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Vorobjova T, Metsküla K, Salumäe L, Uibo O, Heilman K, Uibo R. Immunohistochemical evaluation of LGR5, CD71, CD138 and CXCR3 markers in the small bowel mucosa of participants with celiac disease and persons with normal bowel mucosa. J Mol Histol 2025; 56:64. [PMID: 39747719 DOI: 10.1007/s10735-024-10340-z] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/16/2024] [Accepted: 12/15/2024] [Indexed: 01/04/2025]
Abstract
Celiac disease (CD) is a chronic autoimmune disease of the small bowel mucosa that develops because of the altered immune response to gluten, which leads to intestinal epithelium damage and villous atrophy. However, studies on regeneration of the damaged small bowel mucosa and density of intestinal stem cells (ISC) in CD persons are still scarce. We aimed to evaluate the number of small bowel mucosa cells positive for LGR5, CD138/Syndecan-1, CD71 and CXCR3 in CD and in controls with normal bowel mucosa; to find relationship between these markers and degree of small intestinal atrophy and to compare these results with our previous data about the number of CD103 + , IDO + DCs, FOXP3 + Tregs, enterovirus (EV) density and serum zonulin level. The paraffin sections of the small bowel biopsies were obtained from 26 children with CD (median age 6.5 years), and from 20 controls with normal intestinal mucosa (median age 14.2 years) and from the tissue bank of the Department of Pathology of Tartu University Hospital (from 18 participants with CD including 14 children (median age 13.2 years) and from 11subjects with normal small bowel mucosa, including one child aged 4.8 years. The number of LGR5 + , CD71 + , CD138 + , and CXCR3 + cells was evaluated using immunohistochemistry. The median number of CD138 + and CXCR3 + cells was significantly higher in the small bowel mucosa in CD compared with normal mucosa (p = 0.0002 for CD138 and p = 0.006 for CXCR3). The median number of CD71 + cells was significantly higher in normal small bowel mucosa (p = 0.005). The number of LGR5 + cells did not differ between persons with CD and those with normal small bowel mucosa (p = 0.7). A markedly increased number of CD138 + and CXCR3 + cells in the small bowel mucosa of participants with CD confirms their role in the pathogenesis of this disease. There was no expected marked difference in the density of any of the studied markers between lower or higher grade of small bowel atrophy and level of tTG-IgA in CD.
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Affiliation(s)
- Tamara Vorobjova
- Department of Immunology, Institute of Biomedicine and Translational Medicine, University of Tartu, Ravila 19, 51014, Tartu, Estonia.
| | - Kaja Metsküla
- Department of Immunology, Institute of Biomedicine and Translational Medicine, University of Tartu, Ravila 19, 51014, Tartu, Estonia
| | - Liis Salumäe
- Department of Pathology, Tartu University Hospital, Tartu, Estonia
| | - Oivi Uibo
- Children's Clinic, Tartu University Hospital, Tartu, Estonia
- Centre of Clinical Nutrition, Tartu University Hospital, Tartu, Estonia
- Department of Paediatrics, Institute of Clinical Medicine, University of Tartu, Tartu, Estonia
| | | | - Raivo Uibo
- Department of Immunology, Institute of Biomedicine and Translational Medicine, University of Tartu, Ravila 19, 51014, Tartu, Estonia
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2
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Tan L, Duan X, Mutyala P, Zhou T, Amin S, Zhang T, Herbst B, Askan G, Itkin T, Xiang Z, Michelassi F, Lieberman MD, Iacobuzio-Donahue CA, Leach SD, Evans T, Chen S. A targetable pathway to eliminate TRA-1-60+/TRA-1-81+ chemoresistant cancer cells. J Mol Cell Biol 2023; 15:mjad039. [PMID: 37327088 PMCID: PMC10847630 DOI: 10.1093/jmcb/mjad039] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/19/2022] [Revised: 12/18/2022] [Accepted: 06/14/2023] [Indexed: 06/18/2023] Open
Abstract
Chemoresistance is a primary cause of treatment failure in pancreatic cancer. Identifying cell surface markers specifically expressed in chemoresistant cancer cells (CCCs) could facilitate targeted therapies to overcome chemoresistance. We performed an antibody-based screen and found that TRA-1-60 and TRA-1-81, two 'stemness' cell surface markers, are highly enriched in CCCs. Furthermore, TRA-1-60+/TRA-1-81+ cells are chemoresistant compared to TRA-1-60-/TRA-1-81- cells. Transcriptome profiling identified UGT1A10, shown to be both necessary and sufficient to maintain TRA-1-60/TRA-1-81 expression and chemoresistance. From a high-content chemical screen, we identified Cymarin, which downregulates UGT1A10, eliminates TRA-1-60/TRA-1-81 expression, and increases chemosensitivity both in vitro and in vivo. Finally, TRA-1-60/TRA-1-81 expression is highly specific in primary cancer tissue and positively correlated with chemoresistance and short survival, which highlights their potentiality for targeted therapy. Therefore, we discovered a novel CCC surface marker regulated by a pathway that promotes chemoresistance, as well as a leading drug candidate to target this pathway.
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Affiliation(s)
- Lei Tan
- Department of Surgery, Weill Cornell Medicine, New York, NY 10065, USA
- Center for Energy Metabolism and Reproduction, Institute of Biomedicine and Biotechnology, Shenzhen Institute of Advanced Technology, Chinese Academy of Sciences, Shenzhen 518055, China
| | - Xiaohua Duan
- Department of Surgery, Weill Cornell Medicine, New York, NY 10065, USA
- Center for Genomic Health, Weill Cornell Medicine, New York, NY 10065, USA
| | - Pratyusha Mutyala
- Department of Surgery, Weill Cornell Medicine, New York, NY 10065, USA
| | - Ting Zhou
- The SKI Stem Cell Research Facility, The Center for Stem Cell Biology and Developmental Biology Program, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Sadaf Amin
- Department of Surgery, Weill Cornell Medicine, New York, NY 10065, USA
| | - Tuo Zhang
- Genomic Resource Core Facility, Weill Cornell Medical College, New York, NY 10065, USA
| | - Brian Herbst
- Rubenstein Center for Pancreatic Cancer Research, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Gokce Askan
- Rubenstein Center for Pancreatic Cancer Research, Memorial Sloan Kettering Cancer Center, New York, NY 10065, USA
| | - Tomer Itkin
- Division of Regenerative Medicine, Hartman Institute for Therapeutic Organ Regeneration, Ansary Stem Cell Institute, Department of Medicine, Weill Cornell Medicine, New York, NY 10065, USA
| | - Zhaoying Xiang
- Genomic Resource Core Facility, Weill Cornell Medical College, New York, NY 10065, USA
| | | | | | | | - Steven D Leach
- Dartmouth Cancer Center, Darmouth College, Hanover, NH 03755, USA
| | - Todd Evans
- Department of Surgery, Weill Cornell Medicine, New York, NY 10065, USA
- Center for Genomic Health, Weill Cornell Medicine, New York, NY 10065, USA
| | - Shuibing Chen
- Department of Surgery, Weill Cornell Medicine, New York, NY 10065, USA
- Center for Genomic Health, Weill Cornell Medicine, New York, NY 10065, USA
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3
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Singh A, Khan DUZ, Singh P, Singh AK, Agarwal P. Prognostic utility of microRNA-145 and CD 133 in oral squamous cell carcinoma: A pilot study from Northern India. J Oral Biol Craniofac Res 2022; 13:92-95. [DOI: 10.1016/j.jobcr.2022.11.008] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/13/2022] [Accepted: 11/30/2022] [Indexed: 12/03/2022] Open
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4
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Motohara T, Yoshida GJ, Katabuchi H. The hallmarks of ovarian cancer stem cells and niches: Exploring their harmonious interplay in therapy resistance. Semin Cancer Biol 2021; 77:182-193. [PMID: 33812986 DOI: 10.1016/j.semcancer.2021.03.038] [Citation(s) in RCA: 31] [Impact Index Per Article: 7.8] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 06/01/2020] [Revised: 03/20/2021] [Accepted: 03/29/2021] [Indexed: 12/18/2022]
Abstract
The concept of a "cancer stem cell" has evolved over the past decades, and research on cancer stem cell biology has entered into a stage of remarkable progress. Cancer stem cells are a major determining factor contributing to the establishment of phenotypic and functional intratumoral heterogeneity in synchronization with their surrounding "cancer stem cell niches." They serve as the driving force for cancer initiation, metastasis, and therapeutic resistance in various types of malignancies. In verity, reciprocal interplay between ovarian cancer stem cells and their niches involves a complex but ingeniously orchestrated tumor microenvironment within the intraperitoneal milieu and especially contribute to chemotherapy resistance in patients with advanced ovarian cancer. Herein, we review the principles of our current understanding of the biological features of ovarian cancer stem cells, focusing mainly on the precise mechanisms underlying acquired chemotherapy resistance. Furthermore, we highlight the specific roles of various cancer-associated stromal and immune cells in creating possible cancer stem cell niches that regulate ovarian cancer stemness.
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Affiliation(s)
- Takeshi Motohara
- Department of Obstetrics and Gynecology, Faculty of Life Sciences, Kumamoto University, 1-1-1 Honjo, Chuo-ku, Kumamoto City, Kumamoto, 860-8556, Japan.
| | - Go J Yoshida
- Department of Immunological Diagnosis, Juntendo University School of Medicine, 2-1-1 Hongo, Bunkyo-ku, Tokyo, 113-8421, Japan
| | - Hidetaka Katabuchi
- Department of Obstetrics and Gynecology, Faculty of Life Sciences, Kumamoto University, 1-1-1 Honjo, Chuo-ku, Kumamoto City, Kumamoto, 860-8556, Japan
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5
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de Maria S, Santoro A, Fuggetta MP, Rocchetti R, Cottarelli A, Lanzilli G, Stiuso P, Angelico G, Spadola S, Franco Zannoni G, Rubini C, Emanuelli M, Carmela Pedicillo M, Pannone G, Muzio LL. A possible interplay between HR-HPV and stemness in tumor development: an in vivo investigation of CD133 as a putative marker of cancer stem cell in HPV18-infected KB cell line. APMIS 2020; 128:637-646. [PMID: 32911563 DOI: 10.1111/apm.13078] [Citation(s) in RCA: 5] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/25/2020] [Accepted: 09/08/2020] [Indexed: 12/30/2022]
Abstract
High-risk HPVs (HR-HPVs) are DNA viruses considered as primary etiologic factors in malignancies of the low female genital tract. Their presence has also been documented in oropharyngeal and laryngeal cancers. However, HPV infection is considered a necessary but not sufficient cause of tumoral development; meantime, increasing evidences on the tumorigenic role of cancer stem cells (CSCs) have been documented in the literature. CSCs represent a small subpopulation of neoplastic cells with self-renewal potential, capable of maintaining tumor growth and cell differentiation, also involved in metastatic process, recurrence, and resistance to chemotherapeutic agents. In the present study, performed on KB cell lines, we evaluated the tumor forming potential of CSCs, and their relationship with the HPV infection status. We started our study by identifying the most aggressive cell line on the minimal number of cells being able of growth in vivo in a model of athymic nude mice (BALB/c nu/nu). We used an oral-derived KB cell line separated in the KB-CD133+ and KB-CD133- populations, by using immunomagnetic beads and fluorescence-activated cell sorting (FACS). The separated populations were injected in athymic nude mice (BALB/c nu/nu). Xenograft tumors have been analyzed for tumor size, CD133 expression by immunohistochemistry (IHC) and for DNA HR-HPV integration by in situ hybridization (ISH), comparing CD133-enriched xenograft tumors versus the CD133 non-enriched ones. On standard conditions, the KB cell line has a poor population of glycosylated CD133 marker (<5.0%) when investigated with antibodies versus CD133, and more specifically its glycosylated epitope (AC133). Enriched CD133 KB cells possess a higher capacity of tumor growth in xenograft models of nude mice when compared to KB CD133-negative cells. We observed that the AC133 epitope, extensively used to purifying hematopoietic stem cells, is able to select an epithelial subpopulation of cancer stem cells with aggressive behavior. We retain that CD133 may be a useful target in anticancer strategies including pharmacological and immunological therapies.
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Affiliation(s)
- Salvatore de Maria
- Department of Precision Medicine, University of Campania "Luigi Vanvitelli", Naples, Italy.,GLURES, Academic SPINOFF Ca Foscari University of Venice, Venice, Italy
| | - Angela Santoro
- Unità di Gineco-Patologia e Patologia Mammaria, Dipartimento Scienze della Salute della Donna, del Bambino e di Sanita Pubblica, Fondazione Policlinico Universitario A. Gemelli IRCCS, Roma, Italy
| | - Maria Pia Fuggetta
- GLURES, Academic SPINOFF Ca Foscari University of Venice, Venice, Italy.,Istituto di Farmacologia Traslazionale, Area Ricerca Tor Vergata, CNR, Rome, Italy
| | - Romina Rocchetti
- Dipartimento di Neuroscienze, Sezione di Anatomia Patologica, Università Politecnica delle Marche, Ancona, Italy
| | - Andrea Cottarelli
- Istituto di Farmacologia Traslazionale, Area Ricerca Tor Vergata, CNR, Rome, Italy
| | - Giulia Lanzilli
- Istituto di Farmacologia Traslazionale, Area Ricerca Tor Vergata, CNR, Rome, Italy
| | - Paola Stiuso
- Department of Precision Medicine, University of Campania "Luigi Vanvitelli", Naples, Italy
| | - Giuseppe Angelico
- Unità di Gineco-Patologia e Patologia Mammaria, Dipartimento Scienze della Salute della Donna, del Bambino e di Sanita Pubblica, Fondazione Policlinico Universitario A. Gemelli IRCCS, Roma, Italy
| | - Saveria Spadola
- Unità di Gineco-Patologia e Patologia Mammaria, Dipartimento Scienze della Salute della Donna, del Bambino e di Sanita Pubblica, Fondazione Policlinico Universitario A. Gemelli IRCCS, Roma, Italy
| | - Gian Franco Zannoni
- Unità di Gineco-Patologia e Patologia Mammaria, Dipartimento Scienze della Salute della Donna, del Bambino e di Sanita Pubblica, Fondazione Policlinico Universitario A. Gemelli IRCCS, Roma, Italy.,Istituto di Anatomia Patologica, Università Cattolica del Sacro Cuore, Roma, Italy
| | - Corrado Rubini
- Dipartimento di Neuroscienze, Sezione di Anatomia Patologica, Università Politecnica delle Marche, Ancona, Italy
| | - Monica Emanuelli
- Dipartimento di Neuroscienze, Sezione di Anatomia Patologica, Università Politecnica delle Marche, Ancona, Italy
| | | | - Giuseppe Pannone
- Dipartimento di Scienze Chirurgiche, Università degli Studi di Foggia, Foggia, Italy
| | - Lorenzo Lo Muzio
- Dipartimento di Scienze Chirurgiche, Università degli Studi di Foggia, Foggia, Italy
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Brault J, Vigne B, Meunier M, Beaumel S, Mollin M, Park S, Stasia MJ. NOX4 is the main NADPH oxidase involved in the early stages of hematopoietic differentiation from human induced pluripotent stem cells. Free Radic Biol Med 2020; 146:107-118. [PMID: 31626946 DOI: 10.1016/j.freeradbiomed.2019.10.005] [Citation(s) in RCA: 14] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 09/19/2019] [Revised: 10/08/2019] [Accepted: 10/08/2019] [Indexed: 02/07/2023]
Abstract
Reactive oxygen species (ROS) produced in hematopoietic stem cells (HSCs) are involved in the balance between quiescence, self-renewal, proliferation and differentiation processes. However the role of NOX enzymes on the early stages of hematopoietic differentiation is poorly investigated. For that, we used induced pluripotent stem cells (iPSCs) derived from X-linked Chronic Granulomatous Disease (X0CGD) patients with deficiency in NOX2, and AR220CGD patients with deficiency in p22phox subunit which decreases NOX1, NOX2, NOX3 and NOX4 activities. CD34+ hematopoietic progenitors were obtained after 7, 10 and 13 days of iPS/OP9 co-culture differentiation system. Neither NOX expression nor activity was found in Wild-type (WT), X0CGD and AR220CGD iPSCs. Although NOX2 and NOX4 mRNA were found in WT, X0CGD and AR220CGD iPSC-derived CD34+ cells at day 10 and 13 of differentiation, NOX4 protein was the only NOX enzyme expressed in these cells. A NADPH oxidase activity was measured in WT and X0CGD iPSC-derived CD34+ cells but not in AR220CGD iPSC-derived CD34+ cells because of the absence of p22phox, which is essential for the NOX4 activity. The absence of NOX4 activity and the poor NOX-independent ROS production in AR220CGD iPSC-derived CD34+ cells favored the CD34+ cells production but lowered their hematopoietic potential compared to WT and X0CGD iPSC-derived CD34+ cells. In addition we found a large production of primitive AR220CGD iPSC-derived progenitors at day 7 compared to the WT and X0CGD cell types. In conclusion NOX4 is the major NOX enzyme involved in the early stages of hematopoietic differentiation from iPSCs and its activity can modulate the production, the hematopoietic potential and the phenotype of iPSC-derived CD34+.
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Affiliation(s)
- Julie Brault
- Centre Hospitalier Universitaire Grenoble Alpes, CGD Diagnosis and Research Centre (CDiReC), Grenoble, France.
| | - Bénédicte Vigne
- Centre Hospitalier Universitaire Grenoble Alpes, CGD Diagnosis and Research Centre (CDiReC), Grenoble, France.
| | - Mathieu Meunier
- Centre Hospitalier Universitaire Grenoble Alpes, University Clinic of Hematology, Grenoble, France; CNRS UMR 5309, INSERM, U1209, Université Grenoble Alpes, Institute for Advanced Bioscience, 38700, Grenoble, France.
| | - Sylvain Beaumel
- Centre Hospitalier Universitaire Grenoble Alpes, CGD Diagnosis and Research Centre (CDiReC), Grenoble, France.
| | - Michelle Mollin
- Centre Hospitalier Universitaire Grenoble Alpes, CGD Diagnosis and Research Centre (CDiReC), Grenoble, France.
| | - Sophie Park
- Centre Hospitalier Universitaire Grenoble Alpes, University Clinic of Hematology, Grenoble, France; CNRS UMR 5309, INSERM, U1209, Université Grenoble Alpes, Institute for Advanced Bioscience, 38700, Grenoble, France.
| | - Marie José Stasia
- Centre Hospitalier Universitaire Grenoble Alpes, CGD Diagnosis and Research Centre (CDiReC), Grenoble, France; Univ. Grenoble Alpes, CEA, CNRS, IBS, F-38044, Grenoble, France, Grenoble, France.
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7
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Kato Y, Ohishi T, Yamada S, Itai S, Furusawa Y, Sano M, Nakamura T, Kawada M, Kaneko MK. Anti-CD133 Monoclonal Antibody CMab-43 Exerts Antitumor Activity in a Mouse Xenograft Model of Colon Cancer. Monoclon Antib Immunodiagn Immunother 2019; 38:75-78. [PMID: 30969150 DOI: 10.1089/mab.2019.0002] [Citation(s) in RCA: 13] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/19/2023] Open
Abstract
Cancer stem cells contribute to tumorigenesis, metastasis, recurrence, and chemoresistance. CD133/prominin-1-a pentaspan membrane glycoprotein-has been used as a stem cell biomarker for the isolation of stem-like cells from a variety of normal and pathological tissues. In our previous studies, we developed several anti-CD133 monoclonal antibodies using Cell-Based Immunization and Screening (CBIS) methods, followed by characterization of their efficacy by flow cytometry, western blotting, and immunohistochemical analyses. One of the 100 clones, CMab-43 (IgG2a, kappa), demonstrated a sensitive and specific reaction against colon cancer cells. This study aimed to investigate the antitumor activity of CMab-43. Caco-2 cells (human colon cancer cell line) were subcutaneously implanted into the flanks of nude mice. CMab-43 and control mouse IgG were injected three times into the peritoneal cavity of mice. Tumor formation was observed in the control and CMab-43-treated mice of Caco-2 xenograft models. CMab-43 significantly reduced tumor development of Caco-2 xenograft in comparison with the control mouse IgG on days 12, 14, and 17. Our results cumulatively suggest that CMab-43 is useful for antibody therapy against CD133-expressing colon cancers.
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Affiliation(s)
- Yukinari Kato
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan.,2 New Industry Creation Hatchery Center, Tohoku University, Sendai, Japan
| | - Tomokazu Ohishi
- 3 Institute of Microbial Chemistry (BIKAKEN), Numazu, Microbial Chemistry Research Foundation, Numazu-shi, Japan
| | - Shinji Yamada
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Shunsuke Itai
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Yoshikazu Furusawa
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan.,2 New Industry Creation Hatchery Center, Tohoku University, Sendai, Japan
| | - Masato Sano
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Takuro Nakamura
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan
| | - Manabu Kawada
- 3 Institute of Microbial Chemistry (BIKAKEN), Numazu, Microbial Chemistry Research Foundation, Numazu-shi, Japan
| | - Mika K Kaneko
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine, Sendai, Japan
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8
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Shear stress: An essential driver of endothelial progenitor cells. J Mol Cell Cardiol 2018; 118:46-69. [PMID: 29549046 DOI: 10.1016/j.yjmcc.2018.03.007] [Citation(s) in RCA: 47] [Impact Index Per Article: 6.7] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 01/03/2018] [Revised: 03/08/2018] [Accepted: 03/09/2018] [Indexed: 02/06/2023]
Abstract
The blood flow through vessels produces a tangential, or shear, stress sensed by their innermost layer (i.e., endothelium) and representing a major hemodynamic force. In humans, endothelial repair and blood vessel formation are mainly performed by circulating endothelial progenitor cells (EPCs) characterized by a considerable expression of vascular endothelial growth factor receptor 2 (VEGFR2), CD34, and CD133, pronounced tube formation activity in vitro, and strong reendothelialization or neovascularization capacity in vivo. EPCs have been proposed as a promising agent to induce reendothelialization of injured arteries, neovascularization of ischemic tissues, and endothelialization or vascularization of bioartificial constructs. A number of preconditioning approaches have been suggested to improve the regenerative potential of EPCs, including the use of biophysical stimuli such as shear stress. However, in spite of well-defined influence of shear stress on mature endothelial cells (ECs), articles summarizing how it affects EPCs are lacking. Here we discuss the impact of shear stress on homing, paracrine effects, and differentiation of EPCs. Unidirectional laminar shear stress significantly promotes homing of circulating EPCs to endothelial injury sites, induces anti-thrombotic and anti-atherosclerotic phenotype of EPCs, increases their capability to form capillary-like tubes in vitro, and enhances differentiation of EPCs into mature ECs in a dose-dependent manner. These effects are mediated by VEGFR2, Tie2, Notch, and β1/3 integrin signaling and can be abrogated by means of complementary siRNA/shRNA or selective pharmacological inhibitors of the respective proteins. Although the testing of sheared EPCs for vascular tissue engineering or regenerative medicine applications is still an unaccomplished task, favorable effects of unidirectional laminar shear stress on EPCs suggest its usefulness for their preconditioning.
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Itai S, Fujii Y, Nakamura T, Chang YW, Yanaka M, Saidoh N, Handa S, Suzuki H, Harada H, Yamada S, Kaneko MK, Kato Y. Establishment of CMab-43, a Sensitive and Specific Anti-CD133 Monoclonal Antibody, for Immunohistochemistry. Monoclon Antib Immunodiagn Immunother 2017; 36:231-235. [PMID: 28910211 PMCID: PMC6975129 DOI: 10.1089/mab.2017.0031] [Citation(s) in RCA: 60] [Impact Index Per Article: 7.5] [Reference Citation Analysis] [Abstract] [Key Words] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/15/2022] Open
Abstract
CD133, also known as prominin-1, was first described as a cell surface marker on early progenitor and hematopoietic stem cells. It is a five-domain transmembrane protein composed of an N-terminal extracellular tail, two small cytoplasmic loops, two large extracellular loops containing seven potential glycosylation sites, and a short C-terminal intracellular tail. CD133 has been used as a marker to identify cancer stem cells derived from primary solid tumors and as a prognostic marker of gliomas. Herein, we developed novel anti-CD133 monoclonal antibodies (mAbs) and characterized their efficacy in flow cytometry, Western blot, and immunohistochemical analyses. We expressed the full length of CD133 in LN229 glioblastoma cells, immunized mice with LN229/CD133 cells, and performed the first screening using flow cytometry. After limiting dilution, we established 100 anti-CD133 mAbs, reacting with LN229/CD133 cells but not with LN229 cells. Subsequently, we performed the second and third screening with Western blot and immunohistochemical analyses, respectively. Among 100 mAbs, 11 strongly reacted with CD133 in Western blot analysis. One of 11 clones, CMab-43 (IgG2a, kappa), showed a sensitive and specific reaction against colon cancer cells, warranting the use of CMab-43 in detecting CD133 in pathological analyses of CD133-expressing cancers.
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Affiliation(s)
- Shunsuke Itai
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan .,2 Department of Oral and Maxillofacial Surgery, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University , Tokyo, Japan
| | - Yuki Fujii
- 3 Department of Regional Innovation, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Takuro Nakamura
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Yao-Wen Chang
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Miyuki Yanaka
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Noriko Saidoh
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Saori Handa
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Hiroyoshi Suzuki
- 4 Department of Pathology and Laboratory Medicine, Sendai Medical Center , Sendai, Japan
| | - Hiroyuki Harada
- 2 Department of Oral and Maxillofacial Surgery, Graduate School of Medical and Dental Sciences, Tokyo Medical and Dental University , Tokyo, Japan
| | - Shinji Yamada
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Mika K Kaneko
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan
| | - Yukinari Kato
- 1 Department of Antibody Drug Development, Tohoku University Graduate School of Medicine , Sendai, Japan .,3 Department of Regional Innovation, Tohoku University Graduate School of Medicine , Sendai, Japan .,5 New Industry Creation Hatchery Center, Tohoku University , Sendai, Japan
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10
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Staege MS. Gene Expression Music Algorithm-Based Characterization of the Ewing Sarcoma Stem Cell Signature. Stem Cells Int 2016; 2016:7674824. [PMID: 27446218 PMCID: PMC4944079 DOI: 10.1155/2016/7674824] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 04/08/2016] [Accepted: 06/15/2016] [Indexed: 12/19/2022] Open
Abstract
Gene Expression Music Algorithm (GEMusicA) is a method for the transformation of DNA microarray data into melodies that can be used for the characterization of differentially expressed genes. Using this method we compared gene expression profiles from endothelial cells (EC), hematopoietic stem cells, neuronal stem cells, embryonic stem cells (ESC), and mesenchymal stem cells (MSC) and defined a set of genes that can discriminate between the different stem cell types. We analyzed the behavior of public microarray data sets from Ewing sarcoma ("Ewing family tumors," EFT) cell lines and biopsies in GEMusicA after prefiltering DNA microarray data for the probe sets from the stem cell signature. Our results demonstrate that individual Ewing sarcoma cell lines have a high similarity to ESC or EC. Ewing sarcoma cell lines with inhibited Ewing sarcoma breakpoint region 1-Friend leukemia virus integration 1 (EWSR1-FLI1) oncogene retained the similarity to ESC and EC. However, correlation coefficients between GEMusicA-processed expression data between EFT and ESC decreased whereas correlation coefficients between EFT and EC as well as between EFT and MSC increased after knockdown of EWSR1-FLI1. Our data support the concept of EFT being derived from cells with features of embryonic and endothelial cells.
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Affiliation(s)
- Martin Sebastian Staege
- University Clinic and Polyclinic for Child and Adolescent Medicine, Martin Luther University of Halle-Wittenberg, 06120 Halle (Saale), Germany
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11
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Piscaglia AC, Rutella S, Laterza L, Cesario V, Campanale M, Cazzato IA, Ianiro G, Barbaro F, Di Maurizio L, Bonanno G, Cenci T, Cammarota G, Larocca LM, Gasbarrini A. Circulating hematopoietic stem cells and putative intestinal stem cells in coeliac disease. J Transl Med 2015; 13:220. [PMID: 26160352 PMCID: PMC4498508 DOI: 10.1186/s12967-015-0591-0] [Citation(s) in RCA: 11] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 05/27/2015] [Accepted: 06/30/2015] [Indexed: 12/23/2022] Open
Abstract
Background The intestinal stem cells (ISC) modulation and the role of circulating hematopoietic stem cells (HSC) in coeliac disease (CD) are poorly understood. Our aim was to investigate the longitudinal modifications in peripheral blood HSC traffic and putative ISC density induced by gluten-free diet (GFD) in CD. Methods Thirty-one CD patients and 7 controls were enrolled. Circulating CD133+ and CD34+ HSC were measured by flow cytometry, at enrolment and after 7 days and 1, 3, 6, 12, and 24 months of GFD. Endoscopy was performed at diagnosis and repeated at 6, 12, and 24 months following GFD. We used the Marsh-Oberhuber score to evaluate the histological severity of duodenal damage; immunohistochemistry was employed to measure the intraepithelial lymphoid infiltrate (IEL, CD3+ lymphoid cells) and the putative ISC compartment (CD133+ and Lgr5+ epithelial cells). Results At enrolment, circulating HSCs were significantly increased in CD patients and they further augmented during the first week of GFD, but progressively decreased afterwards. CD patients presented with villous atrophy, abundant IEL and rare ISC residing at the crypt base. Upon GFD, IEL progressively decreased, while ISC density increased, peaking at 12 months. After 24 months of GFD, all patients were asymptomatic and their duodenal mucosa was macroscopically and histologically normal. Conclusions In active CD patients, the ISC niche is depleted and there is an increased traffic of circulating HSC versus non-coeliac subjects. GFD induces a precocious mobilization of circulating HSC, which is followed by the expansion of the local ISC compartment, leading to mucosal healing and clinical remission.
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Affiliation(s)
- Anna Chiara Piscaglia
- Endoscopy and Gastroenterology Unit, State Hospital, Borgo Maggiore, Republic of San Marino. .,Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Sergio Rutella
- Division of Translational Medicine, Clinical Research Centre, Sidra Medical and Research Centre, PO Box 26999, Burj Doha, 8th Floor, Doha, Qatar.
| | - Lucrezia Laterza
- Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Valentina Cesario
- Endoscopy and Gastroenterology Unit, State Hospital, Borgo Maggiore, Republic of San Marino. .,Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Mariachiara Campanale
- Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | | | - Gianluca Ianiro
- Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Federico Barbaro
- Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Luca Di Maurizio
- Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Giuseppina Bonanno
- Institute of Gynecology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Tonia Cenci
- Institute of Pathology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Giovanni Cammarota
- Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Luigi Maria Larocca
- Institute of Pathology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
| | - Antonio Gasbarrini
- Institute of Internal Medicine and Gastroenterology, "A. Gemelli" Hospital, Catholic University, Rome, Italy.
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12
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Park KS, Shin SW, Choi JW, Um SH. Specific protein markers for stem cell cross-talk with neighboring cells in the environment. Int J Stem Cells 2014; 6:75-86. [PMID: 24386551 DOI: 10.15283/ijsc.2013.6.2.75] [Citation(s) in RCA: 2] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Accepted: 07/29/2013] [Indexed: 01/04/2023] Open
Abstract
A stem cell interacts with the neighboring cells in its environment. To maintain a living organism's metabolism, either cell-cell or cell-environment interactions may be significant. Usually, these cells communicate with each other through biological signaling by interactive behaviors of primary proteins or complementary chemicals. The signaling intermediates offer the stem cell's functionality on its metabolism. With the rapid advent of omics technologies, various specific markers by which stem cells cooperate with their surroundings have been discovered and established. In this article, we review several stem cell markers used to communicate with either cancer or immune cells in the human body.
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Affiliation(s)
- Kyung Soo Park
- Department of Chemical and Biomolecular Engineering and Sogang University, Seoul, Korea
| | - Seung Won Shin
- School of Chemical Engineering and Sungkyunkwan University, Suwon
| | - Jeong-Woo Choi
- Department of Chemical and Biomolecular Engineering and Sogang University, Seoul, Korea ; Graduate School of Management of Technology, Sogang University, Seoul, Korea
| | - Soong Ho Um
- School of Chemical Engineering and Sungkyunkwan University, Suwon ; SKKU Advanced Institute of Nanotechnology (SAINT), Sungkyunkwan University, Suwon
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13
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Appaix F, Nissou MF, Sanden BVD, Dreyfus M, Berger F, Issartel JP, Wion D. Brain mesenchymal stem cells: The other stem cells of the brain? World J Stem Cells 2014; 6:134-143. [PMID: 24772240 PMCID: PMC3999771 DOI: 10.4252/wjsc.v6.i2.134] [Citation(s) in RCA: 51] [Impact Index Per Article: 4.6] [Reference Citation Analysis] [Abstract] [Key Words] [Track Full Text] [Download PDF] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Received: 11/12/2013] [Accepted: 02/20/2014] [Indexed: 02/06/2023] Open
Abstract
Multipotent mesenchymal stromal cells (MSC), have the potential to differentiate into cells of the mesenchymal lineage and have non-progenitor functions including immunomodulation. The demonstration that MSCs are perivascular cells found in almost all adult tissues raises fascinating perspectives on their role in tissue maintenance and repair. However, some controversies about the physiological role of the perivascular MSCs residing outside the bone marrow and on their therapeutic potential in regenerative medicine exist. In brain, perivascular MSCs like pericytes and adventitial cells, could constitute another stem cell population distinct to the neural stem cell pool. The demonstration of the neuronal potential of MSCs requires stringent criteria including morphological changes, the demonstration of neural biomarkers expression, electrophysiological recordings, and the absence of cell fusion. The recent finding that brain cancer stem cells can transdifferentiate into pericytes is another facet of the plasticity of these cells. It suggests that the perversion of the stem cell potential of pericytes might play an even unsuspected role in cancer formation and tumor progression.
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14
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Meng J, Chun S, Asfahani R, Lochmüller H, Muntoni F, Morgan J. Human skeletal muscle-derived CD133(+) cells form functional satellite cells after intramuscular transplantation in immunodeficient host mice. Mol Ther 2014; 22:1008-17. [PMID: 24569833 DOI: 10.1038/mt.2014.26] [Citation(s) in RCA: 54] [Impact Index Per Article: 4.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/18/2013] [Accepted: 02/16/2014] [Indexed: 12/28/2022] Open
Abstract
Stem cell therapy is a promising strategy for treatment of muscular dystrophies. In addition to muscle fiber formation, reconstitution of functional stem cell pool by donor cells is vital for long-term treatment. We show here that some CD133(+) cells within human muscle are located underneath the basal lamina of muscle fibers, in the position of the muscle satellite cell. Cultured hCD133(+) cells are heterogeneous and multipotent, capable of forming myotubes and reserve satellite cells in vitro. They contribute to extensive muscle regeneration and satellite cell formation following intramuscular transplantation into irradiated and cryodamaged tibialis anterior muscles of immunodeficient Rag2-/γ chain-/C5-mice. Some donor-derived satellite cells expressed the myogenic regulatory factor MyoD, indicating that they were activated. In addition, when transplanted host muscles were reinjured, there was significantly more newly-regenerated muscle fibers of donor origin in treated than in control, nonreinjured muscles, indicating that hCD133(+) cells had given rise to functional muscle stem cells, which were able to activate in response to injury and contribute to a further round of muscle regeneration. Our findings provide new evidence for the location and characterization of hCD133(+) cells, and highlight that these cells are highly suitable for future clinical application.
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Affiliation(s)
- Jinhong Meng
- The Dubowitz Neuromuscular Centre, UCL Institute of Child Health, London, UK
| | - Soyon Chun
- The Dubowitz Neuromuscular Centre, UCL Institute of Child Health, London, UK
| | - Rowan Asfahani
- The Dubowitz Neuromuscular Centre, UCL Institute of Child Health, London, UK
| | - Hanns Lochmüller
- Institute of Genetic Medicine, Newcastle University, International Centre for Life, Central Parkway, Newcastle upon Tyne, UK
| | - Francesco Muntoni
- The Dubowitz Neuromuscular Centre, UCL Institute of Child Health, London, UK
| | - Jennifer Morgan
- The Dubowitz Neuromuscular Centre, UCL Institute of Child Health, London, UK
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15
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Welte Y, Davies C, Schäfer R, Regenbrecht CRA. Patient derived cell culture and isolation of CD133⁺ putative cancer stem cells from melanoma. J Vis Exp 2013:e50200. [PMID: 23525090 DOI: 10.3791/50200] [Citation(s) in RCA: 12] [Impact Index Per Article: 1.0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/01/2023] Open
Abstract
Despite improved treatments options for melanoma available today, patients with advanced malignant melanoma still have a poor prognosis for progression-free and overall survival. Therefore, translational research needs to provide further molecular evidence to improve targeted therapies for malignant melanomas. In the past, oncogenic mechanisms related to melanoma were extensively studied in established cell lines. On the way to more personalized treatment regimens based on individual genetic profiles, we propose to use patient-derived cell lines instead of generic cell lines. Together with high quality clinical data, especially on patient follow-up, these cells will be instrumental to better understand the molecular mechanisms behind melanoma progression. Here, we report the establishment of primary melanoma cultures from dissected fresh tumor tissue. This procedure includes mincing and dissociation of the tissue into single cells, removal of contaminations with erythrocytes and fibroblasts as well as primary culture and reliable verification of the cells' melanoma origin. Recent reports revealed that melanomas, like the majority of tumors, harbor a small subpopulation of cancer stem cells (CSCs), which seem to exclusively fuel tumor initiation and progression towards the metastatic state. One of the key markers for CSC identification and isolation in melanoma is CD133. To isolate CD133(+) CSCs from primary melanoma cultures, we have modified and optimized the Magnetic-Activated Cell Sorting (MACS) procedure from Miltenyi resulting in high sorting purity and viability of CD133(+) CSCs and CD133(-) bulk, which can be cultivated and functionally analyzed thereafter.
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Affiliation(s)
- Yvonne Welte
- Institute of Pathology, Laboratory of Molecular Tumor Pathology, Charité - Universitätsmedizin Berlin.
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16
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Defining early human NK cell developmental stages in primary and secondary lymphoid tissues. PLoS One 2012; 7:e30930. [PMID: 22319595 PMCID: PMC3272048 DOI: 10.1371/journal.pone.0030930] [Citation(s) in RCA: 46] [Impact Index Per Article: 3.5] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/14/2011] [Accepted: 12/26/2011] [Indexed: 11/19/2022] Open
Abstract
A better understanding of human NK cell development in vivo is crucial to exploit NK cells for immunotherapy. Here, we identified seven distinctive NK cell developmental stages in bone marrow of single donors using 10-color flow cytometry and found that NK cell development is accompanied by early expression of stimulatory co-receptor CD244 in vivo. Further analysis of cord blood (CB), peripheral blood (PB), inguinal lymph node (inLN), liver lymph node (liLN) and spleen (SPL) samples showed diverse distributions of the NK cell developmental stages. In addition, distinctive expression profiles of early development marker CD33 and C-type lectin receptor NKG2A between the tissues, suggest that differential NK cell differentiation may take place at different anatomical locations. Differential expression of NKG2A and stimulatory receptors (e.g. NCR, NKG2D) within the different subsets of committed NK cells demonstrated the heterogeneity of the CD56(bright)CD16⁺/⁻ and CD56(dim)CD16⁺ subsets within the different compartments and suggests that microenvironment may play a role in differential in situ development of the NK cell receptor repertoire of committed NK cells. Overall, differential in situ NK cell development and trafficking towards multiple tissues may give rise to a broad spectrum of mature NK cell subsets found within the human body.
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17
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Olsen Hult LT, Kleiveland CR, Fosnes K, Jacobsen M, Lea T. EP receptor expression in human intestinal epithelium and localization relative to the stem cell zone of the crypts. PLoS One 2011; 6:e26816. [PMID: 22046368 PMCID: PMC3201980 DOI: 10.1371/journal.pone.0026816] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 02/24/2011] [Accepted: 10/05/2011] [Indexed: 01/22/2023] Open
Abstract
There is substantial evidence for PGE2 affecting intestinal epithelial proliferation. PGE2 is also reported to be involved in the regulation of growth and differentiation in adult stem cells, both effects mediated by binding to EP-receptors. We have used the Lgr5 as a marker to scrutinize EP-receptor and COX expression in human intestinal epithelial cells with focus on the stem cell area of the crypts. Normal tissue from ileum and colon, but also duodenal biopsies from patients with untreated celiac disease, were investigated by immunohistochemistry and RT-PCR. The combination of fresh flash-frozen tissue and laser microdissection made it possible to isolate RNA from the epithelial cell layer, only. In the small intestine, Lgr5 labels cells are in the +4 position, while in the colon, Lgr5 positive cells are localized to the crypt bottoms. Epithelial crypt cells of normal small intestine expressed neither EP-receptor mRNA nor COX1/2. However, crypt cells in tissue from patients with untreated celiac disease expressed EP2/4 receptor and COX1 mRNA. In the colon, the situation was different. Epithelial crypt cells from normal colon were found to express EP2/4 receptor and COX1/2 transcripts. Thus, there are distinct differences between normal human small intestine and colon with regard to expression of EP2/4 receptors and COX1/2. In normal colon tissue, PGE2-mediated signaling through EP-receptors 2/4 could be involved in regulation of growth and differentiation of the epithelium, while the lack of EP-receptor expression in the small intestinal tissue exclude the possibility of a direct effect of PGE2 on the crypt epithelial cells.
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Affiliation(s)
- Lene Th. Olsen Hult
- Molecular Cell Biology Group, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, Aas, Norway
| | - Charlotte R. Kleiveland
- Molecular Cell Biology Group, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, Aas, Norway
- * E-mail:
| | - Kjetil Fosnes
- Molecular Cell Biology Group, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, Aas, Norway
| | - Morten Jacobsen
- Molecular Cell Biology Group, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, Aas, Norway
- Ostfold Hospital, Fredrikstad, Norway
| | - Tor Lea
- Molecular Cell Biology Group, Department of Chemistry, Biotechnology and Food Science, Norwegian University of Life Sciences, Aas, Norway
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18
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A PEDF-derived peptide inhibits retinal neovascularization and blocks mobilization of bone marrow-derived endothelial progenitor cells. EXPERIMENTAL DIABETES RESEARCH 2011; 2012:518426. [PMID: 21754923 PMCID: PMC3132462 DOI: 10.1155/2012/518426] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.5] [Reference Citation Analysis] [Abstract] [Track Full Text] [Download PDF] [Figures] [Subscribe] [Scholar Register] [Received: 04/01/2011] [Accepted: 04/27/2011] [Indexed: 01/18/2023]
Abstract
Proliferative diabetic retinopathy is characterized by pathological retinal neovascularization, mediated by both angiogenesis (involving mature endothelial cells) and vasculogenesis (involving bone marrow-derived circulating endothelial progenitor cells (EPCs)). Pigment epithelium-derived factor (PEDF) contains an N-terminal 34-amino acid peptide (PEDF-34) that has antiangiogenic properties. Herein, we present a novel finding that PEDF-34 also possesses antivasculogenic activity. In the oxygen-induced retinopathy (OIR) model using transgenic mice that have Tie2 promoter-driven GFP expression, we quantified Tie2GFP+ cells in bone marrow and peripheral blood by fluorescence-activated cell sorting (FACS). OIR significantly increased the number of circulating Tie2-GFP+ at P16, correlating with the peak progression of neovascularization. Daily intraperitoneal injections of PEDF-34 into OIR mice decreased the number of Tie2-GFP+ cells in the circulation at P16 by 65% but did not affect the number of Tie2-GFP+ cells in the bone marrow. These studies suggest that PEDF-34 attenuates EPC mobilization from the bone marrow into the blood circulation during retinal neovascularization.
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19
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Park DH, Lee JH, Borlongan CV, Sanberg PR, Chung YG, Cho TH. Transplantation of umbilical cord blood stem cells for treating spinal cord injury. Stem Cell Rev Rep 2011; 7:181-94. [PMID: 20532836 DOI: 10.1007/s12015-010-9163-0] [Citation(s) in RCA: 54] [Impact Index Per Article: 3.9] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
Spinal cord injury (SCI) develops primary and secondary damage to neural tissue and this often results in permanent disability of the motor and sensory functions. However, there is currently no effective treatment except methylprednisolone, and the use of methylprednisolone has also been questioned due to its moderate efficacy and the drug's downside. Regenerative medicine has remarkably developed since the discovery of stem cells, and many studies have suggested the potential of cell-based therapies for neural injury. Especially, the therapeutic potential of human umbilical cord blood cells (hUCB cells) for intractable neurological disorders has been demonstrated using in vitro and vivo models. The hUCB cells are immune naïve and they are able to differentiate into other phenotypes, including the neural lineage. Their ability to produce several neurotropic factors and to modulate immune and inflammatory reactions has also been noted. Recent evidence has emerged suggesting alternative pathways of graft-mediated neural repair that involve neurotrophic effects. These effects are caused by the release of various growth factors that promote cell survival, angiogenesis and anti-inflammation, and this is all aside from a cell replacement mechanism. In this review, we present the recent findings on the stemness properties and the therapeutic potential of hUCB as a safe, feasible and effective cellular source for transplantation in SCI. These multifaceted protective and restorative effects from hUCB grafts may be interdependent and they act in harmony to promote therapeutic benefits for SCI. Nevertheless, clinical studies with hUCB are still rare because of the concerns about safety and efficiency. Among these concerns, the major histocompatibility in allogeneic transplantation is an important issue to be addressed in future clinical trials for treating SCI.
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Affiliation(s)
- Dong-Hyuk Park
- Department of Neurosurgery, Korea University Medical Center, Anam Hospital, Korea University College of Medicine, #126, 5-GA, Anam-Dong, Sungbuk-Ku, Seoul 136-705, Korea.
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20
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Kurth I, Franke K, Pompe T, Bornhäuser M, Werner C. Extracellular matrix functionalized microcavities to control hematopoietic stem and progenitor cell fate. Macromol Biosci 2011; 11:739-47. [PMID: 21341371 DOI: 10.1002/mabi.201000432] [Citation(s) in RCA: 38] [Impact Index Per Article: 2.7] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/28/2010] [Revised: 01/12/2011] [Indexed: 11/07/2022]
Abstract
Polymeric microcavities functionalized with extracellular matrix components were used as an experimental in vitro model to investigate principles of hematopoietic stem and progenitor cell (HSPC) fate control. Using human CD133+ HSPC we could demonstrate distinct differences in HSPC cycling and differentiation dependence on the adhesion ligand specificity (i.e., heparin, collagen I) and cytokine levels. The presented microcavity platform provides a powerful in vitro approach to explore the role of exogenous cues in HSPC fate decisions and can therefore be instrumental to progress in stem cell biology and translational research toward new therapies.
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Affiliation(s)
- Ina Kurth
- Max Bergmann Center of Biomaterials Dresden, Leibniz Institute of Polymer Research Dresden, Dresden, Germany
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21
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Dendritic cells with lymphocyte-stimulating activity differentiate from human CD133 positive precursors. Blood 2011; 117:3983-95. [PMID: 21304102 DOI: 10.1182/blood-2010-08-299735] [Citation(s) in RCA: 8] [Impact Index Per Article: 0.6] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/27/2022] Open
Abstract
CD133 is a hallmark of primitive myeloid progenitors. We have addressed whether human cord blood cells selected for CD133 can generate dendritic cells, and Langerhans cells in particular, in conditions that promote that generation from CD34(+) progenitors. Transforming growth factor-β1 (TGF-β1) and anti-TGF-β1 antibody, respectively, were added in some experiments. With TGF-β, monocytoid cells were recognized after 7 days. Immunophenotypically immature dendritic cells were present at day 14. After 4 more days, the cells expressed CD54, CD80, CD83, and CD86 and were potent stimulators in mixed lymphocyte reaction; part of the cells expressed CD1a and langerin, but not Birbeck granules. Without TGF-β, only a small fraction of cells acquired a dendritic shape and expressed the maturation-related antigens, and lymphocytes were poorly stimulated. With anti-TGF-β, the cell growth was greatly hampered, CD54 and langerin were never expressed, and lymphocytes were stimulated weakly. In conclusion, CD133(+) progenitors can give rise in vitro, through definite steps, to mature, immunostimulatory dendritic cells with molecular features of Langerhans cells, although without Birbeck granules. Addition of TGF-β1 helps to stimulate cell growth and promotes the acquisition of mature immunophenotypical and functional features. Neither langerin nor Birbeck granules proved indispensable for lymphocyte stimulation.
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Yu D, Zhang Y, Zou Y, Qin J, Li X, Xiao H, Tao D, Hu J, Gong J. Proliferation characteristics of CD133+ cell population in colorectal cancer. ACTA ACUST UNITED AC 2010; 30:751-6. [PMID: 21181366 DOI: 10.1007/s11596-010-0652-9] [Citation(s) in RCA: 0] [Impact Index Per Article: 0] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 03/26/2010] [Indexed: 10/18/2022]
Abstract
In this study, CD133+ subpopulations were isolated from 41 primary colorectal cancer tissues, the proliferation and cell cycle distribution of the cells were examined without in vitro expansion, and then compared to those of cell lines. The detection of CD133 in colorectal cancer tissues, isolation of CD133+ and CD133- epithelial subpopulations, Ki-67/DNA multiparameter assay and cell volume analysis were flow cytometrically conducted. The results showed that Ki-67 expression was correlated with CD133 level in primary cancer tissues, while cell cycle G2/M phase distribution or clinicopathological characteristics was not. In addition, the CD133+ cells showed larger cell volume and higher Ki-67 expression as compared with CD133- cells. But there was no statistically significant difference in G(2)/M phase distribution between the two subpopulations. Our results demonstrated that the CD133+ subpopulation in colorectal cancer tissue contained more actively cycling and proliferating cells, which was not correlated to clinicopathological factors but might contribute to tumor progression and poor clinical outcome.
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Affiliation(s)
- Dongdong Yu
- Department of General Surgery, Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China.
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23
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Welte Y, Adjaye J, Lehrach HR, Regenbrecht CR. Cancer stem cells in solid tumors: elusive or illusive? Cell Commun Signal 2010; 8:6. [PMID: 20459772 PMCID: PMC2880310 DOI: 10.1186/1478-811x-8-6] [Citation(s) in RCA: 67] [Impact Index Per Article: 4.5] [Reference Citation Analysis] [Abstract] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 01/20/2010] [Accepted: 05/11/2010] [Indexed: 12/21/2022] Open
Abstract
During the past years in vivo transplantation experiments and in vitro colony-forming assays indicated that tumors arise only from rare cells. These cells were shown to bear self-renewal capacities and the ability to recapitulate all cell types within an individual tumor. Due to their phenotypic resemblance to normal stem cells, the term "cancer stem cells" is used. However, some pieces of the puzzle are missing: (a) a stringent definition of cancer stem cells in solid tumors (b) specific markers that only target cells that meet the criteria for a cancer stem cell in a certain type of tumor. These missing parts started an ongoing debate about which is the best method to identify and characterize cancer stem cells, or even if their mere existence is just an artifact caused by the experimental procedures. Recent findings query the cancer stem cell hypothesis for solid tumors itself since it was shown in xenograft transplantation experiments that under appropriate conditions tumor-initiating cells are not rare. In this review we critically discuss the challenges and prospects of the currently used major methods to identify cancer stem cells. Further on, we reflect the present discussion about the existence of cancer stem cells in solid tumors as well as the amount and characteristics of tumor-initiating cells and finally provide new perspectives like the correlation of cancer stem cells and induced pluripotent cells.
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Affiliation(s)
- Yvonne Welte
- Department of Vertebrate Genomics, Max Planck Institute for Molecular Genetics, Ihnestrasse 63-73, 14195 Berlin, Germany.
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24
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Kemp K, Mallam E, Scolding N, Wilkins A. Stem cells in genetic myelin disorders. Regen Med 2010; 5:425-39. [DOI: 10.2217/rme.10.10] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/06/2023] Open
Abstract
The genetic myelin disorders are a range of diseases that manifest with severe neurological problems, often from infancy. It has been postulated for some time that stem cells might be an effective treatment for these disorders, primarily as agents to restore dysfunctional or lost myelin. Stem cells, however, may offer a wider range of therapeutic potential, for instance as vehicles to replace abnormal enzymes or genes, or to provide trophic support for residual CNS tissue. This article will review several of the more common genetic myelin disorders and currently available therapies, including bone marrow transplantation for adrenoleukodystrophy. Specific stem cell subtypes and their relevance to potential therapeutic use will be discussed and stem cell transplantation in animal model studies will also be reviewed.
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Affiliation(s)
- Kevin Kemp
- MS & Stem Cell Laboratories, Burden Centre, Frenchay Hospital, Bristol, UK
- Department of Neurology, Frenchay Hospital, Bristol, UK
| | - Elizabeth Mallam
- MS & Stem Cell Laboratories, Burden Centre, Frenchay Hospital, Bristol, UK
- Department of Neurology, Frenchay Hospital, Bristol, UK
| | - Neil Scolding
- MS & Stem Cell Laboratories, Burden Centre, Frenchay Hospital, Bristol, UK
- Department of Neurology, Frenchay Hospital, Bristol, UK
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25
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Park DH, Borlongan CV, Willing AE, Eve DJ, Cruz LE, Sanberg CD, Chung YG, Sanberg PR. Human Umbilical Cord Blood Cell Grafts for Brain Ischemia. Cell Transplant 2009; 18:985-98. [DOI: 10.3727/096368909x471279] [Citation(s) in RCA: 77] [Impact Index Per Article: 4.8] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/01/2023] Open
Abstract
Irreversible and permanent damage develop immediately adjacent to the region of reduced cerebral blood perfusion in stroke patients. Currently, the proven thrombolytic treatment for stroke, tissue plasminogen activator, is only effective when administered within 3 h after stroke. These disease characteristics should be taken under consideration in developing any therapeutic intervention designed to widen the narrow therapeutic range, especially cell-based therapy. Over the past several years, our group and others have characterized the therapeutic potential of human umbilical cord blood cells for stroke and other neurological disorders using in vitro and vivo models focusing on the cells' ability to differentiate into nonhematopoietic cells including neural lineage, as well as their ability to produce several neurotrophic factors and modulate immune and inflammatory reaction. Rather than the conventional cell replacement mechanism, we advance alternative pathways of graft-mediated brain repair involving neurotrophic effects resulting from release of various growth factors that afford cell survival, angiogenesis, and anti-inflammation. Eventually, these multiple protective and restorative effects from umbilical cord blood cell grafts may be interdependent and act in harmony in promoting therapeutic benefits for stroke.
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Affiliation(s)
- Dong-Hyuk Park
- Center of Excellence for Aging & Brain Repair, Department of Neurosurgery & Brain Repair, University of South Florida College of Medicine, Tampa, FL, USA
- Department of Neurosurgery, Korea University Medical Center, Korea University College of Medicine, Seoul, Korea
| | - Cesar V. Borlongan
- Center of Excellence for Aging & Brain Repair, Department of Neurosurgery & Brain Repair, University of South Florida College of Medicine, Tampa, FL, USA
| | - Alison E. Willing
- Center of Excellence for Aging & Brain Repair, Department of Neurosurgery & Brain Repair, University of South Florida College of Medicine, Tampa, FL, USA
| | - David J. Eve
- Center of Excellence for Aging & Brain Repair, Department of Neurosurgery & Brain Repair, University of South Florida College of Medicine, Tampa, FL, USA
| | - L. Eduardo Cruz
- Cryopraxis and Silvestre Laboratory, Cryopraxis, BioRio, Pólo de Biotechnologia do Rio de Janeiro, Rio di Janiero, Brazil
| | | | - Yong-Gu Chung
- Cryopraxis and Silvestre Laboratory, Cryopraxis, BioRio, Pólo de Biotechnologia do Rio de Janeiro, Rio di Janiero, Brazil
| | - Paul R. Sanberg
- Center of Excellence for Aging & Brain Repair, Department of Neurosurgery & Brain Repair, University of South Florida College of Medicine, Tampa, FL, USA
- Office of Research and Innovation, University of South Florida, Tampa, FL, USA
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26
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Corbeil D, Joester A, Fargeas CA, Jászai J, Garwood J, Hellwig A, Werner HB, Huttner WB. Expression of distinct splice variants of the stem cell marker prominin-1 (CD133) in glial cells. Glia 2009; 57:860-74. [DOI: 10.1002/glia.20812] [Citation(s) in RCA: 45] [Impact Index Per Article: 2.8] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/31/2022]
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Tillman BW, Yazdani SK, Geary RL, Corriere MA, Atala A, Yoo JJ. Efficient Recovery of Endothelial Progenitors for Clinical Translation. Tissue Eng Part C Methods 2009; 15:213-21. [DOI: 10.1089/ten.tec.2008.0416] [Citation(s) in RCA: 14] [Impact Index Per Article: 0.9] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/12/2022] Open
Affiliation(s)
- Bryan W. Tillman
- Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina
- Department of Vascular and Endovascular Surgery, Wake Forest University School of Medicine, Winston-Salem, North Carolina
| | - Saami K. Yazdani
- Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina
- Department of Biomedical Engineering, Wake Forest University Medical Center, Wake Forest University School of Medicine, Winston-Salem, North Carolina
| | - Randolph L. Geary
- Department of Vascular and Endovascular Surgery, Wake Forest University School of Medicine, Winston-Salem, North Carolina
| | - Matthew A. Corriere
- Department of Vascular and Endovascular Surgery, Wake Forest University School of Medicine, Winston-Salem, North Carolina
| | - Anthony Atala
- Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina
| | - James J. Yoo
- Wake Forest Institute for Regenerative Medicine, Wake Forest University School of Medicine, Winston-Salem, North Carolina
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Liang M, Pariente N, Morizono K, Chen ISY. Targeted transduction of CD34+ hematopoietic progenitor cells in nonpurified human mobilized peripheral blood mononuclear cells. J Gene Med 2009; 11:185-96. [PMID: 19152374 DOI: 10.1002/jgm.1290] [Citation(s) in RCA: 17] [Impact Index Per Article: 1.1] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/13/2023] Open
Abstract
BACKGROUND Conventional gene-therapy applications of hematopoietic stem cells (HSCs) involve purification of CD34+ progenitor cells from the mobilized peripheral blood, ex vivo transduction of the gene of interest into them, and reinfusion of the transduced CD34+ progenitor cells into patients. Eliminating the process of purification would save labor, time and money, while enhancing HSCs viability, transplantability and pluripotency. Lentiviral vectors have been widely used in gene therapy because they infect both dividing and nondividing cells and provide sustained transgene expression. One of the exceptions to this rule is quiescent primary lymphocytes, in which reverse transcription of viral DNA is not completed. METHODS In the present study, we tested the possibility of targeting CD34+ progenitor cells within nonpurified human mobilized peripheral blood mononuclear cells (mPBMCs) utilizing vesicular stomatitis virus G (VSV-G) pseudotyped lentiviral vectors, based on the assumption that the CD34+ progenitor cells would be preferentially transduced. To further enhance the specificity of vector transduction, we also examined utilizing a modified Sindbis virus envelope (2.2) pseudotyped lentiviral vector, developed in our laboratory, that allows targeted transduction to specific cell receptors via antibody recognition. RESULTS Both the VSV-G and 2.2 pseudotyped vectors achieved measurable results when they were used to target CD34+ progenitor cells in nonpurified mPBMCs. CONCLUSIONS Overall, the data obtained demonstrate the potential of ex vivo targeting of CD34+ progenitor cells without purification.
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Affiliation(s)
- Min Liang
- Department of Microbiology, Immunology and Molecular Genetics, University of California, Los Angeles, CA, USA
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29
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Kurth I, Franke K, Pompe T, Bornhäuser M, Werner C. Hematopoietic stem and progenitor cells in adhesive microcavities. Integr Biol (Camb) 2009; 1:427-34. [PMID: 20023748 DOI: 10.1039/b903711j] [Citation(s) in RCA: 55] [Impact Index Per Article: 3.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/21/2022]
Abstract
The homeostasis of hematopoietic stem and progenitor cells (HSC) in the bone marrow is regulated by a complex interplay of exogenous signals, including extracellular matrix (ECM) molecules, cell-cell contacts, and cytokines. To investigate the influence of spatial restriction and adhesive interactions on HSC fate decisions, we prepared a set of fibronectin-coated micrometer-sized cavities. Analysis of human CD133+ HSCs isolated after culture on these surfaces revealed that proliferation and differentiation is decreased when HSCs are supported by substrates with small microcavities. Single cell analysis of adherent cells also revealed decreased DNA synthesis and higher levels of HSC marker expression inside the smaller cavities. Increasing the cytokine concentration highlighted the tight balance of adhesion related signals and soluble cues acting on HSC fate decisions. Our results suggest that confining human HSCs in ECM-coated microcavities is a possible method to maintain these cells in a quiescent and immature state, an important advantage for several HSC applications.
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Affiliation(s)
- Ina Kurth
- Leibniz Institute of Polymer Research, Max Bergmann Center of Biomaterials, Dresden, Germany
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30
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Jaime-Pérez JC, Hernández-Alcántara AE, Méndez-Ramírez N, Vázquez-Garza E, Cantú-Rodríguez OG, Gómez-Almaguer D. Mobilization kinetics of CD133+ hematoprogenitor cells for hematopoietic grafting. Transfusion 2009; 49:532-5. [DOI: 10.1111/j.1537-2995.2008.01979.x] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/28/2022]
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31
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Nichols JE, Cortiella J, Lee J, Niles JA, Cuddihy M, Wang S, Bielitzki J, Cantu A, Mlcak R, Valdivia E, Yancy R, McClure ML, Kotov NA. In vitro analog of human bone marrow from 3D scaffolds with biomimetic inverted colloidal crystal geometry. Biomaterials 2008; 30:1071-9. [PMID: 19042018 DOI: 10.1016/j.biomaterials.2008.10.041] [Citation(s) in RCA: 109] [Impact Index Per Article: 6.4] [Reference Citation Analysis] [Abstract] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 08/30/2008] [Accepted: 10/22/2008] [Indexed: 11/28/2022]
Abstract
In vitro replicas of bone marrow can potentially provide a continuous source of blood cells for transplantation and serve as a laboratory model to examine human immune system dysfunctions and drug toxicology. Here we report the development of an in vitro artificial bone marrow based on a 3D scaffold with inverted colloidal crystal (ICC) geometry mimicking the structural topology of actual bone marrow matrix. To facilitate adhesion of cells, scaffolds were coated with a layer of transparent nanocomposite. After seeding with hematopoietic stem cells (HSCs), ICC scaffolds were capable of supporting expansion of CD34+ HSCs with B-lymphocyte differentiation. Three-dimensional organization was shown to be critical for production of B cells and antigen-specific antibodies. Functionality of bone marrow constructs was confirmed by implantation of matrices containing human CD34+ cells onto the backs of severe combined immunodeficiency (SCID) mice with subsequent generation of human immune cells.
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Affiliation(s)
- Joan E Nichols
- Department of Internal Medicine, Division of Infectious Diseases, University of Texas Medical Branch, Galveston, TX 77555, USA.
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32
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Ladewig J, Koch P, Endl E, Meiners B, Opitz T, Couillard-Despres S, Aigner L, Brüstle O. Lineage selection of functional and cryopreservable human embryonic stem cell-derived neurons. Stem Cells 2008; 26:1705-12. [PMID: 18420830 DOI: 10.1634/stemcells.2008-0007] [Citation(s) in RCA: 35] [Impact Index Per Article: 2.1] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 01/31/2023]
Abstract
A major prerequisite for the biomedical application of human embryonic stem cells (hESC) is the derivation of defined and homogeneous somatic cell types. Here we present a human doublecortin (DCX) promoter-based lineage-selection strategy for the generation of purified hESC-derived immature neurons. After transfection of hESC-derived neural precursors with a DCX-enhanced green fluorescent protein construct, fluorescence-activated cell sorting enables the enrichment of immature human neurons at purities of up to 95%. Selected neurons undergo functional maturation and are able to establish synaptic connections. Considering that the applicability of purified hESC-derived neurons would largely benefit from an efficient cryopreservation technique, we set out to devise defined freezing conditions involving caspase inhibition, which yield post-thaw recovery rates of up to 83%. Combined with our lineage-selection procedure this cryopreservation technique enables the generation of human neurons in a ready-to-use format for a large variety of biomedical applications.
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Affiliation(s)
- Julia Ladewig
- Institute of Reconstructive Neurobiology, Life & Brain Center, University of Bonn, Sigmund-Freud-Strasse 25, D-53105 Bonn, Germany
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33
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Ceder JA, Jansson L, Ehrnström RA, Rönnstrand L, Abrahamsson PA. The characterization of epithelial and stromal subsets of candidate stem/progenitor cells in the human adult prostate. Eur Urol 2007; 53:524-31. [PMID: 18053634 DOI: 10.1016/j.eururo.2007.11.028] [Citation(s) in RCA: 22] [Impact Index Per Article: 1.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 09/11/2007] [Accepted: 11/09/2007] [Indexed: 01/16/2023]
Abstract
OBJECTIVES Questions regarding the cell source and mechanisms in the initiation and progression of prostate cancer are today still open for debate. Indeed, our knowledge regarding prostate cell regulation, self-renewal, and cytodifferentiation is presently rather limited. In this study, we investigated these processes in the normal adult human prostate. METHODS Dynamic expression patterns in prostate stem/progenitor cells, intermediate/transit-amplifying cells, and cell lineages were immunohistochemically identified in an in situ explant renewal model of the human normal/benign adult prostate (n=6). RESULTS Cells with a basal phenotype proliferated significantly in explant cultures, whereas luminal cells went into apoptosis. Results further show down-regulation in tissue cultures of the basal and hypothetical stem cell marker Bcl-2 in the majority of cells, except in rare putative epithelial stem cells. Investigation of established (AC133) and novel candidate prostate stem/progenitor markers, including the cell surface receptor tyrosine kinase KIT and its ligand stem cell factor (SCF), showed that these rare epithelial cells are AC133(+)/CD133(low)/Bcl-2(high)/cytokeratin(+)/vimentin(-)/KIT(low)/SCF(low). In addition, we report on a stromal population that expresses the mesenchymal marker vimentin and that is AC133(-)/CD133(high)/Bcl-2(-)/cytokeratin(-)/KIT(high)/SCF(high). CONCLUSIONS We provide evidence for epithelial renewal in response to tissue culture and for basal and epithelial stem/progenitor cell recruitment leading to an expansion of an intermediate luminal precursor phenotype. Data further suggest that SCF regulates prostate epithelial stem/progenitor cells in an autocrine manner and that all or a subset of the identified novel stromal phenotype represents prostate stromal progenitor cells or interstitial pacemaker cells or both.
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Affiliation(s)
- Jens A Ceder
- Lund University, Department of Clinical Sciences, Division of Urological Research, University Hospital MAS, S-205 02 Malmö, Sweden.
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Haddad R, Pflumio F, Vigon I, Visentin G, Auvray C, Fichelson S, Amsellem S. The HOXB4 homeoprotein differentially promotes ex vivo expansion of early human lymphoid progenitors. Stem Cells 2007; 26:312-22. [PMID: 17962697 DOI: 10.1634/stemcells.2007-0721] [Citation(s) in RCA: 13] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
The HOXB4 homeoprotein is known to promote the expansion of mouse and human hematopoietic stem cells (HSCs) and progenitors of the myeloid lineages. However, the putative involvement of HOXB4 in lymphopoiesis and particularly in the expansion of early lymphoid progenitor cells has remained elusive. Based on the ability of the HOXB4 protein to passively enter hematopoietic cells, our group previously designed a long-term culture procedure of human HSCs that allows ex vivo expansion of these cells. Here, this method has been further used to investigate whether HOXB4 could cause similar expansion on cells originating from CD34(+) hematopoietic progenitor cells (HPCs) committed at various levels toward the lymphoid lineages. We provide evidence that HOXB4 protein delivery promotes the expansion of primitive HPCs that generate lymphoid progenitors. Moreover, HOXB4 acts on lymphomyeloid HPCs and committed T/natural killer HPCs but not on primary B-cell progenitors. Our results clarify the effect of HOXB4 in the early stages of human lymphopoiesis, emphasizing the contribution of this homeoprotein in the maintenance of the intrinsic lymphomyeloid differentiation potential of defined HPC subsets. Finally, this study supports the potential use of HOXB4 protein for HSC and HPC expansion in a therapeutic setting and furthers our understanding of the mechanisms of the molecular regulation of hematopoiesis.
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Affiliation(s)
- Rima Haddad
- Institut Cochin, Département d'Hématologie, Paris, France
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35
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Bar EE, Chaudhry A, Lin A, Fan X, Schreck K, Matsui W, Piccirillo S, Vescovi AL, Dimeco F, Olivi A, Eberhart CG. Cyclopamine-mediated hedgehog pathway inhibition depletes stem-like cancer cells in glioblastoma. Stem Cells 2007; 25:2524-33. [PMID: 17628016 PMCID: PMC2610257 DOI: 10.1634/stemcells.2007-0166] [Citation(s) in RCA: 446] [Impact Index Per Article: 24.8] [Reference Citation Analysis] [Abstract] [Key Words] [MESH Headings] [Grants] [Track Full Text] [Download PDF] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Brain tumors can arise following deregulation of signaling pathways normally activated during brain development and may derive from neural stem cells. Given the requirement for Hedgehog in non-neoplastic stem cells, we investigated whether Hedgehog blockade could target the stem-like population in glioblastoma multiforme (GBM). We found that Gli1, a key Hedgehog pathway target, was highly expressed in 5 of 19 primary GBM and in 4 of 7 GBM cell lines. Shh ligand was expressed in some primary tumors, and in GBM-derived neurospheres, suggesting a potential mechanism for pathway activation. Hedgehog pathway blockade by cyclopamine caused a 40%-60% reduction in growth of adherent glioma lines highly expressing Gli1 but not in those lacking evidence of pathway activity. When GBM-derived neurospheres were treated with cyclopamine and then dissociated and seeded in media lacking the inhibitor, no new neurospheres formed, suggesting that the clonogenic cancer stem cells had been depleted. Consistent with this hypothesis, the stem-like fraction in gliomas marked by both aldehyde dehydrogenase activity and Hoechst dye excretion (side population) was significantly reduced or eliminated by cyclopamine. In contrast, we found that radiation treatment of our GBM neurospheres increased the percentage of these stem-like cells, suggesting that this standard therapy preferentially targets better-differentiated neoplastic cells. Most importantly, viable GBM cells injected intracranially following Hedgehog blockade were no longer able to form tumors in athymic mice, indicating that a cancer stem cell population critical for ongoing growth had been removed. Disclosure of potential conflicts of interest is found at the end of this article.
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Affiliation(s)
- Eli E. Bar
- Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Aneeka Chaudhry
- Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Alex Lin
- Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Xing Fan
- Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Karisa Schreck
- Department of Neurosurgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - William Matsui
- Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Sara Piccirillo
- Department of Biotechnology and Biosciences, University of Milan Bicocca, Milan, Italy
| | - Angelo L. Vescovi
- Department of Biotechnology and Biosciences, University of Milan Bicocca, Milan, Italy
| | | | - Alessandro Olivi
- Department of Neurosurgery, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
- Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
| | - Charles G. Eberhart
- Department of Pathology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
- Department of Oncology, Johns Hopkins University School of Medicine, Baltimore, Maryland, USA
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36
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Morgan R, Pirard PM, Shears L, Sohal J, Pettengell R, Pandha HS. Antagonism of HOX/PBX dimer formation blocks the in vivo proliferation of melanoma. Cancer Res 2007; 67:5806-13. [PMID: 17575148 DOI: 10.1158/0008-5472.can-06-4231] [Citation(s) in RCA: 84] [Impact Index Per Article: 4.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/16/2022]
Abstract
Malignant melanoma is a cancer that arises from melanocyte cells in a complex but well-studied process, and which can only be successfully treated prior to metastasis as it is highly resistant to conventional therapies. A number of recent reports have indicated that members of the HOX family of homeodomain-containing transcription factors are deregulated in melanoma, and may actually be required to maintain proliferation. In this report, we describe the use of a novel, cell-permeable antagonist of the interaction between HOX proteins and PBX, a second homeodomain-containing transcription factor that modifies HOX activity. This antagonist can block the growth of murine B16 cells and trigger apoptosis both in vitro and in vivo when administered to mice with flank tumors.
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Affiliation(s)
- Richard Morgan
- Postgraduate Medical School, University of Surrey, Guildford, United Kingdom.
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Piccoli C, D'Aprile A, Ripoli M, Scrima R, Boffoli D, Tabilio A, Capitanio N. The hypoxia-inducible factor is stabilized in circulating hematopoietic stem cells under normoxic conditions. FEBS Lett 2007; 581:3111-9. [PMID: 17568584 DOI: 10.1016/j.febslet.2007.05.077] [Citation(s) in RCA: 47] [Impact Index Per Article: 2.6] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 12/19/2006] [Revised: 05/22/2007] [Accepted: 05/28/2007] [Indexed: 11/15/2022]
Abstract
The hypoxia-inducible factor (HIF) transcriptional system enables cell adaptation to limited O(2) availability, transducing this signal into patho-physiological responses such as angiogenesis, erythropoiesis, vasomotor control, and altered energy metabolism, as well as cell survival decisions. However, other factors beyond hypoxia are known to activate this pleiotropic transcription factor. The aim of this study was to characterize HIF in human hematopoietic stem cells (HSCs) and evidence is provided that granulocyte colony stimulating factor-mobilized CD34+- and CD133+-HSCs express a stabilized cytoplasmic form of HIF-1alpha under normoxic conditions. It is shown that HIF-1alpha stabilization correlates with down-regulation of the tumour suppressor von Hippel-Lindau protein (pVHL) and is positively controlled by NADPH-oxidase-dependent production of reactive oxygen species, indicating a specific O(2)-independent post-transcriptional control of HIF in mobilized HSCs. This novel finding is discussed in the context of the proposed role of HIF as a mediator of progenitor cell recruitment to injured ischemic tissues and/or in the control of the maintenance of the undifferentiated state.
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Affiliation(s)
- Claudia Piccoli
- Department of Biomedical Science, University of Foggia, Foggia, Italy
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38
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Husain SM, Shou Y, Sorrentino BP, Handgretinger R. Isolation, molecular cloning and in vitro expression of rhesus monkey (Macaca mulatta) prominin-1.s1 complementary DNA encoding a potential hematopoietic stem cell antigen. ACTA ACUST UNITED AC 2006; 68:317-24. [PMID: 17026467 DOI: 10.1111/j.1399-0039.2006.00679.x] [Citation(s) in RCA: 4] [Impact Index Per Article: 0.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/29/2022]
Abstract
Human prominin-1 (CD133 or AC133) is an important cell surface marker used to isolate primitive hematopoietic stem cells. The commercially available antibody to human prominin-1 does not recognize rhesus prominin-1. Therefore, we isolated, cloned and characterized the complementary DNA (cDNA) of rhesus prominin-1 gene and determined its coding potential. Following the nomenclature of prominin family of genes, we named this cDNA as rhesus prominin-1.s1. The amino acid sequence data of the putative rhesus prominin-1.s1 could be used in designing antigenic peptides to raise antibodies for use in isolation of pure populations of rhesus prominin-1(+) hematopoietic cells. To the best of our knowledge, there has been no previously published report about the isolation of a prominin-1 cDNA from rhesus monkey (Macaca mulatta).
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Affiliation(s)
- S M Husain
- Division of Stem Cell Transplantation, Department of Hematology-Oncology, St. Jude Children's Research Hospital, Memphis, TN 38105, USA.
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Freund D, Oswald J, Feldmann S, Ehninger G, Corbeil D, Bornhäuser M. Comparative analysis of proliferative potential and clonogenicity of MACS-immunomagnetic isolated CD34+ and CD133+ blood stem cells derived from a single donor. Cell Prolif 2006; 39:325-32. [PMID: 16872366 PMCID: PMC6496560 DOI: 10.1111/j.1365-2184.2006.00386.x] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/30/2022] Open
Abstract
A novel stem cell marker prominin-1 (CD133) has been shown to be expressed on a subpopulation of CD34(+) haematopoietic stem and progenitor cells. The aim of this study was to compare in parallel commercially available CD34(+) and CD133(+) isolation methods based on paramagnetic bead-coupled antibodies using clinical-grade samples of mobilized peripheral blood from 10 individual healthy donors under identical conditions. The CD133 negative fraction from the first selection was used for CD34(+) enrichment to obtain an additional CD34(+)/CD133(-) population. Although no significant difference in total cell expansion between cells isolated from the three procedures was observed in a 7-day cytokine-driven suspension culture, the long-term culture-initiating cell assay demonstrated that cells derived by CD34(+) isolation contain less primitive progenitors than those isolated based on CD133(+) selection. Interestingly, CD34(+)-enriched progenitors, especially the CD34(+)/CD133(-) fraction, contained a significantly higher proportion of erythroid colony-forming cells, whereas the highest content of myeloid colony-forming cells was concentrated in the CD133(+) selected cells. These subtle differences between CD34(+) and CD133(+) immunomagnetic selection will have to be explored for their potential clinical relevance.
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Affiliation(s)
- D Freund
- Medical Clinic and Polyclinic I, University Hospital, Dreseden, Germany
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40
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Fargeas CA, Fonseca AV, Huttner WB, Corbeil D. Prominin-1 (CD133): from progenitor cells to human diseases. ACTA ACUST UNITED AC 2006. [DOI: 10.2217/17460875.1.2.213] [Citation(s) in RCA: 61] [Impact Index Per Article: 3.2] [Reference Citation Analysis] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/08/2023]
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Ninos JM, Jefferies LC, Cogle CR, Kerr WG. The thrombopoietin receptor, c-Mpl, is a selective surface marker for human hematopoietic stem cells. J Transl Med 2006; 4:9. [PMID: 16480521 PMCID: PMC1402332 DOI: 10.1186/1479-5876-4-9] [Citation(s) in RCA: 34] [Impact Index Per Article: 1.8] [Reference Citation Analysis] [Abstract] [Grants] [Track Full Text] [Download PDF] [Figures] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/20/2005] [Accepted: 02/16/2006] [Indexed: 11/10/2022] Open
Abstract
Background Thrombopoietin (TPO), the primary cytokine regulating megakaryocyte proliferation and differentiation, exerts significant influence on other hematopoietic lineages as well, including erythroid, granulocytic and lymphoid lineages. We previously demonstrated that the receptor for TPO, c-mpl, is expressed by a subset of human adult bone marrow hematopoietic stem/progenitor cells (HSC/PC) that are enriched for long-term multilineage repopulating ability in the SCID-hu Bone in vivo model of human hematopoiesis. Methods Here, we employ flow cytometry and an anti-c-mpl monoclonal antibody to comprehensively define the surface expression pattern of c-mpl in four differentiation stages of human CD34+ HSC/PC (I: CD34+38--, II: CD34+38dim, III: CD34+38+, IV: CD34dim38+) for the major sources of human HSC: fetal liver (FL), umbilical cord blood (UCB), adult bone marrow (ABM), and cytokine-mobilized peripheral blood stem cells (mPBSC). We use a surrogate in vivo model of human thymopoiesis, SCID-hu Thy/Liv, to compare the capacity of c-mpl+ vs. c-mpl-- CD34+38--/dim HSC/PC for thymocyte reconstitution. Results For all tissue sources, the percentage of c-mpl+ cells was significantly highest in stage I HSC/PC (FL 72 ± 10%, UCB 67 ± 19%, ABM 82 ± 16%, mPBSC 71 ± 15%), and decreased significantly through stages II, III, and IV ((FL 3 ± 3%, UCB 8 ± 13%, ABM 0.6 ± 0.6%, mPBSC 0.2 ± 0.1%) [ANOVA: P < 0.0001]. The relative median fluorescence intensity of c-mpl expression was similarly highest in stage I, decreasing through stage IV [ANOVA: P < 0.0001]. No significant differences between tissue sources were observed for either % c-mpl+ cells [P = 0.89] or intensity of c-mpl expression [P = 0.21]. Primary Thy/Liv grafts injected with CD34+38--/dimc-mpl+ cells showed slightly higher levels of donor HLA+ thymocyte reconstitution vs. CD34+38--/dimc-mpl---injected grafts and non-injected controls (c-mpl+ vs. c-mpl--: CD2+ 6.8 ± 4.5% vs. 2.8 ± 3.3%, CD4+8-- 54 ± 35% vs. 31 ± 29%, CD4--8+ 29 ± 19% vs. 18 ± 14%). Conclusion These findings support the hypothesis that the TPO receptor, c-mpl, participates in the regulation of primitive human HSC from mid-fetal through adult life. This study extends our previous work documenting human B-lineage, myeloid and CD34+ cell repopulation by c-mpl+ progenitors to show that c-mpl+ HSC/PC are also capable of significant T-lineage reconstitution in vivo. These results suggest that c-mpl merits consideration as a selective surface marker for the identification and isolation of human HSC in both basic research and clinical settings.
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Affiliation(s)
- John M Ninos
- H. Lee Moffitt Cancer Center and Research Institute, Immunology Program, Department of Interdisciplinary Oncology, University of South Florida, SRB-2, 12902 Magnolia Drive, Tampa, FL 33612-9416, USA
| | - Leigh C Jefferies
- AstraZeneca LP, Drug Safety US, FOC NW2-263, Wilmington, Delaware 19850-5437, USA
| | - Christopher R Cogle
- University of Florida, Division of Hematology/Oncology, 1600 SW Archer Road, ARB R4-252, P.O. Box 100277, Gainesville, FL 32610-0277, USA
| | - William G Kerr
- H. Lee Moffitt Cancer Center and Research Institute, Immunology Program, Departments of Interdisciplinary Oncology and Biochemistry, University of South Florida, SRB-2, 12902 Magnolia Drive, Tampa, FL 33612-9416, USA
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Garbuzova-Davis S, Willing AE, Saporta S, Bickford PC, Gemma C, Chen N, Sanberg CD, Klasko SK, Borlongan CV, Sanberg PR. Novel cell therapy approaches for brain repair. PROGRESS IN BRAIN RESEARCH 2006; 157:207-22. [PMID: 17046673 DOI: 10.1016/s0079-6123(06)57014-1] [Citation(s) in RCA: 41] [Impact Index Per Article: 2.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Subscribe] [Scholar Register] [Indexed: 12/14/2022]
Abstract
Numerous reports elucidate that tissue-specific stem cells are phenotypically plastic and their differentiation pathways are not strictly delineated. Although the identity of all the epigenetic factors which may trigger stem cells to make a lineage selection are still unknown, the plasticity of adult stem cells opens new approaches for their application in the treatment of various disorders. There is increasing researcher interest in hematopoietic stem cells for treatment of not only blood-related diseases but also various unrelated disorders including neurodegenerative diseases. Human umbilical cord blood (hUCB) cells, due to their primitive nature and ability to develop into nonhematopoietic cells of various tissue lineages, including neural cells, may be useful as an alternative cell source for cell-based therapies requiring either the replacement of individual cell types and/or substitution of missing substances. Here we focus on recent findings showing the robustness of adult stem cells derived from hUCB and their potential as a source of transplant cells for the treatment of diseased or injured brains and spinal cords. Depending upon the pathological microenvironment in which the hUCB cells are introduced, neuroprotective and/or trophic effects of these cells, from release of various growth or anti-inflammatory factors to moderation of immune-inflammatory effectors, may be more likely than neural replacement. These protective effects may prove essential to maintaining restored tissue integrity over the course of various diseases or injuries.
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Affiliation(s)
- Svitlana Garbuzova-Davis
- Center of Excellence for Aging and Brain Repair, Department of Neurosurgery, College of Medicine, University of South Florida, MDC 78, 12901 Bruce B. Downs Blvd., Tampa, FL 33612, USA.
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43
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Zhang L, Yang R, Han ZC. Transplantation of umbilical cord blood-derived endothelial progenitor cells: a promising method of therapeutic revascularisation. Eur J Haematol 2006; 76:1-8. [PMID: 16343265 DOI: 10.1111/j.1600-0609.2005.00579.x] [Citation(s) in RCA: 33] [Impact Index Per Article: 1.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 02/02/2023]
Abstract
Therapeutic neovascularisation by endothelial progenitor cells (EPCs) mediated vascular regeneration is becoming a novel option for the treatment of ischaemic diseases. Recently, human umbilical cord blood (CB) has been found to contain a large number of EPCs and transplantation of CB EPCs led to a successful salvage of the ischaemic limbs through improvement in blood perfusion, indicating the feasibility of using CB cells for therapeutic revascularisation. This review will summarise recent studies in therapeutic revascularisation using CB cells and discuss the potential clinical utilisation of CB cells in ischaemic diseases.
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Affiliation(s)
- Lei Zhang
- State Key Laboratory of Experimental Hematology, National Research Center of Stem Cell Engineering and Technology, Institute of Hematology, Chinese Academy of Medical Sciences and Peking Union Medical College, Tianjin, China
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44
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Quesenberry PJ, Dooner G, Colvin G, Abedi M. Stem cell biology and the plasticity polemic. Exp Hematol 2005; 33:389-94. [PMID: 15781328 DOI: 10.1016/j.exphem.2004.11.005] [Citation(s) in RCA: 46] [Impact Index Per Article: 2.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Received: 10/21/2004] [Accepted: 11/12/2004] [Indexed: 01/02/2023]
Abstract
Characterization of a cord blood derived unrestricted somatic stem cell (USSC) with capacity to differentiate into hematopoietic and nonhematopoietic tissues in the absence of cell fusion has highlighted the great potential of stem cell plasticity. A great variety of stem cell types have been defined and even the most pure marrow stem cells are highly heterogeneous. Data suggest that stem cells may exist in a continuum with continually and reversibly changing phenotype. These cells also possess a capacity to produce lung, liver, skin, and skeletal muscle under conditions of tissue injury. Arguments raised against the significance of adult marrow to nonmarrow conversions including the importance of cell fusion appear fallacious. We are at the beginning of an exciting and burgeoning field of research with great clinical potential.
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Affiliation(s)
- Peter J Quesenberry
- Department of Research and the Adele R. Decof Cancer Center, Roger Williams Medical Center, Providence, RI 02908, USA.
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45
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Katz AJ, Tholpady A, Tholpady SS, Shang H, Ogle RC. Cell Surface and Transcriptional Characterization of Human Adipose-Derived Adherent Stromal (hADAS) Cells. Stem Cells 2005; 23:412-23. [PMID: 15749936 DOI: 10.1634/stemcells.2004-0021] [Citation(s) in RCA: 605] [Impact Index Per Article: 30.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Grants] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/17/2022]
Abstract
Adult human subcutaneous adipose tissue contains cells with intriguing multilineage developmental plasticity, much like marrow-derived mesenchymal stem cells. Putative stem or progenitor cells from fat have been given many different names in the literature, reflecting an early and evolving consensus regarding their phenotypic characterization. The study reported here used microarrays to evaluate over 170 genes relating to angiogenesis and extracellular matrix in undifferentiated, early-passage human adipose-derived adherent stromal (hADAS) cells isolated from three separate donors. The hADAS populations unanimously transcribed 66% of the screened genes, and 83% were transcribed by at least two of the three populations. The most highly transcribed genes relate to functional groupings such as cell adhesion, matrix proteins, growth factors and receptors, and proteases. The transcriptome of hADAS cells demonstrated by this work reveals many similarities to published profiles of bone marrow mesenchymal stem cells (MSCs). In addition, flow analysis of over 24 hADAS cell surface proteins (n = 7 donors) both confirms and expands on the existing literature and reveals strong intergroup correlation, despite an inconsistent nomenclature and the lack of standardized protocols for cell isolation and culture. Finally, based on flow analysis and reverse transcription polymerase chain reaction studies, our results suggest that hADAS cells do not express several proteins that are implicated as markers of "stemness" in other stem cell populations, including telomerase, CD133, and the membrane transporter ABCG2.
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Affiliation(s)
- Adam J Katz
- Department of Plastic and Reconstructive Surgery, University of Virginia, Charlottesville, VA 22908, USA.
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46
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Wu X, Rabkin-Aikawa E, Guleserian KJ, Perry TE, Masuda Y, Sutherland FWH, Schoen FJ, Mayer JE, Bischoff J. Tissue-engineered microvessels on three-dimensional biodegradable scaffolds using human endothelial progenitor cells. Am J Physiol Heart Circ Physiol 2004; 287:H480-7. [PMID: 15277191 DOI: 10.1152/ajpheart.01232.2003] [Citation(s) in RCA: 172] [Impact Index Per Article: 8.2] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Submit a Manuscript] [Subscribe] [Scholar Register] [Indexed: 11/22/2022]
Abstract
Tissue engineering may offer patients new options when replacement or repair of an organ is needed. However, most tissues will require a microvascular network to supply oxygen and nutrients. One strategy for creating a microvascular network would be promotion of vasculogenesis in situ by seeding vascular progenitor cells within the biopolymeric construct. To pursue this strategy, we isolated CD34(+)/CD133(+) endothelial progenitor cells (EPC) from human umbilical cord blood and expanded the cells ex vivo as EPC-derived endothelial cells (EC). The EPC lost expression of the stem cell marker CD133 but continued to express the endothelial markers KDR/VEGF-R2, VE-cadherin, CD31, von Willebrand factor, and E-selectin. The cells were also shown to mediate calcium-dependent adhesion of HL-60 cells, a human promyelocytic leukemia cell line, providing evidence for a proinflammatory endothelial phenotype. The EPC-derived EC maintained this endothelial phenotype when expanded in roller bottles and subsequently seeded on polyglycolic acid-poly-l-lactic acid (PGA-PLLA) scaffolds, but microvessel formation was not observed. In contrast, EPC-derived EC seeded with human smooth muscle cells formed capillary-like structures throughout the scaffold (76.5 +/- 35 microvessels/mm(2)). These results indicate that 1) EPC-derived EC can be expanded in vitro and seeded on biodegradable scaffolds with preservation of endothelial phenotype and 2) EPC-derived EC seeded with human smooth muscle cells form microvessels on porous PGA-PLLA scaffolds. These properties indicate that EPC may be well suited for creating microvascular networks within tissue-engineered constructs.
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Affiliation(s)
- Xiao Wu
- Department of Surgery, Children's Hospital, Boston, MA 02115, USA
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Sconocchia G, Fujiwara H, Rezvani K, Keyvanfar K, El Ouriaghli F, Grube M, Melenhorst J, Hensel N, Barrett AJ. G-CSF-mobilized CD34+ cells cultured in interleukin-2 and stem cell factor generate a phenotypically novel monocyte. J Leukoc Biol 2004; 76:1214-9. [PMID: 15345723 DOI: 10.1189/jlb.0504278] [Citation(s) in RCA: 17] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/24/2022] Open
Abstract
To study the early stages of development from stem cells of the CD56+ cell population [which includes natural killer (NK) cells], granulocyte-colony stimulating factor-mobilized peripheral blood CD34+ cells from healthy donors were sorted to >99% purity and cultured in the presence of stem cell factor and interleukin (IL)-2. After 3 weeks in culture, the majority of cells acquired CD33, with or without human leukocyte antigen-DR and CD14. In 20 stem cell donors tested, 8.7 +/- 8.8% of cells were CD56+. Two major CD56+ subsets were identified: CD56(bright), mainly CD33- cells (7+/-10%, n=11) with large, granular lymphocyte morphology, and CD56dim, mainly CD33+ (2.5+/-2, n=11) cells with macrophage morphology. The CD56bright population had cytoplasmic granzyme A but lacked killer inhibitory receptor, suggesting they were immature NK cells. The CD56dim, CD33+, population lacked NK markers. They may represent a minor subset of normal monocytes at a developmental stage comparable with the rare CD56+ CD33+ hybrid myeloid/NK cell leukemia. Consistent with a monocyte nature, CD56dimCD33+ proliferated and produced a variety of cytokines upon lipopolysaccharide stimulation, including IL-8, IL-6, monocyte chemoattractant protein-1, and macrophage-derived chemokine but not interferon-gamma. In a short-term cytotoxicity assay, they failed to kill but powerfully inhibited the proliferation of the NK-resistant cell line P815. The generation of CD56+ cells was negatively regulated by hyaluronic acid and IL-4, indicating that extracellular matrix may play an important role in the commitment of CD34+ cells into CD56 myeloid and lymphoid lineages.
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Affiliation(s)
- Giuseppe Sconocchia
- Hematopoietic Stem Cell Transplantation Section, National Heart Lung and Blood Institute, National Institutes of Health, 9000 Rockville Pike, Bethesda MD 20892-0001, USA.
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Kluger Y, Lian Z, Zhang X, Newburger PE, Weissman SM. A panorama of lineage-specific transcription in hematopoiesis. Bioessays 2004; 26:1276-87. [PMID: 15551261 DOI: 10.1002/bies.20144] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.8] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 12/16/2022]
Abstract
The hematopoietic system consists of more than ten differentiated cell types, all of which are derived from a single type of hematopoietic stem cell. The accessibility and interest of this system have made it a model for understanding normal and abnormal differentiation of mammalian cells. Newer techniques have generated a mass of data that requires integrative approaches for analysis and interpretation. The traditional view of the differentiation program holds that a small number of regulators are involved in each stage of cell specification. However, this may not be the case. Recent analyses have shown that almost all substantial subsets of genes, including the set of broadly expressed transcription factors, are expressed in patterns that are unique for each lineage. Further, much of this difference between lineages can be captured in two-dimensional graphs. Understanding the biologic significance, mechanisms and constraints underlying these differences is a challenge for experimentalists and computational biologists alike.
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Affiliation(s)
- Yuval Kluger
- Department of Cell Biology, New York University School of Medicine, New York, USA
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Dimitriou H, Vorgia P, Stiakaki E, Mavroudis D, Markaki EA, Koumantakis E, Kalmanti M. In vitro proliferative and differentiating characteristics of CD133(+) and CD34(+) cord blood cells in the presence of thrombopoietin (TPO) or erythropoietin (EPO). Potential implications for hematopoietic cell transplantation. Leuk Res 2003; 27:1143-51. [PMID: 12921953 DOI: 10.1016/s0145-2126(03)00107-3] [Citation(s) in RCA: 6] [Impact Index Per Article: 0.3] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/19/2022]
Abstract
We investigated the characteristics of cord blood (CD) CD133(+) and CD34(+) cells, by flow cytometry, clonogenic assays and assessment of the replating ability (area under the curve (AUC)) following 7-day liquid culture in the presence of early acting growth factors and either thrombopoietin (TPO) or erythropoietin (EPO). The CD34(+) population showed a more effective proliferation in all parameters tested and TPO proved to be more effective than EPO. On the contrary, the CD133(+) cell fraction retained and expanded more immature elements in a modest but consistent manner with either TPO or EPO. We conclude that CD133(+) and CD34(+) expanded cord blood cells could potentially be used in combination to overcome the shortcomings of cord blood transplantation in older children and adults.
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Affiliation(s)
- Helen Dimitriou
- Department of Pediatric Hematology/Oncology, University Hospital of Heraklion, University of Crete Medical School, Heraklion, 71110 Crete, Greece
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50
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Belkacémi Y, Bouchet S, Frick J, Huchet A, Pène F, Aigueperse J, Gourmelon P, Lopez M, Gorin NC. Monitoring of residual hematopoiesis after total body irradiation in humans as a model for accidental x-ray exposure: dose-effect and failure of ex vivo expansion of residual stem cells in view of autografting. Int J Radiat Oncol Biol Phys 2003; 57:500-7. [PMID: 12957263 DOI: 10.1016/s0360-3016(03)00596-0] [Citation(s) in RCA: 16] [Impact Index Per Article: 0.7] [Reference Citation Analysis] [Abstract] [MESH Headings] [Track Full Text] [Journal Information] [Subscribe] [Scholar Register] [Indexed: 11/23/2022]
Abstract
PURPOSE To evaluate the residual hematopoiesis at different levels of total body irradiation (TBI) dose in bone marrow (BM) and peripheral blood (PB), and to study the dose-effect relationship on hematopoietic immature and mature progenitors. We also investigated the possibility of expanding ex vivo the residual progenitors exposed to different dose levels of TBI. METHODS AND MATERIALS Eight patients treated for AML (n = 3) and myeloma (n = 5) were included. BM and PB samples were harvested before TBI and after doses of: <or=2 Gy, 2.1-5 Gy, and >5 Gy. Mononuclear cells (MNCs) were assayed for burst-forming unit erythroid (BFU-E), granulocyte-forming unit macrophage (CFU-GM), and long-term culture initiating cells (LTC-ICs). Ex vivo expansion: MNCs (after irradiation and controls) were suspended in long-term cultures and expanded with a combination of five cytokines. RESULTS CD34+ cells were detectable at 10 Gy. We observed a significant decrease of CFU-GM and BFU-E, respectively, to 13.5% and 8.5% of baseline values for doses <or=2 Gy and to 8.2% and 4.6% for doses ranging between 2.1 and 5 Gy. No dose effect was observed for residual MNCs. LTC-ICs were not detectable after 0.8 Gy. The expansion was not successful after 1.2 Gy. CONCLUSION This study confirms the significant decrease of human mature and immature progenitors in BM and PB immediately after low-dose TBI. In addition, the lack of expansion suggests that autografting using BM or PB residual stem cells collected and expanded in vitro in case of accidental whole body exposure may be impractical.
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Affiliation(s)
- Yazid Belkacémi
- Department of Radiation Therapy, Centre Oscar Lambret, Lille, France.
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