bluuebunny/crossref_metadata_2025 huggingface dataset:
The presence of mast cell precursors in rat peripheral blood
Authors: Zucker-Franklin D , Grusky G , Hirayama N , Schnipper E
Published: October 2019

Soft agar culture of mononuclear cell fractions prepared from rat peripheral blood yielded numerous colonies consisting of mast cells. The mast cell nature of the cells was established by ultrastructural and histochemical analyses as well as by the demonstration the the colonies contained histamine and that the cells possessed receptors for the Fc component of IgE. Stringent criteria for the distinction of mast cells from monocytes/macrophages that could have metachromatic inclusions were applied. The alcian-blue-safranin technique delineated the maturation of mast cell granules by showing the loss of alcian-blue and increase in safranin-positive organelles presumed to reflect the increase in N-sulfated polysaccharides representing heparin. The mast cells exhibited low or absent reactions for peroxidase, alpha-naphthyl butyrate, periodic acid Schiff, and Sudan black reacting lipid, whereas macrophages stained in parallel were positive for these substances. Since it is known that extracellular conditions may cause variations in phenotypic expression, the observations have led to the hypothesis that mast cells and macrophages may have a common precursor.
Expression of common acute lymphoblastic leukemia antigen (CALLA) by lymphomas of B-cell and T-cell lineage
Authors: Ritz J , Nadler LM , Bhan AK , Notis-McConarty J , Pesando JM , Schlossman SF
Published: October 2019

Previous studies have demonstrated that the common acute lymphoblastic leukemia antigen (CALLA) is expressed by leukemic cells from approximately 80% of patients with non-T-cell ALL and 30%-50% of patients with chronic myelocytic leukemia in blast crisis. A small number of normal bone marrow and fetal liver cells also express CALLA, but the functional role of this molecule is unknown. In the present study, we have used a monoclonal antibody (J5) specific for CALLA to study the expression of this antigen in non-Hodgkin's lymphomas. Within the B-cell lymphomas, it was found the CALLA was expressed by almost all Burkitt's and nodular poorly differentiated lymphocytic lymphomas. Within the T-cell lymphomas, CALLA was expressed in 40% of patients with lymphoblastic lymphoma. Three of 3 Burkitt's lymphoma cell lines and three of eight T-lymphoblast cell lines were also found to express CALLA. Normal spleen, lymph node, and thymus cells were not reactive with J5 antibody. These findings indicate that expression of CALLA is not limited to relatively undifferentiated leukemic lymphoblasts but also occurs in more differentiated lymphoid malignancies. However, normal differentiated lymphoid cells in lymph node, spleen, and thymus, which have a phenotype similar to that of lymphoma cells, do not appear to express CALLA.
The bleeding time is longer than normal in hemophilia
Authors: Eyster ME , Gordon RA , Ballard JO
Published: October 2019

Bleeding times were performed on 71 hemophiliacs using the Simplate II device. Eight patients receiving Indocin or Motrin for hemophilic arthropathy were evaluated separately from the remaining 63 who had a mean bleeding time of 7.65 +/- 3.20 min (1 SD) compared to the control group of 5.35 +/- 1.49 min (p less than 0.005). No difference was found when 26 mild hemophiliacs who had received less than 10,000 U of clotting factor concentrate the previous year and no infusions in at least 3 mo were compared with 28 severe hemophiliacs who had received greater than 20,000 U of clotting factor concentrate the previous year and had been infused within 1 mo of testing. Ten patients (16%) had bleeding times greater than 10 min. Bleeding times remained prolonged on repeat evaluations in 7 of these patients, 3 of whom had mild disease and all of whom had normal platelet aggregations in response to arachidonic acid. We conclude that the bleeding time is longer than normal in hemophilia. This abnormality is not related to disease severity, recent transfusions, or the use of nonsteroidal antiinflammatory drugs.
Enzymatic conjugation of erythrocyte glutathione with 1-chloro-2,4- dinitrobenzene: the fate of glutathione conjugate in erythrocytes and the effect of glutathione depletion on hemoglobin
Authors: Awasthi YC , Garg HS , Dao DD , Partridge CA , Srivastava SK
Published: October 2019

Erythrocyte glutathione (GSH) can be rapidly depleted by incubating the cells with 1-chloro-2,4-dinitrobenzene (CDNB), which forms 2,4- dinitrophenyl-S-glutathione with GSH through the reaction catalyzed by glutathione S-transferase. GSH-CDNB conjugate thus formed stays undegraded within the erythrocytes. This indicates that in the erythrocytes, mercapturic acid pathway is inoperative. Depletion of GSH in the intact erythrocytes by CDNB results in rapid oxidation of large amounts of hemoglobin to methemoglobin. When glutathione S-transferase- free hemolysate of erythrocytes is incubated with CDNB, the depletion of GSH as well as methemoglobin formation are minimal. Glutathione peroxidase and glutathione reductase activities of the erythrocytes are not affected by CDNB. These studies provide a specific enzymatic method for rapid removal of erythrocyte GSH and also indicate that GSH is vital in maintaining a reduced environment within the erythrocytes.
Lymphocyte dysfunction in chronic graft-versus-host disease
Authors: Saxon A , McIntyre RE , Stevens RH , Gale RP
Published: October 2019

Three recipients of HLA-identical bone marrow transplants developed chronic graft-versus-host disease (cGVHD) and hypergammaglobulinemia. All three had evidence of abnormal B-lymphocyte function, including a polyclonal increase in immunoglobulins (Ig), antinuclear antibodies, rheumatoid factor, lymphocytotoxins, and increased immune complexes. T- lymphocyte function was also abnormal, including decreased mitogen reactivity and delayed cutaneous hypersensitivity. The cellular basis of these immune abnormalities was studied in an in vitro system in which we analyzed spontaneous pokeweed mitogen (PWM) driven Ig synthesis. Multiple defects in both T- and B-lymphocyte function were detected. In contrast to normal B cells, circulating B cells from all three patients with cGVHD spontaneously synthesized in vitro greater than 200 ng of IgG and in two of the three greater than 175 ng of IgM. This increase in spontaneous Ig synthesis was not due to a deficiency of regulatory cells, since T cells from the three patients suppressed spontaneous Ig synthesis in a normal fashion. In contrast to this increased spontaneous Ig synthesis, the response of the patients' B cells to PWM-driven Ig synthesis was normal. Using the PWM system we demonstrated several defects in these patients' T cells, including increased suppressor activity and decreased helper cell activity. These data indicate that some patients with cGVHD have multiple defects in both T- and B-cell function that may contribute to their profound immune deficiency.
Nonrandom chromosome abnormalities in acute leukemia and dysmyelopoietic syndromes in patients with previously treated malignant disease
Authors: Rowley JD , Golomb HM , Vardiman JW
Published: October 2019

Cytogenetic studies were performed on 26 patients who developed acute nonlymphocytic leukemia (ANLL) or a dysmyelopoietic syndrome after treatment of a primary malignancy. Fifteen patients had radiotherapy and chemotherapy, seven had only chemotherapy, and four had only radiotherapy. The median times from diagnosis of the initial disease to the development of bone marrow dysfunction for these treatment groups were 50, 46, and 49 mo, respectively. Twenty-five patients had an abnormal karyotype in myeloid cells. Loss of part or all of no. 5 and/or no. 7 was noted in 23 of 25 patients with aneuploidy. Loss of no. 5 was noted only in patients who previously had malignant lymphoma, whereas loss of no. 7 was seen in these patients as well as in those who had other malignancies. Abnormalities of both nos. 5 and 7 occurred in 53% of the patients treated with combined therapy and in only 27% of patients treated with either modality alone. Although these changes are distinctly different from those noted in lymphomas, they are similar to those seen in 25% of aneuploid patients with ANLL de novo.
Autostimulation of growth by human myelogenous leukemia cells (HL-60)
Authors: Brennan JK , Abboud CN , DiPersio JF , Barlow GH , Lichtman MA
Published: October 2019

We have studied the effects of medium conditioned by the human progranulocytic leukemia cell line, HL-60, on the subsequent growth of new inocula of HL-60 cells. When HL-60 cells were cultured at high cell density, optimal growth rate occurred in liquid suspension and confluent colony growth was observed in viscous medium without the addition of conditioned medium. However, when cells were cultured at lower cell density, growth rate was reduced and colony growth was nil unless conditioned medium from HL-60 culture was added. All HL-60 populations studied, including the earliest available passage, 9, both elaborated and responded to HL-60 CM. HL-60 CM did not stimulate normal human or mouse granulocyte-monocyte colony-forming cell (CFU-GM) growth. Conditioned media from other human cell lines varied in the ability to stimulate HL-60 cell and CFU-GM proliferation. Some, such as GCT CM, stimulated both HL-60 cells and normal CFU-GM, whereas others, like HL-60 CM, stimulated only HL-60 growth. The majority of cell line CMs tested did not stimulate either HL-60 or CFU-GM. Chromatography of HL-60 CM on Ultrogel AcA54 showed a single peak of HL-60 stimulating activity of apparent molecular weight 13,000. The ability of HL-60 cells to elaborate this activity provides a possible explanation for their proliferation at higher cell densities. Autostimulation may prove to be important in the high growth potential of other cell populations that undergo unrepressed proliferation.
Production and characterization of monoclonal antibodies to the subunits of human phosphofructokinase: new tools for the immunochemical and genetic analyses of isozymes
Authors: Vora S , Wims LA , Durham S , Morrison SL
Published: October 2019

Recently we have demonstrated that human phosphofructokinase (PFK; ATP: D-fructose-6-P, 1-phosphotransferase; EC.2.7.1.11) is under the control of three structural loci that code for M (muscle-type), L (liver-type), and P (platelet-type) subunits: random tetramerization of these subunits produces various isozymes. In this study, we have produced and characterized BALB/c hybridoma antibodies to the M- and L-type subunits of human PFK. The specific antibodies were detected by an enzyme- immunoprecipitation assay using Staphylococci-bearing protein A as an immunoadsorbent. Of the wells tested using red blood cell (RBC) PFK (M + L), 61% were positive. Only one M-specific hybridoma was identified. The one anti-M and 4 anti-L antibodies were characterized for their biochemical and immunochemical specificities. To define the combining specificities of these antibodies, we compared their reactivity and that of monospecific rabbit anti-M antiserum with muscle and liver PFKs from 15 different vertebrate species. The rabbit anti-M shows strong cross-reactivity with the muscle PFKs from all the species studied. In contrast, the monoclonal anti-M reacts exclusively with muscle PFKs from primates. Two of four anti-L antibodies react only with human L- PFK, whereas the other two react with that from a few other vertebrate species as well. Taken together, these data suggest that primate- specific antibodies recognize evolutionarily, recently acquired antigenic determinants, whereas the antibodies reactive with PFKs from distantly related species recognize conserved determinants. The differential immunoreactivities of muscle and liver PFKs strongly suggest the presence of distinct isozymes in all the vertebrate species studied. These studies demonstrate that it is feasible to produce and characterize monoclonal antibodies that distinguish among isozymes with structural and functional similarities. These antibodies provide sensitive tools in the analyses of isozyme structure, genetics, and related fields.
Human alveolar macrophages express Ia-like antigens
Authors: Hocking W , Billing R , Foon K , Golde D
Published: October 2019

Monoclonal antibody to la-like antigens was used to demonstrate the presence of these antigens on human alveolar macrophages. Immunoprecipitation demonstrated the 27,000 and 34,000 molecular weight peaks that correspond to the la-like antigen subunits. Immunofluorescence confirmed the presence of la-like antigens on alveolar macrophages but not on other bronchoalveolar cells. The presence of alveolar macrophage la-like antigens may be important in cellular interactions.
Tobacco, cocoa, coffee, and ragweed: cross-reacting allergens that activate factor-XII-dependent pathways
Authors: Becker CG , Van Hamont N , Wagner M
Published: October 2019

A glycoprotein antigen has been isolated from cured tobacco leaves (TGP- L) Nicotiana tabacum) and from cigarette smoke condensate (TGP-CSC) to which approximately one-third of human volunteers, smokers and non- smokers, exhibit immediate cutaneous hypersensitivity. TGP-L and TGP- CSC contain polyphenol haptens that activate the factor XII (Hageman factor) dependent pathways of coagulation, fibrinolysis, and kinin generation in normal human plasma. The purpose of this communication is to describe the isolation antigens from cocoa powder (Theobroma cacao), ground coffee (Coffea arabica), and ragweed (Ambrosia eliator) pollen that are immunologically cross-reactive with TGP-L and TGP-CSC, contain similar polyphenol haptens, and are capable of activating factor-XII- dependent pathways in normal human plasma.
The effects of dietary omega 3 fatty acids on platelet composition and function in man: a prospective, controlled study
Authors: Goodnight SH Jr , Harris WS , Connor WE
Published: October 2019

The rarity of atherosclerotic vascular disease and a mild bruising tendency in Greenland Eskimos has been linked to their ingestion of omega 3 fatty acids contained in foods obtained from the sea. Previous studies have shown that feeding salmon oil to normal volunteers resulted in reductions of plasma cholesterol and triglycerides. We wished to learn whether salmon oil feeding would result in the incorporation of omega 3 fatty acids into platelets and whether platelet function or platelet-vessel interaction would be altered. Diets containing salmon oils led to the incorporation of eicosapentaenoic acid (C20:5 omega 3) into platelets (6.1%) with a reduction in arachidonic acid (C20:4 omega 6). The ratio of C20:5/C20:4 increased from 0.0045 on the control diet to 0.3 on the salmon diet. Bleeding times were prolonged (from 6.75 to 10 min, p less than 0.005), platelet retention on glass beads was mildly reduced (from 89% to 78%, p less than 0.0005), and platelet aggregation in response to dilute concentrations of ADP was inhibited in the subjects ingesting the salmon oil. We conclude that in normal subjects dietary omega 2 fatty acids derived from salmon oil are incorporated into platelet phospholipids and that these changes are accompanied by alterations in bleeding time and platelet function.
Existence of a pool of T-lymphocyte colony-forming cells (T-CFC) in human bone marrow and their place in the differentiation of the T- lymphocyte lineage
Authors: Triebel F , Robinson WA , Hayward AR , Goube de Laforest PG
Published: October 2019

The existence and characteristics of bone marrow T-cell progenitors have not yet been established in man. Several pieces of evidence such as the reconstitution of certain immunodeficiencies by bone marrow graft suggest that T-cell precursors are present in the bone marrow. We report the growth of T-cell colonies from bone marrow populations using PHA-stimulated lymphocyte-conditioned medium containing T-cell growth factor (TCGF). Rosetting experiments and complement-dependent cytotoxicity assays with monoclonal antibodies indicate that the bone marrow T colony-forming cells (T-CFC) are E- OKT 3- and la+, i.e., immature progenitors. The colonies derived from these cells have the phenotype of mature T cells: E + OKT 3 + la- with either helper (OKT 4+) and suppressor (OKT 8 +) antigens. These results suggest that a thymic microenvironment may not be necessary for the in vitro proliferation and differentiation of the T-cell lineage in adult humans. These methodologies may permit direct investigation of early phenomena concerning the T-cell lineage, such as the acquisition of self-tolerance, the formation of a repertoire of specificities, and the HLA restriction phenomena that we believe takes place before the thymic maturation.
Long-term suspension cultures of human cord blood myeloid cells
Authors: Salahuddin SZ , Markham PD , Ruscetti FW , Gallo RC
Published: October 2019

A method is described that allows for the routine long-term (greater than 3 mo) growth of normal, immature human myeloid cells in liquid suspension culture. The techniques employ cell separation, utilization of special growth conditions (RPMI with hydrocortisone and vitamin D), and cord blood as the source of leukocytes. These cultures are composed predominantly of immature myeloid cells that have grown for over 3 mo in culture but eventually terminate as differentiated, mature granulocytes and monocyte-macrophages. Application of these techniques to other sources of fresh human leukocytes (bone marrow and adult peripheral blood) resulted in only short-term proliferation of myeloid cells. The techniques described can be used for the routine expansion of immature myeloid and monocytoid cell populations, particularly from newborns, and should be useful for systematic studies of normal myeloid- monocytoid cell growth and differentiation and providing normal control cells in studies employing human leukemic myeloid cells.
Nonspecific esterases of the formed elements: zymograms produced by pH 9.5 polyarcrylamide gel electrophoresis
Authors: Yourno J , Mastropaolo W
Published: October 2019

The formed elements of human blood each contain multiple isoenzymes of nonspecific esterase that hydrolyze short chain alpha naphthyl esters. Zymograms that are characteristic of each type of formed element are obtained by subjecting purified preparations of each to polyacrylamide slab gel electrophoresis at pH 9.5 and subsequent staining of the gels for esterase activity. The most prominent isoenzyme detected is a species of low mobility that is reactive with either acetyl or butyryl esters and is highly sensitive to inhibition by 40 mM sodium fluoride. Also detected are several major acetyl esterases and a single butyryl esterase, all of which are relatively fluoride resistant. The intercellular distribution of isoenzymes varies from element-specific to pancellular. The prominent fluoride-sensitive acetyl, butyryl esterase, is the major isoenzyme of monocyte zymograms, which is consistent with the well known cytochemistry of monocytes. Lesser but significant amount (2%-3% of monocyte levels) of this isoenzyme were also detected in granulocyte zymograms. This system may prove useful in the study of differentiation of blood cells and in the classification of acute leukemias.
Modulation of erythrocyte-endothelial interactions and the vasocclusive severity of sickling disorders
Authors: Hebbel RP , Moldow CF , Steinberg MH
Published: October 2019

The abnormal adherence of sickle erythrocytes to cultured human vascular endothelium varies among patients and correlates with vasocclusive severity within and among the different sickling disorders. For individual patients, the development of an acute vasocclusive event is not accompanied by an increase in the inherent propensity for their erythrocytes to adhere to endothelium. However, the onset of vasocclusion appears to be associated with the appearance of plasma factors, such as fibrinogen, which enhance the adherence of sickle erythrocytes to endothelium. In addition, sickle erythrocytes adhere even more avidly to injured endothelium and under hyperosmolar conditions. Thus, the overall clinical severity of the sickling disorders appears to correlate with the inherent propensity for sickle erythrocytes to adhere to endothelium, and changes in the red cells' environment may help precipitate crises by the sudden facilitation of erythrocyte/endothelial interactions.
The diagnosis of iron deficiency anemia in sickle cell disease
Authors: Vichinsky E , Kleman K , Embury S , Lubin B
Published: October 2019

We determined the prevalence and optimal methods for laboratory diagnosis of iron deficiency anemia in patients with sickle cell disease. Laboratory investigations of 38 nontransfused and 32 transfused patients included transferrin saturation, serum ferritin, mean corpuscular volume (MCV), and free erythrocyte protoporphyrin (FEP). Response to iron supplementation confirmed the diagnosis of iron deficiency anemia in 16% of the nontransfused patients. None of the transfused patients were iron deficient. All iron-deficient patients (mean age 2.4 yr) had a low MCV, serum ferritin less than 25 ng/ml, transferrin saturation less than 15%, and FEP less than 90 micrograms/dl RBC. Following therapy, all parameters improved and the hemoglobin concentration increased greater than 2 g/dl. A serum ferritin below 25 ng/ml was the most reliable screening test for iron deficiency. There were 13% false positive results with transferrin saturation, 3% with MCV, and 62% with FEP. FEP values correlated strongly with reticulocyte counts. The high FEP was in part due to protoporphyrin IX and not completely due to zinc protoporphyrin, which is elevated in iron deficiency. We conclude that iron deficiency anemia is a potential problem in young nontransfused sickle cell patients. Serum ferritin below 25 ng/ml and low MCV are the most useful screening tests.
Opsonized zymosan-stimulated granulocytes-activation and activity of the superoxide-generating system and membrane potential changes
Authors: Cohen HJ , Newburger PE , Chovaniec ME , Whitin JC , Simons ER
Published: October 2019

Phagocytic cells generate superoxide in response to stimulation by opsonized particles. A continuous assay for opsonized zymosan- stimulated granulocyte superoxide production shows that there is a lag time between the addition of particles and the onset of detectable superoxide production. Superoxide production is preceded by membrane potential depolarization. Neither superoxide production nor membrane depolarization occurs in granulocytes from patients with chronic granulomatous disease. The extent of activation by opsonized zymosan is affected by the dose of zymosan from 0.5 to 4.5 mg/ml, but the time necessary for activation (lag time) is not. Similarly, the extent of depolarization but not the time necessary for attaining maximum depolarization is concentration-dependent. Effects of temperature, divalent cations, 2-deoxyglucose, cyanide, and N-ethyl maleimide on superoxide production are similar for granulocytes treated with soluble stimuli and with opsonized zymosan. Thus, zymosan stimulates granulocytes to generate superoxide and undergo membrane depolarization in a manner similar to that elected by soluble stimuli.
The presence and release of alpha 2-antiplasmin from human platelets
Authors: Plow EF , Collen D
Published: October 2019

An antigen immunochemically indistinguishable from plasma alpha 2- antiplasmin, the primary plasmin inhibitor, was detected in human platelets. By radioimmunoassay, 33–114 ng alpha 2-antiplasmin antigen was quantitated in the detergent-soluble extract of 10(9) washed human platelets from 10 normal donors with a mean level of 62 +/- 24 ng/10(9) platelets. Plasma alpha 2-antiplasmin, either in the platelet suspending medium or on the surface of the platelets, could account for less than 8% of the antigen present in the platelet extracts. When stimulated with thrombin, the platelets released alpha 2-antiplasmin antigen without cell lysis, and greater than 85% of the alpha 2- antiplasmin antigen was released at a high thrombin dose. At a lower dose of thrombin, alpha 2-antiplasmin and platelet factor 4 were partially released without concomitant secretion of serotonin. No alpha 2-antiplasmin antigen was detected in extracts or red blood cells, polymorphonuclear leukocytes, and adherent and nonadherent mononuclear cells. Thus, the platelet is the only peripheral blood cell containing significant amounts of alpha 2-antiplasmin.
Platelet von Willebrand's antigen II: active release by aggregating agents and a marker of platelet release reaction in vivo
Authors: Scott JP , Montgomery RR
Published: October 2019

von Willebrand's antigen II (vW AgII), a plasma and platelet antigen immunologically and biochemically distinct from factor-VIII-related antigen (VIIIR:Ag), is decreased in von Willebrand's disease. vW AgII is released from platelets during aggregation and clotting. The release of platelet vW AgII was studied in washed platelets following aggregation by thrombin, collagen, and adenosine diphosphate (ADP). Total platelet vW AgII was 3.39 U/10(9) platelets. Thrombin and collagen yielded release of 85% and 86% of platelet vW AgII, respectively, at the highest concentrations. Release of platelet vW AgII was correlated with the release of 5-hydroxytryptamine (5HT). Release of vW AgII by collagen and thrombin was inhibited by metabolic inhibitors. In addition, collagen release of vW AgII was inhibited by aspirin. In clinical syndromes associated with intravascular platelet destruction, marked elevations of plasma vW AgII were noted. Thus, vW AgII is released by a metabolically active process from platelets during aggregation. In addition, vW AgII appears to be a marker of intravascular platelet destruction.
Ultrastructural localization of nonspecific esterase activity in guinea pig and human monocytes, macrophages, and lymphocytes
Authors: Monahan RA , Dvorak HF , Dvorak AM
Published: October 2019

Using either hexazotized pararosaniline or new fuchsin as coupling agents, we investigated the ultrastructural localization of alpha- naphthyl acetate esterase (ANAE) activity in guinea pig bone marrow and peritoneal exudates, and in human peripheral blood cells. DFP- inhibitable ANAE activity was present on the cell surface of lymphocytes, monocytes, macrophages, neutrophils, eosinophils, basophils, megakaryocytes, platelets, and blasts. Demarcation lines in megakaryocytes and the perinuclear cisternae in normoblasts were also positive. In addition, lymphocytes, monocytes, and macrophages displayed ANAE activity associated with cytoplasmic-vesicle clusters (CVC). Reaction product was always present on the cytoplasmic surfaces of these vesicles and in the adjacent cytoplasm; vesicle interiors were invariably ANAE-negative. Small lymphocytes generally had a single large paranuclear ANAE-positive CVC, whereas mononuclear phagocytes had multiple discrete foci of similar appearing ANAE-positive CVC that sometime became confluent. ANAE activity was also found in the Gall bodies of human lymphocytes and in coated vesicles of macrophages. Cytoplasmic ANAE activity was increased in oil-induced guinea pig peritoneal macrophages. Both surface and cytoplasmic esterase activities had a neutral pH optimum. An identical distribution of reaction product was observed when alpha-naphthyl butyrate was employed as substrate. The function of these esterases, and their relation to known surface and cytoplasmic neutral proteases, awaits further investigation.
Hydrophobic adsorption chromatography of colony-stimulating activities and erythroid-enhancing activity from the human monocyte-like cell line, GCT
Authors: Abboud CN , Brennan JK , Barlow GH , Lichtman MA
Published: October 2019

The human cell line, GCT, secretes hemopoietins into serum-free culture medium. The conditioned medium contains activities that stimulate neutrophil-monocyte, macrophage, eosinophil, and erythroid colony growth in human marrow cultures. We have used hydrophobic adsorption chromatography to separate a neutrophil-monocyte colony-stimulating factor (CSF) from the other colony-stimulating activities. This hydrophobic CSF has no eosinophil-stimulating activity and is virtually devoid of erythroid-stimulating activity.
Somatic cell hybrid analyses of hematopoietic differentiation
Authors: Koeffler HP , Sparkes RS , Billing R , Klein G
Published: October 2019

A differentiated cell expresses an entire set of specialized features. Somatic cell hybridization provides a method to examine control of gene regulation. We studied the expression of tissue-specific features in hybrids between human promyelocytes (HL-60) and human Burkitt's lymphoma cells (P3HR-1). Two hybrid lines, HP-1 and HP-2, and 18 hybrid clones were established and confirmed by karyotype, isozyme, and surface antigen analyses. The hybrids extinguished the 10 myeloid (HL- 60) features that we examined including myeloid morphology, histochemistry, and functions that included response to colony- stimulating factor and ability to differentiate to granulocytes or macrophages. In contrast, the hybrids synthesized immunoglobulin and expressed Epstein-Barr nuclear, early, and viral capsid antigens similar to the P3HR-1 lymphoid parental line. Results are contrasted to the findings when P3HR-1 lymphocytes are fused to human erythroid- myeloid cells (K562). Taken together, our results suggest that phenotypic differences between human myeloid and lymphoid cells in the hematopoietic lineage involve mutually exclusive programs and may possibly be mediated by the activity of diffusible, transacting molecules.
Serum erythropoietin (ESF) titers in polycythemia
Authors: de Klerk G , Rosengarten PC , Vet RJ , Goudsmit R
Published: October 2019

Serum ESF titers were measured in 42 polycythemic patients using the fetal mouse liver cell bioassay. ESF titers in patients with secondary polycythemia differed significantly from those in patients with polycythemia vera (p less than 0.0001). Among the 21 patients with secondary polycythemia, 1 patient had an ESF titer less than 10 mU/ml (the lower limit of sensitivity) and 20 had ESF titers that ranged between 11 and 112 mU/ml, with a mean titer of 56 mU/ml. Among the 21 patients with polycythemia vera, 13 patients had ESF titers less than 10 mU/ml and 8 had ESF titers ranging between 12 and 55 mU/ml, with a mean titer of 26 mU/ml. The mean hemoglobin concentration in the 8 patients with ESF titers greater than 10 mU/ml was significantly below that in the 13 polycythemia vera patients with ESF titers less than 10 mU/ml (p less than 0.03). If ESF titers less than 10 mU/ml had been indicative of polycythemia vera and ESF titers greater than 10 mU/ml had been indicative of secondary polycythemia in patients with hemoglobin concentrations greater than 17.7 g/dl, but not indicative of either condition in patients with hemoglobin concentrations less than 17.7 g/dl, 71.5% of the polycythemic patients in this study would have been diagnosed correctly, 9.5% incorrectly, and in the 19% the diagnosis would have remained uncertain. It was concluded that measurement of serum ESF titers using this in vitro bioassay can be of clinical importance in differentiating between polycythemia vera and secondary polycythemia.
Three patients with structurally abnormal X chromosomes, each with Xq13 breakpoints and a history of idiopathic acquired sideroblastic anemia
Authors: Dewald GW , Pierre RV , Phyliky RL
Published: October 2019

Structural abnormalities of the X chromosome are rarely found in neoplastic disorders. We describe three patients with a history of idiopathic acquired sideroblastic anemia (IASA); each one had an abnormal clone of cells in the bone marrow, characterized by a structurally abnormal X chromosome. In two of these patients, the predominant karyotype was 47,X,2idic(X)(q13); in the other patient, it was 46,X,t(X;11)(q13;p15). Inasmuch as all three of these cases involved chromosome band Xq13, as did two previously published cases, we suggest that band Xq13 may be more prone to structural rearrangement than other X chromosome bands in hematologic disorders. The common Xq13 chromosome breakpoint and clinical presentation (IASA) among these three patients and the occurrence of an X-linked type of sideroblastic anemia may suggest that an association exists between X chromosome abnormalities and IASA. Perhaps alteration of a gene or chromosome structure in or near band Xq13 predisposes to development of IASA. The fact that two of these patients had preleukemia and the third had overt acute leukemia may imply that patients with IASA and X chromosome abnormalities have a poor prognosis. Cases of IASA without associated X chromosome abnormalities are known; thus, if an association between IASA and an abnormal X chromosome does exist, most likely it involves only some patients with IASA.
Reversal of acute ("malignant") myelosclerosis by allogeneic bone marrow transplantation
Authors: Wolf JL , Spruce WE , Bearman RM , Forman SJ , Scott EP , Fahey JL , Farbstein MJ , Rappaport H , Blume KG
Published: October 2019

A 28-yr-old woman with acute malignant myelosclerosis received, as primary treatment, ablative chemotherapy and total body radiation therapy followed by bone marrow transplantation from her histocompatible brother. The patient is now well more than 15 mo after bone marrow transplantation, with normal peripheral blood counts, a normal bone marrow, no evidence of graft-versus-host disease, and is on no therapy. In light of the poor results obtained with conventional chemotherapy in this disease, bone marrow transplantation may represent the treatment of choice for patients who have an appropriate donor.
Specificity of autoantibodies in autoimmune thrombocytopenia
Authors: van Leeuwen EF , van der Ven JT , Engelfriet CP , von dem Borne AE
Published: October 2019

In 42 patients with autoimmune thrombocytopenia (AITP) and a positive direct platelet suspension immunofluorescence test (PSIFT), the antigenic specificity of the autoantibodies was studied. Because the autoantibodies were often not detectable in the serum and additional HLA antibodies may disturb the reaction pattern with the platelet panel, we used eluates prepared from the patients' platelets for this study. Thirty-five patients had antibodies equally reactive with normal platelets, irrespective of their antigenic make-up, but not with the platelets from two Glanzmann's disease patients. Absorption and elution experiments in two patients showed that his was probably not due to the presence of a combination of anti-Zwa and anti-Zwb antibodies. Thus, the majority of autoantibodies against platelets seems to be directed against antigenic determinants not present on Glanzmann's disease platelets, but perhaps located on the platelet-membrane glycoproteins IIb and/or IIIa. In ten patients, antibodies of no, or still unknown, specificity were detected. Three of these had additional antibodies not reactive with the platelets of the two Glanzmann patients.
Cyclic neutropenia (CN): a clue to the control of granulopoiesis
Authors: von Schulthess GK , Mazer NA
Published: October 2019

A simple quantitative feedback model of granulopoiesis is presented and discussed within the framework of existing data on granulopoiesis in both normals and patients with cyclic neutropenia (CN). The model assumes that the controlled compartment is the bone marrow pool of mature neutrophils (PMNs), which sends a negative feedback signal to the mitotic pool of early granulocyte precursors (i.e., CFU-C, myeloblasts, etc.) thus controlling the granulocyte production rate. Three parameters are found to play important roles in determining the response of the system to perturbations. These are: TM, the granulocyte maturation time; a, a parameter reflecting the strength of the negative feedback exerted by mature PMNs on the granulocyte production rate; and b, a parameter describing the leakiness of the bone marrow for PMN egress. It is shown that depending on the relative magnitudes of a and b, the system will either respond to perturbations with a damped oscillation (a less than b: the normal state) or with a sustained oscillation (a greater than b: the CN state). In both cases, the oscillation period is found to approximately equal 2TM. Deductions of the values of a, b, and TM from experimental data are consistent with the predictions of the model and show an increased value of a in CN relative to the normal state. This suggests an overly active feedback mechanism as the pathophysiologic basis of CN. In addition, the model can explain how various therapeutic agent correct CN and also provides insight into why other hematologic cell lines and CSA oscillate in CN.
Serologic profile of alphamethyldopa-induced hemolytic anemia: correlation between cell-bound IgM and hemolysis
Authors: Lalezari P , Louie JE , Fadlallah N
Published: October 2019

Erythrocyte-bound immunoglobulins have been characterized by a PVP- potentiated antiglobulin test in 11 patients who had developed antibodies after treatment with alpha-methyldopa. Serologic profiles were recognized that could distinguish between the hemolyzing and nonhemolyzing patients: IgM antibodies together with the first component of complement (C1q) were demonstrated on erythrocytes of all eight hemolyzing patients. By contrast, these immunoproteins were absent from the cells of nonhemolyzing patients and became undetectable when the hemolyzing patients recovered. IgG and its subclasses were variably present on erythrocytes of all patients regardless of hemolytic activity. Eluates prepared from erythrocytes of the hemolyzing patients were shown to contain both IgG and IgM, and fixes C1q, C3, and C4. Eluates from the nonhemolyzing patients contained only IgG. The IgM antibodies differed from the commonly occurring cold agglutinins in that they were warm-reactive and were mainly concentrated on the patients' cells rather than being free in the serum. Because of their nonagglutinating property, it is suggested that they are monomeric IgM. It is concluded that the high affinity, warm- reactive IgM and not the IgG antibodies are primarily responsible for clinically manifest anemia in patients receiving alphamethyldopa and that the hemolytic activity is probably mediated by the classic pathway of complement activation.
Properties of sulfatides in factor-XII-dependent contact activation
Authors: Tans G , Griffin JH
Published: October 2019

Incubation of normal human plasma with low amounts of sulfatides resulted in the initiation of intrinsic coagulation and the appearance of kallikrein activity. The optimal initiation of procoagulant and kallikrein amidolytic activity was dependent on the presence of factor XII, high molecular weight kininogen, and prekallikrein. Since the activated partial thromboplastin clotting times in prekallikrein- deficient plasma approach normal values upon prolonged incubation with kaolin, this phenomenon of autocorrection was studied and found to be even more pronounced in the presence of sulfatides. Autocorrection was essentially completed in 5 min in the presence of sulfatides, whereas a preincubation of 15–20 min was required in the presence of kaolin. The limited proteolysis of 125I-factor XII in plasma during incubation with activating material or during clotting was determined. Cleavage of factor XII was more rapid and more extensive in the presence of sulfatides than in the presence of kaolin. In prekallikrein-deficient plasma, factor XII cleavage was completed within 5 min in the presence of sulfatides and within 15 min in the presence of kaolin. Thus, the appearance of factor-XII-dependent coagulant activity correlates with the limited proteolysis of factor XII when normal or prekallikrein- deficient plasma is activated by sulfatides or kaolin.
Isolation and structural characterization of the polypeptide subunits of membrane glycoprotein IIb-IIIa from human platelets
Authors: McEver RP , Baenziger JU , Majerus PW
Published: October 2019

We have previously demonstrated the isolation of platelet membrane glycoprotein IIb-IIIa by affinity chromatography with a specific monoclonal antibody. We have now separated the polypeptide subunits IIb and IIIa of the isolated glycoprotein by preparative sodium dodecyl sulfate polyacrylamide gel electrophoresis and have compared their structural features. Both IIb and IIIa contain approximately 15% carbohydrate, but IIIa contains a larger percentage of mannose residues, suggesting the presence of high mannose as well as complex N- linked oligosaccharide chains. The amino acid compositions are sufficiently similar to imply areas of sequence homology between the two subunits. To examine further the relationship between the subunits, we digested a mixture of 125I-IIb and 131I-IIIa with trypsin and then separated the radiolabeled peptides by high performance liquid chromatography. The resultant peptide maps of IIb and IIIa are completely different. This indicates that neither subunit is derived from the other and suggests that polypeptides IIb and IIIa are products of separate genes.
A liquid culture system for murine megakaryocyte progenitor cells
Authors: Nagasawa T , Nakazawa M , Abe T
Published: October 2019

A liquid culture system is described for murine megakaryocyte progenitor cells (CFU-M) in the presence of pokeweed-mitogen-stimulated spleen-cell conditioned medium. There were dose-related responses between the number of CFU-M developed and the number of cells cultured and the dosage of conditioned medium in this liquid culture system. Murine CFU-M were abundantly cloned in this system an the plating efficiency was similar in comparison with that in a plasma clot system. The acetylcholinesterase-positive colonies (more than 4 acetylcholinesterase-positive cells) were clearly seen on day 3 of culture, and they reached a maximum (60.5 +/- 10.7/2 x 10(5) cells) on day 7 of culture. Ultrastructural analyses of megakaryocytic maturation in this system showed that a few megakaryocytes produced platelets that were released in the culture medium on day 5 of culture. This liquid culture system is suitable for the study of the dynamic process of the megakaryocyte-platelet system.
Mechanism of dexamethasone inhibition of chemotactic factor induced granulocyte aggregation
Authors: Oseas RS , Allen J , Yang HH , Baehner RL , Boxer LA
Published: October 2019

The reaction of FMLP with granulocytes causes aggregation and degranulation and enhances adherence to endothelium. To evaluate whether prevention of granule extrusion could impair these granulocyte activities, granulocytes were treated with either dexamethasone or hydrocortisone prior to treatment with FMLP. Dexamethasone was added to suspensions of cytochalasin B-treated granulocytes; it markedly impaired the aggregation response of the granulocytes of FMLP. When cytochalasin-B was not used, granulocyte aggregation in response to FMLP or PMA was inhibited by dexamethasone. Although dexamethasone prevented aggregation of cells following stimulation with FMLP or PMA, it failed to prevent the aggregation of granulocytes induced by rabbit lactoferrin. Adherence of granulocytes to human endothelial monolayers was enhanced by FMLP; dexamethasone inhibited the enhancement. However, with the addition of human lactoferrin to the granulocytes exposed to dexamethasone, the cells were able to adhere as well to endothelium as the cells exposed to FMLP but free of dexamethasone. When cytochalasin- B-treated granulocytes were incubated with dexamethasone or hydrocortisone prior to the addition of FMLP, the subsequent release of lactoferrin was substantially blocked, whereas the release of the primary granule products, lysozyme and beta-glucuronidase, was attenuated but not completely blocked. Thus, corticosteroids might block chemotactic-factor-induced granulocyte aggregation by selectively preventing release of specific granule products that contribute to and sustain aggregation.
Human leukemia cell line K562 responds to erythroid-potentiating activity
Authors: Gauwerky CE , Lusis AJ , Golde DW
Published: October 2019

We report that erythroid-potentiating activity (EPA), known to stimulate the proliferation of normal human erythroid precursors in vitro, has a growth-promoting effect on human K562 erythroleukemia cells and Friend mouse erythroleukemia cells. Detailed studies were carried out using an EPA produced by a human T-lymphoblast line (Mo). Although EPA has not been purified to homogeneity, several observations indicate that the factor elaborated by Mo cells that stimulates erythroleukemia cell growth is the EPA molecule. The erythroleukemia growth factor cofractionates with EPA using gel exclusion chromatography, isoelectric focusing, and ion exchange chromatography. In addition, the activities exhibit similar kinetics of heat inactivation. A granulocyte-macrophage colony-stimulating factor also elaborated by Mo cells had no effect on the growth of the erythroleukemia cells. Other sources of EPA, such as peripheral blood leukocyte-conditioned medium, preparations from urine of anemic patients, and medium conditioned by a human monocyte-like cell line, stimulated erythroleukemia cell growth. Mouse sources of EPA (termed “burst-promoting activity”) stimulated mouse but not human erythroleukemia cells. The availability of cell lines apparently responsive to EPA should prove useful for examining the mode of action of this regulator of erythropoiesis.
Enrichment for CFU-C from murine and human bone marrow using soybean agglutinin
Authors: Reisner Y , Kapoor N , Hodes MZ , O'Reilly RJ , Good RA
Published: October 2019

Mouse bone marrow and spleen cells agglutinated by soybean agglutinin (SBA) or peanut agglutinin (PNA) were previously shown to be enriched for spleen colony-forming cells (CFU-S) and sufficiently depleted of graft-versus-host reaction producing cells to allow hematologic reconstitution of lethally irradiated allogeneic recipient mice. A similar enrichment for cells capable of forming colonies in soft agar culture (CFU-C) has now been found in the SBA-agglutinated fraction of mouse bone marrow cells, in contrast to the finding that in human bone marrow the majority of the CFU-C are in the fraction not agglutinated by SBA. Cytofluorometric studies with fluorescein-labeled SBA (FITC- SBA) revealed that the majority of both mouse and human bone marrow cells bind the lectin. Experiments mixing the human marrow fractions separated by SBA reveal that true enrichment for CFU-C is achieved in the unagglutinated fraction, as opposed to a possible depletion of a suppressor cell population. Granulocytic, monocytic, and mixed cell colonies were all enriched in the SBA-unagglutinated cell fraction from human bone marrow.
Dexamethasone synergistically induces chemotactic peptide receptor expression in HL-60 cells
Authors: Skubitz KM , Zhen YS , August JT
Published: October 2019

We have examined the effect of physiologic concentrations of dexamethasone on the induction of differentiation of the human promyelocytic leukemia cells (HL-60) and the activity of surface receptors for the f-met-leu-phe chemotactic peptide. Dexamethasone at a concentration of 500 nM was markedly synergistic with dimethylformamide and dimethylsulfoxide, but not with trans-retinoic acid, in inducing f- met-leu-phe binding by HL-60 cells. The increase in peptide binding was due to an increase in the activity of peptide receptors rather than an increase in receptor affinity. The effect of dexamethasone was relatively specific for peptide binding as there was little effect when HL-60 differentiation was assessed by cell morphology or the ability of the cells to reduce nitroblue tetrazolium. Dexamethasone alone had no reproducible effect on HL-60 differentiation as assessed by the three criteria tested. The dexamethasone concentration required for half maximal synergism correlated with the known binding constant of the hormone for steroid receptors in HL-60 cells but the effect of dexamethasone was not blocked with the steroid blocking agents 17 alpha methyltestosterone or progesterone.