bluuebunny/crossref_metadata_2025_split huggingface dataset:
Priming effects of granulocyte-macrophage colony-stimulating factor are coupled to cholera toxin-sensitive guanine nucleotide binding protein in human T lymphocytes
Authors: al-Aoukaty A , Giaid A , Sinoff C , Ho AD , Maghazachi AA 
Published: October 2019

Abstract
In addition to the mobilization of neutrophils and monocytes, granulocyte-macrophage colony-stimulating factor (GM-CSF) also mobilizes lymphocytes into peripheral blood. We examined the ability of GM-CSF to induce the proliferation of purified human T cells (CD3+ CD4+ CD56- CD16- B1- MO2-) in two major aspects: (1) the mechanisms of GM- CSF interaction with interleukin-2 (IL-2) causing T-cell proliferation, and (2) the intracellular signals transmitted by GM-CSF in T lymphocytes. We observed that concentrations of GM-CSF between 0.01 ng/mL and 10 ng/mL had a synergistic effect with concentrations of IL-2 between 1 U/mL and 10 U/mL in stimulating T-cell proliferation. This effect of GM-CSF was maximal when it was added at the start of the culture. In situ hybridization showed the presence of mRNA for GM-CSF receptors in T cells. Further analysis showed that GM-CSF induced the expression of IL-2 receptor (IL-2R) on the surface of T lymphocytes. These events coincide with the ability of GM-CSF to increase the intracellular levels of both cyclic 3′,5′-adenosine monophosphate (cAMP) and cyclic 3′,5′-guanosine monophosphate (cGMP) in T cells, to increase the binding of (gamma-35S) GTP to T-cell membranes, and to enhance GTPase activity as determined by increased hydrolysis of 32P- GTP. IL-2 also induced IL-2R expression, cyclic nucleotide secretion, and G-protein activation. However, the presence of IL-2 reduced GM-CSF induction of these activities. Addition of antibodies to the alpha and beta subunits of IL-2R permitted the activation of G protein by GM-CSF even when IL-2 was present. Furthermore, GTP binding and GTPase activity induced by GM-CSF or IL-2 were inhibited by the addition of cholera toxin (CT), but not pertussis toxin (PT). Cumulatively, these results suggest that in T lymphocytes, receptors for GM-CSF or IL-2 may be coupled to the same CT-sensitive G protein, although other possibilities may exist. The role that G proteins play in mediating the intracellular signaling pathways induced by GM-CSF or IL-2 in human T cells is supported by adenosine diphosphate-ribosylation of a 44-kD or a 39-kD G protein in T-cell membranes by CT and PT, respectively.
DNA rearrangements proximal to the EVI1 locus associated with the 3q21q26 syndrome
Authors: Levy ER , Parganas E , Morishita K , Fichelson S , James L , Oscier D , Gisselbrecht S , Ihle JN , Buckle VJ 
Published: October 2019

Abstract
Specific rearrangements involving 3q21 and 3q26 are well documented in acute myeloid leukemia (AML). Aberrant expression of the Ecotropic virus integration-1 (EVI1) gene, located at 3q26, has been reported in individuals with AML and translocations or inversions of chromosome 3 long arm. We have studied six individuals with AML and inv(3)(q21q26) for disruptions to the EVI1 locus by in situ hybridization and long- range mapping. EVI1 transcripts have been detected in the blast cells of the two individuals available for expression studies. We derived a YAC containing the EVI1 gene and showed that it crossed the 3q26 inversion breakpoints in three of four cases examined. Pulsed field analysis detected aberrant fragments 3′ of the EVI1 gene in all six patients. The orientation of the gene was established and the locations of the breakpoints were refined by in situ hybridization using phage clones from this region.
The 87-kD A gamma-globin enhancer-binding protein is a product of the HOXB2(HOX2H) locus
Authors: Sengupta PK , Lavelle DE , DeSimone J 
Published: October 2019

Abstract
Developmental regulation of globin gene expression may be controlled by developmental stage-specific nuclear proteins that influence interactions between the locus control region and local regulatory sequences near individual globin genes. We previously isolated an 87-kD nuclear protein from K562 cells that bound to DNA sequences in the beta- globin locus control region, gamma-globin promoter, and A gamma-globin enhancer. The presence of this protein in fetal globin-expressing cells and its absence in adult globin-expressing cells suggested that it may be a developmental stage-specific factor. A lambda gt11 K562 cDNA clone encoding a portion of the HOXB2 (formerly HOX2H) homeobox gene was isolated on the basis of the ability of its beta-galactosidase fusion protein to bind to the same DNA sequences as the 87-kD K562 protein. Because no other relationship had been established between the 87-kD K562 protein and the HOXB2 protein other than their ability to bind ot the same DNA sequences, we have investigated whether the two proteins are related antigenically. Our data show that antisera produced against the HOXB2-beta-gal fusion protein and a synthetic HOXB2 decapeptide react specifically with an 87-kD protein from K562 nuclear extract, showing that the 87-kD K562 nuclear protein is a product of the HOXB2 locus, and is the first demonstration of cellular HOXB2 protein.
Rickettsia rickettsii infection of cultured human endothelial cells induces tissue factor expression
Authors: Sporn LA , Haidaris PJ , Shi RJ , Nemerson Y , Silverman DJ , Marder VJ 
Published: October 2019

Abstract
Microvascular thrombi underlie many of the clinical manifestations of Rocky Mountain spotted fever (RMSF), a disease characterized by Rickettsia rickettsii infection of vascular endothelial cells. Studies were designed to determine whether R rickettsii-infection of cultured human umbilical vein endothelial cells results in tissue factor (TF) induction, a process that could directly activate coagulation in infected vessels. Whereas uninfected endothelial cell cultures showed essentially undetectable TF mRNA and activity, both TF mRNA and activity were present after R rickettsii infection. TF mRNA levels were transient, peaking at 4 hours after the initiation of infection, whereas the peak of TF activity occurred at 8 hours. Induction of the TF response requires the intracellular presence of R rickettsii organisms, because uninfected rickettsia were ineffective and the response was blocked by inhibiting rickettsial entry using cytochalasin B. TF induction was not mediated by endothelial cell release of soluble factor, because no response was induced using culture medium conditioned by R rickettsii-infected cells. Furthermore, preadsorption of suspensions of R rickettsii with polymyxin B to remove contaminating lipopolysaccharide did not eliminate the TF response. Induction of TF in vital endothelial cells during R rickettsii infection could be the trigger for vascular thrombus formation of RMSF.
Induction of differentiation of promyelocytic NB4 cells by retinoic acid is associated with rapid increase in urokinase activity subsequently downregulated by production of inhibitors
Authors: Tapiovaara H , Matikainen S , Hurme M , Vaheri A 
Published: October 2019

Abstract
13-trans retinoic acid (13-trans RA) is an effective inducer of differentiation of acute promyelocytic (APL) cells both in vivo and in vitro. It is used in the induction of remission of patients with APL. We found, by using the promyelocytic NB4 cell line established from a patient with APL, that the induction of differentiation with RA was accompanied by modulation of the plasminogen activation system. The expression of urokinase (uPA) activity was rapidly increased in the growth medium and at the surface of cells treated with RA. The high uPA activity was counteracted both in the growth medium and at the cell surface by an increased plasminogen activator inhibitor (PAI) production and reduction of uPA synthesis. The expression of uPA receptor and PAI-2 were stimulated and persisted at 48 hours from RA addition. The simultaneous induction of CD11b suggests that differentiation results in increased production of both. APL patients often encounter episodes of disseminated intravascular coagulation that are associated with secondary fibrinolytic events. Our results suggest that downregulation of uPA activity results in the decrease of plasmin on the surface of the differentiated cells, which may reduce the occurrence of fibrinolytic episodes of patients with APL.
An endogenous glycosylphosphatidylinositol-specific phospholipase D releases basic fibroblast growth factor-heparan sulfate proteoglycan complexes from human bone marrow cultures
Authors: Brunner G , Metz CN , Nguyen H , Gabrilove J , Patel SR , Davitz MA , Rifkin DB , Wilson EL 
Published: October 2019

Abstract
Basic fibroblast growth factor (bFGF) is a hematopoietic cytokine that stimulates stromal and stem cell growth. It binds to a glycosylphosphatidylinositol (GPI)-anchored heparan sulfate proteoglycan on human bone marrow (BM) stromal cells. The bFGF- proteoglycan complex is biologically active and is released by addition of exogenous phosphatidylinositol-specific phospholipase C. In this study, we show the presence of an endogenous GPI-specific phospholipase D (GPI-PLD) that releases the bFGF-binding heparan sulfate proteoglycan and the variant surface glycoprotein (a model GPI-anchored protein) from BM cultures. An involvement of proteases in this process is unlikely, because released proteoglycan contained the GPI anchor component, ethanol-amine, and protease inhibitors did not diminish the release. The mechanism of release is likely to involve a GPI-PLD and not a GPI-specific phospholipase C, because the release of variant surface glycoprotein did not reveal an epitope called the cross- reacting determinant that is exposed by phospholipase C-catalyzed GPI anchor cleavage. In addition, phosphatidic acid (which is specifically a product of GPI-PLD-catalyzed anchor cleavage) was generated during the spontaneous release of the GPI-anchored variant surface glycoprotein. We also detected GPI-PLD-specific enzyme activity and mRNA in BM cells. Therefore, we conclude that an endogenous GPI-PLD releases bFGF-heparan sulfate proteoglycan complexes from human BM cultures. This mechanism of GPI anchor cleavage could be relevant for mobilizing biologically active bFGF in BM. An endogenous GPI-PLD could also release other GPI-anchored proteins important for hematopoiesis and other physiologic processes.
Retrovirus-mediated gene transfer of human pyruvate kinase (PK) cDNA into murine hematopoietic cells: implications for gene therapy of human PK deficiency
Authors: Tani K , Yoshikubo T , Ikebuchi K , Takahashi K , Tsuchiya T , Takahashi S , Shimane M , Ogura H , Tojo A , Ozawa K 
Published: October 2019

Abstract
With the advent of recent molecular studies, nonspherocytic hemolytic anemia caused by red blood cell pyruvate kinase (PK) deficiency is now considered to be caused by a structural mutation of the PK-LR gene. Because PK deficiency is a monogenic disorder, the introduction of the normal PK gene into a patient's bone marrow stem cells should cure the disorder. To study the feasibility of gene therapy for PK deficiency, we first constructed the PK retrovirus pMNSM-hPK using human liver-type PK (LPK) cDNA and obtained a producer cell line of E86/AmPK. By using the supernatant of this virus-producer cell, we transduced NIH/3T3 cells, mouse leukemic cells (NFS60, FDCP-2), and human leukemic cells (K562, HEL). The expression of human LPK enzyme activity was ascertained from the retrovirally transduced NIH/3T3 cells. Northern blot analysis demonstrated the expression of the human LPK mRNA in each transduced cell line. Furthermore, bone marrow stem cells (c-kit+, Lin- , Thy-1lo) sorted by fluorescence-activated cell sorting were also transduced by the producer cells in the presence of interleukin-3 and interleukin-6, and were transplanted into lethally irradiated C57BL/6 mice. Polymerase chain reaction analysis demonstrated the expression of human LPK mRNA in both the peripheral blood and hematopoietic organs on day 30 and on day 135 of bone marrow transplantation.
Molecular abnormality of erythrocyte pyruvate kinase deficiency in the Amish
Authors: Kanno H , Ballas SK , Miwa S , Fujii H , Bowman HS 
Published: October 2019

Abstract
We describe the cellular and molecular biologic studies of the erythrocyte pyruvate kinase (PK) deficiency of the Amish deme in Pennsylvania. Nucleotide sequencing of the patient's PK gene showed a point mutation, CGC to CAC, corresponding to no. 1436 from the translational initiation site of the R-type PK (R-PK) mRNA, and it caused a single amino acid substitution from Arg to His at the 479th amino acid residue of the R-PK. The substituted Arg residue is located in the C domain of PK subunit, that is essential for both the intersubunit contact and the allosteric regulation. Because this enzyme shows the catalytic activity only as a dimer or tetramer, it is rational that the structural alteration would result in severe PK deficiency. To elucidate the effect of the PK deficiency on red blood cell (RBC) membrane, we performed the cellular studies of the patients' RBCs. Ouabain-insensitive K+ efflux was increased to 142% to 145% of normal controls and not inhibited by furosemide, as previously observed in HbSC disease RBCs.
Iron-balance and dose-response studies of the oral iron chelator 1,2- dimethyl-3-hydroxypyrid-4-one (L1) in iron-loaded patients with sickle cell disease
Authors: Collins AF , Fassos FF , Stobie S , Lewis N , Shaw D , Fry M , Templeton DM , McClelland RA , Koren G , Olivieri NF 
Published: October 2019

Abstract
Several life-threatening complications of the common disorder sickle cell disease require management with red blood cell transfusions and, hence, long-term iron-chelating therapy. The efficacy of the oral iron chelator 1,2-dimethyl-3-hydroxypyrid-4-one (L1) has not previously been determined in patients with sickle cell disease. We compared the efficacy of L1 to that of standard-dose subcutaneous deferoxamine in four regularly transfused patients with homozygous sickle cell disease, who had evidence of severe iron overload and a history of poor compliance with deferoxamine. Determination of 24-hour urinary iron excretion conducted over 5 days immediately after transfusion showed that the mean daily urinary iron excretion induced by L1 at 75 mg/kg/d (0.48 +/- 0.23 mg/kg) was equivalent to that induced by deferoxamine at 50 mg/kg/d (0.39 +/- 0.06 mg/kg). In two of three patients studied, a significant (P < .025) increase in mean daily urinary iron excretion was achieved when the dose of L1 was increased to 100 mg/kg/d. Total iron balance studies, which quantitated both urinary and stool iron excretion on L1 and deferoxamine, determined that mean total daily iron excretion induced by deferoxamine (0.88 +/- 0.05 mg/kg) was significantly greater (P < .05) than that induced by L1 (0.53 +/- 0.17 mg/kg), attributable to the significantly greater stool iron excretion during deferoxamine treatment (0.50 +/- 0.16 mg/kg/d) compared with that measured during L1 treatment (0.12 +/- 0.08 mg/kg/d, P < .01). Stool iron excretion accounted for a significantly greater percentage of total iron excretion during deferoxamine treatment (59% +/- 20%) than during L1 treatment (23% +/- 14%, P < .01). These iron balance studies are the first to compare total iron excretion induced by L1 with that achieved by deferoxamine. They demonstrate that the mean total daily iron excretion during L1 treatment (0.53 +/- 0.17 mg/kg) is sufficient to maintain net negative iron balance in most regularly transfused patients with sickle cell disease. Because long-term compliance with L1 has been shown previously to be superior to that with deferoxamine in patients with homozygous beta-thalassemia, the use of L1 should increase the long-term effectiveness of iron chelation in patients with sickle cell disease.
Development of progressive kidney damage and myeloma kidney in interleukin-6 transgenic mice
Authors: Fattori E , Della Rocca C , Costa P , Giorgio M , Dente B , Pozzi L , Ciliberto G 
Published: October 2019

Abstract
Interleukin-6 (IL-6) is a pleiotropic cytokine that has been postulated as playing a role in the pathogenesis of multiple myeloma, chronic autoimmune diseases, and alcoholic liver cirrhosis. We generated transgenic mice carrying a fusion between the mouse metallothionein-I (MT-I) gene promoter and the human IL-6 cDNA. MT-I/IL-6 transgenics express IL-6 constitutively in the liver and secrete the cytokine in the blood. They show initially activation of acute-phase response genes and accumulation of alpha 2- and beta-globulins in the plasma, which is followed by polyclonal hypergammaglobulinemia. MT-I/IL-6 transgenics die between 12 to 20 weeks of age. Histologic examination of transgenic animals at different ages and after necropsy showed, as expected from previous studies of IL-6 disregulation in vivo, an increase in the number of megakaryocytes in the spleen and bone marrow and, at later stages, IgG plasmacytosis in the spleen, lymph nodes, and thymus. However, no plasma cell infiltration was detected in other organs. The distinguishing feature of MT-I/IL-6 transgenics is the development of a progressive kidney pathology, in which the initial membranous glomerulonephritis is followed by focal glomerulosclerosis and finally by extensive tubular damage that reproduces the damage observed in patients at terminal stages of multiple myeloma (myeloma kidney). The pathogenetic role of IL-6 overproduction and of the resulting serum protein overload in the kidney damage is discussed.
Reactive oxygen species rapidly increase endothelial ICAM-1 ability to bind neutrophils without detectable upregulation
Authors: Sellak H , Franzini E , Hakim J , Pasquier C 
Published: October 2019

Abstract
We compared the effects of phorbol 12-myristate 13-acetate (PMA) and thrombin with those of nonlytic concentrations of reactive oxygen species (ROS) generated by hypoxanthine (HX)-xanthine oxidase (XO) on the adhesion properties of human umbilical cord vein endothelial cells (HUVEC) to resting polymorphonuclear neutrophils (PMN). PMN adherence to HX-XO-treated HUVEC was increased approximately twofold to 2.5-fold relative to untreated HUVEC, both immediately and after 2 hours. It was not additive to that induced by PMA or thrombin stimulation of HUVEC. ROS-induced adherence was not due to platelet-activating factor (PAF) or P-selectin expression, as it was neither antagonized by BN52021 (PAF receptor antagonist) nor inhibited by anti-P-selectin monoclonal antibody (MoAb), contrary to the increased adhesion of PMA- and thrombin-stimulated HUVEC. PMN preincubated with mannose-6-P or N- acetylneuraminic acid (sialic acid), but not mannose or galactose-6-P, showed reduced adherence to ROS-treated HUVEC, suggesting that carbohydrate molecules were expressed on the latter and served as the ligand for the PMN L-selectin. Intercellular adhesion molecule (ICAM- 1), constitutively present on the surface of resting HUVEC, was involved in the PMN adherence to ROS-treated HUVEC, since this adherence was inhibited by anti-ICAM-1, anti-CD11a, anti-CD11b, and anti-CD18 MoAbs. A non-CD18, non-ICAM-1-dependent mechanism is also involved in this adherence, since effects of these MoAbs were not additive; moreover, combinations of anti-CD18 and anti-ICAM-1 MoAbs with mannose-6-P and sialic acid completely inhibited PMN adherence. The increased binding of PMN to HX-XO-exposed HUVEC observed here involved IC-AM-1, but was independent of its upregulation, and another non-ICAM-1-dependent mechanism, in which carbohydrates expressed on HUVEC recognize L-selectin on PMN.
Altered expression of plasminogen activator inhibitor type 1 in placentas from pregnant women with preeclampsia and/or intrauterine fetal growth retardation
Authors: Estelles A , Gilabert J , Keeton M , Eguchi Y , Aznar J , Grancha S , Espna F , Loskutoff DJ , Schleef RR 
Published: October 2019

Abstract
Elevated plasma levels of type 1 plasminogen activator inhibitor (PAI- 1) have been implicated in mediating the fibrin deposition and occlusive lesions that occur within the placental vasculature in preeclampsia (PE) and intrauterine growth retardation (IUGR). In this report we identify the cells within the normal-appearing villous tissue that are responsible for the local production of PAI-1 in women with PE and IUGR. Levels for another fibrinolytic inhibitor (ie, type 2 plasminogen activator inhibitor [PAI-2]) were determined for comparative purposes. Elevated levels of PAI-1 were detected in placenta extracts from PE/IUGR patients (121 +/- 38 ng/mg, n = 8) when compared with the levels in placenta extracts from normal women (43 +/- 17 ng/mg, n = 10) or women with IUGR but not PE (51 +/- 22 ng/mg, n = 11). Immunohistochemical analysis of paraffin sections showed an increased immunoreactivity for PAI-1 in the placental villous syncytiotrophoblasts from PE/IUGR women compared with the immunostaining of placental samples from the normal or IUGR group. In contrast, antigen levels and immunostaining for PAI-2 were reduced in the placentas harvested from not only the PE/IUGR women (209 +/- 144 ng/mg) but also the IUGR group (169 +/- 106 ng/mg) in comparison with the PAI-2 levels in normal placentas (535 +/- 98 ng/mg). To document that the increased immunoreactivity for PAI-1 in PE/IUGR syncytiotrophoblasts was mediated by an increased production of PAI-1 within these cells, in situ hybridization analysis was performed. A strong positive signal for PAI-1 mRNA in villous syncytiotrophoblasts from PE patients (n = 5) was obtained after 2 weeks of exposure to the NTB2 emulsion in comparison with the weak signal for PAI-1 mRNA that required a 10-week exposure of the normal placenta sections (n = 10). Northern blotting for PAI-1 mRNA showed that both transcripts (ie, 3.2 and 2.3 kb) were elevated in samples of two PE patients in comparison with the PAI-1 mRNA transcripts present in a normal placenta and an IUGR placental sample. These results show increased PAI-1 and mRNA levels in placentas from PE patients and raise the possibility that localized elevated levels of PAI-1 may play a role in the initiation of placental damage, as well as in the thrombotic complications associated with this disease.
Serum CD44 in malignant lymphoma: an association with treatment response
Authors: Ristamaki R , Joensuu H , Salmi M , Jalkanen S 
Published: October 2019

Abstract
CD44, a cell surface glycoprotein, is involved in lymphocyte trafficking from the blood to lymphatic tissues, and is of importance in dissemination of lymphoma. A variant form of CD44 that has additional amino acids in the common protein backbone (CD44v6) also seems to play a role in the metastatic dissemination of malignancies. We measured serum CD44 and CD44v6 in 34 patients with lymphoma and in healthy controls by dot blot assay. Small amounts of both CD44 (range, 10 to 80 ng/mL) and CD44v6 could be detected in sera of all controls. Serum CD44 was elevated in all patients with lymphoma before treatment (range, 70 to > 2,000 ng/mL, P < .0001), and CD44v6 was also slightly elevated. Serum CD44 levels correlated with response to treatment. Patients with complete response achieved similar CD44 levels as the controls, whereas those with progressive disease had increased serum CD44 levels. We conclude that both the standard and the variant form of CD44 are detectable in sera of healthy individuals, and that serum CD44 may be useful in monitoring treatment response in patients with lymphoma.