WorldWideScience

Sample records for leukemic cells derived

  1. Anti-leukemic therapies induce cytogenetic changes of human bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Yeh, Su-Peng; Lo, Wen-Jyi; Lin, Chiao-Lin; Liao, Yu-Min; Lin, Chen-Yuan; Bai, Li-Yuan; Liang, Ji-An; Chiu, Chang-Fang

    2012-02-01

    Both bone marrow hematopoietic cells (BM-HCs) and mesenchymal stem cells (BM-MSCs) may have cytogenetic aberrations in leukemic patients, and anti-leukemic therapy may induce cytogenetic remission of BM-HCs. The impact of anti-leukemic therapy on BM-MSCs remains unknown. Cytogenetic studies of BM-MSCs from 15 leukemic patients with documented cytogenetic abnormalities of BM-HCs were investigated. To see the influence of anti-leukemic therapy on BM-MSCs, cytogenetic studies were carried out in seven of them after the completion of anti-leukemic therapy, including anthracycline/Ara-C-based chemotherapy in two patients, high-dose busulfan/cyclophosphamide-based allogeneic transplantation in two patients, and total body irradiation (TBI)-based allogeneic transplantation in three patients. To simulate the effect of TBI in vitro, three BM-MSCs from one leukemic patient and two normal adults were irradiated using the same dosage and dosing schedule of TBI and cytogenetics were re-examined after irradiation. At the diagnosis of leukemia, two BM-MSCs had cytogenetic aberration, which were completely different to their BM-HCs counterpart. After the completion of anti-leukemic therapy, cytogenetic aberration was no longer detectable in one patient. Unexpectedly, BM-MSCs from three patients receiving TBI-based allogeneic transplantation acquired new, clonal cytogenetic abnormalities after transplantation. Similarly, complex cytogenetic abnormalities were found in all the three BM-MSCs exposed to in vitro irradiation. In conclusion, anti-leukemic treatments induce not only "cytogenetic remission" but also new cytogenetic abnormalities of BM-MSCs. TBI especially exerts detrimental effect on the chromosomal integrity of BM-MSCs and highlights the equal importance of investigating long-term adverse effect of anti-leukemic therapy on BM-MSCs as opposed to beneficial effect on BM-HCs.

  2. Ex vivo assays to study self-renewal, long-term expansion, and leukemic transformation of genetically modified human hematopoietic and patient-derived leukemic stem cells

    NARCIS (Netherlands)

    Sontakke, Pallavi; Carretta, Marco; Capala, Marta; Schepers, Hein; Schuringa, Jan Jacob

    2014-01-01

    With the emergence of the concept of the leukemic stem cell (LSC), assays to study them remain pivotal in understanding (leukemic) stem cell biology. Although the in vivo NOD-SCID or NSG xenotransplantation model is currently still the favored assay of choice in most cases, this system has some

  3. Targeting Toll-like receptor 7/8 enhances uptake of apoptotic leukemic cells by monocyte-derived dendritic cells but interferes with subsequent cytokine-induced maturation.

    Science.gov (United States)

    van den Ancker, Willemijn; van Luijn, Marvin M; Ruben, Jurjen M; Westers, Theresia M; Bontkes, Hetty J; Ossenkoppele, Gert J; de Gruijl, Tanja D; van de Loosdrecht, Arjan A

    2011-01-01

    Therapeutic vaccination with dendritic cells (DC) is an emerging investigational therapy for eradication of minimal residual disease in acute myeloid leukemia. Various strategies are being explored in manufacturing DC vaccines ex vivo, e.g., monocyte-derived DC (MoDC) loaded with leukemia-associated antigens (LAA). However, the optimal source of LAA and the choice of DC-activating stimuli are still not well defined. Here, loading with leukemic cell preparations (harboring both unknown and known LAA) was explored in combination with a DC maturation-inducing cytokine cocktail (CC; IL-1β, IL-6, TNF-α, and PGE(2)) and Toll-like receptor ligands (TLR-L) to optimize uptake. Since heat shock induced apoptotic blasts were more efficiently taken up than lysates, we focused on uptake of apoptotic leukemic cells. Uptake of apoptotic blast was further enhanced by the TLR7/8-L R848 (20-30%); in contrast, CC-induced maturation inhibited uptake. CC, and to a lesser extent R848, enhanced the ability of MoDC to migrate and stimulate T cells. Furthermore, class II-associated invariant chain peptide expression was down-modulated after R848- or CC-induced maturation, indicating enhanced processing and presentation of antigenic peptides. To improve both uptake and maturation, leukemic cells and MoDC were co-incubated with R848 for 24 h followed by addition of CC. However, this approach interfered with CC-mediated MoDC maturation as indicated by diminished migratory and T cell stimulatory capacity, and the absence of IL-12 production. Taken together, our data demonstrate that even though R848 improved uptake of apoptotic leukemic cells, the sequential use of R848 and CC is counter-indicated due to its adverse effects on MoDC maturation.

  4. Leukemic cell labeling with indium-111-oxine

    International Nuclear Information System (INIS)

    Uchida, T.; Takagi, Y.; Matsuda, S.; Yui, T.; Ishibashi, T.; Kimura, H.; Kariyone, S.

    1984-01-01

    Leukemic cells were labeled with In-111-oxine in patients with acute leukemia. In vitro labeling studies revealed that labeling efficiency reached maximum 80.8 +- 3.6% (mean +- 1SD) by 2 times washes after 20 minutes incubation time. Cell viability was assessed by trypan blue exclusion test and in vitro culture of leukemic cells, which showed no cellular damage during labeling procedure. Elution of In-111 from the labeled cells was 10.0 +- 1.2% at 12 hours after labeling. For in vivo leukemic cell kinetic studies, more than 10/sup 8/ leukemic cells separated from Ficoll-Hypacque sedimentation were labeled by 30 minutes of In-111-oxine incubation and two times washes at 37 0 C. In vivo studies were performed in 7 patients with acute myeloblastic, lymphoblastic leukemia and blastic crisis of chronic myelocytic leukemia. Labeled leukemic cells disappeared in single exponential fashion with half life of 9.6 to 31.8 hours. Total leukemic cell pool in peripheral circulation was calculated, which correlated well with peripheral leukemic cell counts (r=0.99). No relationship was observed between total leukemic cell pool and leukemic cell turnover rate. Migration patterns of labeled leukemic cells showed that pulmonary uptake was evident within 15 minutes after the infusion and returned to base-line. Splenic and hepatic uptake showed gradual increase up to 24 hours. Bone marrow accumulation was shown only in 2 cases. Presently, there are no suitable radionuclides for leukemic cell labeling. In-111-oxine labeled leukemic cells would overcome this difficulty

  5. Nature of leukemic stem cells in murine myelogenous leukemia

    International Nuclear Information System (INIS)

    Yoshida, K.; Nemoto, K.; Nishimura, M.; Hayata, I.; Inoue, T.; Seki, M.

    1986-01-01

    We investigated the nature of myelogenous leukemic stem cells in mice. L-8057, a megakaryoblastic leukemia cell line used in this study, produces in vivo and in vitro colonies. By means of typical chromosomal aberrations in L-8057, one can conveniently detect the origin of the cells in each colony derived from a leukemic stem cell. Direct evidence of whether cells from each colony had leukemogenicity in recipient mice was successfully obtained by the colony transplantation assay. Both leukemic colony-forming unit-spleen (L-CFU-s) and leukemic colony-forming unit-culture (L-CFU-c) in L-8057 may have belonged to the same differentiating stage in the stem cells because of their similar radiosensitivity, although some parts of the L-CFU of L-8057 seemed to have lost their capability to regenerate L-CFU-s when the cells were plated in dishes. This leukemic stem cell preserves high self-renewal ability in vitro after 10 passages. In addition, in vitro colony formation by this leukemic cell during the above course of serial passages did not require any additional exogenous stimulators. The same sort of trials have been made on other types of leukemias. Leukemic stem cells showed remarkable variety in their response to stimulating factors and in their self-renewal activity, which suggests that they may have consisted of heterogeneous populations

  6. Nuclear accumulation of SHIP1 mutants derived from AML patients leads to increased proliferation of leukemic cells.

    Science.gov (United States)

    Nalaskowski, Marcus M; Ehm, Patrick; Rehbach, Christoph; Nelson, Nina; Täger, Maike; Modest, Kathrin; Jücker, Manfred

    2018-05-28

    The inositol 5-phosphatase SHIP1 acts as negative regulator of intracellular signaling in myeloid cells and is a tumor suppressor in myeloid leukemogenesis. After relocalization from the cytoplasm to the plasma membrane SHIP1 terminates PI3-kinase mediated signaling processes. Furthermore, SHIP1 is also found in distinct puncta in the cell nucleus and nuclear SHIP1 has a pro-proliferative function. Here we report the identification of five nuclear export signals (NESs) which regulate together with the two known nuclear localization signals (NLSs) the nucleocytoplasmic shuttling of SHIP1. Mutation of NLSs reduced the nuclear import and mutation of NESs decreased the nuclear export of SHIP1 in the acute myeloid leukemia (AML) cell line UKE-1. Interestingly, four SHIP1 mutants (K210R, N508D, V684E, Q1153L) derived from AML patients showed a nuclear accumulation after expression in UKE-1 cells. In addition, overexpression of the AML patient-derived mutation N508D caused an increased proliferation rate of UKE-1 cells in comparison to wild type SHIP1. Furthermore, we identified serine and tyrosine phosphorylation as a molecular mechanism for the regulation of nucleocytoplasmic shuttling of SHIP1 where tyrosine phosphorylation of distinct residues i.e. Y864, Y914, Y1021 reduces nuclear localization, whereas serine phosphorylation at S933 enhances nuclear localization of SHIP1. In summary, our data further implicate nuclear SHIP1 in cellular signaling and suggest that enhanced accumulation of SHIP1 mutants in the nucleus may be a contributory factor of abnormally high proliferation of AML cells. Copyright © 2017. Published by Elsevier Inc.

  7. An evidence for adhesion-mediated acquisition of acute myeloid leukemic stem cell-like immaturities

    International Nuclear Information System (INIS)

    Funayama, Keiji; Shimane, Miyuki; Nomura, Hitoshi; Asano, Shigetaka

    2010-01-01

    For long-term survival in vitro and in vivo of acute myeloid leukemia cells, their adhesion to bone marrow stromal cells is indispensable. However, it is still unknown if these events are uniquely induced by the leukemic stem cells. Here we show that TF-1 human leukemia cells, once they have formed a cobblestone area by adhering to mouse bone marrow-derived MS-5 cells, can acquire some leukemic stem cell like properties in association with a change in the CD44 isoform-expression pattern and with an increase in a set of related microRNAs. These findings strongly suggest that at least some leukemia cells can acquire leukemic stem cell like properties in an adhesion-mediated stochastic fashion.

  8. An evidence for adhesion-mediated acquisition of acute myeloid leukemic stem cell-like immaturities

    Energy Technology Data Exchange (ETDEWEB)

    Funayama, Keiji; Shimane, Miyuki; Nomura, Hitoshi [Department of Integrative Bioscience and Biomedical Engineering, Waseda University, 4-3-1 Ohkubo, Shinjuku-ku, Tokyo 169-8555 (Japan); Asano, Shigetaka, E-mail: asgtkmd@waseda.jp [Department of Integrative Bioscience and Biomedical Engineering, Waseda University, 4-3-1 Ohkubo, Shinjuku-ku, Tokyo 169-8555 (Japan)

    2010-02-12

    For long-term survival in vitro and in vivo of acute myeloid leukemia cells, their adhesion to bone marrow stromal cells is indispensable. However, it is still unknown if these events are uniquely induced by the leukemic stem cells. Here we show that TF-1 human leukemia cells, once they have formed a cobblestone area by adhering to mouse bone marrow-derived MS-5 cells, can acquire some leukemic stem cell like properties in association with a change in the CD44 isoform-expression pattern and with an increase in a set of related microRNAs. These findings strongly suggest that at least some leukemia cells can acquire leukemic stem cell like properties in an adhesion-mediated stochastic fashion.

  9. Molecular dynamics study of lipid bilayers modeling the plasma membranes of normal murine thymocytes and leukemic GRSL cells.

    Science.gov (United States)

    Andoh, Yoshimichi; Okazaki, Susumu; Ueoka, Ryuichi

    2013-04-01

    Molecular dynamics (MD) calculations for the plasma membranes of normal murine thymocytes and thymus-derived leukemic GRSL cells in water have been performed under physiological isothermal-isobaric conditions (310.15K and 1 atm) to investigate changes in membrane properties induced by canceration. The model membranes used in our calculations for normal and leukemic thymocytes comprised 23 and 25 kinds of lipids, respectively, including phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, phosphatidylinositol, sphingomyelin, lysophospholipids, and cholesterol. The mole fractions of the lipids adopted here were based on previously published experimental values. Our calculations clearly showed that the membrane area was increased in leukemic cells, and that the isothermal area compressibility of the leukemic plasma membranes was double that of normal cells. The calculated membranes of leukemic cells were thus considerably bulkier and softer in the lateral direction compared with those of normal cells. The tilt angle of the cholesterol and the conformation of the phospholipid fatty acid tails both showed a lower level of order in leukemic cell membranes compared with normal cell membranes. The lateral radial distribution function of the lipids also showed a more disordered structure in leukemic cell membranes than in normal cell membranes. These observations all show that, for the present thymocytes, the lateral structure of the membrane is considerably disordered by canceration. Furthermore, the calculated lateral self-diffusion coefficient of the lipid molecules in leukemic cell membranes was almost double that in normal cell membranes. The calculated rotational and wobbling autocorrelation functions also indicated that the molecular motion of the lipids was enhanced in leukemic cell membranes. Thus, here we have demonstrated that the membranes of thymocyte leukemic cells are more disordered and more fluid than normal cell membranes. Copyright © 2013

  10. Leukemic Cells "Gas Up" Leaky Bone Marrow Blood Vessels.

    Science.gov (United States)

    Itkin, Tomer; Rafii, Shahin

    2017-09-11

    In this issue of Cancer Cell, Passaro et al. demonstrate how leukemia through aberrant induction of reactive oxygen species and nitric oxide production trigger marrow vessel leakiness, instigating pro-leukemic function. Disrupted tumor blood vessels promote exhaustion of non-malignant stem and progenitor cells and may facilitate leukemia relapse following chemotherapeutic treatment. Copyright © 2017. Published by Elsevier Inc.

  11. Cannabidiol Reduces Leukemic Cell Size – But Is It Important?

    Science.gov (United States)

    Kalenderoglou, Nikoletta; Macpherson, Tara; Wright, Karen L.

    2017-01-01

    The anti-cancer effect of the plant-derived cannabinoid, cannabidiol, has been widely demonstrated both in vivo and in vitro. However, this body of preclinical work has not been translated into clinical use. Key issues around this failure can be related to narrow dose effects, the cell model used and incomplete efficacy. A model of acute lymphoblastic disease, the Jurkat T cell line, has been used extensively to study the cannabinoid system in the immune system and cannabinoid-induced apoptosis. Using these cells, this study sought to investigate the outcome of those remaining viable cells post-treatment with cannabidiol, both in terms of cell size and tracking any subsequent recovery. The phosphorylation status of the mammalian Target of Rapamycin (mTOR) signaling pathway and the downstream target ribosomal protein S6, were measured. The ability of cannabidiol to exert its effect on cell viability was also evaluated in physiological oxygen conditions. Cannabidiol reduced cell viability incompletely, and slowed the cell cycle with fewer cells in the G2/M phase of the cell cycle. Cannabidiol reduced phosphorylation of mTOR, PKB and S6 pathways related to survival and cell size. The remaining population of viable cells that were cultured in nutrient rich conditions post-treatment were able to proliferate, but did not recover to control cell numbers. However, the proportion of viable cells that were gated as small, increased in response to cannabidiol and normally sized cells decreased. This proportion of small cells persisted in the recovery period and did not return to basal levels. Finally, cells grown in 12% oxygen (physiological normoxia) were more resistant to cannabidiol. In conclusion, these results indicate that cannabidiol causes a reduction in cell size, which persists post-treatment. However, resistance to cannabidiol under physiological normoxia for these cells would imply that cannabidiol may not be useful in the clinic as an anti-leukemic agent. PMID

  12. Cannabidiol Reduces Leukemic Cell Size - But Is It Important?

    Science.gov (United States)

    Kalenderoglou, Nikoletta; Macpherson, Tara; Wright, Karen L

    2017-01-01

    The anti-cancer effect of the plant-derived cannabinoid, cannabidiol, has been widely demonstrated both in vivo and in vitro . However, this body of preclinical work has not been translated into clinical use. Key issues around this failure can be related to narrow dose effects, the cell model used and incomplete efficacy. A model of acute lymphoblastic disease, the Jurkat T cell line, has been used extensively to study the cannabinoid system in the immune system and cannabinoid-induced apoptosis. Using these cells, this study sought to investigate the outcome of those remaining viable cells post-treatment with cannabidiol, both in terms of cell size and tracking any subsequent recovery. The phosphorylation status of the mammalian Target of Rapamycin (mTOR) signaling pathway and the downstream target ribosomal protein S6, were measured. The ability of cannabidiol to exert its effect on cell viability was also evaluated in physiological oxygen conditions. Cannabidiol reduced cell viability incompletely, and slowed the cell cycle with fewer cells in the G2/M phase of the cell cycle. Cannabidiol reduced phosphorylation of mTOR, PKB and S6 pathways related to survival and cell size. The remaining population of viable cells that were cultured in nutrient rich conditions post-treatment were able to proliferate, but did not recover to control cell numbers. However, the proportion of viable cells that were gated as small, increased in response to cannabidiol and normally sized cells decreased. This proportion of small cells persisted in the recovery period and did not return to basal levels. Finally, cells grown in 12% oxygen (physiological normoxia) were more resistant to cannabidiol. In conclusion, these results indicate that cannabidiol causes a reduction in cell size, which persists post-treatment. However, resistance to cannabidiol under physiological normoxia for these cells would imply that cannabidiol may not be useful in the clinic as an anti-leukemic agent.

  13. CD25 targeted therapy of chemotherapy resistant leukemic stem cells using DR5 specific TRAIL peptide

    Directory of Open Access Journals (Sweden)

    Jayaprakasam Madhumathi

    2017-03-01

    Full Text Available Chemotherapy resistant leukemic stem cells (LSCs are being targeted as a modern therapeutic approach to prevent disease relapse. LSCs isolated from methotrexate resistant side population (SP of leukemic cell lines HL60 and MOLT4 exhibited high levels of CD25 and TRAIL R2/DR5 which are potential targets. Recombinant immunotoxin conjugating IL2α with TRAIL peptide mimetic was constructed for DR5 receptor specific targeting of LSCs and were tested in total cell population and LSCs. IL2-TRAIL peptide induced apoptosis in drug resistant SP cells from cell lines and showed potent cytotoxicity in PBMCs derived from leukemic patients with an efficacy of 81.25% in AML and 100% in CML, ALL and CLL. IL2-TRAIL peptide showed cytotoxicity in relapsed patient samples and was more effective than TRAIL or IL2-TRAIL proteins. Additionally, DR5 specific IL2-TRAIL peptide was effective in targeting and killing LSCs purified from cell lines [IC50: 952 nM in HL60, 714 nM in MOLT4] and relapsed patient blood samples with higher efficacy (85% than IL2-TRAIL protein (46%. Hence, CD25 and DR5 specific targeting by IL2-TRAIL peptide may be an effective strategy for targeting drug resistant leukemic cells and LSCs.

  14. In vitro gamma irradiation Medical Center of leukemic cells in mice, rats, and guinea pigs

    International Nuclear Information System (INIS)

    Gross, L.; Dreyfuss, Y.; Ehrenreich, T.; Feldman, D.; Limbert, L.M.

    1980-01-01

    In vitro gamma irradiation of virus-induced (Gross) mouse leukemia cells at doses of 350 to 1600 rads (1 rad = 0.01 gray) had no effect on their ability to induce leukemia, usually within 2 weeks, after transplantation into syngeneic mice. However, when cells irradiated at doses of 2000-20,000 rads were transplanted, they induced leukemia after a latency period exceeding 2.5 months, similar to the results observed in mice inoculated with filtered mouse leukemia extracts. Similar results were also obtained after irradiation of leukemic cells derived from rats in which leukemia had been induced by rat-adapted mouse leukemia virus. Apparently, gamma irradiation at a dose of, or exceeding, 2000 rads, inhibits the ability of mouse and rat leukemic cells to induce leukemia after transplantation into syngeneic hosts; however, it does not inactivate the virus carried by such cells nor prevent it from inducing leukemia. [In previous experiments, doses of more than 4,500,000 rads were needed to inactivate the passage A (Gross) leukemia virus carried in either mouse or rat leukemic cells.] In vitro gamma irradiation of L2C guinea pig leukemic cells at doses of 750 to 2500 rads had no apparent effect on their ability to induce leukemia after transplantation into strain 2 guinea pigs. However, irradiation at doses of 3250 to 20,000 rads inactivated their ability to do so. The morphology of mouse, rat, and guinea pig leukemic cells and the virus particles present in such cells was not affected by irradiation at doses of 20,000 rads

  15. Identification of hepatic niche harboring human acute lymphoblastic leukemic cells via the SDF-1/CXCR4 axis.

    Directory of Open Access Journals (Sweden)

    Itaru Kato

    Full Text Available In acute lymphoblastic leukemia (ALL patients, the bone marrow niche is widely known to be an important element of treatment response and relapse. Furthermore, a characteristic liver pathology observed in ALL patients implies that the hepatic microenvironment provides an extramedullary niche for leukemic cells. However, it remains unclear whether the liver actually provides a specific niche. The mechanism underlying this pathology is also poorly understood. Here, to answer these questions, we reconstituted the histopathology of leukemic liver by using patients-derived primary ALL cells into NOD/SCID/Yc (null mice. The liver pathology in this model was similar to that observed in the patients. By using this model, we clearly demonstrated that bile duct epithelial cells form a hepatic niche that supports infiltration and proliferation of ALL cells in the liver. Furthermore, we showed that functions of the niche are maintained by the SDF-1/CXCR4 axis, proposing a novel therapeutic approach targeting the extramedullary niche by inhibition of the SDF-1/CXCR4 axis. In conclusion, we demonstrated that the liver dissemination of leukemia is not due to nonselective infiltration, but rather systematic invasion and proliferation of leukemic cells in hepatic niche. Although the contribution of SDF-1/CXCR4 axis is reported in some cancer cells or leukemic niches such as bone marrow, we demonstrated that this axis works even in the extramedullary niche of leukemic cells. Our findings form the basis for therapeutic approaches that target the extramedullary niche by inhibiting the SDF-1/CXCR4 axis.

  16. Autocrine stimulation of VEGFR-2 activates human leukemic cell growth and migration

    Science.gov (United States)

    Dias, Sergio; Hattori, Koichi; Zhu, Zhenping; Heissig, Beate; Choy, Margaret; Lane, William; Wu, Yan; Chadburn, Amy; Hyjek, Elizabeth; Gill, Muhammad; Hicklin, Daniel J.; Witte, Larry; Moore, M.A.S.; Rafii, Shahin

    2000-01-01

    Emerging data suggest that VEGF receptors are expressed by endothelial cells as well as hematopoietic stem cells. Therefore, we hypothesized that functional VEGF receptors may also be expressed in malignant counterparts of hematopoietic stem cells such as leukemias. We demonstrate that certain leukemias not only produce VEGF but also express functional VEGFR-2 in vivo and in vitro, resulting in the generation of an autocrine loop that may support leukemic cell survival and proliferation. Approximately 50% of freshly isolated leukemias expressed mRNA and protein for VEGFR-2. VEGF165 induced phosphorylation of VEGFR-2 and increased proliferation of leukemic cells, demonstrating these receptors were functional. VEGF165 also induced the expression of MMP-9 by leukemic cells and promoted their migration through reconstituted basement membrane. The neutralizing mAb IMC-1C11, specific to human VEGFR-2, inhibited leukemic cell survival in vitro and blocked VEGF165-mediated proliferation of leukemic cells and VEGF-induced leukemic cell migration. Xenotransplantation of primary leukemias and leukemic cell lines into immunocompromised nonobese diabetic mice resulted in significant elevation of human, but not murine, VEGF in plasma and death of inoculated mice within 3 weeks. Injection of IMC-1C11 inhibited proliferation of xenotransplanted human leukemias and significantly increased the survival of inoculated mice. Interruption of signaling by VEGFRs, particularly VEGFR-2, may provide a novel strategy for inhibiting leukemic cell proliferation. PMID:10953026

  17. Normal and Leukemic Hematopoiesis

    NARCIS (Netherlands)

    Vercauteren, Suzanne Maria

    2003-01-01

    Acute Myeloid Leukemia (AML) is a clonal myeloproliferative disease characterized by an uncontrolled proliferation and block in differentiation of myeloid committed blood cells in the bone marrow. Despite the lack of mature cells derived from the leukemic clone in the majority of AML patients, AML

  18. Correlation of total body potassium and leukemic cell mass in patients with chronic lymphocytic leukemia

    International Nuclear Information System (INIS)

    Chandra, P.; Sawitsky, A.; Chanana, A.D.; Chikkappa, G.; Cohn, S.H.; Rai, K.R.; Cronkity, E.P.

    1979-01-01

    Total body leukemic mass in patients with chronic lymphocytic leukemia (CLL) was measured by quantitation of total body potassium (TBK) with a whole-body counter. In addition, the predicted normal total body potassium (Kp) for each patient was calculated from an empirically derived relationship involving height, weight age, and sex. Both the absolute TBK and the relative excess of total body potassium (TBK/Kp) were related to the stage of disease. Patients in the early stages of CLL were found to have lower TBK and TBK/Kp than patients in the late stages of disease. Both of these parameters increased with the successively advanced stages of the disease. The clinically monitored reduction of leukemic cell mass following therapy was accompanied by reductions in TBK and TBK/Kp. Data presented support the notion that TBK/Kp is a useful indicator of the total body leukemic mass. Futhermore, the results of these studies quantitatively validate the proposed clinical staging system for CLL. Quantitation of TBK by a whole-body counter is an accurate and noninvasive procedure and does not require administration of isotopes

  19. Altered expression of asparagine synthetase mRNA in human leukemic and carcinoma cell lines

    Energy Technology Data Exchange (ETDEWEB)

    Goodwin, L.O.; Guzowski, D.E.; Millan, C.A. [North Shore Univ. Hospital/Cornell Univ. Medical College, Manhasset, NY (United States)] [and others

    1994-09-01

    Asparagine synthetase (AS) is the enzyme responsible for the ATP-dependant conversion of aspartic acid to asparagine. The AS gene is expressed constitutively in most mammalian cells, including cells of the lymphoid lineage, as a 2 kb mRNA. In some leukemic phenotypes, AS expression is abrogated, resulting in no detectable enzyme activity. These cells are rendered sensitive to killing by L-asparaginase, which destroys extracellular asparagine. Prolonged treatment of leukemic cells with this agent can lead to resistance and the reappearance of AS activity, suggesting derepression of the AS gene, which has been shown to be regulated by intracellular levels of asparagine. Modulation of AS expression by asparagine employs cis and trans-acting elements involved in transcriptional and translational regulation. We have cloned and sequenced the human AS gene and surrounding sequence elements as well as the full-length cDNA. Using probes specific to the third and fourth exons of AS, we have identified an additional higher molecular weight mRNA (2.7 kb) in Northern blots derived from a chronic myelogenous leukemia and a colon carcinoma but not in normal lymphocytic or other human cell lines. We speculate that elements present in the cancer-derived mRNAs may be involved in the derepression of AS activity. This hypothesis is being evaluated by RNase protection assays using RNA isolated from a variety of human cell lines to characterize and elucidate the nature of this additional AS encoded message.

  20. Idarubicin induces mTOR-dependent cytotoxic autophagy in leukemic cells

    International Nuclear Information System (INIS)

    Ristic, Biljana; Bosnjak, Mihajlo; Arsikin, Katarina; Mircic, Aleksandar; Suzin-Zivkovic, Violeta; Bogdanovic, Andrija; Perovic, Vladimir; Martinovic, Tamara; Kravic-Stevovic, Tamara; Bumbasirevic, Vladimir; Trajkovic, Vladimir; Harhaji-Trajkovic, Ljubica

    2014-01-01

    We investigated if the antileukemic drug idarubicin induces autophagy, a process of programmed cellular self-digestion, in leukemic cell lines and primary leukemic cells. Transmission electron microscopy and acridine orange staining demonstrated the presence of autophagic vesicles and intracellular acidification, respectively, in idarubicin-treated REH leukemic cell line. Idarubicin increased punctuation/aggregation of microtubule-associated light chain 3B (LC3B), enhanced the conversion of LC3B-I to autophagosome-associated LC3B-II in the presence of proteolysis inhibitors, and promoted the degradation of the selective autophagic target p62, thus indicating the increase in autophagic flux. Idarubicin inhibited the phosphorylation of the main autophagy repressor mammalian target of rapamycin (mTOR) and its downstream target p70S6 kinase. The treatment with the mTOR activator leucine prevented idarubicin-mediated autophagy induction. Idarubicin-induced mTOR repression was associated with the activation of the mTOR inhibitor AMP-activated protein kinase and down-regulation of the mTOR activator Akt. The suppression of autophagy by pharmacological inhibitors or LC3B and beclin-1 genetic knockdown rescued REH cells from idarubicin-mediated oxidative stress, mitochondrial depolarization, caspase activation and apoptotic DNA fragmentation. Idarubicin also caused mTOR inhibition and cytotoxic autophagy in K562 leukemic cell line and leukocytes from chronic myeloid leukemia patients, but not healthy controls. By demonstrating mTOR-dependent cytotoxic autophagy in idarubicin-treated leukemic cells, our results warrant caution when considering combining idarubicin with autophagy inhibitors in leukemia therapy. - Highlights: • Idarubicin induces autophagy in leukemic cell lines and primary leukemic cells. • Idarubicin induces autophagy by inhibiting mTOR in leukemic cells. • mTOR suppression by idarubicin is associated with AMPK activation and Akt blockade.

  1. Idarubicin induces mTOR-dependent cytotoxic autophagy in leukemic cells

    Energy Technology Data Exchange (ETDEWEB)

    Ristic, Biljana [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Bosnjak, Mihajlo [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Arsikin, Katarina [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Mircic, Aleksandar; Suzin-Zivkovic, Violeta [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Bogdanovic, Andrija [Clinic for Hematology, Clinical Centre of Serbia, School of Medicine, University of Belgrade, Belgrade (Serbia); Perovic, Vladimir [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Martinovic, Tamara; Kravic-Stevovic, Tamara; Bumbasirevic, Vladimir [Institute of Histology and Embryology, School of Medicine, University of Belgrade, Belgrade (Serbia); Trajkovic, Vladimir, E-mail: vtrajkovic@med.bg.ac.rs [Institute of Microbiology and Immunology, School of Medicine, University of Belgrade, Dr. Subotica 1, 11000 Belgrade (Serbia); Harhaji-Trajkovic, Ljubica, E-mail: buajk@yahoo.com [Institute for Biological Research, University of Belgrade, Belgrade, Despot Stefan Blvd. 142, 11000 Belgrade (Serbia)

    2014-08-01

    We investigated if the antileukemic drug idarubicin induces autophagy, a process of programmed cellular self-digestion, in leukemic cell lines and primary leukemic cells. Transmission electron microscopy and acridine orange staining demonstrated the presence of autophagic vesicles and intracellular acidification, respectively, in idarubicin-treated REH leukemic cell line. Idarubicin increased punctuation/aggregation of microtubule-associated light chain 3B (LC3B), enhanced the conversion of LC3B-I to autophagosome-associated LC3B-II in the presence of proteolysis inhibitors, and promoted the degradation of the selective autophagic target p62, thus indicating the increase in autophagic flux. Idarubicin inhibited the phosphorylation of the main autophagy repressor mammalian target of rapamycin (mTOR) and its downstream target p70S6 kinase. The treatment with the mTOR activator leucine prevented idarubicin-mediated autophagy induction. Idarubicin-induced mTOR repression was associated with the activation of the mTOR inhibitor AMP-activated protein kinase and down-regulation of the mTOR activator Akt. The suppression of autophagy by pharmacological inhibitors or LC3B and beclin-1 genetic knockdown rescued REH cells from idarubicin-mediated oxidative stress, mitochondrial depolarization, caspase activation and apoptotic DNA fragmentation. Idarubicin also caused mTOR inhibition and cytotoxic autophagy in K562 leukemic cell line and leukocytes from chronic myeloid leukemia patients, but not healthy controls. By demonstrating mTOR-dependent cytotoxic autophagy in idarubicin-treated leukemic cells, our results warrant caution when considering combining idarubicin with autophagy inhibitors in leukemia therapy. - Highlights: • Idarubicin induces autophagy in leukemic cell lines and primary leukemic cells. • Idarubicin induces autophagy by inhibiting mTOR in leukemic cells. • mTOR suppression by idarubicin is associated with AMPK activation and Akt blockade.

  2. Novel therapeutic strategies to target leukemic cells that hijack compartmentalized continuous hematopoietic stem cell niches

    NARCIS (Netherlands)

    Hira, Vashendriya V. V.; van Noorden, Cornelis J. F.; Carraway, Hetty E.; Maciejewski, Jaroslaw P.; Molenaar, Remco J.

    2017-01-01

    Acute myeloid leukemia and acute lymphoblastic leukemia cells hijack hematopoietic stem cell (HSC) niches in the bone marrow and become leukemic stem cells (LSCs) at the expense of normal HSCs. LSCs are quiescent and resistant to chemotherapy and can cause relapse of the disease. HSCs in niches are

  3. The in-vitro study of human blood leukemic cells by pulsed NMR

    International Nuclear Information System (INIS)

    Zulkarnaen, M.; Munawir; Wibowo, Tono; Suyitno, Gogot

    1983-01-01

    The diagram of leukemic cells in human blood has been studied by using the NMR longitudinal relaxation technique. The observation was treated in whole blood, serum and blood cell. Every result was compared with previous observation and show that the values of the proton longitudinal relaxation in the leukemic whole blood almost twice or more that of normal blood, while in the serum and the blood cell, the values are nearly the same. (author)

  4. Novel therapeutic strategies to target leukemic cells that hijack compartmentalized continuous hematopoietic stem cell niches.

    Science.gov (United States)

    Hira, Vashendriya V V; Van Noorden, Cornelis J F; Carraway, Hetty E; Maciejewski, Jaroslaw P; Molenaar, Remco J

    2017-08-01

    Acute myeloid leukemia and acute lymphoblastic leukemia cells hijack hematopoietic stem cell (HSC) niches in the bone marrow and become leukemic stem cells (LSCs) at the expense of normal HSCs. LSCs are quiescent and resistant to chemotherapy and can cause relapse of the disease. HSCs in niches are needed to generate blood cell precursors that are committed to unilineage differentiation and eventually production of mature blood cells, including red blood cells, megakaryocytes, myeloid cells and lymphocytes. Thus far, three types of HSC niches are recognized: endosteal, reticular and perivascular niches. However, we argue here that there is only one type of HSC niche, which consists of a periarteriolar compartment and a perisinusoidal compartment. In the periarteriolar compartment, hypoxia and low levels of reactive oxygen species preserve the HSC pool. In the perisinusoidal compartment, hypoxia in combination with higher levels of reactive oxygen species enables proliferation of progenitor cells and their mobilization into the circulation. Because HSC niches offer protection to LSCs against chemotherapy, we review novel therapeutic strategies to inhibit homing of LSCs in niches for the prevention of dedifferentiation of leukemic cells into LSCs and to stimulate migration of leukemic cells out of niches. These strategies enhance differentiation and proliferation and thus sensitize leukemic cells to chemotherapy. Finally, we list clinical trials of therapies that tackle LSCs in HSC niches to circumvent their protection against chemotherapy. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  5. Effect of dioxin on normal and leukemic human hematopoietic cells

    Energy Technology Data Exchange (ETDEWEB)

    Lambertenghi-Deliliers, G.; Soligo, D. [Univ. degli Studi, Milan (Italy). Dipt. die Ematologia, Ospedale Maggiore Policlinico IRCCS; Fracchiolla, N.S. [Ospedale Maggiore Policlinico IRCCS, Milan (Italy). Dipt. di Ematologia; Servida, F. [Fondazione Matarelli, Milan (Italy); Bertazzi, P.A. [Istituti Clinici di Perfezionamento, Milan (Italy). Dipt. di Medicina del Lavoro

    2004-09-15

    2,3,7,8-Tetrachlorodibenzo-p-dioxin (TCDD) arises from chlorination of phenolic substrates or from partial combustion of organic materials in the presence of chlorine sources. TCDD has a large number of biological effects such as long-lasting skin disease, cardiovascular disease, diabete and cancer. TCDD is the prototypical agonist of the aryl hydrocarbon receptor (AhR), a member of the erb-A family that also includes the receptors for steroids, thyroid hormones, peroxisome proliferators and retinoids. When bound to dioxin, the AhR can bind to DNA and alter the expression of some genes including cytokines and growth factors. In this study, we analyzed the effect of escalating doses of TCDD on human CD34{sup +} progenitor cells from the leukapheresis of normal donors stimulated with G-CSF as well as the human myeloid leukemic cell lines HL60 (promyelocytic leukemia) and K562 (chronic myelogenous leukemia). The possible specific modulation of gene expression induced by the TCDD exposure was then tested by means of microarray analyses.

  6. Analysis of the surface membrane of iodinated leukemic cells by SDS-polyacrylamide gel electrophoresis

    International Nuclear Information System (INIS)

    Ishitani, Kunihiko; Ikeda, Akira; Tamura, Minoru; Takeuchi, Hidekazu; Ihara, Koji

    1980-01-01

    Surface proteins of human leukemic cells were labeled selectively by lactoperoxydase catalysed-iodination and examined by SDS-polyacrylamide gel electrophoresis. The electrophoretic pattern of the surface membranes of cells from a patients with chronic mylogeneous leukemia in blast crisis was of B cell type and showed Ia like antigen. Leukemic cells from a patient with hairly cell leukemia also expressed the pattern of B cell type when tested by this method the technique of iodinating cell surface with lactoperoxidase is useful in characterization of leukemia cells for diagnosis and monitoring of clinical course. (author)

  7. Exploiting mitochondrial dysfunction for effective elimination of imatinib-resistant leukemic cells.

    Directory of Open Access Journals (Sweden)

    Jérome Kluza

    Full Text Available Challenges today concern chronic myeloid leukemia (CML patients resistant to imatinib. There is growing evidence that imatinib-resistant leukemic cells present abnormal glucose metabolism but the impact on mitochondria has been neglected. Our work aimed to better understand and exploit the metabolic alterations of imatinib-resistant leukemic cells. Imatinib-resistant cells presented high glycolysis as compared to sensitive cells. Consistently, expression of key glycolytic enzymes, at least partly mediated by HIF-1α, was modified in imatinib-resistant cells suggesting that imatinib-resistant cells uncouple glycolytic flux from pyruvate oxidation. Interestingly, mitochondria of imatinib-resistant cells exhibited accumulation of TCA cycle intermediates, increased NADH and low oxygen consumption. These mitochondrial alterations due to the partial failure of ETC were further confirmed in leukemic cells isolated from some imatinib-resistant CML patients. As a consequence, mitochondria generated more ROS than those of imatinib-sensitive cells. This, in turn, resulted in increased death of imatinib-resistant leukemic cells following in vitro or in vivo treatment with the pro-oxidants, PEITC and Trisenox, in a syngeneic mouse tumor model. Conversely, inhibition of glycolysis caused derepression of respiration leading to lower cellular ROS. In conclusion, these findings indicate that imatinib-resistant leukemic cells have an unexpected mitochondrial dysfunction that could be exploited for selective therapeutic intervention.

  8. Differentiation-inducing effects of small fruit juices on HL-60 leukemic cells.

    Science.gov (United States)

    Yoshizawa, Y; Kawaii, S; Urashima, M; Fukase, T; Sato, T; Murofushi, N; Nishimura, H

    2000-08-01

    Epidemiological studies indicate that high intakes of fruits and vegetables are associated with a reduced risk of cancer, and several plant-derived drugs have been developed in medical oncology. Since only a small part of the flora has been tested for any kind of bioactivity, we chose small fruits as sources of differentiation-inducing activity against HL-60 leukemic cells. We have prepared juices from various small fruits that grow mainly in the northern part of Japan. Screening of 43 samples indicated that juices of Actinidia polygama Maxim., Rosa rugosa Thunb., Vaccinium smallii A. Gray, and Sorbus sambucifolia Roem. strongly induced differentiation of HL-60 cells to monocyte/macrophage characteristics in a concentration-dependent manner as indicated by histochemical and biochemical examinations.

  9. Proteinase-Activated Receptor 1 (PAR1 regulates leukemic stem cell functions.

    Directory of Open Access Journals (Sweden)

    Nicole Bäumer

    Full Text Available External signals that are mediated by specific receptors determine stem cell fate. The thrombin receptor PAR1 plays an important role in haemostasis, thrombosis and vascular biology, but also in tumor biology and angiogenesis. Its expression and function in hematopoietic stem cells is largely unknown. Here, we analyzed expression and function of PAR1 in primary hematopoietic cells and their leukemic counterparts. AML patients' blast cells expressed much lower levels of PAR1 mRNA and protein than CD34+ progenitor cells. Constitutive Par1-deficiency in adult mice did not affect engraftment or stem cell potential of hematopoietic cells. To model an AML with Par1-deficiency, we retrovirally introduced the oncogene MLL-AF9 in wild type and Par1-/- hematopoietic progenitor cells. Par1-deficiency did not alter initial leukemia development. However, the loss of Par1 enhanced leukemic stem cell function in vitro and in vivo. Re-expression of PAR1 in Par1-/- leukemic stem cells delayed leukemogenesis in vivo. These data indicate that Par1 contributes to leukemic stem cell maintenance.

  10. Proteinase-Activated Receptor 1 (PAR1) regulates leukemic stem cell functions.

    Science.gov (United States)

    Bäumer, Nicole; Krause, Annika; Köhler, Gabriele; Lettermann, Stephanie; Evers, Georg; Hascher, Antje; Bäumer, Sebastian; Berdel, Wolfgang E; Müller-Tidow, Carsten; Tickenbrock, Lara

    2014-01-01

    External signals that are mediated by specific receptors determine stem cell fate. The thrombin receptor PAR1 plays an important role in haemostasis, thrombosis and vascular biology, but also in tumor biology and angiogenesis. Its expression and function in hematopoietic stem cells is largely unknown. Here, we analyzed expression and function of PAR1 in primary hematopoietic cells and their leukemic counterparts. AML patients' blast cells expressed much lower levels of PAR1 mRNA and protein than CD34+ progenitor cells. Constitutive Par1-deficiency in adult mice did not affect engraftment or stem cell potential of hematopoietic cells. To model an AML with Par1-deficiency, we retrovirally introduced the oncogene MLL-AF9 in wild type and Par1-/- hematopoietic progenitor cells. Par1-deficiency did not alter initial leukemia development. However, the loss of Par1 enhanced leukemic stem cell function in vitro and in vivo. Re-expression of PAR1 in Par1-/- leukemic stem cells delayed leukemogenesis in vivo. These data indicate that Par1 contributes to leukemic stem cell maintenance.

  11. Defective quorum sensing of acute lymphoblastic leukemic cells: evidence of collective behavior of leukemic populations as semi-autonomous aberrant ecosystems

    Science.gov (United States)

    Patel, Sapan J; Dao, Su; Darie, Costel C; Clarkson, Bayard D

    2016-01-01

    Quorum sensing (QS) is a generic term used to describe cell-cell communication and collective decision making by bacterial and social insects to regulate the expression of specific genes in controlling cell density and other properties of the populations in response to nutrient supply or changes in the environment. QS mechanisms also have a role in higher organisms in maintaining homeostasis, regulation of the immune system and collective behavior of cancer cell populations. In the present study, we used a p190BCR-ABL driven pre-B acute lymphoblastic leukemia (ALL3) cell line derived from the pleural fluid of a terminally ill patient with ALL to test the QS hypothesis in leukemia. ALL3 cells don’t grow at low density (LD) in liquid media but grow progressively faster at increasingly high cell densities (HD) in contrast to other established leukemic cell lines that grow well at very low starting cell densities. The ALL3 cells at LD are poised to grow but shortly die without additional stimulation. Supernates of ALL3 cells (HDSN) and some other primary cells grown at HD stimulate the growth of the LD ALL3 cells without which they won’t survive. To get further insight into the activation processes we performed microarray analysis of the LD ALL3 cells after stimulation with ALL3 HDSN at days 1, 3, and 6. This screen identified several candidate genes, and we linked them to signaling networks and their functions. We observed that genes involved in lipid, cholesterol, fatty acid metabolism, and B cell activation are most up- or down-regulated upon stimulation of the LD ALL3 cells using HDSN. We also discuss other pathways that are differentially expressed upon stimulation of the LD ALL3 cells. Our findings suggest that the Ph+ ALL population achieves dominance by functioning as a collective aberrant ecosystem subject to defective quorum-sensing regulatory mechanisms. PMID:27429840

  12. IN SILICO MODELLING OF CYTOTOXIC BEHAVIOUR OF ANTI-LEUKEMIC COMPOUNDS ON HL-60 CELL LINE

    Directory of Open Access Journals (Sweden)

    David Ebuka Arthur

    2016-05-01

    Full Text Available This research employs multiple linear regression technique in the modelling of some potent anti-leukemic compounds using paDEL molecular descriptor software calculator, to identify the best relationship between the chemical structure and toxicities of the anticancer datasets against some leukemic cell lines (HL-60. Statistical parameters such as Q2 and R2pred (test set were computed to validate the strength of the model, while Williams plot was used to assess its applicability domain. The mean effects of the molecular descriptors in the models were calculated to illuminate the principal properties of the molecules responsible for their cytotoxicity.

  13. Cannabidiol Reduces Leukemic Cell Size ? But Is It Important?

    OpenAIRE

    Kalenderoglou, Nikoletta; Macpherson, Tara; Wright, Karen L.

    2017-01-01

    The anti-cancer effect of the plant-derived cannabinoid, cannabidiol, has been widely demonstrated both in vivo and in vitro. However, this body of preclinical work has not been translated into clinical use. Key issues around this failure can be related to narrow dose effects, the cell model used and incomplete efficacy. A model of acute lymphoblastic disease, the Jurkat T cell line, has been used extensively to study the cannabinoid system in the immune system and cannabinoid-induced apoptos...

  14. Positive /sup 111/In-granulocyte scintigraphy in a patient with focal leukemic blast cell infiltrations

    Energy Technology Data Exchange (ETDEWEB)

    Syrjaelae, M; Remes, K; Paavonen, T; Liewendahl, K

    1985-06-01

    A patient with acute myeloid leukemia was investigated with /sup 111/In-granulocyte scintigraphy to reveal possible sites of infection. /sup 111/In-granulocytes accumulated in areas of leukemia blast cell infiltration leading to a false-positive scintigram. This possibility must be kept in mind when studying leukemic patients using labeled leukocytes.

  15. Loss of quiescence and self-renewal capacity of hematopoietic stem cell in an in vitro leukemic niche.

    Science.gov (United States)

    Vanegas, Natalia-Del Pilar; Vernot, Jean-Paul

    2017-01-01

    Leukemic and mesenchymal stem cells interact in the leukemic microenvironment and affect each other differently. This interplay has also important implications for the hematopoietic stem cell (HSC) biology and function. This study evaluated human HSC self-renewal potential and quiescence in an in vitro leukemic niche without leukemic cells. A leukemic niche was established by co-culturing mesenchymal stem cells with a fresh conditioned medium obtained from a leukemic (REH) cell line. After 3 days, the REH-conditioned medium was removed and freshly isolated CD34+ at a density of up to 100,000 cells/ml were added to the leukemic niche. CD34+ cell evaluations (cell cycle, self-renewal gene expression and migration capacity) were performed after 3 further days of co-culture. Additionally, we preliminary investigated the soluble factors present in the leukemic niche and their effect on the mesenchymal stem cells. Statistical significance was assessed by Student's t test or the nonparametric test Kolmogorov-Smirnov. By co-culturing normal mesenchymal stem cells with the REH-conditioned medium we showed that hematopoietic stem cells, normally in a quiescent state, enter cell cycle and proliferate. This loss of quiescence was accompanied by an increased expression of Ki-67 and c-Myc, two well-known cell proliferation-associated markers. Two central regulators of quiescence GATA2 and p53 were also down regulated. Importantly, two genes involved in HSC self-renewal, Klf4 and the histone-lysine N -methyltransferase enzyme Ezh2, were severely affected. On the contrary, c-Kit expression, the stem cell factor receptor, was upregulated in hematopoietic stem cells when compared to the normal niche. Interestingly, mesenchymal stem cells incubated with the REH-conditioned medium stopped growing, showed a flattened morphology with the appearance of small vacuoles, and importantly, became positive for the senescence-associated beta-galactosidase activity. Evaluation of the leukemic

  16. Clonal evolution of pre-leukemic hematopoietic stem cells precedes human acute myeloid leukemia.

    Science.gov (United States)

    Majeti, Ravindra

    2014-01-01

    Massively parallel DNA sequencing has uncovered recurrent mutations in many human cancers. In acute myeloid leukemia (AML), cancer genome/exome resequencing has identified numerous recurrently mutated genes with an average of 5 mutations in each case of de novo AML. In order for these multiple mutations to accumulate in a single lineage of cells, they are serially acquired in clones of self-renewing hematopoietic stem cells (HSC), termed pre-leukemic HSC. Isolation and characterization of pre-leukemic HSC have shown that their mutations are enriched in genes involved in regulating DNA methylation, chromatin modifications, and the cohesin complex. On the other hand, genes involved in regulating activated signaling are generally absent. Pre-leukemic HSC have been found to persist in clinical remission and may ultimately give rise to relapsed disease through the acquisition of novel mutations. Thus, pre-leukemic HSC may constitute a key cellular reservoir that must be eradicated for long-term cures. Copyright © 2014 Elsevier Ltd. All rights reserved.

  17. Co-culture with podoplanin+ cells protects leukemic blast cells with leukemia-associated antigens in the tumor microenvironment.

    Science.gov (United States)

    Lee, Ji Yoon; Han, A-Reum; Lee, Sung-Eun; Min, Woo-Sung; Kim, Hee-Je

    2016-05-01

    Podoplanin+ cells are indispensable in the tumor microenvironment. Increasing evidence suggests that podoplanin may support the growth and metastasis of solid tumors; however, to the best of our knowledge no studies have determined whether or not podoplanin serves a supportive role in acute myeloid leukemia (AML). The effects of co‑culture with podoplanin+ cells on the cellular activities of the leukemic cells, such as apoptosis and cell proliferation, in addition to the expression of podoplanin in leukemic cells, were investigated. Due to the fact that genetic abnormalities are the primary cause of leukemogenesis, the overexpression of the fibromyalgia‑like tyrosine kinase‑3 gene in colony forming units was also examined following cell sorting. Podoplanin+ cells were found to play a protective role against apoptosis in leukemic cells and to promote cell proliferation. Tumor‑associated antigens, including Wilms' tumor gene 1 and survivin, were increased when leukemic cells were co‑cultured with podoplanin+ cells. In combination, the present results also suggest that podoplanin+ cells can function as stromal cells for blast cell retention in the AML tumor microenvironment.

  18. Leukemic meningitis in a patient with hairy cell leukemia. A case report

    International Nuclear Information System (INIS)

    Wolfe, D.W.; Scopelliti, J.A.; Boselli, B.D.

    1984-01-01

    Central nervous system involvement has not previously been described in patients with hairy cell leukemia (HCL). A patient is reported who presented with meningeal involvement as his initial symptom of HCL. Diagnosis was established by morphologic and cytochemical studies of his cerebrospinal fluid (CSF) and bone marrow. Treatment with whole-brain irradiation and intrathecal chemotherapy was successful in clearing leukemic cells from the CSF with resolution of symptoms

  19. NKp46 identifies an NKT cell subset susceptible to leukemic transformation in mouse and human

    Science.gov (United States)

    Yu, Jianhua; Mitsui, Takeki; Wei, Min; Mao, Hsiaoyin; Butchar, Jonathan P.; Shah, Mithun Vinod; Zhang, Jianying; Mishra, Anjali; Alvarez-Breckenridge, Christopher; Liu, Xingluo; Liu, Shujun; Yokohama, Akihiko; Trotta, Rossana; Marcucci, Guido; Benson, Don M.; Loughran, Thomas P.; Tridandapani, Susheela; Caligiuri, Michael A.

    2011-01-01

    IL-15 may have a role in the development of T cell large granular lymphocyte (T-LGL) or NKT leukemias. However, the mechanisms of action and the identity of the cell subset that undergoes leukemic transformation remain elusive. Here we show that in both mice and humans, NKp46 expression marks a minute population of WT NKT cells with higher activity and potency to become leukemic. Virtually 100% of T-LGL leukemias in IL-15 transgenic mice expressed NKp46, as did a majority of human T-LGL leukemias. The minute NKp46+ NKT population, but not the NKp46– NKT population, was selectively expanded by overexpression of endogenous IL-15. Importantly, IL-15 transgenic NKp46– NKT cells did not become NKp46+ in vivo, suggesting that NKp46+ T-LGL leukemia cells were the malignant counterpart of the minute WT NKp46+ NKT population. Mechanistically, NKp46+ NKT cells possessed higher responsiveness to IL-15 in vitro and in vivo compared with that of their NKp46– NKT counterparts. Furthermore, interruption of IL-15 signaling using a neutralizing antibody could prevent LGL leukemia in IL-15 transgenic mice. Collectively, our data demonstrate that NKp46 identifies a functionally distinct NKT subset in mice and humans that appears to be directly susceptible to leukemic transformation when IL-15 is overexpressed. Thus, IL-15 signaling and NKp46 may be useful targets in the treatment of patients with T-LGL or NKT leukemia. PMID:21364281

  20. Studies by radioiodination of normal adult, fetal and leukemic cell membranes

    Energy Technology Data Exchange (ETDEWEB)

    Kannourakis, G; Cauchi, M N [Department of Pathology and Immunology, Monash Medical School, Melbourne, Australia

    1978-01-01

    A comparison was made between cord blood lymphocytes, normal adult lymphocytes and leukemic cells after membrane iodination with lactoperoxidase. A double-labeling technique using lactoperoxidase iodination with /sup 125/I and /sup 131/I followed by analysis on polyacrylamide gel electrophoresis revealed a number of membrane differences between leukemic, normal and fetal cells. There was a reduction in the 70,000 molecular weight component in cord blood cells compared to adult lymphocytes, and an increase in membrane peptides with molecular weights of 35,000, 20,000, 9,000 and 4,000. Although smaller molecular weight peptides were also present in chronic lymphatic leukemia as well as acute myeloid leukemia, these were shown to be distinct from fetal type membrane components.

  1. Synergistic apoptosis induction in leukemic cells by the phosphatase inhibitor salubrinal and proteasome inhibitors.

    Directory of Open Access Journals (Sweden)

    Hannes C A Drexler

    Full Text Available Cells adapt to endoplasmic reticulum (ER-stress by arresting global protein synthesis while simultaneously activating specific transcription factors and their downstream targets. These processes are mediated in part by the phosphorylation-dependent inactivation of the translation initiation factor eIF2alpha. Following restoration of homeostasis protein synthesis is resumed when the serine/threonine-protein phosphatase PP1 dephosphorylates and reactivates eIF2alpha. Proteasome inhibitors, used to treat multiple myeloma patients evoke ER-stress and apoptosis by blocking the ER-associated degradation of misfolded proteins (ERAD, however, the role of eIF2alpha phosphorylation in leukemic cells under conditions of proteasome inhibitor-mediated ER stress is currently unclear.Bcr-Abl-positive and negative leukemic cell lines were used to investigate the functional implications of PP1-related phosphatase activities on eIF2alpha phosphorylation in proteasome inhibitor-mediated ER stress and apoptosis. Rather unexpectedly, salubrinal, a recently identified PP1 inhibitor capable to protect against ER stress in various model systems, strongly synergized with proteasome inhibitors to augment apoptotic death of different leukemic cell lines. Salubrinal treatment did not affect the phosphorlyation status of eIF2alpha. Furthermore, the proapoptotic effect of salubrinal occurred independently from the chemical nature of the proteasome inhibitor, was recapitulated by a second unrelated phosphatase inhibitor and was unaffected by overexpression of a dominant negative eIF2alpha S51A variant that can not be phosphorylated. Salubrinal further aggravated ER-stress and proteotoxicity inflicted by the proteasome inhibitors on the leukemic cells since characteristic ER stress responses, such as ATF4 and CHOP synthesis, XBP1 splicing, activation of MAP kinases and eventually apoptosis were efficiently abrogated by the translational inhibitor cycloheximide.Although PP1

  2. Musashi2 modulates K562 leukemic cell proliferation and apoptosis involving the MAPK pathway

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Huijuan; Tan, Shi; Wang, Juan; Chen, Shana; Quan, Jing; Xian, Jingrong; Zhang, Shuai shuai; He, Jingang; Zhang, Ling, E-mail: lingzhang@cqmu.edu.cn

    2014-01-01

    The RNA-binding protein Musashi2 (Msi2) has been identified as a master regulator within a variety of stem cell populations via the regulation of translational gene expression. A recent study has suggested that Msi2 is strongly expressed in leukemic cells of acute myeloid leukemia patients, and elevated Msi2 is associated with poor prognosis. However, the potential role of Msi2 in leukemogenesis is still not well understood. Here, we investigated the effect of Msi2 knockdown on the biological properties of leukemic cells. High expression of Msi2 was found in K562 and KG-1a leukemic cell lines, and low expression was observed in the U937 cell line. We transduced K562 cells with two independent adenoviral shRNA vectors targeting Msi2 and confirmed knockdown of Msi2 at the mRNA and protein levels. Msi2 silencing inhibited cell growth and caused cell cycle arrest by increasing the expression of p21 and decreasing the expression of cyclin D1 and cdk2. In addition, knockdown of Msi2 promoted cellular apoptosis via the upregulation of Bax and downregulation of Bcl-2 expression. Furthermore, Msi2 knockdown resulted in the inactivation of the ERK/MAPK and p38/MAPK pathways, but no remarkable change in p-AKT was observed. These data provide evidence that Msi2 plays an important role in leukemogenesis involving the MAPK signaling pathway, which indicates that Msi2 may be a novel target for leukemia treatment. - Highlights: • Knockdown of Msi2 inhibited K562 cell growth and arrested cell cycle progression. • Knockdown of Msi2 induced K562 cell apoptosis via the regulation of Bax and Bcl-2. • The MAPK pathway was involved in the process of Msi2-mediated leukemogenesis. • Our data indicate that Msi2 is a potential new target for leukemia treatment.

  3. Uncontrolled hypertension secondary to leukemic cell infiltration of kidneys in a hemodialysis patient

    Directory of Open Access Journals (Sweden)

    Kultigin Turkmen

    2010-06-01

    Full Text Available Kultigin Turkmen1, Lutfullah Altintepe2, Ibrahim Guney2, Ismet Aydogdu3, Osman Koc4, Mehmet Ali Erkut5, Halil Zeki Tonbul11Department of Nephrology, Meram School of Medicine, Selcuk University, 2Meram Training and Research Hospital, Selcuk University, 3Department of Hematology, Meram School of Medicine, Selcuk University, 4Department of Radiology, Meram School of Medicine, Selcuk University, 5Department of Hematology, Meram Training and Research Hospital, Selcuk UniversityAbstract: Leukemic infiltration of the kidney is usually silent, and the admission of the patients with renal dysfunction or acute kidney injury is uncommon. We present a 34-year old hemodialysis patient with new onset of uncontrolled hypertension, erythropoietin-resistant anemia, thrombocytopenia, and Bell’s palsy. On admission, his blood pressure (BP was 210/110 mmHg and he had petechiae and purpura at upper and lower extremities. Renal ultrasonography (USG showed bilaterally enlarged kidneys without hydronephrosis, unlike his previous USG, which determined bilaterally atrophic kidneys. Acute lymphoblastic leukemia, hypertensive crisis due to bilateral leukemic cell infiltration of kidneys, tumor lysis syndrome, and leukemic involvement of the facial nerve were diagnosed. Despite intense antihypertensive management, his BP was not controlled. After prednisolone, daunorubicine, and vincristine therapy, the size of kidneys diminished and his BP dropped under normal range. In conclusion, pathological findings such as uncontrolled hypertension, flank pain, skin rashes, and abnormal blood count should be considered carefully, even in patients with end-stage renal disease receiving renal replacement therapy.Keywords: leukemic cell infiltration, uncontrolled hypertension, hemodialysis

  4. Identification of residual leukemic cells by flow cytometry in childhood B-cell precursor acute lymphoblastic leukemia: verification of leukemic state by flow-sorting and molecular/cytogenetic methods

    DEFF Research Database (Denmark)

    Obro, Nina F; Ryder, Lars P; Madsen, Hans O

    2012-01-01

    Reduction in minimal residual disease, measured by real-time quantitative PCR or flow cytometry, predicts prognosis in childhood B-cell precursor acute lymphoblastic leukemia. We explored whether cells reported as minimal residual disease by flow cytometry represent the malignant clone harboring...... clone-specific genomic markers (53 follow-up bone marrow samples from 28 children with B-cell precursor acute lymphoblastic leukemia). Cell populations (presumed leukemic and non-leukemic) were flow-sorted during standard flow cytometry-based minimal residual disease monitoring and explored by PCR and....../or fluorescence in situ hybridization. We found good concordance between flow cytometry and genomic analyses in the individual flow-sorted leukemic (93% true positive) and normal (93% true negative) cell populations. Four cases with discrepant results had plausible explanations (e.g. partly informative...

  5. Anti-leukemic activity of bortezomib and carfilzomib on B-cell precursor ALL cell lines.

    Directory of Open Access Journals (Sweden)

    Kazuya Takahashi

    Full Text Available Prognosis of childhood acute lymphoblastic leukemia (ALL has been dramatically improved. However, prognosis of the cases refractory to primary therapy is still poor. Recent phase 2 study on the efficacy of combination chemotherapy with bortezomib (BTZ, a proteasome inhibitor, for refractory childhood ALL demonstrated favorable clinical outcomes. However, septic death was observed in over 10% of patients, indicating the necessity of biomarkers that could predict BTZ sensitivity. We investigated in vitro BTZ sensitivity in a large panel of ALL cell lines that acted as a model system for refractory ALL, and found that Philadelphia chromosome-positive (Ph+ ALL, IKZF1 deletion, and biallelic loss of CDKN2A were associated with favorable response. Even in Ph-negative ALL cell lines, IKZF1 deletion and bilallelic loss of CDKN2A were independently associated with higher BTZ sensitivity. BTZ showed only marginal cross-resistance to four representative chemotherapeutic agents (vincristine, dexamethasone, l-asparaginase, and daunorubicin in B-cell precursor-ALL cell lines. To improve the efficacy and safety of proteasome inhibitor combination chemotherapy, we also analyzed the anti-leukemic activity of carfilzomib (CFZ, a second-generation proteasome inhibitor, as a substitute for BTZ. CFZ showed significantly higher activity than BTZ in the majority of ALL cell lines except for the P-glycoprotein-positive t(17;19 ALL cell lines, and IKZF1 deletion was also associated with a favorable response to CFZ treatment. P-glycoprotein inhibitors effectively restored the sensitivity to CFZ, but not BTZ, in P-glycoprotein-positive t(17;19 ALL cell lines. P-glycoprotein overexpressing ALL cell line showed a CFZ-specific resistance, while knockout of P-glycoprotein by genome editing with a CRISPR/Cas9 system sensitized P-glycoprotein-positive t(17;19 ALL cell line to CFZ. These observations suggested that IKZF1 deletion could be a useful biomarker to predict good

  6. Demonstration of interleukin-1 beta transcripts in acute myeloblastic leukemic cells by in situ hybridization.

    Science.gov (United States)

    Nakamura, M; Kanakura, Y; Furukawa, Y; Ernst, T J; Griffin, J D

    1990-07-01

    The cells from some patients with acute myeloblastic leukemia will secrete autostimulatory cytokines in tissue culture without the addition of stimulators such as phorbol 12-myristate 13-acetate. Production of interleukin-1 beta (IL-1 beta), for example, has been observed in up to 50% of cases. In order to investigate the nature of the cell secreting IL-1 beta in AML, we used an antisense RNA probe to detect specific IL-1 beta transcripts in individual leukemic cells by in situ hybridization. In fresh, uncultured cells, IL-1 beta transcripts were observed in 1-40% of undifferentiated leukemic blast cells in 17 of 19 cases. In situ hybridization was at least as sensitive as Northern blot analysis in detecting IL-1 beta transcripts. No correlation of IL-1 beta transcript expression with FAB classification was observed. Normal blood and bone marrow mononuclear cells did not contain cells expressing IL-1 beta transcripts. These results support the concept that the regulation of cytokine genes in AML cells is aberrant.

  7. Single cell analysis of normal and leukemic hematopoiesis.

    Science.gov (United States)

    Povinelli, Benjamin J; Rodriguez-Meira, Alba; Mead, Adam J

    2018-02-01

    The hematopoietic system is well established as a paradigm for the study of cellular hierarchies, their disruption in disease and therapeutic use in regenerative medicine. Traditional approaches to study hematopoiesis involve purification of cell populations based on a small number of surface markers. However, such population-based analysis obscures underlying heterogeneity contained within any phenotypically defined cell population. This heterogeneity can only be resolved through single cell analysis. Recent advances in single cell techniques allow analysis of the genome, transcriptome, epigenome and proteome in single cells at an unprecedented scale. The application of these new single cell methods to investigate the hematopoietic system has led to paradigm shifts in our understanding of cellular heterogeneity in hematopoiesis and how this is disrupted in disease. In this review, we summarize how single cell techniques have been applied to the analysis of hematopoietic stem/progenitor cells in normal and malignant hematopoiesis, with a particular focus on recent advances in single-cell genomics, including how these might be utilized for clinical application. Copyright © 2017. Published by Elsevier Ltd.

  8. Resistance of some leukemic blasts to lysis by lymphokine activated killer (LAK) cells

    Energy Technology Data Exchange (ETDEWEB)

    Panayotides, P; Sjoegren, A -M; Reizenstein, P; Porwit, A. Immunopathology Lab., Dept. of Pathology, Karolinska Hospital, Stockholm; Wasserman, J

    1988-01-01

    Peripheral blood mononuclear cells (PBMC) from healthy donors and AML patients in remission were stimulated with phytohemagglutinin (PHA) and recombinant interleukin-2 (IL-2). These stimulated cells (lymphokine activated killer (LAK) cells) showed increased DNA synthesis as measured by /sup 3/H-Thymidine uptake. A synergistic effect of PHA and IL-2 was found. LAK cells' ability to kill acute myeloid leukemia (AML) blasts was investigated by the /sup 51/Cr release assay. LAK cells showed a cytotoxicity (over 10% specific /sup 51/Cr release) against 9/12 leukemic blasts, even at effector/target (E/T) ratios as low as 5:1. However, on average only 22.2% (SD 11.8) and 36.5% (SD 12.5) /sup 51/Cr release were obtained in 4- and 18-hour cytotoxicity assays, respectively, at an E/T ratio of 20:1. Leukemic blasts in 3/12 AML cases and normal PBMC were entirely resistant to lysis, even at an E/T ratio of 80:1. Susceptibility to lysis was not correlated to peanut-agglutinin receptor expression. LAK cells were more cytotoxic towards the K-562 cell line (natural killer activity) than unstimulated PBMC.

  9. Expression of MIF and CD74 in leukemic cell lines: correlation to DR expression destiny.

    Science.gov (United States)

    Georgouli, Mirella; Papadimitriou, Lina; Glymenaki, Maria; Patsaki, Valia; Athanassakis, Irene

    2016-06-01

    Invariant chain (Ii) or CD74 is a non-polymorphic glycoprotein, which apart from its role as a chaperone dedicated to MHCII molecules, is known to be a high-affinity receptor for macrophage migration inhibitory factor (MIF). The present study aimed to define the roles of CD74 and MIF in the immune surveillance escape process. Towards this direction, the cell lines HL-60, Raji, K562 and primary pre-B leukemic cells were examined for expression and secretion of MIF. Flow cytometry analysis detected high levels of MIF and intracellular/membrane CD74 expression in all leukemic cells tested, while MIF secretion was shown to be inversely proportional to intracellular HLA-DR (DR) expression. In the MHCII-negative cells, IFN-γ increased MIF expression and induced its secretion in HL-60 and K562 cells, respectively. In K562 cells, CD74 (Iip33Iip35) was shown to co-precipitate with HLA-DOβ (DOβ), inhibiting thus MIF or DR binding. Induced expression of DOα in K562 (DOα-DOβ+) cells in different transfection combinations decreased MIF expression and secretion, while increasing surface DR expression. Thus, MIF could indeed be part of the antigen presentation process.

  10. Fundamental studies of leukemic cell labeling with /sup 111/In-oxine and their applications to cell kinetics in patients with acute leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Takagi, Yuhkoh; Matsuda, Shin; Uchida, Tatsumi; Kariyone, Shigeo [Fukushima Medical Coll. (Japan)

    1984-04-01

    Fundamental studies of leukemic cell labeling with /sup 111/In-oxine and their applications to leukemic cell kinetics in five patients with acute myeloblastic leukemia (AML) were examined. Labeling efficiency of leukemic cells was 80.3 +- 3.6% for more than 1 x 10/sup 8/ cells at room temperature for 20 minutes of incubation followed by two times washes. Cell viability determined by means of trypanblue exclusion test was 95.3 +- 2.6%. In vitro elution rate of /sup 111/In from the labeled cells during 12 hours was 10.0 +- 1.2%. The disappearance curves of labeled leukemic cells in AMLs followed a single exponential fashion, and the half time of disappearance (T 1/2) ranged from 9.6 to 31.8 hours. Total blood leukemic cell pool (TBLCP) calculated with the dilution principles of radioisotopes correlated significantly with the leukemic cell counts (LC) in the peripheral blood (Y = 0.32 + 1.94X, r = 0.99). In the studies of organ distribution which were observed and analyzed with gamma camera and computer, labeled leukemic cells passed through lungs within 15 minutes. Radioactivity in the spleen increased rapidly for 30 - 60 minutes, then reached a plateau. Hepatic radioactivity showed a temporary decrease during 10 - 60 minutes following the moderate accumulation in initial 10 minutes. In two cases, bone marrow was visualized 24 hours after the injection. Radioactivity of the leukemic cells isolated from the bone marrow at 22 hours after the injection in one case was one third of the radioactivity in leukemic cells obtained from the peripheral blood at the same time.

  11. Fundamental studies of leukemic cell labeling with 111 In-oxine and their applications to cell kinetics in patients with acute leukemia

    International Nuclear Information System (INIS)

    Takagi, Yuhkoh; Matsuda, Shin; Uchida, Tatsumi; Kariyone, Shigeo

    1984-01-01

    Fundamental studies of leukemic cell labeling with 111 In-oxine and their applications to leukemic cell kinetics in five patients with acute myeloblastic leukemia (AML) were examined. Labeling efficiency of leukemic cells was 80.3 +- 3.6% for more than 1 x 10 8 cells at room temperature for 20 minutes of incubation followed by two times washes. Cell viability determined by means of trypanblue exclusion test was 95.3 +- 2.6%. In vitro elution rate of 111 In from the labeled cells during 12 hours was 10.0 +- 1.2%. The disappearance curves of labeled leukemic cells in AMLs followed a single exponential fashion, and the half time of disappearance (T 1/2) ranged from 9.6 to 31.8 hours. Total blood leukemic cell pool (TBLCP) calculated with the dilution principles of radioisotopes correlated significantly with the leukemic cell counts (LC) in the peripheral blood (Y = 0.32 + 1.94X, r = 0.99). In the studies of organ distribution which were observed and analized with gamma camera and computer, labeled leukemic cells passed through lungs within 15 minutes. Radioactivity in the spleen increased rapidly for 30 - 60 minutes, then reached a plateau. Hepatic radioactivity showed a temporary decrease during 10 - 60 minutes following the moderate accumulation in initial 10 minutes. In two cases, bone marrow was visualized 24 hours after the injection. Radioactivity of the leukemic cells isolated from the bone marrow at 22 hours after the injection in one case was one third of the radioactivity in leukemic cells obtained from the peripheral blood at the same time. (author)

  12. Chronic Myelogenous Leukemia Cells Contribute to the Stromal Myofibroblasts in Leukemic NOD/SCID Mouse In Vivo

    Directory of Open Access Journals (Sweden)

    Ryosuke Shirasaki

    2012-01-01

    Full Text Available We recently reported that chronic myelogenous leukemia (CML cells converted into myofibroblasts to create a microenvironment for proliferation of CML cells in vitro. To analyze a biological contribution of CML-derived myofibroblasts in vivo, we observed the characters of leukemic nonobese diabetes/severe combined immunodeficiency (NOD/SCID mouse. Bone marrow nonadherent mononuclear cells as well as human CD45-positive cells obtained from CML patients were injected to the irradiated NOD/SCID mice. When the chimeric BCR-ABL transcript was demonstrated in blood, human CML cells were detected in NOD/SCID murine bone marrow. And CML-derived myofibroblasts composed with the bone marrow-stroma, which produced significant amounts of human vascular endothelial growth factor A. When the parental CML cells were cultured with myofibroblasts separated from CML cell-engrafted NOD/SCID murine bone marrow, CML cells proliferated significantly. These observations indicate that CML cells make an adequate microenvironment for their own proliferation in vivo.

  13. Dendritic cells (DCs) can be successfully generated from leukemic blasts in individual patients with AML or MDS: an evaluation of different methods.

    Science.gov (United States)

    Kremser, Andreas; Dressig, Julia; Grabrucker, Christine; Liepert, Anja; Kroell, Tanja; Scholl, Nina; Schmid, Christoph; Tischer, Johanna; Kufner, Stefanie; Salih, Helmut; Kolb, Hans Jochem; Schmetzer, Helga

    2010-01-01

    Myeloid-leukemic cells (AML, MDS, CML) can be differentiated to leukemia-derived dendritic cell [DC (DCleu)] potentially presenting the whole leukemic antigen repertoire without knowledge of distinct leukemia antigens and are regarded as promising candidates for a vaccination strategy. We studied the capability of 6 serum-free DC culture methods, chosen according to different mechanisms, to induce DC differentiation in 137 cases of AML and 52 cases of MDS. DC-stimulating substances were cytokines ("standard-medium", "MCM-Mimic", "cytokine-method"), bacterial lysates ("Picibanil"), double-stranded RNA ["Poly (I:C)"] or a cytokine bypass method ("Ca-ionophore"). The quality/quantity of DC generated was estimated by flow cytometry studying (co) expressions of "DC"antigens, costimulatory, maturation, and blast-antigens. Comparing these methods on average 15% to 32% DC, depending on methods used, could be obtained from blast-containing mononuclear cells (MNC) in AML/MDS cases with a DC viability of more than 60%. In all, 39% to 64% of these DC were mature; 31% to 52% of leukemic blasts could be converted to DCleu and DCleu-proportions in the suspension were 2% to 70% (13%). Average results of all culture methods tested were comparable, however not every given case of AML could be differentiated to DC with 1 selected method. However performing a pre-analysis with 3 DC-generating methods (MCM-Mimic, Picibanil, Ca-ionophore) we could generate DC in any given case. Functional analyses provided proof, that DC primed T cells to antileukemia-directed cytotoxic cells, although an anti-leukemic reaction was not achieved in every case. In summary our data show that a successful, quantitative DC/DCleu generation is possible with the best of 3 previously tested methods in any given case. Reasons for different functional behaviors of DC-primed T cells must be evaluated to design a practicable DC-based vaccination strategy.

  14. Leukemic optic neuropathy.

    Science.gov (United States)

    Brown, G C; Shields, J A; Augsburger, J J; Serota, F T; Koch, P

    1981-03-01

    The clinical course and ophthalmic manifestations of an eight year old child with acute undifferentiated leukemia and unilateral blindness secondary to leukemic optic nerve head infiltration are described. At autopsy the involved nerve head and peripapillary retina demonstrated massive leukemic cell infiltration and hemorrhagic necrosis. This manifestation of leukemia is quite uncommon and prognosis for life in such cases is poor with existing methods of therapy.

  15. Activation of the complement cascade enhances motility of leukemic cells by downregulating expression of HO-1.

    Science.gov (United States)

    Abdelbaset-Ismail, A; Borkowska-Rzeszotek, S; Kubis, E; Bujko, K; Brzeźniakiewicz-Janus, K; Bolkun, L; Kloczko, J; Moniuszko, M; Basak, G W; Wiktor-Jedrzejczak, W; Ratajczak, M Z

    2017-02-01

    As a crucial arm of innate immunity, the complement cascade (ComC) is involved both in mobilization of normal hematopoietic stem/progenitor cells (HSPCs) from bone marrow (BM) into peripheral blood and in their homing to BM. Despite the fact that ComC cleavage fragments alone do not chemoattract normal HSPCs, we found that leukemia cell lines as well as clonogenic blasts from chronic myeloid leukemia and acute myeloid leukemia patients respond robustly to C3 and C5 cleavage fragments by chemotaxis and increased adhesion. This finding was supported by the detection of C3a and C5a receptors in cells from human malignant hematopoietic cell lines and patient blasts at the mRNA (reverse transcriptase-polymerase chain reaction) and protein level (fluorescence-activated cell sorting), and by the demonstration that these receptors respond to stimulation by C3a and C5a by phosphorylation of p42/44 and p38 mitogen-activated protein kinases (MAPK), and protein kinase B (PKB/AKT). We also found that inducible heme oxygenase 1 (HO-1) is a negative regulator of ComC-mediated trafficking of leukemic cells, and that stimulation of leukemic cells by C3 or C5 cleavage fragments activates p38 MAPK, which downregulates HO-1 expression, rendering cells more mobile. We conclude that activation of the ComC in leukemia/lymphoma patients (for example, as a result of accompanying infections) enhances the motility of malignant cells and contributes to their spread in a p38 MAPK-HO-1-dependent manner. Therefore, inhibition of p38 MAPK or upregulation of HO-1 by small-molecule modulators would have a beneficial effect on ameliorating cell migration-mediated expansion of leukemia/lymphoma cells when the ComC becomes activated.

  16. Activation of the complement cascade enhances motility of leukemic cells by downregulating expression of HO-1

    Science.gov (United States)

    Abdelbaset-Ismail, A; Borkowska-Rzeszotek, S; Kubis, E; Bujko, K; Brzeźniakiewicz-Janus, K; Bolkun, L; Kloczko, J; Moniuszko, M; Basak, G W; Wiktor-Jedrzejczak, W; Ratajczak, M Z

    2017-01-01

    As a crucial arm of innate immunity, the complement cascade (ComC) is involved both in mobilization of normal hematopoietic stem/progenitor cells (HSPCs) from bone marrow (BM) into peripheral blood and in their homing to BM. Despite the fact that ComC cleavage fragments alone do not chemoattract normal HSPCs, we found that leukemia cell lines as well as clonogenic blasts from chronic myeloid leukemia and acute myeloid leukemia patients respond robustly to C3 and C5 cleavage fragments by chemotaxis and increased adhesion. This finding was supported by the detection of C3a and C5a receptors in cells from human malignant hematopoietic cell lines and patient blasts at the mRNA (reverse transcriptase-polymerase chain reaction) and protein level (fluorescence-activated cell sorting), and by the demonstration that these receptors respond to stimulation by C3a and C5a by phosphorylation of p42/44 and p38 mitogen-activated protein kinases (MAPK), and protein kinase B (PKB/AKT). We also found that inducible heme oxygenase 1 (HO-1) is a negative regulator of ComC-mediated trafficking of leukemic cells, and that stimulation of leukemic cells by C3 or C5 cleavage fragments activates p38 MAPK, which downregulates HO-1 expression, rendering cells more mobile. We conclude that activation of the ComC in leukemia/lymphoma patients (for example, as a result of accompanying infections) enhances the motility of malignant cells and contributes to their spread in a p38 MAPK–HO-1-dependent manner. Therefore, inhibition of p38 MAPK or upregulation of HO-1 by small-molecule modulators would have a beneficial effect on ameliorating cell migration-mediated expansion of leukemia/lymphoma cells when the ComC becomes activated. PMID:27451975

  17. Effects of vinegar–egg on growth inhibition, differentiation human leukemic U937 cells and its immunomodulatory activity

    Directory of Open Access Journals (Sweden)

    Shiu-Yu Wang

    2018-04-01

    Full Text Available Vinegar and eggs have rich nutrients. In this study, the mixed form of both derived products, vinegar–egg solution and its products (vinegar–egg concentrate and vinegar–egg condensate were chosen for an assessment of their biological activity. To further our understanding regarding the anticancer and immunomodulatory effects of vinegar–egg, we investigated its effects on the proliferation and differentiation of U937 cells. Vinegar–egg was treated using spray drying, freeze drying and vacuum concentration and used to stimulate human mononuclear cells. The conditioned media obtained from these cultures by filtration were used to treat U937 cells. Three conditioned media inhibited U937 cell growth by 22.1–67.25% more effectively than PHA-treated control (22.53%. CD11b and CD14 expression on the treated U937 cells were 29.1–45.4% and 31.6–47.2%, respectively. High levels of cytokines IL-1β, IFN-γ and TNF-α were detected in the three conditioned media. Vinegar–egg stimulates human mononuclear cells to secrete cytokines, which inhibit the growth of U937 cells and induce their differentiation. Keywords: Cytokines, Differentiation, Immunomodulatory activity, Leukemic U937 cells, Vinegar–egg

  18. Resveratrol protects leukemic cells against cytotoxicity induced by proteasome inhibitors via induction of FOXO1 and p27Kip1

    International Nuclear Information System (INIS)

    Niu, Xiao-Fang; Liu, Bao-Qin; Du, Zhen-Xian; Gao, Yan-Yan; Li, Chao; Li, Ning; Guan, Yifu; Wang, Hua-Qin

    2011-01-01

    It was reported recently that resveratrol could sensitize a number of cancer cells to the antitumoral effects of some conventional chemotherapy drugs. The current study was designed to investigate whether resveratrol could sensitize leukemic cells to proteasome inhibitors. Leukemic cells were treated with MG132 alone or in combination with resveratrol. Cell viability was investigated using MTT assay, and induction of apoptosis and cell cycle distribution was measured using flow cytometry. Western blot and real-time RT-PCR were used to investigate the expression of FOXO1 and p27 Kip1 . CHIP was performed to investigate the binding of FOXO1 to the p27 Kip1 promoter. Resveratrol strongly reduced cytotoxic activities of proteasome inhibitors against leukemic cells. MG132 in combination with resveratrol caused cell cycle blockade at G1/S transition via p27 Kip1 accumulation. Knockdown of p27 Kip1 using siRNA dramatically attenuated the protective effects of resveratrol on cytotoxic actions of proteasome inhibitors against leukemic cells. Resveratrol induced FOXO1 expression at the transcriptional level, while MG132 increased nuclear distribution of FOXO1. MG132 in combination with resveratrol caused synergistic induction of p27 Kip1 through increased recruitment of FOXO1 on the p27 Kip1 promoter. Resveratrol may have the potential to negate the cytotoxic effects of proteasome inhibitors via regulation of FOXO1 transcriptional activity and accumulation of p27 Kip1

  19. [Expression of c-MPL in leukemic stem cells from acute myeloid leukemia patients].

    Science.gov (United States)

    Yu, Pei; Qiu, Shao-Wei; Rao, Qing; Lin, Dong; Xing, Hai-Yan; Tang, Ke-Jing; Tian, Zheng; Wang, Min; Wang, Jian-Xiang

    2012-10-01

    This study was aimed to investigate the expression of c-MPL in acute myeloid leukemia (AML) and the correlation of the c-MPL expression with CD34 and CD38, so as to define the expression of c-MPL in leukemic stem cells. The expression levels of CD34, CD38 and c-MPL were detected by flow cytometry in bone marrow cells from 29 newly diagnosed AML patients. The relationship of c-MPL positive cell ratio with clinical parameters and correlation of c-MPL with CD34 and CD38 expression in AML patients were analyzed. The results showed that expression level of c-MPL in AML patients was significantly higher than that of normal controls (P MPL did not correlate with age, sex, white blood cell count, AML1-ETO fusion gene and remission after chemotherapy, but the expression of c-MPL in M2 and M5 patients was higher than that of normal control (P MPL in CD34 positive AML patients was obviously higher than that in CD34 negative AML patients (P MPL was significantly higher expressed in CD34(+) cells than that in CD34(-) cells (P MPL expression was not significantly different between CD34(+)CD38(-) and CD34(+)CD38(-) cell groups. Positive correlation between c-MPL and CD34 expression was observed (r = 0.380, P = 0.042). It is concluded that expression of c-MPL is higher in AML patients, and positively correlates with the expression level of CD34. The c-MPL expresses in leukemic stem cells.

  20. Leukemic blast cell colony formation in semisolid culture with erythropoietin: a case report of acute poorly differentiated erythroid leukemia.

    Science.gov (United States)

    Tomonaga, M; Jinnai, I; Tagawa, M; Amenomori, T; Nishino, K; Yao, E; Nonaka, H; Kuriyama, K; Yoshida, Y; Matsuo, T

    1987-02-01

    The bone marrow of a patient with acute undifferentiated leukemia developed unique colonies after a 14-day culture in erythropoietin (EPO)-containing methylcellulose. The colonies consisted of 20 to 200 nonhemoglobinized large blast cells. Cytogenetic analysis of single colonies revealed hypotetraploid karyotypes with several marker chromosomes that were identical to those found in directly sampled bone marrow. The concurrently formed erythroid bursts showed only normal karyotypes. No leukemic colony formation was observed in other culture systems with either colony-stimulating activity (CSA) or phytohemagglutinin-stimulated leukocyte-conditioned medium (PHA-LCM). The leukemic colonies exhibited a complete EPO-dose dependency similar to that of the patient's normal BFU-E. Although cytochemical and immunologic marker studies of the bone marrow cells failed to clarify the cell lineage of the leukemic cells with extraordinarily large cell size, ultrastructural study revealed erythroid differentiation such as siderosome formation in the cytoplasm and ferritin particles in the rhophecytosis invaginations. These findings indicate that the patient had poorly differentiated erythroid leukemia and that some of the clonogenic cells might respond to EPO in vitro. Corresponding to this biological feature, the leukemic cells were markedly decreased in number in response to repeated RBC transfusions, and partial remission was obtained. These observations suggest that erythroid leukemia distinct from erythroleukemia (M6) with a myeloblastic component, can develop as a minor entity of human acute leukemia.

  1. GPR84 sustains aberrant β-catenin signaling in leukemic stem cells for maintenance of MLL leukemogenesis.

    Science.gov (United States)

    Dietrich, Philipp A; Yang, Chen; Leung, Halina H L; Lynch, Jennifer R; Gonzales, Estrella; Liu, Bing; Haber, Michelle; Norris, Murray D; Wang, Jianlong; Wang, Jenny Yingzi

    2014-11-20

    β-catenin is required for establishment of leukemic stem cells (LSCs) in acute myeloid leukemia (AML). Targeted inhibition of β-catenin signaling has been hampered by the lack of pathway components amenable to pharmacologic manipulation. Here we identified a novel β-catenin regulator, GPR84, a member of the G protein-coupled receptor family that represents a highly tractable class of drug targets. High GPR84 expression levels were confirmed in human and mouse AML LSCs compared with hematopoietic stem cells (HSCs). Suppression of GPR84 significantly inhibited cell growth by inducing G1-phase cell-cycle arrest in pre-LSCs, reduced LSC frequency, and impaired reconstitution of stem cell-derived mixed-lineage leukemia (MLL) AML, which represents an aggressive and drug-resistant subtype of AML. The GPR84-deficient phenotype in established AML could be rescued by expression of constitutively active β-catenin. Furthermore, GPR84 conferred a growth advantage to Hoxa9/Meis1a-transduced stem cells. Microarray analysis demonstrated that GPR84 significantly upregulated a small set of MLL-fusion targets and β-catenin coeffectors, and downregulated a hematopoietic cell-cycle inhibitor. Altogether, our data reveal a previously unrecognized role of GPR84 in maintaining fully developed AML by sustaining aberrant β-catenin signaling in LSCs, and suggest that targeting the oncogenic GPR84/β-catenin signaling axis may represent a novel therapeutic strategy for AML. © 2014 by The American Society of Hematology.

  2. Generation of TCR-Expressing Innate Lymphoid-like Helper Cells that Induce Cytotoxic T Cell-Mediated Anti-leukemic Cell Response.

    Science.gov (United States)

    Ueda, Norihiro; Uemura, Yasushi; Zhang, Rong; Kitayama, Shuichi; Iriguchi, Shoichi; Kawai, Yohei; Yasui, Yutaka; Tatsumi, Minako; Ueda, Tatsuki; Liu, Tian-Yi; Mizoro, Yasutaka; Okada, Chihiro; Watanabe, Akira; Nakanishi, Mahito; Senju, Satoru; Nishimura, Yasuharu; Kuzushima, Kiyotaka; Kiyoi, Hitoshi; Naoe, Tomoki; Kaneko, Shin

    2018-06-05

    CD4 + T helper (Th) cell activation is essential for inducing cytotoxic T lymphocyte (CTL) responses against malignancy. We reprogrammed a Th clone specific for chronic myelogenous leukemia (CML)-derived b3a2 peptide to pluripotency and re-differentiated the cells into original TCR-expressing T-lineage cells (iPS-T cells) with gene expression patterns resembling those of group 1 innate lymphoid cells. CD4 gene transduction into iPS-T cells enhanced b3a2 peptide-specific responses via b3a2 peptide-specific TCR. iPS-T cells upregulated CD40 ligand (CD40L) expression in response to interleukin-2 and interleukin-15. In the presence of Wilms tumor 1 (WT1) peptide, antigen-specific dendritic cells (DCs) conditioned by CD4-modified CD40L high iPS-T cells stimulated WT1-specific CTL priming, which eliminated WT1 peptide-expressing CML cells in vitro and in vivo. Thus, CD4 modification of CD40L high iPS-T cells generates innate lymphoid helper-like cells inducing bcr-abl-specific TCR signaling that mediates effectiveanti-leukemic CTL responses via DC maturation, showing potential for adjuvant immunotherapy against leukemia. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  3. Cyclopentenyl cytosine induces apoptosis and increases cytarabine-induced apoptosis in a T-lymphoblastic leukemic cell-line

    NARCIS (Netherlands)

    Verschuur, A. C.; Brinkman, J.; van Gennip, A. H.; Leen, R.; Vet, R. J.; Evers, L. M.; Voûte, P. A.; van Kuilenburg, A. B.

    2001-01-01

    Cyclopentenyl cytosine (CPEC) is a nucleoside-analogue that decreases the concentrations of cytidine triphosphate (CTP) and deoxycytidine triphosphate (dCTP) in leukemic cells by inhibiting the enzyme CTP synthetase, resulting in a decreased synthesis of RNA and DNA. Low concentrations of dCTP

  4. Towards The Generation of Functionalized Magnetic Nanowires to Target Leukemic Cells

    KAUST Repository

    Alsharif, Nouf

    2016-04-01

    In recent years, magnetic nanowires (NWs) have been widely used for their therapeutic potential in biomedical applications. The use of iron (Fe) NWs combines two important properties, biocompatibility and remote manipulation by magnetic fields. In addition the NWs can be coated and functionalized to target cells of interest and, upon exposure to an alternating magnetic field, have been shown to induce cell death on several types of adherent cells, including several cancer cell types. For suspension cells, however, using these NWs has been much less effective primarily due to the free-floating nature of the cells minimizing the interaction between them and the NWs. Leukemic cells express higher levels of the cell surface marker CD44 (Braumüller, Gansauge, Ramadani, & Gansauge, 2000), compared to normal blood cells. The goal of this study was to functionalize Fe NWs with a specific monoclonal antibody towards CD44 in order to target leukemic cells (HL-60 cells). This approach is expected to increase the probability of a specific binding to occur between HL-60 cells and Fe NWs. Fe NWs were fabricated with an average diameter of 30-40 nm and a length around 3-4 μm. Then, they were coated with both 3-Aminopropyl-triethoxysilane and bovine serum albumin (BSA) in order to conjugate them with an anti-CD44 antibody (i.e. anti-CD44-iron NWs). The antibody interacts with the amine group in the BSA via the 1-Ethyl-3-3-dimethylaminopropyl-carbodiimide and N-Hydroxysuccinimide coupling. The NWs functionalization was confirmed using a number of approaches including: infrared spectroscopy, Nanodrop to measure the concentration of CD44 antibody, as well as fluorescent-labeled secondary antibody staining to detect the primary CD44 antibody. To confirm that the anti-CD44-iron NWs and bare Fe NWs, in the absence of a magnetic field, were not toxic to HL-60 cells, cytotoxicity assays using XTT (2,3-Bis-2-Methoxy-4-Nitro-5-Sulfophenyl-2H-Tetrazolium-5-Carboxanilide) were performed and

  5. Aberrant methylation of the M-type phospholipase A2 receptor gene in leukemic cells

    International Nuclear Information System (INIS)

    Menschikowski, Mario; Platzbecker, Uwe; Hagelgans, Albert; Vogel, Margot; Thiede, Christian; Schönefeldt, Claudia; Lehnert, Renate; Eisenhofer, Graeme; Siegert, Gabriele

    2012-01-01

    The M-type phospholipase A2 receptor (PLA2R1) plays a crucial role in several signaling pathways and may act as tumor-suppressor. This study examined the expression and methylation of the PLA2R1 gene in Jurkat and U937 leukemic cell lines and its methylation in patients with myelodysplastic syndrome (MDS) or acute leukemia. Sites of methylation of the PLA2R1 locus were identified by sequencing bisulfite-modified DNA fragments. Methylation specific-high resolution melting (MS-HRM) analysis was then carried out to quantify PLA2R1 methylation at 5-CpG sites identified with differences in methylation between healthy control subjects and leukemic patients using sequencing of bisulfite-modified genomic DNA. Expression of PLA2R1 was found to be completely down-regulated in Jurkat and U937 cells, accompanied by complete methylation of PLA2R1 promoter and down-stream regions; PLA2R1 was re-expressed after exposure of cells to 5-aza-2´-deoxycytidine. MS-HRM analysis of the PLA2R1 locus in patients with different types of leukemia indicated an average methylation of 28.9% ± 17.8%, compared to less than 9% in control subjects. In MDS patients the extent of PLA2R1 methylation significantly increased with disease risk. Furthermore, measurements of PLA2R1 methylation appeared useful for predicting responsiveness to the methyltransferase inhibitor, azacitidine, as a pre-emptive treatment to avoid hematological relapse in patients with high-risk MDS or acute myeloid leukemia. The study shows for the first time that PLA2R1 gene sequences are a target of hypermethylation in leukemia, which may have pathophysiological relevance for disease evolution in MDS and leukemogenesis

  6. Interaction between the immune system and acute myeloid leukemia: A model incorporating promotion of regulatory T cell expansion by leukemic cells.

    Science.gov (United States)

    Nishiyama, Yoshiaki; Saikawa, Yutaka; Nishiyama, Nobuaki

    2018-03-01

    Population dynamics of regulatory T cells (Treg) are crucial for the underlying interplay between leukemic and immune cells in progression of acute myeloid leukemia (AML). The goal of this work is to elucidate the dynamics of a model that includes Treg, which can be qualitatively assessed by accumulating clinical findings on the impact of activated immune cell infusion after selective Treg depletion. We constructed an ordinary differential equation model to describe the dynamics of three components in AML: leukemic blast cells, mature regulatory T cells (Treg), and mature effective T cells (Teff), including cytotoxic T lymphocytes. The model includes promotion of Treg expansion by leukemic blast cells, leukemic stem cell and progenitor cell targeting by Teff, and Treg-mediated Teff suppression, and exhibits two coexisting, stable steady states, corresponding to high leukemic cell load at diagnosis or relapse, and to long-term complete remission. Our model is capable of explaining the clinical findings that the survival of patients with AML after allogeneic stem cell transplantation is influenced by the duration of complete remission, and that cut-off minimal residual disease thresholds associated with a 100% relapse rate are identified in AML. Copyright © 2018 Elsevier B.V. All rights reserved.

  7. Profound radiosensitivity in leukemic T-cell lines and T-cell-type acute lymphoblastic leukemia demonstrated by sodium [51Cr]chromate labeling

    International Nuclear Information System (INIS)

    Nakazawa, S.; Minowada, J.; Tsubota, T.; Sinks, L.F.

    1978-01-01

    Radiation sensitivity was determined by measuring spontaneous release from 51 Cr-labeled cells in various lymphoid cell populations. Among six leukemia T-cell lines originating from acute lymphoblastic leukemia, four such lines were found to be highly radiosensitive. In contrast, two of the leukemic T-cell lines and four normal control B-cell lines were not radiosensitive. Thymocytes from six patients and leukemia T-cell blasts from three patients with T-cell leukemia were likewise found to be highly radiosensitive, whereas leukemic blasts from six patients with null-cell (non-T, non-B-cell) acute lymphoblastic leukemia were not radiosensitive. Normal peripheral blood lymphocytes and mitogen-induced normal lymphoblasts were found not to be radiosensitive. The results indicate that measurement of the radiation sensitivity of acute leukemic blasts may have a therapeutic significance in coping with the heterogeneous nature of individual leukemia cases

  8. Hematopoietic stem cells can be separated from leukemic cells in a subgroup of adult acute lymphoblastic leukemia patients.

    Science.gov (United States)

    Wang, Wenwen; Foerner, Elena; Buss, Eike; Jauch, Anna; Eckstein, Volker; Wuchter, Patrick; Ho, Anthony D; Lutz, Christoph

    2017-06-01

    In B-cell acute lymphoblastic leukemia (B-ALL) separation of normal hematopoietic stem cells (HSC) has so far been limited to a subgroup of patients. As aldehyde dehydrogenase (ALDH)-activity is enriched in various stem cells we investigated its value for HSC isolation in adult B-ALL. Based on ALDH-activity patients could be stratified in ALDH-numerous (≥1.9% ALDH +  cells) and ALDH-rare (cells) cases. In ALDH-rare B-ALL clonal-marker negative HSC could be separated by the CD34 + CD38 - ALDH +  phenotype, whereas this separation was not possible in ALDH-numerous B-ALL. Functional analysis confirmed the HSC-potential of isolated cells, which were uniformly CD19-negative. However, addition of ALDH-activity further improved HSC-purity. In summary, we provide a method to separate functionally normal HSC from leukemic cells in a subgroup of B-ALL patients that can be identified prospectively. This protocol thereby facilitates comparative analyses of matched HSC and leukemic cells in order to improve our understanding of leukemia evolution.

  9. Apoptosis induction in MV4-11 and K562 human leukemic cells by Pereskia sacharosa (Cactaceae) leaf crude extract.

    Science.gov (United States)

    Asmaa, Mat Jusoh Siti; Al-Jamal, Hamid Ali Nagi; Ang, Cheng Yong; Asan, Jamaruddin Mat; Seeni, Azman; Johan, Muhammad Farid

    2014-01-01

    Pereskia sacharosa is a genus of cacti widely used in folk medicine for cancer-related treatment. Anti-proliferative effects have been studied in recent years against colon, breast, cervical and lung cancer cell lines, with promising results. We here extended study of anti-proliferative effects to a blood malignancy, leukemia. Two leukemic cell lines, MV4-11 (acute myeloid leukemia) and K562 (chronic myeloid leukemia), were studied. IC50 concentrations were determined and apoptosis and cell cycle regulation were studied by flow cytometric analysis. The expression of apoptosis and cell-cycle related regulatory proteins was assessed by Western blotting. P sacharosa inhibited growth of MV4-11 and K562 cells in a dose-dependent manner. The mode of cell death was via induction of intrinsic apoptotic pathways and cell cycle arrest. There was profound up-regulation of cytochrome c, caspases, p21 and p53 expression and repression of Akt and Bcl-2 expression in treated cells. These results suggest that P sacharosa induces leukemic cell death via apoptosis induction and changes in cell cycle checkpoint, thus deserves further study for anti-leukemic potential.

  10. Chemotherapy impedes in vitro microcirculation and promotes migration of leukemic cells with impact on metastasis

    International Nuclear Information System (INIS)

    Prathivadhi-Bhayankaram, Sruti V.; Ning, Jianhao; Mimlitz, Michael; Taylor, Carolyn; Gross, Erin; Nichols, Michael; Guck, Jochen; Ekpenyong, Andrew E.

    2016-01-01

    Although most cancer drugs target the proliferation of cancer cells, it is metastasis, the complex process by which cancer cells spread from the primary tumor to other tissues and organs of the body where they form new tumors, that leads to over 90% of all cancer deaths. Thus, there is an urgent need for anti-metastasis therapy. Surprisingly, emerging evidence suggests that certain anti-cancer drugs such as paclitaxel and doxorubicin can actually promote metastasis, but the mechanism(s) behind their pro-metastatic effects are still unclear. Here, we use a microfluidic microcirculation mimetic (MMM) platform which mimics the capillary constrictions of the pulmonary and peripheral microcirculation, to determine if in-vivo-like mechanical stimuli can evoke different responses from cells subjected to various cancer drugs. In particular, we show that leukemic cancer cells treated with doxorubicin and daunorubicin, commonly used anti-cancer drugs, have over 100% longer transit times through the device, compared to untreated leukemic cells. Such delays in the microcirculation are known to promote extravasation of cells, a key step in the metastatic cascade. Furthermore, we report a significant (p < 0.01) increase in the chemotactic migration of the doxorubicin treated leukemic cells. Both enhanced retention in the microcirculation and enhanced migration following chemotherapy, are pro-metastatic effects which can serve as new targets for anti-metastatic drugs. - Highlights: • Doxorubicin enhances migration of leukemic cancer cells before cell death. • Doxorubicin and Daunorubicin stiffen and delay cells in mimicked microcirculation. • Some cancer drugs cause changes in cell mechanics that lead to pro-metastatic effects. • Cell mechanics becomes a new target for anti-metastatic drugs.

  11. ROLE OF LEUKEMIC STEM CELLS IN THE CHRONIC MYELOID LEUKEMIA PATHOGENESIS

    Directory of Open Access Journals (Sweden)

    Sviezhentseva IO

    2016-09-01

    Full Text Available The presence of leukemic stem cells (LSC in the bone marrow of patients with chronic myeloid leukemia (CML is the cause of relapses as a result of the treatment with chemotherapeutic agents and target therapy drugs. This is due to the ability of LSC to attach itself to the microenvironment cells and to remain at rest for a long time. Vascular and osteoblasts niche play a very important role in this process. However, for being in G0 phase LSC have direct contact with the cellular elements of bone marrow microenvironment. So LSK contact with mesenchymal cells of bone marrow using the appendixes, connecting components invaginations and lint. The cadherins and integrins are important in the interaction of osteoblasts niche. They are able to activate intracellular signaling cascades that provide resting state of LSK. In addition, a bone marrow niche provides changes of LSC oxidative metabolism, which also plays an important role for cell entry into the G0 phase. Further, LSC also have certain physiological properties, which play an important role in the drug resistance formation, particularly drugs with targeted actions - tyrosine kinase inhibitors. LSK characterized by a high level of BCR-ABL expression and their population can have a lot of point mutations in the bcr-abl gene in the same patient. This leads to the fact that the taken medicines dose does not act against LSK, reducing the number of a whole leukemic cells clone. However, complete LSC elimination from the the patient’s bone marrow need search the main differences between the LSC and normal HSC. After the literature analysis it was found that LSC have several significant differences such as the ability to cause leukemia during the transplantation to immunodeficient animals, this leukemia is morphologically and phenotypically similar to the original tumor, in addition the LSC can be transmitted from animal to animal. In addition, the LSC is also characterized by the mutations presence

  12. NF-κB in T-cell Acute Lymphoblastic Leukemia: Oncogenic Functions in Leukemic and in Microenvironmental Cells

    International Nuclear Information System (INIS)

    Santos, Nuno R. dos; Ghezzo, Marinella N.; Silva, Ricardo C. da; Fernandes, Mónica T.

    2010-01-01

    Two main NF-κB signaling pathways, canonical and noncanonical, performing distinct functions in organisms have been characterized. Identification of mutations in genes encoding components of these NF-κB signaling pathways in lymphoid malignancies confirmed their key role in leukemogenesis. T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of thymocytes that despite significant therapeutic advances can still be fatal. Although mutations in NF-κB genes have not been reported in T-ALL, NF-κB constitutive activation in human T-ALL and in acute T-cell leukemia mouse models has been observed. Although these studies revealed activation of members of both canonical and noncanonical NF-κB pathways in acute T-cell leukemia, only inhibition of canonical NF-κB signaling was shown to impair leukemic T cell growth. Besides playing an important pro-oncogenic role in leukemic T cells, NF-κB signaling also appears to modulate T-cell leukemogenesis through its action in microenvironmental stromal cells. This article reviews recent data on the role of these transcription factors in T-ALL and pinpoints further research crucial to determine the value of NF-κB inhibition as a means to treat T-ALL

  13. NF-κB in T-cell Acute Lymphoblastic Leukemia: Oncogenic Functions in Leukemic and in Microenvironmental Cells

    Energy Technology Data Exchange (ETDEWEB)

    Santos, Nuno R. dos, E-mail: nrsantos@ualg.pt; Ghezzo, Marinella N.; Silva, Ricardo C. da; Fernandes, Mónica T. [IBB-Institute for Biotechnology and Bioengineering, Centre for Molecular and Structural Biomedicine (CBME), University of Algarve, Campus de Gambelas, 8005-139 Faro (Portugal)

    2010-11-05

    Two main NF-κB signaling pathways, canonical and noncanonical, performing distinct functions in organisms have been characterized. Identification of mutations in genes encoding components of these NF-κB signaling pathways in lymphoid malignancies confirmed their key role in leukemogenesis. T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive malignancy of thymocytes that despite significant therapeutic advances can still be fatal. Although mutations in NF-κB genes have not been reported in T-ALL, NF-κB constitutive activation in human T-ALL and in acute T-cell leukemia mouse models has been observed. Although these studies revealed activation of members of both canonical and noncanonical NF-κB pathways in acute T-cell leukemia, only inhibition of canonical NF-κB signaling was shown to impair leukemic T cell growth. Besides playing an important pro-oncogenic role in leukemic T cells, NF-κB signaling also appears to modulate T-cell leukemogenesis through its action in microenvironmental stromal cells. This article reviews recent data on the role of these transcription factors in T-ALL and pinpoints further research crucial to determine the value of NF-κB inhibition as a means to treat T-ALL.

  14. Do endothelial cells belong to the primitive stem leukemic clone in CML? Role of extracellular vesicles.

    Science.gov (United States)

    Ramos, Teresa L; Sánchez-Abarca, Luis Ignacio; López-Ruano, Guillermo; Muntión, Sandra; Preciado, Silvia; Hernández-Ruano, Montserrat; Rosado, Belén; de las Heras, Natalia; Chillón, M Carmen; Hernández-Hernández, Ángel; González, Marcos; Sánchez-Guijo, Fermín; Del Cañizo, Consuelo

    2015-08-01

    The expression of BCR-ABL in hematopoietic stem cells is a well-defined primary event in chronic myeloid leukemia (CML). Some reports have described the presence of BCR-ABL on endothelial cells from CML patients, suggesting the origin of the disease in a primitive hemangioblastic cell. On the other hand, extracellular vesicles (EVs) released by CML leukemic cells are involved in the angiogenesis modulation process. In the current work we hypothesized that EVs released from BCR-ABL(+) cells may carry inside the oncogene that can be transferred to endothelial cells leading to the expression of both BCR-ABL transcript and the oncoprotein. EVs from K562 cells and plasma of newly diagnosed CML patients were isolated by ultracentrifugation. RT-PCR analysis detected the presence of BCR-ABL RNA in the EVs isolated from both K562 cells and plasma of CML patients. The incorporation of these EVs into endothelial cells was demonstrated by flow cytometry and fluorescence microscopy showed that after 24h of incubation most EVs were incorporated. BCR-ABL transcripts were detected in all experiments on endothelial cells incubated with EVs from both sources. The presence of BCR-ABL on endothelial cells incubated with Philadelphia(+) EVs was also confirmed by Western blot assays. In summary, endothelial cells acquire BCR-ABL RNA and the oncoprotein after incubation with EVs released from Ph(+) positive cells (either from K562 cells or from plasma of newly diagnosed CML patients). This results challenge the hypothesis that endothelial cells may be part of the Philadelphia(+) clone in CML. Copyright © 2015 Elsevier Ltd. All rights reserved.

  15. Identification of residual leukemic cells by flow cytometry in childhood B-cell precursor acute lymphoblastic leukemia: verification of leukemic state by flow-sorting and molecular/cytogenetic methods.

    Science.gov (United States)

    Øbro, Nina F; Ryder, Lars P; Madsen, Hans O; Andersen, Mette K; Lausen, Birgitte; Hasle, Henrik; Schmiegelow, Kjeld; Marquart, Hanne V

    2012-01-01

    Reduction in minimal residual disease, measured by real-time quantitative PCR or flow cytometry, predicts prognosis in childhood B-cell precursor acute lymphoblastic leukemia. We explored whether cells reported as minimal residual disease by flow cytometry represent the malignant clone harboring clone-specific genomic markers (53 follow-up bone marrow samples from 28 children with B-cell precursor acute lymphoblastic leukemia). Cell populations (presumed leukemic and non-leukemic) were flow-sorted during standard flow cytometry-based minimal residual disease monitoring and explored by PCR and/or fluorescence in situ hybridization. We found good concordance between flow cytometry and genomic analyses in the individual flow-sorted leukemic (93% true positive) and normal (93% true negative) cell populations. Four cases with discrepant results had plausible explanations (e.g. partly informative immunophenotype and antigen modulation) that highlight important methodological pitfalls. These findings demonstrate that with sufficient experience, flow cytometry is reliable for minimal residual disease monitoring in B-cell precursor acute lymphoblastic leukemia, although rare cases require supplementary PCR-based monitoring.

  16. In vitro anti-leukemic activity and chemical transformation of the 5'-chloro-5'-deoxy derivative of cyclo-cytidine

    International Nuclear Information System (INIS)

    Stankovicova, M.; Bachrata, M.; Sveda, P.; Rauko, P.; Blesova, P.

    1995-01-01

    Hydrochloride of 5'-chloro-5'-deoxy-cytocytidine (Cl-cC) is an analogue of hydrochloride (cC), a pro-drug of the compound with of the compound with the strong anti-leukemic activity arabinosylcytosine (araC). This paper is devoted to the study of its cytotoxic activity in vitro and to the effect of acid alkaline conditions and temperature on its stability. Cl-cC inhibits not only the growth of L1210 leukemia cells in vitro and the DNA synthesis (IC 50 = 0.09 μmol/dm 3 ) but, at the same time, it has a weak effect on RNA synthesis (IC 50 > 250 μmol/dm 3 ) and no effect on proteosynthesis. In alkaline conditions Cl-cC is transformed to 5'-chloro-araC and 2',5'-anhydro-araC but is more stable in acid solutions. (author)

  17. Adhesion molecule profiles of B-cell non-Hodgkin's lymphomas in the leukemic phase

    Directory of Open Access Journals (Sweden)

    D.M. Matos

    2006-10-01

    Full Text Available We evaluated the expression of 10 adhesion molecules on peripheral blood tumor cells of 17 patients with chronic lymphocytic leukemia, 17 with mantle-cell lymphoma, and 13 with nodal or splenic marginal B-cell lymphoma, all in the leukemic phase and before the beginning of any therapy. The diagnosis of B-cell non-Hodgkin's lymphomas was based on cytological, histological, immunophenotypic, and molecular biology methods. The mean fluorescence intensity of the adhesion molecules in tumor cells was measured by flow cytometry of CD19-positive cells and differed amongst the types of lymphomas. Comparison of chronic lymphocytic leukemia and mantle-cell lymphoma showed that the former presented a higher expression of CD11c and CD49c, and a lower expression of CD11b and CD49d adhesion molecules. Comparison of chronic lymphocytic leukemia and marginal B-cell lymphoma showed that the former presented a higher expression of CD49c and a lower expression of CD11a, CD11b, CD18, CD49d, CD29, and CD54. Finally, comparison of mantle-cell lymphoma and marginal B-cell lymphoma showed that marginal B-cell lymphoma had a higher expression of CD11a, CD11c, CD18, CD29, and CD54. Thus, the CD49c/CD49d pair consistently demonstrated a distinct pattern of expression in chronic lymphocytic leukemia compared with mantle-cell lymphoma and marginal B-cell lymphoma, which could be helpful for the differential diagnosis. Moreover, the distinct profiles of adhesion molecules in these diseases may be responsible for their different capacities to invade the blood stream.

  18. Combined Treatment with Low Concentrations of Decitabine and SAHA Causes Cell Death in Leukemic Cell Lines but Not in Normal Peripheral Blood Lymphocytes

    Directory of Open Access Journals (Sweden)

    Barbora Brodská

    2013-01-01

    Full Text Available Epigenetic therapy reverting aberrant acetylation or methylation offers the possibility to target preferentially tumor cells and to preserve normal cells. Combination epigenetic therapy may further improve the effect of individual drugs. We investigated combined action of demethylating agent decitabine and histone deacetylase inhibitor SAHA (Vorinostat on different leukemic cell lines in comparison with peripheral blood lymphocytes. Large decrease of viability, as well as huge p21WAF1 induction, reactive oxygen species formation, and apoptotic features due to combined decitabine and SAHA action were detected in leukemic cell lines irrespective of their p53 status, while essentially no effect was observed in response to the combined drug action in normal peripheral blood lymphocytes of healthy donors. p53-dependent apoptotic pathway was demonstrated to participate in the wtp53 CML-T1 leukemic cell line response, while significant influence of reactive oxygen species on viability decrease has been detected in p53-null HL-60 cell line.

  19. Proteoglycan biosynthesis in murine monocytic leukemic (M1) cells before and after differentiation

    International Nuclear Information System (INIS)

    McQuillan, D.J.; Yanagishita, M.; Hascall, V.C.; Bickel, M.

    1989-01-01

    Murine monocytic leukemic (M1) cells were cultured in the presence of [ 3 H]glucosamine and [ 35 S]sulfate. Labeled proteoglycans were purified by anion exchange chromatography and characterized by gel filtration and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in combination with chemical and enzymatic degradation. M1 cells synthesize a single predominant species of proteoglycan which distributes almost equally between the cell and medium after 17 h labeling. The cell-associated proteoglycan has an overall size of about 135 kDa and contains three to five chondroitin sulfate chains (28-31 kDa each) attached to a chondroitinase-generated core protein of 28 kDa. The synthesis and subsequent secretion of this proteoglycan was enhanced 4-5-fold in cells induced to differentiate into macrophages. This was not a phenomenon of arrest in the G0/G1 stage of the cell cycle, since density inhibited undifferentiated cells arrested at this stage did not increase proteoglycan synthesis. The chondroitin sulfate chains contained exclusively chondroitin 4- and 6-sulfate; however, the ratio of these two disaccharides differed between the medium- and cell-associated proteoglycans, and changed during progression of the cells into a fully differentiated phenotype. Pulse-chase kinetics indicate the presence of two distinct pools of proteoglycan; one that is secreted very rapidly from the cell after a approximately 1-h lag, and a second pool that is turned over in the cell with a half-time of approximately 3.5 h. Subtle differences in the glycosylation patterns of the medium- and cell-associated species are consistent with synthesis of two pools. Papain digestion suggests that the chondroitin sulfate chains are clustered on a small protease resistant peptide. The data suggest that this proteoglycan is similar to the serglycin proteoglycan family

  20. The Leukemic Stem Cell Niche: Adaptation to “Hypoxia” versus Oncogene Addiction

    Directory of Open Access Journals (Sweden)

    Giulia Cheloni

    2017-01-01

    Full Text Available Previous studies based on low oxygen concentrations in the incubation atmosphere revealed that metabolic factors govern the maintenance of normal hematopoietic or leukemic stem cells (HSC and LSC. The physiological oxygen concentration in tissues ranges between 0.1 and 5.0%. Stem cell niches (SCN are placed in tissue areas at the lower end of this range (“hypoxic” SCN, to which stem cells are metabolically adapted and where they are selectively hosted. The data reported here indicated that driver oncogenic proteins of several leukemias are suppressed following cell incubation at oxygen concentration compatible with SCN physiology. This suppression is likely to represent a key positive regulator of LSC survival and maintenance (self-renewal within the SCN. On the other hand, LSC committed to differentiation, unable to stand suppression because of addiction to oncogenic signalling, would be unfit to home in SCN. The loss of oncogene addiction in SCN-adapted LSC has a consequence of crucial practical relevance: the refractoriness to inhibitors of the biological activity of oncogenic protein due to the lack of their molecular target. Thus, LSC hosted in SCN are suited to sustain the long-term maintenance of therapy-resistant minimal residual disease.

  1. Noninvasive identification of subcellular organization and nuclear morphology features associated with leukemic cells using light-scattering spectroscopy

    Science.gov (United States)

    Hsiao, Austin; Hunter, Martin; Greiner, Cherry; Gupta, Sharad; Georgakoudi, Irene

    2011-03-01

    Leukemia is the most common and deadly cancer among children and one of the most prevalent cancers among adults. Improvements in its diagnosis and monitoring of leukemic patients could have a significant impact in their long-term treatment. We demonstrate that light-scattering spectroscopy (LSS)-based approaches could serve as a tool to achieve this goal. Specifically, we characterize the light scattering properties of leukemic (NALM-6) cells and compare them to those of normal lymphocytes and granulocytes in the 440-710 nm range, over +/-4 deg about the exact backscattering direction. We find that the LSS spectra are well described by an inverse power-law wavelength dependence, with a power exponent insensitive to the scattering angle but significantly higher for leukemic cells than for normal leukocytes. This is consistent with differences in the subcellular morphology of these cells, detected in differential interference contrast images. Furthermore, the residual light-scattering signal, extracted after subtracting the inverse power-law fit from the data, can be analyzed assuming a Gaussian distribution of spherical scatterers using Mie theory. This analysis yields scatterer sizes that are consistent with the diameters of cell nuclei and allows the detection of the larger nuclei of NALM-6 cells compared to those of lymphocytes and granulocytes.

  2. Expression of MICA, MICB and NKG2D in human leukemic myelomonocytic and cervical cancer cells

    Directory of Open Access Journals (Sweden)

    Mendoza-Rincon Jorge

    2011-04-01

    Full Text Available Abstract Background Cancer cells are known to secrete the stress molecules MICA and MICB that activate cytotoxicity by lymphocytes and NK cells through their NKG2D receptor as a mechanism of immunological defense. This work was undertaken to evaluate if cancer cells can also express this receptor as a possible mechanisms of depletion of MIC molecules and thus interfere with their immune recognition. Methods Myelomonocytic leukemic (TPH-1 and U-937 and cervical cancer (CALO and INBL cell lines were evaluated by Western Blot, ELISA, flow cytometry and immunocytochemistry to evaluate their capacity to express and secrete MICA and MICB and to be induced to proliferate by these molecules as well as to express their receptor NKG2D. Statistical analysis was performed by two-way ANOVA for time course analysis and Student's t-test for comparison between groups. Values were considered significantly different if p Results THP-1 and U-937 produce and secrete the stress MICA and MICB as shown by Western Blot of lysed cells and by ELISA of their conditioned media. By Western Blot and flow cytometry we found that these cells also express the receptor NKG2D. When THP-1 and U-937 were cultured with recombinant MICA and MICB they exhibited a dose dependent induction for their proliferation. CALO and INBL also produce MICA and MICB and were induced to proliferate by these stress molecules. By Western Blot, flow cytometry and immunocytochemistry we also found that these cells express NKG2D. Conclusions Our novel results that tumor cells can simultaneously secrete MIC molecules and express their receptor, and to be induced for proliferation by these stress molecules, and that tumor epithelial cells can also express the NKG2D receptor that was thought to be exclusive of NK and cytotoxic lymphocytes is discussed as a possible mechanism of immunological escape and of tumor growth induction.

  3. Long Terminal Repeat CRISPR-CAR-Coupled "Universal" T Cells Mediate Potent Anti-leukemic Effects.

    Science.gov (United States)

    Georgiadis, Christos; Preece, Roland; Nickolay, Lauren; Etuk, Aniekan; Petrova, Anastasia; Ladon, Dariusz; Danyi, Alexandra; Humphryes-Kirilov, Neil; Ajetunmobi, Ayokunmi; Kim, Daesik; Kim, Jin-Soo; Qasim, Waseem

    2018-03-06

    Gene editing can be used to overcome allo-recognition, which otherwise limits allogeneic T cell therapies. Initial proof-of-concept applications have included generation of such "universal" T cells expressing chimeric antigen receptors (CARs) against CD19 target antigens combined with transient expression of DNA-targeting nucleases to disrupt the T cell receptor alpha constant chain (TRAC). Although relatively efficient, transgene expression and editing effects were unlinked, yields variable, and resulting T cell populations heterogeneous, complicating dosing strategies. We describe a self-inactivating lentiviral "terminal" vector platform coupling CAR expression with CRISPR/Cas9 effects through incorporation of an sgRNA element into the ΔU3 3' long terminal repeat (LTR). Following reverse transcription and duplication of the hybrid ΔU3-sgRNA, delivery of Cas9 mRNA resulted in targeted TRAC locus cleavage and allowed the enrichment of highly homogeneous (>96%) CAR + (>99%) TCR - populations by automated magnetic separation. Molecular analyses, including NGS, WGS, and Digenome-seq, verified on-target specificity with no evidence of predicted off-target events. Robust anti-leukemic effects were demonstrated in humanized immunodeficient mice and were sustained longer than by conventional CAR + TCR + T cells. Terminal-TRAC (TT) CAR T cells offer the possibility of a pre-manufactured, non-HLA-matched CAR cell therapy and will be evaluated in phase 1 trials against B cell malignancies shortly. Copyright © 2018 The American Society of Gene and Cell Therapy. Published by Elsevier Inc. All rights reserved.

  4. Flow cytometric analysis of expression of interleukin-2 receptor beta chain (p70-75) on various leukemic cells

    International Nuclear Information System (INIS)

    Hoshino, S.; Oshimi, K.; Tsudo, M.; Miyasaka, M.; Teramura, M.; Masuda, M.; Motoji, T.; Mizoguchi, H.

    1990-01-01

    We analyzed the expression of the interleukin-2 receptor (IL-2R) beta chain (p70-75) on various leukemic cells from 44 patients by flow cytometric analysis using the IL-2R beta chain-specific monoclonal antibody, designated Mik-beta 1. Flow cytometric analysis demonstrated the expression of the IL-2R beta chain on granular lymphocytes (GLs) from all eight patients with granular lymphocyte proliferative disorders (GLPDs), on adult T-cell leukemia (ATL) cells from all three patients with ATL, and on T-cell acute lymphoblastic leukemia (T-ALL) cells from one of three patients with T-ALL. Although GLs from all the GLPD patients expressed the IL-2R beta chain alone and not the IL-2R alpha chain (Tac-antigen: p55), ATL and T-ALL cells expressing the beta chain coexpressed the alpha chain. In two of seven patients with common ALL (cALL) and in both patients with B-cell chronic lymphocytic leukemia, the leukemic cells expressed the alpha chain alone. Neither the alpha chain nor the beta chain was expressed on leukemic cells from the remaining 28 patients, including all 18 patients with acute nonlymphocytic leukemia, five of seven patients with cALL, all three patients with multiple myeloma, and two of three patients with T-ALL. These results indicate that three different forms of IL-2R chain expression exist on leukemic cells: the alpha chain alone; the beta chain alone; and both the alpha and beta chains. To examine whether the results obtained by flow cytometric analysis actually reflect functional aspects of the expressed IL-2Rs, we studied the specific binding of 125I-labeled IL-2 (125I-IL-2) to leukemic cells in 18 of the 44 patients. In addition, we performed 125I-IL-2 crosslinking studies in seven patients. The results of IL-2R expression of both 125I-IL-2 binding assay and crosslinking studies were in agreement with those obtained by flow cytometric analysis

  5. Rapid Treatment of Leukostasis in Leukemic Mantle Cell Lymphoma Using Therapeutic Leukapheresis: A Case Report

    Directory of Open Access Journals (Sweden)

    Xuan Duc Nguyen

    2011-01-01

    Full Text Available We describe a case of severe leukocytosis caused by leukemic mantle cell lymphoma (MCL, complicated by leukostasis with myocardial infarction in which leukapheresis was used in the initial management. A 73-year-old male presented to the emergency department because of fatigue and thoracic pain. Blood count revealed 630 × 109/L WBC (white blood cells. The electrocardiogram showed ST-elevation with an increase of troponin and creatinine kinase. The diagnosis was ST-elevation myocardial infarction (STEMI induced and complicated by leukostasis. Immunophenotyping, morphology, cytogenetic and fluorescence-in-situ-hybridization analysis revealed the diagnosis of a blastoid variant of MCL. To remove leukocytes rapidly, leukapheresis was performed in the intensive care unit. Based on the differential blood count with 95% blasts, which were assigned to the lymphocyte population by the automatic hematology analyzer, leukapheresis procedures were then performed with the mononuclear cell standard program on the Spectra cell separator. The patient was treated with daily leukapheresis for 3 days. The WBC count decreased to 174 × 109/L after the third leukapheresis, with a 72% reduction. After the second apheresis, treatment with vincristine, cyclophosphamide, and prednisolone was started. The patient fully recovered in the further course of the treatment. To the best of our knowledge, this is the first report on blastoid MCL with leukostasis associated with a STEMI that was successfully treated by leukapheresis. Effective harvest of circulating lymphoma cells by leukapheresis requires adaptation of instrument settings based on the results of the differential blood count prior to apheresis.

  6. Overcoming of P-glycoprotein mediated vincristine resistance of L1210/VCR mouse leukemic cells could be induced by pentoxifylline but not by theophylline and caffeine

    International Nuclear Information System (INIS)

    Stefankova, Z.; Barancik, M.; Breier, A.

    1996-01-01

    Effects of xanthine derivatives (pentoxifylline (PTX), caffeine, theophylline, 1-methyl-3-isobutylxanthine) on P-glycoprotein mediated vincristine resistance of L1210/VCR mouse leukemic cell sub-line were studied. From the applied xanthines only PTX was found to reverse the vincristine resistance of the above cells. Moreover, only PTX, but not other xanthine, increased the accumulation of [ 3 H]vincristine by L1210/VCR cells. Thus it may be concluded that PTX-induced reversal of vincristine (VCR) resistance could not be explained from the point of known pharmacological effects of PTX that are common for other xanthines such as inhibition of phosphodiesterase activity, calcium mobilizing effect, inhibition of tumor necrosis factor α (TNF), etc. (author)

  7. Alkaloid-rich fraction of Himatanthus lancifolius contains anti-tumor agents against leukemic cells

    Directory of Open Access Journals (Sweden)

    Melissa Pires de Lima

    2010-06-01

    Full Text Available The effects of the alkaloid-rich fraction of Himatanthus lancifolius (Müll. Arg Woodson on normal marrow cells and leukemic cell lines were investigated. After 48 h exposure, the proliferation assay showed significant cell growth inhibition for Daudi (0.1-10 µg/mL, K-562 (1-10 µg/mL, and REH cells (10-100 µg/mL, yet was inert for normal marrow cells. A similar inhibition profile was observed in clonogenic assays. This alkaloid-rich fraction, in which uleine is the main compound, showed no signs of toxicity to any cells up to 10 µg/mL. Cell feature analyses after induction of differentiation showed maintenance of the initial phenotype. Flow cytometric expression of Annexin-V and 7-AAD in K-562 and Daudi cells has indicated that the cells were not undergoing apoptosis or necrosis, suggesting cytostatic activity for tumor cellsOs efeitos da fração rica em alcalóides indólicos de Himatanthus lancifolius (Müll. Arg Woodson sobre células normais de medula óssea e linhagens celulares leucêmicas foram investigados. Após 48 horas de exposição, os ensaios de proliferação demonstraram efeitos inibitórios significativos para as linhagens Daudi (0,1-10 µg/mL, K-562 (1-10 µg/mL e REH (10-100 µg/mL, enquanto mostrou-se inerte sobre células normais de medula óssea. Os perfis de inibição se repetiram nos ensaios clonogênicos. A fração rica em alcalóides, na qual a uleína é a substância majoritária, não demonstrou toxicidade até a dose de 10 µg/mL para nenhuma das células incluídas no estudo. Da mesma forma, não se observou influência dessa fração sobre a diferenciação celular dessas linhagens, mas manutenção de seu estado maturacional inicial. O conjunto de dados descritos associado à baixa co-expressão de anexina-V e 7-AAD sugerem que esta fração exerce atividade citostática para células tumorais.

  8. Indoleamine 2,3-dioxygenase-expressing leukemic dendritic cells impair a leukemia-specific immune response by inducing potent T regulatory cells.

    Science.gov (United States)

    Curti, Antonio; Trabanelli, Sara; Onofri, Chiara; Aluigi, Michela; Salvestrini, Valentina; Ocadlikova, Darina; Evangelisti, Cecilia; Rutella, Sergio; De Cristofaro, Raimondo; Ottaviani, Emanuela; Baccarani, Michele; Lemoli, Roberto M

    2010-12-01

    The immunoregulatory enzyme indoleamine 2,3-dioxygenase, which catalyzes the conversion of tryptophan into kynurenine, is expressed in a significant subset of patients with acute myeloid leukemia, resulting in the inhibition of T-cell proliferation and the induction of regulatory T cells. Acute myeloid leukemia cells can be differentiated into dendritic cells, which have increased immunogenicity and have been proposed as vaccines against leukemia. Leukemic dendritic cells were generated from acute myeloid leukemia cells and used as stimulators in functional assays, including the induction of regulatory T cells. Indoleamine 2,3-dioxygenase expression in leukemic dendritic cells was evaluated at molecular, protein and enzymatic levels. We demonstrate that, after differentiation into dendritic cells, both indoleamine 2,3-dioxygenase-negative and indoleamine 2,3-dioxygenase-positive acute myeloid leukemia samples show induction and up-regulation of indoleamine 2,3-dioxygenase gene and protein, respectively. Indoleamine 2,3-dioxygenase-positive acute myeloid leukemia dendritic cells catabolize tryptophan into kynurenine metabolite and inhibit T-cell proliferation through an indoleamine 2,3-dioxygenase-dependent mechanism. Moreover, indoleamine 2,3-dioxygenase-positive leukemic dendritic cells increase the number of allogeneic and autologous CD4(+)CD25(+) Foxp3(+) T cells and this effect is completely abrogated by the indoleamine 2,3-dioxygenase-inhibitor, 1-methyl tryptophan. Purified CD4(+)CD25(+) T cells obtained from co-culture with indoleamine 2,3-dioxygenase-positive leukemic dendritic cells act as regulatory T cells as they inhibit naive T-cell proliferation and impair the complete maturation of normal dendritic cells. Importantly, leukemic dendritic cell-induced regulatory T cells are capable of in vitro suppression of a leukemia-specific T cell-mediated immune response, directed against the leukemia-associated antigen, Wilms' tumor protein. These data identify

  9. TRIM32 promotes retinoic acid receptor α-mediated differentiation in human promyelogenous leukemic cell line HL60

    International Nuclear Information System (INIS)

    Sato, Tomonobu; Okumura, Fumihiko; Iguchi, Akihiro; Ariga, Tadashi; Hatakeyama, Shigetsugu

    2012-01-01

    Highlights: ► TRIM32 enhanced RARα-mediated transcriptional activity even in the absence of RA. ► TRIM32 stabilized RARα in the human promyelogenous leukemic cell line HL60. ► Overexpression of TRIM32 in HL60 cells induced granulocytic differentiation. ► TRIM32 may function as a coactivator for RARα-mediated transcription in APL cells. -- Abstract: Ubiquitination, one of the posttranslational modifications, appears to be involved in the transcriptional activity of nuclear receptors including retinoic acid receptor α (RARα). We previously reported that an E3 ubiquitin ligase, TRIM32, interacts with several important proteins including RARα and enhances transcriptional activity of RARα in mouse neuroblastoma cells and embryonal carcinoma cells. Retinoic acid (RA), which acts as a ligand to nuclear receptors including RARα, plays crucial roles in development, differentiation, cell cycles and apoptosis. In this study, we found that TRIM32 enhances RARα-mediated transcriptional activity even in the absence of RA and stabilizes RARα in the human promyelogenous leukemic cell line HL60. Moreover, we found that overexpression of TRIM32 in HL60 cells suppresses cellular proliferation and induces granulocytic differentiation even in the absence of RA. These findings suggest that TRIM32 functions as one of the coactivators for RARα-mediated transcription in acute promyelogenous leukemia (APL) cells, and thus TRIM32 may become a potentially therapeutic target for APL.

  10. Elimination of acute muelogenous leukemic cells from marrow and tumor suspensions in the rat with 4-hydroperoxycyclophosphamide

    International Nuclear Information System (INIS)

    Sharkis, S.J.; Santos, G.W.; Colvin, M.

    1980-01-01

    Cell suspensions of normal rat marrow mixed with rat acute myelogenous leukemic cells were prepared and incubated in vitro with graded doses of 4-hydroperoxycyclophosphamide (4HC). The cell suspensions were injected into rats prepared with a lethal dose of total body irradiation. Animals injected with these cells survived fatal irradiation induced aplasia. In a dose related manner 4HC was able to purge tumor cells from the cell mixtures. Thus, animals given cell suspensions incubated with the lower doses of 4HC showed prolonged survived before death from leukemia and animals given cell suspensions incubated with higher doses of 4HC survival lethal irradiation without the subsequent appearance of leukemia. These studies clearly establish that tumor cells may be eliminated from normal marrow suspensions without completely destroying the pluripotent stem cells

  11. Inhibition of time-dependent enhancement of amino acid transport by leukemic leukocytes: a possible index of the sensitivity of cells to drugs

    Energy Technology Data Exchange (ETDEWEB)

    Frengley, P A; Peck, W A; Lichtman, M A

    1975-01-01

    Leukemic leukocytes increase their rates of alpha aminoisobutyric acid (AIB) accumulation when incubated for prolonged periods in amino acid deficient media. The time-dependent increase was prevented by concurrent exposure of cells to cycloheximide or actinomycin D in vitro. In addition, the increase in AIB uptake was not present in leukemic blasts studied in vitro when the cells were obtained from subjects with acute myeloblastic leukemia who had received antileukemic therapy. Cortisol added to cell suspensions in vitro inhibited the development of time-dependent increases in AIB uptake in lymphoid cells, but accentuated the process slightly in myeloblasts. Cortisol administered to a subject with CLL by infusion reduced the time-dependent increase in AIB uptake by CLL cells subsequently studied in vitro. These data indicate that the time-dependent increase in AIB uptake may be a means of testing the sensitivity of leukemic cells to drugs.

  12. Analysis of CTCL cell lines reveals important differences between mycosis fungoides/Sézary syndrome vs. HTLV-1+ leukemic cell lines

    DEFF Research Database (Denmark)

    Netchiporouk, Elena; Gantchev, Jennifer; Tsang, Matthew

    2017-01-01

    HTLV-1 is estimated to affect ~20 million people worldwide and in ~5% of carriers it produces Adult T-Cell Leukemia/Lymphoma (ATLL), which can often masquerade and present with classic erythematous pruritic patches and plaques that are typically seen in Mycosis Fungoides (MF) and Sézary Syndrome...... (SS), the most recognized variants of Cutaneous T-Cell Lymphomas (CTCL). For many years the role of HTLV- 1 in the pathogenesis of MF/SS has been hotly debated. In this study we analyzed CTCL vs. HTLV-1+ leukemic cells. We performed G-banding/spectral karyotyping, extensive gene expression analysis......, TP53 sequencing in the 11 patient-derived HTLV- 1+ (MJ and Hut102) vs. HTLV-1- (Myla, Mac2a, PB2B, HH, H9, Hut78, SZ4, Sez4 and SeAx) CTCL cell lines. We further tested drug sensitivities to commonly used CTCL therapies and studied the ability of these cells to produce subcutaneous xenograft tumors...

  13. Properties of murine leukemia viruses produced by leukemic cells established from NIH Swiss mice with radiation-induced leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Okumoto, Masaaki; Nishikawa, Ryosuke; Takamori, Yasuhiko; Iwai, Yoshiaki; Iwai, Mineko [Radiation Center of Osaka Prefecture, Sakai (Japan); Imai, Shunsuke; Morimoto, Junji; Tsubura, Yoshihiko

    1984-06-01

    Three leukemic cell lines, designated NIH-RL1, NIH-RL2 and NFS-RL1, were established from spleen and thymuses of NIH Swiss and NFS mice with radiation-induced leukemia. The culture fluids of these cell lines contained RNA-dependent DNA polymerase (RDDP) activities associated with particles of buoyant density of 1.15-1.17 (g/cm/sup 3/). The divalent cation reqirement of these enzymes was characteristic for that of murine leukemia viruses. In competition radioimmunoassay, a major core protein, p30, was detected in culture fluid of each leukemic cell line. Competition curves of viral p30 produced by these cell lines revealed that these viruses were very similar to those of xenotropic viruses of NZB mice. These viruses were undetectable both by XC plaque assay using SC-1 cells as an indicator cell, and by mink S/sup +/L/sup -/ focus induction assay. These viruses also lacked productive infectivity to mink lung cells (CCL-64), and were nononcogenic in syngeneic mice when the viruses were intrathymically inoculated.

  14. Indoleamine 2,3-dioxygenase-expressing leukemic dendritic cells impair a leukemia-specific immune response by inducing potent T regulatory cells

    OpenAIRE

    Curti, A; Trabanelli, S; Onofri, C; Aluigi, M; Salvestrini, V; Ocadlikova, D; Evangelisti, C; Rutella, S; De Cristofaro, R; Ottaviani, E; Baccarani, M; Lemoli, RM

    2010-01-01

    Background: The immunoregulatory enzyme indoleamine 2,3-dioxygenase, which catalyzes the conversion of tryptophan into kynurenine, is expressed in a significant subset of patients with acute myeloid leukemia, resulting in the inhibition of T-cell proliferation and the induction of regulatory T cells. Acute myeloid leukemia cells can be differentiated into dendritic cells, which have increased immunogenicity and have been proposed as vaccines against leukemia.\\ud Design and Methods: Leukemic d...

  15. Human Leukemic Cells performing Oxidative Phosphorylation (OXPHOS Generate an Antioxidant Response Independently of Reactive Oxygen species (ROS Production

    Directory of Open Access Journals (Sweden)

    Abrar Ul Haq Khan

    2016-01-01

    Full Text Available Tumor cell metabolism is altered during leukemogenesis. Cells performing oxidative phosphorylation (OXPHOS generate reactive oxygen species (ROS through mitochondrial activity. To limit the deleterious effects of excess ROS, certain gene promoters contain antioxidant response elements (ARE, e.g. the genes NQO-1 and HO-1. ROS induces conformational changes in KEAP1 and releases NRF2, which activates AREs. We show in vitro and in vivo that OXPHOS induces, both in primary leukemic cells and cell lines, de novo expression of NQO-1 and HO-1 and also the MAPK ERK5 and decreases KEAP1 mRNA. ERK5 activates the transcription factor MEF2, which binds to the promoter of the miR-23a–27a–24-2 cluster. Newly generated miR-23a destabilizes KEAP1 mRNA by binding to its 3′UTR. Lower KEAP1 levels increase the basal expression of the NRF2-dependent genes NQO-1 and HO-1. Hence, leukemic cells performing OXPHOS, independently of de novo ROS production, generate an antioxidant response to protect themselves from ROS.

  16. NO-donating aspirin inhibits the growth of leukemic Jurkat cells and modulates β-catenin expression

    International Nuclear Information System (INIS)

    Nath, Niharika; Labaze, Georges; Rigas, Basil; Kashfi, Khosrow

    2004-01-01

    β-Catenin has been implicated in leukemic cell proliferation. We compared the effects of aspirin (ASA) and the ortho, meta, and para positional isomers of NO-donating aspirin (NO-ASA) on cell growth and β-catenin expression in human Jurkat T leukemic cells. Cell growth inhibition was strong: IC 50 for p-, o-, and m- were 20 ± 1.6 (mean ± SEM), 15 ± 1.5, and 200 ± 12 μM, respectively, in contrast to that of ASA (3200 ± 375 μM). The para isomer of NO-ASA degraded β-catenin in a dose- and time-dependent manner coinciding with increasing expression of activated caspase-3. The caspase inhibitor ZVAD blocked β-catenin cleavage by p-NO-ASA and partially reversed cell growth inhibition by p-NO-ASA but not that by ASA. A denitrated analog of p-NO-ASA did not degrade β-catenin indicating the importance of the NO-donating moiety. Our findings suggest that NO-ASA merits further study as an agent against leukemia

  17. Differential Activity of Voltage- and Ca2+-Dependent Potassium Channels in Leukemic T Cell Lines: Jurkat Cells Represent an Exceptional Case

    Directory of Open Access Journals (Sweden)

    Salvador Valle-Reyes

    2018-05-01

    Full Text Available Activation of resting T cells relies on sustained Ca2+ influx across the plasma membrane, which in turn depends on the functional expression of potassium channels, whose activity repolarizes the membrane potential. Depending on the T-cells subset, upon activation the expression of Ca2+- or voltage-activated K+ channels, KCa or Kv, is up-regulated. In this study, by means of patch-clamp technique in the whole cell mode, we have studied in detail the characteristics of Kv and KCa currents in resting and activated human T cells, the only well explored human T-leukemic cell line Jurkat, and two additional human leukemic T cell lines, CEM and MOLT-3. Voltage dependence of activation and inactivation of Kv1.3 current were shifted up to by 15 mV to more negative potentials upon a prolonged incubation in the whole cell mode and displayed little difference at a stable state in all cell lines but CEM, where the activation curve was biphasic, with a high and low potential components. In Jurkat, KCa currents were dominated by apamine-sensitive KCa2.2 channels, whereas only KCa3.1 current was detected in healthy T and leukemic CEM and MOLT-3 cells. Despite a high proliferation potential of Jurkat cells, Kv and KCa currents were unexpectedly small, more than 10-fold lesser as compared to activated healthy human T cells, CEM and MOLT-3, which displayed characteristic Kv1.3high:KCa3.1high phenotype. Our results suggest that Jurkat cells represent perhaps a singular case and call for more extensive studies on primary leukemic T cell lines as well as a verification of the therapeutic potential of specific KCa3.1 blockers to combat acute lymphoblastic T leukemias.

  18. Human TM9SF4 Is a New Gene Down-Regulated by Hypoxia and Involved in Cell Adhesion of Leukemic Cells.

    Directory of Open Access Journals (Sweden)

    Rosa Paolillo

    Full Text Available The transmembrane 9 superfamily protein member 4, TM9SF4, belongs to the TM9SF family of proteins highly conserved through evolution. TM9SF4 homologs, previously identified in many different species, were mainly involved in cellular adhesion, innate immunity and phagocytosis. In human, the function and biological significance of TM9SF4 are currently under investigation. However, TM9SF4 was found overexpressed in human metastatic melanoma and in a small subset of acute myeloid leukemia (AMLs and myelodysplastic syndromes, consistent with an oncogenic function of this gene.In this study, we first analyzed the expression and regulation of TM9SF4 in normal and leukemic cells and identified TM9SF4 as a gene highly expressed in human quiescent CD34+ hematopoietic progenitor cells (HPCs, regulated during monocytic and granulocytic differentiation of HPCs, both lineages giving rise to mature myeloid cells involved in adhesion, phagocytosis and immunity. Then, we found that TM9SF4 is markedly overexpressed in leukemic cells and in AMLs, particularly in M2, M3 and M4 AMLs (i.e., in AMLs characterized by the presence of a more or less differentiated granulocytic progeny, as compared to normal CD34+ HPCs. Proliferation and differentiation of HPCs occurs in hypoxia, a physiological condition in bone marrow, but also a crucial component of cancer microenvironment. Here, we investigated the impact of hypoxia on TM9SF4 expression in leukemic cells and identified TM9SF4 as a direct target of HIF-1α, downregulated in these cells by hypoxia. Then, we found that the hypoxia-mediated downregulation of TM9SF4 expression is associated with a decrease of cell adhesion of leukemic cells to fibronectin, thus demonstrating that human TM9SF4 is a new molecule involved in leukemic cell adhesion.Altogether, our study reports for the first time the expression of TM9SF4 at the level of normal and leukemic hematopoietic cells and its marked expression at the level of AMLs

  19. Isolation of a novel chronic lymphocytic leukemic (CLL) cell line and development of an in vivo mouse model of CLL.

    Science.gov (United States)

    Kellner, Joshua; Wierda, William; Shpall, Elizabeth; Keating, Michael; McNiece, Ian

    2016-01-01

    Leukemic cell lines have become important tools for studies of disease providing a monoclonal cell population that can be extensively expanded in vitro while preserving leukemic cellular characteristics. However, studies of chronic lymphocytic leukemia (CLL) have been impeded in part by the lack of continuous human cell lines. CLL cells have a high spontaneous apoptosis rate in vitro and exhibit minimal proliferation in xenograft models. Therefore, there is a need for development of primary CLL cell lines and we describe the isolation of such a line from the bone marrow of a CLL patient (17p deletion and TP53 mutation) which has been in long term culture for more than 12 months with continuous proliferation. The CLL cell line (termed MDA-BM5) which was generated in vitro with continuous co-culture on autologous stromal cells is CD19+CD5+ and shows an identical pattern of somatic hypermutation as determined in the patient's bone marrow (BM), confirming the origin of the cells from the original CLL clone. MDA-BM5 cells were readily transplantable in NOD/SCID gamma null mice (NSG) with disease developing in the BM, liver and spleen. BM cells from quaternary serial transplantation in NSG mice demonstrated the presence of CD19+CD5+ cells with Ig restricted to lambda which is consistent with the original patient cells. These studies describe a new CLL cell line from a patient with del(17p) that provides a unique model for in vitro and in vivo studies. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Inhibitory effect of turmeric curcuminoids on FLT3 expression and cell cycle arrest in the FLT3-overexpressing EoL-1 leukemic cell line.

    Science.gov (United States)

    Tima, Singkome; Ichikawa, Hideki; Ampasavate, Chadarat; Okonogi, Siriporn; Anuchapreeda, Songyot

    2014-04-25

    Leukemia is a hematologic malignancy with a frequent incidence and high mortality rate. Previous studies have shown that the FLT3 gene is overexpressed in leukemic blast cells, especially in acute myeloid leukemia. In this study, a commercially available curcuminoid mixture (1), pure curcumin (2), pure demethoxycurcumin (3), and pure bisdemethoxycurcumin (4) were investigated for their inhibitory effects on cell growth, FLT3 expression, and cell cycle progression in an FLT3-overexpressing EoL-1 leukemic cell line using an MTT assay, Western blotting, and flow cytometry, respectively. The mixture (1) and compounds 2-4 demonstrated cytotoxic effects with IC50 values ranging from 6.5 to 22.5 μM. A significant decrease in FLT3 protein levels was found after curcuminoid treatment with IC20 doses, especially with mixture 1 and compound 2. In addition, mixture 1 and curcumin (2) showed activity on cell cycle arrest at the G0/G1 phase and decreased the FLT3 and STAT5A protein levels in a dose-dependent manner. Compound 2 demonstrated the greatest potential for inhibiting cell growth, cell cycle progression, and FLT3 expression in EoL-1 cells. This investigation has provided new findings regarding the effect of turmeric curcuminoids on FLT3 expression in leukemic cells.

  1. Brief Report: Human Acute Myeloid Leukemia Reprogramming to Pluripotency Is a Rare Event and Selects for Patient Hematopoietic Cells Devoid of Leukemic Mutations.

    Science.gov (United States)

    Lee, Jong-Hee; Salci, Kyle R; Reid, Jennifer C; Orlando, Luca; Tanasijevic, Borko; Shapovalova, Zoya; Bhatia, Mickie

    2017-09-01

    Induced pluripotent stem cell reprogramming has provided critical insights into disease processes by modeling the genetics and related clinical pathophysiology. Human cancer represents highly diverse genetics, as well as inter- and intra-patient heterogeneity, where cellular model systems capable of capturing this disease complexity would be invaluable. Acute myeloid leukemia (AML) represents one of most heterogeneous cancers and has been divided into genetic subtypes correlated with unique risk stratification over the decades. Here, we report our efforts to induce pluripotency from the heterogeneous population of human patients that represents this disease in the clinic. Using robust optimized reprogramming methods, we demonstrate that reprogramming of AML cells harboring leukemic genomic aberrations is a rare event with the exception of those with de novo mixed-lineage leukemia (MLL) mutations that can be reprogrammed and model drug responses in vitro. Our findings indicate that unlike hematopoietic cells devoid of genomic aberrations, AML cells harboring driver mutations are refractory to reprogramming. Expression of MLL fusion proteins in AML cells did not contribute to induced reprogramming success, which continued to select for patient derived cells devoid of AML patient-specific aberrations. Our study reveals that unanticipated blockades to achieving pluripotency reside within the majority of transformed AML patient cells. Stem Cells 2017;35:2095-2102. © 2017 AlphaMed Press.

  2. Gemtuzumab Ozogamicin (GO Inclusion to Induction Chemotherapy Eliminates Leukemic Initiating Cells and Significantly Improves Survival in Mouse Models of Acute Myeloid Leukemia

    Directory of Open Access Journals (Sweden)

    Cathy C Zhang

    2018-01-01

    Full Text Available Gemtuzumab ozogamicin (GO is an anti-CD33 antibody-drug conjugate for the treatment of acute myeloid leukemia (AML. Although GO shows a narrow therapeutic window in early clinical studies, recent reports detailing a modified dosing regimen of GO can be safely combined with induction chemotherapy, and the combination provides significant survival benefits in AML patients. Here we tested whether the survival benefits seen with the combination arise from the enhanced reduction of chemoresidual disease and leukemic initiating cells (LICs. Herein, we use cell line and patient-derived xenograft (PDX AML models to evaluate the combination of GO with daunorubicin and cytarabine (DA induction chemotherapy on AML blast growth and animal survival. DA chemotherapy and GO as separate treatments reduced AML burden but left significant chemoresidual disease in multiple AML models. The combination of GO and DA chemotherapy eliminated nearly all AML burden and extended overall survival. In two small subsets of AML models, chemoresidual disease following DA chemotherapy displayed hallmark markers of leukemic LICs (CLL1 and CD34. In vivo, the two chemoresistant subpopulations (CLL1+/CD117− and CD34+/CD38+ showed higher ability to self-renewal than their counterpart subpopulations, respectively. CD33 was coexpressed in these functional LIC subpopulations. We demonstrate that the GO and DA induction chemotherapy combination more effectively eliminates LICs in AML PDX models than either single agent alone. These data suggest that the survival benefit seen by the combination of GO and induction chemotherapy, nonclinically and clinically, may be attributed to the enhanced reduction of LICs.

  3. Analysis of myelomonocytic leukemic differentiation by a cell surface marker panel including a fucose-binding lectin from Lotus tetragonolobus.

    Science.gov (United States)

    Elias, L; Van Epps, D E

    1984-06-01

    The fucose-binding lectin from Lotus tetragonolobus ( FBL -L) has been previously shown to bind specifically to normal cells of the myeloid and monocytic lineages. The purpose of this study was to explore the utility of fluoresceinated FBL -L as a leukemia differentiation marker in conjunction with a panel of other frequently used surface markers (Fc receptor, HLA-DR, OKM1, and antimonocyte antibody). FBL -L reacted with leukemic cells in 8/9 cases of clinically recognized acute myeloid leukemia, including myeloid blast crisis of chronic granulocytic leukemia, 3/3 cases of chronic phase chronic myelogenous leukemia, and in 2/7 cases of clinically undifferentiated acute leukemia. Correlations were noted between reactivity with FBL -L, and DR and Fc receptor expression. Among continuous cell lines, FBL -L bound with high intensity to a majority of HL-60 and U937 cells. The less well differentiated myeloblast cell lines, KG-1, KG1a , and HL-60 blast II, exhibited less FBL -L binding than HL-60 and U937. A moderate proportion of K562 cells exhibited low level binding of FBL -L. Several lymphoblastic cell lines exhibited a pattern of low intensity binding that was distinguishable from the high intensity binding pattern of the myeloblastic lines. FBL -L reactivity of U937 was enhanced by induction of differentiation with leukocyte conditioned medium, but not dimethylsulfoxide. Such treatments induced contrasting patterns of change of HL-60 and U937 when labeled with OKM1, alpha-Mono, and HLA-DR. These studies demonstrate the application of FBL -L to analysis and quantitation of myelomonocytic leukemic differentiation.

  4. Evaluation of the stability, bioavailability, and hypersensitivity of the omega-3 derived anti-leukemic prostaglandin: Δ(12-prostaglandin J3.

    Directory of Open Access Journals (Sweden)

    Avinash K Kudva

    Full Text Available Previous studies have demonstrated the ability of an eicosapentaenoic acid (EPA-derived endogenous cyclopentenone prostaglandin (CyPG metabolite, Δ(12-PGJ3, to selectively target leukemic stem cells, but not the normal hematopoietic stems cells, in in vitro and in vivo models of chronic myelogenous leukemia (CML. Here we evaluated the stability, bioavailability, and hypersensitivity of Δ(12-PGJ3. The stability of Δ(12-PGJ3 was evaluated under simulated conditions using artificial gastric and intestinal juice. The bioavailability of Δ(12-PGJ3 in systemic circulation was demonstrated upon intraperitoneal injection into mice by LC-MS/MS. Δ(12-PGJ3 being a downstream metabolite of PGD3 was tested in vitro using primary mouse bone marrow-derived mast cells (BMMCs and in vivo mouse models for airway hypersensitivity. ZK118182, a synthetic PG analog with potent PGD2 receptor (DP-agonist activity and a drug candidate in current clinical trials, was used for toxicological comparison. Δ(12-PGJ3 was relatively more stable in simulated gastric juice than in simulated intestinal juice that followed first-order kinetics of degradation. Intraperitoneal injection into mice revealed that Δ(12-PGJ3 was bioavailable and well absorbed into systemic circulation with a Cmax of 263 µg/L at 12 h. Treatment of BMMCs with ZK118182 for 12 h resulted in increased production of histamine, while Δ(12-PGJ3 did not induce degranulation in BMMCs nor increase histamine. In addition, in vivo testing for hypersensitivity in mice showed that ZK118182 induces higher airways hyperresponsiveness when compared Δ(12-PGJ3 and/or PBS control. Based on the stability studies, our data indicates that intraperitoneal route of administration of Δ(12-PGJ3 was favorable than oral administration to achieve effective pharmacological levels in the plasma against leukemia. Δ(12-PGJ3 failed to increase histamine and IL-4 in BMMCs, which is in agreement with reduced airway

  5. Limiting-dilution analysis for the determination of leukemic cell frequencies after bone marrow decontamination with mafosfamide or merocyanine 540

    International Nuclear Information System (INIS)

    Porcellini, A.; Talevi, N.; Marchetti-Rossi, M.T.; Palazzi, M.; Manna, A.; Sparaventi, G.; Delfini, C.; Valentini, M.

    1987-01-01

    To stimulate a leukemia remission marrow, cell suspensions of normal human bone marrow were mixed with human acute lymphoblastic or myelogenous leukemic cells of the CCRF-SF, Nalm-6, and K-562 lines. The cell mixtures were incubated in vitro with mafosfamide (AZ) or with the photoreactive dye merocyanine 540 (MC-540). A quantity of 10(4) cells of the treated suspensions was dispensed into microculture plates, and graded cell numbers of the line used to contaminate the normal marrow were added. Limiting-dilution analysis was used to estimate the frequency of leukemia cells persisting after treatment with the decontaminating agents. Treatment with AZ or MC-540 produced a total elimination (ie, 6 logs or 5.3 logs respectively) of B cell acute leukemia cells (CCRF-SB), whereas nearly 1.7 logs and 2 logs of K-562 acute myelogenous blasts were still present in the cell mixtures after treatment with MC-540 and AZ, respectively. Treatment of the Nalm-6-contaminated cell mixtures with AZ resulted in 100% elimination of clonogenic cells, whereas nearly 80% decontamination was obtained with MC-540. Our results suggest that treatment with AZ could be an effective method of eliminating clonogenic tumor cells from human bone marrow. MC-540, shown by previous studies to spare sufficient pluripotential stem cells to ensure hemopoietic reconstitution in the murine model and in clinical application, has comparable effects and merits trials for possible clinical use in autologous bone marrow transplantation

  6. Limiting-dilution analysis for the determination of leukemic cell frequencies after bone marrow decontamination with mafosfamide or merocyanine 540

    Energy Technology Data Exchange (ETDEWEB)

    Porcellini, A.; Talevi, N.; Marchetti-Rossi, M.T.; Palazzi, M.; Manna, A.; Sparaventi, G.; Delfini, C.; Valentini, M.

    1987-11-01

    To stimulate a leukemia remission marrow, cell suspensions of normal human bone marrow were mixed with human acute lymphoblastic or myelogenous leukemic cells of the CCRF-SF, Nalm-6, and K-562 lines. The cell mixtures were incubated in vitro with mafosfamide (AZ) or with the photoreactive dye merocyanine 540 (MC-540). A quantity of 10(4) cells of the treated suspensions was dispensed into microculture plates, and graded cell numbers of the line used to contaminate the normal marrow were added. Limiting-dilution analysis was used to estimate the frequency of leukemia cells persisting after treatment with the decontaminating agents. Treatment with AZ or MC-540 produced a total elimination (ie, 6 logs or 5.3 logs respectively) of B cell acute leukemia cells (CCRF-SB), whereas nearly 1.7 logs and 2 logs of K-562 acute myelogenous blasts were still present in the cell mixtures after treatment with MC-540 and AZ, respectively. Treatment of the Nalm-6-contaminated cell mixtures with AZ resulted in 100% elimination of clonogenic cells, whereas nearly 80% decontamination was obtained with MC-540. Our results suggest that treatment with AZ could be an effective method of eliminating clonogenic tumor cells from human bone marrow. MC-540, shown by previous studies to spare sufficient pluripotential stem cells to ensure hemopoietic reconstitution in the murine model and in clinical application, has comparable effects and merits trials for possible clinical use in autologous bone marrow transplantation.

  7. Effects of buffers and pH on in vitro binding of 67Ga by L1210 leukemic cells

    International Nuclear Information System (INIS)

    Glickson, J.D.; Webb, J.; Gams, R.A.

    1974-01-01

    The effect of sodium nitrate and a series of buffers on in vitro 67 Ga binding to L1210 leukemic cells at pH 6.8 +- 0.2 and 37 0 at concentrations of 10 -7 to 10 -2 M has been investigated. The relative ability of these agents to inhibit cellular incorporation of 67 Ga is given. Inhibition probably results from formation of gallium(III) complexes which are either impermeable to the tumor membrane or which compete with intracellular receptor complexes. However, direct interaction of buffers with the cell membrane or with gallium(III) receptors, as well as effects of buffers on cellular metabolism, have not been excluded. A monotonic decrease in the cellular incorporation of 67 Ga occurs between pH 6.2 and 7.8 in the presence of the inert buffer, 10 -2 M morpholinopropane sulfonic acid. (U.S.)

  8. Comparison of edge detection techniques for M7 subtype Leukemic cell in terms of noise filters and threshold value

    Directory of Open Access Journals (Sweden)

    Abdul Salam Afifah Salmi

    2017-01-01

    Full Text Available This paper will focus on the study and identifying various threshold values for two commonly used edge detection techniques, which are Sobel and Canny Edge detection. The idea is to determine which values are apt in giving accurate results in identifying a particular leukemic cell. In addition, evaluating suitability of edge detectors are also essential as feature extraction of the cell depends greatly on image segmentation (edge detection. Firstly, an image of M7 subtype of Acute Myelocytic Leukemia (AML is chosen due to its diagnosing which were found lacking. Next, for an enhancement in image quality, noise filters are applied. Hence, by comparing images with no filter, median and average filter, useful information can be acquired. Each threshold value is fixed with value 0, 0.25 and 0.5. From the investigation found, without any filter, Canny with a threshold value of 0.5 yields the best result.

  9. BCR/ABL downregulates DNA-PK(CS)-dependent and upregulates backup non-homologous end joining in leukemic cells.

    Science.gov (United States)

    Poplawski, Tomasz; Blasiak, Janusz

    2010-06-01

    Non-homologous end joining (NHEJ) and homologous recombination repair (HRR) are the main mechanisms involved in the processing of DNA double strand breaks (DSBs) in humans. We showed previously that the oncogenic tyrosine kinase BCR/ABL stimulated DSBs repair by HRR. To evaluate the role of BCR/ABL in DSBs repair by NHEJ we examined the ability of leukemic BCR/ABL-expressing cell line BV173 to repair DNA damage induced by two DNA topoisomerase II inhibitors: etoposide and sobuzoxane. DNA lesions induced by sobuzoxane are repaired by a NHEJ pathway which is dependent on the catalytic subunit of protein kinase dependent on DNA (DNA-PK(CS); D-NHEJ), whereas damage evoked by etoposide are repaired by two distinct NHEJ pathways, dependent on or independent of DNA-PK(CS) (backup NHEJ, B-NHEJ). Cells incubated with STI571, a highly specific inhibitor of BCR/ABL, displayed resistance to these agents associated with an accelerated kinetics of DSBs repair, as measured by the neutral comet assay and pulsed field gel electrophoresis. However, in a functional NHEJ assay, cells preincubated with STI571 repaired DSBs induced by a restriction enzyme with a lower efficacy than without the preincubation and addition of wortmannin, a specific inhibitor of DNA-PK(CS), did not change efficacy of the NHEJ reaction. We suggest that BCR/ABL switch on B-NHEJ which is more error-prone then D-NHEJ and in such manner contribute to the increase of the genomic instability of leukemic cells.

  10. Fludarabine-mediated circumvention of cytarabine resistance is associated with fludarabine triphosphate accumulation in cytarabine-resistant leukemic cells.

    Science.gov (United States)

    Yamamoto, Shuji; Yamauchi, Takahiro; Kawai, Yasukazu; Takemura, Haruyuki; Kishi, Shinji; Yoshida, Akira; Urasaki, Yoshimasa; Iwasaki, Hiromichi; Ueda, Takanori

    2007-02-01

    The combination of cytarabine (ara-C) with fludarabine is a common approach to treating resistant acute myeloid leukemia. Success depends on a fludarabine triphosphate (F-ara-ATP)-mediated increase in the active intracellular metabolite of ara-C, ara-C 5'-triphosphate (ara-CTP). Therapy-resistant leukemia may exhibit ara-C resistance, the mechanisms of which might induce cross-resistance to fludarabine with reduced F-ara-ATP formation. The present study evaluated the effect of combining ara-C and fludarabine on ara-C-resistant leukemic cells in vitro. Two variant cell lines (R1 and R2) were 8-fold and 10-fold more ara-C resistant, respectively, than the parental HL-60 cells. Reduced deoxycytidine kinase activity was demonstrated in R1 and R2 cells, and R2 cells also showed an increase in cytosolic 5'-nucleotidase II activity. Compared with HL-60 cells, R1 and R2 cells produced smaller amounts of ara-CTP. Both variants accumulated less F-ara-ATP than HL-60 cells and showed cross-resistance to fludarabine nucleoside (F-ara-A). R2 cells, however, accumulated much smaller amounts of F-ara-ATP and were more F-ara-A resistant than R1 cells. In HL-60 and R1 cells, F-ara-A pretreatment followed by ara-C incubation produced F-ara-ATP concentrations sufficient for augmenting ara-CTP production, thereby enhancing ara-C cytotoxicity. No potentiation was observed in R2 cells. Nucleotidase might preferentially degrade F-ara-A monophosphate over ara-C monophosphate, leading to reduced F-ara-ATP production and thereby compromising the F-ara-A-mediated potentiation of ara-C cytotoxicity in R2 cells. Thus, F-ara-A-mediated enhancement of ara-C cytotoxicity depended on F-ara-ATP accumulation in ara-C-resistant leukemic cells but ultimately was associated with the mechanism of ara-C resistance.

  11. Effect of pentoxifylline on P-glycoprotein mediated vincristine resistance of L1210 mouse leukemic cell line

    International Nuclear Information System (INIS)

    Breier, A.; Uhrik, B.; Barancik, M.; Stefankova, Z.; Tribulova, N.

    1994-01-01

    Effect of pentoxifylline (PTX) on vincristine (VCR) resistance of multidrug resistant L1210/VCR mouse leukemic cell line was studied. Reversal effect of PTX (in concentration 50-150 mg dm -3 ) on vincristine resistance, i.e. potentiation of vincristine cytotoxicity on L1210/VCR cells by PTX was found. PTX alone in the above concentration did not exert any significant effect on sensitive or resistant cell lines in the absence of vincristine. Resistance of L1210/VCR cell line was found previously to be accompanied with overexpression of drug transporting P-glycoprotein. Indeed, lower level of 3 H-vincristine accumulation by resistant L1210/VCR cell line in comparison with sensitive L1210 cell line was observed. Accumulation of 3 H-vincristine by L1210/VCR cell line was significantly increased in the presence of PTX. PTX in the same condition did not exert any considerable effect on accumulation of 3 H-vincristine by nonresistant L1210 cells. Observable morphological damage was observed in 1210/VCR cells cultivated in medium containing vincristine (0.2 mg dm -3 ) and pentoxifylline (100 mg dm -3 ) in comparison with the non-damaged cells in the presence of vincristine or pentoxifylline alone. The results obtained indicate that pentoxifylline may be considered as a reversal agent in multidrug resistance. (author)

  12. Lack of correlation between immunologic markers and cell surface ultrastructure in the leukemic phase of lymphoproliferative diseases

    Energy Technology Data Exchange (ETDEWEB)

    Golomb, Harvey M.; Simon, Deberah

    1977-01-01

    In a prospective study of malignant cells from 13 patients with the leukemic phase of lymphoproliferative diseases, we wished to determine whether any correlation between the immunologic markers and the cell surface ultrastructure. Five patients had chronic lymphocytic leukemia, four had malignant lymphomas, poorly differentiated lymphocytic type, two had the Sezary syndrome, and one each had acute prolymphocytic leukemia and acute lymphocytic leukemia. Cell separation and isolation was done at room temperature for all specimens. Immunologic markers tested for were surface immunoglobins, a B-cell property, and E-rosettes, a T-cell property. Three patients had T-cell diseases, 6 had B-cell diseases, and 4 were classified as ''null.'' All but one patient had moderate to large numbers of microvilli on their malignant cells. The single exception had a typical B-cell form of chronic lymphocytic leukemia. There appears to be no correlation between immunologic markers and cell surface ultrastructure; therefore, SEM appears not to be valuable in the diagnosis or classification of immunologic sub-types of certain lymphoproliferative diseases.

  13. Delayed expression of apoptosis in X-irradiated human leukemic MOLT-4 cells transfected with mutant p53

    International Nuclear Information System (INIS)

    Nakano, Hisako; Yonekawa, Hiromichi; Shinohara, Kunio

    2003-01-01

    The effects of X-rays on cell survival, apoptosis, and long-term response in the development of cell death as measured by the dye exclusion test were studied in human leukemic MOLT-4 cells (p53 wild-type) stably transfected with a mutant p53 cDNA expression vector. Cell survival, as determined from colony-forming ability, was increased in an expression level dependent manner, but the increase was partial even with the highest-expressing clone (B3). This contrasts with the prior observation that cell death and apoptosis in B3 are completely inhibited at 24 h after irradiation with 1.8 Gy of X-rays. The examination of B3 cells incubated for longer than 24 h after X-irradiation showed a delay in the induction of cell death and apoptosis. Western blot analysis revealed that the time required to reach the highest level of wild-type p53 protein in B3 was longer than the time in MOLT-4 and that the p53 may be stabilized by the phosphorylation at Ser-15. These results suggest that the introduction of mutant p53 into MOLT-4 merely delays the development of apoptosis, during which the cells could repair the damage induced by X-rays, and results in the partial increase in cell survival. (author)

  14. House dust mite induces expression of intercellular adhesion molecule-1 in EoL-1 human eosinophilic leukemic cells.

    Science.gov (United States)

    Kwon, Byoung Chul; Sohn, Myung Hyun; Kim, Kyung Won; Kim, Eun Soo; Kim, Kyu-Earn; Shin, Myeong Heon

    2007-10-01

    The house dust mite (HDM) is considered to be the most common indoor allergen associated with bronchial asthma. In this study, we investigated whether crude extract of the HDM Dermatophagoides farinae could activate human eosinophilic leukemic cells (EoL-1) to induce upregulation of cell-surface adhesion molecules. When EoL-1 cells were incubated with D. farinae extract, expression of intercellular adhesion molecule-1 (ICAM-1) significantly increased on the cell surfaces compared to cells incubated with medium alone. In contrast, surface expression of CD11b and CD49d in EoL-1 cells was not affected by D. farinae extract. In addition, pretreatment of cells with NF-kappaB inhibitor (MG-132) or JNK inhibitor (SP600125) significantly inhibited ICAM-1 expression promoted by HDM extract. However, neither p38 MAP kinase inhibitor nor MEK inhibitor prevented HDM-induced ICAM-1 expression in EoL-1 cells. These results suggest that crude extract of D. farinae induces ICAM-1 expression in EoL-1 cells through signaling pathways involving both NF-kappaB and JNK.

  15. Effects of in vitro Brevetoxin Exposure on Apoptosis and Cellular Metabolism in a Leukemic T Cell Line (Jurkat

    Directory of Open Access Journals (Sweden)

    John W. Sleasman

    2008-06-01

    Full Text Available Harmful algal blooms (HABs of the toxic dinoflagellate, Karenia brevis, produce red tide toxins, or brevetoxins. Significant health effects associated with red tide toxin exposure have been reported in sea life and in humans, with brevetoxins documented within immune cells from many species. The objective of this research was to investigate potential immunotoxic effects of brevetoxins using a leukemic T cell line (Jurkat as an in vitro model system. Viability, cell proliferation, and apoptosis assays were conducted using brevetoxin congeners PbTx-2, PbTx-3, and PbTx-6. The effects of in vitro brevetoxin exposure on cell viability and cellular metabolism or proliferation were determined using trypan blue and MTT (1-(4,5-dimethylthiazol-2-yl-3,5- diphenylformazan, respectively. Using MTT, cellular metabolic activity was decreased in Jurkat cells exposed to 5 - 10 μg/ml PbTx-2 or PbTx-6. After 3 h, no significant effects on cell viability were observed with any toxin congener in concentrations up to 10 μg/ml. Viability decreased dramatically after 24 h in cells treated with PbTx-2 or -6. Apoptosis, as measured by caspase-3 activity, was significantly increased in cells exposed to PbTx-2 or PbTx-6. In summary, brevetoxin congeners varied in effects on Jurkat cells, with PbTx-2 and PbTx-6 eliciting greater cellular effects compared to PbTx-3.

  16. Glucocorticoid receptors on leukemic cells as evidenced by dexamethasone-induced cytolysis and /sup 3/H-dexamethasone binding

    Energy Technology Data Exchange (ETDEWEB)

    Thraenhardt, H; Haefer, R; Zintl, F

    1987-01-01

    The presence of glucocorticoid receptors on the leukemic cells of 33 patients affected with acute lymphatic leukemia (ALL) and 6 patients affected with acute myeloic leukemia (AML) was investigated by dexamethasone-induced cytolysis and (/sup 3/H)-dexamethasone binding. The tests undertaken proved that after 20 hours of incubation 9 of 26 non-T-non-B-ALL (c-ALL and unclassified ALL) and 2 of AML were lysed with dexamethasone; blood lymphocytes and bone marrow leukocytes of healthy donors, however, were not affected. Non-T-non-B-ALL and AML were able to bind essentially more (/sup 3/H)-dexamethasone than T-ALL. There existed no correlation between dexamethasone binding and dexamethasone-induced cytolysis.

  17. DNA repair and DNA synthesis in leukemic and virus infected cells

    International Nuclear Information System (INIS)

    Tuschl, H.; Altmann, H.; Kovac, R.; Topaloglou, A.; Stacher, A.; Fanta, D.

    1978-09-01

    Autoradiographic determinations of unscheduled DNA synthesis in peripheral lymphocytes of leukemic patients showed strongly different results according to various types of disease of different forms of therapy, respectively. Similar investigations performed with lymphocytes of Herpes simplex infected persons during symptom-free intervals revealed imbalances of the repair system caused by virus infection. BND cellulose chromatography and measurement of 3 H-thymidine incorporation into single- and double stranded DNA fractions showed an increase in velocity of the rejoining process, but a decrease in total incorporation. Because of these results and the demonstration of the supercoiled structure of DNA it is suggested that virusinfections cause a faster rejoining of gaps, but at the same time leave a number of failures within DNA unrecognized. (author)

  18. Characterization of activating mutations of NOTCH3 in T-cell acute lymphoblastic leukemia and anti-leukemic activity of NOTCH3 inhibitory antibodies.

    Science.gov (United States)

    Bernasconi-Elias, P; Hu, T; Jenkins, D; Firestone, B; Gans, S; Kurth, E; Capodieci, P; Deplazes-Lauber, J; Petropoulos, K; Thiel, P; Ponsel, D; Hee Choi, S; LeMotte, P; London, A; Goetcshkes, M; Nolin, E; Jones, M D; Slocum, K; Kluk, M J; Weinstock, D M; Christodoulou, A; Weinberg, O; Jaehrling, J; Ettenberg, S A; Buckler, A; Blacklow, S C; Aster, J C; Fryer, C J

    2016-11-24

    Notch receptors have been implicated as oncogenic drivers in several cancers, the most notable example being NOTCH1 in T-cell acute lymphoblastic leukemia (T-ALL). To characterize the role of activated NOTCH3 in cancer, we generated an antibody that detects the neo-epitope created upon gamma-secretase cleavage of NOTCH3 to release its intracellular domain (ICD3), and sequenced the negative regulatory region (NRR) and PEST (proline, glutamate, serine, threonine) domain coding regions of NOTCH3 in a panel of cell lines. We also characterize NOTCH3 tumor-associated mutations that result in activation of signaling and report new inhibitory antibodies. We determined the structural basis for receptor inhibition by obtaining the first co-crystal structure of a NOTCH3 antibody with the NRR protein and defined two distinct epitopes for NRR antibodies. The antibodies exhibit potent anti-leukemic activity in cell lines and tumor xenografts harboring NOTCH3 activating mutations. Screening of primary T-ALL samples reveals that 2 of 40 tumors examined show active NOTCH3 signaling. We also identified evidence of NOTCH3 activation in 12 of 24 patient-derived orthotopic xenograft models, 2 of which exhibit activation of NOTCH3 without activation of NOTCH1. Our studies provide additional insights into NOTCH3 activation and offer a path forward for identification of cancers that are likely to respond to therapy with NOTCH3 selective inhibitory antibodies.

  19. Leukemic transformation of donor spleen cells following their transplantation into supralethally irradiated mice with pre-existing viral leukemia. [X Radiation

    Energy Technology Data Exchange (ETDEWEB)

    Kuhnert, P M; OKunewick, J P; Erhard, P

    1974-01-01

    Fialkow et al. previously reported leukemia induction in donor-type cells after treating patients for acute lymphoblastic leukemia with total-body irradiation and hematopoietic cell transplantation. Utilizing a murine model and paralleling their treatment protocol, we have documented that induction of leukemia can occur in normal donor cells transplanted into Rauscher viral leukemic mice at 0, 1 and 2 days after irradiation. The induction of leukemia in the grafted cells was verified by: the occurrence of splenomegaly; and secondary spleen cell transplants, whereby the secondary donors were transplanted mice still alive at 30 days and the secondary recipients were normal unirradiated mice. The spleen weights of the grafted leukemic mice were found to be significantly greater than those of the controls and all secondary recipients that received spleen cells from the primary grafted leukemic mice also died of leukemia. Verification that the regenerating hematopoietic tissue was from donor (males) and not host source (females) was accomplished by spleen chromosome preparations taken from randomly selected mice at 14 and at 30 days after cell transplantation. In these preparations, the Y chromosome was clearly distinguishable on the basis of size, shape, and differential staining. The data indicate that induction of leukemia after whole-body irradiation and hematopoietic cell transplantation can occur in immunologically matched donor cells when a viral agent is present and that the incidence of this induction is not affected by a time delay between irradiation and transplant.

  20. PENGARUH EKSTRAK JAMU TERHADAP AKTIVITAS SEL NATURAL KILLER DALAM MELISIS ALUR SEL LEUKIMIA (K-562 SECARA IN VITRO [The Effects of Commercial “Jamu” Extracts on Natural Killer Cell Activity in Lysing Leukemic Cell Line (K-562 in vitro

    Directory of Open Access Journals (Sweden)

    Elisa Veronica D.C. 2

    2002-04-01

    Full Text Available Natural killer (NK cell consitutes white blood cells which specifically functions in lysing tumor and virus invected cells. In this research, a commercial “Jamu” was tested to observe its effect on NK cells activity against leukemic cell lines (K562 in vitro. Jamu was extracted with hot water, diluted and added into cell cultures consisted of a mixture of human peripheric limphocyte cells, as the source of the effector NK cells, and K562 cell line i.e., the target cells which were cell line derived from human leukemia and had been labelled with H3-thymidine. The mixture of the cells were made by culturing the two cells at the ratio of 50:1 and 100 : 1, respectively. The results showed that lysing activity of NK cells in the presence of “Jamu” water extract measured as lysing percentage and lysing index increased only slightly, which were not statiscally significant. It should be considered that the test used in this research represents only a part of the lysing mechanism by NK cells against the target cells. An in vivo test for a period of time will be recessary to elucidate ffurther this NK cell activity.

  1. Novel quinazolinone MJ-29 triggers endoplasmic reticulum stress and intrinsic apoptosis in murine leukemia WEHI-3 cells and inhibits leukemic mice.

    Directory of Open Access Journals (Sweden)

    Chi-Cheng Lu

    Full Text Available The present study was to explore the biological responses of the newly compound, MJ-29 in murine myelomonocytic leukemia WEHI-3 cells in vitro and in vivo fates. We focused on the in vitro effects of MJ-29 on ER stress and mitochondria-dependent apoptotic death in WEHI-3 cells, and to hypothesize that MJ-29 might fully impair the orthotopic leukemic mice. Our results indicated that a concentration-dependent decrease of cell viability was shown in MJ-29-treated cells. DNA content was examined utilizing flow cytometry, whereas apoptotic populations were determined using annexin V/PI, DAPI staining and TUNEL assay. Increasing vital factors of mitochondrial dysfunction by MJ-29 were further investigated. Thus, MJ-29-provaked apoptosis of WEHI-3 cells is mediated through the intrinsic pathway. Importantly, intracellular Ca(2+ release and ER stress-associated signaling also contributed to MJ-29-triggered cell apoptosis. We found that MJ-29 stimulated the protein levels of calpain 1, CHOP and p-eIF2α pathways in WEHI-3 cells. In in vivo experiments, intraperitoneal administration of MJ-29 significantly improved the total survival rate, enhanced body weight and attenuated enlarged spleen and liver tissues in leukemic mice. The infiltration of immature myeloblastic cells into splenic red pulp was reduced in MJ-29-treated leukemic mice. Moreover, MJ-29 increased the differentiations of T and B cells but decreased that of macrophages and monocytes. Additionally, MJ-29-stimulated immune responses might be involved in anti-leukemic activity in vivo. Based on these observations, MJ-29 suppresses WEHI-3 cells in vitro and in vivo, and it is proposed that this potent and selective agent could be a new chemotherapeutic candidate for anti-leukemia in the future.

  2. Documentation of normal and leukemic myelopoietic progenitor cells with high-resolution phase-contrast time-lapse cinematography.

    Science.gov (United States)

    Boll, I T

    2001-08-01

    The high-resolution phase-contrast, time-lapse cinematography using oil immersion lenses and 16-mm film demonstrates the kinetic cell events as maturation, locomotion, mitosis, and apoptosis of cells cultivated at 37 degrees C for up to 10 days. 0.5 v/v frozen-thawed sera with presumably high cytokine concentrations were added to the plasma or agar clot. Vital progenitor cells from human bone marrow and blood have a large, bright, unstructured nucleus with a large nucleolus and a narrow rim of cytoplasm (nuclear/cytoplasmic volume ratio = 0.7). Their nuclei are 6-14 micrometer in diameter and double their volume within 8 h. Many (70%) move at a mean speed of 2 micrometer/min, and many (30%) multiply with alpha-2alpha mitoses, generating progenitor cell families. Various disturbances during the course of mitosis lead to the formation of polyploid cells, thereby yielding the megakaryocytic cell line. Some of the progenitor cells undergo asymmetric alpha-alphan mitoses: One of the two initially identical daughter cells remains a progenitor cell in the morphological sense, whereas the other daughter cell - depending on the size of its mother cell - matures in the same culture medium to form a granulocytopoietic, monocytopoietic or erythrocytopoietic cell line. - In acute myeloid leukemias (AML), the blasts and their nuclei are slightly larger than the corresponding progenitor cells and move faster (5 micrometer/min). Symmetric alpha-2alpha mitoses permit unlimited multiplication of the leukemic blasts if contact with cytotoxic lymphocytes does not render them apoptotic. This results in more stromal cells than normal. Granulocytopenia, monocytopenia, and anemia occur due to the genetic impairment of signaling control for asymmetric alpha-alphan mitoses, and thrombocytopenia occurs due to the reduction in polyploidization. Copyright 2001 S. Karger GmbH, Freiburg

  3. Involvement of CD147 on multidrug resistance through the regulation of P-glycoprotein expression in K562/ADR leukemic cell line

    Directory of Open Access Journals (Sweden)

    Aoranit Somno

    2016-01-01

    Full Text Available The relationship between P-gp and CD147 in the regulation of MDR in leukemic cells has not been reported. This study aimed to investigate the correlation between CD147 and P-gp in the regulation of drug resistance in the K562/ADR leukemic cell line. The results showed that drug-resistant K562/ADR cells expressed significantly higher P-gp and CD147 levels than drug-free K562/ADR cells. To determine the regulatory effect of CD147 on P-gp expression, anti-CD147 antibody MEM-M6/6 significantly decreased P-gp and CD147 mRNA and protein levels. This is the first report to show that CD147 mediates MDR in leukemia through the regulation of P-gp expression.

  4. F-box protein FBXL2 targets cyclin D2 for ubiquitination and degradation to inhibit leukemic cell proliferation

    Science.gov (United States)

    Chen, Bill B.; Glasser, Jennifer R.; Coon, Tiffany A.; Zou, Chunbin; Miller, Hannah L.; Fenton, Moon; McDyer, John F.; Boyiadzis, Michael

    2012-01-01

    Hematologic maligancies exhibit a growth advantage by up-regulation of components within the molecular apparatus involved in cell-cycle progression. The SCF (Skip-Cullin1-F-box protein) E3 ligase family provides homeostatic feedback control of cell division by mediating ubiquitination and degradation of cell-cycle proteins. By screening several previously undescribed E3 ligase components, we describe the behavior of a relatively new SCF subunit, termed FBXL2, that ubiquitinates and destabilizes cyclin D2 protein leading to G0 phase arrest and apoptosis in leukemic and B-lymphoblastoid cell lines. FBXL2 expression was strongly suppressed, and yet cyclin D2 protein levels were robustly expressed in acute myelogenous leukemia (AML) and acute lymphoblastic leukemia (ALL) patient samples. Depletion of endogenous FBXL2 stabilized cyclin D2 levels, whereas ectopically expressed FBXL2 decreased cyclin D2 lifespan. FBXL2 did not bind a phosphodegron within its substrate, which is typical of other F-box proteins, but uniquely targeted a calmodulin-binding signature within cyclin D2 to facilitate its polyubiquitination. Calmodulin competes with the F-box protein for access to this motif where it bound and protected cyclin D2 from FBXL2. Calmodulin reversed FBXL2-induced G0 phase arrest and attenuated FBXL2-induced apoptosis of lymphoblastoid cells. These results suggest an antiproliferative effect of SCFFBXL2 in lymphoproliferative malignancies. PMID:22323446

  5. Oxidative damage of U937 human leukemic cells caused by hydroxyl radical results in singlet oxygen formation.

    Directory of Open Access Journals (Sweden)

    Marek Rác

    Full Text Available The exposure of human cells to oxidative stress leads to the oxidation of biomolecules such as lipids, proteins and nuclei acids. In this study, the oxidation of lipids, proteins and DNA was studied after the addition of hydrogen peroxide and Fenton reagent to cell suspension containing human leukemic monocyte lymphoma cell line U937. EPR spin-trapping data showed that the addition of hydrogen peroxide to the cell suspension formed hydroxyl radical via Fenton reaction mediated by endogenous metals. The malondialdehyde HPLC analysis showed no lipid peroxidation after the addition of hydrogen peroxide, whereas the Fenton reagent caused significant lipid peroxidation. The formation of protein carbonyls monitored by dot blot immunoassay and the DNA fragmentation measured by comet assay occurred after the addition of both hydrogen peroxide and Fenton reagent. Oxidative damage of biomolecules leads to the formation of singlet oxygen as conformed by EPR spin-trapping spectroscopy and the green fluorescence of singlet oxygen sensor green detected by confocal laser scanning microscopy. It is proposed here that singlet oxygen is formed by the decomposition of high-energy intermediates such as dioxetane or tetroxide formed by oxidative damage of biomolecules.

  6. Piperlongumine induces apoptosis and autophagy in leukemic cells through targeting the PI3K/Akt/mTOR and p38 signaling pathways.

    Science.gov (United States)

    Wang, Hongfei; Wang, Yongqiang; Gao, Hongmei; Wang, Bing; Dou, Lin; Li, Yin

    2018-02-01

    Piperlongumine is an alkaloid compound extracted from Piper longum L. It is a chemical substance with various pharmacological effects and medicinal value, including anti-tumor, lipid metabolism regulatory, antiplatelet aggregation and analgesic properties. The present study aimed to understand whether piperlongumine induces the apoptosis and autophagy of leukemic cells, and to identify the mechanism involved. Cell viability and autophagy were detected using MTT, phenazine methyl sulfate and trypan blue exclusion assays. The apoptosis rate was calculated using flow cytometry. The protein expression levels of microtubule-associated protein 1A/1B-light chain 3, Akt and mechanistic target of rapamycin (mTOR) were measured using western blotting. The cell growth of leukemic cells was completely inhibited following treatment with piperlongumine, and marked apoptosis was also induced. Dead cells as a result of autophagy were stained using immunofluorescence and observed under a light microscope. Phosphoinositide 3-kinase (PI3K)/Akt/mTOR signaling was suppressed by treatment with piperlongumine, while p38 signaling and caspase-3 activity were induced by treatment with piperlongumine. It was concluded that piperlongumine induces apoptosis and autophagy in leukemic cells through targeting the PI3K/Akt/mTOR and p38 signaling pathways.

  7. Transfer RNA species in human lymphocytes stimulated by mitogens and in leukemic cells. [/sup 3/H, /sup 14/C, /sup 32/P tracer techniques

    Energy Technology Data Exchange (ETDEWEB)

    Griffin, G.D.; Yang, W.K.; Novelli, G.D.

    1976-01-01

    Transfer ribonucleic acid (tRNA) profiles in human lymphocytes stimulated by various mitogens have been compared with profiles from nonstimulated cells and from leukemic cells using reversed-phase chromatography. Comparisons of (/sup 3/H)- or (/sup 11/C)uridine- or (/sup 32/P)phosphate-labeled tRNAs showed that the greatest changes in tRNA composition upon phytohemagglutinin (PHA) stimulation occurred in the first 8 h after mitogen addition. Stimulation of lymphocytes by pokeweed mitogen, anti-human immunoglobulin, or bacterial lipopolysaccharide resulted in tRNA species which showed distinct differences from each other and also from the tRNAs produced by phytohemagglutinin stimulation. Leukemic lymphocyte tRNAs showed the most extensive differences in profile when compared with chromatograms from non-neoplastic cells stimulated by a variety of mitogens. Specific isoaccepting species of tyrosyl-, aspartyl-, and phenylalanyl-tRNAs were also compared in PHA-stimulated and resting lymphocytes and no differences were found. When these same species were studied in leukemic cells, tyrosyl-tRNA profiles were shifted to elute at a lower salt concentration, while the aspartyl-tRNA profile showed a new peak not present in noncancerous cells.

  8. Genomic Profiling of a Human Leukemic Monocytic Cell-Line (THP-1 Exposed to Alpha Particle Radiation

    Directory of Open Access Journals (Sweden)

    Vinita Chauhan

    2012-01-01

    Full Text Available This study examined alpha (α- particle radiation effects on global changes in gene expression in human leukemic monocytic cells (THP-1 for the purposes of mining for candidate biomarkers that could be used for the development of a biological assessment tool. THP-1 cells were exposed to α-particle radiation at a dose range of 0 to 1.5 Gy. Twenty-four hours and three days after exposure gene expression was monitored using microarray technology. A total of 16 genes were dose responsive and classified as early onset due to their expression 24 h after exposure. Forty-eight transcripts were dose responsive and classified as late-onset as they were expressed 72 h after exposure. Among these genes, 6 genes were time and dose responsive and validated further using alternate technology. These transcripts were upregulated and associated with biological processes related to immune function, organelle stability and cell signalling/communication. This panel of genes merits further validation to determine if they are strong candidate biomarkers indicative of α-particle exposure.

  9. The variable chemotherapeutic response of Malabaricone-A in leukemic and solid tumor cell lines depends on the degree of redox imbalance.

    Science.gov (United States)

    Manna, Alak; De Sarkar, Sritama; De, Soumita; Bauri, Ajay K; Chattopadhyay, Subrata; Chatterjee, Mitali

    2015-07-15

    The 'two-faced' character of reactive oxygen species (ROS) plays an important role in cancer biology by acting as secondary messengers in intracellular signaling cascades, enhancing cell proliferation and survival, thereby sustaining the oncogenic phenotype. Conversely, enhanced generation of ROS can trigger an oxidative assault leading to a redox imbalance translating into an apoptotic cell death. Intrinsically, cancer cells have higher basal levels of ROS which if supplemented by additional oxidative insult by pro-oxidants can be cytotoxic, an example being Malabaricone-A (MAL-A). MAL-A is a plant derived diarylnonanoid, purified from fruit rind of the plant Myristica malabarica whose anti-cancer activity has been demonstrated in leukemic cell lines, the modality of cell death being apoptosis. This study aimed to compare the degree of effectiveness of MAL-A in leukemic vs. solid tumor cell lines. The cytotoxicity of MAL-A was evaluated by the MTS-PMS cell viability assay in leukemic cell lines (MOLT3, K562 and HL-60) and compared with solid tumor cell lines (MCF7, A549 and HepG2); further studies then proceeded with MOLT3 vs. MCF7 and A549. The contribution of redox imbalance in MAL-A induced cytotoxicity was confirmed by pre-incubating cells with an antioxidant, N-acetyl-L-cysteine (NAC) or a thiol depletor, buthionine sulfoximine (BSO). MAL-A induced redox imbalance was quantitated by flow cytometry, by measuring the generation of ROS and levels of non protein thiols using dichlorofluorescein diacetate (CM-H2DCFDA) and 5-chloromethylfluorescein diacetate (CMFDA) respectively. The activities of glutathione peroxidase (GPx), superoxide dismutase, catalase (CAT), NAD(P)H dehydrogenase (quinone 1) NQO1 and glutathione-S-transferase GST were measured spectrophotometrically. The mitochondrial involvement of MAL-A induced cell death was measured by evaluation of cardiolipin peroxidation using 10-N-nonyl acridine orange (NAO), transition pore activity with calcein

  10. Modulation of P-glycoprotein-mediated multidrug resistance in K562 leukemic cells by indole-3-carbinol

    International Nuclear Information System (INIS)

    Arora, Annu; Seth, Kavita; Kalra, Neetu; Shukla, Yogeshwer

    2005-01-01

    Resistance to chemotherapeutic drugs is one of the major problems in the treatment of cancer. P-glycoprotein (P-gp) encoded by the mdr gene is a highly conserved protein, acts as a multidrug transporter, and has a major role in multiple drug resistance (MDR). Targeting of P-gp by naturally occurring compounds is an effective strategy to overcome MDR. Indole-3-carbinol (I3C), a glucosinolates present in cruciferous vegetables, is a promising chemopreventive agent as it is reported to possess antimutagenic, antitumorigenic, and antiestrogenic properties in experimental studies. In the present investigation, the potential of I3C to modulate P-gp expression was evaluated in vinblastine (VBL)-resistant K562 human leukemic cells. The resistant K562 cells (K562/R10) were found to be cross-resistant to vincristine (VCR), doxorubicin (DXR), and other antineoplastic agents. I3C at a nontoxic dose (10 x 10 -3 M) enhanced the cytotoxic effects of VBL time dependently in VBL-resistant human leukemia (K562/R10) cells but had no effect on parent-sensitive cells (K562/S). The Western blot analysis of K 562/R 10 cells showed that I3C downregulates the induced levels of P-gp in resistant cells near to normal levels. The quantitation of immunocytochemically stained K562/R10 cells showed 24%, 48%, and 80% decrease in the levels of P-gp by I3C for 24, 48, and 72 h of incubation. The above features thus indicate that I3C could be used as a novel modulator of P-gp-mediated multidrug resistance in vitro and may be effective as a dietary adjuvant in the treatment of MDR cancers

  11. Correlation of chromosome patterns in leukemic cells of patients with exposure to chemicals and/or radiation

    International Nuclear Information System (INIS)

    Rowley, J.D.

    1989-10-01

    We have identified two new recurring translocations involving chromosome 5; one is a 3;5 translocation and the other involves a rearrangement between chromosomes 5 and 7. The first is t(3;5)(q25.1;q35). We studied five patients with AML and a t(3;5) in their leukemic cells. At diagnosis, four of the patients had a t(3;5) as their sole karyotypic anomaly; the remaining patient had additional structural and numerical abnormalities. Careful cytogenetic analysis indicated that the breakpoints of this rearrangement were 3q25.1 and 5q34, in contrast to the various breakpoints reported in earlier studies (3q21 to 3q25 and 5q31 to 5q35). The karyotypic, morphologic, and clinical characteristics of this group, as well as those of 15 previously reported patients with the t(3;5), were compared to identify any features that might warrant consideration of this anomaly as a specific syndrome. The median age of the group, 37 years, as younger than that of all patients with AML, 49 years. A preceding myelodysplastic syndrome was observed in three patients. We have no information regarding the occupation of most of these patients. Except for acute promyelocytic leukemia, each morphologic subtype occurred in these patients; however, the frequency of erythroleukemia (M6) was much greater than expected. 11 refs., 2 figs., 5 tabs

  12. Predicting radiation effects on the development of leukemic stem cells based on studies of leukemias induced by high- and low-dose-rate radiation

    International Nuclear Information System (INIS)

    Hirouchi, Tokuhisa

    2012-01-01

    One of the most important causes of radiation-induced cancers, particularly leukemia, is gene mutations resulting from single and double strand breaks in the DNA. Tanaka et al. (2003) reported life shortening in specific pathogen free male and female B6C3F1 mice continuously exposed to γ rays at a low dose rate of 20 mGy/22 h/d for 400 days from 8 weeks of age. Early death due to cancer, mostly malignant lymphomas, was observed in both sexes. A significant increase in the incidence of myeloid leukemia, resulting in early death, was also reported in males. It is expected however, that at 20 mGy/22 h/d, which is equivalent to a dose of 15 μGy/min, DNA strand breaks induced in these cells are repaired soon after they occur. Murine leukemias induced by high-dose-rate radiation were also found in males, and 80% of the mice with leukemia had hemizygous deletions in chromosome 2 around the PU.1 gene and they appeared to be derived from DNA strand breaks. Majority of these leukemia showing hemizygous deletions in chromosome 2 revealed point mutations in the remaining alleles resulting in PU.1 inactivation, which was reported to be related to leukemogenesis. These point mutations are assumed to be independent of DNA strand breaks that occur immediately after irradiation, as they appear at later time after irradiation. This review discusses the effect of radiation-induced DNA strand breaks and also mutagenesis induced independently of DNA strand breaks in hematopoietic cells contributing to the development of the first leukemic stem cell. (author)

  13. Towards The Generation of Functionalized Magnetic Nanowires to Target Leukemic Cells

    KAUST Repository

    Alsharif, Nouf

    2016-01-01

    . In addition the NWs can be coated and functionalized to target cells of interest and, upon exposure to an alternating magnetic field, have been shown to induce cell death on several types of adherent cells, including several cancer cell types. For suspension

  14. Leukemic meningitis involving the cauda equina: a case report

    International Nuclear Information System (INIS)

    Lee, Dong Hyun; Kim, Ho Kyun; Lee, Young Hwan

    2008-01-01

    The CNS involvement by leukemia may either be meningeal or parenchymal, although meningeal infiltration of leukemic cells, known as leukemic meningitis is more common. We report a case of leukemic meningitis involving the cauda equina in a patient with an acute lymphoblastic crisis which transformed from the chronic phase of chronic myeloid leukemia. An MR image revealed diffuse enlargement and peripheral ring enhancement of the nerve roots of the cauda equina

  15. Leukemic meningitis involving the cauda equina: a case report

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Dong Hyun; Kim, Ho Kyun; Lee, Young Hwan [School of Medicine, Catholic University of Daegu, Daegu (Korea, Republic of)

    2008-07-15

    The CNS involvement by leukemia may either be meningeal or parenchymal, although meningeal infiltration of leukemic cells, known as leukemic meningitis is more common. We report a case of leukemic meningitis involving the cauda equina in a patient with an acute lymphoblastic crisis which transformed from the chronic phase of chronic myeloid leukemia. An MR image revealed diffuse enlargement and peripheral ring enhancement of the nerve roots of the cauda equina.

  16. Structural features of nucleoli in blood, leukemic, lymphoma and myeloma cells

    Directory of Open Access Journals (Sweden)

    K Smetana

    2010-01-01

    Full Text Available At present, it seems clear that the nucleolus is multifunctional and represents one of the key cell organelles that participate directly or indirectly in cell resting, proliferation, differentiation and maturation states, and possibly also in programmed cell death. Thus, the morphology and cytochemistry of nucleoli may represent a very useful tool not only for the evaluation of nucleolar biosynthetic activities but also for the evaluation of various cell states under physiological, experimental and pathological conditions.

  17. [Effects of PCI-32765 and Dasatinib on the Acute Lymphoblastic Leukemic Cells and Their Mechanisms].

    Science.gov (United States)

    Deng, Yuan; Tao, Shan-Dong; Zhang, Xin; Ma, Jing-Jing; He, Zheng-Mei; Chen, Yue; Deng, Zhi-Kui; Yu, Liang

    2017-02-01

    To investigate the effects of Btk inhibitor (PCI-32765) and BCR-ABL tyrosine kinase inhibitor (Dasatinib) on proliferation and apoptosis of acute lymphoblastic leukemia (ALL) cell lines (Sup-B15, RS4;11) and the possible mechanism. RS4;11 and Sup-B15 cells were treated with PCI-32765 and Dasatinib, the cell proliferation and apoptosis were detected by CCK-8, the Btk and other apoptotic proteins were detected by Western blot. PCI-32765 could inhibit the proliferation of RS4;11 and Sup-B15 cells in a dose-dependent manner, Sup-B15 cells were more sensitive to PCI-32765 than RS4;11 cells, their IC 50 were 3 µmol/L and 8 µmol/L respectively, the difference between them was statistically significant (PPCI-32765(PPCI-32765 or Dasatinib alone group and the combination group at the different time-point (8, 12, 24, 36, 48 and 72 h), the 2 drugs showed a synergistic effect on cells in a time-dependent manner. After being treated with PCI-32765 and Dasatinib, the RS4;11 and Sup-B15 cells showed that cell shrinkage, increase of cytoplasmic density, nuclear pyknosis, deviation and karyorrhexis, and increase of the apoptotic cells in the combination group, while the promotive effect of low dosage dasatinib on apoptosis of RS4;11 cells was not strong. PCI-32765 and Dasatinib could decrease the expression and activity of BCR-ABL, Btk, Lyn, Src in Sup-B15 and RS4;11 cells. PCI-32765 or Dasatinib can inhibit the proliferation and induce the apoptosis of Sup-B15 and RS4;11 cells, PCI-32765 and Dasatinib displayed the synergistic effects. The possible mechanism may be related with the blocking of B cell receptor(BCR) signal pathway, thereby inhibiting the cell proliferation and promoting the cell apoptosis.

  18. Apoptosis induction by Maackia amurensis agglutinin in childhood acute lymphoblastic leukemic cells

    DEFF Research Database (Denmark)

    Kapoor, Sarika; Marwaha, Ram; Majumdar, Siddhartha

    2007-01-01

    acute lymphoblastic leukemia (ALL) as compared to cells from children with non-hematological disorders ("Controls"). MAA recognized a 66 kDa sialoglycoprotein present in membrane fraction of ALL cells. Moreover, MAA induced apoptosis in ALL cells was found to be reduced significantly in presence of GM2...

  19. STAT5-mediated self-renewal of normal hematopoietic and leukemic stem cells

    NARCIS (Netherlands)

    Schepers, Hein; Wierenga, Albertus T. J.; Vellenga, Edo; Schuringa, Jan Jacob

    2012-01-01

    The level of transcription factor activity critically regulates cell fate decisions such as hematopoietic stem cell self-renewal and differentiation. The balance between hematopoietic stem cell self-renewal and differentiation needs to be tightly controlled, as a shift toward differentiation might

  20. Membranes replace irradiated blast cells as growth requirement for leukemic blast progenitors in suspension culture

    International Nuclear Information System (INIS)

    Nara, N.; McCulloch, E.A.

    1985-01-01

    The blast cells of acute myeloblastic leukemia (AML) may be considered as a renewal population, maintained by blast stem cells capable of both self-renewal and the generation of progeny with reduced or absent proliferative potential. This growth requires that two conditions be met: first, the cultures must contain growth factors in media conditioned either by phytohemagglutinin (PHA)-stimulated mononuclear leukocytes (PHA-LCM), or by cells of the continuous bladder carcinoma line HTB9 (HTB9-CM). Second, the cell density must be maintained at 10(6) blasts/ml; this may be achieved by adding irradiated cells to smaller numbers of intact blasts. The authors are concerned with the mechanism of the feeding function. They present evidence that (a) cell-cell contact is required. (b) Blasts are heterogeneous in respect to their capacity to support growth. (c) Fractions containing membranes from blast cells will substitute for intact cells in promoting the generation of new blast progenitors in culture. (d) This membrane function may be specific for AML blasts, since membranes from blasts of lymphoblastic leukemia or normal marrow cells were inactive

  1. Effect of cAMP signaling on expression of glucocorticoid receptor, Bim and Bad in glucocorticoid-sensitive and resistant leukemic and multiple myeloma cells.

    Science.gov (United States)

    Dong, Hongli; Carlton, Michael E; Lerner, Adam; Epstein, Paul M

    2015-01-01

    Stimulation of cAMP signaling induces apoptosis in glucocorticoid-sensitive and resistant CEM leukemic and MM.1 multiple myeloma cell lines, and this effect is enhanced by dexamethasone in both glucocorticoid-sensitive cell types and in glucocorticoid-resistant CEM cells. Expression of the mRNA for the glucocorticoid receptor alpha (GR) promoters 1A3, 1B and 1C, expression of mRNA and protein for GR, and the BH3-only proapoptotic proteins, Bim and Bad, and the phosphorylation state of Bad were examined following stimulation of the cAMP and glucocorticoid signaling pathways. Expression levels of GR promoters were increased by cAMP and glucocorticoid signaling, but GR protein expression was little changed in CEM and decreased in MM.1 cells. Stimulation of these two signaling pathways induced Bim in CEM cells, induced Bad in MM.1 cells, and activated Bad, as indicated by its dephosphorylation on ser112, in both cell types. This study shows that leukemic and multiple myeloma cells, including those resistant to glucocorticoids, can be induced to undergo apoptosis by stimulating the cAMP signaling pathway, with enhancement by glucocorticoids, and the mechanism by which this occurs may be related to changes in Bim and Bad expression, and in all cases, to activation of Bad.

  2. NK-like homeodomain proteins activate NOTCH3-signaling in leukemic T-cells

    International Nuclear Information System (INIS)

    Nagel, Stefan; Scherr, Michaela; MacLeod, Roderick AF; Venturini, Letizia; Przybylski, Grzegorz K; Grabarczyk, Piotr; Meyer, Corinna; Kaufmann, Maren; Battmer, Karin; Schmidt, Christian A; Drexler, Hans G

    2009-01-01

    Homeodomain proteins control fundamental cellular processes in development and in cancer if deregulated. Three members of the NK-like subfamily of homeobox genes (NKLs), TLX1, TLX3 and NKX2-5, are implicated in T-cell acute lymphoblastic leukemia (T-ALL). They are activated by particular chromosomal aberrations. However, their precise function in leukemogenesis is still unclear. Here we screened further NKLs in 24 T-ALL cell lines and identified the common expression of MSX2. The subsequent aim of this study was to analyze the role of MSX2 in T-cell differentiation which may be disturbed by oncogenic NKLs. Specific gene activity was examined by quantitative real-time PCR, and globally by expression profiling. Proteins were analyzed by western blot, immuno-cytology and immuno-precipitation. For overexpression studies cell lines were transduced by lentiviruses. Quantification of MSX2 mRNA in primary hematopoietic cells demonstrated higher levels in CD34+ stem cells as compared to peripheral blood cells and mature CD3+ T-cells. Furthermore, analysis of MSX2 expression levels in T-cell lines after treatment with core thymic factors confirmed their involvement in regulation. These results indicated that MSX2 represents an hematopoietic NKL family member which is downregulated during T-cell development and may functionally substituted by oncogenic NKLs. For functional analysis JURKAT cells were lentivirally transduced, overexpressing either MSX2 or oncogenic TLX1 and NKX2-5, respectively. These cells displayed transcriptional activation of NOTCH3-signaling, including NOTCH3 and HEY1 as analyzed by gene expression profiling and quantitative RT-PCR, and consistently attenuated sensitivity to gamma-secretase inhibitor as analyzed by MTT-assays. Furthermore, in addition to MSX2, both TLX1 and NKX2-5 proteins interacted with NOTCH-pathway repressors, SPEN/MINT/SHARP and TLE1/GRG1, representing a potential mechanism for (de)regulation. Finally, elevated expression of NOTCH3

  3. Detection of Ca2+-induced acetylcholine released from leukemic T-cells using an amperometric microfluidic sensor.

    Science.gov (United States)

    Akhtar, Mahmood H; Hussain, Khalil K; Gurudatt, N G; Shim, Yoon-Bo

    2017-12-15

    A microfluidic structured-dual electrodes sensor comprising of a pair of screen printed carbon electrodes was fabricated to detect acetylcholine, where one of them was used for an enzyme reaction and another for a detection electrode. The former was coated with gold nanoparticles and the latter with a porous gold layer, followed by electropolymerization of 2, 2:5,2-terthiophene-3-(p-benzoic acid) (pTTBA) on both the electrodes. Then, acetylcholinesterase was covalently attached onto the reaction electrode, and hydrazine and choline oxidase were co-immobilized on the detection electrode. The layers of both modified electrodes were characterized employing voltammetry, field emission scanning electron microscopy, X-ray photoelectron spectroscopy, and quartz crystal microscopy. After the modifications of both electrode surfaces, they were precisely faced each other to form a microfluidic channel structure, where H 2 O 2 produced from the sequential enzymatic reactions was reduced by hydrazine to obtain the analytical signal which was analyzed by the detection electrode. The microfluidic sensor at the optimized experimental conditions exhibited a wide dynamic range from 0.7nM to 1500μM with the detection limit of 0.6 ± 0.1nM based on 3s (S/N = 3). The biomedical application of the proposed sensor was evaluated by detecting acetylcholine in human plasma samples. Moreover, the Ca 2+ -induced acetylcholine released in leukemic T-cells was also investigated to show the in vitro detection ability of the designed microfluidic sensor. Interference due to the real component matrix were also studied and long term stability of the designed sensor was evaluated. The analytical performance of the designed sensor was also compared with commercially available ACh detection kit. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Transferrin-polycation-mediated introduction of DNA into human leukemic cells: Stimulation by agents that affect the survival of transfected DNA or modulate transferrin receptor levels

    International Nuclear Information System (INIS)

    Cotten, M.; Laengle-Rouault, F.; Kirlappos, H.; Wagner, E.; Mechtler, K.; Zenke, M.; Beug, H.; Birnstiel, M.L.

    1990-01-01

    The authors have subverted a receptor-mediated endocytosis event to transport genes into human leukemic cells. By coupling the natural iron-delivery protein transferrin to the DNA-binding polycations polylysine or protamine, they have created protein conjugates that bind nucleic acids and carry them into the cell during the normal transferrin cycle. They demonstrate here that this procedure is useful for a human leukemic cell line. They enhanced the rate of gene delivery by (i) increasing the transferrin receptor density through treatment of the cells with the cell permeable iron chelator desferrioxamine, (ii) interfering with the synthesis of heme with succinyl acetone treatment, or (iii) stimulating the degradation of heme with cobalt chloride treatment. Consistent with gene delivery as an endocytosis event, they show that the subsequent expression in K-562 cells of a gene included in the transported DNA depends upon the cellular presence of the lysosomotropic agent chloroquine. By contrast, monensin blocks transferrinfection, as does incubation of the cells at 18 degree C

  5. A micro-Raman spectroscopic investigation of leukemic U-937 cells in aged cultures

    Science.gov (United States)

    Fazio, Enza; Trusso, Sebastiano; Franco, Domenico; Nicolò, Marco Sebastiano; Allegra, Alessandro; Neri, Fortunato; Musolino, Caterina; Guglielmino, Salvatore P. P.

    2016-04-01

    Recently it has been shown that micro-Raman spectroscopy combined with multivariate analysis is able to discriminate among different types of tissues and tumoral cells by the detection of significant alterations and/or reorganizations of complex biological molecules, such as nucleic acids, lipids and proteins. Moreover, its use, being in principle a non-invasive technique, appears an interesting clinical tool for the evaluation of the therapeutical effects and of the disease progression. In this work we analyzed molecular changes in aged cultures of leukemia model U937 cells with respect to fresh cultures of the same cell line. In fact, structural variations of individual neoplastic cells on aging may lead to a heterogeneous data set, therefore falsifying confidence intervals, increasing error levels of analysis and consequently limiting the use of Raman spectroscopy analysis. We found that the observed morphological changes of U937 cells corresponded to well defined modifications of the Raman contributions in selected spectral regions, where markers of specific functional groups, useful to characterize the cell state, are present. A detailed subcellular analysis showed a change in cellular organization as a function of time, and correlated to a significant increase of apoptosis levels. Besides the aforementioned study, Raman spectra were used as input for principal component analysis (PCA) in order to detect and classify spectral changes among U937 cells.

  6. Glucocorticoids and Polyamine Inhibitors Synergize to Kill Human Leukemic CEM Cells1

    Science.gov (United States)

    Miller, Aaron L; Johnson, Betty H; Medh, Rheem D; Townsend, Courtney M; Thompson, E Brad

    2002-01-01

    Abstract Glucocorticoids are well-known apoptotic agents in certain classes of lymphoid cell malignancies. Reduction of intracellular polyamine levels by use of inhibitors that block polyamine synthesis slows or inhibits growth of many cells in vitro. Several such inhibitors have shown efficacy in clinical trials, though the toxicity of some compounds has limited their usefulness. We have tested the effects of combinations of the glucocorticoid dexamethasone (Dex) and two polyamine inhibitors, difluoromethylornithine (DFMO) and methyl glyoxal bis guanylhydrazone (MGBG), on the clonal line of human acute lymphoblastic leukemia cells, CEM-C7-14. Dex alone kills these cells, though only after a delay of at least 24 hours. We also evaluated a partially glucocorticoid-resistant c-Myc-expressing CEM-C7-14 clone. We show that Dex downregulates ornithine decarboxylase (ODC), the rate-limiting enzyme in polyamine synthesis. Pretreatment with the ODC inhibitor DFMO, followed by addition of Dex, enhances steroid-evoked kill slightly. The combination of pretreatment with sublethal concentrations of both DFMO and the inhibitor of S-adenosylmethionine decarboxylase, MGBG, followed by addition of Dex, results in strong synergistic cell kill. Both the rapidity and extent of cell kill are enhanced compared to the effects of Dex alone. These results suggest that use of such combinations in vivo may result in apoptosis of malignant cells with lower overall toxicity. PMID:11922393

  7. Glucocorticoids and Polyamine Inhibitors Synergize to Kill Human Leukemic CEM Cells

    Directory of Open Access Journals (Sweden)

    Aaron L. Miller

    2002-01-01

    Full Text Available Glucocorticoids are well-known apoptotic agents in certain classes of lymphoid cell malignancies. Reduction of intracellular polyamine levels by use of inhibitors that block polyamine synthesis slows or inhibits growth of many cells in vitro. Several such inhibitors have shown efficacy in clinical trials, though the toxicity of some compounds has limited their usefulness. We have tested the effects of combinations of the glucocorticoid dexamethasone. (20Dex and two polyamine inhibitors, difluoromethylornithine. (20DFMO and methyl glyoxal bis guanylhydrazone. (20MGBG, on the clonal line of human acute lymphoblastic leukemia cells, CEM-C7-14. Dex alone kills these cells, though only after a delay of at least 24 hours. We also evaluated a partially glucocorticoid-resistant c-Myc-expressing CEM-C7-14 clone. We show that Dex downregulates ornithine decarboxylase. (20ODC, the rate-limiting enzyme in polyamine synthesis. Pretreatment with the ODC inhibitor DFMO, followed by addition of Dex, enhances steroid-evoked kill slightly. The combination of pretreatment with sublethal concentrations of both DFMO and the inhibitor of S-adenosylmethionine decarboxylase, MGBG, followed by addition of Dex, results in strong synergistic cell kill. Both the rapidity and extent of cell kill are enhanced compared to the effects of Dex alone. These results suggest that use of such combinations in vivo may result in apoptosis of malignant cells with lower overall toxicity.

  8. Siglec-7 tetramers characterize B-cell subpopulations and leukemic blasts.

    Science.gov (United States)

    Gieseke, Friederike; Mang, Philippa; Viebahn, Susanne; Sonntag, Inga; Kruchen, Anne; Erbacher, Annika; Pfeiffer, Matthias; Handgretinger, Rupert; Müller, Ingo

    2012-08-01

    Cell surface glycosylation has important regulatory functions in the maturation, activation, and homeostasis of lymphocytes. The family of human sialic acid-binding immunoglobulin-like lectins (siglecs) comprises inhibitory as well as activating receptors intimately involved in the regulation of immune responses. Analyses of the interaction between siglecs and glycans are hampered by the low affinity of this interaction. Therefore, we expressed siglec-7 in eukaryotic cells, allowing for glycosylation, and oligomerized the protein in analogy to MHC tetramers. Using this tool, flow cytometric analysis of lymphocytes became possible. Sialic acid-dependent binding of siglec-7 tetramers was confirmed by glycan array analysis and loss of siglec tetramer binding after neuraminidase treatment of lymphocytes. In contrast to most lymphocyte subpopulations, which showed high siglec-7 ligand expression, B-cell subpopulations could be further subdivided according to different siglec-7 ligand expression levels. We also analyzed blasts from acute lymphoblastic leukemias of the B-cell lineage as well as the T-cell lineage, since malignant transformation is often associated with aberrant cell surface glycosylation. While pediatric T-ALL blasts highly expressed siglec-7 ligands, siglec-7 ligands were barely detectable on cALL blasts. Taken together, oligomerization of recombinant soluble siglec-7 enabled flow cytometric identification of physiologic lymphocyte subpopulations and malignant blasts. © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  9. Antiproliferative and apoptotic effects of butyrolactone lignans from Arctium lappa on leukemic cells.

    Science.gov (United States)

    Matsumoto, T; Hosono-Nishiyama, K; Yamada, H

    2006-02-01

    In the course of screening for pharmacologically active substances from extracts of crude drugs used traditionally in Sino-Japanese herbal medicines, it was found that the 70 % ethanol extract from the fruits of Arctium lappa L. (Compositae) showed potent antiproliferative activity against B cell hybridoma cell, MH60. By bioassay-guided purification, a new lignan, (+)-7,8-didehydroarctigenin, together with the known lignans (-)-arctigenin and (-)-matairesinol were isolated as the active ingredients from an aqueous ethanolic extract of the fruits of A. lappa. Of these active compounds, (-)-arctigenin showed the most potent antiproliferative activity against MH60 cells (IC (50) : 1.0 microM), and the activity was suggested to be due to apoptosis.

  10. Induction of apoptosis by Citrus paradisi essential oil in human leukemic (HL-60) cells.

    Science.gov (United States)

    Hata, Tomona; Sakaguchi, Ikuyo; Mori, Masahiro; Ikeda, Norikazu; Kato, Yoshiko; Minamino, Miki; Watabe, Kazuhito

    2003-01-01

    Limonene is a primary component of citrus essential oils (EOs) and has been reported to induce apoptosis on tumor cells. Little is known about induction of apoptosis by citrus EOs. In this study, we examined induction of apoptosis by Citrus aurantium var. dulcis (sweet orange) EO, Citrus paradisi (grapefruit) EO and Citrus limon (lemon) EO. These EOs induced apoptosis in HL-60 cells and the apoptosis activities were related to the limonene content of the EOs. Moreover, sweet orange EO and grapefruit EO may contain components besides limonene that have apoptotic activity. To identify the components with apoptotic activity, grapefruit EO was fractionated using silica gel columns, and the components were analyzed by GC-MS. The n-hexane fraction contained limonene, and the dichloromethane fraction (DF) contained aldehyde compounds and nootkatone. Decanal, octanal and citral in the DF showed strong apoptotic activity, suggesting that the aldehyde compounds induced apoptosis strongly in HL-60 cells.

  11. Kinetics of phototoxicity of Fischer's medium for L5178Y leukemic cells

    International Nuclear Information System (INIS)

    Griffin, F.M.; Ashland, G.; Capizzi, R.L.

    1981-01-01

    The uncontrolled exposure of Fischer's medium to cool white fluorescent (CWF) light or other sources emitting near-ultraviolet or visible light absorbance by riboflavin is a crucial random variable in experiments which utilize L5178Y cells and this medium. The radiation effects of CWF light result in the rapid development of toxic photoproducts in the medium which are cytostatic at lower doses of radiation and cytotoxic at higher doses. After a 24-hr suspension in medium irradiated for 3 or 48 hr, the cloning efficiencies of cells subsequently plated in light-protected medium were 87 and 3%, respectively. The corresponding near-ultraviolet doses for these periods of exposure to CWF light were 0.22 x 10(4) for a 3-hr exposure and 3.47 x 10(4) J/sq m for a 48-hr exposure. Cells incubated in lightly irradiated medium resumed growth at nearly normal rates following a 24- to 48-hr period in which no increase in cell numbers occurred. Exposure of medium containing riboflavin, but not tryptophan or tyrosine, to CWF light also produces toxic medium. Tryptophan enhances riboflavin-induced phototoxicity, whereas tyrosine diminishes this effect. As photosusceptibility of this system is very high, Fischer's medium must be fully protected from all sources of light absorbable by riboflavin

  12. Metabolic regulation of hematopoietic and leukemic stem/progenitor cells under homeostatic and stress conditions.

    Science.gov (United States)

    Karigane, Daiki; Takubo, Keiyo

    2017-07-01

    Hematopoietic stem cells (HSCs) exhibit multilineage differentiation and self-renewal activities that maintain the entire hematopoietic system during an organism's lifetime. These abilities are sustained by intrinsic transcriptional programs and extrinsic cues from the microenvironment or niche. Recent studies using metabolomics technologies reveal that metabolic regulation plays an essential role in HSC maintenance. Metabolic pathways provide energy and building blocks for other factors functioning at steady state and in stress. Here we review recent advances in our understanding of metabolic regulation in HSCs relevant to cell cycle quiescence, symmetric/asymmetric division, and proliferation following stress and lineage commitment, and discuss the therapeutic potential of targeting metabolic factors or pathways to treat hematological malignancies.

  13. DNA released by leukemic cells contributes to the disruption of the bone marrow microenvironment

    Czech Academy of Sciences Publication Activity Database

    Dvořáková, Marta; Karafiát, Vít; Pajer, Petr; Kluzáková, E.; Jarkovská, Karla; Peková, S.; Krutílková, L.; Dvořák, Michal

    2013-01-01

    Roč. 32, č. 44 (2013), s. 5201-5209 ISSN 0950-9232 R&D Projects: GA AV ČR KAN200520801; GA MŠk(CZ) LC06061; GA ČR GA204/06/1728; GA ČR GA301/09/1727 Institutional research plan: CEZ:AV0Z50520514 Institutional support: RVO:68378050 ; RVO:67985904 Keywords : acute leukemia * tumor microenvironment * extracellular nucleosomes * extracellular DNA * DNA damage response * cell death Subject RIV: EB - Genetics ; Molecular Biology Impact factor: 8.559, year: 2013

  14. Antileukemic Potential of Momordica charantia Seed Extracts on Human Myeloid Leukemic HL60 Cells

    Directory of Open Access Journals (Sweden)

    Ramani Soundararajan

    2012-01-01

    Full Text Available Momordica charantia (bitter gourd has been used in the traditional system of medicine for the treatment of various diseases. Anticancer activity of M. charantia extracts has been demonstrated by numerous in vitro and in vivo studies. In the present study, we investigated the differentiation inducing potential of fractionated M. charantia seed extracts in human myeloid HL60 cells. We found that the HL60 cells treated with the fractionated seed extracts differentiated into granulocytic lineage as characterized by NBT staining, CD11b expression, and specific esterase activity. The differentiation inducing principle was found to be heat-stable, and organic in nature. The differentiation was accompanied by a downregulation of c-myc transcript, indicating the involvement of c-myc pathway, at least in part, in differentiation. Taken together these results indicate that fractionated extracts of M. charantia seeds possess differentiation inducing activity and therefore can be evaluated for their potential use in differentiation therapy for leukemia in combination with other inducers of differentiation.

  15. Antileukemic Potential of Momordica charantia Seed Extracts on Human Myeloid Leukemic HL60 Cells

    Science.gov (United States)

    Soundararajan, Ramani; Prabha, Punit; Rai, Umesh; Dixit, Aparna

    2012-01-01

    Momordica charantia (bitter gourd) has been used in the traditional system of medicine for the treatment of various diseases. Anticancer activity of M. charantia extracts has been demonstrated by numerous in vitro and in vivo studies. In the present study, we investigated the differentiation inducing potential of fractionated M. charantia seed extracts in human myeloid HL60 cells. We found that the HL60 cells treated with the fractionated seed extracts differentiated into granulocytic lineage as characterized by NBT staining, CD11b expression, and specific esterase activity. The differentiation inducing principle was found to be heat-stable, and organic in nature. The differentiation was accompanied by a downregulation of c-myc transcript, indicating the involvement of c-myc pathway, at least in part, in differentiation. Taken together these results indicate that fractionated extracts of M. charantia seeds possess differentiation inducing activity and therefore can be evaluated for their potential use in differentiation therapy for leukemia in combination with other inducers of differentiation. PMID:22654956

  16. ArtinM Mediates Murine T Cell Activation and Induces Cell Death in Jurkat Human Leukemic T Cells

    Science.gov (United States)

    Oliveira-Brito, Patrícia Kellen Martins; Gonçalves, Thiago Eleutério; Vendruscolo, Patrícia Edivânia; Roque-Barreira, Maria Cristina

    2017-01-01

    The recognition of cell surface glycans by lectins may be critical for the innate and adaptive immune responses. ArtinM, a d-mannose-binding lectin from Artocarpus heterophyllus, activates antigen-presenting cells by recognizing TLR2 N-glycans and induces Th1 immunity. We recently demonstrated that ArtinM stimulated CD4+ T cells to produce proinflammatory cytokines. Here, we further studied the effects of ArtinM on adaptive immune cells. We showed that ArtinM activates murine CD4+ and CD8+ T cells, augmenting their positivity for CD25, CD69, and CD95 and showed higher interleukin (IL)-2 and interferon (IFN)-γ production. The CD4+ T cells exhibited increased T-bet expression in response to ArtinM, and IL-2 production by CD4+ and CD8+ T cells depended on the recognition of CD3εγ-chain glycans by ArtinM. The ArtinM effect on aberrantly-glycosylated neoplastic lymphocytes was studied in Jurkat T cells, in which ArtinM induced IL-2, IFN-γ, and IL-1β production, but decreased cell viability and growth. A higher frequency of AnnexinV- and propidium iodide-stained cells demonstrated the induction of Jurkat T cells apoptosis by ArtinM, and this apoptotic response was reduced by caspases and protein tyrosine kinase inhibitors. The ArtinM effects on murine T cells corroborated with the immunomodulatory property of lectin, whereas the promotion of Jurkat T cells apoptosis may reflect a potential applicability of ArtinM in novel strategies for treating lymphocytic leukemia. PMID:28665310

  17. 5-Aza-2'-deoxycytidine synergistic action with thymidine on leukemic cells and interaction of 5-aza-dCMP with dCMP deaminase

    International Nuclear Information System (INIS)

    Momparler, R.L.; Bartolucci, S.; Bouchard, J.; Momparler, L.F.; Raia, C.A.; Rossi, M.

    1986-01-01

    The authors observe a synergistic antineoplastic effect between 5-AZA-dCR and dTR on leukemia cells in culture. In order to understand the mechanism behind this interaction the authors investigate the effects of dTTP on the deamination of 5-aza-2'-deoxycytidine-5'-monophosphate (5-AZA-dCMP) by dCMP deaminase. The effects of 5-AZA-dCTP on this enzyme is also studied. The incorporation of tritium-5-AZA-Cdr into DNA of leukemic cells was performed. The amount of radioactivity incorproated into DNA was determined by trapping the cells on GF/C glass fiber filters and washing with cold TCA. It is shown that the modulation of the atieoplastic activity of deoxycytidine analogs by allosteric effectors such as dTTP may have the potential to increase the effectiveness of the chemotherapy for acute leukemia

  18. Ethyl acetate extract of Chinese medicinal herb Sarcandra glabra induces growth inhibition on human leukemic HL-60 cells, associated with cell cycle arrest and up-regulation of pro-apoptotic Bax/Bcl-2 ratio.

    Science.gov (United States)

    Li, W Y; Chiu, Lawrence C M; Lam, W S; Wong, W Y; Chan, Y T; Ho, Y P; Wong, Elaine Y L; Wong, Y S; Ooi, Vincent E C

    2007-02-01

    Sarcandra glabra (Thunb.) Nakai, colloquially known as Caoshanhu, is a Chinese medicinal herb with reported anti-tumor, anti-inflammatory, anti-viral and non-specific immunoenhancing properties. Although the plant has been clinically used for treating a variety of diseases, its bioactive ingredients are largely unknown and its mode of action has never been investigated. In this study, the anti-tumor property of ethyl acetate (EA) extract of S. glabra was investigated by determining its in vitro growth-inhibitory effects on a panel of human cancer cell lines of different histotypes. Growth inhibition of the EA extract on the cancer cells seemed to be selective, and the leukemic HL-60 was found to be the most responsive after 48 h of treatment (IC50=58 microg/ml). Flow cytometric studies further illustrated that the extract might interfere with DNA replication and thus arrested the cell cycle at S phase in the leukemic cells, followed by DNA fragmentation and loss of phospholipid asymmetry in the plasma membrane after 72 h of treatment. Concurrently, the pro-apoptotic Bax/Bcl-2 ratio was also up-regulated by more than 178% of the control level. All these findings suggested that the extract had initiated apoptosis to kill the leukemic cells. Results from this pioneer study help to establish a scientific foundation for future research and development of the bioactive ingredients in EA extract of S. glabra as efficacious anti-cancer agents.

  19. C60 Fullerene Effects on Diphenyl-N-(trichloroacetyl)-amidophosphate Interaction with DNA In Silico and Its Cytotoxic Activity Against Human Leukemic Cell Line In Vitro

    Science.gov (United States)

    Grebinyk, A.; Prylutska, S.; Grynyuk, I.; Kolp, B.; Hurmach, V.; Sliva, T.; Amirkhanov, V.; Trush, V.; Matyshevska, O.; Slobodyanik, M.; Prylutskyy, Yu.; Frohme, M.; Ritter, U.

    2018-03-01

    New representative of carbacylamidophosphates - diphenyl-N-(trichloroacetyl)-amidophosphate (HL), which contains two phenoxy substituents near the phosphoryl group, was synthesized, identified by elemental analysis and IR and NMR spectroscopy, and tested as a cytotoxic agent itself and in combination with C60 fullerene. According to molecular simulation results, C60 fullerene and HL could interact with DNA and form a rigid complex stabilized by stacking interactions of HL phenyl groups with C60 fullerene and DNA G nucleotide, as well as by interactions of HL CCl3 group by ion-π bonds with C60 molecule and by electrostatic bonds with DNA G nucleotide. With the use of MTT test, the cytotoxic activity of HL against human leukemic CCRF-CM cells with IC50 value detected at 10 μM concentration at 72 h of cells treatment was shown. Under combined action of 16 μM C60 fullerene and HL, the value of IC50 was detected at lower 5 μM HL concentration and at earlier 48 h period of incubation, besides the cytotoxic effect of HL was observed at a low 2.5 μM concentration at which HL by itself had no influence on cell viability. Binding of C60 fullerene and HL with minor DNA groove with formation of a stable complex is assumed to be one of the possible reasons of their synergistic inhibition of CCRF-CEM cells proliferation. Application of C60 fullerene in combination with 2.5 μM HL was shown to have no harmful effect on structural stability of blood erythrocytes membrane. Thus, combined action of C60 fullerene and HL in a low concentration potentiated HL cytotoxic effect against human leukemic cells and was not followed by hemolytic effect.

  20. Correlation of chromosome patterns in human leukemic cells with exposure to chemicals and/or radiation. Progress report, January 1, 1981-December 31, 1981

    International Nuclear Information System (INIS)

    Rowley, J.D.

    1981-08-01

    The overall aim is to determine whether there is a relationship between exposure to radiation, environmental pollutants, and/or genetic background and the development of ANLL or other hematologic malignancies. I will try to define the factors that influence the development of ANLL as a second malignancy in patients who have been exposed to large doses of radiotherapy and/or chemotherapeutic agents. Two long-term goals are (1) to identify the genes that are located at the sites of consistent translocations, and then to determine the alterations in gene function that are associated with these translocations and (2) to establish the baseline frequency of various chromosome changes (mutations) in myeloid cells and then to analyze the influence of various types of environmental exposure or medical treatment on this baseline mutation rate. Ultimately, it may be possible to determine the extent of mutagenic exposure in various populations through an analysis of the leukemic cells of that populations

  1. Unsupervised explorative data analysis of normal human leukocytes and BCR/ABL positive leukemic cells mid-infrared spectra

    NARCIS (Netherlands)

    Bellisola, G.; Bolomini-Vittori, M.; Cinque, G.; Dumas, P.; Fiorini, Z.; Laudanna, C.; Mirenda, M.; Sandt, C.; Silvestri, G.; Tomasello, L.; Vezzalini, M.; Wehbe, K.; Sorio, C.

    2015-01-01

    We proved the ability of Fourier Transform Infrared microspectroscopy (microFTIR) complemented by Principal Component Analysis (PCA) to detect protein phosphorylation/de-phosphorylation in mammalian cells. We analyzed by microFTIR human polymorphonuclear neutrophil (PMNs) leukocytes, mouse-derived

  2. Human surfactant protein D alters oxidative stress and HMGA1 expression to induce p53 apoptotic pathway in eosinophil leukemic cell line.

    Directory of Open Access Journals (Sweden)

    Lakshna Mahajan

    Full Text Available Surfactant protein D (SP-D, an innate immune molecule, has an indispensable role in host defense and regulation of inflammation. Immune related functions regulated by SP-D include agglutination of pathogens, phagocytosis, oxidative burst, antigen presentation, T lymphocyte proliferation, cytokine secretion, induction of apoptosis and clearance of apoptotic cells. The present study unravels a novel ability of SP-D to reduce the viability of leukemic cells (eosinophilic leukemic cell line, AML14.3D10; acute myeloid leukemia cell line, THP-1; acute lymphoid leukemia cell lines, Jurkat, Raji; and human breast epithelial cell line, MCF-7, and explains the underlying mechanisms. SP-D and a recombinant fragment of human SP-D (rhSP-D induced G2/M phase cell cycle arrest, and dose and time-dependent apoptosis in the AML14.3D10 eosinophilic leukemia cell line. Levels of various apoptotic markers viz. activated p53, cleaved caspase-9 and PARP, along with G2/M checkpoints (p21 and Tyr15 phosphorylation of cdc2 showed significant increase in these cells. We further attempted to elucidate the underlying mechanisms of rhSP-D induced apoptosis using proteomic analysis. This approach identified large scale molecular changes initiated by SP-D in a human cell for the first time. Among others, the proteomics analysis highlighted a decreased expression of survival related proteins such as HMGA1, overexpression of proteins to protect the cells from oxidative burst, while a drastic decrease in mitochondrial antioxidant defense system. rhSP-D mediated enhanced oxidative burst in AML14.3D10 cells was confirmed, while antioxidant, N-acetyl-L-cysteine, abrogated the rhSP-D induced apoptosis. The rhSP-D mediated reduced viability was specific to the cancer cell lines and viability of human PBMCs from healthy controls was not affected. The study suggests involvement of SP-D in host's immunosurveillance and therapeutic potential of rhSP-D in the eosinophilic leukemia and

  3. Stable curcumin-loaded polymeric micellar formulation for enhancing cellular uptake and cytotoxicity to FLT3 overexpressing EoL-1 leukemic cells.

    Science.gov (United States)

    Tima, Singkome; Anuchapreeda, Songyot; Ampasavate, Chadarat; Berkland, Cory; Okonogi, Siriporn

    2017-05-01

    The present study aims to develop a stable polymeric micellar formulation of curcumin (CM) with improved solubility and stability, and that is suitable for clinical applications in leukemia patients. CM-loaded polymeric micelles (CM-micelles) were prepared using poloxamers. The chemical structure of the polymers influenced micellar properties. The best formulation of CM-micelles, namely CM-P407, was obtained from poloxamer 407 at drug to polymer ratio of 1:30 and rehydrated with phosphate buffer solution pH 7.4. CM-P407 exhibited the smallest size of 30.3±1.3nm and highest entrapment efficiency of 88.4±4.1%. When stored at -80°C for 60days, CM-P407 retained high protection of CM and had no significant size change. In comparison with CM solution in dimethyl sulfoxide (CM-DMSO), CM kinetic degradation in both formulations followed a pseudo-first-order reaction, but the half-life of CM in CM-P407 was approx. 200 times longer than in CM-DMSO. Regarding the activity against FLT3 overexpressing EoL-1 leukemic cells, CM-P407 showed higher cytotoxicity than CM-DMSO. Moreover, intracellular uptake to leukemic cells of CM-P407 was 2-3 times greater than that of CM-DMSO. These promising results for CM-P407 will be further investigated in rodents and in clinical studies for leukemia treatment. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. A micro-Raman spectroscopic investigation of leukemic U-937 cells treated with Crotalaria agatiflora Schweinf and the isolated compound madurensine

    Science.gov (United States)

    le Roux, Karlien; Prinsloo, Linda C.; Hussein, Ahmed A.; Lall, Namrita

    In South Africa traditional medicine plays an important role in primary health care and therefore it is very important that the medicinal use of plants is scientifically tested for toxicity and effectiveness. It was established that the ethanolic extract of the leaves of Crotalaria agatiflora, as well as the isolated compound madurensine, is moderately toxic against leukemic U-937 cells. Light microscopic investigations indicated that symptoms of cell death are induced during treatments, but flow cytometry analysis of treated cells, using annexin-V and propidium iodide, showed that apoptosis and necrosis are insignificantly induced. The Raman results suggested that protein extraction and DNA melting occur in the cells during treatment with the ethanolic extracts (IC50 value 73.9 μg/mL), drastically changing the molecular content of the cells. In contrast, treatment with madurensine (IC50 value 136.5 μg/mL), an isolated pyrrolizidine alkaloid from the ethanolic extract of the leaves, did not have the same effect. The results are also compared to that of cells treated with actinomycin D, a compound known to induce apoptosis. The investigation showed that micro-Raman spectroscopy has great promise to be used for initial screening of samples to determine the effects of different treatments on cancerous cell lines together with conventional methods. The results highlight the fact that for many natural products used for medicinal purposes, the therapeutic effect of the crude plant extract tends to be significantly more effective than the particular action of its individual constituents.

  5. Cytotoxic Effect on Human Myeloma Cells and Leukemic Cells by the Agaricus blazei Murill Based Mushroom Extract, Andosan™

    Directory of Open Access Journals (Sweden)

    Jon-Magnus Tangen

    2017-01-01

    Full Text Available Agaricus blazei Murill is an edible mushroom of the Basidiomycetes family, which has been found to contain a number of compounds with antitumor properties, such as proteoglycans and ergosterol. In the present investigation, we show that the commercial mushroom product Andosan, which contains 82.4% Agaricus blazei Murill, together with medicinal mushrooms Hericium erinaceus (14.7% and Grifola frondosa (2.9%, has a cytotoxic effect on primary myeloma cells, other myeloma cell lines, and leukemia cell lines in vitro. Although the exact content and hence the mechanisms of action of the Andosan extract are unknown, we have found in this investigation indications of cell cycle arrest when myeloma cell lines are cultivated with Andosan. This may be one of the possible explanations for the cytotoxic effects of Andosan.

  6. The novel AML stem cell associated antigen CLL-1 aids in discrimination between normal and leukemic stem cells.

    Science.gov (United States)

    van Rhenen, Anna; van Dongen, Guus A M S; Kelder, Angèle; Rombouts, Elwin J; Feller, Nicole; Moshaver, Bijan; Stigter-van Walsum, Marijke; Zweegman, Sonja; Ossenkoppele, Gert J; Jan Schuurhuis, Gerrit

    2007-10-01

    In CD34(+) acute myeloid leukemia (AML), the malignant stem cells reside in the CD38(-) compartment. We have shown before that the frequency of such CD34(+)CD38(-) cells at diagnosis correlates with minimal residual disease (MRD) frequency after chemotherapy and with survival. Specific targeting of CD34(+)CD38(-) cells might thus offer therapeutic options. Previously, we found that C-type lectin-like molecule-1 (CLL-1) has high expression on the whole blast compartment in the majority of AML cases. We now show that CLL-1 expression is also present on the CD34(+)CD38(-) stem- cell compartment in AML (77/89 patients). The CD34(+)CLL-1(+) population, containing the CD34(+)CD38(-)CLL-1(+) cells, does engraft in nonobese diabetic/severe combined immunodeficiency (NOD/SCID) mice with outgrowth to CLL-1(+) blasts. CLL-1 expression was not different between diagnosis and relapse (n = 9). In remission, both CLL-1(-) normal and CLL-1(+) malignant CD34(+)CD38(-) cells were present. A high CLL-1(+) fraction was associated with quick relapse. CLL-1 expression is completely absent both on CD34(+)CD38(-) cells in normal (n = 11) and in regenerating bone marrow controls (n = 6). This AML stem-cell specificity of the anti-CLL-1 antibody under all conditions of disease and the leukemia-initiating properties of CD34(+)CLL-1(+) cells indicate that anti-CLL-1 antibody enables both AML-specific stem-cell detection and possibly antigen-targeting in future.

  7. The cathepsin B inhibitor z-FA-CMK induces cell death in leukemic T cells via oxidative stress.

    Science.gov (United States)

    Liow, K Y; Chow, Sek C

    2018-01-01

    The cathepsin B inhibitor benzyloxycarbonyl-phenylalanine-alanine-chloromethyl ketone (z-FA-CMK) was recently found to induce apoptosis at low concentrations in Jurkat T cells, while at higher concentrations, the cells die of necrosis. In the present study, we showed that z-FA-CMK readily depletes intracellular glutathione (GSH) with a concomitant increase in reactive oxygen species (ROS) generation. The toxicity of z-FA-CMK in Jurkat T cells was completely abrogated by N-acetylcysteine (NAC), suggesting that the toxicity mediated by z-FA-CMK is due to oxidative stress. We found that L-buthionine sulfoximine (BSO) which depletes intracellular GSH through the inhibition of GSH biosynthesis in Jurkat T cells did not promote ROS increase or induce cell death. However, NAC was still able to block z-FA-CMK toxicity in Jurkat T cells in the presence of BSO, indicating that the protective effect of NAC does not involve GSH biosynthesis. This is further corroborated by the protective effect of the non-metabolically active D-cysteine on z-FA-CMK toxicity. Furthermore, in BSO-treated cells, z-FA-CMK-induced ROS increased which remains unchanged, suggesting that the depletion of GSH and increase in ROS generation mediated by z-FA-CMK may be two separate events. Collectively, our results demonstrated that z-FA-CMK toxicity is mediated by oxidative stress through the increase in ROS generation.

  8. Anti-leukemic effect of a synthetic compound, (±) trans-dihydronarciclasine (HYU-01) via cell-cycle arrest and apoptosis in acute myeloid leukemia.

    Science.gov (United States)

    Kim, Seo Ju; Park, Hyun Ki; Kim, Ju Young; Yoon, Jin Sun; Kim, Eun Shil; Cho, Cheon-Gyu; Kim, Byoung Kook; Park, Byeong Bae; Lee, Young Yiul

    2012-10-01

    (±) trans-Dihydronarciclasine, isolated from Chinese medicinal plant Zephyranthes candida, has been shown to possess quite potent anti-tumoral effect against selected human cancer cell lines. However, little is known about the anti-tumoral effect of (±) trans-dihydronarciclasine in acute myeloid leukemia (AML). This study was performed to investigate the effect of a novel synthetic (±) trans-dihydronarciclasine (code name; HYU-01) in AML. The HYU-01 inhibited the proliferation of various AML cell lines including HL-60 as well as primary leukemic blasts in a dose-dependent manner. To investigate the mechanism of the anti-proliferative effect of HYU-01, cell-cycle analysis was attempted in HL-60 cells, resulting in G1 arrest. The expression levels of CDK2, CDK4, CDK6, cyclin E, and cyclin A were decreased in a time-dependent manner. In addition, HYU-01 up-regulated the expression of the p27, and markedly enhanced the binding of p27 with CDK2, 4, and 6, ultimately resulting in the decrease of their kinase activities. Furthermore, HYU-01 induced the apoptosis through the induction of proapoptotic molecules and reduction of antiapoptotic molecules in association with the activation of caspase-3, -8, and -9. These results suggest that HYU-01 may inhibit the proliferation of HL-60 cells, via apoptosis, as well as G1 block in association with the induction of p27. © 2012 The Authors APMIS © 2012 APMIS.

  9. Constitutive STAT3-activation in Sezary syndrome: tyrphostin AG490 inhibits STAT3-activation, interleukin-2 receptor expression and growth of leukemic Sezary cells

    DEFF Research Database (Denmark)

    Eriksen, K W; Kaltoft, K; Mikkelsen, G

    2001-01-01

    are IL-2Ralpha negative. An aberrant expression of IL-2Ralpha has recently been described in cutaneous T-cell lymphoma (CTCL). Here, we study the regulation of IL-2Ralpha expression and STATs in a tumor cell line obtained from peripheral blood from a patient with Sezary syndrome (SS), a leukemic variant...... of CTCL. We show that (1) STAT3 (a transcription factor known to regulate IL-2Ralpha transcription) is constitutively tyrosine-phosphorylated in SS tumor cells, but not in non-malignant T cells; (2) STAT3 binds constitutively to a STAT-binding sequence in the promotor of the IL-2Ralpha gene; (3) the Janus...... kinase inhibitor, tyrphostine AG490, inhibits STAT3 activation, STAT3 DNA binding, and IL-2Ralpha mRNA and protein expression in parallel; and (4) tyrphostine AG490 inhibits IL-2 driven mitogenesis and triggers apoptosis in SS tumor cells. In conclusion, we provide the first example of a constitutive...

  10. Assay of hybrid ribonuclease using a membrane filter-immobilized synthetic hybrid: application to the human leukemic cell

    International Nuclear Information System (INIS)

    Papaphilis, A.D.; Kamper, E.F.

    1985-01-01

    A method for assaying hybrid ribonuclease has been devised which utilizes as substrate the synthetic hybrid [ 3 H]polyriboadenylic acid [poly(rA)]:polydeoxythymidylic acid [poly(dT)] immobilized on the solid matrix of nitrocellulose filters. The hybridization on filter of [ 3 H]poly(rA) to poly(dT) has been explored in terms of efficacy of the process and the response of the product to RNase H. A pulse of uv irradiation of poly(dT) while in dry state on the filter increased its firm binding to the filter in a concentration-dependent manner, resulting in a concomitant increase of the yield of hybrid formation. The filter-immobilized hybrid was 95% resistant to RNase A but sensitive to RNase H. When stored in toluene in the cold the hybrid maintained its stability for over 6 months, as judged by its resistance to RNase A. The method offers a number of advantages over assays that use solution hybrids as substrates and was readily applicable in the screening of leukemic patients, in the leukocytes of which it has demonstrated increased RNase H levels

  11. Leukemic Oral Manifestations and their Management.

    Science.gov (United States)

    Francisconi, Carolina Favaro; Caldas, Rogerio Jardim; Oliveira Martins, Lazara Joyce; Fischer Rubira, Cassia Maria; da Silva Santos, Paulo Sergio

    2016-01-01

    Leukemia is the most common neoplastic disease of the white blood cells which is important as a pediatric malignancy. Oral manifestations occur frequently in leukemic patients and may present as initial evidence of the disease or its relapse. The symptoms include gingival enlargement and bleeding, oral ulceration, petechia, mucosal pallor, noma, trismus and oral infections. Oral lesions arise in both acute and chronic forms of all types of leukemia. These oral manifestations either may be the result of direct infiltration of leukemic cells (primary) or secondary to underlying thrombocytopenia, neutropenia, or impaired granulocyte function. Despite the fact that leukemia has long been known to be associated with oral lesions, the available literature on this topic consists mostly of case reports, without data summarizing the main oral changes for each type of leukemia. Therefore, the present review aimed at describing oral manifestations of all leukemia types and their dental management. This might be useful in early diagnosis, improving patient outcomes.

  12. Activation of store – operated Ca(2+ entry in cisplatin resistant leukemic cells after treatment with photoexcited fullerene C(60 and cisplatin

    Directory of Open Access Journals (Sweden)

    D. V. Franskevych

    2018-04-01

    Full Text Available Ca2+-regulating system in cancer cells is suggested to be remodulated particularly by reduced store-operated Ca2+ entry (SOCE through plasma membrane in order to maintain moderately reduced cytosolic Ca2+ concentration and to avoid apoptosis. The endoplasmic reticulum (ER Ca2+ pool content and the size of SOCE in leukemic wild type (L1210 and resistant to cisplatin (L1210R cells in control, after treatment with either cisplatin (1 µg/ml or photoexcited fulleren C60 (10-5 M alone, or their combination were estimated with the use of Indo-1 AM. The SOCE in resistant to cisplatin L1210R cells was found to be lower than in the wild-type cells. After treatment with cisplatin the decrease of thapsigargin (TG-sensitive ER Ca2+ pool with no significant increase of SOCE was observed in L1210 cells, while no changes were detected in L1210R cells. Photoexcitation of intracellular accumulated fullerene C60 in the visible range of spectrum (410-700 nm was accompanied by increase of SOCE not only in sensitive, but in resistant cells as well. In resistant L1210R cells treated with photoexcited C60 essential effect of cisplatin on Ca2+ homeostasis became obvious: the size of SOCE proved to be higher than after treatment with photoexcited C60 alone. The data obtained allow suggesting­ the influence of photoexcited C60 not only on Ca2+-regulating system, but on those involved in controlling cisplatin entry into drug resistant cancer cells.

  13. Sporadic on/off switching of HTLV-1 Tax expression is crucial to maintain the whole population of virus-induced leukemic cells.

    Science.gov (United States)

    Mahgoub, Mohamed; Yasunaga, Jun-Ichirou; Iwami, Shingo; Nakaoka, Shinji; Koizumi, Yoshiki; Shimura, Kazuya; Matsuoka, Masao

    2018-02-06

    Viruses causing chronic infection artfully manipulate infected cells to enable viral persistence in vivo under the pressure of immunity. Human T-cell leukemia virus type 1 (HTLV-1) establishes persistent infection mainly in CD4+ T cells in vivo and induces leukemia in this subset. HTLV-1-encoded Tax is a critical transactivator of viral replication and a potent oncoprotein, but its significance in pathogenesis remains obscure due to its very low level of expression in vivo. Here, we show that Tax is expressed in a minor fraction of leukemic cells at any given time, and importantly, its expression spontaneously switches between on and off states. Live cell imaging revealed that the average duration of one episode of Tax expression is ∼19 hours. Knockdown of Tax rapidly induced apoptosis in most cells, indicating that Tax is critical for maintaining the population, even if its short-term expression is limited to a small subpopulation. Single-cell analysis and computational simulation suggest that transient Tax expression triggers antiapoptotic machinery, and this effect continues even after Tax expression is diminished; this activation of the antiapoptotic machinery is the critical event for maintaining the population. In addition, Tax is induced by various cytotoxic stresses and also promotes HTLV-1 replication. Thus, it seems that Tax protects infected cells from apoptosis and increases the chance of viral transmission at a critical moment. Keeping the expression of Tax minimal but inducible on demand is, therefore, a fundamental strategy of HTLV-1 to promote persistent infection and leukemogenesis. Copyright © 2018 the Author(s). Published by PNAS.

  14. MicroRNA expression changes during human leukemic HL-60 cell differentiation induced by 4-hydroxynonenal, a product of lipid peroxidation.

    Science.gov (United States)

    Pizzimenti, Stefania; Ferracin, Manuela; Sabbioni, Silvia; Toaldo, Cristina; Pettazzoni, Piergiorgio; Dianzani, Mario Umberto; Negrini, Massimo; Barrera, Giuseppina

    2009-01-15

    4-Hydroxynonenal (HNE) is one of several lipid oxidation products that may have an impact on human pathophysiology. It is an important second messenger involved in the regulation of various cellular processes and exhibits antiproliferative and differentiative properties in various tumor cell lines. The mechanisms by which HNE affects cell growth and differentiation are only partially clarified. Because microRNAs (miRNAs) have the ability to regulate several cellular processes, we hypothesized that HNE, in addition to other mechanisms, could affect miRNA expression. Here, we present the results of a genome-wide miRNA expression profiling of HNE-treated HL-60 leukemic cells. Among 470 human miRNAs, 10 were found to be differentially expressed between control and HNE-treated cells (at p<0.05). Six miRNAs were down-regulated (miR-181a*, miR-199b, miR-202, miR-378, miR-454-3p, miR-575) and 4 were up-regulated (miR-125a, miR-339, miR-663, miR-660). Three of these regulated miRNAs (miR-202, miR-339, miR-378) were further assayed and validated by quantitative real-time RT-PCR. Moreover, consistent with the down-regulation of miR-378, HNE also induced the expression of the SUFU protein, a tumor suppressor recently identified as a target of miR-378. The finding that HNE could regulate the expression of miRNAs and their targets opens new perspectives on the understanding of HNE-controlled pathways. A functional analysis of 191 putative gene targets of miRNAs modulated by HNE is discussed.

  15. The cyclin-dependent kinase inhibitor 5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole induces nongenotoxic, DNA replication-independent apoptosis of normal and leukemic cells, regardless of their p53 status

    International Nuclear Information System (INIS)

    Turinetto, Valentina; Porcedda, Paola; Orlando, Luca; De Marchi, Mario; Amoroso, Antonio; Giachino, Claudia

    2009-01-01

    Current chemotherapy of human cancers focuses on the DNA damage pathway to induce a p53-mediated cellular response leading to either G1 arrest or apoptosis. However, genotoxic treatments may induce mutations and translocations that result in secondary malignancies or recurrent disease. In addition, about 50% of human cancers are associated with mutations in the p53 gene. Nongenotoxic activation of apoptosis by targeting specific molecular pathways thus provides an attractive therapeutic approach. Normal and leukemic cells were evaluated for their sensitivity to 5, 6-dichloro-1-beta-D-ribofuranosylbenzimidazole (DRB) through cell viability and caspase activation tests. The apoptotic pathway induced by DRB was analysed by immunfluorescence and immunoblot analysis. H2AX phosphorylation and cell cycle analysis were performed to study the dependance of apoptosis on DNA damage and DNA replication, respectively. To investigate the role of p53 in DRB-induced apoptosis, specific p53 inhibitors were used. Statistical analysis on cell survival was performed with the test of independence. Here we report that DRB, an inhibitor of the transcriptional cyclin-dependent kinases (CDKs) 7 and 9, triggers DNA replication-independent apoptosis in normal and leukemic human cells regardless of their p53 status and without inducing DNA damage. Our data indicate that (i) in p53-competent cells, apoptosis induced by DRB relies on a cytosolic accumulation of p53 and subsequent Bax activation, (ii) in the absence of p53, it may rely on p73, and (iii) it is independent of ATM and NBS1 proteins. Notably, even apoptosis-resistant leukemic cells such as Raji were sensitive to DRB. Our results indicate that DRB represents a potentially useful cancer chemotherapeutic strategy that employs both the p53-dependent and -independent apoptotic pathways without inducing genotoxic stress, thereby decreasing the risk of secondary malignancies

  16. shRNA-mediated EMMPRIN silencing inhibits human leukemic monocyte lymphoma U937 cell proliferation and increases chemosensitivity to adriamycin.

    Science.gov (United States)

    Gao, Hui; Jiang, Qixiao; Han, Yantao; Peng, Jianjun; Wang, Chunbo

    2015-03-01

    EMMPRIN is a widely distributed cell surface glycoprotein, which plays an important role in tumor progression and confers resistance to some chemotherapeutic drugs. Recent studies have shown that EMMPRIN overexpression indicates poor prognosis in acute myeloid leukemia (AML). However, little was known on the role of EMMPRIN in leukemia. Human leukemia cell line U937 was stably transfected with a EMMPRIN-targeted shRNA-containing vector to investigate the effect of EMMPRIN on cellular functions. EMMPRIN expression was monitored by qRT-PCR and Western blotting. Cell viability and proliferation were determined by trypan blue exclusion and BrdU labeling, respectively. Cell cycle and apoptosis were analyzed by flow cytometry. Cytotoxicity of chemotherapeutic agent adriamycin on cells was assessed by MTT assay. Knockdown of EMMPRIN gene significantly inhibited cell viability and decreased cell proliferation. Fluorescence-activated cell-sorting analysis revealed that the reduced EMMPRIN expression resulted in cell cycle arrest at G1 phase and induced apoptosis. Meanwhile, western blotting analysis showed that EMMPRIN knockdown was associated with downregulation of cell cycle- and apoptosis-related molecules including cyclin D1, cyclin E, as well as increase in cleavage of caspase-3 and PARP. This study also showed that silencing of EMMPRIN sensitized U937 cells to Adriamycin. EMMPRIN is involved in proliferation, growth, and chemosensitivity of human AML line U937, indicating that EMMPRIN may be a promising therapeutic target for AML.

  17. Establishment of a high production system for AIDS retroviruses with a human T-leukemic cell line Molt-4

    International Nuclear Information System (INIS)

    Koyanagi, Yoshio; Harada, Shinji; Yamamoto, Naoki

    1986-01-01

    A cell culture system was developed for the continuous and efficient production of acquired immune deficiency syndrome (AIDS) retrovirus. After infection of a human T-cell line Molt-4 with HTLV-III and LAV the cells grow permanently and produce large amounts of virus continuously. The yields of production of virus were assessed either with reverse transcriptase activity or a newly established biological quantitation assay of active virus. The amounts of virus with this cell system were much higher than those of the H9 cell system. This procedure enabled us first to compare the two viral isolates HTLV-III and LAV directly in the same cell lines. Establishment of the culture system, allowing efficient production of AIDS retroviruses, provides a useful tool for the isolation of the virus from patients with AIDS and for more basic research, such as the mechanisms of immune destruction caused by the virus leading to the occurence of various malignancies (author)

  18. Increased risk of ALL among premature infants is not explained by increased prevalence of pre-leukemic cell clones

    DEFF Research Database (Denmark)

    Lausten-Thomsen, Ulrik; Madsen, Hans O.; Vestergaard, Therese Risom

    2010-01-01

    in the prevalence and magnitude of preleukaemic t(12;21)-positive cells compared to previously published data from mature children could be demonstrated. This indirectly supports the theory that prevalence and quantity of preleukaemic t(12;21)-positive cells peaks at term or early childhood and that exogenous...

  19. Induction of polyploidization in leukemic cell lines and primary bone marrow by Src kinase inhibitor SU6656

    Science.gov (United States)

    Lannutti, Brian J.; Blake, Noel; Gandhi, Manish J.; Reems, Jo Anna; Drachman, Jonathan G.

    2005-01-01

    Megakaryocytes (MKs) undergo successive rounds of endomitosis during differentiation, resulting in polyploidy (typically, 16-64N). Previous studies have demonstrated that this occurs through an interruption of normal cell cycle progression during anaphase. However, the molecular mechanism(s) controlling this unique process is undefined. In the present report, we examine the effect of an Src kinase inhibitor, SU6656, on thrombopoietin (TPO)-induced growth and differentiation. Remarkably, when SU6656 (2.5 μM) was added to a megakaryocytic cell line, UT-7/TPO, the cells ceased cell division but continued to accumulate DNA by endomitosis. During this interval, CD41 and CD61 expression on the cell surface increased. Similar effects on polyploidization and MK differentiation were seen with expanded primary MKs, bone marrow from 2 patients with myelodysplastic syndrome, and other cell lines with MK potential. Our data suggest that SU6656 might be useful as a differentiation-inducing agent for MKs and is an important tool for understanding the molecular basis of MK endomitosis. PMID:15677565

  20. Autocrine Signaling by Wnt-5a Deregulates Chemotaxis of Leukemic Cells and Predicts Clinical Outcome in Chronic Lymphocytic Leukemia

    Czech Academy of Sciences Publication Activity Database

    Janovská, P.; Poppová, L.; Plevová, K.; Plesingerova, E.; Behal, M.; Kaucká, M.; Ovesná, P.; Hlozkova, M.; Borský, M.; Stehlíková, O.; Brychtová, Y.; Doubek, M.; Machalova, M.; Baskar, S.; Kozubík, Alois; Pospíšilová, Š.; Pavlová, Š.; Bryja, Vítězslav

    2016-01-01

    Roč. 22, č. 2 (2016), s. 459-469 ISSN 1078-0432 Institutional support: RVO:68081707 Keywords : receptor tyrosine kinase * cd38 expression * mutation status * cll cells Subject RIV: BO - Biophysics Impact factor: 9.619, year: 2016

  1. Analysis of the interplay between all-trans retinoic acid and histone deacetylase inhibitors in leukemic cells

    DEFF Research Database (Denmark)

    Noack, Katrin; Mahendrarajah, Nisintha; Hennig, Dorle

    2017-01-01

    The treatment of acute promyelocytic leukemia (APL) with all-trans retinoic acid (ATRA) induces granulocytic differentiation. This process renders APL cells resistant to cytotoxic chemotherapies. Epigenetic regulators of the histone deacetylases (HDACs) family, which comprise four classes (I–IV),...

  2. Nuclear topography of beta-like globin gene cluster in IL-3-stimulated human leukemic K-562 cells

    Czech Academy of Sciences Publication Activity Database

    Galiová-Šustáčková, Gabriela; Bártová, Eva; Kozubek, Stanislav

    2004-01-01

    Roč. 33, č. 1 (2004), s. 4-14 ISSN 1079-9796 R&D Projects: GA ČR GA301/01/0186; GA AV ČR KSK5052113; GA AV ČR IAA5004306; GA ČR GA202/04/0907; GA MŠk ME 565 Institutional research plan: CEZ:AV0Z5004920 Keywords : beta-like globin gene cluster * K-562 cells * nuclear topography Subject RIV: BO - Biophysics Impact factor: 2.549, year: 2004

  3. The effect of extracorporeal photopheresis alone or in combination therapy on circulating CD4+Foxp3+CD25- T-cells in patients with leukemic cutaneous T-cell lymphoma

    Science.gov (United States)

    Shiue, Lisa H.; Couturier, Jacob; Lewis, Dorothy E.; Wei, Caimiao; Ni, Xiao; Duvic, Madeleine

    2015-01-01

    Purpose Extracorporeal photopheresis (ECP) alone or in combination therapy is effective for treatment of leukemic cutaneous T-cell lymphoma (L-CTCL), but its mechanism(s) of action remain unclear. This study was designed to investigate the effect of ECP on regulatory T-cell and CD8+ T-cells in L-CTCL patients. Experimental Design Peripheral blood from 18 L-CTCL patients at baseline, Day 2, 1-month, 3-month, and 6-month post-ECP therapy were analyzed by flow cytometry for CD4+CD25+/high, CD4+Foxp3+CD25+/-, CD3+CD8+, CD3+CD8+CD69+, and CD3+CD8+IFN-γ+ T-cells. Clinical responses were assessed and correlated with changes in these T-cell subsets. Results Twelve of 18 patients achieved clinical responses. The average baseline number of CD4+CD25+/high T-cells of PBMCs in L-CTCL patients was normal (2.2%), but increased at 6-month post-therapy (4.3%, p<0.01). The average baseline number of CD4+Foxp3+ T-cells out of CD4+ T-cells in 9 evaluable patients was high (66.8±13.7%), mostly CD25 negative. The levels of CD4+Foxp3+ T cells in responders were higher (n=6, 93.1±5.7%) than non-responders (n=3, 14.2±16.0%, p<0.01), and they declined in parallel with malignant T-cells. The numbers of CD3+CD8+CD69+ and CD3+CD8+ IFN-γ+ T-cells increased at 3-month post-therapy in 5 of 6 patients studied. Conclusions ECP alone or in combination therapy might be effective in L-CTCL patients whose malignant T-cells have a CD4+Foxp3+CD25- phenotype. PMID:25772268

  4. Novel cytotoxic exhibition mode of antimicrobial peptide anoplin in MEL cells, the cell line of murine Friend leukemia virus-induced leukemic cells.

    Science.gov (United States)

    Zhu, Li-Na; Fu, Cai-Yun; Zhang, Shi-Fu; Chen, Wei; Jin, Yuan-Ting; Zhao, Fu-Kun

    2013-09-01

    Anoplin is a recently discovered antimicrobial peptide (AMP) isolated from the venom sac of the spider wasp Anoplius samariensis, and it is one of the shortest α-helical AMP found naturally to date consisting of only ten amino acids. Previous results showed that anoplin exhibits potent antimicrobial activity but little hemolytic activity. In this study, we synthesized anoplin, studied its cytotoxicity in Friend virus-induced leukemia cells [murine erythroleukemia (MEL) cells], and proposed its possible mechanism. Our results showed that anoplin could inhibit the proliferation of MEL cells in a dose-dependent and time-dependent manner via disrupting the integrity of cell membrane, which indicated that anoplin exerts its cytotoxicity efficacy. In addition, the cell cycle distribution of MEL cells was arrested in the G₀/G₁ phase significantly. However, anoplin could not induce obvious apoptosis in MEL cells, as well as anoplin could not induce visible changes on morphology and quantity in the bone marrow cells isolated from normal mice. All of these results indicate that anoplin, as generally believed, is a selective AMP, a value characteristic in the design of safe therapeutic agents. The cytotoxicity of anoplin on MEL cells was mainly attributable to the plasma membrane perturbation and also to the intracellular events such as the arrest of cell cycle. Although this is an initial study that explored the activity of anoplin in vitro rather than in vivo, with the increasing resistance of conventional chemotherapy, there is no doubt that anoplin has desirable feature to be developed as a novel and selective anticancer agent. Copyright © 2013 European Peptide Society and John Wiley & Sons, Ltd.

  5. The novel anticancer agent JNJ-26854165 is active in chronic myeloid leukemic cells with unmutated BCR/ABL and T315I mutant BCR/ABL through promoting proteosomal degradation of BCR/ABL proteins.

    Science.gov (United States)

    You, Liangshun; Liu, Hui; Huang, Jian; Xie, Wanzhuo; Wei, Jueying; Ye, Xiujin; Qian, Wenbin

    2017-01-31

    Chronic myeloid leukemia (CML) is a clonal malignant disease caused by the expression of BCR/ABL. MDM2 (human homolog of the murine double minute-2) inhibitors such as Nutlin-3 have been shown to induce apoptosis in a p53-dependent manner in CML cells and sensitize cells to Imatinib. Here, we demonstrate that JNJ-26854165, an inhibitor of MDM2, inhibits proliferation and triggers cell death in a p53-independent manner in various BCR/ABL-expressing cells, which include primary leukemic cells from patients with CML blast crisis and cells expressing the Imatinib-resistant T315I BCR/ABL mutant. The response to JNJ-26854165 is associated with the downregulation of BCR/ABL dependently of proteosome activation. Moreover, in all tested CML cells, with the exception of T315I mutation cells, combining JNJ-26854165 and tyrosine kinase inhibitor (TKI) Imatinib or PD180970 leads to a synergistic effect. In conclusion, our results suggest that JNJ-26854165, used either alone or in combination with TKIs, represents a promising novel targeted approach to overcome TKI resistance and improve patient outcome in CML.

  6. Interaction of leukemic cells with proteins of the extracellular matrix Interações de células leucêmicas com proteínas da matriz extracelular

    Directory of Open Access Journals (Sweden)

    Adriana Rodrigues-Anjos

    2004-01-01

    Full Text Available The interaction of neoplastic cells with basement membrane molecules is the first step for the dissemination of tumor cells in vivo. Leukemic cells have a great ability to spread in the host, since cells are released from the bone marrow to the circulation. In this study we analysed whether CEM, U937, K562 and HL-60 cells were able to attach to different concentrations of laminin and/or fibronectin and/or type IV collagen. Attachment to type IV collagen was low, but it increased with the addition of laminin and occurred in all four leukemic cell lines. On the other hand, attachment to fibronectin was higher, but it decreased with the addition of laminin in the assays using U937 and HL-60 cells. The combination of type IV collagen and fibronectin was a good substratum for cellular attachment. However, the addition of laminin to this substratum impaired its attachment activity in U937, HL-60 and K562. These data suggest that laminin may control cellular attachment to the extracellular matrix during leukemic dissemination in hosts in different ways.A interação de células neoplásicas com moléculas da membrana basal é a primeira etapa para a disseminação, in vivo, de células tumorais in vivo. As células leucêmicas possuem grande capacidade de espraiamento e disseminação no organismo uma vez que as mesmas são liberadas da medula óssea para a circulação. Neste trabalho avaliamos a capacidade das linhagens celulares CEM, U937, K562 and HL-60 em aderirem a uma matriz extracelular constituída por diferentes concentrações de laminina e,ou fibronectina e sobre colágeno IV. A adesão de todas a linhagens leucêmicas a colágeno IV foi baixa, mas aumentou com a associação à laminina. Por outro lado, as células U937 e HL-60 apresentaram alta ligação à fibronectina porém, foi reduzida com a adição de laminina. A associação de colágeno IV e fibronectina possibilitou um bom substrato para a adesão celular. Entretanto, a adi

  7. Specific inhibition of Wee1 kinase and Rad51 recombinase: A strategy to enhance the sensitivity of leukemic T-cells to ionizing radiation-induced DNA double-strand breaks

    International Nuclear Information System (INIS)

    Havelek, Radim; Cmielova, Jana; Kralovec, Karel; Bruckova, Lenka; Bilkova, Zuzana; Fousova, Ivana; Sinkorova, Zuzana; Vavrova, Jirina; Rezacova, Martina

    2014-01-01

    Highlights: • Pre-treatment with the inhibitors increased the sensitivity of Jurkat cells to irradiation. • Combining both inhibitors together resulted in a G2 cell cycle arrest abrogation in Jurkat. • Jurkat cells pre-treated with inhibitors were positive for γH2AX foci 24 h upon irradiation. • Pre-treatment with Rad51 RI-1 had no effect on apoptosis induction in MOLT-4 cells. • When dosed together, the combination decreased MOLT-4 cell survival. - Abstract: Present-day oncology sees at least two-thirds of cancer patients receiving radiation therapy as a part of their anticancer treatment. The objectives of the current study were to investigate the effects of the small molecule inhibitors of Wee1 kinase II (681641) and Rad51 (RI-1) on cell cycle progression, DNA double-strand breaks repair and apoptosis following ionizing radiation exposure in human leukemic T-cells Jurkat and MOLT-4. Pre-treatment with the Wee1 681641 or Rad51 RI-1 inhibitor alone increased the sensitivity of Jurkat cells to irradiation, however combining both inhibitors together resulted in a further enhancement of apoptosis. Jurkat cells pre-treated with inhibitors were positive for γH2AX foci 24 h upon irradiation. MOLT-4 cells were less affected by inhibitors application prior to ionizing radiation exposure. Pre-treatment with Rad51 RI-1 had no effect on apoptosis induction; however Wee1 681641 increased ionizing radiation-induced cell death in MOLT-4 cells

  8. Quantitative assay for the number of leukemic spleen colony forming unit in radiation-induced murine myeloid leukemia

    Energy Technology Data Exchange (ETDEWEB)

    Nara, N [Tokyo Medical and Dental Univ. (Japan). School of Medicine; Bessho, M

    1981-11-01

    In mice with myelogenous leukemia, leukemic spleen colony forming units were assayed quantitatively. When 5 x 10/sup 3/ - 2 x 10/sup 4/ leukemic cells were transplanted to other mice of the same strain, a rectilinear relationship (p < 0.01) was found between the number of the cells transplanted and that of the colonies formed on the surface of the spleen. From these results, the authors considered that myelogenous leukemia in mice is an adequate model for acute myelogenous leukemia in human adults, and that the quantitative assay of the leukemic colony forming units can be used for sensitivity tests of antileukemic agents.

  9. Patient-Derived Antibody Targets Tumor Cells

    Science.gov (United States)

    An NCI Cancer Currents blog on an antibody derived from patients that killed tumor cells in cell lines of several cancer types and slowed tumor growth in mouse models of brain and lung cancer without evidence of side effects.

  10. 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...

  11. Phenotypic and Functional Alterations of Hematopoietic Stem and Progenitor Cells in an In Vitro Leukemia-Induced Microenvironment

    Directory of Open Access Journals (Sweden)

    Jean-Paul Vernot

    2017-02-01

    Full Text Available An understanding of the cell interactions occurring in the leukemic microenvironment and their functional consequences for the different cell players has therapeutic relevance. By co-culturing mesenchymal stem cells (MSC with the REH acute lymphocytic leukemia (ALL cell line, we have established an in vitro leukemic niche for the functional evaluation of hematopoietic stem/progenitor cells (HSPC, CD34+ cells. We showed that the normal homeostatic control exerted by the MSC over the HSPC is considerably lost in this leukemic microenvironment: HSPC increased their proliferation rate and adhesion to MSC. The adhesion molecules CD54 and CD44 were consequently upregulated in HSPC from the leukemic niche. Consequently, with this adhesive phenotype, HSPC showed less Stromal derived factor-1 (SDF-1-directed migration. Interestingly, multipotency was severely affected with an important reduction in the absolute count and the percentage of primitive progenitor colonies. It was possible to simulate most of these HSPC alterations by incubation of MSC with a REH-conditioned medium, suggesting that REH soluble factors and their effect on MSC are important for the observed changes. Of note, these HSPC alterations were reproduced when primary leukemic cells from an ALL type B (ALL-B patient were used to set up the leukemic niche. These results suggest that a general response is induced in the leukemic niche to the detriment of HSPC function and in favor of leukemic cell support. This in vitro leukemic niche could be a valuable tool for the understanding of the molecular events responsible for HSPC functional failure and a useful scenario for therapeutic evaluation.

  12. Protodioscin, Isolated from the Rhizome of Dioscorea tokoro Collected in Northern Japan is the Major Antiproliferative Compound to HL-60 
Leukemic Cells.

    Science.gov (United States)

    Oyama, Manami; Tokiwano, Tetsuo; Kawaii, Satoru; Yoshida, Yasunori; Mizuno, Kouichi; Oh, Keimei; Yoshizawa, Yuko

    2017-06-01

    The rhizome of Oni-dokoro (a wild yam, Dioscorea tokoro) has extremely bitter taste and is not generally regarded edible;, however, in northern part of Japan, such as Iwate and a part of Aomori, it is used as health promoting food. To clarify the reason, we examined the biologically active compounds in the rhizome collected at Iwate and compared them from the other area in literature. The acetonitrile extract from northern part of Japan was purified by bioassay-guided separation using antiproliferative activity to human leukemia HL-60 cell, and protodioscin (PD) was isolated and identified by instrumental analyses as the major active compound. PD known as a saponin with four sugar moieties, an inhibitor for platelet aggregation, and a low density lipoprotein (LPL) lowering agent, displayed strong growth inhibitory effect to HL-60. The literature search suggested that the rhizome from other area contained dioscin and other saponins with three sugar moieties as their major component. We assume that the edible and health promoting effect of the rhizome in the particular area is partially derived from these different components. We were interested in the differences of utilization in the rhizome of wild yam Dioscorea tokoro, and examined the chemical composition in the rhizome to find protodioscin as antiproliferative compound to HL-60. In the report from other area, the rhizome exhibited dioscin as the major compound. Our study indicated that the protodioscin/dioscin composition varied regionally, although the reason is still needs to be investigated.

  13. An antigen shared by human granulocytes, monocytes, marrow granulocyte precursors and leukemic blasts.

    Science.gov (United States)

    Shumak, K H; Rachkewich, R A

    1983-01-01

    An antibody to human granulocytes was raised in rabbits by immunization with granulocytes pretreated with rabbit antibody to contaminating antigens. The antibody reacted not only with granulocytes but also with monocytes and bone marrow granulocyte precursors including colony-forming units in culture (CFU-C). In tests with leukemic cells, the antibody reacted with blasts from most (8 of 9) patients with acute myelomonoblastic leukemia and from some patients with acute myeloblastic leukemia, morphologically undifferentiated acute leukemia and chronic myelogenous leukemia in blast crisis. The antibody did not react with blasts from patients with acute lymphoblastic leukemia nor with leukemic cells from patients with chronic lymphocytic leukemia.

  14. Adipose-Derived Stem Cells

    DEFF Research Database (Denmark)

    Toyserkani, Navid Mohamadpour; Quaade, Marlene Louise; Sheikh, Søren Paludan

    2015-01-01

    , regulate the inflammatory process, and differentiate into multiple cell types makes them a potential ideal therapy for chronic wounds. The aim of this article was to review all preclinical trials using ASCs in problem wound models. A systematic search was performed and 12 studies were found where different...

  15. Adipose-Derived Stem Cells

    NARCIS (Netherlands)

    Gathier, WA; Türktas, Z; Duckers, HJ

    2015-01-01

    Until recently bone marrow was perceived to be the only significant reservoir of stem cells in the body. However, it is now recognized that there are other and perhaps even more abundant sources, which include adipose tissue. Subcutaneous fat is readily available in most patients, and can easily be

  16. History of myeloid-derived suppressor cells.

    Science.gov (United States)

    Talmadge, James E; Gabrilovich, Dmitry I

    2013-10-01

    Tumour-induced granulocytic hyperplasia is associated with tumour vasculogenesis and escape from immunity via T cell suppression. Initially, these myeloid cells were identified as granulocytes or monocytes; however, recent studies have revealed that this hyperplasia is associated with populations of multipotent progenitor cells that have been identified as myeloid-derived suppressor cells (MDSCs). The study of MDSCs has provided a wealth of information regarding tumour pathobiology, has extended our understanding of neoplastic progression and has modified our approaches to immune adjuvant therapy. In this Timeline article, we discuss the history of MDSCs, their influence on tumour progression and metastasis, and the crosstalk between tumour cells, MDSCs and the host macroenvironment.

  17. Cyanobacteria as a Source for Novel Anti-Leukemic Compounds.

    Science.gov (United States)

    Humisto, Anu; Herfindal, Lars; Jokela, Jouni; Karkman, Antti; Bjørnstad, Ronja; Choudhury, Romi R; Sivonen, Kaarina

    2016-01-01

    Cyanobacteria are an inspiring source of bioactive secondary metabolites. These bioactive agents are a diverse group of compounds which are varying in their bioactive targets, the mechanisms of action, and chemical structures. Cyanobacteria from various environments, especially marine benthic cyanobacteria, are found to be rich sources for the search for novel bioactive compounds. Several compounds with anticancer activities have been discovered from cyanobacteria and some of these have succeeded to enter the clinical trials. Varying anticancer agents are needed to overcome increasing challenges in cancer treatments. Different search methods are used to reveal anticancer compounds from natural products, but cell based methods are the most common. Cyanobacterial bioactive compounds as agents against acute myeloid leukemia are not well studied. Here we examined our new results combined with previous studies of anti-leukemic compounds from cyanobacteria with emphasis to reveal common features in strains producing such activity. We report that cyanobacteria harbor specific anti-leukemic compounds since several studied strains induced apoptosis against AML cells but were inactive against non-malignant cells like hepatocytes. We noted that particularly benthic strains from the Baltic Sea, such as Anabaena sp., were especially potential AML apoptosis inducers. Taken together, this review and re-analysis of data demonstrates the power of maintaining large culture collections for the search for novel bioactivities, and also how anti-AML activity in cyanobacteria can be revealed by relatively simple and low-cost assays.

  18. Influence of x-rays and UV-light on the presence of oncogene proteins in spleen cells of leukemic mice

    International Nuclear Information System (INIS)

    Popovic Hadzija, M.; Poljak Blazi, M.

    1996-01-01

    Proto-oncogenes are involved in growth, defferentiation and proliferation of normal cells, and in process of neoplastic transformation. In genome of normal cells, exist also tumor-suppressor genes, which contribute to cancer when they are inactivated. Those genes are target for carcinogenesis provoked by radiation. However, species specific genetic factors are important in determing which, if any, gene will be transformed by radiation. It is possible to presume that oncogenes are involved in the development of radioresistant phenotype of ML. Because of that, we examined the presence of c-myc protein in ML cells during the growth of ML and after the irradiation of these cells. Also, we examined the presence of tumor-suppressor protein p53, because inactivation or loss of p53 gene is in connection with transformation of cells. ML is strain specific for RFM mice. Spleen cells were tested 9 (nonterminal phase NTP) or 12 days (terminal phase TP) after inoculation of ML. Cells from NTP were also irradiate with x-rays or UV-light. C-myc protein expresse 74.98% spleen cells of healthy RFM mice. Wild type of p53 protein was detected in 60% of these cells, but mp53 was found in only 5.3% of cells. These results could be explained by the role of c-myc and p53 proteins in regulation of biologic processes. A few spleen cells of NTP expressed c-myc (15%) and mp53 (9.6%) proteins. But, in the same phase higher expressions of wp53 protein (30.5%) was found. On the other hand, the number of c-myc positive cells in TP of leukemia explanation lies in connection of c-myc protein and process of programmed cell death (apoptosis). During growth of ML the number of mp53 positive cells increased (to 47.8%), but wp53 positive cells decreased (to13.4%9). Both types of irradiation provoked strong activation of cellular c-myc gene in ML cells of NTP. We found about 95% c-myc positive cells after x-rays and 93% after UV-light

  19. 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.

  20. 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

  1. Radiobiological heterogeneity of leukemic lymphocyte precursors from acute lymphoblastic leukemia patients

    International Nuclear Information System (INIS)

    Uckun, F.M.; Kim, T.H.; Ramsay, N.C.; Min, W.S.; Song, C.W.

    1989-01-01

    The report outlines the authors' findings on the radiobiological features of leukemic lymphocyte precursors from acute lymphoblastic leukemia (ALL) patients. A marked heterogeneity existed between different cell lines, with a remarkable radioresistance and repair capacity in some ALL patients and an acute radiosensitivity in the absence of a detectable repair capacity in others. (U.K.)

  2. Correlation of chromosome patterns in human leukemic cells with exposure to chemicals and/or radiation. Progress report, January 1-July 31, 1986

    International Nuclear Information System (INIS)

    Rowley, J.D.

    1986-08-01

    Two genes for colony stimulating factors (CSF) has been mapped on chromosome five of humans. These CSFs are a family of glycoproteins that are required for the growth and maturation of myeloid progenetor cells in vitro. They are classified according to their cell specificity; thus, M-CSF (CSF-1) and G-CSF primarily stimulate cells committed to the macrophage and granulocyte lineages, respectively. Recently, the genes for both of these factors have been cloned and probes for the clonal genes were used to map their location on normal human chromosomes. Localization of CSF-1 was performed by in situ hybridization of the probe pST-CSF-17 to normal human metaphase chromosomes. This resulted in specific labeling only of chromosome 5. 8 refs., 4 figs., 2 tabs

  3. JS-K, a nitric oxide-releasing prodrug, modulates ß-catenin/TCF signaling in leukemic Jurkat cells: evidence of an S-nitrosylated mechanism.

    Science.gov (United States)

    Nath, Niharika; Chattopadhyay, Mitali; Pospishil, Liliya; Cieciura, Lucyna Z; Goswami, Satindra; Kodela, Ravinder; Saavedra, Joseph E; Keefer, Larry K; Kashfi, Khosrow

    2010-12-01

    β-Catenin is a central player of the Wnt signaling pathway that regulates cell-cell adhesion and may promote leukemia cell proliferation. We examined whether JS-K, an NO-donating prodrug, modulates the Wnt/β-catenin/TCF-4 signaling pathway in Jurkat T-Acute Lymphoblastic Leukemia cells. JS-K inhibited Jurkat T cell growth in a concentration and time-dependent manner. The IC(50)s for cell growth inhibition were 14±0.7 and 9±1.2μM at 24 and 48h, respectively. Treatment of the cells with JS-K for 24h, caused a dose-dependent increase in apoptosis from 16±3.3% at 10μM to 74.8±2% at 100μM and a decrease in proliferation. This growth inhibition was also due, in part, to alterations in the different phases of the cell cycle. JS-K exhibited a dose-dependent cytotoxicity as measured by LDH release at 24h. However, between 2 and 8h, LDH release was less than 20% for any indicated JS-K concentration. The β-catenin/TCF-4 transcriptional inhibitory activity was reduced by 32±8, 63±5, and 93±2% at 2, 10, and 25μM JS-K, respectively, based on luciferase reporter assays. JS-K reduced nuclear β-catenin and cyclin D1 protein levels, but cytosolic β-catenin expression did not change. Based on a time-course assay of S-nitrosylation of proteins by a biotin switch assay, S-nitrsolyation of nuclear β-catenin was determined to precede its degradation. A comparison of the S-nitrosylated nuclear β-catenin to the total nuclear β-catenin showed that β-catenin protein levels were degraded at 24h, while S-nitrosylation of β-catenin occurred earlier at 0-6h. The NO scavenger PTIO abrogated the JS-K mediated degradation of β-catenin demonstrating the need for NO. Copyright © 2010 Elsevier Inc. All rights reserved.

  4. Differential Effects of Tea Extracts on Growth and Cytokine Production by Normal and Leukemic Human Leukocytes

    Directory of Open Access Journals (Sweden)

    Diana Bayer

    2012-04-01

    Full Text Available Background: Tea is one of the world’s most highly consumed beverages, second only to water. It is affordable and abundant and thus has great potential for improving health of those in both developed and developing areas. Green, oolong, and black teas differ in the extent of fermentation and types of bioactive polyphenols produced. Green tea and its major polyphenol decrease growth of some cancer cells and effect production of immune system cytokines. This study compares the effects of different types of tea extracts on viability and cytokine production by normal and leukemic human T lymphocytes. Generation of the toxic reactive oxygen species H2O2 by extracts was also examined.Methods: The Jurkat T lymphoblastic leukemia cells and mitogen-stimulated normal human peripheral blood mononuclear cells were used in this study. Cell viability was determined by (3-4,5-dimethylthiamizol-2-yl-diphenyltetrazolium bromide assay and production of interleukin-2 by Enzyme-Linked ImmunoSorbent Assay. Levels of H2O2 generated by tea extracts were determined using the xylenol-orange method.Results: We found that green, oolong, and black tea extracts differentially effect the growth and viability of T lymphoblastic leukemia cells and normal peripheral blood mononuclear cells, substantially decreasing both growth and viability of leukemic T lymphocytes and having much lesser effects on their normal counterparts. Tea extracts also had differential effects on the production of the T lymphocyte growth factor interleukin-2, significantly decreasing production by leukemic cells while having only minor effects on normal cells. All three extracts induced H2O2 generation, with green and oolong tea extracts having the greatest effect. Leukemic cells were much more susceptible to growth inhibition and killing by H2O2 than normal lymphocytes.Functional Foods in Health and Disease 2012, 2(4:72-85 Conclusions: The three tea extracts studied altered leukemic T lymphocyte

  5. Effects of extremely low frequency electromagnetic field (ELF-EMF) on catalase, cytochrome P450 and nitric oxide synthase in erythro-leukemic cells.

    Science.gov (United States)

    Patruno, Antonia; Tabrez, Shams; Pesce, Mirko; Shakil, Shazi; Kamal, Mohammad A; Reale, Marcella

    2015-01-15

    Extremely low frequency electromagnetic fields (ELF-EMFs) are widely employed in electrical appliances and different equipment such as television sets, mobile phones, computers and microwaves. The molecular mechanism through which ELF-EMFs can influence cellular behavior is still unclear. A hypothesis is that ELF-EMFs could interfere with chemical reactions involving free radical production. Under physiologic conditions, cells maintain redox balance through production of ROS/RNS and antioxidant molecules. The altered balance between ROS generation and elimination plays a critical role in a variety of pathologic conditions including neurodegenerative diseases, aging and cancer. Actually, there is a disagreement as to whether there is a causal or coincidental relationship between ELF-EMF exposure and leukemia development. Increased ROS levels have been observed in several hematopoietic malignancies including acute and chronic myeloid leukemias. In our study, the effect of ELF-EMF exposure on catalase, cytochrome P450 and inducible nitric oxide synthase activity and their expression by Western blot analysis in myelogenous leukemia cell line K562 was evaluated. A significant modulation of iNOS, CAT and Cyt P450 protein expression was recorded as a result of ELF-EMF exposure in both phorbol 12-myristate 13-acetate (PMA)-stimulated and non-stimulated cell lines. Modulation in kinetic parameters of CAT, CYP-450 and iNOS enzymes in response to ELF-EMF indicates an interaction between the ELF-EMF and the enzymological system. These new insights might be important in establishing a mechanistic framework at the molecular level within which the possible effects of ELF-EMF on health can be understood. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Analysis of Tn antigenicity with a panel of new IgM and IgG1 monoclonal antibodies raised against leukemic cells

    DEFF Research Database (Denmark)

    Blixt, Klas Ola; Lavrova, Olga I; Mazurov, Dmitriy V

    2012-01-01

    antigenicity remain poorly understood. As a consequence, a broad variety of anti-Tn monoclonal antibodies (mAbs) have been generated. To further investigate the nature and complexity of the Tn antigen, we generated seven different anti-Tn mAbs of IgM and IgG classes raised against human Jurkat T cells, which...... are Tn-positive due to the low activity of T-synthase and mutation in specific chaperone Cosmc. The binding analysis of anti-Tn mAbs with the array of synthetic saccharides, glycopeptides and O-glycoproteins revealed unexpected differences in specificities of anti-Tn mAbs. IgM mAbs bound the terminal Gal...

  7. Quantification of thymidine kinase (TK1) mRNA in normal and leukemic cells and investigation of structure-function relatiosnhip of recombinant TK1enzyme

    DEFF Research Database (Denmark)

    Kristensen, Tina

    Thymidine kinase (TK) catalyses the ATP-dependent phosphorylation of thymidine to thymidine monophosphate, which is subsequency phosphorylated to thymidine triphosphate and utilized for DNA synthesis. Human cytosolic TK (TKI) is cell cycle regulated, e.g. the TK1 activity increases sharply at the G...... patients with chronic lymphatic leukemia (CLL). 2: Structure-function relationship of recombinant TKI. In the first part a sensitive method (competitive PCR) for quantification of TKI mRNA was established. The TKI mRNA level was quantified in quiescent lymphocytes from control donors (n = 6...... are characterized as being quiescent, the TK activity was in the same range as in quiescent lymphocytes from control donors. However, quantification of the TKI mRNA level shows that all five CLL patients had a very high level (6 to 22 x IO6 copies mg-’ protein) of TKI mRNA, corresponding to the level in dividing...

  8. Correlation of chromosome patterns in human leukemic cells with exposure to chemicals and/or radiation: Comprehensive progress report, January 1986--June 1988

    International Nuclear Information System (INIS)

    Rowley, J.D.

    1988-06-01

    I purchased one of the few available prototypes of the pulse field gel electrophoresis (PFGE) apparatus. We used PFGE and its various modifications to map the human Abelson protooncogene (ABL) and to show that the two alternative first exons (Ia and Ib) are separated by at least 200 kilobases (kb). This has provided the first evidence that alternative splicing from exon Ib to the common splice acceptor site (exon II) could occur over such very large distances. We are actively using vertical field gel electrophoresis, a modification of PFGE, for mapping various DNA probes on chromosome 5. Another major advance has been the development of the polymerase chain reaction (PCR). We are currently using this to define the breakpoints in the BCR gene in the 9; 22 translocation in chronic myeloid leukemia (CML) and in Ph 1 -positive acute lymphoblastic leukemia (ALL). I had expected to be able to describe major progress in cloning the chromosome translocation breakpoints in ANLL, and this has not occurred. Our laboratory knows how to solve the problem. We successfully cloned a new translocation breakpoint in B cell chronic lymphatic leukemia involving Nos. 14 and 19. 22 refs., 2 figs., 5 tabs

  9. Systemic T Cells Immunosuppression of Glioma Stem Cell-Derived Exosomes Is Mediated by Monocytic Myeloid-Derived Suppressor Cells.

    Directory of Open Access Journals (Sweden)

    Rossana Domenis

    Full Text Available A major contributing factor to glioma development and progression is its ability to evade the immune system. Nano-meter sized vesicles, exosomes, secreted by glioma-stem cells (GSC can act as mediators of intercellular communication to promote tumor immune escape. Here, we investigated the immunomodulatory properties of GCS-derived exosomes on different peripheral immune cell populations. Healthy donor peripheral blood mononuclear cells (PBMCs stimulated with anti-CD3, anti-CD28 and IL-2, were treated with GSC-derived exosomes. Phenotypic characterization, cell proliferation, Th1/Th2 cytokine secretion and intracellular cytokine production were analysed by distinguishing among effector T cells, regulatory T cells and monocytes. In unfractionated PBMCs, GSC-derived exosomes inhibited T cell activation (CD25 and CD69 expression, proliferation and Th1 cytokine production, and did not affect cell viability or regulatory T-cell suppression ability. Furthermore, exosomes were able to enhance proliferation of purified CD4+ T cells. In PBMCs culture, glioma-derived exosomes directly promoted IL-10 and arginase-1 production and downregulation of HLA-DR by unstimulated CD14+ monocytic cells, that displayed an immunophenotype resembling that of monocytic myeloid-derived suppressor cells (Mo-MDSCs. Importantly, the removal of CD14+ monocytic cell fraction from PBMCs restored T-cell proliferation. The same results were observed with exosomes purified from plasma of glioblastoma patients. Our results indicate that glioma-derived exosomes suppress T-cell immune response by acting on monocyte maturation rather than on direct interaction with T cells. Selective targeting of Mo-MDSC to treat glioma should be considered with regard to how immune cells allow the acquirement of effector functions and therefore counteracting tumor progression.

  10. 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.

  11. Characterization Of Bovine Adipose-Derived Stem Cells

    OpenAIRE

    Daniel Cebo

    2017-01-01

    Bovine adipose-derived stem cells were obtained from the subcutaneous abdominal adipose tissue. The cells were cultured by the modified tissue-explants method developed in our laboratory and then analyzed using optical microscopy and flow cytometry. These cells were able to replicate in our cell culture conditions. cell Flow cytometry showed that bovine adipose-derived stem cells expressed mesenchymal stem cell markers CD73 and CD90. Meanwhile haematopoietic markers CD45 and CD34 are absent f...

  12. Effects of granulocyte-macrophage colony-stimulating factor and interleukin 6 on the growth of leukemic blasts in suspension culture.

    Science.gov (United States)

    Tsao, C J; Cheng, T Y; Chang, S L; Su, W J; Tseng, J Y

    1992-05-01

    We examined the stimulatory effects of recombinant human granulocyte-macrophage colony-stimulating factor (GM-CSF) and interleukin 6 (IL)-6 on the in vitro proliferation of leukemic blast cells from patients with acute leukemia. Bone marrow or peripheral blood leukemic blast cells were obtained from 21 patients, including 14 cases of acute myeloblastic leukemia (AML), four cases of acute lymphoblastic leukemia (ALL), two cases of acute undifferentiated leukemia, and one case of acute mixed-lineage leukemia. The proliferation of leukemic blast cells was evaluated by measuring the incorporation of 3H-thymidine into cells incubated with various concentrations of cytokines for 3 days. GM-CSF stimulated the DNA synthesis (with greater than 2.0 stimulation index) of blast cells in 9 of 14 (64%) AML cases, two cases of acute undifferentiated leukemia and one case of acute mixed-lineage leukemia. Only two cases of AML blasts responded to IL-6 to grow in the short-term suspension cultures. GM-CSF and IL-6 did not display a synergistic effect on the growth of leukemic cells. Moreover, GM-CSF and IL-6 did not stimulate the proliferation of ALL blast cells. Binding study also revealed the specific binding of GM-CSF on the blast cells of acute undifferentiated leukemia and acute mixed-lineage leukemia. Our results indicated that leukemic blast cells of acute undifferentiated leukemia and acute mixed-lineage leukemia possessed functional GM-CSF receptors.

  13. Adipose-derived stem cells for treatment of chronic ulcers

    DEFF Research Database (Denmark)

    Holm, Jens Selch; Toyserkani, Navid Mohamadpour; Sorensen, Jens Ahm

    2018-01-01

    Chronic ulcers remain a difficult challenge in healthcare systems. While treatment options are limited, stem cells may be a novel alternative. Adipose-derived stem cells (ADSC) have become increasingly popular compared with bone marrow-derived stem cells as they are far easier to harvest...

  14. 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

  15. Stromal cells expressing hedgehog-interacting protein regulate the proliferation of myeloid neoplasms

    International Nuclear Information System (INIS)

    Kobune, M; Iyama, S; Kikuchi, S; Horiguchi, H; Sato, T; Murase, K; Kawano, Y; Takada, K; Ono, K; Kamihara, Y; Hayashi, T; Miyanishi, K; Sato, Y; Takimoto, R; Kato, J

    2012-01-01

    Aberrant reactivation of hedgehog (Hh) signaling has been described in a wide variety of human cancers including cancer stem cells. However, involvement of the Hh-signaling system in the bone marrow (BM) microenvironment during the development of myeloid neoplasms is unknown. In this study, we assessed the expression of Hh-related genes in primary human CD34 + cells, CD34 + blastic cells and BM stromal cells. Both Indian Hh (Ihh) and its signal transducer, smoothened (SMO), were expressed in CD34 + acute myeloid leukemia (AML) and myelodysplastic syndrome (MDS)-derived cells. However, Ihh expression was relatively low in BM stromal cells. Remarkably, expression of the intrinsic Hh-signaling inhibitor, human Hh-interacting protein (HHIP) in AML/MDS-derived stromal cells was markedly lower than in healthy donor-derived stromal cells. Moreover, HHIP expression levels in BM stromal cells highly correlated with their supporting activity for SMO + leukemic cells. Knockdown of HHIP gene in stromal cells increased their supporting activity although control cells marginally supported SMO + leukemic cell proliferation. The demethylating agent, 5-aza-2′-deoxycytidine rescued HHIP expression via demethylation of HHIP gene and reduced the leukemic cell-supporting activity of AML/MDS-derived stromal cells. This indicates that suppression of stromal HHIP could be associated with the proliferation of AML/MDS cells

  16. Heterogeneity of clonogenic cells in acute myeloblastic leukemia.

    Science.gov (United States)

    Sabbath, K D; Ball, E D; Larcom, P; Davis, R B; Griffin, J D

    1985-02-01

    The expression of differentiation-associated surface antigens by the clonogenic leukemic cells from 20 patients with acute myeloblastic leukemia (AML) was studied with a panel of seven cytotoxic monoclonal antibodies (anti-Ia, -MY9, -PM-81, -AML-2-23, -Mol, -Mo2, and -MY3). The surface antigen phenotypes of the clonogenic cells were compared with the phenotypes of the whole leukemic cell population, and with the phenotypes of normal hematopoietic progenitor cells. In each case the clonogenic leukemic cells were found within a distinct subpopulation that was less "differentiated" than the total cell population. Clonogenic leukemic cells from different patients could be divided into three phenotype groups. In the first group (7 of 20 cases), the clonogenic cells expressed surface antigens characteristic of the normal multipotent colony-forming cell (Ia, MY9). These cases tended to have "undifferentiated" (FAB M1) morphology, and the total cell population generally lacked expression of "late" monocyte antigens such as MY3 and Mo2. A second group (seven cases) of clonogenic cells expressed surface antigens characteristic of an "early" (day 14) colony-forming unit granulocyte-monocyte (CFU-GM), and a third group (six cases) was characteristic of a "late" (day 7) CFU-GM. The cases in these latter two groups tended to have myelomonocytic (FAB M4) morphology and to express monocyte surface antigens. These results suggest that the clonogenic cells are a distinct subpopulation in all cases of AML, and may be derived from normal hematopoietic progenitor cells at multiple points in the differentiation pathway. The results further support the possibility that selected monoclonal antibodies have the potential to purge leukemic clonogenic cells from bone marrow in some AML patients without eliminating critical normal progenitor cells.

  17. Graphene and its derivatives for cell biotechnology.

    Science.gov (United States)

    Yang, Mei; Yao, Jun; Duan, Yixiang

    2013-01-07

    Every few years, a novel material with salient and often unique properties emerges and attracts both academic and industrial interest from the scientific community. The latest blockbuster is graphene, an increasingly important nanomaterial with atomically thin sheets of carbon, which has become a shining star and has shown great promise in the field of material science and nanotechnology. In recent years, it has changed from being the exclusive domain of physicists to the new passion of chemists and biologists. Graphene and its derivatives are now at the forefront of nearly every rapidly developing field of science and engineering, including biochemistry, biomedicine and certain cutting-edge interdisciplines that have intense popularity. The aim of this review is, firstly, to provide readers with a comprehensive, systematic and in-depth prospective of graphene's band structure and properties, and secondly, to concentrate on the recent progress in producing graphene-based nanomaterials, including mechanical exfoliation, chemical vapor deposition, plasma enhanced chemical vapor deposition, chemical reduction of graphene oxide, total organic synthesis, electrochemical synthesis and other fabrication strategies widely accepted by research scientists. At the same time, important definitions related to graphene are also introduced. The focus of this Tutorial Review is to emphasize the current situation and significance of using this new kind of two-dimensional material in the hot and emerging fields that are closely related to human life quality, for instance, cell biochemistry, bioimaging along with other frontier areas. Finally, the latest developments and possible impact that affect the heart of the whole scientific community have been discussed. In addition, the future trends along with potential challenges of this rapidly rising layered carbon have been pointed out in this paper.

  18. Anterior cruciate ligament-derived cells have high chondrogenic potential.

    Science.gov (United States)

    Furumatsu, Takayuki; Hachioji, Motomi; Saiga, Kenta; Takata, Naoki; Yokoyama, Yusuke; Ozaki, Toshifumi

    2010-01-01

    Anterior cruciate ligament (ACL)-derived cells have a character different from medial collateral ligament (MCL)-derived cells. However, the critical difference between ACL and MCL is still unclear in their healing potential and cellular response. The objective of this study was to investigate the mesenchymal differentiation property of each ligament-derived cell. Both ligament-derived cells differentiated into adipogenic, osteogenic, and chondrogenic lineages. In chondrogenesis, ACL-derived cells had the higher chondrogenic property than MCL-derived cells. The chondrogenic marker genes, Sox9 and alpha1(II) collagen (Col2a1), were induced faster in ACL-derived pellets than in MCL-derived pellets. Sox9 expression preceded the increase of Col2a1 in both pellet-cultured cells. However, the expression level of Sox9 and a ligament/tendon transcription factor Scleraxis did not parallel the increase of Col2a1 expression along with chondrogenic induction. The present study demonstrates that the balance between Sox9 and Scleraxis have an important role in the chondrogenic differentiation of ligament-derived cells. Copyright 2009 Elsevier Inc. All rights reserved.

  19. Comparison of human adipose-derived stem cells and bone marrow-derived stem cells in a myocardial infarction model

    DEFF Research Database (Denmark)

    Rasmussen, Jeppe; Frøbert, Ole; Holst-Hansen, Claus

    2014-01-01

    Background: Treatment of myocardial infarction with bone marrow-derived mesenchymal stem cells and recently also adipose-derived stem cells has shown promising results. In contrast to clinical trials and their use of autologous bone marrow-derived cells from the ischemic patient, the animal...... myocardial infarction models are often using young donors and young, often immune-compromised, recipient animals. Our objective was to compare bone marrow-derived mesenchymal stem cells with adipose-derived stem cells from an elderly ischemic patient in the treatment of myocardial infarction, using a fully...... grown non-immunecompromised rat model. Methods: Mesenchymal stem cells were isolated from adipose tissue and bone marrow and compared with respect to surface markers and proliferative capability. To compare the regenerative potential of the two stem cell populations, male Sprague-Dawley rats were...

  20. C7a, a Biphosphinic Cyclopalladated Compound, Efficiently Controls the Development of a Patient-Derived Xenograft Model of Adult T Cell Leukemia/Lymphoma

    Directory of Open Access Journals (Sweden)

    Carlos R. Figueiredo

    2011-07-01

    Full Text Available Adult T-cell leukemia/lymphoma (ATLL is a highly aggressive disease that occurs in individuals infected with the human T lymphotropic virus type 1 (HTLV-1. Patients with aggressive ATLL have a poor prognosis because the leukemic cells are resistant to conventional chemotherapy. We have investigated the therapeutic efficacy of a biphosphinic cyclopalladated complex {Pd2 [S(−C2, N-dmpa]2 (μ-dppeCl2}, termed C7a, in a patient-derived xenograft model of ATLL, and investigated the mechanism of C7a action in HTLV-1-positive and negative transformed T cell lines in vitro. In vivo survival studies in immunocompromised mice inoculated with human RV-ATL cells and intraperitoneally treated with C7a led to significantly increased survival of the treated mice. We investigated the mechanism of C7a activity in vitro and found that it induced mitochondrial release of cytochrome c, caspase activation, nuclear condensation and DNA degradation. These results suggest that C7a triggers apoptotic cell death in both HTLV-1 infected and uninfected human transformed T-cell lines. Significantly, C7a was not cytotoxic to peripheral blood mononuclear cells (PBMC from healthy donors and HTLV-1-infected individuals. C7a inhibited more than 60% of the ex vivo spontaneous proliferation of PBMC from HTLV-1-infected individuals. These results support a potential therapeutic role for C7a in both ATLL and HTLV-1-negative T-cell lymphomas.

  1. Induced pluripotent stem cells-derived myeloid-derived suppressor cells regulate the CD8+ T cell response

    Directory of Open Access Journals (Sweden)

    Daniel Joyce

    2018-05-01

    Full Text Available Myeloid-derived suppressor cells (MDSCs are markedly increased in cancer patients and tumor-bearing mice and promote tumor growth and survival by inhibiting host innate and adaptive immunity. In this study, we generated and characterized MDSCs from murine-induced pluripotent stem cells (iPSCs. The iPSCs were co-cultured with OP9 cells, stimulated with GM-CSF, and became morphologically heterologous under co-culturing with hepatic stellate cells. Allogeneic and OVA-specific antigen stimulation demonstrated that iPS-MDSCs have a T-cell regulatory function. Furthermore, a popliteal lymph node assay and autoimmune hepatitis model showed that iPS-MDSCs also regulate immune responsiveness in vivo and have a therapeutic effect against hepatitis. Taken together, our results demonstrated a method of generating functional MDSCs from iPSCs and highlighted the potential of iPS-MDSCs as a key cell therapy resource for transplantation and autoimmune diseases. Keywords: Myeloid-derived suppressor cells, Induced pluripotent stem cells, T cell response

  2. Adipose-derived mesenchymal stem cells and regenerative medicine.

    Science.gov (United States)

    Konno, Masamitsu; Hamabe, Atsushi; Hasegawa, Shinichiro; Ogawa, Hisataka; Fukusumi, Takahito; Nishikawa, Shimpei; Ohta, Katsuya; Kano, Yoshihiro; Ozaki, Miyuki; Noguchi, Yuko; Sakai, Daisuke; Kudoh, Toshihiro; Kawamoto, Koichi; Eguchi, Hidetoshi; Satoh, Taroh; Tanemura, Masahiro; Nagano, Hiroaki; Doki, Yuichiro; Mori, Masaki; Ishii, Hideshi

    2013-04-01

    Adipose tissue-derived mesenchymal stem cells (ADSCs) are multipotent and can differentiate into various cell types, including osteocytes, adipocytes, neural cells, vascular endothelial cells, cardiomyocytes, pancreatic β-cells, and hepatocytes. Compared with the extraction of other stem cells such as bone marrow-derived mesenchymal stem cells (BMSCs), that of ADSCs requires minimally invasive techniques. In the field of regenerative medicine, the use of autologous cells is preferable to embryonic stem cells or induced pluripotent stem cells. Therefore, ADSCs are a useful resource for drug screening and regenerative medicine. Here we present the methods and mechanisms underlying the induction of multilineage cells from ADSCs. © 2013 The Authors Development, Growth & Differentiation © 2013 Japanese Society of Developmental Biologists.

  3. Study of the quantitative, functional, cytogenetic, and immunoregulatory properties of bone marrow mesenchymal stem cells in patients with B-cell chronic lymphocytic leukemia.

    Science.gov (United States)

    Pontikoglou, Charalampos; Kastrinaki, Maria-Christina; Klaus, Mirjam; Kalpadakis, Christina; Katonis, Pavlos; Alpantaki, Kalliopi; Pangalis, Gerassimos A; Papadaki, Helen A

    2013-05-01

    The bone marrow (BM) microenvironment has clearly been implicated in the pathogenesis of B-cell chronic lymphocytic leukemia (B-CLL). However, the potential involvement of BM stromal progenitors, the mesenchymal stem cells (MSCs), in the pathophysiology of the disease has not been extensively investigated. We expanded in vitro BM-MSCs from B-CLL patients (n=11) and healthy individuals (n=16) and comparatively assessed their reserves, proliferative potential, differentiation capacity, and immunoregulatory effects on T- and B-cells. We also evaluated the anti-apoptotic effect of patient-derived MSCs on leukemic cells and studied their cytogenetic characteristics in comparison to BM hematopoietic cells. B-CLL-derived BM MSCs exhibit a similar phenotype, differentiation potential, and ability to suppress T-cell proliferative responses as compared with MSCs from normal controls. Furthermore, they do not carry the cytogenetic abnormalities of the leukemic clone, and they exert a similar anti-apoptotic effect on leukemic cells and healthy donor-derived B-cells, as their normal counterparts. On the other hand, MSCs from B-CLL patients significantly promote normal B-cell proliferation and IgG production, in contrast to healthy-donor-derived MSCs. Furthermore, they have impaired reserves, defective cellular growth due to increased apoptotic cell death and exhibit aberrant production of stromal cell-derived factor 1, B-cell activating factor, a proliferation inducing ligand, and transforming growth factor β1, cytokines that are crucial for the survival/nourishing of the leukemic cells. We conclude that ex vivo expanded B-CLL-derived MSCs harbor intrinsic qualitative and quantitative abnormalities that may be implicated in disease development and/or progression.

  4. Novel levamisole derivative induces extrinsic pathway of apoptosis in cancer cells and inhibits tumor progression in mice.

    Directory of Open Access Journals (Sweden)

    Mahesh Hegde

    Full Text Available BACKGROUND: Levamisole, an imidazo(2,1-bthiazole derivative, has been reported to be a potential antitumor agent. In the present study, we have investigated the mechanism of action of one of the recently identified analogues, 4a (2-benzyl-6-(4'-fluorophenyl-5-thiocyanato-imidazo[2,1-b][1], [3], [4]thiadiazole. MATERIALS AND METHODS: ROS production and expression of various apoptotic proteins were measured following 4a treatment in leukemia cell lines. Tumor animal models were used to evaluate the effect of 4a in comparison with Levamisole on progression of breast adenocarcinoma and survival. Immunohistochemistry and western blotting studies were performed to understand the mechanism of 4a action both ex vivo and in vivo. RESULTS: We have determined the IC(50 value of 4a in many leukemic and breast cancer cell lines and found CEM cells most sensitive (IC(50 5 µM. Results showed that 4a treatment leads to the accumulation of ROS. Western blot analysis showed upregulation of pro-apoptotic proteins t-BID and BAX, upon treatment with 4a. Besides, dose-dependent activation of p53 along with FAS, FAS-L, and cleavage of CASPASE-8 suggest that it induces death receptor mediated apoptotic pathway in CEM cells. More importantly, we observed a reduction in tumor growth and significant increase in survival upon oral administration of 4a (20 mg/kg, six doses in mice. In comparison, 4a was found to be more potent than its parental analogue Levamisole based on both ex vivo and in vivo studies. Further, immunohistochemistry and western blotting studies indicate that 4a treatment led to abrogation of tumor cell proliferation and activation of apoptosis by the extrinsic pathway even in animal models. CONCLUSION: Thus, our results suggest that 4a could be used as a potent chemotherapeutic agent.

  5. The Bone Marrow-Derived Stromal Cells

    DEFF Research Database (Denmark)

    Tencerova, Michaela; Kassem, Moustapha

    2016-01-01

    Bone marrow (BM) microenvironment represents an important compartment of bone that regulates bone homeostasis and the balance between bone formation and bone resorption depending on the physiological needs of the organism. Abnormalities of BM microenvironmental dynamics can lead to metabolic bone...... diseases. BM stromal cells (also known as skeletal or mesenchymal stem cells) [bone marrow stromal stem cell (BMSC)] are multipotent stem cells located within BM stroma and give rise to osteoblasts and adipocytes. However, cellular and molecular mechanisms of BMSC lineage commitment to adipocytic lineage...

  6. Technical Challenges in the Derivation of Human Pluripotent Cells

    Directory of Open Access Journals (Sweden)

    Parinya Noisa

    2011-01-01

    Full Text Available It has long been discovered that human pluripotent cells could be isolated from the blastocyst state of embryos and called human embryonic stem cells (ESCs. These cells can be adapted and propagated indefinitely in culture in an undifferentiated manner as well as differentiated into cell representing the three major germ layers: endoderm, mesoderm, and ectoderm. However, the derivation of human pluripotent cells from donated embryos is limited and restricted by ethical concerns. Therefore, various approaches have been explored and proved their success. Human pluripotent cells can also be derived experimentally by the nuclear reprogramming of somatic cells. These techniques include somatic cell nuclear transfer (SCNT, cell fusion and overexpression of pluripotent genes. In this paper, we discuss the technical challenges of these approaches for nuclear reprogramming, involving their advantages and limitations. We will also highlight the possible applications of these techniques in the study of stem cell biology.

  7. Physiology of stem cell-derived cardiomyocytes

    NARCIS (Netherlands)

    Boer, T.P. de

    2007-01-01

    All chapters in this thesis revolve around the general theme, stem cells and their electrophysiological characteristics and capacity to induce pro-arrhythmia. The first part of this thesis focusses on key aspects that are relevant to possible pro-arrhythmic effects of stem cell transplantation. An

  8. 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.

  9. Senescence and quiescence in adipose-derived stromal cells

    DEFF Research Database (Denmark)

    Søndergaard, Rebekka Harary; Follin, Bjarke; Lund, Lisbeth Drozd

    2017-01-01

    Background aims. Adipose-derived stromal cells (ASCs) are attractive sources for cell-based therapies. The hypoxic niche of ASCs in vivo implies that cells will benefit from hypoxia during in vitro expansion. Human platelet lysate (hPL) enhances ASC proliferation rates, compared with fetal bovine...

  10. Role of glycolysis inhibition and poly(ADP-ribose) polymerase activation in necrotic-like cell death caused by ascorbate/menadione-induced oxidative stress in K562 human chronic myelogenous leukemic cells.

    Science.gov (United States)

    Verrax, Julien; Vanbever, Stéphanie; Stockis, Julie; Taper, Henryk; Calderon, Pedro Buc

    2007-03-15

    Among different features of cancer cells, two of them have retained our interest: their nearly universal glycolytic phenotype and their sensitivity towards an oxidative stress. Therefore, we took advantage of these features to develop an experimental approach by selectively exposing cancer cells to an oxidant insult induced by the combination of menadione (vitamin K(3)) and ascorbate (vitamin C). Ascorbate enhances the menadione redox cycling, increases the formation of reactive oxygen species and kills K562 cells as shown by more than 65% of LDH leakage after 24 hr of incubation. Since both lactate formation and ATP content are depressed by about 80% following ascorbate/menadione exposure, we suggest that the major intracellular event involved in such a cytotoxicity is related to the impairment of glycolysis. Indeed, NAD(+) is rapidly and severely depleted, a fact most probably related to a strong Poly(ADP-ribose) polymerase (PARP) activation, as shown by the high amount of poly-ADP-ribosylated proteins. The addition of N-acetylcysteine (NAC) restores most of the ATP content and the production of lactate as well. The PARP inhibitor dihydroxyisoquinoline (DiQ) was able to partially restore both parameters as well as cell death induced by ascorbate/menadione. These results suggest that the PARP activation induced by the oxidative stress is a major but not the only intracellular event involved in cell death by ascorbate/menadione. Due to the high energetic dependence of cancer cells on glycolysis, the impairment of such an essential pathway may explain the effectiveness of this combination to kill cancer cells. (c) 2006 Wiley-Liss, Inc.

  11. Hemidesmosomal linker proteins regulate cell motility, invasion and tumorigenicity in oral squamous cell carcinoma derived cells.

    Science.gov (United States)

    Chaudhari, Pratik Rajeev; Charles, Silvania Emlit; D'Souza, Zinia Charlotte; Vaidya, Milind Murlidhar

    2017-11-15

    BPAG1e and Plectin are hemidesmosomal linker proteins which anchor intermediate filament proteins to the cell surface through β4 integrin. Recent reports indicate that these proteins play a role in various cellular processes apart from their known anchoring function. However, the available literature is inconsistent. Further, the previous study from our laboratory suggested that Keratin8/18 pair promotes cell motility and tumor progression by deregulating β4 integrin signaling in oral squamous cell carcinoma (OSCC) derived cells. Based on these findings, we hypothesized that linker proteins may have a role in neoplastic progression of OSCC. Downregulation of hemidesmosomal linker proteins in OSCC derived cells resulted in reduced cell migration accompanied by alterations in actin organization. Further, decreased MMP9 activity led to reduced cell invasion in linker proteins knockdown cells. Moreover, loss of these proteins resulted in reduced tumorigenic potential. SWATH analysis demonstrated upregulation of N-Myc downstream regulated gene 1 (NDRG1) in linker proteins downregulated cells as compared to vector control cells. Further, the defects in phenotype upon linker proteins ablation were rescued upon loss of NDRG1 in linker proteins knockdown background. These data together indicate that hemidesmosomal linker proteins regulate cell motility, invasion and tumorigenicity possibly through NDRG1 in OSCC derived cells. Copyright © 2017 Elsevier Inc. All rights reserved.

  12. Skin appendage-derived stem cells: cell biology and potential for wound repair

    OpenAIRE

    Xie, Jiangfan; Yao, Bin; Han, Yutong; Huang, Sha; Fu, Xiaobing

    2016-01-01

    Stem cells residing in the epidermis and skin appendages are imperative for skin homeostasis and regeneration. These stem cells also participate in the repair of the epidermis after injuries, inducing restoration of tissue integrity and function of damaged tissue. Unlike epidermis-derived stem cells, comprehensive knowledge about skin appendage-derived stem cells remains limited. In this review, we summarize the current knowledge of skin appendage-derived stem cells, including their fundament...

  13. Pre leukemic granulocytic sarcoma of vagina: a case report with review of literature

    International Nuclear Information System (INIS)

    Lakshminarasimhan, Srinivasan; Doval, D.C.; Rajashekhar, Usha; Mukherjee, Geethashree; Kannan, V.; Lakshmi Devi; Bapsy, P.P.

    1996-01-01

    Granulocytic sarcoma is an extramedullary tumor of malignant granulocytic progenitor cells, that may precede the onset of acute myeloid leukemia or appear during the leukemic manifestation or blastic crisis of chronic myeloproliferative disorders. A case of granulocytic sarcoma of vagina in a 27 year old woman treated with local radiotherapy is described. After seven months of follow up she developed acute myeloid leukemia. The case has been presented in view of its rarity and discussed in light of the available literature. (author). 13 refs., 1 fig

  14. 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.

  15. 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

  16. Research on human placenta-derived mesenchymal stem cells ...

    African Journals Online (AJOL)

    Research on human placenta-derived mesenchymal stem cells transfected with pIRES2-EGFP-VEGF165 using liposome. ... African Journal of Biotechnology. Journal Home · ABOUT THIS JOURNAL · Advanced Search · Current Issue ...

  17. 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.

  18. Isolation and Osteogenic Differentiation of Rat Periosteum-derived Cells

    OpenAIRE

    Declercq, Heidi Andrea; De Ridder, Leo Isabelle; Cornelissen, Maria Jozefa

    2005-01-01

    Selection of appropriate cultures having an osteogenic potential is a necessity if cell/biomaterial interactions are studied in long-term cultures. Osteoblastic cells derived from rat long bones or calvaria have the disadvantage of being in an advanced differentiation stage which results in terminal differentiation within 21 days. In this regard, less differentiated periosteum-derived osteoprogenitors could be more suitable.

  19. 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.

  20. Stem cell-derived vascular endothelial cells and their potential application in regenerative medicine

    Science.gov (United States)

    Although a 'vascular stem cell' population has not been identified or generated, vascular endothelial and mural cells (smooth muscle cells and pericytes) can be derived from currently known pluripotent stem cell sources, including human embryonic stem cells and induced pluripotent stem cells. We rev...

  1. 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...

  2. 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

  3. Derivation of the human embryonic stem cell line RCM1

    Directory of Open Access Journals (Sweden)

    P.A. De Sousa

    2016-03-01

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

  4. Modeling neurodegenerative diseases with patient-derived induced pluripotent cells

    DEFF Research Database (Denmark)

    Poon, Anna; Zhang, Yu; Chandrasekaran, Abinaya

    2017-01-01

    patient-specific induced pluripotent stem cells (iPSCs) and isogenic controls generated using CRISPR-Cas9 mediated genome editing. The iPSCs are self-renewable and capable of being differentiated into the cell types affected by the diseases. These in vitro models based on patient-derived iPSCs provide...... the possibilities of generating three-dimensional (3D) models using the iPSCs-derived cells and compare their advantages and disadvantages to conventional two-dimensional (2D) models....

  5. Platelet-derived stromal cell-derived factor-1 is required for the transformation of circulating monocytes into multipotential cells.

    Directory of Open Access Journals (Sweden)

    Noriyuki Seta

    Full Text Available BACKGROUND: We previously described a primitive cell population derived from human circulating CD14(+ monocytes, named monocyte-derived multipotential cells (MOMCs, which are capable of differentiating into mesenchymal and endothelial lineages. To generate MOMCs in vitro, monocytes are required to bind to fibronectin and be exposed to soluble factor(s derived from circulating CD14(- cells. The present study was conducted to identify factors that induce MOMC differentiation. METHODS: We cultured CD14(+ monocytes on fibronectin in the presence or absence of platelets, CD14(- peripheral blood mononuclear cells, platelet-conditioned medium, or candidate MOMC differentiation factors. The transformation of monocytes into MOMCs was assessed by the presence of spindle-shaped adherent cells, CD34 expression, and the potential to differentiate in vitro into mesenchymal and endothelial lineages. RESULTS: The presence of platelets or platelet-conditioned medium was required to generate MOMCs from monocytes. A screening of candidate platelet-derived soluble factors identified stromal cell-derived factor (SDF-1 as a requirement for generating MOMCs. Blocking an interaction between SDF-1 and its receptor CXCR4 inhibited MOMC generation, further confirming SDF-1's critical role in this process. Finally, circulating MOMC precursors were found to reside in the CD14(+CXCR4(high cell population. CONCLUSION: The interaction of SDF-1 with CXCR4 is essential for the transformation of circulating monocytes into MOMCs.

  6. Tumor-derived circulating endothelial cell clusters in colorectal cancer.

    KAUST Repository

    Cima, Igor; Kong, Say Li; Sengupta, Debarka; Tan, Iain B; Phyo, Wai Min; Lee, Daniel; Hu, Min; Iliescu, Ciprian; Alexander, Irina; Goh, Wei Lin; Rahmani, Mehran; Suhaimi, Nur-Afidah Mohamed; Vo, Jess H; Tai, Joyce A; Tan, Joanna H; Chua, Clarinda; Ten, Rachel; Lim, Wan Jun; Chew, Min Hoe; Hauser, Charlotte; van Dam, Rob M; Lim, Wei-Yen; Prabhakar, Shyam; Lim, Bing; Koh, Poh Koon; Robson, Paul; Ying, Jackie Y; Hillmer, Axel M; Tan, Min-Han

    2016-01-01

    Clusters of tumor cells are often observed in the blood of cancer patients. These structures have been described as malignant entities for more than 50 years, although their comprehensive characterization is lacking. Contrary to current consensus, we demonstrate that a discrete population of circulating cell clusters isolated from the blood of colorectal cancer patients are not cancerous but consist of tumor-derived endothelial cells. These clusters express both epithelial and mesenchymal markers, consistent with previous reports on circulating tumor cell (CTC) phenotyping. However, unlike CTCs, they do not mirror the genetic variations of matched tumors. Transcriptomic analysis of single clusters revealed that these structures exhibit an endothelial phenotype and can be traced back to the tumor endothelium. Further results show that tumor-derived endothelial clusters do not form by coagulation or by outgrowth of single circulating endothelial cells, supporting a direct release of clusters from the tumor vasculature. The isolation and enumeration of these benign clusters distinguished healthy volunteers from treatment-naïve as well as pathological early-stage (≤IIA) colorectal cancer patients with high accuracy, suggesting that tumor-derived circulating endothelial cell clusters could be used as a means of noninvasive screening for colorectal cancer. In contrast to CTCs, tumor-derived endothelial cell clusters may also provide important information about the underlying tumor vasculature at the time of diagnosis, during treatment, and throughout the course of the disease.

  7. Tumor-derived circulating endothelial cell clusters in colorectal cancer.

    KAUST Repository

    Cima, Igor

    2016-06-29

    Clusters of tumor cells are often observed in the blood of cancer patients. These structures have been described as malignant entities for more than 50 years, although their comprehensive characterization is lacking. Contrary to current consensus, we demonstrate that a discrete population of circulating cell clusters isolated from the blood of colorectal cancer patients are not cancerous but consist of tumor-derived endothelial cells. These clusters express both epithelial and mesenchymal markers, consistent with previous reports on circulating tumor cell (CTC) phenotyping. However, unlike CTCs, they do not mirror the genetic variations of matched tumors. Transcriptomic analysis of single clusters revealed that these structures exhibit an endothelial phenotype and can be traced back to the tumor endothelium. Further results show that tumor-derived endothelial clusters do not form by coagulation or by outgrowth of single circulating endothelial cells, supporting a direct release of clusters from the tumor vasculature. The isolation and enumeration of these benign clusters distinguished healthy volunteers from treatment-naïve as well as pathological early-stage (≤IIA) colorectal cancer patients with high accuracy, suggesting that tumor-derived circulating endothelial cell clusters could be used as a means of noninvasive screening for colorectal cancer. In contrast to CTCs, tumor-derived endothelial cell clusters may also provide important information about the underlying tumor vasculature at the time of diagnosis, during treatment, and throughout the course of the disease.

  8. Benzoxazole derivatives suppress lipopolysaccharide-induced mast cell activation.

    Science.gov (United States)

    Cho, Kyung-Ah; Park, Minhwa; Kim, Yu-Hee; Choo, Hea-Young Park; Lee, Kyung Ho

    2018-05-01

    Mast cells are central regulators of allergic inflammation that function by releasing various proallergic inflammatory mediators, including histamine, eicosanoids and proinflammatory cytokines. Occasionally, bacterial infections may initiate or worsen allergic inflammation. A number of studies have indicated that activation of lipoxygenase in mast cells positive regulates allergic inflammatory responses by generating leukotrienes and proinflammatory cytokines. In the present study, the effects of benzoxazole derivatives on the lipopolysaccharide (LPS)‑induced expression of proinflammatory cytokines, production of histamine and surface expression of co‑stimulatory molecules on bone marrow-derived mast cells (BMMCs) were studied. The benzoxazole derivatives significantly reduced the expression of interleukin (IL)‑1β, IL‑6, IL‑13, tumor necrosis factor‑α, perilipin (PLIN) 2, and PLIN3 in BMMCs treated with LPS. Furthermore, histamine production was suppressed in BMMCs treated with LPS, or treated with phorbol-12-myristate-13-acetate/ionomycin. Benzoxazole derivatives marginally affected the surface expression of cluster of differentiation (CD)80 and CD86 on BMMCs in the presence of LPS, although LPS alone did not increase the expression of those proteins. Therefore, benzoxazole derivatives inhibited the secretion of proinflammatory cytokines in mast cells and may be potential candidate anti‑allergic agents to suppress mast cell activation.

  9. Role of adipose-derived stem cells in wound healing.

    Science.gov (United States)

    Hassan, Waqar Ul; Greiser, Udo; Wang, Wenxin

    2014-01-01

    Impaired wound healing remains a challenge to date and causes debilitating effects with tremendous suffering. Recent advances in tissue engineering approaches in the area of cell therapy have provided promising treatment options to meet the challenges of impaired skin wound healing such as diabetic foot ulcers. Over the last few years, stem cell therapy has emerged as a novel therapeutic approach for various diseases including wound repair and tissue regeneration. Several different types of stem cells have been studied in both preclinical and clinical settings such as bone marrow-derived stem cells, adipose-derived stem cells (ASCs), circulating angiogenic cells (e.g., endothelial progenitor cells), human dermal fibroblasts, and keratinocytes for wound healing. Adipose tissue is an abundant source of mesenchymal stem cells, which have shown an improved outcome in wound healing studies. ASCs are pluripotent stem cells with the ability to differentiate into different lineages and to secrete paracrine factors initiating tissue regeneration process. The abundant supply of fat tissue, ease of isolation, extensive proliferative capacities ex vivo, and their ability to secrete pro-angiogenic growth factors make them an ideal cell type to use in therapies for the treatment of nonhealing wounds. In this review, we look at the pathogenesis of chronic wounds, role of stem cells in wound healing, and more specifically look at the role of ASCs, their mechanism of action and their safety profile in wound repair and tissue regeneration. © 2014 by the Wound Healing Society.

  10. Adipose-Derived Stem Cells and Application Areas

    Directory of Open Access Journals (Sweden)

    Mujde Kivanc

    2015-09-01

    Full Text Available The use of stem cells derived from adipose tissue as an autologous and self-replenishing source for a variety of differentiated cell phenotypes, provides a great deal of promise for reconstructive surgery. The secret of the human body, stem cells are reserved. Stem cells are undifferentiated cells found in the human body placed in any body tissue characteristics that differentiate and win ever known to cross the tissue instead of more than 200 diseases and thus improve and, rejuvenates the tissues. So far, the cord blood of newborn babies are used as a source of stem cells, bone marrow, and twenty years after tooth stem cells in human adipose tissue, scientists studied more than other sources of stem cells in adipose tissue and discovered that. Increase in number of in vitro studies on adult stem cells, depending on many variables is that the stem cells directly to the desired soybean optimization can be performed.. We will conclude by assessing potential avenues for developing this incredibly promising field. The aim of this paper is to review the existing literature on applications of harvest, purification, characterization and cryopreservation of adipose-derived stem cells (ASCs. [Cukurova Med J 2015; 40(3.000: 399-408

  11. Large Scale Production of Stem Cells and Their Derivatives

    Science.gov (United States)

    Zweigerdt, Robert

    Stem cells have been envisioned to become an unlimited cell source for regenerative medicine. Notably, the interest in stem cells lies beyond direct therapeutic applications. They might also provide a previously unavailable source of valuable human cell types for screening platforms, which might facilitate the development of more efficient and safer drugs. The heterogeneity of stem cell types as well as the numerous areas of application suggests that differential processes are mandatory for their in vitro culture. Many of the envisioned applications would require the production of a high number of stem cells and their derivatives in scalable, well-defined and potentially clinical compliant manner under current good manufacturing practice (cGMP). In this review we provide an overview on recent strategies to develop bioprocesses for the expansion, differentiation and enrichment of stem cells and their progenies, presenting examples for adult and embryonic stem cells alike.

  12. 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

  13. Extracellular matrix-derived hydrogels for dental stem cell delivery

    OpenAIRE

    Viswanath, Aiswarya; Vanacker, Julie; Germain, Loic; Leprince, Julien G.; Diogenes, Anibal; Shakesheff, Kevin M.; White, Lisa J.; des Rieux, Anne

    2016-01-01

    Decellularised mammalian extracellular matrices (ECM) have been widely accepted as an ideal substrate for repair and remodelling of numerous tissues in clinical and pre-clinical studies. Recent studies have demonstrated the ability of ECM scaffolds derived from site-specific homologous tissues to direct cell differentiation. The present study investigated the suitability of hydrogels derived from different source tissues: bone, spinal cord and dentine, as suitable carriers to deliver human ap...

  14. 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.

  15. Nestin expression in the cell lines derived from glioblastoma multiforme

    International Nuclear Information System (INIS)

    Veselska, Renata; Kuglik, Petr; Cejpek, Pavel; Svachova, Hana; Neradil, Jakub; Loja, Tomas; Relichova, Jirina

    2006-01-01

    Nestin is a protein belonging to class VI of intermediate filaments that is produced in stem/progenitor cells in the mammalian CNS during development and is consecutively replaced by other intermediate filament proteins (neurofilaments, GFAP). Down-regulated nestin may be re-expressed in the adult organism under certain pathological conditions (brain injury, ischemia, inflammation, neoplastic transformation). Our work focused on a detailed study of the nestin cytoskeleton in cell lines derived from glioblastoma multiforme, because re-expression of nestin together with down-regulation of GFAP has been previously reported in this type of brain tumor. Two cell lines were derived from the tumor tissue of patients treated for glioblastoma multiforme. Nestin and other cytoskeletal proteins were visualized using imunocytochemical methods: indirect immunofluorescence and immunogold-labelling. Using epifluorescence and confocal microscopy, we described the morphology of nestin-positive intermediate filaments in glioblastoma cells of both primary cultures and the derived cell lines, as well as the reorganization of nestin during mitosis. Our most important result came through transmission electron microscopy and provided clear evidence that nestin is present in the cell nucleus. Detailed information concerning the pattern of the nestin cytoskeleton in glioblastoma cell lines and especially the demonstration of nestin in the nucleus represent an important background for further studies of nestin re-expression in relationship to tumor malignancy and invasive potential

  16. Tumorigenicity studies for human pluripotent stem cell-derived products.

    Science.gov (United States)

    Kuroda, Takuya; Yasuda, Satoshi; Sato, Yoji

    2013-01-01

    Human pluripotent stem cells (hPSCs), i.e. human embryonic stem cells and human induced pluripotent stem cells, are able to self-renew and differentiate into multiple cell types. Because of these abilities, numerous attempts have been made to utilize hPSCs in regenerative medicine/cell therapy. hPSCs are, however, also tumorigenic, that is, they can give rise to the progressive growth of tumor nodules in immunologically unresponsive animals. Therefore, assessing and managing the tumorigenicity of all final products is essential in order to prevent ectopic tissue formation, tumor development, and/or malignant transformation elicited by residual pluripotent stem cells after implantation. No detailed guideline for the tumorigenicity testing of hPSC-derived products has yet been issued for regenerative medicine/cell therapy, despite the urgent necessity. Here, we describe the current situations and issues related to the tumorigenicity testing of hPSC-derived products and we review the advantages and disadvantages of several types of tumorigenicity-associated tests. We also refer to important considerations in the execution and design of specific studies to monitor the tumorigenicity of hPSC-derived products.

  17. 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

  18. Curcumin Modulates Pancreatic Adenocarcinoma Cell-Derived Exosomal Function

    Science.gov (United States)

    Osterman, Carlos J. Diaz; Lynch, James C.; Leaf, Patrick; Gonda, Amber; Ferguson Bennit, Heather R.; Griffiths, Duncan; Wall, Nathan R.

    2015-01-01

    Pancreatic cancer has the highest mortality rates of all cancer types. One potential explanation for the aggressiveness of this disease is that cancer cells have been found to communicate with one another using membrane-bound vesicles known as exosomes. These exosomes carry pro-survival molecules and increase the proliferation, survival, and metastatic potential of recipient cells, suggesting that tumor-derived exosomes are powerful drivers of tumor progression. Thus, to successfully address and eradicate pancreatic cancer, it is imperative to develop therapeutic strategies that neutralize cancer cells and exosomes simultaneously. Curcumin, a turmeric root derivative, has been shown to have potent anti-cancer and anti-inflammatory effects in vitro and in vivo. Recent studies have suggested that exosomal curcumin exerts anti-inflammatory properties on recipient cells. However, curcumin’s effects on exosomal pro-tumor function have yet to be determined. We hypothesize that curcumin will alter the pro-survival role of exosomes from pancreatic cancer cells toward a pro-death role, resulting in reduced cell viability of recipient pancreatic cancer cells. The main objective of this study was to determine the functional alterations of exosomes released by pancreatic cancer cells exposed to curcumin compared to exosomes from untreated pancreatic cancer cells. We demonstrate, using an in vitro cell culture model involving pancreatic adenocarcinoma cell lines PANC-1 and MIA PaCa-2, that curcumin is incorporated into exosomes isolated from curcumin-treated pancreatic cancer cells as observed by spectral studies and fluorescence microscopy. Furthermore, curcumin is delivered to recipient pancreatic cancer cells via exosomes, promoting cytotoxicity as demonstrated by Hoffman modulation contrast microscopy as well as AlamarBlue and Trypan blue exclusion assays. Collectively, these data suggest that the efficacy of curcumin may be enhanced in pancreatic cancer cells through

  19. Coelomic epithelium-derived cells in visceral morphogenesis.

    Science.gov (United States)

    Ariza, Laura; Carmona, Rita; Cañete, Ana; Cano, Elena; Muñoz-Chápuli, Ramón

    2016-03-01

    Coelomic cavities of vertebrates are lined by a mesothelium which develops from the lateral plate mesoderm. During development, the coelomic epithelium is a highly active cell layer, which locally is able to supply mesenchymal cells that contribute to the mesodermal elements of many organs and provide signals which are necessary for their development. The relevance of this process of mesenchymal cell supply to the developing organs is becoming clearer because genetic lineage tracing techniques have been developed in recent years. Body wall, heart, liver, lungs, gonads, and gastrointestinal tract are populated by cells derived from the coelomic epithelium which contribute to their connective and vascular tissues, and sometimes to specialized cell types such as the stellate cells of the liver, the Cajal interstitial cells of the gut or the Sertoli cells of the testicle. In this review we collect information about the contribution of coelomic epithelium derived cells to visceral development, their developmental fates and signaling functions. The common features displayed by all these processes suggest that the epithelial-mesenchymal transition of the embryonic coelomic epithelium is an underestimated but key event of vertebrate development, and probably it is shared by all the coelomate metazoans. © 2015 Wiley Periodicals, Inc.

  20. [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.

  1. 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.

  2. Quantitative MR imaging of normal and leukemic bone marrow

    International Nuclear Information System (INIS)

    Hinks, R.S.; Dunlap, H.J.; Poon, P.Y.; Curtis, J.; Henkelman, R.M.

    1986-01-01

    The authors have developed and tested a protocol that allows extraction of reliable T1 and T2 relaxation times from imaging data. They have used these methods to study in vivo the bone marrow of healthy volunteers and patients with acute leukemia. Examinations were performed at 6.25 MHz using an interleaved ISE/SE sequence to calculate T1 and an eight echo (TE = 25) sequence to calculate T2. The results are summarized as follows: In leukemic patients, T1 = 476 +- 115 msec; in leukemic patients in remission, T1 = 290 +- 31 msec; in healthy volunteers, T1 = 329 +- 32 msec. The T2 values were not significantly different for the three groups (105 +- 10 msec). Work is underway to evaluate whether T1 values of bone marrow may be used to monitor patients in remission and to detect the onset of relapse

  3. Myeloid derived suppressor cells as therapeutic target in hematological malignancies

    Directory of Open Access Journals (Sweden)

    Kim eDe Veirman

    2014-12-01

    Full Text Available Myeloid derived suppressor cells (MDSC are a heterogeneous population of immature myeloid cells that accumulate during pathological conditions such as cancer and are associated with a poor clinical outcome. MDSC expansion hampers the host anti-tumor immune response by inhibition of T cell proliferation, cytokine secretion and recruitment of regulatory T cells. In addition, MDSC exert non-immunological functions including the promotion of angiogenesis, tumor invasion and metastasis. Recent years, MDSC are considered as a potential target in solid tumors and hematological malignancies to enhance the effects of currently used immune modulating agents. This review focuses on the characteristics, distribution, functions, cell-cell interactions and targeting of MDSC in hematological malignancies including multiple myeloma, lymphoma and leukemia.

  4. Characterization of goat inner cell mass derived cells in double kinase inhibition condition

    International Nuclear Information System (INIS)

    Wei, Qiang; Xi, Qihui; Liu, Xiaokun; Meng, Kai; Zhao, Xiaoe; Ma, Baohua

    2017-01-01

    The identification of small molecular inhibitors, which were reported to promote the derivation of mouse and human embryonic stem cells (ESCs), provides a potential strategy for the derivation of domesticated ungulate ESCs. In present study, goat inner cell mass (ICM) derived cells in the double inhibition (2i) condition, in which, mitogen-activated protein kinase kinase (MAP2K) and glycogen synthase kinase 3 (GSK3) were inhibited by PD0325901 and BIO respectively, were characterized. The results showed that goat ICM derived cells in 2i medium adding leukaemia inhibitor factor (LIF) possessed a mouse ES-like morphology. But these cells had much compromised proliferation capacity, resulting in difficulty in expansion. In 2i alone medium, goat ICM derived cells possessed primate ES-like morphology. These cells expressed pluripotent markers and could differentiate into derivatives of three germ layers in vitro. However, these cells could not be proliferated in long-term (persisted for 15 passages) because of spontaneously neural differentiation. Additionally, goat ICM derived cells could be inducing differentiated into neural lineage in vitro. Although goat ESCs could not be established in PD0325901 and BIO alone medium, this derivation condition provides a useful research system to find signaling molecular those regulate early embryonic development and pluripotency in goat. - Highlights: • Goat inner cell mass derived cells possessed finite pluripotency in 2i condition. • These cells could not be proliferated in long-term in 2i condition. • These cells could spontaneously and inductively differentiate into neural lineage.

  5. Exosomes Derived From Pancreatic Stellate Cells: MicroRNA Signature and Effects on Pancreatic Cancer Cells.

    Science.gov (United States)

    Takikawa, Tetsuya; Masamune, Atsushi; Yoshida, Naoki; Hamada, Shin; Kogure, Takayuki; Shimosegawa, Tooru

    2017-01-01

    Pancreatic stellate cells (PSCs) interact with pancreatic cancer cells in the tumor microenvironment. Cell constituents including microRNAs may be exported from cells within membranous nanovesicles termed exosomes. Exosomes might play a pivotal role in intercellular communication. This study aimed to clarify the microRNA signature of PSC-derived exosomes and their effects on pancreatic cancer cells. Exosomes were prepared from the conditioned medium of immortalized human PSCs. MicroRNAs were prepared from the exosomes and their source PSCs, and the microRNA expression profiles were compared by microarray. The effects of PSC-derived exosomes on proliferation, migration, and the mRNA expression profiles were examined in pancreatic cancer cells. Pancreatic stellate cell-derived exosomes contained a variety of microRNAs including miR-21-5p. Several microRNAs such as miR-451a were enriched in exosomes compared to their source PSCs. Pancreatic stellate cell-derived exosomes stimulated the proliferation, migration and expression of mRNAs for chemokine (C - X - C motif) ligands 1 and 2 in pancreatic cancer cells. The stimulation of proliferation, migration, and chemokine gene expression by the conditioned medium of PSCs was suppressed by GW4869, an exosome inhibitor. We clarified the microRNA expression profile in PSC-derived exosomes. Pancreatic stellate cell-derived exosomes might play a role in the interactions between PSCs and pancreatic cancer cells.

  6. Macrophages discriminate glycosylation patterns of apoptotic cell-derived microparticles.

    Science.gov (United States)

    Bilyy, Rostyslav O; Shkandina, Tanya; Tomin, Andriy; Muñoz, Luis E; Franz, Sandra; Antonyuk, Volodymyr; Kit, Yuriy Ya; Zirngibl, Matthias; Fürnrohr, Barbara G; Janko, Christina; Lauber, Kirsten; Schiller, Martin; Schett, Georg; Stoika, Rostyslav S; Herrmann, Martin

    2012-01-02

    Inappropriate clearance of apoptotic remnants is considered to be the primary cause of systemic autoimmune diseases, like systemic lupus erythematosus. Here we demonstrate that apoptotic cells release distinct types of subcellular membranous particles (scMP) derived from the endoplasmic reticulum (ER) or the plasma membrane. Both types of scMP exhibit desialylated glycotopes resulting from surface exposure of immature ER-derived glycoproteins or from surface-borne sialidase activity, respectively. Sialidase activity is activated by caspase-dependent mechanisms during apoptosis. Cleavage of sialidase Neu1 by caspase 3 was shown to be directly involved in apoptosis-related increase of surface sialidase activity. ER-derived blebs possess immature mannosidic glycoepitopes and are prioritized by macrophages during clearance. Plasma membrane-derived blebs contain nuclear chromatin (DNA and histones) but not components of the nuclear envelope. Existence of two immunologically distinct types of apoptotic blebs may provide new insights into clearance-related diseases.

  7. Bone marrow-derived CD13+ cells sustain tumor progression

    Science.gov (United States)

    Dondossola, Eleonora; Corti, Angelo; Sidman, Richard L; Arap, Wadih; Pasqualini, Renata

    2014-01-01

    Non-malignant cells found within neoplastic lesions express alanyl (membrane) aminopeptidase (ANPEP, best known as CD13), and CD13-null mice exhibit limited tumor growth and angiogenesis. We have recently demonstrated that a subset of bone marrow-derived CD11b+CD13+ myeloid cells accumulate within neoplastic lesions in several murine models of transplantable cancer to promote angiogenesis. If these findings were confirmed in clinical settings, CD11b+CD13+ myeloid cells could become a non-malignant target for the development of novel anticancer regimens. PMID:25339996

  8. The CNGRC-GG-D(KLAKLAK)2 peptide induces a caspase-independent, Ca2+-dependent death in human leukemic myeloid cells by targeting surface aminopeptidase N/CD13.

    Science.gov (United States)

    Bouchet, Sandrine; Tang, Ruoping; Fava, Fanny; Legrand, Ollivier; Bauvois, Brigitte

    2016-04-12

    The CD13 antigen's binding site for the Asn-Gly-Arg (NGR) motif enables NGR-containing chemotherapeutic drugs to be delivered to CD13-positive tumours. Human CD13-positive acute myeloid leukemia (AML) cells proliferate abnormally and escape death. Here, we show that the CNGRC-GG-D(KLAKLAK)2 peptide induces death in AML cell lines (U937, THP-1, NB4, HL-60) and primary blood cells from AML patients. Cell death was characterized as a caspase-independent mechanism, without DNA fragmentation, but phosphatidylserine externalization and membrane disruption. Our results demonstrate in U937 cells that (i) the NGR-peptide triggers the loss of mitochondrial potential(ΔΨm) and generates superoxide anion (O2-), (ii) N-acetyl-L-cysteine (NAC) and extra/intracellular Ca2+ chelators (BAPTA) prevent both O2- production and cell death, (iii) the Ca2+-channel blocker nifedipine prevents cell death (indicating that Ca2+ influx is the initial death trigger), and (iv) BAPTA, but not NAC, prevents ΔΨm loss (suggesting O2- is a mitochondrial downstream effector). AML cell lines and primary blasts responding to the lethal action of NGR-peptide express promatrix metalloproteinase-12 (proMMP-12) and its substrate progranulin (an 88 kDa cell survival factor). A cell-free assay highlighted proMMP-12 activation by O2-. Accordingly, NGR-peptide's downregulation of 88 kDa progranulin protein was prevented by BAPTA and NAC. Conversely, AML blast resistance to NGR-peptide is associated with the expression of a distinct, 105 kDa progranulin isoform. These results indicate that CNGRC-GG-D(KLAKLAK)2 induces death in AML cells through the Ca2+-mitochondria-O2.-pathway, and support the link between proMMP-12 activation and progranulin cleavage during cell death. Our findings may have implications for the understanding of tumour biology and treatment.

  9. The CNGRC-GG-D(KLAKLAK)2 peptide induces a caspase-independent, Ca2+-dependent death in human leukemic myeloid cells by targeting surface aminopeptidase N/CD13

    OpenAIRE

    Bouchet, Sandrine; Tang, Ruoping; Fava, Fanny; Legrand, Ollivier; Bauvois, Brigitte

    2015-01-01

    The CD13 antigen's binding site for the Asn-Gly-Arg (NGR) motif enables NGR-containing chemotherapeutic drugs to be delivered to CD13-positive tumours. Human CD13-positive acute myeloid leukemia (AML) cells proliferate abnormally and escape death. Here, we show that the CNGRC-GG-D(KLAKLAK)2 peptide induces death in AML cell lines (U937, THP-1, NB4, HL-60) and primary blood cells from AML patients. Cell death was characterized as a caspase-independent mechanism, without DNA fragmentation, but ...

  10. 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.

  11. Myeloid-Derived Suppressor Cells and Therapeutic Strategies in Cancer

    Directory of Open Access Journals (Sweden)

    Hiroshi Katoh

    2015-01-01

    Full Text Available Development of solid cancer depends on escape from host immunosurveillance. Various types of immune cells contribute to tumor-induced immune suppression, including tumor associated macrophages, regulatory T cells, type 2 NKT cells, and myeloid-derived suppressor cells (MDSCs. Growing body of evidences shows that MDSCs play pivotal roles among these immunosuppressive cells in multiple steps of cancer progression. MDSCs are immature myeloid cells that arise from myeloid progenitor cells and comprise a heterogeneous immune cell population. MDSCs are characterized by the ability to suppress both adaptive and innate immunities mainly through direct inhibition of the cytotoxic functions of T cells and NK cells. In clinical settings, the number of circulating MDSCs is associated with clinical stages and response to treatment in several cancers. Moreover, MDSCs are reported to contribute to chemoresistant phenotype. Collectively, targeting MDSCs could potentially provide a rationale for novel treatment strategies in cancer. This review summarizes recent understandings of MDSCs in cancer and discusses promissing clinical approaches in cancer patients.

  12. Rhodacyanine derivative selectively targets cancer cells and overcomes tamoxifen resistance.

    Directory of Open Access Journals (Sweden)

    John Koren

    Full Text Available MKT-077, a rhodacyanine dye, was shown to produce cancer specific cell death. However, complications prevented the use of this compound beyond clinical trials. Here we describe YM-1, a derivative of MKT-077. We found that YM-1 was more cytotoxic and localized differently than MKT-077. YM-1 demonstrated this cytotoxicity across multiple cancer cell lines. This toxicity was limited to cancer cell lines; immortalized cell models were unaffected. Brief applications of YM-1 were found to be non-toxic. Brief treatment with YM-1 restored tamoxifen sensitivity to a refractory tamoxifen-resistant MCF7 cell model. This effect is potentially due to altered estrogen receptor alpha phosphorylation, an outcome precipitated by selective reductions in Akt levels (Akt/PKB. Thus, modifications to the rhodocyanine scaffold could potentially be made to improve efficacy and pharmacokinetic properties. Moreover, the impact on tamoxifen sensitivity could be a new utility for this compound family.

  13. Recruitment of bone marrow derived cells during anti-angiogenic therapy in GBM : Bone marrow derived cell in GBM

    NARCIS (Netherlands)

    Boer, Jennifer C.; Walenkamp, Annemiek M. E.; den Dunnen, Wilfred F. A.

    2014-01-01

    Glioblastoma (GBM) is a highly vascular tumor characterized by rapid and invasive tumor growth, followed by oxygen depletion, hypoxia and neovascularization, which generate a network of disorganized, tortuous and permeable vessels. Recruitment of bone marrow derived cells (BMDC) is crucial for

  14. Derivation and characterization of monkey embryonic stem cells

    Directory of Open Access Journals (Sweden)

    Wolf Don P

    2004-06-01

    Full Text Available Abstract Embryonic stem (ES cell based therapy carries great potential in the treatment of neurodegenerative diseases. However, before clinical application is realized, the safety, efficacy and feasibility of this therapeutic approach must be established in animal models. The rhesus macaque is physiologically and phylogenetically similar to the human, and therefore, is a clinically relevant animal model for biomedical research, especially that focused on neurodegenerative conditions. Undifferentiated monkey ES cells can be maintained in a pluripotent state for many passages, as characterized by a collective repertoire of markers representing embryonic cell surface molecules, enzymes and transcriptional factors. They can also be differentiated into lineage-specific phenotypes of all three embryonic germ layers by epigenetic protocols. For cell-based therapy, however, the quality of ES cells and their progeny must be ensured during the process of ES cell propagation and differentiation. While only a limited number of primate ES cell lines have been studied, it is likely that substantial inter-line variability exists. This implies that diverse ES cell lines may differ in developmental stages, lineage commitment, karyotypic normalcy, gene expression, or differentiation potential. These variables, inherited genetically and/or induced epigenetically, carry obvious complications to therapeutic applications. Our laboratory has characterized and isolated rhesus monkey ES cell lines from in vitro produced blastocysts. All tested cell lines carry the potential to form pluripotent embryoid bodies and nestin-positive progenitor cells. These ES cell progeny can be differentiated into phenotypes representing the endodermal, mesodermal and ectodermal lineages. This review article describes the derivation of monkey ES cell lines, characterization of the undifferentiated phenotype, and their differentiation into lineage-specific, particularly neural, phenotypes

  15. Extracellular matrix-derived hydrogels for dental stem cell delivery.

    Science.gov (United States)

    Viswanath, Aiswarya; Vanacker, Julie; Germain, Loïc; Leprince, Julian G; Diogenes, Anibal; Shakesheff, Kevin M; White, Lisa J; des Rieux, Anne

    2017-01-01

    Decellularized mammalian extracellular matrices (ECM) have been widely accepted as an ideal substrate for repair and remodelling of numerous tissues in clinical and pre-clinical studies. Recent studies have demonstrated the ability of ECM scaffolds derived from site-specific homologous tissues to direct cell differentiation. The present study investigated the suitability of hydrogels derived from different source tissues: bone, spinal cord and dentine, as suitable carriers to deliver human apical papilla derived mesenchymal stem cells (SCAP) for spinal cord regeneration. Bone, spinal cord, and dentine ECM hydrogels exhibited distinct structural, mechanical, and biological characteristics. All three hydrogels supported SCAP viability and proliferation. However, only spinal cord and bone derived hydrogels promoted the expression of neural lineage markers. The specific environment of ECM scaffolds significantly affected the differentiation of SCAP to a neural lineage, with stronger responses observed with spinal cord ECM hydrogels, suggesting that site-specific tissues are more likely to facilitate optimal stem cell behavior for constructive spinal cord regeneration. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 319-328, 2017. © 2016 Wiley Periodicals, Inc.

  16. Myocardial regeneration potential of adipose tissue-derived stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Bai, Xiaowen, E-mail: baixw01@yahoo.com [Department of Molecular Pathology, The University of Texas M.D. Anderson Cancer Center, 1515 Holcombe, Houston, TX 77030 (United States); Alt, Eckhard, E-mail: ealt@mdanderson.org [Department of Molecular Pathology, The University of Texas M.D. Anderson Cancer Center, 1515 Holcombe, Houston, TX 77030 (United States)

    2010-10-22

    Research highlights: {yields} Various tissue resident stem cells are receiving tremendous attention from basic scientists and clinicians and hold great promise for myocardial regeneration. {yields} For practical reasons, human adipose tissue-derived stem cells are attractive stem cells for future clinical application in repairing damaged myocardium. {yields} This review summarizes the characteristics of cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential and the, underlying mechanisms, and safety issues. -- Abstract: Various tissue resident stem cells are receiving attention from basic scientists and clinicians as they hold promise for myocardial regeneration. For practical reasons, adipose tissue-derived stem cells (ASCs) are attractive cells for clinical application in repairing damaged myocardium based on the following advantages: abundant adipose tissue in most patients and easy accessibility with minimally invasive lipoaspiration procedure. Several recent studies have demonstrated that both cultured and freshly isolated ASCs could improve cardiac function in animal model of myocardial infarction. The mechanisms underlying the beneficial effect of ASCs on myocardial regeneration are not fully understood. Growing evidence indicates that transplantation of ASCs improve cardiac function via the differentiation into cardiomyocytes and vascular cells, and through paracrine pathways. Paracrine factors secreted by injected ASCs enhance angiogenesis, reduce cell apoptosis rates, and promote neuron sprouts in damaged myocardium. In addition, Injection of ASCs increases electrical stability of the injured heart. Furthermore, there are no reported cases of arrhythmia or tumorigenesis in any studies regarding myocardial regeneration with ASCs. This review summarizes the characteristics of both cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential, and the

  17. Myocardial regeneration potential of adipose tissue-derived stem cells

    International Nuclear Information System (INIS)

    Bai, Xiaowen; Alt, Eckhard

    2010-01-01

    Research highlights: → Various tissue resident stem cells are receiving tremendous attention from basic scientists and clinicians and hold great promise for myocardial regeneration. → For practical reasons, human adipose tissue-derived stem cells are attractive stem cells for future clinical application in repairing damaged myocardium. → This review summarizes the characteristics of cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential and the, underlying mechanisms, and safety issues. -- Abstract: Various tissue resident stem cells are receiving attention from basic scientists and clinicians as they hold promise for myocardial regeneration. For practical reasons, adipose tissue-derived stem cells (ASCs) are attractive cells for clinical application in repairing damaged myocardium based on the following advantages: abundant adipose tissue in most patients and easy accessibility with minimally invasive lipoaspiration procedure. Several recent studies have demonstrated that both cultured and freshly isolated ASCs could improve cardiac function in animal model of myocardial infarction. The mechanisms underlying the beneficial effect of ASCs on myocardial regeneration are not fully understood. Growing evidence indicates that transplantation of ASCs improve cardiac function via the differentiation into cardiomyocytes and vascular cells, and through paracrine pathways. Paracrine factors secreted by injected ASCs enhance angiogenesis, reduce cell apoptosis rates, and promote neuron sprouts in damaged myocardium. In addition, Injection of ASCs increases electrical stability of the injured heart. Furthermore, there are no reported cases of arrhythmia or tumorigenesis in any studies regarding myocardial regeneration with ASCs. This review summarizes the characteristics of both cultured and freshly isolated stem cells obtained from adipose tissue, their myocardial regeneration potential, and the underlying

  18. Fullerene derivatives as electron acceptors for organic photovoltaic cells.

    Science.gov (United States)

    Mi, Dongbo; Kim, Ji-Hoon; Kim, Hee Un; Xu, Fei; Hwang, Do-Hoon

    2014-02-01

    Energy is currently one of the most important problems humankind faces. Depletion of traditional energy sources such as coal and oil results in the need to develop new ways to create, transport, and store electricity. In this regard, the sun, which can be considered as a giant nuclear fusion reactor, represents the most powerful source of energy available in our solar system. For photovoltaic cells to gain widespread acceptance as a source of clean and renewable energy, the cost per watt of solar energy must be decreased. Organic photovoltaic cells, developed in the past two decades, have potential as alternatives to traditional inorganic semiconductor photovoltaic cells, which suffer from high environmental pollution and energy consumption during production. Organic photovoltaic cells are composed of a blended film of a conjugated-polymer donor and a soluble fullerene-derivative acceptor sandwiched between a poly(3,4-ethylenedioxythiophene):poly(styrenesulfonate)-coated indium tin oxide positive electrode and a low-work-function metal negative electrode. Considerable research efforts aim at designing and synthesizing novel fullerene derivatives as electron acceptors with up-raised lowest unoccupied molecular orbital energy, better light-harvesting properties, higher electron mobility, and better miscibility with the polymer donor for improving the power conversion efficiency of the organic photovoltaic cells. In this paper, we systematically review novel fullerene acceptors synthesized through chemical modification for enhancing the photovoltaic performance by increasing open-circuit voltage, short-circuit current, and fill factor, which determine the performance of organic photovoltaic cells.

  19. Identification of rabbit annulus fibrosus-derived stem cells.

    Directory of Open Access Journals (Sweden)

    Chen Liu

    Full Text Available Annulus fibrosus (AF injuries can lead to substantial deterioration of intervertebral disc (IVD which characterizes degenerative disc disease (DDD. However, treatments for AF repair/regeneration remain challenging due to the intrinsic heterogeneity of AF tissue at cellular, biochemical, and biomechanical levels. In this study, we isolated and characterized a sub-population of cells from rabbit AF tissue which formed colonies in vitro and could self-renew. These cells showed gene expression of typical surface antigen molecules characterizing mesenchymal stem cells (MSCs, including CD29, CD44, and CD166. Meanwhile, they did not express negative markers of MSCs such as CD4, CD8, and CD14. They also expressed Oct-4, nucleostemin, and SSEA-4 proteins. Upon induced differentiation they showed typical osteogenesis, chondrogenesis, and adipogenesis potential. Together, these AF-derived colony-forming cells possessed clonogenicity, self-renewal, and multi-potential differentiation capability, the three criteria characterizing MSCs. Such AF-derived stem cells may potentially be an ideal candidate for DDD treatments using cell therapies or tissue engineering approaches.

  20. The potential of induced pluripotent stem cell derived hepatocytes.

    Science.gov (United States)

    Hannoun, Zara; Steichen, Clara; Dianat, Noushin; Weber, Anne; Dubart-Kupperschmitt, Anne

    2016-07-01

    Orthotopic liver transplantation remains the only curative treatment for liver disease. However, the number of patients who die while on the waiting list (15%) has increased in recent years as a result of severe organ shortages; furthermore the incidence of liver disease is increasing worldwide. Clinical trials involving hepatocyte transplantation have provided encouraging results. However, transplanted cell function appears to often decline after several months, necessitating liver transplantation. The precise aetiology of the loss of cell function is not clear, but poor engraftment and immune-mediated loss appear to be important factors. Also, primary human hepatocytes (PHH) are not readily available, de-differentiate, and die rapidly in culture. Hepatocytes are available from other sources, such as tumour-derived human hepatocyte cell lines and immortalised human hepatocyte cell lines or porcine hepatocytes. However, all these cells suffer from various limitations such as reduced or differences in functions or risk of zoonotic infections. Due to their significant potential, one possible inexhaustible source of hepatocytes is through the directed differentiation of human induced pluripotent stem cells (hiPSCs). This review will discuss the potential applications and existing limitations of hiPSC-derived hepatocytes in regenerative medicine, drug screening, in vitro disease modelling and bioartificial livers. Copyright © 2016 European Association for the Study of the Liver. Published by Elsevier B.V. All rights reserved.

  1. Apple derived cellulose scaffolds for 3D mammalian cell culture.

    Directory of Open Access Journals (Sweden)

    Daniel J Modulevsky

    Full Text Available There are numerous approaches for producing natural and synthetic 3D scaffolds that support the proliferation of mammalian cells. 3D scaffolds better represent the natural cellular microenvironment and have many potential applications in vitro and in vivo. Here, we demonstrate that 3D cellulose scaffolds produced by decellularizing apple hypanthium tissue can be employed for in vitro 3D culture of NIH3T3 fibroblasts, mouse C2C12 muscle myoblasts and human HeLa epithelial cells. We show that these cells can adhere, invade and proliferate in the cellulose scaffolds. In addition, biochemical functionalization or chemical cross-linking can be employed to control the surface biochemistry and/or mechanical properties of the scaffold. The cells retain high viability even after 12 continuous weeks of culture and can achieve cell densities comparable with other natural and synthetic scaffold materials. Apple derived cellulose scaffolds are easily produced, inexpensive and originate from a renewable source. Taken together, these results demonstrate that naturally derived cellulose scaffolds offer a complementary approach to existing techniques for the in vitro culture of mammalian cells in a 3D environment.

  2. 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.

  3. 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...

  4. 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.

  5. Derivation of corneal endothelial cell-like cells from rat neural crest cells in vitro.

    Directory of Open Access Journals (Sweden)

    Chengqun Ju

    Full Text Available The aim of this study was to investigate the feasibility of inducing rat neural crest cells (NCC to differentiate to functional corneal endothelial cell (CEC-like cells in vitro. Rat NCC were induced with adult CEC-derived conditioned medium. Immunofluorescence, flow cytometry and real time RT-PCR assay were used to detect expression of the corneal endothelium differentiation marker N-cadherin and transcription factors FoxC1 and Pitx2. CFDA SE-labeled CEC-like cells were transplanted to the corneal endothelium of a rat corneal endothelium deficiency model, and an eye-down position was maintained for 24 hours to allow cell attachment. The animals were observed for as long as 2 months after surgery and underwent clinical and histological examination. Spindle-like NCC turned to polygonal CEC-like after induction and expressed N-cadherin, FoxC1, Pitx2, zonula occludens-1 and sodium-potassium pump Na(+/K(+ ATPase. The corneas of the experimental group were much clearer than those of the control group and the mean corneal thickness in the experimental group was significantly less than in the control group7, 14, 21 and 28 days after surgery. Confocal microscopy through focusing and histological analysis confirmed that green fluorescence-positive CEC-like cells formed a monolayer covering the Descemet's membrane in the experimental group. In conclusion, CEC-like cells derived from NCCs displayed characters of native CEC, and the induction protocol provides guidance for future human CEC induction from NCC.

  6. Imatinib treatment induces CD5+ B lymphocytes and IgM natural antibodies with anti-leukemic reactivity in patients with chronic myelogenous leukemia.

    Directory of Open Access Journals (Sweden)

    Silvia Catellani

    Full Text Available Imatinib mesylate is a first line treatment of Chronic Myelogenous Leukemia and of a rare form of gastrointestinal stromal cancer, where the response to the drug is also linked to the immune system activation with production of antineoplastic cytokines. In this study, forty patients in the chronic phase of disease, treated with imatinib mesylate, were analyzed. Bone marrow aspirates were drawn at diagnosis, after 3, 6, 12, 18 months for haematological, cytofluorimetric, cytogenetic, biomolecular evaluation and cytokine measurement. Responder and non responder patients were defined according to the European LeukemiaNet recommendations. In responder patients (n = 32, the percentage of bone marrow CD20(+CD5(+sIgM(+ lymphocytes, and the plasma levels of IgM, were significantly higher, at 3 months and up to 9 months, than in non responders. These IgM reacted with O-linked sugars expressed by leukemic cells and could induce tumor cell apoptosis. In responder patients the stromal-derived factor-1 and the B-lymphocyte-activating factor of the tumor necrosis factor family significantly raised in the bone marrow after imatinib administration, together with the bone morphogenetic proteins-2 and -7. All patients with high number of CD20(+CD5(+sIgM(+ cells and high stromal-derived factor-1 and B lymphocyte activating factor levels, underwent complete cytogenetic and/or molecular remission by 12 months. We propose that CD20(+CD5(+sIgM(+ lymphocytes producing anti-carbohydrate antibodies with anti-tumor activity, might contribute to the response to imatinib treatment. As in multivariate analysis bone marrow CD20(+CD5(+sIgM(+ cells and stromal-derived factor-1 and B-lymphocyte-activating factor levels were significantly related to cytogenetical and molecular changes, they might contribute to the definition of the pharmacological response.

  7. Role of bone marrow-derived stem cells, renal progenitor cells and ...

    African Journals Online (AJOL)

    It remains the leading cause of late allograft loss. Bone marrow derived stem cells are undifferentiated cells typically characterized by their capacity for self renewal, ability to give rise to multiple differentiated cellular population, including hematopoietic (HSCs) and mesenchymal stem cells (MSCs). Characterization of HSCs ...

  8. Embryonic stem cell-like cells derived from adult human testis

    NARCIS (Netherlands)

    Mizrak, S. C.; Chikhovskaya, J. V.; Sadri-Ardekani, H.; van Daalen, S.; Korver, C. M.; Hovingh, S. E.; Roepers-Gajadien, H. L.; Raya, A.; Fluiter, K.; de Reijke, Th M.; de la Rosette, J. J. M. C. H.; Knegt, A. C.; Belmonte, J. C.; van der Veen, F.; de rooij, D. G.; Repping, S.; van Pelt, A. M. M.

    2010-01-01

    Given the significant drawbacks of using human embryonic stem (hES) cells for regenerative medicine, the search for alternative sources of multipotent cells is ongoing. Studies in mice have shown that multipotent ES-like cells can be derived from neonatal and adult testis. Here we report the

  9. Derivation of novel human ground state naive pluripotent stem cells.

    Science.gov (United States)

    Gafni, Ohad; Weinberger, Leehee; Mansour, Abed AlFatah; Manor, Yair S; Chomsky, Elad; Ben-Yosef, Dalit; Kalma, Yael; Viukov, Sergey; Maza, Itay; Zviran, Asaf; Rais, Yoach; Shipony, Zohar; Mukamel, Zohar; Krupalnik, Vladislav; Zerbib, Mirie; Geula, Shay; Caspi, Inbal; Schneir, Dan; Shwartz, Tamar; Gilad, Shlomit; Amann-Zalcenstein, Daniela; Benjamin, Sima; Amit, Ido; Tanay, Amos; Massarwa, Rada; Novershtern, Noa; Hanna, Jacob H

    2013-12-12

    Mouse embryonic stem (ES) cells are isolated from the inner cell mass of blastocysts, and can be preserved in vitro in a naive inner-cell-mass-like configuration by providing exogenous stimulation with leukaemia inhibitory factor (LIF) and small molecule inhibition of ERK1/ERK2 and GSK3β signalling (termed 2i/LIF conditions). Hallmarks of naive pluripotency include driving Oct4 (also known as Pou5f1) transcription by its distal enhancer, retaining a pre-inactivation X chromosome state, and global reduction in DNA methylation and in H3K27me3 repressive chromatin mark deposition on developmental regulatory gene promoters. Upon withdrawal of 2i/LIF, naive mouse ES cells can drift towards a primed pluripotent state resembling that of the post-implantation epiblast. Although human ES cells share several molecular features with naive mouse ES cells, they also share a variety of epigenetic properties with primed murine epiblast stem cells (EpiSCs). These include predominant use of the proximal enhancer element to maintain OCT4 expression, pronounced tendency for X chromosome inactivation in most female human ES cells, increase in DNA methylation and prominent deposition of H3K27me3 and bivalent domain acquisition on lineage regulatory genes. The feasibility of establishing human ground state naive pluripotency in vitro with equivalent molecular and functional features to those characterized in mouse ES cells remains to be defined. Here we establish defined conditions that facilitate the derivation of genetically unmodified human naive pluripotent stem cells from already established primed human ES cells, from somatic cells through induced pluripotent stem (iPS) cell reprogramming or directly from blastocysts. The novel naive pluripotent cells validated herein retain molecular characteristics and functional properties that are highly similar to mouse naive ES cells, and distinct from conventional primed human pluripotent cells. This includes competence in the generation

  10. Transfection of bone marrow derived cells with immunoregulatory proteins.

    Science.gov (United States)

    Khantakova, Julia N; Silkov, Alexander N; Tereshchenko, Valeriy P; Gavrilova, Elena V; Maksyutov, Rinat A; Sennikov, Sergey V

    2018-03-23

    In vitro electroporation gene transfer was first performed in 1982. Today, this technology has become one of the major vehicles for non-viral transfection of cells. All non-viral transfections, such as calcium phosphate precipitation, lipofection, and magnetic transfection, have been shown to achieve a transfection efficiency of up to 70% in commonly used cell lines, but not in primary cells. Here we describe the use of electroporation to transfect primary mouse bone marrow-derived cells, such as macrophages (Mφ) and dendritic cells (DCs) with high efficiencies (45%-72%) and minimal cell death. The transfection efficiencies and cell death varied depending on the culture duration of the DCs and Mφ. Moreover, the electroporation efficiency was increased when conditioning medium was used for culturing the cells. Furthermore, we demonstrated that measuring the plasmid-encoded secreted proteins is a highly sensitive method for determining the transfection efficiency. In summary, electroporation with plasmid vectors is an efficient method for producing DCs and Mφ with transient expression of immunoregulatory proteins. Copyright © 2018 Elsevier Ltd. All rights reserved.

  11. Myeloid-derived suppressor cells in breast cancer.

    Science.gov (United States)

    Markowitz, Joseph; Wesolowski, Robert; Papenfuss, Tracey; Brooks, Taylor R; Carson, William E

    2013-07-01

    Myeloid-derived suppressor cells (MDSCs) are a population of immature myeloid cells defined by their suppressive actions on immune cells such as T cells, dendritic cells, and natural killer cells. MDSCs typically are positive for the markers CD33 and CD11b but express low levels of HLADR in humans. In mice, MDSCs are typically positive for both CD11b and Gr1. These cells exert their suppressive activity on the immune system via the production of reactive oxygen species, arginase, and cytokines. These factors subsequently inhibit the activity of multiple protein targets such as the T cell receptor, STAT1, and indoleamine-pyrrole 2,3-dioxygenase. The numbers of MDSCs tend to increase with cancer burden while inhibiting MDSCs improves disease outcome in murine models. MDSCs also inhibit immune cancer therapeutics. In light of the poor prognosis of metastatic breast cancer in women and the correlation of increasing levels of MDSCs with increasing disease burden, the purposes of this review are to (1) discuss why MDSCs may be important in breast cancer, (2) describe model systems used to study MDSCs in vitro and in vivo, (3) discuss mechanisms involved in MDSC induction/function in breast cancer, and (4) present pre-clinical and clinical studies that explore modulation of the MDSC-immune system interaction in breast cancer. MDSCs inhibit the host immune response in breast cancer patients and diminishing MDSC actions may improve therapeutic outcomes.

  12. 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.

  13. Inferring time derivatives including cell growth rates using Gaussian processes

    Science.gov (United States)

    Swain, Peter S.; Stevenson, Keiran; Leary, Allen; Montano-Gutierrez, Luis F.; Clark, Ivan B. N.; Vogel, Jackie; Pilizota, Teuta

    2016-12-01

    Often the time derivative of a measured variable is of as much interest as the variable itself. For a growing population of biological cells, for example, the population's growth rate is typically more important than its size. Here we introduce a non-parametric method to infer first and second time derivatives as a function of time from time-series data. Our approach is based on Gaussian processes and applies to a wide range of data. In tests, the method is at least as accurate as others, but has several advantages: it estimates errors both in the inference and in any summary statistics, such as lag times, and allows interpolation with the corresponding error estimation. As illustrations, we infer growth rates of microbial cells, the rate of assembly of an amyloid fibril and both the speed and acceleration of two separating spindle pole bodies. Our algorithm should thus be broadly applicable.

  14. 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.

  15. Tumor microenvironment derived exosomes pleiotropically modulate cancer cell metabolism.

    Science.gov (United States)

    Zhao, Hongyun; Yang, Lifeng; Baddour, Joelle; Achreja, Abhinav; Bernard, Vincent; Moss, Tyler; Marini, Juan C; Tudawe, Thavisha; Seviour, Elena G; San Lucas, F Anthony; Alvarez, Hector; Gupta, Sonal; Maiti, Sourindra N; Cooper, Laurence; Peehl, Donna; Ram, Prahlad T; Maitra, Anirban; Nagrath, Deepak

    2016-02-27

    Cancer-associated fibroblasts (CAFs) are a major cellular component of tumor microenvironment in most solid cancers. Altered cellular metabolism is a hallmark of cancer, and much of the published literature has focused on neoplastic cell-autonomous processes for these adaptations. We demonstrate that exosomes secreted by patient-derived CAFs can strikingly reprogram the metabolic machinery following their uptake by cancer cells. We find that CAF-derived exosomes (CDEs) inhibit mitochondrial oxidative phosphorylation, thereby increasing glycolysis and glutamine-dependent reductive carboxylation in cancer cells. Through 13C-labeled isotope labeling experiments we elucidate that exosomes supply amino acids to nutrient-deprived cancer cells in a mechanism similar to macropinocytosis, albeit without the previously described dependence on oncogenic-Kras signaling. Using intra-exosomal metabolomics, we provide compelling evidence that CDEs contain intact metabolites, including amino acids, lipids, and TCA-cycle intermediates that are avidly utilized by cancer cells for central carbon metabolism and promoting tumor growth under nutrient deprivation or nutrient stressed conditions.

  16. Tumor microenvironment derived exosomes pleiotropically modulate cancer cell metabolism

    Science.gov (United States)

    Zhao, Hongyun; Yang, Lifeng; Baddour, Joelle; Achreja, Abhinav; Bernard, Vincent; Moss, Tyler; Marini, Juan C; Tudawe, Thavisha; Seviour, Elena G; San Lucas, F Anthony; Alvarez, Hector; Gupta, Sonal; Maiti, Sourindra N; Cooper, Laurence; Peehl, Donna; Ram, Prahlad T; Maitra, Anirban; Nagrath, Deepak

    2016-01-01

    Cancer-associated fibroblasts (CAFs) are a major cellular component of tumor microenvironment in most solid cancers. Altered cellular metabolism is a hallmark of cancer, and much of the published literature has focused on neoplastic cell-autonomous processes for these adaptations. We demonstrate that exosomes secreted by patient-derived CAFs can strikingly reprogram the metabolic machinery following their uptake by cancer cells. We find that CAF-derived exosomes (CDEs) inhibit mitochondrial oxidative phosphorylation, thereby increasing glycolysis and glutamine-dependent reductive carboxylation in cancer cells. Through 13C-labeled isotope labeling experiments we elucidate that exosomes supply amino acids to nutrient-deprived cancer cells in a mechanism similar to macropinocytosis, albeit without the previously described dependence on oncogenic-Kras signaling. Using intra-exosomal metabolomics, we provide compelling evidence that CDEs contain intact metabolites, including amino acids, lipids, and TCA-cycle intermediates that are avidly utilized by cancer cells for central carbon metabolism and promoting tumor growth under nutrient deprivation or nutrient stressed conditions. DOI: http://dx.doi.org/10.7554/eLife.10250.001 PMID:26920219

  17. 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

  18. In vitro proliferation of normal and leukemic human leukocytes controlled by an inhibitory endopeptide. [/sup 3/H-TdR incorporation

    Energy Technology Data Exchange (ETDEWEB)

    Balazs, A; Mann, J; Takacsi-Nagy, L; Zimonyi, I; Molnar, A; Klupp, T [Inst. of Experimental Medicine, Budapest (Hungary); Istvan Municipal Hospital, Budapest (Hungary); Heim Pal Children' s Hospital Budapest, (Hungary))

    1983-01-01

    GI-3, an endogenous inhibitory fraction isolated from leukocytes, selectively inhibits the proliferation of granuloid precursor cells in a non-toxic manner. Its active principle is an acidic chlor-tolidine positive decapeptide. The in vitro effect on normal and acute leukemic human bone marrow and blood cells was examined. A dose dependent inhibition by GI-3 of /sup 3/H-TdR incorporation into myeloid cells of normal bone marrow was found, the sensitivity of human cells being higher than that of rat cells. The proliferation of the target leukemic bone marrow and blood cells was also decreased by the endogenous inhibitor in a dose-dependent manner in untreated subjects as well as in patients in remission or relapse. The rate of inhibition of leukemic cell proliferation in the short-term suspension system examined almost coincided with the action of well-known cytostatics applied for comparison. Beyond its direct cytostatic effect, GI-3 could be used in the differential diagnosis of blastic leukemias, complementing the routine cytochemical methods.

  19. Skin appendage-derived stem cells: cell biology and potential for wound repair.

    Science.gov (United States)

    Xie, Jiangfan; Yao, Bin; Han, Yutong; Huang, Sha; Fu, Xiaobing

    2016-01-01

    Stem cells residing in the epidermis and skin appendages are imperative for skin homeostasis and regeneration. These stem cells also participate in the repair of the epidermis after injuries, inducing restoration of tissue integrity and function of damaged tissue. Unlike epidermis-derived stem cells, comprehensive knowledge about skin appendage-derived stem cells remains limited. In this review, we summarize the current knowledge of skin appendage-derived stem cells, including their fundamental characteristics, their preferentially expressed biomarkers, and their potential contribution involved in wound repair. Finally, we will also discuss current strategies, future applications, and limitations of these stem cells, attempting to provide some perspectives on optimizing the available therapy in cutaneous repair and regeneration.

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

    International Nuclear Information System (INIS)

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

    2011-01-01

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

  1. DENTAL CONSIDERATIONS FOR LEUKEMIC PEDIATRIC PATIENTS: AN UPDATED REVIEW FOR GENERAL DENTAL PRACTITIONER.

    Science.gov (United States)

    Lowal, Kholoud A; Alaizari, Nader Ahmed; Tarakji, Bassel; Petro, Waleed; Hussain, Khaja Amjad; Altamimi, Mohamed Abdullah Alsakran

    2015-10-01

    The early signs of leukemia can usually manifest in the oral cavity due to infiltration of leukemic cells or due to associated decline in normal marrow elements, especially in the acute phase of leukemia, as common lesions at this stage of the disease can be screened and diagnosed by the dentist. Therefore, the dental community should be aware of the oral manifestations of leukemia and oral complications of anticancer treatment. This can eliminate the oral symptoms of the disease and to improve quality of life for these patients. An extensive search in PubMed line using a combination of terms like "leukemia, children, dental, Acute lymphoblastic leukemia, pediatric" for last ten years was made. Reviews and case reports concerned about acute lymphoblastic leukemia in children were all collected and analyzed and data were extracted. Accordingly, the aim of this review is to highlight on the oral presentations of leukemia in children attending dental clinics and the management of its undesirable side effects.

  2. The 6-chromanol derivate SUL-109 enables prolonged hypothermic storage of adipose tissue-derived stem cells

    NARCIS (Netherlands)

    Hajmousa, Ghazaleh; Vogelaar, Pieter; Brouwer, Linda A.; Graaf, Adrianus Cornelis van der; Henning, Robert H.; Krenning, Guido

    Encouraging advances in cell therapy research with adipose derived stem cells (ASC) require an effective short-term preservation method that provides time for quality control and transport of cells from their manufacturing facility to their clinical destination. Hypothermic storage of cells in their

  3. Comparative study of adipose-derived stem cells and bone marrow-derived stem cells in similar microenvironmental conditions

    Energy Technology Data Exchange (ETDEWEB)

    Guneta, Vipra [Division of Materials Technology, School of Materials Science and Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore 639798 (Singapore); Tan, Nguan Soon [School of Biological Science, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore); KK Research Centre, KK Women' s and Children Hospital, 100 Bukit Timah Road, Singapore 229899 (Singapore); Institute of Molecular and Cell Biology, Agency for Science Technology & Research - A*STAR, 61 Biopolis Drive, Proteos, Singapore 138673 (Singapore); Chan, Soon Kiat Jeremy [School of Biological Science, Nanyang Technological University, 60 Nanyang Drive, Singapore 637551 (Singapore); Tanavde, Vivek [Bioinformatics Institute, Agency for Science Technology & Research - A*STAR, 30 Biopolis Street, Matrix, Singapore 138671 (Singapore); Lim, Thiam Chye [Division of Plastic, Reconstructive and Aesthetic Surgery, Department of Surgery, National University Hospital (NUH) and National University of Singapore (NUS), Kent Ridge Wing, Singapore 119074 (Singapore); Wong, Thien Chong Marcus [Plastic, Reconstructive and Aesthetic Surgery Section, Tan Tock Seng Hospital (TTSH), 11, Jalan Tan Tock Seng, Singapore 308433 (Singapore); Choong, Cleo, E-mail: cleochoong@ntu.edu.sg [Division of Materials Technology, School of Materials Science and Engineering, Nanyang Technological University, 50 Nanyang Avenue, Singapore 639798 (Singapore); KK Research Centre, KK Women' s and Children Hospital, 100 Bukit Timah Road, Singapore 229899 (Singapore)

    2016-11-01

    Mesenchymal stem cells (MSCs), which were first isolated from the bone marrow, are now being extracted from various other tissues in the body, including the adipose tissue. The current study presents systematic evidence of how the adipose tissue-derived stem cells (ASCs) and bone marrow-derived mesenchymal stem cells (Bm-MSCs) behave when cultured in specific pro-adipogenic microenvironments. The cells were first characterized and identified as MSCs in terms of their morphology, phenotypic expression, self-renewal capabilities and multi-lineage potential. Subsequently, the proliferation and gene expression profiles of the cell populations cultured on two-dimensional (2D) adipose tissue extracellular matrix (ECM)-coated tissue culture plastic (TCP) and in three-dimensional (3D) AlgiMatrix® microenvironments were analyzed. Overall, it was found that adipogenesis was triggered in both cell populations due to the presence of adipose tissue ECM. However, in 3D microenvironments, ASCs and Bm-MSCs were predisposed to the adipogenic and osteogenic lineages respectively. Overall, findings from this study will contribute to ongoing efforts in adipose tissue engineering as well as provide new insights into the role of the ECM and cues provided by the immediate microenvironment for stem cell differentiation. - Highlights: • Native adipose tissue ECM coated on 2D TCP triggers adipogenesis in both ASCs and Bm-MSCs. • A 3D microenvironment with similar stiffness to adipose tissue induces adipogenic differentiation of ASCs. • ASCs cultured in 3D alginate scaffolds exhibit predisposition to adipogenesis. • Bm-MSCs cultured in 3D alginate scaffolds exhibit predisposition to osteogenesis. • The native microenvironment of the cells affects their differentiation behaviour in vitro.

  4. Isolation of Mature (Peritoneum-Derived Mast Cells and Immature (Bone Marrow-Derived Mast Cell Precursors from Mice.

    Directory of Open Access Journals (Sweden)

    Steffen K Meurer

    Full Text Available Mast cells (MCs are a versatile cell type playing key roles in tissue morphogenesis and host defence against bacteria and parasites. Furthermore, they can enhance immunological danger signals and are implicated in inflammatory disorders like fibrosis. This granulated cell type originates from the myeloid lineage and has similarities to basophilic granulocytes, both containing large quantities of histamine and heparin. Immature murine mast cells mature in their destination tissue and adopt either the connective tissue (CTMC or mucosal (MMC type. Some effector functions are executed by activation/degranulation of MCs which lead to secretion of a typical set of MC proteases (MCPT and of the preformed or newly synthesized mediators from its granules into the local microenvironment. Due to the potential accumulation of mutations in key signalling pathway components of corresponding MC cell-lines, primary cultured MCs are an attractive mean to study general features of MC biology and aspects of MC functions relevant to human disease. Here, we describe a simple protocol for the simultaneous isolation of mature CTMC-like murine MCs from the peritoneum (PMCs and immature MC precursors from the bone marrow (BM. The latter are differentiated in vitro to yield BM-derived MCs (BMMC. These cells display the typical morphological and phenotypic features of MCs, express the typical MC surface markers, and can be propagated and kept in culture for several weeks. The provided protocol allows simple amplification of large quantities of homogenous, non-transformed MCs from the peritoneum and bone marrow-derived mast cells for cell- and tissue-based biomedical research.

  5. Comparative study of adipose-derived stem cells and bone marrow-derived stem cells in similar microenvironmental conditions

    International Nuclear Information System (INIS)

    Guneta, Vipra; Tan, Nguan Soon; Chan, Soon Kiat Jeremy; Tanavde, Vivek; Lim, Thiam Chye; Wong, Thien Chong Marcus; Choong, Cleo

    2016-01-01

    Mesenchymal stem cells (MSCs), which were first isolated from the bone marrow, are now being extracted from various other tissues in the body, including the adipose tissue. The current study presents systematic evidence of how the adipose tissue-derived stem cells (ASCs) and bone marrow-derived mesenchymal stem cells (Bm-MSCs) behave when cultured in specific pro-adipogenic microenvironments. The cells were first characterized and identified as MSCs in terms of their morphology, phenotypic expression, self-renewal capabilities and multi-lineage potential. Subsequently, the proliferation and gene expression profiles of the cell populations cultured on two-dimensional (2D) adipose tissue extracellular matrix (ECM)-coated tissue culture plastic (TCP) and in three-dimensional (3D) AlgiMatrix® microenvironments were analyzed. Overall, it was found that adipogenesis was triggered in both cell populations due to the presence of adipose tissue ECM. However, in 3D microenvironments, ASCs and Bm-MSCs were predisposed to the adipogenic and osteogenic lineages respectively. Overall, findings from this study will contribute to ongoing efforts in adipose tissue engineering as well as provide new insights into the role of the ECM and cues provided by the immediate microenvironment for stem cell differentiation. - Highlights: • Native adipose tissue ECM coated on 2D TCP triggers adipogenesis in both ASCs and Bm-MSCs. • A 3D microenvironment with similar stiffness to adipose tissue induces adipogenic differentiation of ASCs. • ASCs cultured in 3D alginate scaffolds exhibit predisposition to adipogenesis. • Bm-MSCs cultured in 3D alginate scaffolds exhibit predisposition to osteogenesis. • The native microenvironment of the cells affects their differentiation behaviour in vitro.

  6. Interaction between adipose tissue-derived mesenchymal stem cells and regulatory T-cells

    OpenAIRE

    Engela, Anja; Baan, Carla; Peeters, Anna; Weimar, Willem; Hoogduijn, Martin

    2013-01-01

    textabstractMesenchymal stem cells (MSCs) exhibit immunosuppressive capabilities, which have evoked interest in their application as cell therapy in transplant patients. So far it has been unclear whether allogeneic MSCs and host regulatory T-cells (Tregs) functionally influence each other. We investigated the interaction between both cell types using perirenal adipose tissue-derived MSCs (ASCs) from kidney donors and Tregs from blood bank donors or kidney recipients 6 months after transplant...

  7. SETBP1 mutations drive leukemic transformation in ASXL1-mutated MDS.

    Science.gov (United States)

    Inoue, D; Kitaura, J; Matsui, H; Hou, H-A; Chou, W-C; Nagamachi, A; Kawabata, K C; Togami, K; Nagase, R; Horikawa, S; Saika, M; Micol, J-B; Hayashi, Y; Harada, Y; Harada, H; Inaba, T; Tien, H-F; Abdel-Wahab, O; Kitamura, T

    2015-04-01

    Mutations in ASXL1 are frequent in patients with myelodysplastic syndrome (MDS) and are associated with adverse survival, yet the molecular pathogenesis of ASXL1 mutations (ASXL1-MT) is not fully understood. Recently, it has been found that deletion of Asxl1 or expression of C-terminal-truncating ASXL1-MTs inhibit myeloid differentiation and induce MDS-like disease in mice. Here, we find that SET-binding protein 1 (SETBP1) mutations (SETBP1-MT) are enriched among ASXL1-mutated MDS patients and associated with increased incidence of leukemic transformation, as well as shorter survival, suggesting that SETBP1-MT play a critical role in leukemic transformation of MDS. We identify that SETBP1-MT inhibit ubiquitination and subsequent degradation of SETBP1, resulting in increased expression. Expression of SETBP1-MT, in turn, inhibited protein phosphatase 2A activity, leading to Akt activation and enhanced expression of posterior Hoxa genes in ASXL1-mutant cells. Biologically, SETBP1-MT augmented ASXL1-MT-induced differentiation block, inhibited apoptosis and enhanced myeloid colony output. SETBP1-MT collaborated with ASXL1-MT in inducing acute myeloid leukemia in vivo. The combination of ASXL1-MT and SETBP1-MT activated a stem cell signature and repressed the tumor growth factor-β signaling pathway, in contrast to the ASXL1-MT-induced MDS model. These data reveal that SETBP1-MT are critical drivers of ASXL1-mutated MDS and identify several deregulated pathways as potential therapeutic targets in high-risk MDS.

  8. Chromosomal Aberrations Associated with Clonal Evolution and Leukemic Transformation in Fanconi Anemia: Clinical and Biological Implications

    Directory of Open Access Journals (Sweden)

    Stefan Meyer

    2012-01-01

    Full Text Available Fanconi anaemia (FA is an inherited disease with congenital and developmental abnormalities, bone marrow failure, and extreme risk of leukemic transformation. Bone marrow surveillance is an important part of the clinical management of FA and often reveals cytogenetic aberrations. Here, we review bone marrow findings in FA and discuss the clinical and biological implications of chromosomal aberrations associated with leukemic transformation.

  9. Determination of Elements in Normal and Leukemic Human Whole Blood by Neutron Activation Analysis

    Energy Technology Data Exchange (ETDEWEB)

    Brune, D; Frykberg, B; Samsahl, K; Wester, P O

    1961-11-15

    By means of gamma-spectrometry the following elements were simultaneously determined in normal and leukemic human whole blood: Cu, Mn, Zn, Sr, Na, P, Ca, Rb, Cd, Sb, Au, Cs and Fe. Chemical separations were performed according to a group separation method using ion-exchange technique. No significant difference between the concentrations of the elements in normal- and leukemic blood was observed.

  10. Determination of Elements in Normal and Leukemic Human Whole Blood by Neutron Activation Analysis

    International Nuclear Information System (INIS)

    Brune, D.; Frykberg, B.; Samsahl, K.; Wester, P.O.

    1961-11-01

    By means of gamma-spectrometry the following elements were simultaneously determined in normal and leukemic human whole blood: Cu, Mn, Zn, Sr, Na, P, Ca, Rb, Cd, Sb, Au, Cs and Fe. Chemical separations were performed according to a group separation method using ion-exchange technique. No significant difference between the concentrations of the elements in normal- and leukemic blood was observed

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

    Directory of Open Access Journals (Sweden)

    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

  12. 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.

  13. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    Energy Technology Data Exchange (ETDEWEB)

    Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing, E-mail: caijingmmm@hotmail.com; Wang, Zehua, E-mail: zehuawang@163.net

    2015-09-10

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs.

  14. Adipose-derived mesenchymal stem cells promote cell proliferation and invasion of epithelial ovarian cancer

    International Nuclear Information System (INIS)

    Chu, Yijing; Tang, Huijuan; Guo, Yan; Guo, Jing; Huang, Bangxing; Fang, Fang; Cai, Jing; Wang, Zehua

    2015-01-01

    Adipose-derived mesenchymal stem cell (ADSC) is an important component of tumor microenvironment. However, whether ADSCs have a hand in ovarian cancer progression remains unclear. In this study, we investigated the impact of human ADSCs derived from the omentum of normal donors on human epithelial ovarian cancer (EOC) cells in vitro and in vivo. Direct and indirect co-culture models including ADSCs and human EOC cell lines were established and the effects of ADSCs on EOC cell proliferation were evaluated by EdU incorporation and flow cytometry. Transwell migration assays and detection of MMPs were performed to assess the invasion activity of EOC cells in vitro. Mouse models were established by intraperitoneal injection of EOC cells with or without concomitant ADSCs to investigate the role of ADSCs in tumor progression in vivo. We found that ADSCs significantly promoted proliferation and invasion of EOC cells in both direct and indirect co-culture assays. In addition, after co-culture with ADSCs, EOC cells secreted higher levels of matrix metalloproteinases (MMPs), and inhibition of MMP2 and MMP9 partially relieved the tumor-promoting effects of ADSCs in vitro. In mouse xenograft models, we confirmed that ADSCs promoted EOC growth and metastasis and elevated the expression of MMP2 and MMP9. Our findings indicate that omental ADSCs play a promotive role during ovarian cancer progression. - Highlights: • Omental adipose derived stem cells enhanced growth and invasion properties of ovarian cancer cells. • Adipose derived stem cells promoted the growth and metastasis of ovarian cancer in mice models. • Adipose derived stem cells promoted MMPs expression and secretion of ovarian cancer cells. • Elevated MMPs mediated the tumor promoting effects of ADSCs

  15. Adipose tissue-derived stem cells in neural regenerative medicine.

    Science.gov (United States)

    Yeh, Da-Chuan; Chan, Tzu-Min; Harn, Horng-Jyh; Chiou, Tzyy-Wen; Chen, Hsin-Shui; Lin, Zung-Sheng; Lin, Shinn-Zong

    2015-01-01

    Adipose tissue-derived stem cells (ADSCs) have two essential characteristics with regard to regenerative medicine: the convenient and efficient generation of large numbers of multipotent cells and in vitro proliferation without a loss of stemness. The implementation of clinical trials has prompted widespread concern regarding safety issues and has shifted research toward the therapeutic efficacy of stem cells in dealing with neural degeneration in cases such as stroke, amyotrophic lateral sclerosis, Parkinson's disease, Alzheimer's disease, Huntington's disease, cavernous nerve injury, and traumatic brain injury. Most existing studies have reported that cell therapies may be able to replenish lost cells and promote neuronal regeneration, protect neuronal survival, and play a role in overcoming permanent paralysis and loss of sensation and the recovery of neurological function. The mechanisms involved in determining therapeutic capacity remain largely unknown; however, this concept can still be classified in a methodical manner by citing current evidence. Possible mechanisms include the following: 1) the promotion of angiogenesis, 2) the induction of neuronal differentiation and neurogenesis, 3) reductions in reactive gliosis, 4) the inhibition of apoptosis, 5) the expression of neurotrophic factors, 6) immunomodulatory function, and 7) facilitating neuronal integration. In this study, several human clinical trials using ADSCs for neuronal disorders were investigated. It is suggested that ADSCs are one of the choices among various stem cells for translating into clinical application in the near future.

  16. CULTIVATION OF HUMAN LIVER CELLS AND ADIPOSE-DERIVED MESENCHYMAL STROMAL CELLS IN PERFUSION BIOREACTOR

    Directory of Open Access Journals (Sweden)

    Yu. В. Basok

    2018-01-01

    Full Text Available Aim: to show the progress of the experiment of cultivation of human liver cells and adipose-derived mesenchymal stromal cells in perfusion bioreactor.Materials and methods. The cultivation of a cell-engineered construct, consisting of a biopolymer microstructured collagen-containing hydrogel, human liver cells, adipose-derived mesenchymal stromal cells, and William’s E Medium, was performed in a perfusion bioreactor.Results. On the 7th day large cells with hepatocyte morphology – of a polygonal shape and a centrally located round nucleus, – were present in the culture chambers of the bioreactor. The metabolic activity of hepatocytes in cell-engineered constructs was confi rmed by the presence of urea in the culture medium on the seventh day of cultivation in the bioreactor and by the resorption of a biopolymer microstructured collagen-containing hydrogel.

  17. 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.

  18. Generating a non-integrating human induced pluripotent stem cell bank from urine-derived cells.

    Directory of Open Access Journals (Sweden)

    Yanting Xue

    Full Text Available Induced pluripotent stem cell (iPS cell holds great potential for applications in regenerative medicine, drug discovery, and disease modeling. We describe here a practical method to generate human iPS cells from urine-derived cells (UCs under feeder-free, virus-free, serum-free condition and without oncogene c-MYC. We showed that this approach could be applied in a large population with different genetic backgrounds. UCs are easily accessible and exhibit high reprogramming efficiency, offering advantages over other cell types used for the purpose of iPS generation. Using the approach described in this study, we have generated 93 iPS cell lines from 20 donors with diverse genetic backgrounds. The non-viral iPS cell bank with these cell lines provides a valuable resource for iPS cells research, facilitating future applications of human iPS cells.

  19. Comparison of Cytotoxic Activity in Leukemic Lineages Reveals Important Features of β-Hairpin Antimicrobial Peptides.

    Science.gov (United States)

    Buri, Marcus V; Torquato, Heron F Vieira; Barros, Carlos Castilho; Ide, Jaime S; Miranda, Antonio; Paredes-Gamero, Edgar J

    2017-07-01

    Several reports described different modes of cell death triggered by antimicrobial peptides (AMPs) due to direct effects on membrane disruption, and more recently by apoptosis and necrosis-like patterns. Cytotoxic curves of four β-hairpin AMPs (gomesin, protegrin, tachyplesin, and polyphemusin) were obtained from several human leukemic lineages and normal monocytes and Two cell lines were then selected based on their cytotoxic sensitivity. One was sensitive to AMPs (K562) and the other resistant (KG-1) and their effect compared between these lineages. Thus, these lineages were chosen to further investigate biological features related with their cytotoxicities to AMPs. Stimulation with AMPs produced cell death, with activation of caspase-3, in K562 lineage. Increase on the fluidity of plasmatic membrane by reducing cholesterol potentiated cytotoxicity of AMPs in both lineages. Quantification of internal and external gomesin binding to the cellular membrane of both K562 and KG-1 cells showed that more peptide is accumulated inside of K562 cells. Additionally, evaluation of multi-drug resistant pumps activity showed that KG-1 has more activity than K562 lineage. A comparison of intrinsic gene patterns showed great differences between K562 and KG-1, but stimulation with gomesin promoted few changes in gene expression patterns. Differences in internalization process through the plasma membrane, multidrug resistance pumps activity, and gene expression pattern are important features to AMPs regulated cell death. J. Cell. Biochem. 118: 1764-1773, 2017. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.

  20. Adipose-Derived Stem Cells Respond to Increased Osmolarities.

    Directory of Open Access Journals (Sweden)

    Urška Potočar

    Full Text Available Cell therapies present a feasible option for the treatment of degenerated cartilaginous and intervertebral disc (IVD tissues. Microenvironments of these tissues are specific and often differ from the microenvironment of cells that, could be potentially used for therapy, e.g. human adipose-derived stem cells (hASC. To ensure safe and efficient implantation of hASC, it is important to evaluate how microenvironmental conditions at the site of implantation affect the implanted cells. This study has demonstrated that cartilaginous tissue-specific osmolarities ranging from 400-600 mOsm/L affected hASC in a dose- and time-dependent fashion in comparison to 300 mOsm/L. Increased osmolarities resulted in transient (nuclear DNA and actin reorganisation and non-transient, long-term morphological changes (vesicle formation, increase in cell area, and culture morphology, as well as reduced proliferation in monolayer cultures. Increased osmolarities diminished acid proteoglycan production and compactness of chondrogenically induced pellet cultures, indicating decreased chondrogenic potential. Viability of hASC was strongly dependent on the type of culture, with hASC in monolayer culture being more tolerant to increased osmolarity compared to hASC in suspension, alginate-agarose hydrogel, and pellet cultures, thus emphasizing the importance of choosing relevant in vitro conditions according to the specifics of clinical application.

  1. Immune Privilege and Eye-Derived T-Regulatory Cells

    Directory of Open Access Journals (Sweden)

    Hiroshi Keino

    2018-01-01

    Full Text Available Certain cellular components of the eye, such as neural retina, are unable to regenerate and replicate after destructive inflammation. Ocular immune privilege provides the eye with immune protection against intraocular inflammation in order to minimize the risk to vision integrity. The eye and immune system use strategies to maintain the ocular immune privilege by regulating the innate and adaptive immune response, which includes immunological ignorance, peripheral tolerance to eye-derived antigens, and intraocular immunosuppressive microenvironment. In this review, we summarize current knowledge regarding the molecular mechanism responsible for the development and maintenance of ocular immune privilege via regulatory T cells (Tregs, which are generated by the anterior chamber-associated immune deviation (ACAID, and ocular resident cells including corneal endothelial (CE cells, ocular pigment epithelial (PE cells, and aqueous humor. Furthermore, we examined the therapeutic potential of Tregs generated by RPE cells that express transforming growth factor beta (TGF-β, cytotoxic T lymphocyte-associated antigen-2 alpha (CTLA-2α, and retinoic acid for autoimmune uveoretinitis and evaluated a new strategy using human RPE-induced Tregs for clinical application in inflammatory ocular disease. We believe that a better understanding of the ocular immune privilege associated with Tregs might offer a new approach with regard to therapeutic interventions for ocular autoimmunity.

  2. Evaluation of cyanobacteria cell count detection derived from ...

    Science.gov (United States)

    Inland waters across the United States (US) are at potential risk for increased outbreaks of toxic cyanobacteria (Cyano) harmful algal bloom (HAB) events resulting from elevated water temperatures and extreme hydrologic events attributable to climate change and increased nutrient loadings associated with intensive agricultural practices. Current monitoring efforts are limited in scope due to resource limitations, analytical complexity, and data integration efforts. The goals of this study were to validate a new ocean color algorithm for satellite imagery that could potentially be used to monitor CyanoHAB events in near real-time to provide a compressive monitoring capability for freshwater lakes (>100 ha). The algorithm incorporated narrow spectral bands specific to the European Space Agency’s (ESA’s) MEdium Resolution Imaging Spectrometer (MERIS) instrument that were optimally oriented at phytoplankton pigment absorption features including phycocyanin at 620 nm. A validation of derived Cyano cell counts was performed using available in situ data assembled from existing monitoring programs across eight states in the eastern US over a 39-month period (2009–2012). Results indicated that MERIS provided robust estimates for Low (10,000–109,000 cells/mL) and Very High (>1,000,000 cells/mL) cell enumeration ranges (approximately 90% and 83%, respectively). However, the results for two intermediate ranges (110,000–299,000 and 300,000–1,000,000 cells/mL)

  3. 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.

  4. Immune Privilege and Eye-Derived T-Regulatory Cells.

    Science.gov (United States)

    Keino, Hiroshi; Horie, Shintaro; Sugita, Sunao

    2018-01-01

    Certain cellular components of the eye, such as neural retina, are unable to regenerate and replicate after destructive inflammation. Ocular immune privilege provides the eye with immune protection against intraocular inflammation in order to minimize the risk to vision integrity. The eye and immune system use strategies to maintain the ocular immune privilege by regulating the innate and adaptive immune response, which includes immunological ignorance, peripheral tolerance to eye-derived antigens, and intraocular immunosuppressive microenvironment. In this review, we summarize current knowledge regarding the molecular mechanism responsible for the development and maintenance of ocular immune privilege via regulatory T cells (Tregs), which are generated by the anterior chamber-associated immune deviation (ACAID), and ocular resident cells including corneal endothelial (CE) cells, ocular pigment epithelial (PE) cells, and aqueous humor. Furthermore, we examined the therapeutic potential of Tregs generated by RPE cells that express transforming growth factor beta (TGF- β ), cytotoxic T lymphocyte-associated antigen-2 alpha (CTLA-2 α ), and retinoic acid for autoimmune uveoretinitis and evaluated a new strategy using human RPE-induced Tregs for clinical application in inflammatory ocular disease. We believe that a better understanding of the ocular immune privilege associated with Tregs might offer a new approach with regard to therapeutic interventions for ocular autoimmunity.

  5. Induced Pluripotent Stem Cell Derived Mesenchymal Stem Cells for Attenuating Age-Related Bone Loss

    Science.gov (United States)

    2012-07-01

    Mesenchymal stem cell (MSC) differentiation towards the bone forming osteoblastic lineage decreases as a function of age and may contribute to age-related...problem of age-related reduced availability of MSC we propose to examine the bone anabolic potential of induced pluripotent stem cell (iPS) derived MSC

  6. 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.

  7. Adipose-derived mesenchymal stromal cells for chronic myocardial ischemia (MyStromalCell Trial)

    DEFF Research Database (Denmark)

    Qayyum, Abbas Ali; Haack-Sørensen, Mandana; Mathiasen, Anders Bruun

    2012-01-01

    Adipose tissue represents an abundant, accessible source of multipotent adipose-derived stromal cells (ADSCs). Animal studies have suggested that ADSCs have the potential to differentiate in vivo into endothelial cells and cardiomyocytes. This makes ADSCs a promising new cell source...... for regenerative therapy to replace injured tissue by creating new blood vessels and cardiomyocytes in patients with chronic ischemic heart disease. The aim of this special report is to review the present preclinical data leading to clinical stem cell therapy using ADSCs in patients with ischemic heart disease....... In addition, we give an introduction to the first-in-man clinical trial, MyStromalCell Trial, which is a prospective, randomized, double-blind, placebo-controlled study using culture-expanded ADSCs obtained from adipose-derived cells from abdominal adipose tissue and stimulated with VEGF-A(165) the week...

  8. Potential differentiation of islet-like cells from pregnant cow-derived placental stem cells.

    Science.gov (United States)

    Peng, Shao-Yu; Chou, Chien-Wen; Kuo, Yu-Hsuan; Shen, Perng-Chih; Shaw, S W Steven

    2017-06-01

    Type 1 diabetes is an autoimmune disease that destroys islet cells and results in insufficient insulin secretion by pancreatic β-cells. Islet transplantation from donors is an approach used for treating patients with diabetes; however, this therapy is difficult to implement because of the lack of donors. Nevertheless, several stem cells have the potential to differentiate from islet-like cells and enable insulin secretion for treating diabetes in animal models. For example, placenta is considered a waste material and can be harvested noninvasively during delivery without ethical or moral concerns. To date, the differentiation of islet-like cells from cow-derived placental stem cells (CPSCs) has yet to be demonstrated. The investigation of potential differentiation of islet-like cells from CPSCs was conducted by supplementation with nicotinamide, exendin-4, glucose, and poly-d-lysine and was detected through reverse transcription polymerase chain reaction, dithizone staining, and immunocytochemical methods. Our results indicated that CPSCs are established and express mesenchymal stem cell surface antigen markers, such as CD73, CD166, β-integrin, and Oct-4, but not hematopoietic stem cell surface antigen markers, such as CD45. After induction, the CPSCs successfully differentiated into islet-like cells. The CPSC-derived islet-like cells expressed islet cell development-related genes, such as insulin, glucagon, pax-4, Nkx6.1, pax-6, and Fox. Moreover, CPSC-derived islet-like cells can be stained with zinc ions, which are widely distributed in the islet cells and enable insulin secretion. Altogether, islet-like cells have the potential to be differentiated from CPSCs without gene manipulation, and can be used in diabetic animal models in the future for preclinical and drug testing trial investigations. Copyright © 2017. Published by Elsevier B.V.

  9. A novel application of furazolidone: anti-leukemic activity in acute myeloid leukemia.

    Directory of Open Access Journals (Sweden)

    Xueqing Jiang

    Full Text Available Acute myeloid leukemia (AML is the most common malignant myeloid disorder of progenitor cells in myeloid hematopoiesis and exemplifies a genetically heterogeneous disease. The patients with AML also show a heterogeneous response to therapy. Although all-trans retinoic acid (ATRA has been successfully introduced to treat acute promyelocytic leukemia (APL, it is rather ineffective in non-APL AML. In our present study, 1200 off-patent marketed drugs and natural compounds that have been approved by the Food and Drug Administration (FDA were screened for anti-leukemia activity using the retrovirus transduction/transformation assay (RTTA. Furazolidone (FZD was shown to inhibit bone marrow transformation mediated by several leukemia fusion proteins, including AML1-ETO. Furazolidone has been used in the treatment of certain bacterial and protozoan infections in human and animals for more than sixty years. We investigated the anti-leukemic activity of FZD in a series of AML cells. FZD displayed potent antiproliferative properties at submicromolar concentrations and induced apoptosis in AML cell lines. Importantly, FZD treatment of certain AML cells induced myeloid cell differentiation by morphology and flow cytometry for CD11b expression. Furthermore, FZD treatment resulted in increased stability of tumor suppressor p53 protein in AML cells. Our in vitro results suggest furazolidone as a novel therapeutic strategy in AML patients.

  10. [Thiamine and its derivatives in the regulation of cell metabolism].

    Science.gov (United States)

    Tylicki, Adam; Siemieniuk, Magdalena

    2011-07-06

    For over 70 years thiamine (vitamin B1) has aroused the interest of biologists, biochemists and medical doctors because of its multilateral participation in key biochemical and physiological processes. The thiamine molecule is composed of pyrimidine and thiazole rings which are linked by a methylene bridge. It is synthesized by microorganisms, fungi and plants, whereas animals and humans have to obtain it from food. There are several known forms of vitamin B1 inside cells: free thiamine, three phosphate esters (mono-, di-, and triphosphate), and the recently found adenosine thiamine triphosphate. Thiamine has a dual, coenzymatic and non-coenzymatic role. First of all, it is a precursor of thiamin diphosphate, which is a coenzyme for over 20 characterized enzymes which are involved in cell bioenergetic processes leading to the synthesis of ATP. Moreover, these enzymes take part in the biosynthesis of pentose (required for the synthesis of nucleotides), amino acids and other organic compounds of cell metabolism. On the other hand, recent discoveries show the non-coenzymatic role of thiamine derivatives in the process of regulation of gene expression (riboswitches in microorganisms and plants), the stress response, and perhaps so far unknown signal transduction pathways associated with adverse environmental conditions, or transduction of nerve signals with participation of thiamine triphosphate and adenosine thiamine triphosphate. From the clinical point of view thiamine deficiency is related to beri-beri, Parkinson disease, Alzheimer disease, Wernicke-Korsakoff syndrome and other pathologies of the nervous system, and it is successfully applied in medical practice. On the other hand, identifying new synthetic analogues of thiamine which could be used as cytostatics, herbicides or agents preventing deficiency of vitamin B1 is currently the major goal of the research. In this paper we present the current state of knowledge of thiamine and its derivatives, indicating

  11. 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.

  12. Endothelial cell-derived microparticles induce plasmacytoid dendritic cell maturation: potential implications in inflammatory diseases.

    Science.gov (United States)

    Angelot, Fanny; Seillès, Estelle; Biichlé, Sabeha; Berda, Yael; Gaugler, Béatrice; Plumas, Joel; Chaperot, Laurence; Dignat-George, Françoise; Tiberghien, Pierre; Saas, Philippe; Garnache-Ottou, Francine

    2009-11-01

    Increased circulating endothelial microparticles, resulting from vascular endothelium dysfunction, and plasmacytoid dendritic cell activation are both encountered in common inflammatory disorders. The aim of our study was to determine whether interactions between endothelial microparticles and plasmacytoid dendritic cells could contribute to such pathologies. Microparticles generated from endothelial cell lines, platelets or activated T cells were incubated with human plasmacytoid dendritic cells sorted from healthy donor blood or with monocyte-derived dendritic cells. Dendritic cell maturation was evaluated by flow cytometry, cytokine secretion as well as naive T-cell activation and polarization. Labeled microparticles were also used to study cellular interactions. Endothelial microparticles induced plasmacytoid dendritic cell maturation. In contrast, conventional dendritic cells were resistant to endothelial microparticle-induced maturation. In addition to upregulation of co-stimulatory molecules, endothelial microparticle-matured plasmacytoid dendritic cells secreted inflammatory cytokines (interleukins 6 and 8, but no interferon-alpha) and also induced allogeneic naive CD4(+) T cells to proliferate and to produce type 1 cytokines such as interferon-gamma and tumor necrosis factor-alpha. Endothelial microparticle endocytosis by plasmacytoid dendritic cells appeared to be required for plasmacytoid dendritic cell maturation. Importantly, the ability of endothelial microparticles to induce plasmacytoid dendritic cells to mature was specific as microparticles derived from activated T cells or platelets (the major source of circulating microparticules in healthy subjects) did not induce such plasmacytoid dendritic cell maturation. Our data show that endothelial microparticles specifically induce plasmacytoid dendritic cell maturation and production of inflammatory cytokines. This novel activation pathway may be implicated in various inflammatory disorders and

  13. Exosomes derived from mesenchymal non-small cell lung cancer cells promote chemoresistance.

    Science.gov (United States)

    Lobb, Richard J; van Amerongen, Rosa; Wiegmans, Adrian; Ham, Sunyoung; Larsen, Jill E; Möller, Andreas

    2017-08-01

    Non-small cell lung cancer (NSCLC) is the most common lung cancer type and the most common cause of mortality in lung cancer patients. NSCLC is often associated with resistance to chemotherapeutics and together with rapid metastatic spread, results in limited treatment options and poor patient survival. NSCLCs are heterogeneous, and consist of epithelial and mesenchymal NSCLC cells. Mesenchymal NSCLC cells are thought to be responsible for the chemoresistance phenotype, but if and how this phenotype can be transferred to other NSCLC cells is currently not known. We hypothesised that small extracellular vesicles, exosomes, secreted by mesenchymal NSCLC cells could potentially transfer the chemoresistance phenotype to surrounding epithelial NSCLC cells. To explore this possibility, we used a unique human bronchial epithelial cell (HBEC) model in which the parental cells were transformed from an epithelial to mesenchymal phenotype by introducing oncogenic alterations common in NSCLC. We found that exosomes derived from the oncogenically transformed, mesenchymal HBECs could transfer chemoresistance to the parental, epithelial HBECs and increase ZEB1 mRNA, a master EMT transcription factor, in the recipient cells. Additionally, we demonstrate that exosomes from mesenchymal, but not epithelial HBECs contain the ZEB1 mRNA, thereby providing a potential mechanism for the induction of a mesenchymal phenotype in recipient cells. Together, this work demonstrates for the first time that exosomes derived from mesenchymal, oncogenically transformed lung cells can transfer chemoresistance and mesenchymal phenotypes to recipient cells, likely via the transfer of ZEB1 mRNA in exosomes. © 2017 UICC.

  14. Adipose tissue-derived stem cells promote pancreatic cancer cell proliferation and invasion

    International Nuclear Information System (INIS)

    Ji, S.Q.; Cao, J.; Zhang, Q.Y.; Li, Y.Y.; Yan, Y.Q.; Yu, F.X.

    2013-01-01

    To explore the effects of adipose tissue-derived stem cells (ADSCs) on the proliferation and invasion of pancreatic cancer cells in vitro and the possible mechanism involved, ADSCs were cocultured with pancreatic cancer cells, and a cell counting kit (CCK-8) was used to detect the proliferation of pancreatic cancer cells. ELISA was used to determine the concentration of stromal cell-derived factor-1 (SDF-1) in the supernatants. RT-PCR was performed to detect the expression of the chemokine receptor CXCR4 in pancreatic cancer cells and ADSCs. An in vitro invasion assay was used to measure invasion of pancreatic cancer cells. SDF-1 was detected in the supernatants of ADSCs, but not in pancreatic cancer cells. Higher CXCR4 mRNA levels were detected in the pancreatic cancer cell lines compared with ADSCs (109.3±10.7 and 97.6±7.6 vs 18.3±1.7, respectively; P<0.01). In addition, conditioned medium from ADSCs promoted the proliferation and invasion of pancreatic cancer cells, and AMD3100, a CXCR4 antagonist, significantly downregulated these growth-promoting effects. We conclude that ADSCs can promote the proliferation and invasion of pancreatic cancer cells, which may involve the SDF-1/CXCR4 axis

  15. Adipose tissue-derived stem cells promote pancreatic cancer cell proliferation and invasion

    Energy Technology Data Exchange (ETDEWEB)

    Ji, S.Q.; Cao, J. [Department of Liver Surgery I, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai (China); Zhang, Q.Y.; Li, Y.Y. [Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital, Wenzhou Medical College, Wenzhou (China); Yan, Y.Q. [Department of Liver Surgery I, Eastern Hepatobiliary Surgery Hospital, Second Military Medical University, Shanghai (China); Yu, F.X. [Department of Hepatobiliary and Pancreatic Surgery, The First Affiliated Hospital, Wenzhou Medical College, Wenzhou (China)

    2013-09-27

    To explore the effects of adipose tissue-derived stem cells (ADSCs) on the proliferation and invasion of pancreatic cancer cells in vitro and the possible mechanism involved, ADSCs were cocultured with pancreatic cancer cells, and a cell counting kit (CCK-8) was used to detect the proliferation of pancreatic cancer cells. ELISA was used to determine the concentration of stromal cell-derived factor-1 (SDF-1) in the supernatants. RT-PCR was performed to detect the expression of the chemokine receptor CXCR4 in pancreatic cancer cells and ADSCs. An in vitro invasion assay was used to measure invasion of pancreatic cancer cells. SDF-1 was detected in the supernatants of ADSCs, but not in pancreatic cancer cells. Higher CXCR4 mRNA levels were detected in the pancreatic cancer cell lines compared with ADSCs (109.3±10.7 and 97.6±7.6 vs 18.3±1.7, respectively; P<0.01). In addition, conditioned medium from ADSCs promoted the proliferation and invasion of pancreatic cancer cells, and AMD3100, a CXCR4 antagonist, significantly downregulated these growth-promoting effects. We conclude that ADSCs can promote the proliferation and invasion of pancreatic cancer cells, which may involve the SDF-1/CXCR4 axis.

  16. Gold nanoparticles cellular toxicity and recovery: adipose Derived Stromal cells.

    Science.gov (United States)

    Mironava, Tatsiana; Hadjiargyrou, Michael; Simon, Marcia; Rafailovich, Miriam H

    2014-03-01

    Gold nanoparticles (AuNPs) are currently used in numerous medical applications. Herein, we describe their in vitro impact on human adipose-derived stromal cells (ADSCs) using 13 nm and 45 nm citrate-coated AuNPs. In their non-differentiated state, ADSCs were penetrated by the AuNPs and stored in vacuoles. The presence of the AuNPs in ADSCs resulted in increased population doubling times, decreased cell motility and cell-mediated collagen contraction. The degree to which the cells were impacted was a function of particle concentration, where the smaller particles required a sevenfold higher concentration to have the same effect as the larger ones. Furthermore, AuNPs reduced adipogenesis as measured by lipid droplet accumulation and adiponectin secretion. These effects correlated with transient increases in DLK1 and with relative reductions in fibronectin. Upon removal of exogenous AuNPs, cellular NP levels decreased and normal ADSC functions were restored. As adiponectin helps regulate energy metabolism, local fluctuations triggered by AuNPs can lead to systemic changes. Hence, careful choice of size, concentration and clinical application duration of AuNPs is warranted.

  17. 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.

  18. Derivation of novel genetically diverse human embryonic stem cell lines.

    Science.gov (United States)

    Stefanova, Valentina T; Grifo, James A; Hansis, Christoph

    2012-06-10

    Human embryonic stem cells (hESCs) have the potential to revolutionize many biomedical fields ranging from basic research to disease modeling, regenerative medicine, drug discovery, and toxicity testing. A multitude of hESC lines have been derived worldwide since the first 5 lines by Thomson et al. 13 years ago, but many of these are poorly characterized, unavailable, or do not represent desired traits, thus making them unsuitable for application purposes. In order to provide the scientific community with better options, we have derived 12 new hESC lines at New York University from discarded genetically normal and abnormal embryos using the latest techniques. We examined the genetic status of the NYUES lines in detail as well as their molecular and cellular features and DNA fingerprinting profile. Furthermore, we differentiated our hESCs into the tissues most affected by a specific condition or into clinically desired cell types. To our knowledge, a number of characteristics of our hESCs have not been previously reported, for example, mutation for alpha thalassemia X-linked mental retardation syndrome, linkage to conditions with a genetic component such as asthma or poor sperm morphology, and novel combinations of ethnic backgrounds. Importantly, all of our undifferentiated euploid female lines tested to date did not show X chromosome inactivation, believed to result in superior potency. We continue to derive new hESC lines and add them to the NIH registry and other registries. This should facilitate the use of our hESCs and lead to advancements for patient-benefitting applications.

  19. Exosomes derived from pancreatic cancer cells induce activation and profibrogenic activities in pancreatic stellate cells.

    Science.gov (United States)

    Masamune, Atsushi; Yoshida, Naoki; Hamada, Shin; Takikawa, Tetsuya; Nabeshima, Tatsuhide; Shimosegawa, Tooru

    2018-01-01

    Pancreatic cancer cells (PCCs) interact with pancreatic stellate cells (PSCs), which play a pivotal role in pancreatic fibrogenesis, to develop the cancer-conditioned tumor microenvironment. Exosomes are membrane-enclosed nanovesicles, and have been increasingly recognized as important mediators of cell-to-cell communications. The aim of this study was to clarify the effects of PCC-derived exosomes on cell functions in PSCs. Exosomes were isolated from the conditioned medium of Panc-1 and SUIT-2 PCCs. Human primary PSCs were treated with PCC-derived exosomes. PCC-derived exosomes stimulated the proliferation, migration, activation of ERK and Akt, the mRNA expression of α-smooth muscle actin (ACTA2) and fibrosis-related genes, and procollagen type I C-peptide production in PSCs. Ingenuity pathway analysis of the microarray data identified transforming growth factor β1 and tumor necrosis factor as top upstream regulators. PCCs increased the expression of miR-1246 and miR-1290, abundantly contained in PCC-derived exosomes, in PSCs. Overexpression of miR-1290 induced the expression of ACTA2 and fibrosis-related genes in PSCs. In conclusion, PCC-derived exosomes stimulate activation and profibrogenic activities in PSCs. Exosome-mediated interactions between PSCs and PCCs might play a role in the development of the tumor microenvironment. Copyright © 2017 Elsevier Inc. All rights reserved.

  20. Effect of chondrocyte-derived early extracellular matrix on chondrogenesis of placenta-derived mesenchymal stem cells.

    Science.gov (United States)

    Park, Yong-Beom; Seo, Sinji; Kim, Jin-A; Heo, Jin-Chul; Lim, Young-Cheol; Ha, Chul-Won

    2015-06-24

    The extracellular matrix (ECM) surrounding cells contains a variety of proteins that provide structural support and regulate cellular functions. Previous studies have shown that decellularized ECM isolated from tissues or cultured cells can be used to improve cell differentiation in tissue engineering applications. In this study we evaluated the effect of decellularized chondrocyte-derived ECM (CDECM) on the chondrogenesis of human placenta-derived mesenchymal stem cells (hPDMSCs) in a pellet culture system. After incubation with or without chondrocyte-derived ECM in chondrogenic medium for 1 or 3 weeks, the sizes and wet masses of the cell pellets were compared with untreated controls (hPDMSCs incubated in chondrogenic medium without chondrocyte-derived ECM). In addition, histologic analysis of the cell pellets (Safranin O and collagen type II staining) and quantitative reverse transcription-PCR analysis of chondrogenic markers (aggrecan, collagen type II, and SOX9) were carried out. Our results showed that the sizes and masses of hPDMSC pellets incubated with chondrocyte-derived ECM were significantly higher than those of untreated controls. Differentiation of hPDMSCs (both with and without chondrocyte-derived ECM) was confirmed by Safranin O and collagen type II staining. Chondrogenic marker expression and glycosaminoglycan (GAG) levels were significantly higher in hPDMSC pellets incubated with chondrocyte-derived ECM compared with untreated controls, especially in cells precultured with chondrocyte-derived ECM for 7 d. Taken together, these results demonstrate that chondrocyte-derived ECM enhances the chondrogenesis of hPDMSCs, and this effect is further increased by preculture with chondrocyte-derived ECM. This preculture method for hPDMSC chondrogenesis represents a promising approach for cartilage tissue engineering.

  1. 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.

  2. Pericytes derived from adipose-derived stem cells protect against retinal vasculopathy.

    Directory of Open Access Journals (Sweden)

    Thomas A Mendel

    Full Text Available Retinal vasculopathies, including diabetic retinopathy (DR, threaten the vision of over 100 million people. Retinal pericytes are critical for microvascular control, supporting retinal endothelial cells via direct contact and paracrine mechanisms. With pericyte death or loss, endothelial dysfunction ensues, resulting in hypoxic insult, pathologic angiogenesis, and ultimately blindness. Adipose-derived stem cells (ASCs differentiate into pericytes, suggesting they may be useful as a protective and regenerative cellular therapy for retinal vascular disease. In this study, we examine the ability of ASCs to differentiate into pericytes that can stabilize retinal vessels in multiple pre-clinical models of retinal vasculopathy.We found that ASCs express pericyte-specific markers in vitro. When injected intravitreally into the murine eye subjected to oxygen-induced retinopathy (OIR, ASCs were capable of migrating to and integrating with the retinal vasculature. Integrated ASCs maintained marker expression and pericyte-like morphology in vivo for at least 2 months. ASCs injected after OIR vessel destabilization and ablation enhanced vessel regrowth (16% reduction in avascular area. ASCs injected intravitreally before OIR vessel destabilization prevented retinal capillary dropout (53% reduction. Treatment of ASCs with transforming growth factor beta (TGF-β1 enhanced hASC pericyte function, in a manner similar to native retinal pericytes, with increased marker expression of smooth muscle actin, cellular contractility, endothelial stabilization, and microvascular protection in OIR. Finally, injected ASCs prevented capillary loss in the diabetic retinopathic Akimba mouse (79% reduction 2 months after injection.ASC-derived pericytes can integrate with retinal vasculature, adopting both pericyte morphology and marker expression, and provide functional vascular protection in multiple murine models of retinal vasculopathy. The pericyte phenotype demonstrated

  3. 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.

  4. Colloidal silver nanoparticles improve anti-leukemic drug efficacy via amplification of oxidative stress.

    Science.gov (United States)

    Guo, Dawei; Zhang, Junren; Huang, Zhihai; Jiang, Shanxiang; Gu, Ning

    2015-02-01

    Recently, increased reactive oxygen species (ROS) levels and altered redox status in cancer cells have become a novel therapeutic strategy to improve cancer selectivity over normal cells. It has been known that silver nanoparticles (AgNPs) display anti-leukemic activity via ROS overproduction. Hence, we hypothesized that AgNPs could improve therapeutic efficacy of ROS-generating agents against leukemia cells. In the current study, N-(4-hydroxyphenyl)retinamide (4-HPR), a synthetic retinoid, was used as a drug model of ROS induction to investigate its synergistic effect with AgNPs. The data exhibited that AgNPs with uniform size prepared by an electrochemical method could localize in the lysosomes, mitochondria and cytoplasm of SHI-1 cells. More importantly, AgNPs together with 4-HPR could exhibit more cytotoxicity and apoptosis via overproduction of ROS in comparison with that alone. Taken together, these results reveal that AgNPs combined with ROS-generating drugs could potentially enhance therapeutic efficacy against leukemia cells, thereby providing a novel strategy for AgNPs in leukemia therapy. Copyright © 2015. Published by Elsevier B.V.

  5. Comprehensive characterization of genomic instability in pluripotent stem cells and their derived neuroprogenitor cell lines

    Directory of Open Access Journals (Sweden)

    Nestor Luis Lopez Corrales

    2012-12-01

    Full Text Available The genomic integrity of two human pluripotent stem cells and their derived neuroprogenitor cell lines was studied, applying a combination of high-resolution genetic methodologies. The usefulness of combining array-comparative genomic hybridization (aCGH and multiplex fluorescence in situ hybridization (M-FISH techniques should be delineated to exclude/detect a maximum of possible genomic structural aberrations. Interestingly, in parts different genomic imbalances at chromosomal and subchromosomal levels were detected in pluripotent stem cells and their derivatives. Some of the copy number variations were inherited from the original cell line, whereas other modifications were presumably acquired during the differentiation and manipulation procedures. These results underline the necessity to study both pluripotent stem cells and their differentiated progeny by as many approaches as possible in order to assess their genomic stability before using them in clinical therapies.

  6. 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.

  7. Umbilical Cord-Derived Mesenchymal Stem Cells for Hematopoietic Stem Cell Transplantation

    Directory of Open Access Journals (Sweden)

    Yu-Hua Chao

    2012-01-01

    Full Text Available Hematopoietic stem cell transplantation (HSCT is becoming an effective therapeutic modality for a variety of diseases. Mesenchymal stem cells (MSCs can be used to enhance hematopoietic engraftment, accelerate lymphocyte recovery, reduce the risk of graft failure, prevent and treat graft-versus-host disease, and repair tissue damage in patients receiving HSCT. Till now, most MSCs for human clinical application have been derived from bone marrow. However, acquiring bone-marrow-derived MSCs involves an invasive procedure. Umbilical cord is rich with MSCs. Compared to bone-marrow-derived MSCs, umbilical cord-derived MSCs (UCMSCs are easier to obtain without harm to the donor and can proliferate faster. No severe adverse effects were noted in our previous clinical application of UCMSCs in HSCT. Accordingly, application of UCMSCs in humans appears to be feasible and safe. Further studies are warranted.

  8. Safety and immune regulatory properties of canine induced pluripotent stem cell-derived mesenchymal stem cells.

    Science.gov (United States)

    Chow, Lyndah; Johnson, Valerie; Regan, Dan; Wheat, William; Webb, Saiphone; Koch, Peter; Dow, Steven

    2017-12-01

    Mesenchymal stem cells (MSCs) exhibit broad immune modulatory activity in vivo and can suppress T cell proliferation and dendritic cell activation in vitro. Currently, most MSC for clinical usage are derived from younger donors, due to ease of procurement and to the superior immune modulatory activity. However, the use of MSC from multiple unrelated donors makes it difficult to standardize study results and compare outcomes between different clinical trials. One solution is the use of MSC derived from induced pluripotent stem cells (iPSC); as iPSC-derived MSC have nearly unlimited proliferative potential and exhibit in vitro phenotypic stability. Given the value of dogs as a spontaneous disease model for pre-clinical evaluation of stem cell therapeutics, we investigated the functional properties of canine iPSC-derived MSC (iMSC), including immune modulatory properties and potential for teratoma formation. We found that canine iMSC downregulated expression of pluripotency genes and appeared morphologically similar to conventional MSC. Importantly, iMSC retained a stable phenotype after multiple passages, did not form teratomas in immune deficient mice, and did not induce tumor formation in dogs following systemic injection. We concluded therefore that iMSC were phenotypically stable, immunologically potent, safe with respect to tumor formation, and represented an important new source of cells for therapeutic modulation of inflammatory disorders. Copyright © 2017. Published by Elsevier B.V.

  9. Phenotype and Function of CD209+ Bovine Blood Dendritic Cells, Monocyte-Derived-Dendritic Cells and Monocyte-Derived Macrophages.

    Directory of Open Access Journals (Sweden)

    Kun Taek Park

    Full Text Available Phylogenic comparisons of the mononuclear phagocyte system (MPS of humans and mice demonstrate phenotypic divergence of dendritic cell (DC subsets that play similar roles in innate and adaptive immunity. Although differing in phenotype, DC can be classified into four groups according to ontogeny and function: conventional DC (cDC1 and cDC2, plasmacytoid DC (pDC, and monocyte derived DC (MoDC. DC of Artiodactyla (pigs and ruminants can also be sub-classified using this system, allowing direct functional and phenotypic comparison of MoDC and other DC subsets trafficking in blood (bDC. Because of the high volume of blood collections required to study DC, cattle offer the best opportunity to further our understanding of bDC and MoDC function in an outbred large animal species. As reported here, phenotyping DC using a monoclonal antibody (mAb to CD209 revealed CD209 is expressed on the major myeloid population of DC present in blood and MoDC, providing a phenotypic link between these two subsets. Additionally, the present study demonstrates that CD209 is also expressed on monocyte derived macrophages (MoΦ. Functional analysis revealed each of these populations can take up and process antigens (Ags, present them to CD4 and CD8 T cells, and elicit a T-cell recall response. Thus, bDC, MoDC, and MoΦ pulsed with pathogens or candidate vaccine antigens can be used to study factors that modulate DC-driven T-cell priming and differentiation ex vivo.

  10. 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.

  11. Marrow-derived mesenchymal stem cells: role in epithelial tumor cell determination.

    Science.gov (United States)

    Fierro, Fernando A; Sierralta, Walter D; Epuñan, Maria J; Minguell, José J

    2004-01-01

    Marrow stroma represents an advantageous environment for development of micrometastatic cells. Within the cellular structure of marrow stroma, mesenchymal stem cells (MSC) have been postulated as an interacting target for disseminated cancer cells. The studies reported here were performed to gain more information on the interaction of the human breast cancer cell line MCF-7 with human bone marrow-derived MSC cells and to investigate whether this interaction affects tumor cell properties. The results showed that after co-culture with MSC, changes were detected in the morphology, proliferative capacity and aggregation pattern of MCF-7 cells, but these parameters were not affected after the co-culture of MSC cells with a non-tumorigenic breast epithelial cell line, MCF-10. Since the indirect culture of MCF-7 with MSC or its products also resulted in functional changes in the tumor cells, we evaluated whether these effects could be attributed to growth factors produced by MSC cells. It was found that VEGF and IL-6 mimic the effects produced by MSC or its products on the proliferation and aggregation properties of MCF-7, cells, respectively. Thus, it seems that after entry of disseminated tumor cells into the marrow space, their proliferative and morphogenetic organization patterns are modified after interaction with distinct stromal cells and/or with specific signals from the marrow microenvironment.

  12. 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.

  13. Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis

    Energy Technology Data Exchange (ETDEWEB)

    Takahara, Kiyoshi [Department of Urology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Ii, Masaaki, E-mail: masaii@art.osaka-med.ac.jp [Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Inamoto, Teruo; Komura, Kazumasa; Ibuki, Naokazu; Minami, Koichiro; Uehara, Hirofumi; Hirano, Hajime; Nomi, Hayahito; Kiyama, Satoshi [Department of Urology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Asahi, Michio [Department of Pharmacology, Faculty of Medicine, Osaka Medical College, Osaka (Japan); Azuma, Haruhito [Department of Urology, Faculty of Medicine, Osaka Medical College, Osaka (Japan)

    2014-04-18

    Highlights: • AdSC transplantation exhibits inhibitory effect on tumor progressions of PCa cells. • AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway. • High expression of the TGF-β1 gene in AdSCs. - Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferation of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa.

  14. Characterisation of insulin-producing cells differentiated from tonsil derived mesenchymal stem cells.

    Science.gov (United States)

    Kim, So-Yeon; Kim, Ye-Ryung; Park, Woo-Jae; Kim, Han Su; Jung, Sung-Chul; Woo, So-Youn; Jo, Inho; Ryu, Kyung-Ha; Park, Joo-Won

    2015-01-01

    Tonsil-derived (T-) mesenchymal stem cells (MSCs) display mutilineage differentiation potential and self-renewal capacity and have potential as a banking source. Diabetes mellitus is a prevalent disease in modern society, and the transplantation of pancreatic progenitor cells or various stem cell-derived insulin-secreting cells has been suggested as a novel therapy for diabetes. The potential of T-MSCs to trans-differentiate into pancreatic progenitor cells or insulin-secreting cells has not yet been investigated. We examined the potential of human T-MSCs to trans-differentiate into pancreatic islet cells using two different methods based on β-mercaptoethanol and insulin-transferin-selenium, respectively. First, we compared the efficacy of the two methods for inducing differentiation into insulin-producing cells. We demonstrated that the insulin-transferin-selenium method is more efficient for inducing differentiation into insulin-secreting cells regardless of the source of the MSCs. Second, we compared the differentiation potential of two different MSC types: T-MSCs and adipose-derived MSCs (A-MSCs). T-MSCs had a differentiation capacity similar to that of A-MSCs and were capable of secreting insulin in response to glucose concentration. Islet-like clusters differentiated from T-MSCs had lower synaptotagmin-3, -5, -7, and -8 levels, and consequently lower secreted insulin levels than cells differentiated from A-MSCs. These results imply that T-MSCs can differentiate into functional pancreatic islet-like cells and could provide a novel, alternative cell therapy for diabetes mellitus. Copyright © 2015 International Society of Differentiation. Published by Elsevier B.V. All rights reserved.

  15. Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis

    International Nuclear Information System (INIS)

    Takahara, Kiyoshi; Ii, Masaaki; Inamoto, Teruo; Komura, Kazumasa; Ibuki, Naokazu; Minami, Koichiro; Uehara, Hirofumi; Hirano, Hajime; Nomi, Hayahito; Kiyama, Satoshi; Asahi, Michio; Azuma, Haruhito

    2014-01-01

    Highlights: • AdSC transplantation exhibits inhibitory effect on tumor progressions of PCa cells. • AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway. • High expression of the TGF-β1 gene in AdSCs. - Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferation of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa

  16. Mast cells enhance T cell activation: Importance of mast cell-derived TNF

    Science.gov (United States)

    Nakae, Susumu; Suto, Hajime; Kakurai, Maki; Sedgwick, Jonathon D.; Tsai, Mindy; Galli, Stephen J.

    2005-05-01

    Mast cells are not only important effector cells in immediate hypersensitivity reactions and immune responses to pathogens but also can contribute to T cell-mediated disorders. However, the mechanisms by which mast cells might influence T cells in such settings are not fully understood. We find that mast cells can enhance proliferation and cytokine production in multiple T cell subsets. Mast cell-dependent enhancement of T cell activation can be promoted by FcRI-dependent mast cell activation, TNF production by both mast cells and T cells, and mast cell-T cell contact. However, at high concentrations of cells, mast cells can promote T cell activation independent of IgE or TNF. Finally, mast cells also can promote T cell activation by means of soluble factors. These findings identify multiple mechanisms by which mast cells can influence T cell proliferation and cytokine production. allergy | asthma | autoimmunity | cytokines | immune response

  17. Immunologic analyses of mouse cystathionase in normal and leukemic cells

    International Nuclear Information System (INIS)

    Bikel, I.; Faibes, D.; Uren, J.R.; Livingston, D.M.

    1978-01-01

    Rabbit antisera have been raised against mouse liver cystathionase and shown to possess enzyme neutralizing activity. Agar gel double immunodiffusion analyses demonstrated that both mouse liver cystathionase and rat liver cystathionase react with the antisera, the latter enzyme being completely cross-reactive with the former. Following radioiodination of the purified rat liver enzyme, a double antibody radioimmunoassay was developed in which greater than 90% of the labeled protein could be specifically precipitated with the anti-mouse cystathionase antibodies. In this test the purified rat liver and mouse liver enzymes were virtually indistinguishable, generating superimposable competition displacement curves on a protein mass basis. These results indicate that both enzymes are immunologically identical, thus validating the use of the rat in lieu of the murine liver enzyme as radiolabeled tracer in an assay for mouse cystathionase. In addition, competition radioimmunoassays demonstrated that the immunological reactivities of both the purified rat liver and mouse liver enzymes were equally heat sensitive. The sensitivity of the assay was determined to be 1 ng of enzyme protein/0.22 mL of assay mixture, and the assay could be used to detect the presence of enzyme protein in tissue homogenates of single mouse organs. Mouse or rat cross-reactivity with human liver cystathionase was incomplete; but, with the exception of heart and spleen, parallel radioimmunoassay competition displacement curves were obtained for cystathionase from different mouse organs including thymus. Extracts of 7-, 9-, and 10-month-old spontaneous AKR mouse thymomas were tested in the radioimmunoassay along with extracts of age-matched thymuses which were grossly tumor free. A reaction of nonidentity was observed for all of the tumor extracts while a reaction identical with that of the pure liver enzyme was found with all of the normal thymus extracts

  18. HIV-derived vectors for gene therapy targeting dendritic cells.

    Science.gov (United States)

    Rossetti, Maura; Cavarelli, Mariangela; Gregori, Silvia; Scarlatti, Gabriella

    2013-01-01

    Human immunodeficiency virus type 1 (HIV-1)-derived lentiviral vectors (LV) have the potential to mediate stable therapeutic gene transfer. However, similarly to other viral vectors, their benefit is compromised by the induction of an immune response toward transgene-expressing cells that closely mimics antiviral immunity. LV share with the parental HIV the ability to activate dendritic cells (DC), while lack the peculiar ability of subverting DC functions, which is responsible for HIV immune escape. Understanding the interaction between LV and DC, with plasmacytoid and myeloid DC playing fundamental and distinct roles, has paved the way to novel approaches aimed at regulating transgene-specific immune responses. Thanks to the ability to target either DC subsets LV might be a powerful tool to induce immunity (i.e., gene therapy of cancer), cell death (i.e., in HIV/AIDS infection), or tolerance (i.e., gene therapy strategies for monogenic diseases). In this chapter, similarities and differences between the LV-mediated and HIV-mediated induction of immune responses, with specific focus on their interactions with DC, are discussed.

  19. Mesenchymal stem cell-derived microparticles: a promising therapeutic strategy.

    Science.gov (United States)

    Tan, Xi; Gong, Yong-Zhen; Wu, Ping; Liao, Duan-Fang; Zheng, Xi-Long

    2014-08-18

    Mesenchymal stem cells (MSCs) are multipotent stem cells that give rise to various cell types of the mesodermal germ layer. Because of their unique ability to home in on injured and cancerous tissues, MSCs are of great potential in regenerative medicine. MSCs also contribute to reparative processes in different pathological conditions, including cardiovascular diseases and cancer. However, many studies have shown that only a small proportion of transplanted MSCs can actually survive and be incorporated into host tissues. The effects of MSCs cannot be fully explained by their number. Recent discoveries suggest that microparticles (MPs) derived from MSCs may be important for the physiological functions of their parent. Though the physiological role of MSC-MPs is currently not well understood, inspiring results indicate that, in tissue repair and anti-cancer therapy, MSC-MPs have similar pro-regenerative and protective properties as their cellular counterparts. Thus, MSC-MPs represent a promising approach that may overcome the obstacles and risks associated with the use of native or engineered MSCs.

  20. Development of donor-derived thymic lymphomas after allogeneic bone marrow transplantation in AKR/J mice

    International Nuclear Information System (INIS)

    Yasumizu, R.; Hiai, H.; Sugiura, K.

    1988-01-01

    The transplantation of bone marrow cells from BALB/c (but not C57BL/6 and C3H/HeN) mice was observed to lead to the development of thymic lymphomas (leukemias) in AKR/J mice. Two leukemic cell lines, CAK1.3 and CAK4.4, were established from the primary culture of two thymic lymphoma, and surface phenotypes of these cell lines found to be H-2d and Thy-1.2+, indicating that these lymphoma cells are derived from BALB/c donor bone marrow cells. Further analyses of surface markers revealed that CAK1.3 is L3T4+ Lyt2+ IL2R-, whereas CAK4.4 is L3T4- Lyt2- IL2R+. Both CAK1.3 and CAK4.4 were transplantable into BALB/c but not AKR/J mice, further indicating that these cells are of BALB/c bone marrow donor origin. The cells were found to produce XC+-ecotropic viruses, but xenotropic and mink cell focus-forming viruses were undetectable. Inasmuch as thymic lymphomas are derived from bone marrow cells of leukemia-resistant BALB/c strain of mice under the allogeneic environment of leukemia-prone AKR/J mice, this animal model may serve as a useful tool not only for the analysis of leukemic relapse after bone marrow transplantation but also for elucidation of the mechanism of leukemogenesis

  1. Bone marrow-derived mesenchymal stem cells versus adipose-derived mesenchymal stem cells for peripheral nerve regeneration

    Directory of Open Access Journals (Sweden)

    Marcela Fernandes

    2018-01-01

    Full Text Available Studies have confirmed that bone marrow-derived mesenchymal stem cells (MSCs can be used for treatment of several nervous system diseases. However, isolation of bone marrow-derived MSCs (BMSCs is an invasive and painful process and the yield is very low. Therefore, there is a need to search for other alterative stem cell sources. Adipose-derived MSCs (ADSCs have phenotypic and gene expression profiles similar to those of BMSCs. The production of ADSCs is greater than that of BMSCs, and ADSCs proliferate faster than BMSCs. To compare the effects of venous grafts containing BMSCs or ADSCs on sciatic nerve injury, in this study, rats were randomly divided into four groups: sham (only sciatic nerve exposed, Matrigel (MG; sciatic nerve injury + intravenous transplantation of MG vehicle, ADSCs (sciatic nerve injury + intravenous MG containing ADSCs, and BMSCs (sciatic nerve injury + intravenous MG containing BMSCs groups. Sciatic functional index was calculated to evaluate the function of injured sciatic nerve. Morphologic characteristics of nerves distal to the lesion were observed by toluidine blue staining. Spinal motor neurons labeled with Fluoro-Gold were quantitatively assessed. Compared with sham-operated rats, sciatic functional index was lower, the density of small-diameter fibers was significantly increased, and the number of motor neurons significantly decreased in rats with sciatic nerve injury. Neither ADSCs nor BMSCs significantly improved the sciatic nerve function of rats with sciatic nerve injury, increased fiber density, fiber diameters, axonal diameters, myelin sheath thickness, and G ratios (axonal diameter/fiber diameter ratios in the sciatic nerve distal to the lesion site. There was no significant difference in the number of spinal motor neurons among ADSCs, BMSCs and MG groups. These results suggest that neither BMSCs nor ADSCs provide satisfactory results for peripheral nerve repair when using MG as the conductor for

  2. Human adipose tissue-derived mesenchymal stem cells differentiate into insulin, somatostatin, and glucagon expressing cells

    International Nuclear Information System (INIS)

    Timper, Katharina; Seboek, Dalma; Eberhardt, Michael; Linscheid, Philippe; Christ-Crain, Mirjam; Keller, Ulrich; Mueller, Beat; Zulewski, Henryk

    2006-01-01

    Mesenchymal stem cells (MSC) from mouse bone marrow were shown to adopt a pancreatic endocrine phenotype in vitro and to reverse diabetes in an animal model. MSC from human bone marrow and adipose tissue represent very similar cell populations with comparable phenotypes. Adipose tissue is abundant and easily accessible and could thus also harbor cells with the potential to differentiate in insulin producing cells. We isolated human adipose tissue-derived MSC from four healthy donors. During the proliferation period, the cells expressed the stem cell markers nestin, ABCG2, SCF, Thy-1 as well as the pancreatic endocrine transcription factor Isl-1. The cells were induced to differentiate into a pancreatic endocrine phenotype by defined culture conditions within 3 days. Using quantitative PCR a down-regulation of ABCG2 and up-regulation of pancreatic developmental transcription factors Isl-1, Ipf-1, and Ngn3 were observed together with induction of the islet hormones insulin, glucagon, and somatostatin

  3. 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

  4. 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.

  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. A 7 YEAR-7-MONTH OLD BOY WITH LEUKEMIC RETINOPATHY

    Directory of Open Access Journals (Sweden)

    Ni Made Rini Suari

    2013-10-01

    Full Text Available Normal 0 false false false IN X-NONE X-NONE MicrosoftInternetExplorer4 Ocular problems in patient with leukemia which are called leukemic retinopathy and subhyaloid hemorrhage is one of its feature. Subhyaloid hemorrhage in children with acute lymphoblastic leukemia (ALL is rarely happened. We reported a boy 7 year 7 month old, complained sudden blurred vision on his both eyes and diagnosed with acute lymphoblastic leukemia. When patient had complained his vision, result of routine hematology showed anemia, thrombocytopenia, and leukocytosis. Treatment of leukemic retinopathy in this patient was supportive and causal therapy with transfusion of thrombocyte concentrate, hydration for leukocytosis, giving chemotherapy intrathecal methotrexate and systemic (vincristine, daunorubicin, L-asparginase. We found gradually undergone resolution of subhyaloid hemorrhages, visible flame shaped thin, and his vision recovered nearly completely to 6/6 OD and 6/20 OS /* Style Definitions */ table.MsoNormalTable {mso-style-name:"Table Normal"; mso-tstyle-rowband-size:0; mso-tstyle-colband-size:0; mso-style-noshow:yes; mso-style-priority:99; mso-style-qformat:yes; mso-style-parent:""; mso-padding-alt:0cm 5.4pt 0cm 5.4pt; mso-para-margin:0cm; mso-para-margin-bottom:.0001pt; mso-pagination:widow-orphan; font-size:11.0pt; font-family:"Calibri","sans-serif"; mso-ascii-font-family:Calibri; mso-ascii-theme-font:minor-latin; mso-fareast-font-family:"Times New Roman"; mso-fareast-theme-font:minor-fareast; mso-hansi-font-family:Calibri; mso-hansi-theme-font:minor-latin; mso-bidi-font-family:"Times New Roman"; mso-bidi-theme-font:minor-bidi;}

  7. Bone marrow-derived stromal cells are more beneficial cell sources for tooth regeneration compared with adipose-derived stromal cells.

    Science.gov (United States)

    Ye, Lanfeng; Chen, Lin; Feng, Fan; Cui, Junhui; Li, Kaide; Li, Zhiyong; Liu, Lei

    2015-10-01

    Tooth loss is presently a global epidemic and tooth regeneration is thought to be a feasible and ideal treatment approach. Choice of cell source is a primary concern in tooth regeneration. In this study, the odontogenic differentiation potential of two non-dental-derived stem cells, adipose-derived stromal cells (ADSCs) and bone marrow-derived stromal cells (BMSCs), were evaluated both in vitro and in vivo. ADSCs and BMSCs were induced in vitro in the presence of tooth germ cell-conditioned medium (TGC-CM) prior to implantation into the omentum majus of rats, in combination with inactivated dentin matrix (IDM). Real-time quantitative polymerase chain reaction (RT-qPCR) was used to detect the mRNA expression levels of odontogenic-related genes. Immunofluorescence and immunohistochemical assays were used to detect the protein levels of odontogenic-specific genes, such as DSP and DMP-1 both in vitro and in vivo. The results suggest that both ADSCs and BMSCs have odontogenic differentiation potential. However, the odontogenic potential of BMSCs was greater compared with ADSCs, showing that BMSCs are a more appropriate cell source for tooth regeneration. © 2015 International Federation for Cell Biology.

  8. Cell-based lipid flippase assay employing fluorescent lipid derivatives

    DEFF Research Database (Denmark)

    Jensen, Maria Stumph; Costa, Sara; Günther-Pomorski, Thomas

    2016-01-01

    P-type ATPases in the P4 subfamily (P4-ATPases) are transmembrane proteins unique for eukaryotes that act as lipid flippases, i.e., to translocate phospholipids from the exofacial to the cytofacial monolayer of cellular membranes. While initially characterized as aminophospholipid translocases, s...... flippase activities in the plasma membrane of cells, using yeast as an example.......P-type ATPases in the P4 subfamily (P4-ATPases) are transmembrane proteins unique for eukaryotes that act as lipid flippases, i.e., to translocate phospholipids from the exofacial to the cytofacial monolayer of cellular membranes. While initially characterized as aminophospholipid translocases......, studies of individual P4-ATPase family members from fungi, plants, and animals show that P4-ATPases differ in their substrate specificities and mediate transport of a broader range of lipid substrates. Here, we describe an assay based on fluorescent lipid derivatives to monitor and characterize lipid...

  9. Enrichment of skin-derived neural precursor cells from dermal cell populations by altering culture conditions.

    Science.gov (United States)

    Bayati, Vahid; Gazor, Rohoullah; Nejatbakhsh, Reza; Negad Dehbashi, Fereshteh

    2016-01-01

    As stem cells play a critical role in tissue repair, their manipulation for being applied in regenerative medicine is of great importance. Skin-derived precursors (SKPs) may be good candidates for use in cell-based therapy as the only neural stem cells which can be isolated from an accessible tissue, skin. Herein, we presented a simple protocol to enrich neural SKPs by monolayer adherent cultivation to prove the efficacy of this method. To enrich neural SKPs from dermal cell populations, we have found that a monolayer adherent cultivation helps to increase the numbers of neural precursor cells. Indeed, we have cultured dermal cells as monolayer under serum-supplemented (control) and serum-supplemented culture, followed by serum free cultivation (test) and compared. Finally, protein markers of SKPs were assessed and compared in both experimental groups and differentiation potential was evaluated in enriched culture. The cells of enriched culture concurrently expressed fibronectin, vimentin and nestin, an intermediate filament protein expressed in neural and skeletal muscle precursors as compared to control culture. In addition, they possessed a multipotential capacity to differentiate into neurogenic, glial, adipogenic, osteogenic and skeletal myogenic cell lineages. It was concluded that serum-free adherent culture reinforced by growth factors have been shown to be effective on proliferation of skin-derived neural precursor cells (skin-NPCs) and drive their selective and rapid expansion.

  10. 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

  11. 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.

  12. 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.

  13. 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

  14. 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.

  15. Human gingiva-derived mesenchymal stem cells are superior to bone marrow-derived mesenchymal stem cells for cell therapy in regenerative medicine

    Energy Technology Data Exchange (ETDEWEB)

    Tomar, Geetanjali B.; Srivastava, Rupesh K.; Gupta, Navita; Barhanpurkar, Amruta P.; Pote, Satish T. [National Center for Cell Science, University of Pune Campus, Pune 411 007 (India); Jhaveri, Hiral M. [Department of Periodontics and Oral Implantology, Dr. D.Y. Patil Dental College and Hospital, Pune (India); Mishra, Gyan C. [National Center for Cell Science, University of Pune Campus, Pune 411 007 (India); Wani, Mohan R., E-mail: mohanwani@nccs.res.in [National Center for Cell Science, University of Pune Campus, Pune 411 007 (India)

    2010-03-12

    Mesenchymal stem cells (MSCs) are capable of self-renewal and differentiation into multiple cell lineages. Presently, bone marrow is considered as a prime source of MSCs; however, there are some drawbacks and limitations in use of these MSCs for cell therapy. In this study, we demonstrate that human gingival tissue-derived MSCs have several advantages over bone marrow-derived MSCs. Gingival MSCs are easy to isolate, homogenous and proliferate faster than bone marrow MSCs without any growth factor. Importantly, gingival MSCs display stable morphology and do not loose MSC characteristic at higher passages. In addition, gingival MSCs maintain normal karyotype and telomerase activity in long-term cultures, and are not tumorigenic. Thus, we reveal that human gingiva is a better source of MSCs than bone marrow, and large number of functionally competent clinical grade MSCs can be generated in short duration for cell therapy in regenerative medicine and tissue engineering.

  16. Human gingiva-derived mesenchymal stem cells are superior to bone marrow-derived mesenchymal stem cells for cell therapy in regenerative medicine

    International Nuclear Information System (INIS)

    Tomar, Geetanjali B.; Srivastava, Rupesh K.; Gupta, Navita; Barhanpurkar, Amruta P.; Pote, Satish T.; Jhaveri, Hiral M.; Mishra, Gyan C.; Wani, Mohan R.

    2010-01-01

    Mesenchymal stem cells (MSCs) are capable of self-renewal and differentiation into multiple cell lineages. Presently, bone marrow is considered as a prime source of MSCs; however, there are some drawbacks and limitations in use of these MSCs for cell therapy. In this study, we demonstrate that human gingival tissue-derived MSCs have several advantages over bone marrow-derived MSCs. Gingival MSCs are easy to isolate, homogenous and proliferate faster than bone marrow MSCs without any growth factor. Importantly, gingival MSCs display stable morphology and do not loose MSC characteristic at higher passages. In addition, gingival MSCs maintain normal karyotype and telomerase activity in long-term cultures, and are not tumorigenic. Thus, we reveal that human gingiva is a better source of MSCs than bone marrow, and large number of functionally competent clinical grade MSCs can be generated in short duration for cell therapy in regenerative medicine and tissue engineering.

  17. The differentiation potential of adipose tissue-derived mesenchymal stem cells into cell lineage related to male germ cells

    Directory of Open Access Journals (Sweden)

    P. Bräunig

    Full Text Available ABSTRACT The adipose tissue is a reliable source of Mesenchymal stem cells (MSCs showing a higher plasticity and transdifferentiation potential into multilineage cells. In the present study, adipose tissue-derived mesenchymal stem cells (AT-MSCs were isolated from mice omentum and epididymis fat depots. The AT-MSCs were initially compared based on stem cell surface markers and on the mesodermal trilineage differentiation potential. Additionally, AT-MSCs, from both sources, were cultured with differentiation media containing retinoic acid (RA and/or testicular cell-conditioned medium (TCC. The AT-MSCs expressed mesenchymal surface markers and differentiated into adipogenic, chondrogenic and osteogenic lineages. Only omentum-derived AT-MSCs expressed one important gene marker related to male germ cell lineages, after the differentiation treatment with RA. These findings reaffirm the importance of adipose tissue as a source of multipotent stromal-stem cells, as well as, MSCs source regarding differentiation purpose.

  18. Distinct gut-derived lactic acid bacteria elicit divergent dendritic cell-mediated NK cell responses

    DEFF Research Database (Denmark)

    Fink, Lisbeth Nielsen; Zeuthen, Louise Hjerrild; Christensen, Hanne

    2007-01-01

    Lactic acid bacteria (LAB) are abundant in the gastrointestinal tract where they continuously regulate the immune system. NK cells are potently activated by dendritic cells (DCs) matured by inflammatory stimuli, and NK cells are present in the gut epithelium and in mesenteric lymph nodes......, but it is not known how NK-DC interactions are affected by the predominantly non-pathogenic LAB. We demonstrate that human DCs exposed to different strains of gut-derived LAB consistently induce proliferation, cytotoxicity and activation markers in autologous NK cells. On the contrary, strains of LAB differ greatly...... in their ability to induce DC-dependent IFN-gamma production by NK cells. This suggests that DCs stimulated by gut LAB may expand the pool of NK cells and increase their cytotoxic potential. Specific LAB, inducing high levels of IL-12 in DCs, may promote amplification of a type-1 response via potent stimulation...

  19. 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.)

  20. Norcantharidin, Derivative of Cantharidin, for Cancer Stem Cells

    Directory of Open Access Journals (Sweden)

    Chen-Hsi Hsieh

    2013-01-01

    Full Text Available Cancer stem cells (CSCs existing in human cancers have been demonstrated to be a major cause of cancer treatment resistance, invasion, metastasis, and relapse. Self-renewal pathways, Wnt/β-catenin, Sonic hedgehog (Shh, and the Notch signaling pathway play critical roles in developing CSCs and lead to angiogenesis, migration, invasion, and metastasis. Multidrug resistance (MDR is an unfavorable factor causing the failure of treatments against cancer cells. The most important and thoroughly studied mechanism involved in MDR is the active efflux of chemotherapeutic agents through membrane drug transporters. There is growing evidence that Norcantharidin (NCTD, a water-soluble synthetic small molecule derivative of naturally occurring cantharidin from the medicinal insect blister beetle (Mylabris phalerata Pallas, is capable of chemoprevention and tumor inhibition. We summarize investigations into the modulation of self-renewal pathways and MDR in CSCs by NCTD. This review may aid in further investigation of using NCTD to develop more effective strategies for cancer treatment to reduce resistance and recurrence.

  1. Generation of insulin-producing cells from gnotobiotic porcine skin-derived stem cells

    Energy Technology Data Exchange (ETDEWEB)

    Yang, Ji Hoon; Lee, Sung Ho; Heo, Young Tae [Department of Bioscience and Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Uhm, Sang Jun [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of); Lee, Hoon Taek, E-mail: htl3675@konkuk.ac.kr [Department of Animal Biotechnology, Bio-Organ Research Center, Konkuk University, Seoul 143-701 (Korea, Republic of)

    2010-07-09

    A major problem in the treatment of type 1 diabetes mellitus is the limited availability of alternative sources of insulin-producing cells for islet transplantation. In this study, we investigated the effect of bone morphogenetic protein 4 (BMP-4) treatments of gnotobiotic porcine skin-derived stem cells (gSDSCs) on their reprogramming and subsequent differentiation into insulin-producing cells (IPCs). We isolated SDSCs from the ear skin of a gnotobiotic pig. During the proliferation period, the cells expressed stem-cell markers Oct-4, Sox-2, and CD90; nestin expression also increased significantly. The cells could differentiate into IPCs after treatments with activin-A, glucagon-like peptide-1 (GLP-1), and nicotinamide. After 15 days in the differentiation medium, controlled gSDSCs began expressing endocrine progenitor genes and proteins (Ngn3, Neuro-D, PDX-1, NKX2.2, NKX6.1, and insulin). The IPCs showed increased insulin synthesis after glucose stimulation. The results indicate that stem cells derived from the skin of gnotobiotic pigs can differentiate into IPCs under the appropriate conditions in vitro. Our three-stage induction protocol could be applied without genetic modification to source IPCs from stem cells in the skin of patients with diabetes for autologous transplantation.

  2. Generation of insulin-producing cells from gnotobiotic porcine skin-derived stem cells

    International Nuclear Information System (INIS)

    Yang, Ji Hoon; Lee, Sung Ho; Heo, Young Tae; Uhm, Sang Jun; Lee, Hoon Taek

    2010-01-01

    A major problem in the treatment of type 1 diabetes mellitus is the limited availability of alternative sources of insulin-producing cells for islet transplantation. In this study, we investigated the effect of bone morphogenetic protein 4 (BMP-4) treatments of gnotobiotic porcine skin-derived stem cells (gSDSCs) on their reprogramming and subsequent differentiation into insulin-producing cells (IPCs). We isolated SDSCs from the ear skin of a gnotobiotic pig. During the proliferation period, the cells expressed stem-cell markers Oct-4, Sox-2, and CD90; nestin expression also increased significantly. The cells could differentiate into IPCs after treatments with activin-A, glucagon-like peptide-1 (GLP-1), and nicotinamide. After 15 days in the differentiation medium, controlled gSDSCs began expressing endocrine progenitor genes and proteins (Ngn3, Neuro-D, PDX-1, NKX2.2, NKX6.1, and insulin). The IPCs showed increased insulin synthesis after glucose stimulation. The results indicate that stem cells derived from the skin of gnotobiotic pigs can differentiate into IPCs under the appropriate conditions in vitro. Our three-stage induction protocol could be applied without genetic modification to source IPCs from stem cells in the skin of patients with diabetes for autologous transplantation.

  3. 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

  4. Effects of everolimus on macrophage-derived foam cell behavior

    International Nuclear Information System (INIS)

    Hsu, Steven; Koren, Eugen; Chan, Yen; Koscec, Mirna; Sheehy, Alexander; Kolodgie, Frank; Virmani, Renu; Feder, Debra

    2014-01-01

    Purpose: The purpose of this study was to investigate the effects of everolimus on foam cell (FC) viability, mRNA levels, and inflammatory cytokine production to better understand its potential inhibitory effects on atheroma progression. Methods and materials: Human THP1 macrophage-derived FC were formed using acetylated LDL (acLDL, 100 μg/mL) for 72 hours, followed by everolimus treatment (10 -5 –10 -11 M) for 24 hours. FC viability was quantified using fluorescent calcein AM/DAPI staining. FC lysates and media supernatants were analyzed for apoptosis and necrosis using a Cell Death ELISA PLUS assay. FC lysates and media supernatants were also analyzed for inflammatory cytokine (IL1β, IL8, MCP1, TNFα) mRNA levels and protein expression using quantitative reverse transcription real-time polymerase chain reaction (QPCR) and a Procarta® immunoassay, respectively. mRNA levels of autophagy (MAP1LC3), apoptosis (survivin, clusterin), and matrix degradation (MMP1, MMP9) markers were evaluated by Quantigene® Plex assay and verified with QPCR. Additionally, hypercholesterolemic rabbits received everolimus-eluting stents (EES) for 28 or 60 days. RAM-11 immunohistochemical staining was performed to compare %RAM-11 positive area between stented sections and unstented proximal sections. Statistical significance was calculated using one-way ANOVA (p ≤ 0.05). Results: Calcein AM/DAPI staining showed that FC exposed to everolimus (10 -5 M) had significantly decreased viability compared to control. FC apoptosis was significantly increased at a high dose of everolimus (10 -5 M), with no necrotic effects at any dose tested. Everolimus did not affect endothelial (HUVEC) and smooth muscle (HCASMC) cell apoptosis or necrosis. Everolimus (10 -5 M) significantly increased MAP1LC3, caused an increased trend in clusterin (p = 0.10), and significantly decreased survivin and MMP1 mRNA levels in FC. MCP1 cytokine mRNA levels and secreted protein expression was significantly decreased

  5. Effects of everolimus on macrophage-derived foam cell behavior

    Energy Technology Data Exchange (ETDEWEB)

    Hsu, Steven, E-mail: steven.hsu@av.abbott.com [Abbott Vascular, 3200 Lakeside Drive, Santa Clara, CA 95054 (United States); Koren, Eugen; Chan, Yen; Koscec, Mirna; Sheehy, Alexander [Abbott Vascular, 3200 Lakeside Drive, Santa Clara, CA 95054 (United States); Kolodgie, Frank; Virmani, Renu [CVPath Institute, Inc., 19 Firstfield Road, Gaithersburg, MD 20878 (United States); Feder, Debra [Abbott Vascular, 3200 Lakeside Drive, Santa Clara, CA 95054 (United States)

    2014-07-15

    Purpose: The purpose of this study was to investigate the effects of everolimus on foam cell (FC) viability, mRNA levels, and inflammatory cytokine production to better understand its potential inhibitory effects on atheroma progression. Methods and materials: Human THP1 macrophage-derived FC were formed using acetylated LDL (acLDL, 100 μg/mL) for 72 hours, followed by everolimus treatment (10{sup -5}–10{sup -11} M) for 24 hours. FC viability was quantified using fluorescent calcein AM/DAPI staining. FC lysates and media supernatants were analyzed for apoptosis and necrosis using a Cell Death ELISA{sup PLUS} assay. FC lysates and media supernatants were also analyzed for inflammatory cytokine (IL1β, IL8, MCP1, TNFα) mRNA levels and protein expression using quantitative reverse transcription real-time polymerase chain reaction (QPCR) and a Procarta® immunoassay, respectively. mRNA levels of autophagy (MAP1LC3), apoptosis (survivin, clusterin), and matrix degradation (MMP1, MMP9) markers were evaluated by Quantigene® Plex assay and verified with QPCR. Additionally, hypercholesterolemic rabbits received everolimus-eluting stents (EES) for 28 or 60 days. RAM-11 immunohistochemical staining was performed to compare %RAM-11 positive area between stented sections and unstented proximal sections. Statistical significance was calculated using one-way ANOVA (p ≤ 0.05). Results: Calcein AM/DAPI staining showed that FC exposed to everolimus (10{sup -5} M) had significantly decreased viability compared to control. FC apoptosis was significantly increased at a high dose of everolimus (10{sup -5} M), with no necrotic effects at any dose tested. Everolimus did not affect endothelial (HUVEC) and smooth muscle (HCASMC) cell apoptosis or necrosis. Everolimus (10{sup -5} M) significantly increased MAP1LC3, caused an increased trend in clusterin (p = 0.10), and significantly decreased survivin and MMP1 mRNA levels in FC. MCP1 cytokine mRNA levels and secreted protein

  6. Induction of Skin-Derived Precursor Cells from Human Induced Pluripotent Stem Cells.

    Science.gov (United States)

    Sugiyama-Nakagiri, Yoriko; Fujimura, Tsutomu; Moriwaki, Shigeru

    2016-01-01

    The generation of full thickness human skin from dissociated cells is an attractive approach not only for treating skin diseases, but also for treating many systemic disorders. However, it is currently not possible to obtain an unlimited number of skin dermal cells. The goal of this study was to develop a procedure to produce skin dermal stem cells from induced pluripotent stem cells (iPSCs). Skin-derived precursor cells (SKPs) were isolated as adult dermal precursors that could differentiate into both neural and mesodermal progenies and could reconstitute the dermis. Thus, we attempted to generate SKPs from iPSCs that could reconstitute the skin dermis. Human iPSCs were initially cultured with recombinant noggin and SB431542, an inhibitor of activin/nodal and TGFβ signaling, to induce neural crest progenitor cells. Those cells were then treated with SKP medium that included CHIR99021, a WNT signal activator. The induction efficacy from neural crest progenitor cells to SKPs was more than 97%. No other modifiers tested were able to induce those cells. Those human iPSC-derived SKPs (hiPSC-SKPs) showed a similar gene expression signature to SKPs isolated from human skin dermis. Human iPSC-SKPs differentiated into neural and mesodermal progenies, including adipocytes, skeletogenic cell types and Schwann cells. Moreover, they could be induced to follicular type keratinization when co-cultured with human epidermal keratinocytes. We here provide a new efficient protocol to create human skin dermal stem cells from hiPSCs that could contribute to the treatment of various skin disorders.

  7. 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.

  8. Transport of curcumin derivatives in Caco-2 cell monolayers.

    Science.gov (United States)

    Zeng, Zhen; Shen, Zhe L; Zhai, Shuo; Xu, Jia L; Liang, Hui; Shen, Qin; Li, Qing Y

    2017-08-01

    Curcumin (Cur) is a strong natural antioxidant, who can prevent multiple diseases such as anti-cancer, anti-inflammatory, have a resistance to alzheimer's disease and various malignant diseases. But it has poor oral bioavailability due to its poor aqueous solubility, as well as instability. While its novel derivatives (CB and FE), showed better anti-tumor activity, better anti-oxidant activity and better stability than the original drug (Cur). The aim of this study was to study the intestinal transport of Cur, CB and FE using an in vitro Caco-2 cell monolayer model. The results showed that Cur had a lower permeability coefficient (1.13×10 -6 ±0.11×10 -6 cm/s) for apical-to-basolated (AP-BL) transport at 25μM, while the transport rate for AP to BL flux of CB (3.18×10 -6 ±0.31×10 -6 cm/s) and FE (5.28×10 -6 ±0.83×10 -6 cm/s) were significantly greater than that of Cur. The efflux ratio (ER) value at the concentration of 25μM was 1.31 for Cur, 1.26 for CB and 1.33 for FE, suggesting there was no active efflux involved in the translocation across the Caco-2 cell monolayers for the three compounds. Furthermore, the transport flux of CB and FE was in a concentration dependent manner, suggesting the intestinal transport mechanism in them was passive transport. In summary, the results demonstrated that both the intestinal permeability of CB and FE across Caco-2 cell monolayers was significantly improved compare to Cur. Thus they might show a higher oral bioavailability in vivo, and show the potential application in clinic or nutraceutical. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Donor lung derived myeloid and plasmacytoid dendritic cells differentially regulate T cell proliferation and cytokine production

    Directory of Open Access Journals (Sweden)

    Benson Heather L

    2012-03-01

    Full Text Available Abstract Background Direct allorecognition, i.e., donor lung-derived dendritic cells (DCs stimulating recipient-derived T lymphocytes, is believed to be the key mechanism of lung allograft rejection. Myeloid (cDCs and plasmacytoid (pDCs are believed to have differential effects on T cell activation. However, the roles of each DC type on T cell activation and rejection pathology post lung transplantation are unknown. Methods Using transgenic mice and antibody depletion techniques, either or both cell types were depleted in lungs of donor BALB/c mice (H-2d prior to transplanting into C57BL/6 mice (H-2b, followed by an assessment of rejection pathology, and pDC or cDC-induced proliferation and cytokine production in C57BL/6-derived mediastinal lymph node T cells (CD3+. Results Depleting either DC type had modest effect on rejection pathology and T cell proliferation. In contrast, T cells from mice that received grafts depleted of both DCs did not proliferate and this was associated with significantly reduced acute rejection scores compared to all other groups. cDCs were potent inducers of IFNγ, whereas both cDCs and pDCs induced IL-10. Both cell types had variable effects on IL-17A production. Conclusion Collectively, the data show that direct allorecognition by donor lung pDCs and cDCs have differential effects on T cell proliferation and cytokine production. Depletion of both donor lung cDC and pDC could prevent the severity of acute rejection episodes.

  10. Epigenetics targeted protein-vorinostat nanomedicine inducing apoptosis in heterogeneous population of primary acute myeloid leukemia cells including refractory and relapsed cases.

    Science.gov (United States)

    Chandran, Parwathy; Kavalakatt, Anu; Malarvizhi, Giridharan Loghanathan; Vasanthakumari, Divya Rani Vikraman Nair; Retnakumari, Archana Payickattu; Sidharthan, Neeraj; Pavithran, Keechilat; Nair, Shantikumar; Koyakutty, Manzoor

    2014-05-01

    Aberrant epigenetics play a key role in the onset and progression of acute myeloid leukemia (AML). Herein we report in silico modelling based development of a novel, protein-vorinostat nanomedicine exhibiting selective and superior anti-leukemic activity against heterogeneous population of AML patient samples (n=9), including refractory and relapsed cases, and three representative cell lines expressing CD34(+)/CD38(-) stem cell phenotype (KG-1a), promyelocytic phenotype (HL-60) and FLT3-ITD mutation (MV4-11). Nano-vorinostat having ~100nm size exhibited enhanced cellular uptake rendering significantly lower IC50 in AML cell lines and patient samples, and induced enhanced HDAC inhibition, oxidative injury, cell cycle arrest and apoptosis compared to free vorinostat. Most importantly, nanomedicine showed exceptional single-agent activity against the clonogenic proliferative capability of bone marrow derived leukemic progenitors, while remaining non-toxic to healthy bone marrow cells. Collectively, this epigenetics targeted nanomedicine appears to be a promising therapeutic strategy against various French-American-British (FAB) classes of AML. Through the use of a protein-vorinostat agent, exceptional single-agent activity was demonstrated against the clonogenic proliferative capability of bone marrow derived leukemic progenitors, while remaining non-toxic to healthy bone marrow cells. The studied epigenetics targeted nanomedicine approach is a promising therapeutic strategy against various French-American-British classes of acute myeloid leukemia. © 2014 Elsevier Inc. All rights reserved.

  11. Rabbit embryonic stem cell lines derived from fertilized, parthenogenetic or somatic cell nuclear transfer embryos

    International Nuclear Information System (INIS)

    Fang, Zhen F.; Gai, Hui; Huang, You Z.; Li, Shan G.; Chen, Xue J.; Shi, Jian J.; Wu, Li; Liu, Ailian; Xu, Ping; Sheng, Hui Z.

    2006-01-01

    Embryonic stem cells were isolated from rabbit blastocysts derived from fertilization (conventional rbES cells), parthenogenesis (pES cells) and nuclear transfer (ntES cells), and propagated in a serum-free culture system. Rabbit ES (rbES) cells proliferated for a prolonged time in an undifferentiated state and maintained a normal karyotype. These cells grew in a monolayer with a high nuclear/cytoplasm ratio and contained a high level of alkaline phosphate activity. In addition, rbES cells expressed the pluripotent marker Oct-4, as well as EBAF2, FGF4, TDGF1, but not antigens recognized by antibodies against SSEA-1, SSEA-3, SSEA-4, TRA-1-10 and TRA-1-81. All 3 types of ES cells formed embryoid bodies and generated teratoma that contained tissue types of all three germ layers. rbES cells exhibited a high cloning efficiency, were genetically modified readily and were used as nuclear donors to generate a viable rabbit through somatic cell nuclear transfer. In combination with genetic engineering, the ES cell technology should facilitate the creation of new rabbit lines

  12. A modified method of insulin producing cells' generation from bone marrow-derived mesenchymal stem cells.

    Science.gov (United States)

    Czubak, Paweł; Bojarska-Junak, Agnieszka; Tabarkiewicz, Jacek; Putowski, Lechosław

    2014-01-01

    Type 1 diabetes mellitus is a result of autoimmune destruction of pancreatic insulin producing β-cells and so far it can be cured only by insulin injection, by pancreas transplantation, or by pancreatic islet cells' transplantation. The methods are, however, imperfect and have a lot of disadvantages. Therefore new solutions are needed. The best one would be the use of differentiated mesenchymal stem cells (MSCs). In the present study, we investigated the potential of the bone marrow-derived MSCs line for in vitro differentiation into insulin producing cells (IPSs). We applied an 18-day protocol to differentiate MSCs. Differentiating cells formed cell clusters some of which resembled pancreatic islet-like cells. Using dithizone we confirmed the presence of insulin in the cells. What is more, the expression of proinsulin C-peptide in differentiated IPCs was analyzed by flow cytometry. For the first time, we investigated the influence of growth factors' concentration on IPCs differentiation efficiency. We have found that an increase in the concentration of growth factors up to 60 ng/mL of β-FGF/EGF and 30 ng/mL of activin A/β-cellulin increases the percentage of IPCs. Further increase of growth factors does not show any increase of the percentage of differentiated cells. Our findings suggest that the presented protocol can be adapted for differentiation of insulin producing cells from stem cells.

  13. 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

  14. Derivation of keratinocytes from chicken embryonic stem cells: Establishment and characterization of differentiated proliferative cell populations

    Directory of Open Access Journals (Sweden)

    Mathilde Couteaudier

    2015-03-01

    Full Text Available A common challenge in avian cell biology is the generation of differentiated cell-lines, especially in the keratinocyte lineage. Only a few avian cell-lines are available and very few of them show an interesting differentiation profile. During the last decade, mammalian embryonic stem cell-lines were shown to differentiate into almost all lineages, including keratinocytes. Although chicken embryonic stem cells had been obtained in the 1990s, few differentiation studies toward the ectodermal lineage were reported. Consequently, we explored the differentiation of chicken embryonic stem cells toward the keratinocyte lineage by using a combination of stromal induction, ascorbic acid, BMP4 and chicken serum. During the induction period, we observed a downregulation of pluripotency markers and an upregulation of epidermal markers. Three homogenous cell populations were derived, which were morphologically similar to chicken primary keratinocytes, displaying intracellular lipid droplets in almost every pavimentous cell. These cells could be serially passaged without alteration of their morphology and showed gene and protein expression profiles of epidermal markers similar to chicken primary keratinocytes. These cells represent an alternative to the isolation of chicken primary keratinocytes, being less cumbersome to handle and reducing the number of experimental animals used for the preparation of primary cells.

  15. Procedures for Derivation and Characterisation of Human Embryonic Stem Cells from Odense, Denmark

    DEFF Research Database (Denmark)

    Harkness, Linda; Kassem, Moustapha

    2012-01-01

    In 1998, a development occurred in stem cell biology with the fi rst report of the derivation of a human embryonic stem cell (hESC) line. Since then a number of techniques have been used to derive and characterise hESCs. Here, we describe the derivation methods used by our laboratory for isolatio...

  16. 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.

  17. Clinical impact of leukemic blast heterogeneity at diagnosis in cytogenetic intermediate-risk acute myeloid leukemia

    DEFF Research Database (Denmark)

    Hoffmann, Marianne Hutchings; Klausen, Tobias Wirenfeldt; Boegsted, Martin

    2012-01-01

    Individual cellular heterogeneity within the acute myeloid leukemia (AML) bone marrow samples can be observed by multi parametric flow cytometry analysis (MFC) indicating that immunophenotypic screening for leukemic blast subsets may have prognostic impact.......Individual cellular heterogeneity within the acute myeloid leukemia (AML) bone marrow samples can be observed by multi parametric flow cytometry analysis (MFC) indicating that immunophenotypic screening for leukemic blast subsets may have prognostic impact....

  18. Cell surface glycopeptides from human intestinal epithelial cell lines derived from normal colon and colon adenocarcinomas

    International Nuclear Information System (INIS)

    Youakim, A.; Herscovics, A.

    1985-01-01

    The cell surface glycopeptides from an epithelial cell line (CCL 239) derived from normal human colon were compared with those from three cell lines (HCT-8R, HCT-15, and CaCo-2) derived independently from human colonic adenocarcinomas. Cells were incubated with D-[2- 3 H]mannose or L-[5,6- 3 H]fucose for 24 h and treated with trypsin to release cell surface components which were then digested exhaustively with Pronase and fractionated on Bio-Gel P-6 before and after treatment with endo-beta-N-acetylglucosaminidase H. The most noticeable difference between the labeled glycopeptides from the tumor and CCL 239 cells was the presence in the former of an endo-beta-N-acetylglucosaminidase H-resistant high molecular weight glycopeptide fraction which was eluted in the void volume of Bio-Gel P-6. This fraction was obtained with both labeled mannose and fucose as precursors. However, acid hydrolysis of this fraction obtained after incubation with [2- 3 H]mannose revealed that as much as 60-90% of the radioactivity was recovered as fucose. Analysis of the total glycopeptides (cell surface and cell pellet) obtained after incubation with [2- 3 H]mannose showed that from 40-45% of the radioactivity in the tumor cells and less than 10% of the radioactivity in the CCL 239 cells was recovered as fucose. After incubation of the HCT-8R cells with D-[1,6- 3 H]glucosamine and L-[1- 14 C]fucose, strong acid hydrolysis of the labeled glycopeptide fraction excluded from Bio-Gel P-6 produced 3 H-labeled N-acetylglucosamine and N-acetylgalactosamine

  19. Indirubin inhibits cell proliferation, migration, invasion and angiogenesis in tumor-derived endothelial cells

    Directory of Open Access Journals (Sweden)

    Li Z

    2018-05-01

    Full Text Available Zhuohong Li, Chaofu Zhu, Baiping An, Yu Chen, Xiuyun He, Lin Qian, Lan Lan, Shijie Li Department of Oncology, The Affiliated Hospital of Chengdu University of Traditional Chinese Medicine, Chengdu, Sichuan, China Purpose: Hepatocellular carcinoma is one of the most predominant malignancies with high fatality rate and its incidence is rising at an alarming rate because of its resistance to radio- and chemotherapy. Indirubin is the major active anti-tumor ingredient of a traditional Chinese herbal medicine. The present study aimed to analyze the effects of indirubin on cell proliferation, migration, invasion, and angiogenesis of tumor-derived endothelial cells (Td-EC. Methods: Td-EC were derived from human umbilical vein endothelial cells (HUVEC by treating HUVEC with the conditioned medium of human liver cancer cell line HepG2. Cell proliferation, migration, invasion, and angiogenesis were assessed by MTT, wound healing, in vitro cell invasion, and in vitro tube formation assay. Results: Td-EC were successfully obtained from HUVEC cultured with 50% culture supernatant from serum-starved HepG2 cells. Indirubin significantly inhibited Td-EC proliferation in a dose- and time-dependent manner. Indirubin also inhibited Td-EC migration, invasion, and angiogenesis. However, indirubin’s effects were weaker on HUVEC than Td-EC. Conclusion: Indirubin significantly inhibited Td-EC proliferation, migration, invasion, and angiogenesis. Keywords: indirubin, Td-EC, proliferation, migration, invasion, angiogenesis

  20. Role of bone marrow-derived stem cells, renal progenitor cells and stem cell factor in chronic renal allograft nephropathy

    OpenAIRE

    Hayam Abdel Meguid El Aggan; Mona Abdel Kader Salem; Nahla Mohamed Gamal Farahat; Ahmad Fathy El-Koraie; Ghaly Abd Al-Rahim Mohammed Kotb

    2013-01-01

    Introduction: Chronic allograft nephropathy (CAN) is a poorly understood clinico-pathological entity associated with chronic allograft loss due to immunologic and non-immunologic causes. It remains the leading cause of late allograft loss. Bone marrow derived stem cells are undifferentiated cells typically characterized by their capacity for self renewal, ability to give rise to multiple differentiated cellular population, including hematopoietic (HSCs) and mesenchymal stem cells (MSCs). Char...

  1. 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.

  2. 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.

  3. Human iPS Cell-Derived Germ Cells: Current Status and Clinical Potential

    Directory of Open Access Journals (Sweden)

    Tetsuya Ishii

    2014-10-01

    Full Text Available Recently, fertile spermatozoa and oocytes were generated from mouse induced pluripotent (iPS cells using a combined in vitro and in vivo induction system. With regard to germ cell induction from human iPS cells, progress has been made particularly in the male germline, demonstrating in vitro generation of haploid, round spermatids. Although iPS-derived germ cells are expected to be developed to yield a form of assisted reproductive technology (ART that can address unmet reproductive needs, genetic and/or epigenetic instabilities abound in iPS cell generation and germ cell induction. In addition, there is still room to improve the induction protocol in the female germline. However, rapid advances in stem cell research are likely to make such obstacles surmountable, potentially translating induced germ cells into the clinical setting in the immediate future. This review examines the current status of the induction of germ cells from human iPS cells and discusses the clinical potential, as well as future directions.

  4. Human Adipose Derived Stem Cells Induced Cell Apoptosis and S Phase Arrest in Bladder Tumor

    Directory of Open Access Journals (Sweden)

    Xi Yu

    2015-01-01

    Full Text Available The aim of this study was to determine the effect of human adipose derived stem cells (ADSCs on the viability and apoptosis of human bladder cancer cells. EJ and T24 cells were cocultured with ADSCs or cultured with conditioned medium of ADSCs (ADSC-CM, respectively. The cell counting and colony formation assay showed ADSCs inhibited the proliferation of EJ and T24 cells. Cell viability assessment revealed that the secretions of ADSCs, in the form of conditioned medium, were able to decrease cancer cell viability. Wound-healing assay suggested ADSC-CM suppressed migration of T24 and EJ cells. Moreover, the results of the flow cytometry indicated that ADSC-CM was capable of inducing apoptosis of T24 cells and inducing S phase cell cycle arrest. Western blot revealed ADSC-CM increased the expression of cleaved caspase-3 and cleaved PARP, indicating that ADSC-CM induced apoptosis in a caspase-dependent way. PTEN/PI3K/Akt pathway and Bcl-2 family proteins were involved in the mechanism of this reaction. Our study indicated that ADSCs may provide a promising and practicable manner for bladder tumor therapy.

  5. Adipose Tissue-Derived Mesenchymal Stem Cells as a New Host Cell in Latent Leishmaniasis

    Science.gov (United States)

    Allahverdiyev, Adil M.; Bagirova, Melahat; Elcicek, Serhat; Koc, Rabia Cakir; Baydar, Serap Yesilkir; Findikli, Necati; Oztel, Olga N.

    2011-01-01

    Some protozoan infections such as Toxoplasma, Cryptosporidium, and Plasmodium can be transmitted through stem cell transplantations. To our knowledge, so far, there is no study about transmission of Leishmania parasites in stem cell transplantation and interactions between parasites and stem cells in vitro. Therefore, the aim of this study was to investigate the interaction between different species of Leishmania parasites and adipose tissue-derived mesenchymal stem cells (ADMSCs). ADMSCs have been isolated, cultured, characterized, and infected with different species of Leishmania parasites (L. donovani, L. major, L. tropica, and L. infantum). Infectivity was examined by Giemsa staining, microculture, and polymerase chain reaction methods. As a result, infectivity of ADMSCs by Leishmania parasites has been determined for the first time in this study. According to our findings, it is very important that donors are screened for Leishmania parasites before stem cell transplantations in regions where leishmaniasis is endemic. PMID:21896818

  6. Derivation of Stromal (Skeletal, Mesenchymal) Stem-like cells from Human Embryonic Stem Cells

    DEFF Research Database (Denmark)

    Mahmood, Amer; Harkness, Linda; Abdallah, Basem

    2012-01-01

    EBs using BMP2 (bone morphogenic protein 2) combined with standard osteoblast induction medium led to weak osteoblastic induction. Conversely, subcutaneous in vivo implantation of day 20 hEBs in immune deficient mice, mixed with hydroxyapatite/tricalcium phosphate (HA/TCP) as an osteoconductive scaffold......Derivation of bone forming cells (osteoblasts) from human embryonic stem cells (hESC) is a pre-requisite for their use in clinical applications. However, there is no standard protocol for differentiating hESC into osteoblastic cells. The aim of this study was to identify the emergence of a human...... stromal (mesenchymal, skeletal) stem cell (hMSC)-like population, known to be osteoblastic cell precursors and to test their osteoblastic differentiation capacity in ex vivo cultures and in vivo. We cultured hESC in a feeder-free environment using serum replacement and as suspension aggregates (embryoid...

  7. Notch signaling is required for maintaining stem-cell features of neuroprogenitor cells derived from human embryonic stem cells

    Directory of Open Access Journals (Sweden)

    Chung Hyung-Min

    2009-08-01

    Full Text Available Abstract Background Studies have provided important findings about the roles of Notch signaling in neural development. Unfortunately, however, most of these studies have investigated the neural stem cells (NSCs of mice or other laboratory animals rather than humans, mainly owing to the difficulties associated with obtaining human brain samples. It prompted us to focus on neuroectodermal spheres (NESs which are derived from human embryonic stem cell (hESC and densely inhabited by NSCs. We here investigated the role of Notch signaling with the hESC-derived NESs. Results From hESCs, we derived NESs, the in-vitro version of brain-derived neurospheres. NES formation was confirmed by increased levels of various NSC marker genes and the emergence of rosette structures in which neuroprogenitors are known to reside. We found that Notch signaling, which maintains stem cell characteristics of in-vivo-derived neuroprogenitors, is active in these hESC-derived NESs, similar to their in-vivo counterpart. Expression levels of Notch signaling molecules such as NICD, DLLs, JAG1, HES1 and HES5 were increased in the NESs. Inhibition of the Notch signaling by a γ-secretase inhibitor reduced rosette structures, expression levels of NSC marker genes and proliferation potential in the NESs, and, if combined with withdrawal of growth factors, triggered differentiation toward neurons. Conclusion Our results indicate that the hESC-derived NESs, which share biochemical features with brain-derived neurospheres, maintain stem cell characteristics mainly through Notch signaling, which suggests that the hESC-derived NESs could be an in-vitro model for in-vivo neurogenesis.

  8. Blastema from rabbit ear contains progenitor cells comparable to marrow derived mesenchymal stem cells

    Directory of Open Access Journals (Sweden)

    Mohamadreza Baghaban Eslaminejad

    2012-09-01

    Full Text Available Rabbits have the capacity to regenerate holes in their ears by forming a blastema, a tissue that is made up of a group of undifferentiated cells. The purpose of the present study was to isolate and characterize blastema progenitor cells and compare them with marrow mesenchymal stem cells (MSCs. Five New Zealand white male rabbits were used in the present study. A 2-mm hole was created in the animal ears. After 4 days, the blastema ring formed in the periphery of the hole was removed and cultivated. The cells were expanded through several subcultures and compared with the MSCs derived from the marrow of same animal in terms of in vitro differentiation capacity, growth kinetics and culture requirements for optimal proliferation. The primary cultures from both cells tended to be heterogeneous. Fibroblastic cells became progressively dominant with advancing passages. Similar to MSCs blastema passaged-3 cells succeeded to differentiate into bone, cartilage and adipose cell lineages. Even lineage specific genes tended to express in higher level in blastema cells compared to MSCs (p < 0.05. Moreover blastema cells appeared more proliferative; producing more colonies (p < 0.05. While blastema cells showed extensive proliferation in 15% fetal bovine serum (FBS, MSCs displayed higher expansion rate at 10% FBS. In conclusion, blastema from rabbit ear contains a population of fibroblastic cells much similar in characteristic to bone marrow mesenchymal stem cells. However, the two cells were different in the level of lineage-specific gene expression, the growth curve characteristics and the culture requirements.

  9. Characterization and comparison of osteoblasts derived from mouse embryonic stem cells and induced pluripotent stem cells.

    Science.gov (United States)

    Ma, Ming-San; Kannan, Vishnu; de Vries, Anneriek E; Czepiel, Marcin; Wesseling, Evelyn M; Balasubramaniyan, Veerakumar; Kuijer, Roel; Vissink, Arjan; Copray, Sjef C V M; Raghoebar, Gerry M

    2017-01-01

    New developments in stem cell biology offer alternatives for the reconstruction of critical-sized bone defects. One of these developments is the use of induced pluripotent stem (iPS) cells. These stem cells are similar to embryonic stem (ES) cells, but can be generated from adult somatic cells and therefore do not raise ethical concerns. Proper characterization of iPS-derived osteoblasts is important for future development of safe clinical applications of these cells. For this reason, we differentiated mouse ES and iPS cells toward osteoblasts using osteogenic medium and compared their functionality. Immunocytochemical analysis showed significant expression of bone markers (osteocalcin and collagen type I) in osteoblasts differentiated from ES and iPS cells on days 7 and 30. An in vitro mineralization assay confirmed the functionality of osteogenically differentiated ES and iPS cells. Gene expression arrays focusing on osteogenic differentiation were performed in order to compare the gene expression pattern in both differentiated and undifferentiated ES cells and iPS cells. We observed a significant upregulation of osteogenesis-related genes such as Runx2, osteopontin, collagen type I, Tnfsf11, Csf1, and alkaline phosphatase upon osteogenic differentiation of the ES and iPS cells. We further validated the expression of key osteogenic genes Runx2, osteopontin, osteocalcin, collagen type I, and osterix in both differentiated and undifferentiated ES and iPS cells by means of quantified real-time polymerase chain reaction. We conclude that ES and iPS cells are similar in their osteogenic differentiation capacities, as well as in their gene expression patterns.

  10. Cell kinetics, DNA integrity, differentiation, and lipid fingerprinting analysis of rabbit adipose-derived stem cells.

    Science.gov (United States)

    Barretto, Letícia Siqueira de Sá; Lessio, Camila; Sawaki e Nakamura, Ahy Natally; Lo Turco, Edson Guimarães; da Silva, Camila Gonzaga; Zambon, João Paulo; Gozzo, Fábio César; Pilau, Eduardo Jorge; de Almeida, Fernando Gonçalves

    2014-10-01

    Human adipose tissue has been described as a potential alternative reservoir for stem cells. Although studies have been performed in rabbits using autologous adipose-derived stem cells (ADSC), these cells have not been well characterized. The primary objectives of this study were to demonstrate the presence of adipose-derived stem cells isolated from rabbit inguinal fat pads and to characterize them through osteogenic and adipogenic in vitro differentiation and lipid fingerprinting analysis. The secondary objective was to evaluate cell behavior through growth kinetics, cell viability, and DNA integrity. Rabbit ADSCs were isolated to determine the in vitro growth kinetics and cell viability. DNA integrity was assessed by an alkaline Comet assay in passages 0 and 5. The osteogenic differentiation was evaluated by Von Kossa, and Alizarin Red S staining and adipogenic differentiation were assessed by Oil Red O staining. Lipid fingerprinting analyses of control, adipogenic, and osteogenic differentiated cells were performed by MALDI-TOF/MS. We demonstrate that rabbit ADSC have a constant growth rate at the early passages, with increased DNA fragmentation at or after passage 5. Rabbit ADSC viability was similar in passages 2 and 5 (90.7% and 86.6%, respectively), but there was a tendency to decreased cellular growth rate after passage 3. The ADSC were characterized by the expression of surface markers such as CD29 (67.4%) and CD44 (89.4%), using CD 45 (0.77%) as a negative control. ADSC from rabbits were successfully isolated form the inguinal region. These cells were capable to differentiate into osteogenic and adipogenic tissue when they were placed in inductive media. After each passage, there was a trend towards decreased cell growth. On the other hand, DNA fragmentation increased at each passage. ADSC had a different lipid profile when placed in control, adipogenic, or osteogenic media.

  11. Evaluation of a dental pulp-derived cell sheet cultured on amniotic membrane substrate.

    Science.gov (United States)

    Honjo, Ken-ichi; Yamamoto, Toshiro; Adachi, Tetsuya; Amemiya, Takeshi; Mazda, Osam; Kanamura, Narisato; Kita, Masakazu

    2015-01-01

    Mesenchymal stem cells (MSC) are transplanted for periodontal tissue regeneration, and the periodontal ligament (PDL) is regenerated using a cultured cell sheet. This cultured cell sheet is prepared using PDL-derived cells, growth factors, and amniotic membrane (AM). Dental pulp (DP)-derived cells can be easily obtained from extracted wisdom teeth, proliferate rapidly, and are less susceptible to bacterial infection than PDL-derived cells. Thus, to prepare a novel cell sheet, DP-derived cells were cultured on AM as a culture substrate for immunohistochemical examination. Wisdom teeth extracted from three adults were cut along the cement-enamel border. DP tissue was collected, minced, and primarily cultured. After three or four passage cultures, DP-derived cells were cultured on AM, followed by hematoxylin-eosin (H-E) and immunofluorescence staining. DP-derived cells cultured on AM formed a layered structure. Cells positive for vimentin, Ki-67, ZO-1, desmoplakin, CD29, 44, 105 or 146, STRO-1, collagen IV or VII or laminin 5 or α5 chain were localized. DP-derived cells proliferated on AM, while retaining the properties of DP, which allowed the cultured cell sheet to be prepared. In addition, the cultured cell sheet contained MSC, which suggests its potential application in periodontal tissue regeneration.

  12. Chitosan derived co-spheroids of neural stem cells and mesenchymal stem cells for neural regeneration.

    Science.gov (United States)

    Han, Hao-Wei; Hsu, Shan-Hui

    2017-10-01

    Chitosan has been considered as candidate biomaterials for neural applications. The effective treatment of neurodegeneration or injury to the central nervous system (CNS) is still in lack nowadays. Adult neural stem cells (NSCs) represents a promising cell source to treat the CNS diseases but they are limited in number. Here, we developed the core-shell spheroids of NSCs (shell) and mesenchymal stem cells (MSCs, core) by co-culturing cells on the chitosan surface. The NSCs in chitosan derived co-spheroids displayed a higher survival rate than those in NSC homo-spheroids. The direct interaction of NSCs with MSCs in the co-spheroids increased the Notch activity and differentiation tendency of NSCs. Meanwhile, the differentiation potential of MSCs in chitosan derived co-spheroids was significantly enhanced toward neural lineages. Furthermore, NSC homo-spheroids and NSC/MSC co-spheroids derived on chitosan were evaluated for their in vivo efficacy by the embryonic and adult zebrafish brain injury models. The locomotion activity of zebrafish receiving chitosan derived NSC homo-spheroids or NSC/MSC co-spheroids was partially rescued in both models. Meanwhile, the higher survival rate was observed in the group of adult zebrafish implanted with chitosan derived NSC/MSC co-spheroids as compared to NSC homo-spheroids. These evidences indicate that chitosan may provide an extracellular matrix-like environment to drive the interaction and the morphological assembly between NSCs and MSCs and promote their neural differentiation capacities, which can be used for neural regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. 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.

  14. Regulatory effects of sestrin 3 (SESN3 in BCR-ABL expressing cells.

    Directory of Open Access Journals (Sweden)

    Eliza Vakana

    Full Text Available Chronic myeloid leukemia (CML and Ph+ acute lymphoblastic leukemia (ALL are characterized by the presence of the BCR-ABL oncoprotein, which leads to activation of a plethora of pro-mitogenic and pro-survival pathways, including the mTOR signaling cascade. We provide evidence that in BCR-ABL expressing cells, treatment with tyrosine kinase inhibitors (TKIs results in upregulation of mRNA levels and protein expression of sestrin3 (SESN3, a unique cellular inhibitor of mTOR complex 1 (mTORC1. Such upregulation appears to be mediated by regulatory effects on mTOR, as catalytic inhibition of the mTOR kinase also induces SESN3. Catalytic mTOR inhibition also results in upregulation of SESN3 expression in cells harboring the TKI-insensitive T315I-BCR-ABL mutant, which is resistant to imatinib mesylate. Overexpression of SESN3 results in inhibitory effects on different Ph+ leukemic cell lines including KT-1-derived leukemic precursors, indicating that SESN3 mediates anti-leukemic responses in Ph+ cells. Altogether, our findings suggest the existence of a novel mechanism for the generation of antileukemic responses in CML cells, involving upregulation of SESN3 expression.

  15. Transglutaminase 2 expression in acute myeloid leukemia: Association with adhesion molecule expression and leukemic blast motility

    Science.gov (United States)

    Meyer, Stefan; Ravandi-Kashani, Farhad; Borthakur, Gautam; Coombes, Kevin R.; Zhang, Nianxiang; Kornblau, Steven

    2016-01-01

    Acute myeloid leukemia (AML) is a heterogenous disease with differential oncogene association, outcome and treatment regimens. Treatment strategies for AML have improved outcome but despite increased molecular biological information AML is still associated with poor prognosis. Proteomic analysis on the effects of a range of leukemogenic oncogenes showed that the protein transglutaminase 2 (TG2) is expressed at greater levels as a consequence of oncogenic transformation. Further analysis of this observation was performed with 511 AML samples using reverse phase proteomic arrays, demonstrating that TG2 expression was higher at relapse than diagnosis in many cases. In addition elevated TG2 expression correlated with increased expression of numerous adhesion proteins and many apoptosis regulating proteins, two processes related to leukemogenesis. TG2 has previously been linked to drug resistance in cancer and given the negative correlation between TG2 levels and peripheral blasts observed increased TG2 levels may lead to the protection of the leukemic stem cell due to increased adhesion/reduced motility. TG2 may therefore form part of a network of proteins that define poor outcome in AML patients and potentially offer a target to sensitize AML stem cells to drug treatment. PMID:23576428

  16. 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.

  17. 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.

  18. Cytoprotective dibenzoylmethane derivatives protect cells from oxidative stress-induced necrotic cell death.

    Science.gov (United States)

    Hegedűs, Csaba; Lakatos, Petra; Kiss-Szikszai, Attila; Patonay, Tamás; Gergely, Szabolcs; Gregus, Andrea; Bai, Péter; Haskó, György; Szabó, Éva; Virág, László

    2013-06-01

    Screening of a small in-house library of 1863 compounds identified 29 compounds that protected Jurkat cells from hydrogen peroxide-induced cytotoxicity. From the cytoprotective compounds eleven proved to possess antioxidant activity (ABTS radical scavenger effect) and two were found to inhibit poly(ADP-ribosyl)ation (PARylation), a cytotoxic pathway operating in severely injured cells. Four cytoprotective dibenzoylmethane (DBM) derivatives were investigated in more detail as they did not scavenge hydrogen peroxide nor did they inhibit PARylation. These compounds protected cells from necrotic cell death while caspase activation, a parameter of apoptotic cell death was not affected. Hydrogen peroxide activated extracellular signal regulated kinase (ERK1/2) and p38 MAP kinases but not c-Jun N-terminal kinase (JNK). The cytoprotective DBMs suppressed the activation of Erk1/2 but not that of p38. Cytoprotection was confirmed in another cell type (A549 lung epithelial cells), indicating that the cytoprotective effect is not cell type specific. In conclusion we identified DBM analogs as a novel class of cytoprotective compounds inhibiting ERK1/2 kinase and protecting from necrotic cell death by a mechanism independent of poly(ADP-ribose) polymerase inhibition. Copyright © 2013 Elsevier Ltd. All rights reserved.

  19. Adipose derived stem cells in radiotherapy injury: a new frontier

    Directory of Open Access Journals (Sweden)

    Lipi eShukla

    2015-01-01

    Full Text Available Radiotherapy is increasingly used to treat numerous human malignancies. In addition to the beneficial anti-cancer effects, there are a series of undesirable effects on normal host tissues surrounding the target tumour. Whilst the early effects of radiotherapy (desquamation, erythema and hair loss typically resolve, the chronic effects persist as unpredictable and often troublesome sequelae of cancer treatment, long after oncological treatment has been completed. Plastic surgeons are often called upon to treat the problems subsequently arising in irradiated tissues, such as recurrent infection, impaired healing, fibrosis, contracture and/or lymphoedema. Recently, it was anecdotally noted - then validated in more robust animal and human studies - that fat grafting can ameliorate some of these chronic tissue effects. Despite the widespread usage of fat grafting, the mechanism of its action remains poorly understood. This review provides an overview of the current understanding of (i mechanisms of chronic radiation injury and its clinical manifestations; (ii biological properties of fat grafts and their key constituent, Adipose-Derived Stem Cells (ADSCs; (iii the role of ADSCs in radiotherapy-induced soft-tissue injury.

  20. Characteristics of bovine inner cell mass-derived cell lines and their fate in chimeric conceptuses.

    Science.gov (United States)

    Furusawa, Tadashi; Ohkoshi, Katsuhiro; Kimura, Koji; Matsuyama, Shuichi; Akagi, Satoshi; Kaneda, Masahiro; Ikeda, Mitsumi; Hosoe, Misa; Kizaki, Keiichiro; Tokunaga, Tomoyuki

    2013-08-01

    Bovine embryonic stem (ES) cells have the potential to provide significant benefits in a range of agricultural and biomedical applications. Here, we employed a combination of conventional methods using glycogen synthase kinase 3 and mitogen-activated protein kinase inhibitors to establish ES cell lines from in vitro fertilization (IVF) and somatic cell nuclear transfer (SCNT) bovine embryos. Five male cell lines were established from IVF embryos, and two female and three male cell lines from SCNT blastocysts; we named these lines bovine ES cell-like cells (bESLCs). The lines exhibited dome-shaped colonies, stained positively for alkaline phosphatase, and expressed pluripotent stem cell markers such as POU5F1, SOX2, and SSEA-1. The expression levels of these markers, especially for NANOG, varied among the cell lines. A DNA methylation assay showed the POU5F1 promoter region was hypomethylated compared to fibroblast cells. An in vitro differentiation assay showed that endoderm and ectoderm marker genes, but not mesoderm markers, were upregulated in differentiating bESLCs. To examine bESLCs in later embryonic stages, we created 22 chimeric blastocysts with a male bESLC line carrying a GFP marker gene and transferred these to a recipient cow. Four chimeric embryos were subsequently retrieved on Day 13 and retransferred to two recipient cows. One living fetus was obtained at Day 62. GFP signals were not identified in fetal cells by fluorescence microscopy; however, genomic PCR analysis detected the GFP gene in major organs. Clusters of GFP-positive cells were observed in amniotic membranes, suggesting that bESLCs can be categorized as a novel type of ICM-derived cells that can potentially differentiate into epiblast and hypoblast lineages.

  1. Optimization of in vitro cell labeling methods for human umbilical cord-derived mesenchymal stem cells.

    Science.gov (United States)

    Tao, R; Sun, T-J; Han, Y-Q; Xu, G; Liu, J; Han, Y-F

    2014-01-01

    Human umbilical cord-derived mesenchymal stem cells (hUCMSCs) are a novel source of seed cells for cell therapy and tissue engineering. However, in vitro labeling methods for hUCMSCs need to be optimized for better detection of transplanted cells. To identify the most stable and efficient method for labeling hUCMSCs in vitro. hUCMSCs were isolated using a modified enzymatic digestion procedure and cultured. hUCMSCs of passage three (P3) were then labeled with BrdU, PKH26, or lentivirus-GFP and passaged further. Cells from the first labeled passage (LP1), the fourth labeled passage (LP4) and later passages were observed using a fluorescence microscope. The differentiation potential of LP4 cells was assessed by induction with adipogenic and osteogenic medium. Flow cytometry was used to measure the percentage of labeled cells and the percentage of apoptotic or dead cells. The labeling efficiencies of the three hUCMSC-labeling methods were compared in vitro. BrdU, PKH26, and lentivirus-GFP all labeled LP1 cells with high intensity and clarity. However, the BrdU labeling of the LP4 cells was vague and not localized to the cell nuclei; LP9 cells were not detected under a fluorescence microscope. There was also a significant decrease in the fluorescence intensity of PKH26-labeled LP4 cells, and LP11 cells were not detected under a fluorescence microscope. However, the fluorescence of LP4 cells labeled with lentivirus-GFP remained strong, and cells labeled with lentivirus-GFP were detected up to LP14 under a fluorescence microscope. Statistical analyses indicated that percentages of LP1 cells labeled with PKH26 and lentivirus-GFP were significantly higher than that of cells labeled with BrdU (p 0.05) was observed between the death rates of labeled and unlabeled cells. Lentivirus-GFP is a valid method for long-term in vitro labeling, and it may be used as a long-term hUCMSC tracker following transplantation in vivo.

  2. Human umbilical cord blood-derived stem cells and brain-derived neurotrophic factor protect injured optic nerve: viscoelasticity characterization

    Directory of Open Access Journals (Sweden)

    Xue-man Lv

    2016-01-01

    Full Text Available The optic nerve is a viscoelastic solid-like biomaterial. Its normal stress relaxation and creep properties enable the nerve to resist constant strain and protect it from injury. We hypothesized that stress relaxation and creep properties of the optic nerve change after injury. More-over, human brain-derived neurotrophic factor or umbilical cord blood-derived stem cells may restore these changes to normal. To validate this hypothesis, a rabbit model of optic nerve injury was established using a clamp approach. At 7 days after injury, the vitreous body re-ceived a one-time injection of 50 µg human brain-derived neurotrophic factor or 1 × 106 human umbilical cord blood-derived stem cells. At 30 days after injury, stress relaxation and creep properties of the optic nerve that received treatment had recovered greatly, with patho-logical changes in the injured optic nerve also noticeably improved. These results suggest that human brain-derived neurotrophic factor or umbilical cord blood-derived stem cell intervention promotes viscoelasticity recovery of injured optic nerves, and thereby contributes to nerve recovery.

  3. Osteoblasts Protect AML Cells from SDF-1-Induced Apoptosis

    Science.gov (United States)

    Kremer, Kimberly N.; Dudakovic, Amel; McGee-Lawrence, Meghan E.; Philips, Rachael L.; Hess, Allan D.; Smith, B. Douglas; van Wijnen, Andre J.; Karp, Judith E.; Kaufmann, Scott H.; Westendorf, Jennifer J.; Hedin, Karen E.

    2014-01-01

    The bone marrow provides a protective environment for acute myeloid leukemia (AML) cells that often allows leukemic stem cells to survive standard chemotherapeutic regimens. Targeting these leukemic stem cells within the bone marrow is critical for preventing relapse. We recently demonstrated that SDF-1, a chemokine abundant in the bone marrow, induces apoptosis in AML cell lines and in patient samples expressing high levels of its receptor, CXCR4. Here we show that a subset of osteoblast lineage cells within the bone marrow can protect AML cells from undergoing apoptosis in response to the SDF-1 naturally present in that location. In co-culture systems, osteoblasts at various stages of differentiation protected AML cell lines and patient isolates from SDF-1-induced apoptosis. The differentiation of the osteoblast cell lines, MC3T3 and W-20-17, mediated this protection via a cell contact-independent mechanism. In contrast, bone marrow-derived mesenchymal cells, the precursors of osteoblasts, induced apoptosis in AML cells via a CXCR4-dependent mechanism and failed to protect AML cells from exogenously added SDF-1. These results indicate that osteoblasts in the process of differentiation potently inhibit the SDF-1-driven apoptotic pathway of CXCR4-expressing AML cells residing in the bone marrow. Drugs targeting this protective mechanism could potentially provide a new approach to treating AML by enhancing the SDF-1-induced apoptosis of AML cells residing within the bone marrow microenvironment. PMID:24851270

  4. Synovium-derived stem cells: a tissue-specific stem cell for cartilage engineering and regeneration.

    Science.gov (United States)

    Jones, Brendan A; Pei, Ming

    2012-08-01

    Articular cartilage is difficult to heal once injury or disease occurs. Autologous chondrocyte transplantation is a biological treatment with good prognosis, but donor site morbidity and limited cell source are disadvantages. Currently, mesenchymal stem cells (MSCs) are a promising approach for cartilage regeneration. Despite there being various sources, the best candidate for cartilage regeneration is the one with the greatest chondrogenic potential and the least hypertrophic differentiation. These properties are able to insure that the regenerated tissue is hyaline cartilage of high quality. This review article will summarize relevant literature to justify synovium-derived stem cells (SDSCs) as a tissue-specific stem cell for chondrogenesis by comparing synovium and cartilage with respect to anatomical location and functional structure, comparing the growth characterization and chondrogenic capacity of SDSCs and MSCs, evaluating the application of SDSCs in regenerative medicine and diseases, and discussing potential future directions.

  5. Activated T cells sustain myeloid-derived suppressor cell-mediated immune suppression

    Science.gov (United States)

    Damuzzo, Vera; Francescato, Samuela; Pozzuoli, Assunta; Berizzi, Antonio; Mocellin, Simone; Rossi, Carlo Riccardo; Bronte, Vincenzo; Mandruzzato, Susanna

    2016-01-01

    The expansion of myeloid derived suppressor cells (MDSCs), a suppressive population able to hamper the immune response against cancer, correlates with tumor progression and overall survival in several cancer types. We have previously shown that MDSCs can be induced in vitro from precursors present in the bone marrow and observed that these cells are able to actively proliferate in the presence of activated T cells, whose activation level is critical to drive the suppressive activity of MDSCs. Here we investigated at molecular level the mechanisms involved in the interplay between MDSCs and activated T cells. We found that activated T cells secrete IL-10 following interaction with MDSCs which, in turn, activates STAT3 phosphorylation on MDSCs then leading to B7-H1 expression. We also demonstrated that B7-H1+ MDSCs are responsible for immune suppression through a mechanism involving ARG-1 and IDO expression. Finally, we show that the expression of ligands B7-H1 and MHC class II both on in vitro-induced MDSCs and on MDSCs in the tumor microenvironment of cancer patients is paralleled by an increased expression of their respective receptors PD-1 and LAG-3 on T cells, two inhibitory molecules associated with T cell dysfunction. These findings highlight key molecules and interactions responsible for the extensive cross-talk between MDSCs and activated T cells that are at the basis of immune suppression. PMID:26700461

  6. 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.

  7. Bone marrow-derived fibrocytes promote stem cell-like properties of lung cancer cells.

    Science.gov (United States)

    Saijo, Atsuro; Goto, Hisatsugu; Nakano, Mayuri; Mitsuhashi, Atsushi; Aono, Yoshinori; Hanibuchi, Masaki; Ogawa, Hirohisa; Uehara, Hisanori; Kondo, Kazuya; Nishioka, Yasuhiko

    2018-05-01

    Cancer stem cells (CSCs) represent a minor population that have clonal tumor initiation and self-renewal capacity and are responsible for tumor initiation, metastasis, and therapeutic resistance. CSCs reside in niches, which are composed of diverse types of stromal cells and extracellular matrix components. These stromal cells regulate CSC-like properties by providing secreted factors or by physical contact. Fibrocytes are differentiated from bone marrow-derived CD14 + monocytes and have features of both macrophages and fibroblasts. Accumulating evidence has suggested that stromal fibrocytes might promote cancer progression. However, the role of fibrocytes in the CSC niches has not been revealed. We herein report that human fibrocytes enhanced the CSC-like properties of lung cancer cells through secreted factors, including osteopontin, CC-chemokine ligand 18, and plasminogen activator inhibitor-1. The PIK3K/AKT pathway was critical for fibrocytes to mediate the CSC-like functions of lung cancer cells. In human lung cancer specimens, the number of tumor-infiltrated fibrocytes was correlated with high expression of CSC-associated protein in cancer cells. These results suggest that fibrocytes may be a novel cell population that regulates the CSC-like properties of lung cancer cells in the CSC niches. Copyright © 2018. Published by Elsevier B.V.

  8. Cell-derived micro-environment helps dental pulp stem cells promote dental pulp regeneration.

    Science.gov (United States)

    Zhang, Xuexin; Li, Hui; Sun, Jingjing; Luo, Xiangyou; Yang, Hefeng; Xie, Li; Yang, Bo; Guo, Weihua; Tian, Weidong

    2017-10-01

    The function of the dental pulp is closely connected to the extracellular matrix (ECM) structure, and ECM has received significant attention due to its biological functions for regulating cells. As such, the interaction between the ECM niche and cells is worth exploring for potential clinical uses. In this study, dental pulp stem cell (DPSC)-derived ECM (DPM) was prepared through cell culture and decellularization to function as the cell niche, and changes in DPSC behaviour and histological analysis of dental pulp tissue regeneration were evaluated following the DPM culture. DPM promoted the replication of DPSCs and exhibited retention of their mineralization. Then, the DPM-based culture strategy under odontogenic culture medium was further investigated, and the mineralization-related markers showed that DPSCs were regulated towards odontogenic differentiation. Dental pulp-like tissue with well-arranged ECM was harvested after a 2-month subcutaneous implantation in nude mice with DPM application. Additionally, DPSCs cultured on the plastic culture surface showed the up-regulation of mineralization makers in vitro, but there was a disorder in matrix formation and mineralization when the cells were cultured in vivo. DPM-based cultivation could serve as a cell niche and modulate DPSC behaviour, and this method also provided an alternative to harvest tissue-specific ECM and provided a strategy for ECM-cell interaction. © 2017 John Wiley & Sons Ltd.

  9. Isolation and expansion of adipose-derived stem cells for tissue engineering

    DEFF Research Database (Denmark)

    Fink, Trine; Rasmussen, Jeppe Grøndahl; Lund, Pia

    2011-01-01

    For treatment of cardiac failure with bone marrow-derived mesenchymal stem cells, several clinical trials are ongoing. However, more attention is gathering on the use of adipose tissue-derived stem cells (ASCs). This paper describes the optimization of isolation and propagation of ASCs for subseq......For treatment of cardiac failure with bone marrow-derived mesenchymal stem cells, several clinical trials are ongoing. However, more attention is gathering on the use of adipose tissue-derived stem cells (ASCs). This paper describes the optimization of isolation and propagation of ASCs...

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

    Directory of Open Access Journals (Sweden)

    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.

  11. 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.

  12. Induced pluripotent stem cells (iPSC)-derived retinal cells in disease modeling and regenerative medicine.

    Science.gov (United States)

    Rathod, Reena; Surendran, Harshini; Battu, Rajani; Desai, Jogin; Pal, Rajarshi

    2018-02-12

    Retinal degenerative disorders are a leading cause of the inherited, irreversible and incurable vision loss. While various rodent model systems have provided crucial information in this direction, lack of disease-relevant tissue availability and species-specific differences have proven to be a major roadblock. Human induced pluripotent stem cells (iPSC) have opened up a whole new avenue of possibilities not just in understanding the disease mechanism but also potential therapeutic approaches towards a cure. In this review, we have summarized recent advances in the methods of deriving retinal cell types from iPSCs which can serve as a renewable source of disease-relevant cell population for basic as well as translational studies. We also provide an overview of the ongoing efforts towards developing a suitable in vitro model for modeling retinal degenerative diseases. This basic understanding in turn has contributed to advances in translational goals such as drug screening and cell-replacement therapies. Furthermore we discuss gene editing approaches for autologous repair of genetic disorders and allogeneic transplantation of stem cell-based retinal derivatives for degenerative disorders with an ultimate goal to restore vision. It is pertinent to note however, that these exciting new developments throw up several challenges that need to be overcome before their full clinical potential can be realized. Copyright © 2018 Elsevier B.V. All rights reserved.

  13. 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

  14. Generation of Dopamine-Secreting Cells from Human Adipose Tissue-Derived Stem Cells In Vitro.

    Science.gov (United States)

    Soheilifar, Mohammad Hasan; Javeri, Arash; Amini, Hossein; Taha, Masoumeh Fakhr

    2018-03-12

    Several studies have demonstrated the differentiation of human adipose tissue-derived stem cells (hADSCs) to neuronal and glial phenotypes, but directing the fate of these cells toward dopaminergic neurons has not been frequently reported. The aim of this study was to investigate dopaminergic specification of hADSCs in vitro. ADSCs were isolated from subcutaneous abdominal adipose tissue and were characterized. For dopaminergic differentiation, a cocktail of sonic hedgehog, fibroblast growth factor 8, basic fibroblast growth factor, and brain-derived neurotrophic factor were used under a low serum condition. As the control group, the ADSCs were cultured under the same low serum condition without the dopaminergic cocktail. At the end of differentiation period, the cells expressed neuron-specific markers, NES, NSE, and NEFL, and dopaminergic markers, EN1, NURR1, PITX3, VMAT2, TH, and GIRK2 genes. TH, NURR1, and EN1 mRNAs were upregulated in the dopaminergic group compared with the control group. NEFL and TH proteins were also expressed in the differentiated cells. A total of 27.9% of the cells differentiated in dopaminergic induction medium showed positive staining for TH protein. Based on reversed-phase high-performance liquid chromatography analysis, the differentiated cells released a significant amount of dopamine in response to KCl-induced depolarization. In conclusion, results of this study indicate that hADSCs can be induced by a growth factor cocktail to produce dopamine secreting cells with possible applications for future cell replacement therapy of Parkinson's disease.

  15. Efflux protein expression in human stem cell-derived retinal pigment epithelial cells.

    Directory of Open Access Journals (Sweden)

    Kati Juuti-Uusitalo

    Full Text Available Retinal pigment epithelial (RPE cells in the back of the eye nourish photoreceptor cells and form a selective barrier that influences drug transport from the blood to the photoreceptor cells. At the molecular level, ATP-dependent efflux transporters have a major role in drug delivery in human RPE. In this study, we assessed the relative expression of several ATP-dependent efflux transporter genes (MRP1, -2, -3, -4, -5, -6, p-gp, and BCRP, the protein expression and localization of MRP1, MRP4, and MRP5, and the functionality of MRP1 efflux pumps at different maturation stages of undifferentiated human embryonic stem cells (hESC and RPE derived from the hESC (hESC-RPE. Our findings revealed that the gene expression of ATP-dependent efflux transporters MRP1, -3, -4, -5, and p-gp fluctuated during hESC-RPE maturation from undifferentiated hESC to fusiform, epithelioid, and finally to cobblestone hESC-RPE. Epithelioid hESC-RPE had the highest expression of MRP1, -3, -4, and P-gp, whereas the most mature cobblestone hESC-RPE had the highest expression of MRP5 and MRP6. These findings indicate that a similar efflux protein profile is shared between hESC-RPE and the human RPE cell line, ARPE-19, and suggest that hESC-RPE cells are suitable in vitro RPE models for drug transport studies. Embryonic stem cell model might provide a novel tool to study retinal cell differentiation, mechanisms of RPE-derived diseases, drug testing and targeted drug therapy.

  16. Pulmonary leukemic involvement: high-resolution computed tomography evaluation

    International Nuclear Information System (INIS)

    Oliveira, Ana Paola de; Marchiori, Edson; Souza Junior, Arthur Soares

    2004-01-01

    Objective: To evaluate the role of high-resolution computed tomography (HRCT) in patients with leukemia and pulmonary symptoms, to establish the main patterns and to correlate them with the etiology. Materials and Methods: This is a retrospective study of the HRCT of 15 patients with leukemia and pulmonary symptoms. The examinations were performed using a spatial high-resolution protocol and were analyzed by two independent radiologists. Results: The main HRCT patterns found were ground-glass opacity (n=11), consolidation (n=9), airspace nodules (n=3), septal thickening (n=3), tree-in-bud pattern (n=3), and pleural effusion (n=3). Pulmonary infection was the most common finding seen in 12 patients: bacterial pneumonia (n=6), fungal infection (n = 4), pulmonary tuberculosis (n=1) and viral infection (n=1). Leukemic pleural infiltration (n=1), lymphoma (n=1) and pulmonary hemorrhage (n=1) were detected in the other three patients. Conclusion: HRCT is an important tool that may suggest the cause of lung involvement, its extension and in some cases to guide invasive procedures in patients with leukemia. (author)

  17. Comparison of neurosphere-like cell clusters derived from dental follicle precursor cells and retinal Müller cells

    DEFF Research Database (Denmark)

    Beck, Hans Christian; Petersen, Jørgen; Felthaus, Oliver

    2011-01-01

    Unrelated cells such as dental follicle precursor cells (DFPCs) and retinal Müller cells (MCs) make spheres after cultivation in serum-replacement medium (SRM). Until today, the relation and molecular processes of sphere formation from different cell types remain undescribed. Thus in this study we...... compared proteomes of spheres derived from MCs and DFPCs. 73% of 676 identified proteins were similar expressed in both cell types and many of them are expressed in the brain (55%). Moreover proteins are overrepresented that are associated with pathways for neural diseases such as Huntington disease...... or Alzheimer disease. Interestingly up-regulated proteins in DFPCs are involved in the biosynthesis of glycosphingolipids. These lipids are components of gangliosides such as GD3, which is a novel neural stem cell marker. In conclusion spheres from different types of cells have highly similar proteomes...

  18. Sox17-Mediated XEN Cell Conversion Identifies Dynamic Networks Controlling Cell-Fate Decisions in Embryo-Derived Stem Cells

    Directory of Open Access Journals (Sweden)

    Angela C.H. McDonald

    2014-10-01

    Full Text Available Little is known about the gene regulatory networks (GRNs distinguishing extraembryonic endoderm (ExEn stem (XEN cells from those that maintain the extensively characterized embryonic stem cell (ESC. An intriguing network candidate is Sox17, an essential transcription factor for XEN derivation and self-renewal. Here, we show that forced Sox17 expression drives ESCs toward ExEn, generating XEN cells that contribute to ExEn when placed back into early mouse embryos. Transient Sox17 expression is sufficient to drive this fate change during which time cells transit through distinct intermediate states prior to the generation of functional XEN-like cells. To orchestrate this conversion process, Sox17 acts in autoregulatory and feedforward network motifs, regulating dynamic GRNs directing cell fate. Sox17-mediated XEN conversion helps to explain the regulation of cell-fate changes and reveals GRNs regulating lineage decisions in the mouse embryo.

  19. Amniotic fluid-derived mesenchymal stem cells as a novel ...

    African Journals Online (AJOL)

    CLEMENTINA

    2012-06-28

    Jun 28, 2012 ... stem cells (AFMSCs) have many advantages over other stem cells: avoiding much ethical controversy ... showed that induced pluripotent stem cells (iPSCs) have ... disadvantages of ESCs, BM-MSCs and iPSCs have.

  20. Epigenetic and phenotypic profile of fibroblasts derived from induced pluripotent stem cells.

    Directory of Open Access Journals (Sweden)

    Kyle J Hewitt

    2011-02-01

    Full Text Available Human induced pluripotent stem (hiPS cells offer a novel source of patient-specific cells for regenerative medicine. However, the biological potential of iPS-derived cells and their similarities to cells differentiated from human embryonic stem (hES cells remain unclear. We derived fibroblast-like cells from two hiPS cell lines and show that their phenotypic properties and patterns of DNA methylation were similar to that of mature fibroblasts and to fibroblasts derived from hES cells. iPS-derived fibroblasts (iPDK and their hES-derived counterparts (EDK showed similar cell morphology throughout differentiation, and patterns of gene expression and cell surface markers were characteristic of mature fibroblasts. Array-based methylation analysis was performed for EDK, iPDK and their parental hES and iPS cell lines, and hierarchical clustering revealed that EDK and iPDK had closely-related methylation profiles. DNA methylation analysis of promoter regions associated with extracellular matrix (ECM-production (COL1A1 by iPS- and hESC-derived fibroblasts and fibroblast lineage commitment (PDGFRβ, revealed promoter demethylation linked to their expression, and patterns of transcription and methylation of genes related to the functional properties of mature stromal cells were seen in both hiPS- and hES-derived fibroblasts. iPDK cells also showed functional properties analogous to those of hES-derived and mature fibroblasts, as seen by their capacity to direct the morphogenesis of engineered human skin equivalents. Characterization of the functional behavior of ES- and iPS-derived fibroblasts in engineered 3D tissues demonstrates the utility of this tissue platform to predict the capacity of iPS-derived cells before their therapeutic application.

  1. 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.

  2. To the nucleolar density and size in apoptotic human leukemic myeloblasts produced in vitro by Trichostatin A

    Directory of Open Access Journals (Sweden)

    K Smetana

    2009-08-01

    Full Text Available The present study was designed to provide more information on nucleoli in apoptotic cells, which were represented in the present study by cultured leukemic myeloblasts (Kasumi-1 cells. The apoptotic process in these cells was produced by trichostatin A (TSA that is a histone deacetylase inhibitor with strong cytostatic effects. The selected TSA concentration added to cultures facilitated to study apoptotic and notapoptotic cells in one and the same specimen. The nucleolar diameter and density were determined using computer assisted measurement and densitometry in specimens stained for RNA. In comparison with not-apoptotic cells, in apoptotic cells, nucleolar mean diameter did not change significantly and nucleolar RNA density was also not apparently different. On the other hand, the cytoplasmic RNA density in apoptotic cells was markedly reduced. Thus it seemed to be possible that the transcribed RNA remained “frozen” within the nucleolus but its transport to the cytoplasm decreased or stopped. However, the possibility of the RNA degradation in the cytoplasm of apoptotic cells based on the present study cannot be eliminated. At this occasion it should be added that AgNORs reflecting nucleolar biosynthetic and cell proliferation activity in apoptotic cells decreased in number or disappeared. The presented results also indicated that large nucleoli intensely stained for RNA need not be necessarily related to the high nucleolar biosynthetic or cell proliferation activity and may be also present in apoptotic cells responding to the cytostatic treatment.

  3. Skeletal Muscle-derived Hematopoietic Stem Cells: Muscular Dystrophy Therapy by Bone Marrow Transplantation

    OpenAIRE

    Asakura, Atsushi

    2012-01-01

    For postnatal growth and regeneration of skeletal muscle, satellite cells, a self-renewing pool of muscle stem cells, give rise to daughter myogenic precursor cells that contribute to the formation of new muscle fibers. In addition to this key myogenic cell class, adult skeletal muscle also contains hematopoietic stem cell and progenitor cell populations which can be purified as a side population (SP) fraction or as a hematopoietic marker CD45-positive cell population. These muscle-derived he...

  4. Tissue Source and Cell Expansion Condition Influence Phenotypic Changes of Adipose-Derived Stem Cells

    Directory of Open Access Journals (Sweden)

    Lauren H. Mangum

    2017-01-01

    Full Text Available Stem cells derived from the subcutaneous adipose tissue of debrided burned skin represent an appealing source of adipose-derived stem cells (ASCs for regenerative medicine. Traditional tissue culture uses fetal bovine serum (FBS, which complicates utilization of ASCs in human medicine. Human platelet lysate (hPL is one potential xeno-free, alternative supplement for use in ASC culture. In this study, adipogenic and osteogenic differentiation in media supplemented with 10% FBS or 10% hPL was compared in human ASCs derived from abdominoplasty (HAP or from adipose associated with debrided burned skin (BH. Most (95–99% cells cultured in FBS were stained positive for CD73, CD90, CD105, and CD142. FBS supplementation was associated with increased triglyceride content and expression of adipogenic genes. Culture in hPL significantly decreased surface staining of CD105 by 31% and 48% and CD142 by 27% and 35% in HAP and BH, respectively (p<0.05. Culture of BH-ASCs in hPL also increased expression of markers of osteogenesis and increased ALP activity. These data indicate that application of ASCs for wound healing may be influenced by ASC source as well as culture conditions used to expand them. As such, these factors must be taken into consideration before ASCs are used for regenerative purposes.

  5. Tissue Source and Cell Expansion Condition Influence Phenotypic Changes of Adipose-Derived Stem Cells

    Science.gov (United States)

    Mangum, Lauren H.; Stone, Randolph; Wrice, Nicole L.; Larson, David A.; Florell, Kyle F.; Christy, Barbara A.; Herzig, Maryanne C.; Cap, Andrew P.

    2017-01-01

    Stem cells derived from the subcutaneous adipose tissue of debrided burned skin represent an appealing source of adipose-derived stem cells (ASCs) for regenerative medicine. Traditional tissue culture uses fetal bovine serum (FBS), which complicates utilization of ASCs in human medicine. Human platelet lysate (hPL) is one potential xeno-free, alternative supplement for use in ASC culture. In this study, adipogenic and osteogenic differentiation in media supplemented with 10% FBS or 10% hPL was compared in human ASCs derived from abdominoplasty (HAP) or from adipose associated with debrided burned skin (BH). Most (95–99%) cells cultured in FBS were stained positive for CD73, CD90, CD105, and CD142. FBS supplementation was associated with increased triglyceride content and expression of adipogenic genes. Culture in hPL significantly decreased surface staining of CD105 by 31% and 48% and CD142 by 27% and 35% in HAP and BH, respectively (p < 0.05). Culture of BH-ASCs in hPL also increased expression of markers of osteogenesis and increased ALP activity. These data indicate that application of ASCs for wound healing may be influenced by ASC source as well as culture conditions used to expand them. As such, these factors must be taken into consideration before ASCs are used for regenerative purposes. PMID:29138638

  6. Characterization of the myeloid-derived suppressor cell subset regulated by NK cells in malignant lymphoma.

    Science.gov (United States)

    Sato, Yusuke; Shimizu, Kanako; Shinga, Jun; Hidaka, Michihiro; Kawano, Fumio; Kakimi, Kazuhiro; Yamasaki, Satoru; Asakura, Miki; Fujii, Shin-Ichiro

    2015-03-01

    Myeloid-derived suppressor cells (MDSCs) are a heterogeneous population with the ability to suppress immune responses and are currently classified into three distinct MDSC subsets: monocytic, granulocytic and non-monocytic, and non-granulocytic MDSCs. Although NK cells provide an important first-line defense against newly transformed cancer cells, it is unknown whether NK cells can regulate MDSC populations in the context of cancer. In this study, we initially found that the frequency of MDSCs in non-Hodgkin lymphoma (NHL) patients was increased and inversely correlated with that of NK cells, but not that of T cells. To investigate the regulation of MDSC subsets by NK cells, we used an EL4 murine lymphoma model and found the non-monocytic and non-granulocytic MDSC subset, i.e., Gr1 + CD11b + Ly6G med Ly6C med MDSC, is increased after NK cell depletion. The MDSC population that expresses MHC class II, CD80, CD124, and CCR2 is regulated mainly by CD27 + CD11b + NK cells. In addition, this MDSC subset produces some immunosuppressive cytokines, including IL-10 but not nitric oxide (NO) or arginase. We also examined two subsets of MDSCs (CD14 + HLA-DR - and CD14 - HLA-DR - MDSC) in NHL patients and found that higher IL-10-producing CD14 + HLA-DR - MDSC subset can be seen in lymphoma patients with reduced NK cell frequency in peripheral blood. Our analyses of MDSCs in this study may enable a better understanding of how MDSCs manipulate the tumor microenvironment and are regulated by NK cells in patients with lymphoma.

  7. A Bilayer Construct Cont