bluuebunny/crossref_metadata_2025_split huggingface dataset:
Collection, Tumor Contamination, and Engraftment Kinetics of Highly Purified Hematopoietic Progenitor Cells to Support High Dose Therapy in Multiple Myeloma
Authors: Tricot G. , Gazitt Y. , Leemhuis T. , Jagannath S. , Desikan K.R. , Siegel D. , Fassas A. , Tindle S. , Nelson J. , Juttner C. , Tsukamoto A. , Hallagan J. , Atkinson K. , Reading C. , Hoffman R. , Barlogie B. 
Published: October 2019

Abstract
Unfractionated peripheral blood stem cell (PBSC) grafts contain measurable quantities of myeloma cells and are therefore a potential source of relapse posttransplantation. In contrast, fluorescence-activated cell sorting (FACS)-sorted CD34+Thy1+ Lin− peripheral blood cells are substantially enriched for stem cell activity, yet contain virtually no clonal myeloma cells. A study was performed in patients with symptomatic myeloma, who had received 12 months or less of preceding standard chemotherapy, to evaluate the feasibility of large scale purification of primitive hematopoietic stem cells in order to study engraftment kinetics posttransplantation and the degree of tumor cell contamination of this cell population, based on polymerase chain reaction (PCR) analysis for the patient-specific complementarity-determining region III (CDR III). PBSC were mobilized with high dose cyclophosphamide and granulocyte-macrophage colony-stimulating factor (GM-CSF). A combination of elutriation and chemical lysis was used to deplete PBSC collections of monocytes, granulocytes, erythrocytes, and platelets. Subsequently, CD34+ Thy1+ Lin−progenitor cells were purified with high speed cell sorting. Of the 10 evaluable patients, nine met the required minimum criteria of ≥7.2 × 105 cells/kg to support tandem transplants. After high dose melphalan (200 mg/m2) eight engrafted successfully, although granulocyte (absolute neutrophil count [ANC] &gt;0.5 × 109/L, 16 days) and platelet recovery (platelets &gt; 50 × 109/L, 39 days) was substantially delayed when compared with unmanipulated PBSC grafts; one patient required infusion of a reserve graft because of lack of evidence of engraftment by day +28. Three patients proceeded to a second graft with high dose melphalan and total body irradiation; two required infusion of a reserve graft and both died of infectious complications; one showed delayed, but complete, engraftment after this myeloablative regimen. Two of the nine evaluable patients attained a clinical complete remission (CR). The grafts from three patients were tested for tumor contamination and contained no detectable clonal myeloma cells. Larger quantities of purified cells may be required to resolve the problem of delayed engraftment.
Regulation of p21(WAF1) Expression During Normal Myeloid Differentiation
Authors: Steinman Richard A. , Huang Jianping , Yaroslavskiy Beatrice , Goff Julie P. , Ball Edward D. , Nguyen Aline 
Published: October 2019

AbstractThe G1-phase cell-cycle inhibitor p21 has been proposed to mediate growth arrest during differentiation. Upregulation of p21 has been shown in multiple cell lines induced to differentiate; however, the mechanism of p21 induction during normal differentiation is largely unknown. In this report, we use normal hematopoietic precursor cells obtained from umbilical cord to model p21 regulation during differentiation. Myeloid maturation of CD34+ precursor cells is associated with a marked increase in p21 expression at the RNA and protein level. The upregulation of p21 transcripts during differentiation is associated with decreased binding to a highly conserved 44-bp fragment within the p21 promoter. This 44-bp regulatory element binds a novel modulator of p21 expression. It is of considerable interest that, although the binding activity is expressed in p53-negative as well as in p53-positive cells, the DNA sequence recognized by this protein overlaps a PuPuPuC(A/T)(T/A)GPyPyPy consensus sequence for p53.
A New Cytokine-Dependent Monoblastic Cell Line With t(9;11)(p22;q23) Differentiates to Macrophages With Macrophage Colony-Stimulating Factor (M-CSF) and to Osteoclast-Like Cells With M-CSF and Interleukin-4
Authors: Ikeda Takashi , Sasaki Kazunori , Ikeda Kazuma , Yamaoka Genji , Kawanishi Koichi , Kawachi Yasunori , Uchida Tatsumi , Takahara Jiro , Irino Shozo 
Published: October 2019

AbstractMonocytes/macrophages exert a series of important functions in vivo. To facilitate detailed investigation of their functional capacity and the mechanism leading to their differentiation, several cell lines have been established from primary material. We present here a new human monoblastic cell line, designated UG3. UG3 cells are characterized by the following features. (1) UG3 cells harbor the t(9;11)(p22;q23) translocation that results in fusion of the MLL and the AF9 genes and produce the corresponding AF9-MLL and MLL-AF9 fusion transcripts. (2) UG3 cells rely on the presence of exogenous growth factors for viability and proliferation, such as interleukin-3 (IL-3), granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF), or macrophage colony-stimulating factor (M-CSF). (3) When cultured in the presence of G-CSF, UG3 cells differentiate along the granulocytic lineage, as evidenced by segmentation of nuclei and positive staining for neutrophilic alkaline phosphatase and peroxidase. (4) When cultured in the presence of GM-CSF or M-CSF, UG3 cells differentiate into mature macrophages while preserving surface expression of CD14 and CD68 and also start to release cytokines into cell-culture supernatants. Under these culture conditions, UG3 cells also take up acetylated LDL. (5) When cultured in the presence of M-CSF and IL-4, UG3 cells differentiate into osteoclast-like multinucleated giant cells capable of bone resorption and display tartrate-resistant acid phosphatase (TRAP) activity. UG3 cells thus provide features to qualify them as a useful model to further investigate the mechanism underlying these processes and also to further elucidate the functional role of mature monocytes/macrophages or osteoclasts.
Transgenically Produced Human Antithrombin: Structural and Functional Comparison to Human Plasma–Derived Antithrombin
Authors: Edmunds Tim , Van Patten Scott M. , Pollock Julie , Hanson Eric , Bernasconi Richard , Higgins Elizabeth , Manavalan Partha , Ziomek Carol , Meade Harry , McPherson John M. , Cole Edward S. 
Published: October 2019

AbstractRecombinant human antithrombin (rhAT) produced in transgenic goat milk was purified to greater than 99%. The specific activity of the rhAT was identical to human plasma–derived AT (phAT) in an in vitro thrombin inhibition assay. However, rhAT had a fourfold higher affinity for heparin than phAT. The rhAT was analyzed and compared with phAT by reverse phase high-performance liquid chromatography, circular dichroism, fluorophore-assisted carbohydrate electrophoresis (FACE), amino acid sequence, and liquid chromatography/mass spectrography peptide mapping. Based on these analyses, rhAT was determined to be structurally identical to phAT except for differences in glycosylation. Oligomannose structures were found on the Asn 155 site of the transgenic protein, whereas only complex structures were observed on the plasma protein. RhAT contained a GalNAc for galactose substitution on some N-linked oligosaccharides, as well as a high degree of fucosylation. RhAT was less sialylated than phAT and contained both N-acetylneuraminic and N-glycolylneuraminic acid. We postulate that the increase in affinity for heparin found with rhAT resulted from the presence of oligomannose-type structures on the Asn 155 glycosylation site and differences in sialylation.
Follicular Lymphomas Contain a Clonally Linked But Phenotypically Distinct Neoplastic B-Cell Population in the Interfollicular Zone
Authors: Dogan Ahmet , Du Ming-Qing , Aiello Antonella , Diss Tim C. , Ye Hong-Tao , Pan Lang-Xing , Isaacson Peter G. 
Published: October 2019

Abstract
Follicular lymphomas are thought to arise from the follicle center B cells and are characterized by follicular structures that recapitulate many features of normal secondary lymphoid follicles. The neoplastic B cells of follicular lymphoma reside not only in follicles but also in the interfollicular zone in which they form a diffuse infiltrate. We have investigated the frequency, extent, and biological characteristics of this interfollicular component in 30 cases of follicular lymphoma. An interfollicular B-cell infiltrate of variable extent (minimal, moderate, or prominent) was present in all cases. Morphologically interfollicular neoplastic B cells were small centrocyte-like cells with lower grade cytology and lower proliferation fraction compared with the neoplastic follicles. The neoplastic phenotype of these cells (CD20+, light chain restricted) was confirmed in 18 cases. Clonal identity between the follicular and interfollicular components was shown in five cases using microdissection and PCR amplification of immunoglobulin heavy chain genes. Analysis of Ig heavy chain gene sequences showed identical variants of tumor subclones in both follicular and interfollicular compartments, indicating active tumor cell traffic between the two. In six cases in which frozen tissue was available, the immunophenotype of follicular and interfollicular tumor cells were compared using immunohistochemistry. Activation markers such as CD10, CD38, and CD95 and T-cell costimulatory molecules CD80 and CD86, which were expressed by neoplastic follicles, were either downregulated or absent in the interfollicular component in most of the cases. The low-grade cytological features, low proliferation fraction, and downregulation of activation markers in the interfollicular neoplastic B cells suggests that these are resting cells analogous to memory B cells of normal lymphoid tissues. The presence of such a resting tumor cell subpopulation in the majority of follicular lymphomas may partly account for the remarkable resistance to therapy of this disease.
Activation of a Plasma Membrane–Associated Neutral Sphingomyelinase and Concomitant Ceramide Accumulation During IgG-Dependent Phagocytosis in Human Polymorphonuclear Leukocytes
Authors: Hinkovska-Galcheva Vania , Kjeldsen Lars , Mansfield Pamela J. , Boxer Laurence A. , Shayman James A. , Suchard Suzanne J. 
Published: October 2019

AbstractThe sphingomyelin cycle, which plays an important role in regulation of cell growth, differentiation, and apoptosis, involves the formation of ceramide by the action of a membrane-associated, Mg2+-dependent, neutral sphingomyelinase and/or a lysosomal acid sphingomyelinase. In human polymorphonuclear leukocytes (PMNs), ceramide production correlates with and plays a role in the regulation of functional responses such as oxidant release and Fcγ receptor-mediated phagocytosis. To increase our understanding of the sphingomyelin cycle in human PMNs, the cellular location of neutral and acid sphingomyelinases was investigated in resting, formylmethionylleucylphenylalanine (FMLP)-activated, and FMLP-activated PMNs engaged in phagocytosis. In resting PMNs, a Mg2+-dependent, neutral sphingomyelinase was the predominant activity and was localized to the plasma membrane fractions along with the majority of ceramide. Upon FMLP-activation, there was a 1.9-fold increase in this neutral, Mg2+-dependent sphingomyelinase activity, which increased to 2.7-fold subsequent to phagocytosis of IgG opsonized targets. This increase in sphingomyelinase activity was restricted to the plasma membrane fractions, which were also the site of increased ceramide levels. Phospholipase D (PLD) activity, which is a target of ceramide action and is required for phagocytosis, was also found primarily in the plasma membrane fractions of FMLP-activated and phagocytosing PMNs. Our findings indicate that in human PMNs engaged in phagocytosis, the sphingomyelin cycle is restricted to the plasma membrane where intracellular targets of ceramide action, such as PLD, are localized.
Lymphocyte-Specific Protein 1 Expression in Eukaryotic Cells Reproduces the Morphologic and Motile Abnormality of NAD 47/89 Neutrophils
Authors: Howard Thomas H. , Hartwig John , Cunningham Casey 
Published: October 2019

AbstractDespite its name, the actin-binding protein lymphocyte-specific protein1 (LSP1) is found in all hematopoetic cells, and yet its role in cell function remains unclear. Recently, LSP1 was identified as the 47-kD protein overexpressed in the polymorphonuclear neutrophils of patients with a rare neutrophil disorder, neutrophil actin dysfunction with abnormalities of 47-kD and 89-kD proteins (NAD 47/89). These neutrophils are immotile, defective in actin polymerization in response to agonists, and display distinctive, fine, “hairlike” F-actin-rich projections on their cell surfaces. We now show that overexpression of LSP1 produces F-actin bundles that are likely responsible for the morphologic and motile abnormalities characteristic of the NAD 47/89 phenotype. Coincident with LSP1 overexpression, cells from each of several different eukaryotic lines, including a highly motile human melanoma line, develop hairlike surface projections that branch distinctively and contain F-actin and LSP1. The hairlike projections are supported at their core by thick actin bundles, composed of actin filaments of mixed polarity, which periodically anastomose to generate a branching structure. The motility of the melanoma cells is inhibited even at low levels of LSP1 expression. Therefore, these studies show that overexpression of LSP1 alone can recreate the morphologic and motile defects seen in NAD 47/89 and suggest that LSP1 is distinct from other known actin binding proteins in its effect on F-actin network structure.
Adenosine Diphosphate (ADP) and ADP Receptor Play a Major Role in Platelet Activation/Aggregation Induced by Sera From Heparin-Induced Thrombocytopenia Patients
Authors: Polgár János , Eichler Petra , Greinacher Andreas , Clemetson Kenneth J. 
Published: October 2019

AbstractThe molecular basis for heparin-induced thrombocytopenia (HIT), a relatively common complication of heparin therapy, is not yet fully understood. We found that pretreatment of platelets with AR-C66096 (formerly FPL 66096), a specific platelet adenosine diphosphate (ADP) receptor antagonist, at a concentration of 100 to 200 nmol/L that blocked ADP-dependent platelet aggregation, resulted in complete loss of platelet aggregation responses to HIT sera. AR-C66096 also totally inhibited HIT serum-induced dense granule release, as judged by measurement of adenosine triphosphate (ATP) release. Apyrase, added to platelets at a concentration that had only minor effects on thrombin- or arachidonic acid-induced aggregation, also blocked completely HIT serum-induced platelet aggregation. Furthermore, AR-C66096 inhibited platelet aggregation and ATP release induced by cross-linking FcγRIIA with specific antibodies. These data show that released ADP and the platelet ADP receptor play a pivotal role in HIT serum-induced platelet activation/aggregation. The thromboxane receptor inhibitor, Daltroban, had no effect on HIT serum-induced platelet activation whereas GPIIb-IIIa antagonists blocked platelet aggregation but had only a moderate effect on HIT serum-induced dense granule release. Pretreatment of platelets with chondroitinases but not with heparinases resulted in concentration dependent inhibition of HIT serum-induced platelet aggregation. These novel data relating to the mechanism of platelet activation induced by HIT sera suggest that the possibility should be examined that ADP receptor antagonists or compounds that inhibit ADP release may be effective as therapeutic agents for the prevention or treatment of complications associated with heparin therapy.
Molecular Cloning and Characterization of Decay-Accelerating Factor Deficiency in Cromer Blood Group Inab Phenotype
Authors: Wang Li , Uchikawa Makoto , Tsuneyama Hatsue , Tokunaga Katsushi , Tadokoro Kenji , Juji Takeo 
Published: October 2019

AbstractAn additional decay-accelerating factor (DAF) mutation, designated as Inab phenotype in the Cromer blood group system, was recently identified in a 28-year-old Japanese woman (H.A.). The red blood cells of H.A., like those of other Inab phenotype individuals, were negative for Cromer system antigens, Cra, Tca, Dra, UMC, and IFC. The deficiency of DAF on the red blood cells of H.A. has been shown by immunoblotting with a murine monoclonal antibody to DAF. Molecular analysis has shown that H.A. is homozygous for a single nucleotide substitution, C1579→A, at the position 24 bp upstream of the 3′-end of exon 2 of the DAF gene. This substitution causes the activation of a novel cryptic splice site and results in the production of mRNA with a 26 bp deletion. The deletion introduces a reading frame shift and creates a stop codon immediately downstream of the deletion. Translation of mRNA would be terminated at the first amino acid residue of the second short consensus repeat (SCR2) domain (exon 3) of DAF. The functional domains of DAF's complement regulatory activity and the carboxy-terminal signal domains for glycosylphosphatidylinositol (GPI) anchoring are predicted to be lacking in H.A. Thus, there would be no DAF present on the cell surface.
T-Cell–Depleted Allogeneic Bone Marrow Transplantation as Postremission Therapy for Acute Myelogenous Leukemia: Freedom From Relapse in the Absence of Graft-Versus-Host Disease
Authors: Papadopoulos Esperanza B. , Carabasi Matthew H. , Castro-Malaspina Hugo , Childs Barrett H. , Mackinnon Stephen , Boulad Farid , Gillio Alfred P. , Kernan Nancy A. , Small Trudy N. , Szabolcs Paul , Taylor Joanne , Yahalom Joachim , Collins Nancy H. , Bleau Sharon A. , Black Patricia M. , Heller Glenn , Reilly Richard J. O' , Young James W. 
Published: October 2019

Abstract
Thirty-one consecutive patients with acute myelogenous leukemia (AML) in first complete remission and 8 with AML in second complete remission received T cell–depleted allogeneic bone marrow transplants from HLA-identical sibling donors. Patients received myeloablative cytoreduction consisting of hyperfractionated total body irradiation, thiotepa, and cyclophosphamide. Those patients at risk for immune-mediated graft rejection received additional immune suppression with antithymocyte globulin and methylprednisolone in the early peritransplant period. Patients with AML who underwent allogeneic T-cell–depleted bone marrow transplantations (BMT) in first or second remission have achieved respective disease-free survival (DFS) probabilities of 77% (median follow-up at approximately 56 months) and 50% (median follow-up at approximately 48 months). Ten of 31 patients transplanted in first remission were ≥ 40 years old and have attained a DFS at 4 years of 70%. For patients with AML transplanted in first or second remission, the respective cause-specific probabilities of relapse were 3.2% or 12.5%, and those of nonleukemic mortality were 19.4% or 37.5%. There were no cases of immune-mediated graft rejection and no cases of grade II to IV acute graft-versus-host disease (GVHD). All survivors enjoy Karnofsky performance scores (KPS) of 100%, except 2 patients with KPS of 80% to 90%. T-cell–depleted allogeneic BMT can provide durable DFS together with an excellent performance status in the majority of patients with de novo AML. In addition, GVHD is not an obligatory correlate of the graft-versus-leukemia benefit or freedom from relapse afforded by allogeneic BMT administered as postremission therapy for AML. This study provides a basis for prospective comparison with other postremission therapies considered standard in the management of patients with this disease.
Effects of Cytokines on Platelet Production From Blood and Marrow CD34+ Cells
Authors: Norol Françoise , Vitrat Natacha , Cramer Elisabeth , Guichard Josette , Burstein Samuel A. , Vainchenker William , Debili Najet 
Published: October 2019

Abstract
The late stages of megakaryocytopoiesis, consisting of the terminal processes of cytoplasmic maturation and platelet shedding, remain poorly understood. A simple liquid culture system using CD34+ cells in serum-free medium has been developed to study the regulation of platelet production in vitro. Platelets produced in vitro were enumerated by flow cytometry. A truncated form of human Mpl-Ligand conjugated to polyethylene glycol (PEG-rHuMGDF) played a crucial role in both proplatelet formation and platelet production. A combination of stem cell factor (SCF), interleukin-3 (IL-3), and IL-6 was as potent as PEG-rHuMGDF for the growth of megakaryocytes (MKs). However, the number of proplatelet-displaying MKs and platelets was increased 10-fold when PEG-rHuMGDF was used. Peripheral blood mobilized CD34+ cells gave rise to a threefold augmentation of platelets compared with marrow CD34+ cells. This finding was related to the higher proliferative capacity of the former population because the proportion of proplatelet-displaying MKs was similar for both types of CD34+ cells. The production of platelets per MK from CD34+ cells was low, perhaps because of the low ploidy of the cultured MKs. This defect in polyploidization correlated with the degree of proliferation of MK progenitors induced by cytokines. In contrast, ploidy development closer to that observed in marrow MKs was observed in MKs derived from the low proliferative CD34+CD41+ progenitors and was associated with a twofold to threefold increment in platelet production per MK. As shown using this CD34+ CD41+ cell population, PEG-rHuMGDF was required throughout the culture period to potently promote platelet production, but was not involved directly in the process of platelet shedding. IL-3, SCF, and IL-6 alone had a very weak effect on proplatelet formation and platelet shedding. Surprisingly, when used in combination, these cytokines elicited a degree of platelet production which was decreased only 2.4-fold in comparison with PEG-rHuMGDF. This suggests that proplatelet formation may be inhibited by non-MK cells which contaminate the cultures when the entire CD34+ cell population is used. Cultured platelets derived from PEG-rHuMGDF– or cytokine combination-stimulated cultures had similar ultrastructural features and a nearly similar response to activation by thrombin. The data show that this culture system may be useful to study the effects of cytokines and the role of polyploidization on platelet production and function.
Cloning and Characterization of the Human Homolog of Mouse Jak2
Authors: Dalal Ilan , Arpaia Enrico , Dadi Harjit , Kulkarni Shaila , Squire Jerami , Roifman Chaim M. 
Published: October 2019

AbstractMembers of the Jak family play a critical role in signal transduction mediated by cytokine and hormone receptors. In this study, we report the cloning and characterization of human Jak2. The predicted amino acid sequence shows 91% homology to the described murine Jak2, but with a significant difference in the extreme C-terminal sequence. Using the human cDNA as a probe, we localized the gene for human Jak2 to chromosome 9p23-24. Human Jak2 mRNA is highly expressed in the spleen, lymph nodes, and peripheral blood lymphocytes (PBLs). A polyclonal antibody raised against the unique C-terminus of human Jak2 was used to characterize Jak2 protein. Levels of Jak2 protein expression increased significantly in mitogen- and anti-IgM–stimulated B cells and to a lesser degree in activated T cells. In addition, high levels of Jak2 protein were detected in pre-B leukemia cells.