bluuebunny/crossref_metadata_2025 huggingface dataset:
Biologic and prognostic significance of the presence of more than two mu heavy-chain genes in childhood acute lymphoblastic leukemia of B precursor cell origin
Authors: Kitchingman GR , Mirro J , Stass S , Rovigatti U , Melvin SL , Williams DL , Raimondi SC , Murphy SB
Published: October 2019

We examined the arrangement of the mu heavy-chain immunoglobulin (Ig) genes in the leukemic blast cell DNA of 93 children with acute lymphoblastic leukemia (ALL). All cases met morphologic and cytochemical criteria for ALL, lacked detectable T cell surface antigens, and expressed HLA-DR (Ia) antigens. Eighty-three of the 93 patients (89%) were positive for the common acute lymphoblastic leukemia antigen (CALLA), and 20 of 91 (22%) tested had detectable cytoplasmic immunoglobulin. As expected, the heavy-chain lg gene was rearranged in all cases, and the pattern of rearrangements was variable; 23 had one allele rearranged and one in the germ line configuration; 15 had one rearranged and one deleted; and 37 had two rearranged. Unexpectedly, in 18 patients the presence of more than two mu gene-hybridizing bands was detected. Combinations of enzymes and heavy-chain gene probes were used to confirm that the extra bands were not the result of underdigestion of the DNA or DNA restriction site polymorphism. In eight of the 18 patients, we identified an extra chromosome 14 as a possible cause of the extra bands' hybridizing to the mu heavy-chain constant-region probe. In the remaining ten patients, the presence of three or four bands hybridizing with the mu probe suggests the presence of two populations of leukemic cells that may have arisen either by separate leukemic transformation events or by clonal evolution of one clone into two related lines. Although preliminary (2-year follow-up), our data suggest that childhood ALL of B lineage with more than two mu heavy-chain genes, but without extra copies of chromosome 14, may be more resistant to therapy.
Hemoglobin switching during murine embryonic development: evidence for two populations of embryonic erythropoietic progenitor cells
Authors: Wong PM , Chung SW , Reicheld SM , Chui DH
Published: October 2019

Explants of normal mouse embryonic tissues and disaggregated embryonic single cells were cultured in vitro to study the erythropoietic progenitor cells present during embryonic development. The results indicate that there are two populations of erythropoietic progenitor cells committed to different hemoglobin synthetic programs. These progenitor cells are present at an early gestational stage prior to the formation of the fetal hepatic primordium. One population of progenitors can be stimulated by erythropoietin alone to form usually small erythroid colonies after culture for six days in vitro. These erythroblasts primarily synthesize embryonic hemoglobins, but produce some adult hemoglobins as well. The other population of progenitors requires stimulation by both erythropoietin and adult spleen cell- conditioned medium, and usually forms large erythroid colonies after culture for six days in vitro. These erythroblasts produce only adult hemoglobins.
Hydroxyurea induction of hemoglobin F production in sickle cell disease: relationship between cytotoxicity and F cell production
Authors: Dover GJ , Humphries RK , Moore JG , Ley TJ , Young NS , Charache S , Nienhuis AW
Published: October 2019

Initial alterations in fetal hemoglobin (HbF) production among eight sickle cell anemia subjects treated with hydroxyurea (Hu) are summarized. Four of these subjects had been previously treated with 5- azacytidine (5-aza). All subjects treated with Hu (50 mg/kg/d for three to five days) had suppression of their total reticulocyte counts by seven days, whereas the four subjects previously treated with 5-aza (2 mg/kg/d for three to five days) had increased reticulocyte counts at day 7. The effect of Hu on increasing the number of HbF-containing reticulocytes (F reticulocytes) is extremely variable, ranging from ten- to less than onefold differences in maximal posttherapy v pretherapy levels. Recovery from marrow suppression did not result in greater than twofold increases in F reticulocyte counts. Mean day 7 F reticulocyte levels in the four subjects treated with both Hu and 5-aza were 4.1 X 10/microL and 15.4 X 10(4)/microL, respectively. Among Hu-treated subjects, increased F reticulocyte production was correlated with low serum creatinine levels and rapid removal of Hu from the plasma. Furthermore, suppression of CFU-E colony formation on day 2 of therapy with Hu was inversely correlated with maximal F reticulocyte response. We conclude that where Hu treatment results in marrow toxicity (decreased reticulocyte counts, decreased CFU-E colony formation) HbF production is less likely to increase. Those sickle cell anemia subjects with minimal renal dysfunction (serum creatinine level, greater than 1.0 mg/dL) exhibit the most cytotoxicity and least F reticulocyte response to Hu.
Mixed hematologic chimerism after allogeneic marrow transplantation for severe aplastic anemia is associated with a higher risk of graft rejection and a lessened incidence of acute graft-versus-host disease
Authors: Hill RS , Petersen FB , Storb R , Appelbaum FR , Doney K , Dahlberg S , Ramberg R , Thomas ED
Published: October 2019

Ninety-six patients with severe aplastic anemia who received a sex- mismatched, HLA-identical allogeneic sibling marrow transplant had sequential cytogenetic analyses performed to determine the incidence and implications of posttransplant mixed hematologic chimerism. Of the 96 patients, 56 (58.3%) became mixed chimeras with coexisting host and donor cells detected in peripheral blood or marrow 14 days or later after transplant, and 40 patients (41.7%) were complete chimeras with 100% donor-type hematopoietic cells. The incidence of mixed chimerism was independent of prior blood production transfusions and infusion of donor buffy coat. The rejection rate was significantly increased in the mixed chimeric group, particularly in patients not receiving buffy coat (14 of 36 rejecting), although overall, the majority (69.7%) retained their first graft. Rejection was seen almost exclusively in patients exposed to multiple transfusions before transplantation. If patients who reject their first graft are censored, the overall incidence of grades II through IV acute graft-v-host disease (GVHD) was significantly reduced in those with mixed chimerism. Transfused patients with mixed chimerism in particular were less likely to develop grades II through IV acute GVHD. The incidence of chronic GVHD was similar in the two groups and did not significantly influence survival. In this study, mixed chimerism persisted for up to 395 days posttransplant, either the first graft being rejected or, more commonly, hematopoiesis reverting to 100% donor-type cells. Mixed lymphohematopoietic chimerism may persist in patients with aplastic anemia who have received matched allogeneic marrow transplants for significant periods before hematopoiesis reverts to donor cell type.
Immunoglobulin gene rearrangements in remission bone marrow specimens from patients with acute lymphoblastic leukemia
Authors: Zehnbauer BA , Pardoll DM , Burke PJ , Graham ML , Vogelstein B
Published: October 2019

Recombinant DNA probes for the joining (JH) segment of the immunoglobulin heavy chain gene were used to detect molecular rearrangements of this gene in the DNA of bone marrow cells obtained during remission of acute lymphoblastic leukemia (ALL). This molecular approach was optimized and found to exceed the sensitivity of conventional morphologic screening for detecting residual leukemia cells; one leukemic cell in 500 normal nucleated bone marrow cells was easily detected using this approach. In the present study, bone marrow from three of seven patients in complete clinical remission (defined morphologically) contained leukemic cells in these proportions. This analysis may be of use in evaluating the status of clinical remission in selected ALL patients.
Monoclonal antibodies binding to the human neutrophil C3bi receptor have disparate functional effects
Authors: Hickstein DD , Locksley RM , Beatty PG , Smith A , Stone DM , Root RK
Published: October 2019

Three monoclonal antibodies (MAb)--OKMI, 7C3, and 60.3-- immunoprecipitated a common 170-kd neutrophil membrane antigen closely associated with, or identical to, the C3bi receptor (CR3). Despite binding to a common receptor, these antibodies displayed marked differences in their effects on C3bi-mediated neutrophil function as assessed by the binding and ingestion of opsonized zymosan and the subsequent triggering of the respiratory burst. Antibody 7C3 caused a time-dependent, irreversible inhibition of the neutrophil oxidative response to opsonized zymosan that correlated with capping of the bound antibody. In contrast, antibody 60.3 caused an immediate inhibition of the neutrophil oxidative response to opsonized zymosan that required the continuous presence of exogenous antibody to achieve the maximal inhibitory effect. Antibody OKMI demonstrated minimal inhibition of O2- release. Despite their functional differences, binding of either 7C3 or 60.3 led to up-regulation of new antigen, presumably from intracellular sites as previously described using OKMI. Crossed immunoprecipitations of radiolabeled neutrophil lysates indicated that each MAb bound to different antigens near or within the CR3 complex. Thus three MAb binding to the neutrophil CR3 receptor each caused receptor up- regulation but had markedly different functional effects on the cell.
Evaluation by quantitative acid elution and radioimmunoassay of multiple classes of immunoglobulins and serum albumin associated with platelets in idiopathic thrombocytopenic purpura
Authors: Hotchkiss AJ , Leissinger CA , Smith ME , Jordan JV , Kautz CA , Shulman NR
Published: October 2019

Immunoglobulins (Igs) and serum albumin were eluted from normal platelets and platelets from patients with idiopathic thrombocytopenic purpura (ITP) with a quantitative acid elution procedure followed by solid-phase radioimmunoassay (SPRIA). Acid elution was shown to release a reproducible fraction of platelet-associated Igs, and the amounts released per platelet were independent of the platelet concentration over a wide range of concentrations. This procedure is suitable for sensitive, reproducible, and specific quantitation of large numbers of samples. Washed platelets from 13 normal donors contained the following components (expressed in femtograms per platelet, mean +/- 2 SEM): IgG, 1.40 +/- 0.26; IgA, 0.72 +/- 0.36; IgM 0.078 +/- 0.036; albumin 7.7 +/- 1.5. Immunoglobulins and albumin eluted from the platelets of ten ITP patients (two in remission), expressed as femtograms per platelet, mean (range), were: IgG 104 (0.3 to 750); IgA 90 (0.9 to 715); IgM 162 (1.2 to 1,300); and albumin 34 (6.8 to 199). All platelet-associated Igs from thrombocytopenic ITP patients were found to be elevated twofold to 2,300-fold with one Ig class occasionally elevated 50-fold to 100-fold higher than the others. A similar group of ten thrombocytopenic ITP patients was found to have twofold to 26-fold elevations of platelet- associated albumin. This demonstration of increases in multiple classes of Igs as well as serum albumin associated with platelets from ITP patients suggests that some nonimmune process may be contributing to the phenomenon of increased platelet-associated proteins in ITP.
Reevaluation of cytochrome b and flavin adenine dinucleotide in neutrophils from patients with chronic granulomatous disease and description of a family with probable autosomal recessive inheritance of cytochrome b deficiency
Authors: Ohno Y , Buescher ES , Roberts R , Metcalf JA , Gallin JI
Published: October 2019

Chronic granulomatous disease (CGD) is a genetically heterogeneous syndrome characterized by a microbial killing defect of polymorphonuclear leukocytes (PMNs) due to lack of superoxide O2-. 2 generation. Recent studies indicate that the neutrophil O2-.-generating system consists of at least two components, flavoprotein--flavin adenine dinucleotide (FAD)--and cytochrome b. We evaluate the cytochrome b and FAD content in PMN from 30 CGD patients. The method for quantitating cytochrome b was modified by using PMN sonicates incubated with azide plus hydrogen peroxide. With this approach, several absorption peaks corresponding to myeloperoxidase and eosinophil peroxidase, which overlap with peaks of cytochrome b, were obliterated from reduced-minus-oxidized spectra, whereas the peaks of cytochrome b were not and could be readily quantitated. Cytochrome b was detected in PMNs from all 24 normal adults (47.4 +/- 2.9 pmol/7.5 X 10(6) cells), was absent in PMNs from 11 male CGD patients and one female CGD patient but was present in normal amounts in PMNs from nine male and nine female CGD patients. Stimulated nitroblue tetrazolium (NBT) tests performed on PMNs from mothers of CGD patients indicated that cytochrome b deficiency was associated with X-linked inheritance, except in one case in which probable autosomal recessive inheritance was demonstrated. The PMN NBT test of the mother of another male patient without cytochrome b deficiency suggested an X-linked form of inheritance. In related studies, the FAD content in PMN particulate fractions was reduced in 4 of 28 CGD patients studied. All four CGD patients with reduced FAD lacked cytochrome b. However, three patients with cytochrome b deficiency had normal FAD. Thus, the results indicate that PMN cytochrome b deficiency is observed in most X-linked and in some autosomal recessive CGD, that cytochrome b deficiency may be associated with FAD deficiency, and that cytochrome b and FAD are normal in most patients with non-X-linked CGD.
Cytomegalovirus infection after bone marrow transplantation: an association with acute graft-v-host disease
Authors: Miller W , Flynn P , McCullough J , Balfour HH Jr , Goldman A , Haake R , McGlave P , Ramsay N , Kersey J
Published: October 2019

Among 181 patients undergoing allogeneic bone marrow transplantation over a five-year period (1978 through 1982), cytomegalovirus (CMV) infection was a frequent and often lethal complication. Recipient pretransplant serology was the most important predictor of posttransplant CMV infection. CMV infection occurred in 26/137 seronegative recipients and in 28/44 seropositive recipients (P less than .001). Among patients who developed CMV infection, the time to infection was identical in seronegative and seropositive patients (median, 71 days post transplant). Bone marrow donor CMV serology did not significantly influence CMV infection rate. CMV infection was strongly associated with acute graft-v-host disease (AGVHD), occurring in 34/81 patients with AGVHD and 20/100 without GVHD (P less than .001). AGVHD preceded CMV infection by 33.7 days (mean) in patients developing both complications. Patients who developed CMV infections had also received more cellular blood products post transplant. These data suggest that CMV infection may occur through reactivation of latent virus (in seropositive recipients) or through exogenous exposure, possibly through transfused blood products, but that duration of immunoincompetence may be more critical than route of exposure in timing of clinically evident CMV infection. Prophylaxis tailored to the likely infectious source and more effective GVHD prevention both may be critical in preventing CMV infection after bone marrow transplantation.
Macrophage colony formation supported by purified CSF-1 and/or interleukin 3 in serum-free culture: evidence for hierarchical difference in macrophage colony-forming cells
Authors: Koike K , Stanley ER , Ihle JN , Ogawa M
Published: October 2019

Using a serum-free culture system, we examined murine macrophage colony formation from bone marrow cells cultured in the presence of purified CSF-1, interleukin 3 (IL 3) or a combination of the two factors. CSF-1 supported macrophage and neutrophil-macrophage colony formation, whereas IL-3 supported the formation of various types of single lineage and multilineage colonies. CSF-1 supported more macrophage colonies from bone marrow cells of normal mice than IL 3, whereas in cultures of bone marrow cells of 5-fluorouracil-treated mice, IL 3 supported more macrophage colonies. A combination of CSF-1 and IL 3 resulted in granulocyte-macrophage (GM) colony formation that was equal to or greater than the sum of GM colony formation supported by the factors individually. The combination of CSF-1 and IL 3 resulted in significant increases in the size of both macrophage and neutrophil-macrophage colonies. Similar increases in colony size were observed when CSF-1 was added to cultures five days after incubation of marrow cells with IL 3. These data support the concept that some of the macrophage colony- forming cells that respond to IL 3 are more primitive than those that are sensitive to CSF-1.
Separation of the acetylcholinesterase-deficient red cells in paroxysmal nocturnal hemoglobinuria
Authors: Chow FL , Hall SE , Rosse WF , Telen MJ
Published: October 2019

Blood of patients with paroxysmal nocturnal hemoglobinuria (PNH) most often contains two or more populations of erythrocytes--one population with normal sensitivity to lysis by complement (PNH I cells) and a second population of moderately abnormal cells (PNH II cells) or markedly abnormal cells (PNH III cells). PNH II and III cells exhibit moderately and markedly increased sensitivity to lysis by complement, respectively, as well as other membrane defects. We have devised a method for isolating pure, intact PNH II and III cells from mixed populations by use of monoclonal antibodies and cell affinity chromatography. Study of purified cell populations has led to the identification of a further subtype, PNH IIIb, of PNH erythrocytes. PNH IIIb erythrocytes are less sensitive to complement lysis than PNH IIIa cells but are lysed by fluid-phase activation of complement, unlike PNH II erythrocytes.
Synchronized cultures of P falciparum in abnormal red cells: the mechanism of the inhibition of growth in HbCC cells
Authors: Olson JA , Nagel RL
Published: October 2019

It has been previously demonstrated that HbCC cells fail to support growth of P falciparum in asynchronous Jensen-Trager cultures. To define the mechanism of inhibition we have studied synchronous cultures and found that while intraerythrocytic parasite development appeared normal, the liberation of merozoites and/or invasion was impaired. This effect was detected by a normal growth during the first growth cycle but dramatically reduced number of ring forms following the schizont stage. A specific test for the invasion of CC cells by P falciparum merozoites, nevertheless, was normal. The defect found in infected CC cells was not modified by changes in O2 tension (which altered the ligand saturation of Hb C) nor by the extracellular K+ concentration (excluding a K+ leak-dependent mechanism for the growth inhibition). The osmotic lysis of late-staged parasitized red cells revealed that 25% of infected AA cells were lysed when the extracellular medium was 95 mOsm. In contrast, infected CC cells required a decrease to 10 mOsm in the extracellular media to reach 25% lysis. We conclude that CC red cells are unsuitable hosts for the malarial parasite primarily because of their inability to lyse and release merozoites at the appropriate stage of intraerythrocytic development of P falciparum.
Pro-urokinase: a study of its stability in plasma and of a mechanism for its selective fibrinolytic effect
Authors: Pannell R , Gurewich V
Published: October 2019

Highly purified pro-urokinase (pro-UK) or single-chain urokinase-type plasminogen activator (scu-PA) was treated with diisopropylfluorophosphate (1 mmol/L) to eliminate traces of two-chain UK activity. This preparation was found to retain a low activity against a chromogenic substrate (S2444), equivalent to 0.1% to 0.5% of the activity of its plasmin-activated derivative. Evidence is presented that the intrinsic activity of pro-UK (scu-PA) was sufficient to activate plasminogen on a fibrin plate or in buffer and was far more reactive against Lys-plasminogen than against Glu-plasminogen. The relative resistance of Glu-plasminogen to activation was overcome by the addition of lysine (25 mmol/L) to the reaction mixture. By contrast, in plasma, pro-UK (scu-PA) was stable and nonreactive for greater than 72 hours when incubated (37 degrees C). Pro-UK (scu-PA) did not form sodium dodecyl sulfate-stable inhibitor complexes, whereas complexation occurred rapidly with UK. Only at high concentrations of pro-UK (scu-PA) (greater than or equal to 250 IU/mL) did plasminogen activation in plasma occur. The relative inertness of pro-UK (scu-PA) in plasma, in contrast to its low-grade enzymatic activity in buffer, was attributed to the effect of inhibitors. The addition of EDTA or the removal of divalent cations by dialysis was associated with a lower threshold for nonspecific plasminogen activation by pro-UK (scu-PA) in plasma. Replacement of Ca++ but not other cations restored baseline conditions. In the presence of a clot, fibrin-selective plasminogen activation and clot lysis were triggered. Lysis was accompanied by less than 10% conversion of pro-UK (scu-PA) to two-chain UK, suggesting that the intrinsic activity of pro-UK (scu-PA) itself may have been responsible for fibrinolysis, although a contribution by the small amount of UK generated could not be excluded. Similarly, pro-UK (scu- PA) supported clot lysis for several days in the same plasma before the effect dissipated as a result of degradation to UK. When Glu- plasminogen in plasma was replaced by Lys-plasminogen, or when lysine was added to normal plasma, nonselective plasminogen activation and fibrinogenolysis occurred. It was concluded that under the experimental conditions, the fibrin specificity of pro-UK (scu-PA) can be explained by its selective activation of fibrin-bound plasminogen and is due to the latter's Lys-plasminogen-like conformation.(ABSTRACT TRUNCATED AT 400 WORDS)
Characterization of murine peritoneal macrophage receptors for fibrin(ogen) degradation products
Authors: Rajagopalan S , Pizzo SV
Published: October 2019

The binding of human fibrinogen degradation fragments D1, E, X, and Y, as well as fibrin fragment D1 dimer, to mouse peritoneal macrophages was examined. A Scatchard plot of fragment D1 binding was biphasic, suggesting two classes of receptors. Fragments D1, D1 dimer, X, and Y in low concentrations bound to macrophages with high affinity (Kd = 23 to 73 X 10(-11) mol/L). Fragment E bound specifically but at a much lower level than the other fragments. Fragment D1 was able to compete for the binding of radiolabeled fragments X and Y but not radiolabeled fragment E. These studies indicate that fragments D and E are recognized by separate receptor systems but that all of the fibrinogen degradation products that contain the D domain are recognized by the same receptor system.
Inhibition of neutrophil cytolysin production by target cells
Authors: Dallegri F , Patrone F , Ballestrero A , Frumento G , Sacchetti C
Published: October 2019

Neutrophils, triggered by heat-aggregated human IgG (Agg.IgG), were found to lyse chicken red blood cells (CRBC) as determined by a 51Cr release method. The lysis was inhibited by azide, catalase, chloride- free medium and amino acids, suggesting the requirement for myeloperoxidase (MPO), hydrogen peroxide (H2O2), chloride ions (Cl-), and hypochlorous acid (HOC1), respectively. These results indicate that neutrophils lyse CRBC through an HOCl-(ie, MPO-H2O2-Cl-) dependent process. Although HOCl can react with neutrophil-derived nitrogenous (N- ) compounds to yield chloramines, the main and well-characterized chloramines did not play a direct role in the lysis of CRBC in our model system. Thus, it appears that lysis is due either to HOCl or to an unknown compound derived from and with characteristics similar to HOCl. When CRBC were replaced with HRBC targets, no lysis could be observed. Treatment of HRBC with carmustine, to inhibit the glutathione cycle, did not affect the cell resistance to lysis by neutrophils. Conversely, the inhibition of HRBC catalase activity with aminotriazole (AT) made the cells susceptible to neutrophil-mediated HOCl-dependent lysis: this suggests that HRBC escape lysis by neutrophils through an AT-inhibitable, ie catalase-dependent, process. Through an identical catalase-dependent process, HRBC were capable of efficiently preventing the H2O2 and HOCl recovery from Agg.IgG-triggered neutrophils, tested under experimental conditions similar to those used for cytolytic assays. Together, these data suggest that HRBC targets, endowed with high catalase activity, escape neutrophil-mediated lysis by consuming (by catalase) neutrophil-derived H2O2, so that HOCl cannot be produced in amounts sufficient to promote lysis. Parallel experiments, performed with AT-treated CRBC, showed that these cells, endowed with a relatively low catalase content, only partially limit neutrophil cytolytic efficiency by a process qualitatively similar to that observed with HRBC targets. The results provide evidence that target cells can restrain neutrophil cytolytic efficiency by interfering with the MPO-H2O2-Cl system through their catalase activity.
Separation of human megakaryocytes by state of differentiation on continuous gradients of Percoll: size and ploidy analysis of cells identified by monoclonal antibody to glycoprotein IIb/IIIa
Authors: Ishibashi T , Ruggeri ZM , Harker LA , Burstein SA
Published: October 2019

Human bone marrow was separated on continuous Percoll density gradients to analyze the distribution of cells of the megakaryocytic lineage. Megakaryocytes were identified by indirect immunofluorescence using a monoclonal antibody (LJP4) specific to the glycoprotein IIb/IIIa (GPIIb/IIIa) complex of platelets. Neither endothelial cells nor monocytes expressed the epitope identified by this antibody. Simultaneous measurement of size and ploidy were made on 2,359 GPIIb/IIIa-positive cells. The smallest cells were located in the most dense fractions where 81% of all 2N and 66% of 4N cells were found at densities greater than or equal to 1.050 g/mL. The largest cells were detected in the least dense fractions, with 70% of 16N, 78% of 32N, and 100% of 64N cells found at densities less than 1.050 g/mL. Ninety-four percent of all GPIIb/IIIa-positive cells less than 14 micron in diameter were found at densities greater than 1.050 g/mL. An exception to this inverse relationship was observed in the uppermost gradient fractions where an anomalous distribution of size and ploidy was found. Megakaryocytic viability was identified as being greater than 90% in all fractions. The data show that megakaryocytic differentiation as assessed by size and ploidy varies inversely with Percoll density. Separation of marrow on continuous Percoll gradients may be a useful method to separate megakaryocytes at different stages of differentiation.
Spontaneous regression of a monoclonal proliferation of large granular lymphocytes associated with reversal of anemia and neutropenia
Authors: Winton EF , Chan WC , Check I , Colenda KW , Bongiovanni KF , Waldmann TA
Published: October 2019

A 43-year-old male with a phenotypically homogeneous, expanded subset of T cells presented in 1981 with anemia and neutropenia. The surface antigen phenotype of 99% of the peripheral blood lymphocytes was T3+, T8+, T4-, and they were morphologically large granular lymphocytes (LGL). The same cells comprised 37% of the marrow nucleated cells. Eight months after he presented, the peripheral blood T8+, LGL diminished spontaneously, and the anemia and neutropenia completely resolved. The patient remains hematologically normal as of October 1984. To determine if the T8+, LGL represented a clonal expansion, DNA from peripheral blood lymphocytes collected and cryopreserved when the patient was neutropenic and anemic, and when he was hematologically normal, was analyzed for clonal T-cell antigen receptor gene rearrangements. Using Southern blot analysis, a clonal DNA rearrangement was demonstrated, and this clone diminished but was still demonstrable in peripheral blood lymphocytes collected in 1984. The above observations implicate the expanded T8+, LGL in the pathogenesis of the neutropenia and anemia, yet the exact mechanism remains to be elucidated.
A collaborative, double-blind randomized study of cetiedil citrate in sickle cell crisis
Authors: Benjamin LJ , Berkowitz LR , Orringer E , Mankad VN , Prasad AS , Lewkow LM , Chillar RK , Peterson CM
Published: October 2019

We have recently completed a double-blind, placebo-controlled, noncrossover study, the goal of which was to determine whether cetiedil citrate (cetiedil) could affect the course of vaso-occlusive crises in sickle cell disease. Patients, who presented to the emergency room at least 4 but no more than 24 hours after the onset of a painful vasoocclusive crisis severe enough to require hospitalization, were considered candidates for the study. Each patient received either placebo or cetiedil at one of the following three dosages: 0.2, 0.3, or 0.4 mg/kg body weight. The assigned drug dosage was given as a 30 minute intravenous infusion every 8 hours for 4 consecutive days. A total of 67 patients was enrolled in the study. Cetiedil, at its highest dosage (0.4 mg/kg body weight), was found to be significantly superior to placebo both in reducing the number of painful sites present on all 4 treatment days and in shortening the total time in crisis. No serious adverse reactions were observed during the course of the study. We conclude that cetiedil, given at a dosage of 0.4 mg/kg body weight, is therapeutically advantageous for sickle cell crisis.
Fibrinopeptide A and the phosphate content of fibrinogen in venous thromboembolism and disseminated intravascular coagulation
Authors: Leeksma OC , Meijer-Huizinga F , Stoepman-van Dalen EA , van Ginkel CJ , van Aken WG , van Mourik JA
Published: October 2019

Concentrations of plasma fibrinopeptide A (FPA) were measured by radioimmunoassay in 50 patients with venous thromboembolism or disseminated intravascular coagulation or both. A consistent discrepancy was observed in values obtained with two anti-FPA antisera. Analysis of extracts from plasma of these patients by high-performance liquid chromatography (HPLC) revealed the presence of a phosphorylated and an unphosphorylated form of the A peptide. Differences in concentrations of FPA measured with the two antisera could be accounted for by their different reactivity with phosphorylated FPA (FPA-P). The differences were abolished by treatment with alkaline phosphatase. A good correlation was observed between the FPA-P content of free A- peptide material and of fibrinogen in plasma as determined by HPLC (r = .88, P less than .001, n = 11). In patients with elevated FPA levels, the mean FPA-P content of fibrinogen was significantly higher (P less than .002, n = 13) than in patients with normal FPA levels (n = 8) and in healthy controls (n = 14). Phosphorus in fibrinogen did not correlate with fibrinogen degradation products or fibrinogen levels and became normal on adequate anticoagulation. Therefore, blood-clotting activation may lead to a high phosphate content of fibrinogen and of free FPA in plasma.
A monoclonal antibody preventing binding of tissue-type plasminogen activator to fibrin: useful to monitor fibrinogen breakdown during t-PA infusion
Authors: Holvoet P , Lijnen HR , Collen D
Published: October 2019

One (MA-1C8) of 36 monoclonal antibodies obtained by fusion of P3X63- Ag8–6.5.3 myeloma cells with spleen cells of mice immunized with purified human tissue-type plasminogen activator (t-PA) blocked the activity of t-PA on fibrin plates but not on chromogenic substrates. MA- 1C8 at a concentration of 200 micrograms/mL inhibited plasma clot lysis and binding of t-PA to the clot. MA-1C8 had no influence on the activation of plasminogen by t-PA, which obeys Michaelis-Menten kinetics with Km = 105 mumol/L and kcat = 0.05 s-1; however, it abolished the influence of CNBr-digested fibrinogen on Km. These findings confirm that the stimulatory effect of fibrin on the activation of plasminogen by t-PA is mediated by binding of t-PA to fibrin and provide additional support for the kinetic model. Addition of t-PA to pooled fresh human plasma to a concentration of 5 micrograms/mL resulted in extensive fibrinogen breakdown after incubation for one hour at 37 degrees C or during storage at -20 degrees C for one day. In both instances, fibrinogen degradation was completely prevented by addition of MA-1C8 to a concentration of 200 micrograms/mL of plasma. MA-1C8 also effectively prevented in vitro fibrinogen degradation and in vitro plasminogen activation in plasma samples obtained during infusion of recombinant t-PA in patients with thromboembolic disease. Thus, MA-1C8 is a useful tool for discriminating between in vivo and in vitro fibrinolysis during thrombolytic therapy with t-PA.
Use of a BamHI polymorphism in the factor IX gene for the determination of hemophilia B carrier status
Authors: Hay CW , Robertson KA , Yong SL , Thompson AR , Growe GH , MacGillivray RT
Published: October 2019

A BamHI polymorphism has been identified in the human factor IX gene. This polymorphism, which occurs in approximately 6% of X chromosomes, has been used to determine the carrier status of a female in a family with a history of hemophilia B. This family was uninformative for the previously reported TaqI and Xmnl polymorphisms in the factor IX gene.
Carrier detection in hemophilia A: a cooperative international study. I. The carrier phenotype
Authors: Graham JB , Rizza CR , Chediak J , Mannucci PM , Briet E , Ljung R , Kasper CK , Essien EM , Green PP
Published: October 2019

Eight laboratories in six countries cooperated to clarify several issues concerning the phenotypes of heterozygous carriers of hemophilia “A.” Plasma levels of factor VIII (F.VIII:C, formerly VIII:C) and von Willebrand factor (VWF:Ag, formerly VIIIR:Ag) of carriers and normal women were determined by various “in-house” methods; a single lyophilized plasma standard was used for all assays. Analysis of the collated data from 336 carriers (296 obligatory carriers and 40 sporadic carriers) and 137 normal women showed that there was no difference in the F.VIII:C levels of “paternal” carriers (women who had obtained the abnormal gene from their fathers) and “maternal” carriers. Neither was there a difference in the VWF:Ag levels of normal women and either type of carrier. Age was found to have a significant effect on both F.VIII:C and VWF:Ag, values being higher at very young and very old ages, the minima occurring in the 25- to 30-year range. ABO blood type had a striking effect. Women of types A, B, and AB (designated non- O in the study), both normals and carriers, had significantly higher levels of both factors than did women of type O. Analysis by laboratories showed that differences in mean levels of both factors between laboratories were highly significant. It was concluded that age, ABO blood type, and laboratory variation should be taken into account in carrier detection.
Selective inhibition of the growth of human erythroid bursts by monoclonal antibodies against transferrin or the transferrin receptor
Authors: Shannon KM , Larrick JW , Fulcher SA , Burck KB , Pacely J , Davis JC , Ring DB
Published: October 2019

The relative requirements of colonies derived from erythroid (BFU-E) and myeloid (CFU-c) progenitors for transferrin were examined using monoclonal antibodies directed against the transferrin molecule (TF-6) or its cell surface receptor (TFR-A12, TFR1–2B). Growth of erythroid bursts was profoundly reduced at concentrations of all three antibodies that had no effect on CFU-c-derived colonies. When TFR1–2B was layered over cultures established one to seven days previously, further burst development was inhibited, and degeneration of early erythroid colonies was observed. Addition of erythropoietin augmented transferrin receptor expression on cells harvested after 1 to 2 weeks in culture and analyzed by flow cytometry. Recombinant human erythropoietin gave results comparable to those obtained in experiments using human urinary erythropoietin. Analysis of erythroblasts plucked directly from culture plates confirmed the presence of transferrin receptors on BFU-E-derived colonies. Thymidine incorporation was maximal early in the second week of culture and coincided with high transferrin receptor expression. These data demonstrate that transferrin must be available into the second week of culture to support the growth and differentiation of BFU- E-derived erythroid bursts, that the generation of erythroid colonies from BFU-E is more dependent on transferrin than myeloid colony formation from CFU-c, and that erythropoietin modulates the expression of transferrin receptors on growing bursts.
Platelets modulate the proteolysis of factor VIII:C protein by plasmin
Authors: Rick ME , Krizek DM
Published: October 2019

Factor VIII coagulant protein (VIII:C) functions as a critical cofactor with factor IXa, calcium ions, and phospholipid during the activation of factor X. In the course of this reaction, the activity of VIII:C is first increased and then is destroyed by one or more serine proteases that are part of the coagulation sequence. In this study, we have investigated the influence of platelets on the inactivation of VIII:C by plasmin. Platelets were separated from plasma proteins in the presence of granule release inhibitors and were incubated with plasmin and isolated VIII:C or the complex of purified VIII:C/von Willebrand factor (vWF); VIII:C activity and antigen levels were assessed over time. In the presence of platelets, the isolated VIII:C showed an initial increase in VIII:C activity that was not present when platelets were absent, and the VIII:C/vWF showed an increase in VIII:C activity over that seen when platelets were absent. In addition, platelets stabilized VIII:C activity over a one-hour time course when compared with buffer. The VIII:C antigen did not increase and decreased slowly whether platelets were present or absent. Preincubating the platelets with ristocetin, collagen, or plasmin did not alter the results, and experiments using platelets from a patient with severe von Willebrand's disease also showed a pattern similar to that seen with normal platelets. Experiments using fixed platelets or phospholipid vesicles showed that they did not support the activation reaction or delay the inactivation reaction. These studies demonstrate that platelets modulate the activation and inactivation of VIII:C by plasmin, apparently by a mechanism that is independent of the platelet release reaction.
Myosin in adult and neonatal human erythrocyte membranes
Authors: Matovcik LM , Groschel-Stewart U , Schrier SL
Published: October 2019

The heavy chain of myosin can be detected in human red cell membranes by immunoblot analysis with antiplatelet myosin antibodies. Neonatal red cell membranes have more immunoreactive myosin than adult membranes. Membranes from young adult red cells contain more immunoreactive myosin than membranes from old adult red cells. In contrast, young and old neonatal red cells have equivalent mounts of myosin. Erythrocyte myosin is present in a membrane fraction enriched in integral membrane proteins but is not found in cytoskeletal preparations.
Adverse effect of severe marrow fibrosis on hematologic recovery after chemoradiotherapy and allogeneic bone marrow transplantation
Authors: Rajantie J , Sale GE , Deeg HJ , Amos D , Appelbaum F , Storb R , Clift RA , Buckner CD
Published: October 2019

We reviewed the records and marrow biopsy specimens of 75 patients with leukemic, myelodysplastic, or myeloproliferative disorders to determine whether the presence of marrow fibrosis affected engraftment after allogeneic marrow transplantation. While 28 control patients without fibrosis achieved prompt engraftment, two of 32 patients (6%) with mild and five of 15 patients (33%) with severe fibrosis failed. The rate of myeloid recovery was significantly slower and the dependence on platelet and red blood cell transfusions was significantly longer in patients with severe fibrosis than in patients with no fibrosis. A finding of severe marrow fibrosis should therefore be taken into account when evaluating the risks and benefits of marrow transplantation.
Refined chromosome analysis as an independent prognostic indicator in de novo myelodysplastic syndromes
Authors: Yunis JJ , Rydell RE , Oken MM , Arnesen MA , Mayer MG , Lobell M
Published: October 2019

In a study of 56 consecutive adult patients with de novo myelodysplastic syndromes (MDS), all cases were successfully analyzed with two refined chromosome banding techniques. Most patients (44 of 56, 79%) were found to have a chromosome defect. The majority of these patients had a recurrent loss of chromosomal material rather than a reciprocal translocation or inversion, as commonly found in acute leukemia. The three largest chromosomal categories found were associated with a wide range of survival. Twelve patients (21%) had normal chromosomes, a stable clinical course, and long survival (median follow-up time of 49 months, with all patients alive). Nine patients had in common a single chromosome defect resulting in either monosomy 7 or deletion 7q. They had a median survival of 12 months, and four died of acute nonlymphocytic leukemia (ANLL). Of 12 patients with complex defects, 11 had a complete or partial loss of a chromosome 5 and a complete or partial loss of the long arm of a chromosome 7 or 20. They had a poor median survival of four months, and six patients died of ANLL. Although the French-American-British (FAB) classification was also found to have some prognostic value, FAB subgroups were chromosomally heterogeneous and showed less dramatic differences in median survival than the larger chromosomal subgroups. We have shown, for the first time, that a refined chromosomal analysis is an independent prognostic indicator in de novo MDS and may be helpful in establishing therapeutic approaches in this difficult group of heterogeneous disorders.
Expression of p53 in human leukemia and lymphoma
Authors: Prokocimer M , Shaklai M , Bassat HB , Wolf D , Goldfinger N , Rotter V
Published: October 2019

Analysis of fresh human tumors have indicated that patients with B type lymphoproliferative diseases and the majority of patients with acute lymphoblastic leukemia (ALL) express elevated levels of p53 production. It is suggested that in these human malignancies, p53 may provide a novel tool for monitoring cancer activity. Conversely, p53 is not expressed in acute myeloid leukemias, myeloproliferative diseases, or myeloid leukemic cell lines. Analysis of the p53 gene structure indicated the existence of similar patterns of p53 restriction fragments in producer and nonproducer cells, which suggests that the p53 gene is not altered in the latter. However, in one case of acute promyelocytic leukemia (APL), we have observed a rearrangement in the p53 gene. Karyotype analysis has indicated that these APL cells do not contain the typical 15;17 translocation. In other APL patients who exhibit a 15;17 translocation, we found no genomic changes of the p53, suggesting that the p53 gene, which was recently mapped to the short arm of chromosome 17 in the human, is not structurally related to the typical chromosomal break point found in the long arm of chromosome 17 of APL patients.
Effects of 2,3-diphosphoglycerate on the mechanical properties of erythrocyte membrane
Authors: Waugh RE
Published: October 2019

Investigation by Schindler et al and Sheetz and Casaly have indicated that high (approximately 10 mmol/L) concentrations of 2,3- diphosphoglycerate (2,3-DPG) have a destabilizing effect on erythrocyte membrane and the membrane skeleton. We have investigated changes in the membrane mechanical properties that occur at elevated 2,3-DPG levels in both intact cells and ghosts. The membrane shear modulus, viscoelastic recovery time constant, critical force, “plastic” viscosity, and material relaxation time constant were measured by standard micropipette and flow channel techniques. Intact cells showed no change in properties at physiologic ionic strength and 2,3-DPG concentrations of about 20 mmol/L, except for an increase in membrane viscosity resulting from an increased cellular hemoglobin concentration that occurs when the 2,3-DPG concentration is elevated. At ionic strengths 20% below physiologic and 2,3-DPG concentrations of approximately 20 mmol/L, decreases in membrane shear modulus and membrane viscosity were observed. In ghosts, no changes in these properties were observed at a 2,3-DPG concentration of 10 mmol/L and ionic strengths as low as 25% below physiologic, but a decrease in the force required to form tethers (critical force) was observed at physiologic ionic strength. The decrease in membrane shear modulus and viscosity of intact cells and the reduced critical force in ghosts are consistent with the results of other investigators. However, the difference in the effects of 2,3-DPG on ghosts and intact cells indicates that the effects of 2,3-DPG depend strongly on the conditions of the experiment. It appears unlikely that 2,3-DPG affects erythrocyte membrane material properties under physiologic conditions.
Alpha-interferon activates the natural killer system in patients with hairy cell leukemia
Authors: Semenzato G , Pizzolo G , Agostini C , Ambrosetti A , Zambello R , Trentin L , Luca M , Masciarelli M , Chilosi M , Vinante F
Published: October 2019

To elucidate the mechanisms of alpha-interferon's (alpha-INF) therapeutic effect on clinical and laboratory findings in hairy cell leukemia, we sequentially monitored different immunologic parameters in three patients treated with recombinant alpha-INF. The most evident effect of this treatment on the immune system was the recovery of natural killer (NK) cell in vitro activity of peripheral blood lymphocytes, which was severely impaired before therapy. In particular, NK function began to improve after 3 months, and a complete recovery was obtained after 6 months in all cases. This increase parallels the improvement in clinical and laboratory findings.
Carbonic anhydrase: a marker for the erythroid phenotype in acute nonlymphocytic leukemia
Authors: Walloch J , Frankel S , Hrisinko MA , Weil SC
Published: October 2019

Protein markers are often used to corroborate the morphological subtyping of hematopoietic malignancy. Most commonly, surface markers are used for the phenotyping of hematopoietic cells; however, internal proteins have also been used as markers. Glycophorin, hemoglobin A, hemoglobin F, and transferrin have all been used as markers for the erythroid phenotype. We have recently shown that carbonic anhydrase is constitutively and aberrantly expressed in two erythroleukemic cell lines. We here show that it is also present in high levels in primary erythroleukemic blasts and that it is a useful marker for the M6 phenotype when classifying acute nonlymphocytic leukemia.
Spurious detection of terminal deoxynucleotidyl transferase in phytohemagglutinin-stimulated lymphocytes
Authors: Fletcher JA , Bell R , Koekebakker M , Dowers AA , McCaffrey RP , Barr RD
Published: October 2019

Expression of terminal transferase (TdT) is believed to be restricted to primitive lymphoid cells; recently, however, indirect immunofluorescent (IF) assays have been used to demonstrate the apparent presence of TdT in phytohemagglutinin (PHA)-stimulated peripheral blood lymphocytes and in various nonlymphoid malignancies. Using an IF assay, we found that a heteroantiserum to TdT reacted with cultured and PHA-stimulated human peripheral blood mononuclear cells, but we were unable to confirm the presence of TdT in these cells using immunoblotting and biochemical assays. We conclude that the IF results are spurious and most likely represent recognition by the heteroantiserum of inducible protein(s) other than TdT.