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R E Ware

Publications and source records attributed to R E Ware.

At least 73 records · Page 4Linked to original sources
PubMed1995

Characterization of human CD7 transgenic mice.

CD7 is a 40-kDa transmembrane glycoprotein member of the lg gene superfamily expressed on most peripheral blood T lymphocytes and NK cells. CD7 is also expressed on myeloid, NK, B, and T cell precursors during adult hematopoiesis. Because Thy-1 is absent in human thymocytes and peripheral blood T cells and shows structural similarities to the human CD7 gene, we have suggested that human CD7 may be a functional homologue in humans of mouse Thy-1. To study the tissue-specific expression of the CD7 gene utilizing its own promoter, we constructed transgenic mice that contained both the coding and flanking regions of the human CD7 gene. We found that human CD7 was expressed in transgenic mice in T, B, NK, and myeloid lineages and was induced with T cell activation. Unlike the expression of CD7 in humans, the CD7 transgene was present on mature B lymphocytes and macrophages. Like mouse Thy-1, transgenic human CD7 was expressed in immature and mature T cells and in Sca-1+ bone marrow mononuclear cells. Unlike mouse Thy-1, the human CD7 transgene was not expressed in mouse brain or fibroblasts. The human CD7 transgene was expressed during fetal development before mouse Thy-1 in fetal liver mononuclear cells. Expression of the human CD7 transgene did not alter mouse thymopiesis or Thy-1 expression. Taken together, these data demonstrated that the CD7 transgene contained sufficient regulatory regions to direct hematopoietic expression and mitogenic induction. The pattern of CD7 transgene expression more closely resembled that of CD7 in humans than that of mouse Thy-1.

Animals↗
PubMed1995

Immunophenotypic analysis of reticulocytes in paroxysmal nocturnal hemoglobinuria.

The hematologic disorder paroxysmal nocturnal hemoglobinuria (PNH) occurs following an acquired somatic mutation in the Piga gene within a bone marrow stem cell. The progeny of this mutated cell cannot synthesize glycosylphosphatidylinositol (GPI) anchors, with a resultant deficiency in surface expression of all GPI-linked proteins. The protean clinical manifestations of PNH presumably result from the deficiency of these GPI-linked surface proteins. To explain the observation that neutrophils are affected at a significantly higher percentage than circulating erythrocytes and to analyze the proliferative rates of erythroid production in PNH, we studied 25 patients using flow cytometry. The fluorescent dye thiazole orange was used to detect reticulocytes, and CD59 monoclonal antibody was used to identify GPI-deficient cells. In contrast to the mature circulating erythrocytes, the percentage of abnormal reticulocytes was similar to the percentage of affected neutrophils. However, the vast majority of reticulocytes was completely GPI-deficient, ie, were type III cells, even in patients with only modest numbers of circulating type III erythrocytes. In addition, greater than 5% type II reticulocytes were identified in only 3 patients, although greater than 5% type II mature erythrocytes were identified in 10 of 25 patients. The results show that the erythroid and neutrophil bone marrow precursors have an equivalent proliferative advantage in PNH. The data also have important implications for the origin of type-II erythrocytes in PNH.

Adolescent↗
PubMed1995

Phenotypic and functional analysis of lymphocytes in paroxysmal nocturnal hemoglobinuria.

The hematologic disorder paroxysmal nocturnal hemoglobinuria (PNH) arises from a somatic mutation within the Piga gene important for the biosynthesis of glycosylphosphatidylinositol (GPI) anchors. The PNH defect has been identified in all cells of the myeloerythroid lineage, but involvement of the lymphoid lineage in PNH is more controversial. We therefore analyzed lymphocytes from 22 patients with PNH to characterize phenotypically the GPI-deficient population, and to investigate the functional consequences of GPI deficiency. GPI-deficient T lymphocytes, B lymphocytes, and NK cells were identified, but at a lower percentage than granulocytes and erythrocytes. CD8+ lymphocytes were significantly more affected than CD4+ T cells, and CD45RA+ lymphocytes were significantly more affected than CD45RO+ cells. Proliferation assays demonstrated that lymphocytes from PNH patients, either unfractionated or purified GPI-deficient cells, responded normally to in vitro stimuli. When stimulated with phytohemagglutinin (PHA), naive CD45RA+ GPI-deficient T lymphocytes acquired the memory CD45RO+ phenotype. In addition, GPI-deficient T lymphocytes had a relative growth advantage as compared to normal T cells. The results demonstrate that PNH involves the lymphoid as well as the myeloerythroid lineage, and therefore arises from a totipotent bone marrow stem cell. The in vitro growth advantage of GPI-deficient lymphocytes in PNH may have important implications for the pathogenesis of some puzzling clinical aspects of PNH, including predominance of the PNH clone, defective hematopoiesis, and leukemogenesis.

Adolescent↗
PubMed1995

Urticarial vasculitis: an autoimmune disorder following therapy for Hodgkin's disease.

Immunological abnormalities have been described in patients with Hodgkin's disease, both associated with the malignancy itself and occurring secondary to therapy. These abnormalities often manifest as an immunodeficiency state, but can also present as immune dysregulation and autoimmune disease. We report two young patients with Hodgkin's disease who, following successful therapy, developed urticarial vasculitis (UV), a form of cutaneous autoimmune vasculitis. Both patients also had systemic symptoms including fever, an elevated erythrocyte sedimentation rate and serum copper, and abnormal in vitro studies of lymphocyte enumeration and proliferation. Distinguishing UV from recurrent Hodgkin's disease was especially difficult in one patient, and was possible only by lymph node biopsy. One patient has responded well to immunosuppressive therapy, while the other, who has more profound immune dysfunction, has developed a chronic autoimmune disorder. UV may thus occur in patients after therapy for Hodgkin's disease; we hypothesize that immune dysregulation, either associated with the malignancy or resulting from therapy, is important in the pathogenesis of this autoimmune process.

Adolescent↗
PubMed1995

Pitfalls in newborn hemoglobinopathy screening: failure to detect beta(+)-thalassemia.

Although universal newborn screening can reliably identify all infants with sickle cell hemoglobinopathies, the initial screening result must not be considered the definitive diagnosis. We describe 23 infants whose screening phenotype was FS or FC but whose true phenotype included hemoglobin A, establishing a definitive diagnosis of hemoglobin S or hemoglobin C in combination with beta(+)-thalassemia. Higher than expected hemoglobin concentrations or lower than expected mean erythrocyte volumes should suggest concurrent beta(+)-thalassemia.

Anemia, Sickle Cell↗
PubMed1995

Emergency department visits by children with sickle hemoglobinopathies: factors associated with hospital admission.

Children with sickle cell disease frequently present to the emergency department (ED) for evaluation of fever or management of pain. We retrospectively analyzed all ED visits by children with sickle hemoglobinopathies during 1990, excluding those for trauma. Of 146 visits by 56 children, 73 (50%) were classified as "Painful Events," 43 (29%) as "Febrile Events," 20 (14%) as "Painful and Febrile Events," and 10 (7%) as "Other." Hospital admission occurred for 42% of Painful Events versus 70% of Febrile Events (P = 0.008) and 85% of Painful and Febrile Events (P = 0.002). For Painful Events, location of pain was not associated with hospitalization, but pain less than 24 hours in duration before the ED visit (P = 0.002) and administration of intravenous fluids and analgesia (P = 0.001) were associated with admission. For children evaluated for Febrile Events, age less than six years (P = 0.016) and maximum temperature greater than 39 degrees C (P = 0.011) were associated with subsequent hospitalization, but total white blood cell count and absolute neutrophil count were not. For Painful and Febrile Events, pain less than 24 hours (P = 0.029) was associated with hospital admission, but age, maximum temperature, white blood cell count, and absolute neutrophil count were not. Although prospective studies are needed to validate these data, the identification of factors predictive of hospital admission should expedite ED care to sickle cell patients.

Adolescent↗
PubMed1995

Molecular basis of spectrin deficiency in beta spectrin Durham. A deletion within beta spectrin adjacent to the ankyrin-binding site precludes spectrin attachment to the membrane in hereditary spherocytosis.

We describe a spectrin variant characterized by a truncated beta chain and associated with hereditary spherocytosis. The clinical phenotype consists of a moderate hemolytic anemia with striking spherocytosis and mild spiculation of the red cells. We describe the biochemical characteristics of this truncated protein which constitutes only 10% of the total beta spectrin present on the membrane, resulting in spectrin deficiency. Analysis of reticulocyte cDNA revealed the deletion of exons 22 and 23. We show, using Southern blot analysis, that this truncation results from a 4.6-kb genomic deletion. To elucidate the basis for the decreased amount of the truncated protein on the membrane and the overall spectrin deficiency, we show that (a) the mutated gene is efficiently transcribed and its mRNA abundant in reticulocytes, (b) the mutant protein is normally synthesized in erythroid progenitor cells, (c) the stability of the mutant protein in the cytoplasm of erythroblasts parallels that of the normal beta spectrin, and (d) the abnormal protein is inefficiently incorporated into the membrane of erythroblasts. We conclude that the truncation within the beta spectrin leads to inefficient incorporation of the mutant protein into the skeleton despite its normal synthesis and stability. We postulate that this misincorporation results from conformational changes of the beta spectrin subunit affecting the binding of the abnormal heterodimer to ankyrin, and we provide evidence based on binding assays of recombinant synthetic peptides to inside-out-vesicles to support this model.

Ankyrins↗
PubMed1994

Novel mechanisms of brequinar sodium immunosuppression on T cell activation.

Brequinar sodium (BQR) is a novel immunosuppressive agent that acts by inhibiting the activity of dihydroorotate dehydrogenase, the fourth enzyme in the de novo pyrimidine biosynthetic pathway. The activity of BQR as an immunosuppressive agent is believed to be inhibition of antigen-induced lymphocyte proliferation through inhibition of DNA and RNA synthesis. BQR, therefore, has a different mechanism of action than cyclosporine and may potentiate the immunosuppressive effects of cyclosporine. In this study, we determined the effect of BQR on peripheral blood mononuclear cell (PBMC) activation in a series of in vitro culture systems. In these studies, BQR inhibited PHA-stimulated activation in a dose-dependent fashion beginning at 10(-6) M. The immunosuppressive effect of BQR was similar in magnitude to cyclosporine. Proliferation assays suggested an additive immunosuppression by the combination of BQR and cyclosporine. Similar inhibition of CD2-stimulated or CD3-stimulated activation of PBMC was found. The mechanisms of action of BQR were complex. BQR inhibited interleukin 2 protein production in response to mitogen stimulation. Cell surface interleukin 2 receptor expression was inhibited by BQR. BQR inhibited cell cycle progression, preventing progression from G0/G1 into S and G2 + M phases. BQR had no effect on induction of transcripts for the interleukin 2 receptor, but markedly inhibited the production of transcripts for interleukin 2. Thus, our studies indicate that BQR exerts a potent immunosuppression on mitogen-induced PBMC activation through multiple mechanisms. Consequently, BQR may be an effective agent for immunosuppression in organ transplantation or inflammatory diseases.

Biphenyl Compounds↗
PubMed1994

Mutations within the Piga gene in patients with paroxysmal nocturnal hemoglobinuria.

Paroxysmal nocturnal hemoglobinuria (PNH) is an acquired clonal hematologic disorder with multiple and varied clinical manifestations. The biochemical defect in PNH resides in the incomplete enzymatic assembly of glycosylphosphatidylinositol (GPI) anchors used for surface protein attachment. In all patients tested thus far, the defect is at the level of N-acetylglucosamine attachment to phosphatidylinositol (complementation class A defect). A human cDNA, Piga, that repairs cell lines with the class A defect has been recently cloned, making Piga a candidate gene for PNH. In the current study, using highly purified GPI-deficient granulocytes, we have performed Northern blot and reverse transcriptase polymerase chain reaction (RT-PCR) analysis of Piga in four patients with PNH. In each case, we have identified a mutation in the Piga coding sequence: three frameshift mutations were found, and a single nucleotide substitution (missense) mutation was identified. Our results provide convincing evidence that alterations in the Piga gene are responsible for PNH.

Adult↗
PubMed1994

Glycosyl-phosphatidylinositol anchor synthesis in paroxysmal nocturnal hemoglobinuria: partial or complete defect in an early step.

The defect in the biosynthesis of the glycosyl-phosphatidyl inositol (GPI) anchor in paroxysmal nocturnal hemoglobinuria (PNH) appears to be in the initial steps. In biosynthetic studies using [3H]mannose, abnormal granulocytes of eight patients, and B lymphocytes transformed by Epstein-Barr virus of six different patients synthesized dolichyl phosphoryl mannose, but little or no later mannosylated intermediates. When fused with murine cell lines known to be deficient at different biosynthetic steps of the GPI anchor, the GPI-anchor-deficient granulocytes of 21/21 patients and lymphocytes from 6/6 patients complemented all murine cell lines except those of class A; cells of this class are not able to add N-acetylglucosamine to phosphatidylinositol. These studies indicate that the defect in GPI-anchor synthesis in PNH is early in the pathway, and is the same as that of class A mutants, but may be partial in some patients, resulting in the production of small amounts of mannosylated intermediates.

Antigens, CD↗
PubMed1994

Elevated numbers of gamma-delta (gamma delta+) T lymphocytes in children with immune thrombocytopenic purpura.

Immune thrombocytopenic purpura (ITP) in childhood is a heterogeneous clinical disorder characterized by immune-mediated platelet destruction. Although generally considered to involve autoreactive B lymphocytes which produce antiplatelet antibodies, there is increasing evidence that T lymphocytes also play an important role in this autoimmune process. We studied 11 children with acute ITP and 19 children with chronic ITP and observed elevated numbers of TCR gamma delta+ T lymphocytes in several patients. In the three children with the highest elevations (TCR gamma delta+/CD3+ percentage ranging from 37.8 to 48.1% at initial evaluation), the expanded cell population exclusively expressed the surface V delta 2/V gamma 9 heterodimer and had enhanced in vitro proliferation to mycobacterial extracts and IL-2. Analysis of the nucleotide sequences used by these TCR gamma delta+ cells demonstrated a diverse set of VDDJC gene rearrangements, indicating polyclonal expansion of cells reminiscent of a superantigen response. There was a close correlation between the number of TCR gamma delta+ T lymphocytes and the degree of thrombocytopenia in each patient. TCR gamma delta+ T lymphocytes may be important in the pathogenesis of immune-mediated platelet destruction in some children with ITP.

Adolescent↗
PubMed1994

Hepatosplenic T-cell lymphoma: an unusual case of a gamma delta T-cell lymphoma with a blast-like terminal transformation.

We describe a case of a middle-aged women who presented with anemia and mild hepatosplenomegaly and who was found to have an unusual peripheral T-cell lymphoma with only subtle morphologically abnormal but mature-appearing cells noted in the blood and bone marrow. Less than 2 years after diagnosis the patient presented with an increasing white blood cell count to 26 x 10(9)/L, and numerous blasts were noted in the periphery. Flow cytometry studies showed cells with an unusual T-cell phenotype expressing the gamma delta T-cell receptor and restricted expression of the V delta 1 but not the V delta 2 protein, indicating the clonal nature of the proliferation. A clonal T-cell receptor gene rearrangement was seen with a V delta 1 probe. The patient died and was found at autopsy to have extensive hepatic sinusoidal infiltration by abnormal cells. The histopathologic, immunophenotypic, and molecular findings are those of "hepatosplenic T-cell lymphoma." In spite of the striking morphologic change during the course of the patient's disease the same phenotype and clonal rearrangement were found both at initial diagnosis and during terminal phase, indicating that this change represented a blast-like transformation of the patient's original lymphoproliferative disorder.

Female↗
PubMed1994

Hemoglobin C disease in infancy and childhood.

We reviewed the clinical course, physical findings, and hematologic values in 16 pediatric patients with hemoglobin C disease, all but one identified by a newborn hemoglobinopathy screening program. The patients had a few symptoms or physical findings. Height and weight percentiles were normal. Patients typically had a mild hemolytic anemia characterized by microcytosis and target cells.

Adolescent↗
PubMed1994

Gallbladder sludge in children with sickle cell disease.

Ultrasonographic records of 75 children with sickle cell disease and hepatobiliary symptoms were reviewed. Seventeen had gallbladder sludge, nine with concurrent stones and eight with sludge alone. Because all the children eventually had gallstones, we recommend that elective cholecystectomy be performed on patients with gallbladder sludge.

Anemia, Sickle Cell↗
PubMed1994

Acute splenic sequestration in the absence of palpable splenomegaly.

PURPOSE: Acute splenic sequestration in children with sickle hemoglobinopathies is a medical emergency characterized by splenomegaly, anemia, and thrombocytopenia. PATIENTS AND METHODS: We describe a young girl with hemoglobin SC disease who experienced an acute splenic sequestration without palpable splenomegaly. CONCLUSIONS: We emphasize the need for a high index of suspicion for splenic sequestration and the utility of ultrasonography in documenting splenomegaly.

Acute Disease↗
PubMed1993

Phenotypic and clonal analysis of T lymphocytes in childhood immune thrombocytopenic purpura.

In an attempt to identify and characterize T-lymphocyte immunoregulatory abnormalities in immune thrombocytopenic purpura (ITP), we have performed phenotypic and clonal analysis on peripheral T lymphocytes from 23 children with ITP. Quantitation of lymphocyte subpopulations showed that children with acute ITP had higher numbers of CD45RA+ and lower numbers of CD45RO+ T cells than children with chronic ITP or controls, but these differences may be age related. Analysis of T-cell receptor variable beta gene usage identified 2 boys with chronic ITP and elevated numbers of V beta 8+ T cells. Eight T-cell clones were established (6 CD4+, 4B4+ helper-inducer lines and 2 CD8+ lines) that showed in vitro proliferation against allogeneic platelets. The addition of autologous antigen-presenting cells enhanced the proliferation of six clones, but not for two clones that coexpressed natural killer (NK) markers. Four of seven positive clones also had measurable interleukin (IL)-2 secretion following platelet stimulation, providing further evidence for T-cell reactivity. Our results provide the first evidence that patients with ITP may have platelet-reactive T lymphocytes identifiable at the clonal level, supporting the hypothesis that autoreactive peripheral T lymphocytes may mediate or participate in the pathogenesis of this disorder.

Acute Disease↗