bluuebunny/crossref_metadata_2025 huggingface dataset:
Thrombocytopenia associated with pregnancy in a patient with type IIB von Willebrand's disease
Authors: Rick ME , Williams SB , Sacher RA , McKeown LP
Published: October 2019

Thrombocytopenia may accompany variant (type IIB) von Willebrand's disease (vWD) and is thought to result from binding of the abnormal von Willebrand factor (vWF) to the patient's platelets with subsequent platelet aggregate formation and clearance. We have studied a patient with type IIB vWD who became thrombocytopenic during two pregnancies. During the third trimester of pregnancy, her platelet counts dropped to 20,000 to 30,000/microL, and an increase in the intermediate-sized vWF multimers was seen on agarose gel electrophoresis. During this time her platelet-rich plasma showed spontaneous platelet aggregation, and her plasma caused spontaneous aggregation of normal washed platelets. Antibody to platelet glycoprotein Ib completely blocked the spontaneous platelet aggregation, while antibody to platelet glycoprotein IIb/IIIa did not block the response at the concentrations used. Inhibitors of platelet function that elevate platelet cyclic AMP also blocked the response, but aspirin had no effect on the spontaneous platelet aggregation. The patient illustrates that the platelet counts in one individual can vary greatly in type IIB vWD and that the thrombocytopenia that occurs can appear under physiologic conditions that stimulate the endogenous production of the patient's abnormal vWF. The mechanisms leading to spontaneous platelet aggregation and thrombocytopenia appear to be similar to those described for other patients with type IIB vWD.
Initial experience in treating human lymphoma with a chimeric univalent derivative of monoclonal anti-idiotype antibody
Authors: Hamblin TJ , Cattan AR , Glennie MJ , MacKenzie MR , Stevenson FK , Watts HF , Stevenson GT
Published: October 2019

Murine monoclonal anti-idiotype antibody was raised against the surface IgM on the neoplastic cells of a patient with widespread follicular lymphoma. For therapy, a chimeric antibody derivative, FabIgG, was constructed by thioether-linking Fab'gamma, from the monoclonal antibody, to human normal IgG. FabIgG is univalent and thereby avoids rapid antigenic modulation. Its human IgG component is intended to optimize recruitment of effectors and metabolic survival while minimizing immunogenicity. Four intravenous (IV) infusions of 380 to 580 mg of anti-idiotype FabIgG were given over a period of 11 weeks. There was no significant toxicity. On each occasion, the antibody disappeared from the plasma with a half-life (t1/2) of less than 24 hours. The brief survival was evidently due to uptake by tumor, as infused control FabIgG, containing Fab'gamma from an irrelevant antibody, yielded a plasma t1/2 of greater than 10 days. With each therapeutic infusion, there was a fall in the number of circulating neoplastic cells over a 24-hour period. The numbers were largely replenished over the next week, but a net fall became discernible over the entire period of treatment. Four days after each infusion, nodal masses were swollen and tender, subsiding over approximately 8 days. At the end of the treatments, the blood lymphocyte count and nodal and splenic swellings continued to subside, so that by 6 weeks a partial remission with removal of greater than 50% of tumor was judged to have occurred. We did not detect any qualitative change in surface idiotype nor any antibody response to the infused Ig.
Variability of the erythropoietic response in autoimmune hemolytic anemia: analysis of 109 cases
Authors: Liesveld JL , Rowe JM , Lichtman MA
Published: October 2019

One hundred-nine cases of autoimmune hemolysis were reviewed to determine the frequency of reticulocytopenia, the state of the erythroid marrow in reticulocytopenic cases, and the course of reticulocyte production indices with time and glucocorticoid treatment. The mean hematocrit at presentation was 24 mL/dL, but 30% of cases had an initial hematocrit less than 20 mL/dL. Median reticulocyte percentage at diagnosis was 9%, and median reticulocyte production index was 2.8 times basal. Twenty percent of cases had an initial reticulocyte count less than 4%, and 37% had an initial reticulocyte production index less than 2.0 times basal. These reticulocytopenic patients were nearly evenly distributed between warm and cold antibody- mediated cases and between primary and secondary cases. Fifty-four percent of reticulocytopenic cases had a bone marrow examination during hospitalization. Three-fourths of these marrows showed erythroid hyperplasia, and erythroid hypoplasia was seen in only one case. Eighty- eight cases had serial reticulocyte measurements, and in only 15% of patients did the reticulocyte production index remain less than 2.0 times basal. Thus, in most cases, the initially low reticulocyte production index may represent a lag in marrow responsiveness to hemolytic stress. In cases with persistent reticulocytopenia, ineffective erythropoiesis is suggested by the frequency of marrow erythroid hyperplasia. In the cases that were initially reticulocytopenic and demonstrated an increase in reticulocyte production index, the magnitude of this increase was significantly greater in glucocorticoid-treated patients than in those not so treated, indicating that a glucocorticoid sensitive component exists in the marrow erythropoietic response to hemolysis. Awareness of the frequency of an initial reticulocytopenia in cases of autoimmune hemolysis may be important in initial diagnosis and treatment.
Evaluation of circulating malignant cells provides prognostic information in cutaneous T cell lymphoma
Authors: Schechter GP , Sausville EA , Fischmann AB , Soehnlen F , Eddy J , Matthews M , Gazdar A , Guccion J , Munson D , Makuch R
Published: October 2019

Peripheral blood lymphocyte morphology was evaluated prospectively by light microscopy of blood smears and E rosette preparations in 160 patients with cutaneous T cell lymphoma (CTCL). Blood involvement was related to the type of cutaneous T-stage, being present in 90% of patients with erythroderma (T4), 27% of those with cutaneous tumors (T3), 9% of those with generalized (T2), and 0% of those with limited skin plaques (T1). Untreated patients with blood involvement (38 of 105) had a higher frequency of CTCL in lymph nodes and viscera and survival inferior to that of patients with normal or nondiagnostic lymphocyte morphology (P less than .001). Multivariate analysis showed skin stage and age to be the most important pretreatment risk factors for survival. Although blood involvement was not an independent risk factor for the entire group, it appeared to have some adverse influence in the T2/T3 subsets (P = .051). Both lymphocytosis and size distribution of the circulating CTCL cells at initial diagnosis influenced survival. Patients with “mixed cell” cytology (greater than 20% large [greater than 11 microns] CTCL cells), had a worse survival than those with predominantly small circulating CTCL cells (P = .009). The former were more likely to have aggressive features, including lymph node effacement by tumor (P less than .001) and visceral disease (P = .074), than were “small cell” patients. Our data indicate that detailed review of the blood lymphocyte morphology in patients with diagnosed or suspected CTCL is helpful in predicting extent of disease and prognosis.
Chromosome abnormalities in AIDS-associated lymphadenopathy
Authors: Alonso ML , Richardson ME , Metroka CE , Mouradian JA , Koduru PR , Filippa DA , Chaganti RS
Published: October 2019

Cytogenetic studies were performed on direct and 24-hour culture preparations of eight lymph node biopsies from seven patients with acquired immunodeficiency syndrome (AIDS) or AIDS-related complex (ARC)- associated lymphadenopathy in whom histological evidence of lymphoma was not detected. Three of these seven had chromosomal abnormalities, including chromosome instability in one and clonal chromosomal abnormalities in two; one of the latter was a t(8;14)(q24;q32). The remaining five showed normal karyotypes. Epstein-Barr virus (EBV) titers were elevated in all three patients that exhibited chromosome abnormalities, two of whom later developed malignant lymphoma. A control group of five patients with reactive lymphadenopathy not associated with AIDS failed to reveal chromosomal aberrations, but elevated EBV titers were present in two. These data are consistent with current views on the role of EBV and chromosome change in the development of lymphoma in immunodeficient states and suggest that karyotypically abnormal AIDS-related lymphadenopathy represents a prelymphomatous proliferation.
Augmentation of in vitro human marrow erythropoiesis under physiological oxygen tensions is mediated by monocytes and T lymphocytes
Authors: Pennathur-Das R , Levitt L
Published: October 2019

Tissue and marrow (BM) in vivo O2 tensions have been estimated at 23 to 40 mm Hg (3% to 5% O2). We have investigated cellular regulation of burst-forming units-erythroid (BFU-E) under 5% O2. BFU-E from BM mononuclear cells (MNC) were cultured in methylcellulose medium and erythropoietin (Ep) +/- monocyte-conditioned medium (MoCM, a source of burst-promoting activity, BPA) in the presence or absence of autologous T cells (T) and/or monocytes (M phi) under either 5% or 21% O2 after depletion of T (MNC-T), M phi (nonadherent buoyant cells, NAB) or both T and M phi depletion (NAB-T). MNC BFU-E growth under 5% O2 was augmented over 0.1 to 1.5 U/mL of Ep. BFU-E augmentation under 5% O2 was abolished by depletion of BM M phi, T, or both from MNC. The addition of MoCM affected neither a BFU-E increase under 5% O2 nor the abrogation of that increase upon T or M phi depletion. The addition of 5% to 20% M phi or 10% to 20% T to NAB-T failed to restore the BFU-E increase under 5% O2. However, BFU-E augmentation under 5% O2 was reestablished when 10% autologous M phi, 10% T, or 10% T plus 10% M phi were added back to marrow NAB, MNC-T, or NAB-T. BM BFU-E was not augmented in the presence of varying concentrations of catalase, superoxide dismutase, or reduced glutathione at 21% O2; moreover, BFU-E augmentation was maintained at 5% O2 relative to 21% O2 in the presence of each of these antioxidants. CM prepared under 5% or 21% O2 from BM M phi, T, or M phi plus T were assessed for BPA against BM NAB-T using a sensitive BPA assay incorporating delayed Ep addition to cultures. Only CM from mixtures of M phi and T cells at 5% O2 demonstrated potent BPA; little or no BPA was detected with T or M phi CM at 5% O2 and at 21% O2 or T and M phi CM at 21% O2. The sensitivity of NAB-T BFU-E to exogenous BPA was virtually identical at 21% and 5% O2. These results indicate that human BM BFU-E are augmented under 5% O2 and that T cells and M phi together mediate that augmentation by collaborating to produce BPA-like activity in response to physiological O2 tensions.
F-actin content of neonate and adult neutrophils
Authors: Hilmo A , Howard TH
Published: October 2019

We utilized flow cytometric analysis of NBDphallacidin-stained cells to measure F-actin content, expressed as fluorescent channel numbers, and we compared the microfilamentous cytoskeletal organization in neutrophils from healthy neonates (36 to 38 weeks gestational age) and adults. Basal F-actin content in neonate cord blood neutrophils is higher than that of adults. The elevation is intrinsic to the cell and not related to parturition because basal F-actin content of neonatal cells obtained by venipuncture (days 1 to 8 of age) is also elevated (38 +/- 10, N = 15) when compared to adults (21 +/- 7.0, N = 27). The rate of N-formyl-methionyl-leucyl-phenylalanine (fMLP)-induced actin polymerization is similar in adult and neonatal neutrophils and is maximal by 30 to 45 seconds at 25 degrees C. Adult, neonatal cord, and neonatal venipuncture neutrophils increase F-actin content to a similar extent following 0.5 mumol/L fMLP activation (52 +/- 18, N = 27; 58.7 +/- 18, N = 18; 51.5 +/- 7.0, N = 15, respectively). However, the relative increase in F-actin content following fMLP activation is much greater in adult (2.37-fold) than neonatal neutrophils (1.28-fold). This difference is due to the elevated basal F-actin content of neonatal cells. Comparison of distribution of F-actin content among basal, neonatal neutrophils reveals two subpopulations of neutrophils with respect to F-actin content--approximately 25% with F-actin content similar to that of adult neutrophils and 75% with F-actin content greater than that of adult cells. Following fMLP activation, the subpopulations disappear. The results suggest that abnormalities in microfilamentous cytoskeletal organization of neonatal cells may, in part, be responsible for decreased chemotactic response of neonatal neutrophils.
Differences in coagulant and fibrinolytic activities of cultured human endothelial cells derived from omental tissue microvessels and umbilical veins
Authors: Speiser W , Anders E , Preissner KT , Wagner O , Muller-Berghaus G
Published: October 2019

Large vessel and microvascular endothelial cells were compared in their capacity to synthesize and secrete coagulant and fibrinolytic factors. Human omental tissue microvascular endothelial cells (HOTMEC) and human umbilical vein endothelial cells (HUVEC) were isolated, grown to confluency under identical conditions, and studied in primary cultures. After an incubation period of 12 hours in serum-free medium, the conditioned medium of confluent HOTMEC contained 100-fold higher levels of tissue plasminogen activator (tPA) antigen than that of HUVEC. The conditioned media as well as the lysates of both cell types did not contain any free tPA activity, but the free plasminogen activator inhibitor capacity was found intracellularly as well as extracellularly. Although von Willebrand factor was detected in both cell types by immunofluorescence, measurable amounts were only found in HUVEC using an enzyme-linked immunosorbent assay. The kinetics of protein C activation by thrombin on the surface of once-passaged cells were identical for HOTMEC and HUVEC. The present study indicates that cultivated HOTMEC produce larger quantities of tPA than HUVEC do, possess smaller amounts of von Willebrand factor than HUVEC do, and express thrombomodulin for protein C activation as effectively as HUVEC.
Immunochemical characterization of fibrinogen, fibrin I, and fibrin II in human thrombi and atherosclerotic lesions
Authors: Bini A , Fenoglio J Jr , Sobel J , Owen J , Fejgl M , Kaplan KL
Published: October 2019

Arterial thrombi and atherosclerotic lesions were analyzed immunochemically and examined histologically. The extent of in vivo proteolytic cleavage of the amino-terminal end of fibrinogen by thrombin and plasmin was determined and quantitated by specific radioimmunoassays. The samples were treated with cyanogen bromide (CNBr), and the total amount of fibrinogen and fibrin-derived protein was determined as NDSK, the NH2-terminal disulfide knot of fibrinogen. Thrombin-releasable fibrinopeptides A and B were used to quantitate fibrinogen and fibrin I. Previous plasmin cleavage of the B beta chain was inferred from the amount of B beta 1–42 and B beta 15–42 in undigested NDSK. The results obtained in both acute and organized thrombi indicate that approximately 60% of the total protein (as determined by amino acid analysis) was fibrinogen-derived and that 70% to 80% of the fibrinogen-derived material was fibrin II. These findings support the hypothesis that fibrin II as distinct from fibrin I is the predominant component in a thrombus. In samples from normal and atherosclerotic aortas, fibrinogen-derived protein comprised less than 10% of the total protein. Samples from grossly normal aortas contained only fibrinogen and fibrin I. Fibrinogen concentration decreased and fibrin II concentration increased with increasing severity of the lesions, suggesting that increased fibrin II formation is associated with progression of atheromas.
Stimulation by cAMP of erythropoietin secretion by an established human renal carcinoma cell line
Authors: Sherwood JB , Burns ER , Shouval D
Published: October 2019

We used our recently reported stable, transformed human renal carcinoma cell line as a model system to study the role of 3′,5′-adenosine monophosphate (cAMP) in erythropoietin secretion. The erythropoietin produced by these cells is both biologically active and immunologically cross-reactive with purified native human hormone in our radioimmunoassay. Erythropoietin release by these renal carcinoma cells appears to be stimulated by cAMP as well as by the phosphodiesterase inhibitor 3-isobutyl-1-methyl-xanthine (MIX). The response to cAMP involves a rapid and enhanced release of hormone, which occurred within 30 minutes of exposure of the cells to the effector and continued for at least 4 hours. Intracellular erythropoietin was higher in the control cultures than in the cells treated with cAMP, suggesting that cAMP stimulates the release of a storage pool of hormone. The ability of cAMP and MIX to elicit the release of erythropoietin suggests that a cAMP-mediated mechanism is involved in the release of this hormone.
Sardinian delta beta zero-thalassemia: a further example of a C to T substitution at position -196 of the A gamma globin gene promoter
Authors: Ottolenghi S , Giglioni B , Pulazzini A , Comi P , Camaschella C , Serra A , Guerrasio A , Saglio G
Published: October 2019

Selective overexpression (50- to 100-fold) in adult erythroid cells of either G gamma or A gamma fetal globin gene is observed in hereditary conditions known as delta beta zero-thalassemia and hereditary persistence of fetal hemoglobin (HPFH). Recently, a C-T change at position -196 of an overexpressed A gamma globin gene from an Italian HPFH was hypothesized, on the basis of indirect evidence, to represent the cause of the functional defect. We now show that the same mutation is present in a different overexpressed A gamma-globin gene from a Sardinian patient with a different syndrome (delta beta zero- thalassemia). The Sardinian A gamma globin gene differs from both the HPFH and the normal A gamma globin gene at nucleotide 1,560 in the noncoding portion of the third exon, where an A is deleted. In addition, the mutant -196 A gamma-globin gene is linked to a normal beta globin gene in HPFH, and to a beta-thalassemic gene (beta 39CAG-TAG) in delta beta zero-thalassemia. These data strengthen the suggestion that -196 mutation is causally linked to the abnormal phenotype and raise the question of whether the same or multiple mutational events are responsible for the appearance of the -196 mutation in different syndromes.
Studies on the ultrastructure of fibrin lacking fibrinopeptide B (beta- fibrin)
Authors: Mosesson MW , DiOrio JP , Muller MF , Shainoff JR , Siebenlist KR , Amrani DL , Homandberg GA , Soria J , Soria C , Samama M
Published: October 2019

Release of fibrinopeptide B from fibrinogen by copperhead venom procoagulant enzyme results in a form of fibrin (beta-fibrin) with weaker self-aggregation characteristics than the normal product (alpha beta-fibrin) produced by release of fibrinopeptides A (FPA) and B (FPB) by thrombin. We investigated the ultrastructure of these two types of fibrin as well as that of beta-fibrin prepared from fibrinogen Metz (A alpha 16 Arg----Cys), a homozygous dysfibrinogenemic mutant that does not release FPA. At 14 degrees C and physiologic solvent conditions (0.15 mol/L of NaCl, 0.015 mol/L of Tris buffer pH 7.4), the turbidity (350 nm) of rapidly polymerizing alpha beta-fibrin (thrombin 1 to 2 U/mL) plateaued in less than 6 min and formed a “coarse” matrix consisting of anastomosing fiber bundles (mean diameter 92 nm). More slowly polymerizing alpha beta-fibrin (thrombin 0.01 and 0.001 U/mL) surpassed this turbidity after greater than or equal to 60 minutes and concomitantly developed a network of thicker fiber bundles (mean diameters 118 and 186 nm, respectively). Such matrices also contained networks of highly branched, twisting, “fine” fibrils (fiber diameters 7 to 30 nm) that are usually characteristic of matrices formed at high ionic strength and pH. Slowly polymerizing beta-fibrin, like slowly polymerizing alpha beta-fibrin, displayed considerable quantities of fine matrix in addition to an underlying thick cable network (mean fiber diameter 135 nm), whereas rapidly polymerizing beta-fibrin monomer was comprised almost exclusively of wide, poorly anastomosed, striated cables (mean diameter 212 nm). Metz beta-fibrin clots were more fragile than those of normal beta-fibrin and were comprised almost entirely of a fine network. Metz fibrin could be induced, however, to form thick fiber bundles (mean diameter 76 nm) in the presence of albumin at a concentration (500 mumol/L) in the physiologic range and resembled a Metz plasma fibrin clot in that regard. The diminished capacity of Metz beta-fibrin to form thick fiber bundles may be due to impaired use or occupancy of a polymerization site exposed by FPB release. Our results indicate that twisting fibrils are an inherent structural feature of all forms of assembling fibrin, and suggest that mature beta-fibrin or alpha beta-fibrin clots develop from networks of thin fibrils that have the ability to coalesce to form thicker fiber bundles.
Mediastinal lymphoma of clear cell type is a tumor corresponding to terminal steps of B cell differentiation
Authors: Moller P , Moldenhauer G , Momburg F , Lammler B , Eberlein-Gonska M , Kiesel S , Dorken B
Published: October 2019

This article reports eight primary mediastinal tumors occurring in young adults (19 to 43 years, mean 29.4 years), predominantly female (six of eight) adults. Most patients responded badly to aggressive therapy. Progression is presently noted in one patient; five patients died 10, 11, 13, 18, and 22 months after diagnosis. No patient developed leukemia. The tumors were highly proliferative, had a diffuse growth pattern, and comprised clear cells of variable size. They could not be classified histologically, but could, however, be immunohistologically characterized as B cell lymphomas. In all cases, the immunophenotype was LC+, cALLa-, CD19+, CD20+, CD21-, Ig (surface/cytoplasm)-, and PC-1+. In addition, the neoplastic cells exhibited variable defects in the expression of HLA-A,B,C and HLA-DR and inconstant expression of other B cell-restricted/associated antigens. This combination of immunophenotypical and clinical features suggests that the mediastinal clear cell lymphoma (MCCL) is a previously undescribed type of B cell lymphoma corresponding to the terminal steps of B cell differentiation.
Contrasting patterns of DNA strand breakage and ADP-ribosylation- dependent DNA ligation during granulocyte and monocyte differentiation
Authors: Khan Z , Francis GE
Published: October 2019

Previous studies have shown that structural changes in DNA, including the ligation of pre-existing DNA breaks and the opening and closure of new breaks, occur shortly after exposure of granulomonocytic precursors (CFU-GM) to granulocyte-macrophage colony stimulating activity (GM- CSA). Monocytic differentiation of CFU-GM is selectively inhibited by compounds known to inhibit the nuclear enzyme ADP-ribosyl transferase (ADPRT). Since this enzyme, which transfers ADP-ribose units to chromatin proteins, is known to activate DNA ligase, we attempted to determine whether ligation of one or both types of DNA break is required for monocytic differentiation. Breaks in DNA were examined using the nucleoid sedimentation technique in which DNA breaks cause loss of DNA supercoiling in nucleoids and concomitant changes in their sedimentation through neutral sucrose gradients. We here report that two distinct patterns of DNA strand breakage and ligation are associated with differentiation to the granulocyte and monocyte lineages. Monocytic inducers (phorbolester and vitamin D3) predominantly produce closure of pre-existing strand breaks, whereas granulocytic inducers (granulocyte colony stimulating activity, G-CSA; retinoic acid) cause opening and closure of new breaks. Only ligation of the pre-existing breaks is highly sensitive to inhibition by 3- methoxybenzamide (a potent ADPRT inhibitor), and only monocytic differentiation is impaired by addition of this compound. These findings suggest that DNA structural changes may be directly involved in granulocyte-macrophage switching.
Vitamin B12-responsive neonatal megaloblastic anemia and homocystinuria with associated reduced methionine synthase activity
Authors: Hallam LJ , Sawyer M , Clark AC , Van der Weyden MB
Published: October 2019

We present findings on an infant with neonatal megaloblastic anemia, homocystinuria, and neurologic dysfunction that included developmental delay and tonic seizures. There was no methylmalonic aciduria. Cyanocobalamin therapy was accompanied by complete hematologic and neurologic recovery, diminished homocystine excretion, and subsequently normal neurologic development. Cultured fibroblasts and lymphoblasts showed a reduced methionine synthase activity and a growth requirement for methionine. Cobalamin incorporation by the patient's lymphoblasts was normal, but the proportion of cellular methylcobalamin in the patient's lymphoblasts and fibroblasts were markedly reduced and that of adenosylcobalamin normal. The reduced methionine synthase activity was independent of assay reducing (thiol) conditions, but normal levels of activity accompanied culture of the patient's lymphoblasts in medium with markedly increased cobalamin concentration. The characteristics of the reduced methionine synthase of our patient differ significantly from that of the previously described infant with cobalamin E disease and suggest that genetic heterogeneity may characterize this mutation.
Correlation of drug-perturbed marrow cell growth kinetics and intracellular 1-B-D-arabinofuranosylcytosine metabolism with clinical response in adult acute myelogenous leukemia
Authors: Karp JE , Donehower RC , Dole GB , Burke PJ
Published: October 2019

To define the relationship between leukemic cell growth, intracellular metabolism of 1-B-D-arabinofuranosylcytosine (ara-C), and the clinical response to timed sequential induction therapy with ara-C in adult acute myelogenous leukemia (AML), growth kinetic and biochemical pharmacologic determinants were examined in AML bone marrow populations. Leukemic blasts from 45 previously untreated patients obtained prior to therapy were cultured in vitro in autologous pretreatment serum (APS) and in serum containing drug-induced humoral stimulatory activity (HSA). Cell populations cultured in HSA demonstrated both increased proliferation, as measured by both [3H]dThd incorporation into DNA and [3H]dThd leukemic blast labeling index, and greater [3H] ara-C leukemic blast labeling index relative to cells maintained in APS. HSA-cultured marrow cells from the 31 patients who achieved complete remission with ara-C-containing therapy demonstrated enhanced intracellular formation of ara-C 5′-triphosphate over three hours and retention of this active form during one subsequent hour in drug-free medium relative to cells maintained in APS. In contrast, cells from the 14 nonresponsive patients demonstrated no such HSA- induced increases in intracellular ara-C metabolism. These studies of human AML marrow cells identify behavior patterns of ara-C activation and net metabolism in the kinetically perturbed, proliferative state that may discriminate clinical sensitivity from clinical resistance to ara-C-based timed sequential therapy. Sensitive AML populations behave similarly to normal hematopoietic cohorts, with direct linkage of HSA- perturbed growth and pharmacologic parameters, while refractory cells demonstrate uncoupling of these determinants in the growth-stimulated state. These in vitro measurements may further serve as a template for prediction of clinical outcome to timed sequential therapy with ara-C, where both pharmacologic and cytokinetic determinants of response are intrinsic to the success of the designed drug scheduling.
The molecular basis of beta-thalassemia in Lebanon: application to prenatal diagnosis
Authors: Chehab FF , Der Kaloustian V , Khouri FP , Deeb SS , Kan YW
Published: October 2019

A study of the molecular lesions of beta-thalassemia in Lebanon revealed the presence of eight different mutations in 25 patients with Cooley's anemia. The IVS1 position 110 mutation predominated with a frequency of 62% and was almost invariably associated with Mediterranean chromosome haplotype I. Five other mutations commonly found in the Mediterranean area occurred with frequencies of 2% to 8%. In addition a G----C substitution in IVS1 position 5 (a lesion previously found in Chinese and Asian Indians) was demonstrated in a patient with Mediterranean haplotype IX. A new mutation at codon 29 was found in two other patients with haplotype II. The characterization of these beta-thalassemia mutations should allow the implementation of a prenatal diagnosis program in that country.
The regulatory role of interleukin 2-responsive T lymphocytes on human marrow granulopoiesis
Authors: Estrov Z , Roifman C , Mills G , Grunberger T , Gelfand EW , Freedman MH
Published: October 2019

The effect of recombinant interleukin 2 (IL2) on marrow CFU-C colony formation was evaluated to define the role for T lymphocytes in human marrow granulopoiesis. The colony-stimulating factor (CSA) used in our experiments was found to contain IL2. IL2 depletion from CSA resulted in a reduction in CFU-C colony proliferation. Addition of exogenous IL2 caused an increase in CFU-C colony numbers in a dose-dependent manner. This increase could be prevented by anti-Tac, a monoclonal antibody (MoAb) to the IL2 receptor. Moreover, anti-Tac in the absence of exogenous IL2 resulted in an overall decrease in colony numbers. Depletion of either adherent cells or T lymphocytes abolished the effect of IL2 and anti-Tac on colony growth. In the presence of IL2, re- addition of T lymphocytes to the T-depleted marrow or adherent cells to adherent cell-depleted marrow resulted in a significant increase in CFU- C colony numbers, whereas no significant effect was found when IL2- depleted CSA was used. Although T lymphocytes were not themselves essential for CFU-C colony growth, our studies indicate that IL2 and IL2-responsive T cells can regulate in vitro granulopoiesis.
Membrane receptors for interleukin 2 on hematopoietic precursors in chronic myeloid leukemia
Authors: Visani G , Delwel R , Touw I , Bot F , Lowenberg B
Published: October 2019

In this report we present data on the expression of IL2 receptors on chronic phase CML cells. Using an anti IL2 receptor monoclonal antibody (McAb aIL2r) in indirect immunofluorescence we found significant proportions (42.2% +/- 19.7 SD) of the CML cells (previously depleted of E rosetting T cells) to be IL2 receptor positive following incubation in suspension for 18 hours at 37 degrees C. Noninduced cells did not express IL2 receptors. After induction the aIL2r positive and negative cell subpopulations were sorted and analyzed separately for morphology, lineage specific cell surface markers, and clonogenic cell numbers. The IL2 receptor positive CML subpopulations mainly contained blast cells and monocytes and revealed reactivity with myeloid McAbs but not with T cell, B cell, platelet, or erythroid markers. Clonogenic cells (CFU-GEMM, BFUe, and CFU-GM) were selectively recovered from aIL2r positive CML cells and thus were IL2 receptor positive. The addition of recombinant IL2 (rIL2) to CFU-GM and BFUe cultures, in concentrations from 50 to 500 U/mL, did not influence the efficiency of colony formation. Binding of a radiolabeled IL2 preparation to the in vitro activated CML cells indicated the presence of low affinity receptors for IL2. In contrast to CML, normal human marrow cells were consistently aIL2r nonreactive. Thus, IL2 receptor inducibility is a characteristic feature of CML clonogenic cells, which they share with AML, but not with normal marrow progenitors. The role of IL2 receptors in the regulation of proliferation of CML cells requires further investigation.
Further studies of the secretory pathway in thrombin-stimulated human platelets
Authors: White JG , Krumwiede M
Published: October 2019

The pathway followed by secretory products stored in platelet alpha granules during the release reaction remains controversial. Tannic acid has been used in the present study as an electron-dense stain to follow the secretory process in thrombin-stimulated platelets. Preliminary experiments demonstrated that tannic acid precipitates fibrinogen, and binds osmium tetroxide to fibrinogen and fibrin strands. Examination of platelets fixed at short intervals after exposure to thrombin and incubated in solutions containing tannic acid revealed electron-dense deposits of osmium not apparent in resting platelets. Granules and lumina of channels making up the open canalicular system (OCS) were unstained in discoid cells. However, exposure to thrombin at concentrations of 1 to 5 U/mL for thirty seconds or more resulted in intense staining of alpha granules by osmium. Some granules communicated directly with dilated channels of the OCS, and several were frequently connected to the same canaliculus. The electron-dense substance in swollen granules and channels appeared to be in the process of extrusion through narrow or dilated openings of the OCS onto the platelet surface. Granule-to-granule fusion and formation of sealed vacuoles of fused granule products unstained by tannic acid-osmium were not observed. The findings support the concept that secretion by stimulated human platelets results from development of direct communications between granules and channels of the OCS and subsequent extrusion of products through channel pores to the surrounding medium.
Retinoic acid cooperates with tumor necrosis factor and immune interferon in inducing differentiation and growth inhibition of the human promyelocytic leukemic cell line HL-60
Authors: Trinchieri G , Rosen M , Perussia B
Published: October 2019

In this study, we analyzed the effect of tumor necrosis factor (TNF) on retinoic acid (RA)-induced myeloid differentiation of the promyelocytic HL-60 leukemia cell line. We show that low concentrations of the two substances, almost inactive in inducing differentiation when used separately, induce differentiation when added simultaneously to the cell cultures. Cells simultaneously expressing both monocyte/macrophage phenotype (typically induced by TNF) and granulocyte characteristics (typically induced by RA) are induced by a combination of the two factors, indicating that TNF and RA potentiate each other's activity. The results obtained using immune interferon (IFN-gamma) in combination with the two inducers suggest that the mechanism of action of TNF and IFN-gamma are possibly different. The inhibitory effect of RA on the expression of HLA class I antigens and of the high-affinity Fc receptor is potentiated by TNF but completely reversed by rIFN-gamma.
Improved results of treatment of adult acute lymphoblastic leukemia
Authors: Linker CA , Levitt LJ , O'Donnell M , Ries CA , Link MP , Forman SJ , Farbstein MJ
Published: October 2019

We designed a treatment program to improve the outcome for adults with acute lymphoblastic leukemia (ALL). Treatment included a remission- induction phase followed by intensive alternating cycles of non-cross- resistant chemotherapy and prolonged oral maintenance therapy. Eighty- one consecutive previously untreated patients were entered on this study. Ninety-four percent of patients entered complete remission. A Kaplan-Meier analysis predicts that 53% +/- 9% (SEM) of patients in remission will remain free of disease at 3 years. Neither age, sex, WBC count, nor immunophenotype had a significant effect on remission duration. This program of intensive cyclical chemotherapy has improved the disease-free survival of patients with adult ALL.
Chromosome aberrations and clinical features of adult T cell leukemia- lymphoma not associated with human T cell leukemia virus type I
Authors: Shimoyama M , Abe T , Miyamoto K , Minato K , Tobinai K , Nagoshi H , Matsunaga M , Nomura T , Tsubota T , Ohnoshi T
Published: October 2019

Chromosome aberrations and clinical features of three patients with adult T cell leukemia-lymphoma (ATL) not associated with human T cell leukemia virus type I (HTLV-I) are described. From their clinical features, two patients were diagnosed as acute type and one patient was diagnosed as chronic type, which later converted to acute crisis. Clonal and many chromosomal abnormalities were observed before therapy in the two acute type cases and at relapse in the chronic type case. Karyotype aberrations, including trisomy 3, trisomy 7, trisomy 21, del(6)(q21), del(10)(p13), 14q11 translocation, and loss of X chromosome, all of which are frequently found in HTLV-I associated ATL, were also seen in these cases of HTLV-I-negative ATL.
Heparin binding defect in a new antithrombin III variant: Rouen, 47 Arg to His
Authors: Owen MC , Borg JY , Soria C , Soria J , Caen J , Carrell RW
Published: October 2019

Antithrombin III (AT-III) Rouen is a hereditary abnormal antithrombin with normal progressive inhibitory activity and reduced heparin cofactor activity. It was isolated from the plasma of a woman who suffered a sudden idiopathic sensorineural hearing loss and balance impairment. There was no familial history of thrombosis. By heparin- Sepharose chromatography, AT-III Rouen was separated from the normal antithrombin on elution with increasing concentrations of NaCl. AT-III Rouen eluted earlier than is normal at both pH 7.4 and pH 6.0. At the lower pH, the antithrombins bound more avidly to the column, with the abnormal AT-III eluting closer to the normal than at the higher pH. Two- dimensional peptide mapping of tryptic and Staphylococcus aureus V8 protease digests of carboxymethylated antithrombins was performed on thin-layer silica plates. The abnormal peptide was located by tryptophan staining, and amino acid analysis and sequence studies demonstrated a substitution of an arginine at residue 47 for a histidine. Results from this study suggest that replacement of arginine 47 by a partially positively charged histidine has less effect on the heparin binding affinity than dose replacing it with a neutral cysteine side chain as in AT-III Toyama, in which no heparin binding was observed. In addition, heparin binding per se is not a sufficient condition to activate AT-III.
Clinical investigation of human alpha interferon in chronic myelogenous leukemia
Authors: Talpaz M , Kantarjian HM , McCredie KB , Keating MJ , Trujillo J , Gutterman J
Published: October 2019

Fifty-one patients with previously untreated or minimally treated chronic myelogenous leukemia in chronic phase received human alpha interferon 3 to 9 X 10(6) units intramuscularly (IM) daily until complete hematologic remission, then at doses ranging from 3 X 10(6) units every other day to 9 X 10(6) units daily. Forty-one (80%) patients achieved a hematologic response, 36 (71%) of them attaining a complete hematologic remission with normal peripheral WBC and differential counts. Responding patients showed continuous but slow normalization of several other blood and marrow parameters including platelet counts, serum lactic dehydrogenase and B12 levels, and marrow cellularity and maturation index. Suppression of the Philadelphia chromosome on serial cytogenetic studies of marrow metaphases was documented in 20 of the 36 patients who achieved complete hematologic remission (56%; 39% of total group), eight of whom (22%) had a decrease of the Philadelphia chromosome-positive metaphases to less than 35%. These changes were persistent for 6 months or longer in 18 patients, seven of whom had continuous suppression of the Philadelphia chromosome to less than 90% for a median of 30+ months (range 21+ to 39+ months). After a median follow-up period of 37 months, 25 patients remain in continued disease control with interferon therapy. The projected 3-year survival rate is 76%, with a yearly death rate of 6%, 9%, and 9% in the first 3 years. Response, Philadelphia chromosome suppression, and survival were significantly better among patients in the low-risk category compared to intermediate- and high-risk categories, as defined by a multivariate analysis-derived prognostic model. The projected 3- year survival rate was 94% for patients who achieved a complete hematologic remission on interferon therapy and 45% for those who did not. Thirteen patients have developed blastic crisis, six with lymphoid and three with undifferentiated morphology. We conclude that human leukocyte alpha interferon effectively controls chronic myeloid leukemia and allows reappearance of diploid hemopoietic cells in some patients.
Immature dense granules in platelets from mice with platelet storage pool disease
Authors: Reddington M , Novak EK , Hurley E , Medda C , McGarry MP , Swank RT
Published: October 2019

Mepacrine uptake into platelets and bone marrow megakaryocytes was analyzed to further characterize the dense granule defects in a group of seven mouse pigment mutants that have characteristics of platelet storage pool disease (SPD). In contrast to our previous studies using electron microscopy, this method revealed that all mutants had normal numbers of dense granules. However, total mepacrine uptake in all mutant platelets was significantly diminished to less than 50% of normal uptake. Also, the flashing phenomenon observed when normal dense granules are irradiated with ultraviolet light was either greatly diminished or absent when platelets of individual mutants were similarly irradiated. Therefore the principal defect in the mutant platelets is an inability to accumulate dense granule contents rather than an absence of the granules. Mepacrine uptake into megakaryocytes was indistinguishable in normal and mutant mice. This indicates the mutant dense granule defects appear either very late in megakaryocyte development or early in platelet formation in correlation with development of the mature dense granule. By standard transmission electron microscopy we have not been able to detect gross structural or subcellular abnormalities in either platelets or megakaryocytes of mutant mice. It appears all seven mutants produce immature or functionally abnormal dense granules.
The presence of clonogenic cells in high-grade malignant lymphoma: a prognostic factor
Authors: Tweeddale ME , Lim B , Jamal N , Robinson J , Zalcberg J , Lockwood G , Minden MD , Messner HA
Published: October 2019

A culture system has been developed that promotes growth of clonogenic lymphoma cells of some patients with intermediate and high-grade malignant lymphoma. The formation of colonies in bone marrow, lymph nodes, and peripheral blood samples is best supported by human plasma. Colony formation of some patients was dependent upon growth factors, which in this study were added in the form of medium conditioned by phytohemagglutinin (PHA)-stimulated leukocytes (PHA-LCM). Some gave rise to lymphoma colonies without PHA-LCM but improved their frequency with PHA-LCM; others were completely independent of PHA-LCM. Colonies grown in primary cultures were routinely recloned and propagated as Epstein-Barr virus (EBV)-negative cell lines with stable B cell phenotype. The cell lines showed the same immunoglobulin rearrangement pattern as that observed in the primary lymphoma sample. In addition, a significant clinical correlation was observed between culture data and clinical outcome. Survival of patients who formed lymphoma colonies at any time during their clinical course was significantly shorter than survival of patients who did not give rise to colonies (P = 0.0009). The same observation was made when the survival assessment was performed for the subset of patients studied at diagnosis (P = 0.0014).
Initiation and regulation of fibrinolysis in human plasma at the plasminogen activator level
Authors: Wun TC , Capuano A
Published: October 2019

The initiation and regulation of fibrinolysis has been studied by reconstitution of fibrinolytic activity in human plasma in vitro. Depletion of tissue plasminogen activator (tPA) antigen by immunoadsorption of human plasma with anti-tPA Ig Sepharose 4B leads to total loss of spontaneous fibrinolytic activity determined by lysis of a thrombin-induced clot. Addition of physiological concentrations of purified tPA to tPA-depleted plasma restores fibrinolytic activity as a function of the length of time between tPA addition and clotting. Addition of free tPA to tPA-depleted plasma followed by immediate clotting results in a high rate of fibrinolysis. In contrast, when free tPA is allowed to incubate in plasma for 10 to 60 minutes prior to clot formation, the fibrinolytic activity of tPA is gradually lost. The loss of tPA-induced fibrinolytic activity in unclotted plasma is accompanied by decreased partitioning of tPA antigen into fibrin after clotting and is kinetically correlated with the formation of a 100 kilodalton (kDa) tPA complex as demonstrated by SDS-gel electrophoresis and fibrin-agar zymography. These results suggest that free tPA is susceptible to complexation by the plasma inhibitor in the absence of a clot. Fibrin formation renders tPA relatively inaccessible to inhibition. The tPA antigen isolated from stored plasma consists mainly of 100 kDa activity in SDS-gel electrophoresis and zymography, indicating that the tPA complex is resistant to dissociation by SDS. Upon rezymography of the sliced gel, only a 60 kDa tPA activity is found, suggesting that the activity at 100 kDa is at least partly due to free tPA dissociated from the complex during the first zymography. Conversion of tPA complex to enzymatically active free tPA also occurs with brief SDS exposure followed by incubation in the presence of excess Triton X-100 or by hydroxylamine treatment. These results reconcile the apparent discrepancy of the 100 kDA inhibitor-tPA complex manifesting plasminogen activation activity during zymography. The plasma tPA- inhibitor complex is precipitated strongly by antisera against plasminogen activator inhibitors (PAIs) of human Hep G2 hepatoma and HT- 1080 fibrosarcoma cells and weakly by antiserum against bovine aortic endothelial cell PAI but not by antiserum against a placental PAI (PAI- 2) suggesting that the plasma inhibitor is immunologically related to Hep G2, HT-1080 and possibly endothedial cell PAIs. Based on the above findings, a simple model for the initiation and regulation of plasma fibrinolysis at the PA level has been formulated.
Purging leukemic cells from simulated human remission marrow with alkyl- lysophospholipid
Authors: Okamoto S , Olson AC , Vogler WR , Winton EF
Published: October 2019

Alkyl-lysophospholipids (ALP) are analogues of 2- lysophosphatidylcholine that have been reported to have selective antitumor activity. These compounds could potentially be useful in purging bone marrow of leukemic cells in autologous marrow transplantation in acute leukemia. To determine the efficacy of pharmacological purging by ALP, we have designed a human assay system to mimic the conditions expected in the clinical setting of autotransplantation using remission marrow. A simulated remission marrow (SRM) was prepared by mixing normal marrow cells and HL60 cells in a ratio of 1,000:1. The effect of cryopreservation on ALP-treated normal, HL60, and SRM cells was examined. In separate experiments, ALP significantly reduced the number of clonogenic HL60 cells with no effect on normal marrow progenitors. The effect of ALP was more apparent after cryopreservation. Incubation of HL60 cells with 50 micrograms/mL ALP for four hours followed by cryopreservation resulted approximately in a 3 log reduction of clonogenic HL60 cells. ALP also selectively purged the small number of leukemic cells from SRM. In SRM, the data suggested that ALP had indirect cytotoxic activity on leukemic cells by enhancing the cytotoxic activity of monocytes in addition to its direct effect. We found no evidence that clonogenic HL60 cells decreased because of induction of differentiation by ALP. These data indicated that treatment of marrow cells with ALP offers an efficient means to eliminate leukemic cells from the graft.