CFU-E) And Granulocyte-macrophage Research Articles

Transfusion begets anemia

Following trauma, transfusion of aged stored blood is often necessary yet associated with increased morbidity and mortality. Despite blood replacement, many patients have a prolonged anemia requiring further transfusions. The effects of aged blood on bone marrow (BM) hematopoiesis have not been studied, and we hypothesized that stored blood suppresses BM function. Blood from Sprague-Dawley rats was stored for 1, 14, or 28 days with the industry preservative citrate-phosphate-dextrose-adenine-1 (CPDA-1). For in vitro studies, 5% supernatant was incubated with normal rat BM and cultured for erythroid (CFU-E) and granulocyte-macrophage (CFU-GM) colony-forming units. Data were compared with cultures of BM alone, 5% control plasma (negative control), and 12% CPDA-1. For in vivo studies, rats were transfused with stored supernatants (5% estimated blood volume (EBV) over 30 minutes). BM from each recipient was cultured for CFU-E and CFU-GM at 3 hours after transfusion. Data were compared with cultures of BM alone. Difference between groups determined by analysis of variance and Tukey's multiple comparison test. In vitro exposure to CPDA-1, control plasma, or 1-day supernatant (D1) had no effect on BM growth compared with BM alone. In vitro exposure to 14-day (D14) and 28-day (D28) supernatant significantly suppressed CFU-E by 60% and CFU-GM growth by 71% (both p < 0.05) compared with D1 or medial alone. There were no differences between D14 and D28. In vivo exposure to D14 reduced BM CFU-E and CFU-GM growth by 55% (both p < 0.05) compared with D1 supernatant. Plasma from aged blood adversely affects CFU-E and CFU-GM growth in rats. The effect is not mediated by CPDA-1. Transfusion of aged stored blood may contribute to BM dysfunction in critically ill patients, resulting in persistent anemia and the need for further transfusion. This BM dysfunction may also partly explain the observed increased susceptibility to infection.

Value of In Vitro Models for the Assessment of Drug-Induced Haematotoxicity

BackgroundA new antipsychotic compound induced unexpected red cell hypoplasia (reticulocytopenia, red marrow hypoplasia) in rats dosed orally for 7days. Materials and methodsSince an erythropoietin-mediated pathogenesis was excluded, in vitro tests on rat and human bone marrow cells were performed with measurement of formation of late erythroid (CFU-E) and granulocyte-macrophage (CFU-GM) colony-forming units after incubation with the drug. CFU-E together with growth factors were cultured for 2days (rat) or 7days (human) and CFU-GM was cultured for 7days (rat) or 10days (human). ResultsThe drug induced inhibition of erythroid progenitors and myeloid progenitors for both species from 3×10−5 mol/L, with the concentration inhibiting the growth of 50% (IC50) consistent with drug plasma levels measured in rats. ConclusionThese cloning assays on rat bone-marrow cells were shown to be adequate models for determining the haematotoxicity of the agent and to be predictive of human toxicity. With only a small amount of compound required, they can be used as screening tools to detect haematotoxic potential of candidate drugs.

Three-lineage hemopoietic precursor cells and effectiveness of recombinant human erythropoietin in patients with myelodysplastic syndromes.

Four-stem-cell assays, which evaluate megakaryocytic (CFU-Meg), immature and mature erythropoietic (BFU-E, CFU-E), and granulocyte-macrophage (CFU-GM) colony formation, were performed in nine patients with myelodysplastic syndromes (MDS). The CFU-Meg, BFU-E, and CFU-E colony growths were disturbed more often than the CFU-GM colony formation. A CFU-E increase was not recognized in most MDS patients, but a dose-dependent increase of bone marrow CFU-Es in response to erythropoietin (EPO) was recognized only in two refractory anemia (RA) patients whose CFU-Es were more than one tenth of normal controls. One patient with RA and the other with chronic myelomonocytic leukemia (CMML), both of whose bone marrow CFU-Es did not increase at the higher dose of EPO in vitro, were treated with recombinant human EPO (rHuEPO), resulting in no effects. The responsiveness of patients with MDS to various recombinant hemopoietic factors might be predicted by both the residual degree of bone marrow hematopoietic precursor cells and the response of stem cells to the higher doses of each hemopoietic factor.

Retrovirus-Induced Feline Pure Red Blood Cell Aplasia: Pathogenesis and Response to Suramin

AbstractFeline leukemia virus, subgroup C/Sarma (FeLV-C/Sarma) induces pure red blood cell aplasia in cats. Although eryth-roid (BFU-E and CFU-E) and granulocyte/macrophage (CFU-GM) progenitors are infected with this virus, only erythropoie-sis is impaired. Two to 3 weeks before the onset of anemia, CFU-E become undetectable in marrow cultures while earlier erythroid progenitors (BFU-E) persist, suggesting that FeLV-C/Sarma (presumably via its envelope glycoprotein gp70) inhibits the differentiation of BFU-E to CFU-E in vivo. To correlate in vitro observations with the progression of disease, prospective studies were performed in six cats. These studies showed that at the time that the frequencies of CFU-E decreased in marrow cultures, BFU-E no longer responded to hematopoietic growth factor(s), although the responses of CFU-GM were unchanged. In further studies, anemic cats received suramin, a reverse-transcriptase inhibitor with other diverse effects. Within 4 to 14 days, erythro-poiesis improved and up to 1,616 CFU-E were detected per 105 marrow mononuclear cells. However, progenitor cells remained infected, suggesting that suramin modulated erythroid differentiation without inhibiting progenitor infection. These observations led to the hypothesis that the gp70 of FeLV-C/Sarma impairs BFU-E differentiation by interference with ligand/receptor interactions or signal transduction pathways unique to erythroid cells. Understanding this mechanism should provide insights into the interactions controlling early erythropoiesis.

Retrovirus-induced feline pure red blood cell aplasia: pathogenesis and response to suramin

Feline leukemia virus, subgroup C/Sarma (FeLV-C/Sarma) induces pure red blood cell aplasia in cats. Although erythroid (BFU-E and CFU-E) and granulocyte/macrophage (CFU-GM) progenitors are infected with this virus, only erythropoiesis is impaired. Two to 3 weeks before the onset of anemia, CFU-E become undetectable in marrow cultures while earlier erythroid progenitors (BFU-E) persist, suggesting that FeLV-C/Sarma (presumably via its envelope glycoprotein gp70) inhibits the differentiation of BFU-E to CFU-E in vivo. To correlate in vitro observations with the progression of disease, prospective studies were performed in six cats. These studies showed that at the time that the frequencies of CFU-E decreased in marrow cultures, BFU-E no longer responded to hematopoietic growth factor(s), although the responses of CFU-GM were unchanged. In further studies, anemic cats received suramin, a reverse-transcriptase inhibitor with other diverse effects. Within 4 to 14 days, erythropoiesis improved and up to 1,616 CFU-E were detected per 10(5) marrow mononuclear cells. However, progenitor cells remained infected, suggesting that suramin modulated erythroid differentiation without inhibiting progenitor infection. These observations led to the hypothesis that the gp70 of FeLV-C/Sarma impairs BFU-E differentiation by interference with ligand/receptor interactions or signal transduction pathways unique to erythroid cells. Understanding this mechanism should provide insights into the interactions controlling early erythropoiesis.

Comparative distribution of marrow CFU‐e and CFU‐gm progenitors in different anatomic sites in the dog

The interpretation of marrow cloning activity, particularly in serial cultures, is greatly influenced by the reproducibility of the collected marrow samples. In order to establish whether bone marrow cloning activities and precision of the cloning assays are influenced by the site of bone marrow collection in the dog, we studied the incidence of marrow erythroid (CFU-e) and granulocyte-macrophage (CFU-gm) progenitor cells in the iliac crest, sternum, vertebrae, femur, and humerus, using microplasma clot and soft agar culture systems. Marrow samples obtained from the femur and humerus revealed consistently higher cell concentrations than those from the iliac crest, vertebrae, or sternum. Those aspirated from the sternum and vertebrae had lower cell concentrations and were less reproducible. Statistical analysis revealed no significant differences in the incidence of marrow CFU-e and CFU-gm progenitor cells between the femur, humerus, iliac crest or vertebrae. With multiple sampling, the marrow cloning efficiency was consistent and reproducible within the individual dogs. We conclude that the distribution of CFU-e and CFU-gm is comparable throughout the active marrow in the dog and that these sites may be used interchangeably for multiple quantitative analysis of marrow hematopoietic progenitor cells.

Cephalosporin-induced alterations in erythroid (CFU-E) and granulocyte-macrophage (CFU-GM) colony-forming capacity in canine bone marrow

Cephalosporins are among the safest antibiotics. Nevertheless, hematologic abnormalities ranging from mono- to pancytopenia do occur, albeit infrequently, following their therapeutic use. Similar abnormalities to those reported in people have been seen in dogs given high doses of cephalosporins. As part of a study to define the latter thoroughly, we explored the effects of long-term, high-dose cephalosporin administration on canine marrow erythroid (CFU-E) and granulocyte-macrophage (CFU-GM) progenitor cells. Cefazedone (Refosporen, E. Merck, Darmstadt) was administered intravenously at doses of 540 to 840 mg/kg daily to 14 healthy beagle dogs for up to 4 months, or less if hematologic effects were evident earlier. Within 6 to 10 weeks, treated dogs developed pancytopenia ( 5 14 ), thrombocytopenia ( 11 14 ), moderate to severe neutropenia ( 8 14 ), and/or normocytic anemia ( 8 14 ). There was evidence of immune-mediated destruction of peripheral blood cells. All treated dogs exhibited a significant reduction in marrow colony-forming capacity, irrespective of whether peripheral cytopenia was present, with 12 14 showing decreased CFU-GM and 14 14 decreased CFU-E activity. Within a week following cessation of dosing, all affected dogs achieved hematologic remission as defined by restoration of the peripheral blood counts. However, despite this apparent recovery, both CFU-E and CFU-GM activities of the bone marrow remained depressed for at least another 8 moths. We conclude that in dogs prolonged administration of high doses of cefazedone induced a persistent deficit of CFU-E and CFU-GM progenitor cells. The clinical relevance of this, if any, remains to be established.

Cephalosporin-lnduced Alterations in Erythroid (CFU-E) and Granulocyte-Macrophage (CFU-GM) Colony-Forming Capacity in Canine Bone Marrow

Cephalosporin-lnduced Alterations in Erythroid (CFU-E) and Granulocyte-Macrophage (CFU-GM) Colony-Forming Capacity in Canine Bone Marrow

Selective inhibition of erythropoiesis by sera from patients with chronic renal failure.

The pathogenesis of anemia in patients with chronic renal failure was studied by analyzing the effect of uremic sera on the in vitro colony growth of erythroid (CFU-E) and granulocyte-macrophage (CFU-GM) progenitor cells. Uremic sera from 20 of 30 patients inhibited erythroid colony growth below 70% of control even when cultured with normal human bone marrow of the same blood type. On the other hand, only one of the sera inhibited colony growth of CFU-GM as compared with normal sera. On Sephadex G-15 gel filtration, the CFU-E-inhibiting activity appeared in two different fractions: the void volume peak and the delayed eluant before the second peak. The inhibiting activity in the former fraction was noted only in uremic sera. The results of this study suggest the existence of a serum inhibitor(s) of erythropoiesis with a relative molecular mass of more than 1500 Da which are virtually impossible to dialyze by conventional membranes.

Effect of recombinant interferons alpha and gamma on human bone marrow- derived megakaryocytic progenitor cells

Interferons (IFNs) have been shown to suppress the proliferation of human pluripotent hematopoietic progenitor cells, CFU-GEMM, and committed erythroid (BFU-E, CFU-E) and granulocyte-macrophage (CFU-GM) progenitor cells. However, no information is yet available concerning the effect of IFNs on human megakaryocytic progenitor cells CFU-Mk. Furthermore the mechanisms underlying the inhibitory activity of IFNs are still controversial. Therefore highly purified recombinant IFN preparations, rIFN-alpha and rIFN-gamma, were assessed for their influence on in vitro growth of human bone marrow-derived CFU-Mk as well as CFU-GEMM. In addition, the role of hematopoietic accessory cells, that is, adherent cells and T lymphocytes, in the mediation of the suppressive effect of rIFNs was examined. When added to unseparated bone marrow cells, both rIFN preparations significantly inhibited colony formation with 50% inhibition of CFU-Mk occurring at 22 U/mL for rIFN-alpha and 59 U/mL for rIFN-gamma, while 50% inhibition of CFU-GEMM occurred at 59 U/mL for rIFN-alpha and 101 U/mL for rIFN-gamma. The suppressive effect of rIFN-alpha and rIFN-gamma was selectively abolished by monoclonal antibodies (MoAbs) against rIFN-alpha and rIFN- gamma, thus confirming that the inhibitory activity was due to the rIFN preparations used. The antiproliferative effect of rIFN-alpha and rIFN- gamma on CFU-GEMM growth was not associated with a decrease in the percentage of mixed colonies containing megakaryocytic cells as assessed by use of the MoAb C17.28 against platelet glycoprotein IIIa. Removal of adherent cells and T lymphocytes from the target bone marrow cells had no influence on the suppressive effect of rIFN-alpha, whereas it significantly reduced the inhibitory effect of rIFN-gamma on the growth of megakaryocytic colonies and the other hematopoietic progenitors. The data indicate that (1) human megakaryocytopoiesis is markedly inhibited by rIFN-alpha and rIFN-gamma, and (2) the inhibitory effect of rIFN-alpha is due to a direct action on hematopoietic progenitor cells, whereas the effect of rIFN-gamma is mediated to a significant degree through accessory cell populations.

Effect of Recombinant Interferons Alpha and Gamma on Human Bone Marrow–Derived Megakaryocytic Progenitor Cells

AbstractInterferons (IFNs) have been shown to suppress the proliferation of human pluripotent hematopoietic progenitor cells, CFU-GEMM, and committed erythroid (BFU-E, CFU-E) and granulocyte-macrophage (CFU-GM) progenitor cells. However, no information is yet available concerning the effect of IFNs on human megakaryocytic progenitor cells CFU-Mk. Furthermore the mechanisms underlying the inhibitory activity of IFNs are still controversial. Therefore highly purified recombinant IFN preparations, rIFN-alpha and rIFN-gamma, were assessed for their influence on in vitro growth of human bone marrow-derived CFU-Mk as well as CFU-GEMM. In addition, the role of hematopoietic accessory cells, that is, adherent cells and T lymphocytes, in the mediation of the suppressive effect of rIFNs was examined. When added to unseparated bone marrow cells, both rIFN preparations significantly inhibited colony formation with 50% inhibition of CFU-Mk occurring at 22 U/mL for rIFN-alpha and 59 U/mL for rIFN-gamma, while 50% inhibition of CFU-GEMM occurred at 59 U/mL for rIFN-alpha and 101 U/mL for rIFN-gamma. The suppressive effect of rIFN-alpha and rIFN-gamma was selectively abolished by monoclonal antibodies (MoAbs) against rIFN-alpha and rIFN-gamma, thus confirming that the inhibitory activity was due to the rIFN preparations used. The antiproliferative effect of rIFN-alpha and rIFN-gamma on CFU-GEMM growth was not associated with a decrease in the percentage of mixed colonies containing megakaryocytic cells as assessed by use of the MoAb C17.28 against platelet glycoprotein IIIa. Removal of adherent cells and T lymphocytes from the target bone marrow cells had no influence on the suppressive effect of rIFN-alpha, whereas it significantly reduced the inhibitory effect of rIFN-gamma on the growth of megakaryocytic colonies and the other hematopoietic progenitors. The data indicate that (1) human megakaryocytopoiesis is markedly inhibited by rIFN-alpha and rIFN-gamma, and (2) the inhibitory effect of rIFN-alpha is due to a direct action on hematopoietic progenitor cells, whereas the effect of rIFN-gamma is mediated to a significant degree through accessory cell populations.

HLA-DR monoclonal antibodies inhibit the proliferation of normal and chronic granulocytic leukaemia myeloid progenitor cells.

We studied the capacity of murine monoclonal antibodies with HLA-DR specificity to inhibit the proliferation in vitro of erythroid (BFU-E and CFU-E) and granulocyte-macrophage (CFU-GM) progenitor cells in normal bone marrow and the blood of patients with chronic granulocytic leukaemia (CGL). Two IgG2 antibodies (CA 2.06 and L243) inhibited the proliferation of normal BFU-E and CFU-GM at relatively high dilution; a third antibody, DA2, had no effect on either progenitor cell. A complement-fixing monoclonal antibody with T-cell activity (OKT3) produced only minor reduction in progenitor cel proliferation. Further studies with L243 showed that BFU-E and CFU-GM from the blood of patients with CGL were inhibited to the same degree as normal marrow progenitor cells. The inhibition of progenitor cell proliferation by a given antibody was always complement dependent and was therefore presumed to be due to a direct cytotoxic effect. The inhibitory effect of monoclonal antibodies is a valuable approach to the characterization of antigenic determinants on myeloid progenitor cells and the differential cytotoxicity of selected monoclonal antibodies might be exploitable for therapy.