Granulocytic colony forming cells in vitro: I. Patients with drug induced agranulocytosis.
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Biomedical subjects
Publications and source records attributed to H Heimpel.
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The protein composition of the erythrocyte membrane of two patients with CDA I and four patients with CDA II has been investigated by two-dimensional polyacrylamide gel electrophoresis. The two-dimensional patterns of erythrocyte membrane proteins of the patients with CDA were compared with the well-established pattern of normal individuals. No alterations could be detected in the patients with CDA I, but a striking deviation from normal was observed in all patients with CDA II. The major aberrations of the erythrocyte membrane proteins in CDA II are the lack of protein B1.1, the drastically increased concentration of protein B1.4 and the presence of a new component, C2'. These results are discussed in relation to the known morphological and immunological abnormalities of the erythrocyte membrane in CDA II.
The characteristics and the concentration of granulopoietic colony forming cells (CFC) were examined in 22 patients with aplastic anaemia at different stages of their disease. Additionally the ability of the patients' peripheral leucocytes to elaborate factors necessary for colony stimulation in vitro (CSA) was studied. The ability of the patients' cells to generate CSA was shown to be unaffected. However, the incidence of CFC within the marrow and peripheral blood suspensions was significanlty reduced in all patients. The results suggest a reduced compartment size of CFC even in those patients who have recovered from aplastic anaemia. This may indicate that the disturbances in the preceding compartments of the haemopoietic cell renewal system still persist after recovery from the acute bone marrow failure.
The concentration and erythropoietin dependence of erythropoietic progenitor cells (CFU-E) were examined in 13 patients with aplastic anaemia at different stages of their disease. The CFU-E incidence was shown to be quantitatively diminished in aplastic anaemia but tended to recover to normal values if the disease recovered. In addition the CFU-E showed a qualitatively different response to stimulation by erythropoietin, being resistant to low concentrations but responsive to concentrations greater than 0.2 U/ml whereas there was a linear response in the controls up to 0.5 U/ml.
The effect of polymorphonuclear granulocytes (PMN) on colony stimulating activity (CSA) was studied in double layer cultures of human Ficoll Isopaque separated white blood cells (mononuclear cells = MNC). Previously published data have been confirmed that granulocytes are able to enhance or inhibit MNC derived CSA. Further analysis of the mode of action of PMN in vitro indicates that the enhancing activity ascribed to granulocytes coincides with low CSA in MNC basal layers. In contrast, in cultures with high levels of CSA as provided by lysed red blood cell enhancement rather than concentrations of PMN are sufficient to induce inhibition of colony growth. A very similar effect to that achieved with basal layer derived CSA could be obtained with conditioned media of PMN and MNC short term liquid cultures. The data indicate, that enhancement and inhibition of colony growth reflect a specific reactivity of granulocytes (PMN) to a given CSA level in the cultures. These findings are discussed in terms of a speculative role of PMN in a negative feed back control mechanism regulating granulopoiesis in vivo.
A patient suffering from aplastic anaemia was treated by bone-marrow transplantation from an ABO- and HLA-identical, MLC- and CML-negative, unrelated donor. MLC and CML became positive after transplantation indicating that a cellular immune response had developed against lymphocyte determinants not recognized prior to sensitization in vivo. Whether these determinants are governed by genes of the HLA region is unknown at present.
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Of 75 patients with asplastic anaemia treated between 1968 and 1975, 33 were retrospectively considered as potential candidates for allogenegic bone-marrow transplantation on the basis of their age and severity of marrow failure. The prognosis of these patients with conservative treatment was assessed from parameters obtained at the time of the initial diagnosis. Initial peripheral-blood granulocyte or platelet concentrations were not of porgnostic value. In contrast, initial reticulocyte concentrations, allowed separation of the patients into two groups with poor and good prognosis. Low initial reticulocyte concentrations (less than 10 000/mu1) indicated those patients at extremely high risk of succumbing to their marrow aplasia (there were no survivors 36 months after disgnosis). In contrast, 75% of those patients with more than 10 000 reticulocytes per mu1 at diagnosis survived for 3 years. Initial peripheral-blood reticulocyte concentrations thus appear to indicate the extent of the marrow failure in aplastic anaemia more accurately than granulocytes or platelets. Low initial reticulocyte concentrations may indicate, among patients with severe aplastic anaemia, those for whom allogeneic bone-marrow transplantation should be seriously considered; patients with higher initial reticulocyte concentrations may benefit from conservation treatment.
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A controlled randomized multicenter trial was carried out to examine the therapeutic value of the androgen mesterolone in aplastic anemia and anemia of renal failure. the drug was given in a dose of 2 mg/kg orally for 6 months. Control patients received no androgens but were otherwise similarily treated. 31 patients with aplastic anemia could be evaluated. No significant difference was found between androgen treated control cases in respect to bone marrow cellularity, improvement of peripheral blood cell counts or survival. In a group of 14 patients not being hemodialyzed with anemia from renal failure, 3 of the androgen treated and none of the control patients showed progressive and significant improvement of erythropoiesis; however, this was not a statistically significant difference when the both groups of patients were compared. The results do not suggest that the androgen mesterolone is of therapeutic value in the majority of adult patients with aplastic anemia. Possible reasons of the discrepancy to positive results reported in the literature are discussed.
The case of an immunoblastic lymphadenopathy with a leukaemoid reaction has been observed. All the clinical, serological and histological features described by previous authors were present. In addition, there were more than 1000000/mul nucleated peripheral cells, consisting of mature and a few immature granulocytes, a small number of normoblasts and many lymphocytes, "immunoblast-like" lymphoid cells and plasma cells. Polyclonality of the latter was demonstrated by serological methods. Spontaneous remission occurred and was apparently enhanced by the application of corticosteroids.
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In vitro investigations were performed to re-examine the variables that influence the binding of sodium 99TCm-pertechnetate to human red blood cells. Irreversible binding of the radioisotope was observed only when red cells were incubated with tin chloride after their incubation with pertechnetate; binding of pertechnetate is completed within 10 min after its addition to red cells. Composition, temperature and possibly the oxygen saturation of the incubation medium affect the fraction of the isotope that becomes red cell-bound. Using a labelling method derived from these in vitro studies, the usefulness of pertechnetate-labelled autologous red cells for the determination of red cell volumes was studied. Elution of pertechnetate occurred in vivo in an exponential fashion. Using the specific red cell activity, corrected for elution, of blood samples drawn 30 min after injection of 99Tcm-labelled red cells, calculated red cell volumes were almost identical to those determined simultaneously with radiochromate-labelled cells. It is concluded that 99Tcm-pertechnetate is a useful label for red cell volume determination; it may be less reliable when delayed mixing of the labelled red cell tracer in the patient's circulation is expected.
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The expression "congenital dyserythropoietic anemia" (CDA) has been used to characterize the kinetic and morphological aberrations in the proliferation and maturation compartment of erythropoiesis, occurring in a group of hereditary anemias of unknown pathogenesis. The main symptoms of these disorders are moderate or mild anemia, increased hemoglobin turnover, ineffectiveness of erythropoiesis, striking morphological aberrations of the erythroblasts and tendency to secondary hemochromatosis. To date, three types have emerged from this group that may be hereditary nosological entities. They are distinguished not only on a morphological basis, but also by different modes of inheritance and immunological properties of the red cell membrane. A number of additional cases or families have been described which could not be attributed to one of these three types. Comparative investigations of morphological, biochemical and immunological details in CDA on the one hand, and other forms of ineffective erythropoiesis (e.g. thalassemia, refractory anemia) on the other demonstrate the lack of specificity of many of the single changes observed in CDA. These changes may well be secondary phenomenon of intramedullary cell destruction brought about by different underlying pathogenetic mechanisms.
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