WorldWideScience

Sample records for cell lineage commitment

  1. A continuum of cell states spans pluripotency and lineage commitment in human embryonic stem cells.

    Directory of Open Access Journals (Sweden)

    Shelley R Hough

    Full Text Available BACKGROUND: Commitment in embryonic stem cells is often depicted as a binary choice between alternate cell states, pluripotency and specification to a particular germ layer or extraembryonic lineage. However, close examination of human ES cell cultures has revealed significant heterogeneity in the stem cell compartment. METHODOLOGY/PRINCIPAL FINDINGS: We isolated subpopulations of embryonic stem cells using surface markers, then examined their expression of pluripotency genes and lineage specific transcription factors at the single cell level, and tested their ability to regenerate colonies of stem cells. Transcript analysis of single embryonic stem cells showed that there is a gradient and a hierarchy of expression of pluripotency genes in the population. Even cells at the top of the hierarchy generally express only a subset of the stem cell genes studied. Many cells co-express pluripotency and lineage specific genes. Cells along the continuum show a progressively decreasing likelihood of self renewal as their expression of stem cell surface markers and pluripotency genes wanes. Most cells that are positive for stem cell surface markers express Oct-4, but only those towards the top of the hierarchy express the nodal receptor TDGF-1 and the growth factor GDF3. SIGNIFICANCE: These findings on gene expression in single embryonic stem cells are in concert with recent studies of early mammalian development, which reveal molecular heterogeneity and a stochasticity of gene expression in blastomeres. Our work indicates that only a small fraction of the population resides at the top of the hierarchy, that lineage priming (co-expression of stem cell and lineage specific genes characterizes pluripotent stem cell populations, and that extrinsic signaling pathways are upstream of transcription factor networks that control pluripotency.

  2. Downregulation of the transcription factor KLF4 is required for the lineage commitment of T cells

    Institute of Scientific and Technical Information of China (English)

    Xiaomin Wen; Haifeng Liu; Gang Xiao; Xiaolong Liu

    2011-01-01

    The roles of the reprogramming factors Oct4,Sox2,c-Myc and Klf4 in early T cell development are incompletely defined.Here,we show that Klf4 is the only reprogramming factor whose expression is downregulated when early thymic progenitors (ETPs) differentiate into T cells.Enforced expression of Klf4 in uncommitted progenitors severely impaired T cell development mainly at the DN2-to-DN3 transition when T cell lineage commitment occurs and affected the transcription of a variety of genes with crucial functions in early T cell development,including genes involved in microenvironmental signaling (IL-7Rα),Notch target genes (Deltexl),and essential T cell lineage regulatory or inhibitory genes (Bcllla,SpiB,and ldl).The survival of thymocytes and the rearrangement at the Tcrb locus were impaired in the presence of enforced Klf4 expression.The defects in the DN1-to-DN2 and DN2-to-DN3 transitions in Klf4 transgenic mice could not be rescued by the introduction of a TCR transgene,but was partially rescued by restoring the expression of IL-7Rα.Thus,our data indicate that the downregulation of Klf4 is a prerequisite for T cell lineage commitment.

  3. Heterogeneity of natural Foxp3+ T cells: A committed regulatory T-cell lineage and an uncommitted minor population retaining plasticity

    OpenAIRE

    Komatsu, Noriko; Mariotti-Ferrandiz, Maria Encarnita; Wang, Ying; Malissen, Bernard; Waldmann, Herman; Hori, Shohei

    2009-01-01

    Natural regulatory T cells (Treg) represent a distinct lineage of T lymphocytes committed to suppressive functions, and expression of the transcription factor Foxp3 is thought to identify this lineage specifically. Here we report that, whereas the majority of natural CD4+Foxp3+ T cells maintain stable Foxp3 expression after adoptive transfer to lymphopenic or lymphoreplete recipients, a minor fraction enriched within the CD25− subset actually lose it. Some of those Foxp3− T cells adopt effect...

  4. Equine-Induced Pluripotent Stem Cells Retain Lineage Commitment Toward Myogenic and Chondrogenic Fates

    Directory of Open Access Journals (Sweden)

    Mattia Quattrocelli

    2016-01-01

    Full Text Available Induced pluripotent stem cells (iPSCs hold great potential not only for human but also for veterinary purposes. The equine industry must often deal with health issues concerning muscle and cartilage, where comprehensive regenerative strategies are still missing. In this regard, a still open question is whether equine iPSCs differentiate toward muscle and cartilage, and whether donor cell type influences their differentiation potential. We addressed these questions through an isogenic system of equine iPSCs obtained from myogenic mesoangioblasts (MAB-iPSCs and chondrogenic mesenchymal stem cells (MSC-iPSCs. Despite similar levels of pluripotency characteristics, the myogenic differentiation appeared enhanced in MAB-iPSCs. Conversely, the chondrogenic differentiation was augmented in MSC-iPSCs through both teratoma and in vitro differentiation assays. Thus, our data suggest that equine iPSCs can differentiate toward the myogenic and chondrogenic lineages, and can present a skewed differentiation potential in favor of the source cell lineage.

  5. BMP signaling pathway is required for commitment of C3H10T1/2 pluripotent stem cells to the adipocyte lineage

    OpenAIRE

    Huang, Haiyan; Song, Tan-Jing; Li, Xi; Hu, Lingling; He, Qun; Liu, Mei; Lane, M. Daniel; Tang, Qi-Qun

    2009-01-01

    Obesity is accompanied by an increase in both adipocyte number and size. The increase in adipocyte number is the result of recruitment to the adipocyte lineage of pluripotent stem cells present in the vascular stroma of adipose tissue. These pluripotent cells have the potential to undergo commitment and then differentiate into adipocytes, as well as myocytes, osteocytes, and chondrocytes. In this article, we show that both bone morphogenetic protein (BMP)2 and BMP4 can induce commitment of C3...

  6. Activation of GSK3β by Sirt2 is required for early lineage commitment of mouse embryonic stem cell.

    Directory of Open Access Journals (Sweden)

    Xiaoxing Si

    Full Text Available Sirt2, a member of the NAD(+-dependent protein deacetylase family, is increasingly recognized as a critical regulator of the cell cycle, cellular necrosis and cytoskeleton organization. However, its role in embryonic stem cells (ESCs remains unclear. Here we demonstrate that Sirt2 is up-regulated during RA (retinoic acid-induced and embryoid body (EB differentiation of mouse ESCs. Using lentivirus-mediated shRNA methods, we found that knockdown of Sirt2 compromises the differentiation of mouse ESCs into ectoderm while promoting mesoderm and endoderm differentiation. Knockdown of Sirt2 expression also leads to the activation of GSK3β through decreased phosphorylation of the serine at position 9 (Ser9 but not tyrosine at position 216 (Tyr216. Moreover, the constitutive activation of GSK3β during EB differentiation mimics the effect of Sirt2 knockdown, while down-regulation of GSK3β rescues the effect of Sirt2 knockdown on differentiation. In contrast to the effect on lineage differentiation, Sirt2 knockdown and GSK3β up-regulation do not change the self-renewal state of mouse ESCs. Overall, our report reveals a new function for Sirt2 in regulating the proper lineage commitment of mouse ESCs.

  7. A Novel Model System to Study the Role of Catecholamines in Cardiac Lineage Commitment of Embryonic Stem Cells and Functional Response to Proarrhythmic Drugs

    OpenAIRE

    Lehmann, Martin

    2014-01-01

    Embryonic stem (ES) cells are pluripotent cells derived from the inner cell mass of the blastocyst. These cells possess the ability to differentiate into all cell types of the three germ-layers and to proliferate indefinitely. In defined conditions ES cells are committed to the mesodermal lineage and differentiate, amongst other cell types, into cardiomyocytes (CMs). The processes underlying mesodermal and subsequent cardiac differentiation are yet only partially understood. Catecholamine rel...

  8. Even Cancers Want Commitment: Lineage Identity and Medulloblastoma Formation

    OpenAIRE

    Eberhart, Charles G.

    2008-01-01

    In this issue of Cancer Cell, Yang et al. (2008) and Schüller et al. (2008) show that Hedgehog activation in either multipotent neural stem cells or developmentally restricted progenitors causes only medulloblastomas to form. These data suggest that some stem cell-derived tumors must commit to a specific lineage in order to grow.

  9. What Happens in the Thymus Does Not Stay in the Thymus: How T Cells Recycle the CD4+-CD8+ Lineage Commitment Transcriptional Circuitry To Control Their Function.

    Science.gov (United States)

    Vacchio, Melanie S; Bosselut, Rémy

    2016-06-15

    MHC-restricted CD4(+) and CD8(+) T cells are at the core of most adaptive immune responses. Although these cells carry distinct functions, they arise from a common precursor during thymic differentiation, in a developmental sequence that matches CD4 and CD8 expression and functional potential with MHC restriction. Although the transcriptional control of CD4(+)-CD8(+) lineage choice in the thymus is now better understood, less was known about what maintains the CD4(+) and CD8(+) lineage integrity of mature T cells. In this review, we discuss the mechanisms that establish in the thymus, and maintain in postthymic cells, the separation of these lineages. We focus on recent studies that address the mechanisms of epigenetic control of Cd4 expression and emphasize how maintaining a transcriptional circuitry nucleated around Thpok and Runx proteins, the key architects of CD4(+)-CD8(+) lineage commitment in the thymus, is critical for CD4(+) T cell helper functions. PMID:27260768

  10. DEC2 is a negative regulator for the proliferation and differentiation of chondrocyte lineage-committed mesenchymal stem cells.

    Science.gov (United States)

    Sasamoto, Tomoko; Fujimoto, Katsumi; Kanawa, Masami; Kimura, Junko; Takeuchi, Junpei; Harada, Naoko; Goto, Noriko; Kawamoto, Takeshi; Noshiro, Mitsuhide; Suardita, Ketut; Tanne, Kazuo; Kato, Yukio

    2016-09-01

    Differentiated embryo chondrocyte 2 (DEC2) is a basic helix-loop-helix-Orange transcription factor that regulates cell differentiation in various mammalian tissues. DEC2 has been shown to suppress the differentiation of mesenchymal stem cells (MSCs) into myocytes and adipocytes. In the present study, we examined the role of DEC2 in the chondrogenic differentiation of human MSCs. The overexpression of DEC2 exerted minimal effects on the proliferation of MSCs in monolayer cultures with the growth medium under undifferentiating conditions, whereas it suppressed increases in DNA content, glycosaminoglycan content, and the expression of several chondrocyte-related genes, including aggrecan and type X collagen alpha 1, in MSC pellets in centrifuge tubes under chondrogenic conditions. In the pellets exposed to chondrogenesis induction medium, DEC2 overexpression downregulated the mRNA expression of fibroblast growth factor 18, which is involved in the proliferation and differentiation of chondrocytes, and upregulated the expression of p16INK4, which is a cell cycle inhibitor. These findings suggest that DEC2 is a negative regulator of the proliferation and differentiation of chondrocyte lineage-committed mesenchymal cells. PMID:27430159

  11. Snail1 controls epithelial–mesenchymal lineage commitment in focal adhesion kinase–null embryonic cells

    OpenAIRE

    Li, Xiao-Yan; Zhou, Xiaoming; Rowe, R. Grant; Hu, Yuexian; Schlaepfer, David D.; Ilić, Dusko; Dressler, Gregory; Park, Ann; Guan, Jun-Lin; Weiss, Stephen J.

    2011-01-01

    Mouse embryonic cells isolated from focal adhesion kinase (FAK)–null animals at embryonic day 7.5 display multiple defects in focal adhesion remodeling, microtubule dynamics, mechanotransduction, proliferation, directional motility, and invasion. To date, the ability of FAK to modulate cell function has been ascribed largely to its control of posttranscriptional signaling cascades in this embryonic cell population. In this paper, we demonstrate that FAK unexpectedly exerts control over an epi...

  12. Transforming growth factor-beta signaling network regulates plasticity and lineage commitment of lung cancer cells

    OpenAIRE

    Ischenko, I; Liu, J.; Petrenko, O; Hayman, M J

    2014-01-01

    Identification of target cells in lung tumorigenesis and characterization of the signals that control their behavior is an important step toward improving early cancer diagnosis and predicting tumor behavior. We identified a population of cells in the adult lung that bear the EpCAM+CD104+CD49f+CD44+CD24loSCA1+ phenotype and can be clonally expanded in culture, consistent with the properties of early progenitor cells. We show that these cells, rather than being restricted to one tumor type, ca...

  13. Amniotic fluid stem cells morph into a cardiovascular lineage: analysis of a chemically induced cardiac and vascular commitment

    Directory of Open Access Journals (Sweden)

    Maioli M

    2013-09-01

    Full Text Available Margherita Maioli,1–3 Giovanni Contini,1 Sara Santaniello,1,2 Pasquale Bandiera,1 Gianfranco Pigliaru,1,2 Raimonda Sanna,5 Salvatore Rinaldi,3 Alessandro P Delitala,1 Andrea Montella,1,5 Luigi Bagella,1,6 Carlo Ventura2–41Department of Biomedical Sciences, University of Sassari, Sassari, 2Laboratory of Molecular Biology and Stem Cell Engineering, National Institute of Biostructures and Biosystems, Bologna, 3Department of Regenerative Medicine, Rinaldi Fontani Institute, Florence, 4Cardiovascular Department, S Orsola-Malpighi Hospital, University of Bologna, Bologna, 5Facility of Genetic and Developmental Biology, AOU Sassari, Sassari, Italy; 6Sbarro Institute for Cancer Research and Molecular Medicine, Center for Biotechnology, College of Science and Technology, Temple University, Philadelphia, PA, USAAbstract: Mouse embryonic stem cells were previously observed along with mesenchymal stem cells from different sources, after being treated with a mixed ester of hyaluronan with butyric and retinoic acids, to show a significant increase in the yield of cardiogenic and vascular differentiated elements. The aim of the present study was to determine if stem cells derived from primitive fetal cells present in human amniotic fluid (hAFSCs and cultured in the presence of a mixture of hyaluronic (HA, butyric (BU, and retinoic (RA acids show a higher yield of differentiation toward the cardiovascular phenotype as compared with untreated cells. During the differentiation process elicited by exposure to HA + BU + RA, genes controlling pluripotency and plasticity of stem cells, such as Sox2, Nanog, and Oct4, were significantly downregulated at the transcriptional level. At this point, a significant increase in expression of genes controlling the appearance of cardiogenic and vascular lineages in HA + BU + RA-treated cells was observed. The protein expression levels typical of cardiac and vascular phenotypes, evaluated by Western blotting

  14. Identification of human erythroid lineage-committed progenitors.

    Science.gov (United States)

    Mori, Yasuo; Akashi, Koichi; Weissman, Irving L

    2016-05-01

    Elucidating the developmental pathway leading to erythrocytes and being able to isolate their progenitors is crucial to understanding and treating disorders of red cell imbalance such as anemia, myelodysplastic syndrome, and polycythemia vera. Endoglin (CD105) is a key marker for purifying mouse erythroid lineage-committed progenitors (EPs) from bone marrow. Herein, we show that human EPs can also be isolated from adult bone marrow. We identified three subfractions that possessed different expression patterns of CD105 and CD71 within the previously defined human megakaryocyte/erythrocyte progenitor (hMEP; Lineage-CD34(+)CD38(+)IL-3Rα(-)CD45RA(-)) population. Both CD71(-)CD105(-) and CD71(+)CD105(-) MEPs, at least in vitro, retained bipotency for the megakaryocyte (MegK) and erythrocyte (E) lineages, although the latter sub-population had a differentiation potential skewed toward the E-lineage. Notably, the differentiation output of the CD71(+)CD105(+) subset of cells within the MEP population was completely restricted to the E-lineage with the loss of MegK potential; thus, we termed CD71(+)CD105(-) MEPs and CD71(+)CD105(+) cells as E-biased MEPs (E-MEPs) and EPs, respectively. These previously unclassified populations may facilitate understanding of the molecular mechanisms governing human erythroid development and serve as potential therapeutic targets in disorders of the erythroid lineage. PMID:27263782

  15. Braveheart, a Long Noncoding RNA Required for Cardiovascular Lineage Commitment

    OpenAIRE

    Klattenhoff, Carla A.; Scheuermann, Johanna C.; Surface, Lauren E.; Bradley, Robert K.; Fields, Paul A.; Steinhauser, Matthew L.; Ding, Huiming; Torrey, Lillian; Haas, Simon; Abo, Ryan; Tabebordbar, Mohammadsharif; Lee, Richard T.; Burge, Christopher B.; Butty, Vincent; Boyer, Laurie

    2013-01-01

    Long noncoding RNAs (lncRNAs) are often expressed in a development-specific manner, yet little is known about their roles in lineage commitment. Here, we identified Braveheart (Bvht), a heart-associated lncRNA in mouse. Using multiple embryonic stem cell (ESC) differentiation strategies, we show that Bvht is required for progression of nascent mesoderm toward a cardiac fate. We find that Bvht is necessary for activation of a core cardiovascular gene network and functions upstream of mesoderm ...

  16. G9a/GLP-dependent histone H3K9me2 patterning during human hematopoietic stem cell lineage commitment

    OpenAIRE

    Chen, Xiaoji; Skutt-Kakaria, Kyobi; Davison, Jerry; Ou, Yang-Li; Choi, Edward; Malik, Punam; Loeb, Keith; Wood, Brent; Georges, George; Torok-Storb, Beverly; Paddison, Patrick J.

    2012-01-01

    G9a and GLP methyltransferases play key roles in mammalian development via H3K9me1/2 modifications, associated with transcriptional repression. However, little is known about H3K9me2 function in adult cells and tissues. Chen et al. now report that H3K9me2 chromatin territories—absent in primitive cells—are formed de novo during hematopoietic stem and progenitor cell (HSPC) lineage commitment. G9a/GLP activity promotes progressive H3K9me2 genomic patterning that spreads across most genic regio...

  17. Multilineage Priming of Enhancer Repertoires Precedes Commitment to the B and Myeloid Cell Lineages in Hematopoeitic Progenitors

    OpenAIRE

    Mercer, Elinore M.; Lin, Yin C; Benner, Christopher; Jhunjhunwala, Suchit; Dutkowski, Janusz; Flores, Martha; Sigvardsson, Mikael; Ideker, Trey; Glass, Christopher K.; Murre, Cornelis

    2011-01-01

    Recent studies have documented genome-wide binding patterns of transcriptional regulators and their associated epigenetic marks in hematopoietic cell lineages. In order to determine how epigenetic marks are established and maintained during developmental progression, we have generated long-term cultures of hematopoietic progenitors by enforcing the expression of the E-protein antagonist Id2. Hematopoietic progenitors that express Id2 are multipotent and readily differentiate upon withdrawal o...

  18. Integrating extrinsic and intrinsic cues into a minimal model of lineage commitment for hematopoietic progenitors.

    Directory of Open Access Journals (Sweden)

    Santhosh Palani

    2009-09-01

    Full Text Available Autoregulation of transcription factors and cross-antagonism between lineage-specific transcription factors are a recurrent theme in cell differentiation. An equally prevalent event that is frequently overlooked in lineage commitment models is the upregulation of lineage-specific receptors, often through lineage-specific transcription factors. Here, we use a minimal model that combines cell-extrinsic and cell-intrinsic elements of regulation in order to understand how both instructive and stochastic events can inform cell commitment decisions in hematopoiesis. Our results suggest that cytokine-mediated positive receptor feedback can induce a "switch-like" response to external stimuli during multilineage differentiation by providing robustness to both bipotent and committed states while protecting progenitors from noise-induced differentiation or decommitment. Our model provides support to both the instructive and stochastic theories of commitment: cell fates are ultimately driven by lineage-specific transcription factors, but cytokine signaling can strongly bias lineage commitment by regulating these inherently noisy cell-fate decisions with complex, pertinent behaviors such as ligand-mediated ultrasensitivity and robust multistability. The simulations further suggest that the kinetics of differentiation to a mature cell state can depend on the starting progenitor state as well as on the route of commitment that is chosen. Lastly, our model shows good agreement with lineage-specific receptor expression kinetics from microarray experiments and provides a computational framework that can integrate both classical and alternative commitment paths in hematopoiesis that have been observed experimentally.

  19. A boost of BMP4 accelerates the commitment of human embryonic stem cells to the endothelial lineage.

    Science.gov (United States)

    Goldman, Orit; Feraud, Olivier; Boyer-Di Ponio, Julie; Driancourt, Catherine; Clay, Denis; Le Bousse-Kerdiles, Marie-Caroline; Bennaceur-Griscelli, Annelise; Uzan, Georges

    2009-08-01

    Embryoid bodies (EBs) generated during differentiation of human embryonic stem cells (hESCs) contain vascular-like structures, suggesting that commitment of mesoderm progenitors into endothelial cells occurs spontaneously. We showed that bone morphogenetic protein 4 (BMP4), an inducer of mesoderm, accelerates the peak expression of CD133/kinase insert domain-containing receptor (KDR) and CD144/KDR. Because the CD133(+)KDR(+) population could represent endothelial progenitors, we sorted them at day 7 and cultured them in endothelial medium. These cells were, however, unable to differentiate into endothelial cells. Under standard conditions, the CD144(+)KDR(+) population represents up to 10% of the total cells at day 12. In culture, these cells, if sorted, give rise to a homogeneous population with a morphology typical of endothelial cells and express endothelial markers. These endothelial cells derived from the day 12 sorted population were functional, as assessed by different in vitro assays. When EBs were stimulated by BMP4, the CD144(+)KDR(+) peak was shifted to day 7. Most of these cells, however, were CD31(-), becoming CD31(+) in culture. They then expressed von Willebrand factor and were functional. This suggests that, initially, the BMP4-boosted day 7, CD144(+)KDR(+)CD31(-) population represents immature endothelial cells that differentiate into mature endothelial cells in culture. The expression of OCT3/4, a marker of immaturity for hESCs decreases during EB differentiation, decreasing faster following BMP4 induction. We also show that BMP4 inhibits the global expression of GATA2 and RUNX1, two transcription factors involved in hemangioblast formation, at day 7 and day 12. PMID:19544443

  20. Commitment of Satellite Cells Expressing the Calcium Channel α2δ1 Subunit to the Muscle Lineage

    Directory of Open Access Journals (Sweden)

    Tammy Tamayo

    2012-01-01

    Full Text Available Satellite cells can maintain or repair muscle because they possess stem cell properties, making them a valuable option for cell therapy. However, cell transplants into skeletal muscle of patients with muscular dystrophy are limited by donor cell attachment, migration, and survival in the host tissue. Cells used for therapy are selected based on specific markers present in the plasma membrane. Although many markers have been identified, there is a need to find a marker that is expressed at different states in satellite cells, activated, quiescent, or differentiated cell. Furthermore, the marker has to be present in human tissue. Recently we reported that the plasma membrane α2δ1 protein is involved in cell attachment and migration in myoblasts. The α2δ1 subunit forms a part of the L-type voltage-dependent calcium channel in adult skeletal muscle. We found that the α2δ1 subunit is expressed in the majority of newly isolated satellite cells and that it appears earlier than the α1 subunits and at higher levels than the β or γ subunits. We also found that those cells that expressed α2δ1 would differentiate into muscle cells. This evidence indicates that the α2δ1 may be used as a marker of satellite cells that will differentiate into muscle.

  1. Transcriptional, epigenetic and retroviral signatures identify regulatory regions involved in hematopoietic lineage commitment.

    Science.gov (United States)

    Romano, Oriana; Peano, Clelia; Tagliazucchi, Guidantonio Malagoli; Petiti, Luca; Poletti, Valentina; Cocchiarella, Fabienne; Rizzi, Ermanno; Severgnini, Marco; Cavazza, Alessia; Rossi, Claudia; Pagliaro, Pasqualepaolo; Ambrosi, Alessandro; Ferrari, Giuliana; Bicciato, Silvio; De Bellis, Gianluca; Mavilio, Fulvio; Miccio, Annarita

    2016-01-01

    Genome-wide approaches allow investigating the molecular circuitry wiring the genetic and epigenetic programs of human somatic stem cells. Hematopoietic stem/progenitor cells (HSPC) give rise to the different blood cell types; however, the molecular basis of human hematopoietic lineage commitment is poorly characterized. Here, we define the transcriptional and epigenetic profile of human HSPC and early myeloid and erythroid progenitors by a combination of Cap Analysis of Gene Expression (CAGE), ChIP-seq and Moloney leukemia virus (MLV) integration site mapping. Most promoters and transcripts were shared by HSPC and committed progenitors, while enhancers and super-enhancers consistently changed upon differentiation, indicating that lineage commitment is essentially regulated by enhancer elements. A significant fraction of CAGE promoters differentially expressed upon commitment were novel, harbored a chromatin enhancer signature, and may identify promoters and transcribed enhancers driving cell commitment. MLV-targeted genomic regions co-mapped with cell-specific active enhancers and super-enhancers. Expression analyses, together with an enhancer functional assay, indicate that MLV integration can be used to identify bona fide developmentally regulated enhancers. Overall, this study provides an overview of transcriptional and epigenetic changes associated to HSPC lineage commitment, and a novel signature for regulatory elements involved in cell identity. PMID:27095295

  2. A microRNA signature associated with chondrogenic lineage commitment

    Indian Academy of Sciences (India)

    Behnaz Bakhshandeh; Masoud Soleimani; Seyed Hassan Paylakhi; Nasser Ghaemi

    2012-08-01

    Generating appropriate cartilage for clinical applications to heal skeletal tissue loss is a major health concern. In this regard, cell-based approaches offer a potential therapeutic strategy for cartilage repair, although little is known about the precise mechanism of chondrogenesis. Unrestricted somatic stem cell (USSC) is considered as a suitable candidate because of its potential for differentiating into multiple cell types. Recent studies show that microRNAs (miRNAs) are involved in several biological processes including development and differentiation. To identify the chondro-specific miRNA signature, miRNA patterns of USSCs and differentiated chondrocytes were investigated using microarrays and validation by qPCR. Prior to these analyses, chondrogenic commitment of differentiated USSCs was verified by immunocytochemistry, specific staining and evaluation of some main chondrogenic marker genes. Various in silico explorations (for both putative targets and signalling pathways) and empirical analyses (miRNA transfections followed by qPCR of some chondrogenic indicators) were carried out to support our results. Transient modulation of multiple chondro-miRs (such as mir-630, mir-624 and mir-376) with chondrocyte targets (such as TGFbR, MAP3K, collagens, SMADs and cadherins) as mediators of chondrogenic signalling pathways including cell–cell interactions, TGF-beta, and MAPK signalling suggests a mechanism for genetic induction of chondrogenic differentiation. In conclusion, this research reveals more details about the allocation of USSCs into the chondrocytes through identification of miRNA signature which modulates targets and pathways required for chondrogenic lineage and could provide guidelines for future clinical treatments and anti-miRNA therapies.

  3. Transcriptional, epigenetic and retroviral signatures identify regulatory regions involved in hematopoietic lineage commitment

    OpenAIRE

    Romano, Oriana; Peano, Clelia; Tagliazucchi, Guidantonio Malagoli; Petiti, Luca; Poletti, Valentina; Cocchiarella, Fabienne; Rizzi, Ermanno; Severgnini, Marco; Cavazza, Alessia; Rossi, Claudia; Pagliaro, Pasqualepaolo; Ambrosi, Alessandro; Ferrari, Giuliana; Bicciato, Silvio; De Bellis, Gianluca

    2016-01-01

    Genome-wide approaches allow investigating the molecular circuitry wiring the genetic and epigenetic programs of human somatic stem cells. Hematopoietic stem/progenitor cells (HSPC) give rise to the different blood cell types; however, the molecular basis of human hematopoietic lineage commitment is poorly characterized. Here, we define the transcriptional and epigenetic profile of human HSPC and early myeloid and erythroid progenitors by a combination of Cap Analysis of Gene Expression (CAGE...

  4. The Earliest Thymic T Cell Progenitors Sustain B Cell and Myeloid Lineage Potentials

    Science.gov (United States)

    Luc, Sidinh; Luis, Tiago C.; Boukarabila, Hanane; Macaulay, Iain C.; Buza-Vidas, Natalija; Bouriez-Jones, Tiphaine; Lutteropp, Michael; Woll, Petter S.; Loughran, Stephen J.; Mead, Adam J.; Hultquist, Anne; Brown, John; Mizukami, Takuo; Matsuoka, Sahoko; Ferry, Helen; Anderson, Kristina; Duarte, Sara; Atkinson, Deborah; Soneji, Shamit; Domanski, Aniela; Farley, Alison; Sanjuan-Pla, Alejandra; Carella, Cintia; Patient, Roger; de Bruijn, Marella; Enver, Tariq; Nerlov, Claus; Blackburn, Clare; Godin, Isabelle; Jacobsen, Sten Eirik W.

    2012-01-01

    The stepwise commitment from hematopoietic stem cells in the bone marrow (BM) to T lymphocyte-restricted progenitors in the thymus represents a paradigm for understanding the requirement for distinct extrinsic cues during different stages of lineage restriction from multipotent to lineage restricted progenitors. However, the commitment stage at which progenitors migrate from the BM to the thymus remains unclear. Here we provide functional and molecular evidence at the single cell level that the earliest progenitors in the neonatal thymus possessed combined granulocyte-monocyte, T and B lymphocyte, but not megakaryocyte-erythroid lineage potential. These potentials were identical to those of thymus-seeding progenitors in the BM, which were closely related at the molecular level. These findings establish the distinct lineage-restriction stage at which the T lineage commitment transits from the BM to the remote thymus. PMID:22344248

  5. Primordial germ cells: the first cell lineage or the last cells standing?

    OpenAIRE

    Johnson, Andrew D.; Alberio, Ramiro

    2015-01-01

    Embryos of many animal models express germ line determinants that suppress transcription and mediate early germ line commitment, which occurs before the somatic cell lineages are established. However, not all animals segregate their germ line in this manner. The ‘last cell standing’ model describes primordial germ cell (PGC) development in axolotls, in which PGCs are maintained by an extracellular signalling niche, and germ line commitment occurs after gastrulation. Here, we propose that this...

  6. Transcriptional Heterogeneity and Lineage Commitment in Myeloid Progenitors

    DEFF Research Database (Denmark)

    Paul, Franziska; Arkin, Ya'ara; Giladi, Amir;

    2015-01-01

    Within the bone marrow, stem cells differentiate and give rise to diverse blood cell types and functions. Currently, hematopoietic progenitors are defined using surface markers combined with functional assays that are not directly linked with in vivo differentiation potential or gene regulatory...... mechanisms. Here, we comprehensively map myeloid progenitor subpopulations by transcriptional sorting of single cells from the bone marrow. We describe multiple progenitor subgroups, showing unexpected transcriptional priming toward seven differentiation fates but no progenitors with a mixed state...... that in vivo priming may still allow for plasticity given strong perturbations. These data establish a reference model and general framework for studying hematopoiesis at single-cell resolution....

  7. Transcriptional Heterogeneity and Lineage Commitment in Myeloid Progenitors.

    Science.gov (United States)

    Paul, Franziska; Arkin, Ya'ara; Giladi, Amir; Jaitin, Diego Adhemar; Kenigsberg, Ephraim; Keren-Shaul, Hadas; Winter, Deborah; Lara-Astiaso, David; Gury, Meital; Weiner, Assaf; David, Eyal; Cohen, Nadav; Lauridsen, Felicia Kathrine Bratt; Haas, Simon; Schlitzer, Andreas; Mildner, Alexander; Ginhoux, Florent; Jung, Steffen; Trumpp, Andreas; Porse, Bo Torben; Tanay, Amos; Amit, Ido

    2015-12-17

    Within the bone marrow, stem cells differentiate and give rise to diverse blood cell types and functions. Currently, hematopoietic progenitors are defined using surface markers combined with functional assays that are not directly linked with in vivo differentiation potential or gene regulatory mechanisms. Here, we comprehensively map myeloid progenitor subpopulations by transcriptional sorting of single cells from the bone marrow. We describe multiple progenitor subgroups, showing unexpected transcriptional priming toward seven differentiation fates but no progenitors with a mixed state. Transcriptional differentiation is correlated with combinations of known and previously undefined transcription factors, suggesting that the process is tightly regulated. Histone maps and knockout assays are consistent with early transcriptional priming, while traditional transplantation experiments suggest that in vivo priming may still allow for plasticity given strong perturbations. These data establish a reference model and general framework for studying hematopoiesis at single-cell resolution. PMID:26627738

  8. A committed hemopoietic precursor to innate lymphoid cells

    Science.gov (United States)

    Constantinides, Michael G.; McDonald, Benjamin D.; Verhoef, Philip A.; Bendelac, Albert

    2014-01-01

    Innate lymphoid cells (ILC) specialize in the rapid secretion of polarized sets of cytokines and chemokines to combat infection and promote tissue repair at mucosal barriers.1–9 Their diversity and similarities with previously characterized NK cells and lymphoid tissue inducers (LTi) have prompted a provisional classification of all innate lymphocytes into groups 1, 2 and 3 based solely on cytokine properties,10 but their developmental pathways and lineage relationships remain elusive. Using lineage tracing and transfer studies, we identified and characterized a novel subset of lymphoid precursors in fetal liver and adult bone marrow that transiently expressed high amounts of PLZF, a transcription factor previously associated with NKT cell development.11,12 PLZFhigh cells were committed ILC progenitors with multiple ILC1, ILC2 and ILC3 potential at the clonal level. They excluded classical LTi and NK cells, but included a peculiar subset of NK1.1+DX5− ‘NK-like’ cells residing in the liver. Deletion of PLZF markedly altered the development of several ILC subsets, but not LTi or NK cells. PLZFhigh precursors also expressed high amounts of Id2 and GATA3, as well as TOX, a known regulator of PLZF-independent NK and LTi lineages.13 These findings establish novel lineage relationships between ILC, NK and LTi cells, and identify the common precursor to ILC, termed ILCP. They also reveal the broad, defining role of PLZF in the differentiation of innate lymphocytes. PMID:24509713

  9. GATA3 induces human T-cell commitment by restraining Notch activity and repressing NK-cell fate.

    Science.gov (United States)

    Van de Walle, Inge; Dolens, Anne-Catherine; Durinck, Kaat; De Mulder, Katrien; Van Loocke, Wouter; Damle, Sagar; Waegemans, Els; De Medts, Jelle; Velghe, Imke; De Smedt, Magda; Vandekerckhove, Bart; Kerre, Tessa; Plum, Jean; Leclercq, Georges; Rothenberg, Ellen V; Van Vlierberghe, Pieter; Speleman, Frank; Taghon, Tom

    2016-01-01

    The gradual reprogramming of haematopoietic precursors into the T-cell fate is characterized by at least two sequential developmental stages. Following Notch1-dependent T-cell lineage specification during which the first T-cell lineage genes are expressed and myeloid and dendritic cell potential is lost, T-cell specific transcription factors subsequently induce T-cell commitment by repressing residual natural killer (NK)-cell potential. How these processes are regulated in human is poorly understood, especially since efficient T-cell lineage commitment requires a reduction in Notch signalling activity following T-cell specification. Here, we show that GATA3, in contrast to TCF1, controls human T-cell lineage commitment through direct regulation of three distinct processes: repression of NK-cell fate, upregulation of T-cell lineage genes to promote further differentiation and restraint of Notch activity. Repression of the Notch1 target gene DTX1 hereby is essential to prevent NK-cell differentiation. Thus, GATA3-mediated positive and negative feedback mechanisms control human T-cell lineage commitment. PMID:27048872

  10. Transcriptional Repression of Gata3 Is Essential for Early B Cell Commitment

    OpenAIRE

    Banerjee, Anupam; Northrup, Daniel; Boukarabila, Hanane; Jacobsen, Sten Erik W.; Allman, David

    2013-01-01

    Summary The mechanisms underlying the silencing of alternative fate potentials in very early B cell precursors remain unclear. Using gain- and loss-of-function approaches together with a synthetic Zinc-finger polypeptide (6ZFP) engineered to prevent transcription factor binding to a defined cis element, we show that the transcription factor EBF1 promotes B cell lineage commitment by directly repressing expression of the T-cell-lineage-requisite Gata3 gene. Ebf1-deficient lymphoid progenitors ...

  11. Primordial germ cells: the first cell lineage or the last cells standing?

    Science.gov (United States)

    Johnson, Andrew D; Alberio, Ramiro

    2015-08-15

    Embryos of many animal models express germ line determinants that suppress transcription and mediate early germ line commitment, which occurs before the somatic cell lineages are established. However, not all animals segregate their germ line in this manner. The 'last cell standing' model describes primordial germ cell (PGC) development in axolotls, in which PGCs are maintained by an extracellular signalling niche, and germ line commitment occurs after gastrulation. Here, we propose that this 'stochastic' mode of PGC specification is conserved in vertebrates, including non-rodent mammals. We postulate that early germ line segregation liberates genetic regulatory networks for somatic development to evolve, and that it therefore emerged repeatedly in the animal kingdom in response to natural selection. PMID:26286941

  12. Trophoblast lineage cells derived from human induced pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Ying, E-mail: ying.chen@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Wang, Kai; Chandramouli, Gadisetti V.R. [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Knott, Jason G. [Developmental Epigenetics Laboratory, Department of Animal Science, Michigan State University (United States); Leach, Richard, E-mail: Richard.leach@hc.msu.edu [Department of Obstetrics, Gynecology and Reproductive Biology, Michigan State University, 333 Bostwick NE, Grand Rapids, MI 49503 (United States); Department of Obstetrics, Gynecology and Women’s Health, Spectrum Health Medical Group (United States)

    2013-07-12

    Highlights: •Epithelial-like phenotype of trophoblast lineage cells derived from human iPS cells. •Trophoblast lineage cells derived from human iPS cells exhibit trophoblast function. •Trophoblasts from iPS cells provides a proof-of-concept in regenerative medicine. -- Abstract: Background: During implantation, the blastocyst trophectoderm attaches to the endometrial epithelium and continues to differentiate into all trophoblast subtypes, which are the major components of a placenta. Aberrant trophoblast proliferation and differentiation are associated with placental diseases. However, due to ethical and practical issues, there is almost no available cell or tissue source to study the molecular mechanism of human trophoblast differentiation, which further becomes a barrier to the study of the pathogenesis of trophoblast-associated diseases of pregnancy. In this study, our goal was to generate a proof-of-concept model for deriving trophoblast lineage cells from induced pluripotency stem (iPS) cells from human fibroblasts. In future studies the generation of trophoblast lineage cells from iPS cells established from patient’s placenta will be extremely useful for studying the pathogenesis of individual trophoblast-associated diseases and for drug testing. Methods and results: Combining iPS cell technology with BMP4 induction, we derived trophoblast lineage cells from human iPS cells. The gene expression profile of these trophoblast lineage cells was distinct from fibroblasts and iPS cells. These cells expressed markers of human trophoblasts. Furthermore, when these cells were differentiated they exhibited invasive capacity and placental hormone secretive capacity, suggesting extravillous trophoblasts and syncytiotrophoblasts. Conclusion: Trophoblast lineage cells can be successfully derived from human iPS cells, which provide a proof-of-concept tool to recapitulate pathogenesis of patient placental trophoblasts in vitro.

  13. Trophoblast lineage cells derived from human induced pluripotent stem cells

    International Nuclear Information System (INIS)

    Highlights: •Epithelial-like phenotype of trophoblast lineage cells derived from human iPS cells. •Trophoblast lineage cells derived from human iPS cells exhibit trophoblast function. •Trophoblasts from iPS cells provides a proof-of-concept in regenerative medicine. -- Abstract: Background: During implantation, the blastocyst trophectoderm attaches to the endometrial epithelium and continues to differentiate into all trophoblast subtypes, which are the major components of a placenta. Aberrant trophoblast proliferation and differentiation are associated with placental diseases. However, due to ethical and practical issues, there is almost no available cell or tissue source to study the molecular mechanism of human trophoblast differentiation, which further becomes a barrier to the study of the pathogenesis of trophoblast-associated diseases of pregnancy. In this study, our goal was to generate a proof-of-concept model for deriving trophoblast lineage cells from induced pluripotency stem (iPS) cells from human fibroblasts. In future studies the generation of trophoblast lineage cells from iPS cells established from patient’s placenta will be extremely useful for studying the pathogenesis of individual trophoblast-associated diseases and for drug testing. Methods and results: Combining iPS cell technology with BMP4 induction, we derived trophoblast lineage cells from human iPS cells. The gene expression profile of these trophoblast lineage cells was distinct from fibroblasts and iPS cells. These cells expressed markers of human trophoblasts. Furthermore, when these cells were differentiated they exhibited invasive capacity and placental hormone secretive capacity, suggesting extravillous trophoblasts and syncytiotrophoblasts. Conclusion: Trophoblast lineage cells can be successfully derived from human iPS cells, which provide a proof-of-concept tool to recapitulate pathogenesis of patient placental trophoblasts in vitro

  14. Pax6 downregulation mediates abnormal lineage commitment of the ocular surface epithelium in aqueous-deficient dry eye disease.

    Directory of Open Access Journals (Sweden)

    Ying Ting Chen

    Full Text Available Keratinizing squamous metaplasia (SQM of the ocular surface is a blinding consequence of systemic autoimmune disease and there is no cure. Ocular SQM is traditionally viewed as an adaptive tissue response during chronic keratoconjunctivitis sicca (KCS that provokes pathological keratinization of the corneal epithelium and fibrosis of the corneal stroma. Recently, we established the autoimmune regulator-knockout (Aire KO mouse as a model of autoimmune KCS and identified an essential role for autoreactive CD4+ T cells in SQM pathogenesis. In subsequent studies, we noted the down-regulation of paired box gene 6 (Pax6 in both human patients with chronic KCS associated with Sjögren's syndrome and Aire KO mice. Pax6 encodes a pleiotropic transcription factor guiding eye morphogenesis during development. While the postnatal function of Pax6 is largely unknown, we hypothesized that its role in maintaining ocular surface homeostasis was disrupted in the inflamed eye and that loss of Pax6 played a functional role in the initiation and progression of SQM. Adoptive transfer of autoreactive T cells from Aire KO mice to immunodeficient recipients confirmed CD4+ T cells as the principal downstream effectors promoting Pax6 downregulation in Aire KO mice. CD4+ T cells required local signaling via Interleukin-1 receptor (IL-1R1 to provoke Pax6 loss, which prompted a switch from corneal-specific cytokeratin, CK12, to epidermal-specific CK10. The functional role of Pax6 loss in SQM pathogenesis was indicated by the reversal of SQM and restoration of ocular surface homeostasis following forced expression of Pax6 in corneal epithelial cells using adenovirus. Thus, tissue-restricted restoration of Pax6 prevented aberrant epidermal-lineage commitment suggesting adjuvant Pax6 gene therapy may represent a novel therapeutic approach to prevent SQM in patients with chronic inflammatory diseases of the ocular surface.

  15. 采用多功能流式细胞术分析分选造血干细胞和髓系定向分化祖细胞%Sorting and analysis of hematopoietic stem cells and myeloid lineage-committed progenitors using flow cytometry

    Institute of Scientific and Technical Information of China (English)

    崔巍; 许晓东; 许勇钢; 汪玄

    2008-01-01

    目的 探讨富集纯化造血干细胞(HSC)和髓系定向分化祖细胞的新实验方案.方法 根据造血干细胞和定向分化祖细胞在发育过程中表达某些特异性分化抗原的特性,通过免疫磁珠分选技术结合四色和六色流式细胞术分析14只健康小鼠的骨髓造血干细胞、造血祖细胞及定向分化祖细胞系列的表达,并对其进行分选,以进一步通过集落细胞培养和传代试验对分选后细胞的活性进行检测.结果 经上述实验方案分析,14只健康小鼠骨髓造血祖细胞(HPC)的表达率约为HSC的10倍;但其牛成活性远不如造血干细胞,共同髓系祖细胞(CMP)的传代能力仅为HSC的1/2,且次级分化的粒系单核系祖细胞(GMP)和红系巨核系祖细胞(MEP)的生成活性更弱,其传代次数为零.结论 通过多色流式细胞术实验方案可以分析纯化HSC和髓系定向分化祖细胞的表达,并精确计数HSC和祖细胞.%Objective To study the experimental protocol for purification and analysis of hematopoietic stem cells(HSC)and myeloid lineage-committed progenitors.Methods According to differentiation antigen expression pattern on hematopoietic stem cells(HSC) and progenitors during hematopoietic development,HSC and progenitors from bone marrow of 14 healthy mice were analyzed and sorted by magnetic nanoparticles and 4-color or 6-color flow cytometry using multiple antibody panels.Sorted HSC and progenitors were further tested by methylcellulose colony forming unit(CFU)and serial replatingassays.Results The expression of hematopoietic progenitor cells(HPC)was 10-fold higher expression than that of HSC.However,replating activity of common myeloid rogenitors(CMP)was only half of that of HSC.And there was almost 120 replating activity observed in granulocyte/macrophage lineage-restricted progenitors(GMP)and megakaryocyte/erythroeyte lineage-restricted progenitors(MEP).Conclusion Multiparametric flow cytometry could be used to isolate and

  16. Stat3 inhibition in neural lineage cells.

    Science.gov (United States)

    Chiba, Tomohiro; Mack, Laura; Delis, Natalia; Brill, Boris; Groner, Bernd

    2012-06-01

    Abstract Deregulation of signal transducer and activator of transcription 3 (Stat3) is attracting attentions in neurological disorders of elderly populations, e.g., Stat3 is inactivated in hippocampal neurons of Alzheimer's disease (AD) brains, whereas it is often constitutively activated in glioblastoma multiforme (GBM), correlating with poor prognosis. Stat3-inhibiting drugs have been intensively developed for chemotherapy based on the fact that GBM, in many cases, are "addicted" to Stat3 activation. Stat3 inhibitors, however, potentially have unfavorable side effects on postmitotic neurons, normal permanent residents in the central nervous system. It is, therefore, of great importance to address detailed cellular responses of neural lineage cells including normal neurons, astrocytes, and neuronal/glial cancer cell lines to several classes of Stat3 inhibitors focusing on their effective concentrations. Here, we picked up five human and mouse cancer cell lines (Neuro-2a and SH-SY5Y neuroblastoma cell lines and Tu-9648, U-87MG, and U-373MG glioblastoma cell lines) and treated with various Stat3 inhibitors. Among them, Stattic, FLLL31, and resveratrol potently suppressed P-Stat3 and cell viability in all the tested cell lines. Stat3 knockdown or expression of dominant-negative Stat3 further sensitized cells to the inhibitors. Expression of familial AD-related mutant amyloid precursor protein sensitized neuronal cells, not glial cells, to Stat3 inhibitors by reducing P-Stat3 levels. Primary neurons and astrocytes also responded to Stat3 inhibitors with similar sensitivities to those observed in cancer cell lines. Thus, Stat3 inhibitors should be carefully targeted to GBM cells to avoid potential neurotoxicity leading to AD-like neuropsychiatric dysfunctions. PMID:25436682

  17. Lineage Switching in Acute Leukemias: A Consequence of Stem Cell Plasticity?

    Directory of Open Access Journals (Sweden)

    Elisa Dorantes-Acosta

    2012-01-01

    Full Text Available Acute leukemias are the most common cancer in childhood and characterized by the uncontrolled production of hematopoietic precursor cells of the lymphoid or myeloid series within the bone marrow. Even when a relatively high efficiency of therapeutic agents has increased the overall survival rates in the last years, factors such as cell lineage switching and the rise of mixed lineages at relapses often change the prognosis of the illness. During lineage switching, conversions from lymphoblastic leukemia to myeloid leukemia, or vice versa, are recorded. The central mechanisms involved in these phenomena remain undefined, but recent studies suggest that lineage commitment of plastic hematopoietic progenitors may be multidirectional and reversible upon specific signals provided by both intrinsic and environmental cues. In this paper, we focus on the current knowledge about cell heterogeneity and the lineage switch resulting from leukemic cells plasticity. A number of hypothetical mechanisms that may inspire changes in cell fate decisions are highlighted. Understanding the plasticity of leukemia initiating cells might be fundamental to unravel the pathogenesis of lineage switch in acute leukemias and will illuminate the importance of a flexible hematopoietic development.

  18. Single-cell analysis defines the divergence between the innate lymphoid cell lineage and lymphoid tissue-inducer cell lineage.

    Science.gov (United States)

    Ishizuka, Isabel E; Chea, Sylvestre; Gudjonson, Herman; Constantinides, Michael G; Dinner, Aaron R; Bendelac, Albert; Golub, Rachel

    2016-03-01

    The precise lineage relationship between innate lymphoid cells (ILCs) and lymphoid tissue-inducer (LTi) cells is poorly understood. Using single-cell multiplex transcriptional analysis of 100 lymphoid genes and single-cell cultures of fetal liver precursor cells, we identified the common proximal precursor to these lineages and found that its bifurcation was marked by differential induction of the transcription factors PLZF and TCF1. Acquisition of individual effector programs specific to the ILC subsets ILC1, ILC2 and ILC3 was initiated later, at the common ILC precursor stage, by transient expression of mixed ILC1, ILC2 and ILC3 transcriptional patterns, whereas, in contrast, the development of LTi cells did not go through multilineage priming. Our findings provide insight into the divergent mechanisms of the differentiation of the ILC lineage and LTi cell lineage and establish a high-resolution 'blueprint' of their development. PMID:26779601

  19. Lineage-restricted expression of homeobox-containing genes in human hematopoietic cell lines

    International Nuclear Information System (INIS)

    The authors investigated the role of homeobox-containing genes in human hematopoiesis because homeobox genes (i) control cell fate in the Drosophila embryo, (ii) are expressed in specific patterns in human embryos, and (iii) appear to function as transcription factors that control cell phenotype in other mammalian organs. Using four homeobox probes from the HOX2 locus and a previously undescribed homeobox cDNA (PL1), they screened mRNAs from 18 human leukemic cell lines representing erythroid, myeloid, and T- and B-cell lineages. Complex patterns of lineage-restricted expression are observed. No single homeobox gene is expressed in all types of hematopoietic cells, but each cell type exhibits homeobox gene expression. They have demonstrated (i) lineage-restricted expression of five homeobox genes in erythroid and monocytic cell lines; (ii) expression of additional homeobox genes in other cell lineages (HL-60 and lymphoid cells); (iii) expression of one homeobox gene in normal marrow cells; and (iv) modulation of expression during differentiation. These data suggest that these genes play a role in human hematopoietic development and lineage commitment

  20. Nf1 Haploinsufficiency Alters Myeloid Lineage Commitment and Function, Leading to Deranged Skeletal Homeostasis.

    Science.gov (United States)

    Rhodes, Steven D; Yang, Hao; Dong, Ruizhi; Menon, Keshav; He, Yongzheng; Li, Zhaomin; Chen, Shi; Staser, Karl W; Jiang, Li; Wu, Xiaohua; Yang, Xianlin; Peng, Xianghong; Mohammad, Khalid S; Guise, Theresa A; Xu, Mingjiang; Yang, Feng-Chun

    2015-10-01

    Although nullizygous loss of NF1 leads to myeloid malignancies, haploinsufficient loss of NF1 (Nf1) has been shown to contribute to osteopenia and osteoporosis which occurs in approximately 50% of neurofibromatosis type 1 (NF1) patients. Bone marrow mononuclear cells of haploinsufficient NF1 patients and Nf1(+/-) mice exhibit increased osteoclastogenesis and accelerated bone turnover; however, the culprit hematopoietic lineages responsible for perpetuating these osteolytic manifestations have yet to be elucidated. Here we demonstrate that conditional inactivation of a single Nf1 allele within the myeloid progenitor cell population (Nf1-LysM) is necessary and sufficient to promote multiple osteoclast gains-in-function, resulting in enhanced osteoclastogenesis and accelerated osteoclast bone lytic activity in response to proresorptive challenge in vivo. Surprisingly, mice conditionally Nf1 heterozygous in mature, terminally differentiated osteoclasts (Nf1-Ctsk) do not exhibit any of these skeletal phenotypes, indicating a critical requirement for Nf1 haploinsufficiency at a more primitive/progenitor stage of myeloid development in perpetuating osteolytic activity. We further identified p21Ras-dependent hyperphosphorylation of Pu.1 within the nucleus of Nf1 haploinsufficient myelomonocytic osteoclast precursors, providing a novel therapeutic target for the potential treatment of NF1 associated osteolytic manifestations. PMID:25917016

  1. Id1 restrains myeloid commitment, maintaining the self-renewal capacity of hematopoietic stem cells

    OpenAIRE

    Jankovic, Vladimir; Ciarrocchi, Alessia; Boccuni, Piernicola; Deblasio, Tony; Benezra, Robert; Nimer, Stephen D.

    2007-01-01

    Appropriate hematopoietic stem cell (HSC) self-renewal reflects the tight regulation of cell cycle entry and lineage commitment. Here, we show that Id1, a dominant-negative regulator of E protein transcription factors, maintains HSC self-renewal by preserving the undifferentiated state. Id1-deficient HSCs show increased cell cycling, by BrdU incorporation in vivo, but fail to efficiently self-renew, leading to low steady-state HSC numbers and premature exhaustion in serial bone marrow transpl...

  2. Data defining markers of human neural stem cell lineage potential.

    Science.gov (United States)

    Oikari, Lotta E; Okolicsanyi, Rachel K; Griffiths, Lyn R; Haupt, Larisa M

    2016-06-01

    Neural stem cells (NSCs) and neural progenitor cells (NPCs) are self-renewing and multipotent cells, however, NPCs are considered to be more lineage-restricted with a reduced self-renewing capacity. We present data comparing the expression of 21 markers encompassing pluripotency, self-renewal (NSC) as well as neuronal and glial (astrocyte and oligodendrocyte) lineage specification and 28 extracellular proteoglycan (PG) genes and their regulatory enzymes between embryonic stem cell (ESC)-derived human NSCs (hNSC H9 cells, Thermo Fisher) and human cortex-derived normal human NPCs (nhNPCs, Lonza). The data demonstrates expression differences of multiple lineage and proteoglycan-associated genes between hNSC H9 cells and nhNPCs. Data interpretation of markers and proteoglycans defining NSC and neural cell lineage characterisation can be found in "Cell surface heparan sulfate proteoglycans as novel markers of human neural stem cell fate determination" (Oikari et al. 2015) [1]. PMID:26958640

  3. Asynchronous combinatorial action of four regulatory factors activates Bcl11b for T cell commitment.

    Science.gov (United States)

    Kueh, Hao Yuan; Yui, Mary A; Ng, Kenneth K H; Pease, Shirley S; Zhang, Jingli A; Damle, Sagar S; Freedman, George; Siu, Sharmayne; Bernstein, Irwin D; Elowitz, Michael B; Rothenberg, Ellen V

    2016-08-01

    During T cell development, multipotent progenitors relinquish competence for other fates and commit to the T cell lineage by turning on Bcl11b, which encodes a transcription factor. To clarify lineage commitment mechanisms, we followed developing T cells at the single-cell level using Bcl11b knock-in fluorescent reporter mice. Notch signaling and Notch-activated transcription factors collaborate to activate Bcl11b expression irrespectively of Notch-dependent proliferation. These inputs work via three distinct, asynchronous mechanisms: an early locus 'poising' function dependent on TCF-1 and GATA-3, a stochastic-permissivity function dependent on Notch signaling, and a separate amplitude-control function dependent on Runx1, a factor already present in multipotent progenitors. Despite their necessity for Bcl11b expression, these inputs act in a stage-specific manner, providing a multitiered mechanism for developmental gene regulation. PMID:27376470

  4. GATA-3 dose-dependent checkpoints in early T cell commitment1

    OpenAIRE

    Scripture-Adams, Deirdre D.; Damle, Sagar S.; Li, Long; Elihu, Koorosh J.; Qin, Shuyang; Arias, Alexandra M.; Butler, Robert R.; Champhekar, Ameya; Zhang, Jingli A.; Rothenberg, Ellen V.

    2014-01-01

    GATA-3 expression is crucial for T cell development and peaks during commitment to the T-cell lineage, midway through the CD4−CD8− (DN) 1-3 stages. We used RNA interference and conditional deletion to reduce GATA-3 protein acutely at specific points during T-cell differentiation in vitro. Even moderate GATA-3 reduction killed DN1 cells, delayed progression to DN2 stage, skewed DN2 gene regulation, and blocked appearance of DN3 phenotype. Although a Bcl-2 transgene rescued DN1 survival and imp...

  5. Stochastic dynamics of interacting haematopoietic stem cell niche lineages.

    Directory of Open Access Journals (Sweden)

    Tamás Székely

    2014-09-01

    Full Text Available Since we still know very little about stem cells in their natural environment, it is useful to explore their dynamics through modelling and simulation, as well as experimentally. Most models of stem cell systems are based on deterministic differential equations that ignore the natural heterogeneity of stem cell populations. This is not appropriate at the level of individual cells and niches, when randomness is more likely to affect dynamics. In this paper, we introduce a fast stochastic method for simulating a metapopulation of stem cell niche lineages, that is, many sub-populations that together form a heterogeneous metapopulation, over time. By selecting the common limiting timestep, our method ensures that the entire metapopulation is simulated synchronously. This is important, as it allows us to introduce interactions between separate niche lineages, which would otherwise be impossible. We expand our method to enable the coupling of many lineages into niche groups, where differentiated cells are pooled within each niche group. Using this method, we explore the dynamics of the haematopoietic system from a demand control system perspective. We find that coupling together niche lineages allows the organism to regulate blood cell numbers as closely as possible to the homeostatic optimum. Furthermore, coupled lineages respond better than uncoupled ones to random perturbations, here the loss of some myeloid cells. This could imply that it is advantageous for an organism to connect together its niche lineages into groups. Our results suggest that a potential fruitful empirical direction will be to understand how stem cell descendants communicate with the niche and how cancer may arise as a result of a failure of such communication.

  6. The Drosophila cyst stem cell lineage

    OpenAIRE

    Zoller, Richard; Schulz, Cordula

    2012-01-01

    In all animals, germline cells differentiate in intimate contact with somatic cells and interactions between germline and soma are particularly important for germline development and function. In the male gonad of Drosophila melanogaster, the developing germline cells are enclosed by somatic cyst cells. The cyst cells are derived from cyst stem cells (CySCs) of somatic origin and codifferentiate with the germline cells. The fast generation cycle and the genetic tractability of Drosophila has ...

  7. Differentiation into Endoderm Lineage: Pancreatic differentiation from Embryonic Stem Cells

    OpenAIRE

    Lee, Dong Hyeon; Chung, Hyung Min

    2011-01-01

    The endoderm gives rise to digestive and respiratory tracts, thyroid, liver, and pancreas. Representative disease of endoderm lineages is type 1 diabetes resulting from destruction of the insulin-producing β cells. Generation of functional β cells from human embryonic stem (ES) cells in vitro can be practical, renewable cell source for replacement cell therapy for type 1 diabetes. It has been achieved by progressive instructive differentiation through each of the developmental stages. In this...

  8. Architectural protein subclasses shape 3-D organization of genomes during lineage commitment

    OpenAIRE

    Phillips-Cremins, Jennifer E.; Sauria, Michael E. G.; Sanyal, Amartya; Gerasimova, Tatiana I; Lajoie, Bryan R.; Bell, Joshua S. K.; Ong, Chin-Tong; Hookway, Tracy A.; Guo, Changying; Sun, Yuhua; Bland, Michael J.; Wagstaff, William; Dalton, Stephen; McDevitt, Todd C.; Sen, Ranjan

    2013-01-01

    Understanding the topological configurations of chromatin may reveal valuable insights into how the genome and epigenome act in concert to control cell fate during development. Here we generate high-resolution architecture maps across seven genomic loci in embryonic stem cells and neural progenitor cells. We observe a hierarchy of 3-D interactions that undergo marked reorganization at the sub-Mb scale during differentiation. Distinct combinations of CTCF, Mediator, and cohesin show widespread...

  9. Bovine Mammary Epithelial Cell Lineages and Parenchymal Development

    Science.gov (United States)

    Mammary development proceeds from an aggregation of cells in the ventral ectoderm to the establishment of an elaborate tree of alveoli, ducts, and cisternae. However, despite abundant data on endocrine regulation of ruminant mammary growth, we know comparatively little about cell lineages, express...

  10. Independent Stem Cell Lineages Regulate Adipose Organogenesis and Adipose Homeostasis

    Directory of Open Access Journals (Sweden)

    Yuwei Jiang

    2014-11-01

    Full Text Available Adipose tissues have striking plasticity, highlighted by childhood and adult obesity. Using adipose lineage analyses, smooth muscle actin (SMA-mural cell-fate mapping, and conditional PPARγ deletion to block adipocyte differentiation, we find two phases of adipocyte generation that emanate from two independent adipose progenitor compartments: developmental and adult. These two compartments are sequentially required for organ formation and maintenance. Although both developmental and adult progenitors are specified during the developmental period and express PPARγ, they have distinct microanatomical, functional, morphogenetic, and molecular profiles. Furthermore, the two compartments derive from different lineages; whereas adult adipose progenitors fate-map from an SMA+ mural lineage, developmental progenitors do not. Remarkably, the adult progenitor compartment appears to be specified earlier than the developmental cells and then enters the already developmentally formed adipose depots. Thus, two distinct cell compartments control adipose organ development and organ homeostasis, which may provide a discrete therapeutic target for childhood and adult obesity.

  11. Mast cell diversion of T-lineage precursor cells by the essential T-lineage transcription factor GATA-3

    OpenAIRE

    Taghon, Tom; Yui, Mary A.; Rothenberg, Ellen V.

    2007-01-01

    GATA-3 is essential for T cell development from the earliest stages. However, highly abundant GATA-3 can drive T-lineage precursors to a non-T fate, depending on Notch signaling and developmental stage. GATA-3 overexpression blocked pro-T cell survival when Notch-Delta signals were present, but enhanced viability in their absence. In double-negative (DN1) and DN2 but not DN3 fetal thymocytes, GATA-3 overexpression rapidly induced mast cell lineage respecification with high frequency by direct...

  12. Chromatin dynamics in pollen mother cells underpin a common scenario at the somatic-to-reproductive fate transition of both the male and female lineages in Arabidopsis

    OpenAIRE

    She, Wenjing; Baroux, Célia

    2015-01-01

    Unlike animals, where the germline is established early during embryogenesis, plants set aside their reproductive lineage late in development in dedicated floral organs. The specification of pollen mother cells (PMC) committed to meiosis takes place in the sporogenous tissue in anther locules and marks the somatic-to-reproductive cell fate transition toward the male reproductive lineage. Here we show that Arabidopsis PMC differentiation is accompanied by large-scale changes in chromatin organ...

  13. Molecular Genetics of Ameloblast Cell Lineage

    OpenAIRE

    Bei, Marianna

    2009-01-01

    Late tooth morphogenesis is characterized by a series of events that determine crown morphogenesis and the histodifferentiation of epithelial cells into enamel-secreting ameloblasts and of mesenchymal cells into dentin-secreting odontoblasts. Functional ameloblasts are tall, columnar, polarized cells that synthesize and secrete a number of enamel-specific proteins. After depositing the full thickness of enamel matrix, ameloblasts shrink in size and regulate enamel maturation. Amelogenesis imp...

  14. Cytokine-Regulated GADD45G Induces Differentiation and Lineage Selection in Hematopoietic Stem Cells

    Directory of Open Access Journals (Sweden)

    Frederic B. Thalheimer

    2014-07-01

    Full Text Available The balance of self-renewal and differentiation in long-term repopulating hematopoietic stem cells (LT-HSC must be strictly controlled to maintain blood homeostasis and to prevent leukemogenesis. Hematopoietic cytokines can induce differentiation in LT-HSCs; however, the molecular mechanism orchestrating this delicate balance requires further elucidation. We identified the tumor suppressor GADD45G as an instructor of LT-HSC differentiation under the control of differentiation-promoting cytokine receptor signaling. GADD45G immediately induces and accelerates differentiation in LT-HSCs and overrides the self-renewal program by specifically activating MAP3K4-mediated MAPK p38. Conversely, the absence of GADD45G enhances the self-renewal potential of LT-HSCs. Videomicroscopy-based tracking of single LT-HSCs revealed that, once GADD45G is expressed, the development of LT-HSCs into lineage-committed progeny occurred within 36 hr and uncovered a selective lineage choice with a severe reduction in megakaryocytic-erythroid cells. Here, we report an unrecognized role of GADD45G as a central molecular linker of extrinsic cytokine differentiation and lineage choice control in hematopoiesis.

  15. Stochastic differentiation into an osteoclast lineage from cloned macrophage-like cells

    International Nuclear Information System (INIS)

    Highlights: ► The frequency of C7 differentiation into osteoclast was low and constant. ► Only extended C7 cell cultures exponentially increased osteoclast+ cultures. ► C7 cell differentiation into committed osteoclast precursors is on ‘autopilot’. ► The system may maintain the stem cell self-renewal and differentiation. -- Abstract: Differentiation into osteoclasts is induced by a macrophage colony-stimulating factor and receptor activator of nuclear-factor κB ligand. The macrophage-like cell line, C7 has the potential to differentiate into osteoclasts when it is cultured with both factors for 6 days. Although C7 is an established cell line, the frequency of differentiation into this lineage was less than 10%, and the ratio was maintained at a constant level, even after repeated cloning. In this study, to increase the differentiation of C7 cells to osteoclasts, C7 derivative treatments with several activators and/or inhibitors were performed for 3 days prior to setting osteoclast induction analysis; however, a reagent to significantly up-regulate the frequency of differentiation was not found. Only extended cultures for osteoclastogenesis exponentially increased the frequency of osteoclast precursors. It is likely that C7 cell differentiation into committed osteoclast precursors is on ‘autopilot’ rather than requiring specific signals to drive this process.

  16. A mex3 homolog is required for differentiation during planarian stem cell lineage development.

    Science.gov (United States)

    Zhu, Shu Jun; Hallows, Stephanie E; Currie, Ko W; Xu, ChangJiang; Pearson, Bret J

    2015-01-01

    Neoblasts are adult stem cells (ASCs) in planarians that sustain cell replacement during homeostasis and regeneration of any missing tissue. While numerous studies have examined genes underlying neoblast pluripotency, molecular pathways driving postmitotic fates remain poorly defined. In this study, we used transcriptional profiling of irradiation-sensitive and irradiation-insensitive cell populations and RNA interference (RNAi) functional screening to uncover markers and regulators of postmitotic progeny. We identified 32 new markers distinguishing two main epithelial progenitor populations and a planarian homolog to the MEX3 RNA-binding protein (Smed-mex3-1) as a key regulator of lineage progression. mex3-1 was required for generating differentiated cells of multiple lineages, while restricting the size of the stem cell compartment. We also demonstrated the utility of using mex3-1(RNAi) animals to identify additional progenitor markers. These results identified mex3-1 as a cell fate regulator, broadly required for differentiation, and suggest that mex3-1 helps to mediate the balance between ASC self-renewal and commitment. PMID:26114597

  17. Independent Stem Cell Lineages Regulate Adipose Organogenesis and Adipose Homeostasis

    OpenAIRE

    Yuwei Jiang; Daniel C. Berry; Wei Tang; Jonathan M. Graff

    2014-01-01

    Adipose tissues have striking plasticity, highlighted by childhood and adult obesity. Using adipose lineage analyses, smooth muscle actin (SMA)-mural cell-fate mapping, and conditional PPARγ deletion to block adipocyte differentiation, we find two phases of adipocyte generation that emanate from two independent adipose progenitor compartments: developmental and adult. These two compartments are sequentially required for organ formation and maintenance. Although both developmental and adult pr...

  18. Lineage relationship of effector and memory T cells

    Science.gov (United States)

    Restifo, Nicholas P.; Gattinoni, Luca

    2013-01-01

    Adaptive immunity is characterized by the ability to form long-lived immunological memory. Upon re-exposure to antigen, memory T cells respond more rapidly and robustly than naïve T cells, providing better clearance of pathogens. Recent reviews have reinforced the text-book view that memory T cells arise from effector cells. Although this notion is teleologically appealing, emerging data is more consistent with a model where naïve cells directly develop into memory cells without transitioning through an effector stage. A clear understanding of the lineage relationships between memory and effector cells has profound implications for the design of vaccines and for the development of effective T cell-based therapies. PMID:24148236

  19. Foetal stem cell derivation & characterization for osteogenic lineage

    Directory of Open Access Journals (Sweden)

    A Mangala Gowri

    2013-01-01

    Full Text Available Background & objectives: Mesencymal stem cells (MSCs derived from foetal tissues present a multipotent progenitor cell source for application in tissue engineering and regenerative medicine. The present study was carried out to derive foetal mesenchymal stem cells from ovine source and analyze their differentiation to osteogenic linage to serve as an animal model to predict human applications. Methods: Isolation and culture of sheep foetal bone marrow cells were done and uniform clonally derived MSC population was collected. The cells were characterized using cytochemical, immunophenotyping, biochemical and molecular analyses. The cells with defined characteristics were differentiated into osteogenic lineages and analysis for differentiated cell types was done. The cells were analyzed for cell surface marker expression and the gene expression in undifferentiated and differentiated osteoblast was checked by reverse transcriptase PCR (RT PCR analysis and confirmed by sequencing using genetic analyzer. Results: Ovine foetal samples were processed to obtain mononuclear (MNC cells which on culture showed spindle morphology, a characteristic oval body with the flattened ends. MSC population CD45 - /CD14 - was cultured by limiting dilution to arrive at uniform spindle morphology cells and colony forming units. The cells were shown to be positive for surface markers such as CD44, CD54, integrinβ1, and intracellular collagen type I/III and fibronectin. The osteogenically induced MSCs were analyzed for alkaline phosphatase (ALP activity and mineral deposition. The undifferentiated MSCs expressed RAB3B, candidate marker for stemness in MSCs. The osteogenically induced and uninduced MSCs expressed collagen type I and MMP13 gene in osteogenic induced cells. Interpretation & conclusions: The protocol for isolation of ovine foetal bone marrow derived MSCs was simple to perform, and the cultural method of obtaining pure spindle morphology cells was established

  20. B-lymphocyte lineage cells and the respiratory system.

    Science.gov (United States)

    Kato, Atsushi; Hulse, Kathryn E; Tan, Bruce K; Schleimer, Robert P

    2013-04-01

    Adaptive humoral immune responses in the airways are mediated by B cells and plasma cells that express highly evolved and specific receptors and produce immunoglobulins of most isotypes. In some cases, such as autoimmune diseases or inflammatory diseases caused by excessive exposure to foreign antigens, these same immune cells can cause disease by virtue of overly vigorous responses. This review discusses the generation, differentiation, signaling, activation, and recruitment pathways of B cells and plasma cells, with special emphasis on unique characteristics of subsets of these cells functioning within the respiratory system. The primary sensitization events that generate B cells responsible for effector responses throughout the airways usually occur in the upper airways, tonsils, and adenoid structures that make up the Waldeyer ring. On secondary exposure to antigen in the airways, antigen-processing dendritic cells migrate into secondary lymphoid organs, such as lymph nodes, that drain the upper and lower airways, and further B-cell expansion takes place at those sites. Antigen exposure in the upper or lower airways can also drive expansion of B-lineage cells in the airway mucosal tissue and lead to the formation of inducible lymphoid follicles or aggregates that can mediate local immunity or disease. PMID:23540615

  1. B lymphocyte lineage cells and the respiratory system

    Science.gov (United States)

    Kato, Atsushi; Hulse, Kathryn E.; Tan, Bruce K.; Schleimer, Robert P.

    2013-01-01

    Adaptive humoral immune responses in the airways are mediated by B cells and plasma cells that express highly evolved and specific receptors and produce immunoglobulins of most isotypes. In some cases, such as autoimmune diseases or inflammatory diseases caused by excessive exposure to foreign antigens, these same immune cells can cause disease by virtue of overly vigorous responses. This review discusses the generation, differentiation, signaling, activation and recruitment pathways of B cells and plasma cells, with special emphasis on unique characteristics of subsets of these cells functioning within the respiratory system. The primary sensitization events that generate B cells responsible for effector responses throughout the airways usually occur in the upper airways, in tonsils and adenoid structures that make up Waldeyer’s Ring. Upon secondary exposure to antigen in the airways, antigen-processing dendritic cells migrate into secondary lymphoid organs such as lymph nodes that drain the upper and lower airways and further B cell expansion takes place at those sites. Antigen exposure in the upper or lower airways can also drive expansion of B lineage cells in the airway mucosal tissue and lead to the formation of inducible lymphoid follicles or aggregates that can mediate local immunity or disease. PMID:23540615

  2. Identification and prospective isolation of a mesothelial precursor lineage giving rise to smooth muscle cells & fibroblasts for mammalian internal organs, and their vasculature

    OpenAIRE

    Rinkevich, Yuval; Mori, Taisuke; Sahoo, Debashis; Xu, Pin-Xian; Bermingham, John R., Jr.; Weissman, Irving L

    2012-01-01

    Fibroblasts and smooth muscle cells (FSMCs) are principal cell types of connective and adventitial tissues that participate in the development, physiology and pathology of internal organs, with incompletely defined cellular origins. Here, we identify and prospectively isolate from mesothelium a mouse cell lineage that is committed to FSMCs. Mesothelium is an epithelial monolayer covering the vertebrate thoracic and abdominal cavities and internal organs. Time-lapse imaging and transplantation...

  3. Creation of Primary Cell Lines from Lineage-Labeled Mouse Models of Cancer

    Science.gov (United States)

    Rhim, Andrew D.

    2015-01-01

    Frequently, it is necessary to isolate pure populations of cancer cells for downstream assays, such as transcriptional analysis, signaling studies, and the creation of noncontaminated primary cell lines. Genetic lineage labeling with fluorescent reporter alleles allows for the identification of epithelial-derived cells within tumors. This protocol describes a method to isolate lineage-labeled pancreatic epithelial cells for ex vivo analysis, but it can be adapted for any type of lineage-labeled tumor. PMID:25934932

  4. Creation of Primary Cell Lines from Lineage-Labeled Mouse Models of Cancer

    OpenAIRE

    Rhim, Andrew D.

    2015-01-01

    Frequently, it is necessary to isolate pure populations of cancer cells for downstream assays, such as transcriptional analysis, signaling studies, and the creation of noncontaminated primary cell lines. Genetic lineage labeling with fluorescent reporter alleles allows for the identification of epithelial-derived cells within tumors. This protocol describes a method to isolate lineage-labeled pancreatic epithelial cells for ex vivo analysis, but it can be adapted for any type of lineage-label...

  5. Differentiation in Stem Cell Lineages and in Life: Explorations in the Male Germ Line Stem Cell Lineage.

    Science.gov (United States)

    Fuller, Margaret T

    2016-01-01

    I have been privileged to work on cellular differentiation during a great surge of discovery that has revealed the molecular mechanisms and genetic regulatory circuitry that control embryonic development and adult tissue maintenance and repair. Studying the regulation of proliferation and differentiation in the male germ line stem cell lineage has allowed us investigate how the developmental program imposes layers of additional controls on fundamental cellular processes like cell cycle progression and gene expression to give rise to the huge variety of specialized cell types in our bodies. We are beginning to understand how local signals from somatic support cells specify self-renewal versus differentiation in the stem cell niche at the apical tip of the testis. We are discovering the molecular events that block cell proliferation and initiate terminal differentiation at the switch from mitosis to meiosis-a signature event of the germ cell program. Our work is beginning to reveal how the developmental program that sets up the dramatic new cell type-specific transcription program that prepares germ cells for meiotic division and spermatid differentiation is turned on when cells become spermatocytes. I have had the privilege of working with incredible students, postdocs, and colleagues who have discovered, brainstormed, challenged, and refined our science and our ideas of how developmental pathways and cellular mechanisms work together to drive differentiation. PMID:26970629

  6. Optical Imaging for Stem Cell Differentiation to Neuronal Lineage

    Energy Technology Data Exchange (ETDEWEB)

    Hwang, Do Won; Lee, Dong Soo [Seoul National Univ., Seoul (Korea, Republic of)

    2012-03-15

    In regenerative medicine, the prospect of stem cell therapy hold great promise for the recovery of injured tissues and effective treatment of intractable diseases. Tracking stem cell fate provides critical information to understand and evaluate the success of stem cell therapy. The recent emergence of in vivo noninvasive molecular imaging has enabled assessment of the behavior of grafted stem cells in living subjects. In this review, we provide an overview of current optical imaging strategies based on cell or tissue specific reporter gene expression and of in vivo methods to monitor stem cell differentiation into neuronal lineages. These methods use optical reporters either regulated by neuron-specific promoters or containing neuron-specific microRNA binding sites. Both systems revealed dramatic changes in optical reporter imaging signals in cells differentiating a yeast GAL4 amplification system or an engineering-enhanced luciferase reported gene. Furthermore, we propose an advanced imaging system to monitor neuronal differentiation during neurogenesis that uses in vivo multiplexed imaging techniques capable of detecting several targets simultaneously.

  7. Optical Imaging for Stem Cell Differentiation to Neuronal Lineage

    International Nuclear Information System (INIS)

    In regenerative medicine, the prospect of stem cell therapy hold great promise for the recovery of injured tissues and effective treatment of intractable diseases. Tracking stem cell fate provides critical information to understand and evaluate the success of stem cell therapy. The recent emergence of in vivo noninvasive molecular imaging has enabled assessment of the behavior of grafted stem cells in living subjects. In this review, we provide an overview of current optical imaging strategies based on cell or tissue specific reporter gene expression and of in vivo methods to monitor stem cell differentiation into neuronal lineages. These methods use optical reporters either regulated by neuron-specific promoters or containing neuron-specific microRNA binding sites. Both systems revealed dramatic changes in optical reporter imaging signals in cells differentiating a yeast GAL4 amplification system or an engineering-enhanced luciferase reported gene. Furthermore, we propose an advanced imaging system to monitor neuronal differentiation during neurogenesis that uses in vivo multiplexed imaging techniques capable of detecting several targets simultaneously

  8. Evolutionary origins of germline segregation in Metazoa: evidence for a germ stem cell lineage in the coral Orbicella faveolata (Cnidaria, Anthozoa).

    Science.gov (United States)

    Barfield, Sarah; Aglyamova, Galina V; Matz, Mikhail V

    2016-01-13

    The ability to segregate a committed germ stem cell (GSC) lineage distinct from somatic cell lineages is a characteristic of bilaterian Metazoans. However, the occurrence of GSC lineage specification in basally branching Metazoan phyla, such as Cnidaria, is uncertain. Without an independently segregated GSC lineage, germ cells and their precursors must be specified throughout adulthood from continuously dividing somatic stem cells, generating the risk of propagating somatic mutations within the individual and its gametes. To address the potential for existence of a GSC lineage in Anthozoa, the sister-group to all remaining Cnidaria, we identified moderate- to high-frequency somatic mutations and their potential for gametic transfer in the long-lived coral Orbicella faveolata (Anthozoa, Cnidaria) using a 2b-RAD sequencing approach. Our results demonstrate that somatic mutations can drift to high frequencies (up to 50%) and can also generate substantial intracolonial genetic diversity. However, these somatic mutations are not transferable to gametes, signifying the potential for an independently segregated GSC lineage in O. faveolata. In conjunction with previous research on germ cell development in other basally branching Metazoan species, our results suggest that the GSC system may be a Eumetazoan characteristic that evolved in association with the emergence of greater complexity in animal body plan organization and greater specificity of stem cell functions. PMID:26763699

  9. Reciprocal t(9;22 ABL/BCR fusion proteins: leukemogenic potential and effects on B cell commitment.

    Directory of Open Access Journals (Sweden)

    Xiaomin Zheng

    Full Text Available BACKGROUND: t(9;22 is a balanced translocation, and the chromosome 22 breakpoints (Philadelphia chromosome--Ph+ determine formation of different fusion genes that are associated with either Ph+ acute lymphatic leukemia (Ph+ ALL or chronic myeloid leukemia (CML. The "minor" breakpoint in Ph+ ALL encodes p185(BCR/ABL from der22 and p96(ABL/BCR from der9. The "major" breakpoint in CML encodes p210(BCR/ABL and p40(ABL/BCR. Herein, we investigated the leukemogenic potential of the der9-associated p96(ABL/BCR and p40(ABL/BCR fusion proteins and their roles in the lineage commitment of hematopoietic stem cells in comparison to BCR/ABL. METHODOLOGY: All t(9;22 derived proteins were retrovirally expressed in murine hematopoietic stem cells (SL cells and human umbilical cord blood cells (UCBC. Stem cell potential was determined by replating efficiency, colony forming--spleen and competitive repopulating assays. The leukemic potential of the ABL/BCR fusion proteins was assessed by in a transduction/transplantation model. Effects on the lineage commitment and differentiation were investigated by culturing the cells under conditions driving either myeloid or lymphoid commitment. Expression of key factors of the B-cell differentiation and components of the preB-cell receptor were determined by qRT-PCR. PRINCIPAL FINDINGS: Both p96(ABL/BCR and p40(ABL/BCR increased proliferation of early progenitors and the short term stem cell capacity of SL-cells and exhibited own leukemogenic potential. Interestingly, BCR/ABL gave origin exclusively to a myeloid phenotype independently from the culture conditions whereas p96(ABL/BCR and to a minor extent p40(ABL/BCR forced the B-cell commitment of SL-cells and UCBC. CONCLUSIONS/SIGNIFICANCE: Our here presented data establish the reciprocal ABL/BCR fusion proteins as second oncogenes encoded by the t(9;22 in addition to BCR/ABL and suggest that ABL/BCR contribute to the determination of the leukemic phenotype through their

  10. Colon stem cell and crypt dynamics exposed by cell lineage reconstruction.

    Directory of Open Access Journals (Sweden)

    Yitzhak Reizel

    2011-07-01

    Full Text Available Stem cell dynamics in vivo are often being studied by lineage tracing methods. Our laboratory has previously developed a retrospective method for reconstructing cell lineage trees from somatic mutations accumulated in microsatellites. This method was applied here to explore different aspects of stem cell dynamics in the mouse colon without the use of stem cell markers. We first demonstrated the reliability of our method for the study of stem cells by confirming previously established facts, and then we addressed open questions. Our findings confirmed that colon crypts are monoclonal and that, throughout adulthood, the process of monoclonal conversion plays a major role in the maintenance of crypts. The absence of immortal strand mechanism in crypts stem cells was validated by the age-dependent accumulation of microsatellite mutations. In addition, we confirmed the positive correlation between physical and lineage proximity of crypts, by showing that the colon is separated into small domains that share a common ancestor. We gained new data demonstrating that colon epithelium is clustered separately from hematopoietic and other cell types, indicating that the colon is constituted of few progenitors and ruling out significant renewal of colonic epithelium from hematopoietic cells during adulthood. Overall, our study demonstrates the reliability of cell lineage reconstruction for the study of stem cell dynamics, and it further addresses open questions in colon stem cells. In addition, this method can be applied to study stem cell dynamics in other systems.

  11. Tapetal cell fate, lineage and proliferation in the Arabidopsis anther.

    Science.gov (United States)

    Feng, Xiaoqi; Dickinson, Hugh G

    2010-07-01

    The four microsporangia of the flowering plant anther develop from archesporial cells in the L2 of the primordium. Within each microsporangium, developing microsporocytes are surrounded by concentric monolayers of tapetal, middle layer and endothecial cells. How this intricate array of tissues, each containing relatively few cells, is established in an organ possessing no formal meristems is poorly understood. We describe here the pivotal role of the LRR receptor kinase EXCESS MICROSPOROCYTES 1 (EMS1) in forming the monolayer of tapetal nurse cells in Arabidopsis. Unusually for plants, tapetal cells are specified very early in development, and are subsequently stimulated to proliferate by a receptor-like kinase (RLK) complex that includes EMS1. Mutations in members of this EMS1 signalling complex and its putative ligand result in male-sterile plants in which tapetal initials fail to proliferate. Surprisingly, these cells continue to develop, isolated at the locular periphery. Mutant and wild-type microsporangia expand at similar rates and the 'tapetal' space at the periphery of mutant locules becomes occupied by microsporocytes. However, induction of late expression of EMS1 in the few tapetal initials in ems1 plants results in their proliferation to generate a functional tapetum, and this proliferation suppresses microsporocyte number. Our experiments also show that integrity of the tapetal monolayer is crucial for the maintenance of the polarity of divisions within it. This unexpected autonomy of the tapetal 'lineage' is discussed in the context of tissue development in complex plant organs, where constancy in size, shape and cell number is crucial. PMID:20570940

  12. Lineage-related cytotoxicity and clonogenic profile of 1,4-benzoquinone-exposed hematopoietic stem and progenitor cells

    Energy Technology Data Exchange (ETDEWEB)

    Chow, Paik Wah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Abdul Hamid, Zariyantey, E-mail: zyantey@ukm.edu.my [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Chan, Kok Meng [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia); Inayat-Hussain, Salmaan Hussain [Environmental Health and Industrial Safety Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Rajab, Nor Fadilah [Biomedical Science Programme, School of Diagnostic & Applied Health Sciences, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Abdul Muda Aziz, 50300 Kuala Lumpur, Wilayah Persekutuan (Malaysia); Toxicology Laboratory, Faculty of Health Sciences, Universiti Kebangsaan Malaysia, Jalan Raja Muda Abdul Aziz, 50300 Kuala Lumpur (Malaysia)

    2015-04-01

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) are sensitive targets for benzene-induced hematotoxicity and leukemogenesis. The impact of benzene exposure on the complex microenvironment of HSCs and HPCs remains elusive. This study aims to investigate the mechanism linking benzene exposure to targeting HSCs and HPCs using phenotypic and clonogenic analyses. Mouse bone marrow (BM) cells were exposed ex vivo to the benzene metabolite, 1,4-benzoquinone (1,4-BQ), for 24 h. Expression of cellular surface antigens for HSC (Sca-1), myeloid (Gr-1, CD11b), and lymphoid (CD45, CD3e) populations were confirmed by flow cytometry. The clonogenicity of cells was studied using the colony-forming unit (CFU) assay for multilineage (CFU-GM and CFU-GEMM) and single-lineage (CFU-E, BFU-E, CFU-G, and CFU-M) progenitors. 1,4-BQ demonstrated concentration-dependent cytotoxicity in mouse BM cells. The percentage of apoptotic cells increased (p < 0.05) following 1,4-BQ exposure. Exposure to 1,4-BQ showed no significant effect on CD3e{sup +} cells but reduced the total counts of Sca-1{sup +}, CD11b{sup +}, Gr-1{sup +}, and CD45{sup +} cells at 7 and 12 μM (p < 0.05). Furthermore, the CFU assay showed reduced (p < 0.05) clonogenicity in 1,4-BQ-treated cells. 1,4-BQ induced CFU-dependent cytotoxicity by significantly inhibiting colony growth for CFU-E, BFU-E, CFU-G, and CFU-M starting at a low concentration of exposure (5 μM); whereas for the CFU-GM and CFU-GEMM, the inhibition of colony growth was remarkable only at 7 and 12 μM of 1,4-BQ, respectively. Taken together, 1,4-BQ caused lineage-related cytotoxicity in mouse HPCs, demonstrating greater toxicity in single-lineage progenitors than in those of multi-lineage. - Highlights: • We examine 1,4-BQ toxicity targeting mouse hematopoietic cell lineages. • 1,4-BQ induces concentration-dependent cytotoxicity in bone marrow (BM) cells. • 1,4-BQ shows lineage-related toxicity on hematopoietic stem and

  13. Lineage-related cytotoxicity and clonogenic profile of 1,4-benzoquinone-exposed hematopoietic stem and progenitor cells

    International Nuclear Information System (INIS)

    Hematopoietic stem cells (HSCs) and hematopoietic progenitor cells (HPCs) are sensitive targets for benzene-induced hematotoxicity and leukemogenesis. The impact of benzene exposure on the complex microenvironment of HSCs and HPCs remains elusive. This study aims to investigate the mechanism linking benzene exposure to targeting HSCs and HPCs using phenotypic and clonogenic analyses. Mouse bone marrow (BM) cells were exposed ex vivo to the benzene metabolite, 1,4-benzoquinone (1,4-BQ), for 24 h. Expression of cellular surface antigens for HSC (Sca-1), myeloid (Gr-1, CD11b), and lymphoid (CD45, CD3e) populations were confirmed by flow cytometry. The clonogenicity of cells was studied using the colony-forming unit (CFU) assay for multilineage (CFU-GM and CFU-GEMM) and single-lineage (CFU-E, BFU-E, CFU-G, and CFU-M) progenitors. 1,4-BQ demonstrated concentration-dependent cytotoxicity in mouse BM cells. The percentage of apoptotic cells increased (p < 0.05) following 1,4-BQ exposure. Exposure to 1,4-BQ showed no significant effect on CD3e+ cells but reduced the total counts of Sca-1+, CD11b+, Gr-1+, and CD45+ cells at 7 and 12 μM (p < 0.05). Furthermore, the CFU assay showed reduced (p < 0.05) clonogenicity in 1,4-BQ-treated cells. 1,4-BQ induced CFU-dependent cytotoxicity by significantly inhibiting colony growth for CFU-E, BFU-E, CFU-G, and CFU-M starting at a low concentration of exposure (5 μM); whereas for the CFU-GM and CFU-GEMM, the inhibition of colony growth was remarkable only at 7 and 12 μM of 1,4-BQ, respectively. Taken together, 1,4-BQ caused lineage-related cytotoxicity in mouse HPCs, demonstrating greater toxicity in single-lineage progenitors than in those of multi-lineage. - Highlights: • We examine 1,4-BQ toxicity targeting mouse hematopoietic cell lineages. • 1,4-BQ induces concentration-dependent cytotoxicity in bone marrow (BM) cells. • 1,4-BQ shows lineage-related toxicity on hematopoietic stem and progenitors. • 1,4-BQ toxicity is

  14. Lineage Extrinsic and Intrinsic Control of Immunoregulatory Cell Numbers by SHIP

    OpenAIRE

    Collazo, Michelle M.; Paraiso, Kim HT; Park, Mi-Young; Hazen, Amy L.; Kerr, William G.

    2012-01-01

    We previously showed that germline or induced SHIP-deficiency expands immunoregulatory cell numbers in T lymphoid and myeloid lineages. We postulated these increases could be interrelated. Here we show that myeloid specific ablation of SHIP leads to expansion of both myeloid-derived suppressor cell (MDSC) and regulatory T cell (Treg) numbers indicating SHIP-dependent control of Treg numbers by a myeloid cell type. Conversely, T lineage specific ablation of SHIP leads to expansion of Treg numb...

  15. Myeloid and lymphoid contribution to non-haematopoietic lineages through irradiation-induced heterotypic cell fusion

    DEFF Research Database (Denmark)

    Nygren, J.M.; Liuba, K.; Breitbach, M.; Stott, S.; Thorén, Lina Anna Maria; Roell, W.; Geisen, C.; Sasse, P.; Kirik, D.; Bjorklund, A.; Nerlov, C.; Fleischmann, B.K.; Jovinge, S.; Jacobsen, S.E.

    2008-01-01

    Recent studies have suggested that regeneration of non-haematopoietic cell lineages can occur through heterotypic cell fusion with haematopoietic cells of the myeloid lineage. Here we show that lymphocytes also form heterotypic-fusion hybrids with cardiomyocytes, skeletal muscle, hepatocytes and...... is induced by organ-specific injuries or whole-body irradiation, which has been used in previous studies to condition recipients of bone marrow transplants. Our findings demonstrate that blood cells of the lymphoid and myeloid lineages contribute to various non-haematopoietic tissues by forming rare...

  16. Calcium receptor expression and function in oligodendrocyte commitment and lineage progression: potential impact on reduced myelin basic protein in CaR-null mice

    DEFF Research Database (Denmark)

    Chattopadhyay, N.; Espinosa-Jeffrey, A.; Yano, S.; Bandyopadhyay, S.; Brown, E.M.; Vellis, J. de; Tfelt, Jacob

    2008-01-01

    oligodendrocyte > neuron > astrocyte. We next determined the rank order of CaR expression on inducing specification of neural stem cells to the neuronal, oligodendroglial, or astrocytic lineages and found that the relative levels of CaR mRNA expression are OPC > neuron > astrocytes. CaR mRNA expression in cells......R promotes cellular proliferation. We further observed that high Ca(2+) stimulates the mRNA levels of myelin basic protein in preoligodendrocytes, which is also CaR mediated. Finally, myelin basic protein levels were significantly reduced in the cerebellum of CaR-null mice during development. Our results...

  17. Differentiation of Equine Mesenchymal Stromal Cells into Cells of Neural Lineage: Potential for Clinical Applications

    Directory of Open Access Journals (Sweden)

    Claudia Cruz Villagrán

    2014-01-01

    Full Text Available Mesenchymal stromal cells (MSCs are able to differentiate into extramesodermal lineages, including neurons. Positive outcomes were obtained after transplantation of neurally induced MSCs in laboratory animals after nerve injury, but this is unknown in horses. Our objectives were to test the ability of equine MSCs to differentiate into cells of neural lineage in vitro, to assess differences in morphology and lineage-specific protein expression, and to investigate if horse age and cell passage number affected the ability to achieve differentiation. Bone marrow-derived MSCs were obtained from young and adult horses. Following demonstration of stemness, MSCs were neurally induced and microscopically assessed at different time points. Results showed that commercially available nitrogen-coated tissue culture plates supported proliferation and differentiation. Morphological changes were immediate and all the cells displayed a neural crest-like cell phenotype. Expression of neural progenitor proteins, was assessed via western blot or immunofluorescence. In our study, MSCs generated from young and middle-aged horses did not show differences in their ability to undergo differentiation. The effect of cell passage number, however, is inconsistent and further experiments are needed. Ongoing work is aimed at transdifferentiating these cells into Schwann cells for transplantation into a peripheral nerve injury model in horses.

  18. Calcium regulates the commitment of murine erythroleukemia cells to terminal erythroid differentiation

    OpenAIRE

    1981-01-01

    An alteration in the rate of calcium transport appears to be the rate- limiting event for the commitment of murine erythroleukemia (MEL) cells to initiate a program of terminal erythroid differentiation. The dimethyl sulfoxide (DMSO)-induced commitment of MEL cells to erythroid differentiation can be inhibited by treatment of cells with the calcium- chelating agent EGTA. Upon removal of EGTA, cells initiate commitment without the 12-h lag normally observed after treatment with DMSO alone. Tre...

  19. Long-term live cell imaging and automated 4D analysis of drosophila neuroblast lineages.

    Directory of Open Access Journals (Sweden)

    Catarina C F Homem

    Full Text Available The developing Drosophila brain is a well-studied model system for neurogenesis and stem cell biology. In the Drosophila central brain, around 200 neural stem cells called neuroblasts undergo repeated rounds of asymmetric cell division. These divisions typically generate a larger self-renewing neuroblast and a smaller ganglion mother cell that undergoes one terminal division to create two differentiating neurons. Although single mitotic divisions of neuroblasts can easily be imaged in real time, the lack of long term imaging procedures has limited the use of neuroblast live imaging for lineage analysis. Here we describe a method that allows live imaging of cultured Drosophila neuroblasts over multiple cell cycles for up to 24 hours. We describe a 4D image analysis protocol that can be used to extract cell cycle times and growth rates from the resulting movies in an automated manner. We use it to perform lineage analysis in type II neuroblasts where clonal analysis has indicated the presence of a transit-amplifying population that potentiates the number of neurons. Indeed, our experiments verify type II lineages and provide quantitative parameters for all cell types in those lineages. As defects in type II neuroblast lineages can result in brain tumor formation, our lineage analysis method will allow more detailed and quantitative analysis of tumorigenesis and asymmetric cell division in the Drosophila brain.

  20. Dedifferentiation of committed epithelial cells into stem cells in vivo

    OpenAIRE

    Tata, Purushothama Rao; Mou, Hongmei; Pardo-Saganta, Ana; Zhao, Rui; Prabhu, Mythili; Law, Brandon M.; Vinarsky, Vladimir; Josalyn L Cho; Breton, Sylvie; Sahay, Amar; Medoff, Benjamin D.; Rajagopal, Jayaraj

    2013-01-01

    Summary Cellular plasticity contributes to the regenerative capacity of plants, invertebrates, teleost fishes, and amphibians. In vertebrates, differentiated cells are known to revert into replicating progenitors, but these cells do not persist as stable stem cells. We now present evidence that differentiated airway epithelial cells can revert into stable and functional stem cells in vivo. Following the ablation of airway stem cells, we observed a surprising increase in the proliferation of c...

  1. Dedifferentiation of committed epithelial cells into stem cells in vivo

    OpenAIRE

    Tata, Purushothama Rao; Mou, Hongmei; Pardo-Saganta, Ana; Zhao, Rui; Prabhu, Mythili; Law, Brandon M.; Vinarsky, Vladimir; Josalyn L Cho; Breton, Sylvie; Sahay, Amar; Medoff, Benjamin D.; Rajagopal, Jayaraj

    2014-01-01

    Summary Cellular plasticity contributes to the regenerative capacity of plants, invertebrates, teleost fishes, and amphibians. In vertebrates, differentiated cells are known to revert into replicating progenitors, but these cells do not persist as stable stem cells. We now present evidence that differentiated airway epithelial cells can revert into stable and functional stem cells in vivo. Following the ablation of airway stem cells, we observed a surprising increase in the proliferation of c...

  2. To be or not to be a Treg cell: lineage decisions controlled by epigenetic mechanisms.

    OpenAIRE

    Toker, Aras; Huehn, Jochen

    2011-01-01

    Regulatory T (T(reg)) cells are a unique CD4(+) T cell lineage that plays a crucial role in the maintenance of immunological tolerance. The Forkhead box transcription factor Foxp3 is critically involved in T(reg) cell development and responsible for determining the suppressive function of these cells. The majority of Foxp3(+) T(reg) cells are generated during T cell development within the thymus and show features of a stable T cell lineage. New work indicates that both induction and stabiliza...

  3. Interleukin-15 Promotes the Commitment of Cord Blood CD34+ Stem Cells into NK Cells

    Institute of Scientific and Technical Information of China (English)

    张建; 夏青; 孙汭; 田志刚

    2004-01-01

    To explore the effect of rhlL-15 on CB-CD34+ stem cells committing to NK cells, CD34+ stem cells were obtained from cord blood (CB) by magnetic-assisted cell sorting (MACS) method. CD3, CD16 and CD56 molecules expressed on cell surface were detected by flow cytometer. MTF method was used to test the cytotoxicity of NK cells. The results were that stem cell factor (SCF) alone has no effect on CD34+ stem cells. IL-15 stimulated CD34+ stem cells commit to NK cells, and SCF showed strong synergistic effect with IL-15. It was concluded that IL-15 and SCF played different roles during NK cell development, llr15 promoted CD34+ stem cells differentiate to NK cell precursor and SCF improved the effectsof IL-15 on NK cell differentiation.

  4. The similarity between human embryonic stem cell-derived epithelial cells and ameloblast-lineage cells

    Institute of Scientific and Technical Information of China (English)

    Li-Wei Zheng; Logan Linthicum; Pamela K DenBesten; Yan Zhang

    2013-01-01

    This study aimed to compare epithelial cells derived from human embryonic stem cells (hESCs) to human ameloblast-lineage cells (ALCs), as a way to determine their potential use as a cell source for ameloblast regeneration. Induced by various concentrations of bone morphogenetic protein 4 (BMP4), retinoic acid (RA) and lithium chloride (LiCI) for 7 days, hESCs adopted cobble-stone epithelial phenotype (hESC-derived epithelial cells (ES-ECs)) and expressed cytokeratin 14. Compared with ALCs and oral epithelial cells (OE), ES-ECs expressed amelogenesis-associated genes similar to ALCs. ES-ECs were compared with human fetal skin epithelium, human fetal oral buccal mucosal epithelial cells and human ALCs for their expression pattern of cytokeratins as well. ALCs had relatively high expression levels of cytokeratin 76, which ,vas also found to be upregulated in ES-ECs. Based on the present study, with the similarity of gene expression with ALCs, ES-ECs are a promising potential cell source for regeneration, which are not available in erupted human teeth for regeneration of enamel.

  5. Derivation of chondrogenically-committed cells from human embryonic cells for cartilage tissue regeneration.

    Directory of Open Access Journals (Sweden)

    Nathaniel S Hwang

    Full Text Available BACKGROUND: Heterogeneous and uncontrolled differentiation of human embryonic stem cells (hESCs in embryoid bodies (EBs limits the potential use of hESCs for cell-based therapies. More efficient strategies are needed for the commitment and differentiation of hESCs to produce a homogeneous population of specific cell types for tissue regeneration applications. METHODOLOGY/PRINCIPAL FINDINGS: We report here that significant chondrocytic commitment of feeder-free cultured human embryonic stem cells (FF-hESCs, as determined by gene expression and immunostaining analysis, was induced by co-culture with primary chondrocytes. Furthermore, a dynamic expression profile of chondrocyte-specific genes was observed during monolayer expansion of the chondrogenically-committed cells. Chondrogenically-committed cells synergistically responded to transforming growth factor-beta1 (TGF-beta1 and beta1-integrin activating antibody by increasing tissue mass in pellet culture. In addition, when encapsulated in hydrogels, these cells formed cartilage tissue both in vitro and in vivo. In contrast, the absence of chondrocyte co-culture did not result in an expandable cell population from FF-hESCs. CONCLUSIONS/SIGNIFICANCE: The direct chondrocytic commitment of FF-hESCs can be induced by morphogenetic factors from chondrocytes without EB formation and homogenous cartilage tissue can be formed in vitro and in vivo.

  6. RhoA determines lineage fate of mesenchymal stem cells by modulating CTGF–VEGF complex in extracellular matrix

    Science.gov (United States)

    Li, Changjun; Zhen, Gehua; Chai, Yu; Xie, Liang; Crane, Janet L.; Farber, Emily; Farber, Charles R.; Luo, Xianghang; Gao, Peisong; Cao, Xu; Wan, Mei

    2016-01-01

    Mesenchymal stem cells (MSCs) participate in the repair/remodelling of many tissues, where MSCs commit to different lineages dependent on the cues in the local microenvironment. Here we show that TGFβ-activated RhoA/ROCK signalling functions as a molecular switch regarding the fate of MSCs in arterial repair/remodelling after injury. MSCs differentiate into myofibroblasts when RhoA/ROCK is turned on, endothelial cells when turned off. The former is pathophysiologic resulting in intimal hyperplasia, whereas the latter is physiological leading to endothelial repair. Further analysis revealed that MSC RhoA activation promotes formation of an extracellular matrix (ECM) complex consisting of connective tissue growth factor (CTGF) and vascular endothelial growth factor (VEGF). Inactivation of RhoA/ROCK in MSCs induces matrix metalloproteinase-3-mediated CTGF cleavage, resulting in VEGF release and MSC endothelial differentiation. Our findings uncover a novel mechanism by which cell–ECM interactions determine stem cell lineage specificity and offer additional molecular targets to manipulate MSC-involved tissue repair/regeneration. PMID:27126736

  7. D-MEF2: a MADS box transcription factor expressed in differentiating mesoderm and muscle cell lineages during Drosophila embryogenesis.

    Science.gov (United States)

    Lilly, B; Galewsky, S; Firulli, A B; Schulz, R A; Olson, E N

    1994-06-01

    The myocyte enhancer factor (MEF) 2 family of transcription factors has been implicated in the regulation of muscle transcription in vertebrates. We have cloned a protein from Drosophila, termed D-MEF2, that shares extensive amino acid homology with the MADS (MCM1, Agamous, Deficiens, and serum-response factor) domains of the vertebrate MEF2 proteins. D-mef2 gene expression is first detected during Drosophila embryogenesis within mesodermal precursor cells prior to specification of the somatic and visceral muscle lineages. Expression of D-mef2 is dependent on the mesodermal determinants twist and snail but independent of the homeobox-containing gene tinman, which is required for visceral muscle and heart development. D-mef2 expression precedes that of the MyoD homologue, nautilus, and, in contrast to nautilus, D-mef2 appears to be expressed in all somatic and visceral muscle cell precursors. Its temporal and spatial expression patterns suggest that D-mef2 may play an important role in commitment of mesoderm to myogenic lineages. PMID:8202544

  8. Stepwise renal lineage differentiation of mouse embryonic stem cells tracing in vivo development

    International Nuclear Information System (INIS)

    Highlights: ► We induced renal lineages from mESCs by following the in vivo developmental cues. ► We induced nephrogenic intermediate mesoderm by stepwise addition of factors. ► We induced two types of renal progenitor cells by reciprocal conditioned media. ► We propose the potential role of CD24 for the enrichment of renal lineage cells. -- Abstract: The in vitro derivation of renal lineage progenitor cells is essential for renal cell therapy and regeneration. Despite extensive studies in the past, a protocol for renal lineage induction from embryonic stem cells remains unestablished. In this study, we aimed to induce renal lineages from mouse embryonic stem cells (mESC) by following in vivo developmental stages, i.e., the induction of mesoderm (Stage I), intermediate mesoderm (Stage II) and renal lineages (Stage III). For stage I induction, in accordance with known signaling pathways involved in mesoderm development in vivo, i.e., Nodal, bone morphogenic proteins (BMPs) and Wnt, we found that the sequential addition of three factors, i.e., Activin-A (A), a surrogate for Nodal signaling, during days 0–2, A plus BMP-4 (4) during days 2–4, and A4 plus lithium (L), a surrogate for Wnt signaling, during days 4–6, was most effective to induce the mesodermal marker, Brachyury. For stage II induction, the addition of retinoic acid (R) in the continuous presence of A4L during days 6–8 was most effective to induce nephrogenic intermediate mesodermal markers, such as Pax2 and Lim1. Under this condition, more than 30% of cells were stained positive for Pax2, and there was a concomitant decrease in the expression of non-mesodermal markers. For stage III induction, in resemblance to the reciprocal induction between ureteric bud (UB) and metanephric mesenchyme (MM) during kidney development, we found that the exposure to conditioned media derived from UB and MM cells was effective in inducing MM and UB markers, respectively. We also observed the emergence and

  9. A Role for RE-1-Silencing Transcription Factor in Embryonic Stem Cells Cardiac Lineage Specification.

    Science.gov (United States)

    Aksoy, Irene; Marcy, Guillaume; Chen, Jiaxuan; Divakar, Ushashree; Kumar, Vibhor; John-Sanchez, Daniel; Rahmani, Mehran; Buckley, Noel J; Stanton, Lawrence W

    2016-04-01

    During development, lineage specification is controlled by several signaling pathways involving various transcription factors (TFs). Here, we studied the RE-1-silencing transcription factor (REST) and identified an important role of this TF in cardiac differentiation. Using mouse embryonic stem cells (ESC) to model development, we found that REST knockout cells lost the ability to differentiate into the cardiac lineage. Detailed analysis of specific lineage markers expression showed selective downregulation of endoderm markers in REST-null cells, thus contributing to a loss of cardiogenic signals. REST regulates cardiac differentiation of ESCs by negatively regulating the Wnt/β-catenin signaling pathway and positively regulating the cardiogenic TF Gata4. We propose here a new role for REST in cell fate specification besides its well-known repressive role of neuronal differentiation. PMID:26864965

  10. Differentiation of murine embryonic stem and induced pluripotent stem cells to renal lineage in vitro

    International Nuclear Information System (INIS)

    Embryonic stem (ES) cells which have the unlimited proliferative capacity and extensive differentiation potency can be an attractive source for kidney regeneration therapies. Recent breakthroughs in the generation of induced pluripotent stem (iPS) cells have provided with another potential source for the artificially-generated kidney. The purpose of this study is to know how to differentiate mouse ES and iPS cells into renal lineage. We used iPS cells from mouse fibroblasts by transfection of four transcription factors, namely Oct4, Sox2, c-Myc and Klf4. Real-time PCR showed that renal lineage markers were expressed in both ES and iPS cells after the induction of differentiation. It also showed that a tubular specific marker, KSP progressively increased to day 18, although the differentiation of iPS cells was slower than ES cells. The results indicated that renal lineage cells can be differentiated from both murine ES and iPS cells. Several inducing factors were tested whether they influenced on cell differentiation. In ES cells, both of GDNF and BMP7 enhanced the differentiation to metanephric mesenchyme, and Activin enhanced the differentiation of ES cells to tubular cells. Activin also enhanced the differentiation of iPS cells to tubular cells, although the enhancement was lower than in ES cells. ES and iPS cells have a potential to differentiate to renal lineage cells, and they will be an attractive resource of kidney regeneration therapy. This differentiation is enhanced by Activin in both ES and iPS cells.

  11. Differentiation of murine embryonic stem and induced pluripotent stem cells to renal lineage in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Morizane, Ryuji [Department of Internal Medicine, Keio University School of Medicine, Tokyo (Japan); Monkawa, Toshiaki, E-mail: monkawa@sc.itc.keio.ac.jp [Department of Internal Medicine, Keio University School of Medicine, Tokyo (Japan); Itoh, Hiroshi [Department of Internal Medicine, Keio University School of Medicine, Tokyo (Japan)

    2009-12-25

    Embryonic stem (ES) cells which have the unlimited proliferative capacity and extensive differentiation potency can be an attractive source for kidney regeneration therapies. Recent breakthroughs in the generation of induced pluripotent stem (iPS) cells have provided with another potential source for the artificially-generated kidney. The purpose of this study is to know how to differentiate mouse ES and iPS cells into renal lineage. We used iPS cells from mouse fibroblasts by transfection of four transcription factors, namely Oct4, Sox2, c-Myc and Klf4. Real-time PCR showed that renal lineage markers were expressed in both ES and iPS cells after the induction of differentiation. It also showed that a tubular specific marker, KSP progressively increased to day 18, although the differentiation of iPS cells was slower than ES cells. The results indicated that renal lineage cells can be differentiated from both murine ES and iPS cells. Several inducing factors were tested whether they influenced on cell differentiation. In ES cells, both of GDNF and BMP7 enhanced the differentiation to metanephric mesenchyme, and Activin enhanced the differentiation of ES cells to tubular cells. Activin also enhanced the differentiation of iPS cells to tubular cells, although the enhancement was lower than in ES cells. ES and iPS cells have a potential to differentiate to renal lineage cells, and they will be an attractive resource of kidney regeneration therapy. This differentiation is enhanced by Activin in both ES and iPS cells.

  12. Hacking cell differentiation: transcriptional rerouting in reprogramming, lineage infidelity and metaplasia

    OpenAIRE

    Regalo, G.; Leutz, A

    2013-01-01

    Initiating neoplastic cell transformation events are of paramount importance for the comprehension of regeneration and vanguard oncogenic processes but are difficult to characterize and frequently clinically overlooked. In epithelia, pre-neoplastic transformation stages are often distinguished by the appearance of phenotypic features of another differentiated tissue, termed metaplasia. In haemato/lymphopoietic malignancies, cell lineage ambiguity is increasingly recorded. Both, metaplasia and...

  13. Clonal analysis of the cell lineages in the male flower of maize

    Energy Technology Data Exchange (ETDEWEB)

    Dawe, R.K.; Freeling, M. (Univ. of California, Berkeley (USA))

    1990-11-01

    The cell lineages in the male flower of maize were characterized using X-rays and transposable elements to produce clonal sectors differing in anthocyanin pigmentation. Less than 50% of the somatic tassel mutations (caused by reversion of unstable color mutations) that were visible on the anther wall were sexually transmitted by the male gametes, unless the sectors were larger than half the tassel circumference. This result is explained by showing that: (a) both the outer (LI) and inner (LII) lineages of the shoot apical meristem form a cell layer in the bilayered anther wall, and that anther pigmentation can be derived from either cell layer; and that (b) the male germ cells are derived almost exclusively from the LII. Therefore, while reversion events in either the LI or LII are visible on the anther, only the LII events are heritable. Reversion events that occur prior to the organization of the shoot apical meristem however, produce large (usually more than one-half tassel) sectors that include both the outer and inner lineages. In contrast to the high level of cell layer invasion previously reported during leaf development, during anther development less than 10(-3) cells in the LI invade the LII to form male gametes. The strong correlation between cell lineage and cell fate in the maize anther has implications for studies on plant evolution and the genetic improvement of cereals by DNA transformation.

  14. Clonal analysis of the cell lineages in the male flower of maize

    International Nuclear Information System (INIS)

    The cell lineages in the male flower of maize were characterized using X-rays and transposable elements to produce clonal sectors differing in anthocyanin pigmentation. Less than 50% of the somatic tassel mutations (caused by reversion of unstable color mutations) that were visible on the anther wall were sexually transmitted by the male gametes, unless the sectors were larger than half the tassel circumference. This result is explained by showing that: (a) both the outer (LI) and inner (LII) lineages of the shoot apical meristem form a cell layer in the bilayered anther wall, and that anther pigmentation can be derived from either cell layer; and that (b) the male germ cells are derived almost exclusively from the LII. Therefore, while reversion events in either the LI or LII are visible on the anther, only the LII events are heritable. Reversion events that occur prior to the organization of the shoot apical meristem however, produce large (usually more than one-half tassel) sectors that include both the outer and inner lineages. In contrast to the high level of cell layer invasion previously reported during leaf development, during anther development less than 10(-3) cells in the LI invade the LII to form male gametes. The strong correlation between cell lineage and cell fate in the maize anther has implications for studies on plant evolution and the genetic improvement of cereals by DNA transformation

  15. Single-cell protein secretomic signatures as potential correlates to tumor cell lineage evolution and cell-cell interaction

    Directory of Open Access Journals (Sweden)

    Minsuk eKwak

    2013-02-01

    Full Text Available Secreted proteins including cytokines, chemokines and growth factors represent important functional regulators mediating a range of cellular behavior and cell-cell paracrine/autocrine signaling, e.g. in the immunological system, tumor microenvironment or stem cell niche. Detection of these proteins is of great value not only in basic cell biology but also for diagnosis and therapeutic monitoring of human diseases such as cancer. However, due to co-production of multiple effector proteins from a single cell, referred to as polyfunctionality, it is biologically informative to measure a panel of secreted proteins, or secretomic signature, at the level of single cells. Recent evidence further indicates that a genetically-identical cell population can give rise to diverse phenotypic differences. It is known that cytokines, for example, in the immune system define the effector functions and lineage differentiation of immune cells. In this Perspective Article, we hypothesize that protein secretion profile may represent a universal measure to identify the definitive correlate in the larger context of cellular functions to dissect cellular heterogeneity and evolutionary lineage relationship in human cancer.

  16. Lineage relationship of effector and memory T cells

    OpenAIRE

    Restifo, Nicholas P; Gattinoni, Luca

    2013-01-01

    Adaptive immunity is characterized by the ability to form long-lived immunological memory. Upon re-exposure to antigen, memory T cells respond more rapidly and robustly than naïve T cells, providing better clearance of pathogens. Recent reviews have reinforced the text-book view that memory T cells arise from effector cells. Although this notion is teleologically appealing, emerging data is more consistent with a model where naïve cells directly develop into memory cells without transitioning...

  17. Cancer Stem Cells in Brain Tumors and Their Lineage Hierarchy

    OpenAIRE

    Kong, Doo-Sik

    2012-01-01

    Despite recent advances in the development of novel targeted chemotherapies, the prognosis of malignant glioma remains dismal. The chemo-resistance of this tumor is attributed to tumor heterogeneity. To explain this unique chemo- resistance, the concept of cancer stem cells has been evoked. Cancer stem cells, a subpopulation of whole tumor cells, are now regarded as candidate therapeutic targets. Here, the author reviews and discusses the cancer stem cell concept.

  18. Flow-cytometric method for simultaneous analysis of mouse lung epithelial, endothelial, and hematopoietic lineage cells.

    Science.gov (United States)

    Singer, Benjamin D; Mock, Jason R; D'Alessio, Franco R; Aggarwal, Neil R; Mandke, Pooja; Johnston, Laura; Damarla, Mahendra

    2016-05-01

    Flow cytometry is a powerful tool capable of simultaneously analyzing multiple parameters on a cell-by-cell basis. Lung tissue preparation for flow cytometry requires creation of a single-cell suspension, which often employs enzymatic and mechanical dissociation techniques. These practices may damage cells and cause cell death that is unrelated to the experimental conditions under study. We tested methods of lung tissue dissociation and sought to minimize cell death in the epithelial, endothelial, and hematopoietic lineage cellular compartments. A protocol that involved flushing the pulmonary circulation and inflating the lung with Dispase, a bacillus-derived neutral metalloprotease, at the time of tissue harvest followed by mincing, digestion in a DNase and collagenase solution, and filtration before staining with fluorescent reagents concurrently maximized viable yields of epithelial, endothelial, and hematopoietic lineage cells compared with a standard method that did not use enzymes at the time of tissue harvest. Flow cytometry identified each population-epithelial (CD326(+)CD31(-)CD45(-)), endothelial (CD326(-)CD31(+)CD45(-)), and hematopoietic lineage (CD326(-)CD31(-)CD45(+))-and measured cellular viability by 7-aminoactinomycin D (7-AAD) staining. The Dispase method permitted discrimination of epithelial vs. endothelial cell death in a systemic lipopolysaccharide model of increased pulmonary vascular permeability. We conclude that application of a dissociative enzyme solution directly to the cellular compartments of interest at the time of tissue harvest maximized viable cellular yields of those compartments. Investigators could employ this dissociation method to simultaneously harvest epithelial, endothelial, and hematopoietic lineage and other lineage-negative cells for flow-cytometric analysis. PMID:26944088

  19. Chromatin dynamics in Pollen Mother Cells underpin a common scenario at the somatic-to-reproductive fate transition of both the male and female lineages in Arabidopsis

    Directory of Open Access Journals (Sweden)

    Wenjing eShe

    2015-04-01

    Full Text Available Unlike animals, where the germline is established early during embryogenesis, plants set aside their reproductive lineage late in development in dedicated floral organs. The specification of pollen mother cells (PMCs committed to meiosis takes place in the sporogenous tissue in anther locules and marks the somatic-to-reproductive cell fate transition towards the male reproductive lineage. Here we show that Arabidopsis PMCs differentiation is accompanied by large-scale changes in chromatin organization. This is characterized by significant increase in nuclear volume, chromatin decondensation, reduction in heterochromatin, eviction of linker histones and the H2AZ histone variant. These structural alterations are accompanied by dramatic, quantitative changes in histone modifications levels compared to that of surrounding somatic cells that do not share a sporogenic fate. All these changes are highly reminiscent of those we have formerly described in female megaspore mother cells (MMCs. This indicates that chromatin reprogramming is a common underlying scenario in the somatic-to-reproductive cell fate transition in both male and female lineages.

  20. Mechanical Modulation of Nascent Stem Cell Lineage Commitment in Tissue Engineering Scaffolds

    OpenAIRE

    Song, Min Jae; Dean, David; Tate, Melissa L Knothe

    2013-01-01

    Taking inspiration from tissue morphogenesis in utero, this study tests the concept of using tissue engineering scaffolds as delivery devices to modulate emergent structure-function relationships at early stages of tissue genesis. We report on the use of a combined computational fluid dynamics (CFD) modeling, advanced manufacturing methods, and experimental fluid mechanics (micro-piv and strain mapping) for the prospective design of tissue engineering scaffold geometries that deliver spatiall...

  1. Transcriptional repressor Tbx3 is required for the hormone-sensing cell lineage in mammary epithelium.

    Directory of Open Access Journals (Sweden)

    Kamini Kunasegaran

    Full Text Available The transcriptional repressor Tbx3 is involved in lineage specification in several tissues during embryonic development. Germ-line mutations in the Tbx3 gene give rise to Ulnar-Mammary Syndrome (comprising reduced breast development and Tbx3 is required for mammary epithelial cell identity in the embryo. Notably Tbx3 has been implicated in breast cancer, which develops in adult mammary epithelium, but the role of Tbx3 in distinct cell types of the adult mammary gland has not yet been characterized. Using a fluorescent reporter knock-in mouse, we show that in adult virgin mice Tbx3 is highly expressed in luminal cells that express hormone receptors, and not in luminal cells of the alveolar lineage (cells primed for milk production. Flow cytometry identified Tbx3 expression already in progenitor cells of the hormone-sensing lineage and co-immunofluorescence confirmed a strict correlation between estrogen receptor (ER and Tbx3 expression in situ. Using in vivo reconstitution assays we demonstrate that Tbx3 is functionally relevant for this lineage because knockdown of Tbx3 in primary mammary epithelial cells prevented the formation of ER+ cells, but not luminal ER- or basal cells. Interestingly, genes that are repressed by Tbx3 in other cell types, such as E-cadherin, are not repressed in hormone-sensing cells, highlighting that transcriptional targets of Tbx3 are cell type specific. In summary, we provide the first analysis of Tbx3 expression in the adult mammary gland at a single cell level and show that Tbx3 is important for the generation of hormone-sensing cells.

  2. Selective cell targeting and lineage tracing of human induced pluripotent stem cells using recombinant avian retroviruses.

    Science.gov (United States)

    Hildebrand, Laura; Seemann, Petra; Kurtz, Andreas; Hecht, Jochen; Contzen, Jörg; Gossen, Manfred; Stachelscheid, Harald

    2015-12-01

    Human induced pluripotent stem cells (hiPSC) differentiate into multiple cell types. Selective cell targeting is often needed for analyzing gene function by overexpressing proteins in a distinct population of hiPSC-derived cell types and for monitoring cell fate in response to stimuli. However, to date, this has not been possible, as commonly used viruses enter the hiPSC via ubiquitously expressed receptors. Here, we report for the first time the application of a heterologous avian receptor, the tumor virus receptor A (TVA), to selectively transduce TVA(+) cells in a mixed cell population. Expression of the TVA surface receptor via genetic engineering renders cells susceptible for infection by avian leucosis virus (ALV). We generated hiPSC lines with this stably integrated, ectopic TVA receptor gene that expressed the receptor while retaining pluripotency. The undifferentiated hiPSC(TVA+) as well as their differentiating progeny could be infected by recombinant ALV (so-called RCAS virus) with high efficiency. Due to incomplete receptor blocking, even sequential infection of differentiating or undifferentiated TVA(+) cells was possible. In conclusion, the TVA/RCAS system provides an efficient and gentle gene transfer system for hiPSC and extends our possibilities for selective cell targeting and lineage tracing studies. PMID:26109426

  3. Cell lineage relationship in the stomach of normal and genetically manipulated mice

    Directory of Open Access Journals (Sweden)

    Karam S.M.

    1998-01-01

    Full Text Available The oxyntic mucosa of the mouse stomach is lined with a heterogeneous population of cells that form numerous short pits continuous with long tubular glands. Tritiated thymidine radioautography has made it possible to pinpoint the origin of all cell types and to follow the differentiation/migration of different cell lineages along the pit-gland unit. The proliferating multipotent stem cells functionally anchored in the upper glandular region, the isthmus, give rise to three main lineage precursors: 1 pre-pit cells, which migrate upward to the pit while differentiating into mucus-producing pit cells; 2 pre-neck cells, which migrate downward to the glandular neck while differentiating into mucus-producing neck cells that, by approaching the glandular base, gradually change their phenotype into pepsinogen- and intrinsic factor-producing zymogenic cells; 3 pre-parietal cells, which differentiate into acid-producing parietal cells in the isthmus and then undergo bipolar migration towards the pit and the glandular base. Thus, parietal cells are the only cells that complete their differentiation in the isthmus and then migrate to be scattered throughout the pit-gland unit. To determine whether parietal cells play a role in controlling decisions about cell fate within the pit-gland unit, the gastric epithelium has been examined in transgenic mice expressing the H,K-ATPase ß-subunit-1035 to +24/simian virus 40 large T antigen fusion gene. The blockade in parietal cell differentiation in these mice produces an amplification of lineage precursors, a marked depletion of zymogenic cells and an increase in pit cell census. Ablation of parietal cells in another transgenic mouse model expressing the H,K-ATPase ß-subunit-1035 to +24/diphtheria toxin fragment A fusion gene also produces amplification of lineage precursors, and similar effects on zymogenic and pit cell census. These findings strongly suggest that parietal cells produce regulatory signals that

  4. BMP signalling regulates the pre-implantation development of extra-embryonic cell lineages in the mouse embryo

    OpenAIRE

    Graham, Sarah J. L.; Wicher, Krzysztof B.; Jedrusik, Agnieszka; Guo, Guoji; Herath, Wishva; Robson, Paul; Zernicka-Goetz, Magdalena

    2014-01-01

    Pre-implantation development requires the specification and organization of embryonic and extra-embryonic lineages. The separation of these lineages takes place when asymmetric divisions generate inside and outside cells that differ in polarity, position and fate. Here we assess the global transcriptional identities of these precursor cells to gain insight into the molecular mechanisms regulating lineage segregation,. Unexpectedly, this reveals that complementary components of the BMP signall...

  5. Lineage tracing reveals the dynamic contribution of Hes1+ cells to the developing and adult pancreas

    OpenAIRE

    Kopinke, Daniel; Brailsford, Marisa; Shea, Jill E; Leavitt, Rebecca; Scaife, Courtney L.; Murtaugh, L. Charles

    2011-01-01

    Notch signaling regulates numerous developmental processes, often acting either to promote one cell fate over another or else to inhibit differentiation altogether. In the embryonic pancreas, Notch and its target gene Hes1 are thought to inhibit endocrine and exocrine specification. Although differentiated cells appear to downregulate Hes1, it is unknown whether Hes1 expression marks multipotent progenitors, or else lineage-restricted precursors. Moreover, although rare cells of the adult pan...

  6. Committed Tc17 cells are phenotypically and functionally resistant to the effects of IL-27.

    Science.gov (United States)

    El-Behi, Mohamed; Dai, Hong; Magalhaes, Joao G; Hwang, Daniel; Zhang, Guang-Xian; Rostami, Abdolmohamad; Ciric, Bogoljub

    2014-10-01

    IL-17-secreting CD8(+) T cells (Tc17 cells) have been implicated in immunity to infections, cancer, and autoimmune diseases. Thus far, studies on Tc17 cells have primarily investigated their development from naïve precursors, while the biology of committed Tc17 cells has been less characterized, in particular during the effector phase of immune responses. IL-27 is an important regulator of inflammation through the induction of regulatory Tr1 cells, as well as a suppressor of Th17-cell development. IL-27 suppresses the development of Tc17 cells, but its effects on committed Tc17 cells are unknown. Here we demonstrate that even though IL-27 completely inhibited the development of C57BL/6 mouse Tc17 cells, it had little effect on previously committed Tc17 cells. Although committed Tc17 cells were capable of responding to IL-27, it had no effect on expression of RORγt and RORα, or production of various cytokines. Committed Tc17 cells did not express granzyme B and lacked cytotoxicity in vitro, features that remained unaltered by IL-27 treatment. Nonetheless, they efficiently induced diabetes, irrespective of treatment with IL-27 prior to transfer into RIP-mOVA mice. These findings suggest that use of IL-27 to modulate autoimmune diseases might have limited therapeutic efficacy if autoaggressive Tc17 cells have already developed. PMID:25070084

  7. Systemic mastocytosis with associated clonal haematological non-mast cell lineage diseases: a histopathological challenge

    OpenAIRE

    Horny, H-P; Sotlar, K; Sperr, W R; Valent, P

    2004-01-01

    Aims: Although systemic mastocytosis (SM) with an associated clonal haematological non-mast cell lineage disease (SM-AHNMD) is a major subtype of SM, little is known about its frequency among myelogenous neoplasms, and mastocytosis in particular, or about AHNMD subtype frequencies.

  8. Msx2 alters the timing of retinal ganglion cells fate commitment and differentiation

    Energy Technology Data Exchange (ETDEWEB)

    Jiang, Shao-Yun, E-mail: jiangshaoyun@yahoo.com [School of Dentistry, Tianjin Medical University, 12 Qi Xiang Tai Street, Tianjin 300070 (China); Wang, Jian-Tao, E-mail: wangjiantao65@hotmail.com [Eye Center, Tianjin Medical University, 64 Tongan Road, Tianjin 300070 (China); Dohney Eye Institute, Keck School of Medicine, University of Southern California, 1355 San Pablo Street, DOH 314, Los Angeles, CA 90033 (United States)

    2010-05-14

    Timing of cell fate commitment determines distinct retinal cell types, which is believed to be controlled by a tightly coordinated regulatory program of proliferation, cell cycle exit and differentiation. Although homeobox protein Msx2 could induce apoptosis of optic vesicle, it is unclear whether Msx2 regulates differentiation and cell fate commitment of retinal progenitor cells (RPCs) to retinal ganglion cells (RGCs). In this study, we show that overexpression of Msx2 transiently suppressed the expression of Cyclin D1 and blocked cell proliferation. Meanwhile, overexpression of Msx2 delayed the expression of RGC-specific differentiation markers (Math5 and Brn3b), which showed that Msx2 could affect the timing of RGCs fate commitment and differentiation by delaying the timing of cell cycle exit of retinal progenitors. These results indicate Msx2 possesses dual regulatory functions in controlling cell cycle progression of retinal RPCs and timing of RGCs differentiation.

  9. Msx2 alters the timing of retinal ganglion cells fate commitment and differentiation

    International Nuclear Information System (INIS)

    Timing of cell fate commitment determines distinct retinal cell types, which is believed to be controlled by a tightly coordinated regulatory program of proliferation, cell cycle exit and differentiation. Although homeobox protein Msx2 could induce apoptosis of optic vesicle, it is unclear whether Msx2 regulates differentiation and cell fate commitment of retinal progenitor cells (RPCs) to retinal ganglion cells (RGCs). In this study, we show that overexpression of Msx2 transiently suppressed the expression of Cyclin D1 and blocked cell proliferation. Meanwhile, overexpression of Msx2 delayed the expression of RGC-specific differentiation markers (Math5 and Brn3b), which showed that Msx2 could affect the timing of RGCs fate commitment and differentiation by delaying the timing of cell cycle exit of retinal progenitors. These results indicate Msx2 possesses dual regulatory functions in controlling cell cycle progression of retinal RPCs and timing of RGCs differentiation.

  10. Conversion of embryonic stem cells into extraembryonic lineages by CRISPR-mediated activators

    Science.gov (United States)

    Wei, Shu; Zou, Qingjian; Lai, Sisi; Zhang, Quanjun; Li, Li; Yan, Quanmei; Zhou, Xiaoqing; Zhong, Huilin; Lai, Liangxue

    2016-01-01

    The recently emerged CRISPR/Cas9 technique has opened a new perspective on readily editing specific genes. When combined with transcription activators, it can precisely manipulate endogenous gene expression. Here, we enhanced the expression of endogenous Cdx2 and Gata6 genes by CRISPR-mediated activators, thus mouse embryonic stem cells (ESCs) were directly converted into two extraembryonic lineages, i.e., typical trophoblast stem cells (TSCs) and extraembryonic endoderm cells (XENCs), which exhibited characters of TSC or XENC derived from the blastocyst extraembryonic lineages such as cell morphology, specific gene expression, and differentiation ability in vitro and in vivo. This study demonstrates that the cell fate can be effectively manipulated by directly activating of specific endogenous gene expression with CRISPR-mediated activator. PMID:26782778

  11. Near Equilibrium Calculus of Stem Cells in Application to the Airway Epithelium Lineage.

    Science.gov (United States)

    Sun, Zheng; Plikus, Maksim V; Komarova, Natalia L

    2016-07-01

    Homeostatic maintenance of tissues is orchestrated by well tuned networks of cellular signaling. Such networks regulate, in a stochastic manner, fates of all cells within the respective lineages. Processes such as symmetric and asymmetric divisions, differentiation, de-differentiation, and death have to be controlled in a dynamic fashion, such that the cell population is maintained at a stable equilibrium, has a sufficiently low level of stochastic variation, and is capable of responding efficiently to external damage. Cellular lineages in real tissues may consist of a number of different cell types, connected by hierarchical relationships, albeit not necessarily linear, and engaged in a number of different processes. Here we develop a general mathematical methodology for near equilibrium studies of arbitrarily complex hierarchical cell populations, under regulation by a control network. This methodology allows us to (1) determine stability properties of the network, (2) calculate the stochastic variance, and (3) predict how different control mechanisms affect stability and robustness of the system. We demonstrate the versatility of this tool by using the example of the airway epithelium lineage. Recent research shows that airway epithelium stem cells divide mostly asymmetrically, while the so-called secretory cells divide predominantly symmetrically. It further provides quantitative data on the recovery dynamics of the airway epithelium, which can include secretory cell de-differentiation. Using our new methodology, we demonstrate that while a number of regulatory networks can be compatible with the observed recovery behavior, the observed division patterns of cells are the most optimal from the viewpoint of homeostatic lineage stability and minimizing the variation of the cell population size. This not only explains the observed yet poorly understood features of airway tissue architecture, but also helps to deduce the information on the still largely hypothetical

  12. Propagation of human parvovirus B19 in primary culture of erythroid lineage cells derived from fetal liver.

    OpenAIRE

    Yaegashi, N; Shiraishi, H; Takeshita, T.; Nakamura, M.; Yajima, A; Sugamura, K

    1989-01-01

    Erythroid lineage cells derived from fetal liver were demonstrated to be target cells for human parvovirus B19 infection. B19 virus antigen-positive serum was inoculated into primary cultures containing erythroid lineage cells enriched from fetal liver. The B19 virus antigen was detected on about 5% of cells in the culture by immunofluorescence staining, and the stained cells were identified as erythroid lineage cells by double staining with anti-B19 virus-positive serum and anti-erythroid li...

  13. Bridging the gap between postembryonic cell lineages and identified embryonic neuroblasts in the ventral nerve cord of Drosophila melanogaster

    Directory of Open Access Journals (Sweden)

    Oliver Birkholz

    2015-03-01

    Full Text Available The clarification of complete cell lineages, which are produced by specific stem cells, is fundamental for understanding mechanisms, controlling the generation of cell diversity and patterning in an emerging tissue. In the developing Central Nervous System (CNS of Drosophila, neural stem cells (neuroblasts exhibit two periods of proliferation: During embryogenesis they produce primary lineages, which form the larval CNS. After a phase of mitotic quiescence, a subpopulation of them resumes proliferation in the larva to give rise to secondary lineages that build up the CNS of the adult fly. Within the ventral nerve cord (VNC detailed descriptions exist for both primary and secondary lineages. However, while primary lineages have been linked to identified neuroblasts, the assignment of secondary lineages has so far been hampered by technical limitations. Therefore, primary and secondary neural lineages co-existed as isolated model systems. Here we provide the missing link between the two systems for all lineages in the thoracic and abdominal neuromeres. Using the Flybow technique, embryonic neuroblasts were identified by their characteristic and unique lineages in the living embryo and their further development was traced into the late larval stage. This comprehensive analysis provides the first complete view of which embryonic neuroblasts are postembryonically reactivated along the anterior/posterior-axis of the VNC, and reveals the relationship between projection patterns of primary and secondary sublineages.

  14. Hacking cell differentiation: transcriptional rerouting in reprogramming, lineage infidelity and metaplasia.

    Science.gov (United States)

    Regalo, Gonçalo; Leutz, Achim

    2013-08-01

    Initiating neoplastic cell transformation events are of paramount importance for the comprehension of regeneration and vanguard oncogenic processes but are difficult to characterize and frequently clinically overlooked. In epithelia, pre-neoplastic transformation stages are often distinguished by the appearance of phenotypic features of another differentiated tissue, termed metaplasia. In haemato/lymphopoietic malignancies, cell lineage ambiguity is increasingly recorded. Both, metaplasia and biphenotypic leukaemia/lymphoma represent examples of dysregulated cell differentiation that reflect a history of trans-differentiation and/or epigenetic reprogramming. Here we compare the similarity between molecular events of experimental cell trans-differentiation as an emerging therapeutic concept, with lineage confusion, as in metaplasia and dysplasia forecasting tumour development. PMID:23828660

  15. Aberrant chromatin at genes encoding stem cell regulators in human mixed-lineage leukemia

    OpenAIRE

    Guenther, Matthew G.; Lawton, Lee N.; Rozovskaia, Tatiana; Frampton, Garrett M.; Levine, Stuart S.; Thomas L Volkert; Croce, Carlo M.; Nakamura, Tatsuya; Canaani, Eli; Young, Richard A.

    2008-01-01

    Mixed-lineage leukemia (MLL) fusion proteins are potent inducers of leukemia, but how these proteins generate aberrant gene expression programs is poorly understood. Here we show that the MLL-AF4 fusion protein occupies developmental regulatory genes important for hematopoietic stem cell identity and self-renewal in human leukemia cells. These MLL-AF4-bound regions have grossly altered chromatin structure, with histone modifications catalyzed by trithorax group proteins and DOT1 extending acr...

  16. Levels of BDNF Impact Oligodendrocyte Lineage Cells Following a Cuprizone Lesion

    OpenAIRE

    VonDran, Melissa W.; Singh, Harmandeep; Honeywell, Jean Z.; Dreyfus, Cheryl F

    2011-01-01

    Previous work in culture has shown that basal forebrain (BF) oligodendrocyte (OLG) lineage cells respond to BDNF by increasing DNA synthesis and differentiation. Further, in the BF in vivo, reduced levels of BDNF as seen in BDNF +/− mice result in reduced numbers of NG2+ cells and deficits in myelin proteins throughout development and in the adult, suggesting that BDNF impacts the proliferating population of OLGs as well as differentiation in vivo. In this study, to investigate roles BDNF may...

  17. Regulation of T-helper-cell lineage development by osteopontin: the inside story

    OpenAIRE

    Cantor, Harvey; Shinohara, Mari L.

    2009-01-01

    Studies of osteopontin (OPN)-dependent regulation of immune responses have focused on the cytokine activities of the secreted form of this protein. Recent evidence has revealed that an intracellular form of OPN expressed by dendritic cells regulates the expression of pro-inflammatory cytokines and the differentiation of T helper (TH)-cell lineages. In this Opinion article, we discuss the properties of both OPN isoforms and their respective contributions to the immune response. We propose that...

  18. Foxp3 exploits a preexistent enhancer landscape for regulatory T cell lineage specification

    OpenAIRE

    Samstein, Robert M.; Arvey, Aaron; Josefowicz, Steven Z.; Peng, Xiao; Reynolds, Alex; Sandstrom, Richard; Neph, Shane; Sabo, Peter; Kim, Jeong M.; Liao, Will; O. Li, Ming; Leslie, Christina; John A. Stamatoyannopoulos; Rudensky, Alexander Y.

    2012-01-01

    Regulatory T (Treg) cells, whose identity and function are defined by the transcription factor Foxp3, are indispensable for immune homeostasis. It is unclear whether Foxp3 exerts its Treg lineage specification function through active modification of the chromatin landscape and establishment of new enhancers or by exploiting a pre-existing enhancer landscape. Analysis of the chromatin accessibility of Foxp3-bound enhancers in Treg and Foxp3-negative T cells showed that Foxp3 was bound overwhel...

  19. Stepwise renal lineage differentiation of mouse embryonic stem cells tracing in vivo development

    Energy Technology Data Exchange (ETDEWEB)

    Nishikawa, Masaki, E-mail: masakiwestriver@gmail.com [Medical and Research Services, Greater Los Angeles Veterans Affairs Healthcare System at Sepulveda, North Hills, CA (United States); University of California at Los Angeles, David Geffen School of Medicine, Los Angeles, CA 91343 (United States); Yanagawa, Naomi [Medical and Research Services, Greater Los Angeles Veterans Affairs Healthcare System at Sepulveda, North Hills, CA (United States); University of California at Los Angeles, David Geffen School of Medicine, Los Angeles, CA 91343 (United States); Kojima, Nobuhiko [Institute of Industrial Science (IIS), University of Tokyo, 4-6-1 Komaba, Meguro-ku, Tokyo 153-8505 (Japan); Yuri, Shunsuke; Hauser, Peter V.; Jo, Oak D.; Yanagawa, Norimoto [Medical and Research Services, Greater Los Angeles Veterans Affairs Healthcare System at Sepulveda, North Hills, CA (United States); University of California at Los Angeles, David Geffen School of Medicine, Los Angeles, CA 91343 (United States)

    2012-01-13

    Highlights: Black-Right-Pointing-Pointer We induced renal lineages from mESCs by following the in vivo developmental cues. Black-Right-Pointing-Pointer We induced nephrogenic intermediate mesoderm by stepwise addition of factors. Black-Right-Pointing-Pointer We induced two types of renal progenitor cells by reciprocal conditioned media. Black-Right-Pointing-Pointer We propose the potential role of CD24 for the enrichment of renal lineage cells. -- Abstract: The in vitro derivation of renal lineage progenitor cells is essential for renal cell therapy and regeneration. Despite extensive studies in the past, a protocol for renal lineage induction from embryonic stem cells remains unestablished. In this study, we aimed to induce renal lineages from mouse embryonic stem cells (mESC) by following in vivo developmental stages, i.e., the induction of mesoderm (Stage I), intermediate mesoderm (Stage II) and renal lineages (Stage III). For stage I induction, in accordance with known signaling pathways involved in mesoderm development in vivo, i.e., Nodal, bone morphogenic proteins (BMPs) and Wnt, we found that the sequential addition of three factors, i.e., Activin-A (A), a surrogate for Nodal signaling, during days 0-2, A plus BMP-4 (4) during days 2-4, and A4 plus lithium (L), a surrogate for Wnt signaling, during days 4-6, was most effective to induce the mesodermal marker, Brachyury. For stage II induction, the addition of retinoic acid (R) in the continuous presence of A4L during days 6-8 was most effective to induce nephrogenic intermediate mesodermal markers, such as Pax2 and Lim1. Under this condition, more than 30% of cells were stained positive for Pax2, and there was a concomitant decrease in the expression of non-mesodermal markers. For stage III induction, in resemblance to the reciprocal induction between ureteric bud (UB) and metanephric mesenchyme (MM) during kidney development, we found that the exposure to conditioned media derived from UB and MM cells was

  20. Generation of polyhormonal and multipotent pancreatic progenitor lineages from human pluripotent stem cells.

    Science.gov (United States)

    Korytnikov, Roman; Nostro, Maria Cristina

    2016-05-15

    Generation of pancreatic β-cells from human pluripotent stem cells (hPSCs) has enormous importance in type 1 diabetes (T1D), as it is fundamental to a treatment strategy based on cellular therapeutics. Being able to generate β-cells, as well as other mature pancreatic cells, from human embryonic stem cells (hESCs) and human induced pluripotent stem cells (hiPSCs) will also enable the development of platforms that can be used for disease modeling and drug testing for a variety of pancreas-associated diseases, including cystic fibrosis. For this to occur, it is crucial to develop differentiation strategies that are robust and reproducible across cell lines and laboratories. In this article we describe two serum-free differentiation protocols designed to generate specific pancreatic lineages from hPSCs. Our approach employs a variety of cytokines and small molecules to mimic developmental pathways active during pancreatic organogenesis and allows for the in vitro generation of distinct pancreatic populations. The first protocol is designed to give rise to polyhormonal cells that have the potential to differentiate into glucagon-producing cells. The second protocol is geared to generate multipotent pancreatic progenitor cells, which harbor the potential to generate all pancreatic lineages including: monohormonal endocrine cells, acinar, and ductal cells. PMID:26515645

  1. Integrin αv in the mechanical response of osteoblast lineage cells

    Energy Technology Data Exchange (ETDEWEB)

    Kaneko, Keiko [Department of Bone and Joint Disease, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8511 (Japan); Ito, Masako [Medical Work-Life-Balance Center, Nagasaki University Hospital, Nagasaki 852-8501 (Japan); Naoe, Yoshinori [Department of Mechanism of Aging, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8511 (Japan); Lacy-Hulbert, Adam [Department of Pediatrics, Massachusetts General Hospital, Boston, MA 02114 (United States); Ikeda, Kyoji, E-mail: kikeda@ncgg.go.jp [Department of Bone and Joint Disease, National Center for Geriatrics and Gerontology, Obu, Aichi 474-8511 (Japan)

    2014-05-02

    Highlights: • Deletion of integrin αv in osteoblast lineage results in an impaired SOST response to loading in vivo. • c-Src–p130Cas–JNK–YAP/TAZ is activated via integrin αv on osteoblasts in response to FSS. • Deletion of integrin αv in osteoblasts results in impaired responses to mechanical stimulation. • Integrin αv is a key component of the mechanosensing machinery in bone. - Abstract: Although osteoblast lineage cells, especially osteocytes, are thought to be a primary mechanosensory cell in bone, the identity of the mechano-receptor and downstream mechano-signaling pathways remain largely unknown. Here we show using osteoblastic cell model of mechanical stimulation with fluid shear stress that in the absence of integrin αv, phosphorylation of the Src substrate p130Cas and JNK was impaired, culminating in an inhibition of nuclear translocation of YAP/TAZ and subsequent transcriptional activation of target genes. Targeted deletion of the integrin αv in osteoblast lineage cells results in an attenuated response to mechanical loading in terms of Sost gene expression, indicative of a role for integrin αv in mechanoreception in vivo. Thus, integrin αv may be integral to a mechanosensing machinery in osteoblastic cells and involved in activation of a Src–JNK–YAP/TAZ pathway in response to mechanical stimulation.

  2. Hematopoietic and mesenchymal stem cells: polymeric nanoparticle uptake and lineage differentiation

    Directory of Open Access Journals (Sweden)

    Ivonne Brüstle

    2015-02-01

    Full Text Available The combination of stem cell therapy and nanoparticles promises to enhance the effect of cellular therapies by using nanocarriers as drug delivery devices to guide the further differentiation or homing of stem cells. The impact of nanoparticles on primary cell types remains much more elusive as most groups study the nanoparticle–cell interaction in malignant cell lines. Here, we report on the influence of polymeric nanoparticles on human hematopoietic stem cells (hHSCs and mesenchymal stem cells (hMSCs. In this study we systematically investigated the influence of polymeric nanoparticles on the cell functionality and differentiation capacity of hHSCs and hMSCs to obtain a deeper knowledge of the interaction of stem cells and nanoparticles. As model systems of nanoparticles, two sets of either bioinert (polystyrene without carboxylic groups on the surface or biodegradable (PLLA without magnetite particles were analyzed. Flow cytometry and microscopy analysis showed high uptake rates and no toxicity for all four tested particles in hMSCs and hHSCs. During the differentiation process, the payload of particles per cell decreased. The PLLA–Fe particle showed a significant increase in the IL-8 release in hMSCs but not in hHSCs. We assume that this is due to an increase of free intracellular iron ions but obviously also depends on the cell type. For hHSCs and hMSCs, lineage differentiation into erythrocytes, granulocytes, and megakaryocytes or adipocytes, osteocytes and chondrocytes, was not influenced by the particles when analyzed with lineage specific cluster of differentiation markers. On the other hand qPCR analysis showed significant changes in the expression of some (but not all investigated lineage markers for both primary cell types.

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

    OpenAIRE

    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 ...

  4. Expression Patterns of Cancer-Testis Antigens in Human Embryonic Stem Cells and Their Cell Derivatives Indicate Lineage Tracks

    OpenAIRE

    Olga Gordeeva; Tatyana Yakovleva; Galina Poljanskaya; Tatyana Krylova; Anna Koltsova; Nadya Lifantseva

    2011-01-01

    Pluripotent stem cells can differentiate into various lineages but undergo genetic and epigenetic changes during long-term cultivation and, therefore, require regular monitoring. The expression patterns of cancer-testis antigens (CTAs) MAGE-A2, -A3, -A4, -A6, -A8, -B2, and GAGE were examined in undifferentiated human embryonic stem (hES) cells, their differentiated derivatives, teratocarcinoma (hEC) cells, and cancer cell lines of neuroectodermal and mesodermal origin. Undifferentiated hES ce...

  5. cKit Lineage Hemogenic Endothelium-Derived Cells Contribute to Mesenteric Lymphatic Vessels

    Directory of Open Access Journals (Sweden)

    Lukas Stanczuk

    2015-03-01

    Full Text Available Pathological lymphatic diseases mostly affect vessels in specific tissues, yet little is known about organ-specific regulation of the lymphatic vasculature. Here, we show that the vascular endothelial growth factor receptor 3 (VEGFR-3/p110α PI3-kinase signaling pathway is selectively required for the formation of mesenteric lymphatic vasculature. Using genetic lineage tracing, we demonstrate that part of the mesenteric lymphatic vasculature develops from cKit lineage cells of hemogenic endothelial origin through a process we define as lymphvasculogenesis. This is contrary to the current dogma that all mammalian lymphatic vessels form by sprouting from veins. Our results reveal vascular-bed-specific differences in the origin and mechanisms of vessel formation, which may critically underlie organ-specific manifestation of lymphatic dysfunction in disease. The progenitor cells identified in this study may be exploited to restore lymphatic function following cancer surgery, lymphedema, or tissue trauma.

  6. Epigallocatechin Gallate Inhibits Mouse Mesenchymal Stem Cell Differentiation to Adipogenic Lineage.

    Science.gov (United States)

    Chani, Baldeep; Puri, Veena; Chander Sobti, Ranbir; Puri, Sanjeev

    2016-01-01

    Epigallocatechin gallate (EGCG) is a major component of green tea polyphenols having a potent anti-oxidant potential. Besides inhibiting the growth of many cancer cell types and inducing proliferation and differentiation in keratinocytes, it has been shown to promote reduction of body fat. The fact that mesenchymal stem cells (MSCs) have ability to self-renew and differentiate into the cells of mesodermal lineages, such as fat and bone, it is, thus, possible that EGCG may directly be involved in affecting fat metabolism through its effect on mesenchymal stem cells. Hence, with this aim, the present study was designed to determine the effect of EGCG on mouse mesenchymal stem cells, C3H10T1/2 cells differentiation into adipocytes. To understand this process, the cells were incubated with varying concentrations of EGCG (1 μM, 5 μM, 10 μM, 50 μM) in the presence and /or absence of adipogenic medium for 9 days. The results demonstrated that, EGCG inhibited the cells proliferation, migration and also prevented their differentiation to adipogenic lineage. These effects were analyzed through the inhibition of wound healing activity, reduction in Oil red O stained cells, together with decrease in the expression of Adipisin gene following EGCG treatment. These observations thus demonstrated anti-adipogenic effect of EGCG with a possibility of its role in the therapeutic intervention of obesity. PMID:27397998

  7. Notch signalling inhibits CD4 expression during initiation and differentiation of human T cell lineage.

    Directory of Open Access Journals (Sweden)

    Stephen M Carlin

    Full Text Available The Delta/Notch signal transduction pathway is central to T cell differentiation from haemopoietic stem cells (HSCs. Although T cell development is well characterized using expression of cell surface markers, the detailed mechanisms driving differentiation have not been established. This issue becomes central with observations that adult HSCs exhibit poor differentiation towards the T cell lineage relative to neonatal or embryonic precursors. This study investigates the contribution of Notch signalling and stromal support cells to differentiation of adult and Cord Blood (CB human HSCs, using the Notch signalling OP9Delta co-culture system. Co-cultured cells were assayed at weekly intervals during development for phenotype markers using flow cytometry. Cells were also assayed for mRNA expression at critical developmental stages. Expression of the central thymocyte marker CD4 was initiated independently of Notch signalling, while cells grown with Notch signalling had reduced expression of CD4 mRNA and protein. Interruption of Notch signalling in partially differentiated cells increased CD4 mRNA and protein expression, and promoted differentiation to CD4(+ CD8(+ T cells. We identified a set of genes related to T cell development that were initiated by Notch signalling, and also a set of genes subsequently altered by Notch signal interruption. These results demonstrate that while Notch signalling is essential for establishment of the T cell lineage, at later stages of differentiation, its removal late in differentiation promotes more efficient DP cell generation. Notch signalling adds to signals provided by stromal cells to allow HSCs to differentiate to T cells via initiation of transcription factors such as HES1, GATA3 and TCF7. We also identify gene expression profile differences that may account for low generation of T cells from adult HSCs.

  8. Lineage-specific expression of bestrophin-2 and bestrophin-4 in human intestinal epithelial cells

    DEFF Research Database (Denmark)

    Ito, Go; Okamoto, Ryuichi; Murano, Tatsuro;

    2013-01-01

    Intestinal epithelial cells (IECs) regulate the absorption and secretion of anions, such as HCO3(-) or Cl(-). Bestrophin genes represent a newly identified group of calcium-activated Cl(-) channels (CaCCs). Studies have suggested that, among the four human bestrophin-family genes, bestrophin-2...... (BEST2) and bestrophin-4 (BEST4) might be expressed within the intestinal tissue. Consistently, a study showed that BEST2 is expressed by human colonic goblet cells. However, their precise expression pattern along the gastrointestinal tract, or the lineage specificity of the cells expressing these genes...

  9. Glutamate Receptor Agonists and Glutamate Transporter Antagonists Regulate Differentiation of Osteoblast Lineage Cells.

    Science.gov (United States)

    Xie, Wenjie; Dolder, Silvia; Siegrist, Mark; Wetterwald, Antoinette; Hofstetter, Willy

    2016-08-01

    Development and function of osteoblast lineage cells are regulated by a complex microenvironment consisting of the bone extracellular matrix, cells, systemic hormones and cytokines, autocrine and paracrine factors, and mechanical load. Apart from receptors that transduce extracellular signals into the cell, molecular transporters play a crucial role in the cellular response to the microenvironment. Transporter molecules are responsible for cellular uptake of nutritional components, elimination of metabolites, ion transport, and cell-cell communication. In this report, the expression of molecular transporters in osteoblast lineage cells was investigated to assess their roles in cell development and activity. Low-density arrays, covering membrane and vesicular transport molecules, were used to assess gene expression in osteoblasts representing early and late differentiation states. Receptors and transporters for the amino acid glutamate were found to be differentially expressed during osteoblast development. Glutamate is a neurotransmitter in the central nervous system, and the mechanisms of its release, signal transduction, and cellular reabsorption in the synaptic cleft are well understood. Less clear, however, is the control of equivalent processes in peripheral tissues. In primary osteoblasts, inhibition of glutamate transporters with nonselective inhibitors leads to an increase in the concentration of extracellular glutamate. This change was accompanied by a decrease in osteoblast proliferation, stimulation of alkaline phosphatase, and the expression of transcripts encoding osteocalcin. Enzymatic removal of extracellular glutamate abolished these pro-differentiation effects, as did the inhibition of PKC- and Erk1/2-signaling pathways. These findings demonstrate that glutamate signaling promotes differentiation and activation of osteoblast lineage cells. Consequently, the glutamate system may represent a putative therapeutic target to induce an anabolic response

  10. Stem cells and lineages of the intestine: a developmental and evolutionary perspective.

    Science.gov (United States)

    Takashima, Shigeo; Gold, David; Hartenstein, Volker

    2013-03-01

    The intestine consists of epithelial cells that secrete digestive enzymes and mucus (gland cells), absorb food particles (enterocytes), and produce hormones (endocrine cells). Intestinal cells are rapidly turned over and need to be replaced. In cnidarians, mitosis of differentiated intestinal cells accounts for much of the replacement; in addition, migratory, multipotent stem cells (interstitial cells) contribute to the production of intestinal cells. In other phyla, intestinal cell replacement is solely the function of stem cells entering the gut from the outside (such as in case of the neoblasts of platyhelminths) or intestinal stem cells located within the midgut epithelium (as in both vertebrates or arthropods). We will attempt in the following to review important aspects of midgut stem cells in different animal groups: where are they located, what types of lineages do they produce, and how do they develop. We will start out with a comparative survey of midgut cell types found across the animal kingdom; then briefly look at the specification of these cells during embryonic development; and finally focus on the stem cells that regenerate midgut cells during adult life. In a number of model systems, including mouse, zebrafish and Drosophila, the molecular pathways controlling intestinal stem cells proliferation and the specification of intestinal cell types are under intensive investigation. We will highlight findings of the recent literature, focusing on aspects that are shared between the different models and that point at evolutionary ancient mechanisms of intestinal cell formation. PMID:23179635

  11. Characterization of glucose-related metabolic pathways in differentiated rat oligodendrocyte lineage cells.

    Science.gov (United States)

    Amaral, Ana I; Hadera, Mussie G; Tavares, Joana M; Kotter, Mark R N; Sonnewald, Ursula

    2016-01-01

    Although oligodendrocytes constitute a significant proportion of cells in the central nervous system (CNS), little is known about their intermediary metabolism. We have, therefore, characterized metabolic functions of primary oligodendrocyte precursor cell cultures at late stages of differentiation using isotope-labelled metabolites. We report that differentiated oligodendrocyte lineage cells avidly metabolize glucose in the cytosol and pyruvate derived from glucose in the mitochondria. The labelling patterns of metabolites obtained after incubation with [1,2-(13)C]glucose demonstrated that the pentose phosphate pathway (PPP) is highly active in oligodendrocytes (approximately 10% of glucose is metabolized via the PPP as indicated by labelling patterns in phosphoenolpyruvate). Mass spectrometry and magnetic resonance spectroscopy analyses of metabolites after incubation of cells with [1-(13)C]lactate or [1,2-(13)C]glucose, respectively, demonstrated that anaplerotic pyruvate carboxylation, which was thought to be exclusive to astrocytes, is also active in oligodendrocytes. Using [1,2-(13)C]acetate, we show that oligodendrocytes convert acetate into acetyl CoA which is metabolized in the tricarboxylic acid cycle. Analysis of labelling patterns of alanine after incubation of cells with [1,2-(13)C]acetate and [1,2-(13)C]glucose showed catabolic oxidation of malate or oxaloacetate. In conclusion, we report that oligodendrocyte lineage cells at late differentiation stages are metabolically highly active cells that are likely to contribute considerably to the metabolic activity of the CNS. PMID:26352325

  12. Self-organization is a dynamic and lineage-intrinsic property of mammary epithelial cells

    Energy Technology Data Exchange (ETDEWEB)

    Chanson, L. [Ecole Polytechnique Federale de Lausanne (Switzerland). Inst. of Bioengineering; Brownfield, D. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Div.; Univ. of California, Berkeley, CA (United States). Dept. of Bioengineering; Garbe, J. C. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Div.; Kuhn, I. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Div.; Stampfer, M. R. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Div.; Bissell, M. J. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Div.; LaBarge, M. A. [Lawrence Berkeley National Lab. (LBNL), Berkeley, CA (United States). Life Sciences Div.

    2011-02-07

    Loss of organization is a principle feature of cancers; therefore it is important to understand how normal adult multilineage tissues, such as bilayered secretory epithelia, establish and maintain their architectures. The self-organization process that drives heterogeneous mixtures of cells to form organized tissues is well studied in embryology and with mammalian cell lines that were abnormal or engineered. Here we used a micropatterning approach that confined cells to a cylindrical geometry combined with an algorithm to quantify changes of cellular distribution over time to measure the ability of different cell types to self-organize relative to each other. Using normal human mammary epithelial cells enriched into pools of the two principal lineages, luminal and myoepithelial cells, we demonstrated that bilayered organization in mammary epithelium was driven mainly by lineage-specific differential E-cadherin expression, but that P-cadherin contributed specifically to organization of the myoepithelial layer. Disruption of the actomyosin network or of adherens junction proteins resulted in either prevention of bilayer formation or loss of preformed bilayers, consistent with continual sampling of the local microenvironment by cadherins. Together these data show that self-organization is an innate and reversible property of communities of normal adult human mammary epithelial cells.

  13. Does cell lineage in the developing cerebral cortex contribute to its columnar organization?

    Directory of Open Access Journals (Sweden)

    Marcos R Costa

    2010-06-01

    Full Text Available Since the pioneer work of Lorente de Nó, Ramón y Cajal, Brodmann, Mountcastle, Hubel and Wiesel and others, the cerebral cortex has been seen as a jigsaw of anatomic and functional modules involved in the processing of different sets of information. In fact, a columnar distribution of neurons displaying similar functional properties throughout the cerebral cortex has been observed by many researchers. Although it has been suggested that much of the anatomical substrate for such organization would be already specified at early developmental stages, before activity-dependent mechanisms could take place, it is still unclear whether gene expression in the ventricular zone could play a role in the development of discrete functional units, such as minicolumns or columns. Cell lineage experiments using replication-incompetent retroviral vectors have shown that the progeny of a single neuroepithelial/radial glial cell in the dorsal telencephalon is organized into discrete radial clusters of sibling excitatory neurons, which have a higher propensity for developing chemical synapses with each other rather than with neighbouring non-siblings. Here, we will discuss the possibility that the cell lineage of single neuroepithelial/radial glia cells could contribute for the columnar organization of the neocortex by generating radial columns of sibling, interconnected neurons. Borrowing some concepts from the studies on cell-cell recognition and transcription factor networks, we will also touch upon the potential molecular mechanisms involved in the establishment of sibling-neuron circuits.

  14. Tcf7l1 prepares epiblast cells in the gastrulating mouse embryo for lineage specification

    OpenAIRE

    Hoffman, Jackson A.; Wu, Chun-I; Merrill, Bradley J.

    2013-01-01

    The core gene regulatory network (GRN) in embryonic stem cells (ESCs) integrates activities of the pro-self-renewal factors Oct4 (Pou5f1), Sox2 and Nanog with that of an inhibitor of self-renewal, Tcf7l1 (Tcf3). The inhibitor function of Tcf7l1 causes dependence on extracellular Wnt/β-catenin signaling activity, making its embryonic role within the ESC GRN unclear. By analyzing intact mouse embryos, we demonstrate that the function of Tcf7l1 is necessary for specification of cell lineages to ...

  15. Sox17 regulates organ lineage segregation of ventral foregut progenitor cells

    OpenAIRE

    Spence, Jason R.; Lange, Alex W.; Lin, Suh-Chin J.; Kaestner, Klaus H.; Lowy, Andrew M.; Kim, Injune; Whitsett, Jeffrey A.; Wells, James M.

    2009-01-01

    The ventral pancreas, biliary system and liver arise from the posterior ventral foregut, but the cell-intrinsic pathway by which these organ lineages are separated is not known. Here we show that the extrahepatobiliary system shares a common origin with the ventral pancreas and not the liver, as previously thought. These pancreatobiliary progenitor cells coexpress the transcription factors Pdx1 and Sox17 at e8.5 and their segregation into a Pdx1+ ventral pancreas and a Sox17+ biliary primordi...

  16. Cells of renin lineage express hypoxia inducible factor 2α following experimental ureteral obstruction

    OpenAIRE

    Stefanska, Ania; Eng, Diana; Kaverina, Natalya; Pippin, Jeffrey W.; Gross, Kenneth W.; Duffield, Jeremy S.; Shankland, Stuart J.

    2016-01-01

    Background Recent studies indicate that mural cells of the preglomerular vessels, known as cells of renin lineage (CoRL), contribute to repair and regeneration of injured kidney glomeruli. However, their potential roles in tubulointerstitial disease are less understood. The aim of this study was to better understand CoRL number and distribution following UUO so that future mechanistic studies could be undertaken. Methods We mapped the fate of CoRL in adult Ren1cCreER x Rs-tdTomato-R reporter ...

  17. Inhibition of PPARγ in myeloid-lineage cells induces systemic inflammation, immunosuppression, and tumorigenesis

    OpenAIRE

    Wu, Lingyan; Yan, Cong; Czader, Magdalena; Foreman, Oded; Blum, Janice S.; Kapur, Reuben; Du, Hong

    2012-01-01

    Peroxisome proliferator–activated receptor-γ (PPARγ) is an anti-inflammatory molecule. To study its biologic function in myeloid cells, dominant-negative PPARγ (dnPPARγ) was overexpressed in a myeloid-specific bitransgenic mouse model. In this bitransgenic system, overexpression of the dnPPARγ-Flag fusion protein in myeloid-lineage cells abnormally elevated frequencies and total numbers of IL-7Rα−Lin−c-Kit+Sca-1−, Lin−/Scal+/c-Kit+, common myeloid, and granulocyte-monocyte progenitor populati...

  18. Essentials of Th17 cell commitment and plasticity

    Science.gov (United States)

    Restifo, Nicholas P.

    2013-01-01

    CD4+ T helper (Th) cells exist in a variety of epigenetic states that determine their function, phenotype, and capacity for persistence. These polarization states include Th1, Th2, Th17, and Foxp3+ T regulatory cells, as well as the more recently described T follicular helper, Th9, and Th22 cells. Th17 cells express the master transcriptional regulator retinoic acid–related orphan receptor γ thymus and produce canonical interleukin (IL)-17A and IL-17F cytokines. Th17 cells display a great degree of context-dependent plasticity, as they are capable of acquiring functional characteristics of Th1 cells. This late plasticity may contribute to the protection against microbes, plays a role in the development of autoimmunity, and is necessary for antitumor activity of Th17 cells in adoptive cell transfer therapy models. Moreover, plasticity of this subset is associated with higher in vivo survival and self-renewal capacity and less senescence than Th1 polarized cells, which have less plasticity and more phenotypic stability. New findings indicate that subset polarization of CD4+ T cells not only induces characteristic patterns of surface markers and cytokine production but also has a maturational aspect that affects a cell’s ability to survive, respond to secondary stimulation, and form long-term immune memory. PMID:23325835

  19. From Adult Bone Marrow Cells to Other Cell Lineages:Transdifferentiation or Cells Fusion

    Institute of Scientific and Technical Information of China (English)

    2003-01-01

    Recent studies have demonstrated that intravenous transplantation or local injection of bone marrow cells can induce unexpected changes of their fate. The results of these experiments showed that after transplantation or injecton, some of tissue specific somatic cells such as hepatocytes, skeleton, cardiac muscle cells and brain cells expressed the donor cell-specific genes, such as Y chromosome. There are two hypotheses that can explain this phenomenon. One is bone marrow stem cell transdifferentiation and the other is spontaneous cell fusion.

  20. Reduced reactivation from dormancy but maintained lineage choice of human mesenchymal stem cells with donor age.

    Directory of Open Access Journals (Sweden)

    Verena Dexheimer

    Full Text Available UNLABELLED: Mesenchymal stem cells (MSC are promising for cell-based regeneration therapies but up to date it is still controversial whether their function is maintained throughout ageing. Aim of this study was to address whether frequency, activation in vitro, replicative function, and in vitro lineage choice of MSC is maintained throughout ageing to answer the question whether MSC-based regeneration strategies should be restricted to younger individuals. MSC from bone marrow aspirates of 28 donors (5-80 years were characterized regarding colony-forming unit-fibroblast (CFU-F numbers, single cell cloning efficiency (SSCE, osteogenic, adipogenic and chondrogenic differentiation capacity in vitro. Alkaline phosphatase (ALP activity, mineralization, Oil Red O content, proteoglycan- and collagen type II deposition were quantified. While CFU-F frequency was maintained, SSCE and early proliferation rate decreased significantly with advanced donor age. MSC with higher proliferation rate before start of induction showed stronger osteogenic, adipogenic and chondrogenic differentiation. MSC with high osteogenic capacity underwent better chondrogenesis and showed a trend to better adipogenesis. Lineage choice was, however, unaltered with age. CONCLUSION: Ageing influenced activation from dormancy and replicative function of MSC in a way that it may be more demanding to mobilize MSC to fast cell growth at advanced age. Since fast proliferation came along with high multilineage capacity, the proliferation status of expanded MSC rather than donor age may provide an argument to restrict MSC-based therapies to certain individuals.

  1. Regulation of HIV-1 transcription in cells of the monocyte-macrophage lineage

    Directory of Open Access Journals (Sweden)

    Shah Sonia

    2009-12-01

    Full Text Available Abstract Human immunodeficiency virus type 1 (HIV-1 has been shown to replicate productively in cells of the monocyte-macrophage lineage, although replication occurs to a lesser extent than in infected T cells. As cells of the monocyte-macrophage lineage become differentiated and activated and subsequently travel to a variety of end organs, they become a source of infectious virus and secreted viral proteins and cellular products that likely initiate pathological consequences in a number of organ systems. During this process, alterations in a number of signaling pathways, including the level and functional properties of many cellular transcription factors, alter the course of HIV-1 long terminal repeat (LTR-directed gene expression. This process ultimately results in events that contribute to the pathogenesis of HIV-1 infection. First, increased transcription leads to the upregulation of infectious virus production, and the increased production of viral proteins (gp120, Tat, Nef, and Vpr, which have additional activities as extracellular proteins. Increased viral production and the presence of toxic proteins lead to enhanced deregulation of cellular functions increasing the production of toxic cellular proteins and metabolites and the resulting organ-specific pathologic consequences such as neuroAIDS. This article reviews the structural and functional features of the cis-acting elements upstream and downstream of the transcriptional start site in the retroviral LTR. It also includes a discussion of the regulation of the retroviral LTR in the monocyte-macrophage lineage during virus infection of the bone marrow, the peripheral blood, the lymphoid tissues, and end organs such as the brain. The impact of genetic variation on LTR-directed transcription during the course of retrovirus disease is also reviewed.

  2. Lineage tracing of human B cells reveals the in vivo landscape of human antibody class switching.

    Science.gov (United States)

    Horns, Felix; Vollmers, Christopher; Croote, Derek; Mackey, Sally F; Swan, Gary E; Dekker, Cornelia L; Davis, Mark M; Quake, Stephen R

    2016-01-01

    Antibody class switching is a feature of the adaptive immune system which enables diversification of the effector properties of antibodies. Even though class switching is essential for mounting a protective response to pathogens, the in vivo patterns and lineage characteristics of antibody class switching have remained uncharacterized in living humans. Here we comprehensively measured the landscape of antibody class switching in human adult twins using antibody repertoire sequencing. The map identifies how antibodies of every class are created and delineates a two-tiered hierarchy of class switch pathways. Using somatic hypermutations as a molecular clock, we discovered that closely related B cells often switch to the same class, but lose coherence as somatic mutations accumulate. Such correlations between closely related cells exist when purified B cells class switch in vitro, suggesting that class switch recombination is directed toward specific isotypes by a cell-autonomous imprinted state. PMID:27481325

  3. Differentiation of human embryonic stem cells along a hepatocyte lineage and its application in liver regeneration

    Institute of Scientific and Technical Information of China (English)

    2008-01-01

    Hepatocyte transplantation and bioartificial liver(BAL)as alternatives to liver transplantation offer the possibility of effective treatment for many inherited and acquired hepatic disorders.Unfortunately,the limited availability of donated livers and the variability of their derived hepatocytes make it difficult to obtain enough viable human hepatocytes for the hepatocyte-based therapies.Embryonic stem cells (ESCs),which could be isolated directly from the blastocyst inner cell mass,have permanent self-renewal capability and developmental pluripotency and therefore might be an ideal cell source in the treatment of hepatic discords.However,differentiation of hESCS into hepatocytes with significant numbers remains a challenge.This review updates our current understanding of differentiation of ESCs into hepatic lineage cells,their future therapeutic uses and problems in liver regeneration.

  4. The cell cycle of Chlamydomonas reinhardtii: the role of the commitment point

    Czech Academy of Sciences Publication Activity Database

    Oldenhof, H.; Zachleder, Vilém; van den Ende, H.

    2007-01-01

    Roč. 52, č. 1 (2007), s. 53-60. ISSN 0015-5632 Institutional research plan: CEZ:AV0Z50200510 Keywords : chlamydomonas reinhardtii * cell cycle * commitment point Subject RIV: EE - Microbiology, Virology Impact factor: 0.989, year: 2007

  5. NG2 targets tumorigenic Rb inactivation in Pit1-lineage pituitary cells.

    Science.gov (United States)

    Tateno, Toru; Nakano-Tateno, Tae; Ezzat, Shereen; Asa, Sylvia L

    2016-05-01

    The proteoglycan neuron-glial antigen 2 (NG2) is expressed by oligodendrocyte progenitors, pericytes, and some cancerous cells where it is implicated in tumor development. We examined mice with NG2-driven pRb inactivation. Unexpectedly, NG2-Cre:pRb(flox/flox) mice developed pituitary tumors with high penetrance. Adenohypophysial neoplasms developed initially as multifocal lesions; by 1 year, large tumors showed brain invasion. Immunohistochemistry identified these as Pit1-lineage neoplasms, with variable immunoreactivity for growth hormone, prolactin, thyrotropin, and α-subunit of glycoprotein hormones. Other than modest hyperprolactinemia, circulating hormone levels were not elevated. To determine the role of NG2 in the pituitary, we investigated NG2 expression. Immunoreactivity was identified in anterior and posterior lobes but not in the intermediate lobe of the mouse pituitary; in the adenohypophysis, folliculostellate cells had the strongest NG2 immunoreactivity but showed no proliferation in response to Rb inactivation. Pit1-positive adenohypophysial cells were positive for NG2, but corticotroph and gonadotroph cells were negative. RT-PCR revealed NG2 expression in normal human pituitary and human pituitary tumors; immunohistochemistry localized NG2 in nontumorous human adenohypophysis with strongest positivity in folliculostellate cells, and in tumors of all types except corticotrophs. Functional studies in GH4 mammosomatotrophs showed that NG2 increases prolactin (PRL), reduces growth hormone (GH) expression, and enhances cell adhesion without influencing proliferation. In conclusion, NG2-driven pRb inactivation results in pituitary tumors that mimic endocrinologically inactive Pit1-lineage human pituitary tumors. This model identifies a role for NG2 in pituitary cell-type-specific functions and unmasks a protective role from Rb inactivation in folliculostellate cells; it can be used for further research, including preclinical testing of novel therapies

  6. Hyaluronan Is Crucial for Stem Cell Differentiation into Smooth Muscle Lineage.

    Science.gov (United States)

    Simpson, Russell M L; Hong, Xuechong; Wong, Mei Mei; Karamariti, Eirini; Bhaloo, Shirin Issa; Warren, Derek; Kong, Wei; Hu, Yanhua; Xu, Qingbo

    2016-05-01

    Deciphering the extracellular signals that regulate SMC differentiation from stem cells is vital to further our understanding of the pathogenesis of vascular disease and for development of cell-based therapies and tissue engineering. Hyaluronan (HA) has emerged as an important component of the stem cell niche, however its role during stem cell differentiation is a complicated and inadequately defined process. This study aimed to investigate the role of HA in embryonic stem cell (ESC) differentiation toward a SMC lineage. ESCs were seeded on collagen-IV in differentiation medium to generate ESC-derived SMCs (esSMCs). Differentiation coincided with increased HA synthase (HAS) 2 expression, accumulation of extracellular HA and its assembly into pericellular matrices. Inhibition of HA synthesis by 4-methylumbelliferone (4MU), removal of the HA coat by hyaluronidase (HYAL) or HAS2 knockdown led to abrogation of SMC gene expression. HA activates ERK1/2 and suppresses EGFR signaling pathways via its principle receptor, CD44. EGFR inactivation coincided with increased binding to CD44, which was further augmented by addition of high molecular weight (HMW)-HA either exogenously or via HAS2 overexpression through adenoviral gene transfer. HMW-HA-stimulated esSMCs displayed a functional role in vascular tissue engineering ex vivo, vasculogenesis in a matrigel plug model and SMC accumulation in neointimal lesions of vein grafts in mice. These findings demonstrate that HAS2-induced HA synthesis and organization drives ESC-SMC differentiation. Thus, remodeling of the HA microenvironment is a critical step in directing stem cell differentiation toward a vascular lineage, highlighting HA as a potential target for treatment of vascular diseases. Stem Cells 2016;34:1225-1238. PMID:26867148

  7. Wnt Signaling Regulates the Lineage Differentiation Potential of Mouse Embryonic Stem Cells through Tcf3 Down-Regulation

    OpenAIRE

    Yaser Atlasi; Rubina Noori; Claudia Gaspar; Patrick Franken; Andrea Sacchetti; Haleh Rafati; Tokameh Mahmoudi; Charles Decraene; Calin, George A; Merrill, Bradley J.; Riccardo Fodde

    2013-01-01

    Canonical Wnt signaling plays a rate-limiting role in regulating self-renewal and differentiation in mouse embryonic stem cells (ESCs). We have previously shown that mutation in the Apc (adenomatous polyposis coli) tumor suppressor gene constitutively activates Wnt signaling in ESCs and inhibits their capacity to differentiate towards ecto-, meso-, and endodermal lineages. However, the underlying molecular and cellular mechanisms through which Wnt regulates lineage differentiation in mouse ES...

  8. Isolation, characterization and multi-lineage differentiation of stem cells from human exfoliated deciduous teeth

    Science.gov (United States)

    ZHANG, NAN; CHEN, BAOXING; WANG, WEI; CHEN, CHAO; KANG, JIE; DENG, SAMUEL QINNAN; ZHANG, BIN; LIU, SHUWEI; HAN, FABIN

    2016-01-01

    The aim of the present study was to isolate stem cells from human exfoliated deciduous teeth (SHEDs) and identify their phenotypes and multi-lineage differentiation potential. Three SHED cell strains were successfully isolated from three exfoliated deciduous teeth from different human subjects using the outgrowth method. Flow cytometric analysis indicated that SHEDs displayed high expression of the mesenchymal cell markers CD73 and CD90 but low expression of the hematopoietic stem cell marker CD34. PCR analysis illustrated that SHEDs expressed the mesenchymal stem cell markers CD44, CD73 and CD90, the osteoblast markers Alpl, Runx2, CBFA1 and collagen I, the cartilage cell markers Col10a1 and Acan, the adipose cell markers PPARγ2 and LPL, and the neuronal stem cell marker Nestin. In vitro induction experiments demonstrated the potential of the SHEDs for osteogenic, adipogenic and neurogenic differentiation. These SHED cells may be useful for further stem cell research and future therapeutic applications. PMID:27151462

  9. Reversible Immortalization Enables Seamless Transdifferentiation of Primary Fibroblasts into Other Lineage Cells.

    Science.gov (United States)

    Xie, Fei; Gong, Kerui; Li, Ke; Zhang, Mingliang; Chang, Judy C; Jiang, Shizhong; Ye, Lin; Wang, Jiaming; Tan, Yuting; Kan, Yuet Wai

    2016-08-15

    Fibroblasts can be transdifferentiated directly into other somatic cells such as cardiomyocytes, hematopoietic cells, and neurons. An advantage of somatic cell differentiation without first generating induced pluripotent stem cells (iPSCs) is that it avoids contamination of the differentiated cells with residual iPSCs, which may cause teratoma. However, since primary fibroblasts from biopsy undergo senescence during repeated culture, it may be difficult to grow transdifferentiated cells in sufficient numbers for future therapeutic purposes. To circumvent this problem, we reversibly immortalized primary fibroblasts by using the piggyBac transposon to deliver the human telomerase reverse transcriptase (hTERT) gene hTERT plus SV40 Large T. Both approaches enabled fibroblasts to grow continuously without senescence, and neither caused teratoma formation in immunodeficient mice. However, fibroblasts immortalized with hTERT plus SV40 large T antigen accumulated chromosomal rearrangements, whereas fibroblasts immortalized with hTERT retained the normal karyotype. To transdifferentiate hTERT-immortalized fibroblasts into other somatic lineage cells, we transiently transfected them with episomal OCT4 and cultured them under neural cell growth condition with transposase to remove the transposon. Tripotent neural progenitor cells were seamlessly and efficiently generated. Thus, reversible immortalization of primary fibroblasts with hTERT will allow potential autologous cell-based therapeutics that bypass and simulate iPSC generation. PMID:27328768

  10. Bmi1 overexpression in the cerebellar granule cell lineage of mice affects cell proliferation and survival without initiating medulloblastoma formation

    Directory of Open Access Journals (Sweden)

    Hourinaz Behesti

    2013-01-01

    BMI1 is a potent inducer of neural stem cell self-renewal and neural progenitor cell proliferation during development and in adult tissue homeostasis. It is overexpressed in numerous human cancers – including medulloblastomas, in which its functional role is unclear. We generated transgenic mouse lines with targeted overexpression of Bmi1 in the cerebellar granule cell lineage, a cell type that has been shown to act as a cell of origin for medulloblastomas. Overexpression of Bmi1 in granule cell progenitors (GCPs led to a decrease in cerebellar size due to decreased GCP proliferation and repression of the expression of cyclin genes, whereas Bmi1 overexpression in postmitotic granule cells improved cell survival in response to stress by altering the expression of genes in the mitochondrial cell death pathway and of Myc and Lef-1. Although no medulloblastomas developed in ageing cohorts of transgenic mice, crosses with Trp53−/− mice resulted in a low incidence of medulloblastoma formation. Furthermore, analysis of a large collection of primary human medulloblastomas revealed that tumours with a BMI1high TP53low molecular profile are significantly enriched in Group 4 human medulloblastomas. Our data suggest that different levels and timing of Bmi1 overexpression yield distinct cellular outcomes within the same cellular lineage. Importantly, Bmi1 overexpression at the GCP stage does not induce tumour formation, suggesting that BMI1 overexpression in GCP-derived human medulloblastomas probably occurs during later stages of oncogenesis and might serve to enhance tumour cell survival.

  11. Lineage relationship of prostate cancer cell types based on gene expression

    Directory of Open Access Journals (Sweden)

    Ware Carol B

    2011-05-01

    Full Text Available Abstract Background Prostate tumor heterogeneity is a major factor in disease management. Heterogeneity could be due to multiple cancer cell types with distinct gene expression. Of clinical importance is the so-called cancer stem cell type. Cell type-specific transcriptomes are used to examine lineage relationship among cancer cell types and their expression similarity to normal cell types including stem/progenitor cells. Methods Transcriptomes were determined by Affymetrix DNA array analysis for the following cell types. Putative prostate progenitor cell populations were characterized and isolated by expression of the membrane transporter ABCG2. Stem cells were represented by embryonic stem and embryonal carcinoma cells. The cancer cell types were Gleason pattern 3 (glandular histomorphology and pattern 4 (aglandular sorted from primary tumors, cultured prostate cancer cell lines originally established from metastatic lesions, xenografts LuCaP 35 (adenocarcinoma phenotype and LuCaP 49 (neuroendocrine/small cell carcinoma grown in mice. No detectable gene expression differences were detected among serial passages of the LuCaP xenografts. Results Based on transcriptomes, the different cancer cell types could be clustered into a luminal-like grouping and a non-luminal-like (also not basal-like grouping. The non-luminal-like types showed expression more similar to that of stem/progenitor cells than the luminal-like types. However, none showed expression of stem cell genes known to maintain stemness. Conclusions Non-luminal-like types are all representatives of aggressive disease, and this could be attributed to the similarity in overall gene expression to stem and progenitor cell types.

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

    Science.gov (United States)

    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

  13. The intimate relationship between Human cytomegalovirus and the dendritic cell lineage

    Directory of Open Access Journals (Sweden)

    MatthewReeves

    2014-08-01

    Our current understanding of HCMV carriage and reactivation is that cellular differentiation of the CD34+ progenitor cells through the myeloid lineage, resulting in terminal differentiation to either a macrophage or dendritic cell (DC phenotype, is crucial for the reactivation event. In this mini-review, we will focus on the interaction of HCMV with DCs, with a particular emphasis on their role in reactivation, and discuss how the critical regulation of viral major immediate early gene expression appears to be delicately entwined with the activation of cellular pathways in differentiating DCs. Furthermore, we will also explore the possible immune consequences associated with reactivation in a professional antigen presenting cell and potential countermeasures HCMV employs to abrogate these.

  14. A role for mixed lineage kinases in granule cell apoptosis induced by cytoskeletal disruption

    DEFF Research Database (Denmark)

    Müller, Georg Johannes; Geist, Marie Aavang; Veng, Lone Merete;

    2006-01-01

    Microtubule disruption by colchicine induces apoptosis in selected neuronal populations. However, little is known about the upstream death signalling events mediating the neurotoxicity. We investigated first whether colchicine-induced granule cell apoptosis activates the c-Jun N-terminal kinase...... (JNK) pathway. Cultured murine cerebellar granule cells were exposed to 1 microm colchicine for 24 h. Activation of the JNK pathway was detected by western blotting as well as immunocytochemistry using antibodies against phospho-c-Jun (p-c-Jun). Next, adult male rats were injected...... intracerebroventricularly with colchicine (10 microg), and JNK pathway activation in dentate granule cells (DGCs) was detected by antibodies against p-c-Jun. The second part of the study tested the involvement of mixed lineage kinases (MLK) as upstream activators of the JNK pathway in colchicine toxicity, using CEP-1347, a...

  15. Human GATA-3: a lineage-restricted transcription factor that regulates the expression of the T cell receptor alpha gene.

    OpenAIRE

    Ho, I C; Vorhees, P; Marin, N; Oakley, B K; Tsai, S F; Orkin, S H; Leiden, J. M.

    1991-01-01

    In addition to its role in the recognition of foreign antigens, the T cell receptor (TCR) alpha gene serves as a model system for studies of developmentally-regulated, lineage-specific gene expression in T cells. TCR alpha gene expression is restricted to cells of the TCR alpha/beta+ lineage, and is controlled by a T cell-specific transcriptional enhancer located 4.5 kb 3' to the C alpha gene segment. The TCR alpha enhancer contains four nuclear protein binding sites called T alpha 1-T alpha ...

  16. Gata-3 Induces T Helper Cell Type 2 (Th2) Cytokine Expression and Chromatin Remodeling in Committed Th1 Cells

    OpenAIRE

    Lee, Hyun Jun; Takemoto, Naofumi; Kurata, Hirokazu; Kamogawa, Yumiko; Miyatake, Shoichiro; O'Garra, Anne; Arai, Naoko

    2000-01-01

    Committed T helper type 1 (Th1) and Th2 effector cells, resulting from chronic antigenic stimulation in interleukin (IL)-12 and IL-4, are implicated in the pathology of autoimmune and allergic diseases. Committed Th1 cells cannot be induced to change their cytokine profiles in response to antigenic stimulation and Th2 cytokine–inducing conditions. Here, we report that ectopic expression of GATA-3 induced Th2-specific cytokine expression not only in developing Th1 cells but also in otherwise i...

  17. Neurotrophic Effect of Bone Marrow Stromal Cells on Proliferation and Committed Differentiation of Ventral Mesencephalic Precursors

    OpenAIRE

    Xiao-dong Wang; Heng-zhu Zhang; Zhi-gang Gong; Xi-gang Yan; Qing Lan; Qiang Huang

    2011-01-01

    OBJECTIVE To explore the potential neurotrophic effect of bone marrow stromal cells (BMSCs) on cell proliferation and committed neuronal differentiation of ventral mesencephalic precursors (VMPs) in vitro.METHODS Ventral mesencephalic precursors from E11 inbred rat embryos and BMSCs from adult rats were cultured both separately and in co-culture. After a 7-day incubation in vitro, three conditioned culture media were obtained, termed VMP or common medium, BMSC medium, and BMSC+VMP medium. V...

  18. Deletion of the Scl +19 enhancer increases the blood stem cell compartment without affecting the formation of mature blood lineages

    OpenAIRE

    Spensberger, Dominik; Kotsopoulou, Ekaterini; Ferreira, Rita; Broccardo, Cyril; Scott, Linda M.; Fourouclas, Nasios; Ottersbach, Katrin; Green, Anthony R.; Göttgens, Berthold

    2012-01-01

    The stem cell leukemia (Scl)/Tal1 gene is essential for normal blood and endothelial development, and is expressed in hematopoietic stem cells (HSCs), progenitors, erythroid, megakaryocytic, and mast cells. The Scl +19 enhancer is active in HSCs and progenitor cells, megakaryocytes, and mast cells, but not mature erythroid cells. Here we demonstrate that in vivo deletion of the Scl +19 enhancer (Scl Δ19/Δ19 ) results in viable mice with normal Scl expression in mature hematopoietic lineages. ...

  19. Initiation of immune tolerance-controlled HIV gp41 neutralizing B cell lineages.

    Science.gov (United States)

    Zhang, Ruijun; Verkoczy, Laurent; Wiehe, Kevin; Munir Alam, S; Nicely, Nathan I; Santra, Sampa; Bradley, Todd; Pemble, Charles W; Zhang, Jinsong; Gao, Feng; Montefiori, David C; Bouton-Verville, Hilary; Kelsoe, Garnett; Larimore, Kevin; Greenberg, Phillip D; Parks, Robert; Foulger, Andrew; Peel, Jessica N; Luo, Kan; Lu, Xiaozhi; Trama, Ashley M; Vandergrift, Nathan; Tomaras, Georgia D; Kepler, Thomas B; Moody, M Anthony; Liao, Hua-Xin; Haynes, Barton F

    2016-04-27

    Development of an HIV vaccine is a global priority. A major roadblock to a vaccine is an inability to induce protective broadly neutralizing antibodies (bnAbs). HIV gp41 bnAbs have characteristics that predispose them to be controlled by tolerance. We used gp41 2F5 bnAb germline knock-in mice and macaques vaccinated with immunogens reactive with germline precursors to activate neutralizing antibodies. In germline knock-in mice, bnAb precursors were deleted, with remaining anergic B cells capable of being activated by germline-binding immunogens to make gp41-reactive immunoglobulin M (IgM). Immunized macaques made B cell clonal lineages targeted to the 2F5 bnAb epitope, but 2F5-like antibodies were either deleted or did not attain sufficient affinity for gp41-lipid complexes to achieve the neutralization potency of 2F5. Structural analysis of members of a vaccine-induced antibody lineage revealed that heavy chain complementarity-determining region 3 (HCDR3) hydrophobicity was important for neutralization. Thus, gp41 bnAbs are controlled by immune tolerance, requiring vaccination strategies to transiently circumvent tolerance controls. PMID:27122615

  20. Initiation of immune tolerance–controlled HIV gp41 neutralizing B cell lineages

    Science.gov (United States)

    Zhang, Ruijun; Verkoczy, Laurent; Wiehe, Kevin; Alam, S. Munir; Nicely, Nathan I.; Santra, Sampa; Bradley, Todd; Pemble, Charles W.; Zhang, Jinsong; Gao, Feng; Montefiori, David C.; Bouton-Verville, Hilary; Kelsoe, Garnett; Larimore, Kevin; Greenberg, Phillip D.; Parks, Robert; Foulger, Andrew; Peel, Jessica N.; Luo, Kan; Lu, Xiaozhi; Trama, Ashley M.; Vandergrift, Nathan; Tomaras, Georgia D.; Kepler, Thomas B.; Moody, M. Anthony; Liao, Hua-Xin; Haynes, Barton F.

    2016-01-01

    Development of an HIV vaccine is a global priority. A major roadblock to a vaccine is an inability to induce protective broadly neutralizing antibodies (bnAbs). HIV gp41 bnAbs have characteristics that predispose them to be controlled by tolerance. We used gp41 2F5 bnAb germline knock-in mice and macaques vaccinated with immunogens reactive with germline precursors to activate neutralizing antibodies. In germline knock-in mice, bnAb precursors were deleted, with remaining anergic B cells capable of being activated by germline-binding immunogens to make gp41-reactive immunoglobulin M (IgM). Immunized macaques made B cell clonal lineages targeted to the 2F5 bnAb epitope, but 2F5-like antibodies were either deleted or did not attain sufficient affinity for gp41-lipid complexes to achieve the neutralization potency of 2F5. Structural analysis of members of a vaccine-induced antibody lineage revealed that heavy chain complementarity-determining region 3 (HCDR3) hydrophobicity was important for neutralization. Thus, gp41 bnAbs are controlled by immune tolerance, requiring vaccination strategies to transiently circumvent tolerance controls. PMID:27122615

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

    Science.gov (United States)

    Pearson, Bret J; Sánchez Alvarado, Alejandro

    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

  2. Hematopoietic microenvironment. Origin, lineage, and transplantability of the stromal cells in long-term bone marrow cultures from chimeric mice

    Energy Technology Data Exchange (ETDEWEB)

    Perkins, S.; Fleischman, R.A.

    1988-04-01

    Studies of bone marrow transplant patients have suggested that the stromal cells of the in vitro hematopoietic microenvironment are transplantable into conditioned recipients. Moreover, in patients with myeloproliferative disorders, all of the stromal cells, which include presumptive endothelial cells, appear to be derived from hematopoietic precursors. To confirm these findings, we have constructed two chimeric mouse models: (a) traditional radiation chimeras, and (b) fetal chimeras, produced by placental injection of bone marrow into genetically anemic Wx/Wv fetuses, a technique that essentially precludes engraftment of nonhematopoietic cells. Using two-color indirect immunofluorescence, the stromal cells in long-term bone marrow culture derived from these chimeras were analyzed for donor or host origin by strain-specific H-2 antigens, and for cell lineage by a variety of other specific markers. 75-95% of the stromal cells were shown to be hematopoietic cells of the monocyte-macrophage lineage, based upon donor origin, phagocytosis, and expression of specific hematopoietic surface antigens. The remaining 5-25% of the stromal cells were exclusively host in origin. Apart from occasional fat cells, these cells uniformly expressed collagen type IV, laminin, and a surface antigen associated with endothelial cells. Since these endothelial-like cells are not transplantable into radiation or fetal chimeras, they are not derived from hematopoietic stem cells. The contrast between our findings and human studies suggests either unexpected species differences in the origin of stromal lineages or limitations in the previous methodology used to detect nonhematopoietic stromal cells.

  3. Hematopoietic microenvironment. Origin, lineage, and transplantability of the stromal cells in long-term bone marrow cultures from chimeric mice

    International Nuclear Information System (INIS)

    Studies of bone marrow transplant patients have suggested that the stromal cells of the in vitro hematopoietic microenvironment are transplantable into conditioned recipients. Moreover, in patients with myeloproliferative disorders, all of the stromal cells, which include presumptive endothelial cells, appear to be derived from hematopoietic precursors. To confirm these findings, we have constructed two chimeric mouse models: (a) traditional radiation chimeras, and (b) fetal chimeras, produced by placental injection of bone marrow into genetically anemic Wx/Wv fetuses, a technique that essentially precludes engraftment of nonhematopoietic cells. Using two-color indirect immunofluorescence, the stromal cells in long-term bone marrow culture derived from these chimeras were analyzed for donor or host origin by strain-specific H-2 antigens, and for cell lineage by a variety of other specific markers. 75-95% of the stromal cells were shown to be hematopoietic cells of the monocyte-macrophage lineage, based upon donor origin, phagocytosis, and expression of specific hematopoietic surface antigens. The remaining 5-25% of the stromal cells were exclusively host in origin. Apart from occasional fat cells, these cells uniformly expressed collagen type IV, laminin, and a surface antigen associated with endothelial cells. Since these endothelial-like cells are not transplantable into radiation or fetal chimeras, they are not derived from hematopoietic stem cells. The contrast between our findings and human studies suggests either unexpected species differences in the origin of stromal lineages or limitations in the previous methodology used to detect nonhematopoietic stromal cells

  4. Signaling pathways involved in megakaryocyte-mediated proliferation of osteoblast lineage cells.

    Science.gov (United States)

    Cheng, Ying-Hua; Streicher, Drew A; Waning, David L; Chitteti, Brahmananda R; Gerard-O'Riley, Rita; Horowitz, Mark C; Bidwell, Joseph P; Pavalko, Fredrick M; Srour, Edward F; Mayo, Lindsey D; Kacena, Melissa A

    2015-03-01

    Recent studies suggest that megakaryocytes (MKs) may play a significant role in skeletal homeostasis, as evident by the occurrence of osteosclerosis in multiple MK related diseases (Lennert et al., 1975; Thiele et al., 1999; Chagraoui et al., 2006). We previously reported a novel interaction whereby MKs enhanced proliferation of osteoblast lineage/osteoprogenitor cells (OBs) by a mechanism requiring direct cell-cell contact. However, the signal transduction pathways and the downstream effector molecules involved in this process have not been characterized. Here we show that MKs contact with OBs, via beta1 integrin, activate the p38/MAPKAPK2/p90RSK kinase cascade in the bone cells, which causes Mdm2 to neutralizes p53/Rb-mediated check point and allows progression through the G1/S. Interestingly, activation of MAPK (ERK1/2) and AKT, collateral pathways that regulate the cell cycle, remained unchanged with MK stimulation of OBs. The MK-to-OB signaling ultimately results in significant increases in the expression of c-fos and cyclin A, necessary for sustaining the OB proliferation. Overall, our findings show that OBs respond to the presence of MKs, in part, via an integrin-mediated signaling mechanism, activating a novel response axis that de-represses cell cycle activity. Understanding the mechanisms by which MKs enhance OB proliferation will facilitate the development of novel anabolic therapies to treat bone loss associated with osteoporosis and other bone-related diseases. PMID:25160801

  5. Tc17 cells are a proinflammatory, plastic lineage of pathogenic CD8+ T cells that induce GVHD without antileukemic effects.

    Science.gov (United States)

    Gartlan, Kate H; Markey, Kate A; Varelias, Antiopi; Bunting, Mark D; Koyama, Motoko; Kuns, Rachel D; Raffelt, Neil C; Olver, Stuart D; Lineburg, Katie E; Cheong, Melody; Teal, Bianca E; Lor, Mary; Comerford, Iain; Teng, Michele W L; Smyth, Mark J; McCluskey, James; Rossjohn, Jamie; Stockinger, Brigitta; Boyle, Glen M; Lane, Steven W; Clouston, Andrew D; McColl, Shaun R; MacDonald, Kelli P A; Hill, Geoffrey R

    2015-09-24

    IL-17-producing cells are important mediators of graft-versus-host disease (GVHD) after allogeneic stem cell transplantation (SCT). Here we demonstrate that a distinct CD8(+) Tc17 population develops rapidly after SCT but fails to maintain lineage fidelity such that they are unrecognizable in the absence of a fate reporter. Tc17 differentiation is dependent on alloantigen presentation by host dendritic cells (DCs) together with IL-6. Tc17 cells express high levels of multiple prototypic lineage-defining transcription factors (eg, RORγt, T-bet) and cytokines (eg, IL-17A, IL-22, interferon-γ, granulocyte macrophage colony-stimulating factor, IL-13). Targeted depletion of Tc17 early after transplant protects from lethal acute GVHD; however, Tc17 cells are noncytolytic and fail to mediate graft-versus-leukemia (GVL) effects. Thus, the Tc17 differentiation program during GVHD culminates in a highly plastic, hyperinflammatory, poorly cytolytic effector population, which we term "inflammatory iTc17" (iTc17). Because iTc17 cells mediate GVHD without contributing to GVL, therapeutic inhibition of iTc17 development in a clinical setting represents an attractive approach for separating GVHD and GVL. PMID:26206951

  6. Ecto-mesenchymal stem cells from dental pulp are committed to differentiate into active melanocytes

    Directory of Open Access Journals (Sweden)

    F Paino

    2010-10-01

    Full Text Available Dental pulp stem cells (DPSCs are multipotent stem cells derived from neural crest and mesenchyme and have the capacity to differentiate into multiple cell lineages. It has already been demonstrated that DPSCs differentiate into melanocyte-like cells but only when cultivated in a specific melanocyte differentiating medium. In this study we have shown, for the first time, that DPSCs are capable of spontaneously differentiating into mature melanocytes, which display molecular and ultrastructural features of full development, including the expression of melanocyte specific markers and the presence of melanosomes up to the terminal stage of maturation. We have also compared the differentiating features of DPSCs grown in different culture conditions, following the timing of differentiation at molecular and cytochemical levels and found that in all culture conditions full development of these cells was obtained, although at different times. The spontaneous differentiating potential of these cells strongly suggests their possible applications in regenerative medicine.

  7. Chicken globin gene transcription is cell lineage specific during the time of the switch

    International Nuclear Information System (INIS)

    Posttranscriptional silencing of embryonic globin gene expression occurs during hemoglobin switching in chickens. Here the authors use Percoll density gradients to fractionate the red blood cells of 5-9 day embryos in order to determine the cellular source and the timing of this posttranscriptional process. By means of nuclear run-on transcription in vitro they show that it is within mature primitive cells that production of embryonic globin mRNA is terminated posttranscriptionally. In contrast, young definitive cells produce little (or no) embryonic globin mRNA because of regulation at the transcriptional level. Thus the lineage specificity of embryonic and adult globin gene expression is determined transcriptionally, and the posttranscriptional process described by Landes et al. is a property of the senescing primitive cells, not a mechanism operative in the hemoglobin switch. This conclusion is supported by [3H]leucine incorporation experiments on Percoll-fractionated cells which reveal no posttranscriptional silencing of the embryonic genes during the early stages of the switch. In the course of these studies they have noticed a strong transcriptional pause near the second exon of the globin genes which is induced by 5,6-dichloro-1-β-D-ribofuranosylbenzimidazole (DRB) and which resembles a natural pause near that position

  8. Regulatory T cell reprogramming toward a Th2-cell-like lineage impairs oral tolerance and promotes food allergy.

    Science.gov (United States)

    Noval Rivas, Magali; Burton, Oliver T; Wise, Petra; Charbonnier, Louis-Marie; Georgiev, Peter; Oettgen, Hans C; Rachid, Rima; Chatila, Talal A

    2015-03-17

    Oral immunotherapy has had limited success in establishing tolerance in food allergy, reflecting failure to elicit an effective regulatory T (Treg) cell response. We show that disease-susceptible (Il4ra(F709)) mice with enhanced interleukin-4 receptor (IL-4R) signaling exhibited STAT6-dependent impaired generation and function of mucosal allergen-specific Treg cells. This failure was associated with the acquisition by Treg cells of a T helper 2 (Th2)-cell-like phenotype, also found in peripheral-blood allergen-specific Treg cells of food-allergic children. Selective augmentation of IL-4R signaling in Treg cells induced their reprogramming into Th2-like cells and disease susceptibility, whereas Treg-cell-lineage-specific deletion of Il4 and Il13 was protective. IL-4R signaling impaired the capacity of Treg cells to suppress mast cell activation and expansion, which in turn drove Th2 cell reprogramming of Treg cells. Interruption of Th2 cell reprogramming of Treg cells might thus provide candidate therapeutic strategies in food allergy. PMID:25769611

  9. Triennial Lactation Symposium: Bovine mammary epithelial cell lineages and parenchymal development.

    Science.gov (United States)

    Ellis, S; Akers, R M; Capuco, A V; Safayi, S

    2012-05-01

    Mammary development proceeds from an aggregation of cells in the ventral ectoderm to the establishment of an elaborate tree of alveoli, ducts, and cisternae. However, despite abundant data on endocrine regulation of ruminant mammary growth, we know comparatively little about cell lineages, expression of differentiation markers, and plasticity in mammary cell phenotype. Histologic analyses have revealed cell populations with distinct histochemical profiles, but functional assessment of cell populations during development has been limited to analysis of proliferation and frequency estimations of morphotypes. The lack of transplantation models, limited availability of validated antibodies with reactivity to bovine antigens, and similar technical challenges have generally hindered the pace of discovery, but the application of new technologies such as laser microdissection, transcriptional profiling, and multispectral image analysis are yielding important clues into bovine mammary cell ontogeny and developmental regulation. Our analyses have shown that prepubertal ovariectomy affects epithelial architecture, increases the proportion of cells expressing the estrogen receptor, and increases myoepithelial cell development, all concomitant with a dramatic reduction in the mass of parenchymal tissue. Our observations point to a dual role for ovarian secretions in the control of not only the rate of epithelial development, but also the nature of the parenchymal development. The balance of stimulus and inhibition pathways cooperatively regulates mammary growth. The increased reliance on objective staining analyses and quantitative approaches will ensure broader repeatability, application, and extension of the findings regarding the impact of the ovary and other regulatory entities and factors. Advances in understanding the ontogeny of mammary epithelial cells, coupled with established and increasing knowledge of endocrine factors affecting mammary development, may yield

  10. Mixed lineage kinase 3 is required for matrix metalloproteinase expression and invasion in ovarian cancer cells

    International Nuclear Information System (INIS)

    Mixed lineage kinase 3 (MLK3) is a mitogen-activated protein kinase kinase kinase (MAP3K) that activates MAPK signaling pathways and regulates cellular responses such as proliferation, migration and apoptosis. Here we report high levels of total and phospho-MLK3 in ovarian cancer cell lines in comparison to immortalized nontumorigenic ovarian epithelial cell lines. Using small interfering RNA (siRNA)-mediated gene silencing, we determined that MLK3 is required for the invasion of SKOV3 and HEY1B ovarian cancer cells. Furthermore, mlk3 silencing substantially reduced matrix metalloproteinase (MMP)-1, -2, -9 and -12 gene expression and MMP-2 and -9 activities in SKOV3 and HEY1B ovarian cancer cells. MMP-1, -2, -9 and-12 expression, and MLK3-induced activation of MMP-2 and MMP-9 requires both extracellular signal-regulated kinase (ERK) and c-Jun N-terminal kinase (JNK) activities. In addition, inhibition of activator protein-1 (AP-1) reduced MMP-1, MMP-9 and MMP-12 gene expression. Collectively, these findings establish MLK3 as an important regulator of MMP expression and invasion in ovarian cancer cells. -- Highlights: ► Ovarian cancer cell lines have high levels of total and phosphorylated MLK3. ► MLK3 is required for MMP expression and activity in ovarian cancer cells. ► MLK3 is required for invasion of SKOV3 and HEY1B ovarian cancer cells. ► MLK3-dependent regulation of MMP-2 and MMP-9 activities requires ERK and JNK.

  11. Activation of arylhydrocarbon receptor (AhR) in T lineage cells inhibits cellular growth

    Energy Technology Data Exchange (ETDEWEB)

    Nohara, K.; Tomohiro, I.; Chiharu, T. [National Institute for Environmental Studies, Tsukuba (Japan)

    2004-09-15

    Dioxins, including the most toxic congener, 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), exert their toxic effects by binding and activating the arylhydrocarbon receptor (AhR), a liganddependent transcription factor. Upon binding dioxins, the AhR in the cytoplasm is activated and translocated to the nucleus, where it heterodimerizes with another transcription factor, ARNT. The AhR/ARNT heterodimer modulates expressions of various genes by binding xenobiotic responsive elements (XREs) in their enhancer regions or modifies cellular functions through protein-protein interactions. The AhR activation by TCDD exposure induces various immunotoxic reactions including thymus involution and suppression of T cell-dependent antibody production. We have investigated the roles of AhR activation in T lineage cells and their underlying mechanisms by generating transgenic (Tg) mice expressing a constitutively active AhR (CA-AhR) mutant specifically in T cells and by transiently expressing the CA-AhR mutant in Jurkat T cells.

  12. Protocols for in vitro Differentiation of Human Mesenchymal Stem Cells into Osteogenic, Chondrogenic and Adipogenic Lineages.

    Science.gov (United States)

    Ciuffreda, Maria Chiara; Malpasso, Giuseppe; Musarò, Paola; Turco, Valentina; Gnecchi, Massimiliano

    2016-01-01

    Mesenchymal stem cells (MSC) possess high plasticity and the potential to differentiate into several different cell types; this characteristic has implications for cell therapy and reparative biotechnologies. MSC have been originally isolated from the bone marrow (BM-MSC), but they have been found also in other tissues such as adipose tissue, cord blood, synovium, skeletal muscle, and lung. MSC are able to differentiate in vitro and in vivo into several cell types such as bone, osteocytes, chondrocytes, adipocytes, and skeletal myocytes, just to name a few.During the last two decades, an increasing number of studies have proven the therapeutic potential of MSC for the treatment of neurodegenerative diseases, spinal cord and brain injuries, cardiovascular diseases, diabetes mellitus, and diseases of the skeleton. Their immuno-privileged profile allows both autologous and allogeneic use. For all these reasons, the scientific appeal of MSC is constantly on the rise.The identity of MSC is currently based on three main criteria: plastic-adherence capacity, defined epitope profile, and capacity to differentiate in vitro into osteocytes, chondrocytes, and adipocytes. Here, we describe standard protocols for the differentiation of BM-MSC into the osteogenic, chondrogenic, and adipogenic lineages. PMID:27236670

  13. Lipocalin-2 inhibits osteoclast formation by suppressing the proliferation and differentiation of osteoclast lineage cells

    International Nuclear Information System (INIS)

    Lipocalin-2 (LCN2) is a member of the lipocalin superfamily and plays a critical role in the regulation of various physiological processes, such as inflammation and obesity. In this study, we report that LCN2 negatively modulates the proliferation and differentiation of osteoclast precursors, resulting in impaired osteoclast formation. The overexpression of LCN2 in bone marrow-derived macrophages or the addition of recombinant LCN2 protein inhibits the formation of multinuclear osteoclasts. LCN2 suppresses macrophage colony-stimulating factor (M-CSF)-induced proliferation of osteoclast precursor cells without affecting their apoptotic cell death. Interestingly, LCN2 decreases the expression of the M-CSF receptor, c-Fms, and subsequently blocks its downstream signaling cascades. In addition, LCN2 inhibits RANKL-induced osteoclast differentiation and attenuates the expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which are important modulators in osteoclastogenesis. Mechanistically, LCN2 inhibits NF-κB signaling pathways, as demonstrated by the suppression of IκBα phosphorylation, nuclear translocation of p65, and NF-κB transcriptional activity. Thus, LCN2 is an anti-osteoclastogenic molecule that exerts its effects by retarding the proliferation and differentiation of osteoclast lineage cells. - Highlights: • LCN2 expression is regulated during osteoclast development. • LCN2 suppresses M-CSF-mediated osteoclast precursor proliferation. • LCN2 inhibits RANKL-induced osteoclast differentiation

  14. Lineage tracing of human B cells reveals the in vivo landscape of human antibody class switching

    Science.gov (United States)

    Horns, Felix; Vollmers, Christopher; Croote, Derek; Mackey, Sally F; Swan, Gary E; Dekker, Cornelia L; Davis, Mark M; Quake, Stephen R

    2016-01-01

    Antibody class switching is a feature of the adaptive immune system which enables diversification of the effector properties of antibodies. Even though class switching is essential for mounting a protective response to pathogens, the in vivo patterns and lineage characteristics of antibody class switching have remained uncharacterized in living humans. Here we comprehensively measured the landscape of antibody class switching in human adult twins using antibody repertoire sequencing. The map identifies how antibodies of every class are created and delineates a two-tiered hierarchy of class switch pathways. Using somatic hypermutations as a molecular clock, we discovered that closely related B cells often switch to the same class, but lose coherence as somatic mutations accumulate. Such correlations between closely related cells exist when purified B cells class switch in vitro, suggesting that class switch recombination is directed toward specific isotypes by a cell-autonomous imprinted state. DOI: http://dx.doi.org/10.7554/eLife.16578.001 PMID:27481325

  15. Lipocalin-2 inhibits osteoclast formation by suppressing the proliferation and differentiation of osteoclast lineage cells

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Hyun-Ju, E-mail: biohjk@knu.ac.kr [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Yoon, Hye-Jin [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Yoon, Kyung-Ae [Department of Orthopedic Surgery, Skeletal Diseases Genome Research Center, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Gwon, Mi-Ri; Jin Seong, Sook [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Suk, Kyoungho [Department of Pharmacology, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Kim, Shin-Yoon [Department of Orthopedic Surgery, Skeletal Diseases Genome Research Center, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of); Yoon, Young-Ran, E-mail: yry@knu.ac.kr [Department of Molecular Medicine, Cell and Matrix Research Institute, Clinical Trial Center, BK21 Plus KNU Biomedical Convergence Program, School of Medicine, Kyungpook National University, Daegu 700-422 (Korea, Republic of)

    2015-06-10

    Lipocalin-2 (LCN2) is a member of the lipocalin superfamily and plays a critical role in the regulation of various physiological processes, such as inflammation and obesity. In this study, we report that LCN2 negatively modulates the proliferation and differentiation of osteoclast precursors, resulting in impaired osteoclast formation. The overexpression of LCN2 in bone marrow-derived macrophages or the addition of recombinant LCN2 protein inhibits the formation of multinuclear osteoclasts. LCN2 suppresses macrophage colony-stimulating factor (M-CSF)-induced proliferation of osteoclast precursor cells without affecting their apoptotic cell death. Interestingly, LCN2 decreases the expression of the M-CSF receptor, c-Fms, and subsequently blocks its downstream signaling cascades. In addition, LCN2 inhibits RANKL-induced osteoclast differentiation and attenuates the expression of c-Fos and nuclear factor of activated T cells c1 (NFATc1), which are important modulators in osteoclastogenesis. Mechanistically, LCN2 inhibits NF-κB signaling pathways, as demonstrated by the suppression of IκBα phosphorylation, nuclear translocation of p65, and NF-κB transcriptional activity. Thus, LCN2 is an anti-osteoclastogenic molecule that exerts its effects by retarding the proliferation and differentiation of osteoclast lineage cells. - Highlights: • LCN2 expression is regulated during osteoclast development. • LCN2 suppresses M-CSF-mediated osteoclast precursor proliferation. • LCN2 inhibits RANKL-induced osteoclast differentiation.

  16. Identification of novel genes involved in the commitment of endodermal cells to the thymic epithelial cell fate

    OpenAIRE

    Mathieu, Yves D.

    2006-01-01

    The thymus provides the microenvironment for the maturation and selection of the majority of peripheral T cells. Endodermal cells of the ventral aspect of the third pharyngeal pouch (3rdpp) at 10.5 days of mouse gestation (E10.5) adopt a thymic epithelial cell fate while cells of the dorsal part of the 3rdpp give rise to the parathyroid glands. To identify novel genes potentially involved in the commitment of endodermal cells to the thymic epithelial cell fate, the transcriptome o...

  17. Lack of the p42 form of C/EBPα leads to spontaneous immortalization and lineage infidelity of committed myeloid progenitors

    DEFF Research Database (Denmark)

    Schuster, Mikkel B; Frank, Anne-Katrine; Bagger, Frederik O;

    2013-01-01

    transforming events. In this study, we use premalignant cells from a Cebpa mutant AML model, in which the LIC population resembles granulocyte-macrophage progenitors (GMPs), to show that premalignant GMPs undergo spontaneous immortalization with a high clonal frequency when cultured in vitro, suggesting that...

  18. Adipose derived mesenchymal stem cells express keratinocyte lineage markers in a co-culture model.

    Science.gov (United States)

    Irfan-Maqsood, M; Matin, M M; Heirani-Tabasi, A; Bahrami, M; Naderi-Meshkin, H; Mirahmadi, M; Hassanzadeh, H; Sanjar Moussavi, N; Raza-Shah, H; Raeesolmohaddeseen, M; Bidkhori, H; Bahrami, A R

    2016-01-01

    Cutaneous wound healing is a complex type of biological event involving proliferation, differentiation, reprograming, trans/de-differentiation, recruitment, migration, and apoptosis of a number of cells (keratinocytes, fibroblasts, endothelial cells, nerve cells and stem cells) to regenerate a multi-layered tissue that is damaged by either internal or external factors. The exact regeneration mechanism of damaged skin is still unknown but the epithelial and other kinds of stem cells located in skin play crucial roles in the healing process. In this work, a co-culture model composed of adipose derived mesenchymal stem cells and keratinocytes was developed to understand the cellular differentiation behaviour in wound healing. Human mesenchymal stem cells were isolated from waste lipoaspirates. Keratinocytes were isolated from neonatal rats skin as well from human adult skin. Both types of cells were cultured and their culturing behaviour was observed microscopically under regular intervals of time. The identity of both cells was confirmed by flow cytometry and qRT-PCR. Cells were co-cultured under the proposed co-culturing model and the model was observed for 7, 14 and 21 days. The cellular behaviour was studied based on change in morphology, colonization, stratification, migration and expression of molecular markers. Expression of molecular markers was studied at transcriptional level and change in cellular morphology and migration capabilities was observed under the invert microscope regularly. Successfully isolated and characterized mesenchymal stem cells were found to express keratinocyte lineage markers i.e. K5, K10, K14, K18, K19 and Involucrin when co-cultured with keratinocytes after 14 and 21 days. Their expression was found to increase by increasing the time span of cell culturing. The keratinocyte colonies started to disappear after 10 days of culturing which might be due to stratification process initiated by possibly transdifferentiated stem cells. It can

  19. The BCL6 RD2 domain governs commitment of activated B cells to form germinal centers.

    Science.gov (United States)

    Huang, Chuanxin; Gonzalez, David G; Cote, Christine M; Jiang, Yanwen; Hatzi, Katerina; Teater, Matt; Dai, Kezhi; Hla, Timothy; Haberman, Ann M; Melnick, Ari

    2014-09-11

    To understand how the Bcl6 transcriptional repressor functions in the immune system, we disrupted its RD2 repression domain in mice. Bcl6RD2(MUT) mice exhibit a complete loss of germinal center (GC) formation but retain normal extrafollicular responses. Bcl6RD2(MUT) antigen-engaged B cells migrate to the interfollicular zone and interact with cognate T helper cells. However, these cells fail to complete early GC-commitment differentiation and coalesce as nascent GC aggregates. Bcl6 directly binds and represses trafficking receptors S1pr1 and Gpr183 by recruiting Hdac2 through the RD2 domain. Deregulation of these genes impairs B cell migration and may contribute to GC failure in Bcl6RD2(MUT) mice. The development of functional GC-TFH cells was partially impaired in Bcl6RD2(MUT) mice. In contrast to Bcl6(-/-) mice, Bcl6RD2(MUT) animals experience no inflammatory disease or macrophage deregulation. These results reveal an essential role for RD2 repression in early GC commitment and striking biochemical specificity in Bcl6 control of humoral and innate immune-cell phenotypes. PMID:25176650

  20. The BCL6 RD2 Domain Governs Commitment of Activated B Cells to Form Germinal Centers

    Directory of Open Access Journals (Sweden)

    Chuanxin Huang

    2014-09-01

    Full Text Available To understand how the Bcl6 transcriptional repressor functions in the immune system, we disrupted its RD2 repression domain in mice. Bcl6RD2MUT mice exhibit a complete loss of germinal center (GC formation but retain normal extrafollicular responses. Bcl6RD2MUT antigen-engaged B cells migrate to the interfollicular zone and interact with cognate T helper cells. However, these cells fail to complete early GC-commitment differentiation and coalesce as nascent GC aggregates. Bcl6 directly binds and represses trafficking receptors S1pr1 and Gpr183 by recruiting Hdac2 through the RD2 domain. Deregulation of these genes impairs B cell migration and may contribute to GC failure in Bcl6RD2MUT mice. The development of functional GC-TFH cells was partially impaired in Bcl6RD2MUT mice. In contrast to Bcl6−/− mice, Bcl6RD2MUT animals experience no inflammatory disease or macrophage deregulation. These results reveal an essential role for RD2 repression in early GC commitment and striking biochemical specificity in Bcl6 control of humoral and innate immune-cell phenotypes.

  1. An in vitro adherence assay reveals that Helicobacter pylori exhibits cell lineage-specific tropism in the human gastric epithelium.

    OpenAIRE

    Falk, P; Roth, K A; Borén, T; Westblom, T U; Gordon, J I; Normark, S

    1993-01-01

    Helicobacter pylori is a microaerophilic bacterium found in the stomach of asymptomatic humans as well as patients with acid peptic disease and gastric adenocarcinoma. We have developed an in situ adherence assay to examine the cell lineage-specific nature of binding of this organism and to characterize the nature of cell surface receptors that recognize its adhesin. Fluorescein isothiocyanate-labeled H. pylori strains were bound to surface mucous cells present in the pit region of human and ...

  2. Dynamic changes in replication timing and gene expression during lineage specification of human pluripotent stem cells.

    Science.gov (United States)

    Rivera-Mulia, Juan Carlos; Buckley, Quinton; Sasaki, Takayo; Zimmerman, Jared; Didier, Ruth A; Nazor, Kristopher; Loring, Jeanne F; Lian, Zheng; Weissman, Sherman; Robins, Allan J; Schulz, Thomas C; Menendez, Laura; Kulik, Michael J; Dalton, Stephen; Gabr, Haitham; Kahveci, Tamer; Gilbert, David M

    2015-08-01

    Duplication of the genome in mammalian cells occurs in a defined temporal order referred to as its replication-timing (RT) program. RT changes dynamically during development, regulated in units of 400-800 kb referred to as replication domains (RDs). Changes in RT are generally coordinated with transcriptional competence and changes in subnuclear position. We generated genome-wide RT profiles for 26 distinct human cell types, including embryonic stem cell (hESC)-derived, primary cells and established cell lines representing intermediate stages of endoderm, mesoderm, ectoderm, and neural crest (NC) development. We identified clusters of RDs that replicate at unique times in each stage (RT signatures) and confirmed global consolidation of the genome into larger synchronously replicating segments during differentiation. Surprisingly, transcriptome data revealed that the well-accepted correlation between early replication and transcriptional activity was restricted to RT-constitutive genes, whereas two-thirds of the genes that switched RT during differentiation were strongly expressed when late replicating in one or more cell types. Closer inspection revealed that transcription of this class of genes was frequently restricted to the lineage in which the RT switch occurred, but was induced prior to a late-to-early RT switch and/or down-regulated after an early-to-late RT switch. Analysis of transcriptional regulatory networks showed that this class of genes contains strong regulators of genes that were only expressed when early replicating. These results provide intriguing new insight into the complex relationship between transcription and RT regulation during human development. PMID:26055160

  3. Loss of C/EBP alpha cell cycle control increases myeloid progenitor proliferation and transforms the neutrophil granulocyte lineage

    DEFF Research Database (Denmark)

    Porse, Bo T; Bryder, David; Theilgaard-Mönch, Kim; Hasemann, Marie S; Anderson, Kristina; Damgaard, Inge; Jacobsen, Sten Eirik W; Nerlov, Claus

    2005-01-01

    dissociate the ability of C/EBP alpha to block cell cycle progression through E2F inhibition from its function as a transcriptional activator impair the in vivo development of the neutrophil granulocyte and adipose lineages. We now show that such mutations increase the capacity of bone marrow (BM) myeloid...

  4. Stress-stiffening-mediated stem-cell commitment switch in soft responsive hydrogels

    Science.gov (United States)

    Das, Rajat K.; Gocheva, Veronika; Hammink, Roel; Zouani, Omar F.; Rowan, Alan E.

    2016-03-01

    Bulk matrix stiffness has emerged as a key mechanical cue in stem cell differentiation. Here, we show that the commitment and differentiation of human mesenchymal stem cells encapsulated in physiologically soft (~0.2-0.4 kPa), fully synthetic polyisocyanopeptide-based three-dimensional (3D) matrices that mimic the stiffness of adult stem cell niches and show biopolymer-like stress stiffening, can be readily switched from adipogenesis to osteogenesis by changing only the onset of stress stiffening. This mechanical behaviour can be tuned by simply altering the material’s polymer length whilst maintaining stiffness and ligand density. Our findings introduce stress stiffening as an important parameter that governs stem cell fate in a 3D microenvironment, and reveal a correlation between the onset of stiffening and the expression of the microtubule-associated protein DCAMKL1, thus implicating DCAMKL1 in a stress-stiffening-mediated, mechanotransduction pathway that involves microtubule dynamics in stem cell osteogenesis.

  5. High level PHGDH expression in breast is predominantly associated with keratin 5-positive cell lineage independently of malignancy

    DEFF Research Database (Denmark)

    Gromova, Irina; Gromov, Pavel; Honma, Naoko; Kumar, Sudha; Rimm, David; Talman, Maj-Lis Møller; Wielenga, Vera Timmermans; Moreira, José

    2015-01-01

    showed high-level expression of Phgdh in normal CK5-positive mammary epithelial cells, indicating that expression of this protein was not associated with malignancy, but rather with cell lineage. However, proteomic profiling of Phgdh showed it to be expressed in two major protein forms, and that the...... in TNBC samples. One such protein was D-3-phosphoglycerate dehydrogenase (Phgdh), a candidate oncogene. We analysed expression of Phgdh in normal and TNBC mammary tissue samples by 2D gel-based proteomics and immunohistochemistry (IHC), and show here that high-level expression of Phgdh in mammary...... epithelial cells is primarily associated with cell lineage, as we found that Phgdh expression was predominant in CK5-positive cells, normal as well as malignant, thus identifying an association of this protein with the basal phenotype. Quantitative IHC analysis of Phgdh expression in normal breast tissue...

  6. The Lineage Specification of Mesenchymal Stem Cells Is Directed by the Rate of Fluid Shear Stress.

    Science.gov (United States)

    Lu, Juan; Fan, Yijuan; Gong, Xiaoyuan; Zhou, Xin; Yi, Caixia; Zhang, Yinxing; Pan, Jun

    2016-08-01

    The effective regulation of fluid shear stress (FSS) on the lineage specification of mesenchymal stem cells (MSCs) remains to be addressed. We hypothesized that when MSCs are recruited to musculoskeletal system following stimulation, their differentiation into osteogenic or chondrogenic cells is directed by the rate of FSS (ΔSS) through modulation of the mechanosensitive, cation-selective channels (MSCCs), intracellular calcium levels, and F-actin. To this end, MSCs were exposed to laminar FSS linearly increased from 0 to 10 dyn/cm(2) in 0, 2, or 20 min and maintained at 10 dyn/cm(2) for a total of 20 min (termed as ΔSS 0-0', 0-2', and 0-20', respectively, representing more physiological (0-0') and non-physiological (0-2' and 0-20') ΔSS treatments). Our results showed 0-0' facilitated MSC differentiation towards chondrogenic and not osteogenic phenotype, by promoting moderate intracellular calcium concentration ([Ca(2+) ]i ) increase from the calcium channels with the exception of MSCCs or intracellular calcium stores, and F-actin organization. In contrast, 0-2' promoted MSCs towards osteogenic and not chondrogenic phenotype, by inducing significant [Ca(2+) ]i increase mainly from the MSCCs, and F-actin assembly. However, 0-20' elicited the modest osteogenic and chondrogenic phenotypes, as it induced the lowest [Ca(2+) ]i increase mainly from MSCCs, and F-actin assembly. Our results suggest that compared to the more physiological ΔSS, the non-physiological ΔSS favors [Ca(2+) ]i influx from MSCCs. An appropriate non-physiological ΔSS (0-2') even elicits a large [Ca(2+) ]i influx from the MSCCs that reverses the lineage specification of MSCs, providing validation for the high mechanosensitivity of MSCs and guidance for training osteoporosis and osteoarthritis patients. J. Cell. Physiol. 231: 1752-1760, 2016. © 2015 Wiley Periodicals, Inc. PMID:26636289

  7. Murine Mesenchymal Stem Cell Commitment to Differentiation Is Regulated by Mitochondrial Dynamics.

    Science.gov (United States)

    Forni, Maria Fernanda; Peloggia, Julia; Trudeau, Kyle; Shirihai, Orian; Kowaltowski, Alicia J

    2016-03-01

    Mouse skin mesenchymal stem cells (msMSCs) are dermis CD105(+) CD90(+) CD73(+) CD29(+) CD34(-) mesodermal precursors which, after in vitro induction, undergo chondro, adipo, and osteogenesis. Extensive metabolic reconfiguration has been found to occur during differentiation, and the bioenergetic status of a cell is known to be dependent on the quality and abundance of the mitochondrial population, which may be regulated by fusion and fission. However, little is known regarding the impact of mitochondrial dynamics on the differentiation process. We addressed this knowledge gap by isolating MSCs from Swiss female mice, inducing these cells to differentiate into osteo, chondro, and adipocytes and measuring changes in mass, morphology, dynamics, and bioenergetics. Mitochondrial biogenesis was increased in adipogenesis, as evaluated through confocal microscopy, citrate synthase activity, and mtDNA content. The early steps of adipo and osteogenesis involved mitochondrial elongation, as well as increased expression of mitochondrial fusion proteins Mfn1 and 2. Chondrogenesis involved a fragmented mitochondrial phenotype, increased expression of fission proteins Drp1, Fis1, and 2, and enhanced mitophagy. These events were accompanied by profound bioenergetic alterations during the commitment period. Moreover, knockdown of Mfn2 in adipo and osteogenesis and the overexpression of a dominant negative form of Drp1 during chondrogenesis resulted in a loss of differentiation ability. Overall, we find that mitochondrial morphology and its regulating processes of fission/fusion are modulated early on during commitment, leading to alterations in the bioenergetic profile that are important for differentiation. We thus propose a central role for mitochondrial dynamics in the maintenance/commitment of mesenchymal stem cells. PMID:26638184

  8. Tenascin expression patterns and cells of monocyte lineage: relationship in human gliomas.

    Science.gov (United States)

    Kulla, A; Liigant, A; Piirsoo, A; Rippin, G; Asser, T

    2000-01-01

    Stromal extracellular matrix (ECM) components are thought to play an important role in regulating invasion of human gliomas. Macrophages and microglial cells may heavily influence the integrity of the extracellular compartment of gliomas, and the affected ECM may play a key role in regulating migratory activity of both tumor cells and macrophages/microglia. The aim of this investigation was to study immunohistochemically the expression patterns of four ECM components: fibronectin, laminin, collagen IV, and tenascin (TN) in human gliomas, with special attention to TN. Our main goal was to study the possible correlation between TN expression and macrophagic/microglial infiltration in gliomas. Altogether, 90 gliomas were studied. Tumors included 46 glioblastomas, 19 anaplastic gliomas, 22 low grade gliomas, and 3 pilocytic astrocytomas. Vascular TN prevailed in perinecrotic areas of glioblastomas, whereas interstitial TN was more often expressed distant from necrosis and in the ECM of anaplastic and low grade gliomas. Double staining with CD68 and anti-TN antibodies showed that macrophagic/microglial density was significantly higher in TN-positive areas of most of the glioblastomas and anaplastic gliomas, whereas microglial percentage from total number of CD68-positive cells was in most of the cases significantly higher in TN-negative areas. In addition, we saw a morphologically spatial correlation between higher densities of macrophagic/microglial infiltration and TN expression in perinecrotic areas in glioblastomas. Attachment of macrophages to TN-positive basement membrane zones of newly formed stromal blood vessels was evident. On the basis of our results, we conclude that TN may play a crucial role in regulating trafficking of cells of monocyte lineage in human gliomas. PMID:10658911

  9. Oct4 is required for lineage priming in the developing inner cell mass of the mouse blastocyst

    OpenAIRE

    Le Bin, Gloryn Chia; Muñoz-Descalzo, Silvia; Kurowski, Agata; Leitch, Harry; Lou, Xinghua; Mansfield, William; Etienne-Dumeau, Charles; Grabole, Nils; Mulas, Carla; Niwa, Hitoshi; Hadjantonakis, Anna-Katerina; Nichols, Jennifer

    2014-01-01

    The transcription factor Oct4 is required in vitro for establishment and maintenance of embryonic stem cells and for reprogramming somatic cells to pluripotency. In vivo, it prevents the ectopic differentiation of early embryos into trophoblast. Here, we further explore the role of Oct4 in blastocyst formation and specification of epiblast versus primitive endoderm lineages using conditional genetic deletion. Experiments involving mouse embryos deficient for both maternal and zygotic Oct4 sug...

  10. Lineage-Specific Effector Signatures of Invariant NKT Cells Are Shared amongst γδ T, Innate Lymphoid, and Th Cells.

    Science.gov (United States)

    Lee, You Jeong; Starrett, Gabriel J; Lee, Seungeun Thera; Yang, Rendong; Henzler, Christine M; Jameson, Stephen C; Hogquist, Kristin A

    2016-08-15

    Invariant NKT cells differentiate into three predominant effector lineages in the steady state. To understand these lineages, we sorted undifferentiated invariant NK T progenitor cells and each effector population and analyzed their transcriptional profiles by RNAseq. Bioinformatic comparisons were made to effector subsets among other lymphocytes, specifically Th cells, innate lymphoid cells (ILC), and γδ T cells. Myc-associated signature genes were enriched in NKT progenitors, like in other hematopoietic progenitors. Only NKT1 cells, but not NKT2 and NKT17 cells, had transcriptome similarity to NK cells and were also similar to other IFN-γ-producing lineages such as Th1, ILC1, and intraepithelial γδ T cells. NKT2 and NKT17 cells were similar to their analogous subsets of γδ T cells and ILCs, but surprisingly, not to Th2 and Th17 cells. We identified a set of genes common to each effector lineage regardless of Ag receptor specificity, suggesting the use of conserved regulatory cores for effector function. PMID:27385777

  11. Group 3 innate lymphoid cells continuously require the transcription factor GATA3 after commitment

    OpenAIRE

    Zhong, Chao; Cui, Kairong; Wilhelm, Christoph; Hu, Gangqing; Mao, Kairui; Belkaid, Yasmine; Zhao, Keji; Zhu, Jinfang

    2015-01-01

    The transcription factor GATA3 is indispensable for the development of all interleukin-7 receptor α (IL-7Rα)-expressing innate lymphoid cells (ILCs). However, the functional role of low GATA3 expression in committed ILC3s has not been identified. We report that GATA3 regulates homeostasis of ILC3s by controlling IL-7Rα expression. In addition, GATA3 is critical for the development of the NKp46+ ILC3 subset by regulating the balance between the transcription factors T-bet and RORγt. Alhough GA...

  12. Muscle-derived stem cells isolated as non-adherent population give rise to cardiac, skeletal muscle and neural lineages

    International Nuclear Information System (INIS)

    Stem cells with the ability to differentiate in specialized cell types can be extracted from a wide array of adult tissues including skeletal muscle. Here we have analyzed a population of cells isolated from skeletal muscle on the basis of their poor adherence on uncoated or collagen-coated dishes that show multi-lineage differentiation in vitro. When analysed under proliferative conditions, these cells express stem cell surface markers Sca-1 (65%) and Bcrp-1 (80%) but also MyoD (15%), Neuronal β III-tubulin (25%), GFAP (30%) or Nkx2.5 (1%). Although capable of growing as non-attached spheres for months, when given an appropriate matrix, these cells adhere giving rise to skeletal muscle, neuronal and cardiac muscle cell lineages. A similar cell population could not be isolated from either bone marrow or cardiac tissue suggesting their specificity to skeletal muscle. When injected into damaged muscle, these non-adherent muscle-derived cells are retrieved expressing Pax7, in a sublaminar position characterizing satellite cells and participate in forming new myofibers. These data show that a non-adherent stem cell population can be specifically isolated and expanded from skeletal muscle and upon attachment to a matrix spontaneously differentiate into muscle, cardiac and neuronal lineages in vitro. Although competing with resident satellite cells, these cells are shown to significantly contribute to repair of injured muscle in vivo supporting that a similar muscle-derived non-adherent cell population from human muscle may be useful in treatment of neuromuscular disorders

  13. Plasticity of mesenchymal stem cells under microgravity: from cytoskeletal reorganization to commitment shift

    Science.gov (United States)

    Buravkova, Ludmila

    Mesenchymal stem cells (MSCs) can be used to examine osteogenesis of uncommitted cells maintaining the bone differentiation potential such as osteogenic gene expression, osteogenic markers, matrix maturation and mineralization. MSCs are therefore a good model for studying osteogenesis in the space environment. Recent investigations have demonstrated that MSCs change in response to microgravity and, consequently, can be involved in the development of osteopenia detected in space travelers. This is a factor that can limit human space missions due to potential risks of osteoporosis and its aftereffects during and after flight. Simulated microgravity inhibited MSC differentiation towards osteoblasts and accelerated adipocyte development due to cytoskeleton modifications, including its structure and regulation associated with signal transduction cascades. We identified transient changes in the actin cytoskeleton of non-committed human bone marrow MSCs in short-term RPM experiments. In addition, we detected transient changes in the expression of genes encoding actin cytoskeleton proteins and associated elements (ACTA1, ACTG, RHOA, CFL1, VCL). When discussing the microgravity effects on MSC osteogenic differentiation, it should be mentioned the inhibition of Runx2 and ALPL and stimulation of PPARg2 in the MSCs induced for osteogenesis. It is probable that the reciprocal regulation of the two transcription factors is a molecular mechanism underlying progenitor cell response to microgravity. It is very likely that these genes are involved in the universal circuits within which mechanical (or gravity ) signals are sensed by MSCs. Recently, the list of osteogenic markers was extended to include several new proteins as microgravity targets (proteoglycans, osteomodulin, osteoglycin). It can be believed that exposure to microgravity produces similar effects on mature bone cells (osteoblasts) and non-committed osteogenic cells (MSCs). This finds a support in the fact that

  14. A novel lineage restricted, pericyte-like cell line isolated from human embryonic stem cells.

    Science.gov (United States)

    Greenwood-Goodwin, Midori; Yang, Jiwei; Hassanipour, Mohammad; Larocca, David

    2016-01-01

    Pericytes (PCs) are endothelium-associated cells that play an important role in normal vascular function and maintenance. We developed a method comparable to GMP quality protocols for deriving self-renewing perivascular progenitors from the human embryonic stem cell (hESC), line ESI-017. We identified a highly scalable, perivascular progenitor cell line that we termed PC-A, which expressed surface markers common to mesenchymal stromal cells. PC-A cells were not osteogenic or adipogenic under standard differentiation conditions and showed minimal angiogenic support function in vitro. PC-A cells were capable of further differentiation to perivascular progenitors with limited differentiation capacity, having osteogenic potential (PC-O) or angiogenic support function (PC-M), while lacking adipogenic potential. Importantly, PC-M cells expressed surface markers associated with pericytes. Moreover, PC-M cells had pericyte-like functionality being capable of co-localizing with human umbilical vein endothelial cells (HUVECs) and enhancing tube stability up to 6 days in vitro. We have thus identified a self-renewing perivascular progenitor cell line that lacks osteogenic, adipogenic and angiogenic potential but is capable of differentiation toward progenitor cell lines with either osteogenic potential or pericyte-like angiogenic function. The hESC-derived perivascular progenitors described here have potential applications in vascular research, drug development and cell therapy. PMID:27109637

  15. Precursor cells from Atlantic salmon (Salmo salar visceral fat holds the plasticity to differentiate into the osteogenic lineage

    Directory of Open Access Journals (Sweden)

    Elisabeth Ytteborg

    2015-07-01

    Full Text Available In order to study the potential plasticity of Atlantic salmon (Salmo salar precursor cells (aSPCs from the adipogenic mesenchyme cell lineage to differentiate to the osteogenic lineage, aSPCs were isolated and cultivated under either osteogenic or adipogenic promoting conditions. The results strengthen the hypothesis that aSPCs most likely are predestined to the adipogenic lineage, but they also hold the flexibility to turn into other lineages given the right stimuli. This assumption is supported by the fact that the transcription factor pparγ , important for regulation of adiopogenesis, was silent in aSPCs grown in osteogenic media, while runx2, important for osteogenic differentiation, was not expressed in aSPCs cultivated in adipogenic media. After 2 weeks in osteogenic promoting conditions the cells started to deposit extracellular matrix and after 4 weeks, the cells started mineralizing secreted matrix. Microarray analyses revealed large-scale transcriptome responses to osteogenic medium after 2 days, changes remained stable at day 15 and decreased by magnitude at day 30. Induction was observed in many genes involved in osteogenic differentiation, growth factors, regulators of development, transporters and production of extracellular matrix. Transcriptome profile in differentiating adipocytes was markedly different from differentiating osteoblasts with far fewer genes changing activity. The number of regulated genes slowly increased at the mature stage, when adipocytes increased in size and accumulated lipids. This is the first report on in vitro differentiation of aSPCs from Atlantic salmon to mineralizing osteogenic cells. This cell model system provides a new valuable tool for studying osteoblastogenesis in fish.

  16. Insulin-positive, Glut2-low cells present within mouse pancreas exhibit lineage plasticity and are enriched within extra-islet endocrine cell clusters.

    Science.gov (United States)

    Beamish, Christine A; Strutt, Brenda J; Arany, Edith J; Hill, David J

    2016-04-18

    Regeneration of insulin-producing β-cells from resident pancreas progenitors requires an understanding of both progenitor identity and lineage plasticity. One model suggested that a rare β-cell sub-population within islets demonstrated multi-lineage plasticity. We hypothesized that β-cells from young mice (postnatal day 7, P7) exhibit such plasticity and used a model of islet dedifferentiation toward a ductal epithelial-cell phenotype to test this theory. RIPCre;Z/AP(+/+) mice were used to lineage trace the fate of β-cells during dedifferentiation culture by a human placental alkaline phosphatase (HPAP) reporter. There was a significant loss of HPAP-expressing β-cells in culture, but remaining HPAP(+) cells lost insulin expression while gaining expression of the epithelial duct cell marker cytokeratin-19 (Ck19). Flow cytometry and recovery of β-cell subpopulations from whole pancreas vs. islets suggest that the HPAP(+)Ck19(+) cells had derived from insulin-positive, glucose-transporter-2-low (Ins(+)Glut2(LO)) cells, representing 3.5% of all insulin-expressing cells. The majority of these cells were found outside of islets within clusters of <5 β-cells. These insulin(+)Glut2(LO) cells demonstrated a greater proliferation rate in vivo and in vitro as compared to insulin(+)Glut2(+) cells at P7, were retained into adulthood, and a subset differentiated into endocrine, ductal, and neural lineages, illustrating substantial plasticity. Results were confirmed using RIPCre;ROSA- eYFP mice. Quantitative PCR data indicated these cells possess an immature β-cell phenotype. These Ins(+)Glut2(LO) cells may represent a resident population of cells capable of forming new, functional β-cells, and which may be potentially exploited for regenerative therapies in the future. PMID:27010375

  17. A reaction–diffusion mechanism influences cell lineage progression as a basis for formation, regeneration, and stability of intestinal crypts

    Directory of Open Access Journals (Sweden)

    Zhang Lei

    2012-07-01

    Full Text Available Abstract Background Colon crypts, a single sheet of epithelia cells, consist of a periodic pattern of stem cells, transit-amplifying cells, and terminally differentiated cells that constantly renew and turnover. Experimental evidence suggests that Wnt signaling promotes and regulates stem cell division, differentiation, and possible cell migrations while intestinal BMP signaling inhibits stem cell self-renewal and repression in crypt formation. As more molecular details on Wnt and BMP in crypts are being discovered, little is still known about how complex interactions among Wnt, BMP, and different types of cells, and surrounding environments may lead to de novo formation of multiple crypts or how such interactions affect regeneration and stability of crypts. Results We present a mathematical model that contains Wnt and BMP, a cell lineage, and their feedback regulations to study formation, regeneration, and stability of multiple crypts. The computational explorations and linear stability analysis of the model suggest a reaction–diffusion mechanism, which exhibits a short-range activation of Wnt plus a long-range inhibition with modulation of BMP signals in a growing tissue of cell lineage, can account for spontaneous formation of multiple crypts with the spatial and temporal pattern observed in experiments. Through this mechanism, the model can recapitulate some distinctive and important experimental findings such as crypt regeneration and crypt multiplication. BMP is important in maintaining stability of crypts and loss of BMP usually leads to crypt multiplication with a fingering pattern. Conclusions The study provides a mechanism for de novo formation of multiple intestinal crypts and demonstrates a synergetic role of Wnt and BMP in regeneration and stability of intestinal crypts. The proposed model presents a robust framework for studying spatial and temporal dynamics of cell lineages in growing tissues driven by multiple signaling

  18. Directing lineage specification of human mesenchymal stem cells by decoupling electrical stimulation and physical patterning on unmodified graphene

    Science.gov (United States)

    Balikov, Daniel A.; Fang, Brian; Chun, Young Wook; Crowder, Spencer W.; Prasai, Dhiraj; Lee, Jung Bok; Bolotin, Kiril I.; Sung, Hak-Joon

    2016-07-01

    The organization and composition of the extracellular matrix (ECM) have been shown to impact the propagation of electrical signals in multiple tissue types. To date, many studies with electroactive biomaterial substrates have relied upon passive electrical stimulation of the ionic media to affect cell behavior. However, development of cell culture systems in which stimulation can be directly applied to the material - thereby isolating the signal to the cell-material interface and cell-cell contracts - would provide a more physiologically-relevant paradigm for investigating how electrical cues modulate lineage-specific stem cell differentiation. In the present study, we have employed unmodified, directly-stimulated, (un)patterned graphene as a cell culture substrate to investigate how extrinsic electrical cycling influences the differentiation of naïve human mesenchymal stem cells (hMSCs) without the bias of exogenous biochemicals. We first demonstrated that cyclic stimulation does not deteriorate the cell culture media or result in cytotoxic pH, which are critical experiments for correct interpretation of changes in cell behavior. We then measured how the expression of osteogenic and neurogenic lineage-specific markers were altered simply by exposure to electrical stimulation and/or physical patterns. Expression of the early osteogenic transcription factor RUNX2 was increased by electrical stimulation on all graphene substrates, but the mature marker osteopontin was only modulated when stimulation was combined with physical patterns. In contrast, the expression of the neurogenic markers MAP2 and β3-tubulin were enhanced in all electrical stimulation conditions, and were less responsive to the presence of patterns. These data indicate that specific combinations of non-biological inputs - material type, electrical stimulation, physical patterns - can regulate hMSC lineage specification. This study represents a substantial step in understanding how the interplay of

  19. Testicular cell-conditioned medium supports embryonic stem cell differentiation toward germ lineage and to spermatocyte- and oocyte-like cells.

    Science.gov (United States)

    Shah, Syed M; Saini, Neha; Singh, Manoj K; Manik, Radheysham; Singla, Suresh K; Palta, Prabhat; Chauhan, Manmohan S

    2016-08-01

    Testicular cells are believed to secrete various growth factors that activate signaling pathways finally leading to gametogenesis. In vitro gametogenesis is an obscure but paramountly important task primarily because of paucity of the precursor cells and first trimester gonadal tissues. To overcome these limitations for development of in vitro gametes, the present study was designed to induce differentiation of buffalo embryonic stem (ES) cells into germ lineage cells on stimulation by testicular cell-conditioned medium (TCM), on the basis of the assumption that ES cells have the intrinsic property to differentiate into any cell type and TCM would provide the necessary growth factors for differentiation toward germ cell lineage. For this purpose, buffalo ES cells were differentiated as embryoid bodies (EB) in floating cultures and as monolayer adherent cultures in different doses (10%, 20%, and 40%) of TCM for different culture intervals (4, 8, and 14 days), to identify the optimum dose-and-time period. We observed that 40% TCM dose induces highest expression of primordial germ cell-specific (DAZL, VASA, and PLZF), meiotic (SYCP3, MLH1, TNP1/2, and PRM2), spermatocyte-specific (BOULE and TEKT1), and oocyte-specific genes (GDF9 and ZP2/3) for a culture period of 14 days under both floating and adherent differentiation. Immunocytochemical analysis of EBs and adherent cultures revealed presence of primordial germ cell markers (c-KIT, DAZL, and VASA), meiotic markers (SYCP3, MLH1 and PROTAMINE1), spermatocyte markers (ACROSIN and HAPRIN), and oocyte markers (GDF9 and ZP4), indicating progression into post-meiotic gametogenesis. The detection of germ cell-specific proteins in Day 14 EBs like VASA, GDF9, and ZP4 by Western blotting further confirmed germ lineage differentiation. The significantly lower (P propagation, augmentation of reproductive performance in poor breeding buffalo species, and as a model for understanding human germ cell formation. PMID:27056417

  20. Flt3+ macrophage precursors commit sequentially to osteoclasts, dendritic cells and microglia

    Directory of Open Access Journals (Sweden)

    Hanau Daniel

    2002-10-01

    Full Text Available Abstract Background Macrophages, osteoclasts, dendritic cells, and microglia are highly specialized cells that belong to the mononuclear phagocyte system. Functional and phenotypic heterogeneity within the mononuclear phagocyte system may reveal differentiation plasticity of a common progenitor, but developmental pathways leading to such diversity are still unclear. Results Mouse bone marrow cells were expanded in vitro in the presence of Flt3-ligand (FL, yielding high numbers of non-adherent cells exhibiting immature monocyte characteristics. Cells expanded for 6 days, 8 days, or 11 days (day 6-FL, day 8-FL, and day 11-FL cells, respectively exhibited constitutive potential towards macrophage differentiation. In contrast, they showed time-dependent potential towards osteoclast, dendritic, and microglia differentiation that was detected in day 6-, day 8-, and day 11-FL cells, in response to M-CSF and receptor activator of NFκB ligand (RANKL, granulocyte-macrophage colony stimulating-factor (GM-CSF and tumor necrosis factor-α (TNFα, and glial cell-conditioned medium (GCCM, respectively. Analysis of cell proliferation using the vital dye CFSE revealed homogenous growth in FL-stimulated cultures of bone marrow cells, demonstrating that changes in differential potential did not result from sequential outgrowth of specific precursors. Conclusions We propose that macrophages, osteoclasts, dendritic cells, and microglia may arise from expansion of common progenitors undergoing sequential differentiation commitment. This study also emphasizes differentiation plasticity within the mononuclear phagocyte system. Furthermore, selective massive cell production, as shown here, would greatly facilitate investigation of the clinical potential of dendritic cells and microglia.

  1. Lineage-specific interface proteins match up the cell cycle and differentiation in embryo stem cells

    DEFF Research Database (Denmark)

    Re, Angela; Workman, Christopher; Waldron, Levi;

    2014-01-01

    The shortage of molecular information on cell cycle changes along embryonic stem cell (ESC) differentiation prompts an in silico approach, which may provide a novel way to identify candidate genes or mechanisms acting in coordinating the two programs. We analyzed germ layer specific gene expression...... changes during the cell cycle and ESC differentiation by combining four human cell cycle transcriptome profiles with thirteen in vitro human ESC differentiation studies. To detect cross-talk mechanisms we then integrated the transcriptome data that displayed differential regulation with protein...... interaction data. A new class of non-transcriptionally regulated genes was identified, encoding proteins which interact systematically with proteins corresponding to genes regulated during the cell cycle or cell differentiation, and which therefore can be seen as interface proteins coordinating the two...

  2. Cell lineage specific distribution of H3K27 trimethylation accumulation in an in vitro model for human implantation.

    Directory of Open Access Journals (Sweden)

    Gijs Teklenburg

    Full Text Available Female mammals inactivate one of their two X-chromosomes to compensate for the difference in gene-dosage with males that have just one X-chromosome. X-chromosome inactivation is initiated by the expression of the non-coding RNA Xist, which coats the X-chromosome in cis and triggers gene silencing. In early mouse development the paternal X-chromosome is initially inactivated in all cells of cleavage stage embryos (imprinted X-inactivation followed by reactivation of the inactivated paternal X-chromosome exclusively in the epiblast precursors of blastocysts, resulting temporarily in the presence of two active X-chromosomes in this specific lineage. Shortly thereafter, epiblast cells randomly inactivate either the maternal or the paternal X-chromosome. XCI is accompanied by the accumulation of histone 3 lysine 27 trimethylation (H3K27me3 marks on the condensed X-chromosome. It is still poorly understood how XCI is regulated during early human development. Here we have investigated lineage development and the distribution of H3K27me3 foci in human embryos derived from an in-vitro model for human implantation. In this system, embryos are co-cultured on decidualized endometrial stromal cells up to day 8, which allows the culture period to be extended for an additional two days. We demonstrate that after the co-culture period, the inner cell masses have relatively high cell numbers and that the GATA4-positive hypoblast lineage and OCT4-positive epiblast cell lineage in these embryos have segregated. H3K27me3 foci were observed in ∼25% of the trophectoderm cells and in ∼7.5% of the hypoblast cells, but not in epiblast cells. In contrast with day 8 embryos derived from the co-cultures, foci of H3K27me3 were not observed in embryos at day 5 of development derived from regular IVF-cultures. These findings indicate that the dynamics of H3K27me3 accumulation on the X-chromosome in human development is regulated in a lineage specific fashion.

  3. Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells

    Energy Technology Data Exchange (ETDEWEB)

    Sawada, Keigo [Department of Periodontology, Osaka University Graduate School of Dentistry, Osaka (Japan); Takedachi, Masahide, E-mail: takedati@dent.osaka-u.ac.jp [Department of Periodontology, Osaka University Graduate School of Dentistry, Osaka (Japan); Yamamoto, Satomi; Morimoto, Chiaki; Ozasa, Masao; Iwayama, Tomoaki [Department of Periodontology, Osaka University Graduate School of Dentistry, Osaka (Japan); Lee, Chun Man [Medical Center for Translational Research, Osaka University Hospital, Osaka (Japan); Okura, Hanayuki; Matsuyama, Akifumi [Research on Disease Bioresources, Platform of Therapeutics for Rare Disease, National Institute of Biomedical Innovation, Osaka (Japan); Kitamura, Masahiro; Murakami, Shinya [Department of Periodontology, Osaka University Graduate School of Dentistry, Osaka (Japan)

    2015-08-14

    Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response. ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. - Highlights: • ADMPC-derived humoral factors stimulate cytodifferentiation of HPDLs. • ADMPCs secret growth factors including IGFBP6, VEGF and HGF. • IGFBP6 is involved in the promotion effect of ADMPC-CM on HPDL cytodifferentiation.

  4. Trophic factors from adipose tissue-derived multi-lineage progenitor cells promote cytodifferentiation of periodontal ligament cells

    International Nuclear Information System (INIS)

    Stem and progenitor cells are currently being investigated for their applicability in cell-based therapy for periodontal tissue regeneration. We recently demonstrated that the transplantation of adipose tissue-derived multi-lineage progenitor cells (ADMPCs) enhances periodontal tissue regeneration in beagle dogs. However, the molecular mechanisms by which transplanted ADMPCs induce periodontal tissue regeneration remain to be elucidated. In this study, trophic factors released by ADMPCs were examined for their paracrine effects on human periodontal ligament cell (HPDL) function. ADMPC conditioned medium (ADMPC-CM) up-regulated osteoblastic gene expression, alkaline phosphatase activity and calcified nodule formation in HPDLs, but did not significantly affect their proliferative response. ADMPCs secreted a number of growth factors, including insulin-like growth factor binding protein 6 (IGFBP6), hepatocyte growth factor and vascular endothelial growth factor. Among these, IGFBP6 was most highly expressed. Interestingly, the positive effects of ADMPC-CM on HPDL differentiation were significantly suppressed by transfecting ADMPCs with IGFBP6 siRNA. Our results suggest that ADMPCs transplanted into a defect in periodontal tissue release trophic factors that can stimulate the differentiation of HPDLs to mineralized tissue-forming cells, such as osteoblasts and cementoblasts. IGFBP6 may play crucial roles in ADMPC-induced periodontal regeneration. - Highlights: • ADMPC-derived humoral factors stimulate cytodifferentiation of HPDLs. • ADMPCs secret growth factors including IGFBP6, VEGF and HGF. • IGFBP6 is involved in the promotion effect of ADMPC-CM on HPDL cytodifferentiation

  5. Neural induction from ES cells portrays default commitment but instructive maturation.

    Directory of Open Access Journals (Sweden)

    Nibedita Lenka

    Full Text Available The neural induction has remained a debatable issue pertaining to whether it is a mere default process or it involves precise instructive cues. We have chosen the embryonic stem (ES cell model to address this issue. In a devised monoculture strategy, the cell-cell interaction availed through optimum cell plating density could define the niche for the attainment of efficient in vitro neurogenesis from the ES cells. The medium plating density was found ideal in generating optimum number of progenitors and also yielded about 80% mature neurons in a serum free culture set up barring any exogenous inducers. We could also demarcate and quantify the neural stem cells/progenitors among the heterogeneous cell population of differentiating ES cells using nestin intron II driven EGFP expression as a tool. The one week post-plating was determined to be the critical time window for optimum neural progenitor generation from ES cells that helped us further in purifying these cells and in demonstrating their proliferation and multipotent differentiation potential. Seeding cells at varying densities, we could decipher an interesting paradoxical scenario that interlinked both commitment and maturation with the initial plating density having a vital influence on neuronal maturation but not specification and the secretory factors were apparently playing a key role during this process. Thus it was comprehended that, the neural specification was a default process independent of exogenous factors and cellular interaction. Conversely, a defined number of cells at the specification stage itself seemed critical to provide an auto-/paracrine means of signaling threshold for the maturation process to materialize.

  6. Hematopoietic expression of oncogenic BRAF promotes aberrant growth of monocyte-lineage cells resistant to PLX4720

    OpenAIRE

    Kamata, Tamihiro; Dankort, David; Kang, Jing; Giblett, Susan; Pritchard, Catrin A.; McMahon, Martin; Leavitt, Andrew D.

    2013-01-01

    Mutational activation of BRAF leading to expression of the BRAFV600E oncoprotein was recently identified in a high percentage of specific hematopoietic neoplasms in monocyte/histiocyte and mature B-cell lineages. Although BRAFV600E is a driver oncoprotein and pharmacological target in solid tumors such as melanoma, lung and thyroid cancer, it remains unknown whether BRAFV600E is an appropriate therapeutic target in hematopoietic neoplasms. To address this critical question, we generated a mou...

  7. Adipogenic placenta-derived mesenchymal stem cells are not lineage restricted by withdrawing extrinsic factors: developing a novel visual angle in stem cell biology.

    Science.gov (United States)

    Hu, C; Cao, H; Pan, X; Li, J; He, J; Pan, Q; Xin, J; Yu, X; Li, J; Wang, Y; Zhu, D; Li, L

    2016-01-01

    Current evidence implies that differentiated bone marrow mesenchymal stem cells (BMMSCs) can act as progenitor cells and transdifferentiate across lineage boundaries. However, whether this unrestricted lineage has specificities depending on the stem cell type is unknown. Placental-derived mesenchymal stem cells (PDMSCs), an easily accessible and less invasive source, are extremely useful materials in current stem cell therapies. No studies have comprehensively analyzed the transition in morphology, surface antigens, metabolism and multilineage potency of differentiated PDMSCs after their dedifferentiation. In this study, we showed that after withdrawing extrinsic factors, adipogenic PDMSCs reverted to a primitive cell population and retained stem cell characteristics. The mitochondrial network during differentiation and dedifferentiation may serve as a marker of absent or acquired pluripotency in various stem cell models. The new population proliferated faster than unmanipulated PDMSCs and could be differentiated into adipocytes, osteocytes and hepatocytes. The cell adhesion molecules (CAMs) signaling pathway and extracellular matrix (ECM) components modulate cell behavior and enable the cells to proliferate or differentiate during the differentiation, dedifferentiation and redifferentiation processes in our study. These observations indicate that the dedifferentiated PDMSCs are distinguishable from the original PDMSCs and may serve as a novel source in stem cell biology and cell-based therapeutic strategies. Furthermore, whether PDMSCs differentiated into other lineages can be dedifferentiated to a primitive cell population needs to be investigated. PMID:26986509

  8. A monoclonal antibody (8H3) that binds to rat T lineage cells and augments in vitro proliferative responses

    OpenAIRE

    1990-01-01

    A murine monoclonal antibody, designated 8H3, recognizes a cell surface antigen expressed exclusively on rat T lineage cells. 8H3 antibody immunoprecipitated 180-, 120-, and 90-kD components from rat thymocytes as well as splenic T cells under nonreducing conditions. 8H3 antibody specifically inhibited the binding of thymocytes to fibronectin. Furthermore, binding of rat thymocytes to immobilized synthetic peptide Gly-Arg-Gly-Asp-Ser-Pro-Cys-BSA was inhibited by 8H3 antibody as was Gly-Arg-Gl...

  9. In vitro cultured progenitors and precursors of cardiac cell lineages from human normal and post-ischemic hearts

    Directory of Open Access Journals (Sweden)

    F Di Meglio

    2009-08-01

    Full Text Available The demonstration of the presence of dividing primitive cells in damaged hearts has sparked increased interest about myocardium regenerative processes. We examined the rate and the differentiation of in vitro cultured resident cardiac primitive cells obtained from pathological and normal human hearts in order to evaluate the activation of progenitors and precursors of cardiac cell lineages in post-ischemic human hearts. The precursors and progenitors of cardiomyocyte, smooth muscle and endothelial lineage were identified by immunocytochemistry and the expression of characteristic markers was studied by western blot and RT-PCR. The amount of proteins characteristic for cardiac cells (a-SA and MHC, VEGFR-2 and FVIII, SMA for the precursors of cardiomyocytes, endothelial and smooth muscle cells, respectively inclines toward an increase in both a-SA and MHC. The increased levels of FVIII and VEGFR2 are statistically significant, suggesting an important re-activation of neoangiogenesis. At the same time, the augmented expression of mRNA for Nkx 2.5, the trascriptional factor for cardiomyocyte differentiation, confirms the persistence of differentiative processes in terminally injured hearts. Our study would appear to confirm the activation of human heart regeneration potential in pathological conditions and the ability of its primitive cells to maintain their proliferative capability in vitro. The cardiac cell isolation method we used could be useful in the future for studying modifications to the microenvironment that positively influence cardiac primitive cell differentiation or inhibit, or retard, the pathological remodeling and functional degradation of the heart.

  10. High-affinity FRβ-specific CAR T cells eradicate AML and normal myeloid lineage without HSC toxicity.

    Science.gov (United States)

    Lynn, R C; Feng, Y; Schutsky, K; Poussin, M; Kalota, A; Dimitrov, D S; Powell, D J

    2016-06-01

    Acute myeloid leukemia (AML) is an aggressive malignancy, and development of new treatments to prolong remissions is warranted. Chimeric antigen receptor (CAR) T-cell therapies appear promising but on-target, off-tumor recognition of antigen in healthy tissues remains a concern. Here we isolated a high-affinity (HA) folate receptor beta (FRβ)-specific single-chain variable fragment (2.48 nm KD) for optimization of FRβ-redirected CAR T-cell therapy for AML. T cells stably expressing the HA-FRβ CAR exhibited greatly enhanced antitumor activity against FRβ(+) AML in vitro and in vivo compared with a low-affinity FRβ CAR (54.3 nm KD). Using the HA-FRβ immunoglobulin G, FRβ expression was detectable in myeloid-lineage hematopoietic cells; however, expression in CD34(+) hematopoietic stem cells (HSCs) was nearly undetectable. Accordingly, HA-FRβ CAR T cells lysed mature CD14(+) monocytes, while HSC colony formation was unaffected. Because of the potential for elimination of mature myeloid lineage, mRNA CAR electroporation for transient CAR expression was evaluated. mRNA-electroporated HA-FRβ CAR T cells retained effective antitumor activity in vitro and in vivo. Together, our results highlight the importance of antibody affinity in target protein detection and CAR development and suggest that transient delivery of potent HA-FRβ CAR T cells is highly effective against AML and reduces the risk for long-term myeloid toxicity. PMID:26898190

  11. Snai2 and Snai3 transcriptionally regulate cellular fitness and functionality of T cell lineages through distinct gene programs.

    Science.gov (United States)

    Pioli, Peter D; Whiteside, Sarah K; Weis, Janis J; Weis, John H

    2016-05-01

    T lymphocytes are essential contributors to the adaptive immune system and consist of multiple lineages that serve various effector and regulatory roles. As such, precise control of gene expression is essential to the proper development and function of these cells. Previously, we identified Snai2 and Snai3 as being essential regulators of immune tolerance partly due to the impaired function of CD4(+) regulatory T cells in Snai2/3 conditional double knockout mice. Here we extend those previous findings using a bone marrow transplantation model to provide an environmentally unbiased view of the molecular changes imparted onto various T lymphocyte populations once Snai2 and Snai3 are deleted. The data presented here demonstrate that Snai2 and Snai3 transcriptionally regulate the cellular fitness and functionality of not only CD4(+) regulatory T cells but effector CD8(α+) and CD4(+) conventional T cells as well. This is achieved through the modulation of gene sets unique to each cell type and includes transcriptional targets relevant to the survival and function of each T cell lineage. As such, Snai2 and Snai3 are essential regulators of T cell immunobiology. PMID:26831822

  12. Aging alters bone-fat reciprocity by shifting in vivo mesenchymal precursor cell fate towards an adipogenic lineage.

    Science.gov (United States)

    Singh, Lakshman; Brennan, Tracy A; Russell, Elizabeth; Kim, Jung-Hoon; Chen, Qijun; Brad Johnson, F; Pignolo, Robert J

    2016-04-01

    Bone marrow derived mesenchymal progenitor cells (MPCs) play an important role in bone homeostasis. Age-related changes occur in bone resulting in a decrease in bone density and a relative increase in adipocity. Although in vitro studies suggest the existence of an age-related lineage switch between osteogenic and adipogenic fates, stem cell and microenvironmental contributions to this process have not been elucidated in vivo. In order to study the effects of MPC and microenvironmental aging on functional engraftment and lineage switching, transplantation studies were performed under non-myeloablative conditions in old recipients, with donor MPCs derived from young and old green fluorescent protein (GFP) transgenic mice. Robust engraftment by young MPCs or their progeny was observed in the marrow, bone-lining region and in the matrix of young recipients; however, significantly lower engraftment was seen at the same sites in old recipients transplanted with old MPCs. Differentiation of transplanted MPCs strongly favored adipogenesis over osteogenesis in old recipients irrespective of MPC donor age, suggesting that microenvironmental alterations that occur with in vivo aging are predominately responsible for MPC lineage switching. These data indicate that aging alters bone-fat reciprocity and differentiation of mesenchymal progenitors towards an adipogenic fate. PMID:26805026

  13. TGFβ1-Induced Baf60c Regulates both Smooth Muscle Cell Commitment and Quiescence

    Science.gov (United States)

    Sohni, Abhishek; Mulas, Francesca; Ferrazzi, Fulvia; Luttun, Aernout; Bellazzi, Riccardo; Huylebroeck, Danny; Ekker, Stephen C.; Verfaillie, Catherine M.

    2012-01-01

    Smooth muscle cells (SMCs) play critical roles in a number of diseases; however, the molecular mechanism underlying their development is unclear. Although the role of TGFβ1 signaling in SMC development is well established, the downstream molecular signals are not fully understood. We used several rat multipotent adult progenitor cell ((r)MAPC) lines that express levels of Oct4 mRNA similar to hypoblast stem cells (HypoSC), and can differentiate robustly to mesodermal and endodermal cell types. TGFβ1 alone, or with PDGF-BB, induces differentiation of rMAPCs to SMCs, which expressed structural SMC proteins, including α-smooth muscle actin (αSMA), and contribute to the SMC coat of blood vessels in vivo. A genome-wide time-course transcriptome analysis revealed that transcripts of Baf60c, part of the SWI/SNF actin binding chromatin remodeling complex D-3 (SMARCD3/BAF60c), were significantly induced during MAPC-SMC differentiation. We demonstrated that BAF60c is a necessary co-regulator of TGFβ1 mediated induction of SMC genes. Knock-down of Baf60c decreased SMC gene expression in rMAPCs whereas ectopic expression of Baf60c was sufficient to commit rMAPCs to SMCs in the absence of exogenous cytokines. TGFβ1 activates Baf60c via the direct binding of SMAD2/3 complexes to the Baf60c promoter region. Chromatin- and co-immunoprecipitation studies demonstrated that regulation of SMC genes by BAF60c is mediated via interaction with SRF binding CArG box-containing promoter elements in SMC genes. We noted that compared with TGFβ1, Baf60c overexpression in rMAPC yielded SMC with a more immature phenotype. Similarly, Baf60c induced an immature phenotype in rat aortic SMCs marked by increased cell proliferation and decreased contractile marker expression. Thus, Baf60c is important for TGFβ-mediated commitment of primitive stem cells (rMAPCs) to SMCs and is associated with induction of a proliferative state of quiescent SMCs. The MAPC-SMC differentiation system may be

  14. TGFβ1-induced Baf60c regulates both smooth muscle cell commitment and quiescence.

    Directory of Open Access Journals (Sweden)

    Abhishek Sohni

    Full Text Available Smooth muscle cells (SMCs play critical roles in a number of diseases; however, the molecular mechanism underlying their development is unclear. Although the role of TGFβ1 signaling in SMC development is well established, the downstream molecular signals are not fully understood. We used several rat multipotent adult progenitor cell ((rMAPC lines that express levels of Oct4 mRNA similar to hypoblast stem cells (HypoSC, and can differentiate robustly to mesodermal and endodermal cell types. TGFβ1 alone, or with PDGF-BB, induces differentiation of rMAPCs to SMCs, which expressed structural SMC proteins, including α-smooth muscle actin (αSMA, and contribute to the SMC coat of blood vessels in vivo. A genome-wide time-course transcriptome analysis revealed that transcripts of Baf60c, part of the SWI/SNF actin binding chromatin remodeling complex D-3 (SMARCD3/BAF60c, were significantly induced during MAPC-SMC differentiation. We demonstrated that BAF60c is a necessary co-regulator of TGFβ1 mediated induction of SMC genes. Knock-down of Baf60c decreased SMC gene expression in rMAPCs whereas ectopic expression of Baf60c was sufficient to commit rMAPCs to SMCs in the absence of exogenous cytokines. TGFβ1 activates Baf60c via the direct binding of SMAD2/3 complexes to the Baf60c promoter region. Chromatin- and co-immunoprecipitation studies demonstrated that regulation of SMC genes by BAF60c is mediated via interaction with SRF binding CArG box-containing promoter elements in SMC genes. We noted that compared with TGFβ1, Baf60c overexpression in rMAPC yielded SMC with a more immature phenotype. Similarly, Baf60c induced an immature phenotype in rat aortic SMCs marked by increased cell proliferation and decreased contractile marker expression. Thus, Baf60c is important for TGFβ-mediated commitment of primitive stem cells (rMAPCs to SMCs and is associated with induction of a proliferative state of quiescent SMCs. The MAPC-SMC differentiation system

  15. MLL-AF9-mediated immortalization of human hematopoietic cells along different lineages changes during ontogeny

    NARCIS (Netherlands)

    Horton, S J; Jaques, J; Woolthuis, C; van Dijk, J; Mesuraca, M; Huls, G; Morrone, G; Vellenga, E; Schuringa, J J

    2013-01-01

    The MLL-AF9 fusion gene is associated with aggressive leukemias of both the myeloid and lymphoid lineage in infants, whereas in adults, this translocation is mainly associated with acute myeloid leukemia. These observations suggest that differences exist between fetal and adult tissues in terms of t

  16. MLL-AF9-mediated immortalization of human hematopoietic cells along different lineages changes during ontogeny.

    NARCIS (Netherlands)

    Horton, S.J.; Jaques, J.; Woolthuis, C.; Dijk, J. van; Mesuraca, M.; Huls, G.A.; Morrone, G.; Vellenga, E.; Schuringa, J.J.

    2013-01-01

    The MLL-AF9 fusion gene is associated with aggressive leukemias of both the myeloid and lymphoid lineage in infants, whereas in adults, this translocation is mainly associated with acute myeloid leukemia. These observations suggest that differences exist between fetal and adult tissues in terms of t

  17. Enhancement of committed hematopoietic stem cell colony formation by nandrolone decanoate after sublethal whole body irradiation

    International Nuclear Information System (INIS)

    The ability of an anabolic steroid, nandrolone decanoate, to increase committed topoietic stem cell (CFU-gm, CFU-e, and BFU-e) colony formation after sublethal irradiation was evaluated. Immediately after receiving whole body irradiation and on the next two days, each mouse was injected intraperitoneally with nandrolone decanoate (1.25 mg) in propylene glycol. Irradiated control mice received only propylene glycol. Compared to controls, drug-treated mice showed marked peripheral blood leukocytosis and more stable packed red cell volume. Drug-treated mice also demonstrated increased erythropoiesis, as CFU-e/BFU-e concentrations from both marrow (9% to 581%) and spleen (15% to 797%) were elevated. Granulopoiesis was increased similarly, as CFU-gm concentrations from marrow (38% to 685%) and spleen (9% to 373%) were elevated. These results demonstrate that nandrolone decanoate enhances hematopoietic stem cell recovery after sublethal whole body irradiation. This suggests that following hematopoietic suppression, nandrolone decanoate may stimulate the recovery of hematopoiesis at the stem cell level and in peripheral blood

  18. Enhancement of committed hematopoietic stem cell colony formation by nandrolone decanoate after sublethal whole body irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Gallicchio, V.S.; Chen, M.G.; Watts, T.D.

    1984-11-01

    The ability of an anabolic steroid, nandrolone decanoate, to increase committed topoietic stem cell (CFU-gm, CFU-e, and BFU-e) colony formation after sublethal irradiation was evaluated. Immediately after receiving whole body irradiation and on the next two days, each mouse was injected intraperitoneally with nandrolone decanoate (1.25 mg) in propylene glycol. Irradiated control mice received only propylene glycol. Compared to controls, drug-treated mice showed marked peripheral blood leukocytosis and more stable packed red cell volume. Drug-treated mice also demonstrated increased erythropoiesis, as CFU-e/BFU-e concentrations from both marrow (9% to 581%) and spleen (15% to 797%) were elevated. Granulopoiesis was increased similarly, as CFU-gm concentrations from marrow (38% to 685%) and spleen (9% to 373%) were elevated. These results demonstrate that nandrolone decanoate enhances hematopoietic stem cell recovery after sublethal whole body irradiation. This suggests that following hematopoietic suppression, nandrolone decanoate may stimulate the recovery of hematopoiesis at the stem cell level and in peripheral blood.

  19. Human interleukin 7: molecular cloning and growth factor activity on human and murine B-lineage cells.

    OpenAIRE

    Goodwin, R G; Lupton, S; Schmierer, A; Hjerrild, K J; Jerzy, R; Clevenger, W; Gillis, S; Cosman, D; Namen, A E

    1989-01-01

    A cDNA encoding biologically active human interleukin 7 was isolated by hybridization with the homologous murine clone. Nucleotide sequence analysis indicated that this cDNA was capable of encoding a protein of 177 amino acids with a signal sequence of 25 amino acids and a calculated mass of 17.4 kDa for the mature protein. Recombinant human interleukin 7 stimulated the proliferation of murine pre-B cells and was active on cells harvested from human bone marrow that are enriched for B-lineage...

  20. Induction of Functional Hair-Cell-Like Cells from Mouse Cochlear Multipotent Cells

    OpenAIRE

    Quanwen Liu; Yi Shen; Jiarong Chen; Jie Ding; Zihua Tang; Cui Zhang; Jianling Chen; Liang Li; Ping Chen; Jinfu Wang

    2016-01-01

    In this paper, we developed a two-step-induction method of generating functional hair cells from inner ear multipotent cells. Multipotent cells from the inner ear were established and induced initially into progenitor cells committed to the inner ear cell lineage on the poly-L-lysine substratum. Subsequently, the committed progenitor cells were cultured on the mitotically inactivated chicken utricle stromal cells and induced into hair-cell-like cells containing characteristic stereocilia bund...

  1. WASH is required for the differentiation commitment of hematopoietic stem cells in a c-Myc–dependent manner

    OpenAIRE

    Xia, Pengyan; Wang, Shuo; Huang, Guanling; Zhu, Pingping; Li, Man; Ye, Buqing; Du, Ying; Fan, Zusen

    2014-01-01

    Hematopoiesis is fully dependent on hematopoietic stem cells (HSCs) that possess the capacity to self-renew and differentiate into all blood cell lineages. WASH, Wiskott–Aldrich syndrome protein (WASP) and SCAR homologue (WASH) is involved in endosomal sorting as an actin-nucleating protein. Here, we show that conditional WASH deletion in the hematopoietic system causes defective blood production of the host, leading to severe cytopenia and rapid anemia. WASH deficiency causes the accumulatio...

  2. Fas/Fas ligand-mediated apoptosis in different cell lineages and functional compartments of human lymph nodes.

    Science.gov (United States)

    Kokkonen, Tuomo S; Karttunen, Tuomo J

    2010-02-01

    We have optimized an immunohistochemical double-staining method combining immunohistochemical lymphocyte lineage marker detection and apoptosis detection with terminal deoxyribonucleotidyl transferase-mediated dUTP nick end labeling. The method was used to trace Fas-mediated apoptosis in human reactive lymph nodes according to cell lineage and anatomical location. In addition to Fas, we also studied the expression of Fas ligand (FasL), CD3, CD20, CD19, CD23, and CD68 of apoptotic cells. The presence of simultaneous Fas and FasL positivity indicated involvement of activation-induced death in the induction of paracortical apoptosis. FasL expression in the high endothelial venules might be an inductor of apoptosis of Fas-positive lymphoid cells. In addition to B-lymphocyte apoptosis in the germinal centers, there was often a high apoptosis rate of CD23-expressing follicular dendritic cells. In summary, our double-staining method provides valuable new information about the occurrence and mechanisms of apoptosis of different immune cell types in the lymph node compartments. Among other things, we present support for the importance of Fas/FasL-mediated apoptosis in lymph node homeostasis. PMID:19826071

  3. A Distinct Subpopulation of Bone Marrow Mesenchymal Stem Cells, Muse Cells, Directly Commit to the Replacement of Liver Components.

    Science.gov (United States)

    Katagiri, H; Kushida, Y; Nojima, M; Kuroda, Y; Wakao, S; Ishida, K; Endo, F; Kume, K; Takahara, T; Nitta, H; Tsuda, H; Dezawa, M; Nishizuka, S S

    2016-02-01

    Genotyping graft livers by short tandem repeats after human living-donor liver transplantation (n = 20) revealed the presence of recipient or chimeric genotype cases in hepatocytes (6 of 17, 35.3%), sinusoidal cells (18 of 18, 100%), cholangiocytes (15 of 17, 88.2%) and cells in the periportal areas (7 of 8, 87.5%), suggesting extrahepatic cell involvement in liver regeneration. Regarding extrahepatic origin, bone marrow mesenchymal stem cells (BM-MSCs) have been suggested to contribute to liver regeneration but compose a heterogeneous population. We focused on a more specific subpopulation (1-2% of BM-MSCs), called multilineage-differentiating stress-enduring (Muse) cells, for their ability to differentiate into liver-lineage cells and repair tissue. We generated a physical partial hepatectomy model in immunodeficient mice and injected green fluorescent protein (GFP)-labeled human BM-MSC Muse cells intravenously (n = 20). Immunohistochemistry, fluorescence in situ hybridization and species-specific polymerase chain reaction revealed that they integrated into regenerating areas and expressed liver progenitor markers during the early phase and then differentiated spontaneously into major liver components, including hepatocytes (≈74.3% of GFP-positive integrated Muse cells), cholangiocytes (≈17.7%), sinusoidal endothelial cells (≈2.0%), and Kupffer cells (≈6.0%). In contrast, the remaining cells in the BM-MSCs were not detected in the liver for up to 4 weeks. These results suggest that Muse cells are the predominant population of BM-MSCs that are capable of replacing major liver components during liver regeneration. PMID:26663569

  4. Rewiring of human lung cell lineage and mitotic networks in lung adenocarcinomas

    OpenAIRE

    Kim, Il-Jin; Quigley, David; To, Minh D.; Pham, Patrick; Lin, Kevin; Jo, Brian; Jen, Kuang-Yu; Raz, Dan; Kim, Jae; Mao, Jian-Hua; Jablons, David; Balmain, Allan

    2013-01-01

    Analysis of gene expression patterns in normal tissues and their perturbations in tumors can help to identify the functional roles of oncogenes or tumor suppressors and identify potential new therapeutic targets. Here, gene expression correlation networks were derived from 92 normal human lung samples and patient-matched adenocarcinomas. The networks from normal lung show that NKX2-1 is linked to the alveolar type 2 lineage, and identify PEBP4 as a novel marker expressed in alveolar type 2 ce...

  5. Tracking the elusive fibrocyte: identification and characterization of collagen-producing hematopoietic lineage cells during murine wound healing.

    Science.gov (United States)

    Suga, Hirotaka; Rennert, Robert C; Rodrigues, Melanie; Sorkin, Michael; Glotzbach, Jason P; Januszyk, Michael; Fujiwara, Toshihiro; Longaker, Michael T; Gurtner, Geoffrey C

    2014-05-01

    Fibrocytes are a unique population of circulating cells reported to exhibit characteristics of both hematopoietic and mesenchymal cells, and play an important role in wound healing. However, putative fibrocytes have been found to lose expression of hematopoietic surface markers such as CD45 during differentiation, making it difficult to track these cells in vivo with conventional methodologies. In this study, to distinguish hematopoietic and nonhematopoietic cells without surface markers, we took advantage of the gene vav 1, which is expressed solely on hematopoietic cells but not on other cell types, and established a novel transgenic mouse, in which hematopoietic cells are irreversibly labeled with green fluorescent protein and nonhematopoietic cells with red fluorescent protein. Use of single-cell transcriptional analysis in this mouse model revealed two discrete types of collagen I (Col I) expressing cells of hematopoietic lineage recruited into excisional skin wounds. We confirmed this finding on a protein level, with one subset of these Col I synthesizing cells being CD45+ and CD11b+, consistent with the traditional definition of a fibrocyte, while another was CD45- and Cd11b-, representing a previously unidentified population. Both cell types were found to initially peak, then reduce posthealing, consistent with a disappearance from the wound site and not a loss of identifying surface marker expression. Taken together, we have unambiguously identified two cells of hematopoietic origin that are recruited to the wound site and deposit collagen, definitively confirming the existence and natural time course of fibrocytes in cutaneous healing. PMID:24446236

  6. Glioblastoma Stem Cells Respond to Differentiation Cues but Fail to Undergo Commitment and Terminal Cell-Cycle Arrest

    Directory of Open Access Journals (Sweden)

    Helena Carén

    2015-11-01

    Full Text Available Glioblastoma (GBM is an aggressive brain tumor whose growth is driven by stem cell-like cells. BMP signaling triggers cell-cycle exit and differentiation of GBM stem cells (GSCs and, therefore, might have therapeutic value. However, the epigenetic mechanisms that accompany differentiation remain poorly defined. It is also unclear whether cell-cycle arrest is terminal. Here we find only a subset of GSC cultures exhibit astrocyte differentiation in response to BMP. Although overtly differentiated non-cycling astrocytes are generated, they remain vulnerable to cell-cycle re-entry and fail to appropriately reconfigure DNA methylation patterns. Chromatin accessibility mapping identified loci that failed to alter in response to BMP and these were enriched in SOX transcription factor-binding motifs. SOX transcription factors, therefore, may limit differentiation commitment. A similar propensity for cell-cycle re-entry and de-differentiation was observed in GSC-derived oligodendrocyte-like cells. These findings highlight significant obstacles to BMP-induced differentiation as therapy for GBM.

  7. Clinical-scale cultures of cord blood CD34(+) cells to amplify committed progenitors and maintain stem cell activity.

    Science.gov (United States)

    Ivanovic, Zoran; Duchez, Pascale; Chevaleyre, Jean; Vlaski, Marija; Lafarge, Xavier; Dazey, Bernard; Robert-Richard, Elodie; Mazurier, Frédéric; Boiron, Jean-Michel

    2011-01-01

    We developed a clinical-scale cord blood (CB) cell ex vivo procedure to enable an extensive expansion of committed progenitors--colony-forming cells (CFCs) without impairing very primitive hematopoietic stem cells (HSCs). CD34(++) cells, selected from previously cryopreserved and thawed CB units, were cultured in two steps (diluted 1:4 after 6 days) in the presence of stem cell factor (SCF), fms-related tyrosine kinase 3 ligand (Flt-3L), megakaryocyte growth and development factor (MGDF) (100 ng/ml each), granulocyte-colony stimulating factor (G-CSF) (10 ng/ml) in HP01 serum-free medium. HSC activity was evaluated in a serial transplantation assay, by detection of human cells (CD45, CD33, CD19 and CFC of human origin) in bone marrow (BM) of primary and secondary recipient NOD/SCID mice 6-8 weeks after transplantation. A wide amplification of total cells (∼350-fold), CD34(+) cells (∼100-fold), and CFC (∼130-fold) without impairing the HSC activity was obtained. The activity of a particular HSC subpopulation (SRC(CFC)) was even enhanced.Thus, an extensive ex vivo expansion of CFCs is feasible without impairing the activity of HSCs. This result was enabled by associating antioxidant power of medium with an appropriate cytokine cocktail (i.e., mimicking physiologic effects of a weak oxygenation in hematopoietic environment). PMID:21294956

  8. Oct4 is required for lineage priming in the developing inner cell mass of the mouse blastocyst.

    Science.gov (United States)

    Le Bin, Gloryn Chia; Muñoz-Descalzo, Silvia; Kurowski, Agata; Leitch, Harry; Lou, Xinghua; Mansfield, William; Etienne-Dumeau, Charles; Grabole, Nils; Mulas, Carla; Niwa, Hitoshi; Hadjantonakis, Anna-Katerina; Nichols, Jennifer

    2014-03-01

    The transcription factor Oct4 is required in vitro for establishment and maintenance of embryonic stem cells and for reprogramming somatic cells to pluripotency. In vivo, it prevents the ectopic differentiation of early embryos into trophoblast. Here, we further explore the role of Oct4 in blastocyst formation and specification of epiblast versus primitive endoderm lineages using conditional genetic deletion. Experiments involving mouse embryos deficient for both maternal and zygotic Oct4 suggest that it is dispensable for zygote formation, early cleavage and activation of Nanog expression. Nanog protein is significantly elevated in the presumptive inner cell mass of Oct4 null embryos, suggesting an unexpected role for Oct4 in attenuating the level of Nanog, which might be significant for priming differentiation during epiblast maturation. Induced deletion of Oct4 during the morula to blastocyst transition disrupts the ability of inner cell mass cells to adopt lineage-specific identity and acquire the molecular profile characteristic of either epiblast or primitive endoderm. Sox17, a marker of primitive endoderm, is not detected following prolonged culture of such embryos, but can be rescued by provision of exogenous FGF4. Interestingly, functional primitive endoderm can be rescued in Oct4-deficient embryos in embryonic stem cell complementation assays, but only if the host embryos are at the pre-blastocyst stage. We conclude that cell fate decisions within the inner cell mass are dependent upon Oct4 and that Oct4 is not cell-autonomously required for the differentiation of primitive endoderm derivatives, as long as an appropriate developmental environment is established. PMID:24504341

  9. High affinity FRβ-specific CAR T cells eradicate AML and normal yeloid lineage without HSC toxicity

    Science.gov (United States)

    Lynn, Rachel C; Feng, Yang; Schutsky, Keith; Poussin, Mathilde; Kalota, Anna; Dimitrov, Dimiter S; Powell, Daniel J

    2016-01-01

    Acute myeloid leukemia (AML) is an aggressive malignancy, and development of new treatments to prolong remissions is warranted. Chimeric antigen receptor (CAR) T-cell therapies appear promising but on-target, off-tumor recognition of antigen in healthy tissues remains a concern. Here, we isolated a high affinity (HA) folate receptor beta (FRβ)-specific scFv (2.48nM KD) for optimization of FRβ-redirected CAR T-cell therapy for AML. T-cells stably expressing the HA-FRβ CAR exhibited greatly enhanced antitumor activity against FRβ+ AML in vitro and in vivo compared to a low affinity (LA) FRβ CAR (54.3nM KD). Using the HA-FRβ IgG, FRβ expression was detectable in myeloid-lineage hematopoietic cells; however, expression in CD34+ hematopoietic stem cells (HSCs) was nearly undetectable. Accordingly, HA-FRβ CAR T-cells lysed mature CD14+ monocytes, while HSC colony formation was unaffected. Because of the potential for elimination of mature myeloid lineage, mRNA CAR electroporation for transient CAR expression was evaluated. mRNA-electroporated HA-FRβ CAR T-cells retained effective anti-tumor activity in vitro and in vivo. Together, our results highlight the importance of antibody affinity in target protein detection and CAR development and suggest that transient delivery of potent HA-FRβ CAR T-cells is highly effective against AML and reduces the risk for long-term myeloid toxicity. PMID:26898190

  10. Prohibitin 2 regulates the proliferation and lineage-specific differentiation of mouse embryonic stem cells in mitochondria.

    Directory of Open Access Journals (Sweden)

    Megumi Kowno

    Full Text Available BACKGROUND: The pluripotent state of embryonic stem (ES cells is controlled by a network of specific transcription factors. Recent studies also suggested the significant contribution of mitochondria on the regulation of pluripotent stem cells. However, the molecules involved in these regulations are still unknown. METHODOLOGY/PRINCIPAL FINDINGS: In this study, we found that prohibitin 2 (PHB2, a pleiotrophic factor mainly localized in mitochondria, is a crucial regulatory factor for the homeostasis and differentiation of ES cells. PHB2 was highly expressed in undifferentiated mouse ES cells, and the expression was decreased during the differentiation of ES cells. Knockdown of PHB2 induced significant apoptosis in pluripotent ES cells, whereas enhanced expression of PHB2 contributed to the proliferation of ES cells. However, enhanced expression of PHB2 strongly inhibited ES cell differentiation into neuronal and endodermal cells. Interestingly, only PHB2 with intact mitochondrial targeting signal showed these specific effects on ES cells. Moreover, overexpression of PHB2 enhanced the processing of a dynamin-like GTPase (OPA1 that regulates mitochondrial fusion and cristae remodeling, which could induce partial dysfunction of mitochondria. CONCLUSIONS/SIGNIFICANCE: Our results suggest that PHB2 is a crucial mitochondrial regulator for homeostasis and lineage-specific differentiation of ES cells.

  11. Identification of vascular lineage-specific genes by transcriptional profiling of isolated blood vascular and lymphatic endothelial cells.

    Science.gov (United States)

    Hirakawa, Satoshi; Hong, Young-Kwon; Harvey, Natasha; Schacht, Vivien; Matsuda, Kant; Libermann, Towia; Detmar, Michael

    2003-02-01

    In mammals, the lymphatic vascular system develops by budding of lymphatic progenitor endothelial cells from embryonic veins to form a distinct network of draining vessels with important functions in the immune response and in cancer metastasis. However, the lineage-specific molecular characteristics of blood vascular versus lymphatic endothelium have remained poorly defined. We isolated lymphatic endothelial cells (LECs) and blood vascular endothelial cells (BVECs) by immunomagnetic isolation directly from human skin. Cultured LECs but not BVECs expressed the lymphatic markers Prox1 and LYVE-1 and formed LYVE-1-positive vascular tubes after implantation in vivo. Transcriptional profiling studies revealed increased expression of several extracellular matrix and adhesion molecules in BVECs, including versican, collagens, laminin, and N-cadherin, and of the growth factor receptors endoglin and vascular endothelial growth factor receptor-1/Flt-1. Differential immunostains of human skin confirmed the blood vessel-specific expression of these genes. During embryonic development, endoglin expression was gradually down-regulated on lymphatic endothelium whereas vascular endothelial growth factor receptor-1 was absent from lymphatics. We also identified several genes with specific expression in LECs. These results demonstrate that some lineage-specific genes are only expressed during distinct developmental stages and they identify new molecular markers for blood vascular and lymphatic endothelium with important implications for future studies of vascular development and function. PMID:12547715

  12. Diagnosis of a T-lineage acute lymphoblastic leukemia through digitalized cell analysis of the pleural effusion

    Directory of Open Access Journals (Sweden)

    Peruzzi B

    2013-11-01

    Full Text Available Benedetta Peruzzi,1 Ilaria Cutini,2 Anna Maria Grazia Gelli,1 Tommaso Rondelli,1 Marinella Statello,1 Sara Bencini,2 Francesco Mannelli,2 Roberto Caporale,1 Alberto Bosi,2 Alessandra Fanelli1 1General Laboratory Unit (Microscopy and Clinical Cytometry Unit, 2Hematology Unit, Azienda Ospedaliero–Universitaria Careggi, Firenze, Italy Introduction: Pleural effusion as the first clinical manifestation of acute lymphoblastic leukemia (ALL is a relatively rare event. An early and accurate diagnosis of this clinical picture is very important for adequate patient management. Case presentation: We report the atypical onset of T-lineage ALL in a 31-year-old man. The patient was admitted to the emergency room due to lung failure; at that moment, the patient's initial blood count was normal; the chest X-ray radiography showed a massive pleural effusion and a thoracentesis was carried out. Routine investigations performed on the pleural fluid using a new technology system and digitalized cell analysis demonstrated infiltration by immature cells. Therefore, bone marrow aspirate and flow cytometry analyses were performed, leading to the diagnosis of T-lineage ALL. A cord blood transplantation procedure was performed at the first hematological remission following chemotherapy regimens. The patient died of septic shock. Conclusion: The case we reported underlines the usefulness of using automated instruments to identify abnormal lymphoid cells in body fluids. Keywords: pleural effusion, digital morphology, leukemia

  13. Donor-matched mesenchymal stem cells from knee infrapatellar and subcutaneous adipose tissue of osteoarthritic donors display differential chondrogenic and osteogenic commitment

    Directory of Open Access Journals (Sweden)

    S Lopa

    2014-04-01

    Full Text Available Cell-based therapies have recently been proposed for the treatment of degenerative articular pathologies, such as early osteoarthritis, with an emphasis on autologous mesenchymal stem cells (MSCs, as an alternative to terminally differentiated cells. In this study, we performed a donor-matched comparison between infrapatellar fat pad MSCs (IFP-MSCs and knee subcutaneous adipose tissue stem cells (ASCs, as appealing candidates for cell-based therapies that are easily accessible during surgery. IFP-MSCs and ASCs were obtained from 25 osteoarthritic patients undergoing total knee replacement and compared for their immunophenotype and differentiative potential. Undifferentiated IFP-MSCs and ASCs displayed the same immunophenotype, typical of MSCs (CD13+/CD29+/CD44+/CD73+/CD90+/CD105+/CD166+/CD31-/CD45-. IFP-MSCs and ASCs showed similar adipogenic potential, though undifferentiated ASCs had higher LEP expression compared to IFP-MSCs (p < 0.01. Higher levels of calcified matrix (p < 0.05 and alkaline phosphatase (p < 0.05 in ASCs highlighted their superior osteogenic commitment compared to IFP-MSCs. Conversely, IFP-MSCs pellets showed greater amounts of glycosaminoglycans (p < 0.01 and superior expression of ACAN (p < 0.001, SOX9, COMP (p < 0.001 and COL2A1 (p < 0.05 compared to ASCs pellets, revealing a superior chondrogenic potential. This was also supported by lower COL10A1 (p < 0.05 and COL1A1 (p < 0.01 expression and lower alkaline phosphatase release (p < 0.05 by IFP-MSCs compared to ASCs. The observed dissimilarities between IFP-MSCs and ASCs show that, despite expressing similar surface markers, MSCs deriving from different fat depots in the same surgical site possess specific features. Furthermore, the in vitro peculiar commitment of IFP-MSCs and ASCs from osteoarthritic donors towards the chondrogenic or osteogenic lineage may suggest a preferential use for cartilage and bone cell-based treatments, respectively.

  14. GATA-3 promotes T-cell specification by repressing B-cell potential in pro-T cells in mice

    NARCIS (Netherlands)

    M.E. García (Marcos); R.G.J. Klein Wolterink (Roel); F. Lemâitre (Fabrice); C. Le Goff (Carine); M. Hasan (Milena); R.W. Hendriks (Rudi); A. Cumano (Ana); J.P. di Santo (James)

    2013-01-01

    textabstractTranscription factors orchestrate T-lineage differentiation in the thymus. One critical checkpoint involves Notch1 signaling that instructs T-cell commitment at the expense of the B-lineage program. While GATA-3 is required for T-cell specification, its mechanism of action is poorly unde

  15. PROP1 overexpression in corticotrophinomas: evidence for the role of PROP1 in the maintenance of cells committed to corticotrophic differentiation

    Directory of Open Access Journals (Sweden)

    Ricardo V. Araujo

    2013-06-01

    Full Text Available OBJECTIVE: The expression of transcription factors involved in early pituitary development, such as PROP1 and POU1F1, has been detected in pituitary adenoma tissues. In this study, we sought to characterize the transcriptional profiles of PROP1, POU1F1, and TBX19 in functioning and nonfunctioning pituitary adenomas in an attempt to identify their roles in tumorigenesis and hormone hypersecretion. METHODS: RT-qPCR analyses were performed to assess the transcriptional pattern of PROP1, POU1F1, TBX19, and hormone-producing genes in tissue samples of corticotrophinomas (n = 10, somatotrophinomas (n = 8, and nonfunctioning adenomas (n = 6. RESULTS: Compared with normal pituitary tissue, POU1F1 was overexpressed in somatotrophinomas by 3-fold. PROP1 expression was 18-fold higher in corticotrophinomas, 10-fold higher in somatotrophinomas, and 3-fold higher in nonfunctioning adenomas. TBX19 expression was 27-fold higher in corticotrophinomas. Additionally, the level of TBX19 mRNA positively correlated with that of pro-opiomelanocortin (r = 0.49, p = 0.014. CONCLUSIONS: Our data demonstrate that PROP1 is overexpressed in pituitary adenomas, mainly in corticotrophinomas. Together with previously published data showing that patients who harbor PROP1 loss-of-function mutations present a progressive decline in corticotrope function, our results support a role for PROP1 in pituitary tumor development and in the maintenance of cell lineages committed to corticotrophic differentiation.

  16. Single-Cell RNA-Sequencing Reveals a Continuous Spectrum of Differentiation in Hematopoietic Cells

    Directory of Open Access Journals (Sweden)

    Iain C. Macaulay

    2016-02-01

    Full Text Available The transcriptional programs that govern hematopoiesis have been investigated primarily by population-level analysis of hematopoietic stem and progenitor cells, which cannot reveal the continuous nature of the differentiation process. Here we applied single-cell RNA-sequencing to a population of hematopoietic cells in zebrafish as they undergo thrombocyte lineage commitment. By reconstructing their developmental chronology computationally, we were able to place each cell along a continuum from stem cell to mature cell, refining the traditional lineage tree. The progression of cells along this continuum is characterized by a highly coordinated transcriptional program, displaying simultaneous suppression of genes involved in cell proliferation and ribosomal biogenesis as the expression of lineage specific genes increases. Within this program, there is substantial heterogeneity in the expression of the key lineage regulators. Overall, the total number of genes expressed, as well as the total mRNA content of the cell, decreases as the cells undergo lineage commitment.

  17. The Wnt receptor, Lrp5, is expressed by mouse mammary stem cells and is required to maintain the basal lineage.

    Directory of Open Access Journals (Sweden)

    Nisha M Badders

    Full Text Available BACKGROUND: Ectopic Wnt signaling induces increased stem/progenitor cell activity in the mouse mammary gland, followed by tumor development. The Wnt signaling receptors, Lrp5/6, are uniquely required for canonical Wnt activity. Previous data has shown that the absence of Lrp5 confers resistance to Wnt1-induced tumor development. METHODOLOGY/PRINCIPAL FINDINGS: Here, we show that all basal mammary cells express Lrp5, and co-express Lrp6 in a similar fashion. Though Wnt dependent transcription of key target genes is relatively unchanged in mammary epithelial cell cultures, the absence of Lrp5 specifically depletes adult regenerative stem cell activity (to less than 1%. Stem cell activity can be enriched by >200 fold (over 80% of activity, based on high Lrp5 expression alone. Though Lrp5 null glands have apparent normal function, the basal lineage is relatively reduced (from 42% basal/total epithelial cells to 22% and Lrp5-/- mammary epithelial cells show enhanced expression of senescence-associated markers in vitro, as measured by expression of p16(Ink4a and TA-p63. CONCLUSIONS/SIGNIFICANCE: This is the first single biomarker that has been demonstrated to be functionally involved in stem cell maintenance. Together, these results demonstrate that Wnt signaling through Lrp5 is an important component of normal mammary stem cell function.

  18. Stem cells and lineages of the intestine: a developmental and evolutionary perspective

    OpenAIRE

    Takashima, Shigeo; Gold, David; Hartenstein, Volker

    2012-01-01

    The intestine consists of epithelial cells that secrete digestive enzymes and mucus (gland cells), absorb food particles (enterocytes), and produce hormones (endocrine cells). Intestinal cells are rapidly turned over and need to be replaced. In cnidarians, mitosis of differentiated intestinal cells accounts for much of the replacement; in addition, migratory, multipotent stem cells (interstitial cells) contribute to the production of intestinal cells. In other phyla, intestinal cell replaceme...

  19. All-trans retinoic acid promotes neural lineage entry by pluripotent embryonic stem cells via multiple pathways

    Directory of Open Access Journals (Sweden)

    Fang Bo

    2009-07-01

    Full Text Available Abstract Background All-trans retinoic acid (RA is one of the most important morphogens with pleiotropic actions. Its embryonic distribution correlates with neural differentiation in the developing central nervous system. To explore the precise effects of RA on neural differentiation of mouse embryonic stem cells (ESCs, we detected expression of RA nuclear receptors and RA-metabolizing enzymes in mouse ESCs and investigated the roles of RA in adherent monolayer culture. Results Upon addition of RA, cell differentiation was directed rapidly and exclusively into the neural lineage. Conversely, pharmacological interference with RA signaling suppressed this neural differentiation. Inhibition of fibroblast growth factor (FGF signaling did not suppress significantly neural differentiation in RA-treated cultures. Pharmacological interference with extracellular signal-regulated kinase (ERK pathway or activation of Wnt pathway effectively blocked the RA-promoted neural specification. ERK phosphorylation was enhanced in RA-treated cultures at the early stage of differentiation. Conclusion RA can promote neural lineage entry by ESCs in adherent monolayer culture systems. This effect depends on RA signaling and its crosstalk with the ERK and Wnt pathways.

  20. Differentiation of human gingival mesenchymal stem cells into neuronal lineages in 3D bioconjugated injectable protein hydrogel construct for the management of neuronal disorder.

    Science.gov (United States)

    Rao, Suresh Ranga; Subbarayan, Rajasekaran; Dinesh, Murugan Girija; Arumugam, Gnanamani; Raja, Selvaraj Thirupathi Kumara

    2016-01-01

    The success of regeneration attempt is based on an ideal combination of stem cells, scaffolding and growth factors. Tissue constructs help to maintain stem cells in a required area for a desired time. There is a need for easily obtainable cells, potentially autologous stem cells and a biologically acceptable scaffold for use in humans in different difficult situations. This study aims to address these issues utilizing a unique combination of stem cells from gingiva and a hydrogel scaffold, based on a natural product for regenerative application. Human gingival mesenchymal stem cells (HGMSCs) were, with due induction, differentiated to neuronal lineages to overcome the problems associated with birth tissue-related stem cells. The differentiation potential of neuronal lineages was confirmed with suitable specific markers. The properties of mesenchymal stem cells in encapsulated form were observed to be similar to free cells. The encapsulated cells (3D) were then subjected to differentiation into neuronal lineages with suitable inducers, and the morphology and gene expression of transient cells were analyzed. HGMSCs was differentiated into neuronal lineages as both free and encapsulated forms without any significant differences. The presence of Nissl bodies and the neurite outgrowth confirm the differentiation. The advantages of this new combination appear to make it a promising tissue construct for translational application. PMID:26869025

  1. Differentiation of human gingival mesenchymal stem cells into neuronal lineages in 3D bioconjugated injectable protein hydrogel construct for the management of neuronal disorder

    Science.gov (United States)

    Rao, Suresh Ranga; Subbarayan, Rajasekaran; Dinesh, Murugan Girija; Arumugam, Gnanamani; Raja, Selvaraj Thirupathi Kumara

    2016-01-01

    The success of regeneration attempt is based on an ideal combination of stem cells, scaffolding and growth factors. Tissue constructs help to maintain stem cells in a required area for a desired time. There is a need for easily obtainable cells, potentially autologous stem cells and a biologically acceptable scaffold for use in humans in different difficult situations. This study aims to address these issues utilizing a unique combination of stem cells from gingiva and a hydrogel scaffold, based on a natural product for regenerative application. Human gingival mesenchymal stem cells (HGMSCs) were, with due induction, differentiated to neuronal lineages to overcome the problems associated with birth tissue-related stem cells. The differentiation potential of neuronal lineages was confirmed with suitable specific markers. The properties of mesenchymal stem cells in encapsulated form were observed to be similar to free cells. The encapsulated cells (3D) were then subjected to differentiation into neuronal lineages with suitable inducers, and the morphology and gene expression of transient cells were analyzed. HGMSCs was differentiated into neuronal lineages as both free and encapsulated forms without any significant differences. The presence of Nissl bodies and the neurite outgrowth confirm the differentiation. The advantages of this new combination appear to make it a promising tissue construct for translational application. PMID:26869025

  2. Differential expression of genes involved in the epigenetic regulation of cell identity in normal human mammary cell commitment and differentiation

    Directory of Open Access Journals (Sweden)

    Danila Coradini

    2014-10-01

    Full Text Available The establishment and maintenance of mammary epithelial cell identity depends on the activity of a group of proteins, collectively called maintenance proteins, that act as epigenetic regulators of gene transcription through DNA methylation, histone modification, and chromatin remodeling. Increasing evidence indicates that dysregulation of these crucial proteins may disrupt epithelial cell integrity and trigger breast tumor initiation. Therefore, we explored in silico the expression pattern of a panel of 369 genes known to be involved in the establishment and maintenance of epithelial cell identity and mammary gland remodeling in cell subpopulations isolated from normal human mammary tissue and selectively enriched in their content of bipotent progenitors, committed luminal progenitors, and differentiated myoepithelial or differentiated luminal cells. The results indicated that, compared to bipotent cells, differentiated myoepithelial and luminal subpopulations were both characterized by the differential expression of 4 genes involved in cell identity maintenance: CBX6 and PCGF2, encoding proteins belonging to the Polycomb group, and SMARCD3 and SMARCE1, encoding proteins belonging to the Trithorax group. In addition to these common genes, the myoepithelial phenotype was associated with the differential expression of HDAC1, which encodes histone deacetylase 1, whereas the luminal phenotype was associated with the differential expression of SMARCA4 and HAT1, which encode a Trithorax protein and histone acetylase 1, respectively. The luminal compartment was further characterized by the overexpression of ALDH1A3 and GATA3, and the down-regulation of NOTCH4 and CCNB1, with the latter suggesting a block in cell cycle progression at the G2 phase. In contrast, myoepithelial differentiation was associated with the overexpression of MYC and the down-regulation of CCNE1, with the latter suggesting a block in cell cycle progression at the G1 phase.

  3. Differentiation of retinal ganglion cells and photoreceptor precursors from mouse induced pluripotent stem cells carrying an Atoh7/Math5 lineage reporter.

    Directory of Open Access Journals (Sweden)

    Bin-Bin Xie

    Full Text Available The neural retina is a critical component of the visual system, which provides the majority of sensory input in humans. Various retinal degenerative diseases can result in the permanent loss of retinal neurons, especially the light-sensing photoreceptors and the centrally projecting retinal ganglion cells (RGCs. The replenishment of lost RGCs and the repair of optic nerve damage are particularly challenging, as both RGC specification and their subsequent axonal growth and projection involve complex and precise regulation. To explore the developmental potential of pluripotent stem cell-derived neural progenitors, we have established mouse iPS cells that allow cell lineage tracing of progenitors that have expressed Atoh7/Math5, a bHLH transcription factor required for RGC production. These Atoh7 lineage reporter iPS cells encode Cre to replace one copy of the endogenous Atoh7 gene and a Cre-dependent YFP reporter in the ROSA locus. In addition, they express pluripotent markers and are capable of generating teratomas in vivo. Under anterior neural induction and neurogenic conditions in vitro, the Atoh7-Cre/ROSA-YFP iPS cells differentiate into neurons that co-express various RGC markers and YFP, indicating that these neurons are derived from Atoh7-expressing progenitors. Consistent with previous in vivo cell lineage studies, the Atoh7-Cre/ROSA-YFP iPS cells also give rise to a subset of Crx-positive photoreceptor precursors. Furthermore, inhibition of Notch signaling in the iPSC cultures results in a significant increase of YFP-positive RGCs and photoreceptor precursors. Together, these results show that Atoh7-Cre/ROSA-YFP iPS cells can be used to monitor the development and survival of RGCs and photoreceptors from pluripotent stem cells.

  4. Arx and Nkx2.2 compound deficiency redirects pancreatic alpha- and beta-cell differentiation to a somatostatin/ghrelin co-expressing cell lineage

    Directory of Open Access Journals (Sweden)

    Mansouri Ahmed

    2011-08-01

    Full Text Available Abstract Background Nkx2.2 and Arx represent key transcription factors implicated in the specification of islet cell subtypes during pancreas development. Mice deficient for Arx do not develop any alpha-cells whereas beta- and delta-cells are found in considerably higher numbers. In Nkx2.2 mutant animals, alpha- and beta-cell development is severely impaired whereas a ghrelin-expressing cell population is found augmented. Notably, Arx transcription is clearly enhanced in Nkx2.2-deficient pancreata. Hence in order to precise the functional link between both factors we performed a comparative analysis of Nkx2.2/Arx single- and double-mutants but also of Pax6-deficient animals. Results We show that most of the ghrelin+ cells emerging in pancreata of Nkx2.2- and Pax6-deficient mice, express the alpha-cell specifier Arx, but also additional beta-cell related genes. In Nkx2.2-deficient mice, Arx directly co-localizes with iAPP, PC1/3 and Pdx1 suggesting an Nkx2.2-dependent control of Arx in committed beta-cells. The combined loss of Nkx2.2 and Arx likewise results in the formation of a hyperplastic ghrelin+ cell population at the expense of mature alpha- and beta-cells. Surprisingly, such Nkx2.2-/-Arx- ghrelin+ cells also express the somatostatin hormone. Conclusions Our data indicate that Nkx2.2 acts by reinforcing the transcriptional networks initiated by Pax4 and Arx in early committed beta- and alpha-cell, respectively. Our analysis also suggests that one of the coupled functions of Nkx2.2 and Pax4 is to counteract Arx gene activity in early committed beta-cells.

  5. EGF–FGF2 stimulates the proliferation and improves the neuronal commitment of mouse epidermal neural crest stem cells (EPI-NCSCs)

    International Nuclear Information System (INIS)

    Epidermal neural crest stem cells (EPI-NCSCs), which reside in the bulge of hair follicles, are attractive candidates for several applications in cell therapy, drug screening and tissue engineering. As suggested remnants of the embryonic neural crest (NC) in an adult location, EPI-NCSCs are able to generate a wide variety of cell types and are readily accessible by a minimally invasive procedure. Since the combination of epidermal growth factor (EGF) and fibroblast growth factor type 2 (FGF2) is mitogenic and promotes the neuronal commitment of various stem cell populations, we examined its effects in the proliferation and neuronal potential of mouse EPI-NCSCs. By using a recognized culture protocol of bulge whiskers follicles, we were able to isolate a population of EPI-NCSCs, characterized by the migratory potential, cell morphology and expression of phenotypic markers of NC cells. EPI-NCSCs expressed neuronal, glial and smooth muscle markers and exhibited the NC-like fibroblastic morphology. The treatment with the combination EGF and FGF2, however, increased their proliferation rate and promoted the acquisition of a neuronal-like morphology accompanied by reorganization of neural cytoskeletal proteins βIII-tubulin and nestin, as well as upregulation of the pan neuronal marker βIII-tubulin and down regulation of the undifferentiated NC, glial and smooth muscle cell markers. Moreover, the treatment enhanced the response of EPI-NCSCs to neurogenic stimulation, as evidenced by induction of GAP43, and increased expression of Mash-1 in neuron-like cell, both neuronal-specific proteins. Together, the results suggest that the combination of EGF–FGF2 stimulates the proliferation and improves the neuronal potential of EPI-NCSCs similarly to embryonic NC cells, ES cells and neural progenitor/stem cells of the central nervous system and highlights the advantage of using EGF–FGF2 in neuronal differentiation protocols. - Highlights: • EPI-NCSCs express

  6. EGF–FGF{sub 2} stimulates the proliferation and improves the neuronal commitment of mouse epidermal neural crest stem cells (EPI-NCSCs)

    Energy Technology Data Exchange (ETDEWEB)

    Bressan, Raul Bardini; Melo, Fernanda Rosene; Almeida, Patricia Alves; Bittencourt, Denise Avani; Visoni, Silvia; Jeremias, Talita Silva [Departamento de Biologia Celular, Embriologia e Genética, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Campus Universitário – Trindade, 88040-900 Florianópolis SC (Brazil); Costa, Ana Paula; Leal, Rodrigo Bainy [Departamento de Bioquímica, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Campus Universitário – Trindade, 88040-900 Florianópolis SC (Brazil); Trentin, Andrea Gonçalves, E-mail: andrea.trentin@ufsc.br [Departamento de Biologia Celular, Embriologia e Genética, Centro de Ciências Biológicas, Universidade Federal de Santa Catarina, Campus Universitário – Trindade, 88040-900 Florianópolis SC (Brazil)

    2014-09-10

    Epidermal neural crest stem cells (EPI-NCSCs), which reside in the bulge of hair follicles, are attractive candidates for several applications in cell therapy, drug screening and tissue engineering. As suggested remnants of the embryonic neural crest (NC) in an adult location, EPI-NCSCs are able to generate a wide variety of cell types and are readily accessible by a minimally invasive procedure. Since the combination of epidermal growth factor (EGF) and fibroblast growth factor type 2 (FGF{sub 2}) is mitogenic and promotes the neuronal commitment of various stem cell populations, we examined its effects in the proliferation and neuronal potential of mouse EPI-NCSCs. By using a recognized culture protocol of bulge whiskers follicles, we were able to isolate a population of EPI-NCSCs, characterized by the migratory potential, cell morphology and expression of phenotypic markers of NC cells. EPI-NCSCs expressed neuronal, glial and smooth muscle markers and exhibited the NC-like fibroblastic morphology. The treatment with the combination EGF and FGF{sub 2}, however, increased their proliferation rate and promoted the acquisition of a neuronal-like morphology accompanied by reorganization of neural cytoskeletal proteins βIII-tubulin and nestin, as well as upregulation of the pan neuronal marker βIII-tubulin and down regulation of the undifferentiated NC, glial and smooth muscle cell markers. Moreover, the treatment enhanced the response of EPI-NCSCs to neurogenic stimulation, as evidenced by induction of GAP43, and increased expression of Mash-1 in neuron-like cell, both neuronal-specific proteins. Together, the results suggest that the combination of EGF–FGF2 stimulates the proliferation and improves the neuronal potential of EPI-NCSCs similarly to embryonic NC cells, ES cells and neural progenitor/stem cells of the central nervous system and highlights the advantage of using EGF–FGF{sub 2} in neuronal differentiation protocols. - Highlights: • EPI

  7. HIV Skews the Lineage-Defining Transcriptional Profile of Mycobacterium tuberculosis-Specific CD4+ T Cells.

    Science.gov (United States)

    Riou, Catherine; Strickland, Natalie; Soares, Andreia P; Corleis, Björn; Kwon, Douglas S; Wherry, E John; Wilkinson, Robert J; Burgers, Wendy A

    2016-04-01

    HIV-infected persons are at greater risk of developing tuberculosis (TB) even before profound CD4 loss occurs, suggesting that HIV alters CD4(+) T cell functions capable of containing bacterial replication. An effective immune response to Mycobacterium tuberculosis most likely relies on the development of a balanced CD4 response, in which distinct CD4(+) Th subsets act in synergy to control the infection. To define the diversity of M. tuberculosis-specific CD4(+) Th subsets and determine whether HIV infection impacts such responses, the expression of lineage-defining transcription factors T-bet, Gata3, RORγt, and Foxp3 was measured in M. tuberculosis-specific CD4(+) T cells in HIV-uninfected (n = 20) and HIV-infected individuals (n = 20) with latent TB infection. Our results show that, upon 5-d restimulation in vitro, M. tuberculosis-specific CD4(+) T cells from healthy individuals have the ability to exhibit a broad spectrum of Th subsets, defined by specific patterns of transcription factor coexpression. These transcription factor profiles were skewed in HIV-infected individuals where the proportion of T-bet(high)Foxp3(+) M. tuberculosis-specific CD4(+) T cells was significantly decreased (p = 0.002) compared with HIV-uninfected individuals, a change that correlated inversely with HIV viral load (p = 0.0007) and plasma TNF-α (p = 0.027). Our data demonstrate an important balance in Th subset diversity defined by lineage-defining transcription factor coexpression profiles that is disrupted by HIV infection and suggest a role for HIV in impairing TB immunity by altering the equilibrium of M. tuberculosis-specific CD4(+) Th subsets. PMID:26927799

  8. Integrative genomic analyses identify BRF2 as a novel lineage-specific oncogene in lung squamous cell carcinoma.

    Directory of Open Access Journals (Sweden)

    William W Lockwood

    2010-07-01

    Full Text Available BACKGROUND: Traditionally, non-small cell lung cancer is treated as a single disease entity in terms of systemic therapy. Emerging evidence suggests the major subtypes--adenocarcinoma (AC and squamous cell carcinoma (SqCC--respond differently to therapy. Identification of the molecular differences between these tumor types will have a significant impact in designing novel therapies that can improve the treatment outcome. METHODS AND FINDINGS: We used an integrative genomics approach, combing high-resolution comparative genomic hybridization and gene expression microarray profiles, to compare AC and SqCC tumors in order to uncover alterations at the DNA level, with corresponding gene transcription changes, which are selected for during development of lung cancer subtypes. Through the analysis of multiple independent cohorts of clinical tumor samples (>330, normal lung tissues and bronchial epithelial cells obtained by bronchial brushing in smokers without lung cancer, we identified the overexpression of BRF2, a gene on Chromosome 8p12, which is specific for development of SqCC of lung. Genetic activation of BRF2, which encodes a RNA polymerase III (Pol III transcription initiation factor, was found to be associated with increased expression of small nuclear RNAs (snRNAs that are involved in processes essential for cell growth, such as RNA splicing. Ectopic expression of BRF2 in human bronchial epithelial cells induced a transformed phenotype and demonstrates downstream oncogenic effects, whereas RNA interference (RNAi-mediated knockdown suppressed growth and colony formation of SqCC cells overexpressing BRF2, but not AC cells. Frequent activation of BRF2 in >35% preinvasive bronchial carcinoma in situ, as well as in dysplastic lesions, provides evidence that BRF2 expression is an early event in cancer development of this cell lineage. CONCLUSIONS: This is the first study, to our knowledge, to show that the focal amplification of a gene in

  9. PDX1- and NGN3-mediated in vitro reprogramming of human bone marrow-derived mesenchymal stromal cells into pancreatic endocrine lineages

    DEFF Research Database (Denmark)

    Limbert, Catarina; Päth, Günter; Ebert, Regina;

    2011-01-01

    Reprogramming of multipotent adult bone marrow (BM)-derived mesenchymal stromal/stem cells (MSC) (BM-MSC) represents one of several strategies for cell-based therapy of diabetes. However, reprogramming primary BM-MSC into pancreatic endocrine lineages has not yet been consistently demonstrated....

  10. Self-glycolipids modulate dendritic cells changing the cytokine profiles of committed autoreactive T cells.

    Directory of Open Access Journals (Sweden)

    Karsten Buschard

    Full Text Available The impact of glycolipids of non-mammalian origin on autoimmune inflammation has become widely recognized. Here we report that the naturally occurring mammalian glycolipids, sulfatide and β-GalCer, affect the differentiation and the quality of antigen presentation by monocyte-derived dendritic cells (DCs. In response to sulfatide and β-GalCer, monocytes develop into immature DCs with higher expression of HLA-DR and CD86 but lower expression of CD80, CD40 and CD1a and lower production of IL-12 compared to non-modulated DCs. Self-glycolipid-modulated DCs responded to lipopolysaccharide (LPS by changing phenotype but preserved low IL-12 production. Sulfatide, in particular, reduced the capacity of DCs to stimulate autoreactive Glutamic Acid Decarboxylase (GAD65 - specific T cell response and promoted IL-10 production by the GAD65-specific clone. Since sulfatide and β-GalCer induced toll-like receptor (TLR-mediated signaling, we hypothesize that self-glycolipids deliver a (tolerogenic polarizing signal to differentiating DCs, facilitating the maintenance of self-tolerance under proinflammatory conditions.

  11. Skn-1a/Pou2f3 specifies taste receptor cell lineage

    OpenAIRE

    Matsumoto, Ichiro; Ohmoto, Makoto; Narukawa, Masataka; Yoshihara, Yoshihiro; Abe, Keiko

    2011-01-01

    Functional diversification of taste cells is crucial for proper discrimination of taste qualities. We found homeodomain protein Skn-1a/Pou2f3 is expressed in sweet, umami, and bitter taste cells. The Skn-1a–deficient mice lacked electrophysiological and behavioral responses to sweet, umami, and bitter tastes, due to complete absence of sweet, umami, and bitter cells with concomitant expansion of sour cells. Skn-1a is critical for generating and balancing the diverse composition of taste cells.

  12. Common and distinct signals specify the distribution of blood and vascular cell lineages in Xenopus laevis embryos.

    Science.gov (United States)

    Iraha, Fumie; Saito, Yoshinari; Yoshida, Keiko; Kawakami, Masatoki; Izutsu, Yumi; Daar, Ira Owen; Maéno, Mitsugu

    2002-10-01

    In an effort to elucidate the regulatory mechanisms that determine the fate of blood cells and vascular cells in the ventral blood island mesoderm, the embryonic expression of Xtie-2, a Xenopus homolog of the tie-2 receptor tyrosine kinase, was examined. Whole-mount in situ hybridization analysis revealed that Xtie-2 mRNA is expressed at the late tailbud stage within the regions where endothelial precursor cells exist. On the ventral side of embryos, Xtie-2-positive cells are predominantly present just outside the boundary of alpha-globin-positive cells, thus the expression pattern of these two markers seems mutually exclusive. Further experiments revealed that there is a consistent and strong correlation between the induction of Xtie-2 and alpha-globin expression in embryos and explant tissues. First, these two markers displayed overlapping expression in embryos ventralized by the removal of a "dorsal determinant" from the vegetal cytoplasm at the 1-cell stage. Second, expression of both Xtie-2 and alpha-globin were markedly induced in ectodermal explants (animal caps) from embryos co-injected with activin and bone morphogenetic protein (BMP)-4 RNA. Furthermore, both Xtie-2 and alpha-globin messages were strongly positive in dorsal marginal zone explants that had been injected with BMP-4 RNA. In contrast, however, there was a clear distinction in the localization of these two transcripts in embryos dorsalized by LiCl treatment. Distinct localization was also found in the ventral marginal zone (VMZ) explants. Using the VMZ explant system, we demonstrate a role of fibroblast growth factor (FGF) signaling in enhancing the vascular cell marker and reducing the blood cell marker. The present study suggests that the early steps of blood and vascular cell differentiation are regulated by a common BMP-4-dependent signaling; however, distinct factor(s) such as FGF are involved in different distribution of these two cell lineages. PMID:12392573

  13. Cross-differentiation from the CD8 lineage to CD4 T cells in the gut-associated microenvironment with a nonessential role of microbiota.

    Science.gov (United States)

    Lui, Jen Bon; Devarajan, Priyadharshini; Teplicki, Sarah A; Chen, Zhibin

    2015-02-01

    CD4 and CD8 T cell lineages differentiate through respective thymic selection processes. Here, we report cross-differentiation from the CD8 lineage to CD4 T cells, but not vice versa, predominantly in the large-intestine-associated microenvironment. It occurred in the absence or distal presence of cognate antigens. This pathway produced MHC-class-I-restricted CD4(+)Foxp3(+) T(reg) (CI-T(reg)) cells. Blocking T cell-intrinsic TGFβ signaling diminished CI-Treg populations in lamina propria, but it did not preclude the CD8-to-CD4 conversion. Microbiota were not required for the cross-differentiation, but the presence of microbiota led to expansion of the converted CD4 T cell population in the large intestine. CI-T(reg) cells did not promote tolerance to microbiota per se, but they regulated systemic homeostasis of T lymphocytes and protected the large intestine from inflammatory damage. Overall, the clonal conversion from the CD8 lineage to CD4 T cell subsets occurred regardless of "self" or "nonself." This lineage plasticity may promote "selfless" tolerance for immune balance. PMID:25640181

  14. Glucose starvation induces mutation and lineage-dependent adaptive responses in a large collection of cancer cell lines.

    Science.gov (United States)

    He, Ningning; Kim, Nayoung; Jeong, Euna; Lu, Yiling; Mills, Gordon B; Yoon, Sukjoon

    2016-01-01

    Tolerance of glucose deprivation is an important factor for cancer proliferation, survival, migration and progression. To systematically understand adaptive responses under glucose starvation in cancers, we analyzed reverse phase protein array (RPPA) data of 115 protein antibodies across a panel of approximately 170 heterogeneous cancer cell lines, cultured under normal and low glucose conditions. In general, glucose starvation broadly altered levels of many of the proteins and phosphoproteins assessed across the cell lines. Many mTOR pathway components were selectively sensitive to glucose stress, although the change in their levels still varied greatly across the cell line set. Furthermore, lineage- and genotype-based classification of cancer cell lines revealed mutation-specific variation of protein expression and phosphorylation in response to glucose starvation. Decreased AKT phosphorylation (S473) was significantly associated with PTEN mutation under glucose starvation conditions in lung cancer cell lines. The present study (see TCPAportal.org for data resource) provides insight into adaptive responses to glucose deprivation under diverse cellular contexts. PMID:26573869

  15. Transcription Factors and Medium Suitable for Initiating the Differentiation of Human-Induced Pluripotent Stem Cells to the Hepatocyte Lineage.

    Science.gov (United States)

    Tomizawa, Minoru; Shinozaki, Fuminobu; Motoyoshi, Yasufumi; Sugiyama, Takao; Yamamoto, Shigenori; Ishige, Naoki

    2016-09-01

    Transcription factors and culture media were investigated to determine the condition to initiate the differentiation of human-induced pluripotent stem (iPS) cells most efficiently. The expression of genes in human adult liver was compared with that in 201B7 cells (iPS cells) using cDNA microarray analysis. Episomal plasmids expressing transcription factors were constructed. 201B7 cells were transfected with the episomal plasmids and cultured in ReproFF (feeder-free media maintaining pluripotency), Leibovitz-15 (L15), William's E (WE), or Dulbecco's modified Eagle medium/Nutrient F-12 Ham (DF12) for 7 days. RNA was isolated and subjected to real-time quantitative PCR to analyze the expression of alpha-feto protein (AFP) and albumin. cDNA microarray analysis revealed 16 transcription factors that were upregulated in human adult liver relative to that in 201B7 cells. Episomal plasmids expressing these 16 genes were transfected into 201B7 cells. CCAAT/enhancer-binding protein alpha (CEBPA), CCAAT/enhancer-binding protein beta (CEBPB), forkhead box A1 (FOXA1), and forkhead box A3 (FOXA3) up-regulated AFP and down-regulated Nanog. These four genes were further analyzed. The expression of AFP and albumin was the highest in 201B7 cells transfected with the combination of CEBPA, CEBPB, FOXA1, and FOXA3 and cultured in WE. The combination of CEBPA, CEBPB, FOXA1, and FOXA3 was suitable for 201B7 cells to initiate differentiation to the hepatocyte lineage and WE was the most suitable medium for culture after transfection. J. Cell. Biochem. 117: 2001-2009, 2016. © 2016 Wiley Periodicals, Inc. PMID:26773721

  16. Mutually exclusive signaling signatures define the hepatic and pancreatic progenitor cell lineage divergence

    OpenAIRE

    Rodriguez-Seguel, E.; Mah, N.; Naumann, H.; Pongrac, I.M.; Cerda-Esteban, N.; Fontaine, J.-F.; Wang, Y.; Chen, W; Andrade-Navarro, M A; Spagnoli, F. M.

    2013-01-01

    Understanding how distinct cell types arise from multipotent progenitor cells is a major quest in stem cell biology. The liver and pancreas share many aspects of their early development and possibly originate from a common progenitor. However, how liver and pancreas cells diverge from a common endoderm progenitor population and adopt specific fates remains elusive. Using RNA sequencing (RNA-seq), we defined the molecular identity of liver and pancreas progenitors that were isolated from the m...

  17. neurogenin3 is required for the development of the four endocrine cell lineages of the pancreas

    OpenAIRE

    Gradwohl, Gérard; Dierich, Andrée; LeMeur, Marianne; Guillemot, François

    2000-01-01

    In the mammalian pancreas, the endocrine cell types of the islets of Langerhans, including the α-, β-, δ-, and pancreatic polypeptide cells as well as the exocrine cells, derive from foregut endodermal progenitors. Recent genetic studies have identified a network of transcription factors, including Pdx1, Isl1, Pax4, Pax6, NeuroD, Nkx2.2, and Hlxb9, regulating the development of islet cells at different stages, but the molecular mechanisms controlling the specification of pancreatic endocrine ...

  18. A new loss-of-function allele 28y reveals a role of ARGONAUTE1 in limiting asymmetric division of stomatal lineage ground cell

    Institute of Scientific and Technical Information of China (English)

    Kezhen Yangy; Min Jiangy; Jie Le

    2014-01-01

    In Arabidopsis thaliana L., stomata are produced through a series of divisions including asymmetric and symmetric divisions. Asymmetric entry division of meristemoid mother cellproduces two daughter cells, the smal er meristemoid and the larger sister cell, a stomatal lineage ground cell(SLGC). Stomatal lineage ground cells can differentiate into epidermal pavement cells but have the potential to divide asymmetrical y, spacing divisions, to create satel ite meristemoids. Peptide ligands and TOO MANY MOUTHS (TMM) and ERECTA family receptors regulate the initiation of stomatal lineages, activity, and orientation of spacing divisions. Here, we reported that a natural mutant 28y displayed an increased stomatal density and index. Using map-based cloning, we identified mutation in ARGONAUTE1 (AGO1) as the cause of 28y phenotypes. Time-lapse tracing of stomatal lineage cells reveals that stomatal overproduction in 28y is caused by the excessive asymmetric spacing division of SLGCs.Further genetic results demonstrated that AGO1 acts down-stream of TMM and negatively regulates the SPCH transcripts, but in a brassinosteroid-independent manner. Upregulation of AGAMOUS-LIKE16 (AGL16) in 28y mutants suggests that AGO1 is required to restrict AGL16-mediated stomatal spacing divisions, an miRNA pathway in addition to ligand-receptor signaling modules.

  19. The BCL6 RD2 domain governs commitment of activated B-cells to form germinal centers

    Science.gov (United States)

    Huang, Chuanxin; Gonzalez, David G.; Cote, Christine M.; Jiang, Yanwen; Hatzi, Katerina; Teater, Matt; Dai, Kezhi; Hla, Timothy; Haberman, Ann M.; Melnick, Ari

    2014-01-01

    Summary To understand how the Bcl6 transcriptional repressor functions in the immune system we disrupted its RD2 repression domain in mice. Bcl6RD2MUT mice exhibit a complete loss of GC formation but retain normal extrafollicular responses. Bcl6RD2MUT antigen-engaged B-cells migrate to the interfollicular zone and interact with cognate T helper cells. However, these cells fail to complete early GC-commitment differentiation and coalesce as nascent GC aggregates. Bcl6 directly binds and represses trafficking receptors S1pr1 and Gpr183 by recruiting Hdac2 through the RD2 domain. Deregulation of these genes impairs B-cell migration and may contribute to GC failure in Bcl6RD2MUT mice. The development of functional GC-TFH cells was partially impaired in Bcl6RD2MUT mice. In contrast to Bcl6−/− mice, Bcl6RD2MUT animals experience no inflammatory disease or macrophage deregulation. These results reveal an essential role for RD2 repression in early GC commitment and striking biochemical specificity in Bcl6 control of humoral and innate immune-cell phenotypes. PMID:25176650

  20. Gamma Interferon Signaling in Macrophage Lineage Cells Regulates Central Nervous System Inflammation and Chemokine Production ▿

    OpenAIRE

    Lin, Adora A.; Tripathi, Pulak K.; Sholl, Allyson; Jordan, Michael B.; Hildeman, David A.

    2009-01-01

    Intracranial (i.c.) infection of mice with lymphocytic choriomeningitis virus (LCMV) results in anorexic weight loss, mediated by T cells and gamma interferon (IFN-γ). Here, we assessed the role of CD4+ T cells and IFN-γ on immune cell recruitment and proinflammatory cytokine/chemokine production in the central nervous system (CNS) after i.c. LCMV infection. We found that T-cell-depleted mice had decreased recruitment of hematopoietic cells to the CNS and diminished levels of IFN-γ, CCL2 (MCP...

  1. Lineage-specific stem cells, signals and asymmetries during stomatal development.

    Science.gov (United States)

    Han, Soon-Ki; Torii, Keiko U

    2016-04-15

    Stomata are dispersed pores found in the epidermis of land plants that facilitate gas exchange for photosynthesis while minimizing water loss. Stomata are formed from progenitor cells, which execute a series of differentiation events and stereotypical cell divisions. The sequential activation of master regulatory basic-helix-loop-helix (bHLH) transcription factors controls the initiation, proliferation and differentiation of stomatal cells. Cell-cell communication mediated by secreted peptides, receptor kinases, and downstream mitogen-activated kinase cascades enforces proper stomatal patterning, and an intrinsic polarity mechanism ensures asymmetric cell divisions. As we review here, recent studies have provided insights into the intrinsic and extrinsic factors that control stomatal development. These findings have also highlighted striking similarities between plants and animals with regards to their mechanisms of specialized cell differentiation. PMID:27095491

  2. Targeting IκB kinase β in Adipocyte Lineage Cells for Treatment of Obesity and Metabolic Dysfunctions.

    Science.gov (United States)

    Helsley, Robert N; Sui, Yipeng; Park, Se-Hyung; Liu, Zun; Lee, Richard G; Zhu, Beibei; Kern, Philip A; Zhou, Changcheng

    2016-07-01

    IκB kinase β (IKKβ), a central coordinator of inflammation through activation of nuclear factor-κB, has been identified as a potential therapeutic target for the treatment of obesity-associated metabolic dysfunctions. In this study, we evaluated an antisense oligonucleotide (ASO) inhibitor of IKKβ and found that IKKβ ASO ameliorated diet-induced metabolic dysfunctions in mice. Interestingly, IKKβ ASO also inhibited adipocyte differentiation and reduced adiposity in high-fat (HF)-fed mice, indicating an important role of IKKβ signaling in the regulation of adipocyte differentiation. Indeed, CRISPR/Cas9-mediated genomic deletion of IKKβ in 3T3-L1 preadipocytes blocked these cells differentiating into adipocytes. To further elucidate the role of adipose progenitor IKKβ signaling in diet-induced obesity, we generated mice that selectively lack IKKβ in the white adipose lineage and confirmed the essential role of IKKβ in mediating adipocyte differentiation in vivo. Deficiency of IKKβ decreased HF-elicited adipogenesis in addition to reducing inflammation and protected mice from diet-induced obesity and insulin resistance. Further, pharmacological inhibition of IKKβ also blocked human adipose stem cell differentiation. Our findings establish IKKβ as a pivotal regulator of adipogenesis and suggest that overnutrition-mediated IKKβ activation serves as an initial signal that triggers adipose progenitor cell differentiation in response to HF feeding. Inhibition of IKKβ with antisense therapy may represent as a novel therapeutic approach to combat obesity and metabolic dysfunctions. Stem Cells 2016;34:1883-1895. PMID:26991836

  3. Highly Synchronized Expression of Lineage-Specific Genes during In Vitro Hepatic Differentiation of Human Pluripotent Stem Cell Lines.

    Science.gov (United States)

    Ghosheh, Nidal; Olsson, Björn; Edsbagge, Josefina; Küppers-Munther, Barbara; Van Giezen, Mariska; Asplund, Annika; Andersson, Tommy B; Björquist, Petter; Carén, Helena; Simonsson, Stina; Sartipy, Peter; Synnergren, Jane

    2016-01-01

    Human pluripotent stem cells- (hPSCs-) derived hepatocytes have the potential to replace many hepatic models in drug discovery and provide a cell source for regenerative medicine applications. However, the generation of fully functional hPSC-derived hepatocytes is still a challenge. Towards gaining better understanding of the differentiation and maturation process, we employed a standardized protocol to differentiate six hPSC lines into hepatocytes and investigated the synchronicity of the hPSC lines by applying RT-qPCR to assess the expression of lineage-specific genes (OCT4, NANOG, T, SOX17, CXCR4, CER1, HHEX, TBX3, PROX1, HNF6, AFP, HNF4a, KRT18, ALB, AAT, and CYP3A4) which serve as markers for different stages during liver development. The data was evaluated using correlation and clustering analysis, demonstrating that the expression of these markers is highly synchronized and correlated well across all cell lines. The analysis also revealed a distribution of the markers in groups reflecting the developmental stages of hepatocytes. Functional analysis of the differentiated cells further confirmed their hepatic phenotype. Taken together, these results demonstrate, on the molecular level, the highly synchronized differentiation pattern across multiple hPSC lines. Moreover, this study provides additional understanding for future efforts to improve the functionality of hPSC-derived hepatocytes and thereby increase the value of related models. PMID:26949401

  4. Myotube formation is affected by adipogenic lineage cells in a cell-to-cell contact-independent manner

    Energy Technology Data Exchange (ETDEWEB)

    Takegahara, Yuki; Yamanouchi, Keitaro, E-mail: akeita@mail.ecc.u-tokyo.ac.jp; Nakamura, Katsuyuki; Nakano, Shin-ichi; Nishihara, Masugi

    2014-05-15

    Intramuscular adipose tissue (IMAT) formation is observed in some pathological conditions such as Duchenne muscular dystrophy (DMD) and sarcopenia. Several studies have suggested that IMAT formation is not only negatively correlated with skeletal muscle mass but also causes decreased muscle contraction in sarcopenia. In the present study, we examined w hether adipocytes affect myogenesis. For this purpose, skeletal muscle progenitor cells were transfected with siRNA of PPARγ (siPPARγ) in an attempt to inhibit adipogenesis. Myosin heavy chain (MHC)-positive myotube formation was promoted in cells transfected with siPPARγ compared to that of cells transfected with control siRNA. To determine whether direct cell-to-cell contact between adipocytes and myoblasts is a prerequisite for adipocytes to affect myogenesis, skeletal muscle progenitor cells were cocultured with pre- or mature adipocytes in a Transwell coculture system. MHC-positive myotube formation was inhibited when skeletal muscle progenitor cells were cocultured with mature adipocytes, but was promoted when they were cocultured with preadipocytes. Similar effects were observed when pre- or mature adipocyte-conditioned medium was used. These results indicate that preadipocytes play an important role in maintaining skeletal muscle mass by promoting myogenesis; once differentiated, the resulting mature adipocytes negatively affect myogenesis, leading to the muscle deterioration observed in skeletal muscle pathologies. - Highlights: • We examined the effects of pre- and mature adipocytes on myogenesis in vitro. • Preadipocytes and mature adipocytes affect myoblast fusion. • Preadipocytes play an important role in maintaining skeletal muscle mass. • Mature adipocytes lead to muscle deterioration observed in skeletal muscle pathologies.

  5. The roles of ERAS during cell lineage specification of mouse early embryonic development

    OpenAIRE

    Zhao, Zhen-Ao; Yu, Yang; Ma, Huai-Xiao; Wang, Xiao-Xiao; Lu, Xukun; Zhai, Yanhua; Zhang, Xiaoxin; Wang, Haibin; Li, Lei

    2015-01-01

    Eras encodes a Ras-like GTPase protein that was originally identified as an embryonic stem cell-specific Ras. ERAS has been known to be required for the growth of embryonic stem cells and stimulates somatic cell reprogramming, suggesting its roles on mouse early embryonic development. We now report a dynamic expression pattern of Eras during mouse peri-implantation development: its expression increases at the blastocyst stage, and specifically decreases in E7.5 mesoderm. In accordance with it...

  6. Real-Time Lineage Analysis to Study Cell Division Orientation in the Arabidopsis Shoot Meristem.

    Science.gov (United States)

    Tobin, Cory J; Meyerowitz, Elliot M

    2016-01-01

    Cells in the Arabidopsis shoot apical meristem are small and divide frequently throughout the life-time of the organism making them good candidates for studying the mechanisms of cell division in plants. But tracking these cell divisions requires multiple images to be taken of the same specimen over time which means the specimen must stay alive throughout the process. This chapter provides details on how to prepare plants for live imaging, keep them alive and growing through multiple time points, and how to process the data to extract cell boundary coordinates from three-dimensional images. PMID:26659961

  7. Effects of dose rates on radiation-induced replenishment of intestinal stem cells determined by Lgr5 lineage tracing

    International Nuclear Information System (INIS)

    An understanding of the dynamics of intestinal Lgr5+ stem cells is important for elucidating the mechanism of colonic cancer development. We previously established a method for evaluating Lgr5+ stem cells by tamoxifen-dependent Lgr5-lineage tracing and showed that high-dose-rate radiation stimulated replenishment of colonic stem cells. In this study, we evaluated the effects of low-dose-rate radiation on stem cell maintenance. Tamoxifen (4OHT)-injected Lgr5-EGFP-IRES-CreERT2 × ROSA-LSL-LacZ mice were used, LacZ-labeled colonic crypts were enumerated, and the loss of LacZ+ crypts under low-dose-rate radiation was estimated. After 4OHT treatment, the number of LacZ-labeled Lgr5+ stem cells was higher in the colon of infant mice than in adult mice. The percentage of LacZ-labeled crypts in infant mice rapidly decreased after 4OHT treatment. However, the percentage of labeled crypts plateaued at ∼2% at 4 weeks post-treatment and remained unchanged for up to 7 months. Thus, it will be advantageous to evaluate the long-term effects of low-dose-rate radiation. Next, we determined the percentages of LacZ-labeled crypts irradiated with 1 Gy administered at different dose rates. As reported in our previous study, mice exposed to high-dose-rate radiation (30 Gy/h) showed a marked replenishment (P = 0.04). However, mice exposed to low-dose-rate radiation (0.003 Gy/h) did not exhibit accelerated stem-cell replenishment (P = 0.47). These findings suggest the percentage of labeled crypts can serve as a useful indicator of the effects of dose rate on the stem cell pool. (author)

  8. Origin and dynamic lineage characteristics of the developing Drosophila midgut stem cells.

    Science.gov (United States)

    Takashima, Shigeo; Aghajanian, Patrick; Younossi-Hartenstein, Amelia; Hartenstein, Volker

    2016-08-15

    Proliferating intestinal stem cells (ISCs) generate all cell types of the Drosophila midgut, including enterocytes, endocrine cells, and gland cells (e.g., copper cells), throughout the lifetime of the animal. Among the signaling mechanisms controlling the balance between ISC self-renewal and the production of different cell types, Notch (N) plays a pivotal role. In this paper we investigated the emergence of ISCs during metamorphosis and the role of N in this process. Precursors of the Drosophila adult intestinal stem cells (pISCs) can be first detected within the pupal midgut during the first hours after onset of metamorphosis as motile mesenchymal cells. pISCs perform 2-3 rounds of parasynchronous divisions. The first mitosis yields only an increase in pISC number. During the following rounds of mitosis, dividing pISCs give rise to more pISCs, as well as the endocrine cells that populate the midgut of the eclosing fly. Enterocytes do not appear among the pISC progeny until around the time of eclosion. The "proendocrine" gene prospero (pros), expressed from mid-pupal stages onward in pISCs, is responsible to advance the endocrine fate in these cells; following removal of pros, pISCs continue to proliferate, but endocrine cells do not form. Conversely, the onset of N activity that occurs around the stage when pros comes on restricts pros expression among pISCs. Loss of N abrogates proliferation and switches on an endocrine fate among all pISCs. Our results suggest that a switch depending on the activity of N and pros acts at the level of the pISC to decide between continued proliferation and endocrine differentiation. PMID:27321560

  9. Identification of a negative regulatory role for spi-C in the murine B cell lineage.

    Science.gov (United States)

    Li, Stephen K H; Solomon, Lauren A; Fulkerson, Patricia C; DeKoter, Rodney P

    2015-04-15

    Spi-C is an E26 transformation-specific family transcription factor that is highly related to PU.1 and Spi-B. Spi-C is expressed in developing B cells, but its function in B cell development and function is not well characterized. To determine whether Spi-C functions as a negative regulator of Spi-B (encoded by Spib), mice were generated that were germline knockout for Spib and heterozygous for Spic (Spib(-/-)Spic(+/-)). Interestingly, loss of one Spic allele substantially rescued B cell frequencies and absolute numbers in Spib(-/-) mouse spleens. Spib(-/-)Spic(+/-) B cells had restored proliferation compared with Spib(-/-) B cells in response to anti-IgM or LPS stimulation. Investigation of a potential mechanism for the Spib(-/-)Spic(+/-) phenotype revealed that steady-state levels of Nfkb1, encoding p50, were elevated in Spib(-/-)Spic(+/-) B cells compared with Spib(-/-) B cells. Spi-B was shown to directly activate the Nfkb1 gene, whereas Spi-C was shown to repress this gene. These results indicate a novel role for Spi-C as a negative regulator of B cell development and function. PMID:25769919

  10. Blood-borne stem cells differentiate into vascular and cardiac lineages during normal development

    Czech Academy of Sciences Publication Activity Database

    Zhang, N.; Mustin, D.; Reardon, M. W.; Dealmeida, A.; Mozdziak, P.; Mrug, M.; Eisenberg, L. M.; Sedmera, David

    2006-01-01

    Roč. 15, 1 (2006), s. 17-28. ISSN 1547-3287 Grant ostatní: March of Dimes 5-FY02-269; NIH RR16434 Institutional research plan: CEZ:AV0Z50450515 Keywords : stem cells * embryonic development * circulation Subject RIV: EA - Cell Biology Impact factor: 3.076, year: 2006

  11. Lineage tracing of resident tendon progenitor cells during growth and natural healing.

    Directory of Open Access Journals (Sweden)

    Nathaniel A Dyment

    Full Text Available Unlike during embryogenesis, the identity of tissue resident progenitor cells that contribute to postnatal tendon growth and natural healing is poorly characterized. Therefore, we utilized 1 an inducible Cre driven by alpha smooth muscle actin (SMACreERT2, that identifies mesenchymal progenitors, 2 a constitutively active Cre driven by growth and differentiation factor 5 (GDF5Cre, a critical regulator of joint condensation, in combination with 3 an Ai9 Cre reporter to permanently label SMA9 and GDF5-9 populations and their progeny. In growing mice, SMA9+ cells were found in peritendinous structures and scleraxis-positive (ScxGFP+ cells within the tendon midsubstance and myotendinous junction. The progenitors within the tendon midsubstance were transiently labeled as they displayed a 4-fold expansion from day 2 to day 21 but reduced to baseline levels by day 70. SMA9+ cells were not found within tendon entheses or ligaments in the knee, suggesting a different origin. In contrast to the SMA9 population, GDF5-9+ cells extended from the bone through the enthesis and into a portion of the tendon midsubstance. GDF5-9+ cells were also found throughout the length of the ligaments, indicating a significant variation in the progenitors that contribute to tendons and ligaments. Following tendon injury, SMA9+ paratenon cells were the main contributors to the healing response. SMA9+ cells extended over the defect space at 1 week and differentiated into ScxGFP+ cells at 2 weeks, which coincided with increased collagen signal in the paratenon bridge. Thus, SMA9-labeled cells represent a unique progenitor source that contributes to the tendon midsubstance, paratenon, and myotendinous junction during growth and natural healing, while GDF5 progenitors contribute to tendon enthesis and ligament development. Understanding the mechanisms that regulate the expansion and differentiation of these progenitors may prove crucial to improving future repair strategies.

  12. Chimeric-transgenic mice represent a powerful tool for studying how the proliferation and differentiation programs of intestinal epithelial cell lineages are regulated.

    OpenAIRE

    Hermiston, M L; Green, R. P.; Gordon, J I

    1993-01-01

    An in vivo system has been developed for examining the effects of wild-type or mutant proteins on cell fate determination in the mouse intestinal epithelium or on the proliferation and differentiation programs of its component epithelial lineages. This system takes advantage of the fact that at the conclusion of gut morphogenesis, each intestinal crypt is composed of a monoclonal population of cells descended from a single active multipotent stem cell, each villus is supplied by several monoc...

  13. Deleterious effects on MDAMB-231 breast adenocarcinoma cell lineage submitted to Ho-166 radioactive seeds at very low activity

    International Nuclear Information System (INIS)

    Herein, the deleterious effect of ionizing radiation provided by Ho-166 radioactive seeds at low activity were addressed, based on experimental in vitro assays at the MDA MB231 cell lineage, a breast adenocarcinoma, compared to PBMC - peripheral blood cells. The methodology involves of the MDBMB-231 and PBMC expansion in culture in suitable environment in 30mm well plates and T-25 flasks. Seeds were synthesized with Ho-165 incorporated and characterized previously. Activation was processed at IPR1 reactor at the peripheral table, at 8h exposition. Three groups of seeds were tested: 0,34 mCi, 0,12 mCi activity, and control group. Such seeds were placed on culture and held to a period of 05 half-lives of the radionuclide. The biological responses at these exposure were documented by inverse microscopic photographic in time. Also, MTT essay were performed. A fast response in producing deleterious effects at cancer cell was observed even if for the low activity seeds. Also, a biological response dependent to a radial distance of the seed was observed. At conclusion, viability clonogenic control of MDAMB231 is identified at the exposition to Ho-166 ceramic seeds, even if at low activity of 0,1 to 0,3mCi. (author)

  14. Deleterious effects on MDAMB-231 breast adenocarcinoma cell lineage submitted to Ho-166 radioactive seeds at very low activity

    Energy Technology Data Exchange (ETDEWEB)

    Falcao, Patricia L.; Campos, Tarcisio P.R., E-mail: campos@nuclear.ufmg.br [Universidade Federal de Minas Gerais (UFMG), Belo Horizonte, MG (Brazil). Dept. de Engenharia Nuclear; Sarmento, Eduardo V. [Centro de Desenvolvimento de Tecnologia Nuclear (CDTN/CNEN-MG), Belo Horizonte, MG (Brazil); Cuperschmid, Ethel M. [Universidade Federal de Minas Gerais (CEMEMOR/UFMG), Belo Horizonte, BR (Brazil). Fac. de Medicina. Centro de Memoria da Medicina

    2011-07-01

    Herein, the deleterious effect of ionizing radiation provided by Ho-166 radioactive seeds at low activity were addressed, based on experimental in vitro assays at the MDA MB231 cell lineage, a breast adenocarcinoma, compared to PBMC - peripheral blood cells. The methodology involves of the MDBMB-231 and PBMC expansion in culture in suitable environment in 30mm well plates and T-25 flasks. Seeds were synthesized with Ho-165 incorporated and characterized previously. Activation was processed at IPR1 reactor at the peripheral table, at 8h exposition. Three groups of seeds were tested: 0,34 mCi, 0,12 mCi activity, and control group. Such seeds were placed on culture and held to a period of 05 half-lives of the radionuclide. The biological responses at these exposure were documented by inverse microscopic photographic in time. Also, MTT essay were performed. A fast response in producing deleterious effects at cancer cell was observed even if for the low activity seeds. Also, a biological response dependent to a radial distance of the seed was observed. At conclusion, viability clonogenic control of MDAMB231 is identified at the exposition to Ho-166 ceramic seeds, even if at low activity of 0,1 to 0,3mCi. (author)

  15. Lineage-specific STAT5 target gene activation in hematopoietic progenitor cells predicts the FLT3(+)-mediated leukemic phenotype.

    Science.gov (United States)

    Müller, T A; Grundler, R; Istvanffy, R; Rudelius, M; Hennighausen, L; Illert, A L; Duyster, J

    2016-08-01

    Mutations that activate FMS-like tyrosine kinase 3 (FLT3) are frequent occurrences in acute myeloid leukemia. Two distinct types of mutations have been described: internal duplication of the juxtamembranous domain (ITD) and point mutations of the tyrosine kinase domain (TKD). Although both mutations lead to constitutive FLT3 signaling, only FLT3-ITD strongly activates signal transducer and activator of transcription 5 (STAT5). In a murine transplantation model, FLT3-ITD induces a myeloproliferative neoplasm, whereas FLT3-TKD leads to a lymphoid malignancy with significantly longer latency. Here we report that the presence of STAT5 is critical for the development of a myeloproliferative disease by FLT3-ITD in mice. Deletion of Stat5 in FLT3-ITD-induced leukemogenesis leads not only to a significantly longer survival (82 vs 27 days) of the diseased mice, but also to an immunophenotype switch with expansion of the lymphoid cell compartment. Interestingly, we were able to show differential STAT5 activation in FLT3-ITD(+) myeloid and lymphoid murine progenitors. STAT5 target genes such as Oncostatin M were highly expressed in FLT3-ITD(+) myeloid but not in FLT3-ITD(+) lymphoid progenitor cells. Strikingly, FLT3-TKD expression in combination with Oncostatin M is sufficient to reverse the phenotype to a myeloproliferative disease in FLT3-TKD mice. Thus, lineage-specific STAT5 activation in hematopoietic progenitor cells predicts the FLT3(+)-mediated leukemic phenotype in mice. PMID:27046463

  16. Adipose tissue stem cells meet preadipocyte commitment: going back to the future[S

    OpenAIRE

    Cawthorn, William P; Erica L. Scheller; MacDougald, Ormond A.

    2012-01-01

    White adipose tissue (WAT) is perhaps the most plastic organ in the body, capable of regeneration following surgical removal and massive expansion or contraction in response to altered energy balance. Research conducted for over 70 years has investigated adipose tissue plasticity on a cellular level, spurred on by the increasing burden that obesity and associated diseases are placing on public health globally. This work has identified committed preadipocytes in the stromal vascular fraction o...

  17. Micro-/Nano- sized hydroxyapatite directs differentiation of rat bone marrow derived mesenchymal stem cells towards an osteoblast lineage

    Science.gov (United States)

    Huang, Yan; Zhou, Gang; Zheng, Lisha; Liu, Haifeng; Niu, Xufeng; Fan, Yubo

    2012-03-01

    Regenerative medicine consisting of cells and materials provides a new way for the repair and regeneration of tissues and organs. Nano-biomaterials are highlighted due to their advantageous features compared with conventional micro-materials. The aim of this study is to investigate the effects of micro-/nano- sized hydroxyapatite (μ/n-HA) on the osteogenic differentiation of rat bone marrow derived mesenchymal stem cells (rBMSCs). μ/n-HA were prepared by a microwave synthesizer and precipitation method, respectively. Different sizes of μ/n-HA were characterized by IR, XRD, SEM, TEM and co-cultured with rBMSCs. It was shown that rBMSCs expressed higher levels of osteoblast-related markers by n-HA than μ-HA stimulation. The size of HA is an important factor for affecting the osteogenic differentiation of rBMSCs. This provides a new avenue for mechanistic studies of stem cell differentiation and a new approach to obtain more committed differentiated cells.

  18. The FOXD1 lineage of kidney perivascular cells and myofibroblasts: functions and responses to injury

    Science.gov (United States)

    Gomez, Ivan G; Duffield, Jeremy S

    2014-01-01

    Recent studies have identified a poorly appreciated yet extensive population of perivascular mesenchymal cells in the kidney, which are derived from metanephric mesenchyme progenitor cells during nephrogenesis at which time they express the transcription factor FOXD1. Some studies have called these resident fibroblasts, whereas others have called them pericytes. Regardless of nomenclature, many are partially integrated into the capillary basement membrane and contribute in important ways to the homeostasis of peritubular capillaries. Fate-mapping studies using conditional CreER recombinase-mediated tracing of discrete cell cohorts have identified these pericytes and resident fibroblasts as the major precursor population of interstitial myofibroblasts in animal models of kidney disease. Here, we will review the evidence that they are the major population of myofibroblast precursors, highlight some critical functions in homeostasis, and focus on the cell signaling pathways that are important to their differentiation into, and persistence as myofibroblasts. PMID:26312147

  19. Stem-cell dynamics and lineage topology from in vivo fate mapping in the hematopoietic system.

    Science.gov (United States)

    Höfer, Thomas; Barile, Melania; Flossdorf, Michael

    2016-06-01

    In recent years, sophisticated fate-mapping tools have been developed to study the behavior of stem cells in the intact organism. These experimental approaches are beginning to yield a quantitative picture of how cell numbers are regulated during steady state and in response to challenges. Focusing on hematopoiesis and immune responses, we discuss how novel mathematical approaches driven by these fate-mapping data have provided insights into the dynamics and topology of cellular differentiation pathways in vivo. The combination of experiment and theory has allowed to quantify the degree of self-renewal in stem and progenitor cells, shown how native hematopoiesis differs fundamentally from post-transplantation hematopoiesis, and uncovered that the diversification of T lymphocytes during immune responses resembles tissue renewal driven by stem cells. PMID:27107166

  20. Memory lineage relationships in HTLV-1-specific CD8+ cytotoxic T cells

    Science.gov (United States)

    Johnson-Nauroth, Julie M.; Graber, Jerome; Yao, Karen; Jacobson, Steve; Calabresi, Peter A.

    2016-01-01

    Cytotoxic memory T cells play a critical role in combating viral infections; however, in some diseases they may contribute to tissue damage. In HAM/TSP, HTLV-1 Tax 11–19+ cells proliferate spontaneously in vitro and can be tracked using the Tax 11–19 MHC Class I tetramer. Immediately ex vivo, these cells were a mix of CD45RA−/CCR7− TEM and CD45RA+/CCR7− TDiff memory CTL. The subsequent proliferating Tax 11–19 tetramer+ population expressed low levels of IL-7Rα, failed to respond to IL-7 and IL-15, and did not develop a TCM phenotype. Thus, chronic exposure to viral antigen may result in a sustained pool of TEM cells that home to the CNS and mediate the spinal cord pathology seen in this disease. PMID:16740321

  1. Defining the Minimal Factors Required for Erythropoiesis through Direct Lineage Conversion

    Directory of Open Access Journals (Sweden)

    Sandra Capellera-Garcia

    2016-06-01

    Full Text Available Erythroid cell commitment and differentiation proceed through activation of a lineage-restricted transcriptional network orchestrated by a group of well characterized genes. However, the minimal set of factors necessary for instructing red blood cell (RBC development remains undefined. We employed a screen for transcription factors allowing direct lineage reprograming from fibroblasts to induced erythroid progenitors/precursors (iEPs. We show that Gata1, Tal1, Lmo2, and c-Myc (GTLM can rapidly convert murine and human fibroblasts directly to iEPs. The transcriptional signature of murine iEPs resembled mainly that of primitive erythroid progenitors in the yolk sac, whereas addition of Klf1 or Myb to the GTLM cocktail resulted in iEPs with a more adult-type globin expression pattern. Our results demonstrate that direct lineage conversion is a suitable platform for defining and studying the core factors inducing the different waves of erythroid development.

  2. The FOXD1 lineage of kidney perivascular cells and myofibroblasts: functions and responses to injury

    OpenAIRE

    Gomez, Ivan G.; Duffield, Jeremy S.

    2014-01-01

    Recent studies have identified a poorly appreciated yet extensive population of perivascular mesenchymal cells in the kidney, which are derived from metanephric mesenchyme progenitor cells during nephrogenesis at which time they express the transcription factor FOXD1. Some studies have called these resident fibroblasts, whereas others have called them pericytes. Regardless of nomenclature, many are partially integrated into the capillary basement membrane and contribute in important ways to t...

  3. A distinct lineage of CD4 T cells regulates tissue inflammation by producing interleukin 17

    OpenAIRE

    Park, Heon; Li, Zhaoxia; Yang, Xuexian O.; Chang, Seon Hee; Nurieva, Roza; Wang, Yi-hong; Wang, Ying; Hood, Leroy; Zhu, Zhou; Tian, Qiang; Dong, Chen

    2005-01-01

    Interleukin 17 (IL-17) has been linked to autoimmune diseases, although its regulation and function have remained unclear. Here we have evaluated in vitro and in vivo the requirements for the differentiation of naive CD4 T cells into effector T helper cells that produce IL-17. This process required the costimulatory molecules CD28 and ICOS but was independent of the cytokines and transcription factors required for T helper type 1 or type 2 differentiation. Furthermore, both IL-4 and interfero...

  4. Lineage tracing of metastasis in a mouse model for Non-small cell lung cancer (NSCLC)

    OpenAIRE

    Thakur, Chitra

    2012-01-01

    Non-small cell lung cancer (NSCLC) is the deadliest form of lung cancer and has a poor prognosis due to its high rate of metastasis. Notably, metastasis is one of the leading causes of death among cancer patients. Despite the clinical importance, the cellular and molecular mechanisms that govern the initiation, establishment and progression of metastasis remain unclear. Moreover, knowledge gained on metastatic process was largely based on cultured or in vitro manipulated cells that were reint...

  5. Stem Cell Derived Extracellular Matrix Enables Survival and Multi Lineage Differentiation within Superporous Hydrogels

    OpenAIRE

    Köllmer, Melanie; Keskar, Vandana; Hauk, Thomas G.; Collins, John M.; Russell, Brenda; Gemeinhart, Richard A.

    2012-01-01

    Hydrophilic poly(ethylene glycol) diacrylate (PEGDA) hydrogel surfaces resist protein adsorption and are generally thought to be unsuitable for anchorage dependent cells to adhere. Intriguingly, our previous findings revealed that PEGDA superporous hydrogel scaffolds (SPHs) allow anchorage of bone marrow derived human mesenchymal stem cells (hMSCs) and support their long term survival. Therefore, we hypothesized that the physicochemical characteristics of the scaffold impart properties that c...

  6. Rat Blastocysts from Nuclear Injection and Time-Lagged Enucleation and Their Commitment to Embryonic Stem Cells.

    Science.gov (United States)

    Hara, Hiromasa; Goto, Teppei; Takizawa, Akiko; Sanbo, Makoto; Jacob, Howard J; Kobayashi, Toshihiro; Nakauchi, Hiromitsu; Hochi, Shinichi; Hirabayashi, Masumi

    2016-04-01

    Pronucleus-like vesicle formation following premature chromosome condensation (PCC) of the donor cell nucleus is the key event for successful generation of cloned rodents by nuclear transplantation (NT). However in rat cloning, this change is difficult to induce in enucleated recipient oocytes because of their inability to maintain maturation-promoting factor levels. In this study, intact oocytes retrieved from nuclear-visualized H2B-tdTomato knock-in rats were injected with Venus-labeled cell nuclei. Because the incidence of PCC under MG-132 treatment significantly increased with the culture period (0%, 10.8%, 36.8%, and 87.5% at 0, 0.5, 1, and 2 h postinjection, respectively), the metaphase plate of the oocyte was removed 1-2 h after the nuclear injection. The NT-derived rat zygotes (n = 748) were activated with ionomycin/cycloheximide and transferred into temporal host mothers, resulting in the harvest of three blastocysts (0.4%) with Venus fluorescence. Two blastocysts were examined for their potential to commit to NT-derived embryonic stem cells (ntESCs). One ntESC line was established successfully and found to be competent in terms of karyotype, stem cell marker expression, and pluripotency. In conclusion, time-lagged enucleation of visualized oocyte nuclei allows the PCC incidence of donor nuclei and generation of NT blastocysts, and the blastocysts can commit to germline-competent ntESCs. PMID:26990947

  7. A Dorsal SHH-Dependent Domain in the V-SVZ Produces Large Numbers of Oligodendroglial Lineage Cells in the Postnatal Brain

    Directory of Open Access Journals (Sweden)

    Cheuk Ka Tong

    2015-10-01

    Full Text Available Neural stem cells in different locations of the postnatal mouse ventricular-subventricular zone (V-SVZ generate different subtypes of olfactory bulb (OB interneurons. High Sonic hedgehog (SHH signaling in the ventral V-SVZ regulates the production of specific subtypes of neurons destined for the OB. Here we found a transient territory of high SHH signaling in the dorsal V-SVZ beneath the corpus callosum (CC. Using intersectional lineage tracing in neonates to label dorsal radial glial cells (RGCs expressing the SHH target gene Gli1, we demonstrate that this region produces many CC cells in the oligodendroglial lineage and specific subtypes of neurons in the OB. The number of oligodendroglial cells generated correlated with the levels of SHH signaling. This work identifies a dorsal domain of SHH signaling, which is an important source of oligodendroglial cells for the postnatal mammalian forebrain.

  8. Notch-mediated post-translational control of Ngn3 protein stability regulates pancreatic patterning and cell fate commitment

    DEFF Research Database (Denmark)

    Qu, Xiaoling; Afelik, Solomon; Jensen, Jan Nygaard;

    2013-01-01

    involves transcriptional repression as previously shown, but also incorporates a novel post-translational mechanism. In addition to its ability to promote endocrine fate, we provide evidence of a competing ability of Ngn3 in the patterning of multipotent progenitor cells in turn controlling the formation......-producing beta-cells can be significantly enhanced upon induction of a pro-endocrine drive combined with the inhibition of Notch processing....... of ducts. On one hand, Ngn3 cell-intrinsically activates endocrine target genes; on the other, Ngn3 cell-extrinsically promotes lateral signaling via the Dll1>Notch>Hes1 pathway which substantially limits its ability to sustain endocrine formation. Prior to endocrine commitment, the Ngn3-mediated...

  9. A population of human brain cells expressing phenotypic markers of more than one lineage can be induced in vitro to differentiate into mesenchymal cells

    International Nuclear Information System (INIS)

    Proliferating astrocytic cells from germinal, as well as mature areas of brain parenchyma, have the characteristics of neural stem/progenitor cells and are capable of generating both neurons and glia. We previously reported that primary fetal human brain cells, designated as Normal Human Astrocytes (NHA), expressed, in addition to GFAP, Vimentin and Nestin, low levels of βIII-Tubulin, an early neuronal marker, and differentiated into neurons and astrocytes in vitro. Here, we showed that primary NHA cells co-express low levels of mesenchymal markers Fibronectin and Collagen-1 in culture. These cells transitioned into mesenchymal-like cells when cultured in adherent conditions in serum containing media. The mesenchymal-like derivatives of these cells were characterized based on their morphological changes, high expression of Vimentin and extracellular matrix (ECM) proteins, Collagen-1 and Fibronectin, and decline of neural markers. When incubated in osteogenic and adipogenic induction media, the mesenchymal-like cells differentiated into osteoblasts and adipocytes. Furthermore, NHA cells express markers of neural crest cells, SOX-10 and p75. These data support the idea of ectoderm-derived mesenchymal lineages. These findings suggest that a population of primitive fetal brain cells with neural/neural crest/mesenchymal phenotype, resembles the remarkable phenotypic plasticity of neural crest cells, and differentiates into adipocytes and osteocytes under the influence of environmental factors

  10. Jagged1 expression by osteoblast-lineage cells regulates trabecular bone mass and periosteal expansion in mice.

    Science.gov (United States)

    Youngstrom, D W; Dishowitz, M I; Bales, C B; Carr, E; Mutyaba, P L; Kozloff, K M; Shitaye, H; Hankenson, K D; Loomes, K M

    2016-10-01

    through homotypic Jag1 signaling in osteochondral progenitors, but not mature osteoblasts, inhibits periosteal expansion. Therefore, Jag1 signaling within the osteoblast lineage regulates bone metabolism in a compartment-dependent manner. Moreover, loss of Jag1 function in osteoblast lineage cells may contribute to the skeletal phenotype associated with ALGS. PMID:27416809

  11. Cell-lineage regulated myogenesis for dystrophin replacement: a novel therapeutic approach for treatment of muscular dystrophy.

    Science.gov (United States)

    Kimura, En; Han, Jay J; Li, Sheng; Fall, Brent; Ra, Jennifer; Haraguchi, Miki; Tapscott, Stephen J; Chamberlain, Jeffrey S

    2008-08-15

    Duchenne muscular dystrophy (DMD) is characterized in skeletal muscle by cycles of myofiber necrosis and regeneration leading to loss of muscle fibers and replacement with fibrotic connective and adipose tissue. The ongoing activation and recruitment of muscle satellite cells for myofiber regeneration results in loss of regenerative capacity in part due to proliferative senescence. We explored a method whereby new myoblasts could be generated in dystrophic muscles by transplantation of primary fibroblasts engineered to express a micro-dystrophin/enhanced green fluorescent protein (muDys/eGFP) fusion gene together with a tamoxifen-inducible form of the myogenic regulator MyoD [MyoD-ER(T)]. Fibroblasts isolated from mdx(4cv) mice, a mouse model for DMD, were efficiently transduced with lentiviral vectors expressing muDys/eGFP and MyoD-ER(T) and underwent myogenic conversion when exposed to tamoxifen. These cells could also be induced to differentiate into muDys/eGFP-expressing myocytes and myotubes. Transplantation of transduced mdx(4cv) fibroblasts into mdx(4cv) muscles enabled tamoxifen-dependent regeneration of myofibers that express muDys. This lineage control method therefore allows replenishment of myogenic stem cells using autologous fibroblasts carrying an exogenous dystrophin gene. This strategy carries several potential advantages over conventional myoblast transplantation methods including: (i) the relative simplicity of culturing fibroblasts compared with myoblasts, (ii) a readily available cell source and ease of expansion and (iii) the ability to induce MyoD gene expression in vivo via administration of a medication. Our study provides a proof of concept for a novel gene/stem cell therapy technique and opens another potential therapeutic approach for degenerative muscle disorders. PMID:18511457

  12. New Functional Signatures for Understanding Melanoma Biology from Tumor Cell Lineage-Specific Analysis

    Directory of Open Access Journals (Sweden)

    Florian Rambow

    2015-10-01

    Full Text Available Molecular signatures specific to particular tumor types are required to design treatments for resistant tumors. However, it remains unclear whether tumors and corresponding cell lines used for drug development share such signatures. We developed similarity core analysis (SCA, a universal and unsupervised computational framework for extracting core molecular features common to tumors and cell lines. We applied SCA to mRNA/miRNA expression data from various sources, comparing melanoma cell lines and metastases. The signature obtained was associated with phenotypic characteristics in vitro, and the core genes CAPN3 and TRIM63 were implicated in melanoma cell migration/invasion. About 90% of the melanoma signature genes belong to an intrinsic network of transcription factors governing neural development (TFAP2A, DLX2, ALX1, MITF, PAX3, SOX10, LEF1, and GAS7 and miRNAs (211-5p, 221-3p, and 10a-5p. The SCA signature effectively discriminated between two subpopulations of melanoma patients differing in overall survival, and classified MEKi/BRAFi-resistant and -sensitive melanoma cell lines.

  13. Dissecting the molecular signatures of apical cell-type shoot meristems from two ancient land plant lineages.

    Science.gov (United States)

    Frank, Margaret H; Edwards, Molly B; Schultz, Eric R; McKain, Michael R; Fei, Zhangjun; Sørensen, Iben; Rose, Jocelyn K C; Scanlon, Michael J

    2015-08-01

    Shoot apical meristem (SAM) structure varies markedly within the land plants. The SAMs of many seedless vascular plants contain a conspicuous inverted, pyramidal cell called the apical cell (AC), which is unidentified in angiosperms. In this study, we use transcriptomic sequencing with precise laser microdissections of meristem subdomains to define the molecular signatures of anatomically distinct zones from the AC-type SAMs of a lycophyte (Selaginella moellendorffii) and a monilophyte (Equisetum arvense). The two model species for this study represent vascular plant lineages that diverged > 400 million yr ago. Our data comprise comprehensive molecular signatures for the distinct subdomains within AC-type SAMs, an anatomical anomaly whose functional significance has been debated in the botanical literature for over two centuries. Moreover, our data provide molecular support for distinct gene expression programs between the AC-type SAMs of Selaginella and Equisetum, as compared with the SAM transcriptome of the angiosperm maize. The results are discussed in light of the functional significance and evolutionary success of the AC-type SAM within the embryophytes. PMID:25900772

  14. The "Yin" and "Yang" of Cell Cycle Progression and Differentiation in the Oligodendroglial Lineage

    Science.gov (United States)

    Nguyen, Laurent; Borgs, Laurence; Vandenbosch, Renaud; Mangin, Jean-Marie; Beukelaers, Pierre; Moonen, Gustave; Gallo, Vittorio; Malgrange, Brigitte; Belachew, Shibeshih

    2006-01-01

    In white matter disorders such as leukodystrophies (LD), periventricular leucomalacia (PVL), or multiple sclerosis (MS), the hypomyelination or the remyelination failure by oligodendrocyte progenitor cells involves errors in the sequence of events that normally occur during development when progenitors proliferate, migrate through the white…

  15. Of lineage and legacy: The development of mammalian hematopoietic stem cells

    NARCIS (Netherlands)

    E.A. Dzierzak (Elaine); N.A. Speck (Nancy)

    2008-01-01

    textabstractThe hematopoietic system is one of the first complex tissues to develop in the mammalian conceptus. Of particular interest in the field of developmental hematopoiesis is the origin of adult bone marrow hematopoietic stem cells. Tracing their origin is complicated because blood is a mobil

  16. CD19 CAR immune pressure induces B-precursor acute lymphoblastic leukaemia lineage switch exposing inherent leukaemic plasticity.

    Science.gov (United States)

    Jacoby, Elad; Nguyen, Sang M; Fountaine, Thomas J; Welp, Kathryn; Gryder, Berkley; Qin, Haiying; Yang, Yinmeng; Chien, Christopher D; Seif, Alix E; Lei, Haiyan; Song, Young K; Khan, Javed; Lee, Daniel W; Mackall, Crystal L; Gardner, Rebecca A; Jensen, Michael C; Shern, Jack F; Fry, Terry J

    2016-01-01

    Adoptive immunotherapy using chimeric antigen receptor (CAR) expressing T cells targeting the CD19 B lineage receptor has demonstrated marked success in relapsed pre-B-cell acute lymphoblastic leukaemia (ALL). Persisting CAR-T cells generate sustained pressure against CD19 that may drive unique mechanisms of resistance. Pre-B ALL originates from a committed pre-B cell or an earlier progenitor, with potential to reprogram into other hematopoietic lineages. Here we report changes in lineage markers including myeloid conversion in patients following CD19 CAR therapy. Using murine ALL models we study the long-term effects of CD19 CAR-T cells and demonstrate partial or complete lineage switch as a consistent mechanism of CAR resistance depending on the underlying genetic oncogenic driver. Deletion of Pax5 or Ebf1 recapitulates lineage reprogramming occurring during CD19 CAR pressure. Our findings establish lineage switch as a mechanism of CAR resistance exposing inherent plasticity in genetic subtypes of pre-B-cell ALL. PMID:27460500

  17. Angiogenic CXC chemokine expression during differentiation of human mesenchymal stem cells towards the osteoblastic lineage.

    Science.gov (United States)

    Bischoff, D S; Zhu, J H; Makhijani, N S; Kumar, A; Yamaguchi, D T

    2008-02-15

    The potential role of ELR(+) CXC chemokines in early events in bone repair was studied using human mesenchymal stem cells (hMSCs). Inflammation, which occurs in the initial phase of tissue healing in general, is critical to bone repair. Release of cytokines from infiltrating immune cells and injured bone can lead to recruitment of MSCs to the region of repair. CXC chemokines bearing the Glu-Leu-Arg (ELR) motif are also released by inflammatory cells and serve as angiogenic factors stimulating chemotaxis and proliferation of endothelial cells. hMSCs, induced to differentiate with osteogenic medium (OGM) containing ascorbate, beta-glycerophosphate (beta-GP), and dexamethasone (DEX), showed an increase in mRNA and protein secretion of the ELR(+) CXC chemokines CXCL8 and CXCL1. CXCL8 mRNA half-life studies reveal an increase in mRNA stability upon OGM stimulation. Increased expression and secretion is a result of DEX in OGM and is dose-dependent. Inhibition of the glucocorticoid receptor with mifepristone only partially inhibits DEX-stimulated CXCL8 expression indicating both glucocorticoid receptor dependent and independent pathways. Treatment with signal transduction inhibitors demonstrate that this expression is due to activation of the ERK and p38 mitogen-activated protein kinase (MAPK) pathways and is mediated through the G(alphai)-coupled receptors. Angiogenesis assays demonstrate that OGM-stimulated conditioned media containing secreted CXCL8 and CXCL1 can induce angiogenesis of human microvascular endothelial cells in an in vitro Matrigel assay. PMID:17583554

  18. A CRISPR/Cas9-Based System for Reprogramming Cell Lineage Specification

    Directory of Open Access Journals (Sweden)

    Syandan Chakraborty

    2014-12-01

    Full Text Available Gene activation by the CRISPR/Cas9 system has the potential to enable new approaches to science and medicine, but the technology must be enhanced to robustly control cell behavior. We show that the fusion of two transactivation domains to Cas9 dramatically enhances gene activation to a level that is necessary to reprogram cell phenotype. Targeted activation of the endogenous Myod1 gene locus with this system led to stable and sustained reprogramming of mouse embryonic fibroblasts into skeletal myocytes. The levels of myogenic marker expression obtained by the activation of endogenous Myod1 gene were comparable to that achieved by overexpression of lentivirally delivered MYOD1 transcription factor.

  19. Evaluation of hela cell lineage response to β radiation from Holmium-166 embedded in ceramic seeds

    Directory of Open Access Journals (Sweden)

    Eduardo Sarmento Valente

    2011-10-01

    Full Text Available This work studied the effects of β radiation of Ho-166 embedded in ceramic seeds on HeLa cells. Methodology consisted in the production of ceramic seeds with holmium-165 by sol-gel route. Chemical and physical characterizations of the seeds were performed. Subsequently, nuclear characterization was performed by gamma spectrometry. Experimental and theoretical activities were defined and initial dose rate were evaluated by MIRD (Medical Internal Radiation Dose Committee methodology. The seeds were placed in confluent culture flasks and remained for six radionuclide half-lives. Biological results were represented by a clean 6 mm diameter area around the seed where the tumour cells were killed. The initial dose rate was 15.5 Gy. h-1. The maximum absorbed dose was 591.3 Gy. The features of the Ho-166 seeds suggested that such ceramic seeds were suitable for high dose rate brachytherapy.

  20. Characterization of an individual neural crest-like cell lineage in the invertebrate chordate Ciona intestinalis

    OpenAIRE

    Cone, Angela C.

    2008-01-01

    During embryogenesis, all chordate embryos undergo neurulation to form a dorsal, hollow nerve cord. Neural crest cells (NCC), considered a vertebrate innovation, arise during neurulation and later differentiate into a multitude of tissues that account for much of the structural complexity that distinguishes craniates from invertebrate chordates [1, 2]. NCCs are induced and specified at the border of the neural and non-neural ectoderm by a complex network of inductive signals and transcription...

  1. Endoderm lineage labelling using BAC reporter constructs in ES cells and mouse embryos

    OpenAIRE

    Imhof, Sascha

    2009-01-01

    The endoderm is one of the three embryonic germ layers that gives rise to inner organs like lung, liver and pancreas. BAC recombineering was used to generate transgenic embryonic stem cells and mouse lines containing fluorescent reporters under the control of important endodermal genes like Foxa2, Sox17, Hhex, Nkx2.1 and Pdx1. This allowed the analysis of the specific expression patterns of those genes during endoderm and organ formation. In particular, the analysis of the Hhex reporter expre...

  2. Clonal and Lineage Analysis of Melanocyte Stem Cells and Their Progeny in the Zebrafish

    OpenAIRE

    Tryon, Robert C.; Johnson, Stephen L

    2012-01-01

    The study of melanocyte biology in the zebrafish presents a highly tractable system for understanding fundamental principles of developmental biology. Melanocytes are visible in the transparent embryo and in the mature fish following metamorphosis, a physical transformation from the larval to adult form. While early developing larval melanocytes are direct derivatives of the neural crest, the remainder of melanocytes develop from unpigmented precursors, or melanocyte stem cells (MSCs). The To...

  3. E47 regulates hematopoietic stem cell proliferation and energetics but not myeloid lineage restriction

    OpenAIRE

    Yang, Qi; Esplin, Brandt; Borghesi, Lisa

    2011-01-01

    The immune system is replenished by self-renewing hematopoietic stem cells (HSCs) that produce multipotent progenitors (MPPs) with little renewal capacity. E-proteins, the widely expressed basic helix-loop-helix transcription factors, contribute to HSC and MPP activity, but their specific functions remain undefined. Using quantitative in vivo and in vitro approaches, we show that E47 is dispensable for the short-term myeloid differentiation of HSCs but regulates their long-term capabilities. ...

  4. Nuclear receptor steroidogenic factor 1 directs embryonic stem cells toward the steroidogenic lineage.

    OpenAIRE

    Crawford, P A; Sadovsky, Y.; Milbrandt, J

    1997-01-01

    The orphan nuclear receptor steroidogenic factor 1 (SF-1) is expressed in the adrenal gland and gonads and is an important regulator of the expression of cytochrome P-450 steroidogenic enzymes in cultured cells. Targeted disruption of the SF-1 gene in mice shows that it is a critical participant in the genetic program that promotes the development of urogenital mesoderm into the adrenal gland and gonads. To assess the ability of SF-1 to regulate this differentiation pathway, we ectopically ex...

  5. More efficient repair of DNA double-strand breaks in skeletal muscle stem cells compared to their committed progeny

    Directory of Open Access Journals (Sweden)

    Leyla Vahidi Ferdousi

    2014-11-01

    Full Text Available The loss of genome integrity in adult stem cells results in accelerated tissue aging and is possibly cancerogenic. Adult stem cells in different tissues appear to react robustly to DNA damage. We report that adult skeletal stem (satellite cells do not primarily respond to radiation-induced DNA double-strand breaks (DSBs via differentiation and exhibit less apoptosis compared to other myogenic cells. Satellite cells repair these DNA lesions more efficiently than their committed progeny. Importantly, non-proliferating satellite cells and post-mitotic nuclei in the fiber exhibit dramatically distinct repair efficiencies. Altogether, reduction of the repair capacity appears to be more a function of differentiation than of the proliferation status of the muscle cell. Notably, satellite cells retain a high efficiency of DSB repair also when isolated from the natural niche. Finally, we show that repair of DSB substrates is not only very efficient but, surprisingly, also very accurate in satellite cells and that accurate repair depends on the key non-homologous end-joining factor DNA-PKcs.

  6. The gene for erythropoietin receptor is expressed in multipotential hematopoietic and embryonal stem cells: evidence for differentiation stage-specific regulation.

    OpenAIRE

    Heberlein, C; Fischer, K D; Stoffel, M; Nowock, J; Ford, A.; Tessmer, U.; Stocking, C

    1992-01-01

    The principal regulator of erythropoiesis is the glycoprotein erythropoietin, which interacts with a specific cell surface receptor (EpoR). A study aimed at analyzing EpoR gene regulation has shown that both pluripotent embryonal stem cells and early multipotent hematopoietic cells express EpoR transcripts. Commitment to nonerythroid lineages (e.g., macrophage or lymphocytic) results in the shutdown of EpoR gene expression, whereas commitment to the erythroid lineage is concurrent with or fol...

  7. Analysis of folylpoly-γ-glutamate synthetase gene expression in human B-precursor ALL and T-lineage ALL cells

    Directory of Open Access Journals (Sweden)

    Barredo Julio C

    2006-05-01

    Full Text Available Abstract Background Expression of folylpoly-γ-glutamate synthetase (FPGS gene is two- to three-fold higher in B-precursor ALL (Bp- ALL than in T-lineage ALL (T-ALL and correlates with intracellular accumulation of methotrexate (MTX polyglutamates and lymphoblast sensitivity to MTX. In this report, we investigated the molecular regulatory mechanisms directing FPGS gene expression in Bp-ALL and T-ALL cells. Methods To determine FPGS transcription rate in Bp-ALL and T-ALL we used nuclear run-on assays. 5'-RACE was used to uncover potential regulatory regions involved in the lineage differences. We developed a luciferase reporter gene assay to investigate FPGS promoter/enhancer activity. To further characterize the FPGS proximal promoter, we determined the role of the putative transcription binding sites NFY and E-box on FPGS expression using luciferase reporter gene assays with substitution mutants and EMSA. Results FPGS transcription initiation rate was 1.6-fold higher in NALM6 vs. CCRF-CEM cells indicating that differences in transcription rate led to the observed lineage differences in FPGS expression between Bp-ALL and T-ALL blasts. Two major transcripts encoding the mitochondrial/cytosolic and cytosolic isoforms were detected in Bp-ALL (NALM6 and REH whereas in T-ALL (CCRF-CEM cells only the mitochondrial/cytosolic transcript was detected. In all DNA fragments examined for promoter/enhancer activity, we measured significantly lower luciferase activity in NALM6 vs. CCRF-CEM cells, suggesting the need for additional yet unidentified regulatory elements in Bp-ALL. Finally, we determined that the putative transcription factor binding site NFY, but not E-box, plays a role in FPGS transcription in both Bp- and T-lineage. Conclusion We demonstrated that the minimal FPGS promoter region previously described in CCRF-CEM is not sufficient to effectively drive FPGS transcription in NALM6 cells, suggesting that different regulatory elements are required

  8. Analysis of folylpoly-γ-glutamate synthetase gene expression in human B-precursor ALL and T-lineage ALL cells

    International Nuclear Information System (INIS)

    Expression of folylpoly-γ-glutamate synthetase (FPGS) gene is two- to three-fold higher in B-precursor ALL (Bp- ALL) than in T-lineage ALL (T-ALL) and correlates with intracellular accumulation of methotrexate (MTX) polyglutamates and lymphoblast sensitivity to MTX. In this report, we investigated the molecular regulatory mechanisms directing FPGS gene expression in Bp-ALL and T-ALL cells. To determine FPGS transcription rate in Bp-ALL and T-ALL we used nuclear run-on assays. 5'-RACE was used to uncover potential regulatory regions involved in the lineage differences. We developed a luciferase reporter gene assay to investigate FPGS promoter/enhancer activity. To further characterize the FPGS proximal promoter, we determined the role of the putative transcription binding sites NFY and E-box on FPGS expression using luciferase reporter gene assays with substitution mutants and EMSA. FPGS transcription initiation rate was 1.6-fold higher in NALM6 vs. CCRF-CEM cells indicating that differences in transcription rate led to the observed lineage differences in FPGS expression between Bp-ALL and T-ALL blasts. Two major transcripts encoding the mitochondrial/cytosolic and cytosolic isoforms were detected in Bp-ALL (NALM6 and REH) whereas in T-ALL (CCRF-CEM) cells only the mitochondrial/cytosolic transcript was detected. In all DNA fragments examined for promoter/enhancer activity, we measured significantly lower luciferase activity in NALM6 vs. CCRF-CEM cells, suggesting the need for additional yet unidentified regulatory elements in Bp-ALL. Finally, we determined that the putative transcription factor binding site NFY, but not E-box, plays a role in FPGS transcription in both Bp- and T-lineage. We demonstrated that the minimal FPGS promoter region previously described in CCRF-CEM is not sufficient to effectively drive FPGS transcription in NALM6 cells, suggesting that different regulatory elements are required for FPGS gene expression in Bp-cells. Our data indicate

  9. Loss of IKKβ but Not NF-κB p65 Skews Differentiation towards Myeloid over Erythroid Commitment and Increases Myeloid Progenitor Self-Renewal and Functional Long-Term Hematopoietic Stem Cells.

    Science.gov (United States)

    Zhang, Jing; Li, Li; Baldwin, Albert S; Friedman, Alan D; Paz-Priel, Ido

    2015-01-01

    NF-κB is an important regulator of both differentiation and function of lineage-committed hematopoietic cells. Targeted deletion of IκB kinase (IKK) β results in altered cytokine signaling and marked neutrophilia. To investigate the role of IKKβ in regulation of hematopoiesis, we employed Mx1-Cre mediated IKKβ conditional knockout mice. As previously reported, deletion of IKKβ in hematopoietic cells results in neutrophilia, and we now also noted decreased monocytes and modest anemia. Granulocyte-macrophage progenitors (GMPs) accumulated markedly in bone marrow of IKKβ deleted mice whereas the proportion and number of megakaryocyte-erythrocyte progenitors (MEP) decreased. Accordingly, we found a significantly reduced frequency of proerythroblasts and basophilic and polychromatic erythroblasts, and IKKβ-deficient bone marrow cells yielded a significantly decreased number of BFU-E compared to wild type. These changes are associated with elevated expression of C/EBPα, Gfi1, and PU.1 and diminished Gata1, Klf1, and SCL/Tal1 in IKKβ deficient Lineage-Sca1+c-Kit+ (LSK) cells. In contrast, no effect on erythropoiesis or expression of lineage-related transcription factors was found in marrow lacking NF-κB p65. Bone marrow from IKKβ knockout mice has elevated numbers of phenotypic long and short term hematopoietic stem cells (HSC). A similar increase was observed when IKKβ was deleted after marrow transplantation into a wild type host, indicating cell autonomous expansion. Myeloid progenitors from IKKβ- but not p65-deleted mice demonstrate increased serial replating in colony-forming assays, indicating increased cell autonomous self-renewal capacity. In addition, in a competitive repopulation assay deletion of IKKβ resulted in a stable advantage of bone marrow derived from IKKβ knockout mice. In summary, loss of IKKβ resulted in significant effects on hematopoiesis not seen upon NF-κB p65 deletion. These include increased myeloid and reduced erythroid

  10. Thymic and Postthymic Regulation of Naïve CD4+ T-Cell Lineage Fates in Humans and Mice Models

    Directory of Open Access Journals (Sweden)

    José E. Belizário

    2016-01-01

    Full Text Available Our understanding of how thymocytes differentiate into many subtypes has been increased progressively in its complexity. At early life, the thymus provides a suitable microenvironment with specific combination of stromal cells, growth factors, cytokines, and chemokines to induce the bone marrow lymphoid progenitor T-cell precursors into single-positive CD4+ and CD8+ T effectors and CD4+CD25+ T-regulatory cells (Tregs. At postthymic compartments, the CD4+ T-cells acquire distinct phenotypes which include the classical T-helper 1 (Th1, T-helper 2 (Th2, T-helper 9 (Th9, T-helper 17 (Th17, follicular helper T-cell (Tfh, and induced T-regulatory cells (iTregs, such as the regulatory type 1 cells (Tr1 and transforming growth factor-β- (TGF-β- producing CD4+ T-cells (Th3. Tregs represent only a small fraction, 5–10% in mice and 1-2% in humans, of the overall CD4+ T-cells in lymphoid tissues but are essential for immunoregulatory circuits mediating the inhibition and expansion of all lineages of T-cells. In this paper, we first provide an overview of the major cell-intrinsic developmental programs that regulate T-cell lineage fates in thymus and periphery. Next, we introduce the SV40 immortomouse as a relevant mice model for implementation of new approaches to investigate thymus organogenesis, CD4 and CD8 development, and thymus cells tumorogenesis.

  11. Thymic and Postthymic Regulation of Naïve CD4+ T-Cell Lineage Fates in Humans and Mice Models

    Science.gov (United States)

    Belizário, José E.; Brandão, Wesley; Rossato, Cristiano; Peron, Jean Pierre

    2016-01-01

    Our understanding of how thymocytes differentiate into many subtypes has been increased progressively in its complexity. At early life, the thymus provides a suitable microenvironment with specific combination of stromal cells, growth factors, cytokines, and chemokines to induce the bone marrow lymphoid progenitor T-cell precursors into single-positive CD4+ and CD8+ T effectors and CD4+CD25+ T-regulatory cells (Tregs). At postthymic compartments, the CD4+ T-cells acquire distinct phenotypes which include the classical T-helper 1 (Th1), T-helper 2 (Th2), T-helper 9 (Th9), T-helper 17 (Th17), follicular helper T-cell (Tfh), and induced T-regulatory cells (iTregs), such as the regulatory type 1 cells (Tr1) and transforming growth factor-β- (TGF-β-) producing CD4+ T-cells (Th3). Tregs represent only a small fraction, 5–10% in mice and 1-2% in humans, of the overall CD4+ T-cells in lymphoid tissues but are essential for immunoregulatory circuits mediating the inhibition and expansion of all lineages of T-cells. In this paper, we first provide an overview of the major cell-intrinsic developmental programs that regulate T-cell lineage fates in thymus and periphery. Next, we introduce the SV40 immortomouse as a relevant mice model for implementation of new approaches to investigate thymus organogenesis, CD4 and CD8 development, and thymus cells tumorogenesis.

  12. Caspase-dependent inhibition of store-operated Ca{sup 2+} entry into apoptosis-committed Jurkat cells

    Energy Technology Data Exchange (ETDEWEB)

    Onopiuk, Marta; Wierzbicka, Katarzyna; Brutkowski, Wojciech; Szczepanowska, Joanna [Department of Biochemistry, The Nencki Institute of Experimental Biology, Warsaw (Poland); Zablocki, Krzysztof, E-mail: k.zablocki@nencki.gov.pl [Department of Biochemistry, The Nencki Institute of Experimental Biology, Warsaw (Poland)

    2010-08-20

    Activation of T-cells triggers store-operated Ca{sup 2+} entry, which begins a signaling cascade leading to induction of appropriate gene expression and eventually lymphocyte proliferation and differentiation. The simultaneous enhancement of Fas ligand gene expression in activated cells allows the immune response to be limited by committing the activated cells to apoptosis. In apoptotic cells the store-operated calcium entry is significantly inhibited. It has been documented that moderate activation of Fas receptor may cause reversible inhibition of store-operated channels by ceramide released from hydrolyzed sphingomyelin. Here we show that activation of Fas receptor in T-cells results in caspase-dependent decrease of cellular STIM1 and Orai1 protein content. This effect may be responsible for the substantial inhibition of Ca{sup 2+} entry into Jurkat cells undergoing apoptosis. In turn, this inhibition might prevent overloading of cells with calcium and protect them against necrosis. -- Research highlights: {yields} Fas activation reduces STIM1 and Orai1 protein content in caspase dependent manner. {yields} Fas activation partially reduces mitochondrial potential in caspase dependent manner. {yields} Fas stimulation inhibits of store-operated Ca{sup 2+} entry in caspase dependent manner. {yields} Inhibition of Ca{sup 2+} entry in apoptotic cells may protect them from secondary necrosis.

  13. Reassortment compatibility between PB1, PB2, and HA genes of the two influenza B virus lineages in mammalian cells

    Science.gov (United States)

    Kim, Jin Il; Lee, Ilseob; Park, Sehee; Bae, Joon-Yong; Yoo, Kirim; Lemey, Philippe; Park, Mee Sook; Song, Jin-Won; Kee, Sun-Ho; Song, Ki-Joon; Park, Man-Seong

    2016-01-01

    In addition to influenza A subtypes, two distinct lineages of influenza B virus also cause seasonal epidemics to humans. Recently, Dudas et al. have done evolutionary analyses of reassortment patterns of the virus and suggested genetic lineage relationship between PB1, PB2, and HA genes. Using genetic plasmids and reassortant viruses, we here demonstrate that a homologous lineage PB1-PB2 pair exhibits better compatibility than a heterologous one and that the lineage relationship between PB1 and HA is more important for viral replication than that between PB2 and HA. However, co-adaptation of PB1-PB2-HA genes appears to be affected by complete gene constellation. PMID:27270757

  14. MiR-124 is differentially expressed in derivatives of the sympathoadrenal cell lineage and promotes neurite elongation in chromaffin cells.

    Science.gov (United States)

    Shtukmaster, Stella; Narasimhan, Priyanka; El Faitwri, Tehani; Stubbusch, Jutta; Ernsberger, Uwe; Rohrer, Hermann; Unsicker, Klaus; Huber, Katrin

    2016-08-01

    The neural-crest-derived sympathoadrenal cell lineage gives rise to sympathetic neurons and to endocrine chromaffin cells of the adrenal medulla. Both cell types express a largely overlapping set of genes, including those coding for the molecular machinery related to the synthesis and exocytotic release of catecholamines. During their early development, sympathetic neurons and chromaffin cells rely on a shared transcription factor network that controls the establishment of these common features. Despite many similarities, mature sympathetic neurons and chromaffin cells significantly differ regarding their morphology and function. Most prominently, sympathetic neurons possess axons that are absent in mammalian adrenal chromaffin cells. The molecular mechanism underlying the divergent development of sympathoadrenal cells into neuronal and endocrine cells remains elusive. Mutational inactivation of the ribonuclease dicer hints at the importance of microRNAs in this diversification. We show here that miR-124 is detectable in developing sympathetic neurons but absent in chromaffin cell precursors. We further demonstrate that miR-124 promotes neurite elongation when transfected into cultured chromaffin cells indicating its capability to support the establishment of a neuronal morphology in non-neuronal sympathoadrenal cells. Our results also show that treatment of PC12 cells with the neurotrophin nerve growth factor leads to an upregulation of miR-124 expression and that inhibition of miR-124 reduces nerve-growth-factor-induced neurite outgrowth in PC12 cells. Thus, our data indicate that miR-124 contributes to the establishment of specific neuronal features in developing sympathoadrenal cells. PMID:27094431

  15. Deficiency of the ribosome biogenesis gene Sbds in hematopoietic stem and progenitor cells causes neutropenia in mice by attenuating lineage progression in myelocytes.

    Science.gov (United States)

    Zambetti, Noemi A; Bindels, Eric M J; Van Strien, Paulina M H; Valkhof, Marijke G; Adisty, Maria N; Hoogenboezem, Remco M; Sanders, Mathijs A; Rommens, Johanna M; Touw, Ivo P; Raaijmakers, Marc H G P

    2015-10-01

    Shwachman-Diamond syndrome is a congenital bone marrow failure disorder characterized by debilitating neutropenia. The disease is associated with loss-of-function mutations in the SBDS gene, implicated in ribosome biogenesis, but the cellular and molecular events driving cell specific phenotypes in ribosomopathies remain poorly defined. Here, we established what is to our knowledge the first mammalian model of neutropenia in Shwachman-Diamond syndrome through targeted downregulation of Sbds in hematopoietic stem and progenitor cells expressing the myeloid transcription factor CCAAT/enhancer binding protein α (Cebpa). Sbds deficiency in the myeloid lineage specifically affected myelocytes and their downstream progeny while, unexpectedly, it was well tolerated by rapidly cycling hematopoietic progenitor cells. Molecular insights provided by massive parallel sequencing supported cellular observations of impaired cell cycle exit and formation of secondary granules associated with the defect of myeloid lineage progression in myelocytes. Mechanistically, Sbds deficiency activated the p53 tumor suppressor pathway and induced apoptosis in these cells. Collectively, the data reveal a previously unanticipated, selective dependency of myelocytes and downstream progeny, but not rapidly cycling progenitors, on this ubiquitous ribosome biogenesis protein, thus providing a cellular basis for the understanding of myeloid lineage biased defects in Shwachman-Diamond syndrome. PMID:26185170

  16. Multiple Lineages of Human Breast Cancer Stem/Progenitor Cells Identified by Profiling with Stem Cell Markers

    OpenAIRE

    Hwang-Verslues, Wendy W.; Wen-Hung Kuo; Po-Hao Chang; Chi-Chun Pan; Hsing-Hui Wang; Sheng-Ta Tsai; Yung-Ming Jeng; Jin-Yu Shew; Kung, John T.; Chung-Hsuan Chen; Lee, Eva Y-H. P.; King-Jen Chang; Wen-Hwa Lee

    2009-01-01

    Heterogeneity of cancer stem/progenitor cells that give rise to different forms of cancer has been well demonstrated for leukemia. However, this fundamental concept has yet to be established for solid tumors including breast cancer. In this communication, we analyzed solid tumor cancer stem cell markers in human breast cancer cell lines and primary specimens using flow cytometry. The stem/progenitor cell properties of different marker expressing-cell populations were further assessed by in vi...

  17. OxyR-dependent formation of DNA methylation patterns in OpvABOFF and OpvABON cell lineages of Salmonella enterica.

    Science.gov (United States)

    Cota, Ignacio; Bunk, Boyke; Spröer, Cathrin; Overmann, Jörg; König, Christoph; Casadesús, Josep

    2016-05-01

    Phase variation of the Salmonella enterica opvAB operon generates a bacterial lineage with standard lipopolysaccharide structure (OpvAB(OFF)) and a lineage with shorter O-antigen chains (OpvAB(ON)). Regulation of OpvAB lineage formation is transcriptional, and is controlled by the LysR-type factor OxyR and by DNA adenine methylation. The opvAB regulatory region contains four sites for OxyR binding (OBSA-D), and four methylatable GATC motifs (GATC1-4). OpvAB(OFF) and OpvAB(ON) cell lineages display opposite DNA methylation patterns in the opvAB regulatory region: (i) in the OpvAB(OFF) state, GATC1 and GATC3 are non-methylated, whereas GATC2 and GATC4 are methylated; (ii) in the OpvAB(ON) state, GATC2 and GATC4 are non-methylated, whereas GATC1 and GATC3 are methylated. We provide evidence that such DNA methylation patterns are generated by OxyR binding. The higher stability of the OpvAB(OFF) lineage may be caused by binding of OxyR to sites that are identical to the consensus (OBSA and OBSc), while the sites bound by OxyR in OpvAB(ON) cells (OBSB and OBSD) are not. In support of this view, amelioration of either OBSB or OBSD locks the system in the ON state. We also show that the GATC-binding protein SeqA and the nucleoid protein HU are ancillary factors in opvAB control. PMID:26687718

  18. Multiple lineages of human breast cancer stem/progenitor cells identified by profiling with stem cell markers.

    Directory of Open Access Journals (Sweden)

    Wendy W Hwang-Verslues

    Full Text Available Heterogeneity of cancer stem/progenitor cells that give rise to different forms of cancer has been well demonstrated for leukemia. However, this fundamental concept has yet to be established for solid tumors including breast cancer. In this communication, we analyzed solid tumor cancer stem cell markers in human breast cancer cell lines and primary specimens using flow cytometry. The stem/progenitor cell properties of different marker expressing-cell populations were further assessed by in vitro soft agar colony formation assay and the ability to form tumors in NOD/SCID mice. We found that the expression of stem cell markers varied greatly among breast cancer cell lines. In MDA-MB-231 cells, PROCR and ESA, instead of the widely used breast cancer stem cell markers CD44(+/CD24(-/low and ALDH, could be used to highly enrich cancer stem/progenitor cell populations which exhibited the ability to self renew and divide asymmetrically. Furthermore, the PROCR(+/ESA(+ cells expressed epithelial-mesenchymal transition markers. PROCR could also be used to enrich cells with colony forming ability from MB-361 cells. Moreover, consistent with the marker profiling using cell lines, the expression of stem cell markers differed greatly among primary tumors. There was an association between metastasis status and a high prevalence of certain markers including CD44(+/CD24(-/low, ESA(+, CD133(+, CXCR4(+ and PROCR(+ in primary tumor cells. Taken together, these results suggest that similar to leukemia, several stem/progenitor cell-like subpopulations can exist in breast cancer.

  19. A New Avenue to Cure Cancer by Turning Adaptive Immune T Cells to Innate Immune NK Cells via Reprogramming

    Institute of Scientific and Technical Information of China (English)

    Dong-Ming Su; Ramakrishna Vankayalapati

    2010-01-01

    Thymocytes after T-lineage commitment develop in the T-cell pathway. However, in a recent study, Li et al. (2010) demonstrated that inducing to delete Bcl11b gene in these thymocytes, even in mature T cells turns these cells into natural killer (NK) cells during the culture. They called this conversion 'reprogramming', and the reprogrammed killer cells 'ITNK cells'.

  20. Green tea epigallocatechin-3-gallate modulates differentiation of naive CD4+ T cells into specific lineage effector cells

    Science.gov (United States)

    CD4+ T helper (Th) subsets Th1, Th9, and Th17 cells are implicated in inducing autoimmunity whereas regulatory T cells (Treg) have a protective effect. We previously showed that epigallocatechin-3-gallate (EGCG) attenuated experimental autoimmune encephalomyelitis (EAE) and altered CD4+ T cell subpo...

  1. The myocardial regenerative potential of three-dimensional engineered cardiac tissues composed of multiple human iPS cell-derived cardiovascular cell lineages.

    Science.gov (United States)

    Masumoto, Hidetoshi; Nakane, Takeichiro; Tinney, Joseph P; Yuan, Fangping; Ye, Fei; Kowalski, William J; Minakata, Kenji; Sakata, Ryuzo; Yamashita, Jun K; Keller, Bradley B

    2016-01-01

    Human induced pluripotent stem cells (hiPSCs) are a robust source for cardiac regenerative therapy due to their potential to support autologous and allogeneic transplant paradigms. The in vitro generation of three-dimensional myocardial tissue constructs using biomaterials as an implantable hiPSC-derived myocardium provides a path to realize sustainable myocardial regeneration. We generated engineered cardiac tissues (ECTs) from three cellular compositions of cardiomyocytes (CMs), endothelial cells (ECs), and vascular mural cells (MCs) differentiated from hiPSCs. We then determined the impact of cell composition on ECT structural and functional properties. In vitro force measurement showed that CM+EC+MC ECTs possessed preferential electromechanical properties versus ECTs without vascular cells indicating that incorporation of vascular cells augmented tissue maturation and function. The inclusion of MCs facilitated more mature CM sarcomeric structure, preferential alignment, and activated multiple tissue maturation pathways. The CM+EC+MC ECTs implanted onto infarcted, immune tolerant rat hearts engrafted, displayed both host and graft-derived vasculature, and ameliorated myocardial dysfunction. Thus, a composition of CMs and multiple vascular lineages derived from hiPSCs and incorporated into ECTs promotes functional maturation and demonstrates myocardial replacement and perfusion relevant for clinical translation. PMID:27435115

  2. Single-cell sequencing analysis characterizes common and cell-lineage-specific mutations in a muscle-invasive bladder cancer

    DEFF Research Database (Denmark)

    Li, Yingrui; Xu, Xun; Song, Luting;

    2012-01-01

    sequencing of 66 individual tumor cells from a muscle-invasive bladder transitional cell carcinoma (TCC). Analyses of the somatic mutant allele frequency spectrum and clonal structure revealed that the tumor cells were derived from a single ancestral cell, but that subsequent evolution occurred, leading to...... two distinct tumor cell subpopulations. By analyzing recurrently mutant genes in an additional cohort of 99 TCC tumors, we identified genes that might play roles in the maintenance of the ancestral clone and in the muscle-invasive capability of subclones of this bladder cancer, respectively...

  3. The cholesterol-binding protein NPC2 restrains recruitment of stromal macrophage-lineage cells to early-stage lung tumours

    OpenAIRE

    Kamata, Tamihiro; Jin, Hong; Giblett, Susan; Patel, Bipin; Patel, Falguni; Foster, Charles; Pritchard, Catrin

    2015-01-01

    The tumour microenvironment is known to play an integral role in facilitating cancer progression at advanced stages, but its function in some pre-cancerous lesions remains elusive. We have used the V600 EBRAF-driven mouse lung model that develop premalignant lesions to understand stroma–tumour interactions during pre-cancerous development. In this model, we have found that immature macrophage-lineage cells (IMCs) producing PDGFA, TGFβ and CC chemokines are recruited to the stroma of premalign...

  4. A Gene Regulatory Network Cooperatively Controlled by Pdx1 and Sox9 Governs Lineage Allocation of Foregut Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Hung Ping Shih

    2015-10-01

    Full Text Available The generation of pancreas, liver, and intestine from a common pool of progenitors in the foregut endoderm requires the establishment of organ boundaries. How dorsal foregut progenitors activate pancreatic genes and evade the intestinal lineage choice remains unclear. Here, we identify Pdx1 and Sox9 as cooperative inducers of a gene regulatory network that distinguishes the pancreatic from the intestinal lineage. Genetic studies demonstrate dual and cooperative functions for Pdx1 and Sox9 in pancreatic lineage induction and repression of the intestinal lineage choice. Pdx1 and Sox9 bind to regulatory sequences near pancreatic and intestinal differentiation genes and jointly regulate their expression, revealing direct cooperative roles for Pdx1 and Sox9 in gene activation and repression. Our study identifies Pdx1 and Sox9 as important regulators of a transcription factor network that initiates pancreatic fate and sheds light on the gene regulatory circuitry that governs the development of distinct organs from multi-lineage-competent foregut progenitors.

  5. Nestin-positive mesenchymal stem cells favour the astroglial lineage in neural progenitors and stem cells by releasing active BMP4

    Directory of Open Access Journals (Sweden)

    Leprince Pierre

    2004-09-01

    Full Text Available Abstract Background Spontaneous repair is limited after CNS injury or degeneration because neurogenesis and axonal regrowth rarely occur in the adult brain. As a result, cell transplantation has raised much interest as potential treatment for patients with CNS lesions. Several types of cells have been considered as candidates for such cell transplantation and replacement therapies. Foetal brain tissue has already been shown to have significant effects in patients with Parkinson's disease. Clinical use of the foetal brain tissue is, however, limited by ethical and technical problems as it requires high numbers of grafted foetal cells and immunosuppression. Alternatively, several reports suggested that mesenchymal stem cells, isolated from adult bone marrow, are multipotent cells and could be used in autograft approach for replacement therapies. Results In this study, we addressed the question of the possible influence of mesenchymal stem cells on neural stem cell fate. We have previously reported that adult rat mesenchymal stem cells are able to express nestin in defined culture conditions (in the absence of serum and after 25 cell population doublings and we report here that nestin-positive (but not nestin-negative mesenchymal stem cells are able to favour the astroglial lineage in neural progenitors and stem cells cultivated from embryonic striatum. The increase of the number of GFAP-positive cells is associated with a significant decrease of the number of Tuj1- and O4-positive cells. Using quantitative RT-PCR, we demonstrate that mesenchymal stem cells express LIF, CNTF, BMP2 and BMP4 mRNAs, four cytokines known to play a role in astroglial fate decision. In this model, BMP4 is responsible for the astroglial stimulation and oligodendroglial inhibition, as 1 this cytokine is present in a biologically-active form only in nestin-positive mesenchymal stem cells conditioned medium and 2 anti-BMP4 antibodies inhibit the nestin-positive mesenchymal

  6. Lineage tracing in the adult mouse corneal epithelium supports the limbal epithelial stem cell hypothesis with intermittent periods of stem cell quiescence

    Directory of Open Access Journals (Sweden)

    Natalie J. Dorà

    2015-11-01

    Full Text Available The limbal epithelial stem cell (LESC hypothesis proposes that LESCs in the corneal limbus maintain the corneal epithelium both during normal homeostasis and wound repair. The alternative corneal epithelial stem cell (CESC hypothesis proposes that LESCs are only involved in wound repair and CESCs in the corneal epithelium itself maintain the corneal epithelium during normal homeostasis. We used tamoxifen-inducible, CreER-loxP lineage tracing to distinguish between these hypotheses. Clones of labelled cells were induced in adult CAGG-CreER;R26R-LacZ reporter mice and their distributions analysed after different chase periods. Short-lived clones, derived from labelled transient amplifying cells, were shed during the chase period and long-lived clones, derived from stem cells, expanded. At 6 weeks, labelled clones appeared at the periphery, extended centripetally as radial stripes and a few reached the centre by 14 weeks. Stripe numbers depended on the age of tamoxifen treatment. Stripes varied in length, some were discontinuous, few reached the centre and almost half had one end at the limbus. Similar stripes extended across the cornea in CAGG-CreER;R26R-mT/mG reporter mice. The distributions of labelled clones are inconsistent with the CESC hypothesis and support the LESC hypothesis if LESCs cycle between phases of activity and quiescence, each lasting several weeks.

  7. Regulatory T cell reprogramming towards a Th2 cell-like lineage impairs oral tolerance and promotes food allergy

    OpenAIRE

    Rivas, Magali Noval; Burton, Oliver T.; Wise, Petra; Charbonnier, Louis-Marie; Georgiev, Peter; Oettgen, Hans C.; Rachid, Rima; Chatila, Talal

    2015-01-01

    Oral immunotherapy has had limited success in establishing tolerance in food allergy, reflecting failure to elicit an effective regulatory T (Treg) cell response. We show that disease-susceptible mice (Il4raF709) with enhanced IL-4 receptor (IL-4R) signaling exhibited STAT6-dependent impaired generation and function of mucosal allergen-specific Treg cells. This failure was associated with the acquisition by Treg cells of T helper 2 (Th2) cell-like phenotype, also found in peripheral blood all...

  8. GATA3 and the T-cell lineage: essential functions before and after T-helper-2-cell differentiation

    OpenAIRE

    Ho, I-Cheng; Tai, Tzong-Shyuan; Pai, Sung-Yun

    2009-01-01

    Many advances in our understanding of the molecules that regulate the development, differentiation and function of T cells have been made over the past few years. One important regulator of T-cell differentiation is the transcription factor GATA3 (GATA-binding protein 3). Although the main function of GATA3 is to act as a master transcription factor for the differentiation of T helper 2 (TH2) cells, new research has helped to uncover crucial functions of GATA3 in T cells that go beyond TH2-ce...

  9. Blastema cells derived from New Zealand white rabbit's pinna carry stemness properties as shown by differentiation into insulin producing, neural, and osteogenic lineages representing three embryonic germ layers.

    Science.gov (United States)

    Saeinasab, Morvarid; Matin, Maryam M; Rassouli, Fatemeh B; Bahrami, Ahmad Reza

    2016-05-01

    Stem cells (SCs) are known as undifferentiated cells with self-renewal and differentiation capacities. Regeneration is a phenomenon that occurs in a limited number of animals after injury, during which blastema tissue is formed. It has been hypothesized that upon injury, the dedifferentiation of surrounding tissues leads into the appearance of cells with SC characteristics. In present study, stem-like cells (SLCs) were obtained from regenerating tissue of New Zealand white rabbit's pinna and their stemness properties were examined by their capacity to differentiate toward insulin producing cells (IPCs), as well as neural and osteogenic lineages. Differentiation was induced by culture of SLCs in defined medium, and cell fates were monitored by specific staining, RT-PCR and flow cytometry assays. Our results revealed that dithizone positive cells, which represent IPCs, and islet-like structures appeared 1 week after induction of SLCs, and this observation was confirmed by the elevated expression of Ins, Pax6 and Glut4 at mRNA level. Furthermore, SLCs were able to express neural markers as early as 1 week after retinoic acid treatment. Finally, SLCs were able to differentiate into osteogenic lineage, as confirmed by Alizarin Red S staining and RT-PCR studies. In conclusion, SLCs, which could successfully differentiate into cells derived from all three germ layers, can be considered as a valuable model to study developmental biology and regenerative medicine. PMID:25371011

  10. Flt3+ macrophage precursors commit sequentially to osteoclasts, dendritic cells and microglia

    OpenAIRE

    Hanau Daniel; Dumontel Christiane; Perret Magali; Rivollier Aymeric; Destaing Olivier; Domenget Chantal; Soulas Caroline; Grasset Marie-France; Nataf Serge; Jurdic Pierre; Arnaud Sylvie; Servet-Delprat Christine; Gilmore Gary L; Belin Marie-Françoise; Rabourdin-Combe Chantal

    2002-01-01

    Abstract Background Macrophages, osteoclasts, dendritic cells, and microglia are highly specialized cells that belong to the mononuclear phagocyte system. Functional and phenotypic heterogeneity within the mononuclear phagocyte system may reveal differentiation plasticity of a common progenitor, but developmental pathways leading to such diversity are still unclear. Results Mouse bone marrow cells were expanded in vitro in the presence of Flt3-ligand (FL), yielding high numbers of non-adheren...

  11. Divergent functions of hematopoietic transcription factors in lineage priming and differentiation during erythro-megakaryopoiesis.

    Science.gov (United States)

    Pimkin, Maxim; Kossenkov, Andrew V; Mishra, Tejaswini; Morrissey, Christapher S; Wu, Weisheng; Keller, Cheryl A; Blobel, Gerd A; Lee, Dongwon; Beer, Michael A; Hardison, Ross C; Weiss, Mitchell J

    2014-12-01

    Combinatorial actions of relatively few transcription factors control hematopoietic differentiation. To investigate this process in erythro-megakaryopoiesis, we correlated the genome-wide chromatin occupancy signatures of four master hematopoietic transcription factors (GATA1, GATA2, TAL1, and FLI1) and three diagnostic histone modification marks with the gene expression changes that occur during development of primary cultured megakaryocytes (MEG) and primary erythroblasts (ERY) from murine fetal liver hematopoietic stem/progenitor cells. We identified a robust, genome-wide mechanism of MEG-specific lineage priming by a previously described stem/progenitor cell-expressed transcription factor heptad (GATA2, LYL1, TAL1, FLI1, ERG, RUNX1, LMO2) binding to MEG-associated cis-regulatory modules (CRMs) in multipotential progenitors. This is followed by genome-wide GATA factor switching that mediates further induction of MEG-specific genes following lineage commitment. Interaction between GATA and ETS factors appears to be a key determinant of these processes. In contrast, ERY-specific lineage priming is biased toward GATA2-independent mechanisms. In addition to its role in MEG lineage priming, GATA2 plays an extensive role in late megakaryopoiesis as a transcriptional repressor at loci defined by a specific DNA signature. Our findings reveal important new insights into how ERY and MEG lineages arise from a common bipotential progenitor via overlapping and divergent functions of shared hematopoietic transcription factors. PMID:25319996

  12. Freezing tolerance of sea urchin embryonic cells: Differentiation commitment and cytoskeletal disturbances in culture.

    Science.gov (United States)

    Odintsova, Nelly A; Ageenko, Natalya V; Kipryushina, Yulia O; Maiorova, Mariia A; Boroda, Andrey V

    2015-08-01

    This study focuses on the freezing tolerance of sea urchin embryonic cells. To significantly reduce the loss of physiological activity of these cells that occurs after cryopreservation and to study the effects of ultra-low temperatures on sea urchin embryonic cells, we tested the ability of the cells to differentiate into spiculogenic or pigment directions in culture, including an evaluation of the expression of some genes involved in pigment differentiation. A morphological analysis of cytoskeletal disturbances after freezing in a combination of penetrating (dimethyl sulfoxide and ethylene glycol) and non-penetrating (trehalose and polyvinylpyrrolidone) cryoprotectants revealed that the distribution pattern of filamentous actin and tubulin was similar to that in the control cultures. In contrast, very rare spreading cells and a small number of cells with filamentous actin and tubulin were detected after freezing in the presence of only non-penetrating cryoprotectants. The largest number of pigment cells was found in cultures frozen with trehalose or trehalose and dimethyl sulfoxide. The ability to induce the spicule formation was lost in the cells frozen only with non-penetrating cryoprotectants, while it was maximal in cultures frozen in a cryoprotective mixture containing both non-penetrating and penetrating cryoprotectants (particularly, when ethylene glycol was present). Using different markers for cell state assessment, an effective cryopreservation protocol for sea urchin cells was developed: three-step freezing with a low cooling rate (1-2°C/min) and a combination of non-penetrating and penetrating cryoprotectants made it possible to obtain a high level of cell viability (up to 65-80%). PMID:26049089

  13. Sex determining region Y-Box 2 (SOX2 is a potential cell-lineage gene highly expressed in the pathogenesis of squamous cell carcinomas of the lung.

    Directory of Open Access Journals (Sweden)

    Ping Yuan

    Full Text Available BACKGROUND: Non-small cell lung cancer (NSCLC represents the majority (85% of lung cancers and is comprised mainly of adenocarcinomas and squamous cell carcinomas (SCCs. The sequential pathogenesis of lung adenocarcinomas and SCCs occurs through dissimilar phases as the former tumors typically arise in the lung periphery whereas the latter normally arise near the central airway. METHODOLOGY/PRINCIPAL FINDINGS: We assessed the expression of SOX2, an embryonic stem cell transcriptional factor that also plays important roles in the proliferation of basal tracheal cells and whose expression is restricted to the main and central airways and bronchioles of the developing and adult mouse lung, in NSCLC by various methodologies. Here, we found that SOX2 mRNA levels, from various published datasets, were significantly elevated in lung SCCs compared to adenocarcinomas (all p<0.001. Moreover, a previously characterized OCT4/SOX2/NANOG signature effectively separated lung SCCs from adenocarcinomas in two independent publicly available datasets which correlated with increased SOX2 mRNA in SCCs. Immunohistochemical analysis of various histological lung tissue specimens demonstrated marked nuclear SOX2 protein expression in all normal bronchial epithelia, alveolar bronchiolization structures and premalignant lesions in SCC development (hyperplasia, dysplasia and carcinoma in situ and absence of expression in all normal alveoli and atypical adenomatous hyperplasias. Moreover, SOX2 protein expression was greatly higher in lung SCCs compared to adenocarcinomas following analyses in two independent large TMA sets (TMA set I, n = 287; TMA set II, n = 511 both p<0.001. Furthermore, amplification of SOX2 DNA was detected in 20% of lung SCCs tested (n = 40 and in none of the adenocarcinomas (n = 17. CONCLUSIONS/SIGNIFICANCE: Our findings highlight a cell-lineage gene expression pattern for the stem cell transcriptional factor SOX2 in the pathogenesis of lung SCCs and

  14. PARP-1 inhibitors DPQ and PJ-34 negatively modulate proinflammatory commitment of human glioblastoma cells.

    Science.gov (United States)

    Scalia, Marina; Satriano, Cristina; Greca, Rossana; Stella, Anna Maria Giuffrida; Rizzarelli, Enrico; Spina-Purrello, Vittoria

    2013-01-01

    Poly(ADP-ribose) polymerases (PARPs) are recognized as key regulators of cell survival or death. PARP-1 is essential to the repair of DNA single-strand breaks via the base excision repair pathway. The enzyme may be overactivated in response to inflammatory cues, thus depleting cellular energy pools and eventually causing cell death. Accordingly, PARP-1 inhibitors, acting by competing with its physiological substrate NAD(+), have been proposed to play a protective role in a wide range of inflammatory and ischemia/reperfusion-associated diseases. Recently, it has also been reported that PARP-1 regulates proinflammatory mediators, including cytokines, chemokines, adhesion molecules, and enzymes (e.g., iNOS). Furthermore, PARP-1 has been shown to act as a coactivator of NF-κB- and other transcription factors implicated in stress/inflammation, as AP-1, Oct-1, SP-1, HIF, and Stat-1. To further substantiate this hypothesis, we tested the biomolecular effects of PARP-1 inhibitors DPQ and PJ-34 on human glioblastoma cells, induced to a proinflammatory state with lipopolysaccharide and Interferon-γ. PARP-1 expression was evaluated by laser scanning confocal microscopy immunofluorescence (LSM); nitrite production, LDH release and cell viability were also determined. LSM of A-172, SNB-19 and CAS-1 cells demonstrated that DPQ and PJ-34 downregulate PARP-1 expression; they also cause a decrease of LDH release and nitrite production, while increasing cell viability. Similar effects were caused in all three cell lines by N-mono-methyl-arginine, a well known iNOS inhibitor, and by L-carnosine and trehalose, two antioxidant molecules. These results demonstrate that, similar to other well characterized drugs, DPQ and PJ-34 reduce cell inflammation and damage that follow PARP-1 overexpression, while they increase cell survival: this suggests their potential exploitation in clinical Medicine. PMID:23011206

  15. A Gene Regulatory Network Cooperatively Controlled by Pdx1 and Sox9 Governs Lineage Allocation of Foregut Progenitor Cells

    DEFF Research Database (Denmark)

    Shih, Hung Ping; Seymour, Philip A; Patel, Nisha A; Xie, Ruiyu; Wang, Allen; Liu, Patrick P; Yeo, Gene W; Magnuson, Mark A; Sander, Maike

    2015-01-01

    The generation of pancreas, liver, and intestine from a common pool of progenitors in the foregut endoderm requires the establishment of organ boundaries. How dorsal foregut progenitors activate pancreatic genes and evade the intestinal lineage choice remains unclear. Here, we identify Pdx1 and Sox...

  16. The Molecular Signature of HIV-1-Associated Lipomatosis Reveals Differential Involvement of Brown and Beige/Brite Adipocyte Cell Lineages.

    Directory of Open Access Journals (Sweden)

    Rubén Cereijo

    Full Text Available Highly active antiretroviral therapy has remarkably improved quality of life of HIV-1-infected patients. However, this treatment has been associated with the so-called lipodystrophic syndrome, which conveys a number of adverse metabolic effects and morphological alterations. Among them, lipoatrophy of subcutaneous fat in certain anatomical areas and hypertrophy of visceral depots are the most common. Less frequently, lipomatous enlargements of subcutaneous fat at distinct anatomic areas occur. Lipomatous adipose tissue in the dorso-cervical area ("buffalo hump" has been associated with a partial white-to-brown phenotype transition and with increased cell proliferation, but, to date, lipomatous enlargements arising in other parts of the body have not been characterized. In order to establish the main molecular events associated with the appearance of lipomatosis in HIV-1 patients, we analyzed biopsies of lipomatous tissue from "buffalo hump" and from other anatomical areas in patients, in comparison with healthy subcutaneous adipose tissue, using a marker gene expression approach. Both buffalo-hump and non-buffalo-hump lipomatous adipose tissues exhibited similar patterns of non-compromised adipogenesis, unaltered inflammation, non-fibrotic phenotype and proliferative activity. Shorter telomere length, prelamin A accumulation and SA-β-Gal induction, reminiscent of adipocyte senescence, were also common to both types of lipomatous tissues. Buffalo hump biopsies showed expression of marker genes of brown adipose tissue (e.g. UCP1 and, specifically, of "classical" brown adipocytes (e.g. ZIC1 but not of beige/brite adipocytes. No such brown fat-related gene expression occurred in lipomatous tissues at other anatomical sites. In conclusion, buffalo hump and other subcutaneous adipose tissue enlargements from HIV-1-infected patients share a similar lipomatous character. However, a distorted induction of white-to-"classical brown adipocyte" phenotype

  17. Current lineages of the epithelioma papulosum cyprini (EPC) cell line are contaminated with fathead minnow, Pimephales promelas, cells

    Science.gov (United States)

    Winton, J.; Batts, W.; deKinkelin, P.; LeBerre, M.; Bremont, M.; Fijan, N.

    2010-01-01

    Initially established from proliferative skin lesions of the common carp, Cyprinus carpio L., the epithelioma papulosum cyprini (EPC) cell line (Fijan, Sulimanovic, Bearzotti, Muzinic, Zwillenberg, Chilmonczyk, Vautherot & de Kinkelin 1983) has become one of the most widely used tools for research on fish viruses and the diagnosis of fish viral diseases.

  18. Myocardial commitment from human pluripotent stem cells: Rapid production of human heart grafts.

    Science.gov (United States)

    Garreta, Elena; de Oñate, Lorena; Fernández-Santos, M Eugenia; Oria, Roger; Tarantino, Carolina; Climent, Andreu M; Marco, Andrés; Samitier, Mireia; Martínez, Elena; Valls-Margarit, Maria; Matesanz, Rafael; Taylor, Doris A; Fernández-Avilés, Francisco; Izpisua Belmonte, Juan Carlos; Montserrat, Nuria

    2016-08-01

    Genome editing on human pluripotent stem cells (hPSCs) together with the development of protocols for organ decellularization opens the door to the generation of autologous bioartificial hearts. Here we sought to generate for the first time a fluorescent reporter human embryonic stem cell (hESC) line by means of Transcription activator-like effector nucleases (TALENs) to efficiently produce cardiomyocyte-like cells (CLCs) from hPSCs and repopulate decellularized human heart ventricles for heart engineering. In our hands, targeting myosin heavy chain locus (MYH6) with mCherry fluorescent reporter by TALEN technology in hESCs did not alter major pluripotent-related features, and allowed for the definition of a robust protocol for CLCs production also from human induced pluripotent stem cells (hiPSCs) in 14 days. hPSCs-derived CLCs (hPSCs-CLCs) were next used to recellularize acellular cardiac scaffolds. Electrophysiological responses encountered when hPSCs-CLCs were cultured on ventricular decellularized extracellular matrix (vdECM) correlated with significant increases in the levels of expression of different ion channels determinant for calcium homeostasis and heart contractile function. Overall, the approach described here allows for the rapid generation of human cardiac grafts from hPSCs, in a total of 24 days, providing a suitable platform for cardiac engineering and disease modeling in the human setting. PMID:27179434

  19. A programmable synthetic lineage-control network that differentiates human IPSCs into glucose-sensitive insulin-secreting beta-like cells.

    Science.gov (United States)

    Saxena, Pratik; Heng, Boon Chin; Bai, Peng; Folcher, Marc; Zulewski, Henryk; Fussenegger, Martin

    2016-01-01

    Synthetic biology has advanced the design of standardized transcription control devices that programme cellular behaviour. By coupling synthetic signalling cascade- and transcription factor-based gene switches with reverse and differential sensitivity to the licensed food additive vanillic acid, we designed a synthetic lineage-control network combining vanillic acid-triggered mutually exclusive expression switches for the transcription factors Ngn3 (neurogenin 3; OFF-ON-OFF) and Pdx1 (pancreatic and duodenal homeobox 1; ON-OFF-ON) with the concomitant induction of MafA (V-maf musculoaponeurotic fibrosarcoma oncogene homologue A; OFF-ON). This designer network consisting of different network topologies orchestrating the timely control of transgenic and genomic Ngn3, Pdx1 and MafA variants is able to programme human induced pluripotent stem cells (hIPSCs)-derived pancreatic progenitor cells into glucose-sensitive insulin-secreting beta-like cells, whose glucose-stimulated insulin-release dynamics are comparable to human pancreatic islets. Synthetic lineage-control networks may provide the missing link to genetically programme somatic cells into autologous cell phenotypes for regenerative medicine. PMID:27063289

  20. Plasmodium berghei ANKA: erythropoietin activates neural stem cells in an experimental cerebral malaria model

    DEFF Research Database (Denmark)

    Core, Andrew; Hempel, Casper; Kurtzhals, Jørgen A L;

    2011-01-01

    ventricles, relative to control-treatment. 75% of the EPO-treated CM mice displayed migration as nestin(+) NSC. The NSC showed differentiation towards a neural cell lineage as shown by PSA-NCAM binding and NSC maturation and lineage commitment was significantly affected by exogenous EPO and by CM in the sub...

  1. A Role for Ethanol-Induced Oxidative Stress in Controlling Lineage Commitment of Mesenchymal Stromal Cells Through Inhibition of Wnt/β-Catenin Signaling

    OpenAIRE

    Chen, Jin-Ran; Lazarenko, Oxana P.; Shankar, Kartik; Blackburn, Michael L; Badger, Thomas M.; Ronis, Martin J.

    2009-01-01

    The mechanisms by which chronic ethanol intake induces bone loss remain unclear. In females, the skeletal response to ethanol varies depending on physiologic status (e.g., cycling, pregnancy, or lactation). Ethanol-induced oxidative stress appears to be a key event leading to skeletal toxicity. In this study, ethanol-containing liquid diets were fed to postlactational female Sprague-Dawley rats intragastrically for 4 weeks beginning at weaning. Ethanol consumption decreased bone mineral densi...

  2. High cell density and latent membrane protein 1 expression induce cleavage of the mixed lineage leukemia gene at 11q23 in nasopharyngeal carcinoma cell line

    Directory of Open Access Journals (Sweden)

    Sim Sai-Peng

    2010-09-01

    Full Text Available Abstract Background Nasopharyngeal carcinoma (NPC is commonly found in Southern China and South East Asia. Epstein-Barr virus (EBV infection is well associated with NPC and has been implicated in its pathogenesis. Moreover, various chromosome rearrangements were reported in NPC. However, the underlying mechanism of chromosome rearrangement remains unclear. Furthermore, the relationship between EBV and chromosome rearrangement with respect to the pathogenesis of NPC has not been established. We hypothesize that during virus- or stress-induced apoptosis, chromosomes are initially cleaved at the base of the chromatin loop domain structure. Upon DNA repair, cell may survive with rearranged chromosomes. Methods In this study, cells were seeded at various densities to induce apoptosis. Genomic DNA extracted was processed for Southern hybridization. In order to investigate the role of EBV, especially the latent membrane protein 1 (LMP1, LMP1 gene was overexpressed in NPC cells and chromosome breaks were analyzed by inverse polymerase chain (IPCR reaction. Results Southern analysis revealed that high cell density resulted in cleavage of the mixed lineage leukemia (MLL gene within the breakpoint cluster region (bcr. This high cell density-induced cleavage was significantly reduced by caspase inhibitor, Z-DEVD-FMK. Similarly, IPCR analysis showed that LMP1 expression enhanced cleavage of the MLL bcr. Breakpoint analysis revealed that these breaks occurred within the matrix attachment region/scaffold attachment region (MAR/SAR. Conclusions Since MLL locates at 11q23, a common deletion site in NPC, our results suggest a possibility of stress- or virus-induced apoptosis in the initiation of chromosome rearrangements at 11q23. The breakpoint analysis results also support the role of chromatin structure in defining the site of chromosome rearrangement.

  3. Dynamic Control of Enhancer Repertoires Drives Lineage and Stage-Specific Transcription during Hematopoiesis.

    Science.gov (United States)

    Huang, Jialiang; Liu, Xin; Li, Dan; Shao, Zhen; Cao, Hui; Zhang, Yuannyu; Trompouki, Eirini; Bowman, Teresa V; Zon, Leonard I; Yuan, Guo-Cheng; Orkin, Stuart H; Xu, Jian

    2016-01-11

    Enhancers are the primary determinants of cell identity, but the regulatory components controlling enhancer turnover during lineage commitment remain largely unknown. Here we compare the enhancer landscape, transcriptional factor occupancy, and transcriptomic changes in human fetal and adult hematopoietic stem/progenitor cells and committed erythroid progenitors. We find that enhancers are modulated pervasively and direct lineage- and stage-specific transcription. GATA2-to-GATA1 switch is prevalent at dynamic enhancers and drives erythroid enhancer commissioning. Examination of lineage-specific enhancers identifies transcription factors and their combinatorial patterns in enhancer turnover. Importantly, by CRISPR/Cas9-mediated genomic editing, we uncover functional hierarchy of constituent enhancers within the SLC25A37 super-enhancer. Despite indistinguishable chromatin features, we reveal through genomic editing the functional diversity of several GATA switch enhancers in which enhancers with opposing functions cooperate to coordinate transcription. Thus, genome-wide enhancer profiling coupled with in situ enhancer editing provide critical insights into the functional complexity of enhancers during development. PMID:26766440

  4. ISX-9 can potentiate cell proliferation and neuronal commitment in the rat dentate gyrus.

    Science.gov (United States)

    Bettio, Luis E B; Patten, Anna R; Gil-Mohapel, Joana; O'Rourke, Natasha F; Hanley, Ronan P; Kennedy, Samantha; Gopalakrishnan, Karthik; Rodrigues, Ana Lúcia S; Wulff, Jeremy; Christie, Brian R

    2016-09-22

    Adult hippocampal neurogenesis can be modulated by various physiological and pathological conditions, including stress, affective disorders, and several neurological conditions. Given the proposed role of this form of structural plasticity in the functioning of the hippocampus (namely learning and memory and affective behaviors), it is believed that alterations in hippocampal neurogenesis might underlie some of the behavioral deficits associated with these psychiatric and neurological conditions. Thus, the search for compounds that can reverse these deficits with minimal side effects has become a recognized priority. In the present study we tested the pro-neurogenic effects of isoxazole 9 (Isx-9), a small synthetic molecule that has been recently identified through the screening of chemical libraries in stem cell-based assays. We found that administration of Isx-9 for 14days was able to potentiate cell proliferation and increase the number of immature neurons in the hippocampal DG of adult rats. In addition, Isx-9 treatment was able to completely reverse the marked reduction in these initial stages of the neurogenic process observed in vehicle-treated animals (which were submitted to repeated handling and exposure to daily intraperitoneal injections). Based on these results, we recommend that future neurogenesis studies that require repeated handling and manipulation of animals should include a naïve (non-manipulated) control to determine the baseline levels of hippocampal cell proliferation and neuronal differentiation. Overall, these findings demonstrate that Isx-9 is a promising synthetic compound for the mitigation of stress-induced deficits in adult hippocampal neurogenesis. Future studies are thus warranted to evaluate the pro-neurogenic properties of Isx-9 in animal models of affective and neurological disorders associated with impaired hippocampal structural plasticity. PMID:27373772

  5. Enhancer sequences of a retroviral vector determine expression of a gene in multipotent hematopoietic progenitors and committed erythroid cells.

    OpenAIRE

    Holland, C A; Anklesaria, P; Sakakeeny, M A; Greenberger, J.S.

    1987-01-01

    To analyze the transcriptional activity of retroviral enhancer sequences in hematopoietic lineages, we determined the effect of enhancer sequences on the expression of the neomycin resistance gene transferred by two retroviral vectors to primary hematopoietic lineages. We constructed the vector pFr-SV(X). The Moloney murine leukemia virus enhancer region of a vector, pZIP-SV(X), was replaced by a 380-nucleotide-long fragment containing the enhancer sequences of the Friend murine leukemia viru...

  6. Lineage analysis of micromere 4d, a super-phylotypic cell for Lophotrochozoa, in the leech Helobdella and the sludgeworm Tubifex.

    Science.gov (United States)

    Gline, Stephanie E; Nakamoto, Ayaki; Cho, Sung-Jin; Chi, Candace; Weisblat, David A

    2011-05-01

    The super-phylum Lophotrochozoa contains the plurality of extant animal phyla and exhibits a corresponding diversity of adult body plans. Moreover, in contrast to Ecdysozoa and Deuterostomia, most lophotrochozoans exhibit a conserved pattern of stereotyped early divisions called spiral cleavage. In particular, bilateral mesoderm in most lophotrochozoan species arises from the progeny of micromere 4d, which is assumed to be homologous with a similar cell in the embryo of the ancestral lophotrochozoan, more than 650 million years ago. Thus, distinguishing the conserved and diversified features of cell fates in the 4d lineage among modern spiralians is required to understand how lophotrochozoan diversity has evolved by changes in developmental processes. Here we analyze cell fates for the early progeny of the bilateral daughters (M teloblasts) of micromere 4d in the leech Helobdella sp. Austin, a clitellate annelid. We show that the first six progeny of the M teloblasts (em1-em6) contribute five different sets of progeny to non-segmental mesoderm, mainly in the head and in the lining of the digestive tract. The latter feature, associated with cells em1 and em2 in Helobdella, is seen with the M teloblast lineage in a second clitellate species, the sludgeworm Tubifex tubifex and, on the basis of previously published work, in the initial progeny of the M teloblast homologs in molluscan species, suggesting that it may be an ancestral feature of lophotrochozoan development. PMID:21295566

  7. Normal and leukemic stem cells

    Science.gov (United States)

    Pelicci, P G

    2012-01-01

    Studies on hematopoietic stem cells have provided several critical insights in the biology of stem cells in general; as mature blood cells are generally short lived, stem cells are in fact required to guarantee, throughout the life of an organism, the replenishment of differentiated blood cells by the generation of multi-lineage progenitors and precursors committed to individual hematopoietic lineages. Similarly, acute myeloid leukemia has been considered as a model system to study cancer stem cells. This presentation illustrates some recent results obtained by our group with regard to both normal and leukemic stem cells.

  8. B-lymphocyte commitment: identifying the point of no return.

    Science.gov (United States)

    Welinder, Eva; Ahsberg, Josefine; Sigvardsson, Mikael

    2011-10-01

    Even though B-lymphocyte development is one of the best understood models for cell differentiation in the hematopoetic system, recent advances in cell sorting and functional genomics has increased this understanding further. This has suggested that already early lymphoid primed multipotent progenitor cells (LMPPs) express low levels of lymphoid restricted transcripts. The expression of these genes becomes more pronounced when cells enter the FLT-3/IL-7 receptor positive common lymphoid progenitor (CLP) stage. However, the expression of B-lineage specific genes is limited to a B-cell restricted Ly6D surface positive subpopulation of the CLP compartment. The gene expression patterns also reflect differences in lineage potential and while Ly6D negative FLT-3/IL-7 receptor positive cells represents true CLPs with an ability to generate B/T and NK cells, the Ly6D positive cells lack NK cell potential and display a reduced T-cell potential in vivo. These recent findings suggest that the CLP compartment is highly heterogenous and that the point of no return in B-cell development may occur already in B220(-)CD19(-) cells. These findings have allowed for a better understanding of the interplay between transcription factors like EBF-1, PAX-5 and E47, all known as crucial for normal B-cell development. In this review, we aim to provide a comprehensive overview of B-cell fate specification and commitment based on the recent advances in the understanding of molecular networks as well as functional properties of early progenitor populations. PMID:21944938

  9. The intestinal stem cell.

    NARCIS (Netherlands)

    Barker, N.; van de Wetering, M.L.; Clevers, H.

    2008-01-01

    The epithelium of the adult mammalian intestine is in a constant dialog with its underlying mesenchyme to direct progenitor proliferation, lineage commitment, terminal differentiation, and, ultimately, cell death. The epithelium is shaped into spatially distinct compartments that are dedicated to ea

  10. Effect of All-Trans Retinoic Acid (ATRA on Viability, Proliferation, Activation and Lineage-Specific Transcription Factors of CD4+ T Cells

    Directory of Open Access Journals (Sweden)

    Katayoon Bidad

    2011-12-01

    Full Text Available All-trans retinoic acid (ATRA, as an active metabolite of vitamin A, has been shown to affect immune cells. This study was performed to evaluate the effect of ATRA on viability, proliferation, activation and lineage-specific transcription factors of CD4+ T cells. CD4+ T cells were  separated  from  heparinized  blood  of  healthy  donors  and  were  cultured  in conditions, some with, some without ATRA.Viability was assessed by PI  flowcytometry and proliferation was measured by MTT assay. CD69 expression was determined by flowcytometry as a measure of cell activation. Lineage-specific transcription  factors  (FOXP3,  RORγt  and  T-bet  were  examined  by intracellular staining and flowcytometry. High doses of ATRA (0.1-1 mM caused extensive cell death in both PBMCs and CD4+ T cells. Doses of ATRA equal to or lower than 10 µM did not  adversely affect cell viability and proliferation in comparison to  culture medium without ATRA.Doses of ATRA between 10 µM and 1nM significantly increased cell activation when compared  to  culture medium without  ATRA. ATRA could increase FOXP3+  and also FOXP3+RORγt+ T cells while it decreased RORγt+ and T-bet+ T cells. This study showed that doses of ATRA up to 10 µM are safe when using with CD4+  T cells in terms of cell viability, proliferation and activation.We  could  also  show  that  ATRA  diverts  the  human  immune  response  in  neutral conditions (without adding polarizing cytokines by increasing FOXP3+  cells and decreasing RORγt+  cells. ATRA could be regarded as a potential therapy in inflammatory conditions and autoimmunities.

  11. Postembryonic lineages of the Drosophila brain: II. Identification of lineage projection patterns based on MARCM clones

    OpenAIRE

    Wong, Darren C.; Lovick, Jennifer K.; Ngo, Kathy T.; Borisuthirattana, Wichanee; Omoto, Jaison J.; Hartenstein, Volker

    2013-01-01

    The Drosophila central brain is largely composed of lineages, units of sibling neurons derived from a single progenitor cell or neuroblast. During the early embryonic period neuroblast generate the primary neurons that constitute the larval brain. Neuroblasts reactivate in the larva, adding to their lineages a large number of secondary neurons which, according to previous studies in which selected lineages were labeled by stably expressed markers, differentiate during metamorphosis, sending t...

  12. Risk assessment of relapse by lineage-specific monitoring of chimerism in children undergoing allogeneic stem cell transplantation for acute lymphoblastic leukemia

    Science.gov (United States)

    Preuner, Sandra; Peters, Christina; Pötschger, Ulrike; Daxberger, Helga; Fritsch, Gerhard; Geyeregger, Rene; Schrauder, André; von Stackelberg, Arend; Schrappe, Martin; Bader, Peter; Ebell, Wolfram; Eckert, Cornelia; Lang, Peter; Sykora, Karl-Walter; Schrum, Johanna; Kremens, Bernhard; Ehlert, Karoline; Albert, Michael H.; Meisel, Roland; Lawitschka, Anita; Mann, Georg; Panzer-Grümayer, Renate; Güngör, Tayfun; Holter, Wolfgang; Strahm, Brigitte; Gruhn, Bernd; Schulz, Ansgar; Woessmann, Wilhelm; Lion, Thomas

    2016-01-01

    Allogeneic hematopoietic stem cell transplantation is required as rescue therapy in about 20% of pediatric patients with acute lymphoblastic leukemia. However, the relapse rates are considerable, and relapse confers a poor outcome. Early assessment of the risk of relapse is therefore of paramount importance for the development of appropriate measures. We used the EuroChimerism approach to investigate the potential impact of lineage-specific chimerism testing for relapse-risk analysis in 162 pediatric patients with acute lymphoblastic leukemia after allogeneic stem cell transplantation in a multicenter study based on standardized transplantation protocols. Within a median observation time of 4.5 years, relapses have occurred in 41/162 patients at a median of 0.6 years after transplantation (range, 0.13–5.7 years). Prospective screening at defined consecutive time points revealed that reappearance of recipient-derived cells within the CD34+ and CD8+ cell subsets display the most significant association with the occurrence of relapses with hazard ratios of 5.2 (P=0.003) and 2.8 (P=0.008), respectively. The appearance of recipient cells after a period of pure donor chimerism in the CD34+ and CD8+ leukocyte subsets revealed dynamics indicative of a significantly elevated risk of relapse or imminent disease recurrence. Assessment of chimerism within these lineages can therefore provide complementary information for further diagnostic and, potentially, therapeutic purposes aiming at the prevention of overt relapse. This study was registered at clinical.trials.gov with the number NC01423747. PMID:26869631

  13. Acute progression of BCR-FGFR1 induced murine B-lympho/myeloproliferative disorder suggests involvement of lineages at the pro-B cell stage.

    Directory of Open Access Journals (Sweden)

    Mingqiang Ren

    Full Text Available Constitutive activation of FGFR1, through rearrangement with various dimerization domains, leads to atypical myeloproliferative disorders where, although T cell lymphoma are common, the BCR-FGFR1 chimeric kinase results in CML-like leukemia. As with the human disease, mouse bone marrow transduction/transplantation with BCR-FGFR1 leads to CML-like myeloproliferation as well as B-cell leukemia/lymphoma. The murine disease described in this report is virtually identical to the human disease in that both showed bi-lineage involvement of myeloid and B-cells, splenomegaly, leukocytosis and bone marrow hypercellularity. A CD19(+ IgM(- CD43(+ immunophenotype was seen both in primary tumors and two cell lines derived from these tumors. In all primary tumors, subpopulations of these CD19(+ IgM(- CD43(+ were also either B220(+ or B220(-, suggesting a block in differentiation at the pro-B cell stage. The B220(- phenotype was retained in one of the cell lines while the other was B220(+. When the two cell lines were transplanted into syngeneic mice, all animals developed the same B-lymphoblastic leukemia within 2-weeks. Thus, the murine model described here closely mimics the human disease with bilineage myeloid and B-cell leukemia/lymphoma which provides a representative model to investigate therapeutic intervention and a better understanding of the etiology of the disease.

  14. Chung-Yeul-Gue-Soup-Sa-Gan-Tang, traditional Korean medicine, enhances CD4(+) T cell activities and modulates Th1/Th2 lineage development.

    Science.gov (United States)

    Ko, Eunjung; Park, Jae-Woo; Rho, Samwoong; Cho, Chongwoon; Park, Seongkyu; Ko, Seonggyu; Lee, Yongwon; Hong, Moo-Chang; Shin, Min-Kyu; Ryu, Ki-Won; Bae, Hyunsu

    2004-04-01

    Chung-Yeul-Gue-Soup-Sa-Gan-Tang (CYT), a traditional Korea herbal medicine, has been widely used in Korea for the treatment of various immunological disorders, including allergic asthma. In this study, CYT was examined in vitro and tested for possible immunological effects. The results demonstrated that CYT had no mitogenic effects on unstimulated CD4(+) T cells, but rather increased CD4(+) T cell proliferation upon activation with anti-CD3/CD28 antibody. Under the Th0 condition, CYT also enhanced expression of interleukin (IL)-2 in purified murine CD4(+) T cells assayed by real-time PCR, suggesting that CYT moderately increases the activity of helper T cells upon T cell receptor ligation under the neutral condition. However, the Th1 cells were overpopulated following CYT treatment under the Th1 condition, while Th2 cells were under-populated in the Th2 driven condition. In addition, under Th1/Th2-skewed conditions, the levels of IL-4 were considerably decreased, while the expression of T-bet and interferon-gamma were increased with CYT treatment. Thus, CYT enhances Th1 lineage development from naive CD4(+) T cells both by increasing Th1 specific cytokine secretion and repressing Th2 specific cytokine production. These results suggest that CYT is a desirable agent for the correction of Th2 dominant pathological disorders. PMID:15107575

  15. Erythropoietin guides multipotent hematopoietic progenitor cells toward an erythroid fate

    Science.gov (United States)

    Grover, Amit; Mancini, Elena; Moore, Susan; Mead, Adam J.; Atkinson, Deborah; Rasmussen, Kasper D.; O’Carroll, Donal; Jacobsen, Sten Eirik W.

    2014-01-01

    The erythroid stress cytokine erythropoietin (Epo) supports the development of committed erythroid progenitors, but its ability to act on upstream, multipotent cells remains to be established. We observe that high systemic levels of Epo reprogram the transcriptomes of multi- and bipotent hematopoietic stem/progenitor cells in vivo. This induces erythroid lineage bias at all lineage bifurcations known to exist between hematopoietic stem cells (HSCs) and committed erythroid progenitors, leading to increased erythroid and decreased myeloid HSC output. Epo, therefore, has a lineage instructive role in vivo, through suppression of non-erythroid fate options, demonstrating the ability of a cytokine to systematically bias successive lineage choices in favor of the generation of a specific cell type. PMID:24493804

  16. Direct in vivo cell lineage analysis in the retrorsine and 2AAF models of liver injury after genetic labeling in adult and newborn rats.

    Directory of Open Access Journals (Sweden)

    Virginie Pichard

    Full Text Available BACKGROUNDS AND AIMS: When hepatocyte proliferation is impaired, liver regeneration proceeds from the division of non parenchymal hepatocyte progenitors. Oval cells and Small Hepatocyte-like Progenitor Cells (SHPCs represent the two most studied examples of such epithelial cells with putative stem cell capacity. In the present study we wished to compare the origin of SHPCs proliferating after retrorsine administration to the one of oval cells observed after 2-Acetyl-Amino fluorene (2-AAF treatment. METHODOLOGY/PRINCIPAL FINDINGS: We used retroviral-mediated nlslacZ genetic labeling of dividing cells to study the fate of cells in the liver. Labeling was performed either in adult rats before treatment or in newborn animals. Labeled cells were identified and characterised by immunohistochemistry. In adult-labeled animals, labeling was restricted to mature hepatocytes. Retrorsine treatment did not modify the overall number of labeled cells in the liver whereas after 2-AAF administration unlabeled oval cells were recorded and the total number of labeled cells decreased significantly. When labeling was performed in newborn rats, results after retrorsine administration were identical to those obtained in adult-labeled rats. In contrast, in the 2-AAF regimen numerous labeled oval cells were present and were able to generate new labeled hepatocytes. Furthermore, we also observed labeled biliary tracts in 2-AAF treated rats. CONCLUSIONS: Our results strongly suggest that SHPCs are derived from hepatocytes and we confirm that SHPCs and oval cells do not share the same origin. We also show that hepatic progenitors are labeled in newborn rats suggesting future directions for in vivo lineage studies.

  17. Embryonic Stem Cell-Derived Cardiomyocyte Heterogeneity and the Isolation of Immature and Committed Cells for Cardiac Remodeling and Regeneration

    Directory of Open Access Journals (Sweden)

    Kenneth R. Boheler

    2011-01-01

    Full Text Available Pluripotent stem cells represent one promising source for cell replacement therapy in heart, but differentiating embryonic stem cell-derived cardiomyocytes (ESC-CMs are highly heterogeneous and show a variety of maturation states. In this study, we employed an ESC clonal line that contains a cardiac-restricted ncx1 promoter-driven puromycin resistance cassette together with a mass culture system to isolate ESC-CMs that display traits characteristic of very immature CMs. The cells display properties of proliferation, CM-restricted markers, reduced mitochondrial mass, and hypoxia-resistance. Following transplantation into rodent hearts, bioluminescence imaging revealed that immature cells, but not more mature CMs, survived for at least one month following injection. These data and comparisons with more mature cells lead us to conclude that immature hypoxia resistant ESC-CMs can be isolated in mass in vitro and, following injection into heart, form grafts that may mediate long-term recovery of global and regional myocardial contractile function following infarction.

  18. Single-cell lineage tracking analysis reveals that an established cell line comprises putative cancer stem cells and their heterogeneous progeny

    Science.gov (United States)

    Sato, Sachiko; Rancourt, Ann; Sato, Yukiko; Satoh, Masahiko S.

    2016-01-01

    Mammalian cell culture has been used in many biological studies on the assumption that a cell line comprises putatively homogeneous clonal cells, thereby sharing similar phenotypic features. This fundamental assumption has not yet been fully tested; therefore, we developed a method for the chronological analysis of individual HeLa cells. The analysis was performed by live cell imaging, tracking of every single cell recorded on imaging videos, and determining the fates of individual cells. We found that cell fate varied significantly, indicating that, in contrast to the assumption, the HeLa cell line is composed of highly heterogeneous cells. Furthermore, our results reveal that only a limited number of cells are immortal and renew themselves, giving rise to the remaining cells. These cells have reduced reproductive ability, creating a functionally heterogeneous cell population. Hence, the HeLa cell line is maintained by the limited number of immortal cells, which could be putative cancer stem cells. PMID:27003384

  19. Constraints to Progress in Embryonic Stem Cells from Domestic Species

    OpenAIRE

    Muñoz, M. (M.); Trigal, B. (Beatriz); Molina, I.; Díez, C.; Caamaño, J.N. (JN); E. Gómez

    2013-01-01

    Domestic animal embryonic stem cells are of potentially big value in transgenic research and studies of lineage commitment and development. Unfortunately, despite many efforts, validated embryonic stem cell lines in species other than mice and primates are yet to be isolated. Here we review some factors that might help to explain why derivation of domestic animal embryonic stem cells is still unsuccessful.

  20. Determination of osteogenic or adipogenic lineages in muscle-derived stem cells (MDSCs) by a collagen-binding peptide (CBP) derived from bone sialoprotein (BSP)

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Yoon Jung [Dental Regenerative Biotechnology Major, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 110-749 (Korea, Republic of); Lee, Jue Yeon [Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Lee, Seung Jin [Department of Industrial Pharmacy, College of Pharmacy, Ewha Womans University, Seoul (Korea, Republic of); Chung, Chong-Pyoung, E-mail: ccpperio@snu.ac.kr [Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of); Department of Periodontology, School of Dentistry and Dental Research Institute, Seoul National University, Seoul (Korea, Republic of); Park, Yoon Jeong, E-mail: parkyj@snu.ac.kr [Dental Regenerative Biotechnology Major, School of Dentistry and Dental Research Institute, Seoul National University, Seoul 110-749 (Korea, Republic of); Research Institute, Nano Intelligent Biomedical Engineering Corporation (NIBEC), Seoul (Korea, Republic of)

    2012-03-09

    Highlights: Black-Right-Pointing-Pointer CBP sequence is identified from BSP and has collagen binding activity. Black-Right-Pointing-Pointer CBP directly activates the MAPK signaling, especially ERK1/2. Black-Right-Pointing-Pointer CBP increase osteoblastic differentiation by the activation of Runx2. Black-Right-Pointing-Pointer CBP decrease adipogenic differentiation by the inhibition of PPAR{gamma}. -- Abstract: Bone sialoprotein (BSP) is a mineralized, tissue-specific, non-collagenous protein that is normally expressed only in mineralized tissues such as bone, dentin, cementum, and calcified cartilage, and at sites of new mineral formation. The binding of BSP to collagen is thought to be important for initiating bone mineralization and bone cell adhesion to the mineralized matrix. Several recent studies have isolated stem cells from muscle tissue, but their functional properties are still unclear. In this study, we examined the effects of a synthetic collagen-binding peptide (CBP) on the differentiation efficiency of muscle-derived stem cells (MDSCs). The CBP sequence (NGVFKYRPRYYLYKHAYFYPHLKRFPVQ) corresponds to residues 35-62 of bone sialoprotein (BSP), which are located within the collagen-binding domain in BSP. Interestingly, this synthetic CBP inhibited adipogenic differentiation but increased osteogenic differentiation in MDSCs. The CBP also induced expression of osteoblastic marker proteins, including alkaline phosphatase (ALP), type I collagen, Runt-related transcription factor 2 (Runx2), and osteocalcin; prevented adipogenic differentiation in MDSCs; and down-regulated adipose-specific mRNAs, such as adipocyte protein 2 (aP2) and peroxisome proliferator-activated receptor {gamma}. The CBP increased Extracellular signal-regulated kinases (ERK) 1/2 protein phosphorylation, which is important in lineage determination. These observations suggest that this CBP determines the osteogenic or adipogenic lineage in MDSCs by activating ERK1/2. Taken together, a

  1. The levels of IL-17A and of the cytokines involved in Th17 cell commitment are increased in patients with chronic immune thrombocytopenia

    Science.gov (United States)

    Rocha, Andreia Maria Camargos; Souza, Cláudia; Rocha, Gifone Aguiar; de Melo, Fabrício Freire; Clementino, Nelma Cristina Diogo; Marino, Marília Campos Abreu; Bozzi, Adriana; Silva, Maria Luiza; Martins Filho, Olindo Assis; Queiroz, Dulciene Maria Magalhães

    2011-01-01

    Th17 cells have been associated with immune-mediated diseases in humans but it has still not been determined whether they play a role in immune thrombocytopenia. We evaluated representative cytokines of the Th17, Th1, Th2 and Treg cell commitment in the serum of patients with chronic immune thrombocytopenia, as well as the cell source of IL-17A. Higher levels of IL-17A and Th17-related cytokines, and an increased percentage of IL-17A producing CD4+ and neutrophils were observed in patients. The levels of cytokines involved in Th1 cell commitment IFN-γ, IL-2, IL12-p70 and the percentages of Th1 cells were also increased, but IL-4 was not detected. Although the concentrations of IL-10 were higher, the levels of TGF-β were similar in both groups. In conclusion, our results point to a putative role for Th-17 cells/IL-17A cytokine in the pathogenesis of chronic immune thrombocytopenia. PMID:21972211

  2. Shear Stress Induces Differentiation of Endothelial Lineage Cells to Protect Neonatal Brain from Hypoxic-Ischemic Injury through NRP1 and VEGFR2 Signaling

    Directory of Open Access Journals (Sweden)

    Chia-Wei Huang

    2015-01-01

    Full Text Available Neonatal hypoxic-ischemic (HI brain injuries disrupt the integrity of neurovascular structure and lead to lifelong neurological deficit. The devastating damage can be ameliorated by preserving the endothelial network, but the source for therapeutic cells is limited. We aim to evaluate the beneficial effect of mechanical shear stress in the differentiation of endothelial lineage cells (ELCs from adipose-derived stem cells (ASCs and the possible intracellular signals to protect HI injury using cell-based therapy in the neonatal rats. The ASCs expressed early endothelial markers after biochemical stimulation of endothelial growth medium. The ELCs with full endothelial characteristics were accomplished after a subsequential shear stress application for 24 hours. When comparing the therapeutic potential of ASCs and ELCs, the ELCs treatment significantly reduced the infarction area and preserved neurovascular architecture in HI injured brain. The transplanted ELCs can migrate and engraft into the brain tissue, especially in vessels, where they promoted the angiogenesis. The activation of Akt by neuropilin 1 (NRP1 and vascular endothelial growth factor receptor 2 (VEGFR2 was important for ELC migration and following in vivo therapeutic outcomes. Therefore, the current study demonstrated importance of mechanical factor in stem cell differentiation and showed promising protection of brain from HI injury using ELCs treatment.

  3. Microfluidics 3D gel-island chip for single cell isolation and lineage-dependent drug responses study.

    Science.gov (United States)

    Zhang, Zhixiong; Chen, Yu-Chih; Cheng, Yu-Heng; Luan, Yi; Yoon, Euisik

    2016-07-01

    3D cell culture in the extracellular matrix (ECM), which not only provides structural support to cellular constituents, but also initiates regulatory biochemical cues for a variety of important cell functions in tissue, has become more and more important in understanding cancer pathology and drug testing. Although the ECM-gel has been used in cell culture both in bulk and on-chip, previous studies focused on collective cell behavior rather than single-cell heterogeneity. To track the behavior of each individual cell, we have developed a gel-island chip, which can form thousands of islands containing single cells encapsulated by the desired ECM. Optimized by Poisson's distribution, the device can attain 34% single cell capture efficiency of the exact number of single cells per island. A good culture media exchange rate and high cell viability can be achieved in the gel-islands. The cells in the islands can be automatically counted for high-throughput analysis. As a proof of concept, we monitored the proliferation and differentiation of single Notch+ (stem-like) T47D breast cancer cells. The 3D collagen gel environment was found to be favorable for the stem-like phenotype through better self-renewal and de-differentiation (Notch- to Notch+ transition). More interestingly, we found that the Notch- de-differentiated cells were more resistant to doxorubicin and cisplatin than the Notch+ cells. Combining the 3D ECM culture and single cell resolution, the presented platform can automatically analyze the individual cell behaviors of hundreds of cells using a small amount of drug and reagents. PMID:27270563

  4. Study of ADCT-402 in Patients With Relapsed or Refractory B-cell Lineage Non Hodgkin Lymphoma (B-NHL)

    Science.gov (United States)

    2016-07-04

    Non-Hodgkin Lymphoma; Burkitt's Lymphoma; Chronic Lymphocytic Leukemia; Small Lymphocytic Lymphoma; Lymphoma, Large B-Cell, Diffuse; Lymphoma, Follicular; Lymphoma, Mantle-Cell; Lymphoma, Marginal Zone; Waldenstrom Macroglobulinemia

  5. Bacterial-induced cell reprogramming to stem cell-like cells: new premise in host-pathogen interactions

    OpenAIRE

    Hess, Samuel; Rambukkana, Anura

    2014-01-01

    Bacterial pathogens employ a myriad of strategies to alter host tissue cell functions for bacterial advantage during infection. Recent advances revealed a fusion of infection biology with stem cell biology by demonstrating developmental reprogramming of lineage committed host glial cells to progenitor/stem cell-like cells by an intracellular bacterial pathogen Mycobacterium leprae. Acquisition of migratory and immunomodulatory properties of such reprogrammed cells provides an added advantage ...

  6. NF-M (chicken C/EBP beta) induces eosinophilic differentiation and apoptosis in a hematopoietic progenitor cell line.

    OpenAIRE

    Müller, C.; Kowenz-Leutz, E; Grieser-Ade, S; Graf, T.; Leutz, A.

    1995-01-01

    CAAT/enhancer binding proteins (C/EBPs) are transcriptional activators implicated in the differentiation processes of various cell lineages. We have shown earlier that NF-M, the chicken homolog of C/EBP beta, is specifically expressed in myelomonocytic and eosinophilic cells of the hematopoietic system. To investigate the role of NF-M in hematopoietic cell lineage commitment, we constructed a conditional form of the protein by fusing it to the hormone binding domain of the human estrogen rece...

  7. A Multi-Lineage Screen Reveals mTORC1 Inhibition Enhances Human Pluripotent Stem Cell Mesendoderm and Blood Progenitor Production

    Directory of Open Access Journals (Sweden)

    Emanuel Joseph Paul Nazareth

    2016-05-01

    Full Text Available Human pluripotent stem cells (hPSCs exist in heterogeneous micro-environments with multiple subpopulations, convoluting fate-regulation analysis. We patterned hPSCs into engineered micro-environments and screened responses to 400 small-molecule kinase inhibitors, measuring yield and purity outputs of undifferentiated, neuroectoderm, mesendoderm, and extra-embryonic populations. Enrichment analysis revealed mammalian target of rapamycin (mTOR inhibition as a strong inducer of mesendoderm. Dose responses of mTOR inhibitors such as rapamycin synergized with Bone Morphogenetic protein 4 (BMP4 and activin A to enhance the yield and purity of BRACHYURY-expressing cells. Mechanistically, small interfering RNA knockdown of RAPTOR, a component of mTOR complex 1, phenocopied the mesendoderm-enhancing effects of rapamycin. Functional analysis during mesoderm and endoderm differentiation revealed that mTOR inhibition increased the output of hemogenic endothelial cells 3-fold, with a concomitant enhancement of blood colony-forming cells. These data demonstrate the power of our multi-lineage screening approach and identify mTOR signaling as a node in hPSC differentiation to mesendoderm and its derivatives.

  8. A Multi-Lineage Screen Reveals mTORC1 Inhibition Enhances Human Pluripotent Stem Cell Mesendoderm and Blood Progenitor Production.

    Science.gov (United States)

    Nazareth, Emanuel Joseph Paul; Rahman, Nafees; Yin, Ting; Zandstra, Peter William

    2016-05-10

    Human pluripotent stem cells (hPSCs) exist in heterogeneous micro-environments with multiple subpopulations, convoluting fate-regulation analysis. We patterned hPSCs into engineered micro-environments and screened responses to 400 small-molecule kinase inhibitors, measuring yield and purity outputs of undifferentiated, neuroectoderm, mesendoderm, and extra-embryonic populations. Enrichment analysis revealed mammalian target of rapamycin (mTOR) inhibition as a strong inducer of mesendoderm. Dose responses of mTOR inhibitors such as rapamycin synergized with Bone Morphogenetic protein 4 (BMP4) and activin A to enhance the yield and purity of BRACHYURY-expressing cells. Mechanistically, small interfering RNA knockdown of RAPTOR, a component of mTOR complex 1, phenocopied the mesendoderm-enhancing effects of rapamycin. Functional analysis during mesoderm and endoderm differentiation revealed that mTOR inhibition increased the output of hemogenic endothelial cells 3-fold, with a concomitant enhancement of blood colony-forming cells. These data demonstrate the power of our multi-lineage screening approach and identify mTOR signaling as a node in hPSC differentiation to mesendoderm and its derivatives. PMID:27132889

  9. The cholesterol-binding protein NPC2 restrains recruitment of stromal macrophage-lineage cells to early-stage lung tumours.

    Science.gov (United States)

    Kamata, Tamihiro; Jin, Hong; Giblett, Susan; Patel, Bipin; Patel, Falguni; Foster, Charles; Pritchard, Catrin

    2015-09-01

    The tumour microenvironment is known to play an integral role in facilitating cancer progression at advanced stages, but its function in some pre-cancerous lesions remains elusive. We have used the (V600) (E)BRAF-driven mouse lung model that develop premalignant lesions to understand stroma-tumour interactions during pre-cancerous development. In this model, we have found that immature macrophage-lineage cells (IMCs) producing PDGFA, TGFβ and CC chemokines are recruited to the stroma of premalignant lung adenomas through CC chemokine receptor 1 (CCR1)-dependent mechanisms. Stromal IMCs promote proliferation and transcriptional alterations suggestive of epithelial-mesenchymal transition in isolated premalignant lung tumour cells ex vivo, and are required for the maintenance of early-stage lung tumours in vivo. Furthermore, we have found that IMC recruitment to the microenvironment is restrained by the cholesterol-binding protein, Niemann-Pick type C2 (NPC2). Studies on isolated cells ex vivo confirm that NPC2 is secreted from tumour cells and is taken up by IMCs wherein it suppresses secretion of the CCR1 ligand CC chemokine 6 (CCL6), at least in part by facilitating its lysosomal degradation. Together, these findings show that NPC2 secreted by premalignant lung tumours suppresses IMC recruitment to the microenvironment in a paracrine manner, thus identifying a novel target for the development of chemopreventive strategies in lung cancer. PMID:26183450

  10. Lineage relationship of CD8+ T cell subsets is revealed by progressive changes in the epigenetic landscape

    Science.gov (United States)

    Crompton, Joseph G.; Narayanan, Manikandan; Cuddapah, Suresh; Roychoudhuri, Rahul; Ji, Yun; Yang, Wenjing; Patel, Shashank J.; Sukumar, Madhusudhanan; Palmer, Douglas C.; Peng, Weiqun; Wang, Ena; Marincola, Francesco M.; Klebanoff, Christopher A.; Zhao, Keji; Tsang, John S.; Gattinoni, Luca; Restifo, Nicholas P.

    2016-01-01

    To better elucidate epigenetic mechanisms that correlate with the dynamic gene expression program observed upon T-cell differentiation, we investigated the genomic landscape of histone modifications in naive and memory CD8+ T cells. Using a ChIP-Seq approach coupled with global gene expression profiling, we generated genome-wide histone H3 lysine 4 (H3K4me3) and H3 lysine 27 (H3K27me3) trimethylation maps in naive, T memory stem cells, central memory cells, and effector memory cells in order to gain insight into how histone architecture is remodeled during T cell differentiation. We show that H3K4me3 histone modifications are associated with activation of genes, while H3K27me3 is negatively correlated with gene expression at canonical loci and enhancers associated with T-cell metabolism, effector function, and memory. Our results also reveal histone modifications and gene expression signatures that distinguish the recently identified T memory stem cells from other CD8+ T-cell subsets. Taken together, our results suggest that CD8+ lymphocytes undergo chromatin remodeling in a progressive fashion. These findings have major implications for our understanding of peripheral T-cell ontogeny and the formation of immunological memory. PMID:25914936

  11. Hematopoietic microenvironment. Origin, lineage, and transplantability of the stromal cells in long-term bone marrow cultures from chimeric mice.

    OpenAIRE

    Perkins, S; Fleischman, R A

    1988-01-01

    Studies of bone marrow transplant patients have suggested that the stromal cells of the in vitro hematopoietic microenvironment are transplantable into conditioned recipients. Moreover, in patients with myeloproliferative disorders, all of the stromal cells, which include presumptive endothelial cells, appear to be derived from hematopoietic precursors. To confirm these findings, we have constructed two chimeric mouse models: (a) traditional radiation chimeras, and (b) fetal chimeras, produce...

  12. 9-cis-Retinoic Acid Promotes Cell Adhesion Through Integrin Dependent and Independent Mechanisms Across Immune Lineages

    OpenAIRE

    Whelan, Jarrett T.; Chen, Jianming; Miller, Jabin; Morrow, Rebekah L.; Lingo, Joshuah D.; Merrell, Kaitlin; Shaikh, Saame Raza; Bridges, Lance C.

    2012-01-01

    Retinoids are essential in the proper establishment and maintenance of immunity. Although retinoids are implicated in immune related processes, their role in immune cell adhesion has not been well established. In this study, the effect of 9-cis-retinoic acid (9-cis-RA) on human hematopoietic cell adhesion was investigated. 9-cis-RA treatment specifically induced cell adhesion of the human immune cell lines HuT-78, NB4, RPMI 8866, and U937. Due to the prominent role of integrin receptors in me...

  13. Glia Maturation Factor Gamma (GMFG): A Cytokine-Responsive Protein During Hematopoietic Lineage Development and Its Functional Genomics Analysis

    Institute of Scientific and Technical Information of China (English)

    Ying Shi; Ling Chen; Lance A.Liotta; Hong-Hui Wan; Griffin P.Rodgers

    2006-01-01

    Human hematopoiesis was evaluated using the techniques of controlled stem cell differentiation, two-dimensional gel electrophoresis-based proteomics, and functional genomics. We provide the first report that glia maturation factor gamma (GMFG) is a cytokine-responsive protein in erythropoietin-induced and granulocyte-colony stimulating factor-induced hematopoietic lineage development.Results from global functional genomics analysis indicate that GMFG possesses several other features: hematopoietic tissue-specific gene expression, a promoter concentrated with high-score hematopoiesis-specific transcription factors, and possible molecular coevolution with a rudimentary blood/immune system. On the basis of our findings, we hypothesize that GMFG is a hematopoietic-specific protein that may mediate the pluripotentiality and lineage commitment of human hematopoietic stem cells.

  14. Postmarket Requirements and Commitments

    Data.gov (United States)

    U.S. Department of Health & Human Services — Provides information to the public on postmarket requirements and commitments. The phrase postmarket requirements and commitments refers to studies and clinical...

  15. The increased number of Leydig cells by di(2-ethylhexyl) phthalate comes from the differentiation of stem cells into Leydig cell lineage in the adult rat testis

    International Nuclear Information System (INIS)

    Highlights: ► DEHP increases rat Leydig cell number. ► DEHP induces the proliferation of stem Leydig cells. ► DEHP induces the formation of progenitor Leydig cells. - Abstract: The objective of the present study is to determine whether di(2-ethylhexyl) phthalate (DEHP) exposure at adulthood increases rat Leydig cell number and to investigate the possible mechanism. 90-day-old Long–Evans rats were randomly divided into 3 groups, and were gavaged with the corn oil (control) or 10 or 750 mg/kg DEHP daily for 7 days, and then received an intraperitoneal injection of 75 mg/kg ethane dimethanesulfonate (EDS) to eliminate Leydig cells. Serum testosterone concentrations were assessed by RIA, and the mRNA levels of Leydig cell genes were measured by qPCR. EDS eliminated all Leydig cells in the control testis on day 4 post-EDS, as judged by undetectable serum testosterone level and no 3β-hydroxysteroid dehydrogenase positive (3β-HSDpos) cells in the interstitium. However, in DEHP-treated groups, there were detectable serum testosterone concentrations and some oval-shaped 3β-HSDpos cells in the interstitium. These 3β-HSDpos cells were not stained by the antibody against 11β-hydroxysteroid dehydrogenase 1 (11β-HSD1), a marker for Leydig cells at a more advanced stage. The disappearance of mRNAs of Leydig cell biomarkers including Lhcgr, Cyp11a1, Cyp17a1, Insl3 and Hsd11b1 in the control testis was observed on day 4 post-EDS. However, there were detectable concentrations of Lhcgr, Cyp11a1 and Cyp17a1 mRNAs but undetectable concentrations of Insl3, Hsd17b3 and Hsd11b1 in the DEHP-treated testes, indicating that these 3β-HSDpos cells were newly formed progenitor Leydig cells. The mRNA level for nestin (Nes, biomarker for stem Leydig cells) was significantly increased in the control testis on day 4 post-EDS, but not in the DEHP treated testes, suggesting that these nestin positive stem cells were differentiated into progenitor Leydig cells in the DEHP-treated testes

  16. Cell lineage analysis of the mandibular segment of the amphipod Orchestia cavimana reveals that the crustacean paragnaths are sternal outgrowths and not limbs

    Directory of Open Access Journals (Sweden)

    Scholtz Gerhard

    2006-12-01

    Full Text Available Abstract The question of arthropod head segmentation has become one of the central issues in Evolutionary Developmental Biology. The number of theories pertaining to head segments progressively enlarges, old concepts have been revitalized, and nearly every conceivable composition of the arthropod head has at some point received discussion. One contentious issue involves a characteristic mouthpart in crustaceans – the lower lips or the so-called paragnaths. The paragnaths build the posterior border of the mouth region antagonistic to the upper lip – the labrum. We show here the development of the appendage-like structures in the mandibular region of the amphipod crustacean Orchestia cavimana at a high level of cellular resolution. The embryos are examined during development of the mouthparts using in vivo labeling. An invariant cell division pattern of the mandibular segment was detected by 4D-microscopy and a preliminary model for pattern of the first cleavages in the mandibular region created. With this indispensable precondition single ectodermal cells of the grid-like pattern were labeled with DiI – a lipophilic fluorescent dye – to trace cell lineages and determine the clonal composition of the developing mouthparts, especially the mandibular segment. From our data it is evident that the paragnaths are sternal outgrowths of the mandible segment. The assumption of the limb nature of paragnaths and the presence of an additional head segment between the mandibular and the second antennal segments are clearly refuted by our data. Our results show the power of cell lineage and clonal analyses for inferences on the nature, origin and thus homology of morphological structures. With this kind of investigation morphological and gene expression data can be complemented. We discuss notable similarities of paragnath anlagen to those of the hypopharynx complex in myriapods and hexapods. The fact that both structures grow out as two lateral buds

  17. Donor commitment and patient needs.

    Science.gov (United States)

    Bakken, R; van Walraven, A-M; Egeland, T

    2004-01-01

    The article discusses views and recommendations of the World Marrow Donor Association concerning ethical issues related to the donation of hematopoietic stem cell products with respect to recruitment, evaluation, workup, and follow-up of unrelated donors. Particular emphasis is placed upon commitment of individual donors, in particular, with respect to the needs of patients to find HLA-matched donors, who may be asked to donate stem cell and other cell products more than once for given patients. PMID:14628078

  18. Auctions with Limited Commitment

    OpenAIRE

    Qingmin Liu; Konrad Mierendorff; Xianwen Shi

    2013-01-01

    We study auction design in the standard symmetric independent private values environment, where the seller lacks the commitment power to withhold an unsold object off the market. The seller has a single object and can conduct an infinite sequence of standard auctions with reserve prices to maximize her expected profit. In each period, the seller can commit to a reserve price for the current period but cannot commit to future reserve prices. We analyze the problem with limited commitment throu...

  19. Integrative Genomic Analyses Identify BRF2 as a Novel Lineage-Specific Oncogene in Lung Squamous Cell Carcinoma

    OpenAIRE

    Lockwood, William W; Chari, Raj; Coe, Bradley P.; Thu, Kelsie L.; Garnis, Cathie; Malloff, Chad A.; Campbell, Jennifer; Williams, Ariane C.; Hwang, Dorothy; Zhu, Chang-Qi; Buys, Timon P.H.; Yee, John; English, John C.; MacAulay, Calum; Tsao, Ming-sound

    2010-01-01

    Editors' Summary Background Lung cancer is the commonest cause of cancer-related death. Every year, 1.3 million people die from this disease, which is mainly caused by smoking. Most cases of lung cancer are “non-small cell lung cancers” (NSCLCs). Like all cancers, NSCLC starts when cells begin to divide uncontrollably and to move round the body (metastasize) because of changes (mutations) in their genes. These mutations are often in “oncogenes,” genes that, when activated, encourage cell divi...

  20. Empowering self-renewal and differentiation: the role of mitochondria in stem cells

    OpenAIRE

    Rehman, Jalees

    2010-01-01

    Stem cells are characterized by their multi-lineage differentiation potential (pluripotency) and their ability for self-renewal, which permits them to proliferate while avoiding lineage commitment and senescence. Recent studies demonstrate that undifferentiated, pluripotent stem cells display lower levels of mitochondrial mass and oxidative phosphorylation, and instead preferentially use non-oxidative glycolysis as a major source of energy. Hypoxia is a potent suppressor of mitochondrial oxid...

  1. Nox2 and Nox4 influence neonatal c-kit+ cardiac precursor cell status and differentiation

    OpenAIRE

    Nadworny, Alyson S.; Guruju, Mallik R.; Poor, Daniel; Doran, Robert M.; Sharma, Ram V.; Kotlikoff, Michael I.; Davisson, Robin L.

    2013-01-01

    Redox status has emerged as critical in modulating stemness and lineage commitment in several precursor cell types. However, a role for redox genes, specifically NADPH oxidases (Nox), in cardiac precursor cells (CPCs) has not been established. We tested whether CPCs marked by type III receptor tyrosine kinase c-kit (c-kit+) exhibit a unique NADPH oxidase signature that confers precursor status and whether alterations in this profile are functionally linked to changes in lineage specification....

  2. Determining Lineage Pathways from Cellular Barcoding Experiments

    Directory of Open Access Journals (Sweden)

    Leïla Perié

    2014-02-01

    Full Text Available Cellular barcoding and other single-cell lineage-tracing strategies form experimental methodologies for analysis of in vivo cell fate that have been instrumental in several significant recent discoveries. Due to the highly nonlinear nature of proliferation and differentiation, interrogation of the resulting data for evaluation of potential lineage pathways requires a new quantitative framework complete with appropriate statistical tests. Here, we develop such a framework, illustrating its utility by analyzing data from barcoded multipotent cells of the blood system. This application demonstrates that the data require additional paths beyond those found in the classical model, which leads us to propose that hematopoietic differentiation follows a loss of potential mechanism and to suggest further experiments to test this deduction. Our quantitative framework can evaluate the compatibility of lineage trees with barcoded data from any proliferating and differentiating cell system.

  3. Human Mesenchymal Stem Cells Derived From Limb Bud Can Differentiate into All Three Embryonic Germ Layers Lineages

    OpenAIRE

    Jiao, Fei; Wang, Juan; Dong, Zhao-lun; Wu, Min-juan; Zhao, Ting-bao; Li, Dan-Dan; Xin WANG

    2012-01-01

    Mesenchymal stem cells (MSCs) have been isolated from many sources, including adults and fetuses. Previous studies have demonstrated that, compared with their adult counterpart, fetal MSCs with several remarkable advantages may be a better resource for clinical applications. In this study, we successfully isolated a rapidly proliferating cell population from limb bud of aborted fetus and termed them “human limb bud–derived mesenchymal stem cells” (hLB-MSCs). Characteristics of their morpholog...

  4. Improvement of mesenchymal stem cell differentiation into the endoderm lineage by four step sequential method in biocompatible biomaterial

    Science.gov (United States)

    Azandeh, Saeed; Mohammad Gharravi, Anneh; Orazizadeh, Mahmoud; Khodadi, Ali; Hashemi Tabar, Mahmoud

    2016-01-01

    Introduction: The goal of the study described here, was to investigate the potential of umbilical cord derived mesenchymal stem cell (UC-MSCs) into hepatocyte like cells in a sequential 2D and 3D differentiation protocols as alternative therapy. Methods: Mesenchymal stem cells (MSCs) were isolated from the umbilical cord (UC) and CD markers were analyzed by flow cytometry. For hepatic differentiation of UC-MSCs, cells were induced with a sequential 4-step protocol in 3D and 2D culture system. Urea concentration and albumin secretion into the culture medium was quantified by ELISA. Gene expression levels of AFP, ALB, and CK18 were determined by RT-PCR. Data were statistically analyzed by the SPSS software. The difference between the mean was considered significant when p < 0.05. Results: Growth factor dependent morphological changes from elongated fibroblast-like cells to round epithelial cell morphology were observed in 2D culture. Cell proliferation analysis showed round-shaped morphology with clear cytoplasm and nucleus on the alginate scaffold in 3D culture. The mean valuses of albumin production and urea secretion were significantly higher in the 3D Culture system when compared with the 2D culture (p = 0.005 vs p = 0.001), respectively. Treatment of cells with TSA in the final step of differentiation induced an increased expression of CK18 and a decreased expression of αFP in both the 3D and 2D cultures (p = 0.026), but led to a decreased albumin gene expression, and an increased expression in the 2D culture (p = 0.001). Conclusion: Findings of the present study indicated that sequential exposure of UC-MSCs with growth factors in 3D culture improves hepatic differentiation.

  5. Integrative genomic analyses identify BRF2 as a novel lineage-specific oncogene in lung squamous cell carcinoma.

    OpenAIRE

    Lockwood, William W; Raj Chari; Coe, Bradley P.; Thu, Kelsie L.; Cathie Garnis; Malloff, Chad A.; Jennifer Campbell; Williams, Ariane C.; Dorothy Hwang; Chang-Qi Zhu; Buys, Timon P.H.; John Yee; English, John C.; Calum Macaulay; Ming-Sound Tsao

    2010-01-01

    BACKGROUND: Traditionally, non-small cell lung cancer is treated as a single disease entity in terms of systemic therapy. Emerging evidence suggests the major subtypes--adenocarcinoma (AC) and squamous cell carcinoma (SqCC)--respond differently to therapy. Identification of the molecular differences between these tumor types will have a significant impact in designing novel therapies that can improve the treatment outcome. METHODS AND FINDINGS: We used an integrative genomics approach, combin...

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

    Science.gov (United States)

    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

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

    Directory of Open Access Journals (Sweden)

    Sabine Conrad

    2016-01-01

    Full Text Available 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.

  8. Human bone-lineage cell responses to anisotropic Ti6Al4V surfaces are dependent on their maturation state.

    Science.gov (United States)

    Calzado-Martín, Alicia; Crespo, Lara; Saldaña, Laura; Boré, Alba; Gómez-Barrena, Enrique; Vilaboa, Nuria

    2014-09-01

    This article reports on the interactions of human bone cells, mesenchymal stem cells (hMSCs) from bone marrow and osteoblasts (hOBs), with a submicron-grooved Ti6Al4V alloy that promotes cell orientation in the direction of the anisotropy. Adhesion sites, actin and tubulin networks and fibronectin extracellular matrix of both cell types align with the direction of the grooves. hMSCs adhere at a higher rate on the patterned substrate than on the polished alloy, while no differences are found in hOBs attachment. Compared to the flat substrate, RhoA activity is higher in hMSCs and hOB cultured on the grooved alloy and treatment with C3 transferase leads to loss of organization of actin and tubulin cytoskeletons. Rho-associated kinase (ROCK) activity of hMSCs is upregulated on the anisotropic samples, but not affected in hOBs. Treatment with hydroxyfasudil disrupts the alignment of adhesion sites in hMSCs but not in hOBs. When cells are cultured in media that support osteogenic maturation, OPN secretion increases in hMSCs on the anisotropic alloy and it remains unaffected in hOBs. Cell layer calcification proceeds to a same extent in hMSCs cultured on the two metallic surfaces but decreases in hOBs cultured on the patterned samples. Taken together, these results indicate that hOBs are less sensitive than hMSCs to the patterned Ti6Al4V alloy. This effect can be attributed to their different stages of cell maturation and may be mediated, at least in part, through ROCK signaling because its activity increases on hMSCs cultured on the patterned alloy, while hOBs fail to upregulate it. PMID:24136907

  9. Primordial germ cells in an oligochaete annelid are specified according to the birth rank order in the mesodermal teloblast lineage.

    Science.gov (United States)

    Kato, Yukie; Nakamoto, Ayaki; Shiomi, Inori; Nakao, Hajime; Shimizu, Takashi

    2013-07-15

    The primordial germ cells (PGCs) in the oligochaete annelid Tubifex tubifex are descentants of the mesodermal (M) teloblast and are located in the two midbody segments X and XI in which they serve as germline precursors forming the testicular gonad and the ovarian gonad, respectively. During embryogenesis, vasa-expressing cells (termed presumptive PGCs or pre-PGCs) emerge in a variable set of midbody segments including the genital segments (X and XI); at the end of embryogenesis, pre-PGCs are confined to the genital segments, where they become PGCs in the juvenile. Here, using live imaging of pre-PGCs, we have demonstrated that during Tubifex embryogenesis, pre-PGCs (defined by Vasa expression) stay in segments where they have emerged, suggesting that it is unlikely that pre-PGCs move intersegmentally during embryogenesis. Thus, it is apparent that pre-PGCs derived from the 10th and 11th M teloblast-derived primary m blast cells (designated m10 and m11) that give rise, respectively, to segments X and XI are specified in situ as PGCs and that those born in other segments become undetectable at the end of embryogenesis. To address the mechanisms for this segment-specific development of PGCs, we have performed a set of cell-transplantation experiments as well as cell-ablation experiments. When m10 and m11 that are normally located in the mid region of the embryo were placed in positions near the anterior end of the host embryo, these cells formed two consecutive segments, which exhibited Vasa-positive PGC-like cells at early juvenile stage. This suggests that in terms of PGC generation, the fates of m10 and m11 remain unchanged even if they are placed in ectopic positions along the anteroposterior axis. Nor was the fate of m10 and m11 changed even if mesodermal blast cell chains preceding or succeeding m10 and m11 were absent. In a previous study, it was shown that PGC development in segments X and XI occurs normally in the absence of the overlying ectoderm. All this

  10. Conditional deletion of the relaxin receptor gene in cells of smooth muscle lineage affects lower reproductive tract in pregnant mice.

    Science.gov (United States)

    Kaftanovskaya, Elena M; Huang, Zaohua; Lopez, Carolina; Conrad, Kirk; Agoulnik, Alexander I

    2015-04-01

    Relaxin hormone secreted into the circulation during pregnancy was discovered through its effects on pubic symphysis relaxation and parturition. Genetic inactivation of the relaxin gene or its cognate relaxin family peptide receptor 1 (RXFP1) in mice caused failure of parturition and mammary nipple enlargement, as well as increased collagen fiber density in the cervix and vagina. However, the relaxin effect on discrete cells and tissues has yet to be determined. Using transgenic mice with a knockin LacZ reporter in the Rxfp1 allele, we showed strong expression of this gene in vaginal and cervical stromal cells, as well as pubic ligament cells. We produced a floxed Rxfp1 allele that was used in combination with the Tagln-cre transgene to generate mice with a smooth muscle-specific gene knockout. In pregnant females, the ROSA26 reporter activated by Tagln-cre was detected in smooth muscle cells of the cervix, vagina, uterine artery, and in cells of the pubic symphysis. In late pregnant females with conditional gene ablation, the length of pubic symphysis was significantly reduced compared with wild-type or heterozygous Rxfp1(+/-) females. Denser collagen content was revealed by Masson trichrome staining in reproductive tract organs, uterine artery, and pubic symphysis. The cervical and vaginal epithelium was less developed than in heterozygous or wild-type females, although nipple size was normal and the dams were able to nurse their pups. In summary, our data indicate that relaxin/RXFP1 signaling in smooth muscle cells is important for normal collagen turnover and relaxation of the pubic symphysis during pregnancy. PMID:25715795

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

    OpenAIRE

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

    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 ge...

  12. The early human germ cell lineage does not express SOX2 during in vivo development or upon in vitro culture

    DEFF Research Database (Denmark)

    Perrett, Rebecca M; Turnpenny, Lee; Eckert, Judith J;

    2008-01-01

    pluripotent embryonic germ cells (EGCs) are derived, also express POU5F1, NANOG, and SOX2. Thus, a similar expression profile has been predicted for human PGCs. Here we show by RT-PCR, immunoblotting, and immunohistochemistry that human PGCs express POU5F1 and NANOG but not SOX2, with no evidence of...

  13. miRNA Profiles of Monocyte-Lineage Cells Are Consistent with Complicated Roles in HIV-1 Restriction

    Directory of Open Access Journals (Sweden)

    Janice E. Clements

    2012-09-01

    Full Text Available Long-lived HIV-1 reservoirs include tissue macrophages. Monocyte-derived macrophages are more susceptible to infection and more permissive to HIV-1 replication than monocytes for reasons that may include the effects of different populations of miRNAs in these two cell classes. Specifically, miRs-28-3p, -150, -223, -198, and -382 exert direct or indirect negative effects on HIV-1 and are reportedly downmodulated during monocyte-to-macrophage differentiation. Here, new experimental results are presented along with reviews and analysis of published studies and publicly available datasets, supporting a broader role of miRNAs in HIV-1 restriction than would be suggested by a simple and uniform downregulation of anti-HIV miRNAs during monocyte-to-macrophage differentiation. Although miR-223 is downregulated in macrophages, other putatively antiviral miRNAs are more abundant in macrophages than in monocytes or are rare and/or variably present in both cell classes. Our analyses point to the need for further studies to determine miRNA profiles of monocytes and macrophages, including classic and newly identified subpopulations; examine the sensitivity of miRNA profiling to cell isolation and differentiation protocols; and characterize rigorously the antiviral effects of previously reported and novel predicted miRNA-HIV-1 interactions in cell-specific contexts.

  14. A major lineage of enteroendocrine cells coexpress CCK, secretin, GIP, GLP-1, PYY, and neurotensin but not somatostatin

    DEFF Research Database (Denmark)

    Egerod, Kristoffer Lihme; Engelstoft, Maja Storm; Grunddal, Kaare Villum;

    2012-01-01

    peptide precursors in addition to CCK. The coexpression pattern was further confirmed through a cell ablation study based on expression of the human diphtheria toxin receptor under the control of the proglucagon promoter, in which activation of the receptor resulted in a marked reduction not only in GLP-1...

  15. Collagen-Coated Polytetrafluoroethane Membrane Inserts Enhances Chondrogenic Differentiation of Human Cord Blood Multi-Lineage Progenitor Cells

    DEFF Research Database (Denmark)

    Munir, Samir; Søballe, Kjeld; Ulrich-Vinther, Michael;

    standard micromass pellet system, layered on calcium polyphosphate (CPP), and on semi-permeable polytetrafluoroethane membranes with and without collagen type I, II or IV pre-coating. Findings / Results: The MPLC cell line used in this study possessed poor chondrogenic potency overall, but membrane...... culturing resulted in a multicellular layer tissue with formation of more cartilaginous tissue compared to micromass or CPP culture. In the membrane system MLPCs produced pellucid discs, 12 mm in diameter by 1 mm in thickness from 2x10^6 cells. The discs had hyaline-like cartilage extracellular matrix, with...... micromass or CPP cultures. Conclusions: In conclusion, we demonstrate that MLPCs possess’ chondrogenic potency, which increased when cultured scaffold-free on membrane inserts resulting in multicellular-layered hyaline-like cartilage tissue. Evaluating the effect of culturing pre-differentiated MLPCs on CPP...

  16. D-MEF2: a MADS box transcription factor expressed in differentiating mesoderm and muscle cell lineages during Drosophila embryogenesis.

    OpenAIRE

    Lilly, B; Galewsky, S; Firulli, A B; Schulz, R A; Olson, E N

    1994-01-01

    The myocyte enhancer factor (MEF) 2 family of transcription factors has been implicated in the regulation of muscle transcription in vertebrates. We have cloned a protein from Drosophila, termed D-MEF2, that shares extensive amino acid homology with the MADS (MCM1, Agamous, Deficiens, and serum-response factor) domains of the vertebrate MEF2 proteins. D-mef2 gene expression is first detected during Drosophila embryogenesis within mesodermal precursor cells prior to specification of the somati...

  17. Foxn1 regulates lineage progression in cortical and medullary thymic epithelial cells but is dispensable for medullary sublineage divergence

    OpenAIRE

    Nowell, Craig S; Nicholas Bredenkamp; Stéphanie Tetélin; Xin Jin; Christin Tischner; Harsh Vaidya; Sheridan, Julie M.; Frances Hogg Stenhouse; Raphaela Heussen; Andrew J H Smith; C Clare Blackburn

    2011-01-01

    The forkhead transcription factor Foxn1 is indispensable for thymus development, but the mechanisms by which it mediates thymic epithelial cell (TEC) development are poorly understood. To examine the cellular and molecular basis of Foxn1 function, we generated a novel and revertible hypomorphic allele of Foxn1. By varying levels of its expression, we identified a number of features of the Foxn1 system. Here we show that Foxn1 is a powerful regulator of TEC differentiation that is required at ...

  18. miRNA Profiles of Monocyte-Lineage Cells Are Consistent with Complicated Roles in HIV-1 Restriction

    OpenAIRE

    Clements, Janice E.; Sisk, Jeanne M.; Witwer, Kenneth W.

    2012-01-01

    Long-lived HIV-1 reservoirs include tissue macrophages. Monocyte-derived macrophages are more susceptible to infection and more permissive to HIV-1 replication than monocytes for reasons that may include the effects of different populations of miRNAs in these two cell classes. Specifically, miRs-28-3p, -150, -223, -198, and -382 exert direct or indirect negative effects on HIV-1 and are reportedly downmodulated during monocyte-to-macrophage differentiation. Here, new experimental results are ...

  19. Cell-lineage regulated myogenesis for dystrophin replacement: a novel therapeutic approach for treatment of muscular dystrophy

    OpenAIRE

    Kimura, En; Han, Jay J.; Li, Sheng; Fall, Brent; Ra, Jennifer; Haraguchi, Miki; Tapscott, Stephen J.; Jeffrey S. Chamberlain

    2008-01-01

    Duchenne muscular dystrophy (DMD) is characterized in skeletal muscle by cycles of myofiber necrosis and regeneration leading to loss of muscle fibers and replacement with fibrotic connective and adipose tissue. The ongoing activation and recruitment of muscle satellite cells for myofiber regeneration results in loss of regenerative capacity in part due to proliferative senescence. We explored a method whereby new myoblasts could be generated in dystrophic muscles by transplantation of primar...

  20. Glucose starvation induces mutation and lineage-dependent adaptive responses in a large collection of cancer cell lines

    OpenAIRE

    He, Ningning; Kim, Nayoung; JEONG, EUNA; Lu, Yiling; Mills, Gordon B.; Yoon, Sukjoon

    2015-01-01

    Tolerance of glucose deprivation is an important factor for cancer proliferation, survival, migration and progression. To systematically understand adaptive responses under glucose starvation in cancers, we analyzed reverse phase protein array (RPPA) data of 115 protein antibodies across a panel of approximately 170 heterogeneous cancer cell lines, cultured under normal and low glucose conditions. In general, glucose starvation broadly altered levels of many of the proteins and phosphoprotein...

  1. Identification and Characterization of Mouse Otic Sensory Lineage Genes

    Directory of Open Access Journals (Sweden)

    Byron H. Hartman

    2015-03-01

    Full Text Available Vertebrate embryogenesis gives rise to all cell types of an organism through the development of many unique lineages derived from the three primordial germ layers. The otic sensory lineage arises from the otic vesicle, a structure formed through invagination of placodal non-neural ectoderm. This developmental lineage possesses unique differentiation potential, giving rise to otic sensory cell populations including hair cells, supporting cells, and ganglion neurons of the auditory and vestibular organs. Here we present a systematic approach to identify transcriptional features that distinguish the otic sensory lineage (from early otic progenitors to otic sensory populations from other major lineages of vertebrate development. We used a microarray approach to analyze otic sensory lineage populations including microdissected otic vesicles (embryonic day 10.5 as well as isolated neonatal cochlear hair cells and supporting cells at postnatal day 3. Non-otic tissue samples including periotic tissues and whole embryos with otic regions removed were used as reference populations to evaluate otic specificity. Otic populations shared transcriptome-wide correlations in expression profiles that distinguish members of this lineage from non-otic populations. We further analyzed the microarray data using comparative and dimension reduction methods to identify individual genes that are specifically expressed in the otic sensory lineage. This analysis identified and ranked top otic sensory lineage-specific transcripts including Fbxo2, Col9a2, and Oc90, and additional novel otic lineage markers. To validate these results we performed expression analysis on select genes using immunohistochemistry and in situ hybridization. Fbxo2 showed the most striking pattern of specificity to the otic sensory lineage, including robust expression in the early otic vesicle and sustained expression in prosensory progenitors and auditory and vestibular hair cells and supporting

  2. Kaposi's Sarcoma-Associated Herpesvirus OX2 Glycoprotein Activates Myeloid-Lineage Cells To Induce Inflammatory Cytokine Production

    OpenAIRE

    Chung, Young-Hwa; Means, Robert E.; Choi, Joong-Kook; Lee, Bok-Soo; Jae U. Jung

    2002-01-01

    Kaposi's sarcoma is an inflammatory cytokine-mediated angioproliferative disease which is triggered by infection by Kaposi's sarcoma-associated herpesvirus (KSHV). KSHV contains an open reading frame, K14, that has significant homology with cellular OX2, designated viral OX2 (vOX2). In this report, we demonstrate that vOX2 encodes a glycosylated cell surface protein with an apparent molecular mass of 55 kDa. Purified glycosylated vOX2 protein dramatically stimulated primary monocytes, macroph...

  3. The intestinal stem cell

    OpenAIRE

    Barker, Nick; van de Wetering, Marc; Clevers, Hans

    2008-01-01

    The epithelium of the adult mammalian intestine is in a constant dialog with its underlying mesenchyme to direct progenitor proliferation, lineage commitment, terminal differentiation, and, ultimately, cell death. The epithelium is shaped into spatially distinct compartments that are dedicated to each of these events. While the intestinal epithelium represents the most vigorously renewing adult tissue in mammals, the stem cells that fuel this self-renewal process have been identified only rec...

  4. Genome Editing of Lineage Determinants in Human Pluripotent Stem Cells Reveals Mechanisms of Pancreatic Development and Diabetes.

    Science.gov (United States)

    Zhu, Zengrong; Li, Qing V; Lee, Kihyun; Rosen, Bess P; González, Federico; Soh, Chew-Li; Huangfu, Danwei

    2016-06-01

    Directed differentiation of human pluripotent stem cells (hPSCs) into somatic counterparts is a valuable tool for studying disease. However, examination of developmental mechanisms in hPSCs remains challenging given complex multi-factorial actions at different stages. Here, we used TALEN and CRISPR/Cas-mediated gene editing and hPSC-directed differentiation for a systematic analysis of the roles of eight pancreatic transcription factors (PDX1, RFX6, PTF1A, GLIS3, MNX1, NGN3, HES1, and ARX). Our analysis not only verified conserved gene requirements between mice and humans but also revealed a number of previously unsuspected developmental mechanisms with implications for type 2 diabetes. These include a role of RFX6 in regulating the number of pancreatic progenitors, a haploinsufficient requirement for PDX1 in pancreatic β cell differentiation, and a potentially divergent role of NGN3 in humans and mice. Our findings support use of systematic genome editing in hPSCs as a strategy for understanding mechanisms underlying congenital disorders. PMID:27133796

  5. [Lymphopoiesis supported by osteolineage cells].

    Science.gov (United States)

    Katayama, Yoshio

    2016-05-01

    Bone marrow(BM)and thymus are known as the primary lymphoid organs for B and T cells, respectively. However, the cell fate for T cell lineage commitment is already determined in the BM. Thus, the lymphopoiesis is critically controlled in the BM and, according to the recent advances in genetic mouse models, it appears that this process is strictly regulated by a series of osteolineage mesenchymal populations. PMID:27117616

  6. Reconstructing a B-cell clonal lineage. I. Statistical inference of unobserved ancestors [v1; ref status: indexed, http://f1000r.es/z6

    Directory of Open Access Journals (Sweden)

    Thomas B Kepler

    2013-04-01

    Full Text Available One of the key phenomena in the adaptive immune response to infection and immunization is affinity maturation, during which antibody genes are mutated and selected, typically resulting in a substantial increase in binding affinity to the eliciting antigen. Advances in technology on several fronts have made it possible to clone large numbers of heavy-chain light-chain pairs from individual B cells and thereby identify whole sets of clonally related antibodies. These collections could provide the information necessary to reconstruct their own history - the sequence of changes introduced into the lineage during the development of the clone - and to study affinity maturation in detail. But the success of such a program depends entirely on accurately inferring the founding ancestor and the other unobserved intermediates. Given a set of clonally related immunoglobulin V-region genes, the method described here allows one to compute the posterior distribution over their possible ancestors, thereby giving a thorough accounting of the uncertainty inherent in the reconstruction. I demonstrate the application of this method on heavy-chain and light-chain clones, assess the reliability of the inference, and discuss the sources of uncertainty.

  7. Nanotopography Directs Mesenchymal Stem Cells to Osteoblast Lineage through Regulation of microRNA-SMAD-BMP-2 Circuit

    Science.gov (United States)

    KATO, ROGERIO B.; ROY, BHASKAR; DE OLIVEIRA, FABIOLA S.; FERRAZ, EMANUELA P.; DE OLIVEIRA, PAULO T.; KEMPER, AUSTIN G.; HASSAN, MOHAMMAD Q.; ROSA, ADALBERTO L.; BELOTI, MARCIO M.

    2016-01-01

    The aim of this study was to investigate if chemically produced nanotopography on titanium (Ti) surface induces osteoblast differentiation of cultured human bone marrow mesenchymal stem cells (hMSCs) by regulating the expression of microRNAs (miRs). It was demonstrated that Ti with nanotopography induces osteoblast differentiation of hMSCs as evidenced by upregulation of osteoblast specific markers compared with untreated (control) Ti at day 4. At this time-point, miR-sequencing analysis revealed that 20 miRs were upregulated (>2 fold) while 20 miRs were downregulated (>3 fold) in hMSCs grown on Ti with nanotopography compared with control Ti. Three miRs, namely miR-4448, -4708 and -4773, which were significantly downregulated (>5 fold) by Ti with nanotopography affect osteoblast differentiation of hMSCs. These miRs that directly target SMAD1 and SMAD4, both key transducers of the bone morphogenetic protein 2 (BMP-2) osteogenic signal, were upregulated by Ti with nanotopography. Overexpression of miR-4448, -4708 and 4773 in MC3T3-E1 pre-osteoblasts noticeably inhibited gene and protein expression of SMAD1 and SMAD4 and therefore repressed the gene expression of key bone markers. Additionally, it was observed that the treatment with BMP-2 displayed a higher osteogenic effect on MC3T3-E1 cells grown on Ti with nanotopography compared with control Ti, suggesting that the BMP-2 signaling pathway was more effective on this surface. Taken together, these results indicate that a complex regulatory network involving a miR-SMAD-BMP-2 circuit governs the osteoblast differentiation induced by Ti with nanotopography. PMID:24619927

  8. Postembryonic lineages of the Drosophila brain: II. Identification of lineage projection patterns based on MARCM clones.

    Science.gov (United States)

    Wong, Darren C; Lovick, Jennifer K; Ngo, Kathy T; Borisuthirattana, Wichanee; Omoto, Jaison J; Hartenstein, Volker

    2013-12-15

    The Drosophila central brain is largely composed of lineages, units of sibling neurons derived from a single progenitor cell or neuroblast. During the early embryonic period, neuroblasts generate the primary neurons that constitute the larval brain. Neuroblasts reactivate in the larva, adding to their lineages a large number of secondary neurons which, according to previous studies in which selected lineages were labeled by stably expressed markers, differentiate during metamorphosis, sending terminal axonal and dendritic branches into defined volumes of the brain neuropil. We call the overall projection pattern of neurons forming a given lineage the "projection envelope" of that lineage. By inducing MARCM clones at the early larval stage, we labeled the secondary progeny of each neuroblast. For the supraesophageal ganglion excluding mushroom body (the part of the brain investigated in the present work) we obtained 81 different types of clones. Based on the trajectory of their secondary axon tracts (described in the accompanying paper, Lovick et al., 2013), we assigned these clones to specific lineages defined in the larva. Since a labeled clone reveals all aspects (cell bodies, axon tracts, terminal arborization) of a lineage, we were able to describe projection envelopes for all secondary lineages of the supraesophageal ganglion. This work provides a framework by which the secondary neurons (forming the vast majority of adult brain neurons) can be assigned to genetically and developmentally defined groups. It also represents a step towards the goal to establish, for each lineage, the link between its mature anatomical and functional phenotype, and the genetic make-up of the neuroblast it descends from. PMID:23872236

  9. Marriage, cohabitation and commitment

    OpenAIRE

    Iyigun, Murat

    2009-01-01

    This paper combines partner matching with an intra-household allocation model where couples decide if they want to marry or cohabitate. Marriage encourages but does not ensure a higher level of spousal commitment, which in turn can generate a larger marital surplus. Individuals’ marital preferences and commitment costs vary, and sorting equilibria are based on individuals’ marital preferences and propensity to commit. In all equilibria, some married couples are able to cooperate and operate e...

  10. The committed organic consumer

    OpenAIRE

    Geen, Natalie; Firth, Chris

    2006-01-01

    Abstract - UK committed organic consumers are reported to be about 23 per cent of consumers and account for 84 per cent of organic sales. This paper compares the committed organic consumer who is also a member of the UK’s national charity for organic growing (HDRA), to committed organic consumers in the wider UK population. Broad parallels were found between these two groups of consumers although there were differences in purchasing drivers as environmental issues were relatively more impor...

  11. Development of innate lymphoid cells.

    Science.gov (United States)

    Zook, Erin C; Kee, Barbara L

    2016-06-21

    Innate lymphoid cells (ILCs) are a family of immune effector cells that have important roles in host defense, metabolic homeostasis and tissue repair but can also contribute to inflammatory diseases such as asthma and colitis. These cells can be categorized into three groups on the basis of the transcription factors that direct their function and the cytokines they produce, which parallel the effector functions of T lymphocytes. The hierarchy of cell-fate-restriction events that occur as common lymphoid progenitors become committed to each of the ILC lineages further underscores the relationship between these innate immune cells and T lymphocytes. In this Review we discuss the developmental program of ILCs and transcription factors that guide ILC lineage specification and commitment. PMID:27328007

  12. Spatio-temporal Model of Endogenous ROS and Raft-Dependent WNT/Beta-Catenin Signaling Driving Cell Fate Commitment in Human Neural Progenitor Cells

    Science.gov (United States)

    Haack, Fiete; Lemcke, Heiko; Ewald, Roland; Rharass, Tareck; Uhrmacher, Adelinde M.

    2015-01-01

    Canonical WNT/β-catenin signaling is a central pathway in embryonic development, but it is also connected to a number of cancers and developmental disorders. Here we apply a combined in-vitro and in-silico approach to investigate the spatio-temporal regulation of WNT/β-catenin signaling during the early neural differentiation process of human neural progenitors cells (hNPCs), which form a new prospect for replacement therapies in the context of neurodegenerative diseases. Experimental measurements indicate a second signal mechanism, in addition to canonical WNT signaling, being involved in the regulation of nuclear β-catenin levels during the cell fate commitment phase of neural differentiation. We find that the biphasic activation of β-catenin signaling observed experimentally can only be explained through a model that combines Reactive Oxygen Species (ROS) and raft dependent WNT/β-catenin signaling. Accordingly after initiation of differentiation endogenous ROS activates DVL in a redox-dependent manner leading to a transient activation of down-stream β-catenin signaling, followed by continuous auto/paracrine WNT signaling, which crucially depends on lipid rafts. Our simulation studies further illustrate the elaborate spatio-temporal regulation of DVL, which, depending on its concentration and localization, may either act as direct inducer of the transient ROS/β-catenin signal or as amplifier during continuous auto-/parcrine WNT/β-catenin signaling. In addition we provide the first stochastic computational model of WNT/β-catenin signaling that combines membrane-related and intracellular processes, including lipid rafts/receptor dynamics as well as WNT- and ROS-dependent β-catenin activation. The model’s predictive ability is demonstrated under a wide range of varying conditions for in-vitro and in-silico reference data sets. Our in-silico approach is realized in a multi-level rule-based language, that facilitates the extension and modification of the

  13. Spatio-temporal model of endogenous ROS and raft-dependent WNT/beta-catenin signaling driving cell fate commitment in human neural progenitor cells.

    Directory of Open Access Journals (Sweden)

    Fiete Haack

    2015-03-01

    Full Text Available Canonical WNT/β-catenin signaling is a central pathway in embryonic development, but it is also connected to a number of cancers and developmental disorders. Here we apply a combined in-vitro and in-silico approach to investigate the spatio-temporal regulation of WNT/β-catenin signaling during the early neural differentiation process of human neural progenitors cells (hNPCs, which form a new prospect for replacement therapies in the context of neurodegenerative diseases. Experimental measurements indicate a second signal mechanism, in addition to canonical WNT signaling, being involved in the regulation of nuclear β-catenin levels during the cell fate commitment phase of neural differentiation. We find that the biphasic activation of β-catenin signaling observed experimentally can only be explained through a model that combines Reactive Oxygen Species (ROS and raft dependent WNT/β-catenin signaling. Accordingly after initiation of differentiation endogenous ROS activates DVL in a redox-dependent manner leading to a transient activation of down-stream β-catenin signaling, followed by continuous auto/paracrine WNT signaling, which crucially depends on lipid rafts. Our simulation studies further illustrate the elaborate spatio-temporal regulation of DVL, which, depending on its concentration and localization, may either act as direct inducer of the transient ROS/β-catenin signal or as amplifier during continuous auto-/parcrine WNT/β-catenin signaling. In addition we provide the first stochastic computational model of WNT/β-catenin signaling that combines membrane-related and intracellular processes, including lipid rafts/receptor dynamics as well as WNT- and ROS-dependent β-catenin activation. The model's predictive ability is demonstrated under a wide range of varying conditions for in-vitro and in-silico reference data sets. Our in-silico approach is realized in a multi-level rule-based language, that facilitates the extension and

  14. Spatio-temporal model of endogenous ROS and raft-dependent WNT/beta-catenin signaling driving cell fate commitment in human neural progenitor cells.

    Science.gov (United States)

    Haack, Fiete; Lemcke, Heiko; Ewald, Roland; Rharass, Tareck; Uhrmacher, Adelinde M

    2015-03-01

    Canonical WNT/β-catenin signaling is a central pathway in embryonic development, but it is also connected to a number of cancers and developmental disorders. Here we apply a combined in-vitro and in-silico approach to investigate the spatio-temporal regulation of WNT/β-catenin signaling during the early neural differentiation process of human neural progenitors cells (hNPCs), which form a new prospect for replacement therapies in the context of neurodegenerative diseases. Experimental measurements indicate a second signal mechanism, in addition to canonical WNT signaling, being involved in the regulation of nuclear β-catenin levels during the cell fate commitment phase of neural differentiation. We find that the biphasic activation of β-catenin signaling observed experimentally can only be explained through a model that combines Reactive Oxygen Species (ROS) and raft dependent WNT/β-catenin signaling. Accordingly after initiation of differentiation endogenous ROS activates DVL in a redox-dependent manner leading to a transient activation of down-stream β-catenin signaling, followed by continuous auto/paracrine WNT signaling, which crucially depends on lipid rafts. Our simulation studies further illustrate the elaborate spatio-temporal regulation of DVL, which, depending on its concentration and localization, may either act as direct inducer of the transient ROS/β-catenin signal or as amplifier during continuous auto-/parcrine WNT/β-catenin signaling. In addition we provide the first stochastic computational model of WNT/β-catenin signaling that combines membrane-related and intracellular processes, including lipid rafts/receptor dynamics as well as WNT- and ROS-dependent β-catenin activation. The model's predictive ability is demonstrated under a wide range of varying conditions for in-vitro and in-silico reference data sets. Our in-silico approach is realized in a multi-level rule-based language, that facilitates the extension and modification of the

  15. Modern genome-wide genetic approaches to reveal intrinsic properties of stem cells

    NARCIS (Netherlands)

    de Haan, Gerald; Gerrits, Alice; Bystrykh, Leonid

    2006-01-01

    Purpose of review The clinical use of hematopoietic stem cells, which produce all mature blood cell lineages in the circulation, is continuously increasing. Identification of genes and gene networks specifying either sternness or commitment will not only be of major relevance for a fundamental under

  16. Human embryonic stem cells as a model for cardiac gene discovery : from chip to chap

    NARCIS (Netherlands)

    Beqqali, A.

    2008-01-01

    Here we described the use of human embryonic stem cells (hESCs) as a model to obtain insights into commitment to the mesoderm and endoderm lineages and the early steps in human cardiac cell differentiation by means of whole-genome temporal expression profiling. Furthermore, we used it as an approach

  17. Implementing environmental commitments

    Energy Technology Data Exchange (ETDEWEB)

    Victor, D.G.; Skolnikoff, E.B. [IIASA, Laxenburg (Austria)

    1997-08-01

    The article summarises the aims and results of a three-year project conducted at the International Institute for Applied Systems Analysis called `Implementation and effectiveness of international environmental commitments` (IEC) which analysed how many environmental commitments are implemented or how intentions are put into practice. Fourteen case studies were conducted on the following issues: conservation and preservation of fauna and flora; stratospheric ozone depletion; Baltic sea pollution; trade in hazardous chemicals and pesticides; acid rain in Europe; North Sea pollution; whaling; and marine dumping of nuclear waste. The research confirmed previous findings that compliance with legally binding commitments is high. However, IEC results suggest that high compliance reflects mainly that governments make special effort to adopt only those commitments with which they can comply. The influence of those commitments is often low. In contrast, compliance with nonbinding commitments has been low, but such commitments can often have a large influence on behavior. The findings differs sharply from conventional wisdom, which maintains that the most effective international commitments are legally binding. The research suggests more attention is needed to building systems for gathering and reviewing data on implementation and for handling implementation problems. Among IEC`s contributions to policy has been to apply this finding to the design of possible non-compliance procedures within the United Nations Framework Convention on Climate Change.

  18. The Superiority of Allogeneic Hematopoietic Stem Cell Transplantation Over Chemotherapy Alone in the Treatment of Acute Myeloid Leukemia Patients with Mixed Lineage Leukemia (MLL) Rearrangements

    Science.gov (United States)

    Yang, Hua; Huang, Sai; Zhu, Cheng-Ying; Gao, Li; Zhu, Hai-Yan; Lv, Na; Jing, Yu; Yu, Li

    2016-01-01

    Background Acute myeloid leukemia (AML) patients with mixed lineage leukemia (MLL) gene rearrangements always had a very poor prognosis. In this study, we report the incidence of MLL rearrangements in AML patients using gene analysis, as well as the clinical significance and prognostic features of these rearrangements. Material/Methods This retrospective study took place from April 2008 to November 2011 in the People’s Liberation Army General Hospital. A total 433 AML patients were screened by multiple nested reverse transcription polymerase chain reaction (RT-PCR) to determine the incidence of the 11 MLL gene rearrangements. There were 68 cases of MLL gene rearrangements, for a positive rate of 15.7%. A total of 24 patients underwent allogeneic hematopoietic stem cell transplantation (Allo-HSCT), and 34 patients received at least 4 cycles of chemotherapy. Ten patients were lost to follow-up. Results The median follow-up was 29 months. The complete remission (CR) rate was 85.4%. The overall survival (OS) was 57.4±5.9 months for the Allo-HSCT group and 21.0±2.1 months for the chemotherapy group. The Allo-HSCT group had superior survival compared with the chemotherapy group (5-year OS: 59±17% vs. 13±8%, P0.05). Multivariate analysis showed that transplantation, platelets >50×109/L at onset, and CR are associated with a better OS in MLL rearranged AML patients. Patients with thrombocytopenia and extramedullary involvement were prone to relapse. Conclusions Our results suggest that Allo-HSCT is superior to chemotherapy alone for treating MLL rearranged AML patients. Patients treated with Allo-HSCT have a better prognosis and a longer survival. CR is an independent prognostic factor for OS, and extramedullary involvement is an independent prognostic factor for DFS. MLL rearranged AML patients with thrombocytopenia at onset <50×109 had very bad OS and DFS. PMID:27373985

  19. Regional and Stage-Specific Effects of Prospectively Purified Vascular Cells on the Adult V-SVZ Neural Stem Cell Lineage

    OpenAIRE

    Crouch, Elizabeth E.; Liu, Chang; Silva-Vargas, Violeta; Doetsch, Fiona

    2015-01-01

    Adult neural stem cells reside in specialized niches. In the ventricular-subventricular zone (V-SVZ), quiescent neural stem cells (qNSCs) become activated (aNSCs), and generate transit amplifying cells (TACs), which give rise to neuroblasts that migrate to the olfactory bulb. The vasculature is an important component of the adult neural stem cell niche, but whether vascular cells in neurogenic areas are intrinsically different from those elsewhere in the brain is unknown. Moreover, the contri...

  20. Redox and Metabolic Regulation of Stem/Progenitor Cells and Their Niche

    OpenAIRE

    Ushio-Fukai, Masuko; Rehman, Jalees

    2014-01-01

    Stem cells are defined as cells that have the capacity to self-renew and exhibit multipotency or pluripotency, whereas progenitor cells are committed to selected lineages but retain their self-renewal capacity. The stem or progenitor cell niche refers to the microenvironment of the regenerative cells in the bone marrow (BM) or other tissues such as the heart. It can regulate self-renewal, differentiation, migration, and proliferation of regenerative stem/progenitor cells. The precise regulato...

  1. Controlling Self-Renewal and Differentiation of Stem Cells via Mechanical Cues

    OpenAIRE

    Nava, Michele M.; Raimondi, Manuela T.; Riccardo Pietrabissa

    2012-01-01

    The control of stem cell response in vitro, including self-renewal and lineage commitment, has been proved to be directed by mechanical cues, even in the absence of biochemical stimuli. Through integrin-mediated focal adhesions, cells are able to anchor onto the underlying substrate, sense the surrounding microenvironment, and react to its properties. Substrate-cell and cell-cell interactions activate specific mechanotransduction pathways that regulate stem cell fate. Mechanical factors, incl...

  2. PEG10 Activation by Co-Stimulation of CXCR5 and CCR7 Essentially Contributes to Resistance to Apoptosis in CD19+CD34+ B Cells from Patients with B Cell Lineage Acute and Chronic Lymphocytic Leukemia

    Institute of Scientific and Technical Information of China (English)

    ChunsongHu; JeiXiong; LinjeiZhang; BaojunHuang; QiupingZhang; QunLi; MingzhenYang; YaouWu; QunWu; QianShen; QingpingGao; KejianZhang; ZhiminSun; JunyanLin; YouxinJin

    2004-01-01

    We investigated CD19+CD34+ and CD19+CD34 B cells from cord blood (CB) and typical patients with B cell lineage acute and chronic lymphocytic leukemia (B-ALL and B-CLL) in terms of expression and functions of CXCR5/CXCL13 and CCR7/CCL19. CXCR5 and CCR7 were selectively frequent expressed on B-ALL, B-CLL and CB CD19+CD34+ B cells, but not on CD19+CD34- B cells. Instead of induction of impressive chemotactic responsiveness, CXCL13 and CCL19 together induced significant resistance to TNF-α-mediated apoptosis in B-ALL and B-CLL but not CB CD19+CD34+ B cells. B-ALL and B-CLL CD19+CD34+ B cells expressed elevatedlevel of Paternally Expressed Gene 10 (PEG10), and CXCL13 and CCL19 together significantly up-regulated PEG10 expression in the cells. We found that CXCL13 and CCL19 together by means of activation of CXCR5 and CCR7 up-regulated PEG10 expression and function, subsequent stabilized caspase-3 and caspase-8 in B-ALL and B-CLL CD19+CD34+ B cells, and rescued the cells from TNF-α-mediated apoptosis. We suggested that normal lymphocytes, especially naive B and T cells, utilized CXCR5/CXCL13 and CCR7/CCL19 for migration, homing, maturation, and cell homeostasis as well as secondary lymphoid tissues organogenesis. Meanwhile certain malignant cells took advantages of CXCR5/CXCL13 and CCR7/CCL19 for infiltration, resistance to apoptosis, and inappropriate proliferation. Cellular & Molecular Immunology.

  3. PEG10 Activation by Co-Stimulation of CXCR5 and CCR7 Essentially Contributes to Resistance to Apoptosis in CD19+CD34+ B Cells from Patients with B Cell Lineage Acute and Chronic Lymphocytic Leukemia

    Institute of Scientific and Technical Information of China (English)

    Chunsong Hu; Qian Shen; Qingping Gao; Kejian Zhang; Zhimin Sun; Junyan Liu; Youxin Jin; Jinquan Tan; Jei Xiong; Linjei zhang; Baojun Huang; Qiuping Zhang; Qun Li; Mingzhen Yang; Yaou Wu; Qun Wu

    2004-01-01

    We investigated CD19+CD34+ and CD19+CD34- B cells from cord blood (CB) and typical patients with B cell lineage acute and chronic lymphocytic leukemia (B-ALL and B-CLL) in terms of expression and functions of CXCR5/CXCL13 and CCR7/CCL19. CXCR5 and CCR7 were selectively frequent expressed on B-ALL, B-CLL and CB CD19+CD34+ B cells, but not on CD19+CD34- B cells. Instead of induction of impressive chemotactic responsiveness, CXCL13 and CCL19 together induced significant resistance to TNF-α-mediated apoptosis in B-ALL and B-CLL but not CB CD19+CD34+ B cells. B-ALL and B-CLL CD19+CD34+ B cells expressed elevated level of Paternally Expressed Gene 10 (PEG10), and CXCL13 and CCL19 together significantly up-regulated PEG10 expression in the cells. We found that CXCL13 and CCL19 together by means of activation of CXCR5 and CCR7 up-regulated PEG10 expression and function, subsequent stabilized caspase-3 and caspase-8 in B-ALL and B-CLL CD19+CD34+ B cells, and rescued the cells from TNF-α-mediated apoptosis. We suggested that normal lymphocytes, especially na(I)ve B and T cells, utilized CXCR5/CXCL13 and CCR7/CCL19 for migration, homing, maturation, and cell homeostasis as well as secondary lymphoid tissues organogenesis.Meanwhile certain malignant cells took advantages of CXCR5/CXCL13 and CCR7/CCL19 for infiltration,resistance to apoptosis, and inappropriate proliferation.

  4. Acinar phenotype is preserved in human exocrine pancreas cells cultured at low temperature: implications for lineage-tracing of β-cell neogenesis.

    Science.gov (United States)

    Mfopou, Josué K; Houbracken, Isabelle; Wauters, Elke; Mathijs, Iris; Song, Imane; Himpe, Eddy; Baldan, Jonathan; Heimberg, Harry; Bouwens, Luc

    2016-06-01

    The regenerative medicine field is expanding with great successes in laboratory and preclinical settings. Pancreatic acinar cells in diabetic mice were recently converted into β-cells by treatment with ciliary neurotrophic factor (CNTF) and epidermal growth factor (EGF). This suggests that human acinar cells might become a cornerstone for diabetes cell therapy in the future, if they can also be converted into glucose-responsive insulin-producing cells. Presently, studying pancreatic acinar cell biology in vitro is limited by their high plasticity, as they rapidly lose their phenotype and spontaneously transdifferentiate to a duct-like phenotype in culture. We questioned whether human pancreatic acinar cell phenotype could be preserved in vitro by physico-chemical manipulations and whether this could be valuable in the study of β-cell neogenesis. We found that culture at low temperature (4°C) resulted in the maintenance of morphological and molecular acinar cell characteristics. Specifically, chilled acinar cells did not form the spherical clusters observed in controls (culture at 37°C), and they maintained high levels of acinar-specific transcripts and proteins. Five-day chilled acinar cells still transdifferentiated into duct-like cells upon transfer to 37°C. Moreover, adenoviral-mediated gene transfer evidenced an active Amylase promoter in the 7-day chilled acinar cells, and transduction performed in chilled conditions improved acinar cell labelling. Together, our findings indicate the maintenance of human pancreatic acinar cell phenotype at low temperature and the possibility to efficiently label acinar cells, which opens new perspectives for the study of human acinar-to-β-cell transdifferentiation. PMID:26987985

  5. A case of cellular alchemy: lineage reprogramming and its potential in regenerative medicine.

    Science.gov (United States)

    Asuelime, Grace E; Shi, Yanhong

    2012-08-01

    The field of regenerative medicine is rapidly gaining momentum as an increasing number of reports emerge concerning the induced conversions observed in cellular fate reprogramming. While in recent years, much attention has been focused on the conversion of fate-committed somatic cells to an embryonic-like or pluripotent state, there are still many limitations associated with the applications of induced pluripotent stem cell reprogramming, including relatively low reprogramming efficiency, the times required for the reprogramming event to take place, the epigenetic instability, and the tumorigenicity associated with the pluripotent state. On the other hand, lineage reprogramming involves the conversion from one mature cell type to another without undergoing conversion to an unstable intermediate. It provides an alternative approach in regenerative medicine that has a relatively lower risk of tumorigenesis and increased efficiency within specific cellular contexts. While lineage reprogramming provides exciting potential, there is still much to be assessed before this technology is ready to be applied in a clinical setting. PMID:22371436

  6. Distinct functions of the transcription factors Gata3 and ThPOK during intrathymic CD4 T cell differentiation*

    OpenAIRE

    Wang, Lie; Wildt, Kathryn F.; Zhu, Jinfang; Zhang, Xianyu; Feigenbaum, Lionel; Tessarollo, Lino; Paul, William E.; Fowlkes, B.J.; Bosselut, Rémy

    2008-01-01

    The transcription factors Gata3 and Zbtb7b are required for intrathymic CD4 T cell differentiation, but their precise roles in this process remain unclear. Here we show that, contrary to previous findings, Gata3 disruption blocked CD4 T cell lineage differentiation before CD4 lineage commitment, and in some contexts permitted ‘redirection’ of MHC class II-restricted thymocytes into the CD8 lineage. We found that Gata3 promotes Zbtb7b expression, and binds within a region of the Zbtb7b locus e...

  7. Neuroblast lineage identification and lineage-specific Hox gene action during postembryonic development of the subesophageal ganglion in the Drosophila central brain.

    Science.gov (United States)

    Kuert, Philipp A; Hartenstein, Volker; Bello, Bruno C; Lovick, Jennifer K; Reichert, Heinrich

    2014-06-15

    The central brain of Drosophila consists of the supraesophageal ganglion (SPG) and the subesophageal ganglion (SEG), both of which are generated by neural stem cell-like neuroblasts during embryonic and postembryonic development. Considerable information has been obtained on postembryonic development of the neuroblasts and their lineages in the SPG. In contrast, very little is known about neuroblasts, neural lineages, or any other aspect of the postembryonic development in the SEG. Here we characterize the neuroanatomy of the larval SEG in terms of tracts, commissures, and other landmark features as compared to a thoracic ganglion. We then use clonal MARCM labeling to identify all adult-specific neuroblast lineages in the late larval SEG and find a surprisingly small number of neuroblast lineages, 13 paired and one unpaired. The Hox genes Dfd, Scr, and Antp are expressed in a lineage-specific manner in these lineages during postembryonic development. Hox gene loss-of-function causes lineage-specific defects in axonal targeting and reduction in neural cell numbers. Moreover, it results in the formation of novel ectopic neuroblast lineages. Apoptosis block also results in ectopic lineages suggesting that Hox genes are required for lineage-specific termination of proliferation through programmed cell death. Taken together, our findings show that postembryonic development in the SEG is mediated by a surprisingly small set of identified lineages and requires lineage-specific Hox gene action to ensure the correct formation of adult-specific neurons in the Drosophila brain. PMID:24713419

  8. Social Consequences of Commitment

    OpenAIRE

    Alan G. Isaac

    2006-01-01

    This paper begins with a detailed computational introduction to a classic ACE model: an evolutionary prisoner's dilemma. The paper presents a simple but fully coded object oriented implementation of this model. (We use the Python programming language, which is shown to be a natural ally for ACE research). Using these tools, we demonstrate that player type evolution is affected by cardinal payoffs. We then explore a possible social benefit to commitment, where 'commitment'...

  9. Derivation of mesenchymal stromal cells from pluripotent stem cells through a neural crest lineage using small molecule compounds with defined media.

    Directory of Open Access Journals (Sweden)

    Makoto Fukuta

    Full Text Available Neural crest cells (NCCs are an embryonic migratory cell population with the ability to differentiate into a wide variety of cell types that contribute to the craniofacial skeleton, cornea, peripheral nervous system, and skin pigmentation. This ability suggests the promising role of NCCs as a source for cell-based therapy. Although several methods have been used to induce human NCCs (hNCCs from human pluripotent stem cells (hPSCs, such as embryonic stem cells (ESCs and induced pluripotent stem cells (iPSCs, further modifications are required to improve the robustness, efficacy, and simplicity of these methods. Chemically defined medium (CDM was used as the basal medium in the induction and maintenance steps. By optimizing the culture conditions, the combination of the GSK3β inhibitor and TGFβ inhibitor with a minimum growth factor (insulin very efficiently induced hNCCs (70-80% from hPSCs. The induced hNCCs expressed cranial NCC-related genes and stably proliferated in CDM supplemented with EGF and FGF2 up to at least 10 passages without changes being observed in the major gene expression profiles. Differentiation properties were confirmed for peripheral neurons, glia, melanocytes, and corneal endothelial cells. In addition, cells with differentiation characteristics similar to multipotent mesenchymal stromal cells (MSCs were induced from hNCCs using CDM specific for human MSCs. Our simple and robust induction protocol using small molecule compounds with defined media enabled the generation of hNCCs as an intermediate material producing terminally differentiated cells for cell-based innovative medicine.

  10. Foxp3(+) T cells expressing RORγt represent a stable regulatory T-cell effector lineage with enhanced suppressive capacity during intestinal inflammation.

    OpenAIRE

    Yang, B-H; S. Hagemann; Mamareli, P; Lauer, U; Hoffmann, U; Beckstette, M; Föhse, L; Prinz, I.; Pezoldt, J; Suerbaum, S.; Sparwasser, Tim; Hamann, A.; Floess, S; Huehn, J.; Lochner, M

    2016-01-01

    Foxp3 (forkhead box P3 transcription factor)-expressing regulatory T cells (Tregs) are essential for immunological tolerance, best illustrated by uncontrolled effector T-cell responses and autoimmunity upon loss of Foxp3 expression. Tregs can adopt specific effector phenotypes upon activation, reflecting the diversity of functional demands in the different tissues of the body. Here, we report that Foxp3(+)CD4(+) T cells coexpressing retinoic acid-related orphan receptor-γt (RORγt), the master...

  11. Sex determining region Y-box 2 (SOX2) is a potential cell-lineage gene highly expressed in the pathogenesis of squamous cell carcinomas of the lung

    OpenAIRE

    Yuan, Ping; Kadara, Humam; Behrens, Carmen; Tang, Ximing; Woods, Denise; Luisa M Solis; Huang, Jiaoti; Spinola, Monica; Dong, Wenli; Yin, Guosheng; Fujimoto, Junya; Kim, Edward; Xie, Yang; Girard, Luc; Moran, Cesar

    2010-01-01

    Background: Non-small cell lung cancer (NSCLC) represents the majority (85%) of lung cancers and is comprised mainly of adenocarcinomas and squamous cell carcinomas (SCCs). The sequential pathogenesis of lung adenocarcinomas and SCCs occurs through dissimilar phases as the former tumors typically arise in the lung periphery whereas the latter normally arise near the central airway. Methodology/Principal Findings:We assessed the expression of SOX2, an embryonic stem cell transcriptional factor...

  12. PDX1 Binds and Represses Hepatic Genes to Ensure Robust Pancreatic Commitment in Differentiating Human Embryonic Stem Cells

    Directory of Open Access Journals (Sweden)

    Adrian Kee Keong Teo

    2015-04-01

    Full Text Available Inactivation of the Pancreatic and Duodenal Homeobox 1 (PDX1 gene causes pancreatic agenesis, which places PDX1 high atop the regulatory network controlling development of this indispensable organ. However, little is known about the identity of PDX1 transcriptional targets. We simulated pancreatic development by differentiating human embryonic stem cells (hESCs into early pancreatic progenitors and subjected this cell population to PDX1 chromatin immunoprecipitation sequencing (ChIP-seq. We identified more than 350 genes bound by PDX1, whose expression was upregulated on day 17 of differentiation. This group included known PDX1 targets and many genes not previously linked to pancreatic development. ChIP-seq also revealed PDX1 occupancy at hepatic genes. We hypothesized that simultaneous PDX1-driven activation of pancreatic and repression of hepatic programs underlie early divergence between pancreas and liver. In HepG2 cells and differentiating hESCs, we found that PDX1 binds and suppresses expression of endogenous liver genes. These findings rebrand PDX1 as a context-dependent transcriptional repressor and activator within the same cell type.

  13. Effect of nano- and micro-scale topological features on alignment of muscle cells and commitment of myogenic differentiation

    International Nuclear Information System (INIS)

    Skeletal muscle injury can lead to severe motor deficits that adversely affect movement and quality of life. Current surgical treatments for skeletal muscle are hindered by the poor formation of organized myotube bundles at the wound site. Tissue-engineered skeletal muscle constructs to date have been unable to generate high degrees of myotube density and alignment. Generating a suitable in vitro tissue-engineered skeletal muscle construct requires the design of a scaffold that recapitulates the structural combination of nanoscale collagen fibrils and aligned microscale basal lamina tracks present in the native extracellular matrix (ECM). We hypothesized that a 3D aligned tubular porous scaffold containing aligned nanofibers inside the pores can mimic the native muscle tissue environment. We constructed a laminar section of the hypothesized scaffold with aligned chitosan-PCL nanofibers arranged co-axially with the aligned microscale chitosan scaffold bands to mimic the required myogenic environment. A 6-day study of C2C12 mouse myoblast cells cultured on this hybrid scaffold indicated that the nanofibers and scaffold bands in the scaffold played a synergetic role in directing cell orientation, interaction, migration and organization. Our results showed that aligned nanofibers mediated cell alignment and the aligned scaffold bands induced the formation of a more compact assembly of myotube cells as compared to various control substrates including chitosan films, nanofibers, and chitosan bands. The expression levels of both early and late-stage myogenic differentiation genes associated with myogenin and myosin heavy chain, respectively, were higher on the hybrid substrate than on control substrates. Our study suggests that the combination of nano and microscale topological features in the ECM can direct myogenic differentiation, and the hybrid material has the potential to improve the outcome of skeletal tissue engineering. (papers)

  14. NFIB is a governor of epithelial–melanocyte stem cell behaviour in a shared niche

    OpenAIRE

    Chang, Chiung-Ying; Pasolli, H. Amalia; Giannopoulou, Eugenia G.; Guasch, Géraldine; Gronostajski, Richard M; Elemento, Olivier; Fuchs, Elaine

    2013-01-01

    Adult stem cells reside in specialized niches where they receive environmental cues to maintain tissue homeostasis. In mammals, the stem cell niche within hair follicles is home to epithelial hair follicle stem cells and melanocyte stem cells, which sustain cyclical bouts of hair regeneration and pigmentation1–4. To generate pigmented hairs, synchrony is achieved such that upon initiation of a new hair cycle, stem cells of each type activate lineage commitment2,5. Dissecting the inter-stem-ce...

  15. Walking along the Fibroblast Growth Factor 10 Route: A Key Pathway to Understand the Control and Regulation of Epithelial and Mesenchymal Cell-Lineage Formation during Lung Development and Repair after Injury.

    Science.gov (United States)

    El Agha, Elie; Bellusci, Saverio

    2014-01-01

    Basic research on embryonic lung development offers unique opportunities to make important discoveries that will impact human health. Developmental biologists interested in the molecular control of branching morphogenesis have intensively studied the developing lung, with its complex and seemingly stereotyped ramified structure. However, it is also an organ that is linked to a vast array of clinical problems in humans such as bronchopulmonary dysplasia in premature babies and emphysema, chronic obstructive pulmonary disease, fibrosis, and cancer in adults. Epithelial stem/progenitor cells reside in niches where they interact with specific extracellular matrices as well as with mesenchymal cells; the latter are still poorly characterized. Interactions of epithelial stem/progenitor cells with their microenvironments are usually instructive, controlling quiescence versus activation, proliferation, differentiation, and migration. During the past 18 years, Fgf10 has emerged not only as a marker for the distal lung mesenchyme during early lung development, but also as a key player in branching morphogenesis and a critical component of the niche for epithelial stem cells. In this paper, we will present the current knowledge regarding the lineage tree in the lung, with special emphasis on cell-lineage decisions in the lung mesenchyme and the role of Fgf10 in this context. PMID:25298902

  16. Walking along the Fibroblast Growth Factor 10 Route: A Key Pathway to Understand the Control and Regulation of Epithelial and Mesenchymal Cell-Lineage Formation during Lung Development and Repair after Injury

    Directory of Open Access Journals (Sweden)

    Elie El Agha

    2014-01-01

    Full Text Available Basic research on embryonic lung development offers unique opportunities to make important discoveries that will impact human health. Developmental biologists interested in the molecular control of branching morphogenesis have intensively studied the developing lung, with its complex and seemingly stereotyped ramified structure. However, it is also an organ that is linked to a vast array of clinical problems in humans such as bronchopulmonary dysplasia in premature babies and emphysema, chronic obstructive pulmonary disease, fibrosis, and cancer in adults. Epithelial stem/progenitor cells reside in niches where they interact with specific extracellular matrices as well as with mesenchymal cells; the latter are still poorly characterized. Interactions of epithelial stem/progenitor cells with their microenvironments are usually instructive, controlling quiescence versus activation, proliferation, differentiation, and migration. During the past 18 years, Fgf10 has emerged not only as a marker for the distal lung mesenchyme during early lung development, but also as a key player in branching morphogenesis and a critical component of the niche for epithelial stem cells. In this paper, we will present the current knowledge regarding the lineage tree in the lung, with special emphasis on cell-lineage decisions in the lung mesenchyme and the role of Fgf10 in this context.

  17. The effect of glow discharge plasma surface modification of polymers on the osteogenic differentiation of committed human mesenchymal stem cells.

    Science.gov (United States)

    Mwale, Fackson; Wang, Hong Tian; Nelea, Valentin; Luo, Li; Antoniou, John; Wertheimer, Michael R

    2006-04-01

    Little is known of the effect of material surfaces on stem cell differentiation. The present study has addressed the hypothesis that the interaction of mesenchymal stem cells (MSCs) with material surfaces modified by glow discharge plasma is a major regulator of osteogenic differentiation. We found that biaxially oriented polypropylene (BOPP) plasma treated in ammonia significantly reduced up-regulation of expression of osteogenic marker genes, such as alkaline phosphatase (ALP), bone sialoprotein (BSP) and osteocalcin (OC). In contrast, ALP expression was up-regulated when cultured on treated Nylon-6 polyamide (Ny-t) but was substantially reduced when cultured on its pristine counterpart (Ny-p) on day 3. On day 7, ALP expression was down-regulated with MSCs cultured on Ny-t although its expression level was up again on day 14. BSP was expressed weakly on day 3, but was up-regulated when cultured on Ny-t and Ny-p. Its expression reached its maximum on day 14 when cultured on a polystyrene control, while it was cyclically up-regulated on Ny-t. Similarly, there was a slight increase in OC expression when MSCs were cultured on Ny-t and Ny-p on day 3, when compared to control. Thus, the nature of the surface can directly influence MSCs differentiation, ultimately affecting the quality of new tissue formation with BOPP-t suppressing osteogenic differentiation. PMID:16313952

  18. LATS-YAP/TAZ controls lineage specification by regulating TGFβ signaling and Hnf4α expression during liver development

    Science.gov (United States)

    Lee, Da-Hye; Park, Jae Oh; Kim, Tae-Shin; Kim, Sang-Kyum; Kim, Tack-hoon; Kim, Min-chul; Park, Gun Soo; Kim, Jeong-Hwan; Kuninaka, Shinji; Olson, Eric N.; Saya, Hideyuki; Kim, Seon-Young; Lee, Ho; Lim, Dae-Sik

    2016-01-01

    The Hippo pathway regulates the self-renewal and differentiation of various adult stem cells, but its role in cell fate determination and differentiation during liver development remains unclear. Here we report that the Hippo pathway controls liver cell lineage specification and proliferation separately from Notch signalling, using mice and primary hepatoblasts with liver-specific knockout of Lats1 and Lats2 kinase, the direct upstream regulators of YAP and TAZ. During and after liver development, the activation of YAP/TAZ induced by loss of Lats1/2 forces hepatoblasts or hepatocytes to commit to the biliary epithelial cell (BEC) lineage. It increases BEC and fibroblast proliferation by up-regulating TGFβ signalling, but suppresses hepatoblast to hepatocyte differentiation by repressing Hnf4α expression. Notably, oncogenic YAP/TAZ activation in hepatocytes induces massive p53-dependent cell senescence/death. Together, our results reveal that YAP/TAZ activity levels govern liver cell differentiation and proliferation in a context-dependent manner. PMID:27358050

  19. Antibody producing B lineage cells invade the central nervous system predominantly at the time of and triggered by acute Epstein-Barr virus infection: A hypothesis on the origin of intrathecal immunoglobulin synthesis in multiple sclerosis.

    Science.gov (United States)

    Otto, Carolin; Hofmann, Jörg; Ruprecht, Klemens

    2016-06-01

    Patients with multiple sclerosis (MS), a chronic inflammatory disease of the central nervous system (CNS), typically have an intrathecal synthesis of immunoglobulin (Ig)G. Intrathecal IgG is produced by B lineage cells that entered the CNS, but why and when these cells invade the CNS of patients with MS is unknown. The intrathecal IgG response in patients with MS is polyspecific and part of it is directed against different common viruses (e.g. measles virus, rubella virus, varicella zoster virus). Strong and consistent evidence suggests an association of MS and Epstein-Barr virus (EBV) infection and EBV seroprevalence in patients with MS is practically 100%. However, intriguingly, despite of the universal EBV seroprevalence, the frequency of intrathecally produced IgG to EBV in patients with MS is much lower than that of intrathecally produced IgG to other common viruses. The acute phase of primary EBV infection is characterized by a strong polyclonal B cell activation. As typical for humoral immune responses against viruses, EBV specific IgG is produced only with a temporal delay after acute EBV infection. Aiming to put the above facts into a logical structure, we here propose the hypothesis that in individuals going on to develop MS antibody producing B lineage cells invade the CNS predominantly at the time of and triggered by acute primary EBV infection. Because at the time of acute EBV infection EBV IgG producing B lineage cells have not yet occurred, the hypothesis could explain the universal EBV seroprevalence and the low frequency of intrathecally produced IgG to EBV in patients with MS. Evidence supporting the hypothesis could be provided by large prospective follow-up studies of individuals with symptomatic primary EBV infection (infectious mononucleosis). Furthermore, the clarification of the molecular mechanism underlying an EBV induced invasion of B lineage cells into the CNS of individuals going on to develop MS could corroborate it, too. If true, our

  20. Wnt-dependent osteogenic commitment of bone marrow stromal cells using a novel GSK3β inhibitor

    Directory of Open Access Journals (Sweden)

    David A. Cook

    2014-03-01

    Using human BMSCs grown in adipogenic medium, we confirmed that AR28-mediated Wnt signalling caused a significant (p < 0.05 dose-dependent reduction of adipogenic markers. In osteogenic media, including dexamethasone, AR28 caused significant (p < 0.05 decreases in alkaline phosphatase (ALP activity compared to vehicle controls, indicative of a reduced osteogenic response. However, when excluding dexamethasone from the osteogenic media, increases in both ALP and mineralisation were identified following AR28 treatment, which was blocked by mitomycin C. Pre-treatment of BMSCs with AR28 for 7 days before osteogenic induction also increased ALP activity and mineralisation. Furthermore, BMP2-induced osteogenic differentiation was strongly enhanced by AR28 addition within 3 days, but without concomitant changes in cell number, therefore revealing BMP-dependent and independent mechanisms for Wnt-induced osteogenesis.

  1. Genetic Mosaics and the Germ Line Lineage

    Directory of Open Access Journals (Sweden)

    Mark E. Samuels

    2015-04-01

    Full Text Available Genetic mosaics provide information about cellular lineages that is otherwise difficult to obtain, especially in humans. De novo mutations act as cell markers, allowing the tracing of developmental trajectories of all descendants of the cell in which the new mutation arises. De novo mutations may arise at any time during development but are relatively rare. They have usually been observed through medical ascertainment, when the mutation causes unusual clinical signs or symptoms. Mutational events can include aneuploidies, large chromosomal rearrangements, copy number variants, or point mutations. In this review we focus primarily on the analysis of point mutations and their utility in addressing questions of germ line versus somatic lineages. Genetic mosaics demonstrate that the germ line and soma diverge early in development, since there are many examples of combined somatic and germ line mosaicism for de novo mutations. The occurrence of simultaneous mosaicism in both the germ line and soma also shows that the germ line is not strictly clonal but arises from at least two, and possibly multiple, cells in the embryo with different ancestries. Whole genome or exome DNA sequencing technologies promise to expand the range of studies of genetic mosaics, as de novo mutations can now be identified through sequencing alone in the absence of a medical ascertainment. These technologies have been used to study mutation patterns in nuclear families and in monozygotic twins, and in animal model developmental studies, but not yet for extensive cell lineage studies in humans.

  2. Identifying neuronal lineages of Drosophila by sequence analysis of axon tracts

    OpenAIRE

    Cardona, A; Saalfeld, S; Arganda, I; Pereanu, W; Schindelin, J; Hartenstein, V.

    2010-01-01

    The Drosophila brain is formed by an invariant set of lineages, each of which is derived from a unique neural stem cell (neuroblast) and forms a genetic and structural unit of the brain. The task of reconstructing brain circuitry at the level of individual neurons can be made significantly easier by assigning neurons to their respective lineages. In this paper we address the automatization of neuron and lineage identification. We focused on the Drosophila brain lineages at the larval stage wh...

  3. Patterns of growth, axonal extension and axonal arborization of neuronal lineages in the developing Drosophila brain

    OpenAIRE

    Larsen, Camilla; Shy, Diana; Spindler, Shana R; Fung, Siaumin; Pereanu, Wayne; Younossi -Hartenstein, Amelia; Hartenstein, Volker

    2009-01-01

    The Drosophila central brain is composed of approximately 100 paired lineages, with most lineages comprising 100–150 neurons. Most lineages have a number of important characteristics in common. Typically, neurons of a lineage stay together as a coherent cluster and project their axons into a coherent bundle visible from late embryo to adult. Neurons born during the embryonic period form the primary axon tracts (PATs) that follow stereotyped pathways in the neuropile. Apoptotic cell death remo...

  4. Programming CD8+ T cells for effective immunotherapy

    OpenAIRE

    Hinrichs, Christian S.; Gattinoni, Luca; Restifo, Nicholas P

    2006-01-01

    The differentiation state of CD8+ T cells has emerged as a crucial determinant of their ability to respond to tumor and infection. Signals from T-cell receptors, co-stimulatory molecules and cytokine receptors direct the differentiation process. These signals ‘program’ sustained and heritable gene expression patterns that govern progressive differentiation and lineage commitment. The epigenetic mechanisms by which T cells are programmed are just beginning to be elucidated. Understanding the m...

  5. Improved outcome with hematopoietic stem cell transplantation in a poor prognostic subgroup of infants with mixed-lineage-leukemia (MLL)-rearranged acute lymphoblastic leukemia: results from the Interfant-99 Study

    DEFF Research Database (Denmark)

    Mann, Georg; Attarbaschi, Andishe; Schrappe, Martin;

    2010-01-01

    To define a role for hematopoietic stem cell transplantation (HSCT) in infants with acute lymphoblastic leukemia and rearrangements of the mixed-lineage-leukemia gene (MLL(+)), we compared the outcome of MLL(+) patients from trial Interfant-99 who either received chemotherapy only or HSCT. Of 376...... such high-risk criteria, with 87 achieving CR. In this group, HSCT was associated with a 64% reduction in the risk of failure resulting from relapse or death in CR (hazard ratio = 0.36, 95% confidence interval, 0.15-0.86). In the remaining patients, there was no advantage for HSCT over chemotherapy only...

  6. Ancestral reconstruction of tick lineages.

    Science.gov (United States)

    Mans, Ben J; de Castro, Minique H; Pienaar, Ronel; de Klerk, Daniel; Gaven, Philasande; Genu, Siyamcela; Latif, Abdalla A

    2016-06-01

    Ancestral reconstruction in its fullest sense aims to describe the complete evolutionary history of a lineage. This depends on accurate phylogenies and an understanding of the key characters of each parental lineage. An attempt is made to delineate our current knowledge with regard to the ancestral reconstruction of the tick (Ixodida) lineage. Tick characters may be assigned to Core of Life, Lineages of Life or Edges of Life phenomena depending on how far back these characters may be assigned in the evolutionary Tree of Life. These include housekeeping genes, sub-cellular systems, heme processing (Core of Life), development, moulting, appendages, nervous and organ systems, homeostasis, respiration (Lineages of Life), specific adaptations to a blood-feeding lifestyle, including the complexities of salivary gland secretions and tick-host interactions (Edges of Life). The phylogenetic relationships of lineages, their origins and importance in ancestral reconstruction are discussed. Uncertainties with respect to systematic relationships, ancestral reconstruction and the challenges faced in comparative transcriptomics (next-generation sequencing approaches) are highlighted. While almost 150 years of information regarding tick biology have been assembled, progress in recent years indicates that we are in the infancy of understanding tick evolution. Even so, broad reconstructions can be made with relation to biological features associated with various lineages. Conservation of characters shared with sister and parent lineages are evident, but appreciable differences are present in the tick lineage indicating modification with descent, as expected for Darwinian evolutionary theory. Many of these differences can be related to the hematophagous lifestyle of ticks. PMID:26868413

  7. From controlled to committed.

    Science.gov (United States)

    Hess, J C

    1996-02-01

    Most of us agree that people are our most important resource. Yet we spend a minimal amount of time learning more about human behavior, communication, and how our attitudes and behavior impact employee performance. Instead we rely on traditional methods of negative reinforcement in an attempt to control our areas of responsibility. While these methods can render some short-term success, managers and organizations that succeed during these times of change and fierce competition will be those that take the time to understand and capture the power of a committed workforce. The committed workforce is energized, not simply compliant, as a result of having basic human needs for achievement satisfied, belonging to a group, and receiving recognition for its contributions. Committed workers typically describe the manager as one who has the ability to give them a great degree of control over their area of influence. We all know that we don't change our leadership style like we change clothes. Old habits die hard. it takes a personal commitment and lots of practice to rid outselves of habits and behavior that no longer serve our departments and facilities. This commitment, however, is crucial to survival. As managers, we must cope with increasing ambiguity and uncertainty in the workplace. To survive these challenges, we must improve our interpersonal skills and ability to successfully bring out the best in others. I believe that success will continue for managers who not only increase their knowledge and technical ability, but who also inspire their workers to move forward with a collective sense of enthusiasm and purpose. PMID:10154218

  8. Enhancer repertoires are reshaped independently of early priming and heterochromatin dynamics during B cell differentiation

    OpenAIRE

    Choukrallah, Mohamed-Amin; Song, Shuang; Rolink, Antonius G.; Burger, Lukas; Matthias, Patrick

    2015-01-01

    A widely accepted model posits that activation of enhancers during differentiation goes through a priming step prior to lineage commitment. To investigate the chronology of enhancer repertoire establishment during hematopoiesis, we monitored epigenome dynamics during three developmental stages representing hematopoietic stem cells, B-cell progenitors and mature B-cells. We find that only a minority of enhancers primed in stem cells or progenitors become active at later stages. Furthermore, mo...

  9. Mouse Germ Cell Development in-vivo and in-vitro

    OpenAIRE

    Deshira Saiti; Orly Lacham-Kaplan

    2007-01-01

    In mammalian development, primordial germ cells (PGCs) represent the initial population of cells that are committed to the germ cell lineage. PGCs segregate early in development, triggered by signals from the extra-embryonic ectoderm. They are distinguished from surrounding cells by their unique gene expression patterns. Some of the more common genes used to identify them are Blimp1, Oct3/4, Fragilis, Stella, c-Kit, Mvh, Dazl and Gcna1. These genes are involved in regulating their migration a...

  10. Bridging sciatic nerve gap using tissue-engineered nerves constructed with neural tissue-committed stem cells derived from bone marrow

    Institute of Scientific and Technical Information of China (English)

    Zhiying Zhang; Congli Ren; Chuansen Zhang; Fang Liu; Liang Li

    2009-01-01

    BACKGROUND: Schwann cells are the most commonly used cells for tissue-engineered nerves. However, autologous Schwann cells are of limited use in a clinical context, and allogeneic Schwann cells induce immunological rejections. Cells that do not induce immunological rejections and that are relatively easy to acquire are urgently needed for transplantation.OBJECTIVE: To bridge sciatic nerve defects using tissue engineered nerves constructed with neural tissue-committed stem cells (NTCSCs) derived from bone marrow; to observe morphology and function of rat nerves following bridging; to determine the effect of autologous nerve transplantation, which serves as the gold standard for evaluating efficacy of tissue-engineered nerves.DESIGN, TIME AND SETTING: This randomized, controlled, animal experiment was performed in the Anatomical laboratory and Biomedical Institute of the Second Military Medical University of Chinese PLA between September 2004 and April 2006.MATERIALS: Five Sprague Dawley rats, aged 1 month and weighing 100-150 g, were used for cell culture. Sixty Sprague Dawiey rats aged 3 months and weighing 220-250 g, were used to establish neurological defect models. Nestin, neuron-specific enolase (NSE), glial fibrillary acidic protein (GFAP), and S-100 antibodies were provided by Santa Cruz Biotechnology, Inc., USA. Acellular nerve grafts were derived from dogs.METHODS: All rats, each with 1-cm gap created in the right sciatic nerve, were randomly assigned to three groups. Each group comprised 20 rats. Autograft nerve transplantation group: the severed 1-cm length nerve segment was reverted, but with the two ends exchanged; the proximal segment was sutured to the distal sciatic nerve stump and the distal segment to the proximal stump. Blank nerve scaffold transplantation group: a 1-cm length acellular nerve graft was used to bridge the sciatic nerve gap. NTCSC engineered nerve transplantation group: a 1-cm length acellular nerve graft, in which NTCSCs were

  11. Fast and scalable inference of multi-sample cancer lineages.

    KAUST Repository

    Popic, Victoria

    2015-05-06

    Somatic variants can be used as lineage markers for the phylogenetic reconstruction of cancer evolution. Since somatic phylogenetics is complicated by sample heterogeneity, novel specialized tree-building methods are required for cancer phylogeny reconstruction. We present LICHeE (Lineage Inference for Cancer Heterogeneity and Evolution), a novel method that automates the phylogenetic inference of cancer progression from multiple somatic samples. LICHeE uses variant allele frequencies of somatic single nucleotide variants obtained by deep sequencing to reconstruct multi-sample cell lineage trees and infer the subclonal composition of the samples. LICHeE is open source and available at http://viq854.github.io/lichee .

  12. Did HAL Commit Murder?

    OpenAIRE

    DENNETT, Daniel C.

    1996-01-01

    The first robot homicide was committed in 1981, according to my files. I have a yellowed clipping dated 12/9/81 from the Philadelphia Inquirer--not the National Enquirer--with the headline: Robot killed repairman, Japan reports The story was an anti-climax: at the Kawasaki Heavy Industries plant in Akashi, a malfunctioning robotic arm pushed a repairman against a gearwheel-milling machine, crushing him to death. The repairman had failed to follow proper instructions for shutting down ...

  13. Commitment to School and Students

    OpenAIRE

    Çağrı Tuğrul Mart

    2013-01-01

    Teaching necessitates passion, dedication, and commitment. Teachers, critical factors of any education system, play an important role in promoting education by enhancing the abilities of students through their work in schools. Commitment is highly related to teachers’ work performance. Teacher commitment deeply contributes to future of students and schools. Committed teachers make a difference to the success of the school and the learning of the students.

  14. Commitment to School and Students

    Directory of Open Access Journals (Sweden)

    Çağrı Tuğrul Mart

    2013-01-01

    Full Text Available Teaching necessitates passion, dedication, and commitment. Teachers, critical factors of any education system, play an important role in promoting education by enhancing the abilities of students through their work in schools. Commitment is highly related to teachers’ work performance. Teacher commitment deeply contributes to future of students and schools. Committed teachers make a difference to the success of the school and the learning of the students.

  15. GATA3-driven Th2 responses inhibit TGF-beta1-induced FOXP3 expression and the formation of regulatory T cells.

    NARCIS (Netherlands)

    P.Y. Mantel; H. Kuipers (Harmjan); O. Boyman (Onur); C. Rhyner (Claudio); N. Ouaked (Nadia); B. Rückert (Beate); C. Karagiannidis (Christian); B.N.M. Lambrecht (Bart); R.W. Hendriks (Rudi); R. Crameri; C.A. Akdis; K. Blaser (Kurt); C.B. Schmidt-Weber

    2007-01-01

    textabstractTranscription factors act in concert to induce lineage commitment towards Th1, Th2, or T regulatory (Treg) cells, and their counter-regulatory mechanisms were shown to be critical for polarization between Th1 and Th2 phenotypes. FOXP3 is an essential transcription factor for natural, thy

  16. The mapping of neurons and lineage classification of the larvae and adult Drosophila brain in several Gal4 transmitter lines

    OpenAIRE

    Ahad, Sally

    2015-01-01

    In Drosophila, neurons within the central nervous system are grouped into units called lineages. Each lineage contains cells derived from a single neuroblast. A neuroblast is a stem cell divides and forms lineages of neurons. In flies, the lineage can be subdivided into different parts; the neurons that are born first are closest to the neuropile (Spindler and Hartenstein, 2010). There is a birth ordering of neurons. In the embryo, the neuroblasts divide 5 to 6 times and are called primary n...

  17. Mammary development and breast cancer: the role of stem cells

    OpenAIRE

    Ercan, C.; J. van Diest, P.; Vooijs, M.

    2011-01-01

    The mammary gland is a highly regenerative organ that can undergo multiple cycles of proliferation, lactation and involution, a process controlled by stem cells. The last decade much progress has been made in the identification of signaling pathways that function in these stem cells to control self-renewal, lineage commitment and epithelial differentiation in the normal mammary gland. The same signaling pathways that control physiological mammary development and homeostasis are also often fou...

  18. The RNA–Methyltransferase Misu (NSun2) Poises Epidermal Stem Cells to Differentiate

    OpenAIRE

    Sandra Blanco; Agata Kurowski; Jennifer Nichols; Watt, Fiona M.; Salvador Aznar Benitah; Michaela Frye

    2011-01-01

    Homeostasis of most adult tissues is maintained by balancing stem cell self-renewal and differentiation, but whether post-transcriptional mechanisms can regulate this process is unknown. Here, we identify that an RNA methyltransferase (Misu/Nsun2) is required to balance stem cell self-renewal and differentiation in skin. In the epidermis, this methyltransferase is found in a defined sub-population of hair follicle stem cells poised to undergo lineage commitment, and its depletion results in e...

  19. Heterogeneous Differentiation of Human Mesenchymal Stem Cells in Response to Extended Culture in Extracellular Matrices

    OpenAIRE

    Jose A Santiago; Pogemiller, Ryan; Ogle, Brenda M.

    2009-01-01

    Extracellular matrix proteins (ECMs) guide differentiation of adult stem cells, but the temporal distribution of differentiation (i.e., heterogeneity) in a given population has not been investigated. We tested the effect of individual ECM proteins on lineage commitment of human bone marrow–derived mesenchymal stem cells (MSCs) over time. We exposed stem cell populations to ECM proteins representing the primary tissue structures of the body (i.e., collagens type I, III, IV; laminin; and fibron...

  20. Selective Lineage Specification by Mab-19 during Caenorhabditis Elegans Male Peripheral Sense Organ Development

    OpenAIRE

    Sutherlin, M. E.; Emmons, S W

    1994-01-01

    The action of the gene mab-19 is required for specification of a subset of Caenorhabditis elegans male peripheral sense organ (ray) lineages. Two mab-19 alleles, isolated in screens for ray developmental mutations, resulted in males that lacked the three most posterior rays. Cell lineage alterations of male-specific divisions of the most posterior lateral hypodermal (seam) blast cell, T, resulted in the ray loss phenotype in mab-19 mutant animals. Postembryonic seam lineage defects were limit...

  1. A novel PAD4/SOX4/PU.1 signaling pathway is involved in the committed differentiation of acute promyelocytic leukemia cells into granulocytic cells.

    Science.gov (United States)

    Song, Guanhua; Shi, Lulu; Guo, Yuqi; Yu, Linchang; Wang, Lin; Zhang, Xiaoyu; Li, Lianlian; Han, Yang; Ren, Xia; Guo, Qiang; Bi, Kehong; Jiang, Guosheng

    2016-01-19

    All-trans retinoic acid (ATRA) treatment yields cure rates > 80% through proteasomal degradation of the PML-RARα fusion protein that typically promotes acute promyelocytic leukemia (APL). However, recent evidence indicates that ATRA can also promote differentiation of leukemia cells that are PML-RARα negative, such as HL-60 cells. Here, gene expression profiling of HL-60 cells was used to investigate the alternative mechanism of impaired differentiation in APL. The expression of peptidylarginine deiminase 4 (PADI4), encoding PAD4, a protein that post-translationally converts arginine into citrulline, was restored during ATRA-induced differentiation. We further identified that hypermethylation in the PADI4 promoter was associated with its transcriptional repression in HL-60 and NB4 (PML-RARα positive) cells. Functionally, PAD4 translocated into the nucleus upon ATRA exposure and promoted ATRA-mediated differentiation. Mechanistic studies using RNAi knockdown or electroporation-mediated delivery of PADI4, along with chromatin immunoprecipitation, helped identify PU.1 as an indirect target and SOX4 as a direct target of PAD4 regulation. Indeed, PAD4 regulates SOX4-mediated PU.1 expression, and thereby the differentiation process, in a SOX4-dependent manner. Taken together, our results highlight an association between PAD4 and DNA hypermethylation in APL and demonstrate that targeting PAD4 or regulating its downstream effectors may be a promising strategy to control differentiation in the clinic. PMID:26673819

  2. [Bone and Stem Cells. The mechanism of osteogenic differentiation from mesenchymal stem cell].

    Science.gov (United States)

    Ohata, Yasuhisa; Ozono, Keiichi

    2014-04-01

    Osteoblasts and osteocytes originate from pluripotent mesenchymal stem cells. Mesenchymal stem cells commit to osteogenic lineage and differentiate into mature osteoblasts and osteocytes through osteoprogenitor cells and preosteoblasts in response to multiple stimuli. The osteoblast commitment, differentiation, and functions are governed by several transcription factors. Among these transcription factors, runt-related transcription factor 2 (Runx2) is a crucial factor in osteoblast differentiation and controls bone formation. Differentiation toward these osteogenic lineage is controlled by a multitude of cytokines including WNTs, bone morphogenetic protein (BMP) , transforming growth factor-β (TGF-β) , hedgehog, parathyroid hormone (PTH) /parathyroid hormone related protein (PTHrP) , insulin-like growth factor-1 (IGF-1) , fibroblast growth factor (FGF) , and Notch. Although regulation of Runx2 activity is a point of convergence of many of the signal transduction routes, there is also a high degree of cross-talk between these pathways. Thus, the combined action of the signal transduction pathways induced by some cytokines determines the commitment and differentiation of mesenchymal stem cells toward the osteogenic lineage. PMID:24681495

  3. Intracoronary artery transplantation of cardiomyoblast-like cells from human adipose tissue-derived multi-lineage progenitor cells improve left ventricular dysfunction and survival in a swine model of chronic myocardial infarction

    International Nuclear Information System (INIS)

    Highlights: ► We administered human CLCs in a swine model of MI via intracoronary artery. ► Histological studies demonstrated engraftment of hCLCs into the scarred myocardium. ► Echocardiography showed rescue of cardiac function in the hCLCs transplanted swine. ► Transplantation of hCLCs is an effective therapeutics for cardiac regeneration. -- Abstract: Transplantation of human cardiomyoblast-like cells (hCLCs) from human adipose tissue-derived multi-lineage progenitor cells improved left ventricular function and survival of rats with myocardial infarction. Here we examined the effect of intracoronary artery transplantation of human CLCs in a swine model of chronic heart failure. Twenty-four pigs underwent balloon-occlusion of the first diagonal branch followed by reperfusion, with a second balloon-occlusion of the left ascending coronary artery 1 week later followed by reperfusion. Four weeks after the second occlusion/reperfusion, 17 of the 18 surviving animals with severe chronic MI (ejection fraction <35% by echocardiography) were immunosuppressed then randomly assigned to receive either intracoronary artery transplantation of hCLCs hADMPCs or placebo lactic Ringer’s solution with heparin. Intracoronary artery transplantation was followed by the distribution of DiI-stained hCLCs into the scarred myocardial milieu. Echocardiography at post-transplant days 4 and 8 weeks showed rescue and maintenance of cardiac function in the hCLCs transplanted group, but not in the control animals, indicating myocardial functional recovery by hCLCs intracoronary transplantation. At 8 week post-transplantation, 7 of 8 hCLCs transplanted animals were still alive compared with only 1 of the 5 control (p = 0.0147). Histological studies at week 12 post-transplantation demonstrated engraftment of the pre DiI-stained hCLCs into the scarred myocardium and their expression of human specific alpha-cardiac actin. Human alpha cardiac actin-positive cells also expressed cardiac

  4. Intracoronary artery transplantation of cardiomyoblast-like cells from human adipose tissue-derived multi-lineage progenitor cells improve left ventricular dysfunction and survival in a swine model of chronic myocardial infarction

    Energy Technology Data Exchange (ETDEWEB)

    Okura, Hanayuki [The Center for Medical Engineering and Informatics, Osaka University, 2-2 Yamada-oka, Suita, Osaka 565-0879 (Japan); Department of Somatic Stem Cell Therapy and Health Policy, Institute of Biomedical Research and Innovation, Foundation for Biomedical Research and Innovation, 2-2 Minatojima-minamimachi, Chuo-ku, Kobe, Hyogo 650-0047 (Japan); Saga, Ayami; Soeda, Mayumi [Department of Somatic Stem Cell Therapy and Health Policy, Institute of Biomedical Research and Innovation, Foundation for Biomedical Research and Innovation, 2-2 Minatojima-minamimachi, Chuo-ku, Kobe, Hyogo 650-0047 (Japan); Miyagawa, Shigeru; Sawa, Yoshiki [Department of Surgery, Osaka University Graduate School of Medicine, 2-2 Yamada-oka, Suita, Osaka 565-0879 (Japan); Daimon, Takashi [Division of Biostatistics, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya, Hyogo 663-8501 (Japan); Ichinose, Akihiro [Department of Plastic Surgery, Kobe University Hospital, 7-5-2 Kusunoki-cho, Chuo-ku, Kobe, Hyogo (Japan); Matsuyama, Akifumi, E-mail: akifumi-matsuyama@umin.ac.jp [The Center for Medical Engineering and Informatics, Osaka University, 2-2 Yamada-oka, Suita, Osaka 565-0879 (Japan); Department of Plastic Surgery, Kobe University Hospital, 7-5-2 Kusunoki-cho, Chuo-ku, Kobe, Hyogo (Japan); RIKEN Program for Drug Discovery and Medical Technology Platforms, 1-7-22 Suehiro-cho, Tsurumi-ku, Yokohama, Kanagawa 230-0045 (Japan)

    2012-09-07

    Highlights: Black-Right-Pointing-Pointer We administered human CLCs in a swine model of MI via intracoronary artery. Black-Right-Pointing-Pointer Histological studies demonstrated engraftment of hCLCs into the scarred myocardium. Black-Right-Pointing-Pointer Echocardiography showed rescue of cardiac function in the hCLCs transplanted swine. Black-Right-Pointing-Pointer Transplantation of hCLCs is an effective therapeutics for cardiac regeneration. -- Abstract: Transplantation of human cardiomyoblast-like cells (hCLCs) from human adipose tissue-derived multi-lineage progenitor cells improved left ventricular function and survival of rats with myocardial infarction. Here we examined the effect of intracoronary artery transplantation of human CLCs in a swine model of chronic heart failure. Twenty-four pigs underwent balloon-occlusion of the first diagonal branch followed by reperfusion, with a second balloon-occlusion of the left ascending coronary artery 1 week later followed by reperfusion. Four weeks after the second occlusion/reperfusion, 17 of the 18 surviving animals with severe chronic MI (ejection fraction <35% by echocardiography) were immunosuppressed then randomly assigned to receive either intracoronary artery transplantation of hCLCs hADMPCs or placebo lactic Ringer's solution with heparin. Intracoronary artery transplantation was followed by the distribution of DiI-stained hCLCs into the scarred myocardial milieu. Echocardiography at post-transplant days 4 and 8 weeks showed rescue and maintenance of cardiac function in the hCLCs transplanted group, but not in the control animals, indicating myocardial functional recovery by hCLCs intracoronary transplantation. At 8 week post-transplantation, 7 of 8 hCLCs transplanted animals were still alive compared with only 1 of the 5 control (p = 0.0147). Histological studies at week 12 post-transplantation demonstrated engraftment of the pre DiI-stained hCLCs into the scarred myocardium and their expression of

  5. Spatio-temporal model of endogenous ROS and raft-dependent WNT/{beta}-catenin signaling driving cell fate commitment in human neural progenitor cells

    OpenAIRE

    Haack, F.; Lemcke, H.; Ewald, R.(Physikalisches Institut, Universität Bonn, Germany); Rharass, T.; Uhrmacher, A.M.

    2015-01-01

    Canonical WNT/{beta}-catenin signaling is a central pathway in embryonic development, but it is also connected to a number of cancers and developmental disorders. Here we apply a combined in-vitro and in-silico approach to investigate the spatio-temporal regulation of WNT/{beta}-catenin signaling during the early neural differentiation process of human neural progenitors cells (hNPCs), which form a new prospect for replacement therapies in the context of neurodegenerative diseases. Experiment...

  6. Thymic commitment of regulatory T cells is a pathway of TCR-dependent selection that isolates repertoires undergoing positive or negative selection.

    Science.gov (United States)

    Coutinho, A; Caramalho, I; Seixas, E; Demengeot, J

    2005-01-01

    The seminal work of Le Douarin and colleagues (Ohki et al. 1987; Ohki et al. 1988; Salaun et al. 1990; Coutinho et al. 1993) first demonstrated that peripheral tissue-specific tolerance is centrally established in the thymus, by epithelial stromal cells (TEC). Subsequent experiments have shown that TEC-tolerance is dominant and mediated by CD4 regulatory T cells (Treg) that are generated intrathymically by recognition of antigens expressed on TECs (Modigliani et al. 1995; Modigliani et al. 1996a). From these and other observations, in 1996 Modigliani and colleagues derived a general model for the establishment and maintenance of natural tolerance (MM96) (Modigliani et al. 1996b), with two central propositions: (1) T cell receptor (TCR)-dependent sorting of emergent repertoires generates TEC-specific Treg displaying the highest TCR self-affinities below deletion thresholds, thus isolating repertoires undergoing positive and negative selection; (2) Treg are intrathymically committed (and activated) for a unique differentiative pathway with regulatory effector functions. The model explained the embryonic/perinatal time window of natural tolerance acquisition, by developmental programs determining (1) TCR multireactivity, (2) the cellular composition in the thymic stroma (relative abundance of epithelial vs hemopoietic cells), and (3) the dynamics of peripheral lymphocyte pools, built by accumulation of recent thymic emigrants (RTE) that remain recruitable to regulatory functions. We discuss here the MM96 in the light of recent results demonstrating the promiscuous expression of tissue-specific antigens by medullary TECs (Derbinski et al. 2001; Anderson et al. 2002; Gotter et al. 2004) and indicating that Treg represent a unique differentiative pathway (Fontenot et al. 2003; Hori et al. 2003; Khattri et al. 2003), which is adopted by CD4 T cells with high avidity for TEC-antigens (Bensinger et al. 2001; Jordan et al. 2001; Apostolou et al. 2002). In the likelihood that

  7. There is more to a lipid than just being a fat: sphingolipid-guided differentiation of oligodendroglial lineage from embryonic stem cells

    OpenAIRE

    Bieberich, Erhard

    2010-01-01

    Dr. Robert K. Yu's research showed for the first time that the composition of glycosphingolipids is tightly regulated during embryo development. Studies in our group showed that the glycosphingolipid precursor ceramide is also critical for stem cell differentiation and apoptosis. Our new studies suggest that ceramide and its derivative, sphingosine-1-phosphate (S1P), act synergistically on embryonic stem (ES) cell differentiation. When using neural precursor cells (NPCs) derived from ES cells...

  8. Thymic and Postthymic Regulation of Naïve CD4+ T-Cell Lineage Fates in Humans and Mice Models

    OpenAIRE

    Belizário, José E.; Wesley Brandão; Cristiano Rossato; Jean Pierre Peron

    2016-01-01

    Our understanding of how thymocytes differentiate into many subtypes has been increased progressively in its complexity. At early life, the thymus provides a suitable microenvironment with specific combination of stromal cells, growth factors, cytokines, and chemokines to induce the bone marrow lymphoid progenitor T-cell precursors into single-positive CD4+ and CD8+ T effectors and CD4+CD25+ T-regulatory cells (Tregs). At postthymic compartments, the CD4+ T-cells acquire distinct phenotypes w...

  9. Patterns of growth, axonal extension and axonal arborization of neuronal lineages in the developing Drosophila brain.

    Science.gov (United States)

    Larsen, Camilla; Shy, Diana; Spindler, Shana R; Fung, Siaumin; Pereanu, Wayne; Younossi-Hartenstein, Amelia; Hartenstein, Volker

    2009-11-15

    The Drosophila central brain is composed of approximately 100 paired lineages, with most lineages comprising 100-150 neurons. Most lineages have a number of important characteristics in common. Typically, neurons of a lineage stay together as a coherent cluster and project their axons into a coherent bundle visible from late embryo to adult. Neurons born during the embryonic period form the primary axon tracts (PATs) that follow stereotyped pathways in the neuropile. Apoptotic cell death removes an average of 30-40% of primary neurons around the time of hatching. Secondary neurons generated during the larval period form secondary axon tracts (SATs) that typically fasciculate with their corresponding primary axon tract. SATs develop into the long fascicles that interconnect the different compartments of the adult brain. Structurally, we distinguish between three types of lineages: PD lineages, characterized by distinct, spatially separate proximal and distal arborizations; C lineages with arborizations distributed continuously along the entire length of their tract; D lineages that lack proximal arborizations. Arborizations of many lineages, in particular those of the PD type, are restricted to distinct neuropile compartments. We propose that compartments are "scaffolded" by individual lineages, or small groups thereof. Thereby, the relatively small number of primary neurons of each primary lineage set up the compartment map in the late embryo. Compartments grow during the larval period simply by an increase in arbor volume of primary neurons. Arbors of secondary neurons form within or adjacent to the larval compartments, resulting in smaller compartment subdivisions and additional, adult specific compartments. PMID:19538956

  10. Commitment Problems in Conflict Resolution

    OpenAIRE

    Kimbrough, Erik; Rubin, Jared; Sheremeta, Roman; Shields, Timothy

    2015-01-01

    Commitment problems are inherent to non-binding conflict resolution mechanisms, since an unsatisfied party can ignore the resolution and initiate conflict. We provide experimental evidence suggesting that even in the absence of binding contractual agreements individuals often avoid conflict by committing to the outcome of a conflict resolution mechanism. Commitment problems are mitigated to a greater extent for groups that opt-in to the conflict resolution mechanism, but only when opting-in i...

  11. Feedback, Lineages and Self-Organizing Morphogenesis.

    Directory of Open Access Journals (Sweden)

    Sameeran Kunche

    2016-03-01

    Full Text Available Feedback regulation of cell lineage progression plays an important role in tissue size homeostasis, but whether such feedback also plays an important role in tissue morphogenesis has yet to be explored. Here we use mathematical modeling to show that a particular feedback architecture in which both positive and negative diffusible signals act on stem and/or progenitor cells leads to the appearance of bistable or bi-modal growth behaviors, ultrasensitivity to external growth cues, local growth-driven budding, self-sustaining elongation, and the triggering of self-organization in the form of lamellar fingers. Such behaviors arise not through regulation of cell cycle speeds, but through the control of stem or progenitor self-renewal. Even though the spatial patterns that arise in this setting are the result of interactions between diffusible factors with antagonistic effects, morphogenesis is not the consequence of Turing-type instabilities.

  12. FoxA1 directs the lineage and immunosuppressive properties of a novel regulatory T cell population in EAE and MS

    DEFF Research Database (Denmark)

    Liu, Yawei; Carlsson, Robert; Comabella, Manuel;

    2014-01-01

    The defective generation or function of regulatory T (Treg) cells in autoimmune disease contributes to chronic inflammation and tissue injury. We report the identification of FoxA1 as a transcription factor in T cells that, after ectopic expression, confers suppressive properties in a newly ident...

  13. The Drosophila neural lineages: a model system to study brain development and circuitry

    OpenAIRE

    Spindler, Shana R; Hartenstein, Volker

    2010-01-01

    In Drosophila, neurons of the central nervous system are grouped into units called lineages. Each lineage contains cells derived from a single neuroblast. Due to its clonal nature, the Drosophila brain is a valuable model system to study neuron development and circuit formation. To better understand the mechanisms underlying brain development, genetic manipulation tools can be utilized within lineages to visualize, knock down, or over-express proteins. Here, we will introduce the formation an...

  14. Generation and Expansion of highly-pure Motor Neuron Progenitors from Human Pluripotent Stem Cells

    OpenAIRE

    Du, Zhong-Wei; Chen, Hong; Liu, Huisheng; Lu, Jianfeng; Qian, Kun; Huang, Cindy Tzu-Ling.; Zhong, Xiaofen; Fan, Frank; Zhang, Su-Chun

    2015-01-01

    SUMMARY Human pluripotent stem cells (hPSCs) have opened new opportunities for understanding human development, modeling disease processes and developing new therapeutics. However, these applications are hindered by low-efficiency and heterogeneity of target cell types differentiated from hPSCs, such as motor neurons (MNs), as well as our inability to maintain the potency of lineage committed progenitors. Here, by using a combination of small molecules that regulate multiple signaling pathway...

  15. Distinct myeloid progenitor differentiation pathways identified through single cell RNA sequencing

    OpenAIRE

    Drissen, R; Buza-Vidas, N.; Woll, P.; Thongjuea, S.; Gambardella, A; Giustacchini, A.; E. Mancini; Zriwil, A.; Lutteropp], M; Grover, A.; Mead, A.; Sitnicka, E; Jacobsen, SE; Nerlov, C.

    2016-01-01

    According to current models for hematopoiesis, lymphoid-primed multi-potent progenitors (LMPPs; Lin– Sca-1+ c-Kit+ CD34+ Flt3hi) and common myeloid progenitors (CMPs; Lin– Sca- 1+ c-Kit+ CD34+ CD41hi) establish an early branch point for separate lineage commitment pathways from hematopoietic stem cells, with the notable exception that both pathways are proposed to generate all myeloid innate immune cell types through the same myeloidrestricted pre-granul...

  16. The Measurement of Organizational Commitment.

    Science.gov (United States)

    Mowday, Richard T.; And Others

    1979-01-01

    This paper summarizes a stream of research aimed at developing and validating a measure of employee commitment to work organizations. The instrument, developed by Porter and his colleagues, is called the Organizational Commitment Questionnaire. Satisfactory test-retest reliabilities and internal consistency reliabilities were found. (Author)

  17. Organizational Climate and Teacher Commitment

    Science.gov (United States)

    Douglas, Stephen Michael

    2010-01-01

    This study examined the relationship of school climate and teacher commitment in elementary schools in Alabama. A total of 67 elementary schools were surveyed and 1353 teachers voluntarily participated in the study. The instruments used in this study were the Organizational Climate Index (OCI) and the Organizational Commitment Questionnaire (OCQ).…

  18. Statistical secrecy and multibit commitments

    DEFF Research Database (Denmark)

    Damgård, Ivan Bjerre; Pedersen, Torben P.; Pfitzmann, Birgit

    1998-01-01

    nothing about it. One definition is based on the L1-norm distance between probability distributions, the other on information theory. We prove that the two definitions are essentially equivalent. We also show that statistical counterparts of definitions of computational secrecy are essentially equivalent......We present and compare definitions of "statistically hiding" protocols, and we propose a novel statistically hiding commitment scheme. Informally, a protocol statistically hides a secret if a computationally unlimited adversary who conducts the protocol with the owner of the secret learns almost...... to our main definitions. Commitment schemes are an important cryptologic primitive. Their purpose is to commit one party to a certain value, while hiding this value from the other party until some later time. We present a statistically hiding commitment scheme allowing commitment to many bits. The...

  19. Quantum Commitments from Complexity Assumptions

    CERN Document Server

    Chailloux, André; Rosgen, Bill

    2010-01-01

    Bit commitment schemes are at the basis of modern cryptography. Since information-theoretic security is impossible both in the classical and the quantum regime, we need to look at computationally secure commitment schemes. In this paper, we study worst-case complexity assumptions that imply quantum bit-commitment schemes. First, we show that QSZK not included in QMA implies a computationally hiding and statistically binding auxiliary-input quantum commitment scheme. Additionally, we give auxiliary-input commitment schemes using quantum advice that depend on the much weaker assumption that QIP is not included in QMA (which is weaker than PSPACE not included in PP). Finally, we find a quantum oracle relative to which honest-verifier QSZK is not contained in QCMA, the class of languages that can be verified using a classical proof in quantum polynomial time.

  20. Contexts as Shared Commitments.

    Science.gov (United States)

    García-Carpintero, Manuel

    2015-01-01

    Contemporary semantics assumes two influential notions of context: one coming from Kaplan (1989), on which contexts are sets of predetermined parameters, and another originating in Stalnaker (1978), on which contexts are sets of propositions that are "common ground." The latter is deservedly more popular, given its flexibility in accounting for context-dependent aspects of language beyond manifest indexicals, such as epistemic modals, predicates of taste, and so on and so forth; in fact, properly dealing with demonstratives (perhaps ultimately all indexicals) requires that further flexibility. Even if we acknowledge Lewis (1980)'s point that, in a sense, Kaplanian contexts already include common ground contexts, it is better to be clear and explicit about what contexts constitutively are. Now, Stalnaker (1978, 2002, 2014) defines context-as-common-ground as a set of propositions, but recent work shows that this is not an accurate conception. The paper explains why, and provides an alternative. The main reason is that several phenomena (presuppositional treatments of pejoratives and predicates of taste, forces other than assertion) require that the common ground includes non-doxastic attitudes such as appraisals, emotions, etc. Hence the common ground should not be taken to include merely contents (propositions), but those together with attitudes concerning them: shared commitments, as I will defend. PMID:26733087

  1. Who commits matricide?

    Science.gov (United States)

    Singhal, S; Dutta, A

    1992-07-01

    The authors studied sixteen men who committed matricide. Fifteen out of sixteen cases had a diagnosis of schizophrenia and the remaining patient had a diagnosis of schizophrenia with personality disorder. All were single at the time of the matricide. Data indicate an intense conflict-laden and ambivalent relationship between the majority of patients with their mothers. Thirteen out of sixteen cases described their mothers as quite domineering and demanding but the EMBU inventory revealed that the Matricidal group differed from the Control group in how tolerant they saw their parents. The sample as a whole saw mothers were more over-involved, overprotective, tolerant, affectionate, stimulating, performance-orientated and shaming. The matricidal group differed from the control group in the way they viewed the difference between mother and father on various scales, like over-involved, tolerant, affectionate and performance-orientated. The matricidal groups' mothers were found to be more over-involved, tolerant, affectionate, and fathers more abusive. Mothers in the control group were more performance-orientated. PMID:1513219

  2. Non–lineage/stage-restricted effects of a gain-of-function mutation in tyrosine phosphatase Ptpn11 (Shp2) on malignant transformation of hematopoietic cells

    OpenAIRE

    Xu, Dan; Liu, Xia; Yu, Wen-Mei; Meyerson, Howard J.; Guo, Caiying; Gerson, Stanton L.; Qu, Cheng-Kui

    2011-01-01

    Activating mutations in protein tyrosine phosphatase 11 (Ptpn11) have been identified in childhood acute leukemias, in addition to juvenile myelomonocytic leukemia (JMML), which is a myeloproliferative disorder (MPD). It is not clear whether activating mutations of this phosphatase play a causal role in the pathogenesis of acute leukemias. If so, the cell origin of leukemia-initiating stem cells (LSCs) remains to be determined. Ptpn11E76K mutation is the most common and most active Ptpn11 mut...

  3. Lineage tracking of mesenchymal and endothelial progenitors in BMP-induced bone formation.

    Science.gov (United States)

    Kolind, Mille; Bobyn, Justin D; Matthews, Brya G; Mikulec, Kathy; Aiken, Alastair; Little, David G; Kalajzic, Ivo; Schindeler, Aaron

    2015-12-01

    To better understand the relative contributions of mesenchymal and endothelial progenitor cells to rhBMP-2 induced bone formation, we examined the distribution of lineage-labeled cells in Tie2-Cre:Ai9 and αSMA-creERT2:Col2.3-GFP:Ai9 reporter mice. Established orthopedic models of ectopic bone formation in the hind limb and spine fusion were employed. Tie2-lineage cells were found extensively in the ectopic bone and spine fusion masses, but co-staining was only seen with tartrate-resistant acid phosphatase (TRAP) activity (osteoclasts) and CD31 immunohistochemistry (vascular endothelial cells), and not alkaline phosphatase (AP) activity (osteoblasts). To further confirm the lack of a functional contribution of Tie2-lineage cells to BMP-induced bone, we developed conditional knockout mice where Tie2-lineage cells are rendered null for key bone transcription factor osterix (Tie2-cre:Osx(fx/fx) mice). Conditional knockout mice showed no difference in BMP-induced bone formation compared to littermate controls. Pulse labeling of mesenchymal cells with Tamoxifen in mice undergoing spine fusion revealed that αSMA-lineage cells contributed to the osteoblastic lineage (Col2.3-GFP), but not to endothelial cells or osteoclast populations. These data indicate that the αSMA+ and Tie2+ progenitor lineages make distinct cellular contributions to bone formation, angiogenesis, and resorption/remodeling. PMID:26141839

  4. Conditional Deletion of the Relaxin Receptor Gene in Cells of Smooth Muscle Lineage Affects Lower Reproductive Tract in Pregnant Mice1

    Science.gov (United States)

    Kaftanovskaya, Elena M.; Huang, Zaohua; Lopez, Carolina; Conrad, Kirk; Agoulnik, Alexander I.

    2015-01-01

    ABSTRACT Relaxin hormone secreted into the circulation during pregnancy was discovered through its effects on pubic symphysis relaxation and parturition. Genetic inactivation of the relaxin gene or its cognate relaxin family peptide receptor 1 (RXFP1) in mice caused failure of parturition and mammary nipple enlargement, as well as increased collagen fiber density in the cervix and vagina. However, the relaxin effect on discrete cells and tissues has yet to be determined. Using transgenic mice with a knockin LacZ reporter in the Rxfp1 allele, we showed strong expression of this gene in vaginal and cervical stromal cells, as well as pubic ligament cells. We produced a floxed Rxfp1 allele that was used in combination with the Tagln-cre transgene to generate mice with a smooth muscle-specific gene knockout. In pregnant females, the ROSA26 reporter activated by Tagln-cre was detected in smooth muscle cells of the cervix, vagina, uterine artery, and in cells of the pubic symphysis. In late pregnant females with conditional gene ablation, the length of pubic symphysis was significantly reduced compared with wild-type or heterozygous Rxfp1+/− females. Denser collagen content was revealed by Masson trichrome staining in reproductive tract organs, uterine artery, and pubic symphysis. The cervical and vaginal epithelium was less developed than in heterozygous or wild-type females, although nipple size was normal and the dams were able to nurse their pups. In summary, our data indicate that relaxin/RXFP1 signaling in smooth muscle cells is important for normal collagen turnover and relaxation of the pubic symphysis during pregnancy. PMID:25715795

  5. Development of Hematopoietic and Endothelial Cells from Human Embryonic Stem Cells: Lessons from the Studies using Mouse as a Model

    OpenAIRE

    Anna Jezierski; Albert Swedani; Lisheng Wang

    2007-01-01

    The current progress using the human embryonic stem cell (hESC) model system has provided much insight into the early origins of the hematopoietic and endothelial lineages, particularly the elusive hemangioblast. Recently, the cellular hierarchy and molecular regulation controlling hematopoietic commitment have been further elucidated. These findings not only provide new insights into early human development, but also advance the knowledge required to develop techniques capable of generating ...

  6. When Family History Matters: The Importance of Lineage Analyses and Fate Maps for Explaining Animal Development.

    Science.gov (United States)

    Klein, Steven L; Moody, Sally A

    2016-01-01

    Initial interest in understanding how the fertilized egg becomes a multicellular animal suggested two possible answers: either the embryo came from preformed components or it arose through epigenetic processes. Extensive research during the past few decades has identified aspects of development that depend on preformed elements, such as cytoplasmic components and a cell's lineage; it also has identified aspects that depend on epigenetic processes, such as cell interactions and morphogen gradients. These advances have depended on understanding embryonic cell lineage and cell fate. This essay explains how lineage analysis and fate mapping have contributed to our current understanding of embryonic development. PMID:26969974

  7. SIRPA, VCAM1 and CD34 identify discrete lineages during early human cardiovascular development

    OpenAIRE

    Rhys J.P. Skelton; Magdaline Costa; Anderson, David J.; Freya Bruveris; Ben W. Finnin; Katerina Koutsis; Deevina Arasaratnam; White, Anthony J; Arash Rafii; Ng, Elizabeth S.; Elefanty, Andrew G.; Edouard G. Stanley; Pouton, Colin W.; Haynes, John M.; Reza Ardehali

    2014-01-01

    The study of human cardiogenesis would benefit from a detailed cell lineage fate map akin to that established for the haematopoietic lineages. Here we sought to define cell lineage relationships based on the expression of NKX2-5 and the cell surface markers VCAM1, SIRPA and CD34 during human cardiovascular development. Expression of NKX2-5GFP was used to identify cardiac progenitors and cardiomyocytes generated during the differentiation of NKX2-5GFP/w human embryonic stem cells (hESCs). Card...

  8. The Drosophila neural lineages: a model system to study brain development and circuitry.

    Science.gov (United States)

    Spindler, Shana R; Hartenstein, Volker

    2010-06-01

    In Drosophila, neurons of the central nervous system are grouped into units called lineages. Each lineage contains cells derived from a single neuroblast. Due to its clonal nature, the Drosophila brain is a valuable model system to study neuron development and circuit formation. To better understand the mechanisms underlying brain development, genetic manipulation tools can be utilized within lineages to visualize, knock down, or over-express proteins. Here, we will introduce the formation and development of lineages, discuss how one can utilize this model system, offer a comprehensive list of known lineages and their respective markers, and then briefly review studies that have utilized Drosophila neural lineages with a look at how this model system can benefit future endeavors. PMID:20306203

  9. Lineage switch with t(6;11)(q27;q23) from T-cell lymphoblastic lymphoma to acute monoblastic leukemia at relapse.

    Science.gov (United States)

    Higuchi, Yusuke; Tokunaga, Kenji; Watanabe, Yuko; Kawakita, Toshiro; Harada, Naoko; Yamaguchi, Shunichiro; Nosaka, Kisato; Mitsuya, Hiroaki; Asou, Norio

    2016-06-01

    We present a patient with T-cell lymphoblastic lymphoma (T-LBL) harboring t(6;11)(q27;q23) that converted to acute monoblastic leukemia at relapse. A 27-year-old man developed T-LBL with a mediastinal mass. He exhibited several recurrences in the central nervous system and marrow. A fifth relapse occurred in the marrow, with 42.8% blasts with CD4, CD5, CD7, CD10, CD33, CD34, HLA-DR and cytoplasmic (cy) CD3. While achieving complete remission with nelarabine, sixth relapse occurred in the marrow with 6.8% blasts, which had characteristics of monoblastic features, 2 months later. Marrow blasts were positive for myeloperoxidase, CD4, CD33, CD56, CD64, and HLA-DR, but were negative for cyCD3, CD5, CD7, CD10, and CD34. Marrow cells at both the 5th lymphoid and 6th myeloid relapses had t(6;11)(q27;q23) and the same MLL-MLLT4 fusion transcript. In addition, the MLL-MLLT4 fusion sequences documented in the initial mediastinal cells were the same as seen in peripheral blood cells at the 6th relapse. The patient continues 7th remission after one course of gemtuzumab ozogamicin therapy followed by cord blood transplantation for more than 3 years. Sequential phenotypic and cytogenetic studies may yield valuable insights into the mechanism of leukemic recurrence and possible implications for treatment selection. PMID:27268298

  10. Circulating Hematopoietic Stem and Progenitor Cells in Aging Atomic Bomb Survivors.

    Science.gov (United States)

    Kyoizumi, Seishi; Kubo, Yoshiko; Misumi, Munechika; Kajimura, Junko; Yoshida, Kengo; Hayashi, Tomonori; Imai, Kazue; Ohishi, Waka; Nakachi, Kei; Young, Lauren F; Shieh, Jae-Hung; Moore, Malcolm A; van den Brink, Marcel R M; Kusunoki, Yoichiro

    2016-01-01

    It is not yet known whether hematopoietic stem and progenitor cells (HSPCs) are compromised in the aging population of atomic bomb (A-bomb) survivors after their exposure nearly 70 years ago. To address this, we evaluated age- and radiation-related changes in different subtypes of circulating HSPCs among the CD34-positive/lineage marker-negative (CD34(+)Lin(-)) cell population in 231 Hiroshima A-bomb survivors. We enumerated functional HSPC subtypes, including: cobblestone area-forming cells; long-term culture-initiating cells; erythroid burst-forming units; granulocyte and macrophage colony-forming units; and T-cell and natural killer cell progenitors using cell culture. We obtained the count of each HSPC subtype per unit volume of blood and the proportion of each HSPC subtype in CD34(+)Lin(-) cells to represent the lineage commitment trend. Multivariate analyses, using sex, age and radiation dose as variables, showed significantly decreased counts with age in the total CD34(+)Lin(-) cell population and all HSPC subtypes. As for the proportion, only T-cell progenitors decreased significantly with age, suggesting that the commitment to the T-cell lineage in HSPCs continuously declines with age throughout the lifetime. However, neither the CD34(+)Lin(-) cell population, nor HSPC subtypes showed significant radiation-induced dose-dependent changes in counts or proportions. Moreover, the correlations of the proportions among HSPC subtypes in the survivors properly revealed the hierarchy of lineage commitments. Taken together, our findings suggest that many years after exposure to radiation and with advancing age, the number and function of HSPCs in living survivors as a whole may have recovered to normal levels. PMID:26720799

  11. Commitment profiles: combinations of organizational commitment forms and job outcomes

    OpenAIRE

    Wasti, S. Arzu

    2005-01-01

    Although the three-component model of organizational commitment by Meyer and Allen (1991) posits that an employee can experience the three components concurrently, previous research has been largely variable-centered, looking at the antecedents and outcomes of each component separately. Two studies explored how the three components combine to create distinct “profiles” of commitment and the implications of different profiles. In Study 1, six clusters were identified using k-means cluster ana...

  12. The inverse agonist DG172 triggers a PPARβ/δ-independent myeloid lineage shift and promotes GM-CSF/IL-4-induced dendritic cell differentiation.

    Science.gov (United States)

    Lieber, Sonja; Scheer, Frithjof; Finkernagel, Florian; Meissner, Wolfgang; Giehl, Gavin; Brendel, Cornelia; Diederich, Wibke E; Müller-Brüsselbach, Sabine; Müller, Rolf

    2015-02-01

    The stilbene derivative (Z)-2-(2-bromophenyl)-3-{[4-(1-methylpiperazine)amino]phenyl}acrylonitrile (DG172) was developed as a highly selective inhibitory peroxisome proliferator-activated receptor (PPAR)β/δ ligand. Here, we describe a novel PPARβ/δ-independent, yet highly specific, effect of DG172 on the differentiation of bone marrow cells (BMCs). DG172 strongly augmented granulocyte-macrophage-colony-stimulating factor (GM-CSF)-induced differentiation of primary BMCs from Ppard null mice into two specific populations, characterized as mature (CD11c(hi)MHCII(hi)) and immature (CD11c(hi)MHCII(lo)) dendritic cells (DCs). IL-4 synergized with DG172 to shift the differentiation from MHCII(lo) cells to mature DCs in vitro. The promotion of DC differentiation occurred at the expense of differentiation to granulocytic Gr1(+)Ly6B(+) cells. In agreement with these findings, transcriptome analyses showed a strong DG172-mediated repression of genes encoding neutrophilic markers in both differentiating wild-type and Ppard null cells, while macrophage/DC marker genes were up-regulated. DG172 also inhibited the expression of transcription factors driving granulocytic differentiation (Cebpe, Gfi1, and Klf5), and increased the levels of transcription factors promoting macrophage/DC differentiation (Irf4, Irf8, Spib, and Spic). DG172 exerted these effects only at an early stage of BMC differentiation induced by GM-CSF, did not affect macrophage-colony-stimulating factor-triggered differentiation to macrophages and had no detectable PPARβ/δ-independent effect on other cell types tested. Structure-function analyses demonstrated that the 4-methylpiperazine moiety in DG172 is required for its effect on DC differentiation, but is dispensable for PPARβ/δ binding. Based on these data we developed a new compound, (Z)-2-(4-chlorophenyl)-3-[4-(4-methylpiperazine-1-yl)phenyl]acrylonitrile (DG228), which enhances DC differentiation in the absence of significant PPARβ/δ binding. PMID

  13. Characterization of a human hematopoietic progenitor cell capable of forming blast cell containing colonies in vitro.

    OpenAIRE

    J. Brandt; Baird, N; Lu, L; Srour, E; R. HOFFMAN

    1988-01-01

    A hematopoietic cell (CFU-B1) capable of producing blast cell containing colonies in vitro was detected using a semisolid culture system. The CFU-B1 has the capacity for self-renewal and commitment to a number of hematopoietic lineages. Monoclonal antibody to the human progenitor cell antigen-1 (HPCA-1) and a monoclonal antibody against the major histocompatibility class II antigen (HLA-DR) were used with fluorescence activated cell sorting to phenotype the CFU-B1. The CFU-B1 was found to exp...

  14. Conditional Deletion of the Relaxin Receptor Gene in Cells of Smooth Muscle Lineage Affects Lower Reproductive Tract in Pregnant Mice1

    OpenAIRE

    Kaftanovskaya, Elena M.; Huang, Zaohua; Lopez, Carolina; Conrad, Kirk; Agoulnik, Alexander I.

    2015-01-01

    Relaxin hormone secreted into the circulation during pregnancy was discovered through its effects on pubic symphysis relaxation and parturition. Genetic inactivation of the relaxin gene or its cognate relaxin family peptide receptor 1 (RXFP1) in mice caused failure of parturition and mammary nipple enlargement, as well as increased collagen fiber density in the cervix and vagina. However, the relaxin effect on discrete cells and tissues has yet to be determined. Using transgenic mice with a k...

  15. Biliary tree stem/progenitor cells in glands of extrahepatic and intraheptic bile ducts: an anatomical in situ study yielding evidence of maturational lineages

    OpenAIRE

    Carpino, Guido; Cardinale, Vincenzo; Onori, Paolo; Franchitto, Antonio; Berloco, Pasquale Bartolomeo; Rossi, Massimo; Wang, Yunfang; Semeraro, Rossella; Anceschi, Maurizio; Brunelli, Roberto; Alvaro, Domenico; Reid, Lola M.; Gaudio, Eugenio

    2011-01-01

    Stem/progenitors have been identified intrahepatically in canals of Hering and extrahepatically in glands of the biliary tree. Glands of the biliary tree (peribiliary glands: PBGs) are tubulo-alveolar glands with mucinous and serous acini, located deep within intrahepatic and extrahepatic bile ducts. We have shown that biliary tree stem/progenitors (BTSCs) are multipotent, giving rise in vitro and in vivo to hepatocytes, cholangiocytes or pancreatic islets. Cells with the phenotype of BTSCs a...

  16. Lineage analysis of micromere 4d, a super-phylotypic cell for Lophotrochozoa, in the leech Helobdella and the sludgeworm Tubifex

    OpenAIRE

    Gline, Stephanie E.; Nakamoto, Ayaki; Cho, Sung-Jin; Chi, Candace; Weisblat, David A.

    2011-01-01

    The super-phylum Lophotrochozoa contains the plurality of extant animal phyla and exhibits a corresponding diversity of adult body plans. Moreover, in contrast to Ecdysozoa and Deuterostomia, most lophotrochozoans exhibit a conserved pattern of stereotyped early divisions called spiral cleavage. In particular, bilateral mesoderm in most lophotrochozoan species arises from the progeny of micromere 4d, which is assumed to be homologous with a similar cell in the embryo of the ancestral lophotro...

  17. Lysine-specific demethylase-1 (LSD1) is compartmentalized at nuclear chromocenters in early post-mitotic cells of the olfactory sensory neuronal lineage.

    Science.gov (United States)

    Kilinc, Seda; Savarino, Alyssa; Coleman, Julie H; Schwob, James E; Lane, Robert P

    2016-07-01

    Mammalian olfaction depends on the development of specialized olfactory sensory neurons (OSNs) that each express one odorant receptor (OR) protein from a large family of OR genes encoded in the genome. The lysine-specific demethylase-1 (LSD1) protein removes activating H3K4 or silencing H3K9 methylation marks at gene promoters and is required for proper OR regulation. We show that LSD1 protein exhibits variable organization within nuclei of developing OSNs, and tends to consolidate into a single dominant compartment at the edges of chromocenters within nuclei of early post-mitotic cells of the mouse olfactory epithelium (MOE). Using an immortalized cell line derived from developing olfactory placode, we show that consolidation of LSD1 appears to be cell-cycle regulated, with a peak occurrence in early G1. LSD1 co-compartmentalizes with CoREST, a protein known to collaborate with LSD1 to carry out a variety of chromatin-modifying functions. We show that LSD1 compartments co-localize with 1-3 OR loci at the exclusion of most OR genes, and commonly associate with Lhx2, a transcription factor involved in OR regulation. Together, our data suggests that LSD1 is sequestered into a distinct nuclear space that might restrict a histone-modifying function to a narrow developmental time window and/or range of OR gene targets. PMID:26947098

  18. Signalling and transcriptional regulation of early developmental lineage decisions

    DEFF Research Database (Denmark)

    Morgani, Sophie Maria Christina

    Embryonic stem (ES) cells are cell lines isolated from the embryo at a time just prior to implantation into the uterus. In the right cocktail of medium and cytokines, these cell lines can be maintained indefinitely in vitro in a self-renewing state. Initially it was assumed that these cells......’ towards a particular cell fate. These populations are also dynamic in nature, converting from one state to another with fairly rapid kinetics. The main focus of this thesis was to gain a more in depth understanding of the mechanisms regulating heterogeneity and lineage priming in murine ES cells by asking...

  19. LEVELS OF ORGANIZATIONAL COMMITMENT FOR FEDERATION EMPLOYEES

    OpenAIRE

    Hayri DEMİR; Selçuk BUĞDAYCI

    2011-01-01

    This paper deals with the levels of organizational commitment for general secretaries, sport experts and civil servants working in sport federations. 183 persons are included in the research. Data for the research have been collected through “Three Dimensional Organizational Commitment Scale” developed by Meyer, Allen and Smith (1993). In this paper, organizational commitment is examined in three different dimensions as emotional commitment, continuance commitment and normative commitment, an...

  20. Systemic expression of Kaposi sarcoma herpesvirus (KSHV) Vflip in endothelial cells leads to a profound proinflammatory phenotype and myeloid lineage remodeling in vivo.

    OpenAIRE

    Gianna Ballon; Gunkut Akar; Ethel Cesarman

    2015-01-01

    Author Summary Kaposi’s sarcoma (KS) is the most common cancer in men infected with HIV, and also among the most frequent malignancies in Sub-Equatorial Africa. KS is a tumor of endothelial cell origin that is caused by infection with a gamma-herpesvirus, called KS herpesvirus (KSHV) or human herpesvirus 8 (HHV-8). KSHV vFLIP is a viral oncoprotein expressed during latent infection. We report here the generation and characterization of mice expressing KSHV vFLIP in an inducible manner in endo...