Effect of low neutral endopeptidase expression on response to fMLP.

The effects of the low neutral endopeptidase (24.11/CD10) exhibited by cord blood neutrophils on response to the peptide mediator of cell function f-met-leu-phe (fMLP) were investigated. Oxidative radical release (superoxide and hydrogen peroxide) and chemotactic responses to fMLP were determined and compared to the responses of normal adult neutrophils. The effect of fMLP on CD10 expression as measured by flow cytometry also was evaluated. The data show that cord blood neutrophils produce increased amounts of O2- and H2O2 largely because of a prolonged reaction time to fMLP. In addition, adult polymorphonuclear neutrophil leukocytes increase the intensity of their expression of CD10 following fMLP stimulation, whereas cord blood CD10 expression does not change. Evaluation of chemotaxis demonstrated that cord blood neutrophils exhibited a shift in the fMLP dose-response relationship showing relatively better chemotaxis to lower concentrations. In support of this observation, the inhibition of endopeptidase on adult polymorphonuclear neutrophils leukocytes by phosphoramidon was associated with an augmentation of chemotaxis to 10(-9) and 10(-10) mol/L fMLP. These studies demonstrate that cord blood and adult neutrophils respond differently to fMLP and suggest that membrane endopeptidase plays a role in the observed response patterns. The low level of expression of CD10 on cord blood neutrophils and the failure to increase its expression after fMLP stimulation suggests that adult neutrophils have preformed intracellular CD10 that is not present in the newborn. We propose that the lack of endopeptidase on cord blood neutrophils together with other known features of immaturity may play a role in the overall compromised host defense exhibited by the newborn.

Neutrophil function in chronic neutrophilic leukemia: defective respiratory burst in response to phorbol esters.

Functional analyses were performed on neutrophils isolated from 6 patients from two institutions who displayed features of chronic neutrophilic leukemia (CNL). These neutrophils demonstrated a consistent deficiency (44 +/- 8% of control values) in superoxide anion (O2-) production in response to the phorbol ester, phorbol myristate acetate (PMA). O2- production in response to chemotactic peptides was near normal (82.3 +/- 10.7% of control values). Bacterial killing was normal in the two patients studied, and chemotaxis was diminished in response to zymosan-activated plasma and to high concentrations of chemotactic peptides in the patients studied. Cytosolic C kinase activity was decreased in one of the two patients studied. These results suggest that a deficient O2- release in response to PMA is a hallmark of neutrophils in CNL and may provide a diagnostic indicator of this condition.

Associated leukemia and mixed germ cell tumor in a patient with gonadal dysgenesis.

A 16-year-old phenotypic female developed acute myeloblastic leukemia with a fulminant course very shortly after surgery and chemotherapy for a mixed germ cell tumor of the ovary. The karyotype (46, XY, 47, XY + 8) suggested de novo rather than therapy-associated leukemia. The relationship between germ cell tumors and leukemia, their common yolk sac derivation and the role of the Y chromosome are discussed. The idea that XY gonadal dysgenesis may be familial also is discussed.

Inhibition of chemotaxis Ng-monomethyl-L-arginine: a role for cyclic GMP.

The metabolism of L-arginine to nitric oxide (NO) has been shown to be important for the effector functions of many cell types, including polymorphonuclear (PMN) leukocytes. Its effect appears to be mediated at least in part by NO stimulation of soluble guanylate cyclase. We evaluated the role of this pathway in two PMN effector functions: cell movement and microbial killing, using the competitive inhibitor of L-arginine conversion to NO, NG-monomethyl-L-arginine (NMA). We also evaluated the effect of additional L-arginine and dibutyryl cyclic guanosine monophosphate (cGMP) on any NMA-associated changes. Human peripheral blood neutrophils were used and the cells were incubated with and without NMA. Chemotaxis was evaluated using a 48-well micro-Boyden chamber. Microbial killing was evaluated using S aureus strains D2C and 502A. These studies demonstrated that chemotaxis to formyl-methionyl-leucyl-phenylalanine was markedly inhibited in NMA-treated cells. This inhibition could be overcome if L-arginine or dibutyryl cGMP were added with the NMA. In contrast, microbial killing of S aureus was unaffected by NMA. These observations support the hypothesis that the L-arginine metabolism to NO and its effect on the cGMP level may be important for the dynamic changes required for neutrophil chemotaxis.

Microgranular promyelocytic leukemia: a multiparameter examination.

Six cases of microgranular variant acute promyelocytic leukemia (M3v) were studied by use of a multiparameter approach including morphology, cytochemistry, flow cytochemistry, flow cytometry, cytogenetics, and gene rearrangement. Three of six cases demonstrated both myeloid and monocytoid associated surface markers by flow cytometry. One of six cases had strong alpha-naphthyl-butyrate esterase (alpha-NBE) activity in addition to myeloperoxidase activity. There was no correlation between percentage of positive monocytoid surface markers and intensity of cytoplasmic alpha-NBE activity. Four of six cases also had a T-cell-associated surface antigen. Further studies indicated that the T-cell markers appeared to be on the promyelocytes and that the T-B receptor gene was not rearranged. Similarly, cytogenetics studies indicated only one clonal abnormality t(15q+; 17q-). Whether these cases represent true "lineage infidelity" remains to be answered. Future important studies are needed on normal hematopoietic progenitor cells at early stages of development and childhood to study lineage-specific characteristics and to determine whether co-expression normally exists during early development.

The effect of recombinant human granulocyte macrophage colony-stimulating factor on neutrophil activation in patients with refractory carcinoma.

Patients with refractory carcinoma were treated with recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) by intravenous (IV) infusion. During the period of treatment, studies of polymorphonuclear leukocyte superoxide (O2-) release in response to formylmethionylleucylphenylalanine (fMLP) and phorbol myristate acetate (PMA) and studies of chemotaxis in response to fMLP and C5a were performed. We observed that patients receiving rhGM-CSF in vivo exhibited primed O2- release after stimulation both with fMLP and PMA. Chemotaxis, however, was not enhanced by the treatment. These data suggest that host defenses may be enhanced by this treatment and that rhGM-CSF may be a useful therapeutic adjunct in compromised patients.

Inhibition of cell movement and associated changes by hexachlorocyclohexanes due to unregulated intracellular calcium increases.

Hexachlorocyclohexanes (HCCHs) are potent stimulators of polymorphonuclear leukocyte (PMN) oxidative metabolism and of mobilization of calcium from intracellular stores. It was of interest, therefore, to evaluate the effect of HCCHs on PMN orientation and chemotaxis and to determine their effectiveness as chemotaxins. Chemotaxis was evaluated using micro-Boyden chambers, f-actin was quantitated by nitrobenzoxadiazole (NBD)-phallacidin fluorescence, and microtubules were quantitated by observing the concanavalin A (Con A) capping phenomenon. We also evaluated changes in intracellular calcium [Ca2+]i using quin 2 fluorescence. We found that the HCCH isomers were not chemotaxins and that the HCCH isomers that stimulated O2- formation (delta and gamma HCCH) inhibited chemotaxis. This effect was associated with inhibition of orientation. In addition, we found extensive inhibition of both f-actin and Con A cap formation. These effects of HCCH on cell function were associated with marked increases of [Ca2+]i. This work suggests that non-receptor-mediated increases of [Ca2+]i associated with HCCH have divergent effects on cell function and suggests that physiologic responses of PMNs requiring cytoskeletal alterations, such as chemotaxis, depend on the controlled responses of receptor-mediated stimulation.

Characterization of lymphocyte subpopulations in human cord blood using the immunogold staining technique.

Lymphocyte subpopulations in human cord blood have been examined using monoclonal antibodies, visualized with immunogold. The proportions of T11, T4, T8, and B1 cells in cord blood are very similar to values in adult peripheral blood. Some evidence of lymphocyte immaturity in cord blood is suggested by the presence of 12% CALLA-positive cells and the sum of T4 and T8 cells significantly exceeding the number of T11 cells; however, there were no TdT-positive cells. The presence of CALLA-positive lymphocytes in normal cord blood should be borne in mind when investigating blood smears from neonates for congenital leukemia.