Red blood cell regeneration induced by subcutaneous recombinant erythropoietin: iron-deficient erythropoiesis in iron-replete subjects [see comments]

Limited red blood cell (RBC) regeneration often prevents collection of sufficient blood from autologous donors. We studied the effects of subcutaneous recombinant erythropoietin (rEPO) in subjects making frequent blood donations. Six healthy iron-replete male subjects took rEPO (200 U/kg) subcutaneously daily, and donated blood (450 mL) twice a week for 3 weeks. During a control study, these subjects also attempted twice-weekly blood donations without rEPO. Four other males given rEPO, including one with idiopathic hemochromatosis, waited until day 8 to begin blood donations. All healthy subjects took oral ferrous sulfate. Subcutaneous rEPO given with blood donations resulted in a marked reticulocytosis (mean peak value 568 +/- 159 x 10(9)/L v 235 +/- 77 x 10(9)/L, control study; P < .05), and enhanced RBC production at 28 days (1,208 +/- 227 mL v 719 +/- 161 mL, P < .05). rEPO in advance of blood donations was slightly less effective in normal subjects (941 +/- 139 mL, P < .05); however, the subject with hemochromatosis produced substantially more RBCs (1,764 mL) than any normal subject. rEPO-treated normal subjects (but not the rEPO-treated patient with hemochromatosis or untreated controls) produced iron-deficient RBCs with elevated zinc protoporphyrin levels and low hemoglobin content. These cells appeared within 1 week of rEPO administration and before laboratory confirmation of depleted iron stores. Thus, subcutaneous rEPO is an effective stimulant of erythropoiesis in nonanemic blood donors. However, in addition to eventual depletion of iron stores, early functional iron deficiency affects response to the drug.     

Beta 2-integrin and L-selectin are obligatory receptors in neutrophil aggregation [see comments]

We have recently found that antibodies to L-selectin, the homing receptor on neutrophils, are as effective as those to beta 2-integrin at blocking formyl peptide-stimulated aggregation. Therefore, we investigated the requirements for expression of L-selectin and beta 2- integrin on adjacent cells during aggregation. Fluorescence flow cytometry allowed characterization of aggregates on the basis of size and color, as well as antibody binding to these two adhesive molecules. Formyl peptide-stimulated aggregate formation was measured for individual populations fluorescently labeled red (LDS-751) or green (CD44-FITC), and interpopulation red-green cell conjugates. Blocking either the beta 2-integrin or L-selectin adhesive epitope with monoclonal antibody on individual cell populations resulted in an approximately 50% reduction in two-color aggregation as compared with that in unblocked samples. Shedding the L-selectin on a cell population by preincubation with complexes of lipopolysaccharide and its plasma membrane binding protein also decreased aggregation to a control population by approximately 50%. We examined the aggregation of neutrophils from patients genetically deficient in beta 2-integrin and clinically leukocyte adhesion deficient (LAD). LAD adhesion to normal neutrophils was dependent on the expression of L-selectin on LAD cells and beta 2-integrin on normal cells. Thus, the minimum requirement for adhesion between two mixed populations of neutrophils was that one population expressed the beta 2-integrin and the other expressed the L- selectin adhesive epitope.     

Expression of specific granule markers on the cell surface of neutrophil cytoplasts

The widespread assumption that cytoplasts generated from human polymorphonuclear leukocytes (PMNs) are vesicles consisting solely of cytoplasm surrounded by plasma membrane and devoid of granule activity remains to be tested. PMN cytoplasts were prepared by centrifugation of intact cells on a Ficoll step gradient in the presence of cytochalasin B. Two granule membrane markers, Mol, a fluorometrically detectable antigen, and cytochrome b, both of which have been shown to translocate to the plasma membrane during granule release, were compared for their activity in cytoplasts and intact PMNs. We found that the amount of Mol detected on the plasma membrane of intact PMNs, as compared with other membrane markers (such as antigens LFA-1 and beta 2m), increased 1.6- fold upon exposure of PMNs to Ficoll plus cytochalasin B prior to centrifugation. Another twofold increase in Mol expression occurred upon cytoplast preparation. Release of the granule enzymes, vitamin B12- binding protein, and lysozyme were also followed and correlated well (r = .78 and .92) with the amount of Mol antigen present on the cell surface. Cytochrome b was also found to be higher (1.4-fold) on plasma membranes isolated from cytoplasts than on plasma membranes isolated from intact control cells. These results indicate that some fusion of granule membranes and plasma membranes occurred during treatment of PMNs with Ficoll plus cytochalasin b and during cytoplast preparation.     

Familial recurring cardiac myxoma

A 34-year-old woman presented with her fourth occurrence of an atrial myxoma and a right lower lung field mass. Her surgical history was extensive. In addition to resection of three previous atrial myxomas, she had undergone resection of her adrenal glands as a teenager because of Cushing's syndrome, and a hysterectomy at age 26 revealed a myxoid leiomyoma. Family history was remarkable as a maternal uncle and daughter died from embolic complications of left atrial myxomas and her sister previously had a left atrial myxoma resected. The patient underwent uncomplicated removal of the myxoma, and resection of the lung mass revealed a granuloma. A review of typical and atypical aspects of cardiac myxomas is provided including a rare and recently described syndrome of familial cardiac myxoma associated with Cushing's syndrome, spotty skin pigmentation and other myxoid tumors.     

Platelet function studies in coronary artery disease. X. Effect of dipyridamole

To evaluate the effects of dipyridamole on blood platelet function in patients with coronary artery disease, platelet counts and aggregation were examined in aortic and coronary venous blood. Before administration of dipyridamole, platelet counts and aggregation in response to adenosine diphosphate were less (p <0.02) in coronary venous than in aortic blood. Dipyridamole administration (100 mg) resulted in an increase in platelet counts and platelet aggregation in coronary venous blood so that the differences in aortic and coronary venous blood values were eliminated. These phenomena were probably related to inhibitory actions of dipyridamole. on platelet adhesion to atherosclerotic vessels. To further study the mechanism of action, the direct effects of dipyridamole on in vitro platelet aggregation were evaluated. Although dipyridamole, in the concentrations used, had no effect on in vitro platelet aggregation, it greatly potentiated the aggregation inhibitory actions of exogenous prostacyclin. In vivo potentiation of endogenous prostacyclin and inhibitory actions on platelet adhesion are the most likely mechanisms of the potentially beneficial actions of dipyridamole.     

Aplastic anemia: analysis of stromal cell function in long-term marrow cultures

Marrow samples from 89 patients with aplastic anemia (AA) were evaluated for their ability to grow stromal layers in standard long- term marrow cultures (LTMCs). Results were highly variable: 6.8% failed to grow any stromal cells (group I); 42.5% either failed to grow to confluency or appeared to have a decreased number of adipocytes and/or macrophages (group II); and 52.8% appeared as normal confluent cultures with fibroblasts, adipocytes, and macrophages (group III). Analyses of patient data suggested that group I patients had a longer disease duration and poorer survival (P = .07). Enzyme-linked immunosorbent assay analysis of cytokine production was performed on 20 of the normal- appearing AA LTMCs and 12 LTMCs established from normal donors. Significant differences between the AA and control groups were apparent for macrophage inflammatory protein-1 alpha (MIP-1 alpha), interleukin- 1 receptor antagonist (IL-1ra), granulocyte-macrophage colony- stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G- CSF), and leukemia-inhibitory factor (LIF). The most dramatic differences observed were elevated levels of MIP-1 alpha and GM-CSF and decreased levels of IL-1ra, particularly after IL-1 alpha stimulation. In contrast, IL-1 alpha stimulation of AA LTMCs produced levels of IL- 6, LIF, and G-CSF comparable with those of controls. These data suggest that defects exist within the microenvironment of some AA marrows. Whether the majority of these defects are the cause or consequence of aplasia is not clear. However, we speculate that some of these abnormalities may contribute to the maintenance of the hypoplastic state and, in extreme cases, prevent engraftment of donor marrow.