bluuebunny/crossref_metadata_2025 huggingface dataset:
The gene located at chromosome 18 band q21 is rearranged in uncultured diffuse lymphomas as well as follicular lymphomas
Authors: Lee MS , Blick MB , Pathak S , Trujillo JM , Butler JJ , Katz RL , McLaughlin P , Hagemeister FB , Velasquez WS , Goodacre A
Published: October 2019

The karyotypic abnormality t(14;18)(q32;q21) is reported to occur in 75% of follicular lymphomas. This translocation results in the rearrangement of a putative oncogene bcl-2, which resides at chromosome 18 band q21 (the 18q21 gene). Using two human genomic DNA fragments cloned from the chromosome 18 band q21 as probes, we analyzed 65 uncultured human lymphoma samples by the Southern blot technique. The 18q21 gene was rearranged in 18 of 26 (69%) follicular lymphomas, 3 of 5 (60%) follicular lymphomas transformed to large cell lymphomas, 8 of 20 (40%) diffuse large cell lymphomas (DLCLs), and 2 of 7 (29%) small noncleaved cell lymphomas (SNCs). Our analysis detected rearrangement of the 18q21 gene in 10 of 13 (77%) cases in which the t(14;18)(q32;q21) translocation was found by cytogenetic techniques. Our analysis also proved helpful in difficult karyotyping situations: (a) identifying the donor chromosome fragment as chromosome 18 band q21 in 4 of 9 (44%) cases that cytogenetically displayed a 14q+ chromosome of unknown origin, and (b) identifying a rearrangement of chromosome 18 band q21 in 12 of 18 (67%) cases that cytogenetically yielded no cells in metaphase. We also demonstrated three cases of submicroscopic rearrangement of the 18q21 gene. In our studies, patients with DLCLs and rearrangement of the 18q21 gene had a significantly higher incidence of extranodal involvement when compared with patients with DLCLs and no 18q21 gene rearrangement (P = 0.03).
The c-sis gene expression in cells from a patient with acute megakaryoblastic leukemia and Down's syndrome
Authors: Sunami S , Fuse A , Simizu B , Eguchi M , Hayashi Y , Sugita K , Nakazawa S , Okimoto Y , Sato T , Nakajima H
Published: October 2019

The c-sis gene expression in leukemia cells from a patient with acute megakaryoblastic leukemia and Down's syndrome was studied. The leukemia blasts were identified as megakaryoblasts by the platelet peroxidase reaction and the reactivity against antiplatelet monoclonal antibodies. Leukemia cells obtained from peripheral blood or bone marrow specimens before and after initiation of chemotherapy were analyzed for c-sis gene expression by the RNA-DNA dot blot hybridization. Although the level varied, the mRNA of the c-sis gene was detected in all megakaryoblastic leukemia cells obtained at different clinical stages. The c-sis mRNA level in cells obtained after relapse was higher than that obtained before the initiation of therapy. The 25S c-sis mRNA was detected in a megakaryoblastic leukemia cell line established from this patient. The role of the expression of the c-sis gene in acute megakaryoblastic leukemia cells is discussed.
Characterization of a new human diploid myeloid leukemia cell line (PLB- 985) with granulocytic and monocytic differentiating capacity
Authors: Tucker KA , Lilly MB , Heck L Jr , Rado TA
Published: October 2019

A new human diploid cell line, designated PLB-985, has been established from the peripheral blood of a patient with acute nonlymphocytic leukemia (ANLL). Cells of this line are capable of granulocytic and monocytic maturation in the presence of inducing agents. By morphology, the analysis of surface antigens, and cytochemical staining PLB-985 cells are myelomonoblasts. Transmission electron microscopy reveals them to be devoid of neutrophilic primary or secondary granules and to have an open chromatin pattern with frequent nucleoli. The modal karyotype of the line is 46,XX, with no consistent marker chromosomes or recognizable translocations. Myelomonoblasts of this line form colonies in soft agar and induce tumors (chloromas) in nude mice. Growth of the cells in the presence of dimethyl sulfoxide, cis-retinoic acid, or dibutyryl cyclic adenosine monophosphate results in granulocytic maturation as determined by morphology, histochemical staining characteristics, and incorporation of 35S-methionine into the neutrophil primary granule proteinases elastase and cathepsin G. The tumor-promoting phorbol ester phorbol myristate acetate induces PLB-985 cells to differentiate as monocytes. Cells grown in the presence of this agent rapidly become adherent to plastic, display markedly increased phagocytosis of latex particles, stain positively for alpha- naphthyl acetate esterase, and lose the ability to synthesize the neutrophilic proteinases. Induction of differentiation along either pathway is accompanied by a marked decrease in myc oncogene transcription.
Productive infection by B19 parvovirus of human erythroid bone marrow cells in vitro
Authors: Ozawa K , Kurtzman G , Young N
Published: October 2019

B19 parvovirus, the cause of fifth disease and transient aplastic crisis, has been successfully propagated in suspension cultures of human erythroid bone marrow cells obtained from patients with sickle cell disease and stimulated by erythropoietin. B19 inoculation in vitro resulted in a marked decline in identifiable erythroid cells over seven to nine days of incubation. Characteristic giant early erythroid cells were seen on Wright's-Giemsa stain of infected cultures. By in situ hybridization, 30% to 40% of erythroblasts were infected at 48 hours; a similar proportion of cells showed B19 capsid protein by immunofluorescence. B19 DNA was present in erythroblasts but not in the leukocyte fraction of bone marrow. B19 replication, as determined by Southern analysis, and B19 encapsidation, as determined by sensitivity of isolated cell fractions to DNase I, were restricted to the nuclei. B19 DNA was detectable in the nuclei from infected cultures beginning at 18 hours and in the supernatant at 32 hours; B19 genome copy number was estimated at about 25,000 to 30,000/infected cell at 48 hours. Recovery of virus depended on the multiplicity of infection (moi); at low moi, approximately 200x input virus was recovered from total cultures and 50x from the culture supernatants. Virus released into the supernatant was as infectious or more infectious than virus obtained from sera of infected patients. Human erythroid bone marrow culture represents a safe in vitro system for the elucidation of the cellular and molecular biology of the pathogenic B19 parvovirus.
High serum levels of soluble interleukin 2 receptor in patients with B chronic lymphocytic leukemia
Authors: Semenzato G , Foa R , Agostini C , Zambello R , Trentin L , Vinante F , Benedetti F , Chilosi M , Pizzolo G
Published: October 2019

By using an enzyme-linked immunosorbent assay, the presence of the soluble form of the interleukin-2 receptor (sIL-2R) was evaluated in the peripheral blood of 54 patients with B cell chronic lymphocytic leukemia (B-CLL). Serum levels of sIL-2R were correlated with clinical features, relevant hematologic and immunological data, and in some cases, with in vitro functional studies. In 51 patients (94.4%), the levels of sIL-2R were increased as compared with normal age-matched controls (1,781 U/mL +/- 231 v 276 U/mL +/- 26, respectively; P less than .001). Although this increase was observed in all stages of the disease and independently of several hematologic and immunologic parameters, a trend toward lower levels of sIL-2R was documented in patients with a less-invasive disease. When the values were correlated with the functional status of the residual T cell population, it was found that patients with the lowest levels of sIL-2R showed the best mitogenic response and helper capacity. It is suggested that in B-CLL patients the high levels of serum sIL-2R, capable of binding to its ligand, may block the T cell-produced IL-2, thus contributing toward a defective physiological action by this lymphokine. In turn, this defective availability of IL-2 may play a part in the abnormal immunoregulation that is implicated in the hypogammaglobulinemia, susceptibility to infections, and incidence of second neoplasias often observed in this disease.
Pseudo-Bernard-Soulier syndrome: thrombocytopenia caused by autoantibody to platelet glycoprotein Ib
Authors: Devine DV , Currie MS , Rosse WF , Greenberg CS
Published: October 2019

The Bernard-Soulier syndrome is an inherited bleeding disorder that is due to a deficiency in platelet glycoprotein Ib. Bernard-Soulier platelets fail to agglutinate in response to ristocetin despite normal levels of factor VIII:von Willebrand factor. We report a patient who developed severe refractory thrombocytopenia postsurgically while receiving procainamide therapy. Thrombocytopenia was immune mediated since the patient's platelets bore high levels of antiplatelet antibody. Radioimmunoprecipitation studies demonstrated that the autoantibodies had specificity for platelet glycoproteins Ib and V as well as platelet HLA. The patient's plasma as well as purified immunoglobulin G completely inhibited the ristocetin-induced aggregation of normal platelets but did not inhibit adenosine diphosphate-induced aggregation. The laboratory studies revealed that this patient suffered from antibody-mediated thrombocytopenia with unusual characteristics that we have called pseudo-Bernard-Soulier syndrome.
CML patients in blast crisis have breakpoints localized to a specific region of the BCR
Authors: Schaefer-Rego K , Dudek H , Popenoe D , Arlin Z , Mears JG , Bank A , Leibowitz D
Published: October 2019

Chronic myelogenous leukemia (CML) is associated with the Philadelphia (Ph) chromosome, which results from a reciprocal translocation between chromosomes 9 and 22. This activates the abl oncogene by moving it from chromosome 9 and combining it with sequence located on chromosome 22. The new fusion gene, with chromosome 22 sequence at its 5′ end and chromosome 9-abl sequence at its 3′ end, generates a new messenger RNA (mRNA) and protein that are implicated in the pathogenesis of CML. The breakpoint near the c-abl locus on chromosome 9 can occur within a large area. In contrast, the breakpoints on chromosome 22 are concentrated within a 6 kilobase (kb) region termed the breakpoint cluster region (bcr). This study was designed to determine whether chronic-phase and blast crisis patients had identifiable differences in the structure of their Ph chromosomes. Restriction mapping of the chromosome 22 translocation breakpoints performed for 26 patients showed that the breakpoints of eight of the nine patients in blast crisis were in the 3′ portion of the bcr, whereas the breakpoints in the 17 patients in the chronic phase were clustered in the 5′ portion of the bcr. This suggests a strong correlation between a 3′ bcr breakpoint and blast crisis in CML.
Age-related deficiency of the synthesis of platelet activating factor by leukocytes from Zellweger patients
Authors: Sturk A , Schaap MC , Prins A , ten Cate JW , Govaerts LC , Wanders RJ , Heymans HS , Schutgens RB
Published: October 2019

Ca2+-ionophore A23187-induced synthesis of the alkoxyether lipid platelet activating factor (PAF) by leukocytes from Zellweger patients was undetectable in two patients studied at 3 and 4 weeks of age, reduced in a third patient studied at 2 months of age, and in the low normal range in four patients studied between 4 months and 5 years of age. We have previously reported that plasmalogen-type phosphatidylethanolamine (PE) levels of erythrocytes are reduced in Zellweger patients up to 20 weeks of age, but normal in older patients. These levels were reduced in the three patients with abnormal PAF synthesis, and normal in the other four patients. The results suggest a close relationship between the age of the patients at sampling, and both the A23187-induced capacity of leukocytes to synthesize PAF and the plasmalogen PE levels in their erythrocytes.
Increased surface expression of the membrane glycoprotein IIb/IIIa complex induced by platelet activation. Relationship to the binding of fibrinogen and platelet aggregation
Authors: Niiya K , Hodson E , Bader R , Byers-Ward V , Koziol JA , Plow EF , Ruggeri ZM
Published: October 2019

Platelet activation altered the binding of three monoclonal antibodies (monovalent Fab' fragment) directed against the glycoprotein (GP) IIb/IIIa complex. An increased binding of two- to threefold occurred after stimulation with thrombin or phorbol myristate acetate (PMA), with slight but significant increase in the dissociation constants (Kd) of two antibodies (LJ-CP8 and LJ-P9). In contrast, no statistically significant changes were observed with ADP-stimulated platelets. The increased binding of LJ-CP3, but not of the other two antibodies, to activated platelets decreased by 30% to 40% in the presence of EDTA at 22 to 25 degrees C. Platelets stimulated by thrombin or PMA bound more fibrinogen than did those stimulated by ADP, and significant differences in the extent but not in the affinity of fibrinogen binding were observed with various platelet agonists. When the pool of GP IIb/IIIa molecules exposed on the surface of unstimulated platelets was reacted with the monoclonal antibody LJ-CP3 to block ADP-induced fibrinogen binding and platelet aggregation, stimulation with thrombin or PMA still induced substantial binding of antibody and fibrinogen, and aggregation ensued. Therefore, platelets exposed to “strong” agonists exhibit an increased number of surface-oriented epitopes associated with GP IIb/IIIa. The GP IIb/IIIa molecules bearing these newly exposed epitopes are functional in that they can bind fibrinogen and mediate platelet aggregation.
An inhibitory monoclonal antibody against human tissue factor
Authors: Carson SD , Ross SE , Bach R , Guha A
Published: October 2019

We obtained a hybridoma using immune spleen cells from a mouse injected with human brain tissue factor that had been purified on a factor VII- agarose affinity column. This monoclonal IgG1, HTF1–7B8, inhibits tissue factor procoagulant activity. The concentration of HTF1–7B8 producing half-maximal inhibition is influenced by the concentration of factor VIIa, suggesting that the antibody and enzyme compete for the cofactor. The antibody was successfully used to detect both human and bovine tissue factor on nitrocellulose dot blots, indicating that the epitope recognized by this antibody is conserved in both species. This antibody clearly reveals tissue factor on a Western blot. An HTF1–7B8 affinity column was used to purify tissue factor from both human brain and placenta. The electrophoretic mobilities in polyacrylamide gels in the presence of sodium dodecyl sulfate (SDS) and the amino acid compositions of the purified tissue factor from brain and placenta are indistinguishable, as are their specific procoagulant activities in reconstituted systems. This antibody will be useful for immunopurification and characterization of tissue factor structure and mechanism.
Immunoglobulin and T cell receptor gene configuration in acute lymphoblastic leukemia of infancy
Authors: Felix CA , Reaman GH , Korsmeyer SJ , Hollis GF , Dinndorf PA , Wright JJ , Kirsch IR
Published: October 2019

We examined immunoglobulin (Ig) heavy chain, K light chain, and T cell receptor (TCR) gamma and beta gene configuration in the leukemic cells from a series of infants aged less than 1 year with acute lymphoblastic leukemia (ALL). Each of these 11 cases demonstrated leukemic cell surface antigens that have been correlated with a B cell precursor phenotype. Of the 11, lymphoblasts of 4 retained the germline configuration of both Ig and TCR loci, whereas 7 had rearranged the Ig heavy chain gene. Two of these seven showed light chain gene rearrangement. TCB beta chain rearrangement had occurred in only one of the 11 patients' tumors. No TCR gamma chain rearrangements were identified. These results are in contrast to earlier studies of B cell precursor ALL in children in which Ig heavy chain gene rearrangements were evident in every case and approximately 40% showed Ig light chain rearrangement as well. In addition, 45% of cases of B cell precursor ALL of children had rearranged their gamma TCR genes, and 20% had rearranged beta. These data suggest that ALL in infancy represents an earlier stage of B cell development than is found in B cell precursor ALL of children. ALL in the infant age group has been associated with the worst prognosis of all patients with ALL. This study suggests that the disease in infants differs not only clinically, but also at the molecular genetic level, from the disease in children.
ADP-induced platelet aggregation depends on the conformation or availability of the terminal gamma chain sequence of fibrinogen. Study of the reactivity of fibrinogen Paris 1
Authors: Denninger MH , Jandrot-Perrus M , Elion J , Bertrand O , Homandberg GA , Mosesson MW , Guillin MC
Published: October 2019

Fibrinogen Paris I contains a mutant gamma chain that is longer than the normal chain, resulting in altered fibrin polymerization and cross- linking. Because these functions involve the carboxy-terminal region of the gamma chain, we decided to determine whether fibrinogen Paris I or the isolated Paris I gamma chain supports normal ADP-induced platelet aggregation, a function that requires the ultimate 12 residues of the normal gamma chain (400 through 411). Aggregation of ADP-stimulated normal platelets was defective with fibrinogen Paris I and markedly depressed with the gamma Paris I chain. These findings prompted us to characterize the carboxy-terminal structure of the region of the gamma Paris I chain responsible for this activity. The carboxy-terminal cyanogen bromide (CNBr) peptide of the normal gamma chain (385 through 411) or that from gamma Paris I was isolated by differential adsorption to triethylene-tetramine resin or by reverse-phase high-performance liquid chromatography (HPLC). The CNBr peptide from the Paris I gamma chain was identical to that of the normal gamma chain in its retention time on HPLC, its amino acid composition, and its sequence. Thus, the primary structure of the gamma Paris I chain from residue 384 through 411 is normal, indicating that a peptide insertion has occurred upstream from residue 384, resulting in an impairment of those physiologic functions attributable to the carboxy-terminal end of the gamma chain from position 384 (ie, cross-linking, ADP-induced platelet aggregation, and at least a portion of the gamma chain polymerization site). These observations demonstrate that the gamma chain platelet recognition site in the fibrinogen molecule is necessary but not alone sufficient to support normal ADP-induced platelet aggregation. There appears to be an additional requirement for normal conformation of the gamma chain or availability of its terminal sequence during the interaction of fibrinogen with platelets.
An approximately 300 bp deletion involving part of the 5' beta-globin gene region is observed in members of a Turkish family with beta- thalassemia
Authors: Diaz-Chico JC , Yang KG , Kutlar A , Reese AL , Aksoy M , Huisman TH
Published: October 2019

Detailed gene mapping analyses of genomic DNA from two Turkish subjects with a beta-thalassemia trait demonstrated an approximately 300 bp deletion, which is located between the Rsa I restriction site 128 bp 5′ to the Cap site and the Acc I restriction site 284 bp 3′ to the same Cap site; it includes the 5′ beta promoter region, the first exon, and (part of) the IVS-I. Heterozygotes for this and two other beta- thalassemia types, which are also caused by deletions involving 5′ beta promoter sequences, appear to have higher hemoglobin (Hb) A2 levels, perhaps because the loss of this promoter results in an increased transcription of the delta globin gene, as delta and beta promoters may be influenced by the same enhancing sequences 3′ to the beta globin gene.
Immunologic reconstitution after haploidentical bone marrow transplantation for immune deficiency disorders: treatment of bone marrow cells with monoclonal antibody CT-2 and complement
Authors: Moen RC , Horowitz SD , Sondel PM , Borcherding WR , Trigg ME , Billing R , Hong R
Published: October 2019

Patients with congenital T lymphocyte deficiency disorders received transplants with parental bone marrow depleted of mature T cells by the use of an anti-T cell monoclonal antibody (CT-2) and complement. Our results with 16 consecutive patients (20 transplants) showed rapid engraftment of donor cells; cytoreduction (busulfan, cytosine arabinoside [ara-C], cyclophosphamide) was used in six transplants, and marrow ablation was used in six (ara-C, cyclophosphamide, 1,365-cGy total body irradiation). No patient received prophylactic anti-graft-v- host disease (GVHD) therapy posttransplant, and only one patient developed significant GVHD, which involved the skin, liver, and gastrointestinal tract. Seven others showed some manifestations of GVHD, but these were of minimal clinical significance and required only occasional steroid therapy. Overall, eight patients are alive and well; eight did not survive. Polyclonal immunoglobulin synthesis by donor memory B cells was seen shortly after transplantation, with peak donor- derived serum levels seen approximately 2 months after transplantation. After this initial immunoglobulin synthesis waned, another wave of B cell responses developed. This immunoglobulin response appears to be permanent. T cell functions appeared as soon as 3 weeks after transplantation. This experience in a variety of patients with combined immunodeficiency who received transplants with monoclonal antibody T cell-depleted marrow shows gratifying results with a consistent T and B cell benefit.
Induction of proliferation of B prolymphocytic leukemia cells by phorbol ester and native or recombinant interferon-gamma
Authors: Sauerwein RW , van der Meer WG , Aarden LA
Published: October 2019

Phorbol ester phorbol myristate acetate (PMA) induces proliferation in nonmalignant human B cells and B cells from a patient with B prolymphocytic leukemia (B-PLL). Mitogen-free T cell-derived conditioned medium acts synergistically with PMA in inducing proliferation of B-PLL cells but does not enhance the PMA-stimulated outgrowth of nonmalignant B cells. Interleukin 2 (IL-2) has no effect on the outgrowth of B-PLL cells, and monoclonal antibodies against the IL-2 receptor do not influence the response to PMA and conditioned medium. Recombinant interferon-gamma (IFN-gamma), in contrast, is a potent enhancer of PMA-induced proliferation of B-PLL cells. With gel filtration techniques and with the use of anti-IFN-gamma antibodies, it is shown that IFN-gamma in the conditioned medium is responsible for the observed increase in B-PLL cell proliferation. Preincubation of B- PLL cells with IFN-gamma induces responsiveness to PMA, whereas IFN- gamma alone had no effect on these cells when pretreated with PMA. The combined data show that, in the presence of PMA, native and recombinant IFN-gamma are growth factors for B cells from a B-PLL patient and that IL-2 is not involved in this process.
Inhibition of hemopoietic growth factor-induced proliferation by adenosine diphosphate-ribosylation inhibitors
Authors: Colon-Otero G , Sando JJ , Sims JL , McGrath E , Jensen DE , Quesenberry PJ
Published: October 2019

The effects of adenosine diphosphate (ADP) ribosylation inhibitors on hematopoietic growth factor-induced proliferation were examined. Significant inhibition of interleukin-3 (IL-3), colony-stimulating factor 1, and lung conditioned media-induced clonal agar growth of normal murine hematopoietic cells by 10 mmol/L nicotinamide (NAM), 10 mmol/L 3-aminobenzamide (3AB), and 5 mmol/L N1-methylnicotinamide (1MN) was noted. Nicotinic acid, a related compound that does not inhibit ADP ribosylation, failed to inhibit the growth factor-mediated proliferation. NAM (10 mmol/L), 3AB (10 mmol/L), and 1MN (5 mmol/L) also prevented IL-3 and phorbol ester-stimulated 3H-thymidine incorporation into the IL-3-responsive FDC-P1 cell line. Exposure of FDC-P1 cells to 10 mmol/L NAM led to a significant decrease in nuclear poly-(ADP-ribose) levels. Exposure of FDC-P1 cells to 5 mmol/L 1MN did not affect the interaction of the phorbol ester receptor, protein kinase-C (PK-C), with the cell membrane as determined by assay of phorbol ester binding in cytosol and membrane preparations. Nor did it affect the catalytic activity of PK-C as determined by assaying the in vitro phosphorylation of histone H1 by cytosolic kinase preparations from FDC-P1 as well as EL4 thymoma cells. 1MN markedly enhanced the inhibitory effects of phorbol esters on DNA synthesis of EL4 cells even at concentrations (1.25 mmol/L) that had no effects on DNA synthesis in the absence of phorbol esters. Our findings demonstrate that (a) active ADP ribosylation inhibitors interfere with growth factor-induced proliferation of murine hematopoietic cells and (b) the inhibition occurs at a step that follows the activation and translocation of PK-C and is more closely linked to DNA synthesis.
Exposure to gamma-irradiation increases phorbol myristate acetate- induced H2O2 production in human macrophages
Authors: Gallin EK , Green SW
Published: October 2019

Cell number, protein, and phorbol myristate acetate (PMA)-induced H2O2 production were measured in cultured human peripheral blood monocytes for six days after exposure to varying doses of gamma-radiation. Both the number of adherent cells and the protein per dish decreased with increasing radiation doses. The dose of radiation decreasing the number of adherent cells by 37% on days 4 and 6 postirradiation was 29 Gy. Four hours postirradiation there was a small decrease in PMA-induced H2O2 production for doses of 7.5 Gy or greater; levels returned to normal by eight hours and increased at 24 hours postirradiation. By day 4 postirradiation significant increases in PMA-induced H2O2 production were noted at all radiation doses (2.5 to 50 Gy). This increase was not due to a shift in the PMA dose-response curve, a change in the time course of the PMA response, or an effect of decreased cell density on the assay system. Superoxide levels were not significantly changed in cells exposed to 20 Gy. Catalase, glutathione peroxidase, and superoxide dismutase levels also were unchanged. Culturing irradiated cells with gamma-interferon increased PMA-induced H2O2 release, which indicated that irradiated cells retained their capacity to respond to gamma-interferon. These data demonstrate that irradiation affects the PMA-induced H2O2 production of human monocytes in a time- and dose- dependent manner. An increase in the release of reactive oxygen intermediates by the macrophage may play a role in enhancing the deleterious effects of radiation in vivo.
1986 update of HIV seroprevalence, seroconversion, AIDS incidence, and immunologic correlates of HIV infection in patients with hemophilia A and B
Authors: Ragni MV , Winkelstein A , Kingsley L , Spero JA , Lewis JH
Published: October 2019

A cohort of 181 patients with hemophilia A (149) and hemophilia B (32) cared for at the Hemophilia Center of Western Pennsylvania was followed to determine human immunodeficiency virus (HIV) seroprevalence, seroconversion rate, and clinical and immunologic correlates of HIV infection. By December 1986, 82 (45%) were HIV seropositive, and of these, ten (12%) had developed AIDS, 28 (34%) had symptomatic HIV infection (CDC class III, IV), of whom 14 (17%) had AIDS-related complex (ARC), and 44 (54%) had asymptomatic HIV infection (CDC class II). The HIV seropositive group included 82% of those treated with factor VIII concentrate (97% severe, 5% moderate), 48% of those treated with factor IX concentrate (92% severe, 8% moderate), 10% of those treated with cryoprecipitate (67% severe, 33% moderate), and none of those treated with fresh frozen plasma. Based on 77 serially sampled HIV seropositive hemophiliacs (1977 to 1986), peak seroconversion occurred in 1982, with 14% (11 of 77) occurring since 1984. With increasing time from seroconversion, both T4 lymphocyte number and function (the latter measured by growth in soft agar [T colony assay]) progressively declined; T4 number declined to 135 +/- 26/mm3 (SEM), and colony count declined 1193 +/- 537 (control 3851 +/- 387) by 5 years after seroconversion. In those developing AIDS, total T4 fell below 100/mm3 (33 +/- 8/mm3) at diagnosis. In this cohort, the overall AIDS incidence is 5.5% (12% among the HIV seropositive) and in those seropositive 5 or more years, the AIDS incidence approaches 32%.
A delta-globin gene derived from patients with homozygous delta zero- thalassemia functions normally on transient expression in heterologous cells
Authors: Nakamura T , Takihara Y , Ohta Y , Fujita S , Takagi Y , Fukumaki Y
Published: October 2019

Three Japanese individuals with homozygous delta zero-thalassemia from different families were the subjects of molecular genetic analysis. They were homozygous for seven polymorphic sites in the beta-globin gene cluster. Nucleotide sequence analysis of the delta-globin gene cloned from each patient revealed a single nucleotide substitution (T- C) 77 base pairs 5′ to the cap site, just upstream of the CCAAC box of the delta-globin gene. When introduced into COS cells, the gene was expressed at normal levels with proper processing of RNA. These results suggest that the complete suppression of delta-globin chain synthesis in these patients is not due to a defective promoter, a defective RNA processing or a chain terminator mutation, but rather to impaired regulation of gene expression specific to erythroid cells. The region around the CCAAC box may have a significant role in expression of the delta-globin gene in erythroid cells.
Studies with a monoclonal antibody against activated platelets: evidence that a secreted 53,000-molecular weight lysosome-like granule protein is exposed on the surface of activated platelets in the circulation
Authors: Nieuwenhuis HK , van Oosterhout JJ , Rozemuller E , van Iwaarden F , Sixma JJ
Published: October 2019

To define the role of activated platelets we have attempted to prepare monoclonal antibodies specific for activated platelets. The IgG2b antibody of one of the clones, designated 2.28, was studied in more detail. Native platelets from normal individuals bound 650 125I-2.28 molecules/platelet, whereas thrombin-activated platelets bound 12,600 molecules/platelet with high affinity (4.6 nmol/L). Immunoelectrophoretic analysis revealed that 2.28 reacted with a 53,000- mol wt protein. Immunocytochemistry showed that the antigen is located in a special subclass of platelet granules in unstimulated platelets and is exposed on the surface of thrombin-activated platelets. Double- labeling studies with immunogold labels disclosed simultaneous localization of 2.28 binding sites and cathepsin D in the same granules both in megakaryocytes and endothelial cells, thereby indicating that the antigen may be localized in lysosomes. By using flow cytofluorometry, in vivo platelet activation was studied in patients undergoing cardiac surgery with cardiopulmonary bypass. Increased numbers of platelets that expressed the 2.28 antigen on their surface were observed after extracorporeal perfusion. The percentage of 2.28- positive platelets in the circulation was 3.9% +/- 2.7% (SD) in controls (n = 20), 5.5% +/- 3.0% in patients (n = 10) before cardiopulmonary bypass surgery, 24.6% +/- 13.5% after the bypass, and 8.5% in two patients with acute deep venous thrombosis. These data indicate that 2.28 may serve as a useful probe of in vitro and in vivo platelet activation.
A monoclonal anti-human plasma prekallikrein antibody that inhibits activation of prekallikrein by factor XIIa on a surface
Authors: Veloso D , Silver LD , Hahn S , Colman RW
Published: October 2019

Of five IgGI/k murine monoclonal anti-human prekallikrein antibodies produced (MAbs), MAb 13G11 was selected for studying interaction of prekallikrein with factor XII and high-mol-wt kininogen (HMWK) during activation on a surface. Immunoblots from sodium dodecyl sulfate (SDS) gels showed that this MAb recognizes two variants (88 kd and 85 kd) of prekallikrein and kallikrein both in purified proteins and normal plasma. Under reducing conditions, kallikrein exhibits the epitope on the heavy chain but not on the light chains. Preincubation of MAb 13G11 with prekallikrein (added to prekallikrein-deficient plasma) or with normal plasma inhibited surface activation of prekallikrein 60% to 80%, as judged by amidolytic and coagulant assays. In normal plasma, inhibition by the Fab fragments was 87% of that with the entire MAb. Inhibition was not by competition between the MAb and HMWK, since neither binding of 13G11 to prekallikrein (coated on microtiter plates) was inhibited by an excess of HMWK, nor was hydrolysis of HMWK by kallikrein inhibited by 13G11. Using purified proteins in a system mimicking contact activation, inhibition by 13G11 of prekallikrein activation by factor XIIa, HMWK, and kaolin present was approximately 80%. Decreased inhibition (55% to 25%) occurred without HMWK or when kallikrein was used instead of prekallikrein. Kallikrein activity was not inhibited by 13G11 Fab fragments. These results indicate that the effect of 13G11 in plasma was neither dissociation of prekallikrein- HMWK complex nor a direct effect on kallikrein activity. Similar to the results in plasma, activation of prekallikrein, HMWK present, by factor XIIa bound to kaolin, was inhibited approximately 70% by 13G11. The results suggest a previously unrecognized site on the prekallikrein (heavy chain) required for its interaction with factor XIIa, either shared with the 13G11 epitope or located in very close proximity. The inhibition of kallikrein by intact 13G11 indicates that its binding site on the heavy chain is sterically related to the active site (light chain).
Establishment of an interleukin 2-dependent human T cell line from a patient with T cell chronic lymphocytic leukemia who is not infected with human T cell leukemia/lymphoma virus
Authors: Hori T , Uchiyama T , Tsudo M , Umadome H , Ohno H , Fukuhara S , Kita K , Uchino H
Published: October 2019

We established an interleukin 2 (IL-2)-dependent human T cell line, Kit 225, from a patient with T cell chronic lymphocytic leukemia (T-CLL) with OKT3+, -T4+, -T8- phenotype. Southern blot analysis showed that Kit 225 is not infected with human T cell leukemia/lymphoma virus (HTLV) type I or II, and is probably derived from the major clone in the fresh leukemic cells. Kit 225 cells express a large amount of IL 2 receptors constitutively and their growth is absolutely dependent on IL 2. No other stimuli, such as lectins or antigens, are required for maintaining the responsiveness to IL 2. As abnormal IL 2 receptor expression was also seen originally in the fresh leukemic cells, the establishment of this cell line with IL 2 suggests that IL 2-mediated T cell proliferation is involved in the leukemogenesis of some cases of HTLV-negative T-CLL.
Bone marrow matrix modulation of HL-60 phenotype
Authors: Luikart SD , Sackrison JL , Maniglia CA
Published: October 2019

The initiation and maintenance of cellular differentiation for a variety of cell types has been shown to be influenced by the microenvironment. To investigate the influence of bone marrow stroma on leukemic cell differentiation, HL-60 human promyelocytic leukemia cells were grown in the presence of Triton-treated extracellular matrix derived from normal human bone marrow stromal cells. This bone marrow matrix microenvironment had a dramatic impact on the phenotypic expression of this malignant line. HL-60 cellular proliferation, morphology, nonspecific esterase activity, formation of Fc rosettes, and sensitivity to induction by 12-O-tetradecanoyl-phorbol-13-acetate (TPA) were all influenced by the presence of matrix molecules. In contrast, stromal cell-conditioned media did not alter HL-60 phenotype. Thus, HL-60 cells appear to retain responsiveness to a human bone marrow stromal cell-derived matrix despite their ability to grow autonomously. Studies of the interaction of leukemic cells and marrow stroma in vitro may provide important information concerning the regulation of leukemic cell behavior.
Immunohistochemical characterization of a 183 KD myeloid-specific-DNA- binding protein in B5 fixed, paraffin-embedded tissues, and bone marrow aspirates by monoclonal antibody BM-1
Authors: Epstein AL , Samoszuk M , Stathopoulos E , Naeve GS , Clevenger CV , Weil S , Marder RJ
Published: October 2019

A monoclonal antibody, designated BM-1, which is reactive in B5 formalin-fixed, paraffin-embedded tissues, has been generated against a cytoplasmic and nuclear antigen expressed in human myeloid precursor cells and derived leukemias. Using the avidin-biotin-complex immunoperoxidase procedure, BM-1 was found to stain selectively myeloid precursor cells in normal bone marrow and mature granulocytes in the blood. In a screen of 26 normal adult and fetal human organs fixed in B5 formalin, BM-1 was negative in all nonhematopoietic tissues with the exception of tissue granulocytes and scattered cells in the peripheral cortex of the thymus. Likewise a screen of 30 solid tumor cell lines including a spectrum of carcinomas, sarcomas, and neural-derived tumors was negative. BM-1 was also negative with 21 T and B cell lymphomas and 11 Hodgkin's disease tumors. A preliminary study of tumors of the hematopoietic system revealed that BM-1 was reactive with M2 and M3 acute myelogenous leukemias (AML), chronic myelogenous leukemias (CML) and myelomonocytic leukemias, and granulocytic sarcomas. M1, M4, M5, and M6 AML clot preparations were negative in this study, indicating that BM-1 may have a role in the histopathologic diagnosis of myelogenous leukemia. Myeloid leukemic cell lines HL-60, ML-2, KG1, and TPH-1-O showed BM-1 nuclear and/or cytoplasmic reactivity in a subpopulation of cells, but erythroid and lymphoid leukemias and all lymphoma cell lines were negative. Immunoperoxidase studies of a panel of fetal tissues showed BM-1 positive cells in the peripheral cortex of the thymus and portal myelopoietic regions of the liver at 18 weeks gestation. Finally, DNA-cellulose and solid phase radioimmunoassay (RIA) techniques developed in our laboratory demonstrate that the BM-1 antigenic domain is reactive only after binding to eukaryotic but not prokaryotic single- or double-stranded DNA. Immunoblot techniques using a DNA-cellulose purified protein sample revealed that BM-1 recognizes a 183 kD protein. These studies indicate that BM-1 is recognizing a myeloid-specific antigen that, because of its DNA binding characteristics, may have an important role in the differentiation of myeloid cells at the molecular level.
Posttranslational processing of a human myeloid lysosomal protein, myeloperoxidase
Authors: Nauseef WM
Published: October 2019

Myeloperoxidase (MPO) is a lysosomal enzyme present in the azurophilic granules of human neutrophils and monocytes and is important for optimal oxygen-dependent killing of microorganisms. The native molecule is a heterodimer composed of a pair of heavy-light protomers, each containing a 59-kDa and 13.5-kDa subunit. The intracellular processing during biosynthesis of MPO was examined in the human promyelocytic cell line HL-60. Endoglycosidase H and F digestion of immunoprecipitated pro- MPO demonstrated the presence of five N-linked--high-mannose oligosaccharide side chains and no complex mannose units. Incorporation of the threonine analogue beta-hydroxynorvaline produced species approximately 2.5 kDa and approximately 5 kDa smaller than the fully glycosylated pro-MPO, suggesting that two of the glycans were in the asparagine-X-threonine tripeptide sequence. Processing of pro-MPO occurred very rapidly, within approximately five minutes, and was best identified using glucosidase inhibitors. The presence of such inhibitors resulted in synthesis of a 92-kDa glycoprotein rather than the usually identified 89-kDa peptide. Swainsonine, a Golgi mannosidase inhibitor, did not alter the size of the earliest synthesized protein, suggesting that pro-MPO exited the endoplasmic reticulum or cis-Golgi proximal to the site of mannosidases. Intracellular transport and proteolytic maturation of MPO was retarded by weak bases (NH4Cl, chloroquine) or monensin at concentrations shown to raise intralysosomal pH. However, these agents did not qualitatively alter transport nor increase secretion. Thus, although MPO biosynthesis resembled that of other lysosomal enzymes, significant differences exist, including only limited oligosaccharide processing and intracellular transport and proteolytic maturation of pro-MPO that was only retarded by alkalinizing lysosomes without affecting the products or the fraction of pro-MPO secreted. Characterization of the determinants for targeting and of the regulatory factors in processing lysosomal enzymes in myeloid cells will provide insight into the molecular mechanisms underlying common disorders such as myeloperoxidase deficiency.
Relationship between T200 antigen expression and stages of B cell differentiation in resurgent hyperplasia of bone marrow
Authors: Caldwell CW , Patterson WP
Published: October 2019

Using monoclonal antibodies (MoAbs) and dual-parameter flow cytometric techniques, bone marrow mononuclear cells (MMC) from patients with resurgent hyperplasia were analyzed for their coexpression of HLe-1 (T200) and antigens normally associated with particular stages of B cell differentiation. The marrow from those with resurgent hyperplasia contained increased numbers of B cell precursors in multiple stages of differentiation compared to controls, thus providing a useful model system for studies of B cell differentiation. These studies indicate that the quantitative expression of T200 is differentiation-related on normal and malignant B cells and B cell precursors. Immature cells express low amounts of T200, while increasing levels of maturity correlated with increasing amounts of the antigen. This study increases the understanding of relationships between B cell surface antigens and T200 and further demonstrates that B cell hyperplasia occurs commonly in association with bone marrow reactive or resurgent processes. The quantitative, rather than only the qualitative, expression of T200 is therefore a useful marker of B cell differentiation in reactive hyperplasia and in further investigation of B cell malignancy.
Effect of recombinant interferons alpha and gamma on human bone marrow- derived megakaryocytic progenitor cells
Authors: Ganser A , Carlo-Stella C , Greher J , Volkers B , Hoelzer D
Published: October 2019

Interferons (IFNs) have been shown to suppress the proliferation of human pluripotent hematopoietic progenitor cells, CFU-GEMM, and committed erythroid (BFU-E, CFU-E) and granulocyte-macrophage (CFU-GM) progenitor cells. However, no information is yet available concerning the effect of IFNs on human megakaryocytic progenitor cells CFU-Mk. Furthermore the mechanisms underlying the inhibitory activity of IFNs are still controversial. Therefore highly purified recombinant IFN preparations, rIFN-alpha and rIFN-gamma, were assessed for their influence on in vitro growth of human bone marrow-derived CFU-Mk as well as CFU-GEMM. In addition, the role of hematopoietic accessory cells, that is, adherent cells and T lymphocytes, in the mediation of the suppressive effect of rIFNs was examined. When added to unseparated bone marrow cells, both rIFN preparations significantly inhibited colony formation with 50% inhibition of CFU-Mk occurring at 22 U/mL for rIFN-alpha and 59 U/mL for rIFN-gamma, while 50% inhibition of CFU-GEMM occurred at 59 U/mL for rIFN-alpha and 101 U/mL for rIFN-gamma. The suppressive effect of rIFN-alpha and rIFN-gamma was selectively abolished by monoclonal antibodies (MoAbs) against rIFN-alpha and rIFN- gamma, thus confirming that the inhibitory activity was due to the rIFN preparations used. The antiproliferative effect of rIFN-alpha and rIFN- gamma on CFU-GEMM growth was not associated with a decrease in the percentage of mixed colonies containing megakaryocytic cells as assessed by use of the MoAb C17.28 against platelet glycoprotein IIIa. Removal of adherent cells and T lymphocytes from the target bone marrow cells had no influence on the suppressive effect of rIFN-alpha, whereas it significantly reduced the inhibitory effect of rIFN-gamma on the growth of megakaryocytic colonies and the other hematopoietic progenitors. The data indicate that (1) human megakaryocytopoiesis is markedly inhibited by rIFN-alpha and rIFN-gamma, and (2) the inhibitory effect of rIFN-alpha is due to a direct action on hematopoietic progenitor cells, whereas the effect of rIFN-gamma is mediated to a significant degree through accessory cell populations.
Chloramphenicol-induced bone marrow injury: possible role of bacterial metabolites of chloramphenicol
Authors: Jimenez JJ , Arimura GK , Abou-Khalil WH , Isildar M , Yunis AA
Published: October 2019

To explore the potential role of some bacterial metabolites of chloramphenicol (CAP) in CAP-induced hematotoxicity, we examined their cytotoxic effects on bone marrow cells in vitro using a number of cytotoxicity parameters. Among the metabolites tested, dehydro-CAP (DHCAP) and p-nitrophenyl-2-amino-3 hydroxypropanone-HCI (NPAP) were more toxic than CAP. DHCAP was at least as toxic as nitroso-CAP. At concentrations of less than or equal to 10(-4) mol/L, DHCAP caused total irreversible inhibition of myeloid colony (CFU-GM) growth and 80% inhibition of DNA synthesis in human bone marrow. Incubation of human bone marrow cells with 10(-4) mol/L nitroso-CAP or DHCAP for 24 hours resulted in 75% and 65% cell death respectively. Although DHCAP was 10- to 20-fold more cytotoxic than CAP, it was only one third as effective in inhibiting mitochondrial protein synthesis, indicating that DHCAP exerts its toxic effect by alternate mechanisms. The cytotoxicity of DHCAP and its relative stability, compared to the unstable nitroso CAP, suggest that this bacterial metabolite of CAP, and possibly others, may play a significant role in CAP-induced hematotoxicity.
Human leukemic B cell activation: functional consequence of membrane IgM interaction with anti-IgM ligand is an alterable cell characteristic
Authors: Mongini P , Blessinger C , Seremetis S , Winchester R , Rudich S
Published: October 2019

A functional study of several human malignant B cell populations has indicated that occasional leukemic clones are extraordinarily sensitive to signal transduction through membrane IgM. One isolated hairy cell leukemia (HCL) with low background DNA synthesis was stimulated to significant levels of DNA synthesis when cultured with high (100 micrograms/mL) concentrations of soluble anti-IgM ligands. In contrast to the activation of normal peripheral blood polyclonal B cells, this DNA synthesis was completely independent of accessory T cell factors. Although the HCL clone could also be induced to enter S phase by incubation in media supplemented with only activated T cell supernatant, culture of the clone with activated T cell supernatant plus anti-IgM Ab resulted in DNA synthesis that was significantly less than that induced by either activator alone. Factor(s) in T cell supernatant appear to modulate the leukemic clone so that the binding of ligand to membrane IgM is perceived as an inhibitory rather than a stimulatory signal for DNA synthesis. In terms of Ig Fc independence and low ligand dose requirements, anti-IgM-mediated inhibitory signal transduction in the T cell supernatant-activated HCL clone was found to mimic anti-IgM mediated suppression of the spontaneous DNA synthesis of an alternative HCL clone. The functional results suggest that the type of signal transduced anti-Ig ligands may reflect differences in the activation state of receptive leukemic B cells.