bluuebunny/crossref_metadata_2025_split huggingface dataset:
Identification and characterization of a functional receptor for interferon-gamma on a megakaryocytic cell line
Authors: Monte D , Wietzerbin J , Pancre V , Merlin G , Greenberg SM , Kusnierz JP , Capron A , Auriault C 
Published: October 2019

Abstract
We have previously shown that human interferon-gamma (Hu-IFN-gamma) induces platelets to become efficient effector cells, capable of killing young larvae of the parasite Schistosoma mansoni. Recently, binding sites for IFN-gamma on platelets have been characterized. We show here the presence of high-affinity receptors for IFN-gamma on the surface of the human megakaryocytic Dami cell line. Scatchard analysis indicated the presence of about 11,000 binding sites per cell, with a kd of 3 +/- 0.5 x 10(-10) mol/L; the apparent molecular weight of the receptor was 90 Kd. Receptor-bound 125I Hu-recombinant IFN-gamma was rapidly internalized and degraded when the temperature was increased from 4 degrees C to 37 degrees C. The half-life of this receptor was about 7 hours, and pretreatment of cells with IFN-gamma or phorbol myristate acetate had very little effect on the surface receptor number and no detectable effect on IFN-gamma receptor messenger RNA (mRNA) expression. The receptor was functional, because 24 hours of treatment with IFN-gamma led to the increase of HLA class I mRNA expression and to the initiation of HLA class II mRNA expression. These effects were selective because platelet glycoprotein Ib, IIb, or IIIa mRNA expression and cell proliferation were unaffected.
Abundance and stability of erythropoietin receptor mRNA in mouse erythroid progenitor cells
Authors: Wickrema A , Bondurant MC , Krantz SB 
Published: October 2019

Abstract
The abundance and stability of erythropoietin (EPO) receptor mRNA was measured in Friend virus-infected mouse splenic erythroblast (FVA) cells. FVA cells correspond to the colony-forming unit-erythroid (CFU- E) and cluster-forming unit-erythroid stages of differentiation and they mature in vitro into reticulocytes during 45 to 60 hours of culture. After 20 hours, there was a rapid decline in the EPO receptor mRNA level through the 45-hour culture period. Levels of actin and beta- globin mRNAs were monitored as two representative proteins that are actively synthesized in early versus late stages of terminal erythroid differentiation. A general decrease of actin mRNA level was apparent, whereas globin mRNA levels increased throughout the culture period. The greatest decrease in both EPO receptor and actin mRNA was observed when the cell population was at the stage of basophilic and polychromatophilic erythroblasts. The half-life for EPO receptor mRNA was approximately 75 minutes, as determined using the transcriptional inhibitor actinomycin D. The half-life measured at several times during the 48-hour culture period remained constant. These results indicate that EPO receptor mRNA must be transcribed continuously until late in the maturation process of FVA cells in order to maintain the levels seen by Northern analysis. The number of copies of EPO receptor mRNA at 0 hours was determined to be 25 copies per cell. This low number and the decline of EPO receptor mRNA correlate with the low receptor numbers on FVA cells, as well as the decline of binding sites with cell maturation.
Alpha-2-macroglobulin may provide protection from thromboembolic events in antithrombin III-deficient children
Authors: Mitchell L , Piovella F , Ofosu F , Andrew M 
Published: October 2019

Abstract
Antithrombin III (ATIII) deficiency has been implicated in adults as a predisposing factor to thrombosis; however, thromboembolic complications are rare in children with the same deficiency. We hypothesized that because of the elevated levels of plasma alpha-2- macroglobulin (alpha 2M) throughout childhood, plasmas of ATIII- deficient children inhibit thrombin more efficiently than those of ATIII-deficient adults. In total, 14 ATIII-deficient adults (ages 25 to 46 years), 13 ATIII-deficient children (ages 2 to 13 years), 9 normal children (ages 3 to 15 years), and 16 normal adults were studied. We measured thrombin inhibition in these plasmas, as well as the contributions of ATIII, alpha 2M, and heparin cofactor II (HCII) as thrombin inhibitors in each plasma. 125I-alpha-thrombin, 25 nmol/L, was added to each plasma (defibrinated with Arvin at 37 degrees C), and 90 seconds later the free thrombin and thrombin-inhibitor complexes were quantitated after sodium dodecyl sulfate-polyacrylamide gel electrophoresis, autoradiography, and densitometric scanning. Plasma from ATIII-deficient adults inhibited significantly less thrombin (12.8 +/- 0.6 nmol/L) than both normal adults (16.1 +/- 0.3 nmol/L, P less than .01), normal children (15.7 +/- 0.4 nmol/L, P less than .01), or ATIII-deficient children (15.5 +/- 0.3 nmol/L, P less than .01). There was no significant difference between the total concentration of thrombin inhibited by ATIII-deficient children and either normal adult or normal children groups. In addition, plasmas of ATIII-deficient children inhibited thrombin significantly more efficiently than plasma of ATIII-deficient adults (P less than .01). In the ATIII-deficient patients there was a significant correlation between the alpha 2M level and ability to inhibit thrombin (P less than .01), but no correlation between either ATIII or HCII levels and thrombin inhibition. On the addition of heparin (0.4 U/mL) to plasma, all four types of plasma inhibited thrombin to the same extent. Although ATIII was the predominant inhibitor in all heparinized plasmas, HCII inhibited more thrombin in the ATIII-deficient patients than in normal patients (2.8 +/- 0.3 v 1.2 +/- 0.2 nmol/L, P less than .01). We hypothesize that the lower risk of thromboembolic complications in ATIII-deficient children may be due in part to the protective effect of elevated alpha 2M levels during childhood.
Synthetic peptides homologous to human glycophorins of the Miltenberger complex of variants of MNSs blood group system specify the epitopes for Hil, SJL, Hop, and Mur antisera
Authors: Johe KK , Vengelen-Tyler V , Leger R , Blumenfeld OO 
Published: October 2019

Abstract
The antigenic epitopes of the MNSs blood groups are localized on alpha and delta glycophorins (glycophorins A and B) of the erythrocyte surface. Hil, SJL, Mur, and Hop antisera define the Miltenberger (Mi) complex of MiV, MiJ.L., MiIII, and MiVI variant serologic phenotypes of this blood group system. We report here the location of the epitopes for antibodies in these antisera. The antigens of these Mi classes are variant glycophorins that are hybrids of alpha and delta glycophorins in alpha-delta and delta-alpha-delta arrangements. The hybrid junctions give rise to novel polypeptide sequences not present in the parent glycophorins; in MiIII and MiVI this also includes an expressed sequence of the delta pseudoexon. These sequences are identical in the above Mi-glycophorins occurring in erythrocytes that share a common Mi determinant. Four peptides of 10 to 14 amino acids each were constructed to be homologous to the identical sequences; they were designated, “Hil”, “SJL”, “Mur”, and “Hop” to reflect the common determinant. The peptides were tested for inhibition of reaction of appropriate cells with the relevant antisera. The Hil peptide, outlining the alpha-delta s junction region in MiIII, MiV, and MiVI glycophorins, inhibited the reaction of respective erythrocytes (red blood cells [RBCs]) with anti-Hil. The SJL peptide, which differs from the Hil peptide by a single Thr----Met substitution, was specific for inhibition of the reaction of MiJ.L. RBCs with anti-SJL (an example of anti-S specific for such RBCs). The Hop peptide, which corresponds to the delta-alpha junction in MiVI glycophorin, inhibited the hemagglutination of MiVIII RBCs by anti-Hop. MiVI and MiVIII glycophorins share an identical sequence at that site. The Mur peptide, corresponding to a portion of the expressed pseudoexon sequence in MiIII and MiVI glycophorins, was specific for inhibition of the reaction of MiIII and MiVI RBCs with anti-Mur. The peptides had no effect on the hemagglutination of control MNSs RBCs by their respective antisera nor of unrelated Mi classes RBCs by antisera that distinguish these classes. We conclude that the alpha-delta junction in MiIII, MiV, and MiVI glycophorins outlines the epitopes for anti-Hil, the alpha- delta junction in MiJ.L. outlines the epitope for anti-SJL, the delta- alpha junction in MiVI constitutes the epitope for anti-Hop, and the expressed delta pseudoexon sequence in MiIII and MiVI constitutes the epitope for anti-Mur.
Immobilized Arg-Gly-Asp (RGD) peptides of varying lengths as structural probes of the platelet glycoprotein IIb/IIIa receptor
Authors: Beer JH , Springer KT , Coller BS 
Published: October 2019

Abstract
The interactions between ligands containing the recognition sequence arginine-glycine-aspartic acid (RGD) and integrin receptors are important in many cell-cell and cell-protein interactions. The platelet contains five integrin receptors and they contribute significantly to platelet adhesion and aggregation. To investigate the RGD binding domains on platelet integrins, we immobilized a series of RGD peptides containing variable numbers of glycine residues [(G)n-RGDF] on polyacrylonitrile beads and evaluated the ability of the beads to interact with platelets. With native platelets, virtually no interaction occurred with G1-RGDF beads, but the interactions increased as the number of glycine residues increased, plateauing with the G9- RGDF and G11-RGDF beads. ADP pretreatment enhanced the interactions with all of the beads, whereas prostaglandin E1 pretreatment eliminated the interactions with the shortest peptide beads, but only partially inhibited interactions with the longer peptide beads. Monoclonal antibodies to glycoprotein (GP) IIb/IIIa were most effective in inhibiting the interactions, but antibodies to GPIIb/IIIa with similar inhibitory effects on fibrinogen binding varied dramatically in their ability to inhibit the interaction between platelets and immobilized RGD peptides. Our data indicate that the majority of RGD binding sites on GPIIb/IIIa can be reached by peptides that extend out approximately 11 to 32 A from the surface of the bead, and these results are in accord with the dimensions of integrin receptors deduced from electron microscopy. Activation of GPIIb/IIIa facilitates the interactions, but platelet inhibition fails to eliminate the interactions with the longer peptide beads, suggesting that access to the RGD binding site on at least a fraction of the GPIIb/IIIa receptors is always possible for preferred ligands. Finally, we found that the G3-RGDF peptide beads were uniquely sensitive to the activation state of the GPIIb/IIIa receptor.
Origin and fate of iron mobilized by the 3-hydroxypyridin-4-one oral chelators: studies in hypertransfused rats by selective radioiron probes of reticuloendothelial and hepatocellular iron stores
Authors: Zevin S , Link G , Grady RW , Hider RC , Peter HH , Hershko C 
Published: October 2019

Abstract
The mechanism of in vivo iron chelation by 3-hydroxypyridin-4-ones (CP compounds) was studied in hypertransfused rats in which the major storage iron pools in hepatocytes and in the reticuloendothelial (RE) system have been labeled by selective radioiron probes. Both dimethyl-3- hydroxypyridin-4-one (CP 20 or L1) and diethyl-3-hydroxypyridine-4-one (CP 94) have an identical and very high (log beta 3 36) binding constant and selective affinity to iron(III), but the lipid solubility of CP 94 is considerably higher than that of CP 20. Both chelators induced an increase in the fecal excretion of hepatocellular iron with no effect on urinary excretion. In contrast, about one third to one half of the iron mobilized from RE cells was excreted in the urine. The chelating efficiency of CP 20 was comparable with that of deferoxamine (DF), whereas CP 94 was up to eight times more effective than DF. Unlike DF, which had no effect by the oral route, the oral and parenteral effectiveness of both CP compounds was identical. These findings indicate that: (1) lipid solubility is an important determinant of in vivo chelating efficiency; (2) urinary iron excretion induced by the CP compounds is derived from RE cells; (3) part of the iron mobilized from RE cells and all of the iron derived from hepatocytes is excreted through the bile; and (4) contrary to previous observations in cell cultures, there is no in vivo evidence for a diminishing chelating efficiency at the lowest doses used.
bcr-abl-Induced cell lines can switch from mast cell to erythroid or myeloid differentiation in vitro
Authors: Elefanty AG , Cory S 
Published: October 2019

Abstract
The chimeric bcr-abl gene formed by the Philadelphia translocation is thought to initiate chronic myeloid leukemia. Engraftment of mice with bone marrow cells infected with a bcr-abl retrovirus has been shown to elicit multiple hematopoietic disorders, including a clonal but nontransplantable hyperproliferation of erythroid and/or mast cells. Culture of spleen and bone marrow cells from such mice usually yielded mast cell lines, even when erythroid disease dominated the primary animal. The mast cells, which carried the same proviral insert as the primary disease, generally grew slowly and were neither transplantable nor clonogenic in agar until they had been cultured for several months. Unexpectedly, several bcr-abl-induced lines switched in vitro from mast cell to megakaryocytic and/or erythroid character, and one became myeloid. The dramatic phenotypic shifts seem likely to involve changes occurring within progenitor cells maintaining the clone, rather than mutation of mature mast cells. The variant lines exhibited substantial spontaneous differentiation, despite being readily transplantable and therefore fully transformed. The production of hematopoietic growth factors by the mast cell lines and their phenotypic variants may implicate an autocrine loop in their evolution. These novel bcr-abl cell lines should aid in the study of genetic events in the progression from chronic to acute leukemia and facilitate analysis of hematopoietic lineage commitment.
Stromal support enhances cell-free retroviral vector transduction of human bone marrow long-term culture-initiating cells
Authors: Moore KA , Deisseroth AB , Reading CL , Williams DE , Belmont JW 
Published: October 2019

Abstract
Gene transfer into hematopoietic stem cells by cell-free virions is a goal for gene therapy of hematolymphoid disorders. Because the hematopoietic microenvironment provided by the stroma is required for stem cell maintenance both in vivo and in vitro, we reasoned that cell- free transduction of bone marrow cells (BMC) may be aided by stromal support. We used two high-titer replication-defective retroviral vectors to differentially mark progenitor cells. The transducing vector was shown to be a specific DNA fragment by polymerase chain reaction of colony-forming cells derived from progenitors maintained in long-term culture (LTC). BMC were infected separately by cell-free virions with or without pre-established, irradiated, allogeneic stromal layers, and in the presence or absence of exogenous growth factors (GF). The GF assessed were interleukin-3 (IL-3) and IL-6 in combination, leukemia inhibitory factor (LIF), mast cell growth factor (MGF), and LIF and MGF in combination. In addition, we developed a competitive LTC system to directly assess the effect of infection conditions on the transduction of clonogenic progenitors as reflected by the presence of a predominate provirus after maintenance in the same microenvironment. The results show gene transfer into human LTC-initiating cells by cell-free retroviral vector and a beneficial effect of stromal support allowing a transduction efficiency of 64.6% in contrast to 15.8% without a supporting stromal layer. A high transduction rate was achieved independent of stimulation with exogenous GF. We propose that autologous marrow stromal support during the transduction period may have application in clinical gene therapy protocols.
Analysis of the mechanism of anagrelide-induced thrombocytopenia in humans
Authors: Mazur EM , Rosmarin AG , Sohl PA , Newton JL , Narendran A 
Published: October 2019

Abstract
Anagrelide is a new therapeutic compound recently demonstrated to have a rapid and selective thrombocytopenic effect in humans. The effects of anagrelide were evaluated in plasma clot and liquid suspension cultures of optimally stimulated normal human peripheral blood megakaryocyte progenitors in order to determine the mechanism of its thrombocytopenic activity. In plasma clot cultures, at clinically relevant, therapeutic concentrations (5 to 50 ng/mL), anagrelide exerted no significant inhibitory effect on megakaryocyte colony numbers or colony size. Only at anagrelide concentrations of 10 to 500 times therapeutic doses did anagrelide inhibit megakaryocyte colony development: an anagrelide concentration of 5 micrograms/mL reduced colony numbers by 57% and colony size by 31%. In contrast, lower, therapeutic anagrelide concentrations exerted profound effects in liquid culture on megakaryocyte cytoplasmic maturation, size, and DNA content. When present for the entire 12-day culture duration, anagrelide induced left- shifted megakaryocyte maturation and reduced both megakaryocyte ploidy and megakaryocyte diameter. Anagrelide, at concentrations of 5 to 50 ng/mL, shifted the modal cultured megakaryocyte morphologic stage from III to II, reduced the model ploidy value from 16N to 8N, and decreased the mean megakaryocyte diameter by up to 22%, from 27.6 microns to 21.6 microns. Megakaryocyte diameter was significantly reduced in most instances, even when analyzed as a function of morphologic stage. When anagrelide was added to the cultures after 6- and 9-day delays (during the final 6 and 3 days, respectively, of culture), similar inhibitory effects on megakaryocyte maturation stage and ploidy distribution were observed. However, the magnitude of the left-shift in ploidy appeared to be less as the duration of anagrelide exposure was reduced. Conversely, megakaryocyte diameter was not significantly affected by the shorter 3- and 6-day anagrelide exposures. These data indicate that therapeutic concentrations of anagrelide influence primarily the postmitotic phase of megakaryocyte development, decreasing platelet production by reducing megakaryocyte size and ploidy, as well as by disrupting full megakaryocyte maturation. Inhibition of megakaryocyte diameter appears to require more prolonged anagrelide exposure than inhibition of maturation stage and ploidy. The molecular mechanisms responsible for the inhibitory effects of anagrelide on megakaryocytopoiesis remain to be defined.
Growth factor-dependent proliferative stimulation of hematopoietic cells is associated with the modulation of cytoplasmic and nuclear 68- Kd calmodulin-binding protein
Authors: Reddy GP , Reed WC , Deacon DH , Quesenberry PJ 
Published: October 2019

Abstract
Calcium and calmodulin (CaM) are known to play critical roles in controlling cell cycle progression in a variety of cells. We observed that the CaM antagonist, N-(6-aminohexyl)-5-chloro-1- naphthalensulfonamide hydrochloride (W-7), inhibited 3H-thymidine incorporation into DNA of factor-dependent hematopoietic cells. To delineate the role of CaM in proliferation of hematopoietic cells, we have investigated intracellular distribution of specific CaM-binding proteins (CaM-BPs) in response to hematopoietic growth factors in FDC- P1, 32D, NFS-60, and T1165 cells. Each of these cell lines, when deprived of cytokines for 16 to 18 hours, essentially ceased proliferation, even in the presence of fetal calf serum. Concomitant to the cessation of proliferation, there was a dramatic depletion of a specific CaM-BP of about 68 Kd in both their cytoplasmic and nuclear fractions. Within 6 to 12 hours of reexposure to proliferation-specific cytokines, there was a restoration of the nuclear as well as cytoplasmic 68-Kd CaM-BP. Furthermore, such an induction and nuclear localization of the 68-Kd CaM-BP by the cytokines coincided temporally with the progression of synchronized FDC-P1 cells from G1 to S phase. By contrast, colony-stimulating factor-1 (CSF-1)-dependent bone marrow macrophages and BAC-1 cells did not exhibit 68-Kd CaM-BP in the nuclear or cytoplasmic fractions. These studies suggest that while hematopoietic growth factor granulocyte CSF-, granulocyte-macrophage CSF-, interleukin-3 (IL-3)-, or IL-6-, whose receptors are members of the hematopoietin receptor family, induced cell proliferation is associated with a common mechanism involving nuclear localization of the 68-Kd CaM-BP, the CSF-1-induced proliferation seems to involve 68- Kd CaM-BP-independent pathways.
Rapid and simultaneous typing of hemoglobin S, hemoglobin C, and seven Mediterranean beta-thalassemia mutations by covalent reverse dot-blot analysis: application to prenatal diagnosis in Sicily
Authors: Maggio A , Giambona A , Cai SP , Wall J , Kan YW , Chehab FF 
Published: October 2019

Abstract
The molecular lesions causing beta-thalassemia in Sicily can be subdivided into two groups. One that occurs at a 71% frequency and consists of the beta 39, IVS 1,110 and IVS 1,6 mutations and the other group at a 20% frequency comprising the -87, beta s, IVS 1,1 and IVS 2,745 mutations. The identification of all these mutations by polymerase chain reaction (PCR) and conventional dot-blot hybridization has been time consuming and expensive. In this article, we describe the implementation of the reverse dot-blot (RDB) hybridization as a rapid nonradioactive method for the identification of the nine most frequent molecular lesions in the beta-globin gene (-87, beta s, beta c, IVS 1,1, IVS 1,6, IVS 1,110, beta 39, IVS 2,1, IVS 2,745) in Sicily. Sixty prenatal diagnoses were performed by this RDB assay, each of which was confirmed by dot-blot/ASO hybridization; thus demonstrating the accuracy of the RDB. The main advantage of this assay is the rapid typing of an individual's DNA for many mutations in a single working day. Because the mutations in this assay are representative for the Mediterranean region, this mutational panel can also be extended to the screening of beta-thalassemia from other Mediterranean regions.