bluuebunny/crossref_metadata_2025 huggingface dataset:
Clinical pharmacology of low-dose cytosine arabinoside
Authors: Spriggs D , Griffin J , Wisch J , Kufe D
Published: October 2019

Low doses of cytosine arabinoside (ara-C) have recently been administered by intravenous (IV) infusion and intermittent subcutaneous (SC) injection to patients with pre-leukemia and acute leukemia. Our studies have demonstrated that the continuous IV infusion of low-dose (20 mg/m2/d) ara-C produces hematologic improvement in patients with preleukemic syndromes. The present work has monitored plasma ara-C levels in five of these patients. The results demonstrate mean steady- state plasma levels ranging from 1.8 to 6.9 X 10(-8) mol/L. The range for total drug exposure (area under the curve) for the 14-day course was 6.5 to 15.9 X 10(-6) mol/L X hour. These findings have been compared to the pharmacokinetics of ara-C (10 mg/m2) given by bolus SC injection. This dose schedule resulted in peak ara-C levels 15 minutes after injection that were tenfold to 30-fold higher than the mean plasma level achieved during continuous IV infusion in the same patient. Furthermore, there was no detectable plasma ara-C at six hours after bolus injection. The differences in ara-C pharmacology for the continuous IV infusion and bolus SC injection dose schedules may contribute to the variability in response and toxicity achieved with these regimens.
Quantitation of protein 3 content of circulating erythrocytes at the single-cell level
Authors: Jennings LK , Brown LK , Dockter ME
Published: October 2019

The density and size of human erythrocytes has been roughly correlated with cell age, with the denser and smaller cells being older. Observations of this type have led to a hypothesis that the membranes of circulating erythrocytes are dynamic with respect to composition and that material is lost from the membrane during cell maturation and circulation. In this study, flow cytofluorimetry was used to investigate the distribution of the human erythrocyte anion transport protein (protein 3) in heterogeneous samples of circulating red cells. We verified that protein 3 can be specifically and quantitatively labeled in intact human erythrocytes with eosin-5-maleimide, a luminescent probe. Individual cells were accordingly analyzed for size by forward light scattering and for protein 3 content by quantitation of eosin fluorescence. Initial results indicated that the smallest erythrocytes had a protein 3 content equal to that of the largest circulating erythrocytes. This result was independently verified by light scatter-activated cell sorting; direct measurement of cell diameters by microscopy verified that the cell sizes of erythrocytes showing the 10% greatest and 10% smallest light-scattering signal were indeed distinct. Independent analysis of the size-sorted erythrocytes for protein 3 content was accomplished by gel electrophoresis of stroma from 150,000 large and small erythrocytes. Quantitative scanning densitometry of silver-stained gels of prepared stroma showed that protein 3 content of each set of fractionated cells was equal and did not vary as a function of cell size. Taken in combination with the reported correlation between increasing red blood cell age and decreasing cell size, these results indicate that any loss of membranous material during the cell aging process is not random.
Enhanced plasminogen-activator production by leukocytes in the human and murine Chediak-Higashi syndrome
Authors: de Saint Basile G , Fischer A , Dautzenberg MD , Durandy A , Le Deist F , Angles- Cano E , Griscelli C
Published: October 2019

We have studied the coagulation status of eight patients with the Chediak-Higashi syndrome (CHS), both in the chronic and the accelerated phase of the disease. It has been shown that during the accelerated phase there are coagulation abnormalities. These abnormalities include a peripheral thrombocytopenia, minor alterations of liver clotting factors, and mainly a profound hypofibrinogenemia and hypoplasminogenemia, which cause life-threatening bleedings. These disorders are of complex origin, but a fibrinolytic process, possibly primary, appears to play a significant role, since the present evidence for intravascular coagulation is not definitive. The accelerated phase of the CHS is characterized by a visceral infiltration by macrophages and lymphocytes. Therefore, we have investigated the possible role of the macrophages in the fibrinolytic process. We have found an excessive plasminogen activator (PA) production by CHS mononuclear cells in the accelerated phase and to a lesser extent in the chronic phase, except in one patient in whom no anomaly was found. Single-cell studies revealed an increased number of PA-producing cells among the monocyte- macrophage lineage rather than a higher level of production per cell. Polymorphonuclear cells (PMN) from patients with CHS were also shown to contain more PA. Slight but significant abnormalities in PA production were observed in obligatory heterozygotes (five out of nine), indicating the inherited nature of the excessive PA production. Finally, an enhanced PA production was similarly demonstrated using beige mice macrophages. The exacerbated production of PA by macrophages in the accelerated phase of the CHS can account to some extent for the coagulation abnormalities that have been observed.
Zinc-induced platelet aggregation is mediated by the fibrinogen receptor and is not accompanied by release or by thromboxane synthesis
Authors: Heyns A du P , Eldor A , Yarom R , Marx G
Published: October 2019

We demonstrate that zinc (0.1 to 0.3 mmol/L) induces aggregation of washed platelet suspensions. Higher concentrations (1 to 3 mmol/L) of zinc were needed to aggregate platelets in platelet-rich plasma obtained from blood anticoagulated with low-molecular-weight heparin, probably due to the binding of zinc to the plasma proteins. Zinc- induced aggregation of normal washed platelets required added fibrinogen and no aggregation occurred with thrombasthenic platelets or with normal platelets pretreated with a monoclonal antibody (10E5) that blocks the platelet fibrinogen receptor. These data indicate that the platelet membrane fibrinogen receptor-glycoproteins IIb and IIIa mediate the effect of zinc. Zinc-induced aggregation was blocked by the agent TMB-8, which interferes with the internal calcium flux, and by prostacyclin, which elevates platelet cyclic adenosine monophosphate levels. Zinc-induced aggregation was not accompanied by thromboxane synthesis or by the secretion of dense-body serotonin and was not affected by preexposure of platelets to acetylsalicylic acid. Experiments with creatine phosphate/creatine phosphokinase showed that the zinc effect on platelets was independent of extracellular adenosine diphosphate (ADP). Zinc had an additive effect when platelet aggregation was stimulated with subthreshhold concentrations of collagen or ADP. Together with the known effects of nutritional zinc on in vivo bleeding, on platelet aggregation, and on lipid metabolism, the results suggest that zinc may have an important bearing on normal hemostasis, thrombosis, and atherosclerosis.
Late expression of M and N antigens on glycophorin A during erythroid differentiation
Authors: Ekblom M , Gahmberg CG , Andersson LC
Published: October 2019

The M/N blood groups are carried by the major human red cell sialoglycoprotein, glycophorin A. O-glyosidic carbohydrate is needed for the activity, but the M/N specificity is due to amino acid replacements in the NH2-terminal portion of the molecule. We have used monoclonal antibodies specific for M and N blood groups to study their expression during normal erythropoiesis. Here we report that the M/N blood group activities are very weakly or not at all expressed before the polychromatic normoblast stage. Using polyclonal anti-glycophorin A antiserum, it was shown that glycophorin A molecules are already abundantly present on the earliest morphologically recognizable erythroid precursor, the proerythroblast. These findings can be explained by our previous observation that the O-glycosylation of glycophorin A gradually increases during erythroid maturation.
Distinctive lymphocyte subpopulation abnormalities in patients with congenital coagulation disorders who exhibit lymph node enlargement
Authors: Prince HE , Kreiss JK , Kasper CK , Kleinman S , Saunders AM , Waldbeser L , Mandigo G , Kaplan HS
Published: October 2019

The majority of patients with congenital clotting disorders who use clotting factor concentrate exhibit lymphocyte subpopulation abnormalities. A subset of these patients develop lymph node enlargement (LNE), part of the spectrum of clinical disease associated with the acquired immune deficiency syndrome (AIDS). It is therefore important to determine if these patients with LNE exhibit specific immune alterations suggestive of early infection with the AIDS agent. We used one- and two-color immunofluorescence to distinguish the lymphocyte subpopulation alterations associated with concentrate use from those associated with LNE. Patients who use concentrate had elevated levels of Leu-2+ (T suppressor phenotype) cells and Leu-7+ (phenotype of some natural killer) cells. These increased levels were largely caused by a dramatic (2.6-fold) increase in the number of lymphocytes co-expressing Leu-2 and Leu-7 (2+7+). A dose-response effect between amount of concentrate infused during the preceding year and level of 2+7+ cells was observed. Concentrate recipients, as a group, also showed increased levels of T cells expressing Dr antigen (T+Dr+ phenotype, characteristic of activated or immature T cells) and cells expressing the T10 antigen (phenotype of some null cells and activated/immature T cells). Patients with LNE showed a further increase in T10+ cells as well as a distinctive decrease in Leu-3+ (T helper phenotype) lymphocytes. All LNE patients exhibited either low Leu-3+ levels, high T10+ levels, or both. Thus, concentrate use was associated with increased levels of Leu-2+ (particularly 2+7+) cells and T+Dr+ cells, whereas LNE was associated with decreased levels of Leu-3+ cells and high levels of T10+ cells.
Biochemical and functional consequences of dissociation of the platelet membrane glycoprotein IIb-IIIa complex
Authors: Shattil SJ , Brass LF , Bennett JS , Pandhi P
Published: October 2019

The platelet membrane glycoproteins, IIb and IIIa, form a Ca2+- dependent heterodimer complex that functions as the fibrinogen receptor in activated platelets to mediate platelet aggregation. Little is known about factors that affect the IIb-IIIa complex within the platelet membrane. It has been observed that platelets incubated with ethylene glycol tetra-acetic acid (EGTA) at 37 degrees C are unable to aggregate or to bind monoclonal antibodies specific for the IIb-IIIa complex. To determine whether this is due to a dissociation of IIb from IIIa, we developed a method for quantitating the complex on nondenaturing, polyacrylamide gradient gels. Platelets were surface-labeled with 125I and then solubilized and electrophoresed in 0.2% Triton and 10 mmol/L CHAPS. Under these conditions and in the presence of 1 mmol/L Ca2+, glycoproteins IIb and IIIa migrated on the gels as a discrete band at Rf = 0.33. Protein that was eluted from this band bound to an immunoaffinity column specific for the IIb-IIIa complex. In contrast, when the IIb-IIIa complex was solubilized and then dissociated with EGTA, the discrete band at Rf = 0.33 was no longer present, and IIb and IIIa were now found in a broad band at Rf = 0.45 to 0.50. To study IIb and IIIa within the surface membrane, the 125I-labeled platelets were first incubated with 0.5 mmol/L EGTA (1 nmol/L free Ca2+) at 22 degrees C and then solubilized in the absence of EGTA. The IIb and IIIa from these platelets migrated at Rf = 0.33, indicating the presence of the intact IIb-IIIa complex. In contrast, when the platelets were incubated at 37 degrees C for one hour with the EGTA, the discrete band at Rf = 0.33 representing the IIb-IIIa complex gradually disappeared. This phenomenon could not be reversed by adding Ca2+ back to the platelets before solubilization and electrophoresis. This loss of the IIb-IIIa complex from intact platelets was accompanied by (a) a progressive and irreversible decrease in adenosine diphosphate (ADP)-induced platelet aggregation and (b) decreased binding of a complex-dependent monoclonal antibody to the platelets. These studies demonstrate that when platelets are exposed to low Ca2+ at 37 degrees C, the IIb-IIIa heterodimer complexes in their surface membranes are irreversibly disrupted. Because intact IIb-IIIa complexes are required for platelet aggregation, the loss of these complexes may account for the failure of these platelets to aggregate in response to ADP.
Granulated T cell lymphocytosis with neutropenia: malignant or benign chronic lymphoproliferative disorder?
Authors: McKenna RW , Arthur DC , Gajl-Peczalska KJ , Flynn P , Brunning RD
Published: October 2019

The clinical, morphological, immunologic, and cytogenetic features of seven cases of chronic granulated T cell lymphocytosis with neutropenia were studied. The disorder was characterized by moderate blood and bone marrow lymphocytosis, neutropenia, polyclonal hypergammaglobulinemia, splenomegaly, absence of lymphadenopathy, and a chronic, relatively stable clinical course. The proliferative lymphocytes manifested a cytotoxic/suppressor T lymphocyte phenotype. In two of four cases studied, blood lymphocytes showed clonal chromosome abnormalities. One patient treated with pulse steroid therapy had reversal of lymphocytosis and severe neutropenia with subsequent resolution of an intractable infection. The lymphocytosis and neutropenia recurred when steroids were withdrawn. Six of the seven patients were living three months to 17 years from diagnosis; one died at 4.3 years of an unrelated cause. Five of the patients, including the two with lymphocyte chromosome abnormalities, had persistent lymphocytosis and neutropenia from three months to 13 years from diagnosis. In two patients, the disease appears to have undergone spontaneous regression. No differences in clinical presentation or the morphological or immunologic characteristics of the proliferative lymphocytes were apparent between those patients with lymphocyte chromosome abnormalities and persistent disease and those who had a spontaneous regression. The finding of clonal chromosome abnormalities in the blood lymphocytes of two of the patients in this study suggests a neoplastic origin for chronic granulated T cell lymphocytosis with neutropenia. However, apparent spontaneous regression in two patients, one after 11 years, lends support to a chronic reactive or immunoregulatory disorder as the etiology. It is probable that cases of granulated T cell lymphocytosis with neutropenia, although morphologically and immunologically similar, are biologically heterogeneous.
Favism: disordered erythrocyte calcium homeostasis
Authors: De Flora A , Benatti U , Guida L , Forteleoni G , Meloni T
Published: October 2019

The biochemical events that take place during acute hemolysis of G6PD- deficient subjects in favism are far from being elucidated. Evidence is here reported for a constantly and heavily disordered calcium homeostasis in the erythrocytes from seven favic patients. The abnormality, ie, a significantly impaired calcium ATPase activity and a parallel marked increase of intracellular calcium levels, was characteristic of the acute hemolytic crisis although unrelated to the attendant reticulocytosis. Concomitantly, a remarkable decrease of intracellular potassium was also observed. The mean +/- SD Ca2+-ATPase activity in the favic patients was 20.8 +/- 7.8 mumol Pi/g Hb/h compared with 37.2 +/- 8.5 in the matched controls represented by 12 healthy G6PD-deficient subjects (P less than .001). The mean +/- SD intraerythrocytic calcium content was 288 +/- 158 mumol/L of erythrocytes in the favic patients as compared with 22.0 +/- 8.2 in the G6PD-deficient controls (P less than .001). The intraerythrocytic potassium content was 76.6 +/- 19.3 mmol/L of erythrocytes in the favic patients and 106.6 +/- 8.2 in the G6PD-deficient controls (P less than .001). In vitro incubation of normal and G6PD-deficient erythrocytes with divicine, a pyrimidine aglycone present in fava beans and strongly implicated in the pathogenesis of favism, reproduces most of these events, including drop of calcium ATPase, increased intracellular calcium, and leakage of erythrocyte potassium.
Differential interleukin-1 elaboration by density-defined human monocyte subpopulations
Authors: Elias JA , Chien P , Gustilo KM , Schreiber AD
Published: October 2019

Interleukin-1 (IL-1) is an important immunoregulatory peptide produced by monocytes and macrophages. Because mononuclear phagocytes are morphologically and functionally heterogeneous, we examined whether they differ in their ability to elaborate IL-1. We used discontinuous Percoll gradients to obtain five density-defined human blood monocyte subpopulations. Unfractionated monocytes and their subsets were compared for their ability to stimulate thymocyte proliferation. Supernatants obtained from the denser monocytes consistently contained more IL-1 activity than did supernatants from the less dense cells. This difference in IL-1 activity was the result of differences in IL-1 elaboration, not the selective production of an inhibitor of IL-1- induced thymocyte proliferation. These data demonstrate that density- defined human monocyte subpopulations differ in their capacity to elaborate IL-1.
Anti-My-28, an antigranulocyte mouse monoclonal antibody, binds to a sugar sequence in lacto-N-neotetraose
Authors: Spitalnik SL , Schwartz JF , Magnani JL , Roberts DD , Spitalnik PF , Civin CI , Ginsburg V
Published: October 2019

Anti-My-28 is an IgM kappa monoclonal antibody produced by a hybridoma prepared from spleen cells of a mouse immunized with normal human granulocytes. By immunofluorescence it binds to human granulocytes but not to monocytes and lymphocytes. However, after treating cells with neuraminidase, the antibody also binds to lymphocytes and monocytes and to many leukemic cell lines and patient leukemic blast cells. Anti-My- 28 binds to several neutral glycolipids and desialylated gangliosides of leukocytes and erythrocytes as detected by radioimmunoassay and immunostaining of thin-layer chromatograms. It recognizes a sugar sequence in lacto-N-neotetraose, Gal beta 1–4GlcNAc beta 1–3Gal beta 1–4Glc. This tetrasaccharide occurs in the glycolipids paragloboside and sialosylparagloboside, and its distal trisaccharide sequence is found in higher glycolipids and in glycoproteins.
Minactivin expression in human monocyte and macrophage populations
Authors: Stephens RW , Golder JP , Fayle DR , Hume DA , Hapel AJ , Allan W , Fordham CJ , Doe WF
Published: October 2019

Adherent monolayer cultures of human blood monocytes, peritoneal macrophages, bone marrow macrophages, and colonic mucosa macrophages were examined for their ability to produce and secrete minactivin, a specific inactivator of urokinase-type plasminogen activator. All except colonic mucosa macrophages produced and secreted appreciable amounts of minactivin, but only blood monocytes were stimulated by muramyl dipeptide (adjuvant peptide) to increase production. The minactivin from each of these populations could be shown to preferentially inhibit urokinase-type plasminogen activator and not trypsin, plasmin, or “tissue”-type plasminogen activator (HPA66). A plasminogen-activating enzyme present in monocyte cultures appeared unaffected by the presence of minactivin and could be shown to be regulated independently by dexamethasone.
Impaired responsiveness to B cell growth factor in a patient with common variable hypogammaglobulinemia
Authors: Perri RT , Weisdorf DJ
Published: October 2019

Common variable hypogammaglobulinemia (CVH) is a clinical syndrome that includes a diverse group of patients with heterogeneous defects resulting in impaired B cell proliferation and terminal differentiation into mature plasma cells capable of normal immunoglobulin synthesis and secretion. In this study, we report our identification of a previously undescribed intrinsic B cell defect in a patient with CVH. This patient's B cells showed a marked impairment in hemolytic plaque- forming cell (HePFC) formation compared with control B cells (15 v 80 HePFCs per culture, respectively). In addition, this patient's B cells displayed decreased B cell colony formation compared with control B cells (5 +/- 2 v 93 +/- 8, respectively). When examined for their responsiveness to phytohemagglutinin-T cell conditioned media (PHA- TCM), the patient's B cells displayed impaired B cell proliferation compared with control B cells (stimulation index [SI] 1.3 +/- 0.20 v 26 +/- 1.4 with 20% control PHA-TCM [vol/vol]). Impaired proliferation by the patient's B cells persisted with increasing concentrations of B cell growth factor (BCGF). Additionally, PHA-TCM prepared from the patient's T cells when compared with control PHA-TCM consistently showed less support for control B cell proliferation (SI 1.27 +/- 0.21 v 26 +/- 1.4, respectively). In coculture studies of B cell proliferation and immunoglobulin synthesis, patient's T cells showed no evidence of an enhanced suppressive effect or decreased helper effect. This patient's immune defects involve, first, an intrinsic B cell defect characterized by an impaired responsiveness to BCGF's proliferation signal and, second, impaired production of BCGF by the patient's T cells.
Diagnostic and prognostic significance of Sezary cells in peripheral blood smears from patients with cutaneous T cell lymphoma
Authors: Vonderheid EC , Sobel EL , Nowell PC , Finan JB , Helfrich MK , Whipple DS
Published: October 2019

Blood smears stained with Wright-Giemsa were obtained from 124 patients with pathologically confirmed cutaneous T cell lymphoma (CTCL), 70 patients with various other cutaneous disorders, and ten healthy adult volunteers. These were examined in a blinded fashion for atypical lymphocytes with cerebriform nuclei (CLs), which were characterized further according to cell diameter. CLs, comprising up to 15% of lymphocytes in smears, were observed in 20% of the patients with benign dermatitis. CLs, comprising up to 89% of lymphocytes in smears, were found in 22%, 30%, 50%, and 96% of patients with patch, plaque, tumor, and erythrodermic CTCL, respectively. Large-diameter CLs (15 to 20 micron) were observed only in smears from patients with CTCL. Total CL counts above 15 per 100 lymphocytes and/or the presence of large CLs occurred in 33 of 49 (67%) patients with erythrodermic disease and in only two patients with other skin manifestations. Blood smears obtained at the time of cytogenetic studies indicated that a total CL count above 15% was the smear criterion that correlated best with the demonstration of a chromosomally abnormal malignant clone in the blood. The presence of large CLs per se, although also predictive of a malignant clone, was less useful. Multivariate survival analysis showed that the duration of disease before the blood smear and the proportion of large CLs within the total CL population were the covariates that correlated most significantly with survival. We speculate that the reduced survival of patients with increased proportions of large CLs in smears reflects the presence of polyploid malignant lymphocytes in the blood.
A study of liver biopsies and liver disease among hemophiliacs
Authors: Aledort LM , Levine PH , Hilgartner M , Blatt P , Spero JA , Goldberg JD , Bianchi L , Desmet V , Scheuer P , Popper H
Published: October 2019

Hepatic histologic materials (biopsy or autopsy) and associated clinical data from 155 hemophiliacs were collected by an ad hoc hemophilia study group and analyzed retrospectively in an effort to determine the spectrum of liver disease in this population and to examine the relationship between the severity of liver disease and treatment history. Clinical information on the frequency of complications from 126 biopsies in 115 hemophilic patients provided a unique opportunity to assess the safety of liver biopsy in such patients. The incidence of cirrhosis (15%) and chronic active hepatitis (7%) was lower than previously reported. The frequency of severe liver disease (chronic active hepatitis or cirrhosis) in patients receiving large pooled concentrates was no greater than in patients treated principally with cryoprecipitate or plasma. The risks of liver biopsy in this setting are relatively high: clinically significant hemorrhage followed 12.5% of the procedures.
Sustained thrombolysis with DNA-recombinant tissue type plasminogen activator in rabbits
Authors: Agnelli G , Buchanan MR , Fernandez F , Van Ryn J , Hirsh J
Published: October 2019

Tissue type plasminogen activator (t-PA) is an effective thrombolytic agent in experimental animals. The duration of the thrombolytic effect of infused t-PA is unknown. We compared the duration of the thrombolytic effect of t-PA with streptokinase by measuring the lysis of 125I-fibrin-labeled thrombi in rabbit jugular veins at different times after a bolus injection of the fibrinolytic agents. The pharmacodynamics of both thrombolytic agents were determined in rabbits using a sensitive ex vivo fibrinolytic assay. Streptokinase and t-PA were given as a bolus dose of 15,000 U/kg. There was no detectable circulating fibrinolytic activity 30 minutes after the bolus dose of t- PA and 120 minutes after the bolus dose of streptokinase. The t-PA injection produced 34% thrombolysis at 30 minutes, 90% thrombolysis at 120 minutes, and 96% thrombolysis at 240 minutes. The streptokinase injection produced 17% thrombolysis at 30 minutes, 34% at 120 minutes, and 34% at 240 minutes. These observations indicate that the thrombolytic effect of t-PA is sustained beyond its time of clearance from the circulation whereas the thrombolytic effect of streptokinase closely parallels its activity in the circulation.
Autologous bone marrow transplantation for patients with acute lymphoblastic leukemia in second or subsequent remission: results of bone marrow treated with monoclonal antibodies BA-1, BA-2, and BA-3 plus complement
Authors: Ramsay N , LeBien T , Nesbit M , McGlave P , Weisdorf D , Kenyon P , Hurd D , Goldman A , Kim T , Kersey J
Published: October 2019

Autologous bone marrow transplantation (BMT) was utilized as therapy for 23 patients with acute lymphoblastic leukemia (ALL) in second or greater remission. Bone marrow was treated in vitro with a combination of monoclonal antibodies, consisting of BA-1, BA-2, BA-3, and baby rabbit complement (BRC'). All patients were prepared for transplantation with cyclophosphamide and fractionated total body irradiation. Engraftment occurred in all 23 patients. Seven of 23 patients remain relapse-free from six to 32 months (median, 21.4 months) posttransplant. Failures were due to relapse with the exception of one patient who died of infection. This study demonstrates that autologous BMT using in vitro marrow treatment with BA-1, BA-2, BA-3, and BRC' is safe, allows engraftment, and results in prolonged survival for some patients with ALL in second or greater remission.
5-Azacytidine increases HbF production and reduces anemia in sickle cell disease: dose-response analysis of subcutaneous and oral dosage regimens
Authors: Dover GJ , Charache S , Boyer SH , Vogelsang G , Moyer M
Published: October 2019

Varying doses of 5-azacytidine (5-aza) were given to four sickle cell individuals for 500, 200, 100, and 30 days. The percentage of fetal hemoglobin (HbF) containing reticulocytes (F reticulocytes) increased two- to five-fold within five days of 5-aza therapy in all patients, with a two- to three-fold rapid response (less than 48 hours after initial dose) in three patients. Reticulocyte suppression was not observed prior to, during, or after therapy in those patients who responded within 48 hours. Subcutaneous 5-aza was given in 35-day courses consisting of every day, every other day, or three consecutive days a week. No marrow toxicity was observed on any of the regimens. For three patients, the highest average F reticulocyte level was observed on the three consecutive day a week regimen. Oral 5-aza, given with tetrahydrouridine, produced comparable F reticulocyte response. In the two patients treated for more than 100 days, Hb levels increased to 11 to 12 and 9 g/dL, MCV and MCH increased by 25%, and lysate HbF levels peaked at 12% and 20%. Fetal erythroid characteristics (i- antigen, galactokinase activity, and G gamma/A gamma ratios) did not correlate with maximal HbF production. The frequency of vasoocclusive crises appeared to decrease in both patients followed for more than 100 days.
Hypoplastic anemia in B cell chronic lymphocytic leukemia: evolution of T cell-mediated suppression of erythropoiesis in early-stage and late- stage disease
Authors: Mangan KF , D'Alessandro L
Published: October 2019

To define further the role of marrow T suppressor lymphocytes in the pathogenesis of the hypoproliferative anemia in all Rai clinical stages of B cell chronic lymphocytic leukemia (CLL), marrow erythroid progenitor cell (CFU-E and BFU-E) frequency, marrow T gamma lymphocyte frequency per 1,000 nucleated marrow cells, and T cell-erythroid progenitor cell interactions were examined in 30 CLL patients and normal control subjects. As compared with control subjects, decreased numbers of CFU-E and BFU-E were found in patient marrow depleted of neoplastic B cells in all Rai stages of the disease. As a group, Rai stage III through IV patients with or without aplasia (CLL-aplasia) had significantly fewer CFU-E and BFU-E than did Rai O through II stage patients. The numbers of T gamma cells infiltrating CLL marrows were increased 3, 9, and 20 times normal in Rai O through II, Rai III through IV, and CLL-aplasia groups, respectively. Removal of T cells from marrow increased growth of CFU-E and BFU-E in all Rai O through IV patients, but the increase was significant in the CLL-aplasia group only (P less than .05). However, autologous coculture of marrow T cells or T gamma cells but not B cells with marrow B + T-depleted null cells at ratios of 0.2:1 to 1:1 suppressed CFU-E and BFU-E growth in all three patient groups. We conclude that the hypoproliferative anemia occurring in the course of B cell CLL is due to gradual accumulation in the marrow of T gamma lymphocytes which suppress erythroid progenitor cell growth. T gamma cell suppression of erythropoiesis and marrow T gamma cell expansion is detectable in the earliest Rai stages of the disease.
Role of immune interferon in the monocytic differentiation of human promyelocytic cell lines induced by leukocyte conditioned medium
Authors: Dayton ET , Matsumoto-Kobayashi M , Perussia B , Trinchieri G
Published: October 2019

Conditioned medium (CM) from lectin-stimulated human leukocytes contains factors that induce human promyelocytic cell lines to differentiate along the monocytic pathway. In this report, we show that human promyelocytic cell lines are also induced to differentiate along this pathway by immune interferon (IFN gamma). Various preparations of IFN alpha tested did not induce this differentiation. In cultures containing IFN gamma, the cells are induced to coordinately express monocyte markers and functions such as monocyte-specific surface antigens, HLA-DR antigens, nonspecific esterase, receptors for the Fc fragment of IgG, and the ability to mediate antibody-dependent cell- mediated cytotoxicity. Our data indicate that differentiation induced by IFN gamma is not secondary to an arrest of growth of promyelocytic cell lines, but rather that a proportion of cells is induced along a programmed pathway of terminal differentiation similar to that of normal monocytes. CM contains IFN gamma, but its ability to induce differentiation is greater than expected on the basis of its content of IFN gamma. Treatments at 56 degrees C or at pH 2.0, which abolish IFN gamma activity, abrogate the differentiation ability of CM. The antiviral activity and the differentiation activity contained in the CM are coeluted from gel filtration and reverse-phase columns. Monoclonal antibodies anti-IFN gamma, which completely abrogate the differentiation ability of IFN gamma and the antiviral activity in the CM, completely suppress the induction of some monocyte markers by CM, but only reduce the expression of others. When IFN gamma is added to CM, promyelocytic cell lines are induced to differentiate to a much greater extent than that induced by either IFN gamma or IFN gamma- depleted CM alone. These results show that the differentiation activity of leukocyte CM is due to the synergistic effect of IFN gamma and other factors not yet identified.
Surface membrane glycoproteins of wild-type and differentiation-inducer- resistant HL-60 cells
Authors: Felsted RL , Gupta SK , Glover CJ , Gallagher RE
Published: October 2019

Surface membrane glycoproteins (SMGs) of cells from the parental wild- type HL-60 cell line and from three sublines variably cross-resistant to the granulocyte differentiation-inducing effects of retinoic acid (RA), dimethylsulfoxide (DMSO), and certain purine bases (6-thioguanine [6TG] or hypoxanthine) were studied by one-dimensional and two- dimensional gel electrophoresis. After both oligosaccharide (periodate/borotritide) and peptide (1,3,4,6-tetrachloro-3 alpha, 6 alpha-diphenylglycouril) ectolabeling procedures, striking common changes were noted in the gel electrophoretic patterns of the SMGs from the RA- and 6TG-resistant sublines compared to those from the wild-type HL-60 line or the DMSO-resistant subline. Most prominently, this included the presence in the RA- and 6TG-resistant cells of an apparent high molecular weight acidic glycoprotein(s) (mol wt, 200 to 285 kilodaltons [kD]; isoelectric point range [pl], 4.5 to 6.0) not observed in the wild-type or DMSO-resistant cells and, conversely, the presence of a lower molecular weight glycoprotein(s) (mol wt, 120 to 165 kD; pl, 4.2 to 5.9) in the wild-type and DMSO-resistant cells, which was absent or much reduced in the RA- and 6TG-resistant cells. These acidic SMGs did not change as a function of the induction of granulocyte differentiation. However, some other more basic SMGs varied as a function of granulocyte differentiation in both the wild-type and differentiation inducer-resistant sublines, including the loss of the transferrin receptor and the gain of a mol wt 55- to 60-kD neutrophil- associated protein. In the context of previously reported information, these results indicate (1) that the overall pattern of SMG changes in the RA- and 6TG-resistant cells closely resembles that associated with multidrug (pleiotropic) resistance to cytotoxic agents in a variety of mammalian cells; (2) that the RA/6TG resistance-associated SMG changes are not granulocyte differentiation stage-specific; and (3) that either the RA/6TG resistance-associated SMG changes are not related to the resistance mechanism or they are involved in the resistance/cross- resistance mechanism(s) for RA/purine bases but not for DMSO.
A single common electrophoretic abnormality of glucocorticoid receptors in human leukemia cells
Authors: Distelhorst CW , Benutto BM , Griffith RC
Published: October 2019

We determined the mol wt of glucocorticoid receptors in human leukemia cells in order to detect glucocorticoid receptor defects that might cause glucocorticoid resistance. Glucocorticoid receptors in intact cells were affinity labeled with [3H]dexamethasone-21-mesylate and were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Receptors in normal human peripheral blood mononuclear cells and six human leukemia cell lines had mol wt of 97,000. Malignant cells from ten of 25 patients with leukemia contained electrophoretically abnormal glucocorticoid receptors having mol wt of 55,000 in addition to normal size receptors (Mr = 97,000). The receptor abnormality was not restricted to a particular type of leukemia and was seen in cells from both newly diagnosed patients and patients who had received prior chemotherapy, including prednisone. The abnormal receptor was not generated when cells having only normal size receptors were assayed under conditions that favor proteolysis or when cytosol from cells containing the abnormal receptor form was mixed with cytosol from cells containing only normal size receptors. The mol wt of the abnormal receptors in human leukemia cells was the same as the mol wt of receptors in mutant mouse lymphoma cell lines, S49 143R and S49 55R, which have the nuclear transfer-increased phenotype of glucocorticoid resistance. This work describes for the first time a single common electrophoretic abnormality of glucocorticoid receptors in human leukemia cells. Further investigation of glucocorticoid receptor defects in human leukemia cells could lead to an improved understanding of the mechanisms of glucocorticoid resistance in leukemia as well as a method of predicting which patients are likely to be resistant to glucocorticoid therapy.
HTLV-III/LAV antibody status of spouses and household contacts assisting in home infusion of hemophilia patients
Authors: Lawrence DN , Jason JM , Bouhasin JD , McDougal JS , Knutsen AP , Evatt BL , Joist JH
Published: October 2019

Thirty-four adult and pediatric hemophilia A and B patients and 50 nonhemophilic members belonging to 28 families were enrolled in August 1984 in a study of human T cell lymphotropic virus type III/lymphadenopathy-associated virus (HTLV-III/LAV) antibody status and T cell subpopulation numbers. All 50 household contacts, including three spouses of LAV antibody-positive adult hemophiliacs, were immunologically normal and serologically negative with respect to HTLV- III/LAV. Based on Western blot serologic testing of blood samples collected intermittently between July 1981 and August 1984 from 33 representative St Louis hemophiliacs studied during the period from 1981 to 1984, the average time since seroconversion was estimated as 20 months. One spouse of a seropositive hemophiliac and 23 parents of 27 seropositive pediatric hemophiliacs assisted regularly with home infusions. These infusion assistants have collectively experienced 44 person-years of concentrate infusion “exposure” without seroconversion. These results suggest that the likelihood for transmission of HTLV- III/LAV from hemophiliacs to persons assisting in their therapy is extremely low.
Hemophilia B with inhibitor: molecular analysis of the subtotal deletion of the factor IX gene
Authors: Hassan HJ , Leonardi A , Guerriero R , Chelucci C , Cianetti L , Ciavarella N , Ranieri P , Pilolli D , Peschle C
Published: October 2019

The structure of factor IX gene was analyzed in a hemophilia B patient with inhibitor. Genomic DNA, digested with a variety of restriction endonucleases, was hybridized with the cDNA and various genomic factor IX probes. A large subtotal deletion of the gene was observed. The borders of the deletion span from a approximately 125 nucleotide region within the last exon to an unknown domain at least 7.5 kb upstream from the first exon: it thus involves approximately 33 kb of the factor IX locus. The abnormal gene was inherited by the daughter of the propositus, who showed both the normal and the deleted allele.
Ligand state of intraerythrocytic circulating HbC crystals in homozygote CC patients
Authors: Hirsch RE , Raventos-Suarez C , Olson JA , Nagel RL
Published: October 2019

Whole blood and Stractan-Percoll fractions of blood from splenectomized patients with homozygous hemoglobin C (CC) disease were studied under aerobic and anaerobic conditions. Erythrocytes containing typical CC crystals are found in the densest fraction as documented by freeze- fracture electron microscopy. We report that the intraerythrocytic Hb C circulating crystals are in the oxygenated liganded state as demonstrated by melting upon deoxygenation and by absorption spectroscopy. Furthermore, crystals are more likely to form in cells with low concentration of Hb F. Changes of ligand state (which results in melting of the intraerythrocytic crystal) might be involved in the pathophysiology of this disease, removing the danger of vasoocclusive episodes.
DNA sequence variation associated with elevated fetal G gamma globin production
Authors: Gilman JG , Huisman TH
Published: October 2019

The percentage of G gamma chains in the Hb F of SS patients and beta- thalassemia heterozygotes is generally 40%, but some have 60% to 70% G gamma. To test the hypothesis that DNA sequence variation 158 base pairs 5′ of the G gamma gene is associated with this variation in G gamma values, DNA was analyzed using the restriction endonuclease Xmn I (gamma IVS-II probe). Xmn I recognizes the sequence from -157 to -166 only if T is at position -158. Individuals from five families had T at - 158 for G gamma genes in both chromosomes, and the mean G gamma value was 69.7% +/- 4.6% (SD). For 13 families, individuals with T at -158 for the G gamma gene of one chromosome had a G gamma value of 60.6% +/- 5.7%. With one exception, lack of T at -158 was associated with low G gamma values (39.6% +/- 4.0%). In low Hb F G gamma-beta+-HPFH, the Xmn I site was seen 5′ to both G gamma and A gamma genes, which suggests that T at -158 is associated with elevated Hb F; Pst I digestion showed that the A gamma gene T producer G gamma globin, which accounts for high levels of G gamma (87–88%). Calculations show that T at -158 is associated with a three- to 11-fold increase in production per G gamma gene, which is an order of magnitude less than that associated with the previously identified -202 C----G substitution of high Hb F G gamma- beta+-HPFH.
Fetal hemoglobin variants identified in adults through restriction endonuclease gene mapping methodology
Authors: Nakatsuji T , Burnley MS , Huisman TH
Published: October 2019

It has been found possible to detect the presence of some gamma chain abnormal fetal hemoglobins in adults through analysis of genomic DNA with selected restriction endonucleases. These variants are Hb F-Hull (A gamma 121Glu----Lys) which was observed in eight adult members of five families, Hb F-Pendergrass (A gamma 36Pro----Arg) in five adult members of one family, and Hb F-Port Royal (G gamma 125Glu----Ala) in 32 adult members of 17 families. The analyses were extended to include haplotyping, which involved 12 different restriction sites. The Hb F- Port Royal anomaly was only present on a chromosome with two G gamma genes (the 5′-G gamma-G gamma-3′ globin gene arrangement) which may have arisen through gene conversion or point mutations. It appears likely that the mutation resulting in the 125Glu----Ala substitution occurred once on a 5′-G gamma-G gamma-3′ chromosome, while additional base substitutions, gene conversions, and/or cross-over events are responsible for the association of the F-Port Royal anomaly with different chromosomes, as characterized by different haplotypes.
The acetylcholinesterase defect in paroxysmal nocturnal hemoglobinuria: evidence that the enzyme is absent from the cell membrane
Authors: Chow FL , Telen MJ , Rosse WF
Published: October 2019

Paroxysmal nocturnal hemoglobinuria (PNH) is a myelodysplastic disease characterized by erythrocytes that show abnormally increased sensitivity to complement-mediated lysis. Complement-sensitive PNH erythrocyte membranes have previously been shown to lack acetylcholinesterase (AchE) activity, but the molecular basis of this deficiency has been unclear. We have used monoclonal antibodies to four different epitopes on the AchE molecule to show that abnormal PNH erythrocytes failed to bind these antibodies. Moreover, abnormal PNH erythrocytes contained no protein immunoprecipitable by these antibodies, while normal complement-insensitive erythrocytes from PNH patients showed normal amounts of immunoprecipitable AchE which had normal electrophoretic mobility. These data suggest that abnormal PNH erythrocytes lack AchE enzyme activity due to the absence of the AchE molecule from the cell membrane.
Peripheral T cell lymphoma: immunologic and cell-kinetic observations associated with morphological progression
Authors: Winberg CD , Sheibani K , Krance R , Rappaport H
Published: October 2019

Peripheral T cell lymphomas (PTCLs) form a morphologically heterogeneous group of non-Hodgkin's lymphomas that are generally considered to have immunophenotypes associated with mature T cells, usually those of helper T cells. We now describe and correlate the clinical, morphological, immunologic, and cell-kinetic findings based on the evaluation of eight tissue samples obtained at various times from a 13-year-old girl with PTCL. The early morphological expressions of this patient's PTCL were those of diffuse mixed-cell lymphoma and focal large-cell lymphoma (LCL) evolving from the histologic picture of an atypical immune response (AIR). These morphological findings were associated with an immature T cell immunophenotype associated with cortical thymocytes--namely, sheep erythrocyte rosette (sER)+, T11+, Leu-2a+, Leu-3a+, HLA-DR+, OKT6-, OKT9+, OKT10+--and with cell-kinetic findings that showed no evidence of aneuploidy and few cells in S phase. Diffuse pleomorphic LCL developed, which was associated with further dedifferentiation of the neoplastic T cells to the immunophenotype sER-, T11+, Leu-2a-, Leu-3a-, HLA-DR+, OKT6-, OKT9+, OKT10- and with cell-kinetic findings that demonstrated a distinct aneuploid population and a dramatic increase in the percentage of cells in the S phase. The immunophenotype of the PTCL at the time of the patient's death was T11-, Leu-2a-, Leu-3a-, HLA-DR+, OKT6-, OKT9+, OKT10-, an immunophenotype indistinguishable from that of a non-B non-T cell lymphoma. The immunologic findings in this case also suggest that an AIR in some cases may represent a prelymphomatous state or may be a morphological expression of PTCL. These observations indicate that PTCLs may be characterized by rapidly changing clinical, morphological, immunologic, and cell kinetic findings which are best evaluated by multidisciplinary studies.
Effect of natural killer cells on syngeneic bone marrow: in vitro and in vivo studies demonstrating graft failure due to NK cells in an identical twin treated by bone marrow transplantation
Authors: Goss GD , Wittwer MA , Bezwoda WR , Herman J , Rabson A , Seymour L , Derman DP , Mendelow B
Published: October 2019

Bone marrow transplantation for severe idiopathic aplastic anemia was undertaken in a patient, using his monozygotic twin brother as the donor. In spite of the use of syngeneic bone marrow, failure of engraftment occurred on two occasions. In vitro studies demonstrated that natural killer (NK) cells from the recipient markedly inhibited the growth of donor bone marrow granulocyte progenitor cells. On a third attempt, successful bone marrow engraftment was achieved following high-dose cyclophosphamide, which has previously been shown to be inhibitory to NK cells. We conclude that NK cell activity may play an important role in bone marrow failure as well as being responsible for at least some cases of aplastic anemia.