A preliminary investigation into the action of anagrelide: thrombopoietin-c-Mpl receptor interactions.

Objective: Many studies have validated the clinical efficacy of anagrelide to reduce platelet counts in thrombocythemic conditions. With the ability to support human megakaryopoiesis in vitro using thrombopoietin (TPO), specific investigation of changes in platelet levels can be carried out in human systems. Using CD34(+) stem cells and murine BaF3 cells transfected with the human or murine TPO receptor, c-Mpl (BaF3mpl), the effect of anagrelide on cell differentiation, proliferation, and signaling was examined in the presence of TPO.

A randomized controlled trial of darbepoetin alfa administered as a fixed or weight-based dose using a front-loading schedule in patients with anemia who have nonmyeloid malignancies.

Background: The effect of using fixed versus weight-based doses for erythropoietic agents has not been reported previously. To investigate this issue, the authors conducted a randomized Phase II study of darbepoetin alfa administered as either a fixed dose or a weight-based dose using an accelerated correction and maintenance dosing regimen (front-loading).

Methods: During the correction phase, patients with anemia (hemoglobin < 11.

Interleukin-4 elicits apoptosis of developing mast cells via a Stat6-dependent mitochondrial pathway.

Objective: The aim of this study was to assess the effects of interleukin-4 and signal transducer and activator of transcription (Stat)-6 on IL-3+SCF-induced mast cell development.

Patients And Methods: Unseparated mouse bone marrow cells were cultured in IL-3+SCF, giving rise to mast cells and monocytes/macrophages. The addition of IL-4, the use of Stat6-deficient bone marrow cells, and expression of a constitutively active Stat6 mutant were employed to assess the effects of IL-4 and Stat6 on cell viability, proliferation, and differentiation.

Tumor necrosis factor-alpha expressed constitutively in erythroid cells or induced by erythropoietin has negative and stimulatory roles in normal erythropoiesis and erythroleukemia.

Binding of erythropoietin (EPO) to its receptor (EPOR) on erythroid cells induces the activation of numerous signal transduction pathways, including the mitogen-activated protein kinase Jun-N-terminal kinase (JNK). In an effort to understand the regulation of EPO-induced proliferation and JNK activation, we have examined the role of potential autocrine factors in the proliferation of the murine erythroleukemia cell line HCD57. We report here that treatment of these cells with EPO induced the expression and secretion of tumor necrosis factor alpha (TNF-alpha).

Secretion of a unique peptide from interleukin-2-stimulated natural killer cells that induces endomitosis in immature human megakaryocytes.

When interleukin-2 (IL-2) was added to immature, low-ploidy (greater than 80% 2N+4N) megakaryocytes generated in IL-3 and stem cell factor (SCF)-containing liquid cultures of blood mononuclear cells highly enriched in hematopoietic progenitors, a 2- to 6-fold increase in the absolute number of polyploid (more than 8N) megakaryocytes was noted. This effect was found to be indirect and was mediated through natural killer (NK) cells that constitute the major lymphoid cell contaminating day 6 megakaryocyte cell populations. IL-2 had no effect on megakaryocytes generated from CD34(+) cells stimulated with IL-3 and SCF.

Effects of recombinant human thrombopoietin on megakaryocyte colony formation and megakaryocyte ploidy by human CD34+ cells in a serum-free system.

In serum-free cultures of human CD34 cells, recombinant human thrombopoietin (TPO) induced megakaryocyte colony formation a dose-dependent fashion that was further enhanced by the presence of interleukin-3 (IL-3) and stem cell factor (SCF), but not by IL-6, IL-11 or erythropoietin. TPO gave rise to much smaller colonies and at an earlier time than IL_3, indicating that TPO affects predominantly more mature megakaryocytic progenitors. In liquid cultures.

Regulation and specificity of MNDA expression in monocytes, macrophages, and leukemia/B lymphoma cell lines.

The expression of the human myeloid cell nuclear differentiation antigen (MNDA) was observed specifically in cells of the granulocyte-macrophage lineage in our earlier reports. The specificity of MNDA expression for cells in the granulocyte-macrophage lineage was reexamined in cell lines established from patients with Philadelphia chromosome-positive chronic myeloid leukemia. Cell lines that expressed MNDA exhibited myeloid cell features and granulocyte or monocyte differentiation could be induced in vitro, while cell lines exhibiting properties of very early stage cells or multipotential cells did not express MNDA.

Replication and endoreplication in developing megakaryocytes in vitro.

To study the relation in time between replication and endoreplication and the relation between appearance of platelet-specific proteins and endoreplication in maturing megakaryocytes, peripheral blood mononuclear cells highly enriched in hematopoietic progenitors were cultured in liquid cultures and plasma clots in the presence of either interleukin-3 (IL-3) and stem cell factor (SCF) or medium conditioned by blood mononuclear cells stimulated by phytohemagglutinin (PHA). In plasma clots, megakaryocytic (MK) colonies appeared first on day 5 and reached a maximum by day 8, whereas the number of cells per colony increased until day 10, indicating that there was a single wave of MK colony formation. In liquid cultures, the first immunologically recognizable megakaryocytes appeared on day 5 and expressed GPIIb/IIIa and thrombospondin only, but all other platelet-specific protein markers appeared within 24 hours.

The human myeloid cell nuclear differentiation antigen gene is one of at least two related interferon-inducible genes located on chromosome 1q that are expressed specifically in hematopoietic cells.

We have previously shown that the human myeloid cell nuclear differentiation antigen (MNDA) is expressed at both the antigen and mRNA levels specifically in human monocytes and granulocytes and earlier stage cells in the myeloid lineage. A 200 amino acid region of the MNDA is strikingly similar to a region in the proteins encoded by a family of interferon-inducible mouse genes, designated Ifi-201, Ifi-202, Ifi-203, etc, that are not regulated in a cell- or tissue-specific fashion. However, a new member of the Ifi-200 gene family, D3, is induced in mouse mononuclear phagocytes but not in fibroblasts by interferon.

Aplastic anemia and pure red cell aplasia.

The role of known hematopoietic growth factors in the pathogenesis of aplastic anemia and congenital hypoplastic anemia has been extensively studied and no evidence has been obtained that deficiency of these factors contributes to the hypoproliferative state in these disorders. Clonal hematopoiesis seems to be present at least in a small percentage of cases of aplastic anemia, a finding that needs further investigation. Androgens were shown to be beneficial only for women with aplastic anemia treated with antilymphocyte globulin.

Interferon alpha selectively affects expression of the human myeloid cell nuclear differentiation antigen in late stage cells in the monocytic but not the granulocytic lineage.

The human myeloid cell nuclear differentiation antigen (MNDA) is expressed constitutively in cells of the myeloid lineage, appearing in myeloblast cells in some cases of acute myeloid leukemia and consistently being detected in promyelocyte stage cells as well as in all later stage cells including peripheral blood monocytes and granulocytes. The human myeloid leukemia cell lines, HL-60, U937, and THP-1, express similar levels of immunochemically detectable MNDA. Although, the level of MNDA mRNA in primary monocytes is very low it was up-regulated at 6 h following the addition of interferon alpha.

Chronic administration of transforming growth factor-beta suppresses erythropoietin-dependent erythropoiesis and induces tumour necrosis factor in vivo.

Transforming growth factor beta is a known inhibitor of the proliferation and differentiation of early haematopoietic progenitors but has no effect on mature erythroid cells in vitro. Mice injected with rhTGF beta 1 exhibited severe and progressive suppression of erythropoiesis manifested by a decline of reticulocyte count, marrow erythroblasts and marrow and spleen CFU-E, which could be prevented by administration of erythropoietin. This suppression of erythropoiesis was associated with the appearance of tumour necrosis factor in the blood, development of pronounced cachexia and depression of serum erythropoietin levels.

Granulocyte/macrophage colony-stimulating factor stimulates human polymorphonuclear leukocytes to produce interleukin-8 in vitro.

Interleukin-8 (IL-8) is a potent chemotactic factor for polymorphonuclear leukocytes (PMN). Here we examine whether PMN synthesize and release IL-8 in response to stimulation by selected inflammatory cytokines. PMN isolated from normal heparinized peripheral human blood were incubated in RPMI culture medium at 37 degrees C in 5% CO2, with and without granulocyte/macrophage colony-stimulating factor (GM-CSF).

Case report: granulocyte colony-stimulating factor overcomes severe neutropenia of large granular lymphocytosis.

Large granular lymphocytosis (LGL) is characterized by enhanced proliferation of T lymphocytes that have antibody-dependent cell-mediated cytotoxicity or natural killer cell activity and that often produce severe cytopenias, including neutropenia. When a 68-year-old man with seropositive rheumatoid arthritis and severe neutropenia was examined, he was found to have LGL with a T cell gene rearrangement, indicating the presence of a clonal population of T lymphocytes. The patient was admitted with a fever of 102 degrees F and a nonhealing ulcer over the right tibia.

Polycythemia vera. II. Hypersensitivity of bone marrow erythroid, granulocyte-macrophage, and megakaryocyte progenitor cells to interleukin-3 and granulocyte-macrophage colony-stimulating factor.

Polycythemia vera (PV) is a clonal disease of the hematopoietic stem cell characterized by a hyperplasia of marrow erythropoiesis, granulocytopoiesis, and megakaryocytopoiesis. We previously reported that highly purified PV blood burst-forming units-erythroid (BFU-E) are hypersensitive to recombinant human interleukin-3 (rIL-3). Because these cells may be only a subset, and not representative of marrow progenitors, we have now studied partially purified marrow hematopoietic progenitor cells.

Ineffective hematopoiesis in folate-deficient mice.

A folate-free amino acid-based diet provided an opportunity to characterize the effects of folate depletion on growth, tissue folate levels, and hematopoiesis of mice under well-standardized conditions. Weanling mice were fed a folate-free, amino acid-based diet supplemented with either 0 or 2 mg folic acid/kg diet for 35 to 48 days. Folate concentrations were decreased in liver, kidney, serum, and erythrocytes in mice fed the folate-free diet.

Inhibition of human erythroid colony-forming units by interleukin-1 is mediated by gamma interferon.

IL-1 inhibits erythropoiesis in vivo and in vitro. This inhibition was studied by comparing the effect of recombinant human IL-1 (rhIL-1) on highly purified CFU-erythroid (E) generated from peripheral blood burst-forming units-erythroid (BFU-E) (mean purity 44.4%) with its effect on unpurified marrow CFU-E (mean purity 0.

Treatment of refractory pure red cell aplasia with cyclosporine A: disappearance of IgG inhibitor associated with clinical response.

Remissions were obtained in 6/9 evaluable patients with pure red cell asplasia (PRCA) refractory to other immunosuppressive agents who were treated with cyclosporine A (CsA). Four of these patients have remained in continuous remission off all treatment for 4-19 months. Another patient who stopped CsA abruptly relapsed, but responded to reinstitution of therapy.

Inhibition of human colony-forming-unit erythroid by tumor necrosis factor requires accessory cells.

Recombinant tumor necrosis factor (rTNF) inhibits erythropoiesis in vivo and in vitro. To study the mechanism of this inhibition, the effect of rTNF on highly purified human CFU-erythroid (E) (mean purity 63.5%), which were generated from peripheral blood burst-forming units-erythroid (BFU-E), was compared to its effect on unpurified human marrow CFU-E (mean purity 0.

Thrombopoiesis-stimulating factor: its effects on megakaryocyte colony formation in vitro and its relation to human granulocyte-macrophage colony-stimulating factor.

The effects of thrombopoiesis-stimulating factor (TSF) on human marrow megakaryocyte colony formation in vitro were studied by the plasma clot method. TSF was found to stimulate megakaryocyte as well as granulocyte-macrophage colony formation in vitro at optimal concentrations of 200-300 pg/ml of medium containing 2.5% horse serum.

Reversible hepatomegaly and diabetes mellitus in an adult with disseminated histiocytosis X.

Histiocytosis X rarely disseminates in an adult. The authors describe an unusual patients who presented with multiple areas of cutaneous and bone involvement. During the course of his disease he developed massive hepatomegaly.

The pathogenesis of anemia in rheumatoid arthritis: a clinical and laboratory analysis.

Principal concepts concerning the anemia of RA are summarized in Tables 7 and 8. These concepts have been validated by our analysis of 93 anemic RA patients and by our review of the literature. The fact that anemia in RA may have one or more etiologies, occasionally in the same patient, mandates a reasoned approach to the analysis of anemia in every RA patient in whom it may occur.

Treatment of the anemia of rheumatoid arthritis with recombinant human erythropoietin: clinical and in vitro studies.

Two anemic patients with rheumatoid arthritis were treated with recombinant human erythropoietin (EPO) for 5 months. Both patients showed significant increases in hematocrit, red cell volumes, and marrow erythroid and megakaryocyte progenitor cells. No significant toxic effects from EPO were observed.