bluuebunny/crossref_metadata_2025_split huggingface dataset:
Oxidative stress is a critical mediator of the angiotensin II signal in human neutrophils: involvement of mitogen-activated protein kinase, calcineurin, and the transcription factor NF-κB
Authors: El Bekay Rajaa , Álvarez Moisés , Monteseirín Javier , Álba Gonzalo , Chacón Pedro , Vega Antonio , Martín-Nieto José , Jiménez Juan , Pintado Elízabeth , Bedoya Francisco J. , Sobrino Francisco 
Published: April 2003

AbstractNeutrophils are mobilized to the vascular wall during vessel inflammation. Published data are conflicting on phagocytic nicotinamide-adenine dinucleotide phosphate (NADPH) oxidase activation during the hypertensive state, and the capacity of angiotensin II (Ang II) to modulate the intracellular redox status has not been analyzed in neutrophils. We here describe that Ang II highly stimulates endogenous and extracellular O2- production in these cells, consistent with the translocation to the cell membrane of the cytosolic components of NADPH oxidase, p47phox, and p67phox. The Ang II–dependent O2- production was suppressed by specific inhibitors of AT1 receptors, of the p38MAPK and ERK1/2 pathways, and of flavin oxidases. Furthermore, Ang II induced a robust phosphorylation of p38MAPK, ERK1/2, and JNK1/2 (particularly JNK2), which was hindered by inhibitors of NADPH oxidase, tyrosine kinases, and ROS scavengers. Ang II increased cytosolic Ca2+ levels—released mainly from calcium stores—enhanced the synthesis de novo and activity of calcineurin, and stimulated the DNA-binding activity of the transcription factor NF-κB in cultured human neutrophils. Present data demonstrate for the first time a stimulatory role of Ang II in the activation of phagocytic cells, underscore the relevant role of ROS as mediators in this process, and uncover a variety of signaling pathways by which Ang II operates in human neutrophils.
Ontogeny and innate properties of neonatal dendritic cells
Authors: Sun Cheng-Ming , Fiette Laurence , Tanguy Myriam , Leclerc Claude , Lo-Man Richard 
Published: April 2003

AbstractWe investigated whether a developmental immaturity of the dendritic cells (DCs) compartment could contribute to the high susceptibility to infections observed in newborns. DCs are among the first cells to colonize the spleen, but the ontogeny of DC subsets follows distinct steps. At birth, plasmacytoid DCs and CD4-CD8α- DCs are found in the spleen, whereas CD8α+ and CD4+ DCs are not present. Then, the CD8α+ DC compartment quickly develops and reaches an adult size by day 7, whereas the CD4+ DC compartment slowly increases to become predominant by the age of 3 weeks. The production of interleukin (IL)–12p70 by DCs is particularly efficient after birth, reflecting the stronger capacity of the neonatal CD8α- DCs to secrete IL-12 compared with its adult counterpart. Like-wise, neonatal DCs produced type I and II interferons. In vivo, following microbial stimulation, up-regulation of major histocompatibility complexes (MHCs) and of costimulatory molecules on DCs was induced clearly showing the activation of neonatal DCs in the neonatal environment. Therefore, despite a markedly different DC subset composition in early life compared with the adult DC compartment, neonatal DCs are fully competent in their innate immune functions.
Fibrinogen gamma-chain splice variant γ′ alters fibrin formation and structure
Authors: Cooper Amy V. , Standeven Kristina F. , Ariëns Robert A. S. 
Published: March 2003

AbstractFibrinogen γA/γ′ results from alternative splicing of mRNA. This variant, which constitutes approximately 8% to 15% of plasma fibrinogen, contains FXIII and thrombin binding sites. Our objective was to investigate whether γA/γ′ differs in fibrin formation and structure from the more common variant γA/γA. Both variants were separated and purified by anion-exchange chromatography. Fibrin formation and clot structure of the variants and unfractionated fibrinogen were investigated by turbidity and scanning electron microscopy (SEM). Thrombin cleavage of fibrinopeptides was analyzed by high-performance liquid chromatography (HPLC). Turbidity analysis showed significantly altered polymerization rates and overall fiber thickness in γA/γ′ clots compared with γA/γA and unfractionated fibrinogen. This finding was consistent with a range of thrombin concentrations. HPLC demonstrated reduced rates of fibrinopeptide B (FpB) release from γA/γ′ fibrinogen compared with γA/γA. Delayed FpB release was associated with delayed lateral aggregation of protofibrils and significant differences were found on SEM, with γA/γ′ clots consisting of smaller diameter fibers and increased numbers of branch points compared with both γA/γA and unfractionated fibrinogen. These results demonstrate that the γA/γ′ splice variant of fibrinogen directly alters fibrin formation and structure, which may help to explain the increased thrombotic risk associated with this variant.
Gene expression profiling of plasma cells and plasmablasts: toward a better understanding of the late stages of B-cell differentiation
Authors: Tarte Karin , Zhan Fenghuang , De Vos John , Klein Bernard , Shaughnessy John 
Published: April 2003

AbstractPlasma cells (PCs), the end point of B-cell differentiation, are a heterogeneous cell compartment comprising several cell subsets from short-lived highly proliferative plasmablasts to long-lived nondividing fully mature PCs. Whereas the major transcription factors driving the differentiation of B cells to PCs were recently identified, the subtle genetic changes that underlie the transition from plasmablasts to mature PCs are poorly understood. We recently described an in vitro model making it possible to obtain a large number of cells with the morphologic, phenotypic, and functional characteristics of normal polyclonal plasmablastic cells (PPCs). Using Affymetrix microarrays we compared the gene expression profiles of these PPCs with those of mature PCs isolated from tonsils (TPCs) and bone marrow (BMPCs), and with those of B cells purified from peripheral blood (PBB cells) and tonsils (TBCs). Unsupervised principal component analysis clearly distinguished the 5 cell populations on the basis of their differentiation and proliferation status. Detailed statistical analysis allowed the identification of 85 PC genes and 40 B-cell genes, overexpressed, respectively, in the 3 PC subsets or in the 2 B-cell subsets. In addition, several signaling molecules and antiapoptotic proteins were found to be induced in BMPCs compared with PPCs and could be involved in the accumulation and prolonged survival of BMPCs in close contact with specialized stromal microenvironment. These data should help to better understand the molecular events that regulate commitment to a PC fate, mediate PC maintenance in survival niches, and could facilitate PC immortalization in plasma cell dyscrasias.
Unique gene expression profiles of human macrophages and dendritic cells to phylogenetically distinct parasites
Authors: Chaussabel Damien , Semnani Roshanak Tolouei , McDowell Mary Ann , Sacks David , Sher Alan , Nutman Thomas B. 
Published: April 2003

AbstractMonocyte-derived dendritic cells (DCs) and macrophages (Mϕs) generated in vitro from the same individual blood donors were exposed to 5 different pathogens, and gene expression profiles were assessed by microarray analysis. Responses to Mycobacterium tuberculosis and to phylogenetically distinct protozoan (Leishmania major, Leishmania donovani, Toxoplasma gondii) and helminth (Brugia malayi) parasites were examined, each of which produces chronic infections in humans yet vary considerably in the nature of the immune responses they trigger. In the absence of microbial stimulation, DCs and Mϕs constitutively expressed approximately 4000 genes, 96% of which were shared between the 2 cell types. In contrast, the genes altered transcriptionally in DCs and Mϕs following pathogen exposure were largely cell specific. Profiling of the gene expression data led to the identification of sets of tightly coregulated genes across all experimental conditions tested. A newly devised literature-based clustering algorithm enabled the identification of functionally and transcriptionally homogenous groups of genes. A comparison of the responses induced by the individual pathogens by means of this strategy revealed major differences in the functionally related gene profiles associated with each infectious agent. Although the intracellular pathogens induced responses clearly distinct from the extracellular B malayi, they each displayed a unique pattern of gene expression that would not necessarily be predicted on the basis of their phylogenetic relationship. The association of characteristic functional clusters with each infectious agent is consistent with the concept that antigen-presenting cells have prewired signaling patterns for use in the response to different pathogens.
The proapoptotic factor Nix is coexpressed with Bcl-xL during terminal erythroid differentiation
Authors: Aerbajinai Wulin , Giattina Mara , Lee Y. Terry , Raffeld Mark , Miller Jeffery L. 
Published: April 2003

Abstract
Transcriptional profiles of cultured primary human erythroid cells were examined to identify those genes involved in the control of erythroid growth during the terminal phase of maturation. Our in silico screening strategy indicated that a hypoxia-inducible proapoptotic member of the Bcl-2 gene family called Nix is expressed during erythropoiesis. We next performed Northern blot analyses and determined that the 1.4-kb Nix transcript is expressed at lower levels in erythroleukemia cells than reticulocytes. Polymerase chain reaction (PCR)–based transcriptional patterning confirmed the increased expression of Nix during human erythropoiesis with a pattern similar to that of Bcl-xL and glycophorin A and opposite that of Bcl-2. Western blot analyses revealed Nix protein levels that were lower than expected due to increased proteosomal degradation. The expression of Nix and Bcl-xL proteins decreased relative to glyceraldehyde-3-phosphate dehydrogenase (GAPDH) control on the removal of erythropoietin (EPO) from the culture medium. Immunocytochemical analyses demonstrated a similar perinuclear mitochondrial expression pattern for both proteins in hemoglobinized precursors. On the basis of these data, we propose that the proapoptotic factor Nix is a highly regulated effector of growth during terminal erythroid maturation.
Sensitivity toward tyrosine kinase inhibitors varies between different activating mutations of the FLT3 receptor
Authors: Grundler Rebekka , Thiede Christian , Miething Cornelius , Steudel Christine , Peschel Christian , Duyster Justus 
Published: April 2003

AbstractActivating mutations of FLT3 have been detected in patients with acute myeloid leukemia (AML). Two distinct types of FLT3 mutations are most common: internal tandem duplication (ITD) of sequences coding for the juxtamembrane domain and point mutations at codon 835 (Asp835) within the kinase domain. Both types of mutations constitutively activate the tyrosine kinase activity of FLT3 in experimental systems and result in factor-independent proliferation of Ba/F3 and 32D cells. Recently, novel mutations within the activation loop were identified in patients with AML: deletion of isoleucine 836 (Ile836del) and an exchange of isoleucine 836 to methionine plus an arginine insertion (Ile836Met+Arg). To examine whether the Ile836 mutations result in constitutive activation of the FLT3 receptor, we introduced both mutant FLT3 cDNAs transiently into HEK 293 cells. Both mutant FLT3 receptors were constitutively autophosphorylated in the absence of ligand and kinase activity led to constitutive activation of downstream signaling cascades as determined by activation of the STAT5 (signal transducer and activator of transcription 5) pathway. When stably expressed in the growth factor–dependent cell lines Ba/F3 and 32D, both deletion and insertion mutants led to factor-independent proliferation, indicating that both mutants have transforming capabilities. We then examined the sensitivity of the FLT3 ITD, FLT3 Asp835Tyr, and the novel FLT3 receptor mutants toward the kinase inhibitors AG1296, PKC412, and SU5614. We show that these FLT3 kinase inhibitors have distinct inhibitory potencies against different activating FLT3 receptor mutants. These results suggest that it may be useful to determine the exact kind of FLT3 mutation when applying receptor kinase inhibitors in clinical trials.
Single-chain antibody fragments derived from a human synthetic phage-display library bind thrombospondin and inhibit sickle cell adhesion
Authors: Watkins Nicholas A. , Du Lily M. , Scott J. Paul , Ouwehand Willem H. , Hillery Cheryl A. 
Published: April 2003

AbstractThe enhanced adhesion of sickle red blood cells (RBCs) to the vascular endothelium and subendothelial matrix likely plays a significant role in the pathogenesis of vaso-occlusion in sickle cell disease. Sickle RBCs have enhanced adhesion to the plasma and extracellular matrix protein thrombospondin-1 (TSP) under conditions of flow in vitro. In this study, we sought to develop antibodies that bind TSP from a highly diverse library of human single-chain Fv fragments (scFvs) displayed on filamentous phage. Following 3 rounds of phage selection of increasing stringency 6 unique scFvs that bound purified TSP by enzyme-linked immunosorbent assay were isolated. Using an in vitro flow adhesion assay, 3 of the 6 isolated scFvs inhibited the adhesion of sickle RBCs to immobilized TSP by more than 40% compared with control scFvs (P &lt; .001). Furthermore, scFv TSP-A10 partially inhibited sickle RBC adhesion to activated endothelial cells (P &lt; .005). Using TSP proteolytic fragments to map the binding site, we showed that 2 of the inhibitory scFvs bound an epitope in the calcium-binding domain or proximal cell-binding domain of TSP, providing evidence for the role of these domains in the adhesion of sickle RBCs to TSP. In summary, we have isolated a panel of scFvs that specifically bind to TSP and differentially inhibit sickle RBC adhesion to surface-bound TSP under flow conditions. These scFvs will be useful reagents for investigating the role of the calcium and cell-binding domains of TSP in sickle RBC adhesion.
A novel missense mutation in the γ-glutamylcysteine synthetase catalytic subunit gene causes both decreased enzymatic activity and glutathione production
Authors: Hamilton David , Wu Jian Hui , Alaoui-Jamali Moulay , Batist Gerald 
Published: April 2003

Abstractγ-Glutamylcysteine synthetase (γ-GCS) catalyzes the first and rate-limiting step in glutathione (GSH) biosynthesis: the adenosine triphosphate (ATP)–dependent ligation of glutamate and cysteine. γ-GCS consists of a catalytic (γ-GCSH) and modifier (γ-GCSL) subunit. Hereditary deficiency of γ-GCS has been reported in a small number of patients and is associated with low erythrocyte levels of γ-GCS and GSH leading to hemolytic anemia. Here we report a novel γ-GCSH mutation, isolated from the cDNA of 2 related patients diagnosed with γ-GCS deficiency. Each was found to be homozygous for a C&gt;T missense mutation at nucleotide 379, encoding for a predicted Arg127Cys amino acid change. Computerized structure modeling identified that the mutated amino acid lies within a cleft on the protein surface of γ-GCSH, and the border of this cleft was shown to contain Cys249, an evolutionarily conserved residue that has been proven to lie near the binding site of γ-GCSH. Transfection studies showed that the mutation is associated with decreased GSH production, and binding studies using purified recombinant protein showed that the mutant protein has markedly decreased enzymatic activity compared to wild type.
Common lymphoid progenitors rapidly engraft and protect against lethal murine cytomegalovirus infection after hematopoietic stem cell transplantation
Authors: Arber Caroline , BitMansour Andrew , Sparer Timothy E. , Higgins John P. , Mocarski Edward S. , Weissman Irving L. , Shizuru Judith A. , Brown Janice M. Y. 
Published: April 2003

AbstractLymphoid deficiency after allogeneic hematopoietic cell transplantation (HCT) results in increased susceptibility to infection; however, transplantation of mature lymphocytes frequently results in a serious complication known as graft-versus-host disease (GVHD). Here we demonstrate in mice that both congenic as well as allogeneic transplantation of low numbers of highly purified common lymphoid progenitors (CLPs)—a rare population of lymphoid-lineage-committed bone marrow cells—accelerates immune reconstitution after lethal irradiation and rescue with hematopoietic stem cells (HSCs). After congenic transplantation, 3 × 103 CLPs protected against murine cytomegalovirus (MCMV) infection at a level roughly equivalent to 107 unfractionated lymph node cells. In the allogeneic model of matched unrelated donor HSC transplantation, cotransplantation of 3 × 103 CLPs protected thymus-bearing as well as thymectomized hosts from MCMV infection and attenuated disease severity. Immunohistochemistry in combination with antibody depletion of T and natural killer (NK) cells confirmed that CLP-derived as well as residual host lymphocytes contribute to antiviral protection. Importantly, transplantation of allogeneic CLPs provided a durable antiviral immunity without inducing GVHD. These data support the potential for composing grafts with committed progenitors to reduce susceptibility to viral infection following HCT.
Phase 2 study of rituximab in the treatment of cladribine-failed patients with hairy cell leukemia
Authors: Nieva Jorge , Bethel Kelly , Saven Alan 
Published: April 2003

Abstract
Hairy cell leukemia (HCL) is an indolent B-cell neoplasm, strongly expressing CD20. Despite initial very high response rates following cladribine, many patients (pts) ultimately relapse. Having relapsed after prior treatment with cladribine, 24 HCL pts (21 male, 3 female) with a median age of 53.5 years were treated with rituximab at 375 mg/m2 intravenously weekly for 4 weeks. Of the pts, 3 (13%) achieved complete remissions and 3 (13%), partial responses. Thus, 6 (25%) of 24 pts achieved a response following rituximab. At a median follow-up of 14.6 months, 2 responders have relapsed; median time to relapse was not yet reached. The only grade III or IV toxicities demonstrated were culture-negative febrile neutropenia, transient and reversible disseminated intravascular coagulation related to rituximab administration, and a diverticular abscess, each in single patients. Of 18 nonresponders, 9 pts subsequently received other treatments; 5 pts were retreated with cladribine, 3 underwent splenectomy, and 1 received pentostatin. Follow-up data are available on 7 of these 9 patients; all 7 patients achieved improvements in hematologic parameters. Rituximab, administered at this dose and schedule, has only modest single-agent activity in cladribine-failed HCL patients when compared with other agents active in this disease.
The minor histocompatibility antigen HA-3 arises from differential proteasome–mediated cleavage of the lymphoid blast crisis (Lbc) oncoprotein
Authors: Spierings Eric , Brickner Anthony G. , Caldwell Jennifer A. , Zegveld Suzanne , Tatsis Nia , Blokland Els , Pool Jos , Pierce Richard A. , Mollah Sahana , Shabanowitz Jeffrey , Eisenlohr Laurence C. , Van Veelen Peter , Ossendorp Ferry , Hunt Donald F. , Goulmy Els , Engelhard Victor H. 
Published: April 2003

AbstractMinor histocompatibility (H) antigens crucially affect the outcome of human leukocyte antigen (HLA)–identical allogeneic stem cell transplantation (SCT). To understand the basis of alloimmune responses against minor H antigens, identification of minor H peptides and their antigenicity-determining mechanisms is essential. Here we report the identification of HA-3 and its encoding gene. The HA-3 peptide, VTEPGTAQY (HA-3T), is encoded by the lymphoid blast crisis (Lbc) oncogene. We thus show for the first time that a leukemia-associated oncogene can give rise to immunogenic T-cell epitopes that may have participated in antihost and antileukemic alloimmune responses. Genotypic analysis of HA-3- individuals revealed the allelic counterpart VMEPGTAQY (HA-3M). Despite the lack of T-cell recognition of HA-3- cells, the Thr→Met substitution had only a modest effect on peptide binding to HLA-A1 and a minimal impact on recognition by T cells when added exogenously to target cells. This substitution did not influence transporter associated with antigen processing (TAP) transport, but, in contrast to the HA-3T peptide, HA-3M is destroyed by proteasome-mediated digestion. Thus, the immunogenicity of minor H antigens can result from proteasome-mediated destruction of the negative allelic peptide.
Impaired thymic negative selection causes autoimmune graft-versus-host disease
Authors: Teshima Takanori , Reddy Pavan , Liu Chen , Williams Debra , Cooke Kenneth R. , Ferrara James L. M. 
Published: April 2003

AbstractAnimal models with impaired thymic negative selection do not always cause autoimmune diseases despite the development of an autoreactive T-cell repertoire. We investigated the requirements for the de velopment of systemic autoimmune disease by using bone marrow chimeras that lacked expression of major histocompatibility complex (MHC) class II on thymic antigen-presenting cells (APCs), leading to impaired negative selection. We found that impaired negative selection mediated by absence of MHC class II, but not MHC class I, permitted the development of systemic autoimmune disease that is indistinguishable from acute graft-versus-host disease (GVHD). Thymectomy prevented disease, confirming the causal association of the thymus with its development. Adoptive transfer of CD4+ T cells caused GVHD in secondary hosts only when they were irradiated, and cotransfer of peripheral CD4+ and CD8+ T cells from naive mice prevented the disease. These results demonstrate that impaired thymic negative selection can cause lethal autoimmune disease indistinguishable from acute GVHD in the context of a proinflammatory milieu when peripheral regulatory mechanisms are absent.
Successful treatment of murine β-thalassemia using in vivo selection of genetically modified, drug-resistant hematopoietic stem cells
Authors: Persons Derek A. , Allay Esther R. , Sawai Nobukuni , Hargrove Phillip W. , Brent Thomas P. , Hanawa Hideki , Nienhuis Arthur W. , Sorrentino Brian P. 
Published: April 2003

AbstractSuccessful gene therapy of β-thalassemia will require replacement of the abnormal erythroid compartment with erythropoiesis derived from genetically corrected, autologous hematopoietic stem cells (HSCs). However, currently attainable gene transfer efficiencies into human HSCs are unlikely to yield sufficient numbers of corrected cells for a clinical benefit. Here, using a murine model of β-thalassemia, we demonstrate for the first time that selective enrichment in vivo of transplanted, drug-resistant HSCs can be used therapeutically and may therefore be a useful approach to overcome limiting gene transfer. We used an oncoretroviral vector to transfer a methylguanine methyltransferase (MGMT) drug-resistance gene into normal bone marrow cells. These cells were transplanted into β-thalassemic mice given nonmyeloablative pretransplantation conditioning with temozolomide (TMZ) and O6-benzylguanine (BG). A majority of mice receiving 2 additional courses of TMZ/BG demonstrated in vivo selection of the drug-resistant cells and amelioration of anemia, compared with untreated control animals. These results were extended using a novel γ-globin/MGMT dual gene lentiviral vector. Following drug treatment, normal mice that received transduced cells had an average 67-fold increase in γ-globin expressing red cells. These studies demonstrate that MGMT-based in vivo selection may be useful to increase genetically corrected cells to therapeutic levels in patients with β-thalassemia.