bluuebunny/crossref_metadata_2025_split huggingface dataset:
Detection of Cyclin D1 (bcl-1, PRAD1) Overexpression by a Simple Competitive Reverse Transcription-Polymerase Chain Reaction Assay in t(11; 14)(q13; q32)-Bearing B-Cell Malignancies and/or Mantle Cell Lymphoma
Authors: Uchimaru Kaoru , Taniguchi Toshiyasu , Yoshikawa Miwa , Asano Shigetaka , Arnold Andrew , Fujita Toshiro , Motokura Toru 
Published: October 2019

AbstractIn mantle cell lymphoma, the t(11; 14)(q13; q32) and its molecular counterpart, bcl-1 rearrangement, are consistent features and lead to cyclin D1 (bcl-1, PRAD1) proto-oncogene overexpression. In order to detect cyclin D1 overexpression, we developed a simple assay involving a reverse transcription followed by competitive polymerase chain reaction (PCR). A single upstream primer was derived from a homologous region between cyclin D1 and the other D-type cyclins, cyclins D2 and D3, while three downstream primers were specific to their respective D-type cyclins. Because the upstream primer was shared in PCR amplification of the three sequences, each PCR product served as a competitor and the quantification of the target was made by comparison of the intensity of the three products. With this assay we analyzed 45 hematopoietic cell lines and 40 clinical specimens. Cyclin D1 was rarely expressed in lymphoid cell lines except in t(11; 14)(q13; q32)-bearing B-cell malignancies and/or mantle cell lymphoma, which expressed cyclin D1 predominantly. In myeloid cell lines, the levels of cyclin D1 expression varied and never exceeded the sum of cyclin D2 and D3 levels. Cyclin D3 was ubiquitously expressed while cyclins D1 and D2 were differentially used. The observations suggest that human cyclin D3 may play a fundamental role in hematopoiesis and that cyclins D1 and D2 may have different lineage- or differentiation-dependent functions. With this assay, small aliquots of clinical specimens such as 100 μL peripheral blood were enough to detect cyclin D1 overexpression without a well-controlled standard. The technique was validated as highly comparable with Northern analysis. This rapid and reliable detection of cyclin D1 overexpression may have practical clinical utility in the analysis and management of B-cell malignancies.
Interleukin-10 Inhibits Spontaneous Colony-Forming Unit–Granulocyte-Macrophage Growth From Human Peripheral Blood Mononuclear Cells by Suppression of Endogenous Granulocyte-Macrophage Colony-Stimulating Factor Release
Authors: Oehler Leopold , Foedinger Manuela , Koeller Markus , Kollars Marietta , Reiter Elisabeth , Bohle Barbara , Skoupy Sonja , Fritsch Gerhard , Lechner Klaus , Geissler Klaus 
Published: October 2019

AbstractSpontaneous growth of myeloid colonies (colony-forming unit–granulocyte-macrophage [CFU-GM]) can be observed in methylcellulose cultures containing peripheral blood mononuclear cells (PB-MNCs) and is supposedly caused by the release of colony-stimulating factors (CSF ) by accessory cells. Because of its cytokine synthesis-inhibiting effects on T lymphocytes and monocytes, interleukin-10 (IL-10) may be a potential candidate for indirect modulation of hematopoiesis. We studied the effect of recombinant human IL-10 (rhIL-10) on spontaneous growth of myeloid colonies derived from human PB-MNCs. A total of 10 ng/mL of IL-10 almost completely inhibited spontaneous CFU-GM proliferation (by 95.1%; P < .001, n = 7) in unseparated PB-MNCs. This effect was dose-dependent and specific, because a neutralizing anti–IL-10 antibody was able to prevent IL-10–induced suppression of CFU-GM growth. Spontaneous CFU-GM growth, which required the presence of both monocytes (CD14+ cells) and T lymphocytes (CD3+ cells), was also greatly suppressed by a neutralizing anti–granulocyte-macrophage CSF (GM-CSF ) antibody but was only slightly or not at all inhibited by antibodies against G-CSF or IL-3. Moreover, IL-10–suppressed colony growth could be completely restored by the addition of exogenous GM-CSF. Using semiquantitative polymerase chain reaction, we were able to show that GM-CSF transcripts that spontaneously increased in PB-MNCs within 48 hours of culture were markedly reduced by the addition of IL-10. Inhibiton of GM-CSF production in PB-MNCs by IL-10 was also confirmed at the protein level by measuring GM-CSF levels in suspension cultures. Our findings suggest that autonomous CFU-GM growth, resulting from an interaction of monocytes and T lymphocytes, is mainly caused by endogenous GM-CSF release and can be profoundly suppressed by the addition of exogenous IL-10. Considering the strong inhibitory action of IL-10 on GM-CSF production and spontaneous cell growth in vitro, this cytokine may be useful in myeloid malignancies in which autocrine and/or paracrine mechanisms involving GM-CSF are likely to play a pathogenetic role.
Initiation of Murine Embryonic Erythropoiesis: A Spatial Analysis
Authors: Silver Louise , Palis James 
Published: October 2019

AbstractHematopoiesis in the mouse conceptus begins in the visceral yolk (VYS), with primitive erythroblasts first evident in blood islands at the headfold stage (E8.0). VYS erythropoiesis is decreased or abrogated by targeted disruption of the hematopoietic transcription factors tal-1, rbtn2, GATA-1, and GATA-2. To better understand the potential roles of these genes, and to trace the initial temporal and spatial development of mammalian embryonic hematopoiesis, we examined their expression patterns, and that of βH1-globin, in normal mouse conceptuses by means of in situ hybridization. Attention was focused on the 36-hour period from mid-primitive streak to early somite stages (E7.25 to E8.5), when the conceptus undergoes rapid morphologic changes with formation of the yolk sac and blood islands. Each of these genes was expressed in extraembryonic mesoderm, from which blood islands are derived. This VYS expression occurred in a defined temporal sequence: tal-1 and rbtn2 transcripts were detected earlier than the others, followed by GATA-2 and GATA-1, and then by βH1-globin. Transcripts for all of these genes were present in VYS mesoderm cell masses at the neural plate stage (E7.5), indicating commitment of these cells to the erythroid lineage before the appearance of morphologically recognizable erythroblasts. By early somite stages (E8.5), GATA-2 mRNA expression is downregulated in VYS blood islands as terminal primitive erythroid differentiation proceeds. We conclude that primitive mammalian erythropoiesis arises during gastrulation through the ordered temporal expression of tal-1, rbtn2, GATA2, and GATA-1 in a subset of extraembryonic mesoderm cells. During the stages analyzed, tal-1 and rbtn2 expression was also present in posterior embryonic mesoderm, while GATA-1 and GATA-2 expression was evident in extraembryonic tissues of ectodermal origin.
Cell-Cycle–Dependent Regulation of Erythropoietin Receptor Gene
Authors: Komatsu Norio , Kirito Keita , Kashii Yoshifumi , Furukawa Yusuke , Kikuchi Jiro , Suwabe Naruyoshi , Yamamoto Masayuki , Miura Yasusada 
Published: October 2019

AbstractTo understand the regulatory mechanism of erythropoietin (EPO) receptor (EPOR) gene expression, the effect of EPO on the steady-state level of EPOR mRNA was examined using the human EPO-dependent cell line UT-7 as a model system. We found that the treatment of UT-7 cells with EPO resulted in a transient decrease of the EPOR mRNA level. This transient downregulation was also induced by stimulation with granulocyte-macrophage colony-stimulating factor (GM-CSF ), another stimulator of UT-7 cell growth. These results raised the possibility that EPOR gene expression is in part related to cell growth. Moreover, it was found that EPO-induced downregulation of EPOR mRNA level was preceded by a transient downregulation of GATA-1 mRNA. To examine the relationship between the expression of EPOR, GATA-1, and GATA-2 mRNA levels and the cell cycle, logarithmically growing UT-7 cells were centrifugically fractionated according to the cell-cycle phase. Both EPOR and GATA-1 mRNA levels, but not the GATA-2 mRNA level, concomitantly decreased at the G0/G1 phase and increased at the S and G2/M phases. An electrophoretic mobility shift assay (EMSA) showed that in EPO-stimulated UT-7 cells, the dynamic changes in EPOR gene expression paralleled the GATA-1 DNA-binding activity to the oligonucleotide probe containing a GATA-binding site located at the promoter region of the EPOR gene. These findings suggest that the regulation of EPOR mRNA level is mainly associated with GATA-1 gene expression in UT-7 cells undergoing proliferation, and that these serial events are under the control of, or related to, the cell cycle.
Host F1 Mice Pretreated With Granulocyte Colony-Stimulating Factor Accept Parental Bone Marrow Grafts in Hybrid Resistance System
Authors: Iizuka Koho , Kaneko Hideo , Yamada Taketo , Kimura Hiromitsu , Kokai Yasuo , Fujimoto Junichiro 
Published: October 2019

AbstractIn the setting of hybrid resistance, parental C57BL/6 bone marrow (BM) grafts are vigorously rejected by lethally irradiated (C57BL/6xDBA/2) F1 mice. However, F1 mice pretreated by continuous administration of granulocyte colony-stimulating factor (G-CSF ) with a miniosmotic pump before BM grafting developed day-8 splenic colonies of donor origin. This inhibitory effect on rejection was reversible because F1 mice regained the capacity to reject parental BM when the pump ceased functioning. The appearance of only a small number of colonies with the administration of G-CSF soon after BM grafting suggested the importance in producing this inhibitory effect of pre-exposure of host mice to G-CSF. Because G-CSF administration with a syngeneic combination did not influence the number of colonies, an altered distribution of grafted precursors was unlikely. The absence of a reduction in the number of NK1.1-positive cells in G-CSF–treated mice suggested functional impairment of natural killer cells, major effectors in hybrid resistance, but further study is necessary to elucidate the mechanism underlying this phenomenon. However, our results indicate the importance of G-CSF as a regulator in a certain type of immune response and raise the possibility of clinical application in transplantation medicine.
Leptin Stimulates Fetal and Adult Erythroid and Myeloid Development
Authors: Mikhail Adel A. , Beck Eric X , Shafer Alan , Barut Bruce , Smith Gbur Julie , Zupancic Thomas J. , Schweitzer Anthony C. , Cioffi Joseph A. , Lacaud Georges , Ouyang Bin , Keller Gordon , Snodgrass H. Ralph 
Published: October 2019

AbstractThe ob gene product, leptin, has been shown in several studies to be involved in weight control and recombinant leptin recently has entered clinical trials to treat obesity. The leptin receptor (OB-R/B219) is expressed in a variety of protein isoforms not only in the central nervous system, but also in reproductive, and hematopoietic tissues. We reported recently that the OB-R/B219 was associated with a variety of hematopoietic lineages as well as the small fraction of cells containing the long-term reconstituting hematopoietic stem cells. Herein we report that leptin significantly stimulates the proliferation and differentiation of yolk sac cells and fetal liver cells and stimulates directly hematopoietic precursors. Leptin alone can increase the number of macrophage and granulocyte colonies, and leptin plus erythropoietin act synergistically to increase erythroid development. These data show that leptin has a significant, direct effect on early hematopoietic development and can stimulate the differentiation of lineage-restricted precursors of the erythrocytic and myelopoietic lineages. These observations along with a recent report strongly support our previous hypothesis that leptin has an unanticipated important role in hematopoietic and immune system development.
Mouse Strain-Dependent Changes in Frequency and Proliferation of Hematopoietic Stem Cells During Aging: Correlation Between Lifespan and Cycling Activity
Authors: de Haan Gerald , Nijhof Willem , Van Zant Gary 
Published: October 2019

AbstractWe have quantified the frequency and proliferation of five subsets of primitive hematopoietic cells, using the cobblestone area forming cell (CAFC) assay, in marrow of five strains of mice with lifespans ranging from about 500 to 800 days. Stem cell characteristics were determined in young (6 weeks) and old (12 months) mice. We report striking effects of both intrinsic strain lifespan and individual mouse age on stem cell populations. First, the relative and absolute numbers of the most primitive stem cell subsets was threefold to fourfold higher in old than in young mice. Second, a considerable strain-to-strain variation in the number of primitive cells was observed: when absolute frequencies were calculated, there was a trend for longer lifespan to be correlated with a larger stem cell pool. Third, stem cells from old mice had a far lower cycling activity than cells from young mice. However, this was highly strain dependent: short-lived C3H/He and CBA/J mice showed a stronger reduction in cycling activity during aging than long-lived C57BL/6 mice. Finally, a significant negative correlation was demonstrated in young mice between maximal lifespan and proliferative activity. These data show that aging has a major impact on the frequency and cell-cycle kinetics of primitive hematopoietic cell compartments. In addition, the observation that cycling activity of stem cells is related to the maximal lifespan of the mouse strain may open ways to identify the genetic mechanisms of both strain- and age-dependent variation in the structure of primitive hematopoietic cell compartments.
Loss of Inducible Virus in CD45RA Naive Cells After Human Immunodeficiency Virus-1 Entry Accounts for Preferential Viral Replication in CD45RO Memory Cells
Authors: Woods Toni C. , Roberts Beverly D. , Butera Salvatore T. , Folks Thomas M. 
Published: October 2019

AbstractControversy exists concerning the preferential infection and replication of human immunodeficiency virus-1 (HIV-1) within naive (CD45RA+) and memory (CD45RO+) subsets of CD4+ lymphocytes. To explore the susceptibility of these subsets to HIV-1 infection, we purified CD45RA+/CD4+ (RA) and CD45RO+/CD4+ (RO) cells from normal donors and subjected them to a novel monokine activation culture scheme. Following HIV-1 infection and interleukin-2 (IL-2) induction, viral production measured on day 13 was 19-fold greater in RO cultures compared with RA cultures. IL-2–stimulated proliferation in uninfected control cultures was equivalent. To explore the mechanisms by which RA cells were reduced in viral production capacity, RA and RO cells were exposed to HIV-1 followed by treatment with trypsin, and then phytohemagglutinin antigen (PHA)-stimulated at days 4, 7, and 10 postinfection. HIV-1 production in day 4 postinfection RA and RO cultures was analogous, indicating that viral fusion and entry had occurred in both cell types. However, whereas similarly treated day 7 and 10 postinfection RO cultures produced virus, HIV-1 was markedly reduced or lost in the corresponding RA cultures. These results suggest that a temporally labile postfusion HIV-1 complex exists in unstimulated RA cells that requires cellular activation signals beyond that provided by IL-2 alone for productive infection.
CD34+ Cells in the Blood of Patients With Multiple Myeloma Express CD19 and IgH mRNA and Have Patient-Specific IgH VDJ Gene Rearrangements
Authors: Szczepek Agnieszka J. , Bergsagel P.L. , Axelsson Linus , Brown Christopher B. , Belch Andrew R. , Pilarski Linda M. 
Published: October 2019

AbstractPeripheral blood mononuclear cells (PBMC) from patients with multiple myeloma (MM) are here shown to include 23% ± 2% of CD34+ cells, the majority of which coexpress CD19, as identified by a panel of 17 anti-CD34 antibodies. The expression of CD34 mRNA by sorted CD34+ PBMC from MM was confirmed by in situ reverse transcriptase-polymerase chain reaction (RT-PCR) with CD34-specific primers. The majority of CD34+ MM PBMC were CD19+ cells that expressed mRNA for CD19 and for rearranged IgH as identified with consensus IgH VDJ primers, as well as having cytoplasmic Ig, definitively identifying them as B cells, in absolute numbers of 0.06 to 0.69 × 109/L of blood. CD34 is largely absent from normal B cells. To determine the clonal relationship of CD34+ B cells to autologous MM plasma cells, IgH VDJ DNA rearrangements of sorted CD34+ MM blood B cells were amplified by nested PCR using consensus primers followed by Southern blotting with allele-specific oligonucleotides for 7 MM patients, and clonotypic IgH mRNA expression was assessed for 4 MM patients using quantitative patient-specific in situ RT-PCR. For 9 of 11 myeloma patients tested, CD34+ blood B cells included IgH gene rearrangements or expressed IgH mRNA identical to that of autologous bone marrow plasma cells. For 4 of 4 MM patients, 74% to 94% of individual sorted CD34+19+ B cells expressed clonotypic IgH mRNA, as detected by in situ RT-PCR with patient-specific primers. Clonotypic IgH VDJ sequences were absent from B cells of unrelated MM patients and of normal donors. Clonotypic CD34+ B cells were detected before, during, and after treatment, and during relapse. Our results indicate a clonal relationship between CD34+ MM B cells and malignant plasma cells. We speculate that CD34 may play an important role in the biology of myeloma by facilitating extravasation from blood and thus spread of myeloma through the skeletal system.
Mice Bearing a Targeted Interruption of the Homeobox Gene HOXA9 Have Defects in Myeloid, Erythroid, and Lymphoid Hematopoiesis
Authors: Lawrence H. Jeffrey , Helgason Cheryl D. , Sauvageau Guy , Fong Stephen , Izon David J. , Humphries R. Keith , Largman Corey 
Published: October 2019

AbstractSeveral homeobox genes of the HOXA and HOXB clusters are expressed in primitive blood cells, suggesting a role for HOX genes in normal hematopoiesis. The HOXA9 gene is expressed in CD34+ marrow cells and in developing lymphocytes. We examined blood-forming organs of mice homozygous for an interrupted HOXA9 allele to determine if loss of HOX gene function is deleterious to hematopoiesis. HOXA9−/− mice have approximately 30% to 40% reductions in total leukocytes and lymphocytes (P < .001) and a blunted granulocytic response to granulocyte colony-stimulating factor (G-CSF ). Homozygous mice have significantly smaller spleens and thymuses. Myeloid/erythroid and pre-B progenitors in the marrow are significantly reduced, but no significant decreases are noted in mixed colonies, day 12 colony-forming units-spleen (CFU-S), or long-term culture–initiating cells (LTC-IC), suggesting little or no perturbation in earlier progenitors. Heterozygous animals display no hematopoietic defects. The abnormalities in leukocyte production are transplantable, indicating that the defect resides in the hematopoietic cells. These studies demonstrate a physiologic role for a HOX gene in blood cell differentiation, with the greatest apparent influence of HOXA9 at the level of the committed progenitor.
Treatment of Refractory Hodgkin's Disease With an Anti-CD16/CD30 Bispecific Antibody
Authors: Hartmann Frank , Renner Christoph , Jung Wolfram , Deisting Christina , Juwana Marietta , Eichentopf Bertram , Kloft Michael , Pfreundschuh Michael 
Published: October 2019

Abstract
Fifteen patients with refractory Hodgkin's disease were treated in a phase I/II trial with the natural killer (NK)-cell–activating bispecific monoclonal antibody HRS-3/A9, which is directed against the Fcγ-receptor III (CD16 antigen) and the Hodgkin's-associated CD30 antigen, respectively. Median counts of NK cells and of all lymphocyte subsets were considerably decreased in the patients before therapy. HRS-3/A9 was administered 4 times every 3 to 4 days, starting with 1 mg/m2. The treatment was well tolerated, and the maximum tolerated dose was not reached at 64 mg/m2, the highest dose administered because of the limited amounts of HRS-3/A9 available. Side effects were rare and consisted of fever, pain in involved lymph nodes, and a maculopapulous rash. A total of 9 patients developed human antimouse Ig antibodies, and 4 patients developed an allergic reaction after attempted retreatment. A total of 1 complete and 1 partial remission (lasting 16 and 3 months, respectively), 3 minor responses (1 to 11+ months), and 1 mixed response were achieved. There was no clear-cut dose-side effect or dose-response correlation. Our results encourage further clinical trials with this novel immunotherapeutic approach and emphasize the necessity to reduce the immunogenicity of the murine bispecific antibodies.
Altered Formation of Topotecan-Stabilized Topoisomerase I-DNA Adducts in Human Leukemia Cells
Authors: Kaufmann Scott H. , Svingen Phyllis A. , Gore Steven D. , Armstrong Deborah K. , Cheng Yung-Chi , Rowinsky Eric K. 
Published: October 2019

AbstractTopotecan (TPT) is a topoisomerase I (topo I) poison that has shown promising antineoplastic activity in solid tumors and acute leukemia. In the present study, a band depletion assay was used to evaluate the ability of TPT to stabilize topo I-DNA adducts in human leukemia cell lines and in clinical leukemia samples ex vivo. This assay showed that 50% of the cellular topo I in HL-60 human myelomonocytic leukemia cells became covalently bound to DNA at an extracellular TPT concentration of 4 μmol/L. In contrast, in 13 clinical specimens of human leukemia harvested before treatment of patients with TPT, the TPT concentration required to stabilize 50% of the cellular topo I in topo I-DNA complexes ranged from 3 to greater than 100 μmol/L (median, 30 μmol/L). Flow microfluorimetry showed that cellular TPT accumulation varied over only a twofold range and failed to provide evidence for transport-mediated resistance in the clinical samples. These observations raise the possibility that formation of topo I-DNA adducts is diminished in many specimens of refractory/relapsed acute leukemia by a mechanism that might alter topo I sensitivity to TPT.
Endogenous Tumor Necrosis Factor as a Predictor of Doxorubicin Sensitivity in Leukemic Patients
Authors: Kobayashi Daisuke , Watanabe Naoki , Yamauchi Naofumi , Tsuji Naoki , Sato Tsutomu , Niitsu Yoshiro 
Published: October 2019

AbstractWe have previously reported that intracellular tumor necrosis factor (enTNF ) is responsible for resistance, in established cell lines to doxorubicin (DOX), exogenous TNF, and heat stress by inducing manganous superoxide dismutase (MnSOD), thereby scavenging reactive oxygen free radicals. Leukemic cells from 19 patients (6 acute lymphoblastic leukemia, 13 acute myeloid leukemia) were examined for their sensitivity to DOX and TNF in relation to their enTNF expression and MnSOD activity. Sensitivity to DOX and the expression of enTNF or MnSOD activity were inversely correlated. In a case with acquired resistance to chemotherapy which included DOX, enTNF expression and MnSOD activity were increased. Furthermore, in 14 cases treated with a regimen including an anthracycline, 4 cases that failed to respond to chemotherapy showed relatively high amounts of enTNF expression. KG-1 (human acute myelogenous leukemia) cells transfected with a nonsecretory-type TNF expression vector (pTNFΔpro) showed resistance to DOX. A significant increase in MnSOD levels was also noted in the transfectants. TNF antisense cDNA was transfected into isolated leukemic cells from five patients. Sensitivity of the antisense transfectants to DOX was increased, approximately 1.4- to 2.5-fold. These results suggest that enTNF acts as a resistance factor against DOX in leukemia, and that enTNF may be useful as a predictor of DOX sensitivity in leukemia.