undertaken to detect, characterize, and study differentiation potential of stem cells in adult rabbit, sheep, monkey, and menopausal human ovarian surface epithelium (OSE). Two distinct populations of putative stem cells (PSCs) of variable size were detected in scraped OSE, one being smaller and other similar in size to the surrounding red blood cells in the scraped OSE. The smaller 1-3 μm very small embryonic-like PSCs were pluripotent in nature with nuclear Oct-4 and cell surface SSEA-4, whereas the bigger 4-7 μm cells with cytoplasmic localization of Oct-4 and minimal expression of SSEA-4 were possibly the tissue committed progenitor stem cells. Pluripotent gene transcripts of Oct-4, Oct-4A, Nanog, Sox-2, TERT, and Stat-3 in human and sheep OSE were detected by reverse transcriptase-polymerase chain reaction. The PSCs underwent spontaneous differentiation into oocyte-like structures, parthenote-like structures, embryoid body-like structures, cells with neuronal-like phenotype, and embryonic stem cell-like colonies, whereas the epithelial cells transformed into mesenchymal phenotype by epithelial-mesenchymal transition in 3 weeks of OSE culture. Germ cell markers like c-Kit, DAZL, GDF-9, VASA, and ZP4 were immuno-localized in oocyte-like structures. In conclusion, as opposed to the existing view of OSE being a bipotent source of oocytes and granulosa cells, mammalian ovaries harbor distinct very small embryonic-like PSCs and tissue committed progenitor stem cells population that have the potential to develop into oocyte-like structures in vitro, whereas mesenchymal fibroblasts appear to form supporting granulosa-like somatic cells. Research at the single-cell level, including complete gene expression profiling, is required to further confirm whether postnatal oogenesis is a conserved phenomenon in adult mammals.

  8. Human embryonic stem cell research: ethical and legal issues.

    PubMed

    Robertson, J A

    2001-01-01

    The use of human embryonic stem cells to replace damaged cells and tissues promises future hope for the treatment of many diseases. However, many countries now face complex ethical and legal questions as a result of the research needed to develop these cell-replacement therapies. The challenge that must be met is how to permit research on human embryonic tissue to occur while maintaining respect for human life generally.

  9. Evaluating alternative stem cell hypotheses for adult corneal epithelial maintenance

    PubMed Central

    West, John D; Dorà, Natalie J; Collinson, J Martin

    2015-01-01

    In this review we evaluate evidence for three different hypotheses that explain how the corneal epithelium is maintained. The limbal epithelial stem cell (LESC) hypothesis is most widely accepted. This proposes that stem cells in the basal layer of the limbal epithelium, at the periphery of the cornea, maintain themselves and also produce transient (or transit) amplifying cells (TACs). TACs then move centripetally to the centre of the cornea in the basal layer of the corneal epithelium and also replenish cells in the overlying suprabasal layers. The LESCs maintain the corneal epithelium during normal homeostasis and become more active to repair significant wounds. Second, the corneal epithelial stem cell (CESC) hypothesis postulates that, during normal homeostasis, stem cells distributed throughout the basal corneal epithelium, maintain the tissue. According to this hypothesis, LESCs are present in the limbus but are only active during wound healing. We also consider a third possibility, that the corneal epithelium is maintained during normal homeostasis by proliferation of basal corneal epithelial cells without any input from stem cells. After reviewing the published evidence, we conclude that the LESC and CESC hypotheses are consistent with more of the evidence than the third hypothesis, so we do not consider this further. The LESC and CESC hypotheses each have difficulty accounting for one main type of evidence so we evaluate the two key lines of evidence that discriminate between them. Finally, we discuss how lineage-tracing experiments have begun to resolve the debate in favour of the LESC hypothesis. Nevertheless, it also seems likely that some basal corneal epithelial cells can act as long-term progenitors if limbal stem cell function is compromised. Thus, this aspect of the CESC hypothesis may have a lasting impact on our understanding of corneal epithelial maintenance, even if it is eventually shown that stem cells are restricted to the limbus as proposed

  10. Enrichment and characterization of human dermal stem/progenitor cells by intracellular granularity.

    PubMed

    Shim, Joong Hyun; Lee, Tae Ryong; Shin, Dong Wook

    2013-04-15

    Adult stem cells from the dermis would be an attractive cell source for therapeutic purposes as well as studying the process of skin aging. Several studies have reported that human dermal stem/progenitor cells (hDSPCs) with multipotent properties exist within the dermis of adult human skin. However, these cells have not been well characterized, because methods for their isolation or enrichment have not yet been optimized. In the present study, we enriched high side scatter (SSC(high))-hDSPCs from normal human dermal fibroblasts using a structural characteristic, intracellular granularity, as a sorting parameter. The SSC(high)-hDSPCs had high in vitro proliferation properties and expressed high levels of SOX2 and S100B, similar to previously identified mouse SOX2+ hair follicle dermal stem cells. The SSC(high)-hDSPCs could differentiate into not only mesodermal cell types, for example, adipocytes, chondrocytes, and osteoblasts, but also neuroectodermal cell types, such as neural cells. In addition, the SSC(high)-hDSPCs exhibited no significant differences in the expression of nestin, vimentin, SNAI2, TWIST1, versican, and CORIN compared with non-hDSPCs. These cells are therefore different from the previously identified multipotent fibroblasts and skin-derived progenitors. In this study, we suggest that hDSPCs can be enriched by using characteristic of their high intracellular granularity, and these SSC(high)-hDSPCs exhibit high in vitro proliferation and differentiation potentials.

  11. Analysis of Intracellular Calcium Signaling in Human Embryonic Stem Cells.

    PubMed

    Péntek, Adrienn; Pászty, Katalin; Apáti, Ágota

    2016-01-01

    Measurement of changes in intracellular calcium concentration is one of the most common and useful tools for studying signal transduction pathways or cellular responses in basic research and drug screening purposes as well. Increasing number of such applications using human pluripotent stem cells and their derivatives requires development of calcium signal measurements for this special cell type. Here we describe a modified protocol for analysis of calcium signaling events in human embryonic stem cells, which can be used for other pluripotent cell types (such as iPSC) or their differentiated offspring as well.

  12. [In vitro strategies for human gametes production from stem cells].

    PubMed

    Tosca, Lucie; Courtot, Anne-Marie; Bennaceur-Griscelli, Annelise; Tachdjian, Gérard

    2011-10-01

    Embryonic stem cells (ESC) are self-renewal and pluripotent cells that are able to differentiate in vitro into several cell types in favourable conditions. Technical protocols for in vitro gametes production have been developed in mice and human species. The functionality of such differentiated cells is not always analysed and an early meiotic arrest is a current observation. These kinds of experimentations have also been tested from human induced pluripotent stem cells (IPSC). However, differentiation ends shortly at the primordial germ cell stage.

  13. Functional Characterization and Expression Profiling of Human Induced Pluripotent Stem Cell- and Embryonic Stem Cell-Derived Endothelial Cells

    PubMed Central

    Li, Zongjin; Hu, Shijun; Ghosh, Zhumur; Han, Zhongchao

    2011-01-01

    With regard to human induced pluripotent stem cells (hiPSCs), in which adult cells are reprogrammed into embryonic-like cells using defined factors, their functional and transcriptional expression pattern during endothelial differentiation has yet to be characterized. In this study, hiPSCs and human embryonic stem cells (hESCs) were differentiated using the embryoid body method, and CD31+ cells were sorted. Fluorescence activated cell sorting analysis of hiPSC-derived endothelial cells (hiPSC-ECs) and hESC-derived endothelial cells (hESC-ECs) demonstrated similar endothelial gene expression patterns. We showed functional vascular formation by hiPSC-ECs in a mouse Matrigel plug model. We compared the gene profiles of hiPSCs, hESCs, hiPSC-ECs, hESC-ECs, and human umbilical vein endothelial cells (HUVECs) using whole genome microarray. Our analysis demonstrates that gene expression variation of hiPSC-ECs and hESC-ECs contributes significantly to biological differences between hiPSC-ECs and hESC-ECs as well as to the “distances” among hiPSCs, hESCs, hiPSC-ECs, hESC-ECs, and HUVECs. We further conclude that hiPSCs can differentiate into functional endothelial cells, but with limited expansion potential compared with hESC-ECs; thus, extensive studies should be performed to explore the cause and extent of such differences before clinical application of hiPSC-ECs can begin. PMID:21235328

  14. Intrinsic Ability of Adult Stem Cell in Skeletal Muscle: An Effective and Replenishable Resource to the Establishment of Pluripotent Stem Cells

    PubMed Central

    Fujimaki, Shin; Machida, Masanao; Hidaka, Ryo; Asashima, Makoto; Takemasa, Tohru; Kuwabara, Tomoko

    2013-01-01

    Adult stem cells play an essential role in mammalian organ maintenance and repair throughout adulthood since they ensure that organs retain their ability to regenerate. The choice of cell fate by adult stem cells for cellular proliferation, self-renewal, and differentiation into multiple lineages is critically important for the homeostasis and biological function of individual organs. Responses of stem cells to stress, injury, or environmental change are precisely regulated by intercellular and intracellular signaling networks, and these molecular events cooperatively define the ability of stem cell throughout life. Skeletal muscle tissue represents an abundant, accessible, and replenishable source of adult stem cells. Skeletal muscle contains myogenic satellite cells and muscle-derived stem cells that retain multipotent differentiation abilities. These stem cell populations have the capacity for long-term proliferation and high self-renewal. The molecular mechanisms associated with deficits in skeletal muscle and stem cell function have been extensively studied. Muscle-derived stem cells are an obvious, readily available cell resource that offers promise for cell-based therapy and various applications in the field of tissue engineering. This review describes the strategies commonly used to identify and functionally characterize adult stem cells, focusing especially on satellite cells, and discusses their potential applications. PMID:23818907

  15. Stem and progenitor cell dysfunction in human trisomies

    PubMed Central

    Liu, Binbin; Filippi, Sarah; Roy, Anindita; Roberts, Irene

    2015-01-01

    Trisomy 21, the commonest constitutional aneuploidy in humans, causes profound perturbation of stem and progenitor cell growth, which is both cell context dependent and developmental stage specific and mediated by complex genetic mechanisms beyond increased Hsa21 gene dosage. While proliferation of fetal hematopoietic and testicular stem/progenitors is increased and may underlie increased susceptibility to childhood leukemia and testicular cancer, fetal stem/progenitor proliferation in other tissues is markedly impaired leading to the characteristic craniofacial, neurocognitive and cardiac features in individuals with Down syndrome. After birth, trisomy 21-mediated premature aging of stem/progenitor cells may contribute to the progressive multi-system deterioration, including development of Alzheimer's disease. PMID:25520324

  16. Erythropoietic differentiation of a human embryonic stem cell line harbouring the sickle cell anaemia mutation

    PubMed Central

    Pryzhkova, Marina V; Peters, Ann; Zambidis, Elias T

    2012-01-01

    Herein is reported efficient erythropoietic differentiation of a human embryonic stem cell (ESC) line derived from a preimplantation genetic diagnosis (PGD)-screened embryo that harbours the homozygous sickle cell disease (SCD) haemoglobinopathy mutation. This human ESC line possesses typical pluripotency characteristics and forms multilineage teratomas in vivo. SCD-human ESC efficiently differentiated to the haematopoietic lineage under serum-free and stromal co-culture conditions and gave rise to robust primitive and definitive erythrocytes. Expression of embryonic, fetal and adult sickle globin genes in SCD PGD-derived human ESC-derived erythrocytes was confirmed by quantitative real-time PCR, intracytoplasmic fluorescence-activated cell sorting and insitu immunostaining of PGD-derived human ESC teratoma sections. These data introduce important methodologies and paradigms for using patient-specific human ESC to generate normal and haemoglobinopathic erythroid progenitors for biomedical research. PMID:20541472

  17. Intra-Operatively Obtained Human Tissue: Protocols and Techniques for the Study of Neural Stem Cells

    PubMed Central

    Chaichana, Kaisorn; Guerrero-Cazares, Hugo; Capilla-Gonzalez, Vivian; Zamora-Berridi, Grettel; Achanta, Praganthi; Gonzalez-Perez, Oscar; Jallo, George I.; Garcia-Verdugo, Jose Manuel; Quiñones-Hinojosa, Alfredo

    2009-01-01

    The discoveries of neural (NSCs) and brain tumor stem cells (BTSCs) in the adult human brain and in brain tumors, respectively, have led to a new era in neuroscience research. These cells represent novel approaches to studying normal phenomena such as memory and learning, as well as pathological conditions such as Parkinson’s disease, stroke, and brain tumors. This new paradigm stresses the importance of understanding how these cells behave in vitro and in vivo. It also stresses the need to use human-derived tissue to study human disease because animal models may not necessarily accurately replicate the processes that occur in humans. An important, but often underused, source of human tissue and, consequently, both NSCs and BTSCs, is the operating room. This study describes in detail both current and newly developed laboratory techniques, which in our experience are used to process and study human NSCs and BTSCs from tissue obtained directly from the operating room. PMID:19427538

  18. Back to the future: how human induced pluripotent stem cells will transform regenerative medicine

    PubMed Central

    Svendsen, Clive N.

    2013-01-01

    Based on cloning studies in mammals, all adult human cells theoretically contain DNA that is capable of creating a whole new person. Cells are maintained in their differentiated state by selectively activating some genes and silencing. The dogma until recently was that cell differentiation was largely fixed unless exposed to the environment of an activated oocyte. However, it is now possible to activate primitive pluripotent genes within adult human cells that take them back in time to a pluripotent state (termed induced pluripotent stem cells). This technology has grown at an exponential rate over the past few years, culminating in the Nobel Prize in medicine. Discussed here are recent developments in the field as they relate to regenerative medicine, with an emphasis on creating functional cells, editing their genome, autologous transplantation and how this ground-breaking field may eventually impact human aging. PMID:23945396

  19. Improved Method for Ex Ovo-Cultivation of Developing Chicken Embryos for Human Stem Cell Xenografts

    PubMed Central

    Schomann, Timo; Qunneis, Firas; Widera, Darius; Kaltschmidt, Christian; Kaltschmidt, Barbara

    2013-01-01

    The characterization of human stem cells for the usability in regenerative medicine is particularly based on investigations regarding their differentiation potential in vivo. In this regard, the chicken embryo model represents an ideal model organism. However, the access to the chicken embryo is only achievable by windowing the eggshell resulting in limited visibility and accessibility in subsequent experiments. On the contrary, ex ovo-culture systems avoid such negative side effects. Here, we present an improved ex ovo-cultivation method enabling the embryos to survive 13 days in vitro. Optimized cultivation of chicken embryos resulted in a normal development regarding their size and weight. Our ex ovo-approach closely resembles the development of chicken embryos in ovo, as demonstrated by properly developed nervous system, bones, and cartilage at expected time points. Finally, we investigated the usability of our method for trans-species transplantation of adult stem cells by injecting human neural crest-derived stem cells into late Hamburger and Hamilton stages (HH26–HH28/E5—E6) of ex ovo-incubated embryos. We demonstrated the integration of human cells allowing experimentally easy investigation of the differentiation potential in the proper developmental context. Taken together, this ex ovo-method supports the prolonged cultivation of properly developing chicken embryos enabling integration studies of xenografted mammalian stem cells at late developmental stages. PMID:23554818

  20. Investigation of genes important in neurodevelopment disorders in adult human brain.

    PubMed

    Maussion, Gilles; Diallo, Alpha B; Gigek, Carolina O; Chen, Elizabeth S; Crapper, Liam; Théroux, Jean-Francois; Chen, Gary G; Vasuta, Cristina; Ernst, Carl

    2015-10-01

    Several neurodevelopmental disorders (NDDs) are caused by mutations in genes expressed in fetal brain, but little is known about these same genes in adult human brain. Here, we test the hypothesis that genes associated with NDDs continue to have a role in adult human brain to explore the idea that NDD symptoms may be partially a result of their adult function rather than just their neurodevelopmental function. To demonstrate adult brain function, we performed expression analyses and ChIPseq in human neural stem cell(NSC) lines at different developmental stages and adult human brain, targeting two genes associated with NDDs, SATB2 and EHMT1, and the WNT signaling gene TCF7L2, which has not been associated with NDDs. Analysis of DNA interaction sites in neural stem cells reveals high (40-50 %) overlap between proliferating and differentiating cells for each gene in temporal space. Studies in adult brain demonstrate that consensus sites are similar to NSCs but occur at different genomic locations. We also performed expression analyses using BrainSpan data for NDD-associated genes SATB2, EHMT1, FMR1, MECP2, MBD5, CTNND2, RAI1, CHD8, GRIN2A, GRIN2B, TCF4, SCN2A, and DYRK1A and find high expression of these genes in adult brain, at least comparable to developing human brain, confirming that genes associated with NDDs likely have a role in adult tissue. Adult function of genes associated with NDDs might be important in clinical disease presentation and may be suitable targets for therapeutic intervention. PMID:26194112

  1. Investigation of genes important in neurodevelopment disorders in adult human brain.

    PubMed

    Maussion, Gilles; Diallo, Alpha B; Gigek, Carolina O; Chen, Elizabeth S; Crapper, Liam; Théroux, Jean-Francois; Chen, Gary G; Vasuta, Cristina; Ernst, Carl

    2015-10-01

    Several neurodevelopmental disorders (NDDs) are caused by mutations in genes expressed in fetal brain, but little is known about these same genes in adult human brain. Here, we test the hypothesis that genes associated with NDDs continue to have a role in adult human brain to explore the idea that NDD symptoms may be partially a result of their adult function rather than just their neurodevelopmental function. To demonstrate adult brain function, we performed expression analyses and ChIPseq in human neural stem cell(NSC) lines at different developmental stages and adult human brain, targeting two genes associated with NDDs, SATB2 and EHMT1, and the WNT signaling gene TCF7L2, which has not been associated with NDDs. Analysis of DNA interaction sites in neural stem cells reveals high (40-50 %) overlap between proliferating and differentiating cells for each gene in temporal space. Studies in adult brain demonstrate that consensus sites are similar to NSCs but occur at different genomic locations. We also performed expression analyses using BrainSpan data for NDD-associated genes SATB2, EHMT1, FMR1, MECP2, MBD5, CTNND2, RAI1, CHD8, GRIN2A, GRIN2B, TCF4, SCN2A, and DYRK1A and find high expression of these genes in adult brain, at least comparable to developing human brain, confirming that genes associated with NDDs likely have a role in adult tissue. Adult function of genes associated with NDDs might be important in clinical disease presentation and may be suitable targets for therapeutic intervention.

  2. Physicochemical Control of Adult Stem Cell Differentiation: Shedding Light on Potential Molecular Mechanisms

    PubMed Central

    Titushkin, Igor; Sun, Shan; Shin, Jennifer; Cho, Michael

    2010-01-01

    Realization of the exciting potential for stem-cell-based biomedical and therapeutic applications, including tissue engineering, requires an understanding of the cell-cell and cell-environment interactions. To this end, recent efforts have been focused on the manipulation of adult stem cell differentiation using inductive soluble factors, designing suitable mechanical environments, and applying noninvasive physical forces. Although each of these different approaches has been successfully applied to regulate stem cell differentiation, it would be of great interest and importance to integrate and optimally combine a few or all of the physicochemical differentiation cues to induce synergistic stem cell differentiation. Furthermore, elucidation of molecular mechanisms that mediate the effects of multiple differentiation cues will enable the researcher to better manipulate stem cell behavior and response. PMID:20379388

  3. Concise Review: Quiescence in Adult Stem Cells: Biological Significance and Relevance to Tissue Regeneration.

    PubMed

    Rumman, Mohammad; Dhawan, Jyotsna; Kassem, Moustapha

    2015-10-01

    Adult stem cells (ASCs) are tissue resident stem cells responsible for tissue homeostasis and regeneration following injury. In uninjured tissues, ASCs exist in a nonproliferating, reversibly cell cycle-arrested state known as quiescence or G0. A key function of the quiescent state is to preserve stemness in ASCs by preventing precocious differentiation, and thus maintaining a pool of undifferentiated ASCs. Recent evidences suggest that quiescence is an actively maintained state and that excessive or defective quiescence may lead to compromised tissue regeneration or tumorigenesis. The aim of this review is to provide an update regarding the biological mechanisms of ASC quiescence and their role in tissue regeneration.

  4. Human dental pulp stem cells: Applications in future regenerative medicine

    PubMed Central

    Potdar, Pravin D; Jethmalani, Yogita D

    2015-01-01

    Stem cells are pluripotent cells, having a property of differentiating into various types of cells of human body. Several studies have developed mesenchymal stem cells (MSCs) from various human tissues, peripheral blood and body fluids. These cells are then characterized by cellular and molecular markers to understand their specific phenotypes. Dental pulp stem cells (DPSCs) are having a MSCs phenotype and they are differentiated into neuron, cardiomyocytes, chondrocytes, osteoblasts, liver cells and β cells of islet of pancreas. Thus, DPSCs have shown great potentiality to use in regenerative medicine for treatment of various human diseases including dental related problems. These cells can also be developed into induced pluripotent stem cells by incorporation of pluripotency markers and use for regenerative therapies of various diseases. The DPSCs are derived from various dental tissues such as human exfoliated deciduous teeth, apical papilla, periodontal ligament and dental follicle tissue. This review will overview the information about isolation, cellular and molecular characterization and differentiation of DPSCs into various types of human cells and thus these cells have important applications in regenerative therapies for various diseases. This review will be most useful for postgraduate dental students as well as scientists working in the field of oral pathology and oral medicine. PMID:26131314

  5. Nuclear Nox4 Role in Stemness Power of Human Amniotic Fluid Stem Cells

    PubMed Central

    Maraldi, Tullia; Guida, Marianna; Zavatti, Manuela; Resca, Elisa; Bertoni, Laura; La Sala, Giovanni B.; De Pol, Anto

    2015-01-01

    Human amniotic fluid stem cells (AFSC) are an attractive source for cell therapy due to their multilineage differentiation potential and accessibility advantages. However the clinical application of human stem cells largely depends on their capacity to expand in vitro, since there is an extensive donor-to-donor heterogeneity. Reactive oxygen species (ROS) and cellular oxidative stress are involved in many physiological and pathophysiological processes of stem cells, including pluripotency, proliferation, differentiation, and stress resistance. The mode of action of ROS is also dependent on the localization of their target molecules. Thus, the modifications induced by ROS can be separated depending on the cellular compartments they affect. NAD(P)H oxidase family, particularly Nox4, has been known to produce ROS in the nucleus. In the present study we show that Nox4 nuclear expression (nNox4) depends on the donor and it correlates with the expression of transcription factors involved in stemness regulation, such as Oct4, SSEA-4, and Sox2. Moreover nNox4 is linked with the nuclear localization of redox sensitive transcription factors, as Nrf2 and NF-κB, and with the differentiation potential. Taken together, these results suggest that nNox4 regulation may have important effects in stem cell capability through modulation of transcription factors and DNA damage. PMID:26273418

  6. [Human amniotic epithelium (HAE) as a possible source of stem cells (SC)].

    PubMed

    García-López, Guadalupe; García-Castro, Irma Lydia; Avila-González, Daniela; Molina-Hernández, Anayansi; Flores-Herrera, Héctor; Merchant-Larios, Horacio; Díaz-Martínez, Fabián

    2015-01-01

    There have been major recent advances in the field of developmental biology due to the investigation on stem cells (SC). Stem cells are characterized by their capacity of auto-renewal and differentiation to different cellular phenotypes. Based on the developmental stage, they can be classified into two different types: embryonic SCs and adult SCs. It has been widely reported that several problems need to be resolved before their possible clinical applications. As a result, fetal membranes have been suggested as an alternative source of SCs. In the human amniotic epithelium, the presence of markers of pluripotent SC´s has been reported, and its capacity as a feeder layer for expansion of different SC types. Also, fetal membranes are a discarded product after delivery, and thus there are not any ethical issues related to its use. In conclusion, the human amniotic epithelium can be a strong candidate for regenerative medicine.

  7. Correlation between proliferative activity and cellular thickness of human mesenchymal stem cells

    SciTech Connect

    Katsube, Yoshihiro; Hirose, Motohiro Nakamura, Chikashi; Ohgushi, Hajime

    2008-04-04

    A cell's shape is known to be related to its proliferative activity. In particular, large and flat mammalian adult stem cells seem to show slow proliferation, however using quantitative analysis to prove the phenomenon is difficult. We measured the proliferation and cellular thickness of human mesenchymal stem cells (MSCs) by atomic force microscopy and found that MSCs with high proliferative activity were thick while those with low proliferative activity were thin, even though these MSCs were early passage cells. Further, low proliferative MSCs contained many senescence-associated {beta}-galactosidase positive cells together with high senescence-associated gene expression. These findings suggest that the measurement of cellular thickness is useful for estimating the proliferative activity of human MSCs and is expected to be a practical tool for MSC applications in regenerative medicine.

  8. Bioethical aspects of basic research and medical applications of human stem cells.

    PubMed

    Santos, Manuel J; Ventura-Juncá, Patricio

    2012-01-01

    The new discoveries, the extraordinary dynamism in human stem cell (SC) research, and the great expectations of the benefits in clinical treatment of many diseases are on the edge of unparalleled advances in both: 1) the understanding of basic mechanisms of cell differentiation and development and 2) the translation from basic research to new clinical therapies. Human stem cells are obtained from different sources, such as embryo, fetal, and adult tissues, in vitro induction (iPS cells) or transdifferentiation. The evidence that these cells are pluripotent (or multipotent), meaning they have the ability to differentiate into all body tissues or tissues of the same lineage, raises the possibility that they could regenerate diseased or damaged tissue in diseases that until now have had no effective treatments. Human stem cell research and therapy raise important bioethical considerations because of the human nature of these cells and their peculiar characteristics. Here we discuss the bioethical aspects of basic human SC research and the conditions necessary for the translation of basic preclinical research into clinical use of SC.

  9. Smooth Muscle Precursor Cells Derived from Human Pluripotent Stem Cells for Treatment of Stress Urinary Incontinence

    PubMed Central

    Wang, Zhe; Li, Yan Hui; Wei, Yi; Green, Morgaine; Wani, Prachi; Zhang, Pengbo; Pera, Renee Reijo; Chen, Bertha

    2016-01-01

    There is great interest in using stem cells (SC) to regenerate a deficient urethral sphincter in patients with urinary incontinence. The smooth muscle component of the sphincter is a significant contributor to sphincter function. However, current translational efforts for sphincter muscle restoration focus only on skeletal muscle regeneration because they rely on adult mesenchymal SC as cell source. These adult SC do not yield sufficient smooth muscle cells (SMCs) for transplantation. We may be able to overcome this limitation by using pluripotent stem cell (PSC) to derive SMCs. Hence, we sought to investigate whether smooth muscle precursor cells (pSMCs) derived from human PSCs can restore urethral function in an animal model generated by surgical urethrolysis and ovariectomy. Rats were divided into four groups: control (no intervention), sham saline (surgery + saline injection), bladder SMC (surgery + human bladder SMC injection), and treatment (surgery + pSMC injection, which includes human embryonic stem cell (hESC) H9-derived pSMC, episomal reprogrammed induced pluripotent stem cells (iPSCs)-derived pSMC, or viral reprogrammed iPSC-derived pSMC). pSMCs (2 × 106 cells/rat) were injected periurethrally 3 weeks postsurgery. Leak point pressure (LPP) and baseline external urethral sphincter electromyography were measured 5 weeks postinjection. Both iPSC-derived pSMC treatment groups showed significantly higher LPP compared to the sham saline group, consistent with restoration of urethral sphincter function. While the difference between the H9-derived pSMC treatment and sham saline group was not significant, it did show a trend toward restoration of the LPP to the level of intact controls. Our data indicate that pSMCs derived from human PSCs (hESC and iPSC) can restore sphincter function. PMID:26785911

  10. Origins of adult pigmentation: diversity in pigment stem cell lineages and implications for pattern evolution

    PubMed Central

    Spiewak, Jessica E.

    2014-01-01

    Summary Teleosts comprise about half of all vertebrate species and exhibit an extraordinary diversity of adult pigment patterns that function in shoaling, camouflage and mate choice and have played important roles in speciation. Here, we review recent studies that have identified several distinct neural crest lineages, with distinct genetic requirements, that give rise to adult pigment cells in fishes. These lineages include post-embryonic, peripheral nerve associated stem cells that generate black melanophores and iridescent iridophores, cells derived directly from embryonic neural crest cells that generate yellow-orange xanthophores, and bipotent stem cells that generate both melanophores and xanthophores. This complexity in adult chromatophore lineages has implications for our understanding of adult traits, melanoma, and the evolutionary diversification of pigment cell lineages and patterns. PMID:25421288

  11. Origins of adult pigmentation: diversity in pigment stem cell lineages and implications for pattern evolution.

    PubMed

    Parichy, David M; Spiewak, Jessica E

    2015-01-01

    Teleosts comprise about half of all vertebrate species and exhibit an extraordinary diversity of adult pigment patterns that function in shoaling, camouflage, and mate choice and have played important roles in speciation. Here, we review studies that have identified several distinct neural crest lineages, with distinct genetic requirements, that give rise to adult pigment cells in fishes. These lineages include post-embryonic, peripheral nerve-associated stem cells that generate black melanophores and iridescent iridophores, cells derived directly from embryonic neural crest cells that generate yellow-orange xanthophores, and bipotent stem cells that generate both melanophores and xanthophores. This complexity in adult chromatophore lineages has implications for our understanding of adult traits, melanoma, and the evolutionary diversification of pigment cell lineages and patterns.

  12. Stem cell programs are retained in human leukemic lymphoblasts.

    PubMed

    Fan, D; Zhou, X; Li, Z; Li, Z-Q; Duan, C; Liu, T; Zhang, F; Huang, Y; Zhang, Y; Gao, F; Guo, Y; Gupta, R; Chen, G; Enver, T; Tang, J; Hong, D

    2015-04-16

    Leukemic lymphoblasts within different immunophenotypic populations possess stem cell properties. However, whether or not the self-renewal program is retained from stem cells or conferred on progenitors by leukemogenic molecules remains unknown. We have addressed the issue in the context of TEL-AML1-associated acute lymphoblastic leukemia (ALL) by profiling a refined program edited from genes essential for self-renewal of hematopoietic stem cells and B-cell development. Bioinformatic analysis shows that ALL populations are loosely clustered and close to the normal population that contains stem and primitive progenitor cells. This finding indicates that immunophenotypes do not reflect maturation stages in ALL and that the self-renewal program may be retained from stem cells. Results of assessing 'first hit' function of TEL-AML1 in different populations of normal cells demonstrate the molecular model. Therefore, the current study shows a leukemogenic scenario of human ALL in which programs of stem cells are sustained in distinct fractions by leukemogenic mutations.

  13. Differentiation of human adipose-derived stem cells into neuron-like cells by Radix Angelicae Sinensis

    PubMed Central

    Wang, Qiaozhi; Zhou, Lile; Guo, Yong; Liu, Guangyi; Cheng, Jiyan; Yu, Hong

    2013-01-01

    Human adipose tissues are an ideal source of stem cells. It is important to find inducers that can safely and effectively differentiate stem cells into functional neurons for clinical use. In this study, we investigate the use of Radix Angelicae Sinensis as an inducer of neuronal differentiation. Primary human adipose-derived stem cells were obtained from adult subcutaneous fatty tissue, then pre-induced with 10% Radix Angelicae Sinensis injection for 24 hours, and incubated in serum-free Dulbecco's modified Eagle's medium/Nutrient Mixture F-12 containing 40% Radix Angelicae Sinensis to induce its differentiation into neuron-like cells. Butylated hydroxyanisole, a common inducer for neuronal differentiation, was used as the control. After human adipose-derived stem cells differentiated into neuron-like cells under the induction of Radix Angelicae Sinensis for 24 hours, the positive expression of neuron-specific enolase was lower than that of the butylated hydroxyanisole-induced group, and the expression of glial fibrillary acidic protein was negative. After they were induced for 48 hours, the positive expression of neuron specific enolase in human adipose-derived stem cells was significantly higher than that of the butylated hydroxyanisole-induced group. Our experimental findings indicate that Radix Angelicae Sinensis can induce human adipose-derived stem cell differentiation into neuron-like cells and produce less cytotoxicity. PMID:25206657

  14. Imagining STEM Higher Education Futures: Advancing Human Well-Being

    ERIC Educational Resources Information Center

    Walker, Melanie

    2015-01-01

    The paper explores a conceptual approach to the question of what it means to provide a university education that addresses equity, and encourages the formation of STEM graduates oriented to public-good values and with commitments to making professional contributions to society which will advance human well-being. It considers and rejects…

  15. Establishment of immortalized human amniotic mesenchymal stem cells.

    PubMed

    Teng, Zan; Yoshida, Toshiko; Okabe, Motonori; Toda, Ayaka; Higuchi, Osamu; Nogami, Makiko; Yoneda, Noriko; Zhou, Kaixuan; Kyo, Satoru; Kiyono, Touru; Nikaido, Toshio

    2013-01-01

    Human amniotic mesenchymal cells (HAM cells) are known to contain somatic stem cells possessing the characteristics of pluripotency. However, little is known about the biology of these somatic cells because isolated HAM cells from amniotic membrane have a limited lifespan. To overcome this problem, we attempted to prolong the lifespan of HAM cells by infecting retrovirus encoding human papillomavirus type16E6 and E7 (HPV16E6E7), bmi-1, and/or human telomerase reverse transcriptase (hTERT) genes and investigated their characteristics as stem cells. We confirmed the immortalization of the four lines of cultured HAM cells for about 1 year. Immortalized human amnion mesenchymal cells (iHAM cells) have continued to proliferate over 200 population doublings (PDs). iHAM cells were positive for CD73, CD90, CD105, and CD44 and negative for CD34, CD14, CD45, and HLA-DR. They expressed stem cell markers such as Oct3/4, Sox2, Nanog, Klf4, SSEA4, c-myc, vimentin, and nestin. They showed adipogenic, osteogenic, and chondrogenic differentiation abilities after induction. These results suggested that immortalized cell lines with characteristics of stem cells can be established. iHAM cells with an extended lifespan can be used to produce good experimental models both in vitro and in vivo.

  16. Expansion of human cord blood hematopoietic stem cells for transplantation.

    PubMed

    Chou, Song; Chu, Pat; Hwang, William; Lodish, Harvey

    2010-10-01

    A recent Science paper reported a purine derivative that expands human cord blood hematopoietic stem cells in culture (Boitano et al., 2010) by antagonizing the aryl hydrocarbon receptor. Major problems need to be overcome before ex vivo HSC expansion can be used clinically.

  17. Derivation of the human embryonic stem cell line RCM1.

    PubMed

    De Sousa, P A; Tye, B J; Sneddon, S; Bruce, K; Dand, P; Russell, G; Collins, D M; Greenshields, A; McDonald, K; Bradburn, H; Gardner, J; Downie, J M; Courtney, A; Brison, D R

    2016-03-01

    The human embryonic stem cell line RCM-1 was derived from a failed to fertilise egg undergoing parthenogenetic stimulation. The cell line shows normal pluripotency marker expression and differentiation to three germ layers in vitro and in vivo. It has a normal 46XX female karyotype and microsatellite PCR identity, HLA and blood group typing data is available. PMID:27346018

  18. Human neural stem cells promote proliferation of endogenous neural stem cells and enhance angiogenesis in ischemic rat brain

    PubMed Central

    Ryu, Sun; Lee, Seung-Hoon; Kim, Seung U.; Yoon, Byung-Woo

    2016-01-01

    Transplantation of human neural stem cells into the dentate gyrus or ventricle of rodents has been reportedly to enhance neurogenesis. In this study, we examined endogenous stem cell proliferation and angiogenesis in the ischemic rat brain after the transplantation of human neural stem cells. Focal cerebral ischemia in the rat brain was induced by middle cerebral artery occlusion. Human neural stem cells were transplanted into the subventricular zone. The behavioral performance of human neural stem cells-treated ischemic rats was significantly improved and cerebral infarct volumes were reduced compared to those in untreated animals. Numerous transplanted human neural stem cells were alive and preferentially localized to the ipsilateral ischemic hemisphere. Furthermore, 5-bromo-2′-deoxyuridine-labeled endogenous neural stem cells were observed in the subventricular zone and hippocampus, where they differentiated into cells immunoreactive for the neural markers doublecortin, neuronal nuclear antigen NeuN, and astrocyte marker glial fibrillary acidic protein in human neural stem cells-treated rats, but not in the untreated ischemic animals. The number of 5-bromo-2′-deoxyuridine-positive ⁄ anti-von Willebrand factor-positive proliferating endothelial cells was higher in the ischemic boundary zone of human neural stem cells-treated rats than in controls. Finally, transplantation of human neural stem cells in the brains of rats with focal cerebral ischemia promoted the proliferation of endogenous neural stem cells and their differentiation into mature neural-like cells, and enhanced angiogenesis. This study provides valuable insights into the effect of human neural stem cell transplantation on focal cerebral ischemia, which can be applied to the development of an effective therapy for stroke. PMID:27073384

  19. Mesenchymal stem cell like (MSCl) cells generated from human embryonic stem cells support pluripotent cell growth

    SciTech Connect

    Varga, Nora; Vereb, Zoltan; Rajnavoelgyi, Eva; Nemet, Katalin; Uher, Ferenc; Sarkadi, Balazs; Apati, Agota

    2011-10-28

    Highlights: Black-Right-Pointing-Pointer MSC like cells were derived from hESC by a simple and reproducible method. Black-Right-Pointing-Pointer Differentiation and immunosuppressive features of MSCl cells were similar to bmMSC. Black-Right-Pointing-Pointer MSCl cells as feeder cells support the undifferentiated growth of hESC. -- Abstract: Mesenchymal stem cell like (MSCl) cells were generated from human embryonic stem cells (hESC) through embryoid body formation, and isolated by adherence to plastic surface. MSCl cell lines could be propagated without changes in morphological or functional characteristics for more than 15 passages. These cells, as well as their fluorescent protein expressing stable derivatives, efficiently supported the growth of undifferentiated human embryonic stem cells as feeder cells. The MSCl cells did not express the embryonic (Oct4, Nanog, ABCG2, PODXL, or SSEA4), or hematopoietic (CD34, CD45, CD14, CD133, HLA-DR) stem cell markers, while were positive for the characteristic cell surface markers of MSCs (CD44, CD73, CD90, CD105). MSCl cells could be differentiated toward osteogenic, chondrogenic or adipogenic directions and exhibited significant inhibition of mitogen-activated lymphocyte proliferation, and thus presented immunosuppressive features. We suggest that cultured MSCl cells can properly model human MSCs and be applied as efficient feeders in hESC cultures.

  20. Nanotopography Promotes Pancreatic Differentiation of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells.

    PubMed

    Kim, Jong Hyun; Kim, Hyung Woo; Cha, Kyoung Je; Han, Jiyou; Jang, Yu Jin; Kim, Dong Sung; Kim, Jong-Hoon

    2016-03-22

    Although previous studies suggest that nanotopographical features influence properties and behaviors of stem cells, only a few studies have attempted to derive clinically useful somatic cells from human pluripotent stem cells using nanopatterned surfaces. In the present study, we report that polystyrene nanopore-patterned surfaces significantly promote the pancreatic differentiation of human embryonic and induced pluripotent stem cells. We compared different diameters of nanopores and showed that 200 nm nanopore-patterned surfaces highly upregulated the expression of PDX1, a critical transcription factor for pancreatic development, leading to an approximately 3-fold increase in the percentage of differentiating PDX1(+) pancreatic progenitors compared with control flat surfaces. Furthermore, in the presence of biochemical factors, 200 nm nanopore-patterned surfaces profoundly enhanced the derivation of pancreatic endocrine cells producing insulin, glucagon, or somatostatin. We also demonstrate that nanopore-patterned surface-induced upregulation of PDX1 is associated with downregulation of TAZ, suggesting the potential role of TAZ in nanopore-patterned surface-mediated mechanotransduction. Our study suggests that appropriate cytokine treatments combined with nanotopographical stimulation could be a powerful tool for deriving a high purity of desired cells from human pluripotent stem cells. PMID:26900863

  1. Nanotopography Promotes Pancreatic Differentiation of Human Embryonic Stem Cells and Induced Pluripotent Stem Cells.

    PubMed

    Kim, Jong Hyun; Kim, Hyung Woo; Cha, Kyoung Je; Han, Jiyou; Jang, Yu Jin; Kim, Dong Sung; Kim, Jong-Hoon

    2016-03-22

    Although previous studies suggest that nanotopographical features influence properties and behaviors of stem cells, only a few studies have attempted to derive clinically useful somatic cells from human pluripotent stem cells using nanopatterned surfaces. In the present study, we report that polystyrene nanopore-patterned surfaces significantly promote the pancreatic differentiation of human embryonic and induced pluripotent stem cells. We compared different diameters of nanopores and showed that 200 nm nanopore-patterned surfaces highly upregulated the expression of PDX1, a critical transcription factor for pancreatic development, leading to an approximately 3-fold increase in the percentage of differentiating PDX1(+) pancreatic progenitors compared with control flat surfaces. Furthermore, in the presence of biochemical factors, 200 nm nanopore-patterned surfaces profoundly enhanced the derivation of pancreatic endocrine cells producing insulin, glucagon, or somatostatin. We also demonstrate that nanopore-patterned surface-induced upregulation of PDX1 is associated with downregulation of TAZ, suggesting the potential role of TAZ in nanopore-patterned surface-mediated mechanotransduction. Our study suggests that appropriate cytokine treatments combined with nanotopographical stimulation could be a powerful tool for deriving a high purity of desired cells from human pluripotent stem cells.

  2. Sequential cancer mutations in cultured human intestinal stem cells.

    PubMed

    Drost, Jarno; van Jaarsveld, Richard H; Ponsioen, Bas; Zimberlin, Cheryl; van Boxtel, Ruben; Buijs, Arjan; Sachs, Norman; Overmeer, René M; Offerhaus, G Johan; Begthel, Harry; Korving, Jeroen; van de Wetering, Marc; Schwank, Gerald; Logtenberg, Meike; Cuppen, Edwin; Snippert, Hugo J; Medema, Jan Paul; Kops, Geert J P L; Clevers, Hans

    2015-05-01

    Crypt stem cells represent the cells of origin for intestinal neoplasia. Both mouse and human intestinal stem cells can be cultured in medium containing the stem-cell-niche factors WNT, R-spondin, epidermal growth factor (EGF) and noggin over long time periods as epithelial organoids that remain genetically and phenotypically stable. Here we utilize CRISPR/Cas9 technology for targeted gene modification of four of the most commonly mutated colorectal cancer genes (APC, P53 (also known as TP53), KRAS and SMAD4) in cultured human intestinal stem cells. Mutant organoids can be selected by removing individual growth factors from the culture medium. Quadruple mutants grow independently of all stem-cell-niche factors and tolerate the presence of the P53 stabilizer nutlin-3. Upon xenotransplantation into mice, quadruple mutants grow as tumours with features of invasive carcinoma. Finally, combined loss of APC and P53 is sufficient for the appearance of extensive aneuploidy, a hallmark of tumour progression.

  3. Human dermal stem cells differentiate into functional epidermal melanocytes.

    PubMed

    Li, Ling; Fukunaga-Kalabis, Mizuho; Yu, Hong; Xu, Xiaowei; Kong, Jun; Lee, John T; Herlyn, Meenhard

    2010-03-15

    Melanocytes sustain a lifelong proliferative potential, but a stem cell reservoir in glabrous skin has not yet been found. Here, we show that multipotent dermal stem cells isolated from human foreskins lacking hair follicles are able to home to the epidermis to differentiate into melanocytes. These dermal stem cells, grown as three-dimensional spheres, displayed a capacity for self-renewal and expressed NGFRp75, nestin and OCT4, but not melanocyte markers. In addition, cells derived from single-cell clones were able to differentiate into multiple lineages including melanocytes. In a three-dimensional skin equivalent model, sphere-forming cells differentiated into HMB45-positive melanocytes, which migrated from the dermis to the epidermis and aligned singly among the basal layer keratinocytes in a similar fashion to pigmented melanocytes isolated from the epidermis. The dermal stem cells were negative for E-cadherin and N-cadherin, whereas they acquired E-cadherin expression and lost NGFRp75 expression upon contact with epidermal keratinocytes. These results demonstrate that stem cells in the dermis of human skin with neural-crest-like characteristics can become mature epidermal melanocytes. This finding could significantly change our understanding of the etiological factors in melanocyte transformation and pigmentation disorders; specifically, that early epigenetic or genetic alterations leading to transformation may take place in the dermis rather than in the epidermis.

  4. A medium hyperglycosylated podocalyxin enables noninvasive and quantitative detection of tumorigenic human pluripotent stem cells

    PubMed Central

    Tateno, Hiroaki; Onuma, Yasuko; Ito, Yuzuru; Hiemori, Keiko; Aiki, Yasuhiko; Shimizu, Madoka; Higuchi, Kumiko; Fukuda, Masakazu; Warashina, Masaki; Honda, Susumu; Asashima, Makoto; Hirabayashi, Jun

    2014-01-01

    While human pluripotent stem cells are attractive sources for cell-replacement therapies, a major concern remains regarding their tumorigenic potential. Thus, safety assessment of human pluripotent stem cell-based products in terms of tumorigenicity is critical. Previously we have identified a pluripotent stem cell-specific lectin probe rBC2LCN recognizing hyperglycosylated podocalyxin as a cell surface ligand. Here we demonstrate that hyperglycosylated podocalyxin is secreted from human pluripotent stem cells into cell culture supernatants. We establish a sandwich assay system, named the GlycoStem test, targeting the soluble hyperglycosylated podocalyxin using rBC2LCN. The GlycoStem test is sufficiently sensitive and quantitative to detect residual human pluripotent stem cells. This work provides a proof of concept for the noninvasive and quantitative detection of tumorigenic human pluripotent stem cells using cell culture supernatants. The developed method should increase the safety of human pluripotent stem cell-based cell therapies. PMID:24518842

  5. The current state of research with human pluripotent stem cells in Brazil.

    PubMed

    Pranke, Patricia; Chagastelles, Pedro; Sperling, Laura E

    2014-12-01

    This article provides a brief overview of research with human pluripotent stem cells in Brazil, the federal funding supporting this research, and the legislation that allows the isolation of human embryonic stem cells.

  6. Human Pluripotent Stem Cells: Applications and Challenges in Neurological Diseases

    PubMed Central

    Hibaoui, Youssef; Feki, Anis

    2012-01-01

    The ability to generate human pluripotent stem cells (hPSCs) holds great promise for the understanding and the treatment of human neurological diseases in modern medicine. The hPSCs are considered for their in vitro use as research tools to provide relevant cellular model for human diseases, drug discovery, and toxicity assays and for their in vivo use in regenerative medicine applications. In this review, we highlight recent progress, promises, and challenges of hPSC applications in human neurological disease modeling and therapies. PMID:22934023

  7. CHD7 maintains neural stem cell quiescence and prevents premature stem cell depletion in the adult hippocampus.

    PubMed

    Jones, Kieran M; Sarić, Nemanja; Russell, John P; Andoniadou, Cynthia L; Scambler, Peter J; Basson, M Albert

    2015-01-01

    Neural stem/progenitor cells (NSCs) in the hippocampus produce new neurons throughout adult life. NSCs are maintained in a state of reversible quiescence and the failure to maintain the quiescent state can result in the premature depletion of the stem cell pool. The epigenetic mechanisms that maintain this quiescent state have not been identified. Using an inducible knockout mouse model, we show that the chromatin remodeling factor chromodomain-helicase-DNA-binding protein 7 (CHD7) is essential for maintaining NSC quiescence. CHD7 inactivation in adult NSCs results in a loss of stem cell quiescence in the hippocampus, a transient increase in cell divisions, followed by a significant decline in neurogenesis. This loss of NSC quiescence is associated with the premature loss of NSCs in middle-aged mice. We find that CHD7 represses the transcription of several positive regulators of cell cycle progression and is required for full induction of the Notch target gene Hes5 in quiescent NSCs. These findings directly link CHD7 to pathways involved in NSC quiescence and identify the first chromatin-remodeling factor with a role in NSC quiescence and maintenance. As CHD7 haplo-insufficiency is associated with a range of cognitive disabilities in CHARGE syndrome, our observations may have implications for understanding the basis of these deficits.

  8. HOX and TALE signatures specify human stromal stem cell populations from different sources.

    PubMed

    Picchi, Jacopo; Trombi, Luisa; Spugnesi, Laura; Barachini, Serena; Maroni, Giorgia; Brodano, Giovanni Barbanti; Boriani, Stefano; Valtieri, Mauro; Petrini, Mario; Magli, Maria Cristina

    2013-04-01

    Human stromal stem cell populations reside in different tissues and anatomical sites, however a critical question related to their efficient use in regenerative medicine is whether they exhibit equivalent biological properties. Here, we compared cellular and molecular characteristics of stromal stem cells derived from the bone marrow, at different body sites (iliac crest, sternum, and vertebrae) and other tissues (dental pulp and colon). In particular, we investigated whether homeobox genes of the HOX and TALE subfamilies might provide suitable markers to identify distinct stromal cell populations, as HOX proteins control cell positional identity and, together with their co-factors TALE, are involved in orchestrating differentiation of adult tissues. Our results show that stromal populations from different sources, although immunophenotypically similar, display distinct HOX and TALE signatures, as well as different growth and differentiation abilities. Stromal stem cells from different tissues are characterized by specific HOX profiles, differing in the number and type of active genes, as well as in their level of expression. Conversely, bone marrow-derived cell populations can be essentially distinguished for the expression levels of specific HOX members, strongly suggesting that quantitative differences in HOX activity may be crucial. Taken together, our data indicate that the HOX and TALE profiles provide positional, embryological and hierarchical identity of human stromal stem cells. Furthermore, our data suggest that cell populations derived from different body sites may not represent equivalent cell sources for cell-based therapeutical strategies for regeneration and repair of specific tissues.

  9. Decreasing matrix modulus of PEG hydrogels induces a vascular phenotype in human cord blood stem cells

    PubMed Central

    Mahadevaiah, Shruthi; Robinson, Karyn G.; Kharkar, Prathamesh M.; Kiick, Kristi L.; Akins, Robert E.

    2015-01-01

    Adult and congenital cardiovascular diseases are significant health problems that are often managed using surgery. Bypass grafting is a principal therapy, but grafts fail at high rates due to hyperplasia, fibrosis, and atherosclerosis. Biocompatible, cellularized materials that attenuate these complications and encourage healthy microvascularization could reduce graft failure, but an improved understanding of biomaterial effects on human stem cells is needed to reach clinical utility. Our group investigates stem-cell-loaded biomaterials for placement along the adventitia of at-risk vessels and grafts. Here, the effects of substrate modulus on human CD34+ stem cells from umbilical cord blood were evaluated. Cells were isolated by immunomagnetic separation and encapsulated in 3, 4, and 6 weight% PEG hydrogels containing 0.032% gelatin and 0.0044% fibronectin. Gels reached moduli of 0.34, 4.5, and 9.1 kPa. Cell viability approached 100%. Cell morphologies appeared similar across gels, but proliferation was significantly lower in 6 wt% gels. Expression profiling using stem cell signaling arrays indicated enhanced self-renewal and differentiation into vascular endothelium among cells in the lower weight percent gels. Thus, modulus was associated with cell proliferation and function. Gels with moduli in the low kilopascal range may be useful in stimulating cell engraftment and microvascularization of graft adventitia. PMID:26016692

  10. Neural stem cells in the adult ciliary epithelium express GFAP and are regulated by Wnt signaling

    SciTech Connect

    Das, Ani V.; Zhao Xing; James, Jackson; Kim, Min; Cowan, Kenneth H.; Ahmad, Iqbal . E-mail: iahmad@unmc.edu

    2006-01-13

    The identification of neural stem cells with retinal potential in the ciliary epithelium (CE) of the adult mammals is of considerable interest because of their potential for replacing or rescuing degenerating retinal neurons in disease or injury. The evaluation of such a potential requires characterization of these cells with regard to their phenotypic properties, potential, and regulatory mechanisms. Here, we demonstrate that rat CE stem cells/progenitors in neurosphere culture display astrocytic nature in terms of expressing glial intermediate neurofilament protein, GFAP. The GFAP-expressing CE stem cells/progenitors form neurospheres in proliferating conditions and generate neurons when shifted to differentiating conditions. These cells express components of the canonical Wnt pathway and its activation promotes their proliferation. Furthermore, we demonstrate that the activation of the canonical Wnt pathway influences neuronal differentiation of CE stem cells/progenitors in a context dependent manner. Our observations suggest that CE stem cells/progenitors share phenotypic properties and regulatory mechanism(s) with neural stem cells elsewhere in the adult CNS.

  11. Genomic selection for quantitative adult plant stem rust resistance in wheat

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Quantitative adult plant resistance (APR) to stem rust (Puccinia graminis f. sp. tritici) is an important breeding target in wheat (Triticum aestivum L.) and a potential target for genomic selection (GS). To evaluate the relative importance of known APR loci in applying genomic selection, we charact...

  12. Oversight for clinical uses of autologous adult stem cells: lessons from international regulations.

    PubMed

    Lysaght, Tamra; Kerridge, Ian; Sipp, Douglas; Porter, Gerard; Capps, Benjamin J

    2013-12-01

    Autologous adult stem cells (ASCs) are being administered by physicians for indications that have not been demonstrated as safe and effective in formal clinical trials. Examination of regulatory frameworks across five countries suggests that balancing the demands of research with clinical freedom has created structural weaknesses that are being exploited.

  13. Thiamin requirement of the adult human.

    PubMed

    Sauberlich, H E; Herman, Y F; Stevens, C O; Herman, R H

    1979-11-01

    Young adult male subjects maintained on a metabolic ward were fed diets providing controlled intakes of thiamin and either 2800 or 3600 kcal. The higher level of calories was attained by an increased intake of carbohydrates. Constant weights were maintained by the subjects by adjusting daily activity and exercise schedules. Thiamin requirements were evaluated in terms of erythrocyte transketolase activity and urinary excretion of the vitamin. The results of the study revealed that a relationship exists between thiamin requirement and caloric intake and expenditure. Thus, when the calories being utilized were derived primarily from carbohydrate sources, the minimum adult male requirement for thiamin appeared to be 0.30 mg of thiamin per 1000 kcal. Urinary excretion of thiamin and erythrocyte transketolase activity appear to be reasonably reliable reflections of thiamin intakes and thiamin nutritional status. The use of these measurements in nutrition surveys appears justified. The microbiological assay (Lactobacillus viridescens) for measuring thiamin levels in urine samples appears to be a somewhat more sensitive but valid procedure as an alternate for the thiochrome method. Judged from the results of this study, the recommended intake for the adult human of 0.40 mg of thiamin per 1000 kcal by FAO/WHO and the recommended allowance of 0.5 mg per 1000 kcal by the Food and Nutrition Board of the NAS-NRC appear reasonable and amply allow for biological variations and other factors that may influence the requirement for this vitamin.

  14. Modelling human disease with pluripotent stem cells

    PubMed Central

    Park, In-Hyun; Sullivan, Gareth J.

    2013-01-01

    Recent progress in the field of cellular reprogramming has opened up the doors to a new era of disease modelling, as pluripotent stem cells representing a myriad of genetic diseases can now be produced from patient tissue. These cells can be expanded and differentiated to produce a potentially limitless supply of the affected cell type, which can then be used as a tool to improve understanding of disease mechanisms and test therapeutic interventions. This process requires high levels of scrutiny and validation at every stage, but international standards for the characterisation of pluripotent cells and their progeny have yet to be established. Here we discuss the current state of the art with regard to modelling diseases affecting the ectodermal, mesodermal and endodermal lineages, focussing on studies which have demonstrated a disease phenotype in the tissue of interest. We also discuss the utility of pluripotent cell technology for the modelling of cancer and infectious disease. Finally, we spell out the technical and scientific challenges which must be addressed if the field is to deliver on its potential and produce improved patient outcomes in the clinic. PMID:23444871

  15. Role of astrocytes as neural stem cells in the adult brain

    PubMed Central

    Gonzalez-Perez, Oscar; Quiñones-Hinojosa, Alfredo

    2012-01-01

    In the adult mammalian brain, bona fide neural stem cells were discovered in the subventricular zone (SVZ), the largest neurogenic niche lining the striatal wall of the lateral ventricles of the brain. In this region resides a subpopulation of astrocytes that express the glial fibrillary acidic protein (GFAP), nestin and LeX. Astonishingly, these GFAP-expressing progenitors display stem-cell-like features both in vivo and in vitro. Throughout life SVZ astrocytes give rise to interneurons and oligodendrocyte precursors, which populate the olfactory bulb and the white matter, respectively. The role of the progenies of SVZ astrocytes has not been fully elucidated, but some evidence indicates that the new neurons play a role in olfactory discrimination, whereas oligodendrocytes contribute to myelinate white matter tracts. In this chapter, we describe the astrocytic nature of adult neural stem cells, their organization into the SVZ and some of their molecular and genetic characteristics. PMID:23619383

  16. Stem cell research in Germany: ethics of healing vs. human dignity.

    PubMed

    Oduncu, Fuat S

    2003-01-01

    On 25 April 2002, the German Parliament has passed a strict new law referring to stem cell research. This law took effect on July 1, 2002. The so-called embryonic Stem Cell Act ("Stammzellgesetz-StZG") permits the import of embryonic stem (ES) cells isolated from surplus lvF-embryos for research reasons. The production itself of ES cells from human blastocysts has been prohibited by the German Embryo Protection Act of 1990, with the exception of the use of ES cells which exist already. The debate on the legitimate use of ES cells escalated, after the main German research funding agency, the Deutsche Forschungsgemeinschaft (DFG), unexpectedly published new guidelines recommending a restricted use of human ES cells for research. Meanwhile, the debate has ethically divided society, political parties, government and church members into a group supporting and a group rejecting ES cell research. The arguments in favour of such a research can be summarized as arguments derived from a new "ethics of healing" calling for a therapeutic imperative, whereas the arguments against can be summarized as arguments violating the fundamental principle of human dignity as they imply the destruction of human embryos. This article will try to present and evaluate various ethical arguments founded on the latest biological and medical data on the potential use of stem cell technologies. It will finally come to the conclusion that ES cell research is opposed to human dignity, since the procedures of isolating ES cells require the destruction and instrumentalization of human embryos. Human embryos are human beings at a very early stage of their development, fully possessing the ability of completing their development. At this very early stage, human embryos are extremely dependent and fragile, and thus vulnerable corporealities. Vulnerability and human dignity demand the protection of the embryo's corporeal integrity. Hence, this essay will try to propagate research with adult stem (AS

  17. Generating trunk neural crest from human pluripotent stem cells

    PubMed Central

    Huang, Miller; Miller, Matthew L.; McHenry, Lauren K.; Zheng, Tina; Zhen, Qiqi; Ilkhanizadeh, Shirin; Conklin, Bruce R.; Bronner, Marianne E.; Weiss, William A.

    2016-01-01

    Neural crest cells (NCC) are stem cells that generate different lineages, including neuroendocrine, melanocytic, cartilage, and bone. The differentiation potential of NCC varies according to the level from which cells emerge along the neural tube. For example, only anterior “cranial” NCC form craniofacial bone, whereas solely posterior “trunk” NCC contribute to sympathoadrenal cells. Importantly, the isolation of human fetal NCC carries ethical and scientific challenges, as NCC induction typically occur before pregnancy is detectable. As a result, current knowledge of NCC biology derives primarily from non-human organisms. Important differences between human and non-human NCC, such as expression of HNK1 in human but not mouse NCC, suggest a need to study human NCC directly. Here, we demonstrate that current protocols to differentiate human pluripotent stem cells (PSC) to NCC are biased toward cranial NCC. Addition of retinoic acid drove trunk-related markers and HOX genes characteristic of a posterior identity. Subsequent treatment with bone morphogenetic proteins (BMPs) enhanced differentiation to sympathoadrenal cells. Our approach provides methodology for detailed studies of human NCC, and clarifies roles for retinoids and BMPs in the differentiation of human PSC to trunk NCC and to sympathoadrenal lineages. PMID:26812940

  18. In Vivo Targeting of Adult Neural Stem Cells in the Dentate Gyrus by a Split-Cre Approach

    PubMed Central

    Beckervordersandforth, Ruth; Deshpande, Aditi; Schäffner, Iris; Huttner, Hagen B.; Lepier, Alexandra; Lie, Dieter Chichung; Götz, Magdalena

    2014-01-01

    Summary We describe the labeling of adult neural stem cells (aNSCs) in the mouse and human dentate gyrus (DG) by the combinatorial expression of glial fibrillary acidic protein (GFAP) and Prominin1, as revealed by immunohistochemistry. Split-Cre-based genetic fate mapping of these double-positive cells in the adult murine DG reveals their NSC identity, as they are self-renewing and contribute to neurogenesis over several months. Their progeny reacts to stimuli such as voluntary exercise with increased neurogenesis. Prominin1+/GFAP+ cells also exist in the adult human DG, the only region in the human brain for which adult neurogenesis has been consistently reported. Our data, together with previous evidence of such double-positive NSCs in the developing murine brain and in neurogenic regions of vertebrates with widespread neurogenesis, suggest that Prominin1- and GFAP-expressing cells are NSCs in a wide range of species in development and adulthood. PMID:24527389

  19. Human induced pluripotent stem cells: A disruptive innovation.

    PubMed

    De Vos, J; Bouckenheimer, J; Sansac, C; Lemaître, J-M; Assou, S

    2016-01-01

    This year (2016) will mark the 10th anniversary of the discovery of induced pluripotent stem cells (iPSCs). The finding that the transient expression of four transcription factors can radically remodel the epigenome, transcriptome and metabolome of differentiated cells and reprogram them into pluripotent stem cells has been a major and groundbreaking technological innovation. In this review, we discuss the major applications of this technology that we have grouped in nine categories: a model to study cell fate control; a model to study pluripotency; a model to study human development; a model to study human tissue and organ physiology; a model to study genetic diseases in a dish; a tool for cell rejuvenation; a source of cells for drug screening; a source of cells for regenerative medicine; a tool for the production of human organs in animals.

  20. Human cloning, stem cell research. An Islamic perspective.

    PubMed

    Al-Aqeel, Aida I

    2009-12-01

    The rapidly changing technologies that involve human subjects raise complex ethical, legal, social, and religious issues. Recent advances in the field of cloning and stem cell research have introduced new hopes for the treatment of serious diseases. But this promise has raised many complex questions. This field causes debate and challenge, not only among scientists but also among ethicists, religious scholars, governments, and politicians. There is no consensus on the morality of human cloning, even within specific religious traditions. In countries in which religion has a strong influence on political decision making, the moral status of the human embryo is at the center of the debate. Because of the inevitable consequences of reproductive cloning, it is prohibited in Islam. However, stem cell research for therapeutic purposes is permissible with full consideration, and all possible precautions in the pre-ensoulment stages of early fetus development, if the source is legitimate.

  1. Human dental mesenchymal stem cells and neural regeneration.

    PubMed

    Xiao, Li; Tsutsui, Takeki

    2013-09-01

    Nerve tissue presents inherent difficulties for its effective regeneration. Stem cell transplantation is considered an auspicious treatment for neuronal injuries. Recently, human dental mesenchymal stem cells (DMSCs) have received extensive attention in the field of regenerative medicine due to their accessibility and multipotency. Since their origin is within the neural crest, they can be differentiated into neural crest-derived cells including neuron and glia cells both in vitro and in vivo. DMSCs are also able to secrete a wide variety of neurotrophins and chemokines, which promote neuronal cells to survival and differentiation. Experimental evidence has shown that human DMSCs engraftment recovered neuronal tissue damage in animal models of central nervous system injuries. Human DMSCs can be a new hope for treatment of nervous system diseases and deficits such as spinal cord injury, stroke and Parkinson's disease.

  2. Stroke Increases Neural Stem Cells and Angiogenesis in the Neurogenic Niche of the Adult Mouse

    PubMed Central

    Zhang, Rui Lan; Chopp, Michael; Roberts, Cynthia; Liu, Xianshuang; Wei, Min; Nejad-Davarani, Siamak P.; Wang, Xinli; Zhang, Zheng Gang

    2014-01-01

    The unique cellular and vascular architecture of the adult ventricular-subventricular zone (V/SVZ) neurogenic niche plays an important role in regulating neural stem cell function. However, the in vivo identification of neural stem cells and their relationship to blood vessels within this niche in response to stroke remain largely unknown. Using whole-mount preparation of the lateral ventricle wall, we examined the architecture of neural stem cells and blood vessels in the V/SVZ of adult mouse over the course of 3 months after onset of focal cerebral ischemia. Stroke substantially increased the number of glial fibrillary acidic protein (GFAP) positive neural stem cells that are in contact with the cerebrospinal fluid (CSF) via their apical processes at the center of pinwheel structures formed by ependymal cells residing in the lateral ventricle. Long basal processes of these cells extended to blood vessels beneath the ependymal layer. Moreover, stroke increased V/SVZ endothelial cell proliferation from 2% in non-ischemic mice to 12 and 15% at 7 and 14 days after stroke, respectively. Vascular volume in the V/SVZ was augmented from 3% of the total volume prior to stroke to 6% at 90 days after stroke. Stroke-increased angiogenesis was closely associated with neuroblasts that expanded to nearly encompass the entire lateral ventricular wall in the V/SVZ. These data indicate that stroke induces long-term alterations of the neural stem cell and vascular architecture of the adult V/SVZ neurogenic niche. These post-stroke structural changes may provide insight into neural stem cell mediation of stroke-induced neurogenesis through the interaction of neural stem cells with proteins in the CSF and their sub-ependymal neurovascular interaction. PMID:25437857

  3. Hurdles to clinical translation of human induced pluripotent stem cells

    PubMed Central

    Neofytou, Evgenios; O’Brien, Connor Galen; Couture, Larry A.; Wu, Joseph C.

    2015-01-01

    Human pluripotent stem cells are known to have the capacity to renew indefinitely, being intrinsically able to differentiate into many different cell types. These characteristics have generated tremendous enthusiasm about the potential applications of these cells in regenerative medicine. However, major challenges remain with the development and testing of novel experimental stem cell therapeutics in the field. In this Review, we focus on the nature of the preclinical challenges and discuss potential solutions that could help overcome them. Furthermore, we discuss the use of allogeneic versus autologous stem cell products, including a review of their respective advantages and disadvantages, major clinical requirements, quality standards, time lines, and costs of clinical grade development. PMID:26132109

  4. Measurement of human embryonic stem cell in the growing cycle

    NASA Astrophysics Data System (ADS)

    Li, X.; Zhao, L.; Oh, Steve K. W.; Chong, W. K.; Ong, J. K.; Chen, Allen K.; Choo, Andre B. H.

    2008-09-01

    A measurement and imaging system has been developed for in-line continuous measurement of live, unmodified, human embryonic stem cells (hESC). The measurement will not affect cell growth, structure, sterility and suitability for clinical use. The stem cell imaging system (SCIS) can be used to support the optimization of automated stem cell growth for invitro study and for high-volume bio-manufacture. This paper present the experimental and analysis for the optimization of system parameters. A non-linear lighting is developed to obtain a clear images. The individual cluster can be traced from day one to day two. The whole system is calibrated with measurement microscope and haemacytometer. The measurement accuracy is better than 90%.

  5. Biobanking human embryonic stem cell lines: policy, ethics and efficiency.

    PubMed

    Holm, Søren

    2015-12-01

    Stem cell banks curating and distributing human embryonic stem cells have been established in a number of countries and by a number of private institutions. This paper identifies and critically discusses a number of arguments that are used to justify the importance of such banks in policy discussions relating to their establishment or maintenance. It is argued (1) that 'ethical arguments' are often more important in the establishment phase and 'efficiency arguments' more important in the maintenance phase, and (2) that arguments relating to the interests of embryo and gamete donors are curiously absent from the particular stem cell banking policy discourse. This to some extent artificially isolates this discourse from the broader discussions about the flows of reproductive materials and tissues in modern society, and such isolation may lead to the interests of important actors being ignored in the policy making process.

  6. The safety of human pluripotent stem cells in clinical treatment.

    PubMed

    Simonson, Oscar E; Domogatskaya, Anna; Volchkov, Pavel; Rodin, Sergey

    2015-01-01

    Human pluripotent stem cells (hPSCs) have practically unlimited proliferation potential and a capability to differentiate into any cell type in the human body. Since the first derivation in 1998, they have been an attractive source of cells for regenerative medicine. Numerous ethical, technological, and regulatory complications have been hampering hPSC use in clinical applications. Human embryonic stem cells (ESCs), parthenogenetic human ESCs, human nuclear transfer ESCs, and induced pluripotent stem cells are four types of hPSCs that are different in many clinically relevant features such as propensity to epigenetic abnormalities, generation methods, and ability for development of autologous cell lines. Propensity to genetic mutations and tumorigenicity are common features of all pluripotent cells that complicate hPSC-based therapies. Several recent advances in methods of derivation, culturing, and monitoring of hPSCs have addressed many ethical concerns and technological challenges in development of clinical-grade hPSC lines. Generation of banks of such lines may be useful to minimize immune rejection of hPSC-derived allografts. In this review, we discuss different sources of hPSCs available at the moment, various safety risks associated with them, and possible solutions for successful use of hPSCs in the clinic. We also discuss ongoing clinical trials of hPSC-based treatments.

  7. The safety of human pluripotent stem cells in clinical treatment.

    PubMed

    Simonson, Oscar E; Domogatskaya, Anna; Volchkov, Pavel; Rodin, Sergey

    2015-01-01

    Human pluripotent stem cells (hPSCs) have practically unlimited proliferation potential and a capability to differentiate into any cell type in the human body. Since the first derivation in 1998, they have been an attractive source of cells for regenerative medicine. Numerous ethical, technological, and regulatory complications have been hampering hPSC use in clinical applications. Human embryonic stem cells (ESCs), parthenogenetic human ESCs, human nuclear transfer ESCs, and induced pluripotent stem cells are four types of hPSCs that are different in many clinically relevant features such as propensity to epigenetic abnormalities, generation methods, and ability for development of autologous cell lines. Propensity to genetic mutations and tumorigenicity are common features of all pluripotent cells that complicate hPSC-based therapies. Several recent advances in methods of derivation, culturing, and monitoring of hPSCs have addressed many ethical concerns and technological challenges in development of clinical-grade hPSC lines. Generation of banks of such lines may be useful to minimize immune rejection of hPSC-derived allografts. In this review, we discuss different sources of hPSCs available at the moment, various safety risks associated with them, and possible solutions for successful use of hPSCs in the clinic. We also discuss ongoing clinical trials of hPSC-based treatments. PMID:26140342

  8. Planarian MBD2/3 is required for adult stem cell pluripotency independently of DNA methylation.

    PubMed

    Jaber-Hijazi, Farah; Lo, Priscilla J K P; Mihaylova, Yuliana; Foster, Jeremy M; Benner, Jack S; Tejada Romero, Belen; Chen, Chen; Malla, Sunir; Solana, Jordi; Ruzov, Alexey; Aziz Aboobaker, A

    2013-12-01

    Planarian adult stem cells (pASCs) or neoblasts represent an ideal system to study the evolution of stem cells and pluripotency as they underpin an unrivaled capacity for regeneration. We wish to understand the control of differentiation and pluripotency in pASCs and to understand how conserved, convergent or divergent these mechanisms are across the Bilateria. Here we show the planarian methyl-CpG Binding Domain 2/3 (mbd2/3) gene is required for pASC differentiation during regeneration and tissue homeostasis. The genome does not have detectable levels of 5-methylcytosine (5(m)C) and we find no role for a potential DNA methylase. We conclude that MBD proteins may have had an ancient role in broadly controlling animal stem cell pluripotency, but that DNA methylation is not involved in planarian stem cell differentiation.

  9. Planarian MBD2/3 is required for adult stem cell pluripotency independently of DNA methylation☆

    PubMed Central

    Jaber-Hijazi, Farah; Lo, Priscilla J.K.P.; Mihaylova, Yuliana; Foster, Jeremy M.; Benner, Jack S.; Tejada Romero, Belen; Chen, Chen; Malla, Sunir; Solana, Jordi; Ruzov, Alexey; Aziz Aboobaker, A.

    2013-01-01

    Planarian adult stem cells (pASCs) or neoblasts represent an ideal system to study the evolution of stem cells and pluripotency as they underpin an unrivaled capacity for regeneration. We wish to understand the control of differentiation and pluripotency in pASCs and to understand how conserved, convergent or divergent these mechanisms are across the Bilateria. Here we show the planarian methyl-CpG Binding Domain 2/3 (mbd2/3) gene is required for pASC differentiation during regeneration and tissue homeostasis. The genome does not have detectable levels of 5-methylcytosine (5mC) and we find no role for a potential DNA methylase. We conclude that MBD proteins may have had an ancient role in broadly controlling animal stem cell pluripotency, but that DNA methylation is not involved in planarian stem cell differentiation. PMID:24063805

  10. The novel steroidal alkaloids dendrogenin A and B promote proliferation of adult neural stem cells

    SciTech Connect

    Khalifa, Shaden A.M.; Medina, Philippe de; Erlandsson, Anna; El-Seedi, Hesham R.; Silvente-Poirot, Sandrine; Poirot, Marc

    2014-04-11

    Highlights: • Dendrogenin A and B are new aminoalkyl oxysterols. • Dendrogenins stimulated neural stem cells proliferation. • Dendrogenins induce neuronal outgrowth from neurospheres. • Dendrogenins provide new therapeutic options for neurodegenerative disorders. - Abstract: Dendrogenin A (DDA) and dendrogenin B (DDB) are new aminoalkyl oxysterols which display re-differentiation of tumor cells of neuronal origin at nanomolar concentrations. We analyzed the influence of dendrogenins on adult mice neural stem cell proliferation, sphere formation and differentiation. DDA and DDB were found to have potent proliferative effects in neural stem cells. Additionally, they induce neuronal outgrowth from neurospheres during in vitro cultivation. Taken together, our results demonstrate a novel role for dendrogenins A and B in neural stem cell proliferation and differentiation which further increases their likely importance to compensate for neuronal cell loss in the brain.

  11. Large-scale live imaging of adult neural stem cells in their endogenous niche

    PubMed Central

    Dray, Nicolas; Bedu, Sébastien; Vuillemin, Nelly; Alunni, Alessandro; Coolen, Marion; Krecsmarik, Monika; Supatto, Willy; Beaurepaire, Emmanuel; Bally-Cuif, Laure

    2015-01-01

    Live imaging of adult neural stem cells (aNSCs) in vivo is a technical challenge in the vertebrate brain. Here, we achieve long-term imaging of the adult zebrafish telencephalic neurogenic niche and track a population of >1000 aNSCs over weeks, by taking advantage of fish transparency at near-infrared wavelengths and of intrinsic multiphoton landmarks. This methodology enables us to describe the frequency, distribution and modes of aNSCs divisions across the entire germinal zone of the adult pallium, and to highlight regional differences in these parameters. PMID:26395477

  12. Very Small Embryonic-Like Stem Cells: A Potential Developmental Link Between Germinal Lineage and Hematopoiesis in Humans.

    PubMed

    Virant-Klun, Irma

    2016-01-15

    It has been suggested that hematopoietic stem/progenitor cells (HSPCs) could become specified from a population of migrating primordial germ cells (PGCs), precursors of gametes, during embryogenesis. Some recent experimental data demonstrated that the cell population that is usually considered to be PGCs, moving toward the gonadal ridges of an embryo, contains a subset of cells coexpressing several germ cell and hematopoietic markers and possessing hematopoietic activity. Experimental data showed that bone morphogenetic protein 4 (BMP4) generates PGCs from mouse bone marrow-derived pluripotent stem cells. Interestingly, functional reproductive hormone receptors have been identified in HSPCs, thus indicating their potential role in reproductive function. Several reports have demonstrated fertility restoration and germ cell generation after bone marrow transplantation in both animal models and humans. A potential link between HSPCs and germinal lineage might be represented by very small embryonic-like stem cells (VSELs), which have been found in adult human bone marrow, peripheral blood, and umbilical cord blood, express a specific pattern of pluripotency, germinal lineage, and hematopoiesis, and are proposed to persist in adult tissues and organs from the embryonic period of life. Stem cell populations, similar to VSELs, expressing several genes related to pluripotency and germinal lineage, especially to PGCs, have been discovered in adult human reproductive organs, ovaries and testicles, and were related to primitive germ cell-like cell development in vitro, thus supporting the idea of VSELs as a potential link between germinal lineage and hematopoiesis.

  13. Diploid, but not haploid, human embryonic stem cells can be derived from microsurgically repaired tripronuclear human zygotes.

    PubMed

    Fan, Yong; Li, Rong; Huang, Jin; Yu, Yang; Qiao, Jie

    2013-01-15

    Human embryonic stem cells have shown tremendous potential in regenerative medicine, and the recent progress in haploid embryonic stem cells provides new insights for future applications of embryonic stem cells. Disruption of normal fertilized embryos remains controversial; thus, the development of a new source for human embryonic stem cells is important for their usefulness. Here, we investigated the feasibility of haploid and diploid embryo reconstruction and embryonic stem cell derivation using microsurgically repaired tripronuclear human zygotes. Diploid and haploid zygotes were successfully reconstructed, but a large proportion of them still had a tripolar spindle assembly. The reconstructed embryos developed to the blastocyst stage, although the loss of chromosomes was observed in these zygotes. Finally, triploid and diploid human embryonic stem cells were derived from tripronuclear and reconstructed zygotes (from which only one pronucleus was removed), but haploid human embryonic stem cells were not successfully derived from the reconstructed zygotes when two pronuclei were removed. Both triploid and diploid human embryonic stem cells showed the general characteristics of human embryonic stem cells. These results indicate that the lower embryo quality resulting from abnormal spindle assembly contributed to the failure of the haploid embryonic stem cell derivation. However, the successful derivation of diploid embryonic stem cells demonstrated that microsurgical tripronuclear zygotes are an alternative source of human embryonic stem cells. In the future, improving spindle assembly will facilitate the application of triploid zygotes to the field of haploid embryonic stem cells.

  14. Diploid, but not haploid, human embryonic stem cells can be derived from microsurgically repaired tripronuclear human zygotes.

    PubMed

    Fan, Yong; Li, Rong; Huang, Jin; Yu, Yang; Qiao, Jie

    2013-01-15

    Human embryonic stem cells have shown tremendous potential in regenerative medicine, and the recent progress in haploid embryonic stem cells provides new insights for future applications of embryonic stem cells. Disruption of normal fertilized embryos remains controversial; thus, the development of a new source for human embryonic stem cells is important for their usefulness. Here, we investigated the feasibility of haploid and diploid embryo reconstruction and embryonic stem cell derivation using microsurgically repaired tripronuclear human zygotes. Diploid and haploid zygotes were successfully reconstructed, but a large proportion of them still had a tripolar spindle assembly. The reconstructed embryos developed to the blastocyst stage, although the loss of chromosomes was observed in these zygotes. Finally, triploid and diploid human embryonic stem cells were derived from tripronuclear and reconstructed zygotes (from which only one pronucleus was removed), but haploid human embryonic stem cells were not successfully derived from the reconstructed zygotes when two pronuclei were removed. Both triploid and diploid human embryonic stem cells showed the general characteristics of human embryonic stem cells. These results indicate that the lower embryo quality resulting from abnormal spindle assembly contributed to the failure of the haploid embryonic stem cell derivation. However, the successful derivation of diploid embryonic stem cells demonstrated that microsurgical tripronuclear zygotes are an alternative source of human embryonic stem cells. In the future, improving spindle assembly will facilitate the application of triploid zygotes to the field of haploid embryonic stem cells. PMID:23255130

  15. Distinct human stem cell populations in small and large intestine.

    PubMed

    Cramer, Julie M; Thompson, Timothy; Geskin, Albert; LaFramboise, William; Lagasse, Eric

    2015-01-01

    The intestine is composed of an epithelial layer containing rapidly proliferating cells that mature into two regions, the small and the large intestine. Although previous studies have identified stem cells as the cell-of-origin for intestinal epithelial cells, no studies have directly compared stem cells derived from these anatomically distinct regions. Here, we examine intrinsic differences between primary epithelial cells isolated from human fetal small and large intestine, after in vitro expansion, using the Wnt agonist R-spondin 2. We utilized flow cytometry, fluorescence-activated cell sorting, gene expression analysis and a three-dimensional in vitro differentiation assay to characterize their stem cell properties. We identified stem cell markers that separate subpopulations of colony-forming cells in the small and large intestine and revealed important differences in differentiation, proliferation and disease pathways using gene expression analysis. Single cells from small and large intestine cultures formed organoids that reflect the distinct cellular hierarchy found in vivo and respond differently to identical exogenous cues. Our characterization identified numerous differences between small and large intestine epithelial stem cells suggesting possible connections to intestinal disease.

  16. GATAe regulates intestinal stem cell maintenance and differentiation in Drosophila adult midgut.

    PubMed

    Okumura, Takashi; Takeda, Koji; Kuchiki, Megumi; Akaishi, Marie; Taniguchi, Kiichiro; Adachi-Yamada, Takashi

    2016-02-01

    Adult intestinal tissues, exposed to the external environment, play important roles including barrier and nutrient-absorption functions. These functions are ensured by adequately controlled rapid-cell metabolism. GATA transcription factors play essential roles in the development and maintenance of adult intestinal tissues both in vertebrates and invertebrates. We investigated the roles of GATAe, the Drosophila intestinal GATA factor, in adult midgut homeostasis with its first-generated knock-out mutant as well as cell type-specific RNAi and overexpression experiments. Our results indicate that GATAe is essential for proliferation and maintenance of intestinal stem cells (ISCs). Also, GATAe is involved in the differentiation of enterocyte (EC) and enteroendocrine (ee) cells in both Notch (N)-dependent and -independent manner. The results also indicate that GATAe has pivotal roles in maintaining normal epithelial homeostasis of the Drosophila adult midgut through interaction of N signaling. Since recent reports showed that mammalian GATA-6 regulates normal and cancer stem cells in the adult intestinal tract, our data also provide information on the evolutionally conserved roles of GATA factors in stem-cell regulation. PMID:26719127

  17. A single cell bioengineering approach to elucidate mechanisms of adult stem cell self-renewal.

    PubMed

    Gilbert, Penney M; Corbel, Stephane; Doyonnas, Regis; Havenstrite, Karen; Magnusson, Klas E G; Blau, Helen M

    2012-04-01

    The goal of regenerative medicine is to restore form and function to damaged and aging tissues. Adult stem cells, present in tissues such as skeletal muscle, comprise a reservoir of cells with a remarkable capacity to proliferate and repair tissue damage. Muscle stem cells, known as satellite cells, reside in a quiescent state in an anatomically distinct compartment, or niche, ensheathed between the membrane of the myofiber and the basal lamina. Recently, procedures for isolating satellite cells were developed and experiments testing their function upon transplantation into muscles revealed an extraordinary potential to contribute to muscle fibers and access and replenish the satellite cell compartment. However, these properties are rapidly lost once satellite cells are plated in culture. Accordingly, elucidating the role of extrinsic factors in controlling muscle stem cell fate, in particular self-renewal, is critical. Through careful design of bioengineered culture platforms, analysis of specific proteins presented to stem cells is possible. Critical to the success of the approach is single cell analysis, as more rapidly proliferating progenitors may mask the behavior of stem cells that proliferate slowly. Bioengineering approaches provide a potent means of gaining insight into the role of extrinsic factors in the stem cell microenvironment on stem cell function and the mechanisms that control their diverse fates. Ultimately, the multidisciplinary approach presented here will lead to novel therapeutic strategies for degenerative diseases. PMID:22327505

  18. Derivation and differentiation of haploid human embryonic stem cells.

    PubMed

    Sagi, Ido; Chia, Gloryn; Golan-Lev, Tamar; Peretz, Mordecai; Weissbein, Uri; Sui, Lina; Sauer, Mark V; Yanuka, Ofra; Egli, Dieter; Benvenisty, Nissim

    2016-04-01

    Diploidy is a fundamental genetic feature in mammals, in which haploid cells normally arise only as post-meiotic germ cells that serve to ensure a diploid genome upon fertilization. Gamete manipulation has yielded haploid embryonic stem (ES) cells from several mammalian species, but haploid human ES cells have yet to be reported. Here we generated and analysed a collection of human parthenogenetic ES cell lines originating from haploid oocytes, leading to the successful isolation and maintenance of human ES cell lines with a normal haploid karyotype. Haploid human ES cells exhibited typical pluripotent stem cell characteristics, such as self-renewal capacity and a pluripotency-specific molecular signature. Moreover, we demonstrated the utility of these cells as a platform for loss-of-function genetic screening. Although haploid human ES cells resembled their diploid counterparts, they also displayed distinct properties including differential regulation of X chromosome inactivation and of genes involved in oxidative phosphorylation, alongside reduction in absolute gene expression levels and cell size. Surprisingly, we found that a haploid human genome is compatible not only with the undifferentiated pluripotent state, but also with differentiated somatic fates representing all three embryonic germ layers both in vitro and in vivo, despite a persistent dosage imbalance between the autosomes and X chromosome. We expect that haploid human ES cells will provide novel means for studying human functional genomics and development.

  19. Derivation and differentiation of haploid human embryonic stem cells.

    PubMed

    Sagi, Ido; Chia, Gloryn; Golan-Lev, Tamar; Peretz, Mordecai; Weissbein, Uri; Sui, Lina; Sauer, Mark V; Yanuka, Ofra; Egli, Dieter; Benvenisty, Nissim

    2016-04-01

    Diploidy is a fundamental genetic feature in mammals, in which haploid cells normally arise only as post-meiotic germ cells that serve to ensure a diploid genome upon fertilization. Gamete manipulation has yielded haploid embryonic stem (ES) cells from several mammalian species, but haploid human ES cells have yet to be reported. Here we generated and analysed a collection of human parthenogenetic ES cell lines originating from haploid oocytes, leading to the successful isolation and maintenance of human ES cell lines with a normal haploid karyotype. Haploid human ES cells exhibited typical pluripotent stem cell characteristics, such as self-renewal capacity and a pluripotency-specific molecular signature. Moreover, we demonstrated the utility of these cells as a platform for loss-of-function genetic screening. Although haploid human ES cells resembled their diploid counterparts, they also displayed distinct properties including differential regulation of X chromosome inactivation and of genes involved in oxidative phosphorylation, alongside reduction in absolute gene expression levels and cell size. Surprisingly, we found that a haploid human genome is compatible not only with the undifferentiated pluripotent state, but also with differentiated somatic fates representing all three embryonic germ layers both in vitro and in vivo, despite a persistent dosage imbalance between the autosomes and X chromosome. We expect that haploid human ES cells will provide novel means for studying human functional genomics and development. PMID:26982723

  20. Human mesenchymal stem-cell behaviour on direct laser micropatterned electrospun scaffolds with hierarchical structures.

    PubMed

    Li, Huaqiong; Wong, Yee Shan; Wen, Feng; Ng, Kee Woei; Ng, Gary Ka Lai; Venkatraman, Subbu S; Boey, Freddy Yin Chiang; Tan, Lay Poh

    2013-03-01

    Direct laser machining and electrospinning are utilized to obtain a bi-layered hybrid scaffold with hierarchical topographical features to mimic extracellular matrix-like microenvironment of cells. Adult bone marrow derived human mesenchymal stem cells (hMSCs) are cultured in vitro in these hybrid scaffolds, and cell orientation, proliferation, viability, and differentiation are evaluated. The results show that this novel hybrid scaffold not only supports cell growth like traditional scaffolds, but also elicits positive responses from the cells, like lineage commitment and alignment, which are essential features of future scaffolds. PMID:23233197

  1. Persistent production of neurons from adult brain stem cells during recovery after stroke.

    PubMed

    Thored, Pär; Arvidsson, Andreas; Cacci, Emanuele; Ahlenius, Henrik; Kallur, Therése; Darsalia, Vladimer; Ekdahl, Christine T; Kokaia, Zaal; Lindvall, Olle

    2006-03-01

    Neural stem cells in the subventricular zone of adult rodents produce new striatal neurons that may replace those that have died after stroke; however, the neurogenic response has been considered acute and transient, yielding only small numbers of neurons. In contrast, we show herein that striatal neuroblasts are generated without decline at least for 4 months after stroke in adult rats. Neuroblasts formed early or late after stroke either differentiate into mature neurons, which survive for several months, or die through caspase-mediated apoptosis. The directed migration of the new neurons toward the ischemic damage is regulated by stromal cell-derived factor-1alpha and its receptor CXCR4. These results show that endogenous neural stem cells continuously supply the injured adult brain with new neurons, which suggests novel self-repair strategies to improve recovery after stroke. PMID:16210404

  2. Integrating human stem cell expansion and neuronal differentiation in bioreactors

    PubMed Central

    Serra, Margarida; Brito, Catarina; Costa, Eunice M; Sousa, Marcos FQ; Alves, Paula M

    2009-01-01

    Background Human stem cells are cellular resources with outstanding potential for cell therapy. However, for the fulfillment of this application, major challenges remain to be met. Of paramount importance is the development of robust systems for in vitro stem cell expansion and differentiation. In this work, we successfully developed an efficient scalable bioprocess for the fast production of human neurons. Results The expansion of undifferentiated human embryonal carcinoma stem cells (NTera2/cl.D1 cell line) as 3D-aggregates was firstly optimized in spinner vessel. The media exchange operation mode with an inoculum concentration of 4 × 105 cell/mL was the most efficient strategy tested, with a 4.6-fold increase in cell concentration achieved in 5 days. These results were validated in a bioreactor where similar profile and metabolic performance were obtained. Furthermore, characterization of the expanded population by immunofluorescence microscopy and flow cytometry showed that NT2 cells maintained their stem cell characteristics along the bioreactor culture time. Finally, the neuronal differentiation step was integrated in the bioreactor process, by addition of retinoic acid when cells were in the middle of the exponential phase. Neurosphere composition was monitored and neuronal differentiation efficiency evaluated along the culture time. The results show that, for bioreactor cultures, we were able to increase significantly the neuronal differentiation efficiency by 10-fold while reducing drastically, by 30%, the time required for the differentiation process. Conclusion The culture systems developed herein are robust and represent one-step-forward towards the development of integrated bioprocesses, bridging stem cell expansion and differentiation in fully controlled bioreactors. PMID:19772662

  3. Inferring human stem cell behaviour from epigenetic drift.

    PubMed

    Shibata, D

    2009-01-01

    Although conventional experimental manipulations are impractical, it may be possible to infer human stem cell fates by 'reading' histories recorded within their genomes. Genomes are almost perfect copies of copies, and ancestries may be surreptitiously recorded by replication errors that inevitably accumulate. The greater the number of divisions, the greater the number of replication errors ('a molecular clock hypothesis'). Mutations rarely occur during a lifetime, but DNA methylation patterns are also copied after DNA replication and measurably drift with ageing at certain CpG sites in mitotic tissues, such as the colon. Such passenger methylation pattern variation may effectively function as 'epigenetic' somatic cell mitotic clocks. Replication errors can only accumulate in long-lived stem cell lineages, so methylation pattern drift largely records stem cell behaviour. How methylation patterns may encode stem cell ancestries is illustrated with two types of small reproductive units--colon crypt niches with continuous genealogies, and hair follicles with punctuated genealogies. Potentially, the genealogy of any human cell may be inferred by 'reading' its genome. PMID:19031430

  4. Modeling neurodevelopmental disorders using human pluripotent stem cells.

    PubMed

    Telias, Michael; Ben-Yosef, Dalit

    2014-08-01

    Neurodevelopmental disorders (NDs) are impairments that affect the development and growth of the brain and the central nervous system during embryonic and early postnatal life. Genetically manipulated animals have contributed greatly to the advancement of ND research, but many of them differ considerably from the human phenotype. Cellular in vitro models are also valuable, but the availability of human neuronal cells is limited and their lifespan in culture is short. Human pluripotent stem cells (hPSCs), including embryonic stem cells and induced pluripotent stem cells, comprise a powerful tool for studying developmentally regulated diseases, including NDs. We reviewed all recent studies in which hPSCs were used as in vitro models for diseases and syndromes characterized by impairment of neurogenesis or synaptogenesis leading to intellectual disability and delayed neurodevelopment. We analyzed their methodology and results, focusing on the data obtained following in vitro neural differentiation and gene expression and profiling of the derived neurons. Electrophysiological recording of action potentials, synaptic currents and response to neurotransmitters is pivotal for validation of the neuronal fate as well as for assessing phenotypic dysfunctions linked to the disease in question. We therefore focused on the studies which included electrophysiological recordings on the in vitro-derived neurons. Finally, we addressed specific issues that are critical for the advancement of this area of research, specifically in providing a reliable human pre-clinical research model and drug screening platform. PMID:24728983

  5. Identification of a candidate stem cell in human gallbladder.

    PubMed

    Manohar, Rohan; Li, Yaming; Fohrer, Helene; Guzik, Lynda; Stolz, Donna Beer; Chandran, Uma R; LaFramboise, William A; Lagasse, Eric

    2015-05-01

    There are currently no reports of identification of stem cells in human gallbladder. The differences between human gallbladder and intrahepatic bile duct (IHBD) cells have also not been explored. The goals of this study were to evaluate if human fetal gallbladder contains a candidate stem cell population and if fetal gallbladder cells are distinct from fetal IHBD cells. We found that EpCAM+CD44+CD13+ cells represent the cell population most enriched for clonal self-renewal from primary gallbladder. Primary EpCAM+CD44+CD13+ cells gave rise to EpCAM+CD44+CD13+ and EpCAM+CD44+CD13- cells in vitro, and gallbladder cells expanded in vitro exhibited short-term engraftment in vivo. Last, we found that CD13, CD227, CD66, CD26 and CD49b were differentially expressed between gallbladder and IHBD cells cultured in vitro indicating clear phenotypic differences between the two cell populations. Microarray analyses of expanded cultures confirmed that both cell types have unique transcriptional profiles with predicted functional differences in lipid, carbohydrate, nucleic acid and drug metabolism. In conclusion, we have isolated a distinct clonogenic population of epithelial cells from primary human fetal gallbladder with stem cell characteristics and found it to be unique compared to IHBD cells.

  6. Intrahepatic transplantation of CD34+ cord blood stem cells into newborn and adult NOD/SCID mice induce differential organ engraftment.

    PubMed

    Wulf-Goldenberg, Annika; Keil, Marlen; Fichtner, Iduna; Eckert, Klaus

    2012-04-01

    In vivo studies concerning the function of human hematopoietic stem cells (HSC) are limited by relatively low levels of engraftment and the failure of the engrafted HSC preparations to differentiate into functional immune cells after systemic application. In the present paper we describe the effect of intrahepatically transplanted CD34(+) cells from cord blood into the liver of newborn or adult NOD/SCID mice on organ engraftment and differentiation. Analyzing the short and long term time dependency of human cell recruitment into mouse organs after cell transplantation in the liver of newborn and adult NOD/SCID mice by RT-PCR and FACS analysis, a significantly high engraftment was found after transplantation into liver of newborn NOD/SCID mice compared to adult mice, with the highest level of 35% human cells in bone marrow and 4.9% human cells in spleen at day 70. These human cells showed CD19 B-cell, CD34 and CD38 hematopoietic and CD33 myeloid cell differentiation, but lacked any T-cell differentiation. HSC transplantation into liver of adult NOD/SCID mice resulted in minor recruitment of human cells from mouse liver to other mouse organs. The results indicate the usefulness of the intrahepatic application route into the liver of newborn NOD/SCID mice for the investigation of hematopoietic differentiation potential of CD34(+) cord blood stem cell preparations.

  7. The moral value of induced pluripotent stem cells: a Japanese bioethics perspective on human embryo research.

    PubMed

    Sawai, Tsutomu

    2014-11-01

    In contemporary Japan, at least in the field of regenerative medicine, human induced pluripotent stem cells (hiPSCs) are given no moral status and are treated in a purely instrumental way. However, some authors have mentioned the potentiality of hiPSCs in that 'tetraploid complementation' would make it possible to create humans directly from human embryonic stem cells (hESCs) and hiPSCs. A blastocyst consists of inner cell mass (ICM) cells and a trophoblast. The tetraploid complementation technique demonstrates that hESCs and hiPSCs both have the same capacity as ICM cells. If ICM cells, hESCs and hiPSCs were all provided with a trophoblast or a substitute with the same function, which would work as a placenta, they would have the same potential to develop into embryos, fetuses and adult human beings. Thus hiPSCs could be regarded as potential humans. However, no authority or guideline in Japan has specifically considered the status and use of hiPSCs. In this paper, I will address the extent to which the existing recommendations apply to hiPSCs and develop a novel Japanese bioethical perspective on the status of hiPSCs and its implications for hiPSC research, based on the reasoning in the report, 'The fundamental way of thinking in treating the human embryo' presented by the Bioethics Committee of the Council for Science and Technology Policy in 2004, and broader consideration of Japanese culture.

  8. Methods of Liver Stem Cell Therapy in Rodents as Models of Human Liver Regeneration in Hepatic Failure.

    PubMed

    Hashemi Goradel, Nasser; Darabi, Masoud; Shamsasenjan, Karim; Ejtehadifar, Mostafa; Zahedi, Sarah

    2015-09-01

    Cell therapy is a promising intervention for treating liver diseases and liver failure. Different animal models of human liver cell therapy have been developed in recent years. Rats and mice are the most commonly used liver failure models. In fact, rodent models of hepatic failure have shown significant improvement in liver function after cell infusion. With the advent of stem-cell technologies, it is now possible to re-programme adult somatic cells such as skin or hair-follicle cells from individual patients to stem-like cells and differentiate them into liver cells. Such regenerative stem cells are highly promising in the personalization of cell therapy. The present review article will summarize current approaches to liver stem cell therapy with rodent models. In addition, we discuss common cell tracking techniques and how tracking data help to direct liver cell therapy research in animal models of hepatic failure.

  9. The versatile functions of Sox9 in development, stem cells, and human diseases

    PubMed Central

    Jo, Alice; Denduluri, Sahitya; Zhang, Bosi; Wang, Zhongliang; Yin, Liangjun; Yan, Zhengjian; Kang, Richard; Shi, Lewis L.; Mok, James; Lee, Michael J.; Haydon, Rex C.

    2014-01-01

    The transcription factor Sox9 was first discovered in patients with campomelic dysplasia, a haploinsufficiency disorder with skeletal deformities caused by dysregulation of Sox9 expression during chondrogenesis. Since then, its role as a cell fate determiner during embryonic development has been well characterized; Sox9 expression differentiates cells derived from all three germ layers into a large variety of specialized tissues and organs. However, recent data has shown that ectoderm- and endoderm-derived tissues continue to express Sox9 in mature organs and stem cell pools, suggesting its role in cell maintenance and specification during adult life. The versatility of Sox9 may be explained by a combination of post-transcriptional modifications, binding partners, and the tissue type in which it is expressed. Considering its importance during both development and adult life, it follows that dysregulation of Sox9 has been implicated in various congenital and acquired diseases, including fibrosis and cancer. This review provides a summary of the various roles of Sox9 in cell fate specification, stem cell biology, and related human diseases. Ultimately, understanding the mechanisms that regulate Sox9 will be crucial for developing effective therapies to treat disease caused by stem cell dysregulation or even reverse organ damage. PMID:25685828

  10. Effect of Recombinant Human Erythropoietin On the Stemness of Bone Marrow-derived Mesenchymal Stem Cells in vitro

    PubMed Central

    Ye, Lihua; Chen, Long; Yu, Qiang; Cheng, Fanjun

    2010-01-01

    The purpose of this study was to investigate the effects of the recombinant human erythropoietin (rhEPO) on proliferative and multi-differentiation potential of the bone marrow-derived mesenchymal stem cells (MSCs). The MSCs were isolated primarily from bone marrow of adult rat and purified at increasing passage. A purified population of MSCs can be obtained about 2 weeks after the initiation of culture. After three passages (P3-MSCs), bone marrow-derived adherent cells were identified, then different concentrations of rhEPO (0.1, 1, 5, 10, 100 U/ml) was added into the Passage-3 cells which had been identified. The expression of the surface markers in adherent cells was detected by the flow cytometry. The mRNA levels of transcription factors OCT4, SOX2, Nanog and TERT were measured by reverse transcription-polymerase chain reaction (RT-PCR). The results showed that CD29 and CD90 were positive in MSCs, but not CD33, CD44 and CD45, and the cells could differentiate into multiple lineages such as osteocytes and adipocytes. The expression of OCT4, SOX2, TERT, Nanog mRNA were up-regulated by the treatment of EPO. The effect of EPO was the most obvious when its concentration was 5U/mL after 12h. we conclude that MSCs can not only perserve characteristics of stem cells but also maintain its multi-lineage differentiation potential after appropriate treatment of EPO. PMID:24855555

  11. Conditionally reprogrammed cells represent a stem-like state of adult epithelial cells

    PubMed Central

    Suprynowicz, Frank A.; Upadhyay, Geeta; Krawczyk, Ewa; Kramer, Sarah C.; Hebert, Jess D.; Liu, Xuefeng; Yuan, Hang; Cheluvaraju, Chaitra; Clapp, Phillip W.; Boucher, Richard C.; Kamonjoh, Christopher M.; Randell, Scott H.; Schlegel, Richard

    2012-01-01

    The combination of irradiated fibroblast feeder cells and Rho kinase inhibitor, Y-27632, conditionally induces an indefinite proliferative state in primary mammalian epithelial cells. These conditionally reprogrammed cells (CRCs) are karyotype-stable and nontumorigenic. Because self-renewal is a recognized property of stem cells, we investigated whether Y-27632 and feeder cells induced a stem-like phenotype. We found that CRCs share characteristics of adult stem cells and exhibit up-regulated expression of α6 and β1 integrins, ΔNp63α, CD44, and telomerase reverse transcriptase, as well as decreased Notch signaling and an increased level of nuclear β-catenin. The induction of CRCs is rapid (occurs within 2 d) and results from reprogramming of the entire cell population rather than the selection of a minor subpopulation. CRCs do not overexpress the transcription factor sets characteristic of embryonic or induced pluripotent stem cells (e.g., Sox2, Oct4, Nanog, or Klf4). The induction of CRCs is also reversible, and removal of Y-27632 and feeders allows the cells to differentiate normally. Thus, when CRCs from ectocervical epithelium or tracheal epithelium are placed in an air–liquid interface culture system, the cervical cells form a well differentiated stratified squamous epithelium, whereas the tracheal cells form a ciliated airway epithelium. We discuss the diagnostic and therapeutic opportunities afforded by a method that can generate adult stem-like cells in vitro without genetic manipulation. PMID:23169653

  12. Concise review: human umbilical cord stroma with regard to the source of fetus-derived stem cells.

    PubMed

    Can, Alp; Karahuseyinoglu, Sercin

    2007-11-01

    Human umbilical cord (UC) has been a tissue of increasing interest in recent years. Many groups have shown the stem cell potency of stromal cells isolated from the human UC mesenchymal tissue, namely, Wharton's jelly. Since UC is a postnatal organ discarded after birth, the collection of cells does not require an invasive procedure with ethical concerns. Stromal cells, as the dominant cells of this fetus-derived tissue, possess multipotent properties between embryonic stem cells and adult stem cells. They bear a relatively higher proliferation rate and self-renewal capacity. Although they share common surface markers with bone marrow-derived MSCs, they also express certain embryonic stem cell markers, albeit in low levels. Without any spontaneous differentiation, they can be successfully differentiated into mature adipocytes, osteoblasts, chondrocytes, skeletal myocytes, cardiomyocytes, neurons, and endothelial cells. While causing no immunorejection reaction, they effectively function in vivo as dopaminergic neurons, myocytes, and endothelial cells. Given these characteristics, particularly the plasticity and developmental flexibility, UC stromal cells are now considered an alternative source of stem cells and deserve to be examined in long-term clinical trials. This review first aims to document the published findings so far regarding the nature of human UC stroma with special emphasis on the spatial distribution and functional structure of stromal cells and matrix, which serves as a niche for residing cells, and, secondly, to assess the in vitro and in vivo experiments in which differential stem cell potencies were evaluated.

  13. Potential Reparative Role of Resident Adult Renal Stem/Progenitor Cells in Acute Kidney Injury

    PubMed Central

    Sallustio, Fabio; Serino, Grazia; Schena, Francesco Paolo

    2015-01-01

    Abstract Human kidney is particularly susceptible to ischemia and toxins with consequential tubular necrosis and activation of inflammatory processes. This process can lead to the acute renal injury, and even if the kidney has a great capacity for regeneration after tubular damage, in several circumstances, the normal renal repair program may not be sufficient to achieve a successful regeneration. Resident adult renal stem/progenitor cells could participate in this repair process and have the potentiality to enhance the renal regenerative mechanism. This could be achieved both directly, by means of their capacity to differentiate and integrate into the renal tissues, and by means of paracrine factors able to induce or improve the renal repair or regeneration. Recent genetic fate-tracing studies indicated that tubular damage is instead repaired by proliferative duplication of epithelial cells, acquiring a transient progenitor phenotype and by fate-restricted clonal cell progeny emerging from different nephron segments. In this review, we discuss about the properties and the reparative characteristics of high regenerative CD133+/CD24+ cells, with a view to a future application of these cells for the treatment of acute renal injury. PMID:26309808

  14. Inflammatory cues acting on the adult intestinal stem cells and the early onset of cancer (Review)

    PubMed Central

    DE LERMA BARBARO, A.; PERLETTI, G.; BONAPACE, I.M.; MONTI, E.

    2014-01-01

    The observation that cancer often arises at sites of chronic inflammation has prompted the idea that carcinogenesis and inflammation are deeply interwoven. In fact, the current literature highlights a role for chronic inflammation in virtually all the steps of carcinogenesis, including tumor initiation, promotion and progression. The aim of the present article is to review the current literature on the involvement of chronic inflammation in the initiation step and in the very early phases of tumorigenesis, in a type of cancer where adult stem cells are assumed to be the cells of origin of neoplasia. Since the gastrointestinal tract is regarded as the best-established model system to address the liaison between chronic inflammation and neoplasia, the focus of this article will be on intestinal cancer. In fact, the anatomy of the intestinal epithelial lining is uniquely suited to study adult stem cells in their niche, and the bowel crypt is an ideal developmental biology system, as proliferation, differentiation and cell migration are all distributed linearly along the long axis of the crypt. Moreover, crypt stem cells are regarded today as the most likely targets of neoplastic transformation in bowel cancer. More specifically, the present review addresses the molecular mechanisms whereby a state of chronic inflammation could trigger the neoplastic process in the intestine, focusing on the generation of inflammatory cues evoking enhanced proliferation in cells not initiated but at risk of neoplastic transformation because of their stemness. Novel experimental approaches, based on triggering an inflammatory stimulus in the neighbourhood of adult intestinal stem cells, are warranted to address some as yet unanswered questions. A possible approach, the targeted transgenesis of Paneth cells, may be aimed at ‘hijacking’ the crypt stem cell niche from a status characterized by the maintenance of homeostasis to local chronic inflammation, with the prospect of initiating

  15. Surface-Functionalized Silk Fibroin Films as a Platform To Guide Neuron-like Differentiation of Human Mesenchymal Stem Cells.

    PubMed

    Manchineella, Shivaprasad; Thrivikraman, Greeshma; Basu, Bikramjit; Govindaraju, T

    2016-09-01

    Surface interactions at the biomaterial-cellular interface determine the proliferation and differentiation of stem cells. Manipulating such interactions through the surface chemistry of scaffolds renders control over directed stem cell differentiation into the cell lineage of interest. This approach is of central importance for stem cell-based tissue engineering and regenerative therapy applications. In the present study, silk fibroin films (SFFs) decorated with integrin-binding laminin peptide motifs (YIGSR and GYIGSR) were prepared and employed for in vitro adult stem cell-based neural tissue engineering applications. Functionalization of SFFs with short peptides showcased the peptide sequence and nature of functionalization-dependent differentiation of bone marrow-derived human mesenchymal stem cells (hMSCs). Intriguingly, covalently functionalized SFFs with GYIGSR hexapeptide (CL2-SFF) supported hMSC proliferation and maintenance in an undifferentiated pluripotent state and directed the differentiation of hMSCs into neuron-like cells in the presence of a biochemical cue, on-demand. The observed morphological changes were further corroborated by the up-regulation of neuronal-specific marker gene expression (MAP2, TUBB3, NEFL), confirmed through semiquantitative reverse-transcription polymerase chain reaction (RT-PCR) analysis. The enhanced proliferation and on-demand directed differentiation of adult stem cells (hMSCs) by the use of an economically viable short recognition peptide (GYIGSR), as opposed to the integrin recognition protein laminin, establishes the potential of SFFs for neural tissue engineering and regenerative therapy applications. PMID:27518901

  16. Recent insights into the molecular mechanisms involved in aging and the malignant transformation of adult stem/progenitor cells and their therapeutic implications.

    PubMed

    Mimeault, Murielle; Batra, Surinder K

    2009-04-01

    Recent advancements in tissue-resident adult stem/progenitor cell research have revealed that enhanced telomere attrition, oxidative stress, ultraviolet radiation exposure and oncogenic events leading to severe DNA damages and genomic instability may occur in these immature and regenerative cells during chronological aging. Particularly, the alterations in key signaling components controlling their self-renewal capacity and an up-regulation of tumor suppressor gene products such as p16(INK4A), p19(ARF), ataxia-telangiectasia mutated (ATM) kinase, p53 and/or the forkhead box O (FOXOs) family of transcription factors may result in their dysfunctions, growth arrest and senescence or apoptotic death during the aging process. These molecular events may culminate in a progressive decline in the regenerative functions and the number of tissue-resident adult stem/progenitor cells, and age-related disease development. Conversely, the telomerase re-activation and accumulation of numerous genetic and/or epigenetic alterations in adult stem/progenitor cells with advancing age may result in their immortalization and malignant transformation into highly leukemic or tumorigenic cancer-initiating cells and cancer initiation. Therefore, the cell-replacement and gene therapies and molecular targeting of aged and dysfunctional adult stem/progenitor cells including their malignant counterpart, cancer-initiating cells, hold great promise for treating and even curing diverse devastating human diseases. These diseases include premature aging diseases, hematopoietic, cardiovascular, musculoskeletal, pulmonary, ocular, urogenital, neurodegenerative and skin disorders and aggressive and recurrent cancers.

  17. A regulatory argument against human embryonic stem cell research.

    PubMed

    Napier, Stephen

    2009-10-01

    This article explores the plausibility of an argument against embryonic stem cell research based on what the regulations already say about research on pregnant women and fetuses. The center of the argument is the notion of vulnerability and whether such a concept is applicable to human embryos. It is argued that such an argument can be made plausible. The article concludes by responding to several important objections.

  18. Effect of uncontrolled freezing on biological characteristics of human dental pulp stem cells.

    PubMed

    Kumar, Ajay; Bhattacharyya, Shalmoli; Rattan, Vidya

    2015-12-01

    Human dental pulp stem cells (hDPSCs) hold great promise as a source of adult stem cells for utilization in regenerative medicine. Successful storage and post thaw recovery of DPSCs without loss of function is a key issue for future clinical application. Most of the cryopreservation methods use controlled rate freezing and vapor phase nitrogen to store stem cells. But these methods are both expensive and laborious. In this study, we isolated DPSCs from a patient undergoing impacted mandibular third molar extraction. We adopted eight different methods of cryopreservation at -80 °C for long term storage of the DPSC aliquots. Various parameters like proliferation, cell death, cell cycle, retention of stemness markers and differentiation potential were studied post cryopreservation period of 1 year. We observed successful recovery of stem cells in every method and a significant difference in proliferation potential and cell death between samples stored by different methods. However, post thaw, all cells retained their stemness markers. All DPSCs stored by different methods were able to differentiate into osteoblast like cells, adipocytes and neural cells. Based on these parameters we concluded that uncontrolled freezing at a temperature of -80 °C is as effective as controlled freezing using ethanol vessels and other cryopreservation methods. To the best of our knowledge, our study provides the first proof of concept that long term storage in uncontrolled freezing of cells at -80 °C in 10 % DMSO does not affect the revival capacity of hDPSCs. This implies that DPSCs may be used successfully for tissue engineering and cell based therapeutics even after long term, uncontrolled cryopreservation.

  19. Optimized Cryopreservation and Banking of Human Bone-Marrow Fragments and Stem Cells.

    PubMed

    Carnevale, Gianluca; Pisciotta, Alessandra; Riccio, Massimo; De Biasi, Sara; Gibellini, Lara; Ferrari, Adriano; La Sala, Giovanni B; Bruzzesi, Giacomo; Cossarizza, Andrea; de Pol, Anto

    2016-04-01

    Adult mesenchymal stem cells are a promising source for cell therapies and tissue engineering applications. Current procedures for banking of human bone-marrow mesenchymal stem cells (hBM-MSCs) require cell isolation and expansion, and thus the use of large amounts of animal sera. However, animal-derived culture supplements have the potential to trigger infections and severe immune reactions. The aim of this study was to investigate an optimized method for cryopreservation of human bone-marrow fragments for application in cell banking procedures where stem-cell expansion and use are not immediately needed. Whole trabecular fragments enclosing the bone marrow were stored in liquid nitrogen for 1 year in a cryoprotective solution containing a low concentration of dimethyl sulfoxide and a high concentration of human serum (HuS). After thawing, the isolation, colony-forming-unit ability, proliferation, morphology, stemness-related marker expression, cell senescence, apoptosis, and multi-lineage differentiation potential of hBM-MSCs were tested in media containing HuS compared with hBM-MSCs isolated from fresh fragments. Human BM-MSCs isolated from cryopreserved fragments expressed MSC markers until later passages, had a good proliferation rate, and exhibited the capacity to differentiate toward osteogenic, adipogenic, and myogenic lineages similar to hBM-MSCs isolated from fresh fragments. Moreover, the cryopreservation method did not induce cell senescence or cell death. These results imply that minimal processing may be adequate for the banking of tissue samples with no requirement for the immediate isolation and use of hBM-MSCs, thus limiting cost and the risk of contamination, and facilitating banking for clinical use. Furthermore, the use of HuS for cryopreservation and expansion/differentiation has the potential for clinical application in compliance with good manufacturing practice standards.

  20. Generation of folliculogenic human epithelial stem cells from induced pluripotent stem cells

    NASA Astrophysics Data System (ADS)

    Yang, Ruifeng; Zheng, Ying; Burrows, Michelle; Liu, Shujing; Wei, Zhi; Nace, Arben; Guo, Wei; Kumar, Suresh; Cotsarelis, George; Xu, Xiaowei

    2014-01-01

    Epithelial stem cells (EpSCs) in the hair follicle bulge are required for hair follicle growth and cycling. The isolation and propagation of human EpSCs for tissue engineering purposes remains a challenge. Here we develop a strategy to differentiate human iPSCs (hiPSCs) into CD200+/ITGA6+ EpSCs that can reconstitute the epithelial components of the hair follicle and interfollicular epidermis. The hiPSC-derived CD200+/ITGA6+ cells show a similar gene expression signature as EpSCs directly isolated from human hair follicles. Human iPSC-derived CD200+/ITGA6+ cells are capable of generating all hair follicle lineages including the hair shaft, and the inner and outer root sheaths in skin reconstitution assays. The regenerated hair follicles possess a KRT15+ stem cell population and produce hair shafts expressing hair-specific keratins. These results suggest an approach for generating large numbers of human EpSCs for tissue engineering and new treatments for hair loss, wound healing and other degenerative skin disorders.

  1. Paracrine effects of haematopoietic cells on human mesenchymal stem cells

    PubMed Central

    Zhou, Shuanhu

    2015-01-01

    Stem cell function decline during ageing can involve both cell intrinsic and extrinsic mechanisms. Bone and blood formation are intertwined in bone marrow, therefore haematopoietic cells and bone cells could be extrinsic factors for each other. In this study, we assessed the paracrine effects of extrinsic factors from haematopoietic cells on human mesenchymal stem cells (MSCs). Our data showed that haematopoietic cells stimulate proliferation, osteoblast differentiation and inhibit senescence of MSCs; TNF-α, PDGF-β, Wnt1, 4, 6, 7a and 10a, sFRP-3 and sFRP-5 are dominantly expressed in haematopoietic cells; the age-related increase of TNF-α in haematopoietic cells may perform as a negative factor in the interactions of haematopoietic cells on MSCs via TNF-α receptors and then activating NF-κB signaling or Wnt/β-catenin signaling to induce senescence and reduce osteoblast differentiation in MSCs. In conclusion, our data demonstrated that there are paracrine interactions of haematopoietic cells on human MSCs; immunosenescence may be one of the extrinsic mechanisms by which skeletal stem cell function decline during human skeletal ageing. PMID:26030407

  2. A Novel Class of Human Cardiac Stem Cells.

    PubMed

    Moccetti, Tiziano; Leri, Annarosa; Goichberg, Polina; Rota, Marcello; Anversa, Piero

    2015-01-01

    Following the recognition that hematopoietic stem cells improve the outcome of myocardial infarction in animal models, bone marrow mononuclear cells, CD34-positive cells, and mesenchymal stromal cells have been introduced clinically. The intracoronary or intramyocardial injection of these cell classes has been shown to be safe and to produce a modest but significant enhancement in systolic function. However, the identification of resident cardiac stem cells in the human heart (hCSCs) has created great expectation concerning the potential implementation of this category of autologous cells for the management of the human disease. Although phase 1 clinical trials have been conducted with encouraging results, the search for the most powerful hCSC for myocardial regeneration is in its infancy. This manuscript discusses the efforts performed in our laboratory to characterize the critical biological variables that define the growth reserve of hCSCs. Based on the theory of the immortal DNA template, we propose that stem cells retaining the old DNA represent 1 of the most powerful cells for myocardial regeneration. Similarly, the expression of insulin-like growth factor-1 receptors in hCSCs recognizes a cell phenotype with superior replicating reserve. However, the impressive recovery in ventricular hemodynamics and anatomy mediated by clonal hCSCs carrying the "mother" DNA underscores the clinical relevance of this hCSC class for the treatment of human heart failure.

  3. A Novel Class of Human Cardiac Stem Cells

    PubMed Central

    Moccetti, Tiziano; Leri, Annarosa; Goichberg, Polina; Rota, Marcello; Anversa, Piero

    2015-01-01

    Following the recognition that hematopoietic stem cells improve the outcome of myocardial infarction in animal models, bone marrow mononuclear cells, CD34-positive cells and mesenchymal stromal cells have been introduced clinically. The intracoronary or intramyocardial injection of these cell classes has been shown to be safe and to produce a modest but significant enhancement in systolic function. However, the identification of resident cardiac stem cells in the human heart (hCSCs) has created great expectation concerning the potential implementation of this category of autologous cells for the management of the human disease. Although phase 1 clinical trials have been conducted with encouraging results, the search for the most powerful hCSC for myocardial regeneration is in its infancy. This manuscript discusses the efforts performed in our laboratory to characterize the critical biological variables that define the growth reserve of hCSCs. Based on the theory of the immortal DNA template, we propose that stem cells retaining the old DNA represent one of the most powerful cells for myocardial regeneration. Similarly, the expression of insulin-like growth factor-1 receptors in hCSCs recognizes a cell phenotype with superior replicating reserve. However, the impressive recovery in ventricular hemodynamics and anatomy mediated by clonal hCSCs carrying the “mother” DNA underscores the clinical relevance of this hCSC class for the treatment of human heart failure. PMID:25807105

  4. The sexual identity of adult intestinal stem cells controls organ size and plasticity.

    PubMed

    Hudry, Bruno; Khadayate, Sanjay; Miguel-Aliaga, Irene

    2016-02-18

    Sex differences in physiology and disease susceptibility are commonly attributed to developmental and/or hormonal factors, but there is increasing realization that cell-intrinsic mechanisms play important and persistent roles. Here we use the Drosophila melanogaster intestine to investigate the nature and importance of cellular sex in an adult somatic organ in vivo. We find that the adult intestinal epithelium is a cellular mosaic of different sex differentiation pathways, and displays extensive sex differences in expression of genes with roles in growth and metabolism. Cell-specific reversals of the sexual identity of adult intestinal stem cells uncovers the key role this identity has in controlling organ size, reproductive plasticity and response to genetically induced tumours. Unlike previous examples of sexually dimorphic somatic stem cell activity, the sex differences in intestinal stem cell behaviour arise from intrinsic mechanisms that control cell cycle duration and involve a new doublesex- and fruitless-independent branch of the sex differentiation pathway downstream of transformer. Together, our findings indicate that the plasticity of an adult somatic organ is reversibly controlled by its sexual identity, imparted by a new mechanism that may be active in more tissues than previously recognized. PMID:26887495

  5. Quiescent adult neural stem cells are exceptionally sensitive to cosmic radiation.

    PubMed

    Encinas, Juan M; Vazquez, Marcelo E; Switzer, Robert C; Chamberland, Dennis W; Nick, Harry; Levine, Howard G; Scarpa, Philip J; Enikolopov, Grigori; Steindler, Dennis A

    2008-03-01

    Generation of new neurons in the adult brain, a process that is likely to be essential for learning, memory, and mood regulation, is impaired by radiation. Therefore, radiation exposure might have not only such previously expected consequences as increased probability of developing cancer, but might also impair cognitive function and emotional stability. Radiation exposure is encountered in settings ranging from cancer therapy to space travel; evaluating the neurogenic risks of radiation requires identifying the at-risk populations of stem and progenitor cells in the adult brain. Here we have used a novel reporter mouse line to find that early neural progenitors are selectively affected by conditions simulating the space radiation environment. This is reflected both in a decrease in the number of these progenitors in the neurogenic regions and in an increase in the number of dying cells in these regions. Unexpectedly, we found that quiescent neural stem cells, rather than their rapidly dividing progeny, are most sensitive to radiation. Since these stem cells are responsible for adult neurogenesis, their death would have a profound impact on the production of new neurons in the irradiated adult brain. Our finding raises an important concern about cognitive and emotional risks associated with radiation exposure.

  6. The sexual identity of adult intestinal stem cells controls organ size and plasticity

    PubMed Central

    Hudry, Bruno; Khadayate, Sanjay; Miguel-Aliaga, Irene

    2016-01-01

    SUMMARY Sex differences in physiology and disease susceptibility are commonly attributed to developmental and/or hormonal factors, but there is increasing realisation that cell-intrinsic mechanisms play important and persistent roles1,2. Here we use the Drosophila melanogaster intestine to investigate the nature and significance of cellular sex in an adult somatic organ in vivo. We find that the adult intestinal epithelium is a cellular mosaic of different sex differentiation pathways, and displays extensive sex differences in expression of genes with roles in growth and metabolism. Cell-specific reversals of the sexual identity of adult intestinal stem cells uncover its key roles in controlling organ size, its reproductive plasticity and its response to genetically induced tumours. Unlike previous examples of sexually dimorphic somatic stem cell activity, the sex differences in intestinal stem cell behaviour arise from intrinsic mechanisms, which control cell cycle duration and involve a new doublesex- and fruitless-independent branch of the sex differentiation pathway downstream of transformer. Together, our findings indicate that the plasticity of an adult somatic organ is reversibly controlled by its sexual identity, imparted by a new mechanism that may be active in more tissues than previously recognised. PMID:26887495

  7. Empowering Adult Stem Cells for Myocardial Regeneration V2.0: Success in Small Steps.

    PubMed

    Broughton, Kathleen M; Sussman, Mark A

    2016-03-01

    Much has changed since our survey of the landscape for myocardial regeneration powered by adult stem cells 4 years ago.(1) The intervening years since that first review has witnessed an explosive expansion of studies that advance both understanding and implementation of adult stem cells in promoting myocardial repair. Painstaking research from innumerable laboratories throughout the world is prying open doors that may lead to restoration of myocardial structure and function in the wake of pathological injury. This global effort has produced deeper mechanistic comprehension coupled with an evolving appreciation for the complexity of myocardial regeneration in the adult context. Undaunted by both known and (as yet) unknown challenges, pursuit of myocardial regenerative medicine mediated by adult stem cell therapy has gathered momentum fueled by tantalizing clues and visionary goals. This concise review takes a somewhat different perspective than our initial treatise, taking stock of the business sector that has become an integral part of the field while concurrently updating state of affairs in cutting edge research. Looking retrospectively at advancement over the years as all reviews eventually must, the fundamental lesson to be learned is best explained by Jonatan Mårtensson: "Success will never be a big step in the future. Success is a small step taken just now."

  8. Generation of human melanocytes from induced pluripotent stem cells.

    PubMed

    Ohta, Shigeki; Imaizumi, Yoichi; Okada, Yohei; Akamatsu, Wado; Kuwahara, Reiko; Ohyama, Manabu; Amagai, Masayuki; Matsuzaki, Yumi; Yamanaka, Shinya; Okano, Hideyuki; Kawakami, Yutaka

    2011-01-13

    Epidermal melanocytes play an important role in protecting the skin from UV rays, and their functional impairment results in pigment disorders. Additionally, melanomas are considered to arise from mutations that accumulate in melanocyte stem cells. The mechanisms underlying melanocyte differentiation and the defining characteristics of melanocyte stem cells in humans are, however, largely unknown. In the present study, we set out to generate melanocytes from human iPS cells in vitro, leading to a preliminary investigation of the mechanisms of human melanocyte differentiation. We generated iPS cell lines from human dermal fibroblasts using the Yamanaka factors (SOX2, OCT3/4, and KLF4, with or without c-MYC). These iPS cell lines were subsequently used to form embryoid bodies (EBs) and then differentiated into melanocytes via culture supplementation with Wnt3a, SCF, and ET-3. Seven weeks after inducing differentiation, pigmented cells expressing melanocyte markers such as MITF, tyrosinase, SILV, and TYRP1, were detected. Melanosomes were identified in these pigmented cells by electron microscopy, and global gene expression profiling of the pigmented cells showed a high similarity to that of human primary foreskin-derived melanocytes, suggesting the successful generation of melanocytes from iPS cells. This in vitro differentiation system should prove useful for understanding human melanocyte biology and revealing the mechanism of various pigment cell disorders, including melanoma.

  9. Comparison of human adipose-derived stem cells and bone marrow-derived stem cells in a myocardial infarction model.

    PubMed

    Rasmussen, Jeppe Grøndahl; Frøbert, Ole; Holst-Hansen, Claus; Kastrup, Jens; Baandrup, Ulrik; Zachar, Vladimir; Fink, Trine; Simonsen, Ulf

    2014-02-01

    Treatment of myocardial infarction (MI) with bone marrow-derived mesenchymal stem cells and recently also adipose-derived stem cells has shown promising results. In contrast to clinical trials and their use of autologous bone marrow-derived cells from the ischemic patient, the animal MI models are often using young donors and young, often immune-compromised, recipient animals. Our objective was to compare bone marrow-derived mesenchymal stem cells with adipose-derived stem cells from an elderly ischemic patient in the treatment of MI using a fully grown non-immune-compromised rat model. Mesenchymal stem cells were isolated from adipose tissue and bone marrow and compared with respect to surface markers and proliferative capability. To compare the regenerative potential of the two stem cell populations, male Sprague-Dawley rats were randomized to receive intramyocardial injections of adipose-derived stem cells, bone marrow-derived mesenchymal stem cells, or phosphate-buffered saline 1 week following induction of MI. After 4 weeks, left ventricular ejection fraction (LVEF) was improved in the adipose-derived stem cell group, and scar wall thickness was greater compared with the saline group. Adipose-derived as well as bone marrow-derived mesenchymal stem cells prevented left ventricular end diastolic dilation. Neither of the cell groups displayed increased angiogenesis in the myocardium compared with the saline group. Adipose-derived stem cells from a human ischemic patient preserved cardiac function following MI, whereas this could not be demonstrated for bone marrow-derived mesenchymal stem cells, with only adipose-derived stem cells leading to an improvement in LVEF. Neither of the stem cell types induced myocardial angiogenesis, raising the question whether donor age and health have an effect on the efficacy of stem cells used in the treatment of MI.

  10. Comparison of human adipose-derived stem cells and bone marrow-derived stem cells in a myocardial infarction model.

    PubMed

    Rasmussen, Jeppe Grøndahl; Frøbert, Ole; Holst-Hansen, Claus; Kastrup, Jens; Baandrup, Ulrik; Zachar, Vladimir; Fink, Trine; Simonsen, Ulf

    2014-02-01

    Treatment of myocardial infarction (MI) with bone marrow-derived mesenchymal stem cells and recently also adipose-derived stem cells has shown promising results. In contrast to clinical trials and their use of autologous bone marrow-derived cells from the ischemic patient, the animal MI models are often using young donors and young, often immune-compromised, recipient animals. Our objective was to compare bone marrow-derived mesenchymal stem cells with adipose-derived stem cells from an elderly ischemic patient in the treatment of MI using a fully grown non-immune-compromised rat model. Mesenchymal stem cells were isolated from adipose tissue and bone marrow and compared with respect to surface markers and proliferative capability. To compare the regenerative potential of the two stem cell populations, male Sprague-Dawley rats were randomized to receive intramyocardial injections of adipose-derived stem cells, bone marrow-derived mesenchymal stem cells, or phosphate-buffered saline 1 week following induction of MI. After 4 weeks, left ventricular ejection fraction (LVEF) was improved in the adipose-derived stem cell group, and scar wall thickness was greater compared with the saline group. Adipose-derived as well as bone marrow-derived mesenchymal stem cells prevented left ventricular end diastolic dilation. Neither of the cell groups displayed increased angiogenesis in the myocardium compared with the saline group. Adipose-derived stem cells from a human ischemic patient preserved cardiac function following MI, whereas this could not be demonstrated for bone marrow-derived mesenchymal stem cells, with only adipose-derived stem cells leading to an improvement in LVEF. Neither of the stem cell types induced myocardial angiogenesis, raising the question whether donor age and health have an effect on the efficacy of stem cells used in the treatment of MI. PMID:23211469

  11. PTEN deficiency reprogrammes human neural stem cells towards a glioblastoma stem cell-like phenotype.

    PubMed

    Duan, Shunlei; Yuan, Guohong; Liu, Xiaomeng; Ren, Ruotong; Li, Jingyi; Zhang, Weizhou; Wu, Jun; Xu, Xiuling; Fu, Lina; Li, Ying; Yang, Jiping; Zhang, Weiqi; Bai, Ruijun; Yi, Fei; Suzuki, Keiichiro; Gao, Hua; Esteban, Concepcion Rodriguez; Zhang, Chuanbao; Izpisua Belmonte, Juan Carlos; Chen, Zhiguo; Wang, Xiaomin; Jiang, Tao; Qu, Jing; Tang, Fuchou; Liu, Guang-Hui

    2015-01-01

    PTEN is a tumour suppressor frequently mutated in many types of cancers. Here we show that targeted disruption of PTEN leads to neoplastic transformation of human neural stem cells (NSCs), but not mesenchymal stem cells. PTEN-deficient NSCs display neoplasm-associated metabolic and gene expression profiles and generate intracranial tumours in immunodeficient mice. PTEN is localized to the nucleus in NSCs, binds to the PAX7 promoter through association with cAMP responsive element binding protein 1 (CREB)/CREB binding protein (CBP) and inhibits PAX7 transcription. PTEN deficiency leads to the upregulation of PAX7, which in turn promotes oncogenic transformation of NSCs and instates 'aggressiveness' in human glioblastoma stem cells. In a large clinical database, we find increased PAX7 levels in PTEN-deficient glioblastoma. Furthermore, we identify that mitomycin C selectively triggers apoptosis in NSCs with PTEN deficiency. Together, we uncover a potential mechanism of how PTEN safeguards NSCs, and establish a cellular platform to identify factors involved in NSC transformation, potentially permitting personalized treatment of glioblastoma. PMID:26632666

  12. PTEN deficiency reprogrammes human neural stem cells towards a glioblastoma stem cell-like phenotype.

    PubMed

    Duan, Shunlei; Yuan, Guohong; Liu, Xiaomeng; Ren, Ruotong; Li, Jingyi; Zhang, Weizhou; Wu, Jun; Xu, Xiuling; Fu, Lina; Li, Ying; Yang, Jiping; Zhang, Weiqi; Bai, Ruijun; Yi, Fei; Suzuki, Keiichiro; Gao, Hua; Esteban, Concepcion Rodriguez; Zhang, Chuanbao; Izpisua Belmonte, Juan Carlos; Chen, Zhiguo; Wang, Xiaomin; Jiang, Tao; Qu, Jing; Tang, Fuchou; Liu, Guang-Hui

    2015-12-03

    PTEN is a tumour suppressor frequently mutated in many types of cancers. Here we show that targeted disruption of PTEN leads to neoplastic transformation of human neural stem cells (NSCs), but not mesenchymal stem cells. PTEN-deficient NSCs display neoplasm-associated metabolic and gene expression profiles and generate intracranial tumours in immunodeficient mice. PTEN is localized to the nucleus in NSCs, binds to the PAX7 promoter through association with cAMP responsive element binding protein 1 (CREB)/CREB binding protein (CBP) and inhibits PAX7 transcription. PTEN deficiency leads to the upregulation of PAX7, which in turn promotes oncogenic transformation of NSCs and instates 'aggressiveness' in human glioblastoma stem cells. In a large clinical database, we find increased PAX7 levels in PTEN-deficient glioblastoma. Furthermore, we identify that mitomycin C selectively triggers apoptosis in NSCs with PTEN deficiency. Together, we uncover a potential mechanism of how PTEN safeguards NSCs, and establish a cellular platform to identify factors involved in NSC transformation, potentially permitting personalized treatment of glioblastoma.

  13. Nanofibrous substrates support colony formation and maintain stemness of human embryonic stem cells

    PubMed Central

    Gauthaman, Kalamegam; Venugopal, Jayarama Reddy; Yee, Fong Chui; Peh, Gary Swee Lim; Ramakrishna, Seeram; Bongso, Ariff

    2009-01-01

    Inadequate cell numbers in culture is one of the hurdles currently delaying the application of human embryonic stem cells (hESCs) for transplantation therapy. Nanofibrous scaffolds have been effectively used to expand and differentiate non-colony forming multipotent mesenchymal stem cells (MSC) for the repair of tissues or organs. In the present study, we evaluated the influence of nanofibrous scaffolds for hESC proliferation, increase in colony formation, self-renewal properties, undifferentiation and retention of ‘stemness’. Polycaprolactone/collagen (PCL/collagen) and PCL/gelatin nanofibrous scaffolds were fabricated using electrospinning technology. The hESCs were seeded on the nanofibrous scaffolds in the presence or absence of mitomycin-C treated mouse embryonic fibroblasts (MEFs). The hESCs grown on both scaffolds in the presence of the MEFs produced an increase in cell growth of 47.58% (P≤ 0.006) and 40.18% (P≤ 0.005), respectively, over conventional controls of hESCs on MEFs alone. The hESC colonies were also larger in diameter on the scaffolds compared to controls (PCL/collagen, 156.25 ± 7 μM and PCL/gelatin, 135.42 ± 5 μM). Immunohistochemistry of the hESCs grown on the nanofibrous scaffolds with MEFs, demonstrated positive staining for the various stemness-related markers (octamer 4 [OCT-4], tumour rejection antigen-1–60, GCTM-2 and TG-30), and semi-quantitative RT-PCR for the pluripotent stemness genomic markers (NANOG, SOX-2, OCT-4) showed that they were also highly expressed. Continued successful propagation of hESC colonies from nanofibrous scaffolds back to conventional culture on MEFs was also possible. Nanofibrous scaffolds support hESC expansion in an undifferentiated state with retention of stemness characteristics thus having tremendous potential in scaling up cell numbers for transplantation therapy. PMID:19228268

  14. Human pluripotent stem cell-derived limbal epithelial stem cells on bioengineered matrices for corneal reconstruction.

    PubMed

    Mikhailova, Alexandra; Ilmarinen, Tanja; Ratnayake, Anjula; Petrovski, Goran; Uusitalo, Hannu; Skottman, Heli; Rafat, Mehrdad

    2016-05-01

    Corneal epithelium is renewed by limbal epithelial stem cells (LESCs), a type of tissue-specific stem cells located in the limbal palisades of Vogt at the corneo-scleral junction. Acute trauma or inflammatory disorders of the ocular surface can destroy these stem cells, leading to limbal stem cell deficiency (LSCD) - a painful and vision-threatening condition. Treating these disorders is often challenging and complex, especially in bilateral cases with extensive damage. Human pluripotent stem cells (hPSCs) provide new opportunities for corneal reconstruction using cell-based therapy. Here, we investigated the use of hPSC-derived LESC-like cells on bioengineered collagen matrices in serum-free conditions, aiming for clinical applications to reconstruct the corneal epithelium and partially replace the damaged stroma. Differentiation of hPSCs towards LESC-like cells was directed using small-molecule induction followed by maturation in corneal epithelium culture medium. After four to five weeks of culture, differentiated cells were seeded onto bioengineered matrices fabricated as transparent membranes of uniform thickness, using medical-grade porcine collagen type I and a hybrid cross-linking technology. The bioengineered matrices were fully transparent, with high water content and swelling capacity, and parallel lamellar microstructure. Cell proliferation of hPSC-LESCs was significantly higher on bioengineered matrices than on collagen-coated control wells after two weeks of culture, and LESC markers p63 and cytokeratin 15, along with proliferation marker Ki67 were expressed even after 30 days in culture. Overall, hPSC-LESCs retained their capacity to self-renew and proliferate, but were also able to terminally differentiate upon stimulation, as suggested by protein expression of cytokeratins 3 and 12. We propose the use of bioengineered collagen matrices as carriers for the clinically-relevant hPSC-derived LESC-like cells, as a novel tissue engineering approach for

  15. Human brain stem structures respond differentially to noxious heat.

    PubMed

    Ritter, Alexander; Franz, Marcel; Dietrich, Caroline; Miltner, Wolfgang H R; Weiss, Thomas

    2013-01-01

    Concerning the physiological correlates of pain, the brain stem is considered to be one core region that is activated by noxious input. In animal studies, different slopes of skin heating (SSH) with noxious heat led to activation in different columns of the midbrain periaqueductal gray (PAG). The present study aimed at finding a method for differentiating structures in PAG and other brain stem structures, which are associated with different qualities of pain in humans according to the structures that were associated with different behavioral significances to noxious thermal stimulation in animals. Brain activity was studied by functional MRI in healthy subjects in response to steep and shallow SSH with noxious heat. We found differential activation to different SSH in the PAG and the rostral ventromedial medulla (RVM). In a second experiment, we demonstrate that the different SSH were associated with different pain qualities. Our experiments provide evidence that brainstem structures, i.e., the PAG and the RVM, become differentially activated by different SSH. Therefore, different SSH can be utilized when brain stem structures are investigated and when it is aimed to activate these structures differentially. Moreover, percepts of first pain were elicited by shallow SSH whereas percepts of second pain were elicited by steep SSH. The stronger activation of these brain stem structures to SSH, eliciting percepts of second vs. first pain, might be of relevance for activating different coping strategies in response to the noxious input with the two types of SSH.

  16. Derivation of novel human ground state naive pluripotent stem cells.

    PubMed

    Gafni, Ohad; Weinberger, Leehee; Mansour, Abed AlFatah; Manor, Yair S; Chomsky, Elad; Ben-Yosef, Dalit; Kalma, Yael; Viukov, Sergey; Maza, Itay; Zviran, Asaf; Rais, Yoach; Shipony, Zohar; Mukamel, Zohar; Krupalnik, Vladislav; Zerbib, Mirie; Geula, Shay; Caspi, Inbal; Schneir, Dan; Shwartz, Tamar; Gilad, Shlomit; Amann-Zalcenstein, Daniela; Benjamin, Sima; Amit, Ido; Tanay, Amos; Massarwa, Rada; Novershtern, Noa; Hanna, Jacob H

    2013-12-12

    Mouse embryonic stem (ES) cells are isolated from the inner cell mass of blastocysts, and can be preserved in vitro in a naive inner-cell-mass-like configuration by providing exogenous stimulation with leukaemia inhibitory factor (LIF) and small molecule inhibition of ERK1/ERK2 and GSK3β signalling (termed 2i/LIF conditions). Hallmarks of naive pluripotency include driving Oct4 (also known as Pou5f1) transcription by its distal enhancer, retaining a pre-inactivation X chromosome state, and global reduction in DNA methylation and in H3K27me3 repressive chromatin mark deposition on developmental regulatory gene promoters. Upon withdrawal of 2i/LIF, naive mouse ES cells can drift towards a primed pluripotent state resembling that of the post-implantation epiblast. Although human ES cells share several molecular features with naive mouse ES cells, they also share a variety of epigenetic properties with primed murine epiblast stem cells (EpiSCs). These include predominant use of the proximal enhancer element to maintain OCT4 expression, pronounced tendency for X chromosome inactivation in most female human ES cells, increase in DNA methylation and prominent deposition of H3K27me3 and bivalent domain acquisition on lineage regulatory genes. The feasibility of establishing human ground state naive pluripotency in vitro with equivalent molecular and functional features to those characterized in mouse ES cells remains to be defined. Here we establish defined conditions that facilitate the derivation of genetically unmodified human naive pluripotent stem cells from already established primed human ES cells, from somatic cells through induced pluripotent stem (iPS) cell reprogramming or directly from blastocysts. The novel naive pluripotent cells validated herein retain molecular characteristics and functional properties that are highly similar to mouse naive ES cells, and distinct from conventional primed human pluripotent cells. This includes competence in the generation

  17. Derivation of Genea057 human embryonic stem cell line.

    PubMed

    Dumevska, Biljana; Chami, Omar; McKernan, Robert; Goel, Divya; Schmidt, Uli

    2016-01-01

    The Genea057 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated human feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XX karyotype and female allele pattern through traditional karyotyping, CGH and STR analysis. Pluripotency of Genea057 was demonstrated with 97% of cells expressing Nanog, 81% Oct4, 75% Tra1-60 and 97% SSEA4, a PluriTest Pluripotency score of 27.59 and Novelty score of 1.32. The cell line was negative for Mycoplasma and any visible contamination. PMID:27345782

  18. Derivation of Genea042 human embryonic stem cell line.

    PubMed

    Dumevska, Biljana; Chami, Omar; McKernan, Robert; Goel, Divya; Schmidt, Uli

    2016-03-01

    The Genea042 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated human feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XX karyotype and female allele pattern through traditional karyotyping, CGH and STR analysis. Pluripotency of Genea042 was demonstrated with 81% of cells expressing Nanog, 95% Oct4, 53% Tra1-60 and 97% SSEA4, a PluriTest Pluripotency score of 30.06, Novelty score of 1.24 and Alkaline Phosphatase activity. The cell line was negative for Mycoplasma and any visible contamination. PMID:27345994

  19. Derivation of Genea052 human embryonic stem cell line.

    PubMed

    Dumevska, Biljana; Chami, Omar; McKernan, Robert; Goel, Divya; Schmidt, Uli

    2016-03-01

    The Genea052 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated human feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XY karyotype and male allele pattern through CGH and STR analysis. Pluripotency of Genea052 was demonstrated with 85% of cells expressing Nanog, 87% Oct4, 60% Tra1-60 and 97% SSEA4, a PluriTest Pluripotency score of 27.21, Novelty score of 1.2 and tri-lineage teratoma formation. The cell line was negative for Mycoplasma and any visible contamination. PMID:27345996

  20. Derivation of Genea015 human embryonic stem cell line.

    PubMed

    Dumevska, Biljana; Chami, Omar; McKernan, Robert; Goel, Divya; Schmidt, Uli

    2016-03-01

    The Genea015 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated human feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XY karyotype and male Allele pattern through traditional karyotyping, CGH and STR analysis. Pluripotency of Genea015 was demonstrated with 80% of cells expressing Nanog, 97% Oct4, 75% Tra1-60 and 98% SSEA4, a PluriTest Pluripotency score of 29.52, Novelty score of 1.3 and Alkaline Phosphatase activity. The cell line was negative for Mycoplasma and any visible contamination. PMID:27346028

  1. Derivation of Genea047 human embryonic stem cell line.

    PubMed

    Dumevska, Biljana; Chami, Omar; McKernan, Robert; Goel, Divya; Schmidt, Uli

    2016-03-01

    The Genea047 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated human feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XX karyotype and female allele pattern through traditional karyotyping, CGH and STR analysis. Pluripotency of Genea047 was demonstrated with 88% of cells expressing Nanog, 95% Oct4, 59% Tra1-60 and 99% SSEA4, a PluriTest Pluripotency score of 30.86, Novelty score of 1.23 and tri-lineage teratoma formation. The cell line was negative for Mycoplasma and any visible contamination. PMID:27345995

  2. Derivation of Genea043 human embryonic stem cell line.

    PubMed

    Dumevska, Biljana; Chami, Omar; McKernan, Robert; Goel, Divya; Schmidt, Uli

    2016-01-01

    The Genea043 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated human feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XY karyotype and male allele pattern through traditional karyotyping, CGH and STR analysis. Pluripotency of Genea043 was demonstrated with 92% of cells expressing Nanog, 95% Oct4, 61% Tra1-60 and 99% SSEA4, a PluriTest Pluripotency score of 31.74, Novelty score of 1.2 and Alkaline Phosphatase activity. The cell line was negative for Mycoplasma and any visible contamination. PMID:27345801

  3. Derivation of Genea016 human embryonic stem cell line.

    PubMed

    Dumevska, Biljana; Chami, Omar; McKernan, Robert; Goel, Divya; Peura, Teija; Schmidt, Uli

    2016-01-01

    The Genea016 human embryonic stem cell line was derived from a donated, fully commercially consented ART blastocyst, through ICM outgrowth on inactivated human feeders. The line showed pluripotent cell morphology and genomic analysis verified a 46, XX karyotype and female Allele pattern through traditional karyotyping, CGH and STR analysis. Pluripotency of Genea016 was demonstrated with 77% of cells expressing Nanog, 95% Oct4, 53% Tra1-60 and 98% SSEA4, a PluriTest Pluripotency score of 28.4, Novelty score of 1.37 and Alkaline Phosphatase activity. The cell line was negative for Mycoplasma and any visible contamination. PMID:27345780

  4. Engineering human cells and tissues through pluripotent stem cells.

    PubMed

    Jones, Jeffrey R; Zhang, Su-Chun

    2016-08-01

    The utility of human pluripotent stem cells (hPSCs) depends on their ability to produce functional cells and tissues of the body. Two strategies have been developed: directed differentiation of enriched populations of cells that match a regional and functional profile and spontaneous generation of three-dimensional organoids that resemble tissues in the body. Genomic editing of hPSCs and their differentiated cells broadens the use of the hPSC paradigm in studying human cellular function and disease as well as developing therapeutics.

  5. Rethinking Adult Literacy Programs: A Humanities-Based Curriculum.

    ERIC Educational Resources Information Center

    Anania, Joanne

    The Roosevelt University Humanities Enrichment Program tries to acknowledge the adult part of adult literacy. Its instructional materials are of interest and value to the adult student and, therefore, provide incentives for reading and discussion instead of serving merely as skill-building exercises. The materials are drawn from literature,…

  6. Cripto is essential to capture mouse epiblast stem cell and human embryonic stem cell pluripotency.

    PubMed

    Fiorenzano, Alessandro; Pascale, Emilia; D'Aniello, Cristina; Acampora, Dario; Bassalert, Cecilia; Russo, Francesco; Andolfi, Gennaro; Biffoni, Mauro; Francescangeli, Federica; Zeuner, Ann; Angelini, Claudia; Chazaud, Claire; Patriarca, Eduardo J; Fico, Annalisa; Minchiotti, Gabriella

    2016-01-01

    Known molecular determinants of developmental plasticity are mainly transcription factors, while the extrinsic regulation of this process has been largely unexplored. Here we identify Cripto as one of the earliest epiblast markers and a key extracellular determinant of the naive and primed pluripotent states. We demonstrate that Cripto sustains mouse embryonic stem cell (ESC) self-renewal by modulating Wnt/β-catenin, whereas it maintains mouse epiblast stem cell (EpiSC) and human ESC pluripotency through Nodal/Smad2. Moreover, we provide unprecedented evidence that Cripto controls the metabolic reprogramming in ESCs to EpiSC transition. Remarkably, Cripto deficiency attenuates ESC lineage restriction in vitro and in vivo, and permits ESC transdifferentiation into trophectoderm lineage, suggesting that Cripto has earlier functions than previously recognized. All together, our studies provide novel insights into the current model of mammalian pluripotency and contribute to the understanding of the extrinsic regulation of the first cell lineage decision in the embryo. PMID:27586544

  7. Cripto is essential to capture mouse epiblast stem cell and human embryonic stem cell pluripotency

    PubMed Central

    Fiorenzano, Alessandro; Pascale, Emilia; D'Aniello, Cristina; Acampora, Dario; Bassalert, Cecilia; Russo, Francesco; Andolfi, Gennaro; Biffoni, Mauro; Francescangeli, Federica; Zeuner, Ann; Angelini, Claudia; Chazaud, Claire; Patriarca, Eduardo J.; Fico, Annalisa; Minchiotti, Gabriella

    2016-01-01

    Known molecular determinants of developmental plasticity are mainly transcription factors, while the extrinsic regulation of this process has been largely unexplored. Here we identify Cripto as one of the earliest epiblast markers and a key extracellular determinant of the naive and primed pluripotent states. We demonstrate that Cripto sustains mouse embryonic stem cell (ESC) self-renewal by modulating Wnt/β-catenin, whereas it maintains mouse epiblast stem cell (EpiSC) and human ESC pluripotency through Nodal/Smad2. Moreover, we provide unprecedented evidence that Cripto controls the metabolic reprogramming in ESCs to EpiSC transition. Remarkably, Cripto deficiency attenuates ESC lineage restriction in vitro and in vivo, and permits ESC transdifferentiation into trophectoderm lineage, suggesting that Cripto has earlier functions than previously recognized. All together, our studies provide novel insights into the current model of mammalian pluripotency and contribute to the understanding of the extrinsic regulation of the first cell lineage decision in the embryo. PMID:27586544

  8. Endothelial cells derived from human embryonic stem cells

    NASA Astrophysics Data System (ADS)

    Levenberg, Shulamit; Golub, Justin S.; Amit, Michal; Itskovitz-Eldor, Joseph; Langer, Robert

    2002-04-01

    Human embryonic stem cells have the potential to differentiate into various cell types and, thus, may be useful as a source of cells for transplantation or tissue engineering. We describe here the differentiation steps of human embryonic stem cells into endothelial cells forming vascular-like structures. The human embryonic-derived endothelial cells were isolated by using platelet endothelial cell-adhesion molecule-1 (PECAM1) antibodies, their behavior was characterized in vitro and in vivo, and their potential in tissue engineering was examined. We show that the isolated embryonic PECAM1+ cells, grown in culture, display characteristics similar to vessel endothelium. The cells express endothelial cell markers in a pattern similar to human umbilical vein endothelial cells, their junctions are correctly organized, and they have high metabolism of acetylated low-density lipoprotein. In addition, the cells are able to differentiate and form tube-like structures when cultured on matrigel. In vivo, when transplanted into SCID mice, the cells appeared to form microvessels containing mouse blood cells. With further studies, these cells could provide a source of human endothelial cells that could be beneficial for potential applications such as engineering new blood vessels, endothelial cell transplantation into the heart for myocardial regeneration, and induction of angiogenesis for treatment of regional ischemia.

  9. Human embryonic stem cells carrying mutations for severe genetic disorders.

    PubMed

    Frumkin, Tsvia; Malcov, Mira; Telias, Michael; Gold, Veronica; Schwartz, Tamar; Azem, Foad; Amit, Ami; Yaron, Yuval; Ben-Yosef, Dalit

    2010-04-01

    Human embryonic stem cells (HESCs) carrying specific mutations potentially provide a valuable tool for studying genetic disorders in humans. One preferable approach for obtaining these cell lines is by deriving them from affected preimplantation genetically diagnosed embryos. These unique cells are especially important for modeling human genetic disorders for which there are no adequate research models. They can be further used to gain new insights into developmentally regulated events that occur during human embryo development and that are responsible for the manifestation of genetically inherited disorders. They also have great value for the exploration of new therapeutic protocols, including gene-therapy-based treatments and disease-oriented drug screening and discovery. Here, we report the establishment of 15 different mutant human embryonic stem cell lines derived from genetically affected embryos, all donated by couples undergoing preimplantation genetic diagnosis in our in vitro fertilization unit. For further information regarding access to HESC lines from our repository, for research purposes, please email dalitb@tasmc.health.gov.il. PMID:20186514

  10. Human amniotic fluid stem cells as a model for functional studies of genes involved in human genetic diseases or oncogenesis.

    PubMed

    Rosner, Margit; Dolznig, Helmut; Schipany, Katharina; Mikula, Mario; Brandau, Oliver; Hengstschläger, Markus

    2011-09-01

    Besides their putative usage for therapies, stem cells are a promising tool for functional studies of genes involved in human genetic diseases or oncogenesis. For this purpose induced pluripotent stem (iPS) cells can be derived from patients harbouring specific mutations. In contrast to adult stem cells, iPS cells are pluripotent and can efficiently be grown in culture. However, iPS cells are modulated due to the ectopic induction of pluripotency, harbour other somatic mutations accumulated during the life span of the source cells, exhibit only imperfectly cleared epigenetic memory of the source cell, and are often genomically instable. In addition, iPS cells from patients only allow the investigation of mutations, which are not prenatally lethal. Embryonic stem (ES) cells have a high proliferation and differentiation potential, but raise ethical issues. Human embryos, which are not transferred in the course of in vitro fertilization, because of preimplantation genetic diagnosis of a genetic defect, are still rarely donated for the establishment of ES cell lines. In addition, their usage for studies on gene functions for oncogenesis is hampered by the fact the ES cells are already tumorigenic per se. In 2003 amniotic fluid stem (AFS) cells have been discovered, which meanwhile have been demonstrated to harbour the potential to differentiate into cells of all three germ layers. Monoclonal human AFS cell lines derived from amniocenteses have a high proliferative potential, are genomically stable and are not associated with ethical controversies. Worldwide amniocenteses are performed for routine human genetic diagnosis. We here discuss how generation and banking of monoclonal human AFS cell lines with specific chromosomal aberrations or monogenic disease mutations would allow to study the functional consequences of disease causing mutations. In addition, recently a protocol for efficient and highly reproducible siRNA-mediated long-term knockdown of endogenous gene

  11. Cigarette smoking hinders human periodontal ligament-derived stem cell proliferation, migration and differentiation potentials.

    PubMed

    Ng, Tsz Kin; Huang, Li; Cao, Di; Yip, Yolanda Wong-Ying; Tsang, Wai Ming; Yam, Gary Hin-Fai; Pang, Chi Pui; Cheung, Herman S

    2015-01-01

    Cigarette smoking contributes to the development of destructive periodontal diseases and delays its healing process. Our previous study demonstrated that nicotine, a major constituent in the cigarette smoke, inhibits the regenerative potentials of human periodontal ligament-derived stem cells (PDLSC) through microRNA (miRNA) regulation. In this study, we hypothesized that the delayed healing in cigarette smokers is caused by the afflicted regenerative potential of smoker PDLSC. We cultured PDLSC from teeth extracted from smokers and non-smokers. In smoker PDLSC, we found significantly reduced proliferation rate and retarded migration capabilities. Moreover, alkaline phosphatase activity, calcium deposition and acidic polysaccharide staining were reduced after BMP2-induced differentiation. In contrast, more lipid deposition was observed in adipogenic-induced smoker PDLSC. Furthermore, two nicotine-related miRNAs, hsa-miR-1305 (22.08 folds, p = 0.040) and hsa-miR-18b (15.56 folds, p = 0.018), were significantly upregulated in smoker PDLSC, suggesting these miRNAs might play an important role in the deteriorative effects on stem cells by cigarette smoke. Results of this study provide further evidences that cigarette smoking affects the regenerative potentials of human adult stem cells.

  12. Effect of Human Adipose Tissue Mesenchymal Stem Cells on the Regeneration of Ovine Articular Cartilage.

    PubMed

    Zorzi, Alessandro R; Amstalden, Eliane M I; Plepis, Ana Maria G; Martins, Virginia C A; Ferretti, Mario; Antonioli, Eliane; Duarte, Adriana S S; Luzo, Angela C M; Miranda, João B

    2015-11-09

    Cell therapy is a promising approach to improve cartilage healing. Adipose tissue is an abundant and readily accessible cell source. Previous studies have demonstrated good cartilage repair results with adipose tissue mesenchymal stem cells in small animal experiments. This study aimed to examine these cells in a large animal model. Thirty knees of adult sheep were randomly allocated to three treatment groups: CELLS (scaffold seeded with human adipose tissue mesenchymal stem cells), SCAFFOLD (scaffold without cells), or EMPTY (untreated lesions). A partial thickness defect was created in the medial femoral condyle. After six months, the knees were examined according to an adaptation of the International Cartilage Repair Society (ICRS 1) score, in addition to a new Partial Thickness Model scale and the ICRS macroscopic score. All of the animals completed the follow-up period. The CELLS group presented with the highest ICRS 1 score (8.3 ± 3.1), followed by the SCAFFOLD group (5.6 ± 2.2) and the EMPTY group (5.2 ± 2.4) (p = 0.033). Other scores were not significantly different. These results suggest that human adipose tissue mesenchymal stem cells promoted satisfactory cartilage repair in the ovine model.

  13. Label-free nonlinear optical microscopy detects early markers for osteogenic differentiation of human stem cells

    NASA Astrophysics Data System (ADS)

    Hofemeier, Arne D.; Hachmeister, Henning; Pilger, Christian; Schürmann, Matthias; Greiner, Johannes F. W.; Nolte, Lena; Sudhoff, Holger; Kaltschmidt, Christian; Huser, Thomas; Kaltschmidt, Barbara

    2016-05-01

    Tissue engineering by stem cell differentiation is a novel treatment option for bone regeneration. Most approaches for the detection of osteogenic differentiation are invasive or destructive and not compatible with live cell analysis. Here, non-destructive and label-free approaches of Raman spectroscopy, coherent anti-Stokes Raman scattering (CARS) and second harmonic generation (SHG) microscopy were used to detect and image osteogenic differentiation of human neural crest-derived inferior turbinate stem cells (ITSCs). Combined CARS and SHG microscopy was able to detect markers of osteogenesis within 14 days after osteogenic induction. This process increased during continued differentiation. Furthermore, Raman spectroscopy showed significant increases of the PO43‑ symmetric stretch vibrations at 959 cm‑1 assigned to calcium hydroxyapatite between days 14 and 21. Additionally, CARS microscopy was able to image calcium hydroxyapatite deposits within 14 days following osteogenic induction, which was confirmed by Alizarin Red-Staining and RT- PCR. Taken together, the multimodal label-free analysis methods Raman spectroscopy, CARS and SHG microscopy can monitor osteogenic differentiation of adult human stem cells into osteoblasts with high sensitivity and spatial resolution in three dimensions. Our findings suggest a great potential of these optical detection methods for clinical applications including in vivo observation of bone tissue–implant-interfaces or disease diagnosis.

  14. Effect of Human Adipose Tissue Mesenchymal Stem Cells on the Regeneration of Ovine Articular Cartilage

    PubMed Central

    Zorzi, Alessandro R.; Amstalden, Eliane M. I.; Plepis, Ana Maria G.; Martins, Virginia C. A.; Ferretti, Mario; Antonioli, Eliane; Duarte, Adriana S. S.; Luzo, Angela C. M.; Miranda, João B.

    2015-01-01

    Cell therapy is a promising approach to improve cartilage healing. Adipose tissue is an abundant and readily accessible cell source. Previous studies have demonstrated good cartilage repair results with adipose tissue mesenchymal stem cells in small animal experiments. This study aimed to examine these cells in a large animal model. Thirty knees of adult sheep were randomly allocated to three treatment groups: CELLS (scaffold seeded with human adipose tissue mesenchymal stem cells), SCAFFOLD (scaffold without cells), or EMPTY (untreated lesions). A partial thickness defect was created in the medial femoral condyle. After six months, the knees were examined according to an adaptation of the International Cartilage Repair Society (ICRS 1) score, in addition to a new Partial Thickness Model scale and the ICRS macroscopic score. All of the animals completed the follow-up period. The CELLS group presented with the highest ICRS 1 score (8.3 ± 3.1), followed by the SCAFFOLD group (5.6 ± 2.2) and the EMPTY group (5.2 ± 2.4) (p = 0.033). Other scores were not significantly different. These results suggest that human adipose tissue mesenchymal stem cells promoted satisfactory cartilage repair in the ovine model. PMID:26569221

  15. Glycosaminoglycans enhance osteoblast differentiation of bone marrow derived human mesenchymal stem cells.

    PubMed

    Mathews, Smitha; Mathew, Suja Ann; Gupta, Pawan Kumar; Bhonde, Ramesh; Totey, Satish

    2014-02-01

    Extracellular matrix plays an important role in regulating cell growth and differentiation. The biomimetic approach of cell-based tissue engineering is based on mirroring this in vivo micro environment for developing a functional tissue engineered construct. In this study, we treated normal tissue culture plates with selected extracellular matrix components consisting of glycosaminoglycans such as chondroitin-4-sulphate, dermatan sulphate, chondroitin-6-sulphate, heparin and hyaluronic acid. Mesenchymal stem cells isolated from adult human bone marrow were cultured on the glycosaminoglycan treated culture plates to evaluate their regulatory role in cell growth and osteoblast differentiation. Although no significant improvement on human mesenchymal stem cell adhesion and proliferation was observed on the glycosaminoglycan-treated tissue culture plates, there was selective osteoblast differentiation, indicating its potential role in differentiation rather than proliferation. Osteoblast differentiation studies showed high osteogenic potential for all tested glycosaminoglycans except chondroitin-4-sulphate. Osteoblast differentiation-associated genes such as osterix, osteocalcin, integrin binding sialoprotein, osteonectin and collagen, type 1, alpha 1 showed significant upregulation. We identified osterix as the key transcription factor responsible for the enhanced bone matrix deposition observed on hyaluronic acid, heparin and chondroitin-6-sulphate. Hyaluronic acid provided the most favourable condition for osteoblast differentiation and bone matrix synthesis. Our results confirm and emphasise the significant role of extracellular matrix in regulating cell differentiation. To summarise, glycosaminoglycans of extracellular matrix played a significant role in regulating osteoblast differentiation and could be exploited in the biomimetic approach of fabricating or functionalizing scaffolds for stem cell based bone tissue engineering.

  16. Label-free nonlinear optical microscopy detects early markers for osteogenic differentiation of human stem cells

    PubMed Central

    Hofemeier, Arne D.; Hachmeister, Henning; Pilger, Christian; Schürmann, Matthias; Greiner, Johannes F. W.; Nolte, Lena; Sudhoff, Holger; Kaltschmidt, Christian; Huser, Thomas; Kaltschmidt, Barbara

    2016-01-01

    Tissue engineering by stem cell differentiation is a novel treatment option for bone regeneration. Most approaches for the detection of osteogenic differentiation are invasive or destructive and not compatible with live cell analysis. Here, non-destructive and label-free approaches of Raman spectroscopy, coherent anti-Stokes Raman scattering (CARS) and second harmonic generation (SHG) microscopy were used to detect and image osteogenic differentiation of human neural crest-derived inferior turbinate stem cells (ITSCs). Combined CARS and SHG microscopy was able to detect markers of osteogenesis within 14 days after osteogenic induction. This process increased during continued differentiation. Furthermore, Raman spectroscopy showed significant increases of the PO43− symmetric stretch vibrations at 959 cm−1 assigned to calcium hydroxyapatite between days 14 and 21. Additionally, CARS microscopy was able to image calcium hydroxyapatite deposits within 14 days following osteogenic induction, which was confirmed by Alizarin Red-Staining and RT- PCR. Taken together, the multimodal label-free analysis methods Raman spectroscopy, CARS and SHG microscopy can monitor osteogenic differentiation of adult human stem cells into osteoblasts with high sensitivity and spatial resolution in three dimensions. Our findings suggest a great potential of these optical detection methods for clinical applications including in vivo observation of bone tissue–implant-interfaces or disease diagnosis. PMID:27225821

  17. Effects of addictive drugs on adult neural stem/progenitor cells.

    PubMed

    Xu, Chi; Loh, Horace H; Law, Ping-Yee

    2016-01-01

    Neural stem/progenitor cells (NSPCs) undergo a series of developmental processes before giving rise to newborn neurons, astrocytes and oligodendrocytes in adult neurogenesis. During the past decade, the role of NSPCs has been highlighted by studies on adult neurogenesis modulated by addictive drugs. It has been proven that these drugs regulate the proliferation, differentiation and survival of adult NSPCs in different manners, which results in the varying consequences of adult neurogenesis. The effects of addictive drugs on NSPCs are exerted via a variety of different mechanisms and pathways, which interact with one another and contribute to the complexity of NSPC regulation. Here, we review the effects of different addictive drugs on NSPCs, and the related experimental methods and paradigms. We also discuss the current understanding of major signaling molecules, especially the putative common mechanisms, underlying such effects. Finally, we review the future directions of research in this area. PMID:26468052

  18. Differentiated cells are more efficient than adult stem cells for cloning by somatic cell nuclear transfer.

    PubMed

    Sung, Li-Ying; Gao, Shaorong; Shen, Hongmei; Yu, Hui; Song, Yifang; Smith, Sadie L; Chang, Ching-Chien; Inoue, Kimiko; Kuo, Lynn; Lian, Jin; Li, Ao; Tian, X Cindy; Tuck, David P; Weissman, Sherman M; Yang, Xiangzhong; Cheng, Tao

    2006-11-01

    Since the creation of Dolly via somatic cell nuclear transfer (SCNT), more than a dozen species of mammals have been cloned using this technology. One hypothesis for the limited success of cloning via SCNT (1%-5%) is that the clones are likely to be derived from adult stem cells. Support for this hypothesis comes from the findings that the reproductive cloning efficiency for embryonic stem cells is five to ten times higher than that for somatic cells as donors and that cloned pups cannot be produced directly from cloned embryos derived from differentiated B and T cells or neuronal cells. The question remains as to whether SCNT-derived animal clones can be derived from truly differentiated somatic cells. We tested this hypothesis with mouse hematopoietic cells at different differentiation stages: hematopoietic stem cells, progenitor cells and granulocytes. We found that cloning efficiency increases over the differentiation hierarchy, and terminally differentiated postmitotic granulocytes yield cloned pups with the greatest cloning efficiency.

  19. Concise review: programming human pluripotent stem cells into blood.

    PubMed

    Easterbrook, Jennifer; Fidanza, Antonella; Forrester, Lesley M

    2016-06-01

    Blood disorders are treated with cell therapies including haematopoietic stem cell (HSC) transplantation as well as platelet and red blood cell transfusions. However the source of cells is entirely dependent on donors, procedures are susceptible to transfusion-transmitted infections and serious complications can arise in recipients due to immunological incompatibility. These problems could be alleviated if it was possible to produce haematopoietic cells in vitro from an autologous and renewable cell source. The production of haematopoietic cells in the laboratory from human induced pluripotent stem cells (iPSCs) may provide a route to realize this goal but it has proven challenging to generate long-term reconstituting HSCs. To date, the optimization of differentiation protocols has mostly relied on the manipulation of extrinsic signals to mimic the in vivo environment. We review studies that have taken an alternative approach to modulate intrinsic signals by enforced expression of transcription factors. Single and combinations of multiple transcription factors have been used in a variety of contexts to enhance the production of haematopoietic cells from human pluripotent stem cells. This programming approach, together with the recent advances in the production and use of synthetic transcription factors, holds great promise for the production of fully functional HSCs in the future.

  20. hPSCreg--the human pluripotent stem cell registry.

    PubMed

    Seltmann, Stefanie; Lekschas, Fritz; Müller, Robert; Stachelscheid, Harald; Bittner, Marie-Sophie; Zhang, Weiping; Kidane, Luam; Seriola, Anna; Veiga, Anna; Stacey, Glyn; Kurtz, Andreas

    2016-01-01

    The human pluripotent stem cell registry (hPSCreg), accessible at http://hpscreg.eu, is a public registry and data portal for human embryonic and induced pluripotent stem cell lines (hESC and hiPSC). Since their first isolation the number of hESC lines has steadily increased to over 3000 and new iPSC lines are generated in a rapidly growing number of laboratories as a result of their potentially broad applicability in biomedicine and drug testing. Many of these lines are deposited in stem cell banks, which are globally established to store tens of thousands of lines from healthy and diseased donors. The Registry provides comprehensive and standardized biological and legal information as well as tools to search and compare information from multiple hPSC sources and hence addresses a translational research need. To facilitate unambiguous identification over different resources, hPSCreg automatically creates a unique standardized name for each cell line registered. In addition to biological information, hPSCreg stores extensive data about ethical standards regarding cell sourcing and conditions for application and privacy protection. hPSCreg is the first global registry that holds both, manually validated scientific and ethical information on hPSC lines, and provides access by means of a user-friendly, mobile-ready web application. PMID:26400179

  1. hPSCreg—the human pluripotent stem cell registry

    PubMed Central

    Seltmann, Stefanie; Lekschas, Fritz; Müller, Robert; Stachelscheid, Harald; Bittner, Marie-Sophie; Zhang, Weiping; Kidane, Luam; Seriola, Anna; Veiga, Anna; Stacey, Glyn; Kurtz, Andreas

    2016-01-01

    The human pluripotent stem cell registry (hPSCreg), accessible at http://hpscreg.eu, is a public registry and data portal for human embryonic and induced pluripotent stem cell lines (hESC and hiPSC). Since their first isolation the number of hESC lines has steadily increased to over 3000 and new iPSC lines are generated in a rapidly growing number of laboratories as a result of their potentially broad applicability in biomedicine and drug testing. Many of these lines are deposited in stem cell banks, which are globally established to store tens of thousands of lines from healthy and diseased donors. The Registry provides comprehensive and standardized biological and legal information as well as tools to search and compare information from multiple hPSC sources and hence addresses a translational research need. To facilitate unambiguous identification over different resources, hPSCreg automatically creates a unique standardized name for each cell line registered. In addition to biological information, hPSCreg stores extensive data about ethical standards regarding cell sourcing and conditions for application and privacy protection. hPSCreg is the first global registry that holds both, manually validated scientific and ethical information on hPSC lines, and provides access by means of a user-friendly, mobile-ready web application. PMID:26400179

  2. Concise review: programming human pluripotent stem cells into blood.

    PubMed

    Easterbrook, Jennifer; Fidanza, Antonella; Forrester, Lesley M

    2016-06-01

    Blood disorders are treated with cell therapies including haematopoietic stem cell (HSC) transplantation as well as platelet and red blood cell transfusions. However the source of cells is entirely dependent on donors, procedures are susceptible to transfusion-transmitted infections and serious complications can arise in recipients due to immunological incompatibility. These problems could be alleviated if it was possible to produce haematopoietic cells in vitro from an autologous and renewable cell source. The production of haematopoietic cells in the laboratory from human induced pluripotent stem cells (iPSCs) may provide a route to realize this goal but it has proven challenging to generate long-term reconstituting HSCs. To date, the optimization of differentiation protocols has mostly relied on the manipulation of extrinsic signals to mimic the in vivo environment. We review studies that have taken an alternative approach to modulate intrinsic signals by enforced expression of transcription factors. Single and combinations of multiple transcription factors have been used in a variety of contexts to enhance the production of haematopoietic cells from human pluripotent stem cells. This programming approach, together with the recent advances in the production and use of synthetic transcription factors, holds great promise for the production of fully functional HSCs in the future. PMID:26996518

  3. Have you got any cholesterol? Adults' views of human nutrition

    NASA Astrophysics Data System (ADS)

    Schibeci, Renato; Wong, Khoon Yoong

    1994-12-01

    The general aim of our human nutrition project is to develop a health education model grounded in ‘everyday’ or ‘situated’ cognition (Hennessey, 1993). In 1993, we began pilot work to document adult understanding of human nutrition. We used a HyperCard stack as the basis for a series of interviews with 50 adults (25 university students, and 25 adults from offcampus). The interviews were transcribed and analysed using the NUDIST computer program. A summary of the views of these 50 adults on selected aspects of human nutrition is presented in this paper.

  4. Induction of Human Embryonic and Induced Pluripotent Stem Cells Into Urothelium

    PubMed Central

    Osborn, Stephanie L.; Thangappan, Ravikumar; Luria, Ayala; Lee, Justin H.; Nolta, Jan

    2014-01-01

    In vitro generation of human urothelium from stem cells would be a major advancement in the regenerative medicine field, providing alternate nonurologic and/or nonautologous tissue sources for bladder grafts. Such a model would also help decipher the mechanisms of urothelial differentiation and would facilitate investigation of deviated differentiation of normal progenitors into urothelial cancer stem cells, perhaps elucidating areas of intervention for improved treatments. Thus far, in vitro derivation of urothelium from human embryonic stem cells (hESCs) or human induced pluripotent stem (hiPS) cells has not been reported. The goal of this work was to develop an efficient in vitro protocol for the induction of hESCs into urothelium through an intermediary definitive endoderm step and free of matrices and cell contact. During directed differentiation in a urothelial-specific medium (“Uromedium”), hESCs produced up to 60% urothelium, as determined by uroplakin expression; subsequent propagation selected for 90% urothelium. Alteration of the epithelial and mesenchymal cell signaling contribution through noncell contact coculture or conditioned media did not enhance the production of urothelium. Temporospatial evaluation of transcription factors known to be involved in urothelial specification showed association of IRF1, GET1, and GATA4 with uroplakin expression. Additional hESC and hiPS cell lines could also be induced into urothelium using this in vitro system. These results demonstrate that derivation and propagation of urothelium from hESCs and hiPS cells can be efficiently accomplished in vitro in the absence of matrices, cell contact, or adult cell signaling and that the induction process appears to mimic normal differentiation. PMID:24657961

  5. Sensitive Tumorigenic Potential Evaluation of Adult Human Multipotent Neural Cells Immortalized by hTERT Gene Transduction

    PubMed Central

    Jeong, Da Eun; Kim, Sung Soo; Song, Hye Jin; Pyeon, Hee Jang; Kang, Kyeongjin; Hong, Seung-Cheol; Nam, Do-Hyun; Joo, Kyeung Min

    2016-01-01

    Stem cells and therapeutic genes are emerging as a new therapeutic approach to treat various neurodegenerative diseases with few effective treatment options. However, potential formation of tumors by stem cells has hampered their clinical application. Moreover, adequate preclinical platforms to precisely test tumorigenic potential of stem cells are controversial. In this study, we compared the sensitivity of various animal models for in vivo stem cell tumorigenicity testing to identify the most sensitive platform. Then, tumorigenic potential of adult human multipotent neural cells (ahMNCs) immortalized by the human telomerase reverse transcriptase (hTERT) gene was examined as a stem cell model with therapeutic genes. When human glioblastoma (GBM) cells were injected into adult (4–6-week-old) Balb/c-nu, adult NOD/SCID, adult NOG, or neonate (1–2-week-old) NOG mice, the neonate NOG mice showed significantly faster tumorigenesis than that of the other groups regardless of intracranial or subcutaneous injection route. Two kinds of ahMNCs (682TL and 779TL) were primary cultured from surgical samples of patients with temporal lobe epilepsy. Although the ahMNCs were immortalized by lentiviral hTERT gene delivery (hTERT-682TL and hTERT-779TL), they did not form any detectable masses, even in the most sensitive neonate NOG mouse platform. Moreover, the hTERT-ahMNCs had no gross chromosomal abnormalities on a karyotype analysis. Taken together, our data suggest that neonate NOG mice could be a sensitive animal platform to test tumorigenic potential of stem cell therapeutics and that ahMNCs could be a genetically stable stem cell source with little tumorigenic activity to develop regenerative treatments for neurodegenerative diseases. PMID:27391353

  6. Sensitive Tumorigenic Potential Evaluation of Adult Human Multipotent Neural Cells Immortalized by hTERT Gene Transduction.

    PubMed

    Lee, Kee Hang; Nam, Hyun; Jeong, Da Eun; Kim, Sung Soo; Song, Hye Jin; Pyeon, Hee Jang; Kang, Kyeongjin; Hong, Seung-Cheol; Nam, Do-Hyun; Joo, Kyeung Min

    2016-01-01

    Stem cells and therapeutic genes are emerging as a new therapeutic approach to treat various neurodegenerative diseases with few effective treatment options. However, potential formation of tumors by stem cells has hampered their clinical application. Moreover, adequate preclinical platforms to precisely test tumorigenic potential of stem cells are controversial. In this study, we compared the sensitivity of various animal models for in vivo stem cell tumorigenicity testing to identify the most sensitive platform. Then, tumorigenic potential of adult human multipotent neural cells (ahMNCs) immortalized by the human telomerase reverse transcriptase (hTERT) gene was examined as a stem cell model with therapeutic genes. When human glioblastoma (GBM) cells were injected into adult (4-6-week-old) Balb/c-nu, adult NOD/SCID, adult NOG, or neonate (1-2-week-old) NOG mice, the neonate NOG mice showed significantly faster tumorigenesis than that of the other groups regardless of intracranial or subcutaneous injection route. Two kinds of ahMNCs (682TL and 779TL) were primary cultured from surgical samples of patients with temporal lobe epilepsy. Although the ahMNCs were immortalized by lentiviral hTERT gene delivery (hTERT-682TL and hTERT-779TL), they did not form any detectable masses, even in the most sensitive neonate NOG mouse platform. Moreover, the hTERT-ahMNCs had no gross chromosomal abnormalities on a karyotype analysis. Taken together, our data suggest that neonate NOG mice could be a sensitive animal platform to test tumorigenic potential of stem cell therapeutics and that ahMNCs could be a genetically stable stem cell source with little tumorigenic activity to develop regenerative treatments for neurodegenerative diseases. PMID:27391353

  7. Rigid microenvironments promote cardiac differentiation of mouse and human embryonic stem cells

    NASA Astrophysics Data System (ADS)

    Arshi, Armin; Nakashima, Yasuhiro; Nakano, Haruko; Eaimkhong, Sarayoot; Evseenko, Denis; Reed, Jason; Stieg, Adam Z.; Gimzewski, James K.; Nakano, Atsushi

    2013-04-01

    While adult heart muscle is the least regenerative of tissues, embryonic cardiomyocytes are proliferative, with embryonic stem (ES) cells providing an endless reservoir. In addition to secreted factors and cell-cell interactions, the extracellular microenvironment has been shown to play an important role in stem cell lineage specification, and understanding how scaffold elasticity influences cardiac differentiation is crucial to cardiac tissue engineering. Though previous studies have analyzed the role of matrix elasticity on the function of differentiated cardiomyocytes, whether it affects the induction of cardiomyocytes from pluripotent stem cells is poorly understood. Here, we examine the role of matrix rigidity on cardiac differentiation using mouse and human ES cells. Culture on polydimethylsiloxane (PDMS) substrates of varied monomer-to-crosslinker ratios revealed that rigid extracellular matrices promote a higher yield of de novo cardiomyocytes from undifferentiated ES cells. Using a genetically modified ES system that allows us to purify differentiated cardiomyocytes by drug selection, we demonstrate that rigid environments induce higher cardiac troponin T expression, beating rate of foci, and expression ratio of adult α- to fetal β- myosin heavy chain in a purified cardiac population. M-mode and mechanical interferometry image analyses demonstrate that these ES-derived cardiomyocytes display functional maturity and synchronization of beating when co-cultured with neonatal cardiomyocytes harvested from a developing embryo. Together, these data identify matrix stiffness as an independent factor that instructs not only the maturation of already differentiated cardiomyocytes but also the induction and proliferation of cardiomyocytes from undifferentiated progenitors. Manipulation of the stiffness will help direct the production of functional cardiomyocytes en masse from stem cells for regenerative medicine purposes.

  8. Rigid microenvironments promote cardiac differentiation of mouse and human embryonic stem cells

    PubMed Central

    Arshi, Armin; Nakashima, Yasuhiro; Nakano, Haruko; Eaimkhong, Sarayoot; Evseenko, Denis; Reed, Jason; Stieg, Adam Z.; Gimzewski, James K.; Nakano, Atsushi

    2013-01-01

    While adult heart muscle is the least regenerative of tissues, embryonic cardiomyocytes are proliferative, with embryonic stem (ES) cells providing an endless reservoir. In addition to secreted factors and cell-cell interactions, the extracellular microenvironment has been shown to play an important role in stem cell lineage specification, and understanding how scaffold elasticity influences cardiac differentiation is crucial to cardiac tissue engineering. Though previous studies have analyzed the role of the matrix elasticity on the function of differentiated cardiomyocytes, whether it affects the induction of cardiomyocytes from pluripotent stem cells is poorly understood. Here, we examined the role of matrix rigidity on the cardiac differentiation using mouse and human ES cells. Culture on polydimethylsiloxane (PDMS) substrates of varied monomer-to-crosslinker ratios revealed that rigid extracellular matrices promote a higher yield of de novo cardiomyocytes from undifferentiated ES cells. Using an genetically modified ES system that allows us to purify differentiated cardiomyocytes by drug selection, we demonstrate that rigid environments induce higher cardiac troponin T expression, beating rate of foci, and expression ratio of adult α- to fetal β- myosin heavy chain in a purified cardiac population. M-mode and mechanical interferometry image analyses demonstrate that these ES-derived cardiomyocytes display functional maturity and synchronization of beating when co-cultured with neonatal cardiomyocytes harvested from a developing embryo. Together, these data identify matrix stiffness as an independent factor that instructs not only the maturation of the already differentiated cardiomyocytes but also the induction and proliferation of cardiomyocytes from undifferentiated progenitors. Manipulation of the stiffness will help direct the production of functional cardiomyocytes en masse from stem cells for regenerative medicine purposes. PMID:24311969

  9. A model utilizing adult murine stem cells for creation of personalized islets for transplantation.

    PubMed

    Wang, J; Song, L J; Gerber, D A; Fair, J H; Rice, L; LaPaglia, M; Andreoni, K A

    2004-05-01

    Clinical islet cell transplantation has demonstrated great promise for diabetes treatment. Two major obstacles are the organ donor shortage and the immunoresponse. The purpose of this study was to create a model using the patient's own adult stem cell sources, possibly in combination with non-self cells, such as pancreatic, hepatic, or embryonic stem cells, to create "personalized" islets. We hypothesize that the reconstructed islets have the normal capability to produce insulin and glucagon with reduced immunoresponses after transplantation. Stem cells are a proliferating population of master cells that have the ability for self-renewal and multilineage differentiation. The recently developed photolithograph-based, biologic, microelectromechanic system (BioMEMS) technique supplies a useful tool for biomedical applications. Our lab has developed a novel method that integrates the adult stem cell and BioMEMS to reconstruct personalized islets. We selected islet-derived progenitor cells (IPC) for repairing and reconstructing STZ-diabetic islets. A6(+)/PYY(+) or A6(+)/ngn3(+) cells were selected to manipulate the neoislets. After 3 to 4 weeks in culture, the reconstructed cells formed islet-like clusters containing insulin or glucagon producing cells. The pilot results showed the ability of these reconstructed islets to correct hyperglycemia when transplanted into a STZ-diabetic isograft mouse model. Although several technical problems remain with the mouse model, namely, the difficulty to collect enough islets from a single mouse because of animal size, the mouse isograft model is suitable for personalized islet development.

  10. PDGFRα demarcates the cardiogenic clonogenic Sca1+ stem/progenitor cell in adult murine myocardium.

    PubMed

    Noseda, Michela; Harada, Mutsuo; McSweeney, Sara; Leja, Thomas; Belian, Elisa; Stuckey, Daniel J; Abreu Paiva, Marta S; Habib, Josef; Macaulay, Iain; de Smith, Adam J; al-Beidh, Farah; Sampson, Robert; Lumbers, R Thomas; Rao, Pulivarthi; Harding, Sian E; Blakemore, Alexandra I F; Jacobsen, Sten Eirik; Barahona, Mauricio; Schneider, Michael D

    2015-05-18

    Cardiac progenitor/stem cells in adult hearts represent an attractive therapeutic target for heart regeneration, though (inter)-relationships among reported cells remain obscure. Using single-cell qRT-PCR and clonal analyses, here we define four subpopulations of cardiac progenitor/stem cells in adult mouse myocardium all sharing stem cell antigen-1 (Sca1), based on side population (SP) phenotype, PECAM-1 (CD31) and platelet-derived growth factor receptor-α (PDGFRα) expression. SP status predicts clonogenicity and cardiogenic gene expression (Gata4/6, Hand2 and Tbx5/20), properties segregating more specifically to PDGFRα(+) cells. Clonal progeny of single Sca1(+) SP cells show cardiomyocyte, endothelial and smooth muscle lineage potential after cardiac grafting, augmenting cardiac function although durable engraftment is rare. PDGFRα(-) cells are characterized by Kdr/Flk1, Cdh5, CD31 and lack of clonogenicity. PDGFRα(+)/CD31(-) cells derive from cells formerly expressing Mesp1, Nkx2-5, Isl1, Gata5 and Wt1, distinct from PDGFRα(-)/CD31(+) cells (Gata5 low; Flk1 and Tie2 high). Thus, PDGFRα demarcates the clonogenic cardiogenic Sca1(+) stem/progenitor cell.

  11. Spontaneous transformation of adult mesenchymal stem cells from cynomolgus macaques in vitro

    SciTech Connect

    Ren, Zhenhua; Wang, Jiayin; Zhu, Wanwan; Guan, Yunqian; Zou, Chunlin; Chen, Zhiguo; Zhang, Y. Alex

    2011-12-10

    Mesenchymal stem cells (MSCs) have shown potential clinical utility in cell therapy and tissue engineering, due to their ability to proliferate as well as to differentiate into multiple lineages, including osteogenic, adipogenic, and chondrogenic specifications. Therefore, it is crucial to assess the safety of MSCs while extensive expansion ex vivo is a prerequisite to obtain the cell numbers for cell transplantation. Here we show that MSCs derived from adult cynomolgus monkey can undergo spontaneous transformation following in vitro culture. In comparison with MSCs, the spontaneously transformed mesenchymal cells (TMCs) display significantly different growth pattern and morphology, reminiscent of the characteristics of tumor cells. Importantly, TMCs are highly tumorigenic, causing subcutaneous tumors when injected into NOD/SCID mice. Moreover, no multiple differentiation potential of TMCs is observed in vitro or in vivo, suggesting that spontaneously transformed adult stem cells may not necessarily turn into cancer stem cells. These data indicate a direct transformation of cynomolgus monkey MSCs into tumor cells following long-term expansion in vitro. The spontaneous transformation of the cultured cynomolgus monkey MSCs may have important implications for ongoing clinical trials and for models of oncogenesis, thus warranting a more strict assessment of MSCs prior to cell therapy. -- Highlights: Black-Right-Pointing-Pointer Spontaneous transformation of cynomolgus monkey MSCs in vitro. Black-Right-Pointing-Pointer Transformed mesenchymal cells lack multipotency. Black-Right-Pointing-Pointer Transformed mesenchymal cells are highly tumorigenic. Black-Right-Pointing-Pointer Transformed mesenchymal cells do not have the characteristics of cancer stem cells.

  12. Recents patents for isolating, delivering and tracking adult stem cells in regenerative medicine.

    PubMed

    Fierabracci, Alessandra

    2010-06-01

    The field of regenerative medicine offers nowadays the potential to significantly impact a wide spectrum of healthcare issues, from insulin-dependent diabetes mellitus (Type 1 diabetes, T1D) to cardiovascular disease. In tissue engineering biomaterials, biological factors, regeneration competent cells are used in the process of creating functional tissue. Regarding the type of stem or progenitor cells which represents the best candidate for therapy, embryonic stem cells have been considered the master cells capable of differentiating into every type of cells either in vitro or in vivo, in spite of serious ethical concerns. Nevertheless experimental evidence suggests that adult stem cells and even terminally differentiated somatic cells under appropriate microenvironmental treatments can be reprogrammed and contribute to a much wider spectrum of differentiated progeny than previously anticipated. One of the main goals is to exploit novel technologies aiming to isolate, expand and enrich sources of regeneration competent cells, especially adult somatic stem cells. Researchers are also trying to develop innovative strategies for effectively delivering regenerative cell populations and to implement 'tracking' tools to verify their engraftment and destiny in vivo. Here we review recent patents on the field issued over the past five years.

  13. PDGFRα demarcates the cardiogenic clonogenic Sca1+ stem/progenitor cell in adult murine myocardium

    PubMed Central

    Noseda, Michela; Harada, Mutsuo; McSweeney, Sara; Leja, Thomas; Belian, Elisa; Stuckey, Daniel J.; Abreu Paiva, Marta S.; Habib, Josef; Macaulay, Iain; de Smith, Adam J.; al-Beidh, Farah; Sampson, Robert; Lumbers, R. Thomas; Rao, Pulivarthi; Harding, Sian E.; Blakemore, Alexandra I. F.; Eirik Jacobsen, Sten; Barahona, Mauricio; Schneider, Michael D.

    2015-01-01

    Cardiac progenitor/stem cells in adult hearts represent an attractive therapeutic target for heart regeneration, though (inter)-relationships among reported cells remain obscure. Using single-cell qRT–PCR and clonal analyses, here we define four subpopulations of cardiac progenitor/stem cells in adult mouse myocardium all sharing stem cell antigen-1 (Sca1), based on side population (SP) phenotype, PECAM-1 (CD31) and platelet-derived growth factor receptor-α (PDGFRα) expression. SP status predicts clonogenicity and cardiogenic gene expression (Gata4/6, Hand2 and Tbx5/20), properties segregating more specifically to PDGFRα+ cells. Clonal progeny of single Sca1+ SP cells show cardiomyocyte, endothelial and smooth muscle lineage potential after cardiac grafting, augmenting cardiac function although durable engraftment is rare. PDGFRα− cells are characterized by Kdr/Flk1, Cdh5, CD31 and lack of clonogenicity. PDGFRα+/CD31− cells derive from cells formerly expressing Mesp1, Nkx2-5, Isl1, Gata5 and Wt1, distinct from PDGFRα−/CD31+ cells (Gata5 low; Flk1 and Tie2 high). Thus, PDGFRα demarcates the clonogenic cardiogenic Sca1+ stem/progenitor cell. PMID:25980517

  14. Autologous Transplantation of Bone Marrow Adult Stem Cells for the Treatment of Idiopathic Dilated Cardiomyopathy

    PubMed Central

    Westphal, Ricardo João; Bueno, Ronaldo Rocha Loures; Galvão, Paulo Bezerra de Araújo; Zanis Neto, José; Souza, Juliano Mendes; Guérios, Ênio Eduardo; Senegaglia, Alexandra Cristina; Brofman, Paulo Roberto; Pasquini, Ricardo; da Cunha, Claudio Leinig Pereira

    2014-01-01

    Background Morbimortality in patients with dilated idiopathic cardiomyopathy is high, even under optimal medical treatment. Autologous infusion of bone marrow adult stem cells has shown promising preliminary results in these patients. Objective Determine the effectiveness of autologous transplantation of bone marrow adult stem cells on systolic and diastolic left ventricular function, and on the degree of mitral regurgitation in patients with dilated idiopathic cardiomyopathy in functional classes NYHA II and III. Methods We administered 4,54 x 108 ± 0,89 x 108 bone marrow adult stem cells into the coronary arteries of 24 patients with dilated idiopathic cardiomyopathy in functional classes NYHA II and III. Changes in functional class, systolic and diastolic left ventricular function and degree of mitral regurgitation were assessed after 3 months, 6 months and 1 year. Results During follow-up, six patients (25%) improved functional class and eight (33.3%) kept stable. Left ventricular ejection fraction improved 8.9%, 9.7% e 13.6%, after 3, 6 and 12 months (p = 0.024; 0.017 and 0.018), respectively. There were no significant changes neither in diastolic left ventricular function nor in mitral regurgitation degree. A combined cardiac resynchronization and implantable cardioversion defibrillation was implanted in two patients (8.3%). Four patients (16.6%) had sudden death and four patients died due to terminal cardiac failure. Average survival of these eight patients was 2.6 years. Conclusion Intracoronary infusion of bone marrow adult stem cells was associated with an improvement or stabilization of functional class and an improvement in left ventricular ejection fraction, suggesting the efficacy of this intervention. There were no significant changes neither in left ventricular diastolic function nor in the degree of mitral regurgitation. PMID:25590932

  15. Regeneration of plantlets from the callus of stem segments of adult plants of Ficus religiosa L.

    PubMed

    Jaiswal, V S; Narayan, P

    1985-10-01

    Stem segments of adult plants of Ficus religiosa L. cultured on MS medium containing 1.0 mg/l 2,4-D produced callus. Shoots were regenerated when the induced calli were transferred to medium supplemented with 0.05 to 2.0 mg/l BAP. Callus derived shoots produced roots and developed into plantlets when transferred to medium supplemented with 1.0 mg/l NAA. PMID:24253982

  16. Adult Education & Human Resource Development: Overlapping and Disparate Fields

    ERIC Educational Resources Information Center

    Watkins, Karen E.; Marsick, Victoria J.

    2014-01-01

    Adult education and human resource development as fields of practice and study share some roots in common but have grown in different directions in their histories. Adult education's roots focused initially on citizenship for a democratic society, whereas human resource development's roots are in performance at work. While they have…

  17. Site-Specific Genome Engineering in Human Pluripotent Stem Cells

    PubMed Central

    Merkert, Sylvia; Martin, Ulrich

    2016-01-01

    The possibility to generate patient-specific induced pluripotent stem cells (iPSCs) offers an unprecedented potential of applications in clinical therapy and medical research. Human iPSCs and their differentiated derivatives are tools for diseases modelling, drug discovery, safety pharmacology, and toxicology. Moreover, they allow for the engineering of bioartificial tissue and are promising candidates for cellular therapies. For many of these applications, the ability to genetically modify pluripotent stem cells (PSCs) is indispensable, but efficient site-specific and safe technologies for genetic engineering of PSCs were developed only recently. By now, customized engineered nucleases provide excellent tools for targeted genome editing, opening new perspectives for biomedical research and cellular therapies. PMID:27347935

  18. Site-Specific Genome Engineering in Human Pluripotent Stem Cells.

    PubMed

    Merkert, Sylvia; Martin, Ulrich

    2016-01-01

    The possibility to generate patient-specific induced pluripotent stem cells (iPSCs) offers an unprecedented potential of applications in clinical therapy and medical research. Human iPSCs and their differentiated derivatives are tools for diseases modelling, drug discovery, safety pharmacology, and toxicology. Moreover, they allow for the engineering of bioartificial tissue and are promising candidates for cellular therapies. For many of these applications, the ability to genetically modify pluripotent stem cells (PSCs) is indispensable, but efficient site-specific and safe technologies for genetic engineering of PSCs were developed only recently. By now, customized engineered nucleases provide excellent tools for targeted genome editing, opening new perspectives for biomedical research and cellular therapies. PMID:27347935

  19. Dynamic and social behaviors of human pluripotent stem cells

    PubMed Central

    Phadnis, Smruti M.; Loewke, Nathan O.; Dimov, Ivan K.; Pai, Sunil; Amwake, Christine E.; Solgaard, Olav; Baer, Thomas M.; Chen, Bertha; Pera, Renee A. Reijo

    2015-01-01

    Human pluripotent stem cells (hPSCs) can self-renew or differentiate to diverse cell types, thus providing a platform for basic and clinical applications. However, pluripotent stem cell populations are heterogeneous and functional properties at the single cell level are poorly documented leading to inefficiencies in differentiation and concerns regarding reproducibility and safety. Here, we use non-invasive time-lapse imaging to continuously examine hPSC maintenance and differentiation and to predict cell viability and fate. We document dynamic behaviors and social interactions that prospectively distinguish hPSC survival, self-renewal, and differentiation. Results highlight the molecular role of E-cadherin not only for cell-cell contact but also for clonal propagation of hPSCs. Results indicate that use of continuous time-lapse imaging can distinguish cellular heterogeneity with respect to pluripotency as well as a subset of karyotypic abnormalities whose dynamic properties were monitored. PMID:26381699

  20. Enhanced genetic modification of adult growth factor mobilized peripheral blood hematopoietic stem and progenitor cells with rapamycin.

    PubMed

    Li, Lijing; Torres-Coronado, Mónica; Gu, Angel; Rao, Anitha; Gardner, Agnes M; Epps, Elizabeth W; Gonzalez, Nancy; Tran, Chy-Anh; Wu, Xiwei; Wang, Jin-Hui; DiGiusto, David L

    2014-10-01

    Genetic modification of adult human hematopoietic stem and progenitor cells (HSPCs) with lentiviral vectors leads to long-term gene expression in the progeny of the HSPCs and has been used to successfully treat several monogenic diseases. In some cases, the gene-modified cells have a selective growth advantage over nonmodified cells and eventually are the dominant engrafted population. However, in disease indications for which the gene-modified cells do not have a selective advantage, optimizing transduction of HSPC is paramount to successful stem cell-based gene therapy. We demonstrate here that transduction of adult CD34+ HSPCs with lentiviral vectors in the presence of rapamycin, a widely used mTORC1 inhibitor, results in an approximately threefold increase in stable gene marking with minimal effects on HSPC growth and differentiation. Using this approach, we have demonstrated that we can enhance the frequency of gene-modified HSPCs that give rise to clonogenic progeny in vitro without excessive increases in the number of vector copies per cell or changes in integration pattern. The genetic marking of HSPCs and expression of transgenes is durable, and transplantation of gene-modified HSPCs into immunodeficient mice results in high levels of gene marking of the lymphoid and myeloid progeny in vivo. The prior safe clinical history of rapamycin in other applications supports the use of this compound to generate gene-modified autologous HSPCs for our HIV gene therapy clinical trials.

  1. CB1 cannabinoid receptor enrichment in the ependymal region of the adult human spinal cord.

    PubMed

    Paniagua-Torija, Beatriz; Arevalo-Martin, Angel; Ferrer, Isidro; Molina-Holgado, Eduardo; Garcia-Ovejero, Daniel

    2015-12-04

    Cannabinoids are involved in the regulation of neural stem cell biology and their receptors are expressed in the neurogenic niches of adult rodents. In the spinal cord of rats and mice, neural stem cells can be found in the ependymal region, surrounding the central canal, but there is evidence that this region is largely different in adult humans: lacks a patent canal and presents perivascular pseudorosettes, typically found in low grade ependymomas. Using Laser Capture Microdissection, Taqman gene expression assays and immunohistochemistry, we have studied the expression of endocannabinoid system components (receptors and enzymes) at the human spinal cord ependymal region. We observe that ependymal region is enriched in CB1 cannabinoid receptor, due to high CB1 expression in GFAP+ astrocytic domains. However, in human spinal cord levels that retain central canal patency we found ependymal cells with high CB1 expression, equivalent to the CB1(HIGH) cell subpopulation described in rodents. Our results support the existence of ependymal CB1(HIGH) cells across species, and may encourage further studies on this subpopulation, although only in cases when central canal is patent. In the adult human ependyma, which usually shows central canal absence, CB1 may play a different role by modulating astrocyte functions.

  2. CB1 cannabinoid receptor enrichment in the ependymal region of the adult human spinal cord

    PubMed Central

    Paniagua-Torija, Beatriz; Arevalo-Martin, Angel; Ferrer, Isidro; Molina-Holgado, Eduardo; Garcia-Ovejero, Daniel

    2015-01-01

    Cannabinoids are involved in the regulation of neural stem cell biology and their receptors are expressed in the neurogenic niches of adult rodents. In the spinal cord of rats and mice, neural stem cells can be found in the ependymal region, surrounding the central canal, but there is evidence that this region is largely different in adult humans: lacks a patent canal and presents perivascular pseudorosettes, typically found in low grade ependymomas. Using Laser Capture Microdissection, Taqman gene expression assays and immunohistochemistry, we have studied the expression of endocannabinoid system components (receptors and enzymes) at the human spinal cord ependymal region. We observe that ependymal region is enriched in CB1 cannabinoid receptor, due to high CB1 expression in GFAP+ astrocytic domains. However, in human spinal cord levels that retain central canal patency we found ependymal cells with high CB1 expression, equivalent to the CB1HIGH cell subpopulation described in rodents. Our results support the existence of ependymal CB1HIGH cells across species, and may encourage further studies on this subpopulation, although only in cases when central canal is patent. In the adult human ependyma, which usually shows central canal absence, CB1 may play a different role by modulating astrocyte functions. PMID:26634814

  3. Human Finger-Prick Induced Pluripotent Stem Cells Facilitate the Development of Stem Cell Banking

    PubMed Central

    Tan, Hong-Kee; Toh, Cheng-Xu Delon; Ma, Dongrui; Yang, Binxia; Liu, Tong Ming; Lu, Jun; Wong, Chee-Wai; Tan, Tze-Kai; Li, Hu; Syn, Christopher; Tan, Eng-Lee; Lim, Bing; Lim, Yoon-Pin; Cook, Stuart A.

    2014-01-01

    Induced pluripotent stem cells (iPSCs) derived from somatic cells of patients can be a good model for studying human diseases and for future therapeutic regenerative medicine. Current initiatives to establish human iPSC (hiPSC) banking face challenges in recruiting large numbers of donors with diverse diseased, genetic, and phenotypic representations. In this study, we describe the efficient derivation of transgene-free hiPSCs from human finger-prick blood. Finger-prick sample collection can be performed on a “do-it-yourself” basis by donors and sent to the hiPSC facility for reprogramming. We show that single-drop volumes of finger-prick samples are sufficient for performing cellular reprogramming, DNA sequencing, and blood serotyping in parallel. Our novel strategy has the potential to facilitate the development of large-scale hiPSC banking worldwide. PMID:24646489

  4. Human finger-prick induced pluripotent stem cells facilitate the development of stem cell banking.

    PubMed

    Tan, Hong-Kee; Toh, Cheng-Xu Delon; Ma, Dongrui; Yang, Binxia; Liu, Tong Ming; Lu, Jun; Wong, Chee-Wai; Tan, Tze-Kai; Li, Hu; Syn, Christopher; Tan, Eng-Lee; Lim, Bing; Lim, Yoon-Pin; Cook, Stuart A; Loh, Yuin-Han

    2014-05-01

    Induced pluripotent stem cells (iPSCs) derived from somatic cells of patients can be a good model for studying human diseases and for future therapeutic regenerative medicine. Current initiatives to establish human iPSC (hiPSC) banking face challenges in recruiting large numbers of donors with diverse diseased, genetic, and phenotypic representations. In this study, we describe the efficient derivation of transgene-free hiPSCs from human finger-prick blood. Finger-prick sample collection can be performed on a "do-it-yourself" basis by donors and sent to the hiPSC facility for reprogramming. We show that single-drop volumes of finger-prick samples are sufficient for performing cellular reprogramming, DNA sequencing, and blood serotyping in parallel. Our novel strategy has the potential to facilitate the development of large-scale hiPSC banking worldwide.

  5. Generation of Human Epidermis-Derived Mesenchymal Stem Cell-like Pluripotent Cells (hEMSCPCs)

    PubMed Central

    Huang, Bing; Li, Kaijing; Yu, Jie; Zhang, Min; Li, Yongping; Li, Weihua; Wang, Wencong; Guan, Liping; Zhang, Wenxin; Lin, Shaochun; Huang, Xintao; Lin, Liping; Lin, Yongliang; Zhang, Yichi; Song, Xinming; Wang, Zhichong; Ge, Jian

    2013-01-01

    We isolated human epidermis-derived mesenchymal stem cell-like pluripotent cells (hEMSCPCs) and demonstrate efficient harvesting, maintenance in vitro for at least 30 passages, reprogramming into multiple phenotypes in vivo, and integration into adult host tissues after injection into the mouse blastocyst to create chimeras. Cell phenotype was examined by karyotyping, immunostaining, immunofluorescence, and flow cytometry. A nested PCR protocol using primers specific for human SRY genes was designed to detect hEMSCPC-derived cells in female chimeric mice. FISH was used to validate the results of nested PCR. Results indicated that hEMSCPCs were derived from epidermis but were distinct from epidermal cells; they resembled mesenchymal stem cells (MSCs) morphologically and expressed the main markers of MSCs. About half of all female offspring of mice implanted with embryos injected with hEMSCPCs at the blastocyst stage harbored the human Y chromosome and tissue-specific human protein, thereby demonstrating the transdifferentiation of hEMSCPCs. PMID:23733028

  6. Genetic recombination pathways and their application for genome modification of human embryonic stem cells.

    PubMed

    Nieminen, Mikko; Tuuri, Timo; Savilahti, Harri

    2010-10-01

    Human embryonic stem cells are pluripotent cells derived from early human embryo and retain a potential to differentiate into all adult cell types. They provide vast opportunities in cell replacement therapies and are expected to become significant tools in drug discovery as well as in the studies of cellular and developmental functions of human genes. The progress in applying different types of DNA recombination reactions for genome modification in a variety of eukaryotic cell types has provided means to utilize recombination-based strategies also in human embryonic stem cells. Homologous recombination-based methods, particularly those utilizing extended homologous regions and those employing zinc finger nucleases to boost genomic integration, have shown their usefulness in efficient genome modification. Site-specific recombination systems are potent genome modifiers, and they can be used to integrate DNA into loci that contain an appropriate recombination signal sequence, either naturally occurring or suitably pre-engineered. Non-homologous recombination can be used to generate random integrations in genomes relatively effortlessly, albeit with a moderate efficiency and precision. DNA transposition-based strategies offer substantially more efficient random strategies and provide means to generate single-copy insertions, thus potentiating the generation of genome-wide insertion libraries applicable in genetic screens.

  7. Transcriptome of human foetal heart compared with cardiomyocytes from pluripotent stem cells.

    PubMed

    van den Berg, Cathelijne W; Okawa, Satoshi; Chuva de Sousa Lopes, Susana M; van Iperen, Liesbeth; Passier, Robert; Braam, Stefan R; Tertoolen, Leon G; del Sol, Antonio; Davis, Richard P; Mummery, Christine L

    2015-09-15

    Differentiated derivatives of human pluripotent stem cells (hPSCs) are often considered immature because they resemble foetal cells more than adult, with hPSC-derived cardiomyocytes (hPSC-CMs) being no exception. Many functional features of these cardiomyocytes, such as their cell morphology, electrophysiological characteristics, sarcomere organization and contraction force, are underdeveloped compared with adult cardiomyocytes. However, relatively little is known about how their gene expression profiles compare with the human foetal heart, in part because of the paucity of data on the human foetal heart at different stages of development. Here, we collected samples of matched ventricles and atria from human foetuses during the first and second trimester of development. This presented a rare opportunity to perform gene expression analysis on the individual chambers of the heart at various stages of development, allowing us to identify not only genes involved in the formation of the heart, but also specific genes upregulated in each of the four chambers and at different stages of development. The data showed that hPSC-CMs had a gene expression profile similar to first trimester foetal heart, but after culture in conditions shown previously to induce maturation, they cluster closer to the second trimester foetal heart samples. In summary, we demonstrate how the gene expression profiles of human foetal heart samples can be used for benchmarking hPSC-CMs and also contribute to determining their equivalent stage of development.

  8. Regulatory System for Stem/Progenitor Cell Niches in the Adult Rodent Pituitary

    PubMed Central

    Yoshida, Saishu; Kato, Takako; Kato, Yukio

    2016-01-01

    The anterior lobe of the pituitary gland is a master endocrine tissue composed of five types of endocrine cells. Although the turnover rate of pituitary endocrine cells is as low as about 1.6% per day, recent studies have demonstrated that Sex-determining region Y-box 2 (SOX2)+-cells exist as pituitary stem/progenitor cells in the adult anterior lobe and contribute to cell regeneration. Notably, SOX2+-pituitary stem/progenitor cells form two types of niches in this tissue: the marginal cell layer (MCL-niche) and the dense cell clusters scattering in the parenchyma (parenchymal-niche). However, little is known about the mechanisms and factors for regulating the pituitary stem/progenitor cell niches, as well as the functional differences between the two types of niches. Elucidation of the regulatory mechanisms in the niches might enable us to understand the cell regeneration system that acts in accordance with physiological demands in the adult pituitary. In this review, so as to reveal the regulatory mechanisms of the two types of niche, we summarize the regulatory factors and their roles in the adult rodent pituitary niches by focusing on three components: soluble factors, cell surface proteins and extracellular matrixes. PMID:26761002

  9. Adult neurogenesis, neural stem cells and Alzheimer's disease: developments, limitations, problems and promises.

    PubMed

    Taupin, Philippe

    2009-12-01

    Alzheimer's disease (AD) is an irreversible progressive neurodegenerative disease, leading to severe incapacity and death. It is the most common form of dementia among older people. AD is characterized in the brain by amyloid plaques, neurofibrillary tangles, neuronal degeneration, aneuploidy and enhanced neurogenesis and by cognitive, behavioral and physical impairments. Inherited mutations in several genes and genetic, acquired and environmental risk factors have been reported as causes for developing the disease, for which there is currently no cure. Current treatments for AD involve drugs and occupational therapies, and future developments involve early diagnosis and stem cell therapy. In this manuscript, we will review and discuss the recent developments, limitations, problems and promises on AD, particularly related to aneuploidy, adult neurogenesis, neural stem cells (NSCs) and cellular therapy. Though adult neurogenesis may be beneficial for regeneration of the nervous system, it may underly the pathogenesis of AD. Cellular therapy is a promising strategy for AD. Limitations in protocols to establish homogeneous populations of neural progenitor and stem cells and niches for neurogenesis need to be resolved and unlocked, for the full potential of adult NSCs to be realized for therapy.

  10. Could adult hippocampal neurogenesis be relevant for human behavior?

    PubMed Central

    Snyder, Jason S.; Cameron, Heather A.

    2011-01-01

    Although the function of adult neurogenesis is still unclear, tools for directly studying the behavioral role of new hippocampal neurons now exist in rodents. Since similar studies are impossible to do in humans, it is important to assess whether the role of new neurons in rodents is likely to be similar to that in humans. One feature of adult neurogenesis that varies tremendously across species is the number of neurons that are generated, so a key question is whether there are enough neurons generated in humans to impact function. In this review we examine neuroanatomy and circuit function in the hippocampus to ask how many granule neurons are needed to impact hippocampal function and then discuss what is known about numbers of new neurons produced in adult rats and humans. We conclude that relatively small numbers of neurons could affect hippocampal circuits and that the magnitude of adult neurogenesis in adult rats and humans is probably larger than generally believed. PMID:21736900

  11. A murine-ES like state facilitates transgenesis and homologous recombination in human pluripotent stem cells

    PubMed Central

    Buecker, Christa; Chen, Hsu-Hsin; Polo, Jose; Daheron, Laurence; Bu, Lei; Barakat, Tahsin Stefan; Okwieka, Patricia; Porter, Andrew; Gribnau, Joost; Hochedlinger, Konrad; Geijsen, Niels

    2010-01-01

    Murine embryonic stem cells have been shown to exist in two functionally distinct pluripotent states, embryonic stem cells (ES cell)- and epiblast stem cells (EpiSCs), which are defined by the culture growth factor conditions. Human ES cells appear to exist in an epiblast-like state, which in comparison to their murine counterparts, is relatively difficult to propagate and manipulate. As a result, gene targeting is difficult and to-date only a handful of human knock-in or knock-out cell lines exist. We explored whether an alternative stem cell state exists for human stem cells as well, and demonstrate that manipulation of the growth factor milieu allows the derivation of a novel human stem cell type that displays morphological, molecular and functional properties of murine ES cells and facilitates gene targeting. As such, the murine ES-like state provides a powerful tool for the generation of recombinant human pluripotent stem cell lines. PMID:20569691

  12. Differentiation of hepatocytes from induced pluripotent stem cells derived from human hair follicle mesenchymal stem cells.

    PubMed

    Shi, Xu; Lv, Shuang; He, Xia; Liu, Xiaomei; Sun, Meiyu; Li, Meiying; Chi, Guangfan; Li, Yulin

    2016-10-01

    Due to the limitations of organ donors and immune rejection in severe liver diseases, stem cell-based therapy presents a promising application for tissue repair and regeneration. As a novel cell source, mesenchymal stem cells separated from human hair follicles (HF-MSCs) are convenient to obtain and have no age limit. To date, the differentiation of HF-MSCs into hepatocytes has not been reported. In this study, we explored whether HF-MSCs and HF-MSC-derived-induced pluripotent stem cells (HF-iPS) could differentiate into hepatocytes in vitro. Flow cytometry, Oil Red O stain and Alizarin Red stain were used to identify the characteristics of HF-MSCs. The expression of liver-specific gene was detected by immunofluorescence and Quantitative Polymerase Chain Reaction. Periodic Acid-Schiff stain, Indocyanine Green stain and Low-Density Lipoprotein stain were performed to evaluate the functions of induced hepatocyte-like cells (HLCs). HF-MSCs were unable to differentiate into HLCs using previously reported procedures for MSCs from other tissues. However, HF-iPS efficiently induced the generation of HLCs that expressed hepatocyte markers and drug metabolism-related genes. HF-iPS can be used as novel and alternative cellular tools for inducing hepatocytes in vitro, simultaneously benefiting from utilizing HF-MSCs as a noninvasive and convenient cell source for reprogramming.

  13. Decellularized ECM Effects on Human Mesenchymal Stem Cell Stemness and Differentiation

    PubMed Central

    Pattabhi, Sudhakara rao; Martinez, Jessica S.; Keller, Thomas C. S.

    2015-01-01

    Microenvironment extracellular matrices (ECMs) influence cell adhesion, proliferation and differentiation. The ECMs of different microenvironments have distinctive compositions and architectures. This investigation addresses effects ECMs deposited by a variety of cell types and decellularized with a cold-EDTA protocol have on multipotent human mesenchymal stromal/stem cell (hMSC) behavior and differentiation. The cold-EDTA protocol removes intact cells from ECM, with minimal ECM damage and contamination. The decellularized ECMs deposited by cultured hMSCs, osteogenic hMSCs, and two smooth muscle cell (SMC) lines were tested for distinctive effects on the behavior and differentiation of early passage (‘naïve’) hMSC plated and cultured on the decellularized ECMs. Uninduced hMSC decellularized ECM enhanced naïve hMSC proliferation and cell motility while maintaining stemness. Decellularized ECM deposited by osteogenic hMSCs early in the differentiation process stimulated naïve hMSCs osteogenesis and substrate biomineralization in the absence of added dexamethasone, but this osteogenic induction potential was lower in ECMs decellularized later in the osteogenic hMSC differentiation process. Decellularized ECMs deposited by two smooth muscle cell lines induced naïve hMSCs to become smooth muscle cell-like with distinctive phenotypic characteristics of contractile and synthetic smooth muscle cells. This investigation demonstrates a useful approach for obtaining functional cell-deposited ECM and highlights the importance of ECM specificity in influencing stem cell behavior. PMID:25578478

  14. Differentiation of hepatocytes from induced pluripotent stem cells derived from human hair follicle mesenchymal stem cells.

    PubMed

    Shi, Xu; Lv, Shuang; He, Xia; Liu, Xiaomei; Sun, Meiyu; Li, Meiying; Chi, Guangfan; Li, Yulin

    2016-10-01

    Due to the limitations of organ donors and immune rejection in severe liver diseases, stem cell-based therapy presents a promising application for tissue repair and regeneration. As a novel cell source, mesenchymal stem cells separated from human hair follicles (HF-MSCs) are convenient to obtain and have no age limit. To date, the differentiation of HF-MSCs into hepatocytes has not been reported. In this study, we explored whether HF-MSCs and HF-MSC-derived-induced pluripotent stem cells (HF-iPS) could differentiate into hepatocytes in vitro. Flow cytometry, Oil Red O stain and Alizarin Red stain were used to identify the characteristics of HF-MSCs. The expression of liver-specific gene was detected by immunofluorescence and Quantitative Polymerase Chain Reaction. Periodic Acid-Schiff stain, Indocyanine Green stain and Low-Density Lipoprotein stain were performed to evaluate the functions of induced hepatocyte-like cells (HLCs). HF-MSCs were unable to differentiate into HLCs using previously reported procedures for MSCs from other tissues. However, HF-iPS efficiently induced the generation of HLCs that expressed hepatocyte markers and drug metabolism-related genes. HF-iPS can be used as novel and alternative cellular tools for inducing hepatocytes in vitro, simultaneously benefiting from utilizing HF-MSCs as a noninvasive and convenient cell source for reprogramming. PMID:27053247

  15. Defects in ErbB-dependent establishment of adult melanocyte stem cells reveal independent origins for embryonic and regeneration melanocytes.

    PubMed

    Hultman, Keith A; Budi, Erine H; Teasley, Daniel C; Gottlieb, Andrew Y; Parichy, David M; Johnson, Stephen L

    2009-07-01

    Adult stem cells are responsible for maintaining and repairing tissues during the life of an organism. Tissue repair in humans, however, is limited compared to the regenerative capabilities of other vertebrates, such as the zebrafish (Danio rerio). An understanding of stem cell mechanisms, such as how they are established, their self-renewal properties, and their recruitment to produce new cells is therefore important for the application of regenerative medicine. We use larval melanocyte regeneration following treatment with the melanocytotoxic drug MoTP to investigate these mechanisms in Melanocyte Stem Cell (MSC) regulation. In this paper, we show that the receptor tyrosine kinase, erbb3b, is required for establishing the adult MSC responsible for regenerating the larval melanocyte population. Both the erbb3b mutant and wild-type fish treated with the ErbB inhibitor, AG1478, develop normal embryonic melanocytes but fail to regenerate melanocytes after MoTP-induced melanocyte ablation. By administering AG1478 at different time points, we show that ErbB signaling is only required for regeneration prior to MoTP treatment and before 48 hours of development, consistent with a role in establishing MSCs. We then show that overexpression of kitla, the Kit ligand, in transgenic larvae leads to recruitment of MSCs, resulting in overproliferation of melanocytes. Furthermore, kitla overexpression can rescue AG1478-blocked regeneration, suggesting that ErbB signaling is required to promote the progression and specification of the MSC from a pre-MSC state. This study provides evidence that ErbB signaling is required for the establishment of adult MSCs during embryonic development. That this requirement is not shared with the embryonic melanocytes suggests that embryonic melanocytes develop directly, without proceeding through the ErbB-dependent MSC. Moreover, the shared requirement of larval melanocyte regeneration and metamorphic melanocytes that develops at the larval-to-adult

  16. Potential for a pluripotent adult stem cell treatment for acute radiation sickness

    PubMed Central

    Rodgerson, Denis O; Reidenberg, Bruce E; Harris, Alan G; Pecora, Andrew L

    2012-01-01

    Accidental radiation exposure and the threat of deliberate radiation exposure have been in the news and are a public health concern. Experience with acute radiation sickness has been gathered from atomic blast survivors of Hiroshima and Nagasaki and from civilian nuclear accidents as well as experience gained during the development of radiation therapy for cancer. This paper reviews the medical treatment reports relevant to acute radiation sickness among the survivors of atomic weapons at Hiroshima and Nagasaki, among the victims of Chernobyl, and the two cases described so far from the Fukushima Dai-Ichi disaster. The data supporting the use of hematopoietic stem cell transplantation and the new efforts to expand stem cell populations ex vivo for infusion to treat bone marrow failure are reviewed. Hematopoietic stem cells derived from bone marrow or blood have a broad ability to repair and replace radiation induced damaged blood and immune cell production and may promote blood vessel formation and tissue repair. Additionally, a constituent of bone marrow-derived, adult pluripotent stem cells, very small embryonic like stem cells, are highly resistant to ionizing radiation and appear capable of regenerating radiation damaged tissue including skin, gut and lung. PMID:24520532

  17. Potential for a pluripotent adult stem cell treatment for acute radiation sickness.

    PubMed

    Rodgerson, Denis O; Reidenberg, Bruce E; Harris, Alan G; Pecora, Andrew L

    2012-06-20

    Accidental radiation exposure and the threat of deliberate radiation exposure have been in the news and are a public health concern. Experience with acute radiation sickness has been gathered from atomic blast survivors of Hiroshima and Nagasaki and from civilian nuclear accidents as well as experience gained during the development of radiation therapy for cancer. This paper reviews the medical treatment reports relevant to acute radiation sickness among the survivors of atomic weapons at Hiroshima and Nagasaki, among the victims of Chernobyl, and the two cases described so far from the Fukushima Dai-Ichi disaster. The data supporting the use of hematopoietic stem cell transplantation and the new efforts to expand stem cell populations ex vivo for infusion to treat bone marrow failure are reviewed. Hematopoietic stem cells derived from bone marrow or blood have a broad ability to repair and replace radiation induced damaged blood and immune cell production and may promote blood vessel formation and tissue repair. Additionally, a constituent of bone marrow-derived, adult pluripotent stem cells, very small embryonic like stem cells, are highly resistant to ionizing radiation and appear capable of regenerating radiation damaged tissue including skin, gut and lung.

  18. Strategies to Enhance the Effectiveness of Adult Stem Cell Therapy for Ischemic Heart Diseases Affecting the Elderly Patients

    PubMed Central

    Khatiwala, Roshni

    2016-01-01

    Myocardial infarctions and chronic ischemic heart disease both commonly and disproportionately affect elderly patients more than any other patient population. Despite available treatments, heart tissue is often permanently damaged as a result of cardiac injury. This review aims to summarize recent literature proposing the use of modified autologous adult stem cells to promote healing of post-infarct cardiac tissue. This novel cellular treatment involves isolation of adult stem cells from the patient, in vitro manipulation of these stem cells, and subsequent transplantation back into the patient’s own heart to accelerate healing. One of the hindrances affecting this process is that cardiac issues are increasingly common in elderly patients, and stem cells recovered from their tissues tend to be pre-senescent or already in senescence. As a result, harsh in vitro manipulations can cause the aged stem cells to undergo massive in vivo apoptosis after transplantation. The consensus in literature is that inhibition or reversal of senescence onset in adult stem cells would be of utmost benefit. In fact, it is believed that this strategy may lower stem cell mortality and coerce aged stem cells into adopting more resilient phenotypes similar to that of their younger counterparts. This review will discuss a selection of the most efficient and most-recent strategies used experimentally to enhance the effectiveness of current stem cell therapies for ischemic heart diseases. PMID:26779896

  19. Human adipose CD34+ CD90+ stem cells and collagen scaffold constructs grafted in vivo fabricate loose connective and adipose tissues.

    PubMed

    Ferraro, Giuseppe A; De Francesco, Francesco; Nicoletti, Gianfranco; Paino, Francesca; Desiderio, Vincenzo; Tirino, Virginia; D'Andrea, Francesco

    2013-05-01

    Stem cell based therapies for the repair and regeneration of various tissues are of great interest for a high number of diseases. Adult stem cells, instead, are more available, abundant and harvested with minimally invasive procedures. In particular, mesenchymal stem cells (MSCs) are multi-potent progenitors, able to differentiate into bone, cartilage, and adipose tissues. Human adult adipose tissue seems to be the most abundant source of MSCs and, due to its easy accessibility; it is able to give a considerable amount of stem cells. In this study, we selected MSCs co-expressing CD34 and CD90 from adipose tissue. This stem cell population displayed higher proliferative capacity than CD34(-) CD90(-) cells and was able to differentiate in vitro into adipocytes (PPARγ(+) and adiponectin(+)) and endothelial cells (CD31(+) VEGF(+) Flk1(+)). In addition, in methylcellulose without VEGF, it formed a vascular network. The aim of this study was to investigate differentiation potential of human adipose CD34(+) /CD90(+) stem cells loaded onto commercial collagen sponges already used in clinical practice (Gingistat) both in vitro and in vivo. The results of this study clearly demonstrate that human adult adipose and loose connective tissues can be obtained in vivo, highlighting that CD34(+) /CD90 ASCs are extremely useful for regenerative medicine.

  20. Differentiating the stem cell pool of human hair follicle outer root sheath into functional melanocytes.

    PubMed

    Schneider, Marie; Dieckmann, Christina; Rabe, Katrin; Simon, Jan-Christoph; Savkovic, Vuk

    2014-01-01

    Bench-to-Bedside concepts for regenerative therapy place significant weight on noninvasive approaches, with harvesting of the starting material as a header. This is particularly important in autologous treatments, which use one's bodily constituents for therapy. Precisely the stretch between obtaining therapeutic elements invasively and noninvasively places non-intrusive "sampling" rather than "biopsy" in the center of the road map of developing an autologous regenerative therapy. We focus on such a noninvasively available source of adult stem cells that we carry with us throughout our life, available at our fingertips-or shall we say hair roots, by a simple plucking of hair: the human hair follicle. This chapter describes an explant procedure for cultivating melanocytes differentiated from the stem cell pool of the hair follicle Outer Root Sheath (ORS). In vivo, the most abundant derivatives of the heterogeneous ORS stem cell pool are epidermal cells-melanocytes and keratinocytes which complete their differentiation-either spontaneously or upon picking up regenerative cues from damaged skin-and migrate from the ORS towards the adjacent regenerating area of the epidermis. We have taken advantage of the ORS developmental potential by optimizing explant primary culture, expansion and melanogenic differentiation of resident ORS stem cells towards end-stage melanocytes in order to obtain functional melanocytes noninvasively for the purposes of transplantation and use them for the treatment of depigmentation disorders. Our protocol specifies sampling of hair with their ORS, follicle medium-air interface primary culture, stimulation of cell outgrowth, adherent culture and differentiation of ORS stem cells and precursors towards fully functional melanocytes. Along with cultivation, we describe selection techniques for establishing and maintaining a pure melanocyte population and methods suitable for determining melanocyte identity.

  1. Erythroid differentiation of human induced pluripotent stem cells is independent of donor cell type of origin

    PubMed Central

    Dorn, Isabel; Klich, Katharina; Arauzo-Bravo, Marcos J.; Radstaak, Martina; Santourlidis, Simeon; Ghanjati, Foued; Radke, Teja F.; Psathaki, Olympia E.; Hargus, Gunnar; Kramer, Jan; Einhaus, Martin; Kim, Jeong Beom; Kögler, Gesine; Wernet, Peter; Schöler, Hans R.; Schlenke, Peter; Zaehres, Holm

    2015-01-01

    Epigenetic memory in induced pluripotent stem cells, which is related to the somatic cell type of origin of the stem cells, might lead to variations in the differentiation capacities of the pluripotent stem cells. In this context, induced pluripotent stem cells from human CD34+ hematopoietic stem cells might be more suitable for hematopoietic differentiation than the commonly used fibroblast-derived induced pluripotent stem cells. To investigate the influence of an epigenetic memory on the ex vivo expansion of induced pluripotent stem cells into erythroid cells, we compared induced pluripotent stem cells from human neural stem cells and human cord blood-derived CD34+ hematopoietic stem cells and evaluated their potential for differentiation into hematopoietic progenitor and mature red blood cells. Although genome-wide DNA methylation profiling at all promoter regions demonstrates that the epigenetic memory of induced pluripotent stem cells is influenced by the somatic cell type of origin of the stem cells, we found a similar hematopoietic induction potential and erythroid differentiation pattern of induced pluripotent stem cells of different somatic cell origin. All human induced pluripotent stem cell lines showed terminal maturation into normoblasts and enucleated reticulocytes, producing predominantly fetal hemoglobin. Differences were only observed in the growth rate of erythroid cells, which was slightly higher in the induced pluripotent stem cells derived from CD34+ hematopoietic stem cells. More detailed methylation analysis of the hematopoietic and erythroid promoters identified similar CpG methylation levels in the induced pluripotent stem cell lines derived from CD34+ cells and those derived from neural stem cells, which confirms their comparable erythroid differentiation potential. PMID:25326431

  2. Phytoestrogen Metabolism by Adult Human Gut Microbiota.

    PubMed

    Gaya, Pilar; Medina, Margarita; Sánchez-Jiménez, Abel; Landete, José Mᵃ

    2016-08-09

    Phytoestrogens are plant-derived polyphenols with a structure similar to human estrogens. The three main groups of phytoestrogens, isoflavones, ellagitannins, and lignans, are transformed into equol, urolithins, and enterolignans, respectively, by bacteria. These metabolites have more estrogenic/antiestrogenic and antioxidant activities than their precursors, and they are more bioavailable. The aim of this study was to analyze the metabolism of isoflavones, lignans and ellagitannins by gut microbiota, and to study the possible correlation in the metabolism of these three groups of phytoestrogens. In vitro fermentation experiments were performed with feces samples from 14 healthy adult volunteers, and metabolite formation was measured by HPLC-PAD and HPLC-ESI/MS. Only the microbiota of one subject produced equol, while most of them showed production of O-desmethylangolensin (O-DMA). Significant inter-subject differences were observed in the metabolism of dihydrodaidzein and dihydrogenistein, while the glucoside isoflavones and their aglycones showed less variability, except for glycitin. Most subjects produced urolithins M-5 and E. Urolithin D was not detected, while uroltithin B was found in half of the individuals analyzed, and urolithins A and C were detected in two and four subjects, respectively. Enterolactone was found in all subjects, while enterodiol only appeared in five. Isoflavone metabolism could be correlated with the metabolism of lignans and ellagitannins. However, the metabolism of ellagitannins and lignans could not be correlated. This the first study where the metabolism of the three groups together of phytoestrogen, isoflavones, lignans, and ellagitannins by gut microbiota is analyzed.

  3. Induced pluripotent stem cells for modelling human diseases

    PubMed Central

    Unternaehrer, Juli J.; Daley, George Q.

    2011-01-01

    Research into the pathophysiological mechanisms of human disease and the development of targeted therapies have been hindered by a lack of predictive disease models that can be experimentally manipulated in vitro. This review describes the current state of modelling human diseases with the use of human induced pluripotent stem (iPS) cell lines. To date, a variety of neurodegenerative diseases, haematopoietic disorders, metabolic conditions and cardiovascular pathologies have been captured in a Petri dish through reprogramming of patient cells into iPS cells followed by directed differentiation of disease-relevant cells and tissues. However, realizing the true promise of iPS cells for advancing our basic understanding of disease and ultimately providing novel cell-based therapies will require more refined protocols for generating the highly specialized cells affected by disease, coupled with strategies for drug discovery and cell transplantation. PMID:21727133

  4. Generation of functional podocytes from human induced pluripotent stem cells.

    PubMed

    Ciampi, Osele; Iacone, Roberto; Longaretti, Lorena; Benedetti, Valentina; Graf, Martin; Magnone, Maria Chiara; Patsch, Christoph; Xinaris, Christodoulos; Remuzzi, Giuseppe; Benigni, Ariela; Tomasoni, Susanna

    2016-07-01

    Generating human podocytes in vitro could offer a unique opportunity to study human diseases. Here, we describe a simple and efficient protocol for obtaining functional podocytes in vitro from human induced pluripotent stem cells. Cells were exposed to a three-step protocol, which induced their differentiation into intermediate mesoderm, then into nephron progenitors and, finally, into mature podocytes. After differentiation, cells expressed the main podocyte markers, such as synaptopodin, WT1, α-Actinin-4, P-cadherin and nephrin at the protein and mRNA level, and showed the low proliferation rate typical of mature podocytes. Exposure to Angiotensin II significantly decreased the expression of podocyte genes and cells underwent cytoskeleton rearrangement. Cells were able to internalize albumin and self-assembled into chimeric 3D structures in combination with dissociated embryonic mouse kidney cells. Overall, these findings demonstrate the establishment of a robust protocol that, mimicking developmental stages, makes it possible to derive functional podocytes in vitro.

  5. Automated adherent human cell culture (mesenchymal stem cells).

    PubMed

    Thomas, Robert; Ratcliffe, Elizabeth

    2012-01-01

    Human cell culture processes developed at research laboratory scale need to be translated to large-scale production processes to achieve commercial application to a large market. To allow this transition of scale with consistent process performance and control of costs, it will be necessary to reduce manual processing and increase automation. There are a number of commercially available platforms that will reduce manual process intervention and improve process control for different culture formats. However, in many human cell-based applications, there is currently a need to remain close to the development format, usually adherent culture on cell culture plastic or matrix-coated wells or flasks due to deterioration of cell quality in other environments, such as suspension. This chapter presents an example method for adherent automated human stem cell culture using a specific automated flask handling platform, the CompacT SelecT.

  6. Presence of functional gap junctions in human embryonic stem cells.

    PubMed

    Wong, Raymond C B; Pébay, Alice; Nguyen, Linh T V; Koh, Karen L L; Pera, Martin F

    2004-01-01

    Gap junctions are intercellular channels that allow both chemical and electrical signaling between two adjacent cells. Gap junction intercellular communication has been implicated in the regulation of various cellular processes, including cell migration, cell proliferation, cell differentiation, and cell apoptosis. This study aimed to determine the presence and functionality of gap junctions in human embryonic stem cells (hESCs). Using reverse transcription--polymerase chain reaction and immunocytochemistry, we demonstrate that human ES cells express two gap junction proteins, connexin 43 and connexin 45. Western blot analysis revealed the presence of three phosphorylated forms (nonphosphorylated [NP], P1, and P2) of connexin 43, NP being prominent. Moreover, scrape loading/dye transfer assay indicates that human ES cells are coupled through functional gap junctions that are inhibited by protein kinase C activation and extracellular signal-regulated kinase inhibition.

  7. FACS and MACS sorting strategies to isolate and enrich human spermatogonial stem cells

    PubMed Central

    Valli, Hanna; Sukhwani, Meena; Dovey, Serena L.; Peters, Karen A.; Donohue, Julia; Castro, Carlos A.; Chu, Tianjiao; Marshall, Gary R.; Orwig, Kyle E.

    2014-01-01

    Objective Determine the molecular characteristics of human spermatogonia and optimize methods to enrich spermatogonial stem cells (SSCs). Design Laboratory study using human tissues Setting Research institute Patient(s)/Animal(s) Normal adult human testicular tissue. Interventions Human testicular tissue was fixed or digested with enzymes to produce a cell suspension. Human testis cells were fractionated by FACS and MACS. Main Outcome Measure(s) Immunostaining for selected markers, human-to-nude mouse xenotransplantation assay. Results Immunohistochemistry co-staining revealed the relative expression patterns of SALL4, UTF1, ZBTB16, UCHL1 and ENO2 in human undifferentiated spermatogonia as well as the extent of overlap with the differentiation marker, KIT. Whole mount analyses revealed that human undifferentiated spermatogonia (UCHL1+) were typically arranged in clones of 1–4 cells while differentiated spermatogonia (KIT+) were typically arranged in clones of 8 or more cells. The ratio of undifferentiated to differentiated spermatogonia is greater in humans than in rodents. SSC