bluuebunny/crossref_metadata_2025_split huggingface dataset:
Functional Correction of Fanconi Anemia Group A Hematopoietic Cells by Retroviral Gene Transfer
Authors: Fu Kai-Ling , Foe Jerome R. Lo Ten , Joenje Hans , Rao Kathleen W. , Liu Johnson M. , Walsh Christopher E. 
Published: October 2019

AbstractFanconi anemia (FA) is an autosomal recessive genetic disorder characterized by a variety of physical anomalies, bone marrow failure, and an increased risk for malignancy. FA cells exhibit chromosomal instability and are hypersensitive to DNA cross-linking agents such as mitomycin C (MMC). FA is a clinically heterogeneous disorder and can be functionally divided into at least five different complementation groups (A-E). We previously described the use of a retroviral vector expressing the FAC cDNA in the complementation of mutant hematopoietic cells from FA-C patients. This vector is currently being tested in a clinical trial of ex vivo hematopoietic progenitor cell transduction. The FA-A group accounts for over 65% of all FA cases, and the FAA cDNA was recently identified by both expression and positional cloning techniques. We report here the transduction and phenotypic correction of lymphoblastoid cell lines from four unrelated FA-A patients, using two amphotropic FAA retroviral vectors. Expression of the FAA transgene was adequate to normalize cell growth, cell-cycle kinetics, and chromosomal breakage in the presence of MMC. We then analyzed the effect of retroviral vector transduction on hematopoietic progenitor cell growth. After FAA transduction of mutant progenitor cells, either colony number or colony size increased in the presence of MMC. In addition, FAA but not FAC retroviral transduction markedly improved colony growth of progenitor cells derived from an unclassified FA patient. FAA retroviral vectors should be useful for both complementation studies and clinical trials of gene transduction.
Transcriptional Behavior of LCR Enhancer Elements Integrated at the Same Chromosomal Locus by Recombinase-Mediated Cassette Exchange
Authors: Bouhassira Eric E. , Westerman Karen , Leboulch Philippe 
Published: October 2019

Abstract
Efficient integration of transgenes at preselected chromosomal locations was achieved in mammalian cells by recombinase-mediated-cassette-exchange (RMCE), a novel procedure that makes use of the CRE recombinase together with Lox sites bearing different spacer regions. We have applied RMCE to the study of the human β-globin gene Locus Control Region by integrating at the same genetic locus in MEL cells, a LacZ gene driven by the human β-globin promoter linked to HS2 and HS3 alone or in combination with HS4. Expression studies at the cell population level and in individual cells before and after induction of differentiation with hemin or DMSO show that the presence of these enhancers is associated with variegated patterns of expression. We were able to show that the LCR fragments tested act by controlling both the probability of expression and the rate of transcription of the linked β-globin promoter. Both of these factors were also dependent on the state of differentiation of the MELc and on the presence of a second transcription unit located in cis. The ability to manipulate by RMCE constructs integrated into chromosomes should help in the creation of complex, rationally designed, artificial genetic loci.
Interleukin-10 Counterregulates Proinflammatory Cytokine-Induced Inhibition of Neutrophil Apoptosis During Severe Sepsis
Authors: Keel Marius , Ungethüm Udo , Steckholzer Ursula , Niederer Eva , Hartung Thomas , Trentz Otmar , Ertel Wolfgang 
Published: October 2019

AbstractNeutrophils play a key role in the pathophysiology of septic multiple organ dysfunction syndrome (MODS) through excessive release of toxic granule components and reactive oxygen metabolites with consequent tissue destruction. The increase of senescent neutrophils during sepsis indicates a potential breakdown of autoregulatory mechanisms including apoptotic processes to remove activated neutrophils from inflammatory sites. Therefore, neutrophil apoptosis of patients with severe sepsis and its regulatory mechanisms were investigated. Spontaneous neutrophil apoptosis from patients with severe sepsis was significantly reduced in comparison to healthy individuals. Cytokines detected in the circulation during sepsis (tumor necrosis factor-α [TNF-α], interferon-γ [IFN-γ], granulocyte colony-stimulating factor [G-CSF], granulocyte-macrophage colony-stimulating factor [GM-CSF]) inhibited neutrophil apoptosis in both groups, though the effect was more distinct in neutrophils from healthy humans. Addition of lipopolysaccharide (LPS) to neutrophils from healthy humans markedly (P < .05) reduced apoptosis which was partially restored through addition of anti–TNF-antibody. Interleukin-10 (IL-10) counteracted (P < .05) inhibition of neutrophil apoptosis induced by LPS, recombinant human (rh) TNF-α, rhIFN-γ, rhG-CSF, and rhGM-CSF, whereas rhIL-4 or rhIL-13 were ineffective. Reduced neutrophil apoptosis during sepsis was concomitant with increased tyrosine phosphorylation, while IL-10 markedly inhibited tyrosine phosphorylation in LPS-stimulated neutrophils. These results identify proinflammatory cytokines and IL-10 as strong regulators of spontaneous neutrophil apoptosis during sepsis. Inhibition as well as acceleration of neutrophil apoptosis seems to be associated with alterations of signal transduction pathways.
Treatment of Polycythemia Vera: The Use of Hydroxyurea and Pipobroman in 292 Patients Under the Age of 65 Years
Authors: Najean Yves , Rain Jean-Didier 
Published: October 2019

Abstract
Nonradiomimetic drugs, hydroxyurea (HU) and pipobroman (Pi), were administred to relatively young subjects with polycythemia vera (PV) in an attempt to decrease the leukemogenic risk observed in patients treated with 32P. Clinical safety, hematological efficacy, risk of carcinoma or leukemia, and frequency of progression to myelofibrosis have not yet been defined in long-term studies, and no comparative studies of HU and Pi have been conducted. Since 1980, 292 patients with PV diagnosed before the age of 65 years were randomized to receive treatment with HU (25 mg/kg/d, followed by low-dose maintenance) or Pi (1.2 mg/kg/d, followed by low-dose maintenance). Patients were followed until death or until May 1997. Drug tolerance was often poor; leg ulcers and buccal aphthous ulcers (with HU) and gastric pain and diarrhea (with Pi) sometimes required treatment change, mainly in the HU arm. Hematological stability, especially in terms of platelet count, was very often insufficient with HU (45% of cases), but the risk of thrombo-embolic event was similar in both arms. Actuarial survival was similar in the two arms and shorter than that of the reference population. The risk of leukemia was approximately 10% at the 13th year, with no significant difference between the two arms. The risk of carcinoma (when excluding the skin cancers) was similar in both groups. There was a high risk of progression to myelofibrosis in the patients treated by HU, which was significantly higher than with Pi.
Effects of Aerobic Exercise on the Physical Performance and Incidence of Treatment-Related Complications After High-Dose Chemotherapy
Authors: Dimeo Fernando , Fetscher Sebastian , Lange Winand , Mertelsmann Roland , Keul Joseph 
Published: October 2019

Abstract
Loss of physical performance is a universal problem of cancer patients undergoing chemotherapy. We postulated that this impairment can be partially prevented by aerobic exercise. In a randomized study, 33 cancer patients receiving high-dose chemotherapy followed by autologous peripheral blood stem cell transplantation (training group, T) performed an exercise program consisting of biking on an ergometer in the supine position after an interval-training pattern for 30 minutes daily during hospitalization. Patients in the control group (C, n = 37) did not train. Maximal physical performance was assessed with a treadmill test by admission and discharge. Physical performance of the two groups was not different on admission. The decrement in performance during hospitalization was 27% greater in the control group than in the training group (P = .05); this resulted in a significantly higher maximal physical performance at discharge in the trained patients (P = .04). Duration of neutropenia (P = .01) and thrombopenia (P = .06), severity of diarrhea (P = .04), severity of pain (P = .01), and duration of hospitalization (P = .03) were reduced in the training group. We conclude that aerobic exercise can be safely carried out immediately after high-dose chemotherapy and can partially prevent loss of physical performance. Based on the potential significance of the observed outcomes, further studies are warranted to confirm our results.
Leptin Stimulates the Proliferation of Murine Myelocytic and Primitive Hematopoietic Progenitor Cells
Authors: Umemoto Yumi , Tsuji Kohichiro , Yang Feng-Chun , Ebihara Yasuhiro , Kaneko Azusa , Furukawa Susumu , Nakahata Tatsutoshi 
Published: October 2019

AbstractLeptin, the product of obese gene, was originally identified as a factor regulating body-weight homeostasis and energy balance. The present study has shown that leptin acts on murine hematopoiesis in vitro. In the culture of bone marrow cells (BMC) of normal mice, leptin induced only granulocyte-macrophage (GM) colony formation in a dose-dependent manner, and no other types of colonies were detected even in the presence of erythropoietin (Epo). Leptin also induced GM colony formation from BMC of db/db mutant mice whose leptin receptors were incomplete, but the responsiveness was significantly reduced. The effect of leptin on GM colony formation from BMC of normal mice was also observed in serum-free culture, and comparable with that of GM-colony–stimulating factor (CSF ). Although leptin alone supported few colonies from BMC of 5-fluorouracil (5-FU)–treated mice in serum-free culture, remarkable synergism between leptin and stem cell factor (SCF ) was obtained in the colony formation. The addition of leptin to SCF enhanced the SCF-dependent GM colony formation and induced the generation of a number of multilineage colonies in the presence of Epo. When lineage (Lin)−Sca-1+ cells sorted from BMC of 5-FU–treated mice were incubated in serum-free culture, leptin synergized with SCF in the formation of blast cell colonies, which efficiently produced secondary colonies including a large proportion of multilineage colonies in the replating experiment. In serum-free cultures of clone-sorted Lin−c-Kit+Sca-1+ and Lin−c-Kit+Sca-1− cells, although synergism of leptin and SCF was observed in the colony formation from both cells, leptin alone induced the colony formation from Lin−c-Kit+Sca-1−, but not Lin−c-Kit+Sca-1+ cells. These results have shown that leptin stimulates the proliferation of murine myelocytic progenitor cells and synergizes with SCF in the proliferation of primitive hematopoietic progenitors in vitro.
Mechanisms of Stem Cell Factor and Erythropoietin Proliferative Co-signaling in FDC2-ER Cells
Authors: Joneja Bhavana , Chen Hong-Chi , Seshasayee Dhaya , Wrentmore Amy L. , Wojchowski Don M. 
Published: October 2019

AbstractStudies of hematopoietic progenitor cell development in vivo, ex vivo, and in factor-dependent cell lines have shown that c-kit promotes proliferation through synergistic effects with at least certain type 1 cytokine receptors, including the erythropoietin (Epo) receptor. Presently, c-kit is shown to efficiently support both mitogenesis and survival in the FDCP1 cell subline, FDC2. In this system, mitogenic synergy with c-kit was observed for ectopically expressed wild-type Epo receptors (wt-ER), an epidermal growth factor (EGF) receptor/Epo receptor chimera, and a highly truncated Epo receptor construct ER-Bx1. Thus, the Epo receptor cytoplasmic box 1 subdomain appears, at least in part, to mediate mitogenic synergy with c-kit. In studies of potential effectors of this response, Jak2 tyrosine phosphorylation was shown to be induced by Epo, but not by stem cell factor (SCF). In addition and in contrast to signaling in Mo7e and BM6 cell lines, in FDC2-ER cells SCF and Epo each were shown to rapidly activate Pim 1 gene expression. Recently, roles also have been suggested for the nuclear trans-factor GATA-1 in regulating progenitor cell proliferation. In FDC2-ER cells, the ectopic expression of GATA-1 had no detectable effect on Epo inhibition of apoptosis. However, GATA-1 expression did result in a selective and marked inhibition in mitogenic responsiveness to SCF and to a decrease in c-kit transcript expression. These studies of SCF and Epo signaling in FDC2–wt-ER cells serve to functionally map the ERB1 region as a c-kit–interactive domain, suggest that Pim1 might contribute to SCF and Epo mitogenic synergy and support the notion that SCF and Epo may act in opposing ways during red cell differentiation.
Sensitization of Hematopoietic Stem and Progenitor Cells to Trimetrexate Using Nucleoside Transport Inhibitors
Authors: Allay James A. , Spencer H. Trent , Wilkinson Sarah L. , Belt Judith A. , Blakley Raymond L. , Sorrentino Brian P. 
Published: October 2019

Abstract
Antifolates such as methotrexate (MTX) and trimetrexate (TMTX) are widely used in the treatment of cancer and nonmalignant disorders. Transient, yet sometimes severe myelosuppression is an important limitation to the use of these drugs. It has previously been shown that clonogenic myeloid progenitors and colony-forming units-spleen are resistant to antifolates, suggesting that myelotoxicity occurs late in hematopoietic development. The goal of this study was to define the mechanisms by which primitive hematopoietic cells resist the toxic effects of antifolate drugs. To test the hypothesis that myeloid progenitors may salvage extracellular nucleotide precursors to resist TMTX toxicity, a defined liquid culture system was developed to measure TMTX toxicity in expanding progenitor populations. These in vitro experiments showed that both human and murine progenitors can resist TMTX toxicity by importing thymidine and hypoxanthine from the serum. As predicted from these findings, several drugs that block thymidine transport sensitized progenitors to TMTX in vitro, although to differing degrees. These nucleoside transport inhibitors were used to test whether progenitors and hematopoietic stem cells (HSCs) could be sensitized to TMTX in vivo. Treatment of mice with TMTX and nitrobenzylmercaptopurineriboside phosphate (NBMPR-P), a potent transport inhibitor, caused significant depletions of clonogenic progenitors within the bone marrow (20-fold) and spleen (6-fold). Furthermore, NBMPR-P administration dramatically sensitized HSCs to TMTX, with dual-treated mice showing a greater than 90% reduction in bone marrow repopulating activity. These studies demonstrate that both myeloid progenitor cells and HSCs resist TMTX by using nucleotide salvage mechanisms and that these pathways can be pharmacologically blocked in vivo using nucleoside transport inhibitors. These results have important implications regarding the use of transport inhibitors for cancer therapy and for using variants of dihydrofolate reductase for in vivo selection of genetically modified HSCs.
Association of Tissue Factor Pathway Inhibitor With Human Umbilical Vein Endothelial Cells
Authors: Hansen John-Bjarne , Olsen Randi , Webster Paul 
Published: October 2019

AbstractTissue factor pathway inhibitor (TFPI) is a serine protease inhibitor of the extrinsic coagulation system, synthesized in endothelial cells, which has recently been shown to play an important role in the regulation of activated coagulation factors at the endothelial cell surface. In the present study we investigated the subcellular localization and metabolism of TFPI in human umbilical vein endothelial cells (HUVEC). Immunocytochemical labeling of HUVEC with anti-TFPI showed specific labeling associated with the cell surface and with many intracellular organelles including the Golgi complex. Further characterization of these organelles was performed by colocalizing the anti-TFPI with 3-(2,4-dinitroanilino)′-amino-N-methyldipropylamine (DAMP; to demonstrate low pH), mannose phosphate receptor (endosomes), and LAMP 1 (late endocytic compartments). TFPI also colocalized with antibodies to the human transferrin receptor, a marker for early endocytic, recycling compartment. Endogenous TFPI colocalized with biotin in intracellular vesicles during endocytosis after biotinylation of the cell surface, which indicated that TFPI was being co-internalized with the surface biotin. The binding of exogenously added 125I-TFPI increased linearly to HUVEC over the concentration range of 0 to 32 nmol/L without saturation, the binding was not affected by up to a thousand-fold molar excess of unlabeled TFPI, and heparin inhibited the binding dose dependently. An intact C-terminal domain was important for the interaction between TFPI and the cell surface of HUVEC, because less than 10% of a C-terminal truncated form of TFPI (TFPI1-161 ) was bound after addition of equimolar concentrations of full-length TFPI. Exogenously added 125I-TFPI was not degraded in HUVEC during 4 hours at 37°C. The presence of TFPI in endocytic and recycling compartments support the hypothesis that endogenous, membrane-anchored TFPI could be internalized for subsequent recycling back to the cell surface.
Thrombin Stimulation of Platelets Induces Plasminogen Activation Mediated by Endogenous Urokinase-Type Plasminogen Activator
Authors: Lenich Catherine , Liu Jian-Ning , Gurewich Victor 
Published: October 2019

AbstractGene knockout mice studies indicate that urokinase-type plasminogen activator (u-PA) is importantly involved in fibrinolysis, but its physiologic mechanism of action remains poorly understood. We postulated that platelets may be involved in this mechanism, as they carry a novel receptor for u-PA and a portion of the single-chain u-PA (scu-PA) intrinsic to blood is tightly associated with platelets. Therefore, plasminogen activation by platelet-associated u-PA was studied. When washed platelets were incubated with plasminogen, no plasmin was generated as detected by plasmin synthetic substrate (S2403) hydrolysis; however, after the addition of thrombin, but not other agonists, platelet-dependent plasminogen activation occurred. Plasminogen activation was surface-related, being inhibited by blocking platelet fibrinogen receptors or by preventing plasminogen binding to the thrombin-activated platelet surface. U-PA was identified as the only plasminogen activator responsible and enrichment of platelets with exogenous scu-PA significantly augmented plasminogen activation. These findings appeared paradoxical because thrombin inactivates scu-PA. Indeed, zymograms showed inactivation of scu-PA during the first hour of incubation with even the lowest dose of thrombin used (1 u/mL). However, this was followed by a thrombin dose-dependent (1 to 10 u/mL) partial return of u-PA activity. Reactivation of u-PA was not due to the direct action of thrombin, but required platelets and was found to be related to a platelet lysosomal thiol protease, consistent with cathepsin C. In conclusion, a new pathway of plasminogen activation by platelet-associated endogenous or exogenous scu-PA was demonstrated, which is specifically triggered by thrombin activation of platelets. These findings may help explain u-PA–mediated physiological fibrinolysis and have implications for therapeutic thrombolysis with scu-PA.
Fibrin Regulation of Interleukin-8 Gene Expression in Human Vascular Endothelial Cells
Authors: Qi Jiafan , Goralnick Sandra , Kreutzer Donald L. 
Published: October 2019

AbstractRecent studies in our laboratory, as well as others, have suggested that fibrin can regulate cell function in vitro and likely control inflammation in vivo by acting as a potent cell activator. This has led us to hypothesize that during tissue and vascular injury, fibrin can enhance leukocyte recruitment by inducing vascular endothelial cell expression of leukocyte chemotactic factors. To begin to test this hypothesis, we developed an in vitro model of in situ fibrin polymerization on human umbilical vein endothelial cell culture (HUVEC) and determined the ability of fibrin to induce HUVEC expression of the potent leukocyte chemotactic factor interleukin-8 (IL-8). Our initial studies showed that fibrin induced IL-8 expression in a time- and dose-dependent fashion. Fibrin-induced IL-8 expression in HUVEC could be seen as early as 2 hours post-fibrin stimulation. Additionally, fibrin concentrations as low as 30 μg/mL stimulated a detectable level of IL-8 antigen expression from HUVEC. We also showed that this fibrin induced IL-8 had the identical molecular weight and similar antigenic identity as recombinant and monocyte derived IL-8. Northern blot analysis showed that the IL-8 antigen increase seen in fibrin treated HUVEC was due to fibrin induced elevation of steady state mRNA expression in HUVEC. These data clearly support our hypothesis that fibrin is a potent vascular endothelial cell (VEC) activator that can directly contribute to leukocyte recruitment and activation by inducing leukocyte chemotactic factor expression from VEC.
Fas-Independent Apoptosis of Activated T Cells Induced by Antibodies to the HLA Class I α1 Domain
Authors: Genestier Laurent , Paillot Romain , Bonnefoy-Berard Nathalie , Meffre Geneviéve , Flacher Monique , Fèvre David , Liu Yong Jun , Le Bouteiller Philippe , Waldmann Herman , Engelhard Victor H. , Banchereau Jacques , Revillard Jean Pierre 
Published: October 2019

AbstractIn addition to their major function in antigen presentation and natural killer cell activity regulation, HLA class I molecules may modulate T-cell activation and proliferation. Monoclonal antibodies (MoAbs) that recognize distinct epitopes of HLA class I molecules were reported to interfere with T-cell proliferation. We show here that two MoAbs (mouse MoAb90 and rat YTH862) that bind to an epitope of the α1 domain of HLA class I heavy chain induce apoptotic cell death of activated, but not resting, peripheral T lymphocytes. Other reference anti-HLA class I antibodies specific for distinct epitopes of the α1 (B9.12.1), α2 (W6/32), or α3 (TP25.99) domains of the heavy chain decreased T-cell proliferation but had little or no apoptotic effect. Apoptosis shown by DNA fragmentation, phosphatidylserine externalization, and decrease of mitochondrial transmembrane potential was observed whatever the type of T-cell activator. Apoptosis did not result from Fas/Fas-L interaction and distinct though partly overlapping populations of activated T cells were susceptible to Fas– and HLA class I–mediated apoptosis, respectively. Induction of apoptosis did not require HLA class I cross-linking inasmuch as it could be observed with monovalent Fab′ fragments. The data indicate that MoAb90 and YTH862 directed against the α1 domain of HLA class I trigger apoptosis of activated T lymphocytes by a pathway which does not involve Fas-ligand.