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  1. Induced pluripotent stem cell-derived cardiac progenitors differentiate to cardiomyocytes and form biosynthetic tissues.

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    Nicolas Christoforou

    Full Text Available The mammalian heart has little capacity to regenerate, and following injury the myocardium is replaced by non-contractile scar tissue. Consequently, increased wall stress and workload on the remaining myocardium leads to chamber dilation, dysfunction, and heart failure. Cell-based therapy with an autologous, epigenetically reprogrammed, and cardiac-committed progenitor cell source could potentially reverse this process by replacing the damaged myocardium with functional tissue. However, it is unclear whether cardiac progenitor cell-derived cardiomyocytes are capable of attaining levels of structural and functional maturity comparable to that of terminally-fated cardiomyocytes. Here, we first describe the derivation of mouse induced pluripotent stem (iPS cells, which once differentiated allow for the enrichment of Nkx2-5(+ cardiac progenitors, and the cardiomyocyte-specific expression of the red fluorescent protein. We show that the cardiac progenitors are multipotent and capable of differentiating into endothelial cells, smooth muscle cells and cardiomyocytes. Moreover, cardiac progenitor selection corresponds to cKit(+ cell enrichment, while cardiomyocyte cell-lineage commitment is concomitant with dual expression of either cKit/Flk1 or cKit/Sca-1. We proceed to show that the cardiac progenitor-derived cardiomyocytes are capable of forming electrically and mechanically coupled large-scale 2D cell cultures with mature electrophysiological properties. Finally, we examine the cell progenitors' ability to form electromechanically coherent macroscopic tissues, using a physiologically relevant 3D culture model and demonstrate that following long-term culture the cardiomyocytes align, and form robust electromechanical connections throughout the volume of the biosynthetic tissue construct. We conclude that the iPS cell-derived cardiac progenitors are a robust cell source for tissue engineering applications and a 3D culture platform for pharmacological

  2. Mesenchymal stromal cell derived endothelial progenitor treatment in patients with refractory angina

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    Friis, Tina; Haack-Sørensen, Mandana; Mathiasen, Anders B

    2011-01-01

    Abstract Aims. We evaluated the feasibility, safety and efficacy of intra-myocardial injection of autologous mesenchymal stromal cells derived endothelial progenitor cell (MSC) in patients with stable coronary artery disease (CAD) and refractory angina in this first in man trial. Methods and resu......Abstract Aims. We evaluated the feasibility, safety and efficacy of intra-myocardial injection of autologous mesenchymal stromal cells derived endothelial progenitor cell (MSC) in patients with stable coronary artery disease (CAD) and refractory angina in this first in man trial. Methods...... and results. A total of 31 patients with stable CAD, moderate to severe angina and no further revascularization options, were included. Bone marrow MSC were isolated and culture expanded for 6-8 weeks. It was feasible and safe to establish in-hospital culture expansion of autologous MSC and perform intra......-myocardial injection of MSC. After six months follow-up myocardial perfusion was unaltered, but the patients increased exercise capacity (p angina attacks (p Angina Questionnaire (SAQ) evaluations (p

  3. Production of hemizygous and homozygous embryonic stem cell-derived neural progenitor cells from the transgenic alszheimer göttingen minipis

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    Hall, Vanessa Jane; Jacobsen, J.; Gunnarsson, A.

    2011-01-01

    Production of hemizygous and homozygous embryonic stem cell-derived neural progenitor cells from the transgenic alszheimer göttingen minipis......Production of hemizygous and homozygous embryonic stem cell-derived neural progenitor cells from the transgenic alszheimer göttingen minipis...

  4. Low antigenicity of hematopoietic progenitor cells derived from human ES cells

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    Eun-Mi Kim

    2010-02-01

    Full Text Available Eun-Mi Kim1, Nicholas Zavazava1,21Department of Internal Medicine, University of Iowa and Veterans Affairs Medical Center, Iowa City, Iowa, USA; 2Immunology Graduate Program, University of Iowa, Iowa City, Iowa, USAAbstract: Human embryonic stem (hES cells are essential for improved understanding of diseases and our ability to probe new therapies for use in humans. Currently, bone marrow cells and cord blood cells are used for transplantation into patients with hematopoietic malignancies, immunodeficiencies and in some cases for the treatment of autoimmune diseases. However, due to the high immunogenicity of these hematopoietic cells, toxic regimens of drugs are required for preconditioning and prevention of rejection. Here, we investigated the efficiency of deriving hematopoietic progenitor cells (HPCs from the hES cell line H13, after co-culturing with the murine stromal cell line OP9. We show that HPCs derived from the H13 ES cells poorly express major histocompatibility complex (MHC class I and no detectable class II antigens (HLA-DR. These characteristics make hES cell-derived hematopoietic cells (HPCs ideal candidates for transplantation across MHC barriers under minimal immunosuppression.Keywords: human embryonic stem cells, H13, hematopoiesis, OP9 stromal cells, immunogenicity

  5. Transplantation of human embryonic stem cell-derived oligodendrocyte progenitors into rat spinal cord injuries does not cause harm.

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    Cloutier, Frank; Siegenthaler, Monica M; Nistor, Gabriel; Keirstead, Hans S

    2006-07-01

    Demyelination contributes to loss of function following spinal cord injury. We have shown previously that transplantation of human embryonic stem cell-derived oligodendrocyte progenitors into adult rat 200 kD contusive spinal cord injury sites enhances remyelination and promotes recovery of motor function. Previous studies using oligodendrocyte lineage cells have noted a correlation between the presence of demyelinating pathology and the survival and migration rate of the transplanted cells. The present study compared the survival and migration of human embryonic stem cell-derived oligodendrocyte progenitors injected 7 days after a 200 or 50 kD contusive spinal cord injury, as well as the locomotor outcome of transplantation. Our findings indicate that a 200 kD spinal cord injury induces extensive demyelination, whereas a 50 kD spinal cord injury induces no detectable demyelination. Cells transplanted into the 200 kD injury group survived, migrated, and resulted in robust remyelination, replicating our previous studies. In contrast, cells transplanted into the 50 kD injury group survived, exhibited limited migration, and failed to induce remyelination as demyelination in this injury group was absent. Animals that received a 50 kD injury displayed only a transient decline in locomotor function as a result of the injury. Importantly, human embryonic stem cell-derived oligodendrocyte progenitor transplants into the 50 kD injury group did not cause a further decline in locomotion. Our studies highlight the importance of a demyelinating pathology as a prerequisite for the function of transplanted myelinogenic cells. In addition, our results indicate that transplantation of human embryonic stem cell-derived oligodendrocyte progenitor cells into the injured spinal cord is not associated with a decline in locomotor function.

  6. The postischemic environment differentially impacts teratoma or tumor formation after transplantation of human embryonic stem cell-derived neural progenitors

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    Seminatore, Christine; Polentes, Jerome; Ellman, Ditte

    2010-01-01

    Risk of tumorigenesis is a major obstacle to human embryonic and induced pluripotent stem cell therapy. Likely linked to the stage of differentiation of the cells at the time of implantation, formation of teratoma/tumors can also be influenced by factors released by the host tissue. We have...... analyzed the relative effects of the stage of differentiation and the postischemic environment on the formation of adverse structures by transplanted human embryonic stem cell-derived neural progenitors....

  7. Brief report: reconstruction of joint hyaline cartilage by autologous progenitor cells derived from ear elastic cartilage.

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    Mizuno, Mitsuru; Kobayashi, Shinji; Takebe, Takanori; Kan, Hiroomi; Yabuki, Yuichiro; Matsuzaki, Takahisa; Yoshikawa, Hiroshi Y; Nakabayashi, Seiichiro; Ik, Lee Jeong; Maegawa, Jiro; Taniguchi, Hideki

    2014-03-01

    In healthy joints, hyaline cartilage covering the joint surfaces of bones provides cushioning due to its unique mechanical properties. However, because of its limited regenerative capacity, age- and sports-related injuries to this tissue may lead to degenerative arthropathies, prompting researchers to investigate a variety of cell sources. We recently succeeded in isolating human cartilage progenitor cells from ear elastic cartilage. Human cartilage progenitor cells have high chondrogenic and proliferative potential to form elastic cartilage with long-term tissue maintenance. However, it is unknown whether ear-derived cartilage progenitor cells can be used to reconstruct hyaline cartilage, which has different mechanical and histological properties from elastic cartilage. In our efforts to develop foundational technologies for joint hyaline cartilage repair and reconstruction, we conducted this study to obtain an answer to this question. We created an experimental canine model of knee joint cartilage damage, transplanted ear-derived autologous cartilage progenitor cells. The reconstructed cartilage was rich in proteoglycans and showed unique histological characteristics similar to joint hyaline cartilage. In addition, mechanical properties of the reconstructed tissues were higher than those of ear cartilage and equal to those of joint hyaline cartilage. This study suggested that joint hyaline cartilage was reconstructed from ear-derived cartilage progenitor cells. It also demonstrated that ear-derived cartilage progenitor cells, which can be harvested by a minimally invasive method, would be useful for reconstructing joint hyaline cartilage in patients with degenerative arthropathies. © AlphaMed Press.

  8. Patient-specific cardiovascular progenitor cells derived from integration-free induced pluripotent stem cells for vascular tissue regeneration.

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    Hu, Jiang; Wang, Yongyu; Jiao, Jiao; Liu, Zhongning; Zhao, Chao; Zhou, Zhou; Zhang, Zhanpeng; Forde, Kaitlynn; Wang, Lunchang; Wang, Jiangang; Baylink, David J; Zhang, Xiao-Bing; Gao, Shaorong; Yang, Bo; Chen, Y Eugene; Ma, Peter X

    2015-12-01

    Tissue-engineered blood vessels (TEBVs) are promising in regenerating a live vascular replacement. However, the vascular cell source is limited, and it is crucial to develop a scaffold that accommodates new type of vascular progenitor cells and facilitates in vivo lineage specification of the cells into functional vascular smooth muscle cells (VSMCs) to regenerate vascular tissue. In the present study, integration-free human induced pluripotent stem cells (hiPSCs) were established from patient peripheral blood mononuclear cells through episomal vector nucleofection of reprogramming factors. The established hiPSCs were then induced into mesoderm-originated cardiovascular progenitor cells (CVPCs) with a highly efficient directed lineage specification method. The derived CVPCs were demonstrated to be able to differentiate into functional VSMCs. Subcutaneous implantation of CVPCs seeded on macroporous nanofibrous poly(l-lactide) scaffolds led to in vivo VSMC lineage specification and matrix deposition inside the scaffolds. In summary, we established integration-free patient-specific hiPSCs from peripheral blood mononuclear cells, derived CVPCs through directed lineage specification, and developed an advanced scaffold for these progenitor cells to further differentiate in vivo into VSMCs and regenerate vascular tissue in a subcutaneous implantation model. This study has established an efficient patient-specific approach towards in vivo regeneration of vascular tissue. Copyright © 2015 Elsevier Ltd. All rights reserved.

  9. Plasma levels of stromal cell-derived factor-1 (CXCL12) and circulating endothelial progenitor cells in women with idiopathic heavy menstrual bleeding.

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    Elsheikh, E; Andersson, E; Sylvén, C; Ericzon, B-G; Palmblad, J; Mints, M

    2014-01-01

    Do plasma levels of stromal cell-derived factor-1 (CXCL12, sometimes termed SDF-1) and the numbers of circulating endothelial progenitor cells (EPCs), EPC colony-forming units (EPC-CFU) and mature endothelial cells (ECs) differ between women with idiopathic heavy menstrual bleeding of endometrial origin (HMB-E) and controls and are they related to plasma levels of other angiogenic growth factors? Angiogenesis is altered in women with HMB-E, characterized by a reduction in mean plasma levels of CXCL12, a low number of EPCs-CFUs and a high level of circulating ECs. Plasma levels of CXCL12 are significantly higher during the proliferative than the secretory phase of the menstrual cycle in healthy women and exhibit a negative correlation with blood EPC-CFUs. A prospective cohort study in a university hospital setting. Between 2008 and 2009 10 HMB-E patients were recruited from Karolinska University Hospital. Ten healthy women were also included in the analysis. Ten healthy control women and 10 HMB-E patients, all with regular menstrual cycles, provided 4 blood samples during a single menstrual cycle: 2 in the proliferative phase, 1 at ovulation and 1 in the secretory phase. We assessed plasma levels of CXCL12, vascular endothelial growth factor A(165) (VEGFA), basic fibroblast growth factor (bFGF) and granulocyte and granulocyte-macrophage colony-stimulating factors by ELISA. We counted circulating EPC-CFUs by culture, and ECs and EPCs by flow cytometry and immunostaining for cell surface markers. Plasma levels of CXCL12 were significantly lower in HMB-E patients compared with control women (P Market Insurance. The authors have no conflict of interest to declare.

  10. Human-induced pluripotent stem cell-derived cardiomyocytes from cardiac progenitor cells: effects of selective ion channel blockade.

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    Altomare, Claudia; Pianezzi, Enea; Cervio, Elisabetta; Bolis, Sara; Biemmi, Vanessa; Benzoni, Patrizia; Camici, Giovanni G; Moccetti, Tiziano; Barile, Lucio; Vassalli, Giuseppe

    2016-12-01

    Human-induced pluripotent stem cell (hiPSC)-derived cardiomyocytes are likely to revolutionize electrophysiological approaches to arrhythmias. Recent evidence suggests the somatic cell origin of hiPSCs may influence their differentiation potential. Owing to their cardiomyogenic potential, cardiac-stromal progenitor cells (CPCs) are an interesting cellular source for generation of hiPSC-derived cardiomyocytes. The effect of ionic current blockade in hiPSC-derived cardiomyocytes generated from CPCs has not been characterized yet. Human-induced pluripotent stem cell-derived cardiomyocytes were generated from adult CPCs and skin fibroblasts from the same individuals. The effect of selective ionic current blockade on spontaneously beating hiPSC-derived cardiomyocytes was assessed using multi-electrode arrays. Cardiac-stromal progenitor cells could be reprogrammed into hiPSCs, then differentiated into hiPSC-derived cardiomyocytes. Human-induced pluripotent stem cell-derived cardiomyocytes of cardiac origin showed higher upregulation of cardiac-specific genes compared with those of fibroblastic origin. Human-induced pluripotent stem cell-derived cardiomyocytes of both somatic cell origins exhibited sensitivity to tetrodotoxin, a blocker of Na +  current (I Na ), nifedipine, a blocker of L-type Ca 2+  current (I CaL ), and E4031, a blocker of the rapid component of delayed rectifier K +  current (I Kr ). Human-induced pluripotent stem cell-derived cardiomyocytes of cardiac origin exhibited sensitivity to JNJ303, a blocker of the slow component of delayed rectifier K +  current (I Ks ). In hiPSC-derived cardiomyocytes of cardiac origin, I Na , I CaL , I Kr , and I Ks were present as tetrodotoxin-, nifedipine-, E4031-, and JNJ303-sensitive currents, respectively. Although cardiac differentiation efficiency was improved in hiPSCs of cardiac vs. non-cardiac origin, no major functional differences were observed between hiPSC-derived cardiomyocytes of different somatic

  11. Interleukin-3 Does Not Affect the Differentiation of Mast Cells Derived from Human Bone Marrow Progenitors

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    Shimizu, Yuji; Matsumoto, Kenji; Okayama, Yoshimichi; Kentaro, Sakai; Maeno, Toshitaka; Suga, Tatsuo; Miura, Toru; Takai, Shinji; Kurabayashi, Masahiko; Saito, Hirohisa

    2008-01-01

    Although IL-3 is commonly used for culture of human progenitor-derived mast cells together with Stem cell factor (SCF) and IL-6, the effect of IL-3 on human mast cell differentiation has not been well elucidated. Human bone marrow CD34+ progenitors were cultured for up to 12 weeks in the presence of rhSCF and rhIL-6 either with rhIL-3 (IL-3 (+)) or without rhIL-3 (IL-3 (−)) for the initial 1-week of culture. Total cell number increased at 2 weeks in IL-3 (+), as compared to IL-3 (−), but changes in the appearance of mast cells were delayed. When IL-3 was present for the initial 1-week culture, granules looked more mature with IL-3 than without IL-3. However, tryptase and chymase contents, and surface antigen expression (CD18, CD51, CD54, and CD117) were not altered by IL-3. Surface expression and mRNA level of FcεRIα and histamine release by crosslinking of FcεRIα did not differ from one preparation to the next. GeneChip analysis revealed that no significant differences were observed between IL-3 (+) and IL-3 (−) cells either when inactivated or activated by aggregation of FcεRIα. These findings indicate that initial incubation of human bone marrow CD34+ progenitors with IL-3 does not affect the differentiation of mast cells. PMID:18214796

  12. Cancer cells cause vascular endothelial cell (vEC) retraction via 12(S)HETE secretion; the possible role of cancer cell derived microparticle.

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    Uchide, Keiji; Sakon, Masato; Ariyoshi, Hideo; Nakamori, Syouji; Tokunaga, Masaru; Monden, Morito

    2007-02-01

    Cancer cell mediated vascular endothelial cell (vEC) retraction plays a pivotal role in cancer metastasis. The aim of this study is to clarify the biochemical character of vEC retraction factor derived from human breast cancer cell line, MCF-7. In order to estimate vEC retracting activity, transwell chamber assay system was employed. We first tested the effects of trypsin digestion as well as lipid extraction of culture medium (CM). Trypsin digestion of CM resulted in approximately 40% loss of vEC retracting activity and lipid extraction of CM by Brigh and Dyer methods recovered approximately 60% of vEC retracting activity, suggesting that approximately 60% of vEC retracting activity in MCF-7 derived CM is due to lipid. Although Nordihydroguaiaretic acid (NDGA), the specific lipoxygenase inhibitor, suppressed vEC retracting activity in CM, Acetyl salicylic acid (ASA), a specific cyclooxygenase inhibitor, did not affect the activity, suggesting that lipid exerting vEC retracting activity in CM belongs to lipoxygenase mediated arachidonate metabolites. Thin layer chromatography clearly demonstrated that Rf value of lipid vEC retracting factor in CM is identical to 12HETE. Authentic 12(S)HETE, but not 12(R)HETE, showed vEC retracting activity. After the ultracentrifugation of CM, most lipid vEC retracting activity was recovered from the pellet fraction, and flow cytometric analysis using specific antibody against 12(S)HETE clearly showed the association of 12(S)HETE with small particle in CM. These findings suggested the principal involvement of 12(S)HETE in cancer cell derived microparticles in cancer cell mediated vEC retraction.

  13. Human embryonic stem cell derived mesenchymal progenitors express cardiac markers but do not form contractile cardiomyocytes.

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    Christophe M Raynaud

    Full Text Available Mesenchymal progenitors or stromal cells have shown promise as a therapeutic strategy for a range of diseases including heart failure. In this context, we explored the growth and differentiation potential of mesenchymal progenitors (MPs derived in vitro from human embryonic stem cells (hESCs. Similar to MPs isolated from bone marrow, hESC derived MPs (hESC-MPs efficiently differentiated into archetypical mesenchymal derivatives such as chondrocytes and adipocytes. Upon treatment with 5-Azacytidine or TGF-β1, hESC-MPs modified their morphology and up-regulated expression of key cardiac transcription factors such as NKX2-5, MEF2C, HAND2 and MYOCD. Nevertheless, NKX2-5+ hESC-MP derivatives did not form contractile cardiomyocytes, raising questions concerning the suitability of these cells as a platform for cardiomyocyte replacement therapy. Gene profiling experiments revealed that, although hESC-MP derived cells expressed a suite of cardiac related genes, they lacked the complete repertoire of genes associated with bona fide cardiomyocytes. Our results suggest that whilst agents such as TGF-β1 and 5-Azacytidine can induce expression of cardiac related genes, but treated cells retain a mesenchymal like phenotype.

  14. Intraocular Injection of ES Cell-Derived Neural Progenitors Improve Visual Function in Retinal Ganglion Cell-Depleted Mouse Models

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    Mundackal S. Divya

    2017-09-01

    Full Text Available Retinal ganglion cell (RGC transplantation is a promising strategy to restore visual function resulting from irreversible RGC degeneration occurring in glaucoma or inherited optic neuropathies. We previously demonstrated FGF2 induced differentiation of mouse embryonic stem cells (ESC to RGC lineage, capable of retinal ganglion cell layer (GCL integration upon transplantation. Here, we evaluated possible improvement of visual function by transplantation of ES cell derived neural progenitors in RGC depleted glaucoma mice models. ESC derived neural progenitors (ES-NP were transplanted into N-Methyl-D-Aspartate (NMDA injected, RGC-ablated mouse models and a pre-clinical glaucoma mouse model (DBA/2J having sustained higher intra ocular pressure (IOP. Visual acuity and functional integration was evaluated by behavioral experiments and immunohistochemistry, respectively. GFP-expressing ES-NPs transplanted in NMDA-injected RGC-depleted mice differentiated into RGC lineage and possibly integrating into GCL. An improvement in visual acuity was observed after 2 months of transplantation, when compared to the pre-transplantation values. Expression of c-Fos in the transplanted cells, upon light induction, further suggests functional integration into the host retinal circuitry. However, the transplanted cells did not send axonal projections into optic nerve. Transplantation experiments in DBA/2J mouse showed no significant improvement in visual functions, possibly due to both host and transplanted retinal cell death which could be due to an inherent high IOP. We showed that, ES NPs transplanted into the retina of RGC-ablated mouse models could survive, differentiate to RGC lineage, and possibly integrate into GCL to improve visual function. However, for the survival of transplanted cells in glaucoma, strategies to control the IOP are warranted.

  15. Successful immortalization of mesenchymal progenitor cells derived from human placenta and the differentiation abilities of immortalized cells

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    Zhang Xiaohong; Soda, Yasushi; Takahashi, Kenji; Bai, Yuansong; Mitsuru, Ayako; Igura, Koichi; Satoh, Hitoshi; Yamaguchi, Satoru; Tani, Kenzaburo; Tojo, Arinobu; Takahashi, Tsuneo A.

    2006-01-01

    We reported previously that mesenchymal progenitor cells derived from chorionic villi of the human placenta could differentiate into osteoblasts, adipocytes, and chondrocytes under proper induction conditions and that these cells should be useful for allogeneic regenerative medicine, including cartilage tissue engineering. However, similar to human mesenchymal stem cells (hMSCs), though these placental cells can be isolated easily, they are difficult to study in detail because of their limited life span in vitro. To overcome this problem, we attempted to prolong the life span of human placenta-derived mesenchymal cells (hPDMCs) by modifying hTERT and Bmi-1, and investigated whether these modified hPDMCs retained their differentiation capability and multipotency. Our results indicated that the combination of hTERT and Bmi-1 was highly efficient in prolonging the life span of hPDMCs with differentiation capability to osteogenic, adipogenic, and chondrogenic cells in vitro. Clonal cell lines with directional differentiation ability were established from the immortalized parental hPDMC/hTERT + Bmi-1. Interestingly, hPDMC/Bmi-1 showed extended proliferation after long-term growth arrest and telomerase was activated in the immortal hPDMC/Bmi-1 cells. However, the differentiation potential was lost in these cells. This study reports a method to extend the life span of hPDMCs with hTERT and Bmi-1 that should become a useful tool for the study of mesenchymal stem cells

  16. Mesenchymal Stem/Progenitor Cells Derived from Articular Cartilage, Synovial Membrane and Synovial Fluid for Cartilage Regeneration: Current Status and Future Perspectives.

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    Huang, Yi-Zhou; Xie, Hui-Qi; Silini, Antonietta; Parolini, Ornella; Zhang, Yi; Deng, Li; Huang, Yong-Can

    2017-10-01

    Large articular cartilage defects remain an immense challenge in the field of regenerative medicine because of their poor intrinsic repair capacity. Currently, the available medical interventions can relieve clinical symptoms to some extent, but fail to repair the cartilaginous injuries with authentic hyaline cartilage. There has been a surge of interest in developing cell-based therapies, focused particularly on the use of mesenchymal stem/progenitor cells with or without scaffolds. Mesenchymal stem/progenitor cells are promising graft cells for tissue regeneration, but the most suitable source of cells for cartilage repair remains controversial. The tissue origin of mesenchymal stem/progenitor cells notably influences the biological properties and therapeutic potential. It is well known that mesenchymal stem/progenitor cells derived from synovial joint tissues exhibit superior chondrogenic ability compared with those derived from non-joint tissues; thus, these cell populations are considered ideal sources for cartilage regeneration. In addition to the progress in research and promising preclinical results, many important research questions must be answered before widespread success in cartilage regeneration is achieved. This review outlines the biology of stem/progenitor cells derived from the articular cartilage, the synovial membrane, and the synovial fluid, including their tissue distribution, function and biological characteristics. Furthermore, preclinical and clinical trials focusing on their applications for cartilage regeneration are summarized, and future research perspectives are discussed.

  17. Characterization of single cell derived cultures of periosteal progenitor cells to ensure the cell quality for clinical application.

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    Stefan Stich

    Full Text Available For clinical applications of cells and tissue engineering products it is of importance to characterize the quality of the used cells in detail. Progenitor cells from the periosteum are already routinely applied in the clinics for the regeneration of the maxillary bone. Periosteal cells have, in addition to their potential to differentiate into bone, the ability to develop into cartilage and fat. However, the question arises whether all cells isolated from periosteal biopsies are able to differentiate into all three tissue types, or whether there are subpopulations. For an efficient and approved application in bone or cartilage regeneration the clarification of this question is of interest. Therefore, 83 different clonal cultures of freshly isolated human periosteal cells derived from mastoid periosteum biopsies of 4 donors were generated and growth rates calculated. Differentiation capacities of 51 clonal cultures towards the osteogenic, the chondrogenic, and the adipogenic lineage were investigated. Histological and immunochemical stainings showed that 100% of the clonal cultures differentiated towards the osteogenic lineage, while 94.1% demonstrated chondrogenesis, and 52.9% could be stimulated to adipogenesis. For osteogenesis real-time polymerase chain reaction (PCR of BGLAP and RUNX2 and for adipogenesis of FABP4 and PPARG confirmed the results. Overall, 49% of the cells exhibited a tripotent potential, 45.1% showed a bipotent potential (without adipogenic differentiation, 3.9% bipotent (without chondrogenic differentiation, and 2% possessed a unipotent osteogenic potential. In FACS analyses, no differences in the marker profile of undifferentiated clonal cultures with bi- and tripotent differentiation capacity were found. Genome-wide microarray analysis revealed 52 differentially expressed genes for clonal subpopulations with or without chondrogenic differentiation capacity, among them DCN, NEDD9, TGFBR3, and TSLP. For clinical

  18. Epigenetic regulation of cardiac progenitor cells marker c-kit by stromal cell derived factor-1α.

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    Zhongpu Chen

    Full Text Available BACKGROUND: Cardiac progenitor cells (CPCs have been proven suitable for stem cell therapy after myocardial infarction, especially c-kit(+CPCs. CPCs marker c-kit and its ligand, the stem cell factor (SCF, are linked as c-kit/SCF axis, which is associated with the functions of proliferation and differentiation. In our previous study, we found that stromal cell-derived factor-1α (SDF-1α could enhance the expression of c-kit. However, the mechanism is unknown. METHODS AND RESULTS: CPCs were isolated from adult mouse hearts, c-kit(+ and c-kit(- CPCs were separated by magnetic beads. The cells were cultured with SDF-1α and CXCR4-selective antagonist AMD3100, and c-kit expression was measured by qPCR and Western blotting. Results showed that SDF-1α could enhance c-kit expression of c-kit(+CPCs, made c-kit(-CPCs expressing c-kit, and AMD3100 could inhibit the function of SDF-1α. After the intervention of SDF-1α and AMD3100, proliferation and migration of CPCs were measured by CCK-8 and transwell assay. Results showed that SDF-1α could enhance the proliferation and migration of both c-kit(+ and c-kit(- CPCs, and AMD3100 could inhibit these functions. DNA methyltransferase (DNMT mRNA were measured by qPCR, DNMT activity was measured using the DNMT activity assay kit, and DNA methylation was analyzed using Sequenom's MassARRAY platform, after the CPCs were cultured with SDF-1α. The results showed that SDF-1α stimulation inhibited the expression of DNMT1 and DNMT3β, which are critical for the maintenance of regional DNA methylation. Global DNMT activity was also inhibited by SDF-1α. Lastly, SDF-1α treatment led to significant demethylation in both c-kit(+ and c-kit(- CPCs. CONCLUSIONS: SDF-1α combined with CXCR4 could up-regulate c-kit expression of c-kit(+CPCs and make c-kit(-CPCs expressing c-kit, which result in the CPCs proliferation and migration ability improvement, through the inhibition of DNMT1 and DNMT3β expression and global DNMT

  19. An in vitro expansion system for generation of human iPS cell-derived hepatic progenitor-like cells exhibiting a bipotent differentiation potential.

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    Ayaka Yanagida

    Full Text Available Hepatoblasts, hepatic stem/progenitor cells in liver development, have a high proliferative potential and the ability to differentiate into both hepatocytes and cholangiocytes. In regenerative medicine and drug screening for the treatment of severe liver diseases, human induced pluripotent stem (iPS cell-derived mature functional hepatocytes are considered to be a potentially good cell source. However, induction of proliferation of these cells is difficult ex vivo. To circumvent this problem, we generated hepatic progenitor-like cells from human iPS cells using serial cytokine treatments in vitro. Highly proliferative hepatic progenitor-like cells were purified by fluorescence-activated cell sorting using antibodies against CD13 and CD133 that are known cell surface markers of hepatic stem/progenitor cells in fetal and adult mouse livers. When the purified CD13(highCD133(+ cells were cultured at a low density with feeder cells in the presence of suitable growth factors and signaling inhibitors (ALK inhibitor A-83-01 and ROCK inhibitor Y-27632, individual cells gave rise to relatively large colonies. These colonies consisted of two types of cells expressing hepatocytic marker genes (hepatocyte nuclear factor 4α and α-fetoprotein and a cholangiocytic marker gene (cytokeratin 7, and continued to proliferate over long periods of time. In a spheroid formation assay, these cells were found to express genes required for mature liver function, such as cytochrome P450 enzymes, and secrete albumin. When these cells were cultured in a suitable extracellular matrix gel, they eventually formed a cholangiocytic cyst-like structure with epithelial polarity, suggesting that human iPS cell-derived hepatic progenitor-like cells have a bipotent differentiation ability. Collectively these data indicate that this novel procedure using an in vitro expansion system is useful for not only liver regeneration but also for the determination of molecular mechanisms that

  20. Lhx2 expression promotes self-renewal of a distinct multipotential hematopoietic progenitor cell in embryonic stem cell-derived embryoid bodies.

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    Lina Dahl

    Full Text Available The molecular mechanisms regulating the expansion of the hematopoietic system including hematopoietic stem cells (HSCs in the fetal liver during embryonic development are largely unknown. The LIM-homeobox gene Lhx2 is a candidate regulator of fetal hematopoiesis since it is expressed in the fetal liver and Lhx2(-/- mice die in utero due to severe anemia. Moreover, expression of Lhx2 in embryonic stem (ES cell-derived embryoid bodies (EBs can lead to the generation of HSC-like cell lines. To further define the role of this transcription factor in hematopoietic regulation, we generated ES cell lines that enabled tet-inducible expression of Lhx2. Using this approach we observed that Lhx2 expression synergises with specific signalling pathways, resulting in increased frequency of colony forming cells in developing EB cells. The increase in growth factor-responsive progenitor cells directly correlates to the efficiency in generating HSC-like cell lines, suggesting that Lhx2 expression induce self-renewal of a distinct multipotential hematopoietic progenitor cell in EBs. Signalling via the c-kit tyrosine kinase receptor and the gp130 signal transducer by IL-6 is necessary and sufficient for the Lhx2 induced self-renewal. While inducing self-renewal of multipotential progenitor cells, expression of Lhx2 inhibited proliferation of primitive erythroid precursor cells and interfered with early ES cell commitment, indicating striking lineage specificity of this effect.

  1. Culture conditions have an impact on the maturation of traceable, transplantable mouse embryonic stem cell-derived otic progenitor cells.

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    Abboud, Nesrine; Fontbonne, Arnaud; Watabe, Isabelle; Tonetto, Alain; Brezun, Jean Michel; Feron, François; Zine, Azel

    2017-09-01

    The generation of replacement inner ear hair cells (HCs) remains a challenge and stem cell therapy holds the potential for developing therapeutic solutions to hearing and balance disorders. Recent developments have made significant strides in producing mouse otic progenitors using cell culture techniques to initiate HC differentiation. However, no consensus has been reached as to efficiency and therefore current methods remain unsatisfactory. In order to address these issues, we compare the generation of otic and HC progenitors from embryonic stem (ES) cells in two cell culture systems: suspension vs. adherent conditions. In the present study, an ES cell line derived from an Atoh1-green fluorescent protein (GFP) transgenic mouse was used to track the generation of otic progenitors, initial HCs and to compare these two differentiation systems. We used a two-step short-term differentiation method involving an induction period of 5 days during which ES cells were cultured in the presence of Wnt/transforming growth factor TGF-β inhibitors and insulin-like growth factor IGF-1 to suppress mesoderm and reinforce presumptive ectoderm and otic lineages. The generated embryoid bodies were then differentiated in medium containing basic fibroblast growth factor (bFGF) for an additional 5 days using either suspension or adherent culture methods. Upon completion of differentiation, quantitative polymerase chain reaction analysis and immunostaining monitored the expression of otic/HC progenitor lineage markers. The results indicate that cells differentiated in suspension cultures produced cells expressing otic progenitor/HC markers at a higher efficiency compared with the production of these cell types within adherent cultures. Furthermore, we demonstrated that a fraction of these cells can incorporate into ototoxin-injured mouse postnatal cochlea explants and express MYO7A after transplantation. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons

  2. Mobilizing Transit-Amplifying Cell-Derived Ectopic Progenitors Prevents Hair Loss from Chemotherapy or Radiation Therapy.

    Science.gov (United States)

    Huang, Wen-Yen; Lai, Shih-Fan; Chiu, Hsien-Yi; Chang, Michael; Plikus, Maksim V; Chan, Chih-Chieh; Chen, You-Tzung; Tsao, Po-Nien; Yang, Tsung-Lin; Lee, Hsuan-Shu; Chi, Peter; Lin, Sung-Jan

    2017-11-15

    Genotoxicity-induced hair loss from chemotherapy and radiotherapy is often encountered in cancer treatment, and there is a lack of effective treatment. In growing hair follicles (HF), quiescent stem cells (SC) are maintained in the bulge region, and hair bulbs at the base contain rapidly dividing, yet genotoxicity-sensitive transit-amplifying cells (TAC) that maintain hair growth. How genotoxicity-induced HF injury is repaired remains unclear. We report here that HFs mobilize ectopic progenitors from distinct TAC compartments for regeneration in adaptation to the severity of dystrophy induced by ionizing radiation (IR). Specifically, after low-dose IR, keratin 5 + basal hair bulb progenitors, rather than bulge SCs, were quickly activated to replenish matrix cells and regenerated all concentric layers of HFs, demonstrating their plasticity. After high-dose IR, when both matrix and hair bulb cells were depleted, the surviving outer root sheath cells rapidly acquired an SC-like state and fueled HF regeneration. Their progeny then homed back to SC niche and supported new cycles of HF growth. We also revealed that IR induced HF dystrophy and hair loss and suppressed WNT signaling in a p53- and dose-dependent manner. Augmenting WNT signaling attenuated the suppressive effect of p53 and enhanced ectopic progenitor proliferation after genotoxic injury, thereby preventing both IR- and cyclophosphamide-induced alopecia. Hence, targeted activation of TAC-derived progenitor cells, rather than quiescent bulge SCs, for anagen HF repair can be a potential approach to prevent hair loss from chemotherapy and radiotherapy. Cancer Res; 77(22); 6083-96. ©2017 AACR . ©2017 American Association for Cancer Research.

  3. Integration-deficient lentivectors: an effective strategy to purify and differentiate human embryonic stem cell-derived hepatic progenitors.

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    Yang, Guanghua; Si-Tayeb, Karim; Corbineau, Sébastien; Vernet, Rémi; Gayon, Régis; Dianat, Noushin; Martinet, Clémence; Clay, Denis; Goulinet-Mainot, Sylvie; Tachdjian, Gérard; Tachdjian, Gérard; Burks, Deborah; Vallier, Ludovic; Bouillé, Pascale; Dubart-Kupperschmitt, Anne; Weber, Anne

    2013-07-19

    Human pluripotent stem cells (hPSCs) hold great promise for applications in regenerative medicine. However, the safety of cell therapy using differentiated hPSC derivatives must be improved through methods that will permit the transplantation of homogenous populations of a specific cell type. To date, purification of progenitors and mature cells generated from either embryonic or induced pluripotent stem cells remains challenging with use of conventional methods. We used lentivectors encoding green fluorescent protein (GFP) driven by the liver-specific apoliprotein A-II (APOA-II) promoter to purify human hepatic progenitors. We evaluated both integrating and integration-defective lentivectors in combination with an HIV integrase inhibitor. A human embryonic stem cell line was differentiated into hepatic progenitors using a chemically defined protocol. Subsequently, cells were transduced and sorted at day 16 of differentiation to obtain a cell population enriched in hepatic progenitor cells. After sorting, more than 99% of these APOA-II-GFP-positive cells expressed hepatoblast markers such as α-fetoprotein and cytokeratin 19. When further cultured for 16 days, these cells underwent differentiation into more mature cells and exhibited hepatocyte properties such as albumin secretion. Moreover, they were devoid of vector DNA integration. We have developed an effective strategy to purify human hepatic cells from cultures of differentiating hPSCs, producing a novel tool that could be used not only for cell therapy but also for in vitro applications such as drug screening. The present strategy should also be suitable for the purification of a broad range of cell types derived from either pluripotent or adult stem cells.

  4. Maturation and function of human embryonic stem cell-derived pancreatic progenitors in macroencapsulation devices following transplant into mice.

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    Bruin, Jennifer E; Rezania, Alireza; Xu, Jean; Narayan, Kavitha; Fox, Jessica K; O'Neil, John J; Kieffer, Timothy J

    2013-09-01

    Islet transplantation is a promising cell therapy for patients with diabetes, but it is currently limited by the reliance upon cadaveric donor tissue. We previously demonstrated that human embryonic stem cell (hESC)-derived pancreatic progenitor cells matured under the kidney capsule in a mouse model of diabetes into glucose-responsive insulin-secreting cells capable of reversing diabetes. However, the formation of cells resembling bone and cartilage was a major limitation of that study. Therefore, we developed an improved differentiation protocol that aimed to prevent the formation of off-target mesoderm tissue following transplantation. We also examined how variation within the complex host environment influenced the development of pancreatic progenitors in vivo. The hESCs were differentiated for 14 days into pancreatic progenitor cells and transplanted either under the kidney capsule or within Theracyte (TheraCyte, Laguna Hills, CA, USA) devices into diabetic mice. Our revised differentiation protocol successfully eliminated the formation of non-endodermal cell populations in 99% of transplanted mice and generated grafts containing >80% endocrine cells. Progenitor cells developed efficiently into pancreatic endocrine tissue within macroencapsulation devices, despite lacking direct contact with the host environment, and reversed diabetes within 3 months. The preparation of cell aggregates pre-transplant was critical for the formation of insulin-producing cells in vivo and endocrine cell development was accelerated within a diabetic host environment compared with healthy mice. Neither insulin nor exendin-4 therapy post-transplant affected the maturation of macroencapsulated cells. Efficient differentiation of hESC-derived pancreatic endocrine cells can occur in a macroencapsulation device, yielding glucose-responsive insulin-producing cells capable of reversing diabetes.

  5. Treating Diet-Induced Diabetes and Obesity with Human Embryonic Stem Cell-Derived Pancreatic Progenitor Cells and Antidiabetic Drugs

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    Jennifer E. Bruin

    2015-04-01

    Full Text Available Human embryonic stem cell (hESC-derived pancreatic progenitor cells effectively reverse hyperglycemia in rodent models of type 1 diabetes, but their capacity to treat type 2 diabetes has not been reported. An immunodeficient model of type 2 diabetes was generated by high-fat diet (HFD feeding in SCID-beige mice. Exposure to HFDs did not impact the maturation of macroencapsulated pancreatic progenitor cells into glucose-responsive insulin-secreting cells following transplantation, and the cell therapy improved glucose tolerance in HFD-fed transplant recipients after 24 weeks. However, since diet-induced hyperglycemia and obesity were not fully ameliorated by transplantation alone, a second cohort of HFD-fed mice was treated with pancreatic progenitor cells combined with one of three antidiabetic drugs. All combination therapies rapidly improved body weight and co-treatment with either sitagliptin or metformin improved hyperglycemia after only 12 weeks. Therefore, a stem cell-based therapy may be effective for treating type 2 diabetes, particularly in combination with antidiabetic drugs.

  6. Human Induced Pluripotent Stem Cell-Derived Cardiac Progenitor Cells in Phenotypic Screening: A Transforming Growth Factor-β Type 1 Receptor Kinase Inhibitor Induces Efficient Cardiac Differentiation.

    Science.gov (United States)

    Drowley, Lauren; Koonce, Chad; Peel, Samantha; Jonebring, Anna; Plowright, Alleyn T; Kattman, Steven J; Andersson, Henrik; Anson, Blake; Swanson, Bradley J; Wang, Qing-Dong; Brolen, Gabriella

    2016-02-01

    Several progenitor cell populations have been reported to exist in hearts that play a role in cardiac turnover and/or repair. Despite the presence of cardiac stem and progenitor cells within the myocardium, functional repair of the heart after injury is inadequate. Identification of the signaling pathways involved in the expansion and differentiation of cardiac progenitor cells (CPCs) will broaden insight into the fundamental mechanisms playing a role in cardiac homeostasis and disease and might provide strategies for in vivo regenerative therapies. To understand and exploit cardiac ontogeny for drug discovery efforts, we developed an in vitro human induced pluripotent stem cell-derived CPC model system using a highly enriched population of KDR(pos)/CKIT(neg)/NKX2.5(pos) CPCs. Using this model system, these CPCs were capable of generating highly enriched cultures of cardiomyocytes under directed differentiation conditions. In order to facilitate the identification of pathways and targets involved in proliferation and differentiation of resident CPCs, we developed phenotypic screening assays. Screening paradigms for therapeutic applications require a robust, scalable, and consistent methodology. In the present study, we have demonstrated the suitability of these cells for medium to high-throughput screens to assess both proliferation and multilineage differentiation. Using this CPC model system and a small directed compound set, we identified activin-like kinase 5 (transforming growth factor-β type 1 receptor kinase) inhibitors as novel and potent inducers of human CPC differentiation to cardiomyocytes. Significance: Cardiac disease is a leading cause of morbidity and mortality, with no treatment available that can result in functional repair. This study demonstrates how differentiation of induced pluripotent stem cells can be used to identify and isolate cell populations of interest that can translate to the adult human heart. Two separate examples of phenotypic

  7. FZD4 Marks Lateral Plate Mesoderm and Signals with NORRIN to Increase Cardiomyocyte Induction from Pluripotent Stem Cell-Derived Cardiac Progenitors

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    Charles Yoon

    2018-01-01

    Full Text Available The identification of cell surface proteins on stem cells or stem cell derivatives is a key strategy for the functional characterization, isolation, and understanding of stem cell population dynamics. Here, using an integrated mass spectrometry- and microarray-based approach, we analyzed the surface proteome and transcriptome of cardiac progenitor cells (CPCs generated from the stage-specific differentiation of mouse and human pluripotent stem cells. Through bioinformatics analysis, we have identified and characterized FZD4 as a marker for lateral plate mesoderm. Additionally, we utilized FZD4, in conjunction with FLK1 and PDGFRA, to further purify CPCs and increase cardiomyocyte (CM enrichment in both mouse and human systems. Moreover, we have shown that NORRIN presented to FZD4 further increases CM output via proliferation through the canonical WNT pathway. Taken together, these findings demonstrate a role for FZD4 in mammalian cardiac development.

  8. Comparison of direct and indirect radiation effects on osteoclast formation from progenitor cells derived from different hemopoietic sources.

    Science.gov (United States)

    Scheven, B A; Wassenaar, A M; Kawilarang-de Haas, E W; Nijweide, P J

    1987-07-01

    Hemopoietic stem and progenitor cells from different sources differ in radiosensitivity. Recently, we have demonstrated that the multinucleated cell responsible for bone resorption and marrow cavity formation, the osteoclast, is in fact of hemopoietic lineage. In this investigation we have studied the radiosensitivity of osteoclast formation from two different hemopoietic tissues: fetal liver and adult bone marrow. Development of osteoclasts from hemopoietic progenitors was induced by coculture of hemopoietic cell populations with fetal mouse long bones depleted of their own osteoclast precursor pool. During culture, osteoclasts developed from the exogenous cell population and invaded the calcified hypertrophic cartilage of the long bone model, thereby giving rise to the formation of a primitive marrow cavity. To analyze the radiosensitivity of osteoclast formation, either the hemopoietic cells or the bone rudiments were irradiated before coculture. Fetal liver cells were found to be less radiosensitive than bone marrow cells. The D0, Dq values and extrapolation numbers were 1.69 Gy, 5.30 Gy, and 24.40 for fetal liver cells and 1.01 Gy, 1.85 Gy, and 6.02 for bone marrow cells. Irradiation of the (pre)osteoclast-free long bone rudiments instead of the hemopoietic sources resulted in a significant inhibition of osteoclast formation at doses of 4 Gy or more. This indirect effect appeared to be more prominent in the cocultures with fetal than with adult hemopoietic cells. Furthermore, radiation doses of 8.0-10.0 Gy indirectly affected the appearance of other cell types (e.g., granulocytes) in the newly formed but underdeveloped marrow cavity. The results indicate that osteoclast progenitors from different hemopoietic sources exhibit a distinct sensitivity to ionizing irradiation. Radiation injury to long bone rudiments disturbs the osteoclast-forming capacity as well as the hemopoietic microenvironment.

  9. Isolation of Human Induced Pluripotent Stem Cell-Derived Dopaminergic Progenitors by Cell Sorting for Successful Transplantation

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    Daisuke Doi

    2014-03-01

    Full Text Available Human induced pluripotent stem cells (iPSCs can provide a promising source of midbrain dopaminergic (DA neurons for cell replacement therapy for Parkinson’s disease. However, iPSC-derived donor cells inevitably contain tumorigenic or inappropriate cells. Here, we show that human iPSC-derived DA progenitor cells can be efficiently isolated by cell sorting using a floor plate marker, CORIN. We induced DA neurons using scalable culture conditions on human laminin fragment, and the sorted CORIN+ cells expressed the midbrain DA progenitor markers, FOXA2 and LMX1A. When transplanted into 6-OHDA-lesioned rats, the CORIN+ cells survived and differentiated into midbrain DA neurons in vivo, resulting in significant improvement of the motor behavior, without tumor formation. In particular, the CORIN+ cells in a NURR1+ cell-dominant stage exhibited the best survival and function as DA neurons. Our method is a favorable strategy in terms of scalability, safety, and efficiency and may be advantageous for clinical application.

  10. Transcriptional profiling of MEF2-regulated genes in human neural progenitor cells derived from embryonic stem cells

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    Shing Fai Chan

    2015-03-01

    Full Text Available The myocyte enhancer factor 2 (MEF2 family of transcription factors is highly expressed in the brain and constitutes a key determinant of neuronal survival, differentiation, and synaptic plasticity. However, genome-wide transcriptional profiling of MEF2-regulated genes has not yet been fully elucidated, particularly at the neural stem cell stage. Here we report the results of microarray analysis comparing mRNAs isolated from human neural progenitor/stem cells (hNPCs derived from embryonic stem cells expressing a control vector versus progenitors expressing a constitutively-active form of MEF2 (MEF2CA, which increases MEF2 activity. Microarray experiments were performed using the Illumina Human HT-12 V4.0 expression beadchip (GEO#: GSE57184. By comparing vector-control cells to MEF2CA cells, microarray analysis identified 1880 unique genes that were differentially expressed. Among these genes, 1121 genes were up-regulated and 759 genes were down-regulated. Our results provide a valuable resource for identifying transcriptional targets of MEF2 in hNPCs.

  11. Human embryonic stem cell derived islet progenitors mature inside an encapsulation device without evidence of increased biomass or cell escape.

    Science.gov (United States)

    Kirk, Kaitlyn; Hao, Ergeng; Lahmy, Reyhaneh; Itkin-Ansari, Pamela

    2014-05-01

    There are several challenges to successful implementation of a cell therapy for insulin dependent diabetes derived from human embryonic stem cells (hESC). Among these are development of functional insulin producing cells, a clinical delivery method that eliminates the need for chronic immunosuppression, and assurance that hESC derived tumors do not form in the patient. We and others have shown that encapsulation of cells in a bilaminar device (TheraCyte) provides immunoprotection in rodents and primates. Here we monitored human insulin secretion and employed bioluminescent imaging (BLI) to evaluate the maturation, growth, and containment of encapsulated islet progenitors derived from CyT49 hESC, transplanted into mice. Human insulin was detectable by 7 weeks post-transplant and increased 17-fold over the course of 8 weeks, yet during this period the biomass of encapsulated cells remained constant. Remarkably, by 20 weeks post-transplant encapsulated cells secreted sufficient levels of human insulin to ameliorate alloxan induced diabetes. Further, bioluminescent imaging revealed for the first time that hESCs remained fully contained in encapsulation devices for up to 150 days, the longest period tested. Collectively, the data suggest that encapsulated hESC derived islet progenitors hold great promise as an effective and safe cell replacement therapy for insulin dependent diabetes. Copyright © 2014. Published by Elsevier B.V.

  12. F4/80+ Host Macrophages Are a Barrier to Murine Embryonic Stem Cell-Derived Hematopoietic Progenitor Engraftment In Vivo.

    Science.gov (United States)

    Thompson, Heather L; van Rooijen, Nico; McLelland, Bryce T; Manilay, Jennifer O

    2016-01-01

    Understanding how embryonic stem cells and their derivatives interact with the adult host immune system is critical to developing their therapeutic potential. Murine embryonic stem cell-derived hematopoietic progenitors (ESHPs) were generated via coculture with the bone marrow stromal cell line, OP9, and then transplanted into NOD.SCID.Common Gamma Chain (NSG) knockout mice, which lack B, T, and natural killer cells. Compared to control mice transplanted with adult lineage-negative bone marrow (Lin - BM) progenitors, ESHP-transplanted mice attained a low but significant level of donor hematopoietic chimerism. Based on our previous studies, we hypothesized that macrophages might contribute to the low engraftment of ESHPs in vivo . Enlarged spleens were observed in ESHP-transplanted mice and found to contain higher numbers of host F4/80 + macrophages compared to BM-transplanted controls. In vivo depletion of host macrophages using clodronate-loaded liposomes improved the ESHP-derived hematopoietic chimerism in the spleen but not in the BM. F4/80 + macrophages demonstrated a striking propensity to phagocytose ESHP targets in vitro . Taken together, these results suggest that macrophages are a barrier to both syngeneic and allogeneic ESHP engraftment in vivo .

  13. A stromal cell-derived factor-1 releasing matrix enhances the progenitor cell response and blood vessel growth in ischaemic skeletal muscle

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    D Kuraitis

    2011-09-01

    Full Text Available Although many regenerative cell therapies are being developed to replace or regenerate ischaemic muscle, the lack of vasculature and poor persistence of the therapeutic cells represent major limiting factors to successful tissue restoration. In response to ischaemia, stromal cell-derived factor-1 (SDF-1 is up-regulated by the affected tissue to stimulate stem cell-mediated regenerative responses. Therefore, we encapsulated SDF-1 into alginate microspheres and further incorporated these into an injectable collagen-based matrix in order to improve local delivery. Microsphere-matrix impregnation reduced the time for matrix thermogelation, and also increased the viscosity reached. This double-incorporation prolonged the release of SDF-1, which maintained adhesive and migratory bioactivity, attributed to chemotaxis in response to SDF-1. In vivo, treatment of ischaemic hindlimb muscle with microsphere-matrix led to increased mobilisation of bone marrow-derived progenitor cells, and also improved recruitment of angiogenic cells expressing the SDF-1 receptor (CXCR4 from bone marrow and local tissues. Both matrix and SDF-1-releasing matrix were successful at restoring perfusion, but SDF-1 treatment appeared to play an earlier role, as evidenced by arterioles that are phenotypically older and by increased angiogenic cytokine production, stimulating the generation of a qualitative microenvironment for a rapid and therefore more efficient regeneration. These results support the release of implanted SDF-1 as a promising method for enhancing progenitor cell responses and restoring perfusion to ischaemic tissues via neovascularisation.

  14. Human embryonic stem cell-derived oligodendrocyte progenitor cell transplants remyelinate and restore locomotion after spinal cord injury.

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    Keirstead, Hans S; Nistor, Gabriel; Bernal, Giovanna; Totoiu, Minodora; Cloutier, Frank; Sharp, Kelly; Steward, Oswald

    2005-05-11

    Demyelination contributes to loss of function after spinal cord injury, and thus a potential therapeutic strategy involves replacing myelin-forming cells. Here, we show that transplantation of human embryonic stem cell (hESC)-derived oligodendrocyte progenitor cells (OPCs) into adult rat spinal cord injuries enhances remyelination and promotes improvement of motor function. OPCs were injected 7 d or 10 months after injury. In both cases, transplanted cells survived, redistributed over short distances, and differentiated into oligodendrocytes. Animals that received OPCs 7 d after injury exhibited enhanced remyelination and substantially improved locomotor ability. In contrast, when OPCs were transplanted 10 months after injury, there was no enhanced remyelination or locomotor recovery. These studies document the feasibility of predifferentiating hESCs into functional OPCs and demonstrate their therapeutic potential at early time points after spinal cord injury.

  15. Stem/progenitor cells derived from the cochlear sensory epithelium give rise to spheres with distinct morphologies and features.

    Science.gov (United States)

    Diensthuber, Marc; Oshima, Kazuo; Heller, Stefan

    2009-06-01

    Nonmammalian vertebrates regenerate lost sensory hair cells by means of asymmetric division of supporting cells. Inner ear or lateral line supporting cells in birds, amphibians, and fish consequently serve as bona fide stem cells resulting in high regenerative capacity of hair cell-bearing organs. Hair cell regeneration does not happen in the mammalian cochlea, but cells with proliferative capacity can be isolated from the neonatal cochlea. These cells have the ability to form clonal floating colonies, so-called spheres, when cultured in nonadherent conditions. We noticed that the sphere population derived from mouse cochlear sensory epithelium cells was heterogeneous, consisting of morphologically distinct sphere types, hereby classified as solid, transitional, and hollow. Cochlear sensory epithelium-derived stem/progenitor cells initially give rise to small solid spheres, which subsequently transition into hollow spheres, a change that is accompanied by epithelial differentiation of the majority of sphere cells. Only solid spheres, and to a lesser extent, transitional spheres, appeared to harbor self-renewing stem cells, whereas hollow spheres could not be consistently propagated. Solid spheres contained significantly more rapidly cycling Pax-2-expressing presumptive otic progenitor cells than hollow spheres. Islet-1, which becomes upregulated in nascent sensory patches, was also more abundant in solid than in hollow spheres. Likewise, hair cell-like cells, characterized by the expression of multiple hair cell markers, differentiated in significantly higher numbers in cell populations derived from solid spheres. We conclude that cochlear sensory epithelium cell populations initially give rise to small solid spheres that have self-renewing capacity before they subsequently convert into hollow spheres, a process that is accompanied by loss of stemness and reduced ability to spontaneously give rise to hair cell-like cells. Solid spheres might, therefore, represent

  16. Characterization of progenitor cells derived from torn human rotator cuff tendons by gene expression patterns of chondrogenesis, osteogenesis, and adipogenesis.

    Science.gov (United States)

    Nagura, Issei; Kokubu, Takeshi; Mifune, Yutaka; Inui, Atsuyuki; Takase, Fumiaki; Ueda, Yasuhiro; Kataoka, Takeshi; Kurosaka, Masahiro

    2016-03-31

    It is important to regenerate the tendon-to-bone interface after rotator cuff repair to prevent re-tears. The cells from torn human rotator cuff were targeted, and their capacity for multilineage differentiation was investigated. The edges of the rotator cuff were harvested during arthroscopic rotator cuff repair from nine patients, minced into pieces, and cultured on dishes. Adherent cells were cultured, phenotypically characterized. Then expandability, differentiation potential and gene expression were analyzed. Flow cytometry revealed that the mesenchymal stem cells (MSC)-related markers CD29, CD44, CD105, and CD166 were positive. However, CD14, CD34, and CD45 were negative. On RT-PCR analyses, the cells showed osteogenic, adipogenic, and chondrogenic potential after 3 weeks of culture under the respective differentiation conditions. In addition, SOX9, type II collagen, and type X collagen expression patterns during chondrogenesis were similar to those of endochondral ossification at the enthesis. The cells derived from torn human rotator cuff are multipotent mesenchymal stem cells with the ability to undergo multilineage differentiation, suggesting that MSCs form this tissue could be regenerative capacity for potential self-repair.

  17. A chemically defined substrate for the expansion and neuronal differentiation of human pluripotent stem cell-derived neural progenitor cells

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    Yihuan Tsai

    2015-07-01

    Full Text Available Due to the limitation of current pharmacological therapeutic strategies, stem cell therapies have emerged as a viable option for treating many incurable neurological disorders. Specifically, human pluripotent stem cell (hPSC-derived neural progenitor cells (hNPCs, a multipotent cell population that is capable of near indefinite expansion and subsequent differentiation into the various cell types that comprise the central nervous system (CNS, could provide an unlimited source of cells for such cell-based therapies. However the clinical application of these cells will require (i defined, xeno-free conditions for their expansion and neuronal differentiation and (ii scalable culture systems that enable their expansion and neuronal differentiation in numbers sufficient for regenerative medicine and drug screening purposes. Current extracellular matrix protein (ECMP-based substrates for the culture of hNPCs are expensive, difficult to isolate, subject to batch-to-batch variations, and, therefore, unsuitable for clinical application of hNPCs. Using a high-throughput array-based screening approach, we identified a synthetic polymer, poly(4-vinyl phenol (P4VP, that supported the long-term proliferation and self-renewal of hNPCs. The hNPCs cultured on P4VP maintained their characteristic morphology, expressed high levels of markers of multipotency, and retained their ability to differentiate into neurons. Such chemically defined substrates will eliminate critical roadblocks for the utilization of hNPCs for human neural regenerative repair, disease modeling, and drug discovery.

  18. A chemically defined substrate for the expansion and neuronal differentiation of human pluripotent stem cell-derived neural progenitor cells.

    Science.gov (United States)

    Tsai, Yihuan; Cutts, Josh; Kimura, Azuma; Varun, Divya; Brafman, David A

    2015-07-01

    Due to the limitation of current pharmacological therapeutic strategies, stem cell therapies have emerged as a viable option for treating many incurable neurological disorders. Specifically, human pluripotent stem cell (hPSC)-derived neural progenitor cells (hNPCs), a multipotent cell population that is capable of near indefinite expansion and subsequent differentiation into the various cell types that comprise the central nervous system (CNS), could provide an unlimited source of cells for such cell-based therapies. However the clinical application of these cells will require (i) defined, xeno-free conditions for their expansion and neuronal differentiation and (ii) scalable culture systems that enable their expansion and neuronal differentiation in numbers sufficient for regenerative medicine and drug screening purposes. Current extracellular matrix protein (ECMP)-based substrates for the culture of hNPCs are expensive, difficult to isolate, subject to batch-to-batch variations, and, therefore, unsuitable for clinical application of hNPCs. Using a high-throughput array-based screening approach, we identified a synthetic polymer, poly(4-vinyl phenol) (P4VP), that supported the long-term proliferation and self-renewal of hNPCs. The hNPCs cultured on P4VP maintained their characteristic morphology, expressed high levels of markers of multipotency, and retained their ability to differentiate into neurons. Such chemically defined substrates will eliminate critical roadblocks for the utilization of hNPCs for human neural regenerative repair, disease modeling, and drug discovery. Copyright © 2015. Published by Elsevier B.V.

  19. Engrafted human induced pluripotent stem cell-derived anterior specified neural progenitors protect the rat crushed optic nerve.

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    Leila Satarian

    Full Text Available BACKGROUND: Degeneration of retinal ganglion cells (RGCs is a common occurrence in several eye diseases. This study examined the functional improvement and protection of host RGCs in addition to the survival, integration and neuronal differentiation capabilities of anterior specified neural progenitors (NPs following intravitreal transplantation. METHODOLOGY/PRINCIPAL FINDINGS: NPs were produced under defined conditions from human induced pluripotent stem cells (hiPSCs and transplanted into rats whose optic nerves have been crushed (ONC. hiPSCs were induced to differentiate into anterior specified NPs by the use of Noggin and retinoic acid. The hiPSC-NPs were labeled by green fluorescent protein or a fluorescent tracer 1,1' -dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate (DiI and injected two days after induction of ONC in hooded rats. Functional analysis according to visual evoked potential recordings showed significant amplitude recovery in animals transplanted with hiPSC-NPs. Retrograde labeling by an intra-collicular DiI injection showed significantly higher numbers of RGCs and spared axons in ONC rats treated with hiPSC-NPs or their conditioned medium (CM. The analysis of CM of hiPSC-NPs showed the secretion of ciliary neurotrophic factor, basic fibroblast growth factor, and insulin-like growth factor. Optic nerve of cell transplanted groups also had increased GAP43 immunoreactivity and myelin staining by FluoroMyelin™ which imply for protection of axons and myelin. At 60 days post-transplantation hiPSC-NPs were integrated into the ganglion cell layer of the retina and expressed neuronal markers. CONCLUSIONS/SIGNIFICANCE: The transplantation of anterior specified NPs may improve optic nerve injury through neuroprotection and differentiation into neuronal lineages. These NPs possibly provide a promising new therapeutic approach for traumatic optic nerve injuries and loss of RGCs caused by other diseases.

  20. Characterization of calcium signals in human induced pluripotent stem cell-derived dentate gyrus neuronal progenitors and mature neurons, stably expressing an advanced calcium indicator protein.

    Science.gov (United States)

    Vőfély, Gergő; Berecz, Tünde; Szabó, Eszter; Szebényi, Kornélia; Hathy, Edit; Orbán, Tamás I; Sarkadi, Balázs; Homolya, László; Marchetto, Maria C; Réthelyi, János M; Apáti, Ágota

    2018-04-01

    Pluripotent stem cell derived human neuronal progenitor cells (hPSC-NPCs) and their mature neuronal cell culture derivatives may efficiently be used for central nervous system (CNS) drug screening, including the investigation of ligand-induced calcium signalization. We have established hippocampal NPC cultures derived from human induced PSCs, which were previously generated by non-integrating Sendai virus reprogramming. Using established protocols these NPCs were differentiated into hippocampal dentate gyrus neurons. In order to study calcium signaling without the need of dye loading, we have stably expressed an advanced calcium indicator protein (GCaMP6fast) in the NPCs using the Sleeping Beauty transposon system. We observed no significant effects of the long-term GCaMP6 expression on NPC morphology, gene expression pattern or neural differentiation capacity. In order to compare the functional properties of GCaMP6-expressing neural cells and the corresponding parental cells loaded with calcium indicator dye Fluo-4, a detailed characterization of calcium signals was performed. We found that the calcium signals induced by ATP, glutamate, LPA, or proteases - were similar in these two systems. Moreover, the presence of the calcium indicator protein allowed for a sensitive, repeatable detection of changes in calcium signaling during the process of neurogenesis and neuronal maturation. Copyright © 2018 Elsevier Inc. All rights reserved.

  1. Coordinated Proliferation and Differentiation of Human-Induced Pluripotent Stem Cell-Derived Cardiac Progenitor Cells Depend on Bone Morphogenetic Protein Signaling Regulation by GREMLIN 2.

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    Bylund, Jeffery B; Trinh, Linh T; Awgulewitsch, Cassandra P; Paik, David T; Jetter, Christopher; Jha, Rajneesh; Zhang, Jianhua; Nolan, Kristof; Xu, Chunhui; Thompson, Thomas B; Kamp, Timothy J; Hatzopoulos, Antonis K

    2017-05-01

    Heart development depends on coordinated proliferation and differentiation of cardiac progenitor cells (CPCs), but how the two processes are synchronized is not well understood. Here, we show that the secreted Bone Morphogenetic Protein (BMP) antagonist GREMLIN 2 (GREM2) is induced in CPCs shortly after cardiac mesoderm specification during differentiation of human pluripotent stem cells. GREM2 expression follows cardiac lineage differentiation independently of the differentiation method used, or the origin of the pluripotent stem cells, suggesting that GREM2 is linked to cardiogenesis. Addition of GREM2 protein strongly increases cardiomyocyte output compared to established procardiogenic differentiation methods. Our data show that inhibition of canonical BMP signaling by GREM2 is necessary to promote proliferation of CPCs. However, canonical BMP signaling inhibition alone is not sufficient to induce cardiac differentiation, which depends on subsequent JNK pathway activation specifically by GREM2. These findings may have broader implications in the design of approaches to orchestrate growth and differentiation of pluripotent stem cell-derived lineages that depend on precise regulation of BMP signaling.

  2. Coordinated Proliferation and Differentiation of Human-Induced Pluripotent Stem Cell-Derived Cardiac Progenitor Cells Depend on Bone Morphogenetic Protein Signaling Regulation by GREMLIN 2

    Science.gov (United States)

    Bylund, Jeffery B.; Trinh, Linh T.; Awgulewitsch, Cassandra P.; Paik, David T.; Jetter, Christopher; Jha, Rajneesh; Zhang, Jianhua; Nolan, Kristof; Xu, Chunhui; Thompson, Thomas B.; Kamp, Timothy J.

    2017-01-01

    Heart development depends on coordinated proliferation and differentiation of cardiac progenitor cells (CPCs), but how the two processes are synchronized is not well understood. Here, we show that the secreted Bone Morphogenetic Protein (BMP) antagonist GREMLIN 2 (GREM2) is induced in CPCs shortly after cardiac mesoderm specification during differentiation of human pluripotent stem cells. GREM2 expression follows cardiac lineage differentiation independently of the differentiation method used, or the origin of the pluripotent stem cells, suggesting that GREM2 is linked to cardiogenesis. Addition of GREM2 protein strongly increases cardiomyocyte output compared to established procardiogenic differentiation methods. Our data show that inhibition of canonical BMP signaling by GREM2 is necessary to promote proliferation of CPCs. However, canonical BMP signaling inhibition alone is not sufficient to induce cardiac differentiation, which depends on subsequent JNK pathway activation specifically by GREM2. These findings may have broader implications in the design of approaches to orchestrate growth and differentiation of pluripotent stem cell-derived lineages that depend on precise regulation of BMP signaling. PMID:28125926

  3. Intra-Articular Injection of Human Meniscus Stem/Progenitor Cells Promotes Meniscus Regeneration and Ameliorates Osteoarthritis Through Stromal Cell-Derived Factor-1/CXCR4-Mediated Homing

    Science.gov (United States)

    Shen, Weiliang; Chen, Jialin; Zhu, Ting; Chen, Longkun; Zhang, Wei; Fang, Zhi; Heng, Boon Chin; Yin, Zi; Chen, Xiao; Ji, Junfeng

    2014-01-01

    Meniscus injury is frequently encountered in clinical practice. Current surgical therapy involving partial or complete meniscectomy relieves pain in the short-term but often leads to osteoarthritis (OA) in the long-term. In this study, we report a new strategy of articular cartilage protection by intra-articular injection of novel human meniscus stem/progenitor cells (hMeSPCs). We found that hMeSPCs displayed both mesenchymal stem cell characteristics and high expression levels of collagen II. In the rat meniscus injury model, hMeSPC transplantation not only led to more neo-tissue formation and better-defined shape but also resulted in more rounded cells and matured extracellular matrix. Stromal cell-derived factor-1 (SDF-1) enhanced the migration of hMeSPCs, whereas AMD3100 abolished the chemotactic effects of SDF-1 on hMeSPCs, both in vitro and in vivo. In an experimental OA model, transplantation of hMeSPCs effectively protected articular cartilage, as evidenced by reduced expression of OA markers such as collagen I, collagen X, and hypoxia-inducible factor 2α but increased expression of collagen II. Our study demonstrated for the first time that intra-articular injection of hMeSPCs enhanced meniscus regeneration through the SDF-1/CXCR4 axis. Our study highlights a new strategy of intra-articular injection of hMeSPCs for meniscus regeneration. PMID:24448516

  4. Safe and efficient method for cryopreservation of human induced pluripotent stem cell-derived neural stem and progenitor cells by a programmed freezer with a magnetic field.

    Science.gov (United States)

    Nishiyama, Yuichiro; Iwanami, Akio; Kohyama, Jun; Itakura, Go; Kawabata, Soya; Sugai, Keiko; Nishimura, Soraya; Kashiwagi, Rei; Yasutake, Kaori; Isoda, Miho; Matsumoto, Morio; Nakamura, Masaya; Okano, Hideyuki

    2016-06-01

    Stem cells represent a potential cellular resource in the development of regenerative medicine approaches to the treatment of pathologies in which specific cells are degenerated or damaged by genetic abnormality, disease, or injury. Securing sufficient supplies of cells suited to the demands of cell transplantation, however, remains challenging, and the establishment of safe and efficient cell banking procedures is an important goal. Cryopreservation allows the storage of stem cells for prolonged time periods while maintaining them in adequate condition for use in clinical settings. Conventional cryopreservation systems include slow-freezing and vitrification both have advantages and disadvantages in terms of cell viability and/or scalability. In the present study, we developed an advanced slow-freezing technique using a programmed freezer with a magnetic field called Cells Alive System (CAS) and examined its effectiveness on human induced pluripotent stem cell-derived neural stem/progenitor cells (hiPSC-NS/PCs). This system significantly increased cell viability after thawing and had less impact on cellular proliferation and differentiation. We further found that frozen-thawed hiPSC-NS/PCs were comparable with non-frozen ones at the transcriptome level. Given these findings, we suggest that the CAS is useful for hiPSC-NS/PCs banking for clinical uses involving neural disorders and may open new avenues for future regenerative medicine. Copyright © 2015 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  5. Variation in primary and culture-expanded cells derived from connective tissue progenitors in human bone marrow space, bone trabecular surface and adipose tissue.

    Science.gov (United States)

    Qadan, Maha A; Piuzzi, Nicolas S; Boehm, Cynthia; Bova, Wesley; Moos, Malcolm; Midura, Ronald J; Hascall, Vincent C; Malcuit, Christopher; Muschler, George F

    2018-03-01

    Connective tissue progenitors (CTPs) embody the heterogeneous stem and progenitor cell populations present in native tissue. CTPs are essential to the formation and remodeling of connective tissue and represent key targets for tissue-engineering and cell-based therapies. To better understand and characterize CTPs, we aimed to compare the (i) concentration and prevalence, (ii) early in vitro biological behavior and (iii) expression of surface-markers and transcription factors among cells derived from marrow space (MS), trabecular surface (TS), and adipose tissues (AT). Cancellous-bone and subcutaneous-adipose tissues were collected from 8 patients. Cells were isolated and cultured. Colony formation was assayed using Colonyze software based on ASTM standards. Cell concentration ([Cell]), CTP concentration ([CTP]) and CTP prevalence (P CTP ) were determined. Attributes of culture-expanded cells were compared based on (i) effective proliferation rate and (ii) expression of surface-markers CD73, CD90, CD105, SSEA-4, SSEA-3, SSEA-1/CD15, Cripto-1, E-Cadherin/CD324, Ep-CAM/CD326, CD146, hyaluronan and transcription factors Oct3/4, Sox-2 and Nanog using flow cytometry. Mean [Cell], [CTP] and P CTP were significantly different between MS and TS samples (P = 0.03, P = 0.008 and P= 0.0003), respectively. AT-derived cells generated the highest mean total cell yield at day 6 of culture-4-fold greater than TS and more than 40-fold greater than MS per million cells plated. TS colonies grew with higher mean density than MS colonies (290 ± 11 versus 150 ± 11 cell per mm 2 ; P = 0.0002). Expression of classical-mesenchymal stromal cell (MSC) markers was consistently recorded (>95%) from all tissue sources, whereas all the other markers were highly variable. The prevalence and biological potential of CTPs are different between patients and tissue sources and lack variation in classical MSC markers. Other markers are more likely to discriminate differences

  6. Electroacupuncture modulates stromal cell-derived factor-1α expression and mobilization of bone marrow endothelial progenitor cells in focal cerebral ischemia/reperfusion model rats.

    Science.gov (United States)

    Xie, Chenchen; Gao, Xiang; Luo, Yong; Pang, Yueshan; Li, Man

    2016-10-01

    Stromal cell-derived factor-1α(SDF-1α) plays a crucial role in regulating the mobilization, migration and homing of endothelial progenitor cells(EPCs). Electroacupuncture(EA), a modern version of Traditional Chinese Medicine, can improve neurological recovery and angiogenesis in cerebral ischemic area. This study aimed to investigate the effects of electroacupuncture(EA) on the mobilization and migration of bone marrow EPCs and neurological functional recovery in rats model after focal cerebral ischemia/reperfusion and the potentially involved mechanisms. Sprague-Dawley rats received filament occlusion of the right middle cerebral artery for 2h followed by reperfusion for 12h, 1d, 2d, 3d, 7d respectively. Rats were randomly divided into sham group, model group and EA group. After 2h of the reperfusion, EA was given at the "Baihui" (GV 20)/Siguan ("Hegu" (LI 4)/"Taichong" (LR 3)) acupoints in the EA group. Modified neurological severity score (mNSS) was used to assess the neurological functional recovery. EPCs number and SDF-1α level in bone marrow(BM) and peripheral blood(PB) were detected by using fluorescence-activated cell sorting (FACS) analysis and quantitative real time polymerase chain reaction (qRT-PCR) respectively. An mNSS test showed that EA treatment significantly improved the neurological functional outcome. EPCs number in PB and BM were obviously increased in the EA group. After cerebral ischemia, the SDF-1α level was decreased in BM while it was increased in PB, which implied a gradient of SDF-1α among BM and PB after ischemia. It suggested that the forming of SDF-1α concentration gradient can induce the mobilization and homing of EPCs. Eletroacupuncture as a treatment can accelerate and increase the forming of SDF-1α concentration gradient to further induce the mobilization of EPCs and angiogenesis in ischemic brain and improve the neurological function recovery. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Acellular Mouse Kidney ECM can be Used as a Three-Dimensional Substrate to Test the Differentiation Potential of Embryonic Stem Cell Derived Renal Progenitors.

    Science.gov (United States)

    Sambi, Manpreet; Chow, Theresa; Whiteley, Jennifer; Li, Mira; Chua, Shawn; Raileanu, Vanessa; Rogers, Ian M

    2017-08-01

    The development of strategies for tissue regeneration and bio-artificial organ development is based on our understanding of embryogenesis. Differentiation protocols attempt to recapitulate the signaling modalities of gastrulation and organogenesis, coupled with cell selection regimens to isolate the cells of choice. This strategy is impeded by the lack of optimal in vitro culture systems since traditional culture systems do not allow for the three-dimensional interaction between cells and the extracellular matrix. While artificial three-dimensional scaffolds are available, using the natural extracellular matrix scaffold is advantageous because it has a distinct architecture that is difficult to replicate. The adult extracellular matrix is predicted to mediate signaling related to tissue repair not embryogenesis but existing similarities between the two argues that the extracellular matrix will influence the differentiation of stem and progenitor cells. Previous studies using undifferentiated embryonic stem cells grown directly on acellular kidney ECM demonstrated that the acellular kidney supported cell growth but limited differentiation occurred. Using mouse kidney extracellular matrix and mouse embryonic stem cells we report that the extracellular matrix can support the development of kidney structures if the stem cells are first differentiated to kidney progenitor cells before being applied to the acellular organ.

  8. Enrichment of human embryonic stem cell-derived NKX6.1-expressing pancreatic progenitor cells accelerates the maturation of insulin-secreting cells in vivo.

    Science.gov (United States)

    Rezania, Alireza; Bruin, Jennifer E; Xu, Jean; Narayan, Kavitha; Fox, Jessica K; O'Neil, John J; Kieffer, Timothy J

    2013-11-01

    Human embryonic stem cells (hESCs) are considered a potential alternative to cadaveric islets as a source of transplantable cells for treating patients with diabetes. We previously described a differentiation protocol to generate pancreatic progenitor cells from hESCs, composed of mainly pancreatic endoderm (PDX1/NKX6.1-positive), endocrine precursors (NKX2.2/synaptophysin-positive, hormone/NKX6.1-negative), and polyhormonal cells (insulin/glucagon-positive, NKX6.1-negative). However, the relative contributions of NKX6.1-negative versus NKX6.1-positive cell fractions to the maturation of functional β-cells remained unclear. To address this question, we generated two distinct pancreatic progenitor cell populations using modified differentiation protocols. Prior to transplant, both populations contained a high proportion of PDX1-expressing cells (~85%-90%) but were distinguished by their relatively high (~80%) or low (~25%) expression of NKX6.1. NKX6.1-high and NKX6.1-low progenitor populations were transplanted subcutaneously within macroencapsulation devices into diabetic mice. Mice transplanted with NKX6.1-low cells remained hyperglycemic throughout the 5-month post-transplant period whereas diabetes was reversed in NKX6.1-high recipients within 3 months. Fasting human C-peptide levels were similar between groups throughout the study, but only NKX6.1-high grafts displayed robust meal-, glucose- and arginine-responsive insulin secretion as early as 3 months post-transplant. NKX6.1-low recipients displayed elevated fasting glucagon levels. Theracyte devices from both groups contained almost exclusively pancreatic endocrine tissue, but NKX6.1-high grafts contained a greater proportion of insulin-positive and somatostatin-positive cells, whereas NKX6.1-low grafts contained mainly glucagon-expressing cells. Insulin-positive cells in NKX6.1-high, but not NKX6.1-low grafts expressed nuclear MAFA. Collectively, this study demonstrates that a pancreatic endoderm

  9. Transplantation of rat embryonic stem cell-derived retinal progenitor cells preserves the retinal structure and function in rat retinal degeneration.

    Science.gov (United States)

    Qu, Zepeng; Guan, Yuan; Cui, Lu; Song, Jian; Gu, Junjie; Zhao, Hanzhi; Xu, Lei; Lu, Lixia; Jin, Ying; Xu, Guo-Tong

    2015-11-09

    Degenerative retinal diseases like age-related macular degeneration (AMD) are the leading cause of blindness. Cell transplantation showed promising therapeutic effect for such diseases, and embryonic stem cell (ESC) is one of the sources of such donor cells. Here, we aimed to generate retinal progenitor cells (RPCs) from rat ESCs (rESCs) and to test their therapeutic effects in rat model. The rESCs (DA8-16) were cultured in N2B27 medium with 2i, and differentiated to two types of RPCs following the SFEBq method with modifications. For rESC-RPC1, the cells were switched to adherent culture at D10, while for rESC-RPC2, the suspension culture was maintained to D14. Both RPCs were harvested at D16. Primary RPCs were obtained from P1 SD rats, and some of them were labeled with EGFP by infection with lentivirus. To generate Rax::EGFP knock-in rESC lines, TALENs were engineered to facilitate homologous recombination in rESCs, which were cotransfected with the targeting vector and TALEN vectors. The differentiated cells were analyzed with live image, immunofluorescence staining, flow cytometric analysis, gene expression microarray, etc. RCS rats were used to mimic the degeneration of retina and test the therapeutic effects of subretinally transplanted donor cells. The structure and function of retina were examined. We established two protocols through which two types of rESC-derived RPCs were obtained and both contained committed retina lineage cells and some neural progenitor cells (NPCs). These rESC-derived RPCs survived in the host retinas of RCS rats and protected the retinal structure and function in early stage following the transplantation. However, the glia enriched rESC-RPC1 obtained through early and longer adherent culture only increased the b-wave amplitude at 4 weeks, while the longer suspension culture gave rise to evidently neuronal differentiation in rESC-RPC2 which significantly improved the visual function of RCS rats. We have successfully differentiated

  10. Protease-activated receptor-1 negatively regulates proliferation of neural stem/progenitor cells derived from the hippocampal dentate gyrus of the adult mouse

    Directory of Open Access Journals (Sweden)

    Masayuki Tanaka

    2016-07-01

    Full Text Available Thrombin-activated protease-activated receptor (PAR-1 regulates the proliferation of neural cells following brain injury. To elucidate the involvement of PAR-1 in the neurogenesis that occurs in the adult hippocampus, we examined whether PAR-1 regulated the proliferation of neural stem/progenitor cells (NPCs derived from the murine hippocampal dentate gyrus. NPC cultures expressed PAR-1 protein and mRNA encoding all subtypes of PAR. Direct exposure of the cells to thrombin dramatically attenuated the cell proliferation without causing cell damage. This thrombin-induced attenuation was almost completely abolished by the PAR antagonist RWJ 56110, as well as by dabigatran and 4-(2-aminoethylbenzenesulfonyl fluoride (AEBSF, which are selective and non-selective thrombin inhibitors, respectively. Expectedly, the PAR-1 agonist peptide (AP SFLLR-NH2 also attenuated the cell proliferation. The cell proliferation was not affected by the PAR-1 negative control peptide RLLFT-NH2, which is an inactive peptide for PAR-1. Independently, we determined the effect of in vivo treatment with AEBSF or AP on hippocampal neurogenesis in the adult mouse. The administration of AEBSF, but not that of AP, significantly increased the number of newly-generated cells in the hippocampal subgranular zone. These data suggest that PAR-1 negatively regulated adult neurogenesis in the hippocampus by inhibiting the proliferative activity of the NPCs.

  11. CPM Is a Useful Cell Surface Marker to Isolate Expandable Bi-Potential Liver Progenitor Cells Derived from Human iPS Cells.

    Science.gov (United States)

    Kido, Taketomo; Koui, Yuta; Suzuki, Kaori; Kobayashi, Ayaka; Miura, Yasushi; Chern, Edward Y; Tanaka, Minoru; Miyajima, Atsushi

    2015-10-13

    To develop a culture system for large-scale production of mature hepatocytes, liver progenitor cells (LPCs) with a high proliferation potential would be advantageous. We have found that carboxypeptidase M (CPM) is highly expressed in embryonic LPCs, hepatoblasts, while its expression is decreased along with hepatic maturation. Consistently, CPM expression was transiently induced during hepatic specification from human-induced pluripotent stem cells (hiPSCs). CPM(+) cells isolated from differentiated hiPSCs at the immature hepatocyte stage proliferated extensively in vitro and expressed a set of genes that were typical of hepatoblasts. Moreover, the CPM(+) cells exhibited a mature hepatocyte phenotype after induction of hepatic maturation and also underwent cholangiocytic differentiation in a three-dimensional culture system. These results indicated that hiPSC-derived CPM(+) cells share the characteristics of LPCs, with the potential to proliferate and differentiate bi-directionally. Thus, CPM is a useful marker for isolating hiPSC-derived LPCs, which allows development of a large-scale culture system for producing hepatocytes and cholangiocytes. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  12. Comparative transcriptomic analysis of endothelial progenitor cells derived from umbilical cord blood and adult peripheral blood: Implications for the generation of induced pluripotent stem cells

    Directory of Open Access Journals (Sweden)

    Xiugong Gao

    2017-12-01

    Full Text Available Induced pluripotent stem cells (iPSCs offer the potential to generate tissues with ethnic diversity enabling toxicity testing on selected populations. Recently, it has been reported that endothelial progenitor cells (EPCs derived from umbilical cord blood (CB or adult peripheral blood (PB afford a practical and efficient cellular substrate for iPSC generation. However, differences between EPCs from different blood sources have rarely been studied. In the current study, we derived EPCs from blood mononuclear cells (MNCs and reprogrammed EPCs into iPSCs. We also explored differences between CB-EPCs and PB-EPCs at the molecular and cellular levels through a combination of transcriptomic analysis and cell biology techniques. EPC colonies in CB-MNCs emerged 5–7 days earlier, were 3-fold higher in number, and consistently larger in size than in PB-MNCs. Similarly, iPSC colonies generated from CB-EPCs was 2.5-fold higher in number than from PB-EPCs, indicating CB-EPCs have a higher reprogramming efficiency than PB-EPCs. Transcriptomic analysis using microarrays found a total of 1133 genes differentially expressed in CB-EPCs compared with PB-EPCs, with 675 genes upregulated and 458 downregulated. Several canonical pathways were impacted, among which the human embryonic stem cell pluripotency pathway was of particular interest. The differences in the gene expression pattern between CB-EPCs and PB-EPCs provide a molecular basis for the discrepancies seen in their derivation and reprogramming efficiencies, and highlight the advantages of using CB as the cellular source for the generation of iPSCs and their derivative tissues for ethnic-related toxicological applications.

  13. Comparative transcriptomic analysis of endothelial progenitor cells derived from umbilical cord blood and adult peripheral blood: Implications for the generation of induced pluripotent stem cells.

    Science.gov (United States)

    Gao, Xiugong; Yourick, Jeffrey J; Sprando, Robert L

    2017-12-01

    Induced pluripotent stem cells (iPSCs) offer the potential to generate tissues with ethnic diversity enabling toxicity testing on selected populations. Recently, it has been reported that endothelial progenitor cells (EPCs) derived from umbilical cord blood (CB) or adult peripheral blood (PB) afford a practical and efficient cellular substrate for iPSC generation. However, differences between EPCs from different blood sources have rarely been studied. In the current study, we derived EPCs from blood mononuclear cells (MNCs) and reprogrammed EPCs into iPSCs. We also explored differences between CB-EPCs and PB-EPCs at the molecular and cellular levels through a combination of transcriptomic analysis and cell biology techniques. EPC colonies in CB-MNCs emerged 5-7days earlier, were 3-fold higher in number, and consistently larger in size than in PB-MNCs. Similarly, iPSC colonies generated from CB-EPCs was 2.5-fold higher in number than from PB-EPCs, indicating CB-EPCs have a higher reprogramming efficiency than PB-EPCs. Transcriptomic analysis using microarrays found a total of 1133 genes differentially expressed in CB-EPCs compared with PB-EPCs, with 675 genes upregulated and 458 downregulated. Several canonical pathways were impacted, among which the human embryonic stem cell pluripotency pathway was of particular interest. The differences in the gene expression pattern between CB-EPCs and PB-EPCs provide a molecular basis for the discrepancies seen in their derivation and reprogramming efficiencies, and highlight the advantages of using CB as the cellular source for the generation of iPSCs and their derivative tissues for ethnic-related toxicological applications. Published by Elsevier B.V.

  14. Rotary orbital suspension culture of embryonic stem cell-derived neural stem/progenitor cells: impact of hydrodynamic culture on aggregate yield, morphology and cell phenotype.

    Science.gov (United States)

    Laundos, Tiago L; Silva, Joana; Assunção, Marisa; Quelhas, Pedro; Monteiro, Cátia; Oliveira, Carla; Oliveira, Maria J; Pêgo, Ana P; Amaral, Isabel F

    2017-08-01

    Embryonic stem (ES)-derived neural stem/progenitor cells (ES-NSPCs) constitute a promising cell source for application in cell therapies for the treatment of central nervous system disorders. In this study, a rotary orbital hydrodynamic culture system was applied to single-cell suspensions of ES-NSPCs, to obtain homogeneously-sized ES-NSPC cellular aggregates (neurospheres). Hydrodynamic culture allowed the formation of ES-NSPC neurospheres with a narrower size distribution than statically cultured neurospheres, increasing orbital speeds leading to smaller-sized neurospheres and higher neurosphere yield. Neurospheres formed under hydrodynamic conditions (72 h at 55 rpm) showed higher cell compaction and comparable percentages of viable, dead, apoptotic and proliferative cells. Further characterization of cellular aggregates provided new insights into the effect of hydrodynamic shear on ES-NSPC behaviour. Rotary neurospheres exhibited reduced protein levels of N-cadherin and β-catenin, and higher deposition of laminin (without impacting fibronectin deposition), matrix metalloproteinase-2 (MMP-2) activity and percentage of neuronal cells. In line with the increased MMP-2 activity levels found, hydrodynamically-cultured neurospheres showed higher outward migration on laminin. Moreover, when cultured in a 3D fibrin hydrogel, rotary neurospheres generated an increased percentage of neuronal cells. In conclusion, the application of a constant orbital speed to single-cell suspensions of ES-NSPCs, besides allowing the formation of homogeneously-sized neurospheres, promoted ES-NSPC differentiation and outward migration, possibly by influencing the expression of cell-cell adhesion molecules and the secretion of proteases/extracellular matrix proteins. These findings are important when establishing the culture conditions needed to obtain uniformly-sized ES-NSPC aggregates, either for use in regenerative therapies or in in vitro platforms for biomaterial development or

  15. Molecular functions of the LIM-homeobox transcription factor Lhx2 in hematopoietic progenitor cells derived from mouse embryonic stem cells.

    Science.gov (United States)

    Kitajima, Kenji; Kawaguchi, Manami; Iacovino, Michelina; Kyba, Michael; Hara, Takahiko

    2013-12-01

    We previously demonstrated that hematopoietic stem cell (HSC)-like cells are robustly expanded from mouse embryonic stem cells (ESCs) by enforced expression of Lhx2, a LIM-homeobox domain (LIM-HD) transcription factor. In this study, we analyzed the functions of Lhx2 in that process using an ESC line harboring an inducible Lhx2 gene cassette. When ESCs are cultured on OP9 stromal cells, hematopoietic progenitor cells (HPCs) are differentiated and these HPCs are prone to undergo rapid differentiation into mature hematopoietic cells. Lhx2 inhibited differentiation of HPCs into mature hematopoietic cells and this effect would lead to accumulation of HSC-like cells. LIM-HD factors interact with LIM domain binding (Ldb) protein and this interaction abrogates binding of LIM-only (Lmo) protein to Ldb. We found that one of Lmo protein, Lmo2, was unstable due to dissociation of Lmo2 from Ldb1 in the presence of Lhx2. This effect of Lhx2 on the amount of Lmo2 contributed into accumulation of HSC-like cells, since enforced expression of Lmo2 into HSC-like cells inhibited their self-renewal. Expression of Gata3 and Tal1/Scl was increased in HSC-like cells and enforced expression of Lmo2 reduced expression of Gata3 but not Tal1/Scl. Enforced expression of Gata3 into HPCs inhibited mature hematopoietic cell differentiation, whereas Gata3-knockdown abrogated the Lhx2-mediated expansion of HPCs. We propose that multiple transcription factors/cofactors are involved in the Lhx2-mediated expansion of HSC-like cells from ESCs. Lhx2 appears to fine-tune the balance between self-renewal and differentiation of HSC-like cells. © AlphaMed Press.

  16. Male-specific differences in proliferation, neurogenesis, and sensitivity to oxidative stress in neural progenitor cells derived from a rat model of ALS.

    Directory of Open Access Journals (Sweden)

    Ruojia Li

    Full Text Available Amyotrophic Lateral Sclerosis (ALS is a fatal neurodegenerative disease characterized by progressive motor dysfunction and the loss of large motor neurons in the spinal cord and brain stem. A clear genetic link to point mutations in the superoxide dismutase 1 (SOD1 gene has been shown in a small group of familial ALS patients. The exact etiology of ALS is still uncertain, but males have consistently been shown to be at a higher risk for the disease than females. Here we present male-specific effects of the mutant SOD1 transgene on proliferation, neurogenesis, and sensitivity to oxidative stress in rat neural progenitor cells (rNPCs. E14 pups were bred using SOD1(G93A transgenic male rats and wild-type female rats. The spinal cord and cortex tissues were collected, genotyped by PCR using primers for the SOD1(G93A transgene or the male-specific Sry gene, and cultured as neurospheres. The number of dividing cells was higher in male rNPCs compared to female rNPCs. However, SOD1(G93A over-expression significantly reduced cell proliferation in male cells but not female cells. Similarly, male rNPCs produced more neurons compared to female rNPCs, but SOD1(G93A over-expression significantly reduced the number of neurons produced in male cells. Finally we asked whether sex and SOD1(G93A transgenes affected sensitivity to oxidative stress. There was no sex-based difference in cell viability after treatment with hydrogen peroxide or 3-morpholinosydnonimine, a free radical-generating agent. However, increased cytotoxicity by SOD1(G93A over-expression occurred, especially in male rNPCs. These results provide essential information on how the mutant SOD1 gene and sexual dimorphism are involved in ALS disease progression.

  17. The non-alcoholic fraction of beer increases stromal cell derived factor 1 and the number of circulating endothelial progenitor cells in high cardiovascular risk subjects: a randomized clinical trial.

    Science.gov (United States)

    Chiva-Blanch, Gemma; Condines, Ximena; Magraner, Emma; Roth, Irene; Valderas-Martínez, Palmira; Arranz, Sara; Casas, Rosa; Martínez-Huélamo, Miriam; Vallverdú-Queralt, Anna; Quifer-Rada, Paola; Lamuela-Raventos, Rosa M; Estruch, Ramon

    2014-04-01

    Moderate alcohol consumption is associated with a decrease in cardiovascular risk, but fermented beverages seem to confer greater cardiovascular protection due to their polyphenolic content. Circulating endothelial progenitor cells (EPC) are bone-marrow-derived stem cells with the ability to repair and maintain endothelial integrity and function and are considered as a surrogate marker of vascular function and cumulative cardiovascular risk. Nevertheless, no study has been carried out on the effects of moderate beer consumption on the number of circulating EPC in high cardiovascular risk patients. To compare the effects of moderate consumption of beer, non-alcoholic beer and gin on the number of circulating EPC and EPC-mobilizing factors. In this crossover trial, 33 men at high cardiovascular risk were randomized to receive beer (30 g alcohol/d), the equivalent amount of polyphenols in the form of non-alcoholic beer, or gin (30 g alcohol/d) for 4 weeks. Diet and physical exercise were carefully monitored. The number of circulating EPC and EPC-mobilizing factors were determined at baseline and after each intervention. After the beer and non-alcoholic beer interventions, the number of circulating EPC significantly increased by 8 and 5 units, respectively, while no significant differences were observed after the gin period. In correlation, stromal cell derived factor 1 increased significantly after the non-alcoholic and the beer interventions. The non-alcoholic fraction of beer increases the number of circulating EPC in peripheral blood from high cardiovascular risk subjects. http://www.controlled-trials.com/ISRCTN95345245 ISRCTN95345245. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  18. Characterising the developmental profile of human embryonic stem cell-derived medium spiny neuron progenitors and assessing mature neuron function using a CRISPR-generated human DARPP-32WT/eGFP-AMP reporter line.

    Science.gov (United States)

    Hunt, C P J; Pouton, C W; Haynes, J M

    2017-06-01

    In the developing ventral telencephalon, cells of the lateral ganglionic eminence (LGE) give rise to all medium spiny neurons (MSNs). This development occurs in response to a highly orchestrated series of morphogenetic stimuli that pattern the resultant neurons as they develop. Striatal MSNs are characterised by expression of dopamine receptors, dopamine-and cyclic AMP-regulated phosphoprotein (DARPP32) and the neurotransmitter GABA. In this study, we demonstrate that fine tuning Wnt and hedgehog (SHH) signaling early in human embryonic stem cell differentiation can induce a subpallial progenitor molecular profile. Stimulation of TGFβ signaling pathway by activin-A further supports patterning of progenitors to striatal precursors which adopt an LGE-specific gene signature. Moreover, we report that these MSNs also express markers associated with mature neuron function (cannabinoid, adenosine and dopamine receptors). To facilitate live-cell identification we generated a human embryonic stem cell line using CRISPR-mediated gene editing at the DARPP32 locus (DARPP32 WT/eGFP-AMP-LacZ ). The addition of dopamine to MSNs either increased, decreased or had no effect on intracellular calcium, indicating the presence of multiple dopamine receptor subtypes. In summary, we demonstrate greater control over early fate decisions using activin-A, Wnt and SHH to direct differentiation into MSNs. We also generate a DARPP32 reporter line that enables deeper pharmacological profiling and interrogation of complex receptor interactions in human MSNs. Copyright © 2017 Elsevier Ltd. All rights reserved.

  19. Stromal cell-derived factor 1α (SDF-1α)

    DEFF Research Database (Denmark)

    Li, Dana; Bjørnager, Louise; Langkilde, Anne

    2016-01-01

    OBJECTIVES: Stromal cell-derived factor 1a (SDF-1α), is a chemokine and is able to home hematopoietic progenitor cells to injured areas of heart tissue for structural repair. Previous studies have found increased levels of SDF-1α in several cardiac diseases, but only few studies have investigated...

  20. Small-scale screening of anticancer drugs acting specifically on neural stem/progenitor cells derived from human-induced pluripotent stem cells using a time-course cytotoxicity test

    Directory of Open Access Journals (Sweden)

    Hayato Fukusumi

    2018-01-01

    Full Text Available Since the development of human-induced pluripotent stem cells (hiPSCs, various types of hiPSC-derived cells have been established for regenerative medicine and drug development. Neural stem/progenitor cells (NSPCs derived from hiPSCs (hiPSC-NSPCs have shown benefits for regenerative therapy of the central nervous system. However, owing to their intrinsic proliferative potential, therapies using transplanted hiPSC-NSPCs carry an inherent risk of undesired growth in vivo. Therefore, it is important to find cytotoxic drugs that can specifically target overproliferative transplanted hiPSC-NSPCs without damaging the intrinsic in vivo stem-cell system. Here, we examined the chemosensitivity of hiPSC-NSPCs and human neural tissue—derived NSPCs (hN-NSPCs to the general anticancer drugs cisplatin, etoposide, mercaptopurine, and methotrexate. A time-course analysis of neurospheres in a microsphere array identified cisplatin and etoposide as fast-acting drugs, and mercaptopurine and methotrexate as slow-acting drugs. Notably, the slow-acting drugs were eventually cytotoxic to hiPSC-NSPCs but not to hN-NSPCs, a phenomenon not evident in the conventional endpoint assay on day 2 of treatment. Our results indicate that slow-acting drugs can distinguish hiPSC-NSPCs from hN-NSPCs and may provide an effective backup safety measure in stem-cell transplant therapies.

  1. Small-scale screening of anticancer drugs acting specifically on neural stem/progenitor cells derived from human-induced pluripotent stem cells using a time-course cytotoxicity test.

    Science.gov (United States)

    Fukusumi, Hayato; Handa, Yukako; Shofuda, Tomoko; Kanemura, Yonehiro

    2018-01-01

    Since the development of human-induced pluripotent stem cells (hiPSCs), various types of hiPSC-derived cells have been established for regenerative medicine and drug development. Neural stem/progenitor cells (NSPCs) derived from hiPSCs (hiPSC-NSPCs) have shown benefits for regenerative therapy of the central nervous system. However, owing to their intrinsic proliferative potential, therapies using transplanted hiPSC-NSPCs carry an inherent risk of undesired growth in vivo . Therefore, it is important to find cytotoxic drugs that can specifically target overproliferative transplanted hiPSC-NSPCs without damaging the intrinsic in vivo stem-cell system. Here, we examined the chemosensitivity of hiPSC-NSPCs and human neural tissue-derived NSPCs (hN-NSPCs) to the general anticancer drugs cisplatin, etoposide, mercaptopurine, and methotrexate. A time-course analysis of neurospheres in a microsphere array identified cisplatin and etoposide as fast-acting drugs, and mercaptopurine and methotrexate as slow-acting drugs. Notably, the slow-acting drugs were eventually cytotoxic to hiPSC-NSPCs but not to hN-NSPCs, a phenomenon not evident in the conventional endpoint assay on day 2 of treatment. Our results indicate that slow-acting drugs can distinguish hiPSC-NSPCs from hN-NSPCs and may provide an effective backup safety measure in stem-cell transplant therapies.

  2. Progenitor Epithelium

    Science.gov (United States)

    Marty-Santos, Leilani

    2015-01-01

    Insulin-producing β cells within the vertebrate fetal pancreas acquire their fate in a step-wise manner. Whereas the intrinsic factors dictating the transcriptional or epigenetic status of pancreatic lineages have been intensely examined, less is known about cell–cell interactions that might constitute a niche for the developing β cell lineage. It is becoming increasingly clear that understanding and recapitulating these steps may instruct in vitro differentiation of embryonic stem cells and/or therapeutic regeneration. Indeed, directed differentiation techniques have improved since transitioning from 2D to 3D cultures, suggesting that the 3D microenvironment in which β cells are born is critical. However, to date, it remains unknown whether the changing architecture of the pancreatic epithelium impacts the fate of cells therein. An emerging challenge in the field is to elucidate how progenitors are allocated during key events, such as the stratification and subsequent resolution of the pre-pancreatic epithelium, as well as the formation of lumens and branches. Here, we assess the progenitor epithelium and examine how it might influence the emergence of pancreatic multipotent progenitors (MPCs), which give rise to β cells and other pancreatic lineages. PMID:26216134

  3. Endothelial cell-derived matrix promotes the metabolic functional maturation of hepatocyte via integrin-Src signalling.

    Science.gov (United States)

    Guo, Xinyue; Li, Weihong; Ma, Minghui; Lu, Xin; Zhang, Haiyan

    2017-11-01

    The extracellular matrix (ECM) microenvironment is involved in the regulation of hepatocyte phenotype and function. Recently, the cell-derived extracellular matrix has been proposed to represent the bioactive and biocompatible materials of the native ECM. Here, we show that the endothelial cell-derived matrix (EC matrix) promotes the metabolic maturation of human adipose stem cell-derived hepatocyte-like cells (hASC-HLCs) through the activation of the transcription factor forkhead box protein A2 (FOXA2) and the nuclear receptors hepatocyte nuclear factor 4 alpha (HNF4α) and pregnane X receptor (PXR). Reducing the fibronectin content in the EC matrix or silencing the expression of α5 integrin in the hASC-HLCs inhibited the effect of the EC matrix on Src phosphorylation and hepatocyte maturation. The inhibition of Src phosphorylation using the inhibitor PP2 or silencing the expression of Src in hASC-HLCs also attenuated the up-regulation of the metabolic function of hASC-HLCs in a nuclear receptor-dependent manner. These data elucidate integrin-Src signalling linking the extrinsic EC matrix signals and metabolic functional maturation of hepatocyte. This study provides a model for studying the interaction between hepatocytes and non-parenchymal cell-derived matrix. © 2017 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.

  4. Programming Amazon EC2

    CERN Document Server

    Vliet, Jurg

    2011-01-01

    If you plan to use Amazon Web Services to run applications in the cloud, the end-to-end approach in this book will save you needless trial and error. You'll find practical guidelines for designing and building applications with Amazon Elastic Compute Cloud (EC2) and a host of supporting AWS tools, with a focus on critical issues such as load balancing, monitoring, and automation. How do you move an existing application to AWS, or design your application so that it scales effectively? How much storage will you require? Programming Amazon EC2 not only helps you get started, it will also keep y

  5. Niche-dependent development of functional neuronal networks from embryonic stem cell-derived neural populations

    Directory of Open Access Journals (Sweden)

    Siebler Mario

    2009-08-01

    Full Text Available Abstract Background The present work was performed to investigate the ability of two different embryonic stem (ES cell-derived neural precursor populations to generate functional neuronal networks in vitro. The first ES cell-derived neural precursor population was cultivated as free-floating neural aggregates which are known to form a developmental niche comprising different types of neural cells, including neural precursor cells (NPCs, progenitor cells and even further matured cells. This niche provides by itself a variety of different growth factors and extracellular matrix proteins that influence the proliferation and differentiation of neural precursor and progenitor cells. The second population was cultivated adherently in monolayer cultures to control most stringently the extracellular environment. This population comprises highly homogeneous NPCs which are supposed to represent an attractive way to provide well-defined neuronal progeny. However, the ability of these different ES cell-derived immature neural cell populations to generate functional neuronal networks has not been assessed so far. Results While both precursor populations were shown to differentiate into sufficient quantities of mature NeuN+ neurons that also express GABA or vesicular-glutamate-transporter-2 (vGlut2, only aggregate-derived neuronal populations exhibited a synchronously oscillating network activity 2–4 weeks after initiating the differentiation as detected by the microelectrode array technology. Neurons derived from homogeneous NPCs within monolayer cultures did merely show uncorrelated spiking activity even when differentiated for up to 12 weeks. We demonstrated that these neurons exhibited sparsely ramified neurites and an embryonic vGlut2 distribution suggesting an inhibited terminal neuronal maturation. In comparison, neurons derived from heterogeneous populations within neural aggregates appeared as fully mature with a dense neurite network and punctuated

  6. Luminal progenitors restrict their lineage potential during mammary gland development.

    Science.gov (United States)

    Rodilla, Veronica; Dasti, Alessandro; Huyghe, Mathilde; Lafkas, Daniel; Laurent, Cécile; Reyal, Fabien; Fre, Silvia

    2015-02-01

    The hierarchical relationships between stem cells and progenitors that guide mammary gland morphogenesis are still poorly defined. While multipotent basal stem cells have been found within the myoepithelial compartment, the in vivo lineage potential of luminal progenitors is unclear. Here we used the expression of the Notch1 receptor, previously implicated in mammary gland development and tumorigenesis, to elucidate the hierarchical organization of mammary stem/progenitor cells by lineage tracing. We found that Notch1 expression identifies multipotent stem cells in the embryonic mammary bud, which progressively restrict their lineage potential during mammary ductal morphogenesis to exclusively generate an ERαneg luminal lineage postnatally. Importantly, our results show that Notch1-labelled cells represent the alveolar progenitors that expand during pregnancy and survive multiple successive involutions. This study reveals that postnatal luminal epithelial cells derive from distinct self-sustained lineages that may represent the cells of origin of different breast cancer subtypes.

  7. Human Stem Cell Derived Cardiomyocytes: An Alternative ...

    Science.gov (United States)

    Chemical spills and associated deaths in the US has increased 2.6-fold and 16-fold from 1983 to 2012, respectfully. In addition, the number of chemicals to which humans are exposed to in the environment has increased almost 10-fold from 2001 to 2013 within the US. Internationally, a WHO report on the global composite impact of chemicals on health reported that 16% of the total burden of cardiovascular disease was attributed to environmental chemical exposure with 2.5 million deaths per year. Clearly, the cardiovascular system, at all its various developmental and life stages, represents a critical target organ system that can be adversely affected by existing and emerging chemicals (e.g., engineered nanomaterials) in a variety of environmental media. The ability to assess chemical cardiac risk and safety is critically needed but extremely challenging due to the number and categories of chemicals in commerce, as indicated. This presentation\\session will evaluate the use of adult human stem cell derived cardiomyocytes, and existing platforms, as an alternative model to evaluate environmental chemical cardiac toxicity as well as provide key information for the development of predictive adverse outcomes pathways associated with environmental chemical exposures. (This abstract does not represent EPA policy) Rapid and translatable chemical safety screening models for cardiotoxicity current status for informing regulatory decisions, a workshop sponsored by the Society

  8. Postnatal stem/progenitor cells derived from the dental pulp of adult chimpanzee

    Directory of Open Access Journals (Sweden)

    Fillos Dimitri

    2008-04-01

    Full Text Available Background Chimpanzee dental pulp stem/stromal cells (ChDPSCs are very similar to human bone marrow derived mesenchymal stem/stromal cells (hBMSCs as demonstrated by the expression pattern of cell surface markers and their multipotent differentiation capability. Results ChDPSCs were isolated from an incisor and a canine of a forty-seven year old female chimpanzee. A homogenous population of ChDPSCs was established in early culture at a high proliferation rate and verified by the expression pattern of thirteen cell surface markers. The ChDPSCs are multipotent and were capable of differentiating into osteogenic, adipogenic and chondrogenic lineages under appropriate in vitro culture conditions. ChDPSCs also express stem cell (Sox-2, Nanog, Rex-1, Oct-4 and osteogenic (Osteonectin, osteocalcin, osteopontin markers, which is comparable to reported results of rhesus monkey BMSCs (rBMSCs, hBMSCs and hDPSCs. Although ChDPSCs vigorously proliferated during the initial phase and gradually decreased in subsequent passages, the telomere length indicated that telomerase activity was not significantly reduced. Conclusion These results demonstrate that ChDPSCs can be efficiently isolated from post-mortem teeth of adult chimpanzees and are multipotent. Due to the almost identical genome composition of humans and chimpanzees, there is an emergent need for defining the new role of chimpanzee modeling in comparative medicine. Teeth are easy to recover at necropsy and easy to preserve prior to the retrieval of dental pulp for stem/stromal cells isolation. Therefore, the establishment of ChDPSCs would preserve and maximize the applications of such a unique and invaluable animal model, and could advance the understanding of cellular functions and differentiation control of adult stem cells in higher primates.

  9. Postnatal stem/progenitor cells derived from the dental pulp of adult chimpanzee.

    Science.gov (United States)

    Cheng, Pei-Hsun; Snyder, Brooke; Fillos, Dimitri; Ibegbu, Chris C; Huang, Anderson Hsien-Cheng; Chan, Anthony W S

    2008-04-22

    Chimpanzee dental pulp stem/stromal cells (ChDPSCs) are very similar to human bone marrow derived mesenchymal stem/stromal cells (hBMSCs) as demonstrated by the expression pattern of cell surface markers and their multipotent differentiation capability. ChDPSCs were isolated from an incisor and a canine of a forty-seven year old female chimpanzee. A homogenous population of ChDPSCs was established in early culture at a high proliferation rate and verified by the expression pattern of thirteen cell surface markers. The ChDPSCs are multipotent and were capable of differentiating into osteogenic, adipogenic and chondrogenic lineages under appropriate in vitro culture conditions. ChDPSCs also express stem cell (Sox-2, Nanog, Rex-1, Oct-4) and osteogenic (Osteonectin, osteocalcin, osteopontin) markers, which is comparable to reported results of rhesus monkey BMSCs (rBMSCs), hBMSCs and hDPSCs. Although ChDPSCs vigorously proliferated during the initial phase and gradually decreased in subsequent passages, the telomere length indicated that telomerase activity was not significantly reduced. These results demonstrate that ChDPSCs can be efficiently isolated from post-mortem teeth of adult chimpanzees and are multipotent. Due to the almost identical genome composition of humans and chimpanzees, there is an emergent need for defining the new role of chimpanzee modeling in comparative medicine. Teeth are easy to recover at necropsy and easy to preserve prior to the retrieval of dental pulp for stem/stromal cells isolation. Therefore, the establishment of ChDPSCs would preserve and maximize the applications of such a unique and invaluable animal model, and could advance the understanding of cellular functions and differentiation control of adult stem cells in higher primates.

  10. Use of RUNX2 Expression to Identify Osteogenic Progenitor Cells Derived from Human Embryonic Stem Cells

    Science.gov (United States)

    Zou, Li; Kidwai, Fahad K.; Kopher, Ross A.; Motl, Jason; Kellum, Cory A.; Westendorf, Jennifer J.; Kaufman, Dan S.

    2015-01-01

    Summary We generated a RUNX2-yellow fluorescent protein (YFP) reporter system to study osteogenic development from human embryonic stem cells (hESCs). Our studies demonstrate the fidelity of YFP expression with expression of RUNX2 and other osteogenic genes in hESC-derived osteoprogenitor cells, as well as the osteogenic specificity of YFP signal. In vitro studies confirm that the hESC-derived YFP+ cells have similar osteogenic phenotypes to osteoprogenitor cells generated from bone-marrow mesenchymal stem cells. In vivo studies demonstrate the hESC-derived YFP+ cells can repair a calvarial defect in immunodeficient mice. Using the engineered hESCs, we monitored the osteogenic development and explored the roles of osteogenic supplements BMP2 and FGF9 in osteogenic differentiation of these hESCs in vitro. Taken together, this reporter system provides a novel system to monitor the osteogenic differentiation of hESCs and becomes useful to identify soluble agents and cell signaling pathways that mediate early stages of human bone development. PMID:25680477

  11. Use of RUNX2 Expression to Identify Osteogenic Progenitor Cells Derived from Human Embryonic Stem Cells

    Directory of Open Access Journals (Sweden)

    Li Zou

    2015-02-01

    Full Text Available We generated a RUNX2-yellow fluorescent protein (YFP reporter system to study osteogenic development from human embryonic stem cells (hESCs. Our studies demonstrate the fidelity of YFP expression with expression of RUNX2 and other osteogenic genes in hESC-derived osteoprogenitor cells, as well as the osteogenic specificity of YFP signal. In vitro studies confirm that the hESC-derived YFP+ cells have similar osteogenic phenotypes to osteoprogenitor cells generated from bone-marrow mesenchymal stem cells. In vivo studies demonstrate the hESC-derived YFP+ cells can repair a calvarial defect in immunodeficient mice. Using the engineered hESCs, we monitored the osteogenic development and explored the roles of osteogenic supplements BMP2 and FGF9 in osteogenic differentiation of these hESCs in vitro. Taken together, this reporter system provides a novel system to monitor the osteogenic differentiation of hESCs and becomes useful to identify soluble agents and cell signaling pathways that mediate early stages of human bone development.

  12. Decellularized Tissue and Cell-Derived Extracellular Matrices as Scaffolds for Orthopaedic Tissue Engineering

    Science.gov (United States)

    Cheng, Christina W.; Solorio, Loran D.; Alsberg, Eben

    2014-01-01

    The reconstruction of musculoskeletal defects is a constant challenge for orthopaedic surgeons. Musculoskeletal injuries such as fractures, chondral lesions, infections and tumor debulking can often lead to large tissue voids requiring reconstruction with tissue grafts. Autografts are currently the gold standard in orthopaedic tissue reconstruction; however, there is a limit to the amount of tissue that can be harvested before compromising the donor site. Tissue engineering strategies using allogeneic or xenogeneic decellularized bone, cartilage, skeletal muscle, tendon and ligament have emerged as promising potential alternative treatment. The extracellular matrix provides a natural scaffold for cell attachment, proliferation and differentiation. Decellularization of in vitro cell-derived matrices can also enable the generation of autologous constructs from tissue specific cells or progenitor cells. Although decellularized bone tissue is widely used clinically in orthopaedic applications, the exciting potential of decellularized cartilage, skeletal muscle, tendon and ligament cell-derived matrices has only recently begun to be explored for ultimate translation to the orthopaedic clinic. PMID:24417915

  13. Generation and purification of human stem cell-derived cardiomyocytes

    NARCIS (Netherlands)

    Schwach, Verena; Passier, Robert

    2016-01-01

    © 2016 International Society of Differentiation Efficient and reproducible generation and purification of human stem cell-derived cardiomyocytes (CMs) is crucial for regenerative medicine, disease modeling, drug screening and study of developmental events during cardiac specification. Established

  14. EC

    International Nuclear Information System (INIS)

    Anon.

    1992-01-01

    This paper reports on what is being called the Single Europe, a natural evolution to its ultimate of what was once known as simply the European Common Market. So comprehensive is its scope that it will ultimately affect the design of appliances, the siting of tanks, the design of tank trucks, and the creation of harmonized safety rules across the spectrum of the 12-nation membership. Admittedly, the LPG industry is one of the smaller commercial entities in what is now called the European Community, but its members will share in the impact of the revolution that will be felt throughout the economic and social fabric of the member countries

  15. Structural phenotyping of stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Pasqualini, Francesco Silvio; Sheehy, Sean Paul; Agarwal, Ashutosh; Aratyn-Schaus, Yvonne; Parker, Kevin Kit

    2015-03-10

    Structural phenotyping based on classical image feature detection has been adopted to elucidate the molecular mechanisms behind genetically or pharmacologically induced changes in cell morphology. Here, we developed a set of 11 metrics to capture the increasing sarcomere organization that occurs intracellularly during striated muscle cell development. To test our metrics, we analyzed the localization of the contractile protein α-actinin in a variety of primary and stem-cell derived cardiomyocytes. Further, we combined these metrics with data mining algorithms to unbiasedly score the phenotypic maturity of human-induced pluripotent stem cell-derived cardiomyocytes. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  16. Nuclear power in the EC

    International Nuclear Information System (INIS)

    Charrault, J.C.

    1991-01-01

    Nuclear power accounts for some 35% of electricity production in the European Community (EC). Using a mathematical analysis, based on different scenarios, i.e. low/high electricity demand and nuclear moratorium/revival, various demand forecasts are made. A pragmatic approach, considering conventional power generation pollution problems, forecasts a revival of nuclear power

  17. EC6 safety design improvements

    Energy Technology Data Exchange (ETDEWEB)

    Yu, S.; Lee, A.G.; Soulard, M. [Candu Energy Inc., Mississauga, ON (Canada)

    2014-07-01

    The Enhanced CANDU 6 (EC6) builds on the proven high performance design such as the Qinshan CANDU 6 reactor, and has made improvements to safety, operational performance, and has incorporated extensive operational feedback. Completion of all three phases of the pre-licensing design review by the Canadian Regulator - the Canadian Nuclear Safety Commission has provided a higher level of assurance that the EC6 reference design has taken modern regulatory requirements and expectations into account and further confirmed that there are no fundamental barriers to licensing the EC6 design in Canada. The EC6 design is based on the defence-in-depth principles in INSAG-10 and provides further safety features that address the lessons learned from Fukushima. With these safety features, the EC6 design has strengthened accident prevention as the first priority in the defence-in-depth strategy, as outlined in INSAG-10. As well, the EC6 design has incorporated further mitigation measures to provide additional protection of the public and the environment if the preventive measures fail. The EC6 design has an appropriate combination of inherent, passive safety characteristics, engineered features and administrative safety measures to effectively prevent and mitigate severe accident progressions. A strong contributor to the robustness and redundancy of CANDU design is the two-group separation philosophy. This ensures a high degree of independence between safety systems as well as physical separation and functional independence in how fundamental safety functions are provided. This paper will describe the following safety features based on the application of defence-in-depth and design approach to prevent beyond design basis events progressing to severe accidents and to mitigate the consequences if it occurs: Improved steam generator heat sink via a more reliable emergency heat removal system; Increased time before manual field actions are required via enhanced capacity of

  18. Translational applications of adult stem cell-derived organoids

    NARCIS (Netherlands)

    Drost, Jarno; Clevers, Hans

    2017-01-01

    Adult stem cells from a variety of organs can be expanded long-term in vitro as three-dimensional organotypic structures termed organoids. These adult stem cell-derived organoids retain their organ identity and remain genetically stable over long periods of time. The ability to grow organoids from

  19. Glial progenitor cell-based treatment of the childhood leukodystrophies

    DEFF Research Database (Denmark)

    Osório, M. Joana; Goldman, Steven A.

    2016-01-01

    stem cell-derived human neural or glial progenitor cells may comprise a promising strategy for both structural remyelination and metabolic rescue. A broad variety of pediatric white matter disorders, including the primary hypomyelinating disorders, the lysosomal storage disorders, and the broader group...... genetic editing of pluripotent stem cells. Yet these challenges notwithstanding, the promise of glial progenitor cell-based treatment of the childhood myelin disorders offers hope to the many victims of this otherwise largely untreatable class of disease....... and astrocytes are the major affected cell populations, and are either structurally impaired or metabolically compromised through cell-intrinsic pathology, or are the victims of mis-accumulated toxic byproducts of metabolic derangement. In either case, glial cell replacement using implanted tissue or pluripotent...

  20. Differentiation and molecular profiling of human embryonic stem cell-derived corneal epithelial cells.

    Science.gov (United States)

    Brzeszczynska, J; Samuel, K; Greenhough, S; Ramaesh, K; Dhillon, B; Hay, D C; Ross, J A

    2014-06-01

    It has been suggested that the isolation of scalable populations of limbal stem cells may lead to radical changes in ocular therapy. In particular, the derivation and transplantation of corneal stem cells from these populations may result in therapies providing clinical normality of the diseased or damaged cornea. Although feasible in theory, the lack of donor material in sufficient quantity and quality currently limits such a strategy. A potential scalable source of corneal cells could be derived from pluripotent stem cells (PSCs). We developed an in vitro and serum-free corneal differentiation model which displays significant promise. Our stepwise differentiation model was designed with reference to development and gave rise to cells which displayed similarities to epithelial progenitor cells which can be specified to cells displaying a corneal epithelial phenotype. We believe our approach is novel, provides a robust model of human development and in the future, may facilitate the generation of corneal epithelial cells that are suitable for clinical use. Additionally, we demonstrate that following continued cell culture, stem cell-derived corneal epithelial cells undergo transdifferentiation and exhibit squamous metaplasia and therefore, also offer an in vitro model of disease.

  1. Circulating cell-derived microparticles in women with pregnancy loss.

    Science.gov (United States)

    Alijotas-Reig, Jaume; Palacio-Garcia, Carles; Farran-Codina, Immaculada; Zarzoso, Cristina; Cabero-Roura, Luis; Vilardell-Tarres, Miquel

    2011-09-01

    To analyze cell-derived microparticles (cMP) in pregnancy loss (PL), both recurrent miscarriages (RM) and unexplained fetal loss (UFL). Non-matched case-control study was performed at Vall d'Hebron Hospital. Cell-derived microparticles of 53 PL cases, 30 with RM, 16 with UFL, and 7 (RM + UFL), were compared to 38 healthy pregnant women. Twenty healthy non-pregnant women act as controls. Cell-derived microparticles were analyzed through flow cytometry. Results are given as total annexin (A5+), endothelial-(CD144+/CD31+ CD41-), platelet-(CD41+), leukocyte-(CD45+) and CD41- c-MP/μL of plasma. Antiphospholipid antibodies (aPLA) were analyzed according to established methods. Comparing PL versus healthy pregnant, we observed a significant endothelial cMP decrease in PL. When comparing RM subgroup with controls, we observed significant decreases in endothelial cMP. When comparing the PL positive for aPLA versus PL-aPLA-negative, no cMP numbering differences were seen. Pregnancy loss seems to be related to endothelial cell activation and/or consumption. A relationship between aPLA and cMP could not be demonstrated. © 2011 John Wiley & Sons A/S.

  2. In vitro generation of three-dimensional substrate-adherent embryonic stem cell-derived neural aggregates for application in animal models of neurological disorders.

    Science.gov (United States)

    Hargus, Gunnar; Cui, Yi-Fang; Dihné, Marcel; Bernreuther, Christian; Schachner, Melitta

    2012-05-01

    In vitro-differentiated embryonic stem (ES) cells comprise a useful source for cell replacement therapy, but the efficiency and safety of a translational approach are highly dependent on optimized protocols for directed differentiation of ES cells into the desired cell types in vitro. Furthermore, the transplantation of three-dimensional ES cell-derived structures instead of a single-cell suspension may improve graft survival and function by providing a beneficial microenvironment for implanted cells. To this end, we have developed a new method to efficiently differentiate mouse ES cells into neural aggregates that consist predominantly (>90%) of postmitotic neurons, neural progenitor cells, and radial glia-like cells. When transplanted into the excitotoxically lesioned striatum of adult mice, these substrate-adherent embryonic stem cell-derived neural aggregates (SENAs) showed significant advantages over transplanted single-cell suspensions of ES cell-derived neural cells, including improved survival of GABAergic neurons, increased cell migration, and significantly decreased risk of teratoma formation. Furthermore, SENAs mediated functional improvement after transplantation into animal models of Parkinson's disease and spinal cord injury. This unit describes in detail how SENAs are efficiently derived from mouse ES cells in vitro and how SENAs are isolated for transplantation. Furthermore, methods are presented for successful implantation of SENAs into animal models of Huntington's disease, Parkinson's disease, and spinal cord injury to study the effects of stem cell-derived neural aggregates in a disease context in vivo.

  3. Single-cell-derived mesenchymal stem cells overexpressing Csx/Nkx2.5 and GATA4 undergo the stochastic cardiomyogenic fate and behave like transient amplifying cells

    International Nuclear Information System (INIS)

    Yamada, Yoji; Sakurada, Kazuhiro; Takeda, Yukiji; Gojo, Satoshi; Umezawa, Akihiro

    2007-01-01

    Bone marrow-derived stromal cells can give rise to cardiomyocytes as well as adipocytes, osteocytes, and chondrocytes in vitro. The existence of mesenchymal stem cells has been proposed, but it remains unclear if a single-cell-derived stem cell stochastically commits toward a cardiac lineage. By single-cell marking, we performed a follow-up study of individual cells during the differentiation of 9-15c mesenchymal stromal cells derived from bone marrow cells. Three types of cells, i.e., cardiac myoblasts, cardiac progenitors and multipotent stem cells were differentiated from a single cell, implying that cardiomyocytes are generated stochastically from a single-cell-derived stem cell. We also demonstrated that overexpression of Csx/Nkx2.5 and GATA4, precardiac mesodermal transcription factors, enhanced cardiomyogenic differentiation of 9-15c cells, and the frequency of cardiomyogenic differentiation was increased by co-culturing with fetal cardiomyocytes. Single-cell-derived mesenchymal stem cells overexpressing Csx/Nkx2.5 and GATA4 behaved like cardiac transient amplifying cells, and still retained their plasticity in vivo

  4. Retinal vascular injuries and intravitreal human embryonic stem cell-derived haemangioblasts.

    Science.gov (United States)

    Wang, Jin-Da; An, Ying; Zhang, Jing-Shang; Wan, Xiu-Hua; Zhang, Wei; Lanza, Robert; Lu, Shi-Jiang; Jonas, Jost B; Xu, Liang

    2017-09-01

    To investigate whether intravitreally applied haemangioblasts (HB) derived from human embryonic stem cells (hESCs) are helpful for the repair of vascular damage caused in animals by an oxygen-induced retinopathy (OIR), by an induced diabetic retinopathy (DR) or by an induced retinal ischaemia with subsequent reperfusion. Human embryonic stem cell-derived HBs were transplanted intravitreally into C57BL/6J mice (OIR model), into male Wistar rats with an induced DR and into male Wistar rats undergoing induced retinal ischaemia with subsequent reperfusion. Control groups of animals received an intravitreal injection of endothelial cells (ECs) or phosphate-buffered saline (PBS). We examined the vasculature integrity in the mice with OIR, the blood-retina barrier in the rats with induced DR, and retinal thickness and retinal ganglion cell density in retina flat mounts of the rats with the retinal ischaemic-reperfusion retinopathy. In the OIR model, the study group versus control groups showed a significantly (p < 0.001) smaller retinal avascular area [5.1 ± 2.7%;n = 18 animals versus 12.2 ± 2.8% (PBS group; n = 10 animals) and versus 11.8 ± 3.7% (EC group; n = 8 animals)] and less retinal neovascularization [6.3 ± 2.5%;n = 18 versus 15.2 ± 6.3% (n = 10; PBS group) and versus 15.8 ± 3.3% (n = 8; EC group)]. On retinal flat mounts, hESC-HBs were integrated into damaged retinal vessels and stained positive for PECAM (CD31) as EC marker. In the DR model, the study group versus the EC control group showed a significantly (p = 0.001) better blood-retina barrier function as measured at 2 days after the intravitreal injections [study group: 20.2 ± 12.8 μl/(g × hr); n = 6; versus EC control group: 52.9 ± 9.9 μl/(g × hr; n = 6)]. In the retinal ischaemia-reperfusion model, the groups did not differ significantly in retinal thickness and retinal ganglion cell density at 2, 5 and 7 days after baseline. By integrating into

  5. Thrombin-specific inactivation of endothelial cell derived plasminogen activator

    International Nuclear Information System (INIS)

    Highsmith, R.F.; Gallaher, M.J.

    1986-01-01

    Although thrombin (T) has diverse functions in the overall hemostatic mechanism, relatively little is known about its direct effect on components of the fibrinolytic enzyme system. The authors have investigated the interaction of T with plasminogen activators (PA) derived from bovine aortic endothelial cells (EC) in culture (2-5th passage, preconfluent monolayers). Varying concentrations of purified bovine or human thrombin were added to EC-conditioned media (CM). CM + T mixtures were assayed at various times for PA activity using purified plasminogen and a sensitive 125 I-fibrinogenolytic or caseinolytic assay. T (5 nM), but not plasmin or trypsin at equivalent concentrations, resulted in a time-dependent inhibition of the PA activity in CM. T had no effect on the PA activity of urokinase, streptokinase or preformed plasmin. The ability of T to inactivate the EC-derived PA was abolished by prior treatment of T with active site-directed reagents. SDS-PAGE and zymography with copolymerized fibrinogen and plasminogen revealed further specificity in that only one of the multiple-molecular weight forms of PA present in EC-CM was inactivated by T. The authors conclude that in a highly specific fashion, T inactivates the predominant PA present in EC-CM by limited proteolysis. Thus, another potentially important function of T is suggested which may have particular significance in the temporal regulation of coagulation and fibrinolysis at the blood-endothelium interface

  6. Thrombin-specific inactivation of endothelial cell derived plasminogen activator

    Energy Technology Data Exchange (ETDEWEB)

    Highsmith, R.F.; Gallaher, M.J.

    1986-03-05

    Although thrombin (T) has diverse functions in the overall hemostatic mechanism, relatively little is known about its direct effect on components of the fibrinolytic enzyme system. The authors have investigated the interaction of T with plasminogen activators (PA) derived from bovine aortic endothelial cells (EC) in culture (2-5th passage, preconfluent monolayers). Varying concentrations of purified bovine or human thrombin were added to EC-conditioned media (CM). CM + T mixtures were assayed at various times for PA activity using purified plasminogen and a sensitive /sup 125/I-fibrinogenolytic or caseinolytic assay. T (5 nM), but not plasmin or trypsin at equivalent concentrations, resulted in a time-dependent inhibition of the PA activity in CM. T had no effect on the PA activity of urokinase, streptokinase or preformed plasmin. The ability of T to inactivate the EC-derived PA was abolished by prior treatment of T with active site-directed reagents. SDS-PAGE and zymography with copolymerized fibrinogen and plasminogen revealed further specificity in that only one of the multiple-molecular weight forms of PA present in EC-CM was inactivated by T. The authors conclude that in a highly specific fashion, T inactivates the predominant PA present in EC-CM by limited proteolysis. Thus, another potentially important function of T is suggested which may have particular significance in the temporal regulation of coagulation and fibrinolysis at the blood-endothelium interface.

  7. Linking incomplete reprogramming to the improved pluripotency of murine embryonal carcinoma cell-derived pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Gang Chang

    Full Text Available Somatic cell nuclear transfer (SCNT has been proved capable of reprogramming various differentiated somatic cells into pluripotent stem cells. Recently, induced pluripotent stem cells (iPS have been successfully derived from mouse and human somatic cells by the over-expression of a combination of transcription factors. However, the molecular mechanisms underlying the reprogramming mediated by either the SCNT or iPS approach are poorly understood. Increasing evidence indicates that many tumor pathways play roles in the derivation of iPS cells. Embryonal carcinoma (EC cells have the characteristics of both stem cells and cancer cells and thus they might be the better candidates for elucidating the details of the reprogramming process. Although previous studies indicate that EC cells cannot be reprogrammed into real pluripotent stem cells, the reasons for this remain unclear. Here, nuclei from mouse EC cells (P19 were transplanted into enucleated oocytes and pluripotent stem cells (P19 NTES cells were subsequently established. Interestingly, P19 NTES cells prolonged the development of tetraploid aggregated embryos compared to EC cells alone. More importantly, we found that the expression recovery of the imprinted H19 gene was dependent on the methylation state in the differential methylation region (DMR. The induction of Nanog expression, however, was independent of the promoter region DNA methylation state in P19 NTES cells. A whole-genome transcriptome analysis further demonstrated that P19 NTES cells were indeed the intermediates between P19 cells and ES cells and many interesting genes were uncovered that may be responsible for the failed reprogramming of P19 cells. To our knowledge, for the first time, we linked incomplete reprogramming to the improved pluripotency of EC cell-derived pluripotent stem cells. The candidate genes we discovered may be useful not only for understanding the mechanisms of reprogramming, but also for deciphering the

  8. Characterisation of human induced pluripotent stem cell-derived endothelial cells under shear stress using an easy-to-use microfluidic cell culture system.

    Science.gov (United States)

    Ohtani-Kaneko, Rsituko; Sato, Kenjiro; Tsutiya, Atsuhiro; Nakagawa, Yuka; Hashizume, Kazutoshi; Tazawa, Hidekatsu

    2017-10-09

    Induced pluripotent stem cell-derived endothelial cells (iPSC-ECs) can contribute to elucidating the pathogenesis of heart and vascular diseases and developing their treatments. Their precise characteristics in fluid flow however remain unclear. Therefore, the aim of the present study is to characterise these features. We cultured three types of ECs in a microfluidic culture system: commercially available human iPS-ECs, human umbilical vein endothelial cells (HUVECs) and human umbilical artery endothelial cells (HUAECs). We then examined the mRNA expression levels of endothelial marker gene cluster of differentiation 31 (CD31), fit-related receptor tyrosine kinase (Flk-1), and the smooth muscle marker gene smooth muscle alpha-actin, and investigated changes in plasminogen activator inhibitor-1 (PAI-1) secretion and intracellular F-actin arrangement following heat stress. We also compared expressions of the arterial and venous marker genes ephrinB2 and EphB4, and the endothelial gap junction genes connexin (Cx) 37, 40, and 43 under fluidic shear stress to determine their arterial or venous characteristics. We found that iPS-ECs had similar endothelial marker gene expressions and exhibited similar increases in PAI-1 secretion under heat stress as HUVECs and HUAECs. In addition, F-actin arrangement in iPSC-ECs also responded to heat stress, as previously reported. However, they had different expression patterns of arterial and venous marker genes and Cx genes under different fluidic shear stress levels, showing that iPSC-ECs exhibit different characteristics from arterial and venous ECs. This microfluidic culture system equipped with variable shear stress control will provide an easy-to-use assay tool to examine characteristics of iPS-ECs generated by different protocols in various laboratories and contribute to basic and applied biomedical researches on iPS-ECs.

  9. Β-amyloid 1-42 oligomers impair function of human embryonic stem cell-derived forebrain cholinergic neurons.

    Directory of Open Access Journals (Sweden)

    Linn Wicklund

    Full Text Available Cognitive impairment in Alzheimer's disease (AD patients is associated with a decline in the levels of growth factors, impairment of axonal transport and marked degeneration of basal forebrain cholinergic neurons (BFCNs. Neurogenesis persists in the adult human brain, and the stimulation of regenerative processes in the CNS is an attractive prospect for neuroreplacement therapy in neurodegenerative diseases such as AD. Currently, it is still not clear how the pathophysiological environment in the AD brain affects stem cell biology. Previous studies investigating the effects of the β-amyloid (Aβ peptide on neurogenesis have been inconclusive, since both neurogenic and neurotoxic effects on progenitor cell populations have been reported. In this study, we treated pluripotent human embryonic stem (hES cells with nerve growth factor (NGF as well as with fibrillar and oligomeric Aβ1-40 and Aβ1-42 (nM-µM concentrations and thereafter studied the differentiation in vitro during 28-35 days. The process applied real time quantitative PCR, immunocytochemistry as well as functional studies of intracellular calcium signaling. Treatment with NGF promoted the differentiation into functionally mature BFCNs. In comparison to untreated cells, oligomeric Aβ1-40 increased the number of functional neurons, whereas oligomeric Aβ1-42 suppressed the number of functional neurons. Interestingly, oligomeric Aβ exposure did not influence the number of hES cell-derived neurons compared with untreated cells, while in contrast fibrillar Aβ1-40 and Aβ1-42 induced gliogenesis. These findings indicate that Aβ1-42 oligomers may impair the function of stem cell-derived neurons. We propose that it may be possible for future AD therapies to promote the maturation of functional stem cell-derived neurons by altering the brain microenvironment with trophic support and by targeting different aggregation forms of Aβ.

  10. Progenitors of white dwarfs

    International Nuclear Information System (INIS)

    Drilling, J.S.; Schoenberner, D.

    1985-01-01

    Direct observational evidence is presented which indicates that the immediate progenitors of white dwarfs are the central stars of planetary nebulae (approximately 70%), other post-AGB objects (approximately 30%), and post-HB objects not massive enough to climb the AGB (approximately 0.3%). The combined birth rate for these objects is in satisfactory agreement with the death rate of main-sequence stars and the birth rate of white dwarfs

  11. Neural stem cell-derived exosomes mediate viral entry

    Directory of Open Access Journals (Sweden)

    Sims B

    2014-10-01

    Full Text Available Brian Sims,1,2,* Linlin Gu,3,* Alexandre Krendelchtchikov,3 Qiana L Matthews3,4 1Division of Neonatology, Department of Pediatrics, 2Department of Cell, Developmental, and Integrative Biology, 3Division of Infectious Diseases, Department of Medicine, 4Center for AIDS Research, University of Alabama at Birmingham, Birmingham, AL, USA *These authors contributed equally to this work Background: Viruses enter host cells through interactions of viral ligands with cellular receptors. Viruses can also enter cells in a receptor-independent fashion. Mechanisms regarding the receptor-independent viral entry into cells have not been fully elucidated. Exosomal trafficking between cells may offer a mechanism by which viruses can enter cells.Methods: To investigate the role of exosomes on cellular viral entry, we employed neural stem cell-derived exosomes and adenovirus type 5 (Ad5 for the proof-of-principle study. Results: Exosomes significantly enhanced Ad5 entry in Coxsackie virus and adenovirus receptor (CAR-deficient cells, in which Ad5 only had very limited entry. The exosomes were shown to contain T-cell immunoglobulin mucin protein 4 (TIM-4, which binds phosphatidylserine. Treatment with anti-TIM-4 antibody significantly blocked the exosome-mediated Ad5 entry.Conclusion: Neural stem cell-derived exosomes mediated significant cellular entry of Ad5 in a receptor-independent fashion. This mediation may be hampered by an antibody specifically targeting TIM-4 on exosomes. This set of results will benefit further elucidation of virus/exosome pathways, which would contribute to reducing natural viral infection by developing therapeutic agents or vaccines. Keywords: neural stem cell-derived exosomes, adenovirus type 5, TIM-4, viral entry, phospholipids

  12. Circulating red cell-derived microparticles in human malaria.

    Science.gov (United States)

    Nantakomol, Duangdao; Dondorp, Arjen M; Krudsood, Srivicha; Udomsangpetch, Rachanee; Pattanapanyasat, Kovit; Combes, Valery; Grau, Georges E; White, Nicholas J; Viriyavejakul, Parnpen; Day, Nicholas P J; Chotivanich, Kesinee

    2011-03-01

    In patients with falciparum malaria, plasma concentrations of cell-derived microparticles correlate with disease severity. Using flow cytometry, we quantified red blood cell-derived microparticles (RMPs) in patients with malaria and identified the source and the factors associated with production. RMP concentrations were increased in patients with Plasmodium falciparum (n = 29; median, 457 RMPs/μL [range, 13-4,342 RMPs/μL]), Plasmodium vivax (n = 5; median, 409 RMPs/μL [range, 281-503/μL]), and Plasmodium malariae (n = 2; median, 163 RMPs/μL [range, 127-200 RMPs/μL]) compared with those in healthy subjects (n = 11; median, 8 RMPs/μL [range, 3-166 RMPs/μL]; P = .01). RMP concentrations were highest in patients with severe falciparum malaria (P = .01). Parasitized red cells produced >10 times more RMPs than did unparasitized cells, but the overall majority of RMPs still derived from uninfected red blood cells (URBCs). In cultures, RMP production increased as the parasites matured. Hemin and parasite products induced RMP production in URBCs, which was inhibited by N-acetylcysteine, suggesting heme-mediated oxidative stress as a pathway for the generation of RMPs.

  13. Activity-dependent long-term plasticity of afferent synapses on grafted stem/progenitor cell-derived neurons

    DEFF Research Database (Denmark)

    Sørensen, Andreas Toft; Rogelius, Nina; Lundberg, Cecilia

    2011-01-01

    Stem cell-based cell replacement therapies aiming at restoring injured or diseased brain function ultimately rely on the capability of transplanted cells to promote functional recovery. The mechanisms by which stem cell-based therapies for neurological conditions can lead to functional recovery...

  14. The Postischemic Environment Differentially Impacts Teratoma or Tumor Formation After Transplantation of Human Embryonic Stem Cell-Derived Neural Progenitors

    Czech Academy of Sciences Publication Activity Database

    Seminatore, CH.; Polentes, J.; Ellman, D.; Kozubenko, Nataliya; Itier, V.; Tine, S.; Tritschler, L.; Brenot, M.; Guidou, E.; Blondeau, J.; Lhuillier, M.; Bugi, A.; Aubry, L.; Jendelová, Pavla; Syková, Eva; Perrier, A. L.; Finsen, B.; Onteniente, B.

    2010-01-01

    Roč. 41, č. 1 (2010), s. 153-159 ISSN 0039-2499 Institutional research plan: CEZ:AV0Z50390703 Keywords : brain transplantation * human embryonic stem cells * neural differentiation Subject RIV: FH - Neurology Impact factor: 5.756, year: 2010

  15. Embryonic stem cell-derived microvesicles reprogram hematopoietic progenitors: evidence for horizontal transfer of mRNA and protein delivery

    Czech Academy of Sciences Publication Activity Database

    Ratajczak, J.; Miekus, K.; Kucia, M.; Zhang, J.; Reca, R.; Dvořák, Petr; Rtajczak, M.Z.

    2006-01-01

    Roč. 20, - (2006), s. 847-856 ISSN 0887-6924 R&D Projects: GA ČR GA301/03/1122 Institutional research plan: CEZ:AV0Z50390512 Keywords : Microvesicles * Embryonic stem cells * Stem cell expansion Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 6.146, year: 2006

  16. Transcriptome profile and cytogenetic analysis of immortalized neuronally restricted progenitor cells derived from the porcine olfactory bulb

    Science.gov (United States)

    Recently, we established and phenotypically characterized an immortalized porcine olfactory bulb neuroblast cell line, OBGF400 (Uebing-Czipura et al., 2008). To facilitate the future application of these cells in studies of neurological dysfunction and neuronal replacement therapies, a comprehensive...

  17. Special report: EC oil market

    International Nuclear Information System (INIS)

    Anon.

    1992-01-01

    A European Commission report on the EC's oil market has conceded that the Community will not meet its official objective of reducing oil's share of energy consumption to 40% by 1995. The paper, a 'Communication' to the European Council entitled ''The Oil Market and the Refining Industry in the Community: Recent Developments and Prospects'' says oil will ''continue to account for a major share - of the order of 45% -of the Community's energy consumption'' up to the year 2000. Nonetheless, the report's authors insist the Commission has proposed ''a number of measures'' which could reduce the potential consumption of fossil fuels and help limit CO2 emissions. The report confirms that though lower in 1990 than 1980 at 530mt, oil demand picked up in the second half of the 1980s, reflecting world trends. Little impression has been made on the proportion of the Community's crude supply that is imported. However, one area where the report did find energy objectives successes was that of diversifying sources of crude oil supply. A major source of concern for the Commission at one stage in the second half of the 1980s was the possibility of massive imports of finished oil products from refineries in the oil producing countries. However, the fact that this threat did not materialise is taken as a vindication of the Commission's assessment in previous analyses that ''an open Community market should be maintained''. (author)

  18. Masses of supernova progenitors

    International Nuclear Information System (INIS)

    Tinsley, B.M.

    1977-01-01

    The possible nature and masses of supernovae progenitors, and the bearing of empirical results on some unsolved theoretical problems concerning the origin of supernovae, are discussed. The author concentrates on two main questions: what is the lower mass limit for stars to die explosively and what stars initiate type I supernovae. The evidence considered includes local supernova rates, empirical estimates of msub(w) (the upper mass limit for death as a white dwarf), the distributions of supernovae among stellar populations in galaxies and the colors of supernova producing galaxies. (B.D.)

  19. Multipotent embryonic isl1+ progenitor cells lead to cardiac, smooth muscle, and endothelial cell diversification.

    Science.gov (United States)

    Moretti, Alessandra; Caron, Leslie; Nakano, Atsushi; Lam, Jason T; Bernshausen, Alexandra; Chen, Yinhong; Qyang, Yibing; Bu, Lei; Sasaki, Mika; Martin-Puig, Silvia; Sun, Yunfu; Evans, Sylvia M; Laugwitz, Karl-Ludwig; Chien, Kenneth R

    2006-12-15

    Cardiogenesis requires the generation of endothelial, cardiac, and smooth muscle cells, thought to arise from distinct embryonic precursors. We use genetic fate-mapping studies to document that isl1(+) precursors from the second heart field can generate each of these diverse cardiovascular cell types in vivo. Utilizing embryonic stem (ES) cells, we clonally amplified a cellular hierarchy of isl1(+) cardiovascular progenitors, which resemble the developmental precursors in the embryonic heart. The transcriptional signature of isl1(+)/Nkx2.5(+)/flk1(+) defines a multipotent cardiovascular progenitor, which can give rise to cells of all three lineages. These studies document a developmental paradigm for cardiogenesis, where muscle and endothelial lineage diversification arises from a single cell-level decision of a multipotent isl1(+) cardiovascular progenitor cell (MICP). The discovery of ES cell-derived MICPs suggests a strategy for cardiovascular tissue regeneration via their isolation, renewal, and directed differentiation into specific mature cardiac, pacemaker, smooth muscle, and endothelial cell types.

  20. Characterization of mesenchymal stem cells derived from equine adipose tissue

    Directory of Open Access Journals (Sweden)

    A.M. Carvalho

    2013-08-01

    Full Text Available Stem cell therapy has shown promising results in tendinitis and osteoarthritis in equine medicine. The purpose of this work was to characterize the adipose-derived mesenchymal stem cells (AdMSCs in horses through (1 the assessment of the capacity of progenitor cells to perform adipogenic, osteogenic and chondrogenic differentiation; and (2 flow cytometry analysis using the stemness related markers: CD44, CD90, CD105 and MHC Class II. Five mixed-breed horses, aged 2-4 years-old were used to collect adipose tissue from the base of the tail. After isolation and culture of AdMSCs, immunophenotypic characterization was performed through flow cytometry. There was a high expression of CD44, CD90 and CD105, and no expression of MHC Class II markers. The tri-lineage differentiation was confirmed by specific staining: adipogenic (Oil Red O, osteogenic (Alizarin Red, and chondrogenic (Alcian Blue. The equine AdMSCs are a promising type of adult progenitor cell for tissue engineering in veterinary medicine.

  1. Balancing Ethical Pros and Cons of Stem Cell Derived Gametes.

    Science.gov (United States)

    Segers, Seppe; Mertes, Heidi; de Wert, Guido; Dondorp, Wybo; Pennings, Guido

    2017-07-01

    In this review we aim to provide an overview of the most important ethical pros and cons of stem cell derived gametes (SCD-gametes), as a contribution to the debate about reproductive tissue engineering. Derivation of gametes from stem cells holds promising applications both for research and for clinical use in assisted reproduction. We explore the ethical issues connected to gametes derived from embryonic stem cells (both patient specific and non-patient specific) as well as those related to gametes derived from induced pluripotent stem cells. The technology of SCD-gametes raises moral concerns of how reproductive autonomy relates to issues of embryo destruction, safety, access, and applications beyond clinical infertility.

  2. Stem-cell-derived products: an FDA update.

    Science.gov (United States)

    Moos, Malcolm

    2008-12-01

    The therapeutic potential of products derived from stem cells of various types has prompted increasing research and development and public attention. Initiation of human clinical trials in the not-too-distant future is now a realistic possibility. It is, therefore, important to weigh the potential benefits against known, theoretical and totally unsuspected risks in light of current knowledge to ensure that subjects participating in these trials are afforded the most reasonable balance possible between potential risks and potential benefits. There are no apparent differences in fundamental, qualitative biological characteristics between stem-cell-derived products and other cellular therapies regulated by the United States Food and Drug Administration (FDA). Existing authorities can, therefore, be applied. Nevertheless, these products do have properties that require careful evaluation.

  3. Delivery of chemotherapeutic drugs in tumour cell-derived microparticles.

    Science.gov (United States)

    Tang, Ke; Zhang, Yi; Zhang, Huafeng; Xu, Pingwei; Liu, Jing; Ma, Jingwei; Lv, Meng; Li, Dapeng; Katirai, Foad; Shen, Guan-Xin; Zhang, Guimei; Feng, Zuo-Hua; Ye, Duyun; Huang, Bo

    2012-01-01

    Cellular microparticles are vesicular plasma membrane fragments with a diameter of 100-1,000 nanometres that are shed by cells in response to various physiological and artificial stimuli. Here we demonstrate that tumour cell-derived microparticles can be used as vectors to deliver chemotherapeutic drugs. We show that tumour cells incubated with chemotherapeutic drugs package these drugs into microparticles, which can be collected and used to effectively kill tumour cells in murine tumour models without typical side effects. We describe several mechanisms involved in this process, including uptake of drug-containing microparticles by tumour cells, synthesis of additional drug-packaging microparticles by these cells that contribute to the cytotoxic effect and the inhibition of drug efflux from tumour cells. This study highlights a novel drug delivery strategy with potential clinical application.

  4. Cell-derived microparticles in haemostasis and vascular medicine.

    Science.gov (United States)

    Burnier, Laurent; Fontana, Pierre; Kwak, Brenda R; Angelillo-Scherrer, Anne

    2009-03-01

    Considerable interest for cell-derived microparticles has emerged, pointing out their essential role in haemostatic response and their potential as disease markers, but also their implication in a wide range of physiological and pathological processes. They derive from different cell types including platelets - the main source of microparticles - but also from red blood cells, leukocytes and endothelial cells, and they circulate in blood. Despite difficulties encountered in analyzing them and disparities of results obtained with a wide range of methods, microparticle generation processes are now better understood. However, a generally admitted definition of microparticles is currently lacking. For all these reasons we decided to review the literature regarding microparticles in their widest definition, including ectosomes and exosomes, and to focus mainly on their role in haemostasis and vascular medicine.

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2013-07-12

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

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

    International Nuclear Information System (INIS)

    Chen, Ying; Wang, Kai; Chandramouli, Gadisetti V.R.; Knott, Jason G.; Leach, Richard

    2013-01-01

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

  7. Cell-derived microparticles promote coagulation after moderate exercise.

    Science.gov (United States)

    Sossdorf, Maik; Otto, Gordon P; Claus, Ralf A; Gabriel, Holger H W; Lösche, Wolfgang

    2011-07-01

    Cell-derived procoagulant microparticles (MP) might be able to contribute to exercise-induced changes in blood hemostasis. This study aimed to examine (i) the concentration and procoagulant activity of cell-derived MP after a moderate endurance exercise and (ii) the differences in the release, clearance, and activity of MP before and after exercise between trained and untrained individuals. All subjects performed a single bout of physical exercise on a bicycle ergometer for 90 min at 80% of their individual anaerobic threshold. MP were identified and quantified by flow cytometry measurements. Procoagulant activity of MP was measured by a prothrombinase activity assay as well as tissue factor-induced fibrin formation in MP-containing plasma. At baseline, no differences were observed for the absolute number and procoagulant activities of MP between trained and untrained subjects. However, trained individuals had a lower number of tissue factor-positive monocyte-derived MP compared with untrained individuals. In trained subjects, exercise induced a significant increase in the number of MP derived from platelets, monocytes, and endothelial cells, with maximum values at 45 min after exercise and returned to basal levels at 2 h after exercise. Untrained subjects revealed a similar increase in platelet-derived MP, but their level was still increased at 2 h after exercise, indicating a reduced clearance compared with trained individuals. Procoagulant activities of MP were increased immediately after exercise and remained elevated up to 2 h after exercise. We conclude that increased levels of MP were found in healthy individuals after an acute bout of exercise, that the amount of circulating MP contributes to an exercise-induced increase of hemostatic potential, and that there were differences in kinetic and dynamic characteristics between trained and untrained individuals.

  8. Hmga2 regulates self-renewal of retinal progenitors.

    Science.gov (United States)

    Parameswaran, Sowmya; Xia, Xiaohuan; Hegde, Ganapati; Ahmad, Iqbal

    2014-11-01

    In vertebrate retina, histogenesis occurs over an extended period. To sustain the temporal generation of diverse cell types, retinal progenitor cells (RPCs) must self-renew. However, self-renewal and regulation of RPCs remain poorly understood. Here, we demonstrate that cell-extrinsic factors coordinate with the epigenetic regulator high-mobility group AT-hook 2 (Hmga2) to regulate self-renewal of late retinal progenitor cells (RPCs). We observed that a small subset of RPCs was capable of clonal propagation and retained multipotentiality of parents in the presence of endothelial cells (ECs), known self-renewal regulators in various stem cell niches. The self-renewing effects, also observed in vivo, involve multiple intercellular signaling pathways, engaging Hmga2. As progenitors exhaust during retinal development, expression of Hmga2 progressively decreases. Analyses of Hmga2-expression perturbation, in vitro and in vivo, revealed that Hmga2 functionally helps to mediate cell-extrinsic influences on late-retinal progenitor self-renewal. Our results provide a framework for integrating the diverse intercellular influences elicited by epigenetic regulators for self-renewal in a dynamic stem cell niche: the developing vertebrate retina. © 2014. Published by The Company of Biologists Ltd.

  9. Leukemia inhibitory factor (LIF) enhances MAP2 + and HUC/D + neurons and influences neurite extension during differentiation of neural progenitors derived from human embryonic stem cells.

    Science.gov (United States)

    Leukemia Inhibitory Factor (L1F), a member of the Interleukin 6 cytokine family, has a role in differentiation of Human Neural Progenitor (hNP) cells in vitro. hNP cells, derived from Human Embryonic Stem (hES) cells, have an unlimited capacity for self-renewal in monolayer cultu...

  10. Dual EC : a standardized back door

    NARCIS (Netherlands)

    Bernstein, D.J.; Lange, T.; Niederhagen, R.F.

    2015-01-01

    Dual EC is an algorithm to compute pseudorandom numbers starting from some random input. Dual EC was standardized by NIST, ANSI, and ISO among other algorithms to generate pseudorandom numbers. For a long time this algorithm was considered suspicious -- the entity designing the algorithm could have

  11. Characterization of transcription factor networks involved in umbilical cord blood CD34+ stem cells-derived erythropoiesis.

    Directory of Open Access Journals (Sweden)

    Biaoru Li

    Full Text Available Fetal stem cells isolated from umbilical cord blood (UCB possess a great capacity for proliferation and differentiation and serve as a valuable model system to study gene regulation. Expanded knowledge of the molecular control of hemoglobin synthesis will provide a basis for rational design of therapies for β-hemoglobinopathies. Transcriptome data are available for erythroid progenitors derived from adult stem cells, however studies to define molecular mechanisms controlling globin gene regulation during fetal erythropoiesis are limited. Here, we utilize UCB-CD34+ stem cells induced to undergo erythroid differentiation to characterize the transcriptome and transcription factor networks (TFNs associated with the γ/β-globin switch during fetal erythropoiesis. UCB-CD34+ stem cells grown in the one-phase liquid culture system displayed a higher proliferative capacity than adult CD34+ stem cells. The γ/β-globin switch was observed after day 42 during fetal erythropoiesis in contrast to adult progenitors where the switch occurred around day 21. To gain insights into transcription factors involved in globin gene regulation, microarray analysis was performed on RNA isolated from UCB-CD34+ cell-derived erythroid progenitors harvested on day 21, 42, 49 and 56 using the HumanHT-12 Expression BeadChip. After data normalization, Gene Set Enrichment Analysis identified transcription factors (TFs with significant changes in expression during the γ/β-globin switch. Forty-five TFs were silenced by day 56 (Profile-1 and 30 TFs were activated by day 56 (Profile-2. Both GSEA datasets were analyzed using the MIMI Cytoscape platform, which discovered TFNs centered on KLF4 and GATA2 (Profile-1 and KLF1 and GATA1 for Profile-2 genes. Subsequent shRNA studies in KU812 leukemia cells and human erythroid progenitors generated from UCB-CD34+ cells supported a negative role of MAFB in γ-globin regulation. The characteristics of erythroblasts derived from UCB-CD34

  12. Immune surveillance properties of human NK cell-derived exosomes.

    Science.gov (United States)

    Lugini, Luana; Cecchetti, Serena; Huber, Veronica; Luciani, Francesca; Macchia, Gianfranco; Spadaro, Francesca; Paris, Luisa; Abalsamo, Laura; Colone, Marisa; Molinari, Agnese; Podo, Franca; Rivoltini, Licia; Ramoni, Carlo; Fais, Stefano

    2012-09-15

    Exosomes are nanovesicles released by normal and tumor cells, which are detectable in cell culture supernatant and human biological fluids, such as plasma. Functions of exosomes released by "normal" cells are not well understood. In fact, several studies have been carried out on exosomes derived from hematopoietic cells, but very little is known about NK cell exosomes, despite the importance of these cells in innate and adaptive immunity. In this paper, we report that resting and activated NK cells, freshly isolated from blood of healthy donors, release exosomes expressing typical protein markers of NK cells and containing killer proteins (i.e., Fas ligand and perforin molecules). These nanovesicles display cytotoxic activity against several tumor cell lines and activated, but not resting, immune cells. We also show that NK-derived exosomes undergo uptake by tumor target cells but not by resting PBMC. Exosomes purified from plasma of healthy donors express NK cell markers, including CD56+ and perforin, and exert cytotoxic activity against different human tumor target cells and activated immune cells as well. The results of this study propose an important role of NK cell-derived exosomes in immune surveillance and homeostasis. Moreover, this study supports the use of exosomes as an almost perfect example of biomimetic nanovesicles possibly useful in future therapeutic approaches against various diseases, including tumors.

  13. Cells derived from young bone marrow alleviate renal aging.

    Science.gov (United States)

    Yang, Hai-Chun; Rossini, Michele; Ma, Li-Jun; Zuo, Yiqin; Ma, Ji; Fogo, Agnes B

    2011-11-01

    Bone marrow-derived stem cells may modulate renal injury, but the effects may depend on the age of the stem cells. Here we investigated whether bone marrow from young mice attenuates renal aging in old mice. We radiated female 12-mo-old 129SvJ mice and reconstituted them with bone marrow cells (BMC) from either 8-wk-old (young-to-old) or 12-mo-old (old-to-old) male mice. Transfer of young BMC resulted in markedly decreased deposition of collagen IV in the mesangium and less β-galactosidase staining, an indicator of cell senescence. These changes paralleled reduced expression of plasminogen activator inhibitor-1 (PAI-1), PDGF-B (PDGF-B), the transdifferentiation marker fibroblast-specific protein-1 (FSP-1), and senescence-associated p16 and p21. Tubulointerstitial and glomerular cells derived from the transplanted BMC did not show β-galactosidase activity, but after 6 mo, there were more FSP-1-expressing bone marrow-derived cells in old-to-old mice compared with young-to-old mice. Young-to-old mice also exhibited higher expression of the anti-aging gene Klotho and less phosphorylation of IGF-1 receptor β. Taken together, these data suggest that young bone marrow-derived cells can alleviate renal aging in old mice. Direct parenchymal reconstitution by stem cells, paracrine effects from adjacent cells, and circulating anti-aging molecules may mediate the aging of the kidney.

  14. Large-scale generation of cell-derived nanovesicles

    Science.gov (United States)

    Jo, W.; Kim, J.; Yoon, J.; Jeong, D.; Cho, S.; Jeong, H.; Yoon, Y. J.; Kim, S. C.; Gho, Y. S.; Park, J.

    2014-09-01

    Exosomes are enclosed compartments that are released from cells and that can transport biological contents for the purpose of intercellular communications. Research into exosomes is hindered by their rarity. In this article, we introduce a device that uses centrifugal force and a filter with micro-sized pores to generate a large quantity of cell-derived nanovesicles. The device has a simple polycarbonate structure to hold the filter, and operates in a common centrifuge. Nanovesicles are similar in size and membrane structure to exosomes. Nanovesicles contain intracellular RNAs ranging from microRNA to mRNA, intracellular proteins, and plasma membrane proteins. The quantity of nanovesicles produced using the device is 250 times the quantity of naturally secreted exosomes. Also, the quantity of intracellular contents in nanovesicles is twice that in exosomes. Nanovesicles generated from murine embryonic stem cells can transfer RNAs to target cells. Therefore, this novel device and the nanovesicles that it generates are expected to be used in exosome-related research, and can be applied in various applications such as drug delivery and cell-based therapy.

  15. [Cell-derived microparticles unveil their fibrinolytic and proteolytic function].

    Science.gov (United States)

    Doeuvre, Loïc; Angles-Cano, Eduardo

    2009-01-01

    Cell-derived microparticles (MP) are membrane microvesicles, 0.1-1 microm in size, shed by cells following activation or during apoptosis in a variety of pathological conditions. MPs released by blood cells or by vascular endothelial cells display molecular signatures that allow their identification and functional characterization. In addition, they provide tissue factor (TF) and a procoagulant phospholipid surface. Therefore, at present, the most strongly established applied research on MPs is their procoagulant activity as a determinant of thrombotic risk in various clinical conditions. Previous studies have indicated that MPs derived from malignant cells express matrix metalloproteinases, urokinase and its receptor (uPA/uPAR) that, in the presence of plasminogen, may act in concert to degrade extracellular matrix proteins. Recently, it was shown that MPs from TNFa-stimulated endothelial cells served as a surface for interaction with plasminogen and its conversion into plasmin by the uPA/uPAR system expressed at their surface. This capacity of MPs to promote plasmin generation confers them a new profibrinolytic and proteolytic function that may be of relevance in fibrinolysis, cell migration, angiogenesis, dissemination of malignant cells, cell detachment and apoptosis.

  16. The in Vitro Assessment of Biochemical Factors in Hepatocyte like Cells Derived from Umbilical Cord Blood Stem Cells

    Directory of Open Access Journals (Sweden)

    A KHoramroodi

    2009-10-01

    Full Text Available Introduction & Objective: Umbilical cord blood (UCB is a source of Hematopoietic Stem Cells (HSC and progenitor cells that can reconstitute the hematopoietic system in patients with malignant and nonmalignant disorders. Mesenchymal stem cell-derived from umbilical cord blood (UCB have been differentiated to some kind of cells, such as osteobblast, adipoblast and chondroblast in Vitro. This study examined the differentiation of Umbilical Cord Blood (UCB derived stem cells to functional hepatocytes. Materials & Methods: The present study was an experimental study which was carried out in the Payam-e-Noor University of Tehran in cooperation with Hamedan University of Medical Sciences in 2008. Umbilical cord blood (UCB was obtained from Fatemieh hospital (Hamadan, Iran. Stem cells were isolated from the cord blood by combining density gradient centrifugation with plastic adherence. When the isolated cells reached 80% confluence, they differentiated to hepatocyte like cells. The medium which was used was consists of DMEM and 10% Fetal Bovine Serum (FBS supplemented with 20 ng/mL Hepatocyte Growth Factor (HGF, 10 ng/mL basic Fibroblast Growth Factor (bFGF and 20 ng/mL Oncostatin M (OSM.The medium was changed every 3 days and stored for Albumin (ALB, Alpha Fetoprotein (AFP, Alkaline Phosphatase (ALP, and urea assay. Finally PAS stain was done to study Glycogen storage in the differentiated cell. Results: Measurement of biochemical factors in different days showed that concentration of albumin (ALB, alpha fetoprotein (AFP, alkaline phosphatase (ALP, and Urea gradually increased. Also, PAS staining showed the storage of glycogen in these cells. Conclusion: Stem cell-derived from human umbilical cord blood (HUCB is a new source of cell types for cell transplantation therapy of hepatic diseases and under certain conditions these cells can differentiate into liver cells.

  17. Canada files WTO complaint against EC.

    Science.gov (United States)

    2000-01-01

    In December 1998, Canada filed a complaint alleging that the European Communities (EC) had adopted regulations that amounted to a scheme to extend patent terms, limited to pharmaceutical and agricultural chemical products.

  18. Special Issue: Selected papers from ECS'97

    DEFF Research Database (Denmark)

    1999-01-01

    This Special Issue of Analog Integrated Circuits and Signal Processing presents extended versions of selected papers from the First Electronic Circuits and Systems Conference (ECS'97) which was held on September 4-5, 1997, in Bratislava, Slovakia.......This Special Issue of Analog Integrated Circuits and Signal Processing presents extended versions of selected papers from the First Electronic Circuits and Systems Conference (ECS'97) which was held on September 4-5, 1997, in Bratislava, Slovakia....

  19. The EC discourse on vocational training

    DEFF Research Database (Denmark)

    Cort, Pia

    2009-01-01

    This article traces the EC vocational training policy historically and describes the discursive alignments which brought the policy from a ‘common vocational training policy' as laid down in Article 128, in the Treaty of Rome to the Lisbon Lifelong Learning strategy. The argument is that vocational...... EC vocational training policy (Ball, What is policy? Texts, trajectories and toolboxes. Discourse, 13(2), 1993)....

  20. The EC-ASEAN COGEN programme

    Energy Technology Data Exchange (ETDEWEB)

    Pennington, M.; Lacrosse, L. [Asian Institute of Technology (Thailand); Schenkel, Y. [Centre de Recherches Agronomiques (Belgium)

    1996-12-31

    The EC-ASEAN COGEN Programme is a co-operation programme between the European Commission (EC) and the Association of South East Asian Nations (ASEAN) co-ordinated by the Asian Institute of Technology (AIT), Bangkok, Thailand. Its aim is to accelerate the implementation of proven technologies generating heat and/or power from wood and agroindustrial residues through partnerships between European and ASEAN companies. (orig.)

  1. The EC-ASEAN COGEN programme

    Energy Technology Data Exchange (ETDEWEB)

    Pennington, M; Lacrosse, L [Asian Institute of Technology (Thailand); Schenkel, Y [Centre de Recherches Agronomiques (Belgium)

    1997-12-31

    The EC-ASEAN COGEN Programme is a co-operation programme between the European Commission (EC) and the Association of South East Asian Nations (ASEAN) co-ordinated by the Asian Institute of Technology (AIT), Bangkok, Thailand. Its aim is to accelerate the implementation of proven technologies generating heat and/or power from wood and agroindustrial residues through partnerships between European and ASEAN companies. (orig.)

  2. PET imaging of adoptive progenitor cell therapies

    International Nuclear Information System (INIS)

    Gelovani, Juri G.

    2008-01-01

    is proposed to circumvent the major limitation of in vitro radiolabeling - the eventual radiolabel decay. Stable transduction of stem cells in vitro would allow for the selection of high quality stem cells with optimal functional parameters of the transduced reporter systems. The use of a long-lived radioisotope 124I to label a highly specific reporter gene probe will allow for ex vivo labeling of stem cells and their imaging immediately after injection and during the following next week. The use of short-lived radioisotopes (i.e., 18F) to label highly specific reporter gene probes will allow repetitive PET imaging for the assessment of to stem cell migration, targeting, differentiation, and long-term viability of stem cell-derived tissues. Qualifications of the research team and resources. An established research team of experts in various disciplines has been assembled at MD Anderson Cancer Center (MDACC) over the past two years including the PI, senior co-investigators and collaborators. The participants of this team are recognized internationally to be among the leaders in their corresponding fields of research and clinical medicine. The resources at MDACC are exceptionally well developed and have been recently reinforced by the installation of a microPET and microSPECT/CT cameras, and a 7T MRI system for high resolution animal imaging; and by integrating a synthetic chemistry core for the development and production of precursors for radiolabeling.

  3. PET imaging of adoptive progenitor cell therapies.

    Energy Technology Data Exchange (ETDEWEB)

    Gelovani, Juri G.

    2008-05-13

    stem cell imaging is proposed to circumvent the major limitation of in vitro radiolabeling – the eventual radiolabel decay. Stable transduction of stem cells in vitro would allow for the selection of high quality stem cells with optimal functional parameters of the transduced reporter systems. The use of a long-lived radioisotope 124I to label a highly specific reporter gene probe will allow for ex vivo labeling of stem cells and their imaging immediately after injection and during the following next week. The use of short-lived radioisotopes (i.e., 18F) to label highly specific reporter gene probes will allow repetitive PET imaging for the assessment of to stem cell migration, targeting, differentiation, and long-term viability of stem cell-derived tissues. Qualifications of the research team and resources. An established research team of experts in various disciplines has been assembled at MD Anderson Cancer Center (MDACC) over the past two years including the PI, senior co-investigators and collaborators. The participants of this team are recognized internationally to be among the leaders in their corresponding fields of research and clinical medicine. The resources at MDACC are exceptionally well developed and have been recently reinforced by the installation of a microPET and microSPECT/CT cameras, and a 7T MRI system for high resolution animal imaging; and by integrating a synthetic chemistry core for the development and production of precursors for radiolabeling.

  4. ES-cell derived hematopoietic cells induce transplantation tolerance.

    Directory of Open Access Journals (Sweden)

    Sabrina Bonde

    Full Text Available BACKGROUND: Bone marrow cells induce stable mixed chimerism under appropriate conditioning of the host, mediating the induction of transplantation tolerance. However, their strong immunogenicity precludes routine use in clinical transplantation due to the need for harsh preconditioning and the requirement for toxic immunosuppression to prevent rejection and graft-versus-host disease. Alternatively, embryonic stem (ES cells have emerged as a potential source of less immunogenic hematopoietic progenitor cells (HPCs. Up till now, however, it has been difficult to generate stable hematopoietic cells from ES cells. METHODOLOGY/PRINCIPAL FINDINGS: Here, we derived CD45(+ HPCs from HOXB4-transduced ES cells and showed that they poorly express MHC antigens. This property allowed their long-term engraftment in sublethally irradiated recipients across MHC barriers without the need for immunosuppressive agents. Although donor cells declined in peripheral blood over 2 months, low level chimerism was maintained in the bone marrow of these mice over 100 days. More importantly, chimeric animals were protected from rejection of donor-type cardiac allografts. CONCLUSIONS: Our data show, for the first time, the efficacy of ES-derived CD45(+ HPCs to engraft in allogenic recipients without the use of immunosuppressive agents, there by protecting cardiac allografts from rejection.

  5. Sphingosine-1-Phosphate (S1P) Signaling in Neural Progenitors.

    Science.gov (United States)

    Callihan, Phillip; Alqinyah, Mohammed; Hooks, Shelley B

    2018-01-01

    Sphingosine-1-phosphate (S1P) and its receptors are important in nervous system development. Reliable in vitro human model systems are needed to further define specific roles for S1P signaling in neural development. We have described S1P-regulated signaling, survival, and differentiation in a human embryonic stem cell-derived neuroepithelial progenitor cell line (hNP1) that expresses functional S1P receptors. These cells can be further differentiated to a neuronal cell type and therefore represent a good model system to study the role of S1P signaling in human neural development. The following sections describe in detail the culture and differentiation of hNP1 cells and two assays to measure S1P signaling in these cells.

  6. Characterization and modulation of canine mast cell derived eicosanoids

    Science.gov (United States)

    Lin, Tzu-Yin; London, Cheryl A.

    2013-01-01

    Mast cells play an important role in both innate and acquired immunity as well as several pathological conditions including allergy, arthritis and neoplasia. They influence these processes by producing a variety of mediators including cytokines, chemokines and eicosanoids. Very little is currently known about the spectrum of inflammatory mediators, particularly eicosanoids (prostaglandins and leukotrienes), produced by canine mast cells. This is important since modulating mast cell derived eicosanoids may help in the treatment of autoimmune and inflammatory disorders. The purpose of this study was to investigate the spectrum of eicosanoids produced by normal canine mast cells and to evaluate the effects of cytokines and non-steroidal anti-inflammatory mediators (NSAIDS) on eicosanoid production and release. Canine bone marrow derived cultured mast cells (cBMCMCs) expressed COX-1, COX-2, and 5-LOX and synthesized and released PGD2, PGE2, LTB4, and LTC4 following activation by a variety of stimuli. The selective COX-2 NSAIDs carprofen (Rimadyl®) and deracoxib (Deramaxx®) inhibited PGD2 and PGE2 production but only slightly inhibited LTB4 and LTC4. The mixed COX-1/COX-2 inhibitor piroxicam blocked PGD2 and PGE2 production, but upregulated LTC4 following treatment while tepoxilan (Zubrin®), a pan COX/LOX inhibitor, markedly reduced the production of all eicosanoids. The LOX inhibitor nordihydroguaiaretic acid (NDGA) prevented LTB4/LTC4 release and BMBMC degranulation. Pre-incubation of cBMCMCs with IL-4 and SCF sensitized these cells to degranulation in response to substance P. In conclusion, canine BMCMCs produce an array of eicosanoids similar to those produced by mast cells from other species. Tepoxilan appeared to be the most effective NSAID for blocking eicosanoid production and thus may be useful for modulating mast cell mediated responses in dogs. PMID:20036014

  7. Influence of red blood cell-derived microparticles upon vasoregulation.

    Science.gov (United States)

    Said, Ahmed S; Doctor, Allan

    2017-10-01

    Here we review recent data and the evolving understanding of the role of red blood cell-derived microparticles (RMPs) in normal physiology and in disease progression. Microparticles (MPs) are small membrane vesicles derived from various parent cell types. MPs are produced in response to a variety of stimuli through several cytoskeletal and membrane phospholipid changes. MPs have been investigated as potential biomarkers for multiple disease processes and are thought to have biological effects, most notably in: promotion of coagulation, production and handling of reactive oxygen species, immune modulation, angiogenesis, and in apoptosis. Specifically, RMPs are produced normally during RBC maturation and their production is accelerated during processing and storage for transfusion. Several factors during RBC storage are known to trigger RMP production, including: increased intracellular calcium, increased potassium leakage, and energy failure with ATP depletion. Of note, RMP composition differs from that of intact RBCs, and the nature and composition of RMP components are affected by both storage duration and the character of storage solutions. Recognised RMP bioactivities include: promotion of coagulation, immune modulation, and promotion of endothelial adhesion, as well as influence upon vasoregulation via nitric oxide (NO) scavenging. Of particular relevance, RMPs are more avid NO scavengers than intact RBCs and this feature has been proposed as a mechanism for the impaired oxygen delivery homeostasis that has been observed following transfusion. Preliminary human studies demonstrate that circulating RMP abundance increases with RBC transfusion and is associated with altered plasma vasoactivity and abnormal vasoregulation. In summary, RMPs are submicron particles released from stored RBCs, with demonstrated vasoactive properties that appear to disturb oxygen delivery homeostasis. The clinical impact of RMPs in transfusion recipients is an area of continued

  8. Red cell-derived microparticles (RMP) as haemostatic agent.

    Science.gov (United States)

    Jy, Wenche; Johansen, Max E; Bidot, Carlos; Horstman, Lawrence L; Ahn, Yeon S

    2013-10-01

    Among circulating cell-derived microparticles, those derived from red cells (RMP) have been least well investigated. To exploit potential haemostatic benefit of RMP, we developed a method of producing them in quantity, and here report on their haemostatic properties. High-pressure extrusion of washed RBC was employed to generate RMP. RMP were identified and enumerated by flow cytometry. Their size distribution was assessed by Doppler electrophoretic light scattering analysis (DELSA). Interaction with platelets was studied by platelet aggregometry, and shear-dependent adhesion by Diamed IMPACT-R. Thrombin generation and tissue factor (TF) expression was also measured. The effect of RMP on blood samples of patients with bleeding disorders was investigated ex vivo by thromboelastography (TEG). Haemostatic efficacy in vivo was assessed by measuring reduction of blood loss and bleeding time in rats and rabbits. RMP have mean diameter of 0.45 µm and 50% of them exhibit annexin V binding, a proxy for procoagulant phospholipids (PL). No TF could be detected by flow cytometry. At saturating concentrations of MPs, RMP generated thrombin robustly but after longer delay compared to PMP and EMP. RMP enhanced platelet adhesion and aggregation induced by low-dose ADP or AA. In TEG study, RMP corrected or improved haemostatic defects in blood of patients with platelet and coagulation disorders. RMP reduced bleeding time and blood loss in thrombocytopenic rabbits (busulfan-treated) and in Plavix-treated rats. In conclusion, RMP has broad haemostatic activity, enhancing both primary (platelet) and secondary (coagulation) haemostasis, suggesting potential use as haemostatic agent for treatment of bleeding.

  9. Human neural progenitors express functional lysophospholipid receptors that regulate cell growth and morphology

    Directory of Open Access Journals (Sweden)

    Callihan Phillip

    2008-12-01

    Full Text Available Abstract Background Lysophospholipids regulate the morphology and growth of neurons, neural cell lines, and neural progenitors. A stable human neural progenitor cell line is not currently available in which to study the role of lysophospholipids in human neural development. We recently established a stable, adherent human embryonic stem cell-derived neuroepithelial (hES-NEP cell line which recapitulates morphological and phenotypic features of neural progenitor cells isolated from fetal tissue. The goal of this study was to determine if hES-NEP cells express functional lysophospholipid receptors, and if activation of these receptors mediates cellular responses critical for neural development. Results Our results demonstrate that Lysophosphatidic Acid (LPA and Sphingosine-1-phosphate (S1P receptors are functionally expressed in hES-NEP cells and are coupled to multiple cellular signaling pathways. We have shown that transcript levels for S1P1 receptor increased significantly in the transition from embryonic stem cell to hES-NEP. hES-NEP cells express LPA and S1P receptors coupled to Gi/o G-proteins that inhibit adenylyl cyclase and to Gq-like phospholipase C activity. LPA and S1P also induce p44/42 ERK MAP kinase phosphorylation in these cells and stimulate cell proliferation via Gi/o coupled receptors in an Epidermal Growth Factor Receptor (EGFR- and ERK-dependent pathway. In contrast, LPA and S1P stimulate transient cell rounding and aggregation that is independent of EGFR and ERK, but dependent on the Rho effector p160 ROCK. Conclusion Thus, lysophospholipids regulate neural progenitor growth and morphology through distinct mechanisms. These findings establish human ES cell-derived NEP cells as a model system for studying the role of lysophospholipids in neural progenitors.

  10. Differentiation of oligodendrocyte progenitor cells from dissociated monolayer and feeder-free cultured pluripotent stem cells.

    Science.gov (United States)

    Yamashita, Tomoko; Miyamoto, Yuki; Bando, Yoshio; Ono, Takashi; Kobayashi, Sakurako; Doi, Ayano; Araki, Toshihiro; Kato, Yosuke; Shirakawa, Takayuki; Suzuki, Yutaka; Yamauchi, Junji; Yoshida, Shigetaka; Sato, Naoya

    2017-01-01

    Oligodendrocytes myelinate axons and form myelin sheaths in the central nervous system. The development of therapies for demyelinating diseases, including multiple sclerosis and leukodystrophies, is a challenge because the pathogenic mechanisms of disease remain poorly understood. Primate pluripotent stem cell-derived oligodendrocytes are expected to help elucidate the molecular pathogenesis of these diseases. Oligodendrocytes have been successfully differentiated from human pluripotent stem cells. However, it is challenging to prepare large amounts of oligodendrocytes over a short amount of time because of manipulation difficulties under conventional primate pluripotent stem cell culture methods. We developed a proprietary dissociated monolayer and feeder-free culture system to handle pluripotent stem cell cultures. Because the dissociated monolayer and feeder-free culture system improves the quality and growth of primate pluripotent stem cells, these cells could potentially be differentiated into any desired functional cells and consistently cultured in large-scale conditions. In the current study, oligodendrocyte progenitor cells and mature oligodendrocytes were generated within three months from monkey embryonic stem cells. The embryonic stem cell-derived oligodendrocytes exhibited in vitro myelinogenic potency with rat dorsal root ganglion neurons. Additionally, the transplanted oligodendrocyte progenitor cells differentiated into myelin basic protein-positive mature oligodendrocytes in the mouse corpus callosum. This preparative method was used for human induced pluripotent stem cells, which were also successfully differentiated into oligodendrocyte progenitor cells and mature oligodendrocytes that were capable of myelinating rat dorsal root ganglion neurons. Moreover, it was possible to freeze, thaw, and successfully re-culture the differentiating cells. These results showed that embryonic stem cells and human induced pluripotent stem cells maintained in a

  11. Flow Cytometric Quantification of Peripheral Blood Cell β-Adrenergic Receptor Density and Urinary Endothelial Cell-Derived Microparticles in Pulmonary Arterial Hypertension.

    Directory of Open Access Journals (Sweden)

    Jonathan A Rose

    Full Text Available Pulmonary arterial hypertension (PAH is a heterogeneous disease characterized by severe angiogenic remodeling of the pulmonary artery wall and right ventricular hypertrophy. Thus, there is an increasing need for novel biomarkers to dissect disease heterogeneity, and predict treatment response. Although β-adrenergic receptor (βAR dysfunction is well documented in left heart disease while endothelial cell-derived microparticles (Ec-MPs are established biomarkers of angiogenic remodeling, methods for easy large clinical cohort analysis of these biomarkers are currently absent. Here we describe flow cytometric methods for quantification of βAR density on circulating white blood cells (WBC and Ec-MPs in urine samples that can be used as potential biomarkers of right heart failure in PAH. Biotinylated β-blocker alprenolol was synthesized and validated as a βAR specific probe that was combined with immunophenotyping to quantify βAR density in circulating WBC subsets. Ec-MPs obtained from urine samples were stained for annexin-V and CD144, and analyzed by a micro flow cytometer. Flow cytometric detection of alprenolol showed that βAR density was decreased in most WBC subsets in PAH samples compared to healthy controls. Ec-MPs in urine was increased in PAH compared to controls. Furthermore, there was a direct correlation between Ec-MPs and Tricuspid annular plane systolic excursion (TAPSE in PAH patients. Therefore, flow cytometric quantification of peripheral blood cell βAR density and urinary Ec-MPs may be useful as potential biomarkers of right ventricular function in PAH.

  12. Flow Cytometric Quantification of Peripheral Blood Cell β-Adrenergic Receptor Density and Urinary Endothelial Cell-Derived Microparticles in Pulmonary Arterial Hypertension.

    Science.gov (United States)

    Rose, Jonathan A; Wanner, Nicholas; Cheong, Hoi I; Queisser, Kimberly; Barrett, Patrick; Park, Margaret; Hite, Corrine; Naga Prasad, Sathyamangla V; Erzurum, Serpil; Asosingh, Kewal

    2016-01-01

    Pulmonary arterial hypertension (PAH) is a heterogeneous disease characterized by severe angiogenic remodeling of the pulmonary artery wall and right ventricular hypertrophy. Thus, there is an increasing need for novel biomarkers to dissect disease heterogeneity, and predict treatment response. Although β-adrenergic receptor (βAR) dysfunction is well documented in left heart disease while endothelial cell-derived microparticles (Ec-MPs) are established biomarkers of angiogenic remodeling, methods for easy large clinical cohort analysis of these biomarkers are currently absent. Here we describe flow cytometric methods for quantification of βAR density on circulating white blood cells (WBC) and Ec-MPs in urine samples that can be used as potential biomarkers of right heart failure in PAH. Biotinylated β-blocker alprenolol was synthesized and validated as a βAR specific probe that was combined with immunophenotyping to quantify βAR density in circulating WBC subsets. Ec-MPs obtained from urine samples were stained for annexin-V and CD144, and analyzed by a micro flow cytometer. Flow cytometric detection of alprenolol showed that βAR density was decreased in most WBC subsets in PAH samples compared to healthy controls. Ec-MPs in urine was increased in PAH compared to controls. Furthermore, there was a direct correlation between Ec-MPs and Tricuspid annular plane systolic excursion (TAPSE) in PAH patients. Therefore, flow cytometric quantification of peripheral blood cell βAR density and urinary Ec-MPs may be useful as potential biomarkers of right ventricular function in PAH.

  13. Earmuff restricts progenitor cell potential by attenuating the competence to respond to self-renewal factors.

    Science.gov (United States)

    Janssens, Derek H; Komori, Hideyuki; Grbac, Daniel; Chen, Keng; Koe, Chwee Tat; Wang, Hongyan; Lee, Cheng-Yu

    2014-03-01

    Despite expressing stem cell self-renewal factors, intermediate progenitor cells possess restricted developmental potential, which allows them to give rise exclusively to differentiated progeny rather than stem cell progeny. Failure to restrict the developmental potential can allow intermediate progenitor cells to revert into aberrant stem cells that might contribute to tumorigenesis. Insight into stable restriction of the developmental potential in intermediate progenitor cells could improve our understanding of the development and growth of tumors, but the mechanisms involved remain largely unknown. Intermediate neural progenitors (INPs), generated by type II neural stem cells (neuroblasts) in fly larval brains, provide an in vivo model for investigating the mechanisms that stably restrict the developmental potential of intermediate progenitor cells. Here, we report that the transcriptional repressor protein Earmuff (Erm) functions temporally after Brain tumor (Brat) and Numb to restrict the developmental potential of uncommitted (immature) INPs. Consistently, endogenous Erm is detected in immature INPs but undetectable in INPs. Erm-dependent restriction of the developmental potential in immature INPs leads to attenuated competence to respond to all known neuroblast self-renewal factors in INPs. We also identified that the BAP chromatin-remodeling complex probably functions cooperatively with Erm to restrict the developmental potential of immature INPs. Together, these data led us to conclude that the Erm-BAP-dependent mechanism stably restricts the developmental potential of immature INPs by attenuating their genomic responses to stem cell self-renewal factors. We propose that restriction of developmental potential by the Erm-BAP-dependent mechanism functionally distinguishes intermediate progenitor cells from stem cells, ensuring the generation of differentiated cells and preventing the formation of progenitor cell-derived tumor-initiating stem cells.

  14. Functional Differences in Engineered Myocardium from Embryonic Stem Cell-Derived versus Neonatal Cardiomyocytes

    NARCIS (Netherlands)

    Feinberg, Adam W.; Ripplinger, Crystal M.; van der Meer, Peter; Sheehy, Sean P.; Domian, Ibrahim; Chien, Kenneth R.; Parker, Kevin Kit

    2013-01-01

    Stem cell-derived cardiomyocytes represent unique tools for cell-and tissue-based regenerative therapies, drug discovery and safety, and studies of fundamental heart-failure mechanisms. However, the degree to which stem cell-derived cardiomyocytes compare to mature cardiomyocytes is often debated.

  15. Mesenchymal progenitor cells for the osteogenic lineage.

    Science.gov (United States)

    Ono, Noriaki; Kronenberg, Henry M

    2015-09-01

    Mesenchymal progenitors of the osteogenic lineage provide the flexibility for bone to grow, maintain its function and homeostasis. Traditionally, colony-forming-unit fibroblasts (CFU-Fs) have been regarded as surrogates for mesenchymal progenitors; however, this definition cannot address the function of these progenitors in their native setting. Transgenic murine models including lineage-tracing technologies based on the cre-lox system have proven to be useful in delineating mesenchymal progenitors in their native environment. Although heterogeneity of cell populations of interest marked by a promoter-based approach complicates overall interpretation, an emerging complexity of mesenchymal progenitors has been revealed. Current literatures suggest two distinct types of bone progenitor cells; growth-associated mesenchymal progenitors contribute to explosive growth of bone in early life, whereas bone marrow mesenchymal progenitors contribute to the much slower remodeling process and response to injury that occurs mainly in adulthood. More detailed relationships of these progenitors need to be studied through further experimentation.

  16. Human Induced Pluripotent Stem Cell-Derived Macrophages Share Ontogeny with MYB-Independent Tissue-Resident Macrophages

    Directory of Open Access Journals (Sweden)

    Julian Buchrieser

    2017-02-01

    Full Text Available Tissue-resident macrophages, such as microglia, Kupffer cells, and Langerhans cells, derive from Myb-independent yolk sac (YS progenitors generated before the emergence of hematopoietic stem cells (HSCs. Myb-independent YS-derived resident macrophages self-renew locally, independently of circulating monocytes and HSCs. In contrast, adult blood monocytes, as well as infiltrating, gut, and dermal macrophages, derive from Myb-dependent HSCs. These findings are derived from the mouse, using gene knockouts and lineage tracing, but their applicability to human development has not been formally demonstrated. Here, we use human induced pluripotent stem cells (iPSCs as a tool to model human hematopoietic development. By using a CRISPR-Cas9 knockout strategy, we show that human iPSC-derived monocytes/macrophages develop in an MYB-independent, RUNX1-, and SPI1 (PU.1-dependent fashion. This result makes human iPSC-derived macrophages developmentally related to and a good model for MYB-independent tissue-resident macrophages, such as alveolar and kidney macrophages, microglia, Kupffer cells, and Langerhans cells.

  17. Foetal stem cell derivation & characterization for osteogenic lineage

    Directory of Open Access Journals (Sweden)

    A Mangala Gowri

    2013-01-01

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

  18. A review of human pluripotent stem cell-derived cardiomyocytes for high-throughput drug discovery, cardiotoxicity screening, and publication standards.

    Science.gov (United States)

    Mordwinkin, Nicholas M; Burridge, Paul W; Wu, Joseph C

    2013-02-01

    Drug attrition rates have increased in past years, resulting in growing costs for the pharmaceutical industry and consumers. The reasons for this include the lack of in vitro models that correlate with clinical results and poor preclinical toxicity screening assays. The in vitro production of human cardiac progenitor cells and cardiomyocytes from human pluripotent stem cells provides an amenable source of cells for applications in drug discovery, disease modeling, regenerative medicine, and cardiotoxicity screening. In addition, the ability to derive human-induced pluripotent stem cells from somatic tissues, combined with current high-throughput screening and pharmacogenomics, may help realize the use of these cells to fulfill the potential of personalized medicine. In this review, we discuss the use of pluripotent stem cell-derived cardiomyocytes for drug discovery and cardiotoxicity screening, as well as current hurdles that must be overcome for wider clinical applications of this promising approach.

  19. Human embryonic stem cell-derived pancreatic endoderm alleviates diabetic pathology and improves reproductive outcome in C57BL/KsJ-Lep(db/+) gestational diabetes mellitus mice.

    Science.gov (United States)

    Xing, Baoheng; Wang, Lili; Li, Qin; Cao, Yalei; Dong, Xiujuan; Liang, Jun; Wu, Xiaohua

    2015-07-01

    Gestational diabetes mellitus is a condition commonly encountered during mid to late pregnancy with pathologic manifestations including hyperglycemia, hyperinsulinemia, insulin resistance, and fetal maldevelopment. The cause of gestational diabetes mellitus can be attributed to both genetic and environmental factors, hence complicating its diagnosis and treatment. Pancreatic progenitors derived from human embryonic stem cells were shown to be able to effectively treat diabetes in mice. In this study, we have developed a system of treating diabetes using human embryonic stem cell-derived pancreatic endoderm in a mouse model of gestational diabetes mellitus. Human embryonic stem cells were differentiated in vitro into pancreatic endoderm, which were then transplanted into db/+ mice suffering from gestational diabetes mellitus. The transplant greatly improved glucose metabolism and reproductive outcome of the females compared with the control groups. Our findings support the feasibility of using differentiated human embryonic stem cells for treating gestational diabetes mellitus patients. Copyright © 2015 Elsevier Inc. All rights reserved.

  20. Study of mesanchymal stem cells derived from human umbilical cord vein wall and determining the Process of differentiation to cartilage and bone

    Directory of Open Access Journals (Sweden)

    MohammadAli Zare

    2015-01-01

    Full Text Available Background: Mesenchymal stem cells (MSCs comprise a rare population of multipotent progenitors capable of supporting hematopoiesis and differentiating into three (osteogenic, adipogenic, and chondrogenic or more (myogenic, cardiomyogenic, etc. lineages. Due to this ability, MSCs appear to be an attractive tool in the context of tissue engineering and cell-based therapy. Currently, bone marrow represents the main source of MSCs for both experimental and clinical studies. The purpose of this study was isolation and quantitative comparison of mesenchymal stem cells derived from umbilical vein. Materials and Methods: In this study, 35 samples of umbilical cord of healthy full- term newborn were studied. Results: The cells had fibroblastoid like appearance and had revealed the potential to differentiate into three linage of bone, Adipose and cartilage. Surface markers for mesenchymal nature were their demonstratives. Conclusion: Based on our findings the mesenchymal stem cells, from umbilical vein wall can be isolated, cultured and differentiated into three categories of bone, cartilage and adipose.

  1. ST ECS error handling and detector safety

    CERN Document Server

    Esperante Pereira, D

    2010-01-01

    This note describes the automatic operation sequences that the ECS implements in order to operate the ST detector in a safe and reliable way. First, the start-up sequences to set the detector in a well known state are described. In addition the possible ST detector states based on the status of the different detector elements will be defined. The states are of crucial interest for the safe operation of the detector when the LHC beam is ready. Besides, the ECS automatic actions taken in case of abnormal status of any of the detector elements are reported. Next, the operational alarm ranges and the error recovery mechanisms are mentioned.

  2. Energy conservation in the EC glass industry

    Energy Technology Data Exchange (ETDEWEB)

    Waal, H. de [TNO Institute of Applied Physics, Delft (Netherlands)

    1994-12-31

    The data presented in this survey are based mainly on a recent study, performed by the Energy Technology Support Unit ETSU. Harwell Laboratory, United Kingdom, in the context of the EC-Thermie programme. Also, use has been made of a paper `Glass Manufacture, energy and CO{sub 2}-emissions`, presented by G.J. Copley of the British Glass Manufacturers Confederation, Sheffield, United Kingdom, presented at the Thermie Seminar in Wiesbaden, 1992. A third source of information has been the data collected by the CPIV, the European Glass Manufacturers Federation on the present and future economic situation of the EC Glass Industry. (orig.)

  3. Public Availability to ECS Collected Datasets

    Science.gov (United States)

    Henderson, J. F.; Warnken, R.; McLean, S. J.; Lim, E.; Varner, J. D.

    2013-12-01

    Coastal nations have spent considerable resources exploring the limits of their extended continental shelf (ECS) beyond 200 nm. Although these studies are funded to fulfill requirements of the UN Convention on the Law of the Sea, the investments are producing new data sets in frontier areas of Earth's oceans that will be used to understand, explore, and manage the seafloor and sub-seafloor for decades to come. Although many of these datasets are considered proprietary until a nation's potential ECS has become 'final and binding' an increasing amount of data are being released and utilized by the public. Data sets include multibeam, seismic reflection/refraction, bottom sampling, and geophysical data. The U.S. ECS Project, a multi-agency collaboration whose mission is to establish the full extent of the continental shelf of the United States consistent with international law, relies heavily on data and accurate, standard metadata. The United States has made it a priority to make available to the public all data collected with ECS-funding as quickly as possible. The National Oceanic and Atmospheric Administration's (NOAA) National Geophysical Data Center (NGDC) supports this objective by partnering with academia and other federal government mapping agencies to archive, inventory, and deliver marine mapping data in a coordinated, consistent manner. This includes ensuring quality, standard metadata and developing and maintaining data delivery capabilities built on modern digital data archives. Other countries, such as Ireland, have submitted their ECS data for public availability and many others have made pledges to participate in the future. The data services provided by NGDC support the U.S. ECS effort as well as many developing nation's ECS effort through the U.N. Environmental Program. Modern discovery, visualization, and delivery of scientific data and derived products that span national and international sources of data ensure the greatest re-use of data and

  4. EC Transmission Line Risk Identification and Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Bigelow, Tim S [ORNL

    2012-04-01

    The purpose of this document is to assist in evaluating and planning for the cost, schedule, and technical project risks associated with the delivery and operation of the EC (Electron cyclotron) transmission line system. In general, the major risks that are anticipated to be encountered during the project delivery phase associated with the implementation of the Procurement Arrangement for the EC transmission line system are associated with: (1) Undefined or changing requirements (e.g., functional or regulatory requirements) (2) Underperformance of prototype, first unit, or production components during testing (3) Unavailability of qualified vendors for critical components Technical risks associated with the design and operation of the system are also identified.

  5. Rapid procedure to calibrate EC-10 and EC-20 capacitance sensors ...

    African Journals Online (AJOL)

    A rapid calibration procedure for EC-10 and EC-20 sensors is introduced to promote the commercial use of these sensors for hydroponic irrigation management in coir. The method is comprised of taking one sensor reading, by a sensor installed under hydroponic crop production conditions, and one gravimetric sample, ...

  6. Tracking differentiating neural progenitors in pluripotent cultures using microRNA-regulated lentiviral vectors.

    Science.gov (United States)

    Sachdeva, Rohit; Jönsson, Marie E; Nelander, Jenny; Kirkeby, Agnete; Guibentif, Carolina; Gentner, Bernhard; Naldini, Luigi; Björklund, Anders; Parmar, Malin; Jakobsson, Johan

    2010-06-22

    In this study, we have used a microRNA-regulated lentiviral reporter system to visualize and segregate differentiating neuronal cells in pluripotent cultures. Efficient suppression of transgene expression, specifically in undifferentiated pluripotent cells, was achieved by using a lentiviral vector expressing a fluorescent reporter gene regulated by microRNA-292. Using this strategy, it was possible to track progeny from murine ES, human ES cells, and induced pluripotent stem cells as they differentiated toward the neural lineage. In addition, this strategy was successfully used to FACS purify neuronal progenitors for molecular analysis and transplantation. FACS enrichment reduced tumor formation and increased survival of ES cell-derived neuronal progenitors after transplantation. The properties and versatility of the microRNA-regulated vectors allows broad use of these vectors in stem cell applications.

  7. Myocardial regeneration by transplantation of modified endothelial progenitor cells expressing SDF-1 in a rat model

    DEFF Research Database (Denmark)

    Schuh, A.; Kroh, A.; Konschalla, S.

    2012-01-01

    Cell based therapy has been shown to attenuate myocardial dysfunction after myocardial infarction (MI) in different acute and chronic animal models. It has been further shown that stromal-cell derived factor-1a (SDF-1a) facilitates proliferation and migration of endogenous progenitor cells...... into injured tissue. The aim of the present study was to investigate the role of exogenously applied and endogenously mobilized cells in a regenerative strategy for MI therapy. Lentivirally SDF-1a-infected endothelial progenitor cells (EPCs) were injected after 90 min. of ligation and reperfusion of the left...... compared to medium control. Intracoronary application of cells did not lead to significant differences compared to medium injected control hearts. Histology showed a significantly elevated rate of apoptotic cells and augmented proliferation after transplantation of EPCs and EPCs + SDF-1 alpha in infarcted...

  8. Endothelial progenitor cell subsets and preeclampsia: Findings and controversies

    Directory of Open Access Journals (Sweden)

    Armin Attar

    2017-10-01

    Full Text Available Vascular remodeling is an essential component of gestation. Endothelial progenitor cells (EPCs play an important role in the regulation of vascular homeostasis. The results of studies measuring the number of EPCs in normal pregnancies and in preeclampsia have been highly controversial or even contradictory because of some variations in technical issues and different methodologies enumerating three distinct subsets of EPCs: circulating angiogenic cells (CAC, colony forming unit endothelial cells (CFU-ECs, and endothelial colony-forming cells (ECFCs. In general, most studies have shown an increase in the number of CACs in the maternal circulation with a progression in the gestational age in normal pregnancies, while functional capacities measured by CFU-ECs and ECFCs remain intact. In the case of preeclampsia, mobilization of CACs and ECFCs occurs in the peripheral blood of pregnant women, but the functional capacities shown by culture of the derived colony-forming assays (CFU-EC and ECFC assays are altered. Furthermore, the number of all EPC subsets will be reduced in umbilical cord blood in the case of preeclampsia. As EPCs play an important role in the homeostasis of vascular networks, the difference in their frequency and functionality in normal pregnancies and those with preeclampsia can be expected. In this review, there was an attempt to provide a justification for these controversies.

  9. Wnt inhibition promotes vascular specification of embryonic cardiac progenitors.

    Science.gov (United States)

    Reichman, David E; Park, Laura; Man, Limor; Redmond, David; Chao, Kenny; Harvey, Richard P; Taketo, Makoto M; Rosenwaks, Zev; James, Daylon

    2018-01-08

    Several studies have demonstrated a multiphasic role for Wnt signaling during embryonic cardiogenesis and developed protocols that enrich for cardiac derivatives during in vitro differentiation of human pluripotent stem cells (hPSCs). However, few studies have investigated the role of Wnt signaling in the specification of cardiac progenitor cells (CPCs) toward downstream fates. Using transgenic mice and hPSCs, we tracked endothelial cells (ECs) that originated from CPCs expressing NKX2.5. Analysis of EC-fated CPCs at discrete phenotypic milestones during hPSC differentiation identified reduced Wnt activity as a hallmark of EC specification, and the enforced activation or inhibition of Wnt reduced or increased, respectively, the degree of vascular commitment within the CPC population during both hPSC differentiation and mouse embryogenesis. Wnt5a, which has been shown to exert an inhibitory influence on Wnt signaling during cardiac development, was dynamically expressed during vascular commitment of hPSC-derived CPCs, and ectopic Wnt5a promoted vascular specification of hPSC-derived and mouse embryonic CPCs. © 2018. Published by The Company of Biologists Ltd.

  10. Radiosensitivity of glial progenitor cells of the perinatal and adult rat optic nerve studied by an in vitro clonogenic assay

    International Nuclear Information System (INIS)

    Maazen, R.W.M. van der; Verhagen, I.; Kleiboer, B.J.; Kogel, A.J. van der

    1991-01-01

    The cellular basis of radiation-induced demyelination and white matter necrosis of the central nervous system (CNS), is poorly understood. Glial cells responsible for myelination in the CNS might be the target cells of this type of damage. Glial cells with stem cell properties derived from the perinatal and adult rat CNS can be cultured in vitro. These cells are able to differentiate into oligodendrocytes or type-2 astrocytes (O-2A) depending on the culture conditions. Growth factors produced by monolayers of type-1 astrocytes inhibit premature differentiation of O-2A progenitor cells and allow colony formation. A method which employs these monolayers of type-1 astrocytes to culture O-2A progenitor cells has been adapted to allow the analysis of colonies of surviving cells after X-irradiation. In vitro survival curves were obtained for glial progenitor cells derived from perinatal and adult optic nerves. The intrinsic radiosensitivity of perinatal and adult O-2A progenitor cells showed a large difference. Perinatal O-2A progenitor cells are quite radiosensitive, in contrast to adult O-2A progenitor cells. For both cell types an inverse relationship was found between the dose and the size of colonies derived from surviving cells. Surviving O-2A progenitor cells maintain their ability to differentiate into oligo-dendrocytes or type-2 astrocytes. This system to assess radiation-induced damage to glial progenitor cells in vitro systems to have a great potential in unraveling the cellular basis of radiation-induced demyelinating syndromes of the CNS. (author). 28 refs.; 4 figs.; 1 tab

  11. Preclinical Studies of Induced Pluripotent Stem Cell-Derived Astrocyte Transplantation in ALS

    Science.gov (United States)

    2012-10-01

    Pluripotent Stem Cell -Derived Astrocyte Transplantation in ALS PRINCIPAL INVESTIGATOR: Nicholas J. Maragakis, M.D...Pluripotent Stem Cell -Derived Astrocyte Transplantation in ALS 5b. GRANT NUMBER W81XWH-10-1-0520 5c. PROGRAM ELEMENT NUMBER 6. AUTHOR(S) 5d...into astrocytes following transplantation. 15. SUBJECT TERMS Stem Cells , iPS cells, astrocytes, familial ALS 16. SECURITY CLASSIFICATION OF

  12. Stem Cell-Derived Exosome in Cardiovascular Diseases: Macro Roles of Micro Particles.

    Science.gov (United States)

    Yuan, Ye; Du, Weijie; Liu, Jiaqi; Ma, Wenya; Zhang, Lai; Du, Zhimin; Cai, Benzhi

    2018-01-01

    The stem cell-based therapy has emerged as the promising therapeutic strategies for cardiovascular diseases (CVDs). Recently, increasing evidence suggest stem cell-derived active exosomes are important communicators among cells in the heart via delivering specific substances to the adjacent/distant target cells. These exosomes and their contents such as certain proteins, miRNAs and lncRNAs exhibit huge beneficial effects on preventing heart damage and promoting cardiac repair. More importantly, stem cell-derived exosomes are more effective and safer than stem cell transplantation. Therefore, administration of stem cell-derived exosomes will expectantly be an alternative stem cell-based therapy for the treatment of CVDs. Furthermore, modification of stem cell-derived exosomes or artificial synthesis of exosomes will be the new therapeutic tools for CVDs in the future. In addition, stem cell-derived exosomes also have been implicated in the diagnosis and prognosis of CVDs. In this review, we summarize the current advances of stem cell-derived exosome-based treatment and prognosis for CVDs, including their potential benefits, underlying mechanisms and limitations, which will provide novel insights of exosomes as a new tool in clinical therapeutic translation in the future.

  13. Potential roles of cell-derived microparticles in ischemic brain disease.

    Science.gov (United States)

    Horstman, Lawrence L; Jy, Wenche; Bidot, Carlos J; Nordberg, Mary L; Minagar, Alireza; Alexander, J Steven; Kelley, Roger E; Ahn, Yeon S

    2009-10-01

    The objective of this study is to review the role of cell-derived microparticles in ischemic cerebrovascular diseases. An extensive PubMed search of literature pertaining to this study was performed in April 2009 using specific keyword search terms related to cell-derived microparticles and ischemic stroke. Some references are not cited here as it is not possible to be all inclusive or due to space limitation. Cell-derived microparticles are small membranous vesicles released from the plasma membranes of platelets, leukocytes, red cells and endothelial cells in response to diverse biochemical agents or mechanical stresses. They are the main carriers of circulating tissue factor, the principal initiator of intravascular thrombosis, and are implicated in a variety of thrombotic and inflammatory disorders. This review outlines evidence suggesting that cell-derived microparticles are involved predominantly with microvascular, as opposed to macrovascular, thrombosis. More specifically, cell-derived microparticles may substantially contribute to ischemic brain disease in several settings, as well as to neuroinflammatory conditions. If further work confirms this hypothesis, novel therapeutic strategies for minimizing cell-derived microparticles-mediated ischemia are available or can be developed, as discussed.

  14. EC-fans and fan control systems; EC-ventilatoren en ventilatorregelsystemen

    Energy Technology Data Exchange (ETDEWEB)

    Havenaar, D.

    2011-07-15

    Saving of energy can be achieved by the design and application of energy efficient components such as EC-fans (EC stands for Electronically Commutated technology), which are used in different types of cooling-refrigeration and air conditioning systems. In this article attention is paid to two products of Ziehl-Abigg: the Icontrol and the Fcontrol frequency converters. [Dutch] Het beperken van het energiegebruik en het verminderen van de uitstoot van het broeikasgas CO2 is voor de sector een belangrijke opgave. Door het ontwerpen en toepassen van energie-efficiente componenten zoals EC-ventilatoren (EC staat voor Electronically Commutated technologie) in allerlei koel-, vries- en luchtbehandelingsinstallaties kan veel bespaard worden in het energiegebruik. In dit artikel wordt aandacht besteed aan 2 producten van Ziehl-Abigg: de Icontrol en de Fcontrol frequentie omvormers.

  15. Differentiation and functional incorporation of embryonic stem cell-derived GABAergic interneurons in the dentate gyrus of mice with temporal lobe epilepsy.

    Science.gov (United States)

    Maisano, Xu; Litvina, Elizabeth; Tagliatela, Stephanie; Aaron, Gloster B; Grabel, Laura B; Naegele, Janice R

    2012-01-04

    Cell therapies for neurological disorders require an extensive knowledge of disease-associated neuropathology and procedures for generating neurons for transplantation. In many patients with severe acquired temporal lobe epilepsy (TLE), the dentate gyrus exhibits sclerosis and GABAergic interneuron degeneration. Mounting evidence suggests that therapeutic benefits can be obtained by transplanting fetal GABAergic progenitors into the dentate gyrus in rodents with TLE, but the scarcity of human fetal cells limits applicability in patient populations. In contrast, virtually limitless quantities of neural progenitors can be obtained from embryonic stem (ES) cells. ES cell-based therapies for neurological repair in TLE require evidence that the transplanted neurons integrate functionally and replace cell types that degenerate. To address these issues, we transplanted mouse ES cell-derived neural progenitors (ESNPs) with ventral forebrain identities into the hilus of the dentate gyrus of mice with TLE and evaluated graft differentiation, mossy fiber sprouting, cellular morphology, and electrophysiological properties of the transplanted neurons. In addition, we compared electrophysiological properties of the transplanted neurons with endogenous hilar interneurons in mice without TLE. The majority of transplanted ESNPs differentiated into GABAergic interneuron subtypes expressing calcium-binding proteins parvalbumin, calbindin, or calretinin. Global suppression of mossy fiber sprouting was not observed; however, ESNP-derived neurons formed dense axonal arborizations in the inner molecular layer and throughout the hilus. Whole-cell hippocampal slice electrophysiological recordings and morphological analyses of the transplanted neurons identified five basic types; most with strong after-hyperpolarizations and smooth or sparsely spiny dendritic morphologies resembling endogenous hippocampal interneurons. Moreover, intracellular recordings of spontaneous EPSCs indicated that

  16. Cathepsin L is required for endothelial progenitor cell-induced neovascularization

    Energy Technology Data Exchange (ETDEWEB)

    Urbich, Carmen; Heeschen, Christopher; Aicher, Alexandra; Sasaki, Ken-ichiro; Bruhl, Thomas; Hofmann, Wolf K.; Peters, Christoph; Reinheckel, Thomas; Pennacchio, Len A.; Abolmaali, Nasreddin D.; Chavakis, Emmanouil; Zeiher, Andreas M.; Dimmeler, Stefanie

    2004-01-15

    Infusion of endothelial progenitor cells (EPCs), but not of mature endothelial cells (ECs), promotes neovascularization after ischemia. We performed a gene expression profiling of EPCs and ECs to identify genes, which might be important for the neovascularization capacity of EPCs. Intriguingly, the protease cathepsin L (CathL) was highly expressed in EPCs as opposed to ECs and is essential for matrix degradation and invasion by EPCs in vitro. CathL deficient mice showed impaired functional recovery after hind limb ischemia supporting the concept for an important role of CathL in postnatal neovascularization. Infused CathL deficient progenitor cells failed to home to sites of ischemia and to augment neovascularization. In contrast, over expression of CathL in mature ECs significantly enhanced their invasive activity and induced their neovascularization capacity in vivo. Taken together, CathL plays a crucial role for the integration of circulating EPCs into the ischemic tissue and is required for neovascularization mediated by EPCs.

  17. EC tells Bonn let in UK coal

    Energy Technology Data Exchange (ETDEWEB)

    1992-06-23

    The European Commission has told the German federal government to open the door for coal imports from other EC member countries. As an initial step, the Commission is suggesting that some of the power station coal which was to be sourced from German stockpiles under the November 1991 agreement between Rurhkohle AG, the state mining company and the generators, be supplied by other member states such as the UK. The implications of this move for the German coal industry are discussed. 2 tabs.

  18. Identification of human embryonic progenitor cell targeting peptides using phage display.

    Directory of Open Access Journals (Sweden)

    Paola A Bignone

    Full Text Available Human pluripotent stem (hPS cells are capable of differentiation into derivatives of all three primary embryonic germ layers and can self-renew indefinitely. They therefore offer a potentially scalable source of replacement cells to treat a variety of degenerative diseases. The ability to reprogram adult cells to induced pluripotent stem (iPS cells has now enabled the possibility of patient-specific hPS cells as a source of cells for disease modeling, drug discovery, and potentially, cell replacement therapies. While reprogramming technology has dramatically increased the availability of normal and diseased hPS cell lines for basic research, a major bottleneck is the critical unmet need for more efficient methods of deriving well-defined cell populations from hPS cells. Phage display is a powerful method for selecting affinity ligands that could be used for identifying and potentially purifying a variety of cell types derived from hPS cells. However, identification of specific progenitor cell-binding peptides using phage display may be hindered by the large cellular heterogeneity present in differentiating hPS cell populations. We therefore tested the hypothesis that peptides selected for their ability to bind a clonal cell line derived from hPS cells would bind early progenitor cell types emerging from differentiating hPS cells. The human embryonic stem (hES cell-derived embryonic progenitor cell line, W10, was used and cell-targeting peptides were identified. Competition studies demonstrated specificity of peptide binding to the target cell surface. Efficient peptide targeted cell labeling was accomplished using multivalent peptide-quantum dot complexes as detected by fluorescence microscopy and flow cytometry. The cell-binding peptides were selective for differentiated hPS cells, had little or no binding on pluripotent cells, but preferential binding to certain embryonic progenitor cell lines and early endodermal hPS cell derivatives. Taken

  19. Vascular niche promotes hematopoietic multipotent progenitor formation from pluripotent stem cells

    Science.gov (United States)

    Gori, Jennifer L.; Butler, Jason M.; Chan, Yan-Yi; Chandrasekaran, Devikha; Poulos, Michael G.; Ginsberg, Michael; Nolan, Daniel J.; Elemento, Olivier; Wood, Brent L.; Adair, Jennifer E.; Rafii, Shahin; Kiem, Hans-Peter

    2015-01-01

    Pluripotent stem cells (PSCs) represent an alternative hematopoietic stem cell (HSC) source for treating hematopoietic disease. The limited engraftment of human PSC–derived (hPSC-derived) multipotent progenitor cells (MPP) has hampered the clinical application of these cells and suggests that MPP require additional cues for definitive hematopoiesis. We hypothesized that the presence of a vascular niche that produces Notch ligands jagged-1 (JAG1) and delta-like ligand-4 (DLL4) drives definitive hematopoiesis. We differentiated hes2 human embryonic stem cells (hESC) and Macaca nemestrina–induced PSC (iPSC) line-7 with cytokines in the presence or absence of endothelial cells (ECs) that express JAG1 and DLL4. Cells cocultured with ECs generated substantially more CD34+CD45+ hematopoietic progenitors compared with cells cocultured without ECs or with ECs lacking JAG1 or DLL4. EC-induced cells exhibited Notch activation and expressed HSC-specific Notch targets RUNX1 and GATA2. EC-induced PSC-MPP engrafted at a markedly higher level in NOD/SCID/IL-2 receptor γ chain–null (NSG) mice compared with cytokine-induced cells, and low-dose chemotherapy-based selection further increased engraftment. Long-term engraftment and the myeloid-to-lymphoid ratio achieved with vascular niche induction were similar to levels achieved for cord blood–derived MPP and up to 20-fold higher than those achieved with hPSC-derived MPP engraftment. Our findings indicate that endothelial Notch ligands promote PSC-definitive hematopoiesis and production of long-term engrafting CD34+ cells, suggesting these ligands are critical for HSC emergence. PMID:25664855

  20. The chemokine receptor CCR1 is identified in mast cell-derived exosomes.

    Science.gov (United States)

    Liang, Yuting; Qiao, Longwei; Peng, Xia; Cui, Zelin; Yin, Yue; Liao, Huanjin; Jiang, Min; Li, Li

    2018-01-01

    Mast cells are important effector cells of the immune system, and mast cell-derived exosomes carrying RNAs play a role in immune regulation. However, the molecular function of mast cell-derived exosomes is currently unknown, and here, we identify differentially expressed genes (DEGs) in mast cells and exosomes. We isolated mast cells derived exosomes through differential centrifugation and screened the DEGs from mast cell-derived exosomes, using the GSE25330 array dataset downloaded from the Gene Expression Omnibus database. Biochemical pathways were analyzed by Gene ontology (GO) annotation and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway on the online tool DAVID. DEGs-associated protein-protein interaction networks (PPIs) were constructed using the STRING database and Cytoscape software. The genes identified from these bioinformatics analyses were verified by qRT-PCR and Western blot in mast cells and exosomes. We identified 2121 DEGs (843 up and 1278 down-regulated genes) in HMC-1 cell-derived exosomes and HMC-1 cells. The up-regulated DEGs were classified into two significant modules. The chemokine receptor CCR1 was screened as a hub gene and enriched in cytokine-mediated signaling pathway in module one. Seven genes, including CCR1, CD9, KIT, TGFBR1, TLR9, TPSAB1 and TPSB2 were screened and validated through qRT-PCR analysis. We have achieved a comprehensive view of the pivotal genes and pathways in mast cells and exosomes and identified CCR1 as a hub gene in mast cell-derived exosomes. Our results provide novel clues with respect to the biological processes through which mast cell-derived exosomes modulate immune responses.

  1. An Abbreviated Protocol for In Vitro Generation of Functional Human Embryonic Stem Cell-Derived Beta-Like Cells.

    Directory of Open Access Journals (Sweden)

    Mohammad Massumi

    Full Text Available The ability to yield glucose-responsive pancreatic beta-cells from human pluripotent stem cells in vitro will facilitate the development of the cell replacement therapies for the treatment of Type 1 Diabetes. Here, through the sequential in vitro targeting of selected signaling pathways, we have developed an abbreviated five-stage protocol (25-30 days to generate human Embryonic Stem Cell-Derived Beta-like Cells (ES-DBCs. We showed that Geltrex, as an extracellular matrix, could support the generation of ES-DBCs more efficiently than that of the previously described culture systems. The activation of FGF and Retinoic Acid along with the inhibition of BMP, SHH and TGF-beta led to the generation of 75% NKX6.1+/NGN3+ Endocrine Progenitors. The inhibition of Notch and tyrosine kinase receptor AXL, and the treatment with Exendin-4 and T3 in the final stage resulted in 35% mono-hormonal insulin positive cells, 1% insulin and glucagon positive cells and 30% insulin and NKX6.1 co-expressing cells. Functionally, ES-DBCs were responsive to high glucose in static incubation and perifusion studies, and could secrete insulin in response to successive glucose stimulations. Mitochondrial metabolic flux analyses using Seahorse demonstrated that the ES-DBCs could efficiently metabolize glucose and generate intracellular signals to trigger insulin secretion. In conclusion, targeting selected signaling pathways for 25-30 days was sufficient to generate ES-DBCs in vitro. The ability of ES-DBCs to secrete insulin in response to glucose renders them a promising model for the in vitro screening of drugs, small molecules or genes that may have potential to influence beta-cell function.

  2. A novel method to generate and culture human mast cells: Peripheral CD34+ stem cell-derived mast cells (PSCMCs).

    Science.gov (United States)

    Schmetzer, Oliver; Valentin, Patricia; Smorodchenko, Anna; Domenis, Rossana; Gri, Giorgia; Siebenhaar, Frank; Metz, Martin; Maurer, Marcus

    2014-11-01

    The identification and characterization of human mast cell (MC) functions are hindered by the shortage of MC populations suitable for investigation. Here, we present a novel technique for generating large numbers of well differentiated and functional human MCs from peripheral stem cells (=peripheral stem cell-derived MCs, PSCMCs). Innovative and key features of this technique include 1) the use of stem cell concentrates, which are routinely discarded by blood banks, as the source of CD34+ stem cells, 2) cell culture in serum-free medium and 3) the addition of LDL as well as selected cytokines. In contrast to established and published protocols that use CD34+ or CD133+ progenitor cells from full blood, we used a pre-enriched cell population obtained from stem cell concentrates, which yielded up to 10(8) differentiated human MCs per batch after only three weeks of culture starting with 10(6) total CD34+ cells. The total purity on MCs (CD117+, FcεR1+) generated by this method varied between 55 and 90%, of which 4-20% were mature MCs that contain tryptase and chymase and show expression of FcεRI and CD117 in immunohistochemistry. PSCMCs showed robust histamine release in response to stimulation with anti-FcεR1 or IgE/anti-IgE, and increased proliferation and differentiation in response to IL-1β or IFN-γ. Taken together, this new protocol of the generation of large numbers of human MCs provides for an innovative and suitable option to investigate the biology of human MCs. Copyright © 2014 Elsevier B.V. All rights reserved.

  3. The poster as modernist progenitor

    Directory of Open Access Journals (Sweden)

    Katherine Hauser

    2015-12-01

    Full Text Available Ruth E. Iskin’s The Poster: Art, Advertising. Design, and Collecting, 1860s-1900s positions the late-nineteenth-century advertising poster as the progenitor of valued modernist practices typically attached solely to photography and film. Modernist biases separating high art from mass culture account for scholars ignoring posters, however the poster ushered in an innovative reductive graphic style as well as pioneered the notion of multiple originals.

  4. PROGENITORS OF RECOMBINING SUPERNOVA REMNANTS

    Energy Technology Data Exchange (ETDEWEB)

    Moriya, Takashi J., E-mail: takashi.moriya@ipmu.jp [Kavli Institute for the Physics and Mathematics of the Universe, Todai Institutes for Advanced Study, University of Tokyo, Kashiwanoha 5-1-5, Kashiwa, Chiba 277-8583 (Japan)

    2012-05-01

    Usual supernova remnants have either ionizing plasma or plasma in collisional ionization equilibrium, i.e., the ionization temperature is lower than or equal to the electron temperature. However, the existence of recombining supernova remnants, i.e., supernova remnants with ionization temperature higher than the electron temperature, has been recently confirmed. One suggested way to have recombining plasma in a supernova remnant is to have a dense circumstellar medium at the time of the supernova explosion. If the circumstellar medium is dense enough, collisional ionization equilibrium can be established in the early stage of the evolution of the supernova remnant and subsequent adiabatic cooling, which occurs after the shock wave gets out of the dense circumstellar medium, makes the electron temperature lower than the ionization temperature. We study the circumstellar medium around several supernova progenitors and show which supernova progenitors can have a circumstellar medium dense enough to establish collisional ionization equilibrium soon after the explosion. We find that the circumstellar medium around red supergiants (especially massive ones) and the circumstellar medium dense enough to make Type IIn supernovae can establish collisional ionization equilibrium soon after the explosion and can evolve to become recombining supernova remnants. Wolf-Rayet stars and white dwarfs have the possibility to be recombining supernova remnants but the fraction is expected to be very small. As the occurrence rate of the explosions of red supergiants is much higher than that of Type IIn supernovae, the major progenitors of recombining supernova remnants are likely to be red supergiants.

  5. Application of ECS to a hybrid automobile; `ECS` no hybrid jidosha eno tekiyo

    Energy Technology Data Exchange (ETDEWEB)

    Shimizu, M.; Okamura, T.

    1997-01-30

    This paper describes a simulation for a case when an energy capacitor system (ECS) is mounted in a hybrid electric vehicle in place of secondary batteries. The ECS is electric power storage equipment consisted of capacitors and electronic circuits, and capable of quick charge and discharge at high efficiency. Its energy density per unit weight is about the same as lead-acid batteries, but its output density is greater than any type of batteries. Being free of deterioration due to charge and discharge, its cycle life is almost limitless. In order to suppress heat generation, a switching converter is used to control the output to about 300W. However, internal resistance of an ECS is smaller than in an equivalent secondary battery, and internal loss and heat generation stay as low as 1/5 to 1/10. Discussions have been given on cases of using the ECS in small passenger cars, small buses/trucks, and large buses. Since the ECS can use the stored power to about 94% of the capacity, and its cycle life is sufficiently long, it can operate with charge capacity as small as 1/4 of a lead-acid battery. The weight can also be reduced largely. Regenerative power storage efficiency is high, fuel consumption can be improved, heat generation is less, and no forced cooling is required. 9 refs., 4 figs., 1 tab.

  6. Permeation of Telone EC through protective gloves.

    Science.gov (United States)

    Zainal, Hanaa; Que Hee, Shane S

    2005-09-30

    Telone is a potent fumigant that is based on the chlorinated unsaturated hydrocarbon, 1,3-dichloropropene (1,3-DCP). It is often applied without dilution and so poses severe inhalation and air pollution threats. Urinary metabolites of 1,3-DCP have been detected after Telone skin exposure, so that preventing dermal exposure is also important. The objective of the study was to assess if nitrile and multi-layer ("laminated") gloves provide adequate protection against Telone EC formulation. To accomplish this, disposable (Safeskin) and chemically resistant (Sol-Vex) nitrile and laminated (Barrier mark and Silver Shield) glove materials were challenged by Telone EC with hexane liquid collection in an ASTM-type I-PTC-600 permeation cell. Analyses of cis- and trans-1,3-DCP in the collection fluid at specified times were performed on a moderately polar capillary column by gas chromatography-electron capture detection. Telone EC caused microholes in both nitrile materials, though the chemically protective material was degraded slower than the disposable nitrile. The laminated gloves offered limited protection. Silver Shield protected best because 1.5-2.3 mg 1,3-DCP permeated by 8 h relative to 2.5-7.6 mg for Barrier, implying about 2.5 times more protection for 8 h. Even for Silver Shield, the extent of protection was inadequate as illustrated by a risk assessment of the skin exposure situation. The normalized breakthrough times for both types of laminated gloves varied between 27 and 60 min. It is recommended that Viton gloves still be worn for protection.

  7. Involvement of multiple myeloma cell-derived exosomes in osteoclast differentiation

    Science.gov (United States)

    Raimondi, Lavinia; De Luca, Angela; Amodio, Nicola; Manno, Mauro; Raccosta, Samuele; Taverna, Simona; Bellavia, Daniele; Naselli, Flores; Fontana, Simona; Schillaci, Odessa; Giardino, Roberto; Fini, Milena; Tassone, Pierfrancesco; Santoro, Alessandra; De Leo, Giacomo; Giavaresi, Gianluca; Alessandro, Riccardo

    2015-01-01

    Bone disease is the most frequent complication in multiple myeloma (MM) resulting in osteolytic lesions, bone pain, hypercalcemia and renal failure. In MM bone disease the perfect balance between bone-resorbing osteoclasts (OCs) and bone-forming osteoblasts (OBs) activity is lost in favour of OCs, thus resulting in skeletal disorders. Since exosomes have been described for their functional role in cancer progression, we here investigate whether MM cell-derived exosomes may be involved in OCs differentiation. We show that MM cells produce exosomes which are actively internalized by Raw264.7 cell line, a cellular model of osteoclast formation. MM cell-derived exosomes positively modulate pre-osteoclast migration, through the increasing of CXCR4 expression and trigger a survival pathway. MM cell-derived exosomes play a significant pro-differentiative role in murine Raw264.7 cells and human primary osteoclasts, inducing the expression of osteoclast markers such as Cathepsin K (CTSK), Matrix Metalloproteinases 9 (MMP9) and Tartrate-resistant Acid Phosphatase (TRAP). Pre-osteoclast treated with MM cell-derived exosomes differentiate in multinuclear OCs able to excavate authentic resorption lacunae. Similar results were obtained with exosomes derived from MM patient's sera. Our data indicate that MM-exosomes modulate OCs function and differentiation. Further studies are needed to identify the OCs activating factors transported by MM cell-derived exosomes. PMID:25944696

  8. Development of EC technology in JAERI

    International Nuclear Information System (INIS)

    Sakamoto, Keishi; Takahashi, Koji; Kasugai, Atsushi; Hayashi, Kenichi; Ikeda, Yoshitaka; Kajiwara, Ken; Fujii, Tsuneyuki; Imai, Tsuyoshi; Kariya, Tsuyoshi; Mitsunaka, Yoshika

    2003-01-01

    Recent progress of electron cyclotron (EC) heating and current drive technologies in JAERI is reported. In 170 GHz gyrotron development, 0.9 MW/9.2 sec (efficiency: 43%), 0.5 MW/30 sec (46%), etc, have been demonstrated. As for 110 GHz gyrotron, 1 MW/5 sec and 1.2 MW/4.1 sec were obtained. Using four 110 GHz gyrotrons, a 3 MW power injection into JT- 60U plasma was carried out. In parallel, a launcher design and its development that includes neutron irradiation of the launcher components and a remote steering launcher are underway for ITER application. (authors)

  9. Adhesion behavior of endothelial progenitor cells to endothelial cells in simple shear flow

    Science.gov (United States)

    Gong, Xiao-Bo; Li, Yu-Qing; Gao, Quan-Chao; Cheng, Bin-Bin; Shen, Bao-Rong; Yan, Zhi-Qiang; Jiang, Zong-Lai

    2011-12-01

    The adhesion of endothelial progenitor cells (EPCs) on endothelial cells (ECs) is one of the critical physiological processes for the regenesis of vascular vessels and the prevention of serious cardiovascular diseases. Here, the rolling and adhesion behavior of EPCs on ECs was studied numerically. A two-dimensional numerical model was developed based on the immersed boundary method for simulating the rolling and adhesion of cells in a channel flow. The binding force arising from the catch bond of a receptor and ligand pair was modeled with stochastic Monte Carlo method and Hookean spring model. The effect of tumor necrosis factor alpha (TNF- α) on the expression of the number of adhesion molecules in ECs was analyzed experimentally. A flow chamber system with CCD camera was set up to observe the top view of the rolling of EPCs on the substrate cultivated with ECs. Numerical results prove that the adhesion of EPC on ECs is closely related to membrane stiffness of the cell and shear rate of the flow. It also suggests that the adhesion force between EPC and EC by P-selectin glycoprotein ligand-1 only is not strong enough to bond the cell onto vessel walls unless contributions of other catch bond are considered. Experimental results demonstrate that TNF- α enhanced the expressions of VCAM, ICAM, P-selectin and E-selectin in ECs, which supports the numerical results that the rolling velocity of EPC on TNF- α treated EC substrate decreases obviously compared with its velocity on the untreated one. It is found that because the adhesion is affected by both the rolling velocity and the deformability of the cell, an optimal stiffness of EPC may exist at a given shear rate of flow for achieving maximum adhesion rates.

  10. EC Competition law and environmental protection

    Energy Technology Data Exchange (ETDEWEB)

    Rantala, M.

    1996-12-31

    Due to the increasing attention paid on environmental protection, by the European Community, the Member States and the consumers, undertakings are forced to adjust their policies to meet the new demands posed by public authorities and the market. This has an inevitable impact on competition. No matter how acceptable the goals were, the means are to be assessed also from the competition law point of view. The objective of this study is to analyze the situations where conflicts may take place, the main sources of research being the principles of law relating to the Treaty ,the judgements of the European Court of Justice, the Commission Decisions and other official documents of the Community as well as legal literature. This study classifies the Community and Member State instruments which are used for environmental protection into four categories. These are legislative, market-based, horizontal and financial supporting instruments. Undertakings` environmental practices, which are capable of affecting competition, are examined under Articles 85 and 86 EC, some attention being paid also to Article 90 EC. The touching points of competition policy and environmental protection in such cases are scrutinized under three headings. These are the two, in this case, most relevant public instruments, ie. legislative and market-based measures, and the voluntary actions of undertakings. (42 refs.)

  11. Cardiotoxicity evaluation using human embryonic stem cells and induced pluripotent stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Zhao, Qi; Wang, Xijie; Wang, Shuyan; Song, Zheng; Wang, Jiaxian; Ma, Jing

    2017-03-09

    Cardiotoxicity remains an important concern in drug discovery. Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) have become an attractive platform to evaluate cardiotoxicity. However, the consistency between human embryonic stem cell-derived cardiomyocytes (hESC-CMs) and human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) in prediction of cardiotoxicity has yet to be elucidated. Here we screened the toxicities of four representative drugs (E-4031, isoprenaline, quinidine, and haloperidol) using both hESC-CMs and hiPSC-CMs, combined with an impedance-based bioanalytical method. It showed that both hESC-CMs and hiPSC-CMs can recapitulate cardiotoxicity and identify the effects of well-characterized compounds. The combined platform of hPSC-CMs and an impedance-based bioanalytical method could improve preclinical cardiotoxicity screening, holding great potential for increasing drug development accuracy.

  12. Wnt signaling positively regulates endothelial cell fate specification in the Fli1a-positive progenitor population via Lef1.

    Science.gov (United States)

    Hübner, Kathleen; Grassme, Kathrin S; Rao, Jyoti; Wenke, Nina K; Zimmer, Cordula L; Korte, Laura; Mu Ller, Katja; Sumanas, Saulius; Greber, Boris; Herzog, Wiebke

    2017-10-01

    During vertebrate embryogenesis, vascular endothelial cells (ECs) and primitive erythrocytes become specified within close proximity in the posterior lateral plate mesoderm (LPM) from a common progenitor. However, the signaling cascades regulating the specification into either lineage remain largely elusive. Here, we analyze the contribution of β-catenin dependent Wnt signaling to EC and erythrocyte specification during zebrafish embryogenesis. We generated novel β-catenin dependent Wnt signaling reporters which, by using destabilized fluorophores (Venus-Pest, dGFP), specifically allow us to detect Wnt signaling responses in narrow time windows as well as in spatially restricted domains, defined by Cre recombinase expression (Tg(axin2 BAC :Venus-Pest) mu288 ; Tg(14TCF:loxP-STOP-loxP-dGFP) mu202 ). We therefore can detect β-catenin dependent Wnt signaling activity in a subset of the Fli1a-positive progenitor population. Additionally, we show that mesodermal Wnt3a-mediated signaling via the transcription factor Lef1 positively regulates EC specification (defined by kdrl expression) at the expense of primitive erythrocyte specification (defined by gata1 expression) in zebrafish embryos. Using mesoderm derived from human embryonic stem cells, we identified the same principle of Wnt signaling dependent EC specification in conjunction with auto-upregulation of LEF1. Our data indicate a novel role of β-catenin dependent Wnt signaling in regulating EC specification during vasculogenesis. Copyright © 2017. Published by Elsevier Inc.

  13. T cell-derived Lymphotoxin is Essential for anti-HSV-1 Humoral Immune Response.

    Science.gov (United States)

    Yang, Kaiting; Liang, Yong; Sun, Zhichen; Xue, Diyuan; Xu, Hairong; Zhu, Mingzhao; Fu, Yang-Xin; Peng, Hua

    2018-05-09

    B cell-derived lymphotoxin (LT) is required for the development of follicular dendritic cell clusters for the formation of primary and secondary lymphoid follicles, but the role of T cell-derived LT in antibody response has not been well demonstrated. We observed that lymphotoxin-β-receptor (LTβR) signaling is essential for optimal humoral immune response and protection against an acute HSV-1 infection. Blocking the LTβR pathway caused poor maintenance of germinal center B (GC-B) cells and follicular helper T (Tfh) cells. Using bone marrow chimeric mice and adoptive transplantation, we determined that T cell-derived LT played an indispensable role in the humoral immune response to HSV-1. Up-regulation of IFNγ by the LTβR-Ig blockade impairs the sustainability of Tfh-like cells, thus leading to an impaired humoral immune response. Our findings have identified a novel role of T cell-derived LT in the humoral immune response against HSV-1 infection. IMPORTANCE Immunocompromised people are susceptible for HSV-1 infection and lethal recurrence, which could be inhibited by anti-HSV-1 humoral immune response in the host. This study sought to explore the role of T cell-derived LT in the anti-HSV-1 humoral immune response using LT-LTβR signaling deficient mice and the LTβR-Ig blockade. The data indicate that the T cell-derived LT may play an essential role in sustaining Tfh-like cells and ensure Tfh-like cells' migration into primary or secondary follicles for further maturation. This study provides insights for vaccine development against infectious diseases. Copyright © 2018 American Society for Microbiology.

  14. Solving the puzzle of pluripotent stem cell-derived cardiomyocyte maturation: piece by piece.

    Science.gov (United States)

    Lundy, David J; Lee, Desy S; Hsieh, Patrick C H

    2017-03-01

    There is a growing need for in vitro models which can serve as platforms for drug screening and basic research. Human adult cardiomyocytes cannot be readily obtained or cultured, and so pluripotent stem cell-derived cardiomyocytes appear to be an attractive option. Unfortunately, these cells are structurally and functionally immature-more comparable to foetal cardiomyocytes than adult. A recent study by Ruan et al ., provides new insights into accelerating the maturation process and takes us a step closer to solving the puzzle of pluripotent stem cell-derived cardiomyocyte maturation.

  15. Classification of follicular cell-derived thyroid cancer by global RNA profiling

    DEFF Research Database (Denmark)

    Rossing, Maria

    2013-01-01

    The incidence of thyroid cancer is increasing worldwide and thyroid nodules are a frequent clinical finding. Diagnosing follicular cell-derived cancers is, however, challenging both histopathologically and especially cytopathologically. The advent of high-throughput molecular technologies has...... profiling of follicular cell-derived thyroid cancers....... prompted many researchers to explore the transcriptome and, in recent years, also the miRNome in order to generate new molecular classifiers capable of classifying thyroid tumours more accurately than by conventional cytopathological and histopathological methods. This has led to a number of molecular...

  16. Differentiation, Evaluation, and Application of Human Induced Pluripotent Stem Cell-Derived Endothelial Cells.

    Science.gov (United States)

    Lin, Yang; Gil, Chang-Hyun; Yoder, Mervin C

    2017-11-01

    The emergence of induced pluripotent stem cell (iPSC) technology paves the way to generate large numbers of patient-specific endothelial cells (ECs) that can be potentially delivered for regenerative medicine in patients with cardiovascular disease. In the last decade, numerous protocols that differentiate EC from iPSC have been developed by many groups. In this review, we will discuss several common strategies that have been optimized for human iPSC-EC differentiation and subsequent studies that have evaluated the potential of human iPSC-EC as a cell therapy or as a tool in disease modeling. In addition, we will emphasize the importance of using in vivo vessel-forming ability and in vitro clonogenic colony-forming potential as a gold standard with which to evaluate the quality of human iPSC-EC derived from various protocols. © 2017 American Heart Association, Inc.

  17. Macrophages control vascular stem/progenitor cell plasticity through tumor necrosis factor-α-mediated nuclear factor-κB activation.

    Science.gov (United States)

    Wong, Mei Mei; Chen, Yikuan; Margariti, Andriani; Winkler, Bernhard; Campagnolo, Paola; Potter, Claire; Hu, Yanhua; Xu, Qingbo

    2014-03-01

    Vascular lineage differentiation of stem/progenitor cells can contribute to both tissue repair and exacerbation of vascular diseases such as in vein grafts. The role of macrophages in controlling vascular progenitor differentiation is largely unknown and may play an important role in graft development. This study aims to identify the role of macrophages in vascular stem/progenitor cell differentiation and thereafter elucidate the mechanisms that are involved in the macrophage- mediated process. We provide in vitro evidence that macrophages can induce endothelial cell (EC) differentiation of the stem/progenitor cells while simultaneously inhibiting their smooth muscle cell differentiation. Mechanistically, both effects were mediated by macrophage-derived tumor necrosis factor-α (TNF-α) via TNF-α receptor 1 and canonical nuclear factor-κB activation. Although the overexpression of p65 enhanced EC (or attenuated smooth muscle cell) differentiation, p65 or TNF-α receptor 1 knockdown using lentiviral short hairpin RNA inhibited EC (or rescued smooth muscle cell) differentiation in response to TNF-α. Furthermore, TNF-α-mediated EC differentiation was driven by direct binding of nuclear factor-κB (p65) to specific VE-cadherin promoter sequences. Subsequent experiments using an ex vivo decellularized vessel scaffold confirmed an increase in the number of ECs and reduction in smooth muscle cell marker expression in the presence of TNF-α. The lack of TNF-α in a knockout mouse model of vein graft decreased endothelialization and significantly increased thrombosis formation. Our study highlights the role of macrophages in directing vascular stem/progenitor cell lineage commitment through TNF-α-mediated TNF-α receptor 1 and nuclear factor-κB activation that is likely required for endothelial repair in vascular diseases such as vein graft.

  18. Ca2+-currents in human induced pluripotent stem cell-derived cardiomyocytes - effects of two different culture conditions

    Directory of Open Access Journals (Sweden)

    Ahmet Umur Uzun

    2016-09-01

    Full Text Available Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM provide a unique opportunity to study human heart physiology and pharmacology and repair injured hearts. The suitability of hiPSC-CM critically depends on how closely they share physiological properties of human adult cardiomyocytes (CM. Here we investigated whether a 3D engineered heart tissue (EHT culture format favors maturation and addressed the L-type Ca2+-current (ICa,L as a readout. The results were compared with hiPSC-CM cultured in conventional monolayer (ML and to our previous data from human adult atrial and ventricular CM obtained when identical patch-clamp protocols were used. HiPSC-CM were 2-3 fold smaller than adult CM, independently of culture format (capacitance ML 45±1 pF (n=289, EHT 45±1 pF (n=460, atrial CM 87±3 pF (n=196, ventricular CM 126±8 pF (n=50. Only 88% of ML cells showed ICa, but all EHT. Basal ICa density was 10±1 pA/pF (n=207 for ML and 12±1 pA/pF (n=361 for EHT and was larger than in adult CM (7±1 pA/pF (p<0.05, n=196 for atrial CM and 6±1 pA/pF (p<0.05, n=47 for ventricular CM. However, ML and EHT showed robust T-type Ca2+-currents (ICa,T. While (--Bay K 8644, that activates ICa,L directly, increased ICa,L to the same extent in ML and EHT, β1- and β2-adrenoceptor effects were marginal in ML, but of same size as (--Bay K 8644 in EHT. The opposite was true for serotonin receptors. Sensitivity to β1 and β2-adrenoceptor stimulation was the same in EHT as in adult CM (-logEC50: 5.9 and 6.1 for norepinephrine (NE and epinephrine (Epi, respectively, but very low concentrations of Rp-8-Br-cAMPS were sufficient to suppress effects (-logEC50: 5.3 and 5.3 respectively for NE and Epi. Taken together, hiPSC-CM express ICa,L at the same density as human adult CM, but, in contrast, possess robust ICa,T. Increased effects of catecholamines in EHT suggest more efficient maturation.

  19. (EC+β+) decay of 130Ce

    International Nuclear Information System (INIS)

    Xu Shuwei; Zhang Tianmei; Xie Yuanxiang; Ma Ruichang; Ge Yuanxiu; Guo Yingxiang; Wang Chunfang; Li Zhankui; Guo Bing; Xing Jianping; Guo Tianrui; Zhu Shaofei; Xu Wang; Du Jinzhou

    1996-01-01

    The nuclide 130 Ce was produced by a ( 16 O, 4n) reaction on an enriched 118 Sn target. Reaction products were transported to a shielded location by using a helium-jet tape transport system. A 22.9 min activity in chemically separated cerium sample was identified as 130 Ce. The (EC+β + ) decay scheme of 130 Ce was proposed for the first time. This scheme includes 108 γ-lines, 107 γ-lines among them being new. More than 13 1 + low-lying states of 130 La are populated in the decay of 130 Ce. Two new isomers with half-life of 77±10 ns and 17±5 ns were observed by means of delayed γ-γ coincidence measurements. (orig.). With 5 figs., 3 tabs

  20. Endothelial jagged-2 sustains hematopoietic stem and progenitor reconstitution after myelosuppression.

    Science.gov (United States)

    Guo, Peipei; Poulos, Michael G; Palikuqi, Brisa; Badwe, Chaitanya R; Lis, Raphael; Kunar, Balvir; Ding, Bi-Sen; Rabbany, Sina Y; Shido, Koji; Butler, Jason M; Rafii, Shahin

    2017-12-01

    Angiocrine factors, such as Notch ligands, supplied by the specialized endothelial cells (ECs) within the bone marrow and splenic vascular niche play an essential role in modulating the physiology of adult hematopoietic stem and progenitor cells (HSPCs). However, the relative contribution of various Notch ligands, specifically jagged-2, to the homeostasis of HSPCs is unknown. Here, we show that under steady state, jagged-2 is differentially expressed in tissue-specific vascular beds, but its expression is induced in hematopoietic vascular niches after myelosuppressive injury. We used mice with EC-specific deletion of the gene encoding jagged-2 (Jag2) to demonstrate that while EC-derived jagged-2 was dispensable for maintaining the capacity of HSPCs to repopulate under steady-state conditions, by activating Notch2 it did contribute to the recovery of HSPCs in response to myelosuppressive conditions. Engraftment and/or expansion of HSPCs was dependent on the expression of endothelial-derived jagged-2 following myeloablation. Additionally, jagged-2 expressed in bone marrow ECs regulated HSPC cell cycle and quiescence during regeneration. Endothelial-deployed jagged-2 triggered Notch2/Hey1, while tempering Notch2/Hes1 signaling in HSPCs. Collectively, these data demonstrate that EC-derived jagged-2 activates Notch2 signaling in HSPCs to promote hematopoietic recovery and has potential as a therapeutic target to accelerate balanced hematopoietic reconstitution after myelosuppression.

  1. Consumption of food in EC countries

    International Nuclear Information System (INIS)

    Ven-Breken, T.J. van de; Brenot, J.; Bonnefous, S.; Noordijk, H.; Leenhouts, H.P.

    1990-08-01

    Annual consumption of 10 classes of food and drink is reported for the total population of each EC-country. National food consumption data are generalized by distinguishing groups of countries with similar diet. Group 1: UK and Ireland (low consumption of fruit, high consumption of potatoes and fresh dairy products). Group 2: Italy and Greece (high consumption of cereals, vegetables and fruit, low consumption of fresh dairy products). Group 3: France, Belgium, Luxembourg, Netherlands, FRG and Spain (intermediate consumption of most foodstuffs). Portugal and Denmark could not be classified in any group. Changes in food consumption related to differences in age, sex, social class, regional habits and level of urbanisation are reported.In some cases, variation of annual food consumption over the different seasons on the relative contribution of food from local origin to the total diet, could be estimated. Differences of region, age and level of urbanisation appear to have the largest influence on consumed amounts. As for age, the values for the different age-groups range from 50 to 80 percent of the adult diet. Regional consumption may diverge from 70-130 percent of the average national consumption, with values of 200 percent for potatoes and fruit. Consumption in urban areas compared with the entire country diverges from 70-100 percent. In rural areas the values diverge from 100-140 percent. Self-support of vegetables and milk seems to be important for most countries. Of the other products, self-support of wheat and cheese is only of importance in the south of the EC. When combining the effects of the different factors, a range of consumed amounts for specific population groups of 50 to 200 percent of the average national consumption was determined. (author). 38 refs.; 13 figs.; 75 tabs

  2. Generation of Oligodendrogenic Spinal Neural Progenitor Cells From Human Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Khazaei, Mohamad; Ahuja, Christopher S; Fehlings, Michael G

    2017-08-14

    This unit describes protocols for the efficient generation of oligodendrogenic neural progenitor cells (o-NPCs) from human induced pluripotent stem cells (hiPSCs). Specifically, detailed methods are provided for the maintenance and differentiation of hiPSCs, human induced pluripotent stem cell-derived neural progenitor cells (hiPS-NPCs), and human induced pluripotent stem cell-oligodendrogenic neural progenitor cells (hiPSC-o-NPCs) with the final products being suitable for in vitro experimentation or in vivo transplantation. Throughout, cell exposure to growth factors and patterning morphogens has been optimized for both concentration and timing, based on the literature and empirical experience, resulting in a robust and highly efficient protocol. Using this derivation procedure, it is possible to obtain millions of oligodendrogenic-NPCs within 40 days of initial cell plating which is substantially shorter than other protocols for similar cell types. This protocol has also been optimized to use translationally relevant human iPSCs as the parent cell line. The resultant cells have been extensively characterized both in vitro and in vivo and express key markers of an oligodendrogenic lineage. © 2017 by John Wiley & Sons, Inc. Copyright © 2017 John Wiley and Sons, Inc.

  3. Regulatory Systems in Bone Marrow for Hematopoietic Stem/Progenitor Cells Mobilization and Homing

    Directory of Open Access Journals (Sweden)

    P. Alvarez

    2013-01-01

    Full Text Available Regulation of hematopoietic stem cell release, migration, and homing from the bone marrow (BM and of the mobilization pathway involves a complex interaction among adhesion molecules, cytokines, proteolytic enzymes, stromal cells, and hematopoietic cells. The identification of new mechanisms that regulate the trafficking of hematopoietic stem/progenitor cells (HSPCs cells has important implications, not only for hematopoietic transplantation but also for cell therapies in regenerative medicine for patients with acute myocardial infarction, spinal cord injury, and stroke, among others. This paper reviews the regulation mechanisms underlying the homing and mobilization of BM hematopoietic stem/progenitor cells, investigating the following issues: (a the role of different factors, such as stromal cell derived factor-1 (SDF-1, granulocyte colony-stimulating factor (G-CSF, and vascular cell adhesion molecule-1 (VCAM-1, among other ligands; (b the stem cell count in peripheral blood and BM and influential factors; (c the therapeutic utilization of this phenomenon in lesions in different tissues, examining the agents involved in HSPCs mobilization, such as the different forms of G-CSF, plerixafor, and natalizumab; and (d the effects of this mobilization on BM-derived stem/progenitor cells in clinical trials of patients with different diseases.

  4. A novel method of mouse ex utero transplantation of hepatic progenitor cells into the fetal liver

    International Nuclear Information System (INIS)

    Shikanai, Mima; Asahina, Kinji; Iseki, Sachiko; Teramoto, Kenichi; Nishida, Tomohiro; Shimizu-Saito, Keiko; Ota, Masato; Eto, Kazuhiro; Teraoka, Hirobumi

    2009-01-01

    Avoiding the limitations of the adult liver niche, transplantation of hepatic stem/progenitor cells into fetal liver is desirable to analyze immature cells in a hepatic developmental environment. Here, we established a new monitor tool for cell fate of hepatic progenitor cells transplanted into the mouse fetal liver by using ex utero surgery. When embryonic day (ED) 14.5 hepatoblasts were injected into the ED14.5 fetal liver, the transplanted cells expressed albumin abundantly or α-fetoprotein weakly, and contained glycogen in the neonatal liver, indicating that transplanted hepatoblasts can proliferate and differentiate in concord with surrounding recipient parenchymal cells. The transplanted cells became mature in the liver of 6-week-old mice. Furthermore, this method was applicable to transplantation of hepatoblast-like cells derived from mouse embryonic stem cells. These data indicate that this unique technique will provide a new in vivo experimental system for studying cell fate of hepatic stem/progenitor cells and liver organogenesis.

  5. A novel mechanism of bacterial toxin transfer within host blood cell-derived microvesicles.

    Directory of Open Access Journals (Sweden)

    Anne-lie Ståhl

    2015-02-01

    Full Text Available Shiga toxin (Stx is the main virulence factor of enterohemorrhagic Escherichia coli, which are non-invasive strains that can lead to hemolytic uremic syndrome (HUS, associated with renal failure and death. Although bacteremia does not occur, bacterial virulence factors gain access to the circulation and are thereafter presumed to cause target organ damage. Stx was previously shown to circulate bound to blood cells but the mechanism by which it would potentially transfer to target organ cells has not been elucidated. Here we show that blood cell-derived microvesicles, shed during HUS, contain Stx and are found within patient renal cortical cells. The finding was reproduced in mice infected with Stx-producing Escherichia coli exhibiting Stx-containing blood cell-derived microvesicles in the circulation that reached the kidney where they were transferred into glomerular and peritubular capillary endothelial cells and further through their basement membranes followed by podocytes and tubular epithelial cells, respectively. In vitro studies demonstrated that blood cell-derived microvesicles containing Stx undergo endocytosis in glomerular endothelial cells leading to cell death secondary to inhibited protein synthesis. This study demonstrates a novel virulence mechanism whereby bacterial toxin is transferred within host blood cell-derived microvesicles in which it may evade the host immune system.

  6. Single-Cell Functional Analysis of Stem-Cell Derived Cardiomyocytes on Micropatterned Flexible Substrates

    NARCIS (Netherlands)

    Kijlstra, Jan David; Hu, Dongjian; van der Meer, Peter; Domian, Ibrahim J

    2017-01-01

    Human pluripotent stem-cell derived cardiomyocytes (hPSC-CMs) hold great promise for applications in human disease modeling, drug discovery, cardiotoxicity screening, and, ultimately, regenerative medicine. The ability to study multiple parameters of hPSC-CM function, such as contractile and

  7. Application of stem cell-derived retinal pigmented epithelium in retinal degenerative diseases: present and future

    Directory of Open Access Journals (Sweden)

    Mingyue Luo

    2018-01-01

    Full Text Available As a constituent of blood-retinal barrier and retinal outer segment (ROS scavenger, retinal pigmented epithelium (RPE is fundamental to normal function of retina. Malfunctioning of RPE contributes to the onset and advance of retinal degenerative diseases. Up to date, RPE replacement therapy is the only possible method to completely reverse retinal degeneration. Transplantation of human RPE stem cell-derived RPE (hRPESC-RPE has shown some good results in animal models. With promising results in terms of safety and visual improvement, human embryonic stem cell-derived RPE (hESC-RPE can be expected in clinical settings in the near future. Despite twists and turns, induced pluripotent stem cell-derived RPE (iPSC-RPE is now being intensely investigated to overcome genetic and epigenetic instability. By far, only one patient has received iPSC-RPE transplant, which is a hallmark of iPSC technology development. During follow-up, no major complications such as immunogenicity or tumorigenesis have been observed. Future trials should keep focusing on the safety of stem cell-derived RPE (SC-RPE especially in long period, and better understanding of the nature of stem cell and the molecular events in the process to generate SC-RPE is necessary to the prosperity of SC-RPE clinical application.

  8. Derivation and characterization of a pig embryonic stem cell-derived exocrine pancreatic cell line

    Science.gov (United States)

    The establishment and initial characterization of a pig embryonic stem cell-derived pancreatic cell line, PICM-31, and a colony-cloned derivative cell line, PICM-31A, is described. The cell lines were propagated for several months at split ratios of 1:3 or 1:5 at each passage on STO feeder cells af...

  9. Bioinspired Cell-Derived Nanovesicles versus Exosomes as Drug Delivery Systems : A Cost-Effective Alternative

    NARCIS (Netherlands)

    Goh, Wei Jiang; Zou, Shui; Ong, Wei Yi; Torta, Federico; Alexandra, Alvarez Fernandez; Schiffelers, Raymond M.; Storm, Gert; Wang, Jiong-Wei; Czarny, Bertrand M S; Pastorin, Giorgia

    2017-01-01

    Cell Derived Nanovesicles (CDNs) have been developed from the rapidly expanding field of exosomes, representing a class of bioinspired Drug Delivery Systems (DDS). However, translation to clinical applications is limited by the low yield and multi-step approach in isolating naturally secreted

  10. Bioinspired Cell-Derived Nanovesicles versus Exosomes as Drug Delivery Systems : a Cost-Effective Alternative

    NARCIS (Netherlands)

    Goh, Wei Jiang; Zou, Shui; Ong, Wei Yi; Torta, Federico; Alexandra, Alvarez Fernandez; Schiffelers, Raymond M; Storm, G; Wang, Jiong Wei; Czarny, Bertrand; Pastorin, Giorgia

    2017-01-01

    Cell Derived Nanovesicles (CDNs) have been developed from the rapidly expanding field of exosomes, representing a class of bioinspired Drug Delivery Systems (DDS). However, translation to clinical applications is limited by the low yield and multi-step approach in isolating naturally secreted

  11. Application of stem cell-derived retinal pigmented epithelium in retinal degenerative diseases: present and future.

    Science.gov (United States)

    Luo, Mingyue; Chen, Youxin

    2018-01-01

    As a constituent of blood-retinal barrier and retinal outer segment (ROS) scavenger, retinal pigmented epithelium (RPE) is fundamental to normal function of retina. Malfunctioning of RPE contributes to the onset and advance of retinal degenerative diseases. Up to date, RPE replacement therapy is the only possible method to completely reverse retinal degeneration. Transplantation of human RPE stem cell-derived RPE (hRPESC-RPE) has shown some good results in animal models. With promising results in terms of safety and visual improvement, human embryonic stem cell-derived RPE (hESC-RPE) can be expected in clinical settings in the near future. Despite twists and turns, induced pluripotent stem cell-derived RPE (iPSC-RPE) is now being intensely investigated to overcome genetic and epigenetic instability. By far, only one patient has received iPSC-RPE transplant, which is a hallmark of iPSC technology development. During follow-up, no major complications such as immunogenicity or tumorigenesis have been observed. Future trials should keep focusing on the safety of stem cell-derived RPE (SC-RPE) especially in long period, and better understanding of the nature of stem cell and the molecular events in the process to generate SC-RPE is necessary to the prosperity of SC-RPE clinical application.

  12. Tumor cell-derived microparticles: a new form of cancer vaccine.

    Science.gov (United States)

    Zhang, Huafeng; Huang, Bo

    2015-08-01

    For cancer vaccines, tumor antigen availability is currently not an issue due to technical advances. However, the generation of optimal immune stimulation during vaccination is challenging. We have recently demonstrated that tumor cell-derived microparticles (MP) can function as a new form of potent cancer vaccine by efficiently activating type I interferon pathway in a cGAS/STING dependent manner.

  13. Analysis of Cell-Derived Microparticles with Highly Precise Nanotechnological Methods

    DEFF Research Database (Denmark)

    Cherré, Solène; Østergaard, Ole; Heegaard, Niels H.H.

    2014-01-01

    Cell-derived microparticles have gained a broad interest in the past years. Being released by blood cells upon activation or induction of apoptosis, they have a great potential as novel diagnostic markers and their investigation can bring new knowledge into the pathogenesis of various diseases. H...

  14. Progenitor's Signatures in Type Ia Supernova Remnants

    NARCIS (Netherlands)

    Chiotellis, A.; Kosenko, D.; Schure, K.M.; Vink, J.

    2013-01-01

    The remnants of Type Ia supernovae (SNe Ia) can provide important clues about their progenitor histories. We discuss two well-observed supernova remnants (SNRs) that are believed to have resulted from SNe Ia, and use various tools to shed light on the possible progenitor histories. We find that

  15. Pulpal progenitors and dentin repair.

    Science.gov (United States)

    Harichane, Y; Hirata, A; Dimitrova-Nakov, S; Granja, I; Goldberg, A; Kellermann, O; Poliard, A

    2011-07-01

    Mesenchymal stem cells are present in the dental pulp. They have been shown to contribute to dentin-like tissue formation in vitro and to participate in bone repair after a mandibular lesion. However, their capacity to contribute efficiently to reparative dentin formation after pulp lesion has never been explored. After pulp exposure, we have identified proliferative cells within 3 zones. In the crown, zone I is near the cavity, and zone II corresponds to the isthmus between the mesial and central pulp. In the root, zone III, near the apex, at a distance from the inflammatory site, contains mitotic stromal cells which may represent a source of progenitor cells. Stem-cell-based strategies are promising treatments for tissue injury in dentistry. Our experiments focused on (1) location of stem cells induced to leave their quiescent state early after pulp injury and (2) implantation of pulp progenitors, a substitute for classic endodontic treatments, paving the way for pulp stem-cell-based therapies.

  16. SH2-inositol phosphatase 1 negatively influences early megakaryocyte progenitors.

    Directory of Open Access Journals (Sweden)

    Lia E Perez

    Full Text Available The SH2-containing-5'inositol phosphatase-1 (SHIP influences signals downstream of cytokine/chemokine receptors that play a role in megakaryocytopoiesis, including thrombopoietin, stromal-cell-derived-Factor-1/CXCL-12 and interleukin-3. We hypothesize that SHIP might control megakaryocytopoiesis through effects on proliferation of megakaryocyte progenitors (MKP and megakaryocytes (MK.Herein, we report the megakaryocytic phenotype and MK functional assays of hematopoietic organs of two strains of SHIP deficient mice with deletion of the SHIP promoter/first exon or the inositol phosphatase domain. Both SHIP deficient strains exhibit a profound increase in MKP numbers in bone marrow (BM, spleen and blood as analyzed by flow cytometry (Lin(-c-Kit+CD41+ and functional assays (CFU-MK. SHIP deficient MKP display increased phosphorylation of Signal Transducers and Activators of Transcription 3 (STAT-3, protein kinase B (PKB/AKT and extracellular signal-regulated kinases (ERKs. Despite increased MKP content, total body number of mature MK (Lin(-c-kit(-CD41+ are not significantly changed as SHIP deficient BM contains reduced MK while spleen MK numbers are increased. Reduction of CXCR-4 expression in SHIP deficient MK may influence MK localization to the spleen instead of the BM. Endomitosis, process involved in MK maturation, was preserved in SHIP deficient MK. Circulating platelets and red blood cells are also reduced in SHIP deficient mice.SHIP may play an important role in regulation of essential signaling pathways that control early megakaryocytopoiesis in vivo.

  17. Transplanting oligodendrocyte progenitors into the adult CNS

    International Nuclear Information System (INIS)

    Franklin, R.J.M.; Blakemore, W.F.; Cambridge Univ.

    1997-01-01

    This review covers a number of aspects of the behaviour of oligodendrocyte progenitors following transplantation into the adult CNS. First, an account is given of the ability of transplanted oligodendrocyte progenitors, grown in tissue culture in the presence of PDGF and bFGF, to extensively remyelinate focal areas of persistent demyelination. Secondly, we describe how transplanted clonal cell lines of oligodendrocyte progenitors will differentiate in to astrocytes as will oligodendrocytes following transplantation into pathological environments in which both oligodendrocytes and astrocytes are absent, thereby manifesting the bipotentially demonstrable in vitro but not during development. Finally, a series of studies examining the migratory behaviour of transplanted oligodendrocyte progenitors (modelled using the oligodendrocyte progenitor cell line CG4) are described. (author)

  18. Effects of an anti-oxidative ACAT inhibitor on apoptosis/necrosis and cholesterol accumulation under oxidative stress in THP-1 cell-derived foam cells.

    Science.gov (United States)

    Miike, Tomohiro; Shirahase, Hiroaki; Jino, Hiroshi; Kunishiro, Kazuyoshi; Kanda, Mamoru; Kurahashi, Kazuyoshi

    2008-01-02

    THP-1 cell-derived foam cells were exposed to oxidative stress through combined treatment with acetylated LDL (acLDL) and copper ions (Cu2+). The foam cells showed caspase-dependent apoptotic changes on exposure to oxidative stress for 6 h, and necrotic changes with the leakage of LDH after 24 h. KY-455, an anti-oxidative ACAT inhibitor, and ascorbic acid (VC) but not YM-750, an ACAT inhibitor, prevented apoptotic and necrotic changes. These preventive effects of KY-455 and VC were accompanied by the inhibition of lipid peroxidation in culture medium containing acLDL and Cu2+, suggesting the involvement of oxidized acLDL in apoptosis and necrosis. Foam cells accumulated esterified cholesterol (EC) for 24 h in the presence of acLDL without Cu2+, which was suppressed by KY-455 and YM-750. Foam cells showed necrotic changes and died in the presence of acLDL and Cu2+. KY-455 but not YM-750 prevented cell death and reduced the amount of EC accumulated. The foam cells treated with VC further accumulated EC without necrotic changes for 24 h even in the presence of acLDL and Cu2+. YM-750 as well as KY-455 inhibited lipid accumulation when co-incubated with VC in foam cells exposed to oxidative stress. It is concluded that an anti-oxidative ACAT inhibitor or the combination of an antioxidant and an ACAT inhibitor protects foam cells from oxidative stress and effectively reduces cholesterol levels, which would be a promising approach in anti-atherosclerotic therapy.

  19. Panax Notoginseng Saponins Promote Endothelial Progenitor Cell Mobilization and Attenuate Atherosclerotic Lesions in Apolipoprotein E Knockout Mice

    Directory of Open Access Journals (Sweden)

    Ya Liu

    2013-09-01

    Full Text Available Background: Endothelial progenitor cells (EPCs derived from the bone marrow (BM play a key role in the homeostasis of vascular repair by enhanced reendothelialization. Panax notoginseng saponins (PNS, a highly valued traditional Chinese medicine, has been shown to reduce morbidity and mortality from coronary artery disease. The present research was designed to explore the contribution of progenitor cells to the progression of atherosclerotic plaques and the possible modulatory role of PNS in this process. Methods: PNS (60 or 120 mg/kg via intraperitoneal injection was administered over 8 weeks in apolipoprotein E knockout mice on an atherogenic diet. The sizes and histochemical alteration of atherosclerotic lesions and numbers of EPCs in BM and peripheral blood were analyzed. The expression of chemokine stromal cell-derived factor 1α (SDF-1α and its receptor, CXCR4, was monitored as well. Results: PNS significantly reduced the lesion area and intima-to-media ratio compared to vehicle treatment. PNS also augmented endothelialization and reduced the smooth muscle cell (SMCs content of the lesions. The number of c-kit and sca-1 double-positive progenitor cells and flk-1 and sca-1 double-positive progenitor cells were significantly increased in the BM and the peripheral blood of the PNS-treated groups. PNS treatment increased the plasma levels of SDF-1α and SCF as well as the BM levels of matrix metalloproteinase-9 (MMP-9. Moreover, the mRNA levels of SDF-1α and protein levels of CXCR4 were both increased in the BM of mice treated with PNS, while SDF-1α expression decreased. Conclusion: PNS reduce the size of atherosclerotic plaques, and this effect appears to involve progenitor cell mobilization. SDF-1α-CXCR4 interactions and the possible modulatory role of PNS in this process may contribute to the increased progenitor cell mobilization.

  20. Current status of ITER EC design

    International Nuclear Information System (INIS)

    Bosia, G.

    2003-01-01

    The ITER-FEAT Electron Cyclotron System employs one equatorial launcher and three upper port launchers to inject 20 + 20 MW of mm-wave power at 170 GHz through slots in the plasma-facing neutron and radiation shield. The equatorial launcher includes 3 sets of toroidally steer-able mirrors, each collecting 8 RF beams from 24 circular cross-section corrugated wave-guides. In the upper-port launcher, eight beams, reflected in pairs by four toroidally angled (∼30 angle) and poloidally steer-able (∼ 10 angle) mirrors focus of the RF power on the plasma m = 2 and m = 3/2 plasma flux rational surfaces, for neo-classical tearing mode(s) (NTM) control. Conceptual studies of remote steering that would possibly allow removing the beam- steering equipment from the vacuum vessel and locating them in more protected positions have been carried out. All EC wave transmission is based on low losses circular cross section corrugated and evacuated wave guides each connected to a 170 GHz, 1 MW CW Gyrotron tube oscillator featuring an overall efficiency of ∼ 50%, obtained by depressed-collector energy recovery. In the paper, requirements, issues and the current status of the design is reviewed. (authors)

  1. PAR1 inhibition suppresses the self-renewal and growth of A2B5-defined glioma progenitor cells and their derived gliomas in vivo

    DEFF Research Database (Denmark)

    Auvergne, R.; Wu, C.; Connell, A.

    2016-01-01

    Glioblastoma (GBM) remains the most common and lethal intracranial tumor. In a comparison of gene expression by A2B5-defined tumor-initiating progenitor cells (TPCs) to glial progenitor cells derived from normal adult human brain, we found that the F2R gene encoding PAR1 was differentially...... overexpressed by A2B5-sorted TPCs isolated from gliomas at all stages of malignant development. In this study, we asked if PAR1 is causally associated with glioma progression. Lentiviral knockdown of PAR1 inhibited the expansion and self-renewal of human GBM-derived A2B5(+) TPCs in vitro, while pharmacological...

  2. Concomitant multipotent and unipotent dental pulp progenitors and their respective contribution to mineralised tissue formation

    Directory of Open Access Journals (Sweden)

    S Dimitrova-Nakov

    2012-05-01

    Full Text Available Upon in vitro induction or in vivo implantation, the stem cells of the dental pulp display hallmarks of odontoblastic, osteogenic, adipogenic or neuronal cells. However, whether these phenotypes result from genuine multipotent cells or from coexistence of distinct progenitors is still an open question. Furthermore, determining whether a single cell-derived progenitor is capable of undergoing a differentiation cascade leading to tissue repair in situ is important for the development of cell therapy strategies. Three clonal pulp precursor cell lines (A4, C5, H8, established from embryonic ED18 first molars of mouse transgenic for a recombinant plasmid adeno-SV40, were induced to differentiate towards the odonto/osteogenic, chondrogenic or adipogenic programme. Expression of phenotypic markers of each lineage was evaluated by RT-PCR, histochemistry or immunocytochemistry. The clones were implanted into mandibular incisors or calvaria of adult mice. The A4 clone was capable of being recruited towards at least 3 mesodermal lineages in vitro and of contributing to dentin-like or bone formation, in vivo, thus behaving as a multipotent cell. In contrast, the C5 and H8 clones displayed a more restricted potential. Flow cytometric analysis revealed that isolated monopotent and multipotent clones could be distinguished by a differential expression of CD90. Altogether, isolation of these clonal lines allowed demonstrating the coexistence of multipotential and restricted-lineage progenitors in the mouse pulp. These cells may further permit unravelling specificities of the different types of pulp progenitors, hence facilitating the development of cell-based therapies of the dental pulp or other cranio-facial tissues.

  3. Altered calcium handling and increased contraction force in human embryonic stem cell derived cardiomyocytes following short term dexamethasone exposure

    NARCIS (Netherlands)

    Kosmidis, Georgios; Bellin, Milena; Ribeiro, Marcelo C.; van Meer, Berend; Ward-van Oostwaard, Dorien; Passier, Robert; Tertoolen, Leon G. J.; Mummery, Christine L.; Casini, Simona

    2015-01-01

    One limitation in using human pluripotent stem cell derived cardiomyocytes (hPSC-CMs) for disease modeling and cardiac safety pharmacology is their immature functional phenotype compared with adult cardiomyocytes. Here, we report that treatment of human embryonic stem cell derived cardiomyocytes

  4. GPM GROUND VALIDATION ENVIRONMENT CANADA (EC) RADIOSONDE GCPEX V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The GPM Ground Validation Environment Canada (EC) Radiosonde GCPEx dataset provides measurements of pressure, temperature, humidity, and winds collected by Vaisala...

  5. EC/β+ decay of 161Er

    International Nuclear Information System (INIS)

    Kalinnikov, V.G.; Ibraheem, Y.S.; Stegailov, V.I.

    2004-01-01

    Full text: The study of odd-proton nuclei in rare-earth region has been performed to get more information on the nuclear structure of these nuclei. In the framework of a program at the ISOL complex of YASNAPP-2 at JINR to study the decay of odd-nuclei, 161 Er has been investigated. The experiments have been focused on the 161 Ho level scheme populated in the EC/β + decay of 161 Er for which a few results have been previously reported [1,2]. Spectra of single γ-ray, γ-γ coincidence and internal conversion electrons were measured. There are appreciable disagreements between results of some γ-ray intensities obtained by us and in [2]. The greatest difference was for relative γ-ray intensity of the 11.28 keV, which has reported value I γ ∼ 10 (I γ 211 =1000) in [2], but this leads to an overpopulation of the 211 keV level by 14% of the decay [3]. The relative intensities of transitions with Eγ γ211 =1000). Inaccuracy of Iγ in [2] is probably due to a sudden change in the efficiency curve of their Ge(Li) detector in the low energy region while our HPGe detector feature a much smoother variation in the efficiency curve in this γ-ray energy region. Besides, the levels earlier introduced by us in [4], three new levels in 161 Ho were proposed from γ-γ coincidences at energies 693.2 keV, 859.6 keV, and 957.97 keV. Spins, parities and Nilsson quantum characteristics of some 161 Ho levels were established

  6. An expandable embryonic stem cell-derived Purkinje neuron progenitor population that exhibits in vivo maturation in the adult mouse cerebellum

    NARCIS (Netherlands)

    Higuera, Gustavo A; Iaffaldano, Grazia; Bedar, Meiwand; Shpak, Guy; Broersen, Robin; Munshi, Shashini T; Dupont, Catherine; Gribnau, Joost; de Vrij, Femke M S; Kushner, Steven A; De Zeeuw, Chris I

    2017-01-01

    The directed differentiation of patient-derived induced pluripotent stem cells into cell-type specific neurons has inspired the development of therapeutic discovery for neurodegenerative diseases. Many forms of ataxia result from degeneration of cerebellar Purkinje cells, but thus far it has not

  7. An expandable embryonic stem cell-derived Purkinje neuron progenitor population that exhibits in vivo maturation in the adult mouse cerebellum

    NARCIS (Netherlands)

    G.A. Higuera (Gustavo A.); Iaffaldano, G. (Grazia); Bedar, M. (Meiwand); G. Shpak (Guy); R. Broersen (Robin); S.T. Munshi (Shashini T.); Dupont, C. (Catherine); J.H. Gribnau (Joost); F.M.S. Vrij (Femke); S.A. Kushner (Steven); C.I. de Zeeuw (Chris)

    2017-01-01

    textabstractThe directed differentiation of patient-derived induced pluripotent stem cells into cell-type specific neurons has inspired the development of therapeutic discovery for neurodegenerative diseases. Many forms of ataxia result from degeneration of cerebellar Purkinje cells, but thus far it

  8. Mutation analysis with random DNA identifiers (MARDI) catalogs Pig-a mutations in heterogeneous pools of CD48-deficient T cells derived from DMBA-treated rats.

    Science.gov (United States)

    Revollo, Javier R; Crabtree, Nathaniel M; Pearce, Mason G; Pacheco-Martinez, M Monserrat; Dobrovolsky, Vasily N

    2016-03-01

    Identification of mutations induced by xenotoxins is a common task in the field of genetic toxicology. Mutations are often detected by clonally expanding potential mutant cells and genotyping each viable clone by Sanger sequencing. Such a "clone-by-clone" approach requires significant time and effort, and sometimes is even impossible to implement. Alternative techniques for efficient mutation identification would greatly benefit both basic and regulatory genetic toxicology research. Here, we report the development of Mutation Analysis with Random DNA Identifiers (MARDI), a novel high-fidelity Next Generation Sequencing (NGS) approach that circumvents clonal expansion and directly catalogs mutations in pools of mutant cells. MARDI uses oligonucleotides carrying Random DNA Identifiers (RDIs) to tag progenitor DNA molecules before PCR amplification, enabling clustering of descendant DNA molecules and eliminating NGS- and PCR-induced sequencing artifacts. When applied to the Pig-a cDNA analysis of heterogeneous pools of CD48-deficient T cells derived from DMBA-treated rats, MARDI detected nearly all Pig-a mutations that were previously identified by conventional clone-by-clone analysis and discovered many additional ones consistent with DMBA exposure: mostly A to T transversions, with the mutated A located on the non-transcribed DNA strand. © 2015 Wiley Periodicals, Inc.

  9. Polysialic acid modification of the synaptic cell adhesion molecule SynCAM 1 in human embryonic stem cell-derived oligodendrocyte precursor cells

    Directory of Open Access Journals (Sweden)

    Sebastian Werneburg

    2015-05-01

    Full Text Available Oligodendrocyte precursor cells (OPCs are the progenitors of myelinating oligodendrocytes in brain development and repair. Successful myelination depends on the control of adhesiveness during OPC migration and axon contact formation. The decoration of cell surface proteins with the glycan polysialic acid (polySia is a key regulatory element of OPC interactions during development and under pathological conditions. By far the major protein carrier of polySia is the neural cell adhesion molecule NCAM, but recently, polysialylation of the synaptic cell adhesion molecule SynCAM 1 has been detected in the developing mouse brain. In mice, polySia-SynCAM 1 is associated with cells expressing NG2, a marker of a heterogeneous precursor cell population, which is the primary source for oligodendrocytes in development and myelin repair but can also give rise to astrocytes and possibly neurons. It is not yet clear if polySia-SynCAM 1 is expressed by OPCs and its occurrence in humans is elusive. By generating uniform human embryonic stem cell-derived OPC cultures, we demonstrate that polySia is present on human OPCs but down-regulated during differentiation into myelin basic protein-positive oligodendrocytes. PolySia on NCAM resides on the isoforms NCAM-180 and NCAM-140, and SynCAM 1 is identified as a novel polySia acceptor in human OPCs.

  10. Evaluation of mRNA expression levels and electrophysiological function of neuron-like cells derived from canine bone marrow stromal cells.

    Science.gov (United States)

    Nakano, Rei; Edamura, Kazuya; Sugiya, Hiroshi; Narita, Takanori; Okabayashi, Ken; Moritomo, Tadaaki; Teshima, Kenji; Asano, Kazushi; Nakayama, Tomohiro

    2013-10-01

    To investigate the in vitro differentiation of canine bone marrow stromal cells (BMSCs) into functional, mature neurons. Bone marrow from 6 adult dogs. BMSCs were isolated from bone marrow and chemically induced to develop into neurons. The morphology of the BMSCs during neuronal induction was monitored, and immunocytochemical analyses for neuron markers were performed after the induction. Real-time PCR methods were used to evaluate the mRNA expression levels of markers for neural stem or progenitor cells, neurons, and ion channels, and western blotting was used to assess the expression of neuronal proteins before and after neuronal induction. The electrophysiological properties of the neuron-like cells induced from canine BMSCs were evaluated with fluorescent dye to monitor Ca(2)+ influx. Canine BMSCs developed a neuron-like morphology after neuronal induction. Immunocytochemical analysis revealed that these neuron-like cells were positive for neuron markers. After induction, the cells' mRNA expression levels of almost all neuron and ion channel markers increased, and the protein expression levels of nestin and neurofilament-L increased significantly. However, the neuron-like cells derived from canine BMSCs did not have the Ca(2)+ influx characteristic of spiking neurons. Although canine BMSCs had neuron-like morphological and biochemical properties after induction, they did not develop the electrophysiological characteristics of neurons. Thus, these results have suggested that canine BMSCs could have the capacity to differentiate into a neuronal lineage, but the differentiation protocol used may have been insufficient to induce development into functional neurons.

  11. Involvement of multiple myeloma cell-derived exosomes in osteoclast differentiation

    OpenAIRE

    Raimondi, Lavinia; De Luca, Angela; Amodio, Nicola; Manno, Mauro; Raccosta, Samuele; Taverna, Simona; Bellavia, Daniele; Naselli, Flores; Fontana, Simona; Schillaci, Odessa; Giardino, Roberto; Fini, Milena; Tassone, Pierfrancesco; Santoro, Alessandra; De Leo, Giacomo

    2015-01-01

    Bone disease is the most frequent complication in multiple myeloma (MM) resulting in osteolytic lesions, bone pain, hypercalcemia and renal failure. In MM bone disease the perfect balance between bone-resorbing osteoclasts (OCs) and bone-forming osteoblasts (OBs) activity is lost in favour of OCs, thus resulting in skeletal disorders. Since exosomes have been described for their functional role in cancer progression, we here investigate whether MM cell-derived exosomes may be involved in OCs ...

  12. Potential of cancer cell-derived exosomes in clinical application: a review of recent research advances.

    Science.gov (United States)

    Sun, Yu; Liu, Jing

    2014-06-01

    Exosomes are 30- to 100-nm, membrane-bound vesicles that are released by most types of cells, including tumor cells. Exosomes contain a great variety of bioactive molecules, including signal peptides, microRNA, lipids, and DNA. In cancer, tumor cells aberrantly secrete large quantities of exosomes to transport paracrine signals or to contribute to tumor-environment interaction at a distance. The goal of this review was to discuss the recent advances on the mechanism of cancer-derived exosomes in tumor regulation. Pertinent articles and abstracts were identified through searches of PubMed for literature published from 1983 to December 2013. Search terms included exosome, tumor, cancer, diagnosis, and therapy. All of the exposed evidence points to communication between cancer cells and their surroundings, either mediated by cancer cell-derived exosomes or by stromal cell-derived exosomes. This communication probably supports tumor proliferation, motility, invasion, angiogenesis, and premetastatic niche preparation. In addition, recent research implies that cancer cell-derived exosomes play a suppressive role in cancer-directed immune response. The biomarkers detected in bodily fluid-derived exosomes imply a potential for exosomes in cancer diagnosis. Also, exosomes could be used as a vehicle to selectively deliver therapeutic nucleic-acid drugs or conventional drugs for tumor therapy. The tolerability and feasibility of cancer exosomes in diagnosis and therapy need to be further evaluated. Copyright © 2014 Elsevier HS Journals, Inc. All rights reserved.

  13. Functional neuromuscular junctions formed by embryonic stem cell-derived motor neurons.

    Directory of Open Access Journals (Sweden)

    Joy A Umbach

    Full Text Available A key objective of stem cell biology is to create physiologically relevant cells suitable for modeling disease pathologies in vitro. Much progress towards this goal has been made in the area of motor neuron (MN disease through the development of methods to direct spinal MN formation from both embryonic and induced pluripotent stem cells. Previous studies have characterized these neurons with respect to their molecular and intrinsic functional properties. However, the synaptic activity of stem cell-derived MNs remains less well defined. In this study, we report the development of low-density co-culture conditions that encourage the formation of active neuromuscular synapses between stem cell-derived MNs and muscle cells in vitro. Fluorescence microscopy reveals the expression of numerous synaptic proteins at these contacts, while dual patch clamp recording detects both spontaneous and multi-quantal evoked synaptic responses similar to those observed in vivo. Together, these findings demonstrate that stem cell-derived MNs innervate muscle cells in a functionally relevant manner. This dual recording approach further offers a sensitive and quantitative assay platform to probe disorders of synaptic dysfunction associated with MN disease.

  14. Progenitor cells in pulmonary vascular remodeling

    Science.gov (United States)

    Yeager, Michael E.; Frid, Maria G.; Stenmark, Kurt R.

    2011-01-01

    Pulmonary hypertension is characterized by cellular and structural changes in the walls of pulmonary arteries. Intimal thickening and fibrosis, medial hypertrophy and fibroproliferative changes in the adventitia are commonly observed, as is the extension of smooth muscle into the previously non-muscularized vessels. A majority of these changes are associated with the enhanced presence of α-SM-actin+ cells and inflammatory cells. Atypical abundances of functionally distinct endothelial cells, particularly in the intima (plexiform lesions), and also in the perivascular regions, are also described. At present, neither the origin(s) of these cells nor the molecular mechanisms responsible for their accumulation, in any of the three compartments of the vessel wall, have been fully elucidated. The possibility that they arise from either resident vascular progenitors or bone marrow–derived progenitor cells is now well established. Resident vascular progenitor cells have been demonstrated to exist within the vessel wall, and in response to certain stimuli, to expand and express myofibroblastic, endothelial or even hematopoietic markers. Bone marrow–derived or circulating progenitor cells have also been shown to be recruited to sites of vascular injury and to assume both endothelial and SM-like phenotypes. Here, we review the data supporting the contributory role of vascular progenitors (including endothelial progenitor cells, smooth muscle progenitor cells, pericytes, and fibrocytes) in vascular remodeling. A more complete understanding of the processes by which progenitor cells modulate pulmonary vascular remodeling will undoubtedly herald a renaissance of therapies extending beyond the control of vascular tonicity and reduction of pulmonary artery pressure. PMID:22034593

  15. On the practical exploitability of dual EC in TLS implementations

    NARCIS (Netherlands)

    Checkoway, S.; Fredrikson, M.; Niederhagen, R.F.; Everspaugh, A.; Green, M.; Lange, T.; Ristenpart, T.; Bernstein, D.J.; Maskiewicz, J,; Shacham, H.

    2014-01-01

    This paper analyzes the actual cost of attacking TLS implementations that use NIST’s Dual EC pseudorandom number generator, assuming that the attacker generated the constants used in Dual EC. It has been known for several years that an attacker generating these constants and seeing a long enough

  16. CXC chemokine receptor 3 expression on CD34(+) hematopoietic progenitors from human cord blood induced by granulocyte-macrophage colony-stimulating factor

    DEFF Research Database (Denmark)

    Jinquan, T; Quan, S; Jacobi, H H

    2000-01-01

    -induced CD34(+) progenitor chemotaxis. These chemotactic attracted CD34(+) progenitors are colony-forming units-granulocyte-macrophage. gamma IP-10 and Mig also induced GM-CSF-stimulated CD34(+) progenitor adhesion and aggregation by means of CXCR3, a finding confirmed by the observation that anti-CXCR3 m......Ab blocked these functions of gammaIP-10 and Mig but not of chemokine stromal cell-derived factor 1 alpha. gamma IP-10-induced and Mig-induced up-regulation of integrins (CD49a and CD49b) was found to play a crucial role in adhesion of GM-CSF-stimulated CD34(+) progenitors. Moreover, gamma IP-10 and Mig...... stimulated CXCR3 redistribution and cellular polarization in GM-CSF-stimulated CD34(+) progenitors. These results indicate that CXCR3-gamma IP-10 and CXCR3-Mig receptor-ligand pairs, as well as the effects of GM-CSF on them, may be especially important in the cytokine/chemokine environment...

  17. Reversible immortalisation enables genetic correction of human muscle progenitors and engineering of next-generation human artificial chromosomes for Duchenne muscular dystrophy.

    Science.gov (United States)

    Benedetti, Sara; Uno, Narumi; Hoshiya, Hidetoshi; Ragazzi, Martina; Ferrari, Giulia; Kazuki, Yasuhiro; Moyle, Louise Anne; Tonlorenzi, Rossana; Lombardo, Angelo; Chaouch, Soraya; Mouly, Vincent; Moore, Marc; Popplewell, Linda; Kazuki, Kanako; Katoh, Motonobu; Naldini, Luigi; Dickson, George; Messina, Graziella; Oshimura, Mitsuo; Cossu, Giulio; Tedesco, Francesco Saverio

    2018-02-01

    Transferring large or multiple genes into primary human stem/progenitor cells is challenged by restrictions in vector capacity, and this hurdle limits the success of gene therapy. A paradigm is Duchenne muscular dystrophy (DMD), an incurable disorder caused by mutations in the largest human gene: dystrophin. The combination of large-capacity vectors, such as human artificial chromosomes (HACs), with stem/progenitor cells may overcome this limitation. We previously reported amelioration of the dystrophic phenotype in mice transplanted with murine muscle progenitors containing a HAC with the entire dystrophin locus (DYS-HAC). However, translation of this strategy to human muscle progenitors requires extension of their proliferative potential to withstand clonal cell expansion after HAC transfer. Here, we show that reversible cell immortalisation mediated by lentivirally delivered excisable hTERT and Bmi1 transgenes extended cell proliferation, enabling transfer of a novel DYS-HAC into DMD satellite cell-derived myoblasts and perivascular cell-derived mesoangioblasts. Genetically corrected cells maintained a stable karyotype, did not undergo tumorigenic transformation and retained their migration ability. Cells remained myogenic in vitro (spontaneously or upon MyoD induction) and engrafted murine skeletal muscle upon transplantation. Finally, we combined the aforementioned functions into a next-generation HAC capable of delivering reversible immortalisation, complete genetic correction, additional dystrophin expression, inducible differentiation and controllable cell death. This work establishes a novel platform for complex gene transfer into clinically relevant human muscle progenitors for DMD gene therapy. © 2017 The Authors. Published under the terms of the CC BY 4.0 license.

  18. Induced Pluripotent Stem Cell-Derived Neural Cells Survive and Mature in the Nonhuman Primate Brain

    Directory of Open Access Journals (Sweden)

    Marina E. Emborg

    2013-03-01

    Full Text Available The generation of induced pluripotent stem cells (iPSCs opens up the possibility for personalized cell therapy. Here, we show that transplanted autologous rhesus monkey iPSC-derived neural progenitors survive for up to 6 months and differentiate into neurons, astrocytes, and myelinating oligodendrocytes in the brains of MPTP-induced hemiparkinsonian rhesus monkeys with a minimal presence of inflammatory cells and reactive glia. This finding represents a significant step toward personalized regenerative therapies.

  19. Sex Differences in Maturation of Human Embryonic Stem Cell-Derived β Cells in Mice.

    Science.gov (United States)

    Saber, Nelly; Bruin, Jennifer E; O'Dwyer, Shannon; Schuster, Hellen; Rezania, Alireza; Kieffer, Timothy J

    2018-04-01

    Pancreatic progenitors derived from human embryonic stem cells (hESCs) are now in clinical trials for insulin replacement in patients with type 1 diabetes. Animal studies indicate that pancreatic progenitor cells can mature into a mixed population of endocrine cells, including glucose-responsive β cells several months after implantion. However, it remains unclear how conditions in the recipient may influence the maturation and ultimately the function of these hESC-derived cells. Here, we investigated the effects of (1) pregnancy on the maturation of human stage 4 (S4) pancreatic progenitor cells and (2) the impact of host sex on both S4 cells and more mature stage 7 (S7) pancreatic endocrine cells implanted under the kidney capsule of immunodeficient SCID-beige mice. Pregnancy led to increased proliferation of endogenous pancreatic β cells, but did not appear to affect proliferation or maturation of S4 cells at midgestation. Interestingly, S4 and S7 cells both acquired glucose-stimulated C-peptide secretion in females before males. Moreover, S4 cells lowered fasting blood glucose levels in females sooner than in males, whereas the responses with S7 cells were similar. These data indicate that the host sex may impact the maturation of hESC-derived cells in vivo and that this effect can be minimized by more advanced differentiation of the cells before implantation.

  20. The negative impact of Wnt signaling on megakaryocyte and primitive erythroid progenitors derived from human embryonic stem cells

    Directory of Open Access Journals (Sweden)

    Prasuna Paluru

    2014-03-01

    Full Text Available The Wnt gene family consists of structurally related genes encoding secreted signaling molecules that have been implicated in many developmental processes, including regulation of cell fate and patterning during embryogenesis. Previously, we found that Wnt signaling is required for primitive or yolk sac-derived-erythropoiesis using the murine embryonic stem cell (ESC system. Here, we examine the effect of Wnt signaling on the formation of early hematopoietic progenitors derived from human ESCs. The first hematopoietic progenitor cells in the human ESC system express the pan-hematopoietic marker CD41 and the erythrocyte marker, glycophorin A or CD235. We have developed a novel serum-free, feeder-free, adherent differentiation system that can efficiently generate large numbers of CD41 + CD235+ cells. We demonstrate that this cell population contains progenitors not just for primitive erythroid and megakaryocyte cells but for the myeloid lineage as well and term this population the primitive common myeloid progenitor (CMP. Treatment of mesoderm-specified cells with Wnt3a led to a loss of hematopoietic colony-forming ability while the inhibition of canonical Wnt signaling with DKK1 led to an increase in the number of primitive CMPs. Canonical Wnt signaling also inhibits the expansion and/or survival of primitive erythrocytes and megakaryocytes, but not myeloid cells, derived from this progenitor population. These findings are in contrast to the role of Wnt signaling during mouse ESC differentiation and demonstrate the importance of the human ESC system in studying species-specific differences in development.

  1. Differentiation and Application of Induced Pluripotent Stem Cell-Derived Vascular Smooth Muscle Cells.

    Science.gov (United States)

    Maguire, Eithne Margaret; Xiao, Qingzhong; Xu, Qingbo

    2017-11-01

    Vascular smooth muscle cells (VSMCs) play a role in the development of vascular disease, for example, neointimal formation, arterial aneurysm, and Marfan syndrome caused by genetic mutations in VSMCs, but little is known about the mechanisms of the disease process. Advances in induced pluripotent stem cell technology have now made it possible to derive VSMCs from several different somatic cells using a selection of protocols. As such, researchers have set out to delineate key signaling processes involved in triggering VSMC gene expression to grasp the extent of gene regulatory networks involved in phenotype commitment. This technology has also paved the way for investigations into diseases affecting VSMC behavior and function, which may be treatable once an identifiable culprit molecule or gene has been repaired. Moreover, induced pluripotent stem cell-derived VSMCs are also being considered for their use in tissue-engineered blood vessels as they may prove more beneficial than using autologous vessels. Finally, while several issues remains to be clarified before induced pluripotent stem cell-derived VSMCs can become used in regenerative medicine, they do offer both clinicians and researchers hope for both treating and understanding vascular disease. In this review, we aim to update the recent progress on VSMC generation from stem cells and the underlying molecular mechanisms of VSMC differentiation. We will also explore how the use of induced pluripotent stem cell-derived VSMCs has changed the game for regenerative medicine by offering new therapeutic avenues to clinicians, as well as providing researchers with a new platform for modeling of vascular disease. © 2017 American Heart Association, Inc.

  2. Association of mast cell-derived VEGF and proteases in Dengue shock syndrome.

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    Takahisa Furuta

    Full Text Available BACKGROUND: Recent in-vitro studies have suggested that mast cells are involved in Dengue virus infection. To clarify the role of mast cells in the development of clinical Dengue fever, we compared the plasma levels of several mast cell-derived mediators (vascular endothelial cell growth factor [VEGF], soluble VEGF receptors [sVEGFRs], tryptase, and chymase and -related cytokines (IL-4, -9, and -17 between patients with differing severity of Dengue fever and healthy controls. METHODOLOGY/PRINCIPAL FINDINGS: The study was performed at Children's Hospital No. 2, Ho Chi Minh City, and Vinh Long Province Hospital, Vietnam from 2002 to 2005. Study patients included 103 with Dengue fever (DF, Dengue hemorrhagic fever (DHF, and Dengue shock syndrome (DSS, as diagnosed by the World Health Organization criteria. There were 189 healthy subjects, and 19 febrile illness patients of the same Kinh ethnicity. The levels of mast cell-derived mediators and -related cytokines in plasma were measured by ELISA. VEGF and sVEGFR-1 levels were significantly increased in DHF and DSS compared with those of DF and controls, whereas sVEGFR-2 levels were significantly decreased in DHF and DSS. Significant increases in tryptase and chymase levels, which were accompanied by high IL-9 and -17 concentrations, were detected in DHF and DSS patients. By day 4 of admission, VEGF, sVEGFRs, and proteases levels had returned to similar levels as DF and controls. In-vitro VEGF production by mast cells was examined in KU812 and HMC-1 cells, and was found to be highest when the cells were inoculated with Dengue virus and human Dengue virus-immune serum in the presence of IL-9. CONCLUSIONS: As mast cells are an important source of VEGF, tryptase, and chymase, our findings suggest that mast cell activation and mast cell-derived mediators participate in the development of DHF. The two proteases, particularly chymase, might serve as good predictive markers of Dengue disease severity.

  3. Circulating cell-derived microparticles in patients with minimally symptomatic obstructive sleep apnoea.

    Science.gov (United States)

    Ayers, L; Ferry, B; Craig, S; Nicoll, D; Stradling, J R; Kohler, M

    2009-03-01

    Moderate-severe obstructive sleep apnoea (OSA) has been associated with several pro-atherogenic mechanisms and increased cardiovascular risk, but it is not known if minimally symptomatic OSA has similar effects. Circulating cell-derived microparticles have been shown to have pro-inflammatory, pro-coagulant and endothelial function-impairing effects, as well as to predict subclinical atherosclerosis and cardiovascular risk. In 57 patients with minimally symptomatic OSA, and 15 closely matched control subjects without OSA, AnnexinV-positive, platelet-, leukocyte- and endothelial cell-derived microparticles were measured by flow cytometry. In patients with OSA, median (interquartile range) levels of AnnexinV-positive microparticles were significantly elevated compared with control subjects: 2,586 (1,566-3,964) microL(-1) versus 1,206 (474-2,501) microL(-1), respectively. Levels of platelet-derived and leukocyte-derived microparticles were also significantly higher in patients with OSA (2,267 (1,102-3,592) microL(-1) and 20 (14-31) microL(-1), respectively) compared with control subjects (925 (328-2,068) microL(-1) and 15 (5-23) microL(-1), respectively). Endothelial cell-derived microparticle levels were similar in patients with OSA compared with control subjects (13 (8-25) microL(-1) versus 11 (6-17) microL(-1)). In patients with minimally symptomatic obstructive sleep apnoea, levels of AnnexinV-positive, platelet- and leukocyte-derived microparticles are elevated when compared with closely matched control subjects without obstructive sleep apnoea. These findings suggest that these patients may be at increased cardiovascular risk, despite being minimally symptomatic.

  4. EC6 safety enhancement - including impact of Fukushima lessons learned

    Energy Technology Data Exchange (ETDEWEB)

    Yu, S.; Zemdegs, R.; Boyle, S.; Soulard, M., E-mail: stephen.yu@candu.com [Candu Energy Inc., Mississauga, Ontario (Canada)

    2012-09-15

    The Enhanced CANDU 6 (EC6) is the new Generation III CANDU reactor design that meets the most up to date regulatory requirements and customer expectations. EC6 builds on the proven high performance design inch as the Qinshan CANDU 6 units and has made improvements to safety and operational performance, and has incorporated extensive operational feedback including Fukushima. The Fukushima Dai-ichi March 11, 2011 event has demonstrated the importance of defence-in-depth considerations for beyond-design basis events, including severe accidents. The EC6 design is based on the defence-in-depth principles and provides further design features that address the lessons learned from Fukushima. (author)

  5. Nuclear power plant performance in the EC in 1978

    International Nuclear Information System (INIS)

    Anon.

    1979-01-01

    According to the report presented by the Statistical Office of the Ec, nuclear electricity generation in the nine EC countries has been increased in 1978 by 11% to a net value of 115 TWh as compared to the year before. Installed capacity went up by 18.6% to a net value of 24. 000 MWe at the end of the year. The mean availability period has been close to the hitherto best value of 6000 h/a. The share of nuclear energy in total electricity generation in the EC rose by 0.5% to 10.5%. (orig.) [de

  6. Drug-loaded nanoparticles induce gene expression in human pluripotent stem cell derivatives

    Science.gov (United States)

    Gajbhiye, Virendra; Escalante, Leah; Chen, Guojun; Laperle, Alex; Zheng, Qifeng; Steyer, Benjamin; Gong, Shaoqin; Saha, Krishanu

    2013-12-01

    Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained higher fibroblast proliferation levels and MMP activity. The results demonstrate that the PEG-H40-DXC nanoparticle system provides an effective tool to controlling gene expression in human stem cell derivatives.Tissue engineering and advanced manufacturing of human stem cells requires a suite of tools to control gene expression spatiotemporally in culture. Inducible gene expression systems offer cell-extrinsic control, typically through addition of small molecules, but small molecule inducers typically contain few functional groups for further chemical modification. Doxycycline (DXC), a potent small molecule inducer of tetracycline (Tet) transgene systems, was conjugated to a hyperbranched dendritic polymer (Boltorn H40) and subsequently reacted with polyethylene glycol (PEG). The resulting PEG-H40-DXC nanoparticle exhibited pH-sensitive drug release behavior and successfully controlled gene expression in stem-cell-derived fibroblasts with a Tet-On system. While free DXC inhibited fibroblast proliferation and matrix metalloproteinase (MMP) activity, PEG-H40-DXC nanoparticles maintained

  7. Brain derived neurotrophic factor contributes to the cardiogenic potential of adult resident progenitor cells in failing murine heart.

    Directory of Open Access Journals (Sweden)

    Rasmita Samal

    Full Text Available Resident cardiac progenitor cells show homing properties when injected into the injured but not to the healthy myocardium. The molecular background behind this difference in behavior needs to be studied to elucidate how adult progenitor cells can restore cardiac function of the damaged myocardium. Since the brain derived neurotrophic factor (BDNF moderates cardioprotection in injured hearts, we focused on delineating its regulatory role in the damaged myocardium.Comparative gene expression profiling of freshly isolated undifferentiated Sca-1 progenitor cells derived either from heart failure transgenic αMHC-CyclinT1/Gαq overexpressing mice or wildtype littermates revealed transcriptional variations. Bdnf expression was up regulated 5-fold during heart failure which was verified by qRT-PCR and confirmed at protein level. The migratory capacity of Sca-1 cells from transgenic hearts was improved by 15% in the presence of 25 ng/ml BDNF. Furthermore, BDNF-mediated effects on Sca-1 cells were studied via pulsed Stable Isotope Labeling of Amino acids in Cell Culture (pSILAC proteomics approach. After BDNF treatment significant differences between newly synthesized proteins in Sca-1 cells from control and transgenic hearts were observed for CDK1, SRRT, HDGF, and MAP2K3 which are known to regulate cell cycle, survival and differentiation. Moreover BDNF repressed the proliferation of Sca-1 cells from transgenic hearts.Comparative profiling of resident Sca-1 cells revealed elevated BDNF levels in the failing heart. Exogenous BDNF (i stimulated migration, which might improve the homing ability of Sca-1 cells derived from the failing heart and (ii repressed the cell cycle progression suggesting its potency to ameliorate heart failure.

  8. Hydroxyurea inhibits parvovirus B19 replication in erythroid progenitor cells.

    Science.gov (United States)

    Bonvicini, Francesca; Bua, Gloria; Conti, Ilaria; Manaresi, Elisabetta; Gallinella, Giorgio

    2017-07-15

    Parvovirus B19 (B19V) infection is restricted to erythroid progenitor cells (EPCs) of the human bone marrow, leading to transient arrest of erythropoiesis and severe complications mainly in subjects with underlying hematological disorders or with immune system deficits. Currently, there are no specific antiviral drugs for B19V treatment, but identification of compounds inhibiting B19V replication can be pursued by a drug repositioning strategy. In this frame, the present study investigates the activity of hydroxyurea (HU), the only disease-modifying therapy approved for sickle cell disease (SCD), towards B19V replication in the two relevant cellular systems, the UT7/EpoS1 cell line and EPCs. Results demonstrate that HU inhibits B19V replication with EC 50 values of 96.2µM and 147.1µM in UT7/EpoS1 and EPCs, respectively, providing experimental evidence of the antiviral activity of HU towards B19V replication, and confirming the efficacy of a drug discovery process by drug repositioning strategy. The antiviral activity occurs in vitro at concentrations lower than those affecting cellular DNA replication and viability, and at levels measured in plasma samples of SCD patients undergoing HU therapy. HU might determine a dual beneficial effect on SCD patients, not only for the treatment of the disease but also towards a virus responsible for severe complications. Copyright © 2017 Elsevier Inc. All rights reserved.

  9. Heterogeneity of limbal basal epithelial progenitor cells.

    Science.gov (United States)

    Hayashida, Yasutaka; Li, Wei; Chen, Ying-Ting; He, Hua; Chen, Szu-yu; Kheirkah, Ahmad; Zhu, Ying-Tien; Matsumoto, Yukihiro; Tseng, Scheffer C G

    2010-11-01

    Although corneal epithelial stem cells (SCs) are located at the limbus between the cornea and the conjunctiva, not all limbal basal epithelial cells are SCs. Using 2 dispase digestions to remove different amounts of limbal basal epithelial cells for cross-sections, flat mounts, and cytospin preparations, double immunostaining to pancytokeratins (PCK) and vimentin (Vim) identified 3 p63+ epithelial progenitors such as PCK-/Vim+, PCK/Vim, and PCK-/Vim+ and 1 p63+ mesenchymal cell, PCK-/Vim+. PCK-/Vim- progenitors had the smallest cell size were 10-20 times more enriched on collagen I-coated dishes in the 5-minute rapid adherent fraction that contained the highest percentage of p63+ cells but the lowest percentage of cytokeratin12+ cells, and gave rise to high Ki67 labeling and vivid clonal growth. In contrast, PCK+/Vim+ and PCK+/Vim- progenitors were found more in the slow-adherent fraction and yielded poor clonal growth. PCK/Vim progenitors and clusters of PCK-/Vim+ mesenchymal cells, which were neither melanocytes nor Langerhans cells, were located in the limbal basal region. Therefore, differential expression of PCK and Vim helps identify small PCK-/Vim- cells as the most likely candidate for SCs among a hierarchy of heterogeneous limbal basal progenitors, and their close association with PCK-/Vim+ presumed "niche" cells.

  10. Roles of major organizations in EC for decision making. EC no ishi kettei ni okeru shuyo kikan no yakuwari

    Energy Technology Data Exchange (ETDEWEB)

    Kodama, M.

    1992-12-01

    This paper considers authorization powers held by the major organizations in the EC, with the Maastricht Treaty kept in mind. One of the objectives of establishing the EC is to harmonize and develop the economic activities in the community on the whole, and promote making closer the relations among the member nations. The EC's supernationality has two aspects: finances (using financial sources from externally common tariffs as the EC's independent financial resources) and legislations (member nations delegating their own powers to the international organization). Organizations involving in the EC legislations are the European Assembly, the board of ministers, and the EC Committee. The Assembly has deciding powers at high levels on the EC budget decision, as well as vetoing power on the budget. The board of ministers is provided with deciding powers to assure achievement of the objectives defined in the official requirements. The Committee has a supernational character that each committee member can act independently free from being subjected to orders from his or her native country. Practically, however, the authorization relationship among the organizations has confrontations over the interests of the organizations and member nations. There have been such movements in this situation as the Luxemburg compromises, actions at the agricultural ministers meeting, and preparation of the unified European protocol. 2 figs., 4 tabs.

  11. What is the potential of oligodendrocyte progenitor cells to successfully treat human spinal cord injury?

    Directory of Open Access Journals (Sweden)

    Yeung Trevor M

    2011-09-01

    Full Text Available Abstract Background Spinal cord injury is a serious and debilitating condition, affecting millions of people worldwide. Long seen as a permanent injury, recent advances in stem cell research have brought closer the possibility of repairing the spinal cord. One such approach involves injecting oligodendrocyte progenitor cells, derived from human embryonic stem cells, into the injured spinal cord in the hope that they will initiate repair. A phase I clinical trial of this therapy was started in mid 2010 and is currently underway. Discussion The theory underlying this approach is that these myelinating progenitors will phenotypically replace myelin lost during injury whilst helping to promote a repair environment in the lesion. However, the importance of demyelination in the pathogenesis of human spinal cord injury is a contentious issue and a body of literature suggests that it is only a minor factor in the overall injury process. Summary This review examines the validity of the theory underpinning the on-going clinical trial as well as analysing published data from animal models and finally discussing issues surrounding safety and purity in order to assess the potential of this approach to successfully treat acute human spinal cord injury.

  12. Culture of hESC-derived pancreatic progenitors in alginate-based scaffolds.

    Science.gov (United States)

    Formo, Kjetil; Cho, Candy H-H; Vallier, Ludovic; Strand, Berit L

    2015-12-01

    The effect of alginate-based scaffolds with added basement membrane proteins on the in vitro development of hESC-derived pancreatic progenitors was investigated. Cell clusters were encapsulated in scaffolds containing the basement membrane proteins collagen IV, laminin, fibronectin, or extracellular matrix-derived peptides, and maintained in culture for up to 46 days. The cells remained viable throughout the experiment with no signs of central necrosis. Whereas nonencapsulated cells aggregated into larger clusters, some of which showed signs of morphological changes and tissue organization, the alginate matrix stabilized the cluster size and displayed more homogeneous cell morphologies, allowing culture for long periods of time. For all conditions tested, a stable or declining expression of insulin and PDX1 and an increase in glucagon and somatostatin over time indicated a progressive reduction in beta cell-related gene expression. Alginate scaffolds can provide a chemically defined, xeno-free and easily scalable alternative for culture of pancreatic progenitors. Although no increase in insulin and PDX1 gene expression after alginate-immobilized cell culture was seen in this study, further optimization of the matrix physicochemical and biological properties and of the medium composition may still be a relevant strategy to promote the stabilization or maturation of stem cell-derived beta cells. © 2015 Wiley Periodicals, Inc.

  13. Sympathetic neurons modulate the beat rate of pluripotent cell-derived cardiomyocytes in vitro.

    Science.gov (United States)

    Takeuchi, Akimasa; Shimba, Kenta; Mori, Masahide; Takayama, Yuzo; Moriguchi, Hiroyuki; Kotani, Kiyoshi; Lee, Jong-Kook; Noshiro, Makoto; Jimbo, Yasuhiko

    2012-12-01

    Although stem cell-derived cardiomyocytes have great potential for the therapy of heart failure, it is unclear whether their function after grafting can be controlled by the host sympathetic nervous system, a component of the autonomic nervous system (ANS). Here we demonstrate the formation of functional connections between rat sympathetic superior cervical ganglion (SCG) neurons and pluripotent (P19.CL6) cell-derived cardiomyocytes (P19CMs) in compartmentalized co-culture, achieved using photolithographic microfabrication techniques. Formation of synapses between sympathetic neurons and P19CMs was confirmed by immunostaining with antibodies against β-3 tubulin, synapsin I and cardiac troponin-I. Changes in the beat rate of P19CMs were triggered after electrical stimulation of the co-cultured SCG neurons, and were affected by the pulse frequency of the electrical stimulation. Such changes in the beat rate were prevented when propranolol, a β-adrenoreceptor antagonist, was added to the culture medium. These results suggest that the beat rate of differentiated cardiomyocytes can be modulated by electrical stimulation of connected sympathetic neurons.

  14. Data on importance of hematopoietic cell derived Lipocalin 2 against gut inflammation

    Directory of Open Access Journals (Sweden)

    Piu Saha

    2016-09-01

    Full Text Available The data herein is related to the research article entitled “Microbiota-inducible innate immune siderophore binding protein Lipocalin 2 is critical for intestinal homeostasis” (Singh et al., 2016 [1]. In the present article, we monitored dextran sodium sulfate (DSS-induced colitis development upon Lipocalin 2 (Lcn2 neutralization, and examined the survival of Lcn2 deficient (Lcn2KO mice and their WT littermates upon DSS challenge. To dissect the relative contribution of immune and non-immune cells-derived Lcn2 in mediating protection against gut inflammation, we generated respective bone marrow chimera and evaluated their susceptibility to IL-10 receptor neutralization-induced chronic colitis.Neutralization of Lcn2 in WT mice resulted in exacerbated DSS-induced colitis. Notably, mice lacking Lcn2 exhibited 100% mortality whereas only 20% mortality was observed in WT mice upon DSS challenge. Further, data from bone marrow chimera showed that immune cell-derived Lcn2 is the major contributor in conferring protection against colitis. Keywords: Siderocalin, Gut microbiota, Inflammatory bowel disease, Bone marrow chimeras, Colitis

  15. G{sub 2} radiosensitivity of cells derived from cancer-prone individuals

    Energy Technology Data Exchange (ETDEWEB)

    Darroudi, F.; Vyas, R.C.; Vermeulen, S.; Natarajan, A.T. [J.A. Cohen Institute of Radiopathology and Radiation Protection, Interuniversity Institute, Leiden (Netherlands)

    1995-04-01

    The potential of enhanced chromatid damage, observed after X-irradiation of G{sub 2} phase, has been used to detect individuals genetically predisposed to cancer, utilising fibroblasts/lymphocytes from these patients as well as fibroblasts derived from human tumours. Fibroblasts and/or lymphocyte samples of two autosomal recessive syndromes (xeroderma pigmentosum (XP), Fanconi`s anaemia (FA)) and one congenital or acquired disorder, aplastic anaemia (AA), were employed for the G{sub 2} radiosensitivity assay. In addition, we have estimated the frequencies of spontaneously occurring chromosomal aberrations as well as G{sub 2} radiosensitivity of eight samples of fibroblasts/fibroblast-like cells (two normal, two colorectal carcinoma, two Wilms` tumour, one retinoblastoma and one polyposis coli), and three samples of lymphocytes (two normal and one from a lymphoma patient). The results obtained indicate that there were no differences between fibroblast cells derived from patients or tumours, except FA patients, in the frequency of spontaneously occurring chromosomal aberrations when compared to normal cells. Following X-irradiation we did not observe any significantly increased G{sub 2} radiosensitivity in FA and XP cells. Lymphocytes from AA and lymphoma patients, and all tumour cell lines except retinoblastoma, responded with increased frequencies of aberrations following G{sub 2} X-irradiation in comparison to cells derived from normal individuals. In our hands, the G{sub 2} sensitivity assay could not always discriminate cells from cancer-prone individuals from those of controls.

  16. Functions of Müller cell-derived vascular endothelial growthfactor in diabetic retinopathy

    Institute of Scientific and Technical Information of China (English)

    2015-01-01

    Müller cells are macroglia and play many essentialroles as supporting cells in the retina. To respond topathological changes in diabetic retinopathy (DR), amajor complication in the eye of diabetic patients,retinal Müller glia produce a high level of vascularendothelial growth factor (VEGF or VEGF-A). As VEGFis expressed by multiple retinal cell-types and Müllerglia comprise only a small portion of cells in the retina,it has been a great challenge to reveal the function ofVEGF or other globally expressed proteins produced byMüller cells. With the development of conditional genetargeting tools, it is now possible to dissect the functionof Müller cell-derived VEGF in vivo . By using conditionalgene targeting approach, we demonstrate that Müllerglia are a major source of retinal VEGF in diabetic miceand Müller cell-derived VEGF plays a significant role inthe alteration of protein expression and peroxynitration,which leads to retinal inflammation, neovascularization,vascular leakage, and vascular lesion, key pathologicalchanges in DR. Therefore, Müller glia are a potentialcellular target for the treatment of DR, a leading causeof blindness.

  17. Functional Properties of Human Stem Cell-Derived Neurons in Health and Disease

    Directory of Open Access Journals (Sweden)

    Jason P. Weick

    2016-01-01

    Full Text Available Stem cell-derived neurons from various source materials present unique model systems to examine the fundamental properties of central nervous system (CNS development as well as the molecular underpinnings of disease phenotypes. In order to more accurately assess potential therapies for neurological disorders, multiple strategies have been employed in recent years to produce neuronal populations that accurately represent in vivo regional and transmitter phenotypes. These include new technologies such as direct conversion of somatic cell types into neurons and glia which may accelerate maturation and retain genetic hallmarks of aging. In addition, novel forms of genetic manipulations have brought human stem cells nearly on par with those of rodent with respect to gene targeting. For neurons of the CNS, the ultimate phenotypic characterization lies with their ability to recapitulate functional properties such as passive and active membrane characteristics, synaptic activity, and plasticity. These features critically depend on the coordinated expression and localization of hundreds of ion channels and receptors, as well as scaffolding and signaling molecules. In this review I will highlight the current state of knowledge regarding functional properties of human stem cell-derived neurons, with a primary focus on pluripotent stem cells. While significant advances have been made, critical hurdles must be overcome in order for this technology to support progression toward clinical applications.

  18. Cell-derived microparticles and vascular pregnancy complications: a systematic and comprehensive review.

    Science.gov (United States)

    Alijotas-Reig, Jaume; Palacio-Garcia, Carles; Llurba, Elisa; Vilardell-Tarres, Miquel

    2013-02-01

    To assess current studies on the relationship between cell-derived microparticles (cMP) and recurrent miscarriages (RM) and pre-eclampsia (PE), and review the relationships between cMP and inflammatory and clot pathways, antiphospholipid antibodies (aPL), cytokines, and pregnancy complications. Systematic and comprehensive review of the literature from January 2000 to January 2012. Vall d'Hebron University Hospital. Women with recurrent miscarriages or PE, healthy nonpregnant women, and healthy pregnant women. None. Comparison of cMP numbers and types among groups. Platelet and endothelial cMP are increased in women with normal pregnancies compared with nonpregnant healthy women. Only five case-control studies regarding cMP and RM and 16 on cMP and PE were found to match our objective. Three of five articles referring to RM showed differences in cMP numbering, and 13 of 16 on cMP and PE showed differences in some type of cMP compared with controls. Cell-derived microparticles were raised in normal pregnancy. Recurrent miscarriage seems to be related to endothelial and platelet cell activation and/or consumption. An increase in almost all cMP types was observed in PE. A relationship between cMP and endothelial activation and proinflammatory status seems to exist. Copyright © 2013 American Society for Reproductive Medicine. Published by Elsevier Inc. All rights reserved.

  19. Haemopoietic progenitor cells in human peripheral blood

    International Nuclear Information System (INIS)

    Zwaan, F.E.

    1980-01-01

    The purpose of the investigation reported is to purify haemopoietic progenitor cells from human peripheral blood using density gradient centrifugation in order to isolate a progenitor cell fraction without immunocompetent cells. The purification technique of peripheral blood flow colony forming unit culture (CFU-c) by means of density gradient centrifugation and a combined depletion of various rosettes is described. The results of several 'in vitro' characteristics of purified CFU-c suspensions and of the plasma clot diffusion chamber culture technique are presented. Irradiation studies revealed that for both human bone marrow and peripheral blood the CFU-c were less radioresistant than clusters. Elimination of monocytes (and granulocytes) from the test suspensions induced an alteration in radiosensitivity pararmeters. The results obtained with the different techniques are described by analysing peripheral progenitor cell activity in myeloproliferative disorders. (Auth.)

  20. Human induced pluripotent stem cells differentiation into oligodendrocyte progenitors and transplantation in a rat model of optic chiasm demyelination.

    Directory of Open Access Journals (Sweden)

    Alireza Pouya

    Full Text Available BACKGROUND: This study aims to differentiate human induced pluripotent stem cells (hiPSCs into oligodendrocyte precursors and assess their recovery potential in a demyelinated optic chiasm model in rats. METHODOLOGY/PRINCIPAL FINDINGS: We generated a cell population of oligodendrocyte progenitors from hiPSCs by using embryoid body formation in a defined medium supplemented with a combination of factors, positive selection and mechanical enrichment. Real-time polymerase chain reaction and immunofluorescence analyses showed that stage-specific markers, Olig2, Sox10, NG2, PDGFRα, O4, A2B5, GalC, and MBP were expressed following the differentiation procedure, and enrichment of the oligodendrocyte lineage. These results are comparable with the expression of stage-specific markers in human embryonic stem cell-derived oligodendrocyte lineage cells. Transplantation of hiPSC-derived oligodendrocyte progenitors into the lysolecithin-induced demyelinated optic chiasm of the rat model resulted in recovery from symptoms, and integration and differentiation into oligodendrocytes were detected by immunohistofluorescence staining against PLP and MBP, and measurements of the visual evoked potentials. CONCLUSIONS/SIGNIFICANCE: These results showed that oligodendrocyte progenitors generated efficiently from hiPSCs can be used in future biomedical studies once safety issues have been overcome.

  1. Activated Fps/Fes partially rescues the in vivo developmental potential of Flk1-deficient vascular progenitor cells.

    Science.gov (United States)

    Haigh, Jody J; Ema, Masatsugu; Haigh, Katharina; Gertsenstein, Marina; Greer, Peter; Rossant, Janet; Nagy, Andras; Wagner, Erwin F

    2004-02-01

    Relatively little is known about the modulators of the vascular endothelial growth factor A (VEGF-A)/Flk1 signaling cascade. To functionally characterize this pathway, VEGF-A stimulation of endothelial cells was performed. VEGF-A-mediated Flk1 activation resulted in increased translocation of the endogenous Fps/Fes cytoplasmic tyrosine kinase to the plasma membrane and increased tyrosine phosphorylation, suggesting a role for Fps/Fes in VEGF-A/Flk1 signaling events. Addition of a myristoylation consensus sequence to Fps/Fes resulted in VEGF-A-independent membrane localization of Fps/Fes in endothelial cells. Expression of the activated Fps/Fes protein in Flk1-deficient embryonic stem (ES) cells rescued their contribution to the developing vascular endothelium in vivo by using ES cell-derived chimeras. Activated Fps/Fes contributed to this rescue event by restoring the migratory potential to Flk1 null progenitors, which is required for movement of hemangioblasts from the primitive streak region into the yolk sac proper. Activated Fps/Fes in the presence of Flk1 increased the number of hemangioblast colonies in vitro and increased the number of mesodermal progenitors in vivo. These results suggest that Fps/Fes may act synergistically with Flk1 to modulate hemangioblast differentiation into the endothelium. We have also demonstrated that activated Fps/Fes causes hemangioma formation in vivo, independently of Flk1, as a result of increasing vascular progenitor density.

  2. ESC-Derived BDNF-Overexpressing Neural Progenitors Differentially Promote Recovery in Huntington's Disease Models by Enhanced Striatal Differentiation

    Directory of Open Access Journals (Sweden)

    Tina Zimmermann

    2016-10-01

    Full Text Available Huntington's disease (HD is characterized by fatal motoric failures induced by loss of striatal medium spiny neurons. Neuronal cell death has been linked to impaired expression and axonal transport of the neurotrophin BDNF (brain-derived neurotrophic factor. By transplanting embryonic stem cell-derived neural progenitors overexpressing BDNF, we combined cell replacement and BDNF supply as a potential HD therapy approach. Transplantation of purified neural progenitors was analyzed in a quinolinic acid (QA chemical and two genetic HD mouse models (R6/2 and N171-82Q on the basis of distinct behavioral parameters, including CatWalk gait analysis. Explicit rescue of motor function by BDNF neural progenitors was found in QA-lesioned mice, whereas genetic mouse models displayed only minor improvements. Tumor formation was absent, and regeneration was attributed to enhanced neuronal and striatal differentiation. In addition, adult neurogenesis was preserved in a BDNF-dependent manner. Our findings provide significant insight for establishing therapeutic strategies for HD to ameliorate neurodegenerative symptoms.

  3. Tissue engineering and the use of stem/progenitor cells for airway epithelium repair

    Directory of Open Access Journals (Sweden)

    GM Roomans

    2010-06-01

    Full Text Available Stem/progenitor cells can be used to repair defects in the airway wall, resulting from e.g., tumors, trauma, tissue reactions following long-time intubations, or diseases that are associated with epithelial damage. Several potential sources of cells for airway epithelium have been identified. These can be divided into two groups. The first group consists of endogenous progenitor cells present in the respiratory tract. This group can be subdivided according to location into (a a ductal cell type in the submucosal glands of the proximal trachea, (b basal cells in the intercartilaginous zones of the lower trachea and bronchi, (c variant Clara cells (Clarav-cells in the bronchioles and (d at the junctions between the bronchioles and the alveolar ducts, and (e alveolar type II cells. This classification of progenitor cell niches is, however, controversial. The second group consists of exogenous stem cells derived from other tissues in the body. This second group can be subdivided into: (a embryonic stem (ES cells, induced pluripotent stem (iPS cells, or amniotic fluid stem cells, (b side-population cells from bone marrow or epithelial stem cells present in bone marrow or circulation and (c fat-derived mesenchymal cells. Airway epithelial cells can be co-cultured in a system that includes a basal lamina equivalent, extracellular factors from mesenchymal fibroblasts, and in an air-liquid interface system. Recently, spheroid-based culture systems have been developed. Several clinical applications have been suggested: cystic fibrosis, acute respiratory distress syndrome, chronic obstructive lung disease, pulmonary fibrosis, pulmonary edema, and pulmonary hypertension. Clinical applications so far are few, but include subglottic stenosis, tracheomalacia, bronchiomalacia, and emphysema.

  4. AKT signaling mediates IGF-I survival actions on otic neural progenitors.

    Directory of Open Access Journals (Sweden)

    Maria R Aburto

    Full Text Available BACKGROUND: Otic neurons and sensory cells derive from common progenitors whose transition into mature cells requires the coordination of cell survival, proliferation and differentiation programmes. Neurotrophic support and survival of post-mitotic otic neurons have been intensively studied, but the bases underlying the regulation of programmed cell death in immature proliferative otic neuroblasts remains poorly understood. The protein kinase AKT acts as a node, playing a critical role in controlling cell survival and cell cycle progression. AKT is activated by trophic factors, including insulin-like growth factor I (IGF-I, through the generation of the lipidic second messenger phosphatidylinositol 3-phosphate by phosphatidylinositol 3-kinase (PI3K. Here we have investigated the role of IGF-dependent activation of the PI3K-AKT pathway in maintenance of otic neuroblasts. METHODOLOGY/PRINCIPAL FINDINGS: By using a combination of organotypic cultures of chicken (Gallus gallus otic vesicles and acoustic-vestibular ganglia, Western blotting, immunohistochemistry and in situ hybridization, we show that IGF-I-activation of AKT protects neural progenitors from programmed cell death. IGF-I maintains otic neuroblasts in an undifferentiated and proliferative state, which is characterised by the upregulation of the forkhead box M1 (FoxM1 transcription factor. By contrast, our results indicate that post-mitotic p27(Kip-positive neurons become IGF-I independent as they extend their neuronal processes. Neurons gradually reduce their expression of the Igf1r, while they increase that of the neurotrophin receptor, TrkC. CONCLUSIONS/SIGNIFICANCE: Proliferative otic neuroblasts are dependent on the activation of the PI3K-AKT pathway by IGF-I for survival during the otic neuronal progenitor phase of early inner ear development.

  5. Identification of the homing molecules that escort pluripotent stem cells-derived hematopoietic stem cells to their niches and human activated T-cells to inflammatory sites.

    KAUST Repository

    Ali, Amal

    2017-12-01

    Hematopoietic cells exploit the multistep paradigm of cell migration to ultimately enable them to perform their function. This process is dictated by the ability of adhesion molecules on the circulating hematopoietic cells to find their counter-receptors on endothelial cells. Of those molecules, the selectin family and their respective ligands induce the initial transient interactions between circulating cells and the opposing endothelium. In this thesis, I focused on studying E-selectin mediated cellular migration in two hematopoietic cell types, namely human hematopoietic stem and progenitor cells (HSPCs) and human T-lymphocytes. HSPCs derived from pluripotent sources theoretically offers a novel, unlimited source for hematopoietic stem cell transplantation therapy. In vitro pluripotent stem cell derived- hematopoietic stem/progenitor cells (ES/iPS-HSPCs) behave much like somatic HSPCs in that they exhibit clonal expansion and multilineage hematopoietic capacity. However, unlike somatic sources, ES/iPS-HSPCs do not give rise to effective hematopoietic repopulation, which may be due to insufficient HSPCs homing to the bone marrow. HSPCs exploit E- and P-selectin to home and engraft into bone marrow niches. Thus, one of my objectives in this thesis was to study the expression of E-selectin ligands associated with ES/iPS-HSPCs. I showed that ES/iPS-HSPCs lack functional E-selectin ligand(s). In an effort to enhance the interaction between Eselectin and ES/iPS-HSPCs, we decorated the cell surface with sialyl-Lewis x (sLex) using the ex-vivo glycan engineering technology. However, this decoration did not improve the engraftment capacity of ES/iPS-HSPCs, in vivo. Induction of E-selectin expression during inflammation is key to recruitment of immune cells and therefore I also focused on analyzing the expression of E-selectin ligands on activated human T-cells. I identified several novel glycoproteins that may function as E-selectin ligands. Specifically, I compared the

  6. X Inactivation and Progenitor Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ruben Agrelo

    2011-04-01

    Full Text Available In mammals, silencing of one of the two X chromosomes is necessary to achieve dosage compensation. The 17 kb non-coding RNA called Xist triggers X inactivation. Gene silencing by Xist can only be achieved in certain contexts such as in cells of the early embryo and in certain hematopoietic progenitors where silencing factors are present. Moreover, these epigenetic contexts are maintained in cancer progenitors in which SATB1 has been identified as a factor related to Xist-mediated chromosome silencing.

  7. Induced pluripotent stem cell-derived neural cells survive and mature in the nonhuman primate brain.

    Science.gov (United States)

    Emborg, Marina E; Liu, Yan; Xi, Jiajie; Zhang, Xiaoqing; Yin, Yingnan; Lu, Jianfeng; Joers, Valerie; Swanson, Christine; Holden, James E; Zhang, Su-Chun

    2013-03-28

    The generation of induced pluripotent stem cells (iPSCs) opens up the possibility for personalized cell therapy. Here, we show that transplanted autologous rhesus monkey iPSC-derived neural progenitors survive for up to 6 months and differentiate into neurons, astrocytes, and myelinating oligodendrocytes in the brains of MPTP-induced hemiparkinsonian rhesus monkeys with a minimal presence of inflammatory cells and reactive glia. This finding represents a significant step toward personalized regenerative therapies. Copyright © 2013 The Authors. Published by Elsevier Inc. All rights reserved.

  8. A NEW CLASS OF LUMINOUS TRANSIENTS AND A FIRST CENSUS OF THEIR MASSIVE STELLAR PROGENITORS

    International Nuclear Information System (INIS)

    Thompson, Todd A.; Prieto, Jose L.; Stanek, K. Z.; Beacom, John F.; Kochanek, Christopher S.; Kistler, Matthew D.

    2009-01-01

    The progenitors of SN 2008S and the 2008 luminous transient in NGC 300 were deeply dust-enshrouded massive stars, with extremely red mid-infrared (MIR) colors and relatively low bolometric luminosities (∼5 x 10 4 L sun ). The transients were optically faint compared to normal core-collapse supernovae (ccSNe), with peak absolute visual magnitudes of -13 ∼> M V ∼> -15, and their spectra exhibit narrow Balmer and [Ca II] emission lines. These events are unique among transient-progenitor pairs and hence constitute a new class. Additional members of this class may include the M85 transient, SN 1999bw, 2002bu, and others. Whether they are true supernovae or bright massive-star eruptions, we argue that their rate is of order ∼20% of the ccSN rate in star-forming galaxies. This fact is remarkable in light of the observation that a very small fraction of all massive stars in any one galaxy, at any moment, have the infrared colors of the progenitors of SN 2008S and the NGC 300 transient. We show this by extracting MIR and optical luminosity, color, and variability properties of massive stars in M33 using archival imaging. We find that the fraction of massive stars with colors consistent with the progenitors of SN 2008S and the NGC 300 transient is ∼ -4 . In fact, only ∼ 4 yr before explosion, be it death or merely eruption. We discuss the implications of this finding for the evolution and census of 'low-mass' massive stars (i.e., ∼8-12 M sun ), and we connect it with theoretical discussions of electron-capture supernovae (ecSNe) near this mass range. Other potential mechanisms, including the explosive birth of massive white dwarfs and massive star outbursts, are also discussed. A systematic census with (warm) Spitzer of galaxies in the local universe (D ∼< 10 Mpc) for analogous progenitors would significantly improve our knowledge of this channel to massive stellar explosions, and potentially to others with obscured progenitors.

  9. Cataclysmic Variables as Supernova Ia Progenitors

    Directory of Open Access Journals (Sweden)

    Stella Kafka

    2012-06-01

    Full Text Available Although the identification of the progenitors of type Ia supernovae (SNeIa remains controversial, it is generally accepted that they originate from binary star systems in which at least one component is a carbon-oxygen white dwarf (WD; those systems are grouped under the wide umbrella of cataclysmic variables. Current theories for SNeIa progenitors hold that, either via Roche lobe overflow of the companion or via a wind, the WD accumulates hydrogen or helium rich material which is then burned to C and O onto the WD’s surface. However, the specifics of this scenario are far from being understood or defined, allowing for a wealth of theories fighting for attention and a dearth of observations to support them. I discuss the latest attempts to identify and study those controversial SNeIa progenitors. I also introduce the most promising progenitor in hand and I present observational diagnostics that can reveal more members of the category.

  10. Cardiac Progenitor Cell Extraction from Human Auricles

    KAUST Repository

    Di Nardo, Paolo; Pagliari, Francesca

    2017-01-01

    by precursor cells mostly embedded into the heart apex and in the atria. We have shown that an elective region of progenitor cell embedding is represented by the auricles, non-contractile atria appendages that can be easily sampled without harming the patient

  11. Electrophysiological properties and calcium handling of embryonic stem cell-derived cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Jae Boum Youm

    2016-03-01

    Full Text Available Embryonic stem cell-derived cardiomyocytes (ESC-CMs hold great interest in many fields of research including clinical applications such as stem cell and gene therapy for cardiac repair or regeneration. ESC-CMs are also used as a platform tool for pharmacological tests or for investigations of cardiac remodeling. ESC-CMs have many different aspects of morphology, electrophysiology, calcium handling, and bioenergetics compared with adult cardiomyocytes. They are immature in morphology, similar to sinus nodal-like in the electrophysiology, higher contribution of trans-sarcolemmal Ca2+ influx to Ca2+ handling, and higher dependence on anaerobic glycolysis. Here, I review a detailed electrophysiology and Ca2+ handling features of ESC-CMs during differentiation into adult cardiomyocytes to gain insights into how all the developmental changes are related to each other to display cardinal features of developing cardiomyocytes.

  12. Generation of a Motor Nerve Organoid with Human Stem Cell-Derived Neurons

    Directory of Open Access Journals (Sweden)

    Jiro Kawada

    2017-11-01

    Full Text Available During development, axons spontaneously assemble into a fascicle to form nerves and tracts in the nervous system as they extend within a spatially constrained path. However, understanding of the axonal fascicle has been hampered by lack of an in vitro model system. Here, we report generation of a nerve organoid composed of a robust fascicle of axons extended from a spheroid of human stem cell-derived motor neurons within our custom-designed microdevice. The device is equipped with a narrow channel providing a microenvironment that facilitates the growing axons to spontaneously assemble into a unidirectional fascicle. The fascicle was specifically made with axons. We found that it was electrically active and elastic and could serve as a model to evaluate degeneration of axons in vitro. This nerve organoid model should facilitate future studies on the development of the axonal fascicle and drug screening for diseases affecting axon fascicles.

  13. Purification of human induced pluripotent stem cell-derived neural precursors using magnetic activated cell sorting.

    Science.gov (United States)

    Rodrigues, Gonçalo M C; Fernandes, Tiago G; Rodrigues, Carlos A V; Cabral, Joaquim M S; Diogo, Maria Margarida

    2015-01-01

    Neural precursor (NP) cells derived from human induced pluripotent stem cells (hiPSCs), and their neuronal progeny, will play an important role in disease modeling, drug screening tests, central nervous system development studies, and may even become valuable for regenerative medicine treatments. Nonetheless, it is challenging to obtain homogeneous and synchronously differentiated NP populations from hiPSCs, and after neural commitment many pluripotent stem cells remain in the differentiated cultures. Here, we describe an efficient and simple protocol to differentiate hiPSC-derived NPs in 12 days, and we include a final purification stage where Tra-1-60+ pluripotent stem cells (PSCs) are removed using magnetic activated cell sorting (MACS), leaving the NP population nearly free of PSCs.

  14. Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes Afford New Opportunities in Inherited Cardiovascular Disease Modeling

    Directory of Open Access Journals (Sweden)

    Daniel R. Bayzigitov

    2016-01-01

    Full Text Available Fundamental studies of molecular and cellular mechanisms of cardiovascular disease pathogenesis are required to create more effective and safer methods of their therapy. The studies can be carried out only when model systems that fully recapitulate pathological phenotype seen in patients are used. Application of laboratory animals for cardiovascular disease modeling is limited because of physiological differences with humans. Since discovery of induced pluripotency generating induced pluripotent stem cells has become a breakthrough technology in human disease modeling. In this review, we discuss a progress that has been made in modeling inherited arrhythmias and cardiomyopathies, studying molecular mechanisms of the diseases, and searching for and testing drug compounds using patient-specific induced pluripotent stem cell-derived cardiomyocytes.

  15. Induced Pluripotent Stem Cell-Derived Endothelial Cells in Insulin Resistance and Metabolic Syndrome.

    Science.gov (United States)

    Carcamo-Orive, Ivan; Huang, Ngan F; Quertermous, Thomas; Knowles, Joshua W

    2017-11-01

    Insulin resistance leads to a number of metabolic and cellular abnormalities including endothelial dysfunction that increase the risk of vascular disease. Although it has been particularly challenging to study the genetic determinants that predispose to abnormal function of the endothelium in insulin-resistant states, the possibility of deriving endothelial cells from induced pluripotent stem cells generated from individuals with detailed clinical phenotyping, including accurate measurements of insulin resistance accompanied by multilevel omic data (eg, genetic and genomic characterization), has opened new avenues to study this relationship. Unfortunately, several technical barriers have hampered these efforts. In the present review, we summarize the current status of induced pluripotent stem cell-derived endothelial cells for modeling endothelial dysfunction associated with insulin resistance and discuss the challenges to overcoming these limitations. © 2017 American Heart Association, Inc.

  16. DNA repair ability of cultured cells derived from mouse embryos in comparison with human cells

    International Nuclear Information System (INIS)

    Yaki, T.

    1982-01-01

    DNA repair in mouse cells derived from embryos of 3 inbred strains were investigated in comparison with that in human cells. The levels of unscheduled DNA synthesis after UV irradiation appeared to change at different passages, but capacities of host-cell reactivation of UV-irradiated herpes simplex virus were always reduced to the same levels as those in xeroderma pigmentosum cells. This implied that mouse cells are reduced in excision-repair capacities and that the apparently high levels of unscheduled DNA synthesis at certain passages are not quantitatively related to high levels of cell survival. Essentially no differences in DNA repair were noted among 3 strains - BALB/c, C3H/He and C57BL/10. (orig.)

  17. PAX7 Targets, CD54, Integrin α9β1, and SDC2, Allow Isolation of Human ESC/iPSC-Derived Myogenic Progenitors

    Directory of Open Access Journals (Sweden)

    Alessandro Magli

    2017-06-01

    Full Text Available Pluripotent stem (PS-cell-derived cell types hold promise for treating degenerative diseases. However, PS cell differentiation is intrinsically heterogeneous; therefore, clinical translation requires the development of practical methods for isolating progenitors from unwanted and potentially teratogenic cells. Muscle-regenerating progenitors can be derived through transient PAX7 expression. To better understand the biology, and to discover potential markers for these cells, here we investigate PAX7 genomic targets and transcriptional changes in human cells undergoing PAX7-mediated myogenic commitment. We identify CD54, integrin α9β1, and Syndecan2 (SDC2 as surface markers on PAX7-induced myogenic progenitors. We show that these markers allow for the isolation of myogenic progenitors using both fluorescent- and CGMP-compatible magnetic-based sorting technologies and that CD54+α9β1+SDC2+ cells contribute to long-term muscle regeneration in vivo. These findings represent a critical step toward enabling the translation of PS-cell-based therapies for muscle diseases.

  18. Chronic inhibition of tumor cell-derived VEGF enhances the malignant phenotype of colorectal cancer cells

    International Nuclear Information System (INIS)

    Yamagishi, Naoko; Teshima-Kondo, Shigetada; Masuda, Kiyoshi; Nishida, Kensei; Kuwano, Yuki; Dang, Duyen T; Dang, Long H; Nikawa, Takeshi; Rokutan, Kazuhito

    2013-01-01

    Vascular endothelial growth factor-a (VEGF)-targeted therapies have become an important treatment for a number of human malignancies. The VEGF inhibitors are actually effective in several types of cancers, however, the benefits are transiently, and the vast majority of patients who initially respond to the therapies will develop resistance. One of possible mechanisms for the acquired resistance may be the direct effect(s) of VEGF inhibitors on tumor cells expressing VEGF receptors (VEGFR). Thus, we investigated here the direct effect of chronic VEGF inhibition on phenotype changes in human colorectal cancer (CRC) cells. To chronically inhibit cancer cell-derived VEGF, human CRC cell lines (HCT116 and RKO) were chronically exposed (2 months) to an anti-VEGF monoclonal antibody (mAb) or were disrupted the Vegf gene (VEGF-KO). Effects of VEGF family members were blocked by treatment with a VEGF receptor tyrosine kinase inhibitor (VEGFR-TKI). Hypoxia-induced apoptosis under VEGF inhibited conditions was measured by TUNEL assay. Spheroid formation ability was assessed using a 3-D spheroid cell culture system. Chronic inhibition of secreted/extracellular VEGF by an anti-VEGF mAb redundantly increased VEGF family member (PlGF, VEGFR1 and VEGFR2), induced a resistance to hypoxia-induced apoptosis, and increased spheroid formation ability. This apoptotic resistance was partially abrogated by a VEGFR-TKI, which blocked the compensate pathway consisted of VEGF family members, or by knockdown of Vegf mRNA, which inhibited intracellular function(s) of all Vegf gene products. Interestingly, chronic and complete depletion of all Vegf gene products by Vegf gene knockout further augmented these phenotypes in the compensate pathway-independent manner. These accelerated phenotypes were significantly suppressed by knockdown of hypoxia-inducible factor-1α that was up-regulated in the VEGF-KO cell lines. Our findings suggest that chronic inhibition of tumor cell-derived VEGF

  19. MicroRNA and protein profiling of brain metastasis competent cell-derived exosomes.

    Directory of Open Access Journals (Sweden)

    Laura Camacho

    Full Text Available Exosomes are small membrane vesicles released by most cell types including tumor cells. The intercellular exchange of proteins and genetic material via exosomes is a potentially effective approach for cell-to-cell communication and it may perform multiple functions aiding to tumor survival and metastasis. We investigated microRNA and protein profiles of brain metastatic (BM versus non-brain metastatic (non-BM cell-derived exosomes. We studied the cargo of exosomes isolated from brain-tropic 70W, MDA-MB-231BR, and circulating tumor cell brain metastasis-selected markers (CTC1BMSM variants, and compared them with parental non-BM MeWo, MDA-MB-231P and CTC1P cells, respectively. By performing microRNA PCR array we identified one up-regulated (miR-210 and two down-regulated miRNAs (miR-19a and miR-29c in BM versus non-BM exosomes. Second, we analyzed the proteomic content of cells and exosomes isolated from these six cell lines, and detected high expression of proteins implicated in cell communication, cell cycle, and in key cancer invasion and metastasis pathways. Third, we show that BM cell-derived exosomes can be internalized by non-BM cells and that they effectively transport their cargo into cells, resulting in increased cell adhesive and invasive potencies. These results provide a strong rationale for additional investigations of exosomal proteins and miRNAs towards more profound understandings of exosome roles in brain metastasis biogenesis, and for the discovery and application of non-invasive biomarkers for new therapies combating brain metastasis.

  20. Improved flow cytometric assessment reveals distinct microvesicle (cell-derived microparticle signatures in joint diseases.

    Directory of Open Access Journals (Sweden)

    Bence György

    Full Text Available INTRODUCTION: Microvesicles (MVs, earlier referred to as microparticles, represent a major type of extracellular vesicles currently considered as novel biomarkers in various clinical settings such as autoimmune disorders. However, the analysis of MVs in body fluids has not been fully standardized yet, and there are numerous pitfalls that hinder the correct assessment of these structures. METHODS: In this study, we analyzed synovial fluid (SF samples of patients with osteoarthritis (OA, rheumatoid arthritis (RA and juvenile idiopathic arthritis (JIA. To assess factors that may confound MV detection in joint diseases, we used electron microscopy (EM, Nanoparticle Tracking Analysis (NTA and mass spectrometry (MS. For flow cytometry, a method commonly used for phenotyping and enumeration of MVs, we combined recent advances in the field, and used a novel approach of differential detergent lysis for the exclusion of MV-mimicking non-vesicular signals. RESULTS: EM and NTA showed that substantial amounts of particles other than MVs were present in SF samples. Beyond known MV-associated proteins, MS analysis also revealed abundant plasma- and immune complex-related proteins in MV preparations. Applying improved flow cytometric analysis, we demonstrate for the first time that CD3(+ and CD8(+ T-cell derived SF MVs are highly elevated in patients with RA compared to OA patients (p=0.027 and p=0.009, respectively, after Bonferroni corrections. In JIA, we identified reduced numbers of B cell-derived MVs (p=0.009, after Bonferroni correction. CONCLUSIONS: Our results suggest that improved flow cytometric assessment of MVs facilitates the detection of previously unrecognized disease-associated vesicular signatures.

  1. SUPERNOVA REMNANT PROGENITOR MASSES IN M31

    Energy Technology Data Exchange (ETDEWEB)

    Jennings, Zachary G.; Williams, Benjamin F.; Dalcanton, Julianne J.; Gilbert, Karoline M.; Fouesneau, Morgan; Weisz, Daniel R. [Department of Astronomy, University of Washington Seattle, Box 351580, WA 98195 (United States); Murphy, Jeremiah W. [Department of Astrophysical Sciences, Princeton University, Princeton, NJ 08544 (United States); Dolphin, Andrew E., E-mail: zachjenn@uw.edu, E-mail: adolphin@raytheon.com [Raytheon, 1151 East Hermans Road, Tucson, AZ 85706 (United States)

    2012-12-10

    Using Hubble Space Telescope photometry, we age-date 59 supernova remnants (SNRs) in the spiral galaxy M31 and use these ages to estimate zero-age main-sequence masses (M{sub ZAMS}) for their progenitors. To accomplish this, we create color-magnitude diagrams (CMDs) and employ CMD fitting to measure the recent star formation history of the regions surrounding cataloged SNR sites. We identify any young coeval population that likely produced the progenitor star, then assign an age and uncertainty to that population. Application of stellar evolution models allows us to infer the M{sub ZAMS} from this age. Because our technique is not contingent on identification or precise location of the progenitor star, it can be applied to the location of any known SNRs. We identify significant young star formation around 53 of the 59 SNRs and assign progenitor masses to these, representing a factor of {approx}2 increase over currently measured progenitor masses. We consider the remaining six SNRs as either probable Type Ia candidates or the result of core-collapse progenitors that have escaped their birth sites. In general, the distribution of recovered progenitor masses is bottom-heavy, showing a paucity of the most massive stars. If we assume a single power-law distribution, dN/dM{proportional_to}M{sup {alpha}}, then we find a distribution that is steeper than a Salpeter initial mass function (IMF) ({alpha} = -2.35). In particular, we find values of {alpha} outside the range -2.7 {>=} {alpha} {>=} -4.4 to be inconsistent with our measured distribution at 95% confidence. If instead we assume a distribution that follows a Salpeter IMF up to some maximum mass, then we find that values of M{sub Max} > 26 are inconsistent with the measured distribution at 95% confidence. In either scenario, the data suggest that some fraction of massive stars may not explode. The result is preliminary and requires more SNRs and further analysis. In addition, we use our distribution to estimate a

  2. EC6 design features and pre-project licensing review

    Energy Technology Data Exchange (ETDEWEB)

    Yu, S.; Lee, A.G.; Dinh, N.B.; Soulard, M. [CANDU Energy Inc., Mississauga, Ontario, (Canada)

    2013-07-01

    The Enhanced CANDU 6 (EC6) is the new Generation III CANDU reactor design that meets the most up to date Canadian regulatory requirements and customer expectations. Candu Energy Inc. is finalizing development of the EC6 which incorporates the CANDU 6's well-proven features, and adds enhancements that strengthened reactor safety margin and improved operability. The EC6 builds on the proven high performance design and the defence-in-depth features of CANDU 6 units, and has incorporated extensive operational feedback including lessons learned from Fukushima. This paper will provide status of the engineering program including progress on the pre-licensing review of the EC6 design by the Canadian Regulator, CNSC, and will also highlight the design and safety enhancements incorporated in the EC6 product. Safety enhancements to meet safety goals and to improve robustness of systems to respond to design basis accidents and beyond design basis accidents include: new severe accident recovery and heat removal system; improved emergency heat removal system; faster shutoff rods with improved safety margins; mechanical guaranteed shutdown rods; daily load cycling capability; robust containment with containment filter venting system; and improved backed-up electrical supply and cooling services. (author)

  3. Different Angiogenic Potentials of Mesenchymal Stem Cells Derived from Umbilical Artery, Umbilical Vein, and Wharton’s Jelly

    Directory of Open Access Journals (Sweden)

    Lu Xu

    2017-01-01

    Full Text Available Human mesenchymal stem cells derived from the umbilical cord (UC are a favorable source for allogeneic cell therapy. Here, we successfully isolated the stem cells derived from three different compartments of the human UC, including perivascular stem cells derived from umbilical arteries (UCA-PSCs, perivascular stem cells derived from umbilical vein (UCV-PSCs, and mesenchymal stem cells derived from Wharton’s jelly (WJ-MSCs. These cells had the similar phenotype and differentiation potential toward adipocytes, osteoblasts, and neuron-like cells. However, UCA-PSCs and UCV-PSCs had more CD146+ cells than WJ-MSCs (P<0.05. Tube formation assay in vitro showed the largest number of tube-like structures and branch points in UCA-PSCs among the three stem cells. Additionally, the total tube length in UCA-PSCs and UCV-PSCs was significantly longer than in WJ-MSCs (P<0.01. Microarray, qRT-PCR, and Western blot analysis showed that UCA-PSCs had the highest expression of the Notch ligand Jagged1 (JAG1, which is crucial for blood vessel maturation. Knockdown of Jagged1 significantly impaired the angiogenesis in UCA-PSCs. In summary, UCA-PSCs are promising cell populations for clinical use in ischemic diseases.

  4. Finite impulse testing (FIT) system for Emergency Cooling System (ECS) in Dhruva

    International Nuclear Information System (INIS)

    Punekar, Parag; Ramkumar, N.; Kulkarni, U.S.; Darbhea, M.D.; Bharadhwaj, G; Jangra, L.R.; Geetha, Patil; Das, Shantanu; Sonnis, S.T.; Trevedi, P.; Patil, M.B.; Biswas, B.B.

    2006-01-01

    Finite Impulse Testing (FIT) system for Emergency Cooling System (ECS) is used to check healthiness of ECS logic circuits in an online mode. The ECS is an important safety system that ensures the cooling of reactor core during shutdown state of Main Coolant Pumps (MCPs), and hence FIT-ECS that monitors the health of ECS logic circuits in an online (real time) mode is an important part of it. Based on a Safety Related Unusual Occurrence in ECS system due to the malfunction of its earlier single channel FIT system, the new FIT-ECS system has been designed with new features and is commissioned. The FIT-ECS system feeds the simulated input signals (fine impulses of nominal width 575 μS) to the ECS logic circuits and read the outputs. These output (predicted) signals from ECS logic circuit are processed in the FIT-ECS system and in event of any discrepancy, the FIT-ECS system displays fault signature on local panel, detailed information of the fault on Operator Console (OC), and generates an alarm 'ECS logic Fail' in the control room. FIT-ECS also monitors the inputs and outputs of ECS logic circuit. All the information required is stored as a database that can be subsequently displayed in various formats. ECS system is designated as Category I-A system and is a hardwired system and FIT-ECS monitors the healthiness of the logics of the ECS System is a computerized system. As per IEC 1226, FIT-ECS is categorized as Category I-B system. This paper provides technical information on FIT-ECS system design, its important features, the testing carried on the FIT-ECS system, interconnections of FIT-ECS and ECS and the commissioning experience of FIT-ECS system. (author)

  5. Characterization of Hemagglutinin Negative Botulinum Progenitor Toxins

    Directory of Open Access Journals (Sweden)

    Suzanne R. Kalb

    2017-06-01

    Full Text Available Botulism is a disease involving intoxication with botulinum neurotoxins (BoNTs, toxic proteins produced by Clostridium botulinum and other clostridia. The 150 kDa neurotoxin is produced in conjunction with other proteins to form the botulinum progenitor toxin complex (PTC, alternating in size from 300 kDa to 500 kDa. These progenitor complexes can be classified into hemagglutinin positive or hemagglutinin negative, depending on the ability of some of the neurotoxin-associated proteins (NAPs to cause hemagglutination. The hemagglutinin positive progenitor toxin complex consists of BoNT, nontoxic non-hemagglutinin (NTNH, and three hemagglutinin proteins; HA-70, HA-33, and HA-17. Hemagglutinin negative progenitor toxin complexes contain BoNT and NTNH as the minimally functional PTC (M-PTC, but not the three hemagglutinin proteins. Interestingly, the genome of hemagglutinin negative progenitor toxin complexes comprises open reading frames (orfs which encode for three proteins, but the existence of these proteins has not yet been extensively demonstrated. In this work, we demonstrate that these three proteins exist and form part of the PTC for hemagglutinin negative complexes. Several hemagglutinin negative strains producing BoNT/A, /E, and /F were found to contain the three open reading frame proteins. Additionally, several BoNT/A-containing bivalent strains were examined, and NAPs from both genes, including the open reading frame proteins, were associated with BoNT/A. The open reading frame encoded proteins are more easily removed from the botulinum complex than the hemagglutinin proteins, but are present in several BoNT/A and /F toxin preparations. These are not easily removed from the BoNT/E complex, however, and are present even in commercially-available purified BoNT/E complex.

  6. Accelerator-Driven Systems - the iThEC strategy

    International Nuclear Information System (INIS)

    Revol, Jean-Pierre

    2015-01-01

    The International Thorium Energy Committee (iThEC) created in Geneva in 2012 is concerned about the energetic future of our society, which is totally organized on the basis of a fossil fuel economy. This has to change because of obvious geopolitical reasons, because of the impact this economy has on the environment, and because of the rather short time on the human time scale, it will take for fossil fuels to run out. One of the main goals of iThEC is to work on reversing the current negative perception of nuclear energy in Europe, which is the main hurdle to a more serious R and D effort in this field. iThEC provides better information to the public on thorium possibilities, supports thorium ADS technology developments, any R and D regarding thorium, and actively engages the scientific, political and business leadership on the subject

  7. Origin of hemopoietic stromal progenitor cells in chimeras

    International Nuclear Information System (INIS)

    Chertkov, J.L.; Drize, N.J.; Gurevitch, O.A.; Samoylova, R.S.

    1985-01-01

    Intravenously injected bone marrow cells do not participate in the regeneration of hemopoietic stromal progenitors in irradiated mice, nor in the curetted parts of the recipient's marrow. The hemopoietic stromal progenitors in allogeneic chimeras are of recipient origin. The adherent cell layer (ACL) of long-term cultures of allogeneic chimera bone marrow contains only recipient hemopoietic stromal progenitors. However, in ectopic hemopoietic foci produced by marrow implantation under the renal capsule and repopulated by the recipient hemopoietic cells after irradiation and reconstitution by syngeneic hemopoietic cells, the stromal progenitors were of implant donor origin, as were stromal progenitors of the ACL in long-term cultures of hemopoietic cells from ectopic foci. Our results confirm that the stromal and hemopoietic progenitors differ in origin and that hemopoietic stromal progenitors are not transplantable by the intravenous route in mice

  8. EC environmental legislation and the European oil industry

    International Nuclear Information System (INIS)

    Luding, W.G.

    1993-01-01

    EUROPIA has 32 ordinary member companies which own and operate more than 95% of EC refining capacity, plus three associate member companies from EFTA countries. A large part of EUROPIA's efforts is devoted to environmental issues. Among its initial actions, EUROPIA developed Guiding Principles for environmental management which have been adopted by all member companies. This article can only highlight some of the key areas of EC environmental policy of interest to the oil industry. Most of the comments will refer to air pollution issues but, of course, water and ground pollution and waste are equally important subjects. (orig./HS)

  9. Ecological audits under EC regulations: background and impact in practice

    International Nuclear Information System (INIS)

    Walther, T.

    1995-01-01

    There are publications discussing the practical aspects of the implementation of the EC Directive on ecological audits, but little has been published yet on the theoretical background of the directive. The contribution in hand is intended to elucidate the theoretical basis of this instrument in the context of the system of law, explains the term of ''sustainable development'' as a key term, and reveals problems encountered in the evaluation of environmental performance. This information will contribute to a better understanding of the meaning and purposes of the EC Directive, and will thus point out ways and means for direct implementation. (orig.) [de

  10. ECS: Efficient Communication Scheduling for Underwater Sensor Networks

    Directory of Open Access Journals (Sweden)

    Lu Hong

    2011-03-01

    Full Text Available TDMA protocols have attracted a lot of attention for underwater acoustic sensor networks (UWSNs, because of the unique characteristics of acoustic signal propagation such as great energy consumption in transmission, long propagation delay and long communication range. Previous TDMA protocols all allocated transmission time to nodes based on discrete time slots. This paper proposes an efficient continuous time scheduling TDMA protocol (ECS for UWSNs, including the continuous time based and sender oriented conflict analysis model, the transmission moment allocation algorithm and the distributed topology maintenance algorithm. Simulation results confirm that ECS improves network throughput by 20% on average, compared to existing MAC protocols.

  11. Genes and proteins of Escherichia coli (GenProtEc).

    Science.gov (United States)

    Riley, M; Space, D B

    1996-01-01

    GenProtEc is a database of Escherichia coli genes and their gene products, classified by type of function and physiological role and with citations to the literature for each. Also present are data on sequence similarities among E.coli proteins with PAM values, percent identity of amino acids, length of alignment and percent aligned. The database is available as a PKZip file by ftp from mbl.edu/pub/ecoli.exe. The program runs under MS-DOS on IMB-compatible machines. GenProtEc can also be accessed through the World Wide Web at URL http://mbl.edu/html/ecoli.html.

  12. Rapid Cellular Phenotyping of Human Pluripotent Stem Cell-Derived Cardiomyocytes using a Genetically Encoded Fluorescent Voltage Sensor

    Directory of Open Access Journals (Sweden)

    Jordan S. Leyton-Mange

    2014-02-01

    Full Text Available In addition to their promise in regenerative medicine, pluripotent stem cells have proved to be faithful models of many human diseases. In particular, patient-specific stem cell-derived cardiomyocytes recapitulate key features of several life-threatening cardiac arrhythmia syndromes. For both modeling and regenerative approaches, phenotyping of stem cell-derived tissues is critical. Cellular phenotyping has largely relied upon expression of lineage markers rather than physiologic attributes. This is especially true for cardiomyocytes, in part because electrophysiological recordings are labor intensive. Likewise, most optical voltage indicators suffer from phototoxicity, which damages cells and degrades signal quality. Here we present the use of a genetically encoded fluorescent voltage indicator, ArcLight, which we demonstrate can faithfully report transmembrane potentials in human stem cell-derived cardiomyocytes. We demonstrate the application of this fluorescent sensor in high-throughput, serial phenotyping of differentiating cardiomyocyte populations and in screening for drug-induced cardiotoxicity.

  13. Identification and purification of human induced pluripotent stem cell-derived atrial-like cardiomyocytes based on sarcolipin expression.

    Directory of Open Access Journals (Sweden)

    Rebecca Josowitz

    Full Text Available The use of human stem cell-derived cardiomyocytes to study atrial biology and disease has been restricted by the lack of a reliable method for stem cell-derived atrial cell labeling and purification. The goal of this study was to generate an atrial-specific reporter construct to identify and purify human stem cell-derived atrial-like cardiomyocytes. We have created a bacterial artificial chromosome (BAC reporter construct in which fluorescence is driven by expression of the atrial-specific gene sarcolipin (SLN. When purified using flow cytometry, cells with high fluorescence specifically express atrial genes and display functional calcium handling and electrophysiological properties consistent with atrial cardiomyocytes. Our data indicate that SLN can be used as a marker to successfully monitor and isolate hiPSC-derived atrial-like cardiomyocytes. These purified cells may find many applications, including in the study of atrial-specific pathologies and chamber-specific lineage development.

  14. Functional integration of grafted neural stem cell-derived dopaminergic neurons monitored by optogenetics in an in vitro Parkinson model

    DEFF Research Database (Denmark)

    Tønnesen, Jan; Parish, Clare L; Sørensen, Andreas T

    2011-01-01

    Intrastriatal grafts of stem cell-derived dopamine (DA) neurons induce behavioral recovery in animal models of Parkinson's disease (PD), but how they functionally integrate in host neural circuitries is poorly understood. Here, Wnt5a-overexpressing neural stem cells derived from embryonic ventral...... of post-synaptic currents, and functional expression of DA D₂ autoreceptors. These properties resembled those recorded from identical cells in acute slices of intrastriatal grafts in the 6-hydroxy-DA-induced mouse PD model and from DA neurons in intact substantia nigra. Optogenetic activation...... using optogenetics that ectopically grafted stem cell-derived DA neurons become functionally integrated in the DA-denervated striatum. Further optogenetic dissection of the synaptic wiring between grafted and host neurons will be crucial to clarify the cellular and synaptic mechanisms underlying...

  15. CD133+ cells derived from skeletal muscles of Duchenne muscular dystrophy patients have a compromised myogenic and muscle regenerative capability.

    Science.gov (United States)

    Meng, Jinhong; Muntoni, Francesco; Morgan, Jennifer

    2018-05-12

    Cell-mediated gene therapy is a possible means to treat muscular dystrophies like Duchenne muscular dystrophy. Autologous patient stem cells can be genetically-corrected and transplanted back into the patient, without causing immunorejection problems. Regenerated muscle fibres derived from these cells will express the missing dystrophin protein, thus improving muscle function. CD133+ cells derived from normal human skeletal muscle contribute to regenerated muscle fibres and form muscle stem cells after their intra-muscular transplantation into an immunodeficient mouse model. But it is not known whether CD133+ cells derived from DMD patient muscles have compromised muscle regenerative function. To test this, we compared CD133+ cells derived from DMD and normal human muscles. DMD CD133+ cells had a reduced capacity to undergo myogenic differentiation in vitro compared with CD133+ cells derived from normal muscle. In contrast to CD133+ cells derived from normal human muscle, those derived from DMD muscle formed no satellite cells and gave rise to significantly fewer muscle fibres of donor origin, after their intra-muscular transplantation into an immunodeficient, non-dystrophic, mouse muscle. DMD CD133+ cells gave rise to more clones of smaller size and more clones that were less myogenic than did CD133+ cells derived from normal muscle. The heterogeneity of the progeny of CD133+ cells, combined with the reduced proliferation and myogenicity of DMD compared to normal CD133+ cells, may explain the reduced regenerative capacity of DMD CD133+ cells. Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Giant Panda (Ailuropoda melanoleuca) Buccal Mucosa Tissue as a Source of Multipotent Progenitor Cells.

    Science.gov (United States)

    Prescott, Hilary M A; Manning, Craig; Gardner, Aaron; Ritchie, William A; Pizzi, Romain; Girling, Simon; Valentine, Iain; Wang, Chengdong; Jahoda, Colin A B

    2015-01-01

    means that we can only postulate why we were unable to obtain m-SKPs from the lion and red panda cultures. However the giant panda observations point to the value of archiving cells from rare species, and the possibilities for later progenitor cell derivation.

  17. Giant Panda (Ailuropoda melanoleuca Buccal Mucosa Tissue as a Source of Multipotent Progenitor Cells.

    Directory of Open Access Journals (Sweden)

    Hilary M A Prescott

    sample numbers means that we can only postulate why we were unable to obtain m-SKPs from the lion and red panda cultures. However the giant panda observations point to the value of archiving cells from rare species, and the possibilities for later progenitor cell derivation.

  18. Podoplanin-expressing Cells Derived from Bone Marrow Play a Crucial Role in Postnatal Lymphatic Neovascularization

    Science.gov (United States)

    Lee, Ji Yoon; Park, Changwon; Cho, Yong Pil; Lee, Eugine; Kim, Hyongbum; Kim, Pilhan; Yun, Seok H.; Yoon, Young-sup

    2010-01-01

    Background Emerging evidence has suggested a contribution of bone marrow (BM) cells to lymphatic vessel formation; however, the exact phenotype of the cells with lymphatic endothelial progenitor cell (LEPC) function has yet to be identified. Here we investigate the identity of BM-derived LEPCs and their role in lymphatic neovascularization. Methods and Results Culture of BM-mononuclear cells (MNCs) in the presence of VEGFA, VEGFC and EGF resulted in expression of lymphatic endothelial cell (LEC) markers. Among these cells, podoplanin+ cells were isolated by magnetic-labeled cell separation system (MACS) and characterized by FACS and immunocytochemistry. These podoplanin+ cells highly express markers for LECs, hematopoietic lineages, and stem/progenitor cells, and upon further cultivation, generate LECs. We further confirmed that podoplanin+ cells exist in small numbers in BM and peripheral blood (PB) of normal mice, but are significantly (15 fold) augmented upon lymphangiogenic stimuli such as tumor implantation. Next, to evaluate the potential of podoplanin+ cells for the formation of new lymphatic vessels in vivo, we injected culture-isolated or freshly isolated BM-derived podoplanin+ cells into wound and tumor models. Immunohistochemistry demonstrated that the injected cells were incorporated into the lymphatic vasculature, displayed LEC phenotypes, and increased lymphatic vascular density in tissues, suggesting lymphvasculogenesis. Podoplanin+ cells also expressed high levels of lymphangiogenic cytokines and increased proliferation of LECs during co-culture, suggesting a lymphangiogenic or paracrine role. Conclusions Our results provide compelling evidence that BM-derived podoplanin+ cells, a previously unrecognized cell type, function as LEPCs and participate in postnatal lymphatic neovascularization through both lymphvasculogenesis and lymphangiogenesis. PMID:20855662

  19. Focus on Extracellular Vesicles: Therapeutic Potential of Stem Cell-Derived Extracellular Vesicles

    Directory of Open Access Journals (Sweden)

    Bin Zhang

    2016-02-01

    Full Text Available The intense research focus on stem and progenitor cells could be attributed to their differentiation potential to generate new cells to replace diseased or lost cells in many highly intractable degenerative diseases, such as Alzheimer disease, multiple sclerosis, and heart diseases. However, experimental and clinical studies have increasingly attributed the therapeutic efficacy of these cells to their secretion. While stem and progenitor cells secreted many therapeutic molecules, none of these molecules singly or in combination could recapitulate the functional effects of stem cell transplantations. Recently, it was reported that extracellular vesicles (EVs could recapitulate the therapeutic effects of stem cell transplantation. Based on the observations reported thus far, the prevailing hypothesis is that stem cell EVs exert their therapeutic effects by transferring biologically active molecules such as proteins, lipids, mRNA, and microRNA from the stem cells to injured or diseased cells. In this respect, stem cell EVs are similar to EVs from other cell types. They are both primarily vehicles for intercellular communication. Therefore, the differentiating factor is likely due to the composition of their cargo. The cargo of EVs from different cell types are known to include a common set of proteins and also proteins that reflect the cell source of the EVs and the physiological or pathological state of the cell source. Hence, elucidation of the stem cell EV cargo would provide an insight into the multiple physiological or biochemical changes necessary to affect the many reported stem cell-based therapeutic outcomes in a variety of experimental models and clinical trials.

  20. Do Decision 2003/54/EC and Decision no. 1229/2003/EC result in a European market for electricity?

    International Nuclear Information System (INIS)

    Wenting, F.

    2004-01-01

    In the summer of 2003 the European Parliament and the European Council issued a new Directive with respect to the internal market for electricity (Directive 2003/54/EC of the European Parliament and of the Council of 26 June 2003 concerning common rules for the internal market in electricity) and Guidelines for trans-European networks in the energy sector (Decision no. 1229/2003/EC). The question is whether these decisions will lead to a European and free market for electricity. It is concluded that this is not yet the case [nl

  1. Use of human stem cell derived cardiomyocytes to examine sunitinib mediated cardiotoxicity and electrophysiological alterations

    Energy Technology Data Exchange (ETDEWEB)

    Cohen, J.D., E-mail: jennifer.cohen@roche.com [Early and Investigative Safety, Nonclinical Safety, Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110 (United States); Babiarz, J.E., E-mail: joshua.babiarz@roche.com [Early and Investigative Safety, Nonclinical Safety, Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110 (United States); Abrams, R.M., E-mail: rory.abrams@roche.com [Early and Investigative Safety, Nonclinical Safety, Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110 (United States); Guo, L., E-mail: liang.guo@roche.com [Early and Investigative Safety, Nonclinical Safety, Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110 (United States); Kameoka, S., E-mail: sei.kameoka@roche.com [Early and Investigative Safety, Nonclinical Safety, Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110 (United States); Chiao, E., E-mail: eric.chiao@roche.com [Early and Investigative Safety, Nonclinical Safety, Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110 (United States); Taunton, J., E-mail: taunton@cmp.ucsf.edu [Howard Hughes Medical Institute, Cellular and Molecular Pharmacology, University California San Francisco, San Francisco, CA 94158 (United States); Kolaja, K.L., E-mail: kyle.kolaja@roche.com [Early and Investigative Safety, Nonclinical Safety, Hoffmann-La Roche, 340 Kingsland Street, Nutley, NJ 07110 (United States)

    2011-11-15

    Sunitinib, an oral tyrosine kinase inhibitor approved to treat advanced renal cell carcinoma and gastrointestinal stroma tumor, is associated with clinical cardiac toxicity. Although the precise mechanism of sunitinib cardiotoxicity is not known, both the key metabolic energy regulator, AMP-activated protein kinase (AMPK), and ribosomal S 6 kinase (RSK) have been hypothesized as causative, albeit based on rodent models. To study the mechanism of sunitinib-mediated cardiotoxicity in a human model, induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) having electrophysiological and contractile properties of native cardiac tissue were investigated. Sunitinib was cardiotoxic in a dose-dependent manner with an IC{sub 50} in the low micromolar range, observed by a loss of cellular ATP, an increase in oxidized glutathione, and induction of apoptosis in iPSC-CMs. Pretreatment of iPSC-CMs with AMPK activators AICAR or metformin, increased the phosphorylation of pAMPK-T172 and pACC-S79, but only marginally attenuated sunitinib mediated cell death. Furthermore, additional inhibitors of AMPK were not directly cytotoxic to iPSC-CMs up to 250 {mu}M concentrations. Inhibition of RSK with a highly specific, irreversible, small molecule inhibitor (RSK-FMK-MEA) did not induce cytotoxicity in iPSC-CMs below 250 {mu}M. Extensive electrophysiological analysis of sunitinib and RSK-FMK-MEA mediated conduction effects were performed. Taken together, these findings suggest that inhibition of AMPK and RSK are not a major component of sunitinib-induced cardiotoxicity. Although the exact mechanism of cardiotoxicity of sunitinib is not known, it is likely due to inhibition of multiple kinases simultaneously. These data highlight the utility of human iPSC-CMs in investigating the potential molecular mechanisms underlying drug-induced cardiotoxicity. -- Highlights: Black-Right-Pointing-Pointer Cytoxic effect of sunitinib on human stem cell derived cardiomyocytes Black

  2. Use of human stem cell derived cardiomyocytes to examine sunitinib mediated cardiotoxicity and electrophysiological alterations

    International Nuclear Information System (INIS)

    Cohen, J.D.; Babiarz, J.E.; Abrams, R.M.; Guo, L.; Kameoka, S.; Chiao, E.; Taunton, J.; Kolaja, K.L.

    2011-01-01

    Sunitinib, an oral tyrosine kinase inhibitor approved to treat advanced renal cell carcinoma and gastrointestinal stroma tumor, is associated with clinical cardiac toxicity. Although the precise mechanism of sunitinib cardiotoxicity is not known, both the key metabolic energy regulator, AMP-activated protein kinase (AMPK), and ribosomal S 6 kinase (RSK) have been hypothesized as causative, albeit based on rodent models. To study the mechanism of sunitinib-mediated cardiotoxicity in a human model, induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) having electrophysiological and contractile properties of native cardiac tissue were investigated. Sunitinib was cardiotoxic in a dose-dependent manner with an IC 50 in the low micromolar range, observed by a loss of cellular ATP, an increase in oxidized glutathione, and induction of apoptosis in iPSC-CMs. Pretreatment of iPSC-CMs with AMPK activators AICAR or metformin, increased the phosphorylation of pAMPK-T172 and pACC-S79, but only marginally attenuated sunitinib mediated cell death. Furthermore, additional inhibitors of AMPK were not directly cytotoxic to iPSC-CMs up to 250 μM concentrations. Inhibition of RSK with a highly specific, irreversible, small molecule inhibitor (RSK-FMK-MEA) did not induce cytotoxicity in iPSC-CMs below 250 μM. Extensive electrophysiological analysis of sunitinib and RSK-FMK-MEA mediated conduction effects were performed. Taken together, these findings suggest that inhibition of AMPK and RSK are not a major component of sunitinib-induced cardiotoxicity. Although the exact mechanism of cardiotoxicity of sunitinib is not known, it is likely due to inhibition of multiple kinases simultaneously. These data highlight the utility of human iPSC-CMs in investigating the potential molecular mechanisms underlying drug-induced cardiotoxicity. -- Highlights: ► Cytoxic effect of sunitinib on human stem cell derived cardiomyocytes ► Sunitinib causes ATP depletion, LDH release, GSH

  3. EC-LEDS Mexico: Advancing Clean Energy Goals

    Energy Technology Data Exchange (ETDEWEB)

    2016-07-01

    EC-LEDS works with the government of Mexico to help meet its goals of reducing greenhouse gas emissions from the energy sector. The program targets specific, highly technical areas where Mexico has indicated the program can add value and make an impact.

  4. Ketone EC50 values in the Microtox test.

    Science.gov (United States)

    Chen, H F; Hee, S S

    1995-03-01

    The Microtox EC50 values for the following ketones are reported in the following homologous series: straight chain methyl ketones (acetone, 2-butanone, 2-pentanone, 2-hepatonone, 2-octanone, 2-decanone, and 2-tridecanone); methyl ketones substituted at one alpha carbon (3-methyl-2-butanone; 3,3-dimethyl-2-butanone); methyl substituted at two alpha carbons (2,4-dimethyl-3-pentanone; 2,2,4,4-tetramethyl-3-pentanone); phenyl groups replacing methyl in acetone (acetophenone; benzophenone); methyl groups substituted at the alpha carbons of cyclohexanone; and 2,3- 2,4-, and 2,5-hexanediones, most for the first time. While there were linear relationships between log EC50 and MW for the straight chain methyl ketones, and for methyl substitution at the alpha carbon for methyl ketones, there were no other linear relationships. As molecular weight increased, the EC50 values of soluble ketones decreased; as distance between two carbonyl groups decreased so too did EC50 values. Thus, for the ketones the geometry around the carbonyl group is an important determinant of toxicity as well as MW, water solubility, and octanol/water coefficient.

  5. EC-LEDS Supports the Low-Carbon Transition

    Energy Technology Data Exchange (ETDEWEB)

    2016-09-01

    EC-LEDS is a flagship U.S. government-led effort that assists countries to create and implement low emission development strategies, or LEDS -- development frameworks that promote sustainable social and economic development while reducing greenhouse gas emissions over the medium to long term.

  6. Endothelial Cells Promote Expansion of Long-Term Engrafting Marrow Hematopoietic Stem and Progenitor Cells in Primates.

    Science.gov (United States)

    Gori, Jennifer L; Butler, Jason M; Kunar, Balvir; Poulos, Michael G; Ginsberg, Michael; Nolan, Daniel J; Norgaard, Zachary K; Adair, Jennifer E; Rafii, Shahin; Kiem, Hans-Peter

    2017-03-01

    Successful expansion of bone marrow (BM) hematopoietic stem and progenitor cells (HSPCs) would benefit many HSPC transplantation and gene therapy/editing applications. However, current expansion technologies have been limited by a loss of multipotency and self-renewal properties ex vivo. We hypothesized that an ex vivo vascular niche would provide prohematopoietic signals to expand HSPCs while maintaining multipotency and self-renewal. To test this hypothesis, BM autologous CD34 + cells were expanded in endothelial cell (EC) coculture and transplanted in nonhuman primates. CD34 + C38 - HSPCs cocultured with ECs expanded up to 17-fold, with a significant increase in hematopoietic colony-forming activity compared with cells cultured with cytokines alone (colony-forming unit-granulocyte-erythroid-macrophage-monocyte; p < .005). BM CD34 + cells that were transduced with green fluorescent protein lentivirus vector and expanded on ECs engrafted long term with multilineage polyclonal reconstitution. Gene marking was observed in granulocytes, lymphocytes, platelets, and erythrocytes. Whole transcriptome analysis indicated that EC coculture altered the expression profile of 75 genes in the BM CD34 + cells without impeding the long-term engraftment potential. These findings show that an ex vivo vascular niche is an effective platform for expansion of adult BM HSPCs. Stem Cells Translational Medicine 2017;6:864-876. © 2016 The Authors Stem Cells Translational Medicine published by Wiley Periodicals, Inc. on behalf of AlphaMed Press.

  7. Endothelial Cells Promote Expansion of Long‐Term Engrafting Marrow Hematopoietic Stem and Progenitor Cells in Primates

    Science.gov (United States)

    Gori, Jennifer L.; Butler, Jason M.; Kunar, Balvir; Poulos, Michael G.; Ginsberg, Michael; Nolan, Daniel J.; Norgaard, Zachary K.; Adair, Jennifer E.; Rafii, Shahin

    2016-01-01

    Abstract Successful expansion of bone marrow (BM) hematopoietic stem and progenitor cells (HSPCs) would benefit many HSPC transplantation and gene therapy/editing applications. However, current expansion technologies have been limited by a loss of multipotency and self‐renewal properties ex vivo. We hypothesized that an ex vivo vascular niche would provide prohematopoietic signals to expand HSPCs while maintaining multipotency and self‐renewal. To test this hypothesis, BM autologous CD34+ cells were expanded in endothelial cell (EC) coculture and transplanted in nonhuman primates. CD34+C38− HSPCs cocultured with ECs expanded up to 17‐fold, with a significant increase in hematopoietic colony‐forming activity compared with cells cultured with cytokines alone (colony‐forming unit‐granulocyte‐erythroid‐macrophage‐monocyte; p < .005). BM CD34+ cells that were transduced with green fluorescent protein lentivirus vector and expanded on ECs engrafted long term with multilineage polyclonal reconstitution. Gene marking was observed in granulocytes, lymphocytes, platelets, and erythrocytes. Whole transcriptome analysis indicated that EC coculture altered the expression profile of 75 genes in the BM CD34+ cells without impeding the long‐term engraftment potential. These findings show that an ex vivo vascular niche is an effective platform for expansion of adult BM HSPCs. Stem Cells Translational Medicine 2017;6:864–876 PMID:28297579

  8. THE AGES OF TYPE Ia SUPERNOVA PROGENITORS

    International Nuclear Information System (INIS)

    Brandt, Timothy D.; Aubourg, Eric; Strauss, Michael A.; Tojeiro, Rita; Heavens, Alan; Jimenez, Raul

    2010-01-01

    Using light curves and host galaxy spectra of 101 Type Ia supernovae (SNe Ia) with redshift z ∼ 2.4 Gyr. We find that each channel contributes roughly half of the Type Ia rate in our reference sample. We also construct the average spectra of high-stretch and low-stretch SN Ia host galaxies, and find that the difference of these spectra looks like a main-sequence B star with nebular emission lines indicative of star formation. This supports our finding that there are two populations of SNe Ia, and indicates that the progenitors of high-stretch supernovae are at the least associated with very recent star formation in the last few tens of Myr. Our results provide valuable constraints for models of Type Ia progenitors and may help improve the calibration of SNe Ia as standard candles.

  9. Interneuron progenitor transplantation to treat CNS dysfunction

    Directory of Open Access Journals (Sweden)

    Muhammad O Chohan

    2016-08-01

    Full Text Available Due to the inadequacy of endogenous repair mechanisms diseases of the nervous system remain a major challenge to scientists and clinicians. Stem cell based therapy is an exciting and viable strategy that has been shown to ameliorate or even reverse symptoms of CNS dysfunction in preclinical animal models. Of particular importance has been the use of GABAergic interneuron progenitors as a therapeutic strategy. Born in the neurogenic niches of the ventral telencephalon, interneuron progenitors retain their unique capacity to disperse, integrate and induce plasticity in adult host circuitries following transplantation. Here we discuss the potential of interneuron based transplantation strategies as it relates to CNS disease therapeutics. We also discuss mechanisms underlying their therapeutic efficacy and some of the challenges that face the field.

  10. Neural Progenitors Adopt Specific Identities by Directly Repressing All Alternative Progenitor Transcriptional Programs.

    Science.gov (United States)

    Kutejova, Eva; Sasai, Noriaki; Shah, Ankita; Gouti, Mina; Briscoe, James

    2016-03-21

    In the vertebrate neural tube, a morphogen-induced transcriptional network produces multiple molecularly distinct progenitor domains, each generating different neuronal subtypes. Using an in vitro differentiation system, we defined gene expression signatures of distinct progenitor populations and identified direct gene-regulatory inputs corresponding to locations of specific transcription factor binding. Combined with targeted perturbations of the network, this revealed a mechanism in which a progenitor identity is installed by active repression of the entire transcriptional programs of other neural progenitor fates. In the ventral neural tube, sonic hedgehog (Shh) signaling, together with broadly expressed transcriptional activators, concurrently activates the gene expression programs of several domains. The specific outcome is selected by repressive input provided by Shh-induced transcription factors that act as the key nodes in the network, enabling progenitors to adopt a single definitive identity from several initially permitted options. Together, the data suggest design principles relevant to many developing tissues. Copyright © 2016 The Authors. Published by Elsevier Inc. All rights reserved.

  11. Observational Investigations of the Progenitors of Supernovae

    Science.gov (United States)

    Lyman, J. D.

    2014-03-01

    Supernovae (SNe) are the spectacular deaths of stars and have shaped the universe we see today. Their far-reaching influence affects the chemical and dynamical evolution of galaxies, star formation, neutron star and black hole formation, and they are largely responsible for most of the heavy elements that make up the universe, including around 90 per cent of the reader. They also provide laboratories of nuclear and particle physics far beyond what we can construct on Earth and act as probes of extreme density and energy. This thesis presents new research into understanding the nature of the progenitor systems of various types of SNe, as well as presenting results that will allow their study to be more productive in the future, through use of automated pipelines and methods to increase the science value of discovered SNe. An environmental study of two peculiar types of transients ('Calcium-rich' and '2002cx-like'), which may not be true SNe, reveals extremely different ages of the exploding systems that will constrain the current theoretical effort into discovering the progenitor systems. The GRB-SN 120422A/2012bz is investigated and found to be an extremely luminous and energetic SN, even amongst the infamously bright GRB-SNe. A method is presented that allows an accurate reconstruction of the bolometric light curve of a core-collapse SN, which relies on only two optical filter observations - this will hugely reduce the observational cost of constructing bolometric light curves, a tool of great importance when hoping to constrain the nature of a SN explosion and hence its progenitor. Finally, this method is utilised to construct the largest bolometric CCSN bolometric light curve sample to date, and these are analytically modelled to reveal population statistics of the explosions, thus informing on the nature of the progenitors.

  12. EVOLUTION OF PROGENITORS FOR ELECTRON CAPTURE SUPERNOVAE

    International Nuclear Information System (INIS)

    Takahashi, Koh; Umeda, Hideyuki; Yoshida, Takashi

    2013-01-01

    We provide progenitor models for electron capture supernovae (ECSNe) with detailed evolutionary calculation. We include minor electron capture nuclei using a large nuclear reaction network with updated reaction rates. For electron capture, the Coulomb correction of rates is treated and the contribution from neutron-rich isotopes is taken into account in each nuclear statistical equilibrium (NSE) composition. We calculate the evolution of the most massive super asymptotic giant branch stars and show that these stars undergo off-center carbon burning and form ONe cores at the center. These cores become heavier up to the critical mass of 1.367 M ☉ and keep contracting even after the initiation of O+Ne deflagration. Inclusion of minor electron capture nuclei causes convective URCA cooling during the contraction phase, but the effect on the progenitor evolution is small. On the other hand, electron capture by neutron-rich isotopes in the NSE region has a more significant effect. We discuss the uniqueness of the critical core mass for ECSNe and the effect of wind mass loss on the plausibility of our models for ECSN progenitors.

  13. Puerarin Facilitates T-Tubule Development of Murine Embryonic Stem Cell-Derived Cardiomyocytes

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    Lu Wang

    2014-07-01

    Full Text Available Aims: The embryonic stem cell-derived cardiomyocytes (ES-CM is one of the promising cell sources for repopulation of damaged myocardium. However, ES-CMs present immature structure, which impairs their integration with host tissue and functional regeneration. This study used murine ES-CMs as an in vitro model of cardiomyogenesis to elucidate the effect of puerarin, the main compound found in the traditional Chinese medicine the herb Radix puerariae, on t-tubule development of murine ES-CMs. Methods: Electron microscope was employed to examine the ultrastructure. The investigation of transverse-tubules (t-tubules was performed by Di-8-ANEPPS staining. Quantitative real-time PCR was utilized to study the transcript level of genes related to t-tubule development. Results: We found that long-term application of puerarin throughout cardiac differentiation improved myofibril array and sarcomeres formation, and significantly facilitated t-tubules development of ES-CMs. The transcript levels of caveolin-3, amphiphysin-2 and junctophinlin-2, which are crucial for the formation and development of t-tubules, were significantly upregulated by puerarin treatment. Furthermore, puerarin repressed the expression of miR-22, which targets to caveolin-3. Conclusion: Our data showed that puerarin facilitates t-tubule development of murine ES-CMs. This might be related to the repression of miR-22 by puerarin and upregulation of Cav3, Bin1 and JP2 transcripts.

  14. Comparison of tumour age response to radiation for cells derived from tissue culture or solid tumours

    International Nuclear Information System (INIS)

    Keng, P.C.; Siemann, D.W.; Rochester Univ., NY; Rochester Univ., NY; Wheeler, K.T.

    1984-01-01

    Direct comparison of the cell age response of 9L and KHT tumour cells derived either from tissue culture or solid tumours was achieved. Cells from dissociated KHT and 9L tumours (the latter implanted either subcutaneously or intracerebrally) and cells from tissue culture were separated into homogenous sized populations by centrifugal elutriation. In both tumour models these homogeneous sized populations correspond to populations enriched at different stages of the cell cycle. The survival of these elutriated cell populations was measured after a single dose of Cs-137 gamma rays. For cells isolated from 9L solid tumours, there was little variation in radiosensitivity throughout the cell cycle; however, a very small but significant increase in resistance was found in late G 1 cells. This lack of a large variation in radiosensitivity through the cell cycle for 9L cells from solid tumours also was seen in 9L cells growing in monolayer tissue culture. When similar experiments were performed using the KHT sarcoma tumour model, the results showed that KHT cells in vitro exhibited a fairly conventional increase in radioresistance in both mid G 1 and late S. However, the cell age response of KHT cells from solid tumours was different; particularly in the late S and G 2 + M phases. (author)

  15. Efficient Generation of Human Embryonic Stem Cell-Derived Corneal Endothelial Cells by Directed Differentiation.

    Directory of Open Access Journals (Sweden)

    Kathryn L McCabe

    Full Text Available To generate human embryonic stem cell derived corneal endothelial cells (hESC-CECs for transplantation in patients with corneal endothelial dystrophies.Feeder-free hESC-CECs were generated by a directed differentiation protocol. hESC-CECs were characterized by morphology, expression of corneal endothelial markers, and microarray analysis of gene expression.hESC-CECs were nearly identical morphologically to primary human corneal endothelial cells, expressed Zona Occludens 1 (ZO-1 and Na+/K+ATPaseα1 (ATPA1 on the apical surface in monolayer culture, and produced the key proteins of Descemet's membrane, Collagen VIIIα1 and VIIIα2 (COL8A1 and 8A2. Quantitative PCR analysis revealed expression of all corneal endothelial pump transcripts. hESC-CECs were 96% similar to primary human adult CECs by microarray analysis.hESC-CECs are morphologically similar, express corneal endothelial cell markers and express a nearly identical complement of genes compared to human adult corneal endothelial cells. hESC-CECs may be a suitable alternative to donor-derived corneal endothelium.

  16. Functional Studies of Missense TREM2 Mutations in Human Stem Cell-Derived Microglia

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    Philip W. Brownjohn

    2018-04-01

    Full Text Available Summary: The derivation of microglia from human stem cells provides systems for understanding microglial biology and enables functional studies of disease-causing mutations. We describe a robust method for the derivation of human microglia from stem cells, which are phenotypically and functionally comparable with primary microglia. We used stem cell-derived microglia to study the consequences of missense mutations in the microglial-expressed protein triggering receptor expressed on myeloid cells 2 (TREM2, which are causal for frontotemporal dementia-like syndrome and Nasu-Hakola disease. We find that mutant TREM2 accumulates in its immature form, does not undergo typical proteolysis, and is not trafficked to the plasma membrane. However, in the absence of plasma membrane TREM2, microglia differentiate normally, respond to stimulation with lipopolysaccharide, and are phagocytically competent. These data indicate that dementia-associated TREM2 mutations have subtle effects on microglia biology, consistent with the adult onset of disease in individuals with these mutations. : Brownjohn and colleagues report methods to generate microglia from induced pluripotent human stem cells, which they demonstrate are highly similar to cultured primary human microglia. Microglia differentiated from patient-derived stem cells carrying neurological disease-causing mutations in the TREM2 receptor differentiate normally and respond appropriately to pathogenic stimuli, despite the absence of functional TREM2 receptor on the plasma membrane. Keywords: dementia, microglia, TREM2, Nasu-Hakola disease, frontotemporal dementia, iPSC-microglia, neuroinflammation

  17. Potential Therapies by Stem Cell-Derived Exosomes in CNS Diseases: Focusing on the Neurogenic Niche

    Directory of Open Access Journals (Sweden)

    Alejandro Luarte

    2016-01-01

    Full Text Available Neurodegenerative disorders are one of the leading causes of death and disability and one of the biggest burdens on health care systems. Novel approaches using various types of stem cells have been proposed to treat common neurodegenerative disorders such as Alzheimer’s Disease, Parkinson’s Disease, or stroke. Moreover, as the secretome of these cells appears to be of greater benefit compared to the cells themselves, the extracellular components responsible for its therapeutic benefit have been explored. Stem cells, as well as most cells, release extracellular vesicles such as exosomes, which are nanovesicles able to target specific cell types and thus to modify their function by delivering proteins, lipids, and nucleic acids. Exosomes have recently been tested in vivo and in vitro as therapeutic conveyors for the treatment of diseases. As such, they could be engineered to target specific populations of cells within the CNS. Considering the fact that many degenerative brain diseases have an impact on adult neurogenesis, we discuss how the modulation of the adult neurogenic niches may be a therapeutic target of stem cell-derived exosomes. These novel approaches should be examined in cellular and animal models to provide better, more effective, and specific therapeutic tools in the future.

  18. Analysis of mammary specific gene locus regulation in differentiated cells derived by somatic cell fusion

    International Nuclear Information System (INIS)

    Robinson, Claire; Kolb, Andreas F.

    2009-01-01

    The transcriptional regulation of a gene is best analysed in the context of its normal chromatin surroundings. However, most somatic cells, in contrast to embryonic stem cells, are refractory to accurate modification by homologous recombination. We show here that it is possible to introduce precise genomic modifications in ES cells and to analyse the phenotypic consequences in differentiated cells by using a combination of gene targeting, site-specific recombination and somatic cell fusion. To provide a proof of principle, we have analysed the regulation of the casein gene locus in mammary gland cells derived from modified murine ES cells by somatic cell fusion. A β-galactosidase reporter gene was inserted in place of the β-casein gene and the modified ES cells, which do not express the reporter gene, were fused with the mouse mammary gland cell line HC11. The resulting cell clones expressed the β-galactosidase gene to a similar extent and with similar hormone responsiveness as the endogenous gene. However, a reporter gene under the control of a minimal β-casein promoter (encompassing the two consensus STAT5 binding sites which mediate the hormone response of the casein genes) was unable to replicate expression levels or hormone responsiveness of the endogenous gene when inserted into the same site of the casein locus. As expected, these results implicate sequences other than the STAT5 sites in the regulation of the β-casein gene

  19. Mesenchymal Stem Cell-Derived Factors Restore Function to Human Frataxin-Deficient Cells.

    Science.gov (United States)

    Kemp, Kevin; Dey, Rimi; Cook, Amelia; Scolding, Neil; Wilkins, Alastair

    2017-08-01

    Friedreich's ataxia is an inherited neurological disorder characterised by mitochondrial dysfunction and increased susceptibility to oxidative stress. At present, no therapy has been shown to reduce disease progression. Strategies being trialled to treat Friedreich's ataxia include drugs that improve mitochondrial function and reduce oxidative injury. In addition, stem cells have been investigated as a potential therapeutic approach. We have used siRNA-induced knockdown of frataxin in SH-SY5Y cells as an in vitro cellular model for Friedreich's ataxia. Knockdown of frataxin protein expression to levels detected in patients with the disorder was achieved, leading to decreased cellular viability, increased susceptibility to hydrogen peroxide-induced oxidative stress, dysregulation of key anti-oxidant molecules and deficiencies in both cell proliferation and differentiation. Bone marrow stem cells are being investigated extensively as potential treatments for a wide range of neurological disorders, including Friedreich's ataxia. The potential neuroprotective effects of bone marrow-derived mesenchymal stem cells were therefore studied using our frataxin-deficient cell model. Soluble factors secreted by mesenchymal stem cells protected against cellular changes induced by frataxin deficiency, leading to restoration in frataxin levels and anti-oxidant defences, improved survival against oxidative stress and stimulated both cell proliferation and differentiation down the Schwann cell lineage. The demonstration that mesenchymal stem cell-derived factors can restore cellular homeostasis and function to frataxin-deficient cells further suggests that they may have potential therapeutic benefits for patients with Friedreich's ataxia.

  20. Three-dimensional bioprinting of stem-cell derived tissues for human regenerative medicine.

    Science.gov (United States)

    Skeldon, Gregor; Lucendo-Villarin, Baltasar; Shu, Wenmiao

    2018-07-05

    Stem cell technology in regenerative medicine has the potential to provide an unlimited supply of cells for drug testing, medical transplantation and academic research. In order to engineer a realistic tissue model using stem cells as an alternative to human tissue, it is essential to create artificial stem cell microenvironment or niches. Three-dimensional (3D) bioprinting is a promising tissue engineering field that offers new opportunities to precisely place stem cells within their niches layer-by-layer. This review covers bioprinting technologies, the current development of 'bio-inks' and how bioprinting has already been applied to stem-cell culture, as well as their applications for human regenerative medicine. The key considerations for bioink properties such as stiffness, stability and biodegradation, biocompatibility and printability are highlighted. Bioprinting of both adult and pluriopotent stem cells for various types of artificial tissues from liver to brain has been reviewed. 3D bioprinting of stem-cell derived tissues for human regenerative medicine is an exciting emerging area that represents opportunities for new research, industries and products as well as future challenges in clinical translation.This article is part of the theme issue 'Designer human tissue: coming to a lab near you'. © 2018 The Author(s).

  1. Steps toward Maturation of Embryonic Stem Cell-Derived Cardiomyocytes by Defined Physical Signals

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    Nian Shen

    2017-07-01

    Full Text Available Cardiovascular disease remains a leading cause of mortality and morbidity worldwide. Embryonic stem cell-derived cardiomyocytes (ESC-CMs may offer significant advances in creating in vitro cardiac tissues for disease modeling, drug testing, and elucidating developmental processes; however, the induction of ESCs to a more adult-like CM phenotype remains challenging. In this study, we developed a bioreactor system to employ pulsatile flow (1.48 mL/min, cyclic strain (5%, and extended culture time to improve the maturation of murine and human ESC-CMs. Dynamically-cultured ESC-CMs showed an increased expression of cardiac-associated proteins and genes, cardiac ion channel genes, as well as increased SERCA activity and a Raman fingerprint with the presence of maturation-associated peaks similar to primary CMs. We present a bioreactor platform that can serve as a foundation for the development of human-based cardiac in vitro models to verify drug candidates, and facilitates the study of cardiovascular development and disease.

  2. Autologous Pluripotent Stem Cell-Derived β-Like Cells for Diabetes Cellular Therapy.

    Science.gov (United States)

    Millman, Jeffrey R; Pagliuca, Felicia W

    2017-05-01

    Development of stem cell technologies for cell replacement therapy has progressed rapidly in recent years. Diabetes has long been seen as one of the first applications for stem cell-derived cells because of the loss of only a single cell type-the insulin-producing β-cell. Recent reports have detailed strategies that overcome prior hurdles to generate functional β-like cells from human pluripotent stem cells in vitro, including from human induced pluripotent stem cells (hiPSCs). Even with this accomplishment, addressing immunological barriers to transplantation remains a major challenge for the field. The development of clinically relevant hiPSC derivation methods from patients and demonstration that these cells can be differentiated into β-like cells presents a new opportunity to treat diabetes without immunosuppression or immunoprotective encapsulation or with only targeted protection from autoimmunity. This review focuses on the current status in generating and transplanting autologous β-cells for diabetes cell therapy, highlighting the unique advantages and challenges of this approach. © 2017 by the American Diabetes Association.

  3. Appearance of differentiated cells derived from polar body nuclei in the silkworm, Bombyx mori

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    Hiroki eSakai

    2013-09-01

    Full Text Available AbstractIn Bombyx mori, polar body nuclei are observed until 9h after egg lying, however, the fate of polar body nuclei remains unclear. To examine the fate of polar body nuclei, we employed a mutation of serosal cell pigmentation, pink-eyed white egg (pe. The heterozygous pe/+pe females produced black serosal cells in white eggs, while pe/pe females did not produce black serosal cells in white eggs. These results suggest that the appearance of black serosal cells in white eggs depends on the genotype (pe/ +pe of the mother. Because the polar body nuclei had +pe genes in the white eggs laid by a pe/ +pe female, polar body nuclei participate in development and differentiate into functional cell (serosal cells. Analyses of serosal cells pigmentation indicated that approximately 30% of the eggs contained polar-body-nucleus-derived cells. These results demonstrate that polar-body-nucleus-derived cells appeared at a high frequency under natural conditions. Approximately 80% of polar-body-nucleus-derived cells appeared near the anterior pole and the dorsal side, which is opposite to where embryogenesis occurs. The number of cells derived from the polar body nuclei was very low. Approximately 26 % of these eggs contained only one black serosal cell. PCR-based analysis revealed that the polar-body-nucleus-derived cells disappeared in late embryonic stages (stage 25. Overall, polar-body-nuclei-derived cells were unlikely to contribute to embryos.

  4. Engineered Microenvironments for the Maturation and Observation of Human Embryonic Stem Cell Derived Cardiomyocytes

    Science.gov (United States)

    Salick, Max R.

    The human heart is a dynamic system that undergoes substantial changes as it develops and adapts to the body's growing needs. To better understand the physiology of the heart, researchers have begun to produce immature heart muscle cells, or cardiomyocytes, from pluripotent stem cell sources with remarkable efficiency. These stem cell-derived cardiomyocytes hold great potential in the understanding and treatment of heart disease; however, even after prolonged culture, these cells continue to exhibit an immature phenotype, as indicated by poor sarcomere organization and calcium handling, among other features. The lack of maturation that is observed in these cardiomyocytes greatly limits their applicability towards drug screening, disease modeling, and cell therapy applications. The mechanical environment surrounding a cell has been repeatedly shown to have a large impact on that cell's behavior. For this reason, we have implemented micropatterning methods to mimic the level of alignment that occurs in the heart in vivo in order to study how this alignment may help the cells to produce a more mature sarcomere phenotype. It was discovered that the level of sarcomere organization of a cardiomyocyte can be strongly influenced by the micropattern lane geometry on which it adheres. Steps were taken to optimize this micropattern platform, and studies of protein organization, gene expression, and myofibrillogenesis were conducted. Additionally, a set of programs was developed to provide quantitative analysis of the level of sarcomere organization, as well as to assist with several other tissue engineering applications.

  5. Induced Pluripotent Stem Cell-Derived Red Blood Cells and Platelet Concentrates: From Bench to Bedside.

    Science.gov (United States)

    Focosi, Daniele; Amabile, Giovanni

    2017-12-27

    Red blood cells and platelets are anucleate blood components indispensable for oxygen delivery and hemostasis, respectively. Derivation of these blood elements from induced pluripotent stem (iPS) cells has the potential to develop blood donor-independent and genetic manipulation-prone products to complement or replace current transfusion banking, also minimizing the risk of alloimmunization. While the production of erythrocytes from iPS cells has challenges to overcome, such as differentiation into adult-type phenotype that functions properly after transfusion, platelet products are qualitatively and quantitatively approaching a clinically-applicable level owing to advances in expandable megakaryocyte (MK) lines, platelet-producing bioreactors, and novel reagents. Guidelines that assure the quality of iPS cells-derived blood products for clinical application represent a novel challenge for regulatory agencies. Considering the minimal risk of tumorigenicity and the expected significant demand of such products, ex vivo production of iPS-derived blood components can pave the way for iPS translation into the clinic.

  6. Effect of strenuous physical exercise on circulating cell-derived microparticles.

    Science.gov (United States)

    Chaar, Vicky; Romana, Marc; Tripette, Julien; Broquere, Cédric; Huisse, Marie-Geneviève; Hue, Olivier; Hardy-Dessources, Marie-Dominique; Connes, Philippe

    2011-01-01

    Strenuous exercise is associated with an inflammatory response involving the activation of several types of blood cells. In order to document the specific activation of these cell types, we studied the effect of three maximal exercise tests conducted to exhaustion on the quantitative and qualitative pattern of circulating cell-derived microparticles and inflammatory molecules in healthy subjects. This study mainly indicated that the plasma concentration of microparticles from platelets and polymorphonuclear neutrophils (PMN) was increased immediately after the strenuous exercise. In addition, the increase in plasma concentration of microparticles from PMN and platelets was still observed after 2 hours of recovery. A similar pattern was observed for the IL-6 plasma level. In contrast, no change was observed for either soluble selectins or plasma concentration of microparticles from red blood cells, monocytes and endothelial cells. In agreement, sVCAM-1 and sICAM-1 levels were not changed by the exercise. We conclude that a strenuous exercise is accompanied by platelet- and PMN-derived microparticle production that probably reflects the activation of these two cell types.

  7. A simple clot based assay for detection of procoagulant cell-derived microparticles.

    Science.gov (United States)

    Patil, Rucha; Ghosh, Kanjaksha; Shetty, Shrimati

    2016-05-01

    Cell-derived microparticles (MPs) are important biomarkers in many facets of medicine. However, the MP detection methods used till date are costly and time consuming. The main aim of this study was to standardize an in-house clot based screening method for MP detection which would not only be specific and sensitive, but also inexpensive. Four different methods of MP assessment were performed and the results correlated. Using the flow cytometry technique as the gold standard, 25 samples with normal phosphatidylserine (PS) expressing MP levels and 25 samples with elevated levels were selected, which was cross checked by the commercial STA Procoag PPL clotting time (CT) assay. A simple recalcification time and an in-house clot assay were the remaining two tests. The in-house test measures the CT after the addition of calcium chloride to MP rich plasma, following incubation with Russell viper venom and phospholipid free plasma. The CT obtained by the in-house assay significantly correlated with the results obtained by flow cytometry (R2=0.87, p<0.01). Though preliminary, the in-house assay seems to be efficient, inexpensive and promising. It could definitely be utilized routinely for procoagulant MP assessment in various clinical settings.

  8. Apoptotic-cell-derived membrane microparticles and IFN-α induce an inflammatory immune response.

    Science.gov (United States)

    Niessen, Anna; Heyder, Petra; Krienke, Stefan; Blank, Norbert; Tykocinski, Lars-Oliver; Lorenz, Hanns-Martin; Schiller, Martin

    2015-07-15

    A dysregulation in the clearance of apoptotic material is considered a major pathogenetic factor for the emergence of autoimmune diseases. Apoptotic-cell-derived membrane microparticles (AdMPs), which are released from the cell surface during apoptosis, have been implicated in the pathogenesis of autoimmunity. Also of importance are cytokines, such as interferon-α (IFN-α), which is known to be a major player in patients with systemic lupus erythematosus (SLE). This study investigates the combined effect of AdMPs and IFN-α on professional phagocytes. In the presence of IFN-α, phagocytosis of AdMPs by human monocytes was significantly increased in a dose-dependent manner. The combination of AdMPs and raised IFN-α concentrations resulted in an increase in the secretion of pro-inflammatory cytokines and an upregulation of surface molecule expression involved in antigen uptake. In addition, macrophage polarisation was shifted towards a more inflammatory type of cell. The synergism between IFN-α and AdMPs seemed to be mediated by an upregulation of phosphorylated STAT1. Our results indicate that IFN-α, together with AdMPs, amplify the initiation and maintenance of inflammation. This mechanism might especially play a crucial role in disorders with a defective clearance of apoptotic material. © 2015. Published by The Company of Biologists Ltd.

  9. Early incorporation of cell-derived cholesterol into pre-beta-migrating high-density lipoprotein

    International Nuclear Information System (INIS)

    Castro, G.R.; Fielding, C.J.

    1988-01-01

    Cultures of human skin fibroblasts were labeled to high cholesterol specific activity with [ 3 H]cholesterol and incubated briefly (1-3 min) with normal human plasma. The plasma was fractionated by two-dimensional agarose-polyacrylamide gel electrophoresis and the early appearance of cholesterol label among plasma lipoproteins determined. A major part of the label at 1-min incubation was in a pre-beta-migrating apo A-I lipoprotein fraction with a molecular weight of ca. 70,000. Label was enriched about 30-fold in this fraction relative to its content of apo A-I (1-2% of total apo A-I). The proportion of label in this lipoprotein was strongly correlated with its concentration in plasma. Further incubation (2 min) in the presence of unlabeled cells demonstrated transfer of label from this fraction to a higher molecular weight pre-beta apo A-I species, to low-density lipoprotein, and to the alpha-migrating apo A-I that made up the bulk (96%) of total apo A-I in plasma. The data suggest that a significant part of cell-derived cholesterol is transferred specifically to a pre-beta-migrating lipoprotein A-I species as part of a cholesterol transport transfer sequence in plasma

  10. Potential of laryngeal muscle regeneration using induced pluripotent stem cell-derived skeletal muscle cells.

    Science.gov (United States)

    Dirja, Bayu Tirta; Yoshie, Susumu; Ikeda, Masakazu; Imaizumi, Mitsuyoshi; Nakamura, Ryosuke; Otsuki, Koshi; Nomoto, Yukio; Wada, Ikuo; Hazama, Akihiro; Omori, Koichi

    2016-01-01

    Conclusion Induced pluripotent stem (iPS) cells may be a new potential cell source for laryngeal muscle regeneration in the treatment of vocal fold atrophy after recurrent laryngeal nerve paralysis. Objectives Unilateral vocal fold paralysis can lead to degeneration, atrophy, and loss of force of the thyroarytenoid muscle. At present, there are some treatments such as thyroplasty, arytenoid adduction, and vocal fold injection. However, such treatments cannot restore reduced mass of the thyroarytenoid muscle. iPS cells have been recognized as supplying a potential resource for cell transplantation. The aim of this study was to assess the effectiveness of the use of iPS cells for the regeneration of laryngeal muscle through the evaluation of both in vitro and in vivo experiments. Methods Skeletal muscle cells were generated from tdTomato-labeled iPS cells using embryoid body formation. Differentiation into skeletal muscle cells was analyzed by gene expression and immunocytochemistry. The tdTomato-labeled iPS cell-derived skeletal muscle cells were transplanted into the left atrophied thyroarytenoid muscle. To evaluate the engraftment of these cells after transplantation, immunohistochemistry was performed. Results The tdTomato-labeled iPS cells were successfully differentiated into skeletal muscle cells through an in vitro experiment. These cells survived in the atrophied thyroarytenoid muscle after transplantation.

  11. Novel Application of Stem Cell-Derived Neurons to Evaluate the Time-and Dose-Dependent Progression of Excitotoxic Injury

    Science.gov (United States)

    2013-05-14

    3. DATES COVERED (From - To) 4. TITLE AND SUBTITLE Novel Application of Stem Cell -Derived Neurons to Evaluate the Time- and Dose-Dependent 5a...01.10.RC.021). 14. ABSTRACT See reprint. 15. SUBJECT TERMS Mouse stem cell -derived neurons, NMDA receptor, AMPA receptor, Neurotoxicity, Apoptosis...area code) 410-436-8044 Standard Form 298 (Rev. 8-98) Prescribed by ANSI Std. Z39.18 Novel Application of Stem Cell -Derived Neurons to Evaluate the

  12. Community effect triggers terminal differentiation of myogenic cells derived from muscle satellite cells by quenching Smad signaling

    Energy Technology Data Exchange (ETDEWEB)

    Yanagisawa, Michiko [Department of Regenerative Medicine, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 35 Gengo, Morioka, Oobu, Aichi 474-8522 (Japan); Aging Research, Nagoya University Graduate School of Medicine, 65 Tsurumai-cho, Showa-ku, Nagoya, Aichi 466-8550 (Japan); Mukai, Atsushi; Shiomi, Kosuke [Department of Regenerative Medicine, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 35 Gengo, Morioka, Oobu, Aichi 474-8522 (Japan); Song, Si-Yong [Institute of Neuroscience, Faculty of Pharmaceutical Sciences at Kagawa, Tokushima Bunri University, 1314-1 Shido, Sanuki-shi, Kagawa 769-2193 (Japan); Hashimoto, Naohiro, E-mail: nao@ncgg.go.jp [Department of Regenerative Medicine, National Institute for Longevity Sciences, National Center for Geriatrics and Gerontology, 35 Gengo, Morioka, Oobu, Aichi 474-8522 (Japan)

    2011-01-15

    A high concentration of bone morphogenetic proteins (BMPs) stimulates myogenic progenitor cells to undergo heterotopic osteogenic differentiation. However, the physiological role of the Smad signaling pathway during terminal muscle differentiation has not been resolved. We report here that Smad1/5/8 was phosphorylated and activated in undifferentiated growing mouse myogenic progenitor Ric10 cells without exposure to any exogenous BMPs. The amount of phosphorylated Smad1/5/8 was severely reduced during precocious myogenic differentiation under the high cell density culture condition even in growth medium supplemented with a high concentration of serum. Inhibition of the Smad signaling pathway by dorsomorphin, an inhibitor of Smad activation, or noggin, a specific antagonist of BMP, induced precocious terminal differentiation of myogenic progenitor cells in a cell density-dependent fashion even in growth medium. In addition, Smad1/5/8 was transiently activated in proliferating myogenic progenitor cells during muscle regeneration in rats. The present results indicate that the Smad signaling pathway is involved in a critical switch between growth and differentiation of myogenic progenitor cells both in vitro and in vivo. Furthermore, precocious cell density-dependent myogenic differentiation suggests that a community effect triggers the terminal muscle differentiation of myogenic cells by quenching the Smad signaling.

  13. Community effect triggers terminal differentiation of myogenic cells derived from muscle satellite cells by quenching Smad signaling

    International Nuclear Information System (INIS)

    Yanagisawa, Michiko; Mukai, Atsushi; Shiomi, Kosuke; Song, Si-Yong; Hashimoto, Naohiro

    2011-01-01

    A high concentration of bone morphogenetic proteins (BMPs) stimulates myogenic progenitor cells to undergo heterotopic osteogenic differentiation. However, the physiological role of the Smad signaling pathway during terminal muscle differentiation has not been resolved. We report here that Smad1/5/8 was phosphorylated and activated in undifferentiated growing mouse myogenic progenitor Ric10 cells without exposure to any exogenous BMPs. The amount of phosphorylated Smad1/5/8 was severely reduced during precocious myogenic differentiation under the high cell density culture condition even in growth medium supplemented with a high concentration of serum. Inhibition of the Smad signaling pathway by dorsomorphin, an inhibitor of Smad activation, or noggin, a specific antagonist of BMP, induced precocious terminal differentiation of myogenic progenitor cells in a cell density-dependent fashion even in growth medium. In addition, Smad1/5/8 was transiently activated in proliferating myogenic progenitor cells during muscle regeneration in rats. The present results indicate that the Smad signaling pathway is involved in a critical switch between growth and differentiation of myogenic progenitor cells both in vitro and in vivo. Furthermore, precocious cell density-dependent myogenic differentiation suggests that a community effect triggers the terminal muscle differentiation of myogenic cells by quenching the Smad signaling.

  14. Poly(trimethylene carbonate) as an elastic biodegradable film for human embryonic stem cell-derived retinal pigment epithelial cells

    NARCIS (Netherlands)

    Sorkio, Anni; Haimi, Suvi; Verdoold, Vincent; Juuti-Uusitalo, Kati; Grijpma, Dirk; Skottman, Heli

    2017-01-01

    Human embryonic stem cell-derived retinal pigment epithelial (hESC-RPE) cell therapies show tremendous potential for the treatment of retinal degenerative diseases. A tissue engineering approach, where cells are delivered to the subretinal space on a biodegradable carrier as a sheet, shows great

  15. Poly(trimethylene carbonate) as an elastic biodegradable film for human embryonic stem cell-derived retinal pigment epithelial cells

    NARCIS (Netherlands)

    Sorkio, Anni; Haimi, Suvi; Verdoold, Vincent; Juuti-Uusitalo, Kati; Grijpma, Dirk; Skottman, Heli

    Human embryonic stem cell-derived retinal pigment epithelial (hESC-RPE) cell therapies show tremendous potential for the treatment of retinal degenerative diseases. A tissue engineering approach, where cells are delivered to the subretinal space on a biodegradable carrier as a sheet, shows great

  16. Feeder-cell-independent culture of the pig-embryonic-stem-cell-derived exocrine pancreatic cell line, PICM-31

    Science.gov (United States)

    The adaptation to feeder-independent growth of a pig embryonic stem cell-derived pancreatic cell line is described. The parental PICM-31 cell line, previously characterized as an exocrine pancreas cell line, was colony-cloned two times in succession resulting in the subclonal cell line, PICM-31A1. P...

  17. Induced pluripotent stem cell-derived neuron as a human model for testing environmentally induced developmental neurotoxicity

    Science.gov (United States)

    Induced pluripotent stem cell-derived neurons as a human model for testing environmentally induced developmental neurotoxicity Ingrid L. Druwe1, Timothy J. Shafer2, Kathleen Wallace2, Pablo Valdivia3 ,and William R. Mundy2. 1University of North Carolina, Curriculum in Toxicology...

  18. Stromal cell-derived factor-1 alpha (SDF-1 alpha) improves neural recovery after spinal cord contusion in rats

    NARCIS (Netherlands)

    Zendedel, A.; Nobakht, M.; Bakhtiyari, M.; Beyer, C.; Kipp, M.; Baazm, M.; Joghataie, M.T.

    2012-01-01

    Stromal cell-derived factor-1 alpha (SDF-1α) is an important cytokine, implicated in the control of stem cell trafficking and bone marrow-derived stem cell mobilization. Generally, SDF-1α regulates multiple physiological processes such as embryonic development and organ homeostasis. There is also

  19. Evaluation of the size segregation of elemental carbon (EC) emission in Europe: Influence on the simulation of EC long-range transportation

    NARCIS (Netherlands)

    Chen, Y.; Cheng, Y.F.; Nordmann, S.; Birmili, W.; Denier Van Der Gon, H.A.C.; Ma, N.; Wolke, R.; Wehner, B.; Sun, J.; Spindler, G.; Mu, Q.; Pöschl, U.; Su, H.; Wiedensohler, A.

    2016-01-01

    Elemental Carbon (EC) has a significant impact on human health and climate change. In order to evaluate the size segregation of EC emission in the EUCAARI inventory and investigate its influence on the simulation of EC long-range transportation in Europe, we used the fully coupled online Weather

  20. [Analysis of factors related to the number of mesenchymal stem cells derived from synovial fluid of the temporomandibular joint].

    Science.gov (United States)

    Sun, Y P; Zheng, Y H; Zhang, Z G

    2017-06-09

    Objective: To analyze related factors on the number of mesenchymal stem cells in the synovial fluid of the temporomandibular joint (TMJ) and provide an research basis for understanding of the source and biological role of mesenchymal stem cells derived from synovial fluid in TMJ. Methods: One hundred and twenty-two synovial fluid samples from 91 temporomandibular disorders (TMD) patients who visited in Department of TMJ Center, Guanghua School of Stomatology, Hospital of Stomatology, Sun Yat-sen University from March 2013 to December 2013 were collected in this study, and 6 TMJ synovial fluid samples from 6 normal volunteers who were studying in the North Campus of Sun Yat-sen University were also collected, so did their clinical information. Then the relation between the number of mesenchymal stem cells derived from synovial fluid and the health status of the joints, age of donor, disc perforation, condylar bony destruction, blood containing and visual analogue scale score of pain were investigated using Mann-Whitney U test and Spearman rank correlation test. Results: The number of mesenchymal stem cells derived from synovial fluid had no significant relation with visual analogue scale score of pain ( r= 0.041, P= 0.672), blood containing ( P= 0.063), condylar bony destruction ( P= 0.371). Linear correlation between the number of mesenchymal stem cells derived from synovial fluid and age of donor was very week ( r= 0.186, P= 0.043). The number of mesenchymal stem cells up-regulated when the joint was in a disease state ( P= 0.001). The disc perforation group had more mesenchymal stem cells in synovial fluid than without disc perforation group ( P= 0.042). Conclusions: The number of mesenchymal stem cells derived from synovial fluid in TMJ has no correlation with peripheral blood circulation and condylar bony destruction, while has close relation with soft tissue structure damage of the joint.

  1. Coculturing with endothelial cells promotes in vitro maturation and electrical coupling of human embryonic stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Pasquier, Jennifer; Gupta, Renuka; Rioult, Damien; Hoarau-Véchot, Jessica; Courjaret, Raphael; Machaca, Khaled; Al Suwaidi, Jassim; Stanley, Edouard G; Rafii, Shahin; Elliott, David A; Abi Khalil, Charbel; Rafii, Arash

    2017-06-01

    Pluripotent human embryonic stem cells (hESC) are a promising source of repopulating cardiomyocytes. We hypothesized that we could improve maturation of cardiomyocytes and facilitate electrical interconnections by creating a model that more closely resembles heart tissue; that is, containing both endothelial cells (ECs) and cardiomyocytes. We induced cardiomyocyte differentiation in the coculture of an hESC line expressing the cardiac reporter NKX2.5-green fluorescent protein (GFP), and an Akt-activated EC line (E4 + ECs). We quantified spontaneous beating rates, synchrony, and coordination between different cardiomyocyte clusters using confocal imaging of Fura Red-detected calcium transients and computer-assisted image analysis. After 8 days in culture, 94% ± 6% of the NKX2-5GFP + cells were beating when hESCs embryonic bodies were plated on E4 + ECs compared with 34% ± 12.9% for controls consisting of hESCs cultured on BD Matrigel (BD Biosciences) without ECs at Day 11 in culture. The spatial organization of beating areas in cocultures was different. The GFP + cardiomyocytes were close to the E4 + ECs. The average beats/min of the cardiomyocytes in coculture was faster and closer to physiologic heart rates compared with controls (50 ± 14 [n = 13] vs 25 ± 9 [n = 8]; p < 0.05). The coculture with ECs led to synchronized beating relying on the endothelial network, as illustrated by the loss of synchronization upon the disruption of endothelial bridges. The coculturing of differentiating cardiomyocytes with Akt-activated ECs but not EC-conditioned media results in (1) improved efficiency of the cardiomyocyte differentiation protocol and (2) increased maturity leading to better intercellular coupling with improved chronotropy and synchrony. Copyright © 2017. Published by Elsevier Inc.

  2. Human pluripotent stem cell-derived erythropoietin-producing cells ameliorate renal anemia in mice.

    Science.gov (United States)

    Hitomi, Hirofumi; Kasahara, Tomoko; Katagiri, Naoko; Hoshina, Azusa; Mae, Shin-Ichi; Kotaka, Maki; Toyohara, Takafumi; Rahman, Asadur; Nakano, Daisuke; Niwa, Akira; Saito, Megumu K; Nakahata, Tatsutoshi; Nishiyama, Akira; Osafune, Kenji

    2017-09-27

    The production of erythropoietin (EPO) by the kidneys, a principal hormone for the hematopoietic system, is reduced in patients with chronic kidney disease (CKD), eventually resulting in severe anemia. Although recombinant human EPO treatment improves anemia in patients with CKD, returning to full red blood cell production without fluctuations does not always occur. We established a method to generate EPO-producing cells from human induced pluripotent stem cells (hiPSCs) by modifying previously reported hepatic differentiation protocols. These cells showed increased EPO expression and secretion in response to low oxygen conditions, prolyl hydroxylase domain-containing enzyme inhibitors, and insulin-like growth factor 1. The EPO protein secreted from hiPSC-derived EPO-producing (hiPSC-EPO) cells induced the erythropoietic differentiation of human umbilical cord blood progenitor cells in vitro. Furthermore, transplantation of hiPSC-EPO cells into mice with CKD induced by adenine treatment improved renal anemia. Thus, hiPSC-EPO cells may be a useful tool for clarifying the mechanisms of EPO production and may be useful as a therapeutic strategy for treating renal anemia. Copyright © 2017 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.

  3. Selective labelling of stromal cell-derived factor 1α with carboxyfluorescein to study receptor internalisation.

    Science.gov (United States)

    Bellmann-Sickert, Kathrin; Baumann, Lars; Beck-Sickinger, Annette G

    2010-10-01

    SDF1α plays an important role in the regeneration of injured tissue after ischemia or stroke by inducing the migration of progenitor cells. In order to study the function of this therapeutically relevant chemokine site-specific protein labelling is of great interest. However, modification of SDF1α is complicated because of its complex tertiary structure. Here, we describe the first site-specific fluorescent modification of SDF1α by EPL. We recombinantly expressed SDF1α (1-49) by intein-mediated protein expression. The C-terminal peptide SDF1α (50-68) was synthesised by SPPS and selectively labelled with carboxyfluorescein at Lys(56). In a cell migration assay, M-[K(56)(CF)]SDF1α showed a clear potency to induce chemotaxis of human T-cell leukaemia cells. Microscopic analysis on HEK293 cells transfected with the CXCR4 revealed specific binding of the fluorescent ligand. Furthermore, receptor-induced internalisation of the ligand could be visualised. These results show that site-specific modification of SDF1α yields in a biologically functional molecule that allows the characterisation of CXCR4 production of cells on a molecular level. © 2010 European Peptide Society and John Wiley & Sons, Ltd.

  4. [Characterization of stem cells derived from the neonatal auditory sensory epithelium].

    Science.gov (United States)

    Diensthuber, M; Heller, S

    2010-11-01

    In contrast to regenerating hair cell-bearing organs of nonmammalian vertebrates the adult mammalian organ of Corti appears to have lost its ability to maintain stem cells. The result is a lack of regenerative ability and irreversible hearing loss following auditory hair cell death. Unexpectedly, the neonatal auditory sensory epithelium has recently been shown to harbor cells with stem cell features. The origin of these cells within the cochlea's sensory epithelium is unknown. We applied a modified neurosphere assay to identify stem cells within distinct subregions of the neonatal mouse auditory sensory epithelium. Sphere cells were characterized by multiple markers and morphologic techniques. Our data reveal that both the greater and the lesser epithelial ridge contribute to the sphere-forming stem cell population derived from the auditory sensory epithelium. These self-renewing sphere cells express a variety of markers for neural and otic progenitor cells and mature inner ear cell types. Stem cells can be isolated from specific regions of the auditory sensory epithelium. The distinct features of these cells imply a potential application in the development of a cell replacement therapy to regenerate the damaged sensory epithelium.

  5. Human induced pluripotent stem cell-derived models to investigate human cytomegalovirus infection in neural cells.

    Directory of Open Access Journals (Sweden)

    Leonardo D'Aiuto

    Full Text Available Human cytomegalovirus (HCMV infection is one of the leading prenatal causes of congenital mental retardation and deformities world-wide. Access to cultured human neuronal lineages, necessary to understand the species specific pathogenic effects of HCMV, has been limited by difficulties in sustaining primary human neuronal cultures. Human induced pluripotent stem (iPS cells now provide an opportunity for such research. We derived iPS cells from human adult fibroblasts and induced neural lineages to investigate their susceptibility to infection with HCMV strain Ad169. Analysis of iPS cells, iPS-derived neural stem cells (NSCs, neural progenitor cells (NPCs and neurons suggests that (i iPS cells are not permissive to HCMV infection, i.e., they do not permit a full viral replication cycle; (ii Neural stem cells have impaired differentiation when infected by HCMV; (iii NPCs are fully permissive for HCMV infection; altered expression of genes related to neural metabolism or neuronal differentiation is also observed; (iv most iPS-derived neurons are not permissive to HCMV infection; and (v infected neurons have impaired calcium influx in response to glutamate.

  6. Oral keratinocyte stem/progenitor cells: specific markers, molecular signaling pathways and potential uses.

    Science.gov (United States)

    Calenic, Bogdan; Greabu, Maria; Caruntu, Constantin; Tanase, Cristiana; Battino, Maurizio

    2015-10-01

    Oral keratinocyte stem cells reside in the basal layers of the oral epithelium, representing a minor population of cells with a great potential to self-renew and proliferate over the course of their lifetime. As a result of the potential uses of oral keratinocyte stem cells in regenerative medicine and the key roles they play in tissue homeostasis, inflammatory conditions, wound healing and tumor initiation and progression, intense scientific efforts are currently being undertaken to identify, separate and reprogram these cells. Although currently there is no specific marker that can characterize and isolate oral keratinocyte stem cells, several suggestions have been made. Thus, different stem/progenitor-cell subpopulations have been categorized based on combinations of positive and/or negative membrane-surface markers, which include integrins, clusters of differentiation and cytokeratins. Important advances have also been made in understanding the molecular pathways that govern processes such as self-renewal, differentiation, proliferation, wound healing and programmed cell death. A thorough understanding of stem-cell biology and the molecular players that govern cellular fate is paramount in the quest for using stem-cell-derived therapies in the treatment of various oral pathologies. The current review focuses on recent advances in understanding the molecular signaling pathways coordinating the behavior of these cells and in identifying suitable markers used for their isolation and characterization. Special emphasis will also be placed on the roles played by oral keratinocyte stem and progenitor cells in normal and diseased oral tissues and on their potential uses in the fields of general medicine and dentistry. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  7. Characterization of stem and progenitor cells in the dental pulp of erupted and unerupted murine molars

    Science.gov (United States)

    Balic, Anamaria; Aguila, H. Leonardo; Caimano, Melissa J.; Francone, Victor P.; Mina, Mina

    2010-01-01

    In the past few years there have been significant advances in the identification of putative stem cells also referred to as “mesenchymal stem cells” (MSC) in dental tissues including the dental pulp. It is thought that MSC in dental pulp share certain similarities with MSC isolated from other tissues. However, cells in dental pulp are still poorly characterized. This study focused on the characterization of progenitor and stem cells in dental pulps of erupted and unerupted mice molars. Our study showed that dental pulps from unerupted molars contain a significant number of cells expressing CD90+/CD45-, CD117+/CD45-, Sca-1+/CD45- and little if any CD45+ cells. Our in vitro functional studies showed that dental pulp cells from unerupted molars displayed extensive osteo-dentinogenic potential but were unable to differentiate into chondrocytes and adipocytes. Dental pulp from erupted molars displayed a reduced number of cells, contained higher percentage of CD45+ and lower percentage of cells expressing CD90+/CD45-, CD117+/CD45- as compared to unerupted molars. In vitro functional assays demonstrated the ability of a small fraction of cells to differentiate into odontoblasts, osteoblasts, adipocytes and chondrocytes. There was a significant reduction in the osteo-dentinogenic potential of the pulp cells derived from erupted molars compared to unerupted molars. Furthermore, the adipogenic and chondrogenic differentiation of pulp cells from erupted molars was dependent on a long induction period and infrequent. Based on these findings we propose that the dental pulp of the erupted molars contain a small population of multipotent cells, whereas the dental pulp of the unerupted molars does not contain multipotent cells but is enriched in osteo-dentinogenic progenitors engaged in the formation of coronal and radicular odontoblasts. PMID:20193787

  8. Comparison of immunodulatory properties of dental pulp stem cells derived from healthy and inflamed teeth.

    Science.gov (United States)

    Yazid, Farinawati Binti; Gnanasegaran, Nareshwaran; Kunasekaran, Wijenthiran; Govindasamy, Vijayendran; Musa, Sabri

    2014-12-01

    The aim of this study was to investigate the immunodulatory properties of dental pulp stem cells derived from healthy (SCD) and inflamed pulp deciduous (SCDIP) tissues. The overall hypothesis is that SCDIP possess equal immune properties with SCD and could be used as an alternative tissue source in regenerative medicine. An intra-oral examination was carried out to assess the status of the pulp tissues and group them according to healthy or inflamed. Primary cells were established from these groups, and basic mesenchymal stem cells (MSC) characterizations were conducted. The expression of human leukocyte antigen (HLA), namely HLA-G, HLA-DR, and HLA-ABC were examined in both cell lines using flow cytometry. We further compared the immunosuppressive effects of SCD and SCDIP on phytohemagglutinin-induced T cell proliferation. Supernatants were tested for cytokine profiling using multiplex array. While SCD exhibited typical MSC characteristics, SCDIP on the other hand, did not. Compared with SCDIP, SCD effectively suppresses mitogen-induced T cells proliferation in a dose-dependent manner, as well as express a higher percentage of HLA-ABC and HLA-G. In addition, levels of several cytokines, such as TNF-α, TNF-β, and IL-2, were drastically suppressed in SCD than SCDIP. Furthermore, a high level of IL-10, an important anti-inflammatory cytokine, was present in SCD compared with SCDIP. These findings suggest that SCDIP is highly dysfunctional in terms of their stemness and immunomodulatory properties. SCDIP is not a viable therapeutic cell source especially when used in graft versus host disease (GvHD) and organ rejection.

  9. Generation of induced pluripotent stem cell-derived mice by reprogramming of a mature NKT cell.

    Science.gov (United States)

    Ren, Yue; Dashtsoodol, Nyambayar; Watarai, Hiroshi; Koseki, Haruhiko; Quan, Chengshi; Taniguchi, Masaru

    2014-10-01

    NKT cells are characterized by their expression of an NKT-cell-specific invariant antigen-receptor α chain encoded by Vα14Jα18 gene segments. These NKT cells bridge the innate and acquired immune systems to mediate effective and augmented responses; however, the limited number of NKT cells in vivo hampers their analysis. Here, two lines of induced pluripotent stem cell-derived mice (NKT-iPSC-derived mice) were generated by reprogramming of mature NKT cells, where one harbors both rearranged Vα14Jα18 and Vβ7 genes and the other carries rearranged Vα14Jα18 on both alleles but germline Vβ loci. The analysis of NKT-iPSC-derived mice showed a significant increase in NKT cell numbers with relatively normal frequencies of functional subsets, but significantly enhanced in some cases, and acquired functional NKT cell maturation in peripheral lymphoid organs. NKT-iPSC-derived mice also showed normal development of other immune cells except for the absence of γδT cells and disturbed development of conventional CD4 αβT cells. These results suggest that the NKT-iPSC-derived mice are a better model for NKT cell development and function study rather than transgenic mouse models reported previously and also that the presence of a pre-rearranged Vα14Jα18 in the natural chromosomal context favors the developmental fate of NKT cells. © The Author 2014. Published by Oxford University Press on behalf of The Japanese Society for Immunology.

  10. Slow conduction in mixed cultured strands of primary ventricular cells and stem cell-derived cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Jan Pavel Kucera

    2015-09-01

    Full Text Available Modern concepts for the treatment of myocardial diseases focus on novel cell therapeutic strategies involving stem cell-derived cardiomyocytes (SCMs. However, functional integration of SCMs requires similar electrophysiological properties as primary cardiomyocytes (PCMs and the ability to establish intercellular connections with host myocytes in order to contribute to the electrical and mechanical activity of the heart. The aim of this project was to investigate the properties of cardiac conduction in a co-culture approach using SCMs and PCMs in cultured cell strands. Murine embryonic SCMs were pooled with fetal ventricular cells and seeded in predefined proportions on microelectrode arrays to form patterned strands of mixed cells. Conduction velocity (CV was measured during steady state pacing. SCM excitability was estimated from action potentials measured in single cells using the patch clamp technique. Experiments were complemented with computer simulations of conduction using a detailed model of cellular architecture in mixed cell strands.CV was significantly lower in strands composed purely of SCMs (5.5±1.5 cm/s, n=11 as compared to PCMs (34.9±2.9 cm/s, n=21 at similar refractoriness (100% SCMs: 122±25 ms, n=9; 100% PCMs: 139±67 ms, n=14. In mixed strands combining both cell types, CV was higher than in pure SCMs strands, but always lower than in 100% PCM strands. Computer simulations demonstrated that both intercellular coupling and electrical excitability limit CV.These data provide evidence that in cultures of murine ventricular cardiomyocytes, SCMs cannot restore CV to control levels resulting in slow conduction, which may lead to reentry circuits and arrhythmias.

  11. Stromal cell-derived factor 1 regulates the actin organization of chondrocytes and chondrocyte hypertrophy.

    Directory of Open Access Journals (Sweden)

    Koichi Murata

    Full Text Available Stromal cell-derived factor 1 (SDF-1/CXCL12/PBSF plays important roles in the biological and physiological functions of haematopoietic and mesenchymal stem cells. This chemokine regulates the formation of multiple organ systems during embryogenesis. However, its roles in skeletal development remain unclear. Here we investigated the roles of SDF-1 in chondrocyte differentiation. We demonstrated that SDF-1 protein was expressed at pre-hypertrophic and hypertrophic chondrocytes in the newly formed endochondral callus of rib fracture as well as in the growth plate of normal mouse tibia by immunohistochemical analysis. Using SDF-1(-/- mouse embryo, we histologically showed that the total length of the whole humeri of SDF-1(-/- mice was significantly shorter than that of wild-type mice, which was contributed mainly by shorter hypertrophic and calcified zones in SDF-1(-/- mice. Actin cytoskeleton of hypertrophic chondrocytes in SDF-1(-/- mouse humeri showed less F-actin and rounder shape than that of wild-type mice. Primary chondrocytes from SDF-1(-/- mice showed the enhanced formation of philopodia and loss of F-actin. The administration of SDF-1 to primary chondrocytes of wild-type mice and SDF-1(-/- mice promoted the formation of actin stress fibers. Organ culture of embryonic metatarsals from SDF-1(-/- mice showed the growth delay, which was recovered by an exogenous administration of SDF-1. mRNA expression of type X collagen in metatarsals and in primary chondrocytes of SDF-1(-/- mouse embryo was down-regulated while the administration of SDF-1 to metatarsals recovered. These data suggests that SDF-1 regulates the actin organization and stimulates bone growth by mediating chondrocyte hypertrophy.

  12. Stromal cell-derived factor 1 regulates the actin organization of chondrocytes and chondrocyte hypertrophy.

    Science.gov (United States)

    Murata, Koichi; Kitaori, Toshiyuki; Oishi, Shinya; Watanabe, Naoki; Yoshitomi, Hiroyuki; Tanida, Shimei; Ishikawa, Masahiro; Kasahara, Takashi; Shibuya, Hideyuki; Fujii, Nobutaka; Nagasawa, Takashi; Nakamura, Takashi; Ito, Hiromu

    2012-01-01

    Stromal cell-derived factor 1 (SDF-1/CXCL12/PBSF) plays important roles in the biological and physiological functions of haematopoietic and mesenchymal stem cells. This chemokine regulates the formation of multiple organ systems during embryogenesis. However, its roles in skeletal development remain unclear. Here we investigated the roles of SDF-1 in chondrocyte differentiation. We demonstrated that SDF-1 protein was expressed at pre-hypertrophic and hypertrophic chondrocytes in the newly formed endochondral callus of rib fracture as well as in the growth plate of normal mouse tibia by immunohistochemical analysis. Using SDF-1(-/-) mouse embryo, we histologically showed that the total length of the whole humeri of SDF-1(-/-) mice was significantly shorter than that of wild-type mice, which was contributed mainly by shorter hypertrophic and calcified zones in SDF-1(-/-) mice. Actin cytoskeleton of hypertrophic chondrocytes in SDF-1(-/-) mouse humeri showed less F-actin and rounder shape than that of wild-type mice. Primary chondrocytes from SDF-1(-/-) mice showed the enhanced formation of philopodia and loss of F-actin. The administration of SDF-1 to primary chondrocytes of wild-type mice and SDF-1(-/-) mice promoted the formation of actin stress fibers. Organ culture of embryonic metatarsals from SDF-1(-/-) mice showed the growth delay, which was recovered by an exogenous administration of SDF-1. mRNA expression of type X collagen in metatarsals and in primary chondrocytes of SDF-1(-/-) mouse embryo was down-regulated while the administration of SDF-1 to metatarsals recovered. These data suggests that SDF-1 regulates the actin organization and stimulates bone growth by mediating chondrocyte hypertrophy.

  13. Developmental changes in electrophysiological characteristics of human induced Pluripotent Stem Cell-derived cardiomyocytes

    Science.gov (United States)

    Ben-Ari, Meital; Naor, Shulamit; Zeevi-Levin, Naama; Schick, Revital; Ben Jehuda, Ronen; Reiter, Irina; Raveh, Amit; Grijnevitch, Inna; Barak, Omri; Rosen, Michael R.; Weissman, Amir; Binah, Ofer

    2016-01-01

    Background Previous studies proposed that throughout differentiation of human induced Pluripotent Stem Cell-derived cardiomyocytes (iPSC-CMs) only 3 types of action potentials (AP) exist: nodal, atrial and ventricular-like. Objective To investigate whether there are precisely 3 phenotypes or a continuum exists among them, we tested 2 hypotheses: (1) during culture development a cardiac precursor cell is present that - depending on age - can evolve into the 3 phenotypes. (2) The predominant pattern is early prevalence of nodal phenotype, transient appearance of atrial phenotype, evolution to ventricular phenotype, and persistence of transitional phenotypes. Methods To test these hypotheses we: (1) performed FACS analysis of nodal, atrial and ventricular markers; (2) recorded AP from 280 7-to-95 day old iPSC-CMs; (3) analyzed AP characteristics. Results The major findings were: (1) FACS analysis of 30 and 60-day old cultures showed that an iPSC-CMs population shifts from nodal into atrial/ventricular phenotype, while including significant transitional populations.(2) The AP population did not consist of 3 distinct phenotypes; (3) Culture aging was associated with a shift from nodal to ventricular dominance, with a transient (57–70 days) appearance of atrial phenotype; (4) Beat Rate Variability was more prominent in nodal than ventricular cardiomyocytes while If density increased in older cultures. Conclusions From the onset of development the iPSC-CMs population includes nodal, atrial and ventricular AP and a broad spectrum of transitional phenotypes. The most readily distinguishable phenotype is atrial which appears only transiently, yet dominates at 57–70 days of evolution. PMID:27639456

  14. Developmental changes in electrophysiological characteristics of human-induced pluripotent stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Ben-Ari, Meital; Naor, Shulamit; Zeevi-Levin, Naama; Schick, Revital; Ben Jehuda, Ronen; Reiter, Irina; Raveh, Amit; Grijnevitch, Inna; Barak, Omri; Rosen, Michael R; Weissman, Amir; Binah, Ofer

    2016-12-01

    Previous studies proposed that throughout differentiation of human induced Pluripotent Stem Cell-derived cardiomyocytes (iPSC-CMs), only 3 types of action potentials (APs) exist: nodal-, atrial-, and ventricular-like. To investigate whether there are precisely 3 phenotypes or a continuum exists among them, we tested 2 hypotheses: (1) During culture development a cardiac precursor cell is present that-depending on age-can evolve into the 3 phenotypes. (2) The predominant pattern is early prevalence of a nodal phenotype, transient appearance of an atrial phenotype, evolution to a ventricular phenotype, and persistence of transitional phenotypes. To test these hypotheses, we (1) performed fluorescence-activated cell sorting analysis of nodal, atrial, and ventricular markers; (2) recorded APs from 280 7- to 95-day-old iPSC-CMs; and (3) analyzed AP characteristics. The major findings were as follows: (1) fluorescence-activated cell sorting analysis of 30- and 60-day-old cultures showed that an iPSC-CMs population shifts from the nodal to the atrial/ventricular phenotype while including significant transitional populations; (2) the AP population did not consist of 3 phenotypes; (3) culture aging was associated with a shift from nodal to ventricular dominance, with a transient (57-70 days) appearance of the atrial phenotype; and (4) beat rate variability was more prominent in nodal than in ventricular cardiomyocytes, while pacemaker current density increased in older cultures. From the onset of development in culture, the iPSC-CMs population includes nodal, atrial, and ventricular APs and a broad spectrum of transitional phenotypes. The most readily distinguishable phenotype is atrial, which appears only transiently yet dominates at 57-70 days of evolution. Copyright © 2016 Heart Rhythm Society. Published by Elsevier Inc. All rights reserved.

  15. Modeling chemotherapeutic neurotoxicity with human induced pluripotent stem cell-derived neuronal cells.

    Directory of Open Access Journals (Sweden)

    Heather E Wheeler

    Full Text Available There are no effective agents to prevent or treat chemotherapy-induced peripheral neuropathy (CIPN, the most common non-hematologic toxicity of chemotherapy. Therefore, we sought to evaluate the utility of human neuron-like cells derived from induced pluripotent stem cells (iPSCs as a means to study CIPN. We used high content imaging measurements of neurite outgrowth phenotypes to compare the changes that occur to iPSC-derived neuronal cells among drugs and among individuals in response to several classes of chemotherapeutics. Upon treatment of these neuronal cells with the neurotoxic drug paclitaxel, vincristine or cisplatin, we identified significant differences in five morphological phenotypes among drugs, including total outgrowth, mean/median/maximum process length, and mean outgrowth intensity (P < 0.05. The differences in damage among drugs reflect differences in their mechanisms of action and clinical CIPN manifestations. We show the potential of the model for gene perturbation studies by demonstrating decreased expression of TUBB2A results in significantly increased sensitivity of neurons to paclitaxel (0.23 ± 0.06 decrease in total neurite outgrowth, P = 0.011. The variance in several neurite outgrowth and apoptotic phenotypes upon treatment with one of the neurotoxic drugs is significantly greater between than within neurons derived from four different individuals (P < 0.05, demonstrating the potential of iPSC-derived neurons as a genetically diverse model for CIPN. The human neuron model will allow both for mechanistic studies of specific genes and genetic variants discovered in clinical studies and for screening of new drugs to prevent or treat CIPN.

  16. Functional cardiotoxicity assessment of cosmetic compounds using human-induced pluripotent stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Chaudhari, Umesh; Nemade, Harshal; Sureshkumar, Poornima; Vinken, Mathieu; Ates, Gamze; Rogiers, Vera; Hescheler, Jürgen; Hengstler, Jan Georg; Sachinidis, Agapios

    2018-01-01

    There is a large demand of a human relevant in vitro test system suitable for assessing the cardiotoxic potential of cosmetic ingredients and other chemicals. Using human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), we have already established an in vitro cardiotoxicity assay and identified genomic biomarkers of anthracycline-induced cardiotoxicity in our previous work. Here, five cosmetic ingredients were studied by the new hiPSC-CMs test; kojic acid (KJA), triclosan (TS), triclocarban (TCC), 2,7-naphthalenediol (NPT), and basic red 51 (BR51) based on cytotoxicity as well as ATP assays, beating rate, and genomic biomarkers to determine the lowest observed effect concentration (LOEC) and no observed effect concentration (NOEC). The LOEC for beating rate were 400, 10, 3, >400, and 3 µM for KJA, TS, TCC, NPT, and BR51, respectively. The corresponding concentrations for cytotoxicity or ATP depletion were similar, with the exception of TS and TCC, where the cardiomyocyte-beating assay showed positive results at non-cytotoxic concentrations. Functional analysis also showed that the individual compounds caused different effects on hiPSC-CMs. While exposure to KJA, TS, TCC, and BR51 induced significant arrhythmic beating, NPT slightly decreased cell viability, but did not influence beating. Gene expression studies showed that TS and NPT caused down-regulation of cytoskeletal and cardiac ion homeostasis genes. Moreover, TS and NPT deregulated genomic biomarkers known to be affected also by anthracyclines. The present study demonstrates that hiPSC-CMs can be used to determine LOECs and NOECs in vitro, which can be compared to human blood concentrations to determine margins of exposure. Our in vitro assay, which so far has been tested with several anthracyclines and cosmetics, still requires validation by larger numbers of positive and negative controls, before it can be recommended for routine analysis.

  17. Targeting eradication of malignant cells derived from human bone marrow mesenchymal stromal cells

    International Nuclear Information System (INIS)

    Yang, Yingbin; Cai, Shaoxi; Yang, Li; Yu, Shuhui; Jiang, Jiahuan; Yan, Xiaoqing; Zhang, Haoxing; Liu, Lan; Liu, Qun; Du, Jun; Cai, Shaohui; Sung, K.L. Paul

    2010-01-01

    Human bone marrow mesenchymal stromal cells (hBMSC) have been shown to participate in malignant transformation. However, hampered by the low frequency of malignant transformation of hBMSC, we do not yet know how to prevent malignant transformation of implanted hBMSC. In this study, in order to establish a model for the eradication of hBMSC-derived malignant cells, a gene fusion consisting of a human telomerase (hTERT) promoter modified with both c-Myc and myeloid zinc finger protein2 (MZF-2) binding elements and followed by the E. coli cytosine deaminase (CD) and luciferase genes was stably transferred into hBMSC via lentiviral transduction; n-phosphonacelyl-L-aspartic acid (PALA) selection was used to generate malignant cell colonies derived from transduced hBMSC after treatment with the carcinogenic reagent BPDE. Cells that were amplified after PALA selection were used for transplantation and 5-FC pro-drug cytotoxicity tests. The results showed that PALA-resistant malignant cells could be generated from hBMSC co-induced with lentiviral transduction and treatment with Benzo(a)pyrene Diol Epoxide (BPDE); the modification of c-Myc and MZF-2 binding elements could remarkably enhance the transcriptional activities of the hTERT promoter in malignant cells, whereas transcriptional activity was depressed in normal hBMSC; malignant cells stably expressing CD under the control of the modified hTERT promoter could be eliminated by 5-FC administration. This study has provided a method for targeted eradication of malignant cells derived from hBMSC.

  18. Proarrhythmia risk prediction using human induced pluripotent stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Yamazaki, Daiju; Kitaguchi, Takashi; Ishimura, Masakazu; Taniguchi, Tomohiko; Yamanishi, Atsuhiro; Saji, Daisuke; Takahashi, Etsushi; Oguchi, Masao; Moriyama, Yuta; Maeda, Sanae; Miyamoto, Kaori; Morimura, Kaoru; Ohnaka, Hiroki; Tashibu, Hiroyuki; Sekino, Yuko; Miyamoto, Norimasa; Kanda, Yasunari

    2018-04-01

    Human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) are expected to become a useful tool for proarrhythmia risk prediction in the non-clinical drug development phase. Several features including electrophysiological properties, ion channel expression profile and drug responses were investigated using commercially available hiPSC-CMs, such as iCell-CMs and Cor.4U-CMs. Although drug-induced arrhythmia has been extensively examined by microelectrode array (MEA) assays in iCell-CMs, it has not been fully understood an availability of Cor.4U-CMs for proarrhythmia risk. Here, we evaluated the predictivity of proarrhythmia risk using Cor.4U-CMs. MEA assay revealed linear regression between inter-spike interval and field potential duration (FPD). The hERG inhibitor E-4031 induced reverse-use dependent FPD prolongation. We next evaluated the proarrhythmia risk prediction by a two-dimensional map, which we have previously proposed. We determined the relative torsade de pointes risk score, based on the extent of FPD with Fridericia's correction (FPDcF) change and early afterdepolarization occurrence, and calculated the margins normalized to free effective therapeutic plasma concentrations. The drugs were classified into three risk groups using the two-dimensional map. This risk-categorization system showed high concordance with the torsadogenic information obtained by a public database CredibleMeds. Taken together, these results indicate that Cor.4U-CMs can be used for drug-induced proarrhythmia risk prediction. Copyright © 2018 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

  19. Knockout of endothelial cell-derived endothelin-1 attenuates skin fibrosis but accelerates cutaneous wound healing.

    Directory of Open Access Journals (Sweden)

    Katsunari Makino

    Full Text Available Endothelin (ET-1 is known for the most potent vasoconstrictive peptide that is released mainly from endothelial cells. Several studies have reported ET-1 signaling is involved in the process of wound healing or fibrosis as well as vasodilation. However, little is known about the role of ET-1 in these processes. To clarify its mechanism, we compared skin fibrogenesis and wound repair between vascular endothelial cell-specific ET-1 knockout mice and their wild-type littermates. Bleomycin-injected fibrotic skin of the knockout mice showed significantly decreased skin thickness and collagen content compared to that of wild-type mice, indicating that bleomycin-induced skin fibrosis is attenuated in the knockout mice. The mRNA levels of transforming growth factor (TGF-β were decreased in the bleomycin-treated skin of ET-1 knockout mice. On the other hand, skin wound healing was accelerated in ET-1 knockout mice, which was indicated by earlier granulation tissue reduction and re-epithelialization in these mice. The mRNA levels of TGF-β, tumor necrosis factor (TNF-α and connective tissue growth factor (CTGF were reduced in the wound of ET-1 knockout mice. In endothelial ET-1 knockout mouse, the expression of TNF-α, CTGF and TGF-β was down-regulated. Bosentan, an antagonist of dual ET receptors, is known to attenuate skin fibrosis and accelerate wound healing in systemic sclerosis, and such contradictory effect may be mediated by above molecules. The endothelial cell-derived ET-1 is the potent therapeutic target in fibrosis or wound healing, and investigations of the overall regulatory mechanisms of these pathological conditions by ET-1 may lead to a new therapeutic approach.

  20. Usefulness of cardiotoxicity assessment using calcium transient in human induced pluripotent stem cell-derived cardiomyocytes.

    Science.gov (United States)

    Watanabe, Hitoshi; Honda, Yayoi; Deguchi, Jiro; Yamada, Toru; Bando, Kiyoko

    2017-01-01

    Monitoring dramatic changes in intracellular calcium ion levels during cardiac contraction and relaxation, known as calcium transient, in human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) would be an attractive strategy for assessing compounds on cardiac contractility. In addition, as arrhythmogenic compounds are known to induce characteristic waveform changes in hiPSC-CMs, it is expected that calcium transient would allow evaluation of not only compound-induced effects on cardiac contractility, but also compound arrhythmogenic potential. Using a combination of calcium transient in hiPSC-CMs and a fast kinetic fluorescence imaging detection system, we examined in this study changes in calcium transient waveforms induced by a series of 17 compounds that include positive/negative inotropic agents as well as cardiac ion channel activators/inhibitors. We found that all positive inotropic compounds induced an increase in peak frequency and/or peak amplitude. The effects of a negative inotropic compound could clearly be detected in the presence of a β-adrenergic receptor agonist. Furthermore, most arrhythmogenic compounds raised the ratio of peak decay time to peak rise time (D/R ratio) in calcium transient waveforms. Compound concentrations at which these parameters exceeded cutoff values correlated well with systemic exposure levels at which arrhythmias were reported to be evoked. In conclusion, we believe that peak analysis of calcium transient and determination of D/R ratio are reliable methods for assessing compounds' cardiac contractility and arrhythmogenic potential, respectively. Using these approaches would allow selection of compounds with low cardiotoxic potential at the early stage of drug discovery.

  1. Targeting eradication of malignant cells derived from human bone marrow mesenchymal stromal cells

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Yingbin [Key Laboratory of Biorheological Science and Technology, Ministry of Education, Bioengineering College, Chongqing University, Chongqing 400044 (China); School of Life Science, Southwest University, Chongqing 400715 (China); Cai, Shaoxi, E-mail: sxcai@cqu.edu.cn [Key Laboratory of Biorheological Science and Technology, Ministry of Education, Bioengineering College, Chongqing University, Chongqing 400044 (China); Yang, Li [Key Laboratory of Biorheological Science and Technology, Ministry of Education, Bioengineering College, Chongqing University, Chongqing 400044 (China); College of Pharmacy, Jinan University, Guangzhou 510632 (China); Yu, Shuhui [Key Laboratory of Biorheological Science and Technology, Ministry of Education, Bioengineering College, Chongqing University, Chongqing 400044 (China); Library of Southwest University, Chongqing 400715 (China); Jiang, Jiahuan; Yan, Xiaoqing [Key Laboratory of Biorheological Science and Technology, Ministry of Education, Bioengineering College, Chongqing University, Chongqing 400044 (China); Zhang, Haoxing [School of Life Science, Southwest University, Chongqing 400715 (China); Liu, Lan [Department of Laboratory of Medicine, Children' s Hospital of Chongqin Medical University, Chongqing 400014 (China); Liu, Qun [College of Life Science and Technology, Southwest University for Nationalities, Chengdu 610041 (China); Du, Jun [Center of Microbiology, Biochemistry, and Pharmacology, School of Pharmaceutical Science, Sun Yat-Sen University, Guangzhou 510080 (China); Cai, Shaohui [College of Pharmacy, Jinan University, Guangzhou 510632 (China); Sung, K.L. Paul [Key Laboratory of Biorheological Science and Technology, Ministry of Education, Bioengineering College, Chongqing University, Chongqing 400044 (China); Departments of Orthopaedic Surgery and Bioengineering, University of California, SD 0412 (United States)

    2010-12-10

    Human bone marrow mesenchymal stromal cells (hBMSC) have been shown to participate in malignant transformation. However, hampered by the low frequency of malignant transformation of hBMSC, we do not yet know how to prevent malignant transformation of implanted hBMSC. In this study, in order to establish a model for the eradication of hBMSC-derived malignant cells, a gene fusion consisting of a human telomerase (hTERT) promoter modified with both c-Myc and myeloid zinc finger protein2 (MZF-2) binding elements and followed by the E. coli cytosine deaminase (CD) and luciferase genes was stably transferred into hBMSC via lentiviral transduction; n-phosphonacelyl-L-aspartic acid (PALA) selection was used to generate malignant cell colonies derived from transduced hBMSC after treatment with the carcinogenic reagent BPDE. Cells that were amplified after PALA selection were used for transplantation and 5-FC pro-drug cytotoxicity tests. The results showed that PALA-resistant malignant cells could be generated from hBMSC co-induced with lentiviral transduction and treatment with Benzo(a)pyrene Diol Epoxide (BPDE); the modification of c-Myc and MZF-2 binding elements could remarkably enhance the transcriptional activities of the hTERT promoter in malignant cells, whereas transcriptional activity was depressed in normal hBMSC; malignant cells stably expressing CD under the control of the modified hTERT promoter could be eliminated by 5-FC administration. This study has provided a method for targeted eradication of malignant cells derived from hBMSC.

  2. Profiling ethanol-targeted transcription factors in human carcinoma cell-derived embryoid bodies.

    Science.gov (United States)

    Mandal, Chanchal; Halder, Debasish; Chai, Jin Choul; Lee, Young Seek; Jung, Kyoung Hwa; Chai, Young Gyu

    2016-01-15

    Fetal alcohol spectrum disorder is a collective term that represents fetal abnormalities associated with maternal alcohol consumption. Prenatal alcohol exposure and related anomalies are well characterized, but the molecular mechanism behind this phenomenon is not yet understood. Few insights have been gained from genetic and epigenetic studies of fetal alcohol spectrum disorder. Our aim was to profile the important molecular regulators of ethanol-related alterations of the genome. For this purpose, we have analyzed the gene expression pattern of human carcinoma cell-derived embryoid bodies in the absence or presence of ethanol. A cDNA microarray analysis was used to profile mRNA expression in embryoid bodies at day 7 with or without ethanol treatment. A total of 493 differentially expressed genes were identified in response to 50 mM ethanol exposure. Of these, 111 genes were up-regulated, and 382 were down-regulated. Gene ontology term enrichment analysis revealed that these genes are involved in important biological processes: neurological system processes, cognition, behavior, sensory perception of smell, taste and chemical stimuli and synaptic transmission. Similarly, the enrichment of disease-related genes included relevant categories such as neurological diseases, developmental disorders, skeletal and muscular disorders, and connective tissue disorders. Furthermore, we have identified a group of 26 genes that encode transcription factors. We validated the relative gene expression of several transcription factors using quantitative real time PCR. We hope that our study substantially contributes to the understanding of the molecular mechanisms underlying the pathology of alcohol-mediated anomalies and facilitates further research. Copyright © 2015 Elsevier B.V. All rights reserved.

  3. Liver cell-derived microparticles activate hedgehog signaling and alter gene expression in hepatic endothelial cells.

    Science.gov (United States)

    Witek, Rafal P; Yang, Liu; Liu, Renshui; Jung, Youngmi; Omenetti, Alessia; Syn, Wing-Kin; Choi, Steve S; Cheong, Yeiwon; Fearing, Caitlin M; Agboola, Kolade M; Chen, Wei; Diehl, Anna Mae

    2009-01-01

    Angiogenesis contributes to vascular remodeling during cirrhosis. In cirrhotic livers, cholangiocytes, and myofibroblastic hepatic stellate cells (MF-HSC) produce Hedgehog (Hh) ligands. During embryogenesis Hh ligands are released from ligand-producing cells in microparticles and activate Hh signaling in endothelial cells. We studied whether adult liver cell-derived microparticles contain Hh ligands that alter hepatic sinusoidal endothelial cells (SEC). MF-HSC and cholangiocytes were exposed to platelet-derived growth factor to induce Hh ligands; microparticles were isolated from medium, analyzed by transmission electron microscopy and immunoblots, and applied to Hh-reporter-containing cells. Microparticles were obtained from serum and bile of rats after bile duct ligation (BDL) or sham surgery and applied to normal primary liver SEC with or without cyclopamine, an Hh signaling inhibitor. Effects on SEC gene expression were evaluated by quantitative reverse-transcription polymerase chain reaction and immunoblotting. Hh target gene expression and SEC activation markers were compared in primary SEC and in liver sections from healthy and BDL rats. Platelet-derived growth factor-treated MF-HSC and cholangiocytes released exosome-enriched microparticles containing biologically-active Hh ligands. BDL increased release of Hh-containing exosome-enriched microparticles into plasma and bile. Transmission electron microscopy and immunoblots revealed similarities among microparticles from all sources; all microparticles induced similar Hh-dependent changes in SEC gene expression. SEC from healthy livers did not express Hh target genes or activation markers, but both were up-regulated in SEC after BDL. Hh-containing exosome-enriched microparticles released from liver cells alter hepatic SEC gene expression, suggesting a novel mechanism for cirrhotic vasculopathy.

  4. Apheresis platelet concentrates contain platelet-derived and endothelial cell-derived microparticles.

    Science.gov (United States)

    Rank, A; Nieuwland, R; Liebhardt, S; Iberer, M; Grützner, S; Toth, B; Pihusch, R

    2011-02-01

    Microparticles (MP) are membrane vesicles with thrombogenic and immunomodulatory properties. We determined MP subgroups from resting platelets, activated platelets and endothelial cells in donors and apheresis platelet concentrates (PC). MP were double stained with annexin V and CD61 (platelet-derived MP; PMP), P-selectin or CD63 (MP from activated platelets) and CD144 plus E-selectin (endothelial cell-derived MP; EMP) and detected by flow cytometry in platelet donors (n=36) and apheresis PC (n=11; Trima™). PC contained MP, mainly from resting platelets [93% (90-95)], and minor fractions of PMP from activated platelets [P-selectin(+) or CD63(+); 4·8% (3·2-7·7) and 2·6% (2·0-4·0)]. Compared to donors, levels of annexin V+ MP, PMP, P-selectin(+) and CD63(+) MP were 1·7-, 2·3-, 8·6- and 3·1-fold higher in PC (all P<0·05). During storage (1-5 days), levels of annexin V+ MP and PMP did not increase, although small increases in the fraction of P-selectin(+) or CD63(+) MP occurred (both P<0·05). PC also contained EMP, which were 2·6- to 3·7-fold enriched in PC compared to donors (P<0·05). Transfusion of apheresis PC also results in transfusion of HLA-carrying PMP and EMP. This might counteract the aim of reducing transfused HLA load by leucodepletion. The increases in PMP exposing P-selectin or CD63 reflect mild platelet activation during storage. We conclude that in leucodepleted platelet apheresis using fluidized particle bed technology, MP are harvested mainly from the donor by apheresis. Improvement in apheresis technology might reduce MP load. © 2010 The Author(s). Vox Sanguinis © 2010 International Society of Blood Transfusion.

  5. A novel broadband impedance method for detection of cell-derived microparticles.

    Science.gov (United States)

    Lvovich, Vadim; Srikanthan, Sowmya; Silverstein, Roy L

    2010-10-15

    A novel label-free method is presented to detect and quantify cell-derived microparticles (MPs) by the electrochemical potential-modulated electrochemical impedance spectroscopy (EIS). MPs are present in elevated concentrations during pathological conditions and play a major role in the establishment and pathogenesis of many diseases. Considering this, accurate detection and quantification of MPs is very important in clinical diagnostics and therapeutics. A combination of bulk solution electrokinetic sorting and interfacial impedance responses allows achieving detection limits as low as several MPs per μL. By fitting resulting EIS spectra with an equivalent electrical circuit, the bulk solution electrokinetic and interfacial impedance responses were characterized. In the bulk solution two major relaxations were prominent-β-relaxation in low MHz region due to the MP capacitive membrane bridging, and α-relaxation at ∼10 kHz due to counter ions diffusion. At low frequencies (10-0.1 Hz) at electrochemical potentials exceeding -100 mV, a facile interfacial Faradaic process of oxidation in MPs coupled with diffusion and non-Faradaic double layer charging dominate, probably due to oxidation of phospholipids and/or proteins on the MP surface and MP lysis. Buffer influence on the MP detection demonstrated that a relatively low conductivity Tyrode's buffer background solution is preferential for the MP electrokinetic separation and characterization. This study also demonstrated that standard laboratory methods such as flow cytometry underestimate MP concentrations, especially those with smaller average sizes, by as much as a factor of 2-40. Copyright © 2010 Elsevier B.V. All rights reserved.

  6. Predicting carotid artery disease and plaque instability from cell-derived microparticles.

    Science.gov (United States)

    Wekesa, A L; Cross, K S; O'Donovan, O; Dowdall, J F; O'Brien, O; Doyle, M; Byrne, L; Phelan, J P; Ross, M D; Landers, R; Harrison, M

    2014-11-01

    Cell-derived microparticles (MPs) are small plasma membrane-derived vesicles shed from circulating blood cells and may act as novel biomarkers of vascular disease. We investigated the potential of circulating MPs to predict (a) carotid plaque instability and (b) the presence of advanced carotid disease. This pilot study recruited carotid disease patients (aged 69.3 ± 1.2 years [mean ± SD], 69% male, 90% symptomatic) undergoing endarterectomy (n = 42) and age- and sex-matched controls (n = 73). Plaques were classified as stable (n = 25) or unstable (n = 16) post surgery using immunohistochemistry. Blood samples were analysed for MP subsets and molecular biomarkers. Odds ratios (OR) are expressed per standard deviation biomarker increase. Endothelial MP (EMP) subsets, but not any vascular, inflammatory, or proteolytic molecular biomarker, were higher (p < .05) in the unstable than the stable plaque patients. The area under the receiver operator characteristic curve for CD31(+)41(-) EMP in discriminating an unstable plaque was 0.73 (0.56-0.90, p < .05). CD31(+)41(-) EMP predicted plaque instability (OR = 2.19, 1.08-4.46, p < .05) and remained significant in a multivariable model that included transient ischaemic attack symptom status. Annexin V(+) MP, platelet MP (PMP) subsets, and C-reactive protein were higher (p < .05) in cases than controls. Annexin V(+) MP (OR = 3.15, 1.49-6.68), soluble vascular cell adhesion molecule-1 (OR = 1.64, 1.03-2.59), and previous smoking history (OR = 3.82, 1.38-10.60) independently (p < .05) predicted the presence of carotid disease in a multivariable model. EMP may have utility in predicting plaque instability in carotid patients and annexin V(+) MPs may predict the presence of advanced carotid disease in aging populations, independent of established biomarkers. Copyright © 2014 European Society for Vascular Surgery. Published by Elsevier Ltd. All rights reserved.

  7. Measurement of circulating cell-derived microparticles by flow cytometry: sources of variability within the assay.

    Science.gov (United States)

    Ayers, Lisa; Kohler, Malcolm; Harrison, Paul; Sargent, Ian; Dragovic, Rebecca; Schaap, Marianne; Nieuwland, Rienk; Brooks, Susan A; Ferry, Berne

    2011-04-01

    Circulating cell-derived microparticles (MPs) have been implicated in several disease processes and elevated levels are found in many pathological conditions. The detection and accurate measurement of MPs, although attracting widespread interest, is hampered by a lack of standardisation. The aim of this study was to establish a reliable flow cytometric assay to measure distinct subtypes of MPs in disease and to identify any significant causes of variability in MP quantification. Circulating MPs within plasma were identified by their phenotype (platelet, endothelial, leukocyte and annexin-V positivity (AnnV+). The influence of key variables (i.e. time between venepuncture and centrifugation, washing steps, the number of centrifugation steps, freezing/long-term storage and temperature of thawing) on MP measurement were investigated. Increasing time between venepuncture and centrifugation leads to increased MP levels. Washing samples results in decreased AnnV+MPs (P=0.002) and platelet-derived MPs (PMPs) (P=0.002). Double centrifugation of MPs prior to freezing decreases numbers of AnnV+MPs (P=0.0004) and PMPs (P=0.0004). A single freeze thaw cycle of samples led to an increase in AnnV+MPs (P=0.0020) and PMPs (P=0.0039). Long-term storage of MP samples at -80° resulted in decreased MP levels. This study found that minor protocol changes significantly affected MP levels. This is one of the first studies attempting to standardise a method for obtaining and measuring circulating MPs. Standardisation will be essential for successful development of MP technologies, allowing direct comparison of results between studies and leading to a greater understanding of MPs in disease. Crown Copyright © 2010. Published by Elsevier Ltd. All rights reserved.

  8. Flow rate calibration to determine cell-derived microparticles and homogeneity of blood components.

    Science.gov (United States)

    Noulsri, Egarit; Lerdwana, Surada; Kittisares, Kulvara; Palasuwan, Attakorn; Palasuwan, Duangdao

    2017-08-01

    Cell-derived microparticles (MPs) are currently of great interest to screening transfusion donors and blood components. However, the current approach to counting MPs is not affordable for routine laboratory use due to its high cost. The current study aimed to investigate the potential use of flow-rate calibration for counting MPs in whole blood, packed red blood cells (PRBCs), and platelet concentrates (PCs). The accuracy of flow-rate calibration was investigated by comparing the platelet counts of an automated counter and a flow-rate calibrator. The concentration of MPs and their origins in whole blood (n=100), PRBCs (n=100), and PCs (n=92) were determined using a FACSCalibur. The MPs' fold-changes were calculated to assess the homogeneity of the blood components. Comparing the platelet counts conducted by automated counting and flow-rate calibration showed an r 2 of 0.6 (y=0.69x+97,620). The CVs of the within-run and between-run variations of flow-rate calibration were 8.2% and 12.1%, respectively. The Bland-Altman plot showed a mean bias of -31,142platelets/μl. MP enumeration revealed both the difference in MP levels and their origins in whole blood, PRBCs, and PCs. Screening the blood components demonstrated high heterogeneity of the MP levels in PCs when compared to whole blood and PRBCs. The results of the present study suggest the accuracy and precision of flow-rate calibration for enumerating MPs. This flow-rate approach is affordable for assessing the homogeneity of MPs in blood components in routine laboratory practice. Copyright © 2017 Elsevier Ltd. All rights reserved.

  9. Red blood cell-derived microparticles isolated from blood units initiate and propagate thrombin generation.

    Science.gov (United States)

    Rubin, Olivier; Delobel, Julien; Prudent, Michel; Lion, Niels; Kohl, Kid; Tucker, Erik I; Tissot, Jean-Daniel; Angelillo-Scherrer, Anne

    2013-08-01

    Red blood cell-derived microparticles (RMPs) are small phospholipid vesicles shed from RBCs in blood units, where they accumulate during storage. Because microparticles are bioactive, it could be suggested that RMPs are mediators of posttransfusion complications or, on the contrary, constitute a potential hemostatic agent. This study was performed to establish the impact on coagulation of RMPs isolated from blood units. Using calibrated automated thrombography, we investigated whether RMPs affect thrombin generation (TG) in plasma. We found that RMPs were not only able to increase TG in plasma in the presence of a low exogenous tissue factor (TF) concentration, but also to initiate TG in plasma in absence of exogenous TF. TG induced by RMPs in the absence of exogenous TF was neither affected by the presence of blocking anti-TF nor by the absence of Factor (F)VII. It was significantly reduced in plasma deficient in FVIII or F IX and abolished in FII-, FV-, FX-, or FXI-deficient plasma. TG was also totally abolished when anti-XI 01A6 was added in the sample. Finally, neither Western blotting, flow cytometry, nor immunogold labeling allowed the detection of traces of TF antigen. In addition, RMPs did not comprise polyphosphate, an important modulator of coagulation. Taken together, our data show that RMPs have FXI-dependent procoagulant properties and are able to initiate and propagate TG. The anionic surface of RMPs might be the site of FXI-mediated TG amplification and intrinsic tenase and prothrombinase complex assembly. © 2012 American Association of Blood Banks.

  10. socio-ec(h)o: Ambient Intelligence and Gameplay

    OpenAIRE

    Wakkary, Ron

    2005-01-01

    The socio-ec(h)o project aims to research a generalized ambient intelligent software platform and design models for responsive environments based on the concept of ambient intelligent "ecologies" and group gameplay. The benefits of the research include a software-architecture, ambient intelligence inference engine, and interaction design models for gameplay and responsive environments. The paper will discuss the results of our prototypes for games in responsive environments. These prototypes ...

  11. Real time control of the sawtooth period using EC launchers

    International Nuclear Information System (INIS)

    Paley, J I; Felici, F; Coda, S; Goodman, T P; Piras, F

    2009-01-01

    Tokamak plasmas operating at high performance are limited by several MHD instabilities. The sawtooth instability limits the core plasma pressure and can drive the neoclassical tearing mode unstable, but also prevents accumulation of impurities in the core. Electron cyclotron heating and current drive systems can be used to modify the local current profile and therefore tailor the sawtooth period. This paper reports on demonstrations of continuous real time feedback control of the sawtooth period by varying the EC injection angle.

  12. Present status of developing petroleum-substituting energy (EC)

    Science.gov (United States)

    1993-03-01

    The EC has had approximately 50% of its total energy demand supplied by imports from the exterior of the Community. Hence, it is getting important to develop oil-substituting renewable energy as well as to curtail the emission of carbon dioxide. In consideration of these situations, the results of investigation on the energy policy of the European Community are described. The policy comprises three courses: European Energy Charter, formation of an open European Community energy market, and environmental conservation. Particularly, concerning the reduction of carbon dioxide emission, the EC Council has decided to introduce carbon dioxide taxation so as to suppress the carbon dioxide emission in the year 2000 to the 1990 level. The arrangement for its introduction, however, encountered with difficulties because of the opposition of various countries other than the European Community and the industrial world of the European Community. Legislation of the investment promotion law for energy saving and the construction of infrastructure footing and an information network are ineffective due to the sluggish fuel price and economic recession. A plurality of EC member countries are advancing a comprehensive activity within the framework of the joint programs of research, development, and demonstration set for the renewable energy policy ensuring energy supply and environmental harmonization.

  13. Hematopoietic stem cell-derived exosomes promote hematopoietic differentiation of mouse embryonic stem cells in vitro via inhibiting the miR126/Notch1 pathway.

    Science.gov (United States)

    Liao, Feng-Ling; Tan, Lin; Liu, Hua; Wang, Jin-Ju; Ma, Xiao-Tang; Zhao, Bin; Chen, Yanfang; Bihl, Ji; Yang, Yi; Chen, Ri-Ling

    2018-04-01

    Cell-derived exosomes (EXs) can modulate target cell differentiation via microRNAs (miRs) that they carried. Previous studies have shown that miR126 is highly expressed in hematopoietic stem cells (HSCs) and plays a role in hematopoiesis via modulating the Notch pathway that participates in progenitors' cell fate decisions. In this study we investigated whether HSC-derived EXs (HSC-EXs) could affect the differentiation of mouse embryonic stem cells (ESCs) into HSCs. We prepared HSC-EXs con , HSC-EXs sc and HSC-EXs miR126 from control HSCs and the HSCs transfected with scramble control or miR126 mimics, respectively. HSC-EXs were isolated by ultracentrifugation and analyzed using nanoparticle tracking analysis. We incubated the collected EXs with mouse ESCs over a 10-d differentiation induction period, during which HSC-EXs and a Notch pathway activator (Jagged1, 100 ng/mL) were added to the cultures every 3 d. After the 10-d differentiation period, the expression levels of miR126, SSEA1, CD117, Sca1, Notch1 and Hes1 in ESCs were assessed. The generated HSCs were validated by flow cytometry using antibodies against HSC markers (CD117, CD34 and Sca1). Our results revealed that: (1) transfection with miR126 mimics significantly increased miR126 levels in HSC-EXs miR126 . (2) HSC-EX co-culture promoted mouse ESCs differentiation into HSCs with the most prominent effect found in the HSC-EXs miR126 co-culture. (3) HSC differentiation was verified by reduced SSEA1 expression and increased CD117 and Sca1 expression. (4) All the effects caused by HSC-EXs were accompanied by significant reduction of Notch1 and Hes1 expression, thus inhibition of the Notch1/Hes1 pathway, whereas activation of Notch by Jagged1 abolished the effects of HSC-EXs miR126 . In conclusion, HSC-EXs promote hematopoietic differentiation of mouse ESCs in vitro by inhibiting the miR126/Notch1 pathway.

  14. Human Breast Adipose-Derived Stem Cells Transfected with the Stromal Cell-Derived Factor-1 Receptor CXCR4 Exhibit Enhanced Viability in Human Autologous Free Fat Grafts

    Directory of Open Access Journals (Sweden)

    Fang-tian Xu

    2014-11-01

    Full Text Available Background: The main complication of autologous free fat tissue transplantation is fat resorption and calcification due to the ischemic necrosis of fat. The promotion of transplant neovascularization soon after autologous free fat grafts may reduce these outcomes. In adulthood, stromal cell-derived factor-1 (SDF-1 and its membrane receptor C-X-C chemokine receptor type 4 (CXCR4 are involved in the homing and migration of multiple stem cell types, neovascularization, and cell proliferation. We hypothesized that CXCR4 may improve the long-term survival of free fat tissue transplants by recruiting endothelial progenitor cells (EPCs and may therefore improve graft revascularization. In this study, we aimed to determine the effect of human breast adipose-derived stem cells (HBASCs transfected with the CXCR4 gene on the survival rate of human autologous free fat transplants in nude mice. Methods: Human breast adipose-derived stem cells (HBASCs were expanded ex vivo for 3 passages, labeled with green fluorescent protein (GFP and transfected with CXCR4 or left untransfected. Autologous fat tissues were mixed with the GFP-labeled, CXCR4-transfected HBASCs (group A, GFP-labeled HBASCs (group B, the known vascularization-promoting agent VEGF (group C, or medium (group D and then injected subcutaneously into 32 nude mice at 4 spots in a random fashion. Six months later, the transplanted tissue volume and histology were evaluated, and neo-vascularization was quantified by counting the capillaries. CXCR4 and SDF-1α mRNA expression in the transplants was determined using real-time quantitative PCR analysis (qPCR. Results: The data revealed that the control (group D transplant volume survival was 28.3 ± 4.5%. Mixing CXCR4-transfected (group A and untransfected (group B HBASCs significantly increased transplant volume survival (79.5 ± 8.3% and 67.2 ± 5.9%, respectively, whereas VEGF-transfected HBASCs (group C were less effective (41.2 ± 5.1%. Histological

  15. Business-to-business electronic commerce systems and services. Smart EC solution; Kigyoka nrenkei system solution system. Smart EC solution

    Energy Technology Data Exchange (ETDEWEB)

    Setoguchi, T.; Manchu, Y.; Katsumata, M. [Toshiba Corp., Tokyo (Japan)

    2000-04-01

    Toshiba provides a range of information technology (IT) solutions called SmartEC Solution, which includes business-to-business electronic commerce systems and services based on international standards and industrial know-how, especially our electronic data interchange (EDI) know-how as a manufacturer. These IT solutions are supplied as services covering strategy planning, system integration, and application service provider based on five types of business-to-business electronic commerce. (author)

  16. Acellular therapeutic approach for heart failure: in vitro production of extracellular vesicles from human cardiovascular progenitors.

    Science.gov (United States)

    El Harane, Nadia; Kervadec, Anaïs; Bellamy, Valérie; Pidial, Laetitia; Neametalla, Hany J; Perier, Marie-Cécile; Lima Correa, Bruna; Thiébault, Léa; Cagnard, Nicolas; Duché, Angéline; Brunaud, Camille; Lemitre, Mathilde; Gauthier, Jeanne; Bourdillon, Alexandra T; Renault, Marc P; Hovhannisyan, Yeranuhi; Paiva, Solenne; Colas, Alexandre R; Agbulut, Onnik; Hagège, Albert; Silvestre, Jean-Sébastien; Menasché, Philippe; Renault, Nisa K E

    2018-05-21

    We have shown that extracellular vesicles (EVs) secreted by embryonic stem cell-derived cardiovascular progenitor cells (Pg) recapitulate the therapeutic effects of their parent cells in a mouse model of chronic heart failure (CHF). Our objectives are to investigate whether EV released by more readily available cell sources are therapeutic, whether their effectiveness is influenced by the differentiation state of the secreting cell, and through which mechanisms they act. The total EV secreted by human induced pluripotent stem cell-derived cardiovascular progenitors (iPSC-Pg) and human induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) were isolated by ultracentrifugation and characterized by Nanoparticle Tracking Analysis, western blot, and cryo-electron microscopy. In vitro bioactivity assays were used to evaluate their cellular effects. Cell and EV microRNA (miRNA) content were assessed by miRNA array. Myocardial infarction was induced in 199 nude mice. Three weeks later, mice with left ventricular ejection fraction (LVEF) ≤ 45% received transcutaneous echo-guided injections of iPSC-CM (1.4 × 106, n = 19), iPSC-Pg (1.4 × 106, n = 17), total EV secreted by 1.4 × 106 iPSC-Pg (n = 19), or phosphate-buffered saline (control, n = 17) into the peri-infarct myocardium. Seven weeks later, hearts were evaluated by echocardiography, histology, and gene expression profiling, blinded to treatment group. In vitro, EV were internalized by target cells, increased cell survival, cell proliferation, and endothelial cell migration in a dose-dependent manner and stimulated tube formation. Extracellular vesicles were rich in miRNAs and most of the 16 highly abundant, evolutionarily conserved miRNAs are associated with tissue-repair pathways. In vivo, EV outperformed cell injections, significantly improving cardiac function through decreased left ventricular volumes (left ventricular end systolic volume: -11%, P < 0.001; left

  17. Generation of Functional Lentoid Bodies From Human Induced Pluripotent Stem Cells Derived From Urinary Cells.

    Science.gov (United States)

    Fu, Qiuli; Qin, Zhenwei; Jin, Xiuming; Zhang, Lifang; Chen, Zhijian; He, Jiliang; Ji, Junfeng; Yao, Ke

    2017-01-01

    The pathological mechanisms underlying cataract formation remain largely unknown on account of the lack of appropriate in vitro cellular models. The aim of this study is to develop a stable in vitro system for human lens regeneration using pluripotent stem cells. Isolated human urinary cells were infected with four Yamanaka factors to generate urinary human induced pluripotent stem cells (UiPSCs), which were induced to differentiate into lens progenitor cells and lentoid bodies (LBs). The expression of lens-specific markers was examined by real-time PCR, immunostaining, and Western blotting. The structure and magnifying ability of LBs were investigated using transmission electron microscopy and observing the magnification of the letter "X," respectively. We developed a "fried egg" differentiation method to generate functional LBs from UiPSCs. The UiPSC-derived LBs exhibited crystalline lens-like morphology and a transparent structure and expressed lens-specific markers αA-, αB-, β-, and γ-crystallin and MIP. During LB differentiation, the placodal markers SIX1, EYA1, DLX3, PAX6, and the specific early lens markers SOX1, PROX1, FOXE3, αA-, and αB-crystallin were observed at certain time points. Microscopic examination revealed the presence of lens epithelial cells adjacent to the lens capsule as well as both immature and mature fiber-like cells. Optical analysis further demonstrated the magnifying ability (1.7×) of the LBs generated from UiPSCs. Our study provides the first evidence toward generating functional LBs from UiPSCs, thereby establishing an in vitro system that can be used to study human lens development and cataractogenesis and perhaps even be useful for drug screening.

  18. The skeletal cell-derived molecule sclerostin drives bone marrow adipogenesis.

    Science.gov (United States)

    Fairfield, Heather; Falank, Carolyne; Harris, Elizabeth; Demambro, Victoria; McDonald, Michelle; Pettitt, Jessica A; Mohanty, Sindhu T; Croucher, Peter; Kramer, Ina; Kneissel, Michaela; Rosen, Clifford J; Reagan, Michaela R

    2018-02-01

    The bone marrow niche is a dynamic and complex microenvironment that can both regulate, and be regulated by the bone matrix. Within the bone marrow (BM), mesenchymal stromal cell (MSC) precursors reside in a multi-potent state and retain the capacity to differentiate down osteoblastic, adipogenic, or chondrogenic lineages in response to numerous biochemical cues. These signals can be altered in various pathological states including, but not limited to, osteoporotic-induced fracture, systemic adiposity, and the presence of bone-homing cancers. Herein we provide evidence that signals from the bone matrix (osteocytes) determine marrow adiposity by regulating adipogenesis in the bone marrow. Specifically, we found that physiologically relevant levels of Sclerostin (SOST), which is a Wnt-inhibitory molecule secreted from bone matrix-embedded osteocytes, can induce adipogenesis in 3T3-L1 cells, mouse ear- and BM-derived MSCs, and human BM-derived MSCs. We demonstrate that the mechanism of SOST induction of adipogenesis is through inhibition of Wnt signaling in pre-adipocytes. We also demonstrate that a decrease of sclerostin in vivo, via both genetic and pharmaceutical methods, significantly decreases bone marrow adipose tissue (BMAT) formation. Overall, this work demonstrates a direct role for SOST in regulating fate determination of BM-adipocyte progenitors. This provides a novel mechanism for which BMAT is governed by the local bone microenvironment, which may prove relevant in the pathogenesis of certain diseases involving marrow adipose. Importantly, with anti-sclerostin therapy at the forefront of osteoporosis treatment and a greater recognition of the role of BMAT in disease, these data are likely to have important clinical implications. © 2017 Wiley Periodicals, Inc.

  19. Molecular and in vivo characterization of cancer-propagating cells derived from MYCN-dependent medulloblastoma.

    Directory of Open Access Journals (Sweden)

    Zai Ahmad

    Full Text Available Medulloblastoma (MB is the most common malignant pediatric brain tumor. While the pathways that are deregulated in MB remain to be fully characterized, amplification and/or overexpression of the MYCN gene, which is has a critical role in cerebellar development as a regulator of neural progenitor cell fate, has been identified in several MB subgroups. Phenotypically, aberrant expression of MYCN is associated with the large-cell/anaplastic MB variant, which accounts for 5-15% of cases and is associated with aggressive disease and poor clinical outcome. To better understand the role of MYCN in MB in vitro and in vivo and to aid the development of MYCN-targeted therapeutics we established tumor-derived neurosphere cell lines from the GTML (Glt1-tTA/TRE-MYCN-Luc genetically engineered mouse model. A fraction of GTML neurospheres were found to be growth factor independent, expressed CD133 (a marker of neural stem cells, failed to differentiate upon MYCN withdrawal and were highly tumorigenic when orthotopically implanted into the cerebellum. Principal component analyzes using single cell RNA assay data suggested that the clinical candidate aurora-A kinase inhibitor MLN8237 converts GTML neurospheres to resemble non-MYCN expressors. Correlating with this, MLN8237 significantly extended the survival of mice bearing GTML MB allografts. In summary, our results demonstrate that MYCN plays a critical role in expansion and survival of aggressive MB-propagating cells, and establish GTML neurospheres as an important resource for the development of novel therapeutic strategies.

  20. IAP-Based Cell Sorting Results in Homogeneous Transplantable Dopaminergic Precursor Cells Derived from Human Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Daniela Lehnen

    2017-10-01

    Full Text Available Human pluripotent stem cell (hPSC-derived mesencephalic dopaminergic (mesDA neurons can relieve motor deficits in animal models of Parkinson's disease (PD. Clinical translation of differentiation protocols requires standardization of production procedures, and surface-marker-based cell sorting is considered instrumental for reproducible generation of defined cell products. Here, we demonstrate that integrin-associated protein (IAP is a cell surface marker suitable for enrichment of hPSC-derived mesDA progenitor cells. Immunomagnetically sorted IAP+ mesDA progenitors showed increased expression of ventral midbrain floor plate markers, lacked expression of pluripotency markers, and differentiated into mature dopaminergic (DA neurons in vitro. Intrastriatal transplantation of IAP+ cells sorted at day 16 of differentiation in a rat model of PD resulted in functional recovery. Grafts from sorted IAP+ mesDA progenitors were more homogeneous in size and DA neuron density. Thus, we suggest IAP-based sorting for reproducible prospective enrichment of mesDA progenitor cells in clinical cell replacement strategies.

  1. IAP-Based Cell Sorting Results in Homogeneous Transplantable Dopaminergic Precursor Cells Derived from Human Pluripotent Stem Cells.

    Science.gov (United States)

    Lehnen, Daniela; Barral, Serena; Cardoso, Tiago; Grealish, Shane; Heuer, Andreas; Smiyakin, Andrej; Kirkeby, Agnete; Kollet, Jutta; Cremer, Harold; Parmar, Malin; Bosio, Andreas; Knöbel, Sebastian

    2017-10-10

    Human pluripotent stem cell (hPSC)-derived mesencephalic dopaminergic (mesDA) neurons can relieve motor deficits in animal models of Parkinson's disease (PD). Clinical translation of differentiation protocols requires standardization of production procedures, and surface-marker-based cell sorting is considered instrumental for reproducible generation of defined cell products. Here, we demonstrate that integrin-associated protein (IAP) is a cell surface marker suitable for enrichment of hPSC-derived mesDA progenitor cells. Immunomagnetically sorted IAP + mesDA progenitors showed increased expression of ventral midbrain floor plate markers, lacked expression of pluripotency markers, and differentiated into mature dopaminergic (DA) neurons in vitro. Intrastriatal transplantation of IAP + cells sorted at day 16 of differentiation in a rat model of PD resulted in functional recovery. Grafts from sorted IAP + mesDA progenitors were more homogeneous in size and DA neuron density. Thus, we suggest IAP-based sorting for reproducible prospective enrichment of mesDA progenitor cells in clinical cell replacement strategies. Copyright © 2017 Miltenyi Biotec GmbH. Published by Elsevier Inc. All rights reserved.

  2. The influence of gender- and age-related differences in the radiosensitivity of hematopoietic progenitor cells detected in steady-state human peripheral blood

    International Nuclear Information System (INIS)

    Kato, Kengo; Kashiwakura, Ikuo; Kuwabara, Mikinori

    2011-01-01

    To investigate the importance of gender and aging on the individual radiosensitivity of lineage-committed myeloid hematopoietic stem/progenitor cells (HSPCs) detected in mononuclear cells (MNCs) of steady-state human peripheral blood (PB), the clonogenic survival of HPCs, including colony-forming unit-granulocyte macrophage; burst-forming unit-erythroid; colony-forming unit-granulocyte-erythroid-macrophage-megakaryocyte cells derived from MNCs exposed to 0.5 Gy and 2 Gy X-irradiation were estimated. MNCs were prepared from the buffy-coats of 59 healthy individual blood donors. The results showed that large individual differences exist in the number of HSPCs, as well as in the surviving fraction of cells. Furthermore, the number of progenitor cells strongly correlated with their surviving fraction, suggesting that the radiosensitivity of hematopoietic progenitor cells decreases with the number of cells in the 10 5 cells population. A statistically significant negative correlation was observed between the surviving fraction observed at a dose of 0.5 Gy and the age of an individual, however, none of these correlations were observed after 2 Gy irradiation. No statistically significant difference was observed in individual radiosensitivity between males and females at either radiation dose. The present results indicated a correlation between the individual responsiveness of HSPCs to ionizing irradiation, especially to low dose irradiation, and aging. (author)

  3. Spatially defined InsP3-mediated signaling in embryonic stem cell-derived cardiomyocytes.

    Directory of Open Access Journals (Sweden)

    Nidhi Kapoor

    Full Text Available The functional role of inositol 1,4,5-trisphosphate (InsP3 signaling in cardiomyocytes is not entirely understood but it was linked to an increased propensity for triggered activity. The aim of this study was to determine how InsP3 receptors can translate Ca(2+ release into a depolarization of the plasma membrane and consequently arrhythmic activity. We used embryonic stem cell-derived cardiomyocytes (ESdCs as a model system since their spontaneous electrical activity depends on InsP3-mediated Ca(2+ release. [InsP3]i was monitored with the FRET-based InsP3-biosensor FIRE-1 (Fluorescent InsP3 Responsive Element and heterogeneity in sub-cellular [InsP3]i was achieved by targeted expression of FIRE-1 in the nucleus (FIRE-1nuc or expression of InsP3 5-phosphatase (m43 localized to the plasma membrane. Spontaneous activity of ESdCs was monitored simultaneously as cytosolic Ca(2+ transients (Fluo-4/AM and action potentials (current clamp. During diastole, the diastolic depolarization was paralleled by an increase of [Ca(2+]i and spontaneous activity was modulated by [InsP3]i. A 3.7% and 1.7% increase of FIRE-1 FRET ratio and 3.0 and 1.5 fold increase in beating frequency was recorded upon stimulation with endothelin-1 (ET-1, 100 nmol/L or phenylephrine (PE, 10 µmol/L, respectively. Buffering of InsP3 by FIRE-1nuc had no effect on the basal frequency while attenuation of InsP3 signaling throughout the cell (FIRE-1, or at the plasma membrane (m43 resulted in a 53.7% and 54.0% decrease in beating frequency. In m43 expressing cells the response to ET-1 was completely suppressed. Ca(2+ released from InsP3Rs is more effective than Ca(2+ released from RyRs to enhance INCX. The results support the hypothesis that in ESdCs InsP3Rs form a functional signaling domain with NCX that translates Ca(2+ release efficiently into a depolarization of the membrane potential.

  4. Evaluation of nefazodone-induced cardiotoxicity in human induced pluripotent stem cell-derived cardiomyocytes

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Sujeong, E-mail: crystalee@gmail.com [Next-generation Pharmaceutical Research Center, Korea Institute of Toxicology, Korea Research Institute of Chemical Technology, 141 Gajeong-ro, Yuseong-gu, Daejeon 305-343 (Korea, Republic of); Lee, Hyang-Ae, E-mail: hyangaelee@gmail.com [Next-generation Pharmaceutical Research Center, Korea Institute of Toxicology, Korea Research Institute of Chemical Technology, 141 Gajeong-ro, Yuseong-gu, Daejeon 305-343 (Korea, Republic of); Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, 110-799 (Korea, Republic of); Human and Environmental Toxicology Program, University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon 305-350 (Korea, Republic of); Choi, Sung Woo, E-mail: djmaya@snu.ac.kr [Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, 110-799 (Korea, Republic of); Kim, Sung Joon, E-mail: sjoonkim@snu.ac.kr [Department of Biomedical Sciences, Seoul National University College of Medicine, Seoul, 110-799 (Korea, Republic of); Kim, Ki-Suk, E-mail: idkks00@gmail.com [Next-generation Pharmaceutical Research Center, Korea Institute of Toxicology, Korea Research Institute of Chemical Technology, 141 Gajeong-ro, Yuseong-gu, Daejeon 305-343 (Korea, Republic of); Human and Environmental Toxicology Program, University of Science and Technology, 217 Gajeong-ro, Yuseong-gu, Daejeon 305-350 (Korea, Republic of)

    2016-04-01

    The recent establishment of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), which express the major cardiac ion channels and recapitulate spontaneous mechanical and electrical activities, may provide a possible solution for the lack of in vitro human-based cardiotoxicity testing models. Cardiotoxicity induced by the antidepressant nefazodone was previously revealed to cause an acquired QT prolongation by hERG channel blockade. To elucidate the cellular mechanisms underlying the cardiotoxicity of nefazodone beyond hERG, its effects on cardiac action potentials (APs) and ion channels were investigated using hiPSC-CMs with whole-cell patch clamp techniques. In a proof of principle study, we examined the effects of cardioactive channel blockers on the electrophysiological profile of hiPSC-CMs in advance of the evaluation of nefazodone. Nefazodone dose-dependently prolonged the AP duration at 90% (APD{sub 90}) and 50% (APD{sub 50}) repolarization, reduced the maximum upstroke velocity (dV/dt{sub max}) and induced early after depolarizations. Voltage-clamp studies of hiPSC-CMs revealed that nefazodone inhibited various voltage-gated ion channel currents including I{sub Kr}, I{sub Ks}, I{sub Na}, and I{sub Ca}. Among them, I{sub Kr} and I{sub Na} showed relatively higher sensitivity to nefazodone, consistent with the changes in the AP parameters. In summary, hiPSC-CMs enabled an integrated approach to evaluate the complex interactions of nefazodone with cardiac ion channels. These results suggest that hiPSC-CMs can be an effective model for detecting drug-induced arrhythmogenicity beyond the current standard assay of heterologously expressed hERG K{sup +} channels. - Highlights: • Nefazodone prolonged APD and decreased upstroke velocity of APs in hiPSC-CMs. • Nefazodone inhibited cardiac ion channels, especially I{sub Kr} and I{sub Na}, in hiPSC-CMs. • Nefazodone-induced AP changes are mainly the result of I{sub Kr} and I{sub Na} inhibition

  5. Evaluation of Optogenetic Electrophysiology Tools in Human Stem Cell-Derived Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Susann Björk

    2017-11-01

    Full Text Available Current cardiac drug safety assessments focus on hERG channel block and QT prolongation for evaluating arrhythmic risks, whereas the optogenetic approach focuses on the action potential (AP waveform generated by a monolayer of human cardiomyocytes beating synchronously, thus assessing the contribution of several ion channels on the overall drug effect. This novel tool provides arrhythmogenic sensitizing by light-induced pacing in combination with non-invasive, all-optical measurements of cardiomyocyte APs and will improve assessment of drug-induced electrophysiological aberrancies. With the help of patch clamp electrophysiology measurements, we aimed to investigate whether the optogenetic modifications alter human cardiomyocytes' electrophysiology and how well the optogenetic analyses perform against this gold standard. Patch clamp electrophysiology measurements of non-transduced stem cell-derived cardiomyocytes compared to cells expressing the commercially available optogenetic constructs Optopatch and CaViar revealed no significant changes in action potential duration (APD parameters. Thus, inserting the optogenetic constructs into cardiomyocytes does not significantly affect the cardiomyocyte's electrophysiological properties. When comparing the two methods against each other (patch clamp vs. optogenetic imaging we found no significant differences in APD parameters for the Optopatch transduced cells, whereas the CaViar transduced cells exhibited modest increases in APD-values measured with optogenetic imaging. Thus, to broaden the screen, we combined optogenetic measurements of membrane potential and calcium transients with contractile motion measured by video motion tracking. Furthermore, to assess how optogenetic measurements can predict changes in membrane potential, or early afterdepolarizations (EADs, cells were exposed to cumulating doses of E-4031, a hERG potassium channel blocker, and drug effects were measured at both spontaneous and

  6. Lethal and mutagenic effects of radiation and chemicals on cultured fish cells derived the erythrophoroma of goldfish (Carassius auratus)

    Energy Technology Data Exchange (ETDEWEB)

    Mitani, H. (Tokyo Univ. (Japan). Inst. of Zoology)

    1983-01-01

    GEM 199 cells derived from an eryhtrophoroma of goldfish (Carassius auratus), which had a high plating efficiency, were used to investigate the lethal and mutational effects of radiations (UV and ..gamma..-rays) and chemicals (4NQO and MNNG). The cells were more resistant to rays than mammalian cells and CAF-MM1 cells derived from the normal fin tissue of goldfish. They were also more resistant to UV-irradiation than CAF-MM1 cells. Photoreactivation after UV-irradiation was present in GEM 199 cells for both survival and mutation. The initial shoulder of the survival curve of UV-irradiated cells was reduced greatly by caffeine, suggesting a high activity of the post-replication repair. The spontaneous mutation frequency to ouabain resistance was 1-5x10/sup -6/ clones per viable cell. MNNG was effective in inducing ouabain-resistant mutation, while 4NQO and ..gamma..-rays did not induce mutation.

  7. EC6 for Darlington - a managed-enhancements replication project

    International Nuclear Information System (INIS)

    Soulard, M.R.

    2011-01-01

    CANDU 6 design has a well-deserved reputation for lifetime performance and on-time, on-schedule construction. Candu Energy is finalizing development of an upgraded reactor product, the Enhanced CANDU 6 (EC6), which incorporates the CANDU 6's well-proven features, and adds enhancements that make the reactor even more safe and easier to operate. The EC6 is the only mid-sized reactor (700 MWe class) with a proven pedigree that meets modern reactor expectations and regulatory standards. It is sized for smaller grids and also has outstanding fuel-cycle capability. Some of the key features incorporated into the EC6 include increasing the plant's power output, shortening the overall project schedule, addressing obsolescence issues, optimizing maintenance outages and incorporating lessons learnt through feedback obtained from the operating plants. The EC6 utilizes modern computers and a distributed control system housed in an advanced control room which, along with automated testing and on-line diagnostics, make the plant easier and safer to operate, with minimal operator intervention. An array of health monitoring equipment are also installed to foretell impending equipment problems, which can be acted upon, avoiding complications that could result in forced shutdowns. Improvements to the fire protection system and enhanced security features will further protect the assets. Containment and seismic capability are upgraded to meet modern standards. The first deployment of the EC6 is anticipated in Canada; other markets are also being pursued. The EC6 reactor is designed for a target lifetime capacity factor of 92% including 94% year-to-year and a 1% Forced Loss Rate. Since the number and duration of maintenance outages impact plant capacity factors, periodic short duration maintenance outages of less than a month once every 36 months is a key target of the EC6 reactor. This objective is achieved by automating a number of tasks such as shutdown systems testing. The majority

  8. Fabrication of endothelial progenitor cell capture surface via DNA aptamer modifying dopamine/polyethyleneimine copolymer film

    Energy Technology Data Exchange (ETDEWEB)

    Li, Xin; Deng, Jinchuan; Yuan, Shuheng; Wang, Juan; Luo, Rifang; Chen, Si [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China); Wang, Jin, E-mail: jinxxwang@263.net [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); Huang, Nan [Key Lab. of Advanced Technology for Materials of Education Ministry, Southwest Jiaotong University, Chengdu 610031 (China); School of Material Science and Engineering, Southwest Jiaotong University, Chengdu 610031 (China)

    2016-11-15

    Highlights: • The dopamine/PEI film with controlled amine density was successfully prepared. • The DNA aptamer was assembled onto the film via electrostatic incorporation. • The A@DPfilmscanspecificallyandeffectivelycaptureEPCs. • The A@DP film can support the survival of ECs, control the hyperplasia of SMCs. • The dynamic/co-culture models are useful for studying cells competitive adhesion. - Abstract: Endothelial progenitor cells (EPCs) are mainly located in bone marrow and circulate, and play a crucial role in repairmen of injury endothelium. One of the most promising strategies of stents designs were considered to make in-situ endothelialization in vivo via EPC-capture biomolecules on a vascular graft to capture EPCs directly from circulatory blood. In this work, an EPC specific aptamer with a 34 bases single strand DNA sequence was conjugated onto the stent surface via dopamine/polyethyleneimine copolymer film as a platform and linker. The assembled density of DNA aptamer could be regulated by controlling dopamine percentage in this copolymer film. X-ray photoelectron spectroscopy (XPS), water contact angle (WCA) and fluorescence test confirmed the successful immobilization of DNA aptamer. To confirm its biofunctionality and cytocompatibility, the capturing cells ability of the aptamer modified surface and the effects on the growth behavior of human umbilical vein endothelial cells (HUVECs), smooth muscle cells (SMCs) were investigated. The aptamer functionalized sample revealed a good EPC-capture ability, and had a cellular friendly feature for both EPC and EC growth, while not stimulated the hyperplasia of SMCs. And, the co-culture experiment of three types of cells confirmed the specificity capturing of EPCs to aptamer modified surface, rather than ECs and SMCs. These data suggested that this aptamer functionalized surface may have a large potentiality for the application of vascular grafts with targeted endothelialization.

  9. Fabrication of endothelial progenitor cell capture surface via DNA aptamer modifying dopamine/polyethyleneimine copolymer film

    International Nuclear Information System (INIS)

    Li, Xin; Deng, Jinchuan; Yuan, Shuheng; Wang, Juan; Luo, Rifang; Chen, Si; Wang, Jin; Huang, Nan

    2016-01-01

    Highlights: • The dopamine/PEI film with controlled amine density was successfully prepared. • The DNA aptamer was assembled onto the film via electrostatic incorporation. • The A@DPfilmscanspecificallyandeffectivelycaptureEPCs. • The A@DP film can support the survival of ECs, control the hyperplasia of SMCs. • The dynamic/co-culture models are useful for studying cells competitive adhesion. - Abstract: Endothelial progenitor cells (EPCs) are mainly located in bone marrow and circulate, and play a crucial role in repairmen of injury endothelium. One of the most promising strategies of stents designs were considered to make in-situ endothelialization in vivo via EPC-capture biomolecules on a vascular graft to capture EPCs directly from circulatory blood. In this work, an EPC specific aptamer with a 34 bases single strand DNA sequence was conjugated onto the stent surface via dopamine/polyethyleneimine copolymer film as a platform and linker. The assembled density of DNA aptamer could be regulated by controlling dopamine percentage in this copolymer film. X-ray photoelectron spectroscopy (XPS), water contact angle (WCA) and fluorescence test confirmed the successful immobilization of DNA aptamer. To confirm its biofunctionality and cytocompatibility, the capturing cells ability of the aptamer modified surface and the effects on the growth behavior of human umbilical vein endothelial cells (HUVECs), smooth muscle cells (SMCs) were investigated. The aptamer functionalized sample revealed a good EPC-capture ability, and had a cellular friendly feature for both EPC and EC growth, while not stimulated the hyperplasia of SMCs. And, the co-culture experiment of three types of cells confirmed the specificity capturing of EPCs to aptamer modified surface, rather than ECs and SMCs. These data suggested that this aptamer functionalized surface may have a large potentiality for the application of vascular grafts with targeted endothelialization.

  10. High-Content Electrophysiological Analysis of Human Pluripotent Stem Cell-Derived Cardiomyocytes (hPSC-CMs).

    Science.gov (United States)

    Kong, Chi-Wing; Geng, Lin; Li, Ronald A

    2018-01-01

    Considerable interest has been raised to develop human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) as a model for drug discovery and cardiotoxicity screening. High-content electrophysiological analysis of currents generated by transmembrane cell surface ion channels has been pursued to complement such emerging applications. Here we describe practical procedures and considerations for accomplishing successful assays of hPSC-CMs using an automated planar patch-clamp system.

  11. Transplant of stem cells derived from bone marrow and granulocytic growth factor in acute and chronic ischemic myocardiopathy

    International Nuclear Information System (INIS)

    Senior Juan M; Cuellar Francisco; Velasquez Oscar; Velasquez Margarita; Navas Claudia M; Ortiz Sergio; Delgado Juan A; Guillerrno, Blanco; Londono Juan L; Coronado Manuel A; Gomez Francisco; Alzate, Fernando Leon; Zuluaga Alejandra

    2007-01-01

    Recent studies have shown the safety and efficacy of the stem cells derived from bone marrow (BMC) implant with concomitant administration of stimulating factor of granulocyte colonies in patients with acute myocardial infarction with ST segment elevation and in chronic ischemic cardiopathy. An open prospective (before and after) design was made to evaluate the safety and efficacy of cell therapy associated to growth factor administration. The first experience with this kind of therapy is reported. Methodology: this is a 6 months follow-up report of patients with acute and chronic ischemic cardiopathy to who transplant of stem cells derived from bone marrow mobilized with granulocyte colonies growth stimulating factor via coronary arteries or epicardium was realized. Two groups of patients were included: Ten patients with anterior wall infarct and 2. Five patients with chronic ischemic cardiopathy, all with extensive necrosis demonstrated by absence of myocardial viability through nuclear medicine and ejection fraction of less than 40%. Results: significant improvement of ejection fraction from 29.44 ± 3.36 to 37.6 ± 5.3 with p<0.001 and decrease of ventricular systolic and diastolic volume without statistical significance (p =0.31 and 0.4 respectively) were demonstrated. Exercise capacity evidenced by increment in the six minutes test, exercise time and the MET number achieved, increased in a significant way. There were significant changes in the perfusion defect from the second follow-up month and no complications directly related to the stem cells derived from bone marrow transplant or the use of stimulating granulocyte colony factor were presented. Conclusions: this is the first experience of stem cells derived from bone marrow transplant associated to the administration of stimulating granulocyte growth colony factor in which recovery of left ventricular function was demonstrated, as well as improvement in exercise capacity and in the perfusion defect

  12. Retinal progenitor cell xenografts to the pig retina

    DEFF Research Database (Denmark)

    Warfvinge, Karin; Kiilgaard, Jens Folke; Lavik, Erin B

    2005-01-01

    To investigate the survival, integration, and differentiation of mouse retinal progenitor cells after transplantation to the subretinal space of adult pigs.......To investigate the survival, integration, and differentiation of mouse retinal progenitor cells after transplantation to the subretinal space of adult pigs....

  13. Clonal Heterogeneity in the Neuronal and Glial Differentiation of Dental Pulp Stem/Progenitor Cells

    Directory of Open Access Journals (Sweden)

    Fraser I. Young

    2016-01-01

    Full Text Available Cellular heterogeneity presents an important challenge to the development of cell-based therapies where there is a fundamental requirement for predictable and reproducible outcomes. Transplanted Dental Pulp Stem/Progenitor Cells (DPSCs have demonstrated early promise in experimental models of spinal cord injury and stroke, despite limited evidence of neuronal and glial-like differentiation after transplantation. Here, we report, for the first time, on the ability of single cell-derived clonal cultures of murine DPSCs to differentiate in vitro into immature neuronal-like and oligodendrocyte-like cells. Importantly, only DPSC clones with high nestin mRNA expression levels were found to successfully differentiate into Map2 and NF-positive neuronal-like cells. Neuronally differentiated DPSCs possessed a membrane capacitance comparable with primary cultured striatal neurons and small inward voltage-activated K+ but not outward Na+ currents were recorded suggesting a functionally immature phenotype. Similarly, only high nestin-expressing clones demonstrated the ability to adopt Olig1, Olig2, and MBP-positive immature oligodendrocyte-like phenotype. Together, these results demonstrate that appropriate markers may be used to provide an early indication of the suitability of a cell population for purposes where differentiation into a specific lineage may be beneficial and highlight that further understanding of heterogeneity within mixed cellular populations is required.

  14. Differentiation of Inflammation-Responsive Astrocytes from Glial Progenitors Generated from Human Induced Pluripotent Stem Cells

    Directory of Open Access Journals (Sweden)

    Renata Santos

    2017-06-01

    Full Text Available Astrocyte dysfunction and neuroinflammation are detrimental features in multiple pathologies of the CNS. Therefore, the development of methods that produce functional human astrocytes represents an advance in the study of neurological diseases. Here we report an efficient method for inflammation-responsive astrocyte generation from induced pluripotent stem cells (iPSCs and embryonic stem cells. This protocol uses an intermediate glial progenitor stage and generates functional astrocytes that show levels of glutamate uptake and calcium activation comparable with those observed in human primary astrocytes. Stimulation of stem cell-derived astrocytes with interleukin-1β or tumor necrosis factor α elicits a strong and rapid pro-inflammatory response. RNA-sequencing transcriptome profiling confirmed that similar gene expression changes occurred in iPSC-derived and primary astrocytes upon stimulation with interleukin-1β. This protocol represents an important tool for modeling in-a-dish neurological diseases with an inflammatory component, allowing for the investigation of the role of diseased astrocytes in neuronal degeneration.

  15. Differentiation of Inflammation-Responsive Astrocytes from Glial Progenitors Generated from Human Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Santos, Renata; Vadodaria, Krishna C; Jaeger, Baptiste N; Mei, Arianna; Lefcochilos-Fogelquist, Sabrina; Mendes, Ana P D; Erikson, Galina; Shokhirev, Maxim; Randolph-Moore, Lynne; Fredlender, Callie; Dave, Sonia; Oefner, Ruth; Fitzpatrick, Conor; Pena, Monique; Barron, Jerika J; Ku, Manching; Denli, Ahmet M; Kerman, Bilal E; Charnay, Patrick; Kelsoe, John R; Marchetto, Maria C; Gage, Fred H

    2017-06-06

    Astrocyte dysfunction and neuroinflammation are detrimental features in multiple pathologies of the CNS. Therefore, the development of methods that produce functional human astrocytes represents an advance in the study of neurological diseases. Here we report an efficient method for inflammation-responsive astrocyte generation from induced pluripotent stem cells (iPSCs) and embryonic stem cells. This protocol uses an intermediate glial progenitor stage and generates functional astrocytes that show levels of glutamate uptake and calcium activation comparable with those observed in human primary astrocytes. Stimulation of stem cell-derived astrocytes with interleukin-1β or tumor necrosis factor α elicits a strong and rapid pro-inflammatory response. RNA-sequencing transcriptome profiling confirmed that similar gene expression changes occurred in iPSC-derived and primary astrocytes upon stimulation with interleukin-1β. This protocol represents an important tool for modeling in-a-dish neurological diseases with an inflammatory component, allowing for the investigation of the role of diseased astrocytes in neuronal degeneration. Copyright © 2017 The Authors. Published by Elsevier Inc. All rights reserved.

  16. Functional integration of grafted neural stem cell-derived dopaminergic neurons monitored by optogenetics in an in vitro Parkinson model.

    Directory of Open Access Journals (Sweden)

    Jan Tønnesen

    Full Text Available Intrastriatal grafts of stem cell-derived dopamine (DA neurons induce behavioral recovery in animal models of Parkinson's disease (PD, but how they functionally integrate in host neural circuitries is poorly understood. Here, Wnt5a-overexpressing neural stem cells derived from embryonic ventral mesencephalon of tyrosine hydroxylase-GFP transgenic mice were expanded as neurospheres and transplanted into organotypic cultures of wild type mouse striatum. Differentiated GFP-labeled DA neurons in the grafts exhibited mature neuronal properties, including spontaneous firing of action potentials, presence of post-synaptic currents, and functional expression of DA D₂ autoreceptors. These properties resembled those recorded from identical cells in acute slices of intrastriatal grafts in the 6-hydroxy-DA-induced mouse PD model and from DA neurons in intact substantia nigra. Optogenetic activation or inhibition of grafted cells and host neurons using channelrhodopsin-2 (ChR2 and halorhodopsin (NpHR, respectively, revealed complex, bi-directional synaptic interactions between grafted cells and host neurons and extensive synaptic connectivity within the graft. Our data demonstrate for the first time using optogenetics that ectopically grafted stem cell-derived DA neurons become functionally integrated in the DA-denervated striatum. Further optogenetic dissection of the synaptic wiring between grafted and host neurons will be crucial to clarify the cellular and synaptic mechanisms underlying behavioral recovery as well as adverse effects following stem cell-based DA cell replacement strategies in PD.

  17. Nano-zymography Using Laser-Scanning Confocal Microscopy Unmasks Proteolytic Activity of Cell-Derived Microparticles.

    Science.gov (United States)

    Briens, Aurélien; Gauberti, Maxime; Parcq, Jérôme; Montaner, Joan; Vivien, Denis; Martinez de Lizarrondo, Sara

    2016-01-01

    Cell-derived microparticles (MPs) are nano-sized vesicles released by activated cells in the extracellular milieu. They act as vectors of biological activity by carrying membrane-anchored and cytoplasmic constituents of the parental cells. Although detection and characterization of cell-derived MPs may be of high diagnostic and prognostic values in a number of human diseases, reliable measurement of their size, number and biological activity still remains challenging using currently available methods. In the present study, we developed a protocol to directly image and functionally characterize MPs using high-resolution laser-scanning confocal microscopy. Once trapped on annexin-V coated micro-wells, we developed several assays using fluorescent reporters to measure their size, detect membrane antigens and evaluate proteolytic activity (nano-zymography). In particular, we demonstrated the applicability and specificity of this method to detect antigens and proteolytic activities of tissue-type plasminogen activator (tPA), urokinase and plasmin at the surface of engineered MPs from transfected cell-lines. Furthermore, we were able to identify a subset of tPA-bearing fibrinolytic MPs using plasma samples from a cohort of ischemic stroke patients who received thrombolytic therapy and in an experimental model of thrombin-induced ischemic stroke in mice. Overall, this method is promising for functional characterization of cell-derived MPs.

  18. Cell-Derived Microparticles in Patients with Type 2 Diabetes Mellitus: a Systematic Review and Meta-Analysis

    Directory of Open Access Journals (Sweden)

    Sheyu Li

    2016-11-01

    Full Text Available Background/Aims: The aim of this study was to assess the association between circulating cell-derived microparticles (MPs and type 2 diabetes mellitus (T2DM. Methods: A literature search was performed systematically in PubMed and Embase to identify available case-control or cross-sectional studies that compared different types of cell-derived MPs in patients with T2DM and non-diabetic controls. Pooled standardized mean differences (SMDs of each MP type were pooled using meta-analysis. Results: Forty-eight studies involving 2,460 patients with T2DM and 1,880 non-diabetic controls were included for systematic review and 34 of which were included for quantitative study by meta-analysis. In the overall analysis, the levels of circulating total MPs (TMPs, platelet-derived MPs (PMPs, monocyte-derived MPs (MMPs and endothelium-derived MPs (EMPs were significantly higher in T2DM patients than those in controls (TMPs: SMD, 0.64; 95%CI, 0.12∼1.15; P=0.02; PMPs: SMD, 1.19; 95%CI, 0.88∼1.50; P Conclusions: The counts of TMPs, PMPs, MMPs and EMPs elevated in patients with T2DM. And cell-derived MPs may play a role in the pathogenesis of T2DM.

  19. Selective uptake of boronophenylalanine by glioma stem/progenitor cells

    International Nuclear Information System (INIS)

    Sun, Ting; Zhou, Youxin; Xie, Xueshun; Chen, Guilin; Li, Bin; Wei, Yongxin; Chen, Jinming; Huang, Qiang; Du, Ziwei

    2012-01-01

    The success of boron neutron capture therapy (BNCT) depends on the amount of boron in cells and the tumor/blood and tumor/(normal tissue) boron concentration ratios. For the first time, measurements of boron uptake in both stem/progenitor and differentiated glioma cells were performed along with measurements of boron biodistribution in suitable animal models. In glioma stem/progenitor cells, the selective accumulation of boronophenylalanine (BPA) was lower, and retention of boron after BPA removal was longer than in differentiated glioma cells in vitro. However, boron biodistribution was not statistically significantly different in mice with xenografts. - Highlights: ► Uptake of BPA was analyzed in stem/progenitor and differentiated glioma cells. ► Selective accumulation of BPA was lower in glioma stem/progenitor cells. ► Retention of boron after BPA removal was longer in glioma stem/progenitor cells. ► Boron biodistribution was not statistically different in mice with xenografts.

  20. Determination of primary combustion source organic carbon-to-elemental carbon (OC / EC ratio using ambient OC and EC measurements: secondary OC-EC correlation minimization method

    Directory of Open Access Journals (Sweden)

    C. Wu

    2016-05-01

    Full Text Available Elemental carbon (EC has been widely used as a tracer to track the portion of co-emitted primary organic carbon (OC and, by extension, to estimate secondary OC (SOC from ambient observations of EC and OC. Key to this EC tracer method is to determine an appropriate OC / EC ratio that represents primary combustion emission sources (i.e., (OC / ECpri at the observation site. The conventional approaches include regressing OC against EC within a fixed percentile of the lowest (OC / EC ratio data (usually 5–20 % or relying on a subset of sampling days with low photochemical activity and dominated by local emissions. The drawback of these approaches is rooted in its empirical nature, i.e., a lack of clear quantitative criteria in the selection of data subsets for the (OC / ECpri determination. We examine here a method that derives (OC / ECpri through calculating a hypothetical set of (OC / ECpri and SOC followed by seeking the minimum of the coefficient of correlation (R2 between SOC and EC. The hypothetical (OC / ECpri that generates the minimum R2(SOC,EC then represents the actual (OC / ECpri ratio if variations of EC and SOC are independent and (OC / ECpri is relatively constant in the study period. This Minimum R Squared (MRS method has a clear quantitative criterion for the (OC / ECpri calculation. This work uses numerically simulated data to evaluate the accuracy of SOC estimation by the MRS method and to compare with two commonly used methods: minimum OC / EC (OC / ECmin and OC / EC percentile (OC / EC10 %. Log-normally distributed EC and OC concentrations with known proportion of SOC are numerically produced through a pseudorandom number generator. Three scenarios are considered, including a single primary source, two independent primary sources, and two correlated primary sources. The MRS method consistently yields the most accurate SOC estimation. Unbiased SOC estimation by OC

  1. Integration of IC/EC systems in ITER

    International Nuclear Information System (INIS)

    Gassmann, T.; Beaumont, B.; Baruah, U.K.; Bonicelli, T.; Chiocchio, S.; Cox, D.; Darbos, C.; Decamps, H.; Denisov, G.; Henderson, M.; Kazarian, F.; Lamalle, P.U.; Mukherjee, A.; Rasmussen, D.; Saibene, G.; Sartori, R.; Sakamoto, K.; Tanga, A.

    2010-01-01

    The RF heating and current drive (H and CD) systems that are to be installed in ITER during the construction phase, are the electron cyclotron (EC) and ion cyclotron (IC) systems. They are complex assemblies of high voltage power supplies (HVPS), RF generators, transmission lines and antennas. Their design and integration are constrained by many interfaces, both internal, between the subsystems, and external, with the other ITER systems. In addition, some components must be compatible with a nuclear environment and are classified as Safety Important Component. This paper describes the processes implemented in ITER to ensure proper integration.

  2. Environmental Quality Standards in the EC-Water Framework Directive

    DEFF Research Database (Denmark)

    Jirka, Gerhard H.; Burrows, Richard; Larsen, Torben

    2004-01-01

    The "combined approach" in the new EC-Water Framework Directive(WFD) consisting of environmental quality standards in addition to emission limit values promises improvements in the quality characteristics of surface water. However, the specification of where in the water body the environmental...... quality standards apply is missing in the WFD. The omission will limit its administrative implementation. A clear mixing zone regulation is needed so that the quality objectives of the WFD are not jeopardized. This need is demonstrated using the examples of point source discharges into rivers and coastal...

  3. Vanadate impedes adipogenesis in mesenchymal stem cells derived from different depots within bone.

    Directory of Open Access Journals (Sweden)

    Frans Alexander Jacobs

    2016-08-01

    Full Text Available Glucocorticoid induced osteoporosis (GIO is associated with an increase in bone marrow adiposity which skews the differentiation of mesenchymal stem cell (MSC progenitors away from osteoblastogenesis and towards adipogenesis. We have previously found that vanadate, a non-specific protein tyrosine phosphatase inhibitor, prevents GIO in rats, but it was unclear whether vanadate directly influenced adipogenesis in bone-derived MSCs. For the present study, we investigated the effect of vanadate on adipogenesis in primary rat MSCs derived from bone marrow (bmMSCs and from the proximal end of the femur (pfMSCs. By passage 3 after isolation, both cell populations expressed the MSC cell surface markers CD90 and CD106, but not the haematopoietic marker CD45. However, although variable, expression of the fibroblast marker CD26 was higher in pfMSCs than in bmMSCs. Differentiation studies using osteogenic and adipogenic induction media (OM and AM, respectively demonstrated that pfMSCs rapidly accumulated lipid droplets within 1 week of exposure to AM, while bmMSCs isolated from the same femur only formed lipid droplets after 3 weeks of AM treatment. Conversely, pfMSCs exposed to OM produced mineralized extracellular matrix (ECM after 3 weeks, compared to 1 week for OM-treated bmMSCs. Vanadate (10 µM added to AM resulted in a significant reduction in AM-induced intracellular lipid accumulation and expression of adipogenic gene markers (PPARγ2, aP2, adipsin in both pfMSCs and bmMSCs. Pharmacological concentrations of glucocorticoids (1 µM alone did not induce lipid accumulation in either bmMSCs or pfMSCs, but resulted in significant cell death in pfMSCs. Our findings demonstrate the existence of at least two fundamentally different MSC depots within the femur, and highlights the presence of MSCs capable of rapid adipogenesis within the proximal femur, an area prone to osteoporotic fractures. In addition, our results suggest that the increased bone marrow

  4. Impairment of circulating endothelial progenitors in Down syndrome

    Directory of Open Access Journals (Sweden)

    Costa Valerio

    2010-09-01

    Full Text Available Abstract Background Pathological angiogenesis represents a critical issue in the progression of many diseases. Down syndrome is postulated to be a systemic anti-angiogenesis disease model, possibly due to increased expression of anti-angiogenic regulators on chromosome 21. The aim of our study was to elucidate some features of circulating endothelial progenitor cells in the context of this syndrome. Methods Circulating endothelial progenitors of Down syndrome affected individuals were isolated, in vitro cultured and analyzed by confocal and transmission electron microscopy. ELISA was performed to measure SDF-1α plasma levels in Down syndrome and euploid individuals. Moreover, qRT-PCR was used to quantify expression levels of CXCL12 gene and of its receptor in progenitor cells. The functional impairment of Down progenitors was evaluated through their susceptibility to hydroperoxide-induced oxidative stress with BODIPY assay and the major vulnerability to the infection with human pathogens. The differential expression of crucial genes in Down progenitor cells was evaluated by microarray analysis. Results We detected a marked decrease of progenitors' number in young Down individuals compared to euploid, cell size increase and some major detrimental morphological changes. Moreover, Down syndrome patients also exhibited decreased SDF-1α plasma levels and their progenitors had a reduced expression of SDF-1α encoding gene and of its membrane receptor. We further demonstrated that their progenitor cells are more susceptible to hydroperoxide-induced oxidative stress and infection with Bartonella henselae. Further, we observed that most of the differentially expressed genes belong to angiogenesis, immune response and inflammation pathways, and that infected progenitors with trisomy 21 have a more pronounced perturbation of immune response genes than infected euploid cells. Conclusions Our data provide evidences for a reduced number and altered

  5. Enhanced adhesion of early endothelial progenitor cells to radiation-induced senescence-like vascular endothelial cells in vitro

    International Nuclear Information System (INIS)

    Sermsathanasawadi, N.; Inoue, Yoshinori; Iwai, Takehisa; Ishii, Hideto; Yoshida, Masayuki; Igarashi, Kaori; Miura, Masahiko

    2009-01-01

    The effects of ionizing radiation (IR) on tumor neovascularization are still unclear. We previously reported that vascular endothelial cells (ECs) expressing the IR-induced senescence-like (IRSL) phenotype exhibit a significant decrease in angiogenic activity in vitro. In this study, we examined the effects of the IRSL phenotype on adhesion to early endothelial progenitor cells (early EPCs). Adhesion of human peripheral blood-derived early EPCs to human umbilical vein endothelial cells (HUVECs) expressing the IRSL phenotype was evaluated by an adhesion assay under static conditions. It was revealed that the IRSL HUVECs supported significantly more adhesion of early EPCs than normal HUVECs. Expressions of ICAM-1, VCAM-1 and E-selectin were up-regulated in IRSL HUVECs. Pre-treatment of IRSL HUVECs with adhesion-blocking monoclonal antibodies against E-selectin and VCAM-1 significantly reduced early EPC adhesion to IRSL HUVECs, suggesting a potential role for the E-selectin and VCAM-1 in the adhesion between IRSL ECs and early EPCs. Therefore, the IRSL phenotype expressed in ECs may enhance neovascularization via increased homing of early EPCs. Our findings are first to implicate the complex effects of this phenotype on tumor neovascularization following irradiation. (author)

  6. Archive of Geosample Data and Information from the U.S. Extended Continental Shelf (ECS) Project.

    Data.gov (United States)

    National Oceanic and Atmospheric Administration, Department of Commerce — Geological samples collected as part of the U.S. Extended Continental Shelf (ECS) Project are curated by the U.S. Geological Survey, a partner in the ECS Project. To...

  7. Everyone talk about EC technology - What's about this technology? AC versus AC technology; Alle reden von EC-Technologie - was bringt sie wirklich? AC- versus EC-Motorentechnik

    Energy Technology Data Exchange (ETDEWEB)

    Anon.

    2010-12-15

    Meanwhile, the EC motor technology is the state of the art in fans. In the consciousness of a possible energy conservation, the EC motor technology is demanded to a large extent with the new installations of the customers. But how high are the possible savings actually? A comparative measurement at a condenser of Guentner AG (Fuerstenfeldbruck, Federal Republic of Germany) supports information.

  8. Attenuation of hind-limb ischemia in mice with endothelial-like cells derived from different sources of human stem cells.

    Directory of Open Access Journals (Sweden)

    Wing-Hon Lai

    Full Text Available Functional endothelial-like cells (EC have been successfully derived from different cell sources and potentially used for treatment of cardiovascular diseases; however, their relative therapeutic efficacy remains unclear. We differentiated functional EC from human bone marrow mononuclear cells (BM-EC, human embryonic stem cells (hESC-EC and human induced pluripotent stem cells (hiPSC-EC, and compared their in-vitro tube formation, migration and cytokine expression profiles, and in-vivo capacity to attenuate hind-limb ischemia in mice. Successful differentiation of BM-EC was only achieved in 1/6 patient with severe coronary artery disease. Nevertheless, BM-EC, hESC-EC and hiPSC-EC exhibited typical cobblestone morphology, had the ability of uptaking DiI-labeled acetylated low-density-lipoprotein, and binding of Ulex europaeus lectin. In-vitro functional assay demonstrated that hiPSC-EC and hESC-EC had similar capacity for tube formation and migration as human umbilical cord endothelial cells (HUVEC and BM-EC (P>0.05. While increased expression of major angiogenic factors including epidermal growth factor, hepatocyte growth factor, vascular endothelial growth factor, placental growth factor and stromal derived factor-1 were observed in all EC cultures during hypoxia compared with normoxia (P<0.05, the magnitudes of cytokine up-regulation upon hypoxic were more dramatic in hiPSC-EC and hESC-EC (P<0.05. Compared with medium, transplanting BM-EC (n = 6, HUVEC (n = 6, hESC-EC (n = 8 or hiPSC-EC (n = 8 significantly attenuated severe hind-limb ischemia in mice via enhancement of neovascularization. In conclusion, functional EC can be generated from hECS and hiPSC with similar therapeutic efficacy for attenuation of severe hind-limb ischemia. Differentiation of functional BM-EC was more difficult to achieve in patients with cardiovascular diseases, and hESC-EC or iPSC-EC are readily available as "off-the-shelf" format for the treatment

  9. Attenuation of Hind-Limb Ischemia in Mice with Endothelial-Like Cells Derived from Different Sources of Human Stem Cells

    Science.gov (United States)

    Chan, Yau-Chi; Ng, Joyce H. L.; Au, Ka-Wing; Wong, Lai-Yung; Siu, Chung-Wah; Tse, Hung-Fat

    2013-01-01

    Functional endothelial-like cells (EC) have been successfully derived from different cell sources and potentially used for treatment of cardiovascular diseases; however, their relative therapeutic efficacy remains unclear. We differentiated functional EC from human bone marrow mononuclear cells (BM-EC), human embryonic stem cells (hESC-EC) and human induced pluripotent stem cells (hiPSC-EC), and compared their in-vitro tube formation, migration and cytokine expression profiles, and in-vivo capacity to attenuate hind-limb ischemia in mice. Successful differentiation of BM-EC was only achieved in 1/6 patient with severe coronary artery disease. Nevertheless, BM-EC, hESC-EC and hiPSC-EC exhibited typical cobblestone morphology, had the ability of uptaking DiI-labeled acetylated low-density-lipoprotein, and binding of Ulex europaeus lectin. In-vitro functional assay demonstrated that hiPSC-EC and hESC-EC had similar capacity for tube formation and migration as human umbilical cord endothelial cells (HUVEC) and BM-EC (P>0.05). While increased expression of major angiogenic factors including epidermal growth factor, hepatocyte growth factor, vascular endothelial growth factor, placental growth factor and stromal derived factor-1 were observed in all EC cultures during hypoxia compared with normoxia (P<0.05), the magnitudes of cytokine up-regulation upon hypoxic were more dramatic in hiPSC-EC and hESC-EC (P<0.05). Compared with medium, transplanting BM-EC (n = 6), HUVEC (n = 6), hESC-EC (n = 8) or hiPSC-EC (n = 8) significantly attenuated severe hind-limb ischemia in mice via enhancement of neovascularization. In conclusion, functional EC can be generated from hECS and hiPSC with similar therapeutic efficacy for attenuation of severe hind-limb ischemia. Differentiation of functional BM-EC was more difficult to achieve in patients with cardiovascular diseases, and hESC-EC or iPSC-EC are readily available as “off-the-shelf” format for the treatment of

  10. ESCAPE: an integrated climate model for the EC

    International Nuclear Information System (INIS)

    Rotmans, J.

    1992-01-01

    A framework has been developed for the evaluation of policy options for climate change, called ESCAPE (Evaluation of Strategies to address Climate change by Adapting to and Preventing Emissions). ESCAPE consists of a suite of linked models which enables scenarios of greenhouse gas emissions to be constructed and their impact on global and regional climate and sea level and sectors of the European economy to be assessed. Conclusions resulting from simulations with the ESCAPE 1.1 model include: the major problem of a climate change for the EC is a sea level rise; Greece, Italy, Portugal and Spain will be faced with higher costs in the agricultural sector; worldwide implementation of an EC carbon tax leads to about 12% lower worldwide CO 2 emissions; to stabilize CO 2 emissions an Ecotax of 18 dollars per barrel would be required; and in all cases the rate of global temperature increase will be above the rate of 0.1 degree C per decade for the coming 40 years. 2 figs

  11. Translation and validation of the Dutch version of the Effective Consumer Scale (EC-17)

    NARCIS (Netherlands)

    ten Klooster, Peter M.; Taal, Erik; Tjin-Kam-Jet-Siemons, Liseth; Oostveen, J.C.M.; Oostveen, Johanna C.M.; Harmsen, Etelka J.; Tugwell, Peter S.; Rader, Tamara; Lyddiatt, Anne; van de Laar, Mart A F J

    2013-01-01

    PURPOSE: The Effective Consumer Scale (EC-17) measures the skills of musculoskeletal patients in managing their own healthcare. The objectives of this study were to translate the EC-17 into Dutch and to further evaluate its psychometric properties. METHODS: The EC-17 was translated and cognitively

  12. Cardiac Progenitor Cell Extraction from Human Auricles

    KAUST Repository

    Di Nardo, Paolo

    2017-02-22

    For many years, myocardial tissue has been considered terminally differentiated and, thus, incapable of regenerating. Recent studies have shown, instead, that cardiomyocytes, at least in part, are slowly substituted by new cells originating by precursor cells mostly embedded into the heart apex and in the atria. We have shown that an elective region of progenitor cell embedding is represented by the auricles, non-contractile atria appendages that can be easily sampled without harming the patient. The protocol here reported describes how from auricles a population of multipotent, cardiogenic cells can be isolated, cultured, and differentiated. Further studies are needed to fully exploit this cell population, but, sampling auricles, it could be possible to treat cardiac patients using their own cells circumventing rejection or organ shortage limitations.

  13. Cytokinetics and Regulation of Progenitor Cells

    Energy Technology Data Exchange (ETDEWEB)

    Lajtha, L. G. [Paterson Laboratories, Christie Hospital and Holt Radium Institute, Manchester (United Kingdom)

    1967-07-15

    Full text: In spite of great differences in the life-span of fully differentiated haemic cells, the cellular kinetics of their production appears to be similar. Recent evidence indicates a common ultimate stem cell for most of the cells in the peripheral blood. The various pathways of differentiation, however, result in transient dividing and differentiating cell populations which differ from each other not only in their specific biochemical processes but also in the manner of control and kinetic pattern of their proliferation. The population best understood is the erythroid progenitor series of cells, primarily because it has the greatest number of experimentally measurable parameters at the present. This will be discussed in detail and comparisons will be made with the myeloid and lymphoid progenitor populations. The fine structure of the bone-marrow stem cell population will be examined in particular, with regard to the suitability or otherwise of the current stem cell models to explain the kinetic pattern of all the peripheral blood elements after perturbations of their steady-state values. Four different assay methods of bone-marrow stem cells have been examined with regard to the kinetic pattern following perturbation of the steady-state system, e.g. by irradiation. Basically, the stem cell assays fall into two categories: those depending on grafting haemopoietic cells into suitably treated recipients, and those in which recovery of the population is allowed in the animal in which the perturbation was produced, without handling the cells. Evidence is accumulating which indicates that in the grafting techniques, a selective loss of stem cells may occur, . especially stem cells in cell cycle, hence in early stages of recovery of the population unduly low numerical values might be noted. In view of this observation, the concept of the colony-forming cell may have to be revised and instead the colony-forming property of the stem cell introduced. (author)

  14. A COMPREHENSIVE PROGENITOR MODEL FOR SNe Ia

    International Nuclear Information System (INIS)

    Meng, X.; Yang, W.

    2010-01-01

    Although the nature of the progenitor of Type Ia supernovae (SNe Ia) is still unclear, the single-degenerate (SD) channel for the progenitor is currently accepted, in which a carbon-oxygen white dwarf (CO WD) accretes hydrogen-rich material from its companion, increases its mass to the Chandrasekhar mass limit, and then explodes as an SN Ia. The companion may be a main sequence or a slightly evolved star (WD + MS), or a red giant star (WD + RG). Incorporating the effect of mass stripping and accretion-disk instability on the evolution of the WD binary, we carried out binary stellar evolution calculations for more than 1600 close WD binaries. As a result, the initial parameter spaces for SNe Ia are presented in an orbital period-secondary mass (log P i , M i 2 ) plane. We confirmed that in a WD + MS system, the initial companion leading to SNe Ia may have mass from 1 M sun to 5 M sun . The initial WD mass for SNe Ia from WD + MS channel is as low as 0.565 M sun , while the lowest WD mass from the WD + RG channel is 1.0 M sun . Adopting the above results, we studied the birth rate of SNe Ia via a binary population synthesis approach. We found that the Galactic SNe Ia birth rate from SD model is (2.55-2.9) x 10 -3 yr -1 (including WD + He star channel), which is slightly smaller than that from observation. If a single starburst is assumed, the distribution of the delay time of SNe Ia from the SD model may be a weak bimodality, where WD + He channel contributes to SNe Ia with delay time shorter than 10 8 yr and WD + RG channel to those with age longer than 6 Gyr.

  15. Cytokinetics and Regulation of Progenitor Cells

    International Nuclear Information System (INIS)

    Lajtha, L.G.

    1967-01-01

    Full text: In spite of great differences in the life-span of fully differentiated haemic cells, the cellular kinetics of their production appears to be similar. Recent evidence indicates a common ultimate stem cell for most of the cells in the peripheral blood. The various pathways of differentiation, however, result in transient dividing and differentiating cell populations which differ from each other not only in their specific biochemical processes but also in the manner of control and kinetic pattern of their proliferation. The population best understood is the erythroid progenitor series of cells, primarily because it has the greatest number of experimentally measurable parameters at the present. This will be discussed in detail and comparisons will be made with the myeloid and lymphoid progenitor populations. The fine structure of the bone-marrow stem cell population will be examined in particular, with regard to the suitability or otherwise of the current stem cell models to explain the kinetic pattern of all the peripheral blood elements after perturbations of their steady-state values. Four different assay methods of bone-marrow stem cells have been examined with regard to the kinetic pattern following perturbation of the steady-state system, e.g. by irradiation. Basically, the stem cell assays fall into two categories: those depending on grafting haemopoietic cells into suitably treated recipients, and those in which recovery of the population is allowed in the animal in which the perturbation was produced, without handling the cells. Evidence is accumulating which indicates that in the grafting techniques, a selective loss of stem cells may occur, . especially stem cells in cell cycle, hence in early stages of recovery of the population unduly low numerical values might be noted. In view of this observation, the concept of the colony-forming cell may have to be revised and instead the colony-forming property of the stem cell introduced. (author)

  16. THE PROGENITOR OF THE TYPE IIb SN 2008ax REVISITED

    Energy Technology Data Exchange (ETDEWEB)

    Folatelli, Gastón; Bersten, Melina C.; Benvenuto, Omar G. [Instituto de Astrofísica de La Plata (Argentina); Kuncarayakti, Hanindyo [Millennium Institute of Astrophysics (MAS), Casilla 36-D, Santiago (Chile); Maeda, Keiichi; Nomoto, Ken’ichi, E-mail: gaston@fcaglp.unlp.edu.ar [Kavli Institute for the Physics and Mathematics of the Universe (WPI), The University of Tokyo, Kashiwa, Chiba 277-8583 (Japan)

    2015-10-01

    Hubble Space Telescope observations of the site of the supernova (SN) SN 2008ax obtained in 2011 and 2013 reveal that the possible progenitor object detected in pre-explosion images was in fact multiple. Four point sources are resolved in the new, higher-resolution images. We identify one of the sources with the fading SN. The other three objects are consistent with single supergiant stars. We conclude that their light contaminated the previously identified progenitor candidate. After subtraction of these stars, the progenitor appears to be significantly fainter and bluer than previously measured. Post-explosion photometry at the SN location indicates that the progenitor object has disappeared. If single, the progenitor is compatible with a supergiant star of B to mid-A spectral type, while a Wolf–Rayet (W-R) star would be too luminous in the ultraviolet to account for the observations. Moreover, our hydrodynamical modeling shows that the pre-explosion mass was 4–5 M{sub ⊙} and the radius was 30–50 R{sub ⊙}, which is incompatible with a W-R progenitor. We present a possible interacting binary progenitor computed with our evolutionary models that reproduces all the observational evidence. A companion star as luminous as an O9–B0 main-sequence star may have remained after the explosion.

  17. Ionizing radiation induces apoptosis in hematopoietic stem and progenitor cells

    International Nuclear Information System (INIS)

    Meng, A.; Zhou, D.; Geiger, H.; Zant, G.V.

    2003-01-01

    The aims of this study was to determine if ionizing radiation (IR) induces apoptosis in hematopoietic stem (HSC) and progenitor cells. Lin-cells were isolated from mouse bone marrow (BM) and pretreated with vehicle or 100 μM z-VAD 1 h prior to exposure to 4 Gy IR. The apoptotic and/or necrotic responses of these cells to IR were analyzed by measuring the annexin V and/or 7-AAD staining in HSC and progenitor populations using flow cytometry, and hematopoietic function of these cells was determined by CAFC assay. Exposure of Lin-cells to IR selectively decreased the numbers of HSC and progenitors in association with an increase in apoptosis in a time-dependent manner. Pretreatment of Lin- cells with z-VAD significantly inhibited IR-induced apoptosis and the decrease in the numbers of HSC and progenitors. However, IR alone or in combination with z-VAD did not lead to a significant increase in necrotic cell death in either HSC or progenitors. In addition, pretreatment of BM cells with z-VAD significantly attenuated IR-induced reduction in the frequencies of day-7, -28 and -35 CAFC. Exposure of HSC and progenitors to IR induces apoptosis. The induction of HSC and progenitor apoptosis contributes to IR-induced suppression of their hematopoietic function

  18. THE PROGENITOR OF THE TYPE IIb SN 2008ax REVISITED

    International Nuclear Information System (INIS)

    Folatelli, Gastón; Bersten, Melina C.; Benvenuto, Omar G.; Kuncarayakti, Hanindyo; Maeda, Keiichi; Nomoto, Ken’ichi

    2015-01-01

    Hubble Space Telescope observations of the site of the supernova (SN) SN 2008ax obtained in 2011 and 2013 reveal that the possible progenitor object detected in pre-explosion images was in fact multiple. Four point sources are resolved in the new, higher-resolution images. We identify one of the sources with the fading SN. The other three objects are consistent with single supergiant stars. We conclude that their light contaminated the previously identified progenitor candidate. After subtraction of these stars, the progenitor appears to be significantly fainter and bluer than previously measured. Post-explosion photometry at the SN location indicates that the progenitor object has disappeared. If single, the progenitor is compatible with a supergiant star of B to mid-A spectral type, while a Wolf–Rayet (W-R) star would be too luminous in the ultraviolet to account for the observations. Moreover, our hydrodynamical modeling shows that the pre-explosion mass was 4–5 M ⊙ and the radius was 30–50 R ⊙ , which is incompatible with a W-R progenitor. We present a possible interacting binary progenitor computed with our evolutionary models that reproduces all the observational evidence. A companion star as luminous as an O9–B0 main-sequence star may have remained after the explosion

  19. Evaluation of the size segregation of elemental carbon (EC emission in Europe: influence on the simulation of EC long-range transportation

    Directory of Open Access Journals (Sweden)

    Y. Chen

    2016-02-01

    Full Text Available Elemental Carbon (EC has a significant impact on human health and climate change. In order to evaluate the size segregation of EC emission in the EUCAARI inventory and investigate its influence on the simulation of EC long-range transportation in Europe, we used the fully coupled online Weather Research and Forecasting/Chemistry model (WRF-Chem at a resolution of 2 km focusing on a region in Germany, in conjunction with a high-resolution EC emission inventory. The ground meteorology conditions, vertical structure and wind pattern were well reproduced by the model. The simulations of particle number and/or mass size distributions were evaluated with observations at the central European background site Melpitz. The fine mode particle concentration was reasonably well simulated, but the coarse mode was substantially overestimated by the model mainly due to the plume with high EC concentration in coarse mode emitted by a nearby point source. The comparisons between simulated EC and Multi-angle Absorption Photometers (MAAP measurements at Melpitz, Leipzig-TROPOS and Bösel indicated that the coarse mode EC (ECc emitted from the nearby point sources might be overestimated by a factor of 2–10. The fraction of ECc was overestimated in the emission inventory by about 10–30 % for Russia and 5–10 % for Eastern Europe (e.g., Poland and Belarus. This incorrect size-dependent EC emission results in a shorter atmospheric life time of EC particles and inhibits the long-range transport of EC. A case study showed that this effect caused an underestimation of 20–40 % in the EC mass concentration in Germany under eastern wind pattern.

  20. Relationship between spontaneous γH2AX foci formation and progenitor functions in circulating hematopoietic stem and progenitor cells among atomic-bomb survivors.

    Science.gov (United States)

    Kajimura, Junko; Kyoizumi, Seishi; Kubo, Yoshiko; Misumi, Munechika; Yoshida, Kengo; Hayashi, Tomonori; Imai, Kazue; Ohishi, Waka; Nakachi, Kei; Weng, Nan-Ping; Young, Lauren F; Shieh, Jae-Hung; Moore, Malcolm A; van den Brink, Marcel R M; Kusunoki, Yoichiro

    2016-05-01

    Accumulated DNA damage in hematopoietic stem cells is a primary mechanism of aging-associated dysfunction in human hematopoiesis. About 70 years ago, atomic-bomb (A-bomb) radiation induced DNA damage and functional decreases in the hematopoietic system of A-bomb survivors in a radiation dose-dependent manner. The peripheral blood cell populations then recovered to a normal range, but accompanying cells derived from hematopoietic stem cells still remain that bear molecular changes possibly caused by past radiation exposure and aging. In the present study, we evaluated radiation-related changes in the frequency of phosphorylated (Ser-139) H2AX (γH2AX) foci formation in circulating CD34-positive/lineage marker-negative (CD34+Lin-) hematopoietic stem and progenitor cells (HSPCs) among 226Hiroshima A-bomb survivors. An association between the frequency of γH2AX foci formation in HSPCs and the radiation dose was observed, but the γH2AX foci frequency was not significantly elevated by past radiation. We found a negative correlation between the frequency of γH2AX foci formation and the length of granulocyte telomeres. A negative interaction effect between the radiation dose and the frequency of γH2AX foci was suggested in a proportion of a subset of HSPCs as assessed by the cobblestone area-forming cell assay (CAFC), indicating that the self-renewability of HSPCs may decrease in survivors who were exposed to a higher radiation dose and who had more DNA damage in their HSPCs. Thus, although many years after radiation exposure and with advancing age, the effect of DNA damage on the self-renewability of HSPCs may be modified by A-bomb radiation exposure. Copyright © 2016 Elsevier B.V. All rights reserved.

  1. "EC to go" takes off at Maryland sites.

    Science.gov (United States)

    2000-03-01

    Baltimore-based Planned Parenthood of Maryland and the Baltimore City Health Department have joined forces in "EC to Go," which distributes free emergency contraceptive pills (ECPs) through the seven affiliate sites of Planned Parenthood and the three family planning centers of the city. The distribution program was started in October 1999 and funds were provided by an undisclosed area foundation. Although the program is still in its infancy, it has recorded some 800 prescriptions of ECPs in the last fiscal year, and 600 prescriptions have been logged in just the first 6 months of the current fiscal year. To inform the public about the program, Planned Parenthood developed newspaper advertisements, a 60-second radio spot, and coupon distributions, all of which emphasize the fact that emergency contraception is a higher dose of birth control, which can prevent pregnancy if taken within 72 hours of unprotected sex.

  2. Environmental policies and regulations on the electricity market in EC

    International Nuclear Information System (INIS)

    Boucher, J.

    1990-01-01

    The use of natural resources like ground and water, the emissions of noxious gases and particles, the thermal rejections, the production of waste and the noise are the main environmental impacts associated with the production of electricity. Emission regulations and standards have been selected so far by the European Community (EC) and the different Member States to deal with the abatement problem. They simply consist in defining some ceilings or regulations for the different types of noxious gases and particles which are emitted by power plants and forcing the power generation companies to comply with these norms within some pre-determined time period. They can also take the form of a general constraint on the sulfur content of the fuel, for example. Tables of emission limits for various gases from existing and new plants are presented. 3 figs., 8 tabs

  3. Should there be an EC carbon/energy tax?

    International Nuclear Information System (INIS)

    Helm, D.

    1993-01-01

    The European Commission has suggested that an EC-wide carbon/energy tax should be introduced to meet the Community's commitment to reduce emissions of carbon dioxide, and to bear down on criticisms of sulphur dioxide and other harmful energy externalities. The UK government to data has pursued at best an agnostic position. Although the White Paper, This Common Inheritance, (DOE, 1990) gave prominence to the use of market-based instruments in advancing environmental objectives, their use in air pollution has been limited to the tax differential on unleaded petrol, the commitment to increase petrol tax in real terms in successive budgets and the imposition of VAT on domestic fuel in a phased series of stages. (Author)

  4. Should there be an EC carbon/energy tax

    Energy Technology Data Exchange (ETDEWEB)

    Helm, D. (New College, Oxford (United Kingdom))

    1993-12-01

    The European Commission has suggested that an EC-wide carbon/energy tax should be introduced to meet the Community's commitment to reduce emissions of carbon dioxide, and to bear down on criticisms of sulphur dioxide and other harmful energy externalities. The UK government to data has pursued at best an agnostic position. Although the White Paper, This Common Inheritance, (DOE, 1990) gave prominence to the use of market-based instruments in advancing environmental objectives, their use in air pollution has been limited to the tax differential on unleaded petrol, the commitment to increase petrol tax in real terms in successive budgets and the imposition of VAT on domestic fuel in a phased series of stages. (Author)

  5. Integrating Amazon EC2 with the CMS production framework

    International Nuclear Information System (INIS)

    Melo, Andrew; Sheldon, Paul

    2012-01-01

    As cloud middleware and cloud providers have become more robust, various experiments with experience in Grid submission have begun to investigate the possibility of taking previously Grid-Enabled applications and making them compatible with Cloud Computing. Successful implementation will allow for dynamic scaling of the available hardware resources, providing access to peak-load handling capabilities and possibly resulting in lower costs to the experiment. Here we discuss current work within the CMS collaboration at the LHC to both perform computation on EC2, both for production and analysis use-cases. We also discuss break-even points between dedicated and cloud resources using real-world costs derived from a CMS site.

  6. NATIONAL MINORITIES IN THE LAW OF THE EC/EU

    Directory of Open Access Journals (Sweden)

    Daniel Šmihula

    2008-09-01

    Full Text Available In the law of the EC/EU the protection of national minorities is still a marginal matter. The EU has relied on general international law and on a European regional system of international law and, in case of necessity, accepted their norms. But in the 1990s there began a process of “de-economisation of the European integration” and the importance of national minorities became higher. Protection of the national minorities has not become a generally accepted legally binding principle of the EU, although in several legal acts issues of national minorities are mentioned. On the other hand, the political relevance of national minorities´ protection is very high. The importance of protection of national minorities in future will probably grow. It is a result of the adoption of the Charter of Fundamental Rights of the EU (2000 and of the discussions regarding the European constitution and the Treaty of Lisbon.

  7. Integrating Amazon EC2 with the CMS Production Framework

    CERN Document Server

    Melo, Andrew Malone

    2011-01-01

    As cloud middleware and cloud providers have become more robust, various experiments with experience in Grid submission have begun to investigate the possibility of taking previously Grid-Enabled applications and making them compatible with Cloud Computing. Successful implementation will allow for dynamic scaling of the available hardware resources, providing access to peak-load handling capabilities and possibly resulting in lower costs to the experiment. Here we discuss current work within the CMS collaboration at the LHC to both perform computation on EC2, both for production and analysis use-cases. We also discuss break-even points between dedicated and cloud resources using real-world costs derived from a CMS site.

  8. The EC CAST project (carbon-14 source term)

    International Nuclear Information System (INIS)

    Williams, S. J.

    2015-01-01

    Carbon-14 is a key radionuclide in the assessment of the safety of underground geological disposal facilities for radioactive wastes. It is possible for carbon-14 to be released from waste packages in a variety of chemical forms, both organic and inorganic, and as dissolved or gaseous species The EC CAST (CArbon-14 Source Term) project aims to develop understanding of the generation and release of carbon-14 from radioactive waste materials under conditions relevant to packaging and disposal. It focuses on the release of carbon-14 from irradiated metals (steels and zirconium alloys), from irradiated graphite and from spent ion-exchange resins. The CAST consortium brings together 33 partners. CAST commenced in October 2013 and this paper describes progress to March 2015. The main activities during this period were reviews of the current status of knowledge, the identification and acquisition of suitable samples and the design of experiments and analytical procedures. (authors)

  9. Completely ES cell-derived mice produced by tetraploid complementation using inner cell mass (ICM deficient blastocysts.

    Directory of Open Access Journals (Sweden)

    Duancheng Wen

    Full Text Available Tetraploid complementation is often used to produce mice from embryonic stem cells (ESCs by injection of diploid (2n ESCs into tetraploid (4n blastocysts (ESC-derived mice. This method has also been adapted to mouse cloning and the derivation of mice from induced pluripotent stem (iPS cells. However, the underlying mechanism(s of the tetraploid complementation remains largely unclear. Whether this approach can give rise to completely ES cell-derived mice is an open question, and has not yet been unambiguously proven. Here, we show that mouse tetraploid blastocysts can be classified into two groups, according to the presence or absence of an inner cell mass (ICM. We designate these as type a (presence of ICM at blastocyst stage or type b (absence of ICM. ESC lines were readily derived from type a blastocysts, suggesting that these embryos retain a pluripotent epiblast compartment; whereas the type b blastocysts possessed very low potential to give rise to ESC lines, suggesting that they had lost the pluripotent epiblast. When the type a blastocysts were used for tetraploid complementation, some of the resulting mice were found to be 2n/4n chimeric; whereas when type b blastocysts were used as hosts, the resulting mice are all completely ES cell-derived, with the newborn pups displaying a high frequency of abdominal hernias. Our results demonstrate that completely ES cell-derived mice can be produced using ICM-deficient 4n blastocysts, and provide evidence that the exclusion of tetraploid cells from the fetus in 2n/4n chimeras can largely be attributed to the formation of ICM-deficient blastocysts.

  10. Cell-Derived Microparticles in Patients with Type 2 Diabetes Mellitus: a Systematic Review and Meta-Analysis.

    Science.gov (United States)

    Li, Sheyu; Wei, Jia; Zhang, Chenghui; Li, Xiaodan; Meng, Wentong; Mo, Xianming; Zhang, Qianying; Liu, Qilin; Ren, Kaiyun; Du, Rong; Tian, Haoming; Li, Jianwei

    2016-01-01

    The aim of this study was to assess the association between circulating cell-derived microparticles (MPs) and type 2 diabetes mellitus (T2DM). A literature search was performed systematically in PubMed and Embase to identify available case-control or cross-sectional studies that compared different types of cell-derived MPs in patients with T2DM and non-diabetic controls. Pooled standardized mean differences (SMDs) of each MP type were pooled using meta-analysis. Forty-eight studies involving 2,460 patients with T2DM and 1,880 non-diabetic controls were included for systematic review and 34 of which were included for quantitative study by meta-analysis. In the overall analysis, the levels of circulating total MPs (TMPs), platelet-derived MPs (PMPs), monocyte-derived MPs (MMPs) and endothelium-derived MPs (EMPs) were significantly higher in T2DM patients than those in controls (TMPs: SMD, 0.64; 95%CI, 0.12∼1.15; P=0.02; PMPs: SMD, 1.19; 95%CI, 0.88∼1.50; P <0.00001; MMPs: SMD, 0.92; 95%CI, 0.66∼1.17; P <0.00001; EMPs: SMD, 0.73; 95%CI, 0.50∼0.96; P <0.00001). Meanwhile, no significant difference was shown in leukocyte-derived MPs (LMPs) level between diabetic and non-diabetic groups (SMD, 0.37; 95%CI, -0.15∼0.89; P=0.17). The counts of TMPs, PMPs, MMPs and EMPs elevated in patients with T2DM. And cell-derived MPs may play a role in the pathogenesis of T2DM. © 2016 The Author(s) Published by S. Karger AG, Basel.

  11. Evaluation of reliability of EC inspection of VVER SG tubes

    International Nuclear Information System (INIS)

    Stanic, D.

    2001-01-01

    Evaluation of eddy current data collected during inspection of VVER steam generators is very complex task because of numerous parameters which have affect on eddy current signals. That was the reason that recently ago INETEC has started related scientific project in order to evaluate the reliability of eddy current (EC) inspection of VVER steam generator (SG) tubing. In the scope of project the following objectives will be investigated: 1. Determination of POD (Probability of detection) of various types degradation cracks, where their basic parameters are variables (basic parameters are depth, length, width, orientation, number) on three different sets of tubes (clean ideal tubes, tubes with pilgering, tubes electroplated with copper) 2. Sizing quality (accuracy, repeatability) (same data sets as defined in 1.) 3. Effect of fill factor on POD and sizing quality. 4. Effect of tube bends on POD and sizing quality. 5. Effect of other tube geometry variations on POD and sizing quality (tube ovality, transition zone region, expanded (rolled) part of tube, dents, dings). Investigation will start with bobbin probe technique which is the most used technique for general purpose VVER tube examination. Since INETEC is the only world company which successfully developed and applied rotating probe technique for VVER SG tubes, scope of the project will be extended on rotating probe technique utilizing 'pancake' and 'point' coil. Method reliability will be investigated first on the huge set of EDM notches representing various defect morphologies and simulating different factors, and the second part will be investigated on sets of degradation defects obtained by artificial corrosion. In the scope of the project the measures for enhancing the method reliability have to be determined. This considers the proper definition of parameters of examination system, as well as establishment of the suitable analysis procedures. This article presents the temporary results of the first part of

  12. Completion Report for Well ER-EC-1

    International Nuclear Information System (INIS)

    Townsend, M.J.

    2000-01-01

    Well ER-EC-1 was drilled for the U.S. Department of Energy, Nevada Operations Office in support of the Nevada Environmental Restoration Project at the Nevada Test Site, Nye County, Nevada. This well was drilled in the spring of 1999 as part of the U.S. Department of Energy's hydrogeologic investigation well program in the Western Pahute Mesa - Oasis Valley region just west of the Test Site. A 44.5-centimeter surface hole was drilled and cased off to the depth 675.1 meters below the surface. The hole diameter was then decreased to 31.1 centimeters for drilling to a total depth of 1,524.0 meters. A preliminary composite, static, water level was measured at the depth of approximately 566.3 meters prior to installation of the completion string. One completion string with three isolated, slotted intervals was installed in the well. Detailed lithologic descriptions with preliminary stratigraphic assignments are included in the report. These are based on composite drill cuttings collected every 3 meters and 31 sidewall samples taken at various depths below 680 meters, supplemented by geophysical log data. Detailed chemical and mineralogical studies of rock samples are in progress. The well penetrated Tertiary-age lava and tuff of the Timber Mountain Group, the Paintbrush Group, the Calico Hills Formation, the Crater Flat Group, and the Volcanics of Quartz Mountain. The preliminary geologic interpretation of data from Well ER-EC-1 indicates the presence of a structural trough or bench filled with a thick section of post-Rainier Mesa lava. These data also suggest that this site is located on a buried structural ridge that may separate the Silent Canyon and Timber Mountain caldera complexes

  13. Developmental regulation of ecdysone receptor (EcR and EcR-controlled gene expression during pharate-adult development of honeybees (Apis mellifera.

    Directory of Open Access Journals (Sweden)

    Tathyana Rachel Palo Mello

    2014-12-01

    Full Text Available Major developmental transitions in multicellular organisms are driven by steroid hormones. In insects, these, together with juvenile hormone (JH, control development, metamorphosis, reproduction and aging, and are also suggested to play an important role in caste differentiation of social insects. Here, we aimed to determine how EcR transcription and ecdysteroid titers are related during honeybee postembryonic development and what may actually be the role of EcR in caste development of this social insect. In addition, we expected that knocking-down EcR gene expression would give us information on the participation of the respective protein in regulating downstream targets of EcR. We found that in Apis mellifera females, EcR-A is the predominantly expressed variant in postembryonic development, while EcR-B transcript levels are higher in embryos, indicating an early developmental switch in EcR function. During larval and pupal stages, EcR-B expression levels are very low, while EcR-A transcripts are more variable and abundant in workers compared to queens. Strikingly, these transcript levels are opposite to the ecdysteroid titer profile. 20-hydroxyecdysone (20E application experiments revealed that low 20E levels induce EcR expression during development, whereas high ecdysteroid titers seem to be repressive. By means of RNAi-mediated knockdown (KD of both EcR transcript variants we detected the differential expression of 234 poly-A+ transcripts encoding genes such as CYPs, MRJPs and certain hormone response genes (Kr-h1 and ftz-f1. EcR-KD also promoted the differential expression of 70 miRNAs, including highly conserved ones (e.g. miR-133 and miR-375, as well honeybee-specific ones (e.g. miR-3745 and miR-3761. Our results put in evidence a broad spectrum of EcR-controlled gene expression during postembryonic development of honeybees, revealing new facets of EcR biology in this social insect.

  14. SAFETY OF CELL-DERIVED SUBUNIT ADJUVANTED INFLUENZA VACCINE FOR CHILDREN VACCINATION: DOUBLE-BLIND RANDOMIZED CLINICAL TRIAL

    Directory of Open Access Journals (Sweden)

    S.M. Kharit

    2010-01-01

    Full Text Available This article presents the safety data for cell-derived inactivated subunit adjuvanted influenza vaccine «Grippol Neo» in children 3–17 years old in comparison with reference egg-derived inactivated subunit vaccine «Grippol plus». Good test vaccine tolerability and high efficacy profile is demonstrated. Based on the results obtained vaccine «Grippol Neo» is recommended for mass influenza prophylaxis in pediatry, including National Immunization Schedule.Key words: children, influenza, vaccination, «Grippol Neo».(Voprosy sovremennoi pediatrii — Current Pediatrics. – 2010;9(4:44-49

  15. Generation and characterization of human iPSC line generated from mesenchymal stem cells derived from adipose tissue.

    Science.gov (United States)

    Zapata-Linares, Natalia; Rodriguez, Saray; Mazo, Manuel; Abizanda, Gloria; Andreu, Enrique J; Barajas, Miguel; Prosper, Felipe; Rodriguez-Madoz, Juan R

    2016-01-01

    In this work, mesenchymal stem cells derived from adipose tissue (ADSCs) were used for the generation of the human-induced pluripotent stem cell line G15.AO. Cell reprogramming was performed using retroviral vectors containing the Yamanaka factors, and the generated G15.AO hiPSC line showed normal karyotype, silencing of the exogenous reprogramming factors, induction of the typical pluripotency-associated markers, alkaline phosphatase enzymatic activity, and in vivo and in vitro differentiation ability to the three germ layers. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.

  16. Enteric nervous system specific deletion of Foxd3 disrupts glial cell differentiation and activates compensatory enteric progenitors.

    Science.gov (United States)

    Mundell, Nathan A; Plank, Jennifer L; LeGrone, Alison W; Frist, Audrey Y; Zhu, Lei; Shin, Myung K; Southard-Smith, E Michelle; Labosky, Patricia A

    2012-03-15

    The enteric nervous system (ENS) arises from the coordinated migration, expansion and differentiation of vagal and sacral neural crest progenitor cells. During development, vagal neural crest cells enter the foregut and migrate in a rostro-to-caudal direction, colonizing the entire gastrointestinal tract and generating the majority of the ENS. Sacral neural crest contributes to a subset of enteric ganglia in the hindgut, colonizing the colon in a caudal-to-rostral wave. During this process, enteric neural crest-derived progenitors (ENPs) self-renew and begin expressing markers of neural and glial lineages as they populate the intestine. Our earlier work demonstrated that the transcription factor Foxd3 is required early in neural crest-derived progenitors for self-renewal, multipotency and establishment of multiple neural crest-derived cells and structures including the ENS. Here, we describe Foxd3 expression within the fetal and postnatal intestine: Foxd3 was strongly expressed in ENPs as they colonize the gastrointestinal tract and was progressively restricted to enteric glial cells. Using a novel Ednrb-iCre transgene to delete Foxd3 after vagal neural crest cells migrate into the midgut, we demonstrated a late temporal requirement for Foxd3 during ENS development. Lineage labeling of Ednrb-iCre expressing cells in Foxd3 mutant embryos revealed a reduction of ENPs throughout the gut and loss of Ednrb-iCre lineage cells in the distal colon. Although mutant mice were viable, defects in patterning and distribution of ENPs were associated with reduced proliferation and severe reduction of glial cells derived from the Ednrb-iCre lineage. Analyses of ENS-lineage and differentiation in mutant embryos suggested activation of a compensatory population of Foxd3-positive ENPs that did not express the Ednrb-iCre transgene. Our findings highlight the crucial roles played by Foxd3 during ENS development including progenitor proliferation, neural patterning, and glial

  17. Extensive characterization and comparison of endothelial cells derived from dermis and adipose tissue : Potential use in tissue engineering

    NARCIS (Netherlands)

    Monsuur, H.N.; Weijers, E.M.; Niessen, F.B.; Gefen, A.; Koolwijk, P.; Gibbs, S.; van den Broek, L.J.

    2016-01-01

    Tissue-engineered constructs need to become quickly vascularized in order to ensure graft take. One way of achieving this is to incorporate endothelial cells (EC) into the construct. The adipose tissue stromal vascular fraction (adipose-SVF) might provide an alternative source for endothelial cells

  18. Increased IgG on cell-derived plasma microparticles in systemic lupus erythematosus is associated with autoantibodies and complement activation

    DEFF Research Database (Denmark)

    Nielsen, Christoffer T; Østergaard, Ole; Stener, Line

    2012-01-01

    To quantify immunoglobulin and C1q on circulating cell-derived microparticles (MPs) in patients with systemic lupus erythematosus (SLE) and to determine whether immunoglobulin and C1q levels are correlated with clinical and serologic parameters.......To quantify immunoglobulin and C1q on circulating cell-derived microparticles (MPs) in patients with systemic lupus erythematosus (SLE) and to determine whether immunoglobulin and C1q levels are correlated with clinical and serologic parameters....

  19. Reporter-Based Isolation of Developmental Myogenic Progenitors

    Directory of Open Access Journals (Sweden)

    Eyemen Kheir

    2018-04-01

    Full Text Available The formation and activity of mammalian tissues entail finely regulated processes, involving the concerted organization and interaction of multiple cell types. In recent years the prospective isolation of distinct progenitor and stem cell populations has become a powerful tool in the hands of developmental biologists and has rendered the investigation of their intrinsic properties possible. In this protocol, we describe how to purify progenitors with different lineage history and degree of differentiation from embryonic and fetal skeletal muscle by fluorescence-activated cell sorting (FACS. The approach takes advantage of a panel of murine strains expressing fluorescent reporter genes specifically in the myogenic progenitors. We provide a detailed description of the dissection procedures and of the enzymatic dissociation required to maximize the yield of mononucleated cells for subsequent FACS-based purification. The procedure takes ~6–7 h to complete and allows for the isolation and the subsequent molecular and phenotypic characterization of developmental myogenic progenitors.

  20. Endothelial progenitor cells in chronic obstructive pulmonary disease and emphysema

    Science.gov (United States)

    Tracy, Russell P.; Parikh, Megha A.; Hoffman, Eric A.; Shimbo, Daichi; Austin, John H. M.; Smith, Benjamin M.; Hueper, Katja; Vogel-Claussen, Jens; Lima, Joao; Gomes, Antoinette; Watson, Karol; Kawut, Steven; Barr, R. Graham

    2017-01-01

    Endothelial injury is implicated in the pathogenesis of COPD and emphysema; however the role of endothelial progenitor cells (EPCs), a marker of endothelial cell repair, and circulating endothelial cells (CECs), a marker of endothelial cell injury, in COPD and its subphenotypes is unresolved. We hypothesized that endothelial progenitor cell populations would be decreased in COPD and emphysema and that circulating endothelial cells would be increased. Associations with other subphenotypes were examined. The Multi-Ethnic Study of Atherosclerosis COPD Study recruited smokers with COPD and controls age 50–79 years without clinical cardiovascular disease. Endothelial progenitor cell populations (CD34+KDR+ and CD34+KDR+CD133+ cells) and circulating endothelial cells (CD45dimCD31+CD146+CD133-) were measured by flow cytometry. COPD was defined by standard spirometric criteria. Emphysema was assessed qualitatively and quantitatively on CT. Full pulmonary function testing and expiratory CTs were measured in a subset. Among 257 participants, both endothelial progenitor cell populations, and particularly CD34+KDR+ endothelial progenitor cells, were reduced in COPD. The CD34+KDR+CD133+ endothelial progenitor cells were associated inversely with emphysema extent. Both endothelial progenitor cell populations were associated inversely with extent of panlobular emphysema and positively with diffusing capacity. Circulating endothelial cells were not significantly altered in COPD but were inversely associated with pulmonary microvascular blood flow on MRI. There was no consistent association of endothelial progenitor cells or circulating endothelial cells with measures of gas trapping. These data provide evidence that endothelial repair is impaired in COPD and suggest that this pathological process is specific to emphysema. PMID:28291826

  1. Functional Recovery Secondary to Neural Stem/Progenitor Cells Transplantation Combined with Treadmill Training in Mice with Chronic Spinal Cord Injury

    DEFF Research Database (Denmark)

    Tashiro, Syoichi; Nishimura, Soraya; Iwai, Hiroki

    are chiefly developed to improve the effect of regenerative therapy for this refractory state, physical training also have attracted the attention as a desirable candidate to combine with cell transplantation. Recently, we have reported that the addition of treadmill training enhances the effect of NS...... in the combined therapy group. Further investigation revealed that NS/PC transplantation improved spinal conductivity and central pattern generator activity, and that training promoted the appropriate inhibitory motor control including spasticity. The combined therapy enhanced these independent effects of each......Rapid progress in stem cell medicine is being realized in neural regeneration also in spinal cord injury (SCI). Researchers have reported remarkable functional recovery with various cell sources including induced Pluripotent Stem cell derived neural stem/progenitor cells (NS/PCs), especially...

  2. Altered calcium handling and increased contraction force in human embryonic stem cell derived cardiomyocytes following short term dexamethasone exposure

    Energy Technology Data Exchange (ETDEWEB)

    Kosmidis, Georgios; Bellin, Milena; Ribeiro, Marcelo C.; Meer, Berend van; Ward-van Oostwaard, Dorien [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands); Passier, Robert [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands); MIRA, University of Twente (Netherlands); Tertoolen, Leon G.J.; Mummery, Christine L. [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands); Casini, Simona, E-mail: s.casini@amc.uva.nl [Department of Anatomy and Embryology, Leiden University Medical Center, Leiden (Netherlands)

    2015-11-27

    One limitation in using human pluripotent stem cell derived cardiomyocytes (hPSC-CMs) for disease modeling and cardiac safety pharmacology is their immature functional phenotype compared with adult cardiomyocytes. Here, we report that treatment of human embryonic stem cell derived cardiomyocytes (hESC-CMs) with dexamethasone, a synthetic glucocorticoid, activated glucocorticoid signaling which in turn improved their calcium handling properties and contractility. L-type calcium current and action potential properties were not affected by dexamethasone but significantly faster calcium decay, increased forces of contraction and sarcomeric lengths, were observed in hESC-CMs after dexamethasone exposure. Activating the glucocorticoid pathway can thus contribute to mediating hPSC-CMs maturation. - Highlights: • Dexamethasone accelerates Ca{sup 2+} transient decay in hESC-CMs. • Dexamethasone enhances SERCA and NCX function in hESC-CMs. • Dexamethasone increases force of contraction and sarcomere length in hESC-CMs. • Dexamethasone does not alter I{sub Ca,L} and action potential characteristics in hESC-CMs.

  3. Defined MicroRNAs Induce Aspects of Maturation in Mouse and Human Embryonic-Stem-Cell-Derived Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Desy S. Lee

    2015-09-01

    Full Text Available Pluripotent-cell-derived cardiomyocytes have great potential for use in research and medicine, but limitations in their maturity currently constrain their usefulness. Here, we report a method for improving features of maturation in murine and human embryonic-stem-cell-derived cardiomyocytes (m/hESC-CMs. We found that coculturing m/hESC-CMs with endothelial cells improves their maturity and upregulates several microRNAs. Delivering four of these microRNAs, miR-125b-5p, miR-199a-5p, miR-221, and miR-222 (miR-combo, to m/hESC-CMs resulted in improved sarcomere alignment and calcium handling, a more negative resting membrane potential, and increased expression of cardiomyocyte maturation markers. Although this could not fully phenocopy all adult cardiomyocyte characteristics, these effects persisted for two months following delivery of miR-combo. A luciferase assay demonstrated that all four miRNAs target ErbB4, and siRNA knockdown of ErbB4 partially recapitulated the effects of miR-combo. In summary, a combination of miRNAs induced via endothelial coculture improved ESC-CM maturity, in part through suppression of ErbB4 signaling.

  4. Therapeutic potential of CAR-T cell-derived exosomes: a cell-free modality for targeted cancer therapy.

    Science.gov (United States)

    Tang, Xiang-Jun; Sun, Xu-Yong; Huang, Kuan-Ming; Zhang, Li; Yang, Zhuo-Shun; Zou, Dan-Dan; Wang, Bin; Warnock, Garth L; Dai, Long-Jun; Luo, Jie

    2015-12-29

    Chimeric antigen receptor (CAR)-based T-cell adoptive immunotherapy is a distinctively promising therapy for cancer. The engineering of CARs into T cells provides T cells with tumor-targeting capabilities and intensifies their cytotoxic activity through stimulated cell expansion and enhanced cytokine production. As a novel and potent therapeutic modality, there exists some uncontrollable processes which are the potential sources of adverse events. As an extension of this impactful modality, CAR-T cell-derived exosomes may substitute CAR-T cells to act as ultimate attackers, thereby overcoming some limitations. Exosomes retain most characteristics of parent cells and play an essential role in intercellular communications via transmitting their cargo to recipient cells. The application of CAR-T cell-derived exosomes will make this cell-based therapy more clinically controllable as it also provides a cell-free platform to diversify anticancer mediators, which responds effectively to the complexity and volatility of cancer. It is believed that the appropriate application of both cellular and exosomal platforms will make this effective treatment more practicable.

  5. Evaluation of Stem Cell-Derived Red Blood Cells as a Transfusion Product Using a Novel Animal Model.

    Science.gov (United States)

    Shah, Sandeep N; Gelderman, Monique P; Lewis, Emily M A; Farrel, John; Wood, Francine; Strader, Michael Brad; Alayash, Abdu I; Vostal, Jaroslav G

    2016-01-01

    Reliance on volunteer blood donors can lead to transfusion product shortages, and current liquid storage of red blood cells (RBCs) is associated with biochemical changes over time, known as 'the storage lesion'. Thus, there is a need for alternative sources of transfusable RBCs to supplement conventional blood donations. Extracorporeal production of stem cell-derived RBCs (stemRBCs) is a potential and yet untapped source of fresh, transfusable RBCs. A number of groups have attempted RBC differentiation from CD34+ cells. However, it is still unclear whether these stemRBCs could eventually be effective substitutes for traditional RBCs due to potential differences in oxygen carrying capacity, viability, deformability, and other critical parameters. We have generated ex vivo stemRBCs from primary human cord blood CD34+ cells and compared them to donor-derived RBCs based on a number of in vitro parameters. In vivo, we assessed stemRBC circulation kinetics in an animal model of transfusion and oxygen delivery in a mouse model of exercise performance. Our novel, chronically anemic, SCID mouse model can evaluate the potential of stemRBCs to deliver oxygen to tissues (muscle) under resting and exercise-induced hypoxic conditions. Based on our data, stem cell-derived RBCs have a similar biochemical profile compared to donor-derived RBCs. While certain key differences remain between donor-derived RBCs and stemRBCs, the ability of stemRBCs to deliver oxygen in a living organism provides support for further development as a transfusion product.

  6. Immune cell-derived c3 is required for autoimmune diabetes induced by multiple low doses of streptozotocin.

    Science.gov (United States)

    Lin, Marvin; Yin, Na; Murphy, Barbara; Medof, M Edward; Segerer, Stephan; Heeger, Peter S; Schröppel, Bernd

    2010-09-01

    The complement system contributes to autoimmune injury, but its involvement in promoting the development of autoimmune diabetes is unknown. In this study, our goal was to ascertain the role of complement C3 in autoimmune diabetes. Susceptibility to diabetes development after multiple low-dose streptozotocin treatment in wild-type (WT) and C3-deficient mice was analyzed. Bone marrow chimeras, luminex, and quantitative reverse transcription PCR assays were performed to evaluate the phenotypic and immunologic impact of C3 in the development of this diabetes model. Coincident with the induced elevations in blood glucose levels, we documented alternative pathway complement component gene expression within the islets of the diabetic WT mice. When we repeated the experiments with C3-deficient mice, we observed complete resistance to disease, as assessed by the absence of histologic insulitis and the absence of T-cell reactivity to islet antigens. Studies of WT chimeras bearing C3-deficient bone marrow cells showed that bone marrow cell-derived C3, and not serum C3, is involved in the induction of diabetes in this model. The data reveal a key role for immune cell-derived C3 in the pathogenesis of murine multiple low-dose streptozotocin-induced diabetes and support the concept that immune cell mediated diabetes is in part complement-dependent.

  7. Innate immune cell-derived microparticles facilitate hepatocarcinoma metastasis by transferring integrin α(M)β₂ to tumor cells.

    Science.gov (United States)

    Ma, Jingwei; Cai, Wenqian; Zhang, Yi; Huang, Chunmei; Zhang, Huafeng; Liu, Jing; Tang, Ke; Xu, Pingwei; Katirai, Foad; Zhang, Jianmin; He, Wei; Ye, Duyun; Shen, Guan-Xin; Huang, Bo

    2013-09-15

    Mechanisms by which tumor cells metastasize to distant organs still remain enigmatic. Immune cells have been assumed to be the root of metastasis by their fusing with tumor cells. This fusion theory, although interpreting tumor metastasis analogically and intriguingly, is arguable to date. We show in this study an alternative explanation by immune cell-derived microparticles (MPs). Upon stimulation by PMA or tumor cell-derived supernatants, immune cells released membrane-based MPs, which were taken up by H22 tumor cells, leading to tumor cell migration in vitro and metastasis in vivo. The underlying molecular basis was involved in integrin α(M)β₂ (CD11b/CD18), which could be effectively relayed from stimulated innate immune cells to MPs, then to tumor cells. Blocking either CD11b or CD18 led to significant decreases in MP-mediated tumor cell metastasis. This MP-mediated transfer of immune phenotype to tumor cells might also occur in vivo. These findings suggest that tumor cells may usurp innate immune cell phenotypes via MP pathway for their metastasis, providing new insight into tumor metastatic mechanism.

  8. Altered calcium handling and increased contraction force in human embryonic stem cell derived cardiomyocytes following short term dexamethasone exposure

    International Nuclear Information System (INIS)

    Kosmidis, Georgios; Bellin, Milena; Ribeiro, Marcelo C.; Meer, Berend van; Ward-van Oostwaard, Dorien; Passier, Robert; Tertoolen, Leon G.J.; Mummery, Christine L.; Casini, Simona

    2015-01-01

    One limitation in using human pluripotent stem cell derived cardiomyocytes (hPSC-CMs) for disease modeling and cardiac safety pharmacology is their immature functional phenotype compared with adult cardiomyocytes. Here, we report that treatment of human embryonic stem cell derived cardiomyocytes (hESC-CMs) with dexamethasone, a synthetic glucocorticoid, activated glucocorticoid signaling which in turn improved their calcium handling properties and contractility. L-type calcium current and action potential properties were not affected by dexamethasone but significantly faster calcium decay, increased forces of contraction and sarcomeric lengths, were observed in hESC-CMs after dexamethasone exposure. Activating the glucocorticoid pathway can thus contribute to mediating hPSC-CMs maturation. - Highlights: • Dexamethasone accelerates Ca"2"+ transient decay in hESC-CMs. • Dexamethasone enhances SERCA and NCX function in hESC-CMs. • Dexamethasone increases force of contraction and sarcomere length in hESC-CMs. • Dexamethasone does not alter I_C_a_,_L and action potential characteristics in hESC-CMs.

  9. Invited review: mesenchymal progenitor cells in intramuscular connective tissue development.

    Science.gov (United States)

    Miao, Z G; Zhang, L P; Fu, X; Yang, Q Y; Zhu, M J; Dodson, M V; Du, M

    2016-01-01

    The abundance and cross-linking of intramuscular connective tissue contributes to the background toughness of meat, and is thus undesirable. Connective tissue is mainly synthesized by intramuscular fibroblasts. Myocytes, adipocytes and fibroblasts are derived from a common pool of progenitor cells during the early embryonic development. It appears that multipotent mesenchymal stem cells first diverge into either myogenic or non-myogenic lineages; non-myogenic mesenchymal progenitors then develop into the stromal-vascular fraction of skeletal muscle wherein adipocytes, fibroblasts and derived mesenchymal progenitors reside. Because non-myogenic mesenchymal progenitors mainly undergo adipogenic or fibrogenic differentiation during muscle development, strengthening progenitor proliferation enhances the potential for both intramuscular adipogenesis and fibrogenesis, leading to the elevation of both marbling and connective tissue content in the resulting meat product. Furthermore, given the bipotent developmental potential of progenitor cells, enhancing their conversion to adipogenesis reduces fibrogenesis, which likely results in the overall improvement of marbling (more intramuscular adipocytes) and tenderness (less connective tissue) of meat. Fibrogenesis is mainly regulated by the transforming growth factor (TGF) β signaling pathway and its regulatory cascade. In addition, extracellular matrix, a part of the intramuscular connective tissue, provides a niche environment for regulating myogenic differentiation of satellite cells and muscle growth. Despite rapid progress, many questions remain in the role of extracellular matrix on muscle development, and factors determining the early differentiation of myogenic, adipogenic and fibrogenic cells, which warrant further studies.

  10. Biology and flow cytometry of proangiogenic hematopoietic progenitors cells.

    Science.gov (United States)

    Rose, Jonathan A; Erzurum, Serpil; Asosingh, Kewal

    2015-01-01

    During development, hematopoiesis and neovascularization are closely linked to each other via a common bipotent stem cell called the hemangioblast that gives rise to both hematopoietic cells and endothelial cells. In postnatal life, this functional connection between the vasculature and hematopoiesis is maintained by a subset of hematopoietic progenitor cells endowed with the capacity to differentiate into potent proangiogenic cells. These proangiogenic hematopoietic progenitors comprise a specific subset of bone marrow (BM)-derived cells that homes to sites of neovascularization and possess potent paracrine angiogenic activity. There is emerging evidence that this subpopulation of hematopoietic progenitors plays a critical role in vascular health and disease. Their angiogenic activity is distinct from putative "endothelial progenitor cells" that become structural cells of the endothelium by differentiation into endothelial cells. Proangiogenic hematopoietic progenitor cell research requires multidisciplinary expertise in flow cytometry, hematology, and vascular biology. This review provides a comprehensive overview of proangiogenic hematopoietic progenitor cell biology and flow cytometric methods to detect these cells in the peripheral blood circulation and BM. © 2014 International Society for Advancement of Cytometry.

  11. LINKING TYPE Ia SUPERNOVA PROGENITORS AND THEIR RESULTING EXPLOSIONS

    International Nuclear Information System (INIS)

    Foley, Ryan J.; Kirshner, Robert P.; Simon, Joshua D.; Burns, Christopher R.; Gal-Yam, Avishay; Hamuy, Mario; Morrell, Nidia I.; Phillips, Mark M.; Shields, Gregory A.; Sternberg, Assaf

    2012-01-01

    Comparing the ejecta velocities at maximum brightness and narrow circumstellar/interstellar Na D absorption line profiles of a sample of 23 Type Ia supernovae (SNe Ia), we determine that the properties of SN Ia progenitor systems and explosions are intimately connected. As demonstrated by Sternberg et al., half of all SNe Ia with detectable Na D absorption at the host-galaxy redshift in high-resolution spectroscopy have Na D line profiles with significant blueshifted absorption relative to the strongest absorption component, which indicates that a large fraction of SN Ia progenitor systems have strong outflows. In this study, we find that SNe Ia with blueshifted circumstellar/interstellar absorption systematically have higher ejecta velocities and redder colors at maximum brightness relative to the rest of the SN Ia population. This result is robust at a 98.9%-99.8% confidence level, providing the first link between the progenitor systems and properties of the explosion. This finding is further evidence that the outflow scenario is the correct interpretation of the blueshifted Na D absorption, adding additional confirmation that some SNe Ia are produced from a single-degenerate progenitor channel. An additional implication is that either SN Ia progenitor systems have highly asymmetric outflows that are also aligned with the SN explosion or SNe Ia come from a variety of progenitor systems where SNe Ia from systems with strong outflows tend to have more kinetic energy per unit mass than those from systems with weak or no outflows.

  12. Depletion of NAD pool contributes to impairment of endothelial progenitor cell mobilization in diabetes.

    Science.gov (United States)

    Wang, Pei; Yang, Xi; Zhang, Zheng; Song, Jie; Guan, Yun-Feng; Zou, Da-Jin; Miao, Chao-Yu

    2016-06-01

    The impaired mobilization of endothelial progenitor cells (EPCs) from bone marrow (BM) critically contributes to the diabetes-associated vascular complications. Here, we investigated the relationship between the nicotinamide phosphoribosyltransferase (NAMPT)-controlled nicotinamide adenine dinucleotide (NAD) metabolism and the impaired mobilization of BM-derived EPCs in diabetic condition. The NAMPT-NAD pool in BM and BM-derived EPCs in wild-type (WT) and diabetic db/db mice was determined. Nicotinamide, a natural substrate for NAD biosynthesis, was administrated for 2weeks in db/db mice to examine the influence of enhancing NAD pool on BM and blood EPCs number. The modulations of stromal cell-derived factor-1α (SDF-1α) and endothelial nitric oxide synthase (eNOS) protein in BM were measured using immunoblotting. The EPCs intracellular NAMPT level and NAD concentration, as well as the blood EPCs number, were compared between 9 healthy people and 16 patients with type 2 diabetes mellitus (T2DM). The T2DM patients were treated with nicotinamide for two weeks and then the blood EPCs number was determined. Moreover, the association between blood EPCs numbers and EPCs intracellular NAD(+)/NAMPT protein levels in 21 healthy individuals was determined. We found that NAD concentration and NAMPT expression in BM and BM-derived EPCs of db/db mice were significantly lower than those in WT mice BM. Enhancing NAD pool not only increased the EPCs intracellular NAD concentration and blood EPCs number, but also improved post-ischemic wound healing and blood reperfusion in db/db mice with hind-limb ischemia model. Enhancing NAD pool rescued the impaired modulations of stromal cell-derived factor-1α (SDF-1α) and endothelial nitric oxide synthase (eNOS) protein levels in db/db mice BM upon hind-limb ischemia. In addition, enhancing NAD pool significantly inhibited PARP and caspase-3 activates in db/db mice BM. The intracellular NAMPT-NAD pool was positively associated with blood

  13. EcPV2 DNA in equine genital squamous cell carcinomas and normal genital mucosa.

    Science.gov (United States)

    Bogaert, Lies; Willemsen, Anouk; Vanderstraeten, Eva; Bracho, Maria A; De Baere, Cindy; Bravo, Ignacio G; Martens, Ann

    2012-07-06

    Squamous cell carcinoma (SCC) represents the most common genital malignant tumor in horses. Similar to humans, papillomaviruses (PVs) have been proposed as etiological agents and recently Equine papillomavirus type 2 (EcPV2) has been identified in a subset of genital SCCs. The goals of this study were (1) to determine the prevalence of EcPV2 DNA in tissue samples from equine genital SCCs, penile intraepithelial neoplasia (PIN) and penile papillomas, using EcPV2-specific PCR, (2) to examine the prevalence of latent EcPV2 infection in healthy genital mucosa and (3) to determine genetic variability within EcPV2 and to disentangle phylogenetic relationships of EcPV2 among PVs. EcPV2 DNA was detected in all but one penile SCC (15/16), in all PIN lesions (8/8) and penile papillomas (4/4). Additionally, EcPV2 DNA was demonstrated in one of two metastasized lymph nodes, one contact metastasis in the mouth, two vaginal and one anal lesion. In healthy horses, EcPV2 DNA was detected in 10% (4/39) of penile swabs but in none of vulvovaginal swabs (0/20). This study confirms the presence of EcPV2 DNA in equine genital SCCs and shows its involvement in anal lesions, a lymph node and contact metastases. Latent EcPV2 presence was also shown in normal male genital mucosa. We found that different EcPV2 variants cocirculate among horses and that EcPV2 is related to the Delta+Zeta PVs and is only a very distant relative of high-risk human PVs causing genital cancer. Thus, similar viral tropism and similar malignant outcome of the infection do not imply close evolutionary relationship. Copyright © 2012 Elsevier B.V. All rights reserved.

  14. Plant Life Management of the EC6 Concrete Containment Structure

    Energy Technology Data Exchange (ETDEWEB)

    Abrishami, Homayoun; Ricciuti, Rick; Khan, Azhar [CANDU Energy Inc., Mississauga (Canada)

    2012-03-15

    Aging of reinforced concrete structures due to service conditions, aggressive environments, or accidents may cause their strength, serviceability and durability to decrease over time. Due to the complex nature of safety-related structures in nuclear power plants in comparison to other structures, they possess a number of characteristics that make them comparison to other structures, they possess a number of characteristics that make them unique. These characteristics are: thick concrete cross-sections, heavy reinforcement, often one-side access only, subjected to such ageing stresses as irradiation and elevated temperature, in addition to other typical ageing mechanisms (i. e., exposure to freeze/thaw cycles, aggressive chemicals, etc.) that typically affects other types of non-nuclear structures. For a new plant, the Plant Life Management Program (PLiM) should start in the design process and then continues through construction, plant operation and decommissioning. Hence PLiM must provide not only Ageing Management program (AMP) but also provide requirements on material characteristic and the design criteria as well. The purpose of this paper is to present the Plant Life Management (PLiM) strategy for the concrete containment structure of EC6 (Enhanced CANDU 6) Nuclear Power Plant designed by CANDU Energy Inc. The EC6 is designed for 100-year plant life including a 60-year operating life and an additional 40-year decommissioning period of time. The approach adopted for the PLiM strategy of the concrete containment structure is a preventive one, key areas being: 1) design methodology, 2) material performance and 3) life cycle management and ageing management program. In addition to strength and serviceability, durability is a major consideration during the design phase, service life and up to the completion of decommissioning. Factors affecting durability design include: a) concrete performance, b) structural application, and c) consideration of environmental

  15. Plant Life Management of the EC6 Concrete Containment Structure

    International Nuclear Information System (INIS)

    Abrishami, Homayoun; Ricciuti, Rick; Khan, Azhar

    2012-01-01

    Aging of reinforced concrete structures due to service conditions, aggressive environments, or accidents may cause their strength, serviceability and durability to decrease over time. Due to the complex nature of safety-related structures in nuclear power plants in comparison to other structures, they possess a number of characteristics that make them comparison to other structures, they possess a number of characteristics that make them unique. These characteristics are: thick concrete cross-sections, heavy reinforcement, often one-side access only, subjected to such ageing stresses as irradiation and elevated temperature, in addition to other typical ageing mechanisms (i. e., exposure to freeze/thaw cycles, aggressive chemicals, etc.) that typically affects other types of non-nuclear structures. For a new plant, the Plant Life Management Program (PLiM) should start in the design process and then continues through construction, plant operation and decommissioning. Hence PLiM must provide not only Ageing Management program (AMP) but also provide requirements on material characteristic and the design criteria as well. The purpose of this paper is to present the Plant Life Management (PLiM) strategy for the concrete containment structure of EC6 (Enhanced CANDU 6) Nuclear Power Plant designed by CANDU Energy Inc. The EC6 is designed for 100-year plant life including a 60-year operating life and an additional 40-year decommissioning period of time. The approach adopted for the PLiM strategy of the concrete containment structure is a preventive one, key areas being: 1) design methodology, 2) material performance and 3) life cycle management and ageing management program. In addition to strength and serviceability, durability is a major consideration during the design phase, service life and up to the completion of decommissioning. Factors affecting durability design include: a) concrete performance, b) structural application, and c) consideration of environmental

  16. Clues on Type Ia Supernovae Progenitors

    International Nuclear Information System (INIS)

    Piersanti, Luciano; Tornambe, Amedeo

    2005-01-01

    We show that in the framework of canonical stellar evolution it is hard, if not impossible, to determine the growth in mass of a CO White Dwarf, up to the Chandrasekhar limit by means of mass transfer from its companion in a binary system. This is the case either if matter is accreted from a normal companion with an H-rich envelope or if direct CO accretion occurs from a CO WD companion. At variance, we show that if the effects of rotation are taken into account in modeling the accretion process, a CO WD can increase its mass at the expenses of the degenerate CO companion up and beyond 1.4 M· , so that an explosive event of the type Ia class is naturally produced. This theoretical finding revives the Double Degenerate scenario for type Ia SNe progenitors. In such a case the internal spread in the observational properties of type Ia SNe may be interpreted as a consequence of different total masses; hence differences between SNe Ia in nearby elliptical galaxies and the majority of those in spirals should be expected and the current use of type Ia SNe as cosmological distance indicators should be justified

  17. TWEAK induces liver progenitor cell proliferation

    Science.gov (United States)

    Jakubowski, Aniela; Ambrose, Christine; Parr, Michael; Lincecum, John M.; Wang, Monica Z.; Zheng, Timothy S.; Browning, Beth; Michaelson, Jennifer S.; Baestcher, Manfred; Wang, Bruce; Bissell, D. Montgomery; Burkly, Linda C.

    2005-01-01

    Progenitor (“oval”) cell expansion accompanies many forms of liver injury, including alcohol toxicity and submassive parenchymal necrosis as well as experimental injury models featuring blocked hepatocyte replication. Oval cells can potentially become either hepatocytes or biliary epithelial cells and may be critical to liver regeneration, particularly when hepatocyte replication is impaired. The regulation of oval cell proliferation is incompletely understood. Herein we present evidence that a TNF family member called TWEAK (TNF-like weak inducer of apoptosis) stimulates oval cell proliferation in mouse liver through its receptor Fn14. TWEAK has no effect on mature hepatocytes and thus appears to be selective for oval cells. Transgenic mice overexpressing TWEAK in hepatocytes exhibit periportal oval cell hyperplasia. A similar phenotype was obtained in adult wild-type mice, but not Fn14-null mice, by administering TWEAK-expressing adenovirus. Oval cell expansion induced by 3,5-diethoxycarbonyl-1,4-dihydrocollidine (DDC) was significantly reduced in Fn14-null mice as well as in adult wild-type mice with a blocking anti-TWEAK mAb. Importantly, TWEAK stimulated the proliferation of an oval cell culture model. Finally, we show increased Fn14 expression in chronic hepatitis C and other human liver diseases relative to its expression in normal liver, which suggests a role for the TWEAK/Fn14 pathway in human liver injury. We conclude that TWEAK has a selective mitogenic effect for liver oval cells that distinguishes it from other previously described growth factors. PMID:16110324

  18. Harmine stimulates proliferation of human neural progenitors

    Directory of Open Access Journals (Sweden)

    Vanja Dakic

    2016-12-01

    Full Text Available Harmine is the β-carboline alkaloid with the highest concentration in the psychotropic plant decoction Ayahuasca. In rodents, classical antidepressants reverse the symptoms of depression by stimulating neuronal proliferation. It has been shown that Ayahuasca presents antidepressant effects in patients with depressive disorder. In the present study, we investigated the effects of harmine in cell cultures containing human neural progenitor cells (hNPCs, 97% nestin-positive derived from pluripotent stem cells. After 4 days of treatment, the pool of proliferating hNPCs increased by 71.5%. Harmine has been reported as a potent inhibitor of the dual specificity tyrosine-phosphorylation-regulated kinase (DYRK1A, which regulates cell proliferation and brain development. We tested the effect of analogs of harmine, an inhibitor of DYRK1A (INDY, and an irreversible selective inhibitor of monoamine oxidase (MAO but not DYRK1A (pargyline. INDY but not pargyline induced proliferation of hNPCs similarly to harmine, suggesting that inhibition of DYRK1A is a possible mechanism to explain harmine effects upon the proliferation of hNPCs. Our findings show that harmine enhances proliferation of hNPCs and suggest that inhibition of DYRK1A may explain its effects upon proliferation in vitro and antidepressant effects in vivo.

  19. Role of liver progenitors in liver regeneration.

    Science.gov (United States)

    Best, Jan; Manka, Paul; Syn, Wing-Kin; Dollé, Laurent; van Grunsven, Leo A; Canbay, Ali

    2015-02-01

    During massive liver injury and hepatocyte loss, the intrinsic regenerative capacity of the liver by replication of resident hepatocytes is overwhelmed. Treatment of this condition depends on the cause of liver injury, though in many cases liver transplantation (LT) remains the only curative option. LT for end stage chronic and acute liver diseases is hampered by shortage of donor organs and requires immunosuppression. Hepatocyte transplantation is limited by yet unresolved technical difficulties. Since currently no treatment is available to facilitate liver regeneration directly, therapies involving the use of resident liver stem or progenitor cells (LPCs) or non-liver stem cells are coming to fore. LPCs are quiescent in the healthy liver, but may be activated under conditions where the regenerative capacity of mature hepatocytes is severely impaired. Non-liver stem cells include embryonic stem cells (ES cells) and mesenchymal stem cells (MSCs). In the first section, we aim to provide an overview of the role of putative cytokines, growth factors, mitogens and hormones in regulating LPC response and briefly discuss the prognostic value of the LPC response in clinical practice. In the latter section, we will highlight the role of other (non-liver) stem cells in transplantation and discuss advantages and disadvantages of ES cells, induced pluripotent stem cells (iPS), as well as MSCs.

  20. Hepatic progenitors for liver disease: current position

    Directory of Open Access Journals (Sweden)

    Alice Conigliaro

    2010-02-01

    Full Text Available Alice Conigliaro1, David A Brenner2, Tatiana Kisseleva21University “La Sapienza”, Dipartimento di Biotecnologie Cellulari ed Ematologia Policlinico Umberto I, V Clinica Medica, Rome, Italy; 2Department of Medicine, University of California, San Diego, La Jolla, CA, USAAbstract: Liver regeneration restores the original functionality of hepatocytes and cholangiocytes in response to injury. It is regulated on several levels, with different cellular populations contributing to this process, eg, hepatocytes, liver precursor cells, intrahepatic stem cells. In response to injury, mature hepatocytes have the capability to proliferate and give rise to new hepatocytes and cholangiocytes. Meanwhile, liver precursor cells (oval cells have become the most recognized bipotential precursor cells in the damaged liver. They rapidly proliferate, change their cellular composition, and differentiate into hepatocytes and cholangiocytes to compensate for the cellular loss and maintain liver homeostasis. There is a growing body of evidence that oval cells originate from the intrahepatic stem cell(s, which in turn give(s rise to epithelial, including oval cells, and/or other hepatic cells of nonepithelial origin. Since there is a close relationship between the liver and hematopoiesis, bone marrow derived cells can also contribute to liver regeneration by the fusion of myeloid cells with damaged hepatocytes, or differentiation of mesenchymal stem cells into hepatocyte-like cells. The current review discusses the contribution of different cells to liver regeneration and their characteristics.Keywords: hepatic progenitor, liver disease, liver precursor cells, oval cells, hepatocytes, intrahepatic stem cells, cholangiocytes

  1. Measurement of stray EC radiation on W7-AS

    Science.gov (United States)

    Gandini, F.; Hirsch, M.; Cirant, S.; Erckmann, V.; Granucci, G.; Kasparek, W.; Laqua, H. P.; Muzzini, V.; Nowak, S.; Radau, S.

    2001-10-01

    In the framework of a collaboration between IFP-CNR Milano, IPP Garching/Greifswald and IPF Stuttgart, a set of four millimeterwave probes has been installed in W7-AS stellarator at selected positions of the inner vessel wall. Their purpose is to observe RF stray radiation during operation in presence of strong level of Electron Cyclotron (EC) waves, used for plasma start-up, heating and current drive. The aim of these measurements is to benchmark two complementary theoretical models for the distribution of the stray radiation in the vessel. From these codes, quantitative predictions are expected for the spatial distribution of the RF wall load and the RF-impact on in-vessel components in large future devices such as W7-X and, possibly, ITER. This input is important to optimize the wall armour and select rf-compatible in-vessel materials. We present first measurements from different heating and startup scenarios, with up to 800 kW of injected power at 140 GHz and different launching geometries. An analysis of measurements performed on FTU using a previous version of sniffer probe is also presented.

  2. EC infuses Serbian nuclear relic cleanup with critical donation

    International Nuclear Information System (INIS)

    2008-01-01

    . In addition to the radiological legacy at Vinca, security had long been a source of concern. The reactor has been offline since 1984, and much of the dangerous material and facilities were inadequately protected. Thanks to funding and other support activities provided by the Serbian government and the USA for security upgrades and police support, overall site security has been substantially improved over the past two years. Yet more needs to be done, and time is running out. Along with the EC's recent donation, an additional $25 million must be raised by 2010 to meet a crucial deadline. The fuel needs to be shipped back to Russia by the end of 2010 or the job falls off the shipping schedule, and potential funding for VIND would also dry up should the 2010 target date be missed. VIND is the largest one-house programme within the IAEA, and the EC contribution is the largest single contribution ever received for a Technical Cooperation (TC) national project. 'The EC support was absolutely crucial to the life of the programme, but we have quite a way to go to find the remaining $25 million for the project,' said Kelly. 'The EC contribution serves as a great example and encourages other potential donors to invest in an important and successful project.' Background: Located on the outskirts of Belgrade, the 'Institute for Nuclear Sciences (Vinca)' was set up as a research centre in the former Yugoslavia in the 1950s. A civilian nuclear research reactor loaded with high-enriched uranium was housed at the site. The area was also a central radioactive waste collection and consolidation centre for the former Yugoslavia. The grounds at Vinca accumulated all the former country's dangerous radioactive waste and other radioactive sources for nearly 45 years, and though the reactor went offline in 1984, the radioactive waste and sources received from around the country continued to pile up. International concern about Vinca mushroomed in the 1990s after the break-up of the former

  3. Reshaping Text Data for Efficient Processing on Amazon EC2

    Directory of Open Access Journals (Sweden)

    Gabriela Turcu

    2011-01-01

    Full Text Available Text analysis tools are nowadays required to process increasingly large corpora which are often organized as small files (abstracts, news articles, etc.. Cloud computing offers a convenient, on-demand, pay-as-you-go computing environment for solving such problems. We investigate provisioning on the Amazon EC2 cloud from the user perspective, attempting to provide a scheduling strategy that is both timely and cost effective. We derive an execution plan using an empirically determined application performance model. A first goal of our performance measurements is to determine an optimal file size for our application to consume. Using the subset-sum first fit heuristic we reshape the input data by merging files in order to match as closely as possible the desired file size. This also speeds up the task of retrieving the results of our application, by having the output be less segmented. Using predictions of the performance of our application based on measurements on small data sets, we devise an execution plan that meets a user specified deadline while minimizing cost.

  4. Electrosynthesis of Clozapine Drug Derivative via an EC Electrochemical Mechanism

    Directory of Open Access Journals (Sweden)

    Esmail Tammari

    2017-12-01

    Full Text Available The fact that oxidation, as one of the main routes of phase I metabolism of drugs, follows by conjugation reactions, and also formation of nitrenium ion as one of the clozapine oxidation products, directed us to investigate the oxidation of clozapine (CLZ in the presence of nucleophile. The oxidation of clozapine (CLZ has been studied on a glassy carbon electrode in the absence and presence of 2-thiobarbituric acid (TBA as nucleophile in aqueous medium by means of cyclic voltammetry and on the graphite rods in controlled-potential coulometry. Cyclic voltammetry studies were realized for CLZ in the pHs 1.0 to 8.0. Results indicate that the electrochemical behavior of CLZ depends on the pH. Based on the obtained electrochemical results, an ECE mechanism was proposed to explain the electrochemical oxidation of CLZ. The results revealed that oxidized CLZ participates in Michael type addition reaction with TBA and via an EC mechanism converts to the corresponding new dibenzodiazepin derivatives. The product has been characterized by IR, 1H NMR, 13C NMR and MS.

  5. Improved design of an ITER equatorial EC launcher

    International Nuclear Information System (INIS)

    Takahashi, K.; Kajiwara, K.; Kobayashi, N.; Kasugai, A.; Sakamoto, K.

    2008-01-01

    New design improvements for an ITER equatorial EC launcher are described. A design of the front shield modules was evaluated analytically. The results indicate that these modules can withstand the electromagnetic forces induced by plasma disruption and vertical plasma motion. The steering mirror mock-up including the spiral cooling tubes was fabricated based on the present design with application of the hot isostatic pressing technique to bond the copper alloy mirror body and the embedded stainless steel cooling tubes. The test of water flow was carried out and the expected flow rate was successfully obtained. A quasi-optical (QO) transmission layout has been proposed instead of the waveguide lines. The advantage of this option is possibly to increase the reliability in terms of fabrication and refurbishment. According to the beam propagation analysis, it is verified that transmission loss at the QO region is 1.1%-2.2%, which is comparable to the reference design. The peak heat load on the steering mirror surface could be reduced by 55%, which relaxes the mirror design

  6. Measurement of stray EC radiation on W7-AS

    International Nuclear Information System (INIS)

    Gandini, F.; Cirant, S.; Granucci, G.; Muzzini, V.; Nowak, S.; Hirsch, M.; Erckmann, V.; Laqua, H.P.; Radau, S.; Kasparek, W.

    2001-01-01

    In the framework of a collaboration between IFP-CNR Milano, IPP Garching/Greifswald and IPF Stuttgart, a set of four millimeterwave probes has been installed in W7-AS stellarator at selected positions of the inner vessel wall. Their purpose is to observe RF stray radiation during operation in presence of strong level of Electron Cyclotron (EC) waves, used for plasma start-up, heating and current drive. The aim of these measurements is to benchmark two complementary theoretical models for the distribution of the stray radiation in the vessel. From these codes, quantitative predictions are expected for the spatial distribution of the RF wall load and the RF-impact on in-vessel components in large future devices such as W7-X and, possibly, ITER. This input is important to optimize the wall armour and select rf-compatible in-vessel materials. We present first measurements from different heating and startup scenarios, with up to 800 kW of injected power at 140 GHz and different launching geometries. An analysis of measurements performed on FTU using a previous version of sniffer probe is also presented

  7. Jung's dissociable psyche and the ec-static self.

    Science.gov (United States)

    Austin, Sue

    2009-11-01

    Much of Jung's later work assumes that the self is an a priori phenomenon in which centripetal dynamics dominate. There is, however, another current in Jung's writings which recognizes the self to be an emergent phenomenon. This view is increasingly prevalent in post-Jungian discourse, and Louis Zinkin's exploration of a post-Jungian-constructivist model of the self can be seen as part of this tendency. My paper privileges an emergent understanding of the self by focusing on the 'unravelling', 'de-centring', centrifugal experiences of otherness in the psyche. It offers a post-Jungian reading of a number of writers who have been influenced by the psychoanalyst Jean Laplanche and proposes a model of the self which focuses on our fantasies, terrors and longings about coming undone and bringing others undone. This model is then linked to Judith Butler's understanding of the self as an ec-static phenomenon, in which the self is, of necessity, outside itself, such that 'there is no final moment in which my return to myself takes place'. I suggest that Jung's early clinical researches into the dissociability of the psyche and the clinical tools which he developed as a result of this work are especially suitable for engaging with these emergent, centrifugal dynamics.

  8. The ITER EC H and CD Upper Launcher: Maintenance concepts

    International Nuclear Information System (INIS)

    Ronden, D.M.S.; Baar, M. de; Chavan, R.; Elzendoorn, B.S.Q.; Grossetti, G.; Heemskerk, C.J.M.; Koning, J.F.; Landis, J.-D.; Spaeh, P.; Strauss, D.

    2013-01-01

    Highlights: ► We explain how an overall maintenance strategy defines individual maintenance tasks. ► Concepts are presented for replacement strategies of the in-vessel optical components. ► Vertical placement of the Upper Launcher in the Hot Cell may simplify maintenance. -- Abstract: Maintenance of the ITER EC H and CD Upper Launcher (UL) shall be performed through the use of Remote Handling (RH) in the ITER Hot Cell Facility (HCF). The UL design will have to be fully compliant with ITER RH maintenance requirements and the set of RH tooling and services available in the HCF. This paper describes the development of an overall maintenance strategy for the UL, starting from a listing of all conceivable maintenance operations, including hands-on tasks. Components for which design concepts are discussed in this paper are the Blanket Shield Module (BSM), the steering mirror (M4), the mid optics (M1, M2) and the waveguide (WG) feed-through plate. Aspects related to RH documentation, overall maintenance strategy and design concepts for optimizing the maintainability of the UL are presented

  9. EC4IT - A Guide to European Commission Projects for CERN/IT department

    CERN Document Server

    Jones, R

    2012-01-01

    European Commission (EC) projects represent an important source of external funding for a number of activities in which IT department is involved. The EC has a set of procedures and formalities for the preparation, negotiation, execution and completion of projects that are different to those normally used at CERN. This document gives an overview of the administrative, financial, reporting and managerial steps involved in participating in an EC project.

  10. and alanine (EC. 2.6.1.2) transaminases, and alkaline phosphatase

    African Journals Online (AJOL)

    The activities of aspartate (E.C. 2.6.1.1) and alanine (E.C. 2.6.1.2) transaminases (AST and ALT, respectively), and alkaline phosphatase (ALP) (E.C. 3.1.3.1) were determined in erythrocytes obtained from 20 HbAA, 15 HbAS and 12 HbSS human subjects. The results showed that the three enzymes had different levels of ...

  11. Direct contact with endoderm-like cells efficiently induces cardiac progenitors from mouse and human pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Hideki Uosaki

    Full Text Available RATIONALE: Pluripotent stem cell-derived cardiac progenitor cells (CPCs have emerged as a powerful tool to study cardiogenesis in vitro and a potential cell source for cardiac regenerative medicine. However, available methods to induce CPCs are not efficient or require high-cost cytokines with extensive optimization due to cell line variations. OBJECTIVE: Based on our in-vivo observation that early endodermal cells maintain contact with nascent pre-cardiac mesoderm, we hypothesized that direct physical contact with endoderm promotes induction of CPCs from pluripotent cells. METHOD AND RESULT: To test the hypothesis, we cocultured mouse embryonic stem (ES cells with the endodermal cell line End2 by co-aggregation or End2-conditioned medium. Co-aggregation resulted in strong induction of Flk1(+ PDGFRa(+ CPCs in a dose-dependent manner, but the conditioned medium did not, indicating that direct contact is necessary for this process. To determine if direct contact with End2 cells also promotes the induction of committed cardiac progenitors, we utilized several mouse ES and induced pluripotent (iPS cell lines expressing fluorescent proteins under regulation of the CPC lineage markers Nkx2.5 or Isl1. In agreement with earlier data, co-aggregation with End2 cells potently induces both Nkx2.5(+ and Isl1(+ CPCs, leading to a sheet of beating cardiomyocytes. Furthermore, co-aggregation with End2 cells greatly promotes the induction of KDR(+ PDGFRa(+ CPCs from human ES cells. CONCLUSIONS: Our co-aggregation method provides an efficient, simple and cost-effective way to induce CPCs from mouse and human pluripotent cells.

  12. GPM GROUND VALIDATION ENVIRONMENT CANADA (EC) MANUAL PRECIPITATION MEASUREMENTS GCPEX V1

    Data.gov (United States)

    National Aeronautics and Space Administration — The GPM Ground Validation Environment Canada (EC) Manual Precipitation Measurements GCPEx dataset was collected during the GPM Cold-season Precipitation Experiment...

  13. Kinetic study on ligand exchange reaction between EC and 99mTc-GH

    International Nuclear Information System (INIS)

    Wu Chunying; Luo Shineng; Fang Ping; Huang Heyun; Xie Minhao; Meng Hong

    1995-01-01

    The ligand exchange reaction between EC and 99m Tc-GH and its influence factors such as concentrations of EC and pH were described. The concentration of EC has no influence on the exchange reaction rate constant, while pH is the most important influence factor. The rate constants of ligand exchange reaction at different pH values were determined. The results showed that in order to make the labelling yield of 99m Tc-EC higher than 90%, pH of the reaction must be higher than 8

  14. Reciprocal upregulation of Notch signaling molecules in hematopoietic progenitor and mesenchymal stromal cells

    Directory of Open Access Journals (Sweden)

    Kikuchi Y

    2011-01-01

    Full Text Available Although mesenchymal stem cells (MSCs play pivotal supportive roles in hematopoiesis, how they interact with hematopoietic stem cells (HSCs is not well understood. We investigated the interaction between HSCs and surrogate MSCs (C3H10T1/2 stromal cells, focusing on the molecular events induced by cell contact of these bipartite populations. C3H10T1/2 is a mesenchymal stromal cell line that can be induced to differentiate into preadipocytes (A54 and myoblasts (M1601. The stromal cell derivatives were cocultured with murine HSCs (Lineage-Sca1+, and gene expression profiles in stromal cells and HSCs were compared before and after the coculture. HSCs gave rise to cobblestone areas only on A54 cells, with ninefold more progenitors than on M1601 or undifferentiated C3H10T1/2 cells. Microarray-based screening and a quantitative reverse transcriptase directed-polymerase chain reaction showed that the levels of Notch ligands (Jagged1 and Delta-like 3 were increased in A54 cells upon interaction with HSCs. On the other hand, the expression of Notch1 and Hes1 was upregulated in the HSCs cocultured with A54 cells. A transwell assay revealed that the reciprocal upregulation was dependent on cell-to-cell contact. The result suggested that in the hematopoietic niche, HSCs help MSCs to produce Notch ligands, and in turn, MSCs help HSCs to express Notch receptor. Such a reciprocal upregulation would reinforce the downstream signaling to determine the fate of hematopoietic cell lineage. Clarification of the initiating events on cell contact should lead to the identification of specific molecular targets to facilitate HSC engraftment in transplantation therapy.

  15. Sources of Hematopoietic Stem and Progenitor Cells and Methods to Optimize Yields for Clinical Cell Therapy.

    Science.gov (United States)

    Panch, Sandhya R; Szymanski, James; Savani, Bipin N; Stroncek, David F

    2017-08-01

    Bone marrow (BM) aspirates, mobilized peripheral blood, and umbilical cord blood (UCB) have developed as graft sources for hematopoietic stem and progenitor cells (HSPCs) for stem cell transplantation and other cellular therapeutics. Individualized techniques are necessary to enhance graft HSPC yields and cell quality from each graft source. BM aspirates yield adequate CD34 + cells but can result in relative delays in engraftment. Granulocyte colony-stimulating factor (G-CSF)-primed BM HSPCs may facilitate faster engraftment while minimizing graft-versus-host disease in certain patient subsets. The levels of circulating HSPCs are enhanced using mobilizing agents, such as G-CSF and/or plerixafor, which act via the stromal cell-derived factor 1/C-X-C chemokine receptor type 4 axis. Alternate niche pathway mediators, including very late antigen-4/vascular cell adhesion molecule-1, heparan sulfate proteoglycans, parathyroid hormone, and coagulation cascade intermediates, may offer promising alternatives for graft enhancement. UCB grafts have been expanded ex vivo with cytokines, notch-ligand, or mesenchymal stromal cells, and most studies demonstrated greater quantities of CD34 + cells ex vivo and improved short-term engraftment. No significant changes were observed in long-term repopulating potential or in patient survival. Early phase clinical trials using nicotinamide and StemReginin1 may offer improved short- and long-term repopulating ability. Breakthroughs in genome editing and stem cell reprogramming technologies may hasten the generation of pooled, third-party HSPC grafts. This review elucidates past, present, and potential future approaches to HSPC graft optimization. Published by Elsevier Inc.

  16. Red wine consumption improves in vitro migration of endothelial progenitor cells in young, healthy individuals.

    Science.gov (United States)

    Hamed, Saher; Alshiek, Jonia; Aharon, Anat; Brenner, Benjamin; Roguin, Ariel

    2010-07-01

    Endothelial progenitor cells (EPCs) contribute to the maintenance of vascular endothelial function. The moderate consumption of red wine provides cardiovascular protection. We investigated the underlying molecular mechanism of EPC migration in young, healthy individuals who drank red wine. Fourteen healthy volunteers consumed 250 mL red wine daily for 21 consecutive days. Vascular endothelial function, plasma stromal cell-derived factor 1alpha (SDF1alpha) concentrations, and the number, migration, and nitric oxide production of EPCs were determined before and after the daily consumption of red wine. EPCs were glucose stressed to study the effect of red wine on EPC migration, proliferation, and senescence and to study the expressions of CXC chemokine receptor 4 (CXCR4) and members of the Pi3K/Akt/eNOS (phosphatidylinositol 3-kinase/protein kinase B/endothelial nitric oxide synthase) signaling pathway by Western blotting. Daily red wine consumption for 21 consecutive days significantly enhanced vascular endothelial function. Although plasma SDF1alpha concentrations were unchanged, EPC count and migration were significantly increased after this 21-d consumption period. Red wine increased the migration, proliferation, CXCR4 expression, and activity of the Pi3K/Akt/eNOS signaling pathway and decreased the extent of apoptosis in glucose-stressed EPCs. The results of the present study indicate that red wine exerts its effect through the up-regulation of CXCR4 expression and activation of the SDF1alpha/CXCR4/Pi3K/Akt/eNOS signaling pathway, which results in increased EPC migration and proliferation and decreased extent of apoptosis. Our findings suggest that these effects could be linked to the mechanism of cardiovascular protection that is associated with the regular consumption of red wine.

  17. Role of endothelial progenitor cells and inflammatory cytokines in healing of diabetic foot ulcers.

    Directory of Open Access Journals (Sweden)

    Francesco Tecilazich

    Full Text Available To evaluate changes in endothelial progenitor cells (EPCs and cytokines in patients with diabetic foot ulceration (DFU in association with wound healing.We studied healthy subjects, diabetic patients not at risk of DFU, at risk of DFU and with active DFU. We prospectively followed the DFU patients over a 12-week period. We also investigated similar changes in diabetic rabbit and mouse models of wound healing.All EPC phenotypes except the kinase insert domain receptor (KDR(+CD133(+ were reduced in the at risk and the DFU groups compared to the controls. There were no major EPC differences between the control and not at risk group, and between the at risk and DFU groups. Serum stromal-cell derived factor-1 (SDF-1 and stem cell factor (SCF were increased in DFU patients. DFU patients who healed their ulcers had lower CD34(+KDR(+ count at visits 3 and 4, serum c-reactive protein (CRP and granulocyte-macrophage colony-stimulating factor (GM-CSF at visit 1, interleukin-1 (IL-1 at visits 1 and 4. EPCs tended to be higher in both diabetic animal models when compared to their non-diabetic counterparts both before and ten days after wounding.Uncomplicated diabetes does not affect EPCs. EPCs are reduced in patients at risk or with DFU while complete wound healing is associated with CD34(+KDR(+ reduction, suggesting possible increased homing. Low baseline CRP, IL-1α and GM-CSF serum levels were associated with complete wound healing and may potentially serve as prognostic markers of DFU healing. No animal model alone is representative of the human condition, indicating the need for multiple experimental models.

  18. Controversies in Cardiovascular Research: Induced pluripotent stem cell-derived cardiomyocytes – boutique science or valuable arrhythmia model?

    Science.gov (United States)

    Knollmann, Björn C

    2013-01-01

    As part of the series on Controversies in Cardiovascular Research, the article reviews the strengths and limitations of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CM) as models of cardiac arrhythmias. Specifically, the article attempts to answer the following questions: Which clinical arrhythmias can be modeled by iPSC-CM? How well can iPSC-CM model adult ventricular myocytes? What are the strengths and limitations of published iPSC-CM arrhythmia models? What new mechanistic insight has been gained? What is the evidence that would support using iPSC-CM to personalize anti-arrhythmic drug therapy? The review also discusses the pros and cons of using the iPSC-CM technology for modeling specific genetic arrhythmia disorders such as long QT syndrome, Brugada Syndrome or Catecholaminergic Polymorphic Ventricular Tachycardia. PMID:23569106

  19. An Abbreviated Protocol for In Vitro Generation of Functional Human Embryonic Stem Cell-Derived Beta-Like Cells

    DEFF Research Database (Denmark)

    Massumi, Mohammad; Pourasgari, Farzaneh; Nalla, Amarnadh

    2016-01-01

    developed an abbreviated five-stage protocol (25-30 days) to generate human Embryonic Stem Cell-Derived Beta-like Cells (ES-DBCs). We showed that Geltrex, as an extracellular matrix, could support the generation of ES-DBCs more efficiently than that of the previously described culture systems......The ability to yield glucose-responsive pancreatic beta-cells from human pluripotent stem cells in vitro will facilitate the development of the cell replacement therapies for the treatment of Type 1 Diabetes. Here, through the sequential in vitro targeting of selected signaling pathways, we have...... positive cells, 1% insulin and glucagon positive cells and 30% insulin and NKX6.1 co-expressing cells. Functionally, ES-DBCs were responsive to high glucose in static incubation and perifusion studies, and could secrete insulin in response to successive glucose stimulations. Mitochondrial metabolic flux...

  20. Matrigel Mattress: A Method for the Generation of Single Contracting Human-Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

    Science.gov (United States)

    Feaster, Tromondae K; Cadar, Adrian G; Wang, Lili; Williams, Charles H; Chun, Young Wook; Hempel, Jonathan E; Bloodworth, Nathaniel; Merryman, W David; Lim, Chee Chew; Wu, Joseph C; Knollmann, Björn C; Hong, Charles C

    2015-12-04

    The lack of measurable single-cell contractility of human-induced pluripotent stem cell-derived cardiac myocytes (hiPSC-CMs) currently limits the utility of hiPSC-CMs for evaluating contractile performance for both basic research and drug discovery. To develop a culture method that rapidly generates contracting single hiPSC-CMs and allows quantification of cell shortening with standard equipment used for studying adult CMs. Single hiPSC-CMs were cultured for 5 to 7 days on a 0.4- to 0.8-mm thick mattress of undiluted Matrigel (mattress hiPSC-CMs) and compared with hiPSC-CMs maintained on a control substrate (method enables the rapid generation of robustly contracting hiPSC-CMs and enhances maturation. This new method allows quantification of contractile performance at the single-cell level, which should be valuable to disease modeling, drug discovery, and preclinical cardiotoxicity testing. © 2015 American Heart Association, Inc.

  1. Transformation of cell-derived microparticles into quantum-dot-labeled nanovectors for antitumor siRNA delivery.

    Science.gov (United States)

    Chen, Gang; Zhu, Jun-Yi; Zhang, Zhi-Ling; Zhang, Wei; Ren, Jian-Gang; Wu, Min; Hong, Zheng-Yuan; Lv, Cheng; Pang, Dai-Wen; Zhao, Yi-Fang

    2015-01-12

    Cell-derived microparticles (MPs) have been recently recognized as critical intercellular information conveyors. However, further understanding of their biological behavior and potential application has been hampered by the limitations of current labeling techniques. Herein, a universal donor-cell-assisted membrane biotinylation strategy was proposed for labeling MPs by skillfully utilizing the natural membrane phospholipid exchange of their donor cells. This innovative strategy conveniently led to specific, efficient, reproducible, and biocompatible quantum dot (QD) labeling of MPs, thereby reliably conferring valuable traceability on MPs. By further loading with small interference RNA, QD-labeled MPs that had inherent cell-targeting and biomolecule-conveying ability were successfully employed for combined bioimaging and tumor-targeted therapy. This study provides the first reliable and biofriendly strategy for transforming biogenic MPs into functionalized nanovectors. © 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  2. Integrated Analysis of Contractile Kinetics, Force Generation, and Electrical Activity in Single Human Stem Cell-Derived Cardiomyocytes

    Directory of Open Access Journals (Sweden)

    Jan David Kijlstra

    2015-12-01

    Full Text Available The quantitative analysis of cardiomyocyte function is essential for stem cell-based approaches for the in vitro study of human cardiac physiology and pathophysiology. We present a method to comprehensively assess the function of single human pluripotent stem cell-derived cardiomyocyte (hPSC-CMs through simultaneous quantitative analysis of contraction kinetics, force generation, and electrical activity. We demonstrate that statistical analysis of movies of contracting hPSC-CMs can be used to quantify changes in cellular morphology over time and compute contractile kinetics. Using a biomechanical model that incorporates substrate stiffness, we calculate cardiomyocyte force generation at single-cell resolution and validate this approach with conventional traction force microscopy. The addition of fluorescent calcium indicators or membrane potential dyes allows the simultaneous analysis of contractility and calcium handling or action potential morphology. Accordingly, our approach has the potential for broad application in the study of cardiac disease, drug discovery, and cardiotoxicity screening.

  3. Completion Report for Well ER-EC-8

    International Nuclear Information System (INIS)

    2004-01-01

    Well ER-EC-8 was drilled for the U.S. Department of Energy, National Nuclear Security Administration Nevada Site Office in support of the Nevada Environmental Restoration Project at the Nevada Test Site, Nye County, Nevada. This well was drilled in the summer of 1999 as part of the U.S. Department of Energy's hydrogeologic investigation program in the Western Pahute Mesa - Oasis Valley region just west of the Nevada Test Site. A 44.5-centimeter surface hole was drilled and cased off to a depth of 129.8 meters below the surface. The hole diameter was then decreased to 31.1 centimeters for drilling to a total depth of 609.6 meters. One completion string with three isolated slotted intervals was installed in the well. A preliminary composite, static water level was measured at the depth of 98.4 meters, 24 days after installation of the completion string. Detailed lithologic descriptions with stratigraphic assignments are included in the report. These are based on evaluation of composite drill cuttings collected every 3 meters, and 20 sidewall samples taken at various depths below 157.9 meters, supplemented by geophysical log data and results of detailed chemical and mineralogical studies of rock samples. Drilling began in Tertiary-age tuff of the Thirsty Canyon Group, and penetrated tuffs of the Beatty Wash Formation, tuff of Buttonhook Wash, and the upper portion of the Ammonia Tanks Tuff. The geologic interpretation of data from this well helps define the location of the western margin of the Timber Mountain caldera complex in the southwestern Nevada volcanic field. Geologic and hydrologic data from the well will aid in development of models to predict groundwater flow and contaminant migration within and near the Nevada Test Site

  4. Completion report for Well ER-EC-6

    Energy Technology Data Exchange (ETDEWEB)

    M. J. Townsend

    2000-05-01

    Well ER-EC-6 was drilled for the U.S. Department of Energy, Nevada Operations Office in support of the Nevada Environmental Restoration Project at the Nevada Test Site, Nye County, Nevada. This well was drilled in the spring of 1999 as part of the DOE's hydrogeologic investigation well program in the Western Pahute Mesa - Oasis Valley region just west of the Nevada Test Site. A 66-centimeter surface hole was drilled and cased off to the depth of 485.1 meters below the surface. The hole diameter was then decreased to 31.1 centimeters for drilling to a total depth of 1,524.0 meters. A preliminary composite, static, water level was measured at the depth of approximately 434.6 meters prior to installation of the completion string. One completion string with four isolated, slotted intervals was installed in the well. Detailed lithologic descriptions with preliminary stratigraphic assignments are included in the report. These are based on composite drill cuttings collected every 3 meters and 33 sidewall samples taken at various depths below 504.4 meters, supplemented by geophysical log data. Detailed chemical and mineralogical studies of rock samples are in progress. The well penetrated Tertiary-age lava and tuff of the Timber Mountain Group, the Paintbrush Group, the Calico Hills Formation, and the Volcanics of Quartz Mountain. Intense hydrothermal alteration was observed below the depth of 640 m. The preliminary geologic interpretation indicates that this site may be located on a buried structural ridge that separates the Silent Canyon and Timber Mountain caldera complexes.

  5. Andrographolide - A promising therapeutic agent, negatively regulates glial cell derived neurodegeneration of prefrontal cortex, hippocampus and working memory impairment.

    Science.gov (United States)

    Das, Sudeshna; Mishra, K P; Ganju, Lilly; Singh, S B

    2017-12-15

    Over activation of glial cell derived innate immune factors induces neuro-inflammation that results in neurodegenerative disease, like working memory impairment. In this study, we have investigated the role of andrographolide, a major constituent of Andrographis paniculata plant, in reduction of reactive glial cell derived working memory impairment. Real time PCR, Western bloting, flow cytometric and immunofluorescence studies demonstrated that andrographolide inhibited lipopolysaccharide (LPS)-induced overexpression of HMGB1, TLR4, NFκB, COX-2, iNOS, and release of inflammatory mediators in primary mix glial culture, adult mice prefrontal cortex and hippocampus region. Active microglial and reactive astrocytic makers were also downregulated after andrographolide treatment. Andrographolide suppressed overexpression of microglial MIP-1α, P2X7 receptor and its downstream signaling mediators including-inflammasome NLRP3, caspase1 and mature IL-1β. Furthermore, in vivo maze studies suggested that andrographolide treatment reversed LPS-induced behavioural and working memory disturbances including regulation of expression of protein markers like PKC, p-CREB, amyloid beta, APP, p-tau, synapsin and PSD-95. Andrographolide, by lowering expression of pro apoptotic genes and enhancing the expression of anti-apoptotic gene showed its anti-apoptotic nature that in turn reduces neurodegeneration. Morphology studies using Nissl and FJB staining also showed the neuroprotective effect of andrographolide in the prefrontal cortex region. The above studies indicated that andrographolide prevented neuroinflammation-associated neurodegeneration and improved synaptic plasticity markers in cortical as well as hippocampal region which suggests that andrographolide could be a novel pharmacological countermeasure for the treatment of neuroinflammation and neurological disorders related to memory impairment. Copyright © 2017 Elsevier B.V. All rights reserved.

  6. Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

    International Nuclear Information System (INIS)

    Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki; Kondo, Ayami; Nakata, Kazuhiko; Mogi, Makio

    2015-01-01

    A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CM treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration

  7. Polyphosphate induces matrix metalloproteinase-3-mediated proliferation of odontoblast-like cells derived from induced pluripotent stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Ozeki, Nobuaki; Hase, Naoko; Yamaguchi, Hideyuki; Hiyama, Taiki; Kawai, Rie [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya, Aichi 464-8651 (Japan); Kondo, Ayami [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan); Nakata, Kazuhiko [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya, Aichi 464-8651 (Japan); Mogi, Makio, E-mail: makio@dpc.agu.ac.jp [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan)

    2015-05-01

    Inorganic polyphosphate [Poly(P)] may represent a physiological source of phosphate and has the ability to induce bone differentiation in osteoblasts. We previously reported that cytokine-induced matrix metalloproteinase (MMP)-3 accelerates the proliferation of purified odontoblast-like cells. In this study, MMP-3 small interfering RNA (siRNA) was transfected into odontoblast-like cells derived from induced pluripotent stem cells to investigate whether MMP-3 activity is induced by Poly(P) and/or is associated with cell proliferation and differentiation into odontoblast-like cells. Treatment with Poly(P) led to an increase in both cell proliferation and additional odontoblastic differentiation. Poly(P)-treated cells showed a small but significant increase in dentin sialophosphoprotein (DSPP) and dentin matrix protein-1 (DMP-1) mRNA expression, which are markers of mature odontoblasts. The cells also acquired additional odontoblast-specific properties including adoption of an odontoblastic phenotype typified by high alkaline phosphatase (ALP) activity and a calcification capacity. In addition, Poly(P) induced expression of MMP-3 mRNA and protein, and increased MMP-3 activity. MMP-3 siRNA-mediated disruption of the expression of these effectors potently suppressed the expression of odontoblastic biomarkers ALP, DSPP, and DMP-1, and blocked calcification. Interestingly, upon siRNA-mediated silencing of MMP-3, we noted a potent and significant decrease in cell proliferation. Using specific siRNAs, we revealed that a unique signaling cascade, Poly(P)→MMP-3→DSPP and/or DMP-1, was intimately involved in the proliferation of odontoblast-like cells. - Highlights: • Polyphosphate increases proliferation of iPS cell-derived odontoblast-like cells. • Polyphosphate-induced MMP-3 results in an increase of cell proliferation. • Induced cell proliferation involves MMP-3, DSPP, and/or DMP-1 sequentially. • Induced MMP-3 also results in an increase of odontoblastic

  8. Unique proliferation response in odontoblastic cells derived from human skeletal muscle stem cells by cytokine-induced matrix metalloproteinase-3

    Energy Technology Data Exchange (ETDEWEB)

    Ozeki, Nobuaki; Hase, Naoko; Kawai, Rie; Yamaguchi, Hideyuki; Hiyama, Taiki [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya 464-8651, Aichi (Japan); Kondo, Ayami [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan); Nakata, Kazuhiko [Department of Endodontics, School of Dentistry, Aichi Gakuin University, 2-11 Suemori-dori, Chikusa-ku, Nagoya 464-8651, Aichi (Japan); Mogi, Makio, E-mail: makio@dpc.agu.ac.jp [Department of Medicinal Biochemistry, School of Pharmacy, Aichi Gakuin University, 1-100 Kusumoto, Chikusa-ku, Nagoya 464-8650 (Japan)

    2015-02-01

    A pro-inflammatory cytokine mixture (CM: interleukin (IL)-1β, tumor necrosis factor-α and interferon-γ) and IL-1β-induced matrix metalloproteinase (MMP)-3 activity have been shown to increase the proliferation of rat dental pulp cells and murine stem cell-derived odontoblast-like cells. This suggests that MMP-3 may regulate wound healing and regeneration in the odontoblast-rich dental pulp. Here, we determined whether these results can be extrapolated to human dental pulp by investigating the effects of CM-induced MMP-3 up-regulation on the proliferation and apoptosis of purified odontoblast-like cells derived from human skeletal muscle stem cells. We used siRNA to specifically reduce MMP-3 expression. We found that CM treatment increased MMP-3 mRNA and protein levels as well as MMP-3 activity. Cell proliferation was also markedly increased, with no changes in apoptosis, upon treatment with CM and following the application of exogenous MMP-3. Endogenous tissue inhibitors of metalloproteinases were constitutively expressed during all experiments and unaffected by MMP-3. Although treatment with MMP-3 siRNA suppressed cell proliferation, it also unexpectedly increased apoptosis. This siRNA-mediated increase in apoptosis could be reversed by exogenous MMP-3. These results demonstrate that cytokine-induced MMP-3 activity regulates cell proliferation and suppresses apoptosis in human odontoblast-like cells. - Highlights: • Pro-inflammatory cytokines induce MMP-3 activity in human odontoblast-like cells. • Increased MMP-3 activity can promote cell proliferation in odontoblasts. • Specific loss of MMP-3 increases apoptosis in odontoblasts. • MMP-3 has potential as a promising new target for pupal repair and regeneration.

  9. Nonanaplastic follicular cell-derived thyroid carcinoma: mitosis and necrosis in long-term follow-up.

    Science.gov (United States)

    Skansing, Daniel Bräuner; Londero, Stefano Christian; Asschenfeldt, Pia; Larsen, Stine Rosenkilde; Godballe, Christian

    2017-06-01

    Nonanaplastic follicular cell-derived thyroid carcinoma (NAFCTC) includes differentiated- (DTC) and poorly differentiated thyroid carcinoma (PDTC). DTC has an excellent prognosis, while PDTC is situated between DTC and anaplastic carcinomas. Short-term studies suggest that PDTC patients diagnosed only on tumor necrosis and/or mitosis have a prognosis similar to those diagnosed according to the TURIN proposal. The purpose of this study was to evaluate prognosis for NAFCTC based on long-term follow-up illuminating the significance of tumor necrosis and mitosis. A cohort of 225 patients with NAFCTC was followed more than 20 years. Age, sex, distant metastasis, histology, tumor size, extrathyroidal invasion, lymph node metastasis, tumor necrosis and mitosis were examined as possible prognostic factors. Median follow-up time for patients alive was 28 years (range 20-43 years). Age, distant metastasis, extrathyroidal invasion, tumor size, tumor necrosis and mitosis were independent prognostic factors in multivariate analysis for overall survival (OS). In disease specific survival (DSS) age was not significant. Using only necrosis and/or mitosis as criteria for PDTC the 5-, 10- and 20-year OS for DTC was 87, 79 and 69%, respectively. In DSS it was 95, 92 and 90%. For PDTC the 5-, 10- and 20-year OS was 57, 40 and 25%, respectively. In DSS it was 71, 55 and 48%. Tumor necrosis and mitosis are highly significant prognostic indicators in analysis of long time survival of nonanaplastic follicular cell-derived thyroid carcinoma indicating that a simplification of the actually used criteria for poorly differentiated carcinomas may be justified.

  10. Extracellular Matrix-Mediated Maturation of Human Pluripotent Stem Cell-Derived Cardiac Monolayer Structure and Electrophysiological Function.

    Science.gov (United States)

    Herron, Todd J; Rocha, Andre Monteiro Da; Campbell, Katherine F; Ponce-Balbuena, Daniela; Willis, B Cicero; Guerrero-Serna, Guadalupe; Liu, Qinghua; Klos, Matt; Musa, Hassan; Zarzoso, Manuel; Bizy, Alexandra; Furness, Jamie; Anumonwo, Justus; Mironov, Sergey; Jalife, José

    2016-04-01

    Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) monolayers generated to date display an immature embryonic-like functional and structural phenotype that limits their utility for research and cardiac regeneration. In particular, the electrophysiological function of hPSC-CM monolayers and bioengineered constructs used to date are characterized by slow electric impulse propagation velocity and immature action potential profiles. Here, we have identified an optimal extracellular matrix for significant electrophysiological and structural maturation of hPSC-CM monolayers. hPSC-CM plated in the optimal extracellular matrix combination have impulse propagation velocities ≈2× faster than previously reported (43.6±7.0 cm/s; n=9) and have mature cardiomyocyte action potential profiles, including hyperpolarized diastolic potential and rapid action potential upstroke velocity (146.5±17.7 V/s; n=5 monolayers). In addition, the optimal extracellular matrix promoted hypertrophic growth of cardiomyocytes and the expression of key mature sarcolemmal (SCN5A, Kir2.1, and connexin43) and myofilament markers (cardiac troponin I). The maturation process reported here relies on activation of integrin signaling pathways: neutralization of β1 integrin receptors via blocking antibodies and pharmacological blockade of focal adhesion kinase activation prevented structural maturation. Maturation of human stem cell-derived cardiomyocyte monolayers is achieved in a 1-week period by plating cardiomyocytes on PDMS (polydimethylsiloxane) coverslips rather than on conventional 2-dimensional cell culture formats, such as glass coverslips or plastic dishes. Activation of integrin signaling and focal adhesion kinase is essential for significant maturation of human cardiac monolayers. © 2016 American Heart Association, Inc.

  11. Evaluation of Stem Cell-Derived Red Blood Cells as a Transfusion Product Using a Novel Animal Model.

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    Sandeep N Shah

    Full Text Available Reliance on volunteer blood donors can lead to transfusion product shortages, and current liquid storage of red blood cells (RBCs is associated with biochemical changes over time, known as 'the storage lesion'. Thus, there is a need for alternative sources of transfusable RBCs to supplement conventional blood donations. Extracorporeal production of stem cell-derived RBCs (stemRBCs is a potential and yet untapped source of fresh, transfusable RBCs. A number of groups have attempted RBC differentiation from CD34+ cells. However, it is still unclear whether these stemRBCs could eventually be effective substitutes for traditional RBCs due to potential differences in oxygen carrying capacity, viability, deformability, and other critical parameters. We have generated ex vivo stemRBCs from primary human cord blood CD34+ cells and compared them to donor-derived RBCs based on a number of in vitro parameters. In vivo, we assessed stemRBC circulation kinetics in an animal model of transfusion and oxygen delivery in a mouse model of exercise performance. Our novel, chronically anemic, SCID mouse model can evaluate the potential of stemRBCs to deliver oxygen to tissues (muscle under resting and exercise-induced hypoxic conditions. Based on our data, stem cell-derived RBCs have a similar biochemical profile compared to donor-derived RBCs. While certain key differences remain between donor-derived RBCs and stemRBCs, the ability of stemRBCs to deliver oxygen in a living organism provides support for further development as a transfusion product.

  12. The role of tumor cell-derived connective tissue growth factor (CTGF/CCN2) in pancreatic tumor growth.

    Science.gov (United States)

    Bennewith, Kevin L; Huang, Xin; Ham, Christine M; Graves, Edward E; Erler, Janine T; Kambham, Neeraja; Feazell, Jonathan; Yang, George P; Koong, Albert; Giaccia, Amato J

    2009-02-01

    Pancreatic cancer is highly aggressive and refractory to existing therapies. Connective tissue growth factor (CTGF/CCN2) is a fibrosis-related gene that is thought to play a role in pancreatic tumor progression. However, CCN2 can be expressed in a variety of cell types, and the contribution of CCN2 derived from either tumor cells or stromal cells as it affects the growth of pancreatic tumors is unknown. Using genetic inhibition of CCN2, we have discovered that CCN2 derived from tumor cells is a critical regulator of pancreatic tumor growth. Pancreatic tumor cells derived from CCN2 shRNA-expressing clones showed dramatically reduced growth in soft agar and when implanted s.c. We also observed a role for CCN2 in the growth of pancreatic tumors implanted orthotopically, with tumor volume measurements obtained by positron emission tomography imaging. Mechanistically, CCN2 protects cells from hypoxia-mediated apoptosis, providing an in vivo selection for tumor cells that express high levels of CCN2. We found that CCN2 expression and secretion was increased in hypoxic pancreatic tumor cells in vitro, and we observed colocalization of CCN2 and hypoxia in pancreatic tumor xenografts and clinical pancreatic adenocarcinomas. Furthermore, we found increased CCN2 staining in clinical pancreatic tumor tissue relative to stromal cells surrounding the tumor, supporting our assertion that tumor cell-derived CCN2 is important for pancreatic tumor growth. Taken together, these data improve our understanding of the mechanisms responsible for pancreatic tumor growth and progression, and also indicate that CCN2 produced by tumor cells represents a viable therapeutic target for the treatment of pancreatic cancer.

  13. Circulating levels of cell-derived microparticles are reduced by mild hypobaric hypoxia: data from a randomised controlled trial.

    Science.gov (United States)

    Ayers, Lisa; Stoewhas, Anne-Christin; Ferry, Berne; Latshang, Tsogyal D; Lo Cascio, Christian M; Sadler, Ross; Stadelmann, Katrin; Tesler, Noemi; Huber, Reto; Achermann, Peter; Bloch, Konrad E; Kohler, Malcolm

    2014-05-01

    Hypoxia is known to induce the release of microparticles in vitro. However, few publications have addressed the role of hypoxia in vivo on circulating levels of microparticles. This randomised, controlled, crossover trial aimed to determine the effect of mild hypoxia on in vivo levels of circulating microparticles in healthy individuals. Blood was obtained from 51 healthy male volunteers (mean age of 26.9 years) at baseline altitude (490 m) and after 24 and 48 h at moderate altitude (2,590 m). The order of altitude exposure was randomised. Flow cytometry was used to assess platelet-poor plasma for levels of circulating microparticles derived from platelets, endothelial cells, leucocytes, granulocytes, monocytes, red blood cells and procoagulant microparticles. Mean (standard deviation) oxygen saturation was significantly lower on the first and second day after arrival at 2,590 m, 91.0 (2.0) and 92.0 (2.0) %, respectively, compared to 490 m, 96 (1.0) %, p microparticles (annexin V+ -221/μl 95 % CI -370.8/-119.0, lactadherin+ -202/μl 95 % CI -372.2/-93.1), platelet-derived microparticles (-114/μl 95 % CI -189.9/-51.0) and red blood cell-derived microparticles (-81.4 μl 95 % CI -109.9/-57.7) after 48 h at moderate altitude was found. Microparticles derived from endothelial cells, granulocytes, monocytes and leucocytes were not significantly altered by exposure to moderate altitude. In healthy male individuals, mild hypobaric hypoxia, induced by a short-term stay at moderate altitude, is associated with lower levels of procoagulant microparticles, platelet-derived microparticles and red blood cell-derived microparticles, suggesting a reduction in thrombotic potential.

  14. Renal blood flow and oxygenation drive nephron progenitor differentiation.

    Science.gov (United States)

    Rymer, Christophe