bluuebunny/crossref_metadata_2025 huggingface dataset:
Correlation between the stimulation of human neutrophil function by monoclonal antibody and by colony-stimulating factor
Authors: Vadas MA , Clarke C , Nicola NA , Lopez AF
Published: October 2019

Purified human neutrophils from 48 individuals were tested for their capacity to kill antibody-coated target cells in vitro in the absence or presence of stimulating agents. The agents used to stimulate cytotoxic capacity were the monoclonal antibody (MAb) WEM-G1, colony- stimulating factor (CSF-alpha), or mononuclear cell supernatant (MNC- SN). There existed an heterogeneity among the neutrophils of different individuals in the capacity to kill target cells both in the unstimulated (“resting”) or the stimulated state. A positive correlation was found between the ability of neutrophils to kill in the “resting” state and their capacity to be stimulated by MAb WEM-G1, CSF- alpha, or MNC-SN. Furthermore, a strong positive correlation in the ability of neutrophils to be stimulated by the MAb WEM-G1 and either CSF-alpha (r = .76) or MNC-SN (r = .68), as well as between CSF-alpha and MNC-SN (r = .79) was demonstrated. No correlation was seen, however, between stimulation of neutrophil function in vitro and total blood leukocyte counts, neutrophil counts, monocyte counts, or intensity of binding of MAb WEM-G1. The observation that neutrophils respond to a similar extent to different types of stimulators, -such as cytokines (CSF-alpha and MNC-SN) and MAb, suggests that these two factors may be operating through a common mechanism and the degree of stimulation may reflect an intrinsic responsiveness of neutrophils that differs among individuals. Our results also suggest a potential clinical use of WEM-G1 in measuring neutrophil functional capacity in vitro and predicting the capacity to respond to CSF-like cytokines.
Human platelet fibrinogen: purification and hemostatic properties
Authors: Kunicki TJ , Newman PJ , Amrani DL , Mosesson MW
Published: October 2019

Conditions were developed in which 80% to 90% of platelet fibrinogen could be routinely purified in nondegraded form from the fluid phase of platelet suspensions stimulated with the calcium ionophore, A23187, in the presence of calcium, leupeptin, and prostaglandin E1. Fibrinogen was separated from other released proteins by chromatography on diethylaminoethanol (DEAE)-cellulose using a continuous pH and ionic strength gradient. Purified platelet fibrinogen, greater than 98% homogeneous by immunoelectrophoresis and sodium-dodecyl sulfate- polyacrylamide gel electrophoresis (SDS-PAGE), consisted of intact A alpha, B beta and gamma A chains, but not gamma' chains, and was 95% to 96% clottable. Platelet fibrinogen was shown to compete for the binding of radiolabeled plasma fibrinogen to ADP-activated platelets in a manner identical to that of unlabeled plasma fibrinogen itself. Also, at equivalent protein concentrations, platelet and plasma fibrinogens supported platelet aggregation to an equivalent extent. Based upon these results, we conclude that there is no significant difference between platelet and plasma fibrinogen with respect to their size, their clottability, their affinity for the activated platelet fibrinogen receptor, or their capacity to support subsequent platelet aggregation.
Differential protection of normal and malignant human myeloid progenitors (CFU-GM) from Ara-C toxicity using cycloheximide
Authors: Slapak CA , Fine RL , Richman CM
Published: October 2019

Cycloheximide, a reversible protein synthesis inhibitor, is thought to block DNA replication in normal cells by preventing synthesis of a labile protein. In animal systems, cycloheximide protects normal cells from cytotoxic S-phase specific agents, such as cytosine arabinoside (Ara-C). Malignant cells appear not to be susceptible to cycloheximide- induced cycle arrest and, subsequently, may not be protected from Ara-C cytotoxicity. The effect of cycloheximide on granulocyte/macrophage progenitors (CFU-GM) after in vitro Ara-C exposure was examined using normal human bone marrow, malignant progenitors from patients with chronic myelogenous leukemia (CML), and clonogenic cells from the human acute nonlymphocytic leukemia cell lines HL-60 and KG-1. Mononuclear or clonogenic cells were incubated for one hour with cycloheximide, followed by the addition, for three or 17 hours, of Ara-C before being plated in a methylcellulose culture system. CFU-GM survival was significantly increase if normal cells were treated with cycloheximide before Ara-C exposure. Similar cycloheximide pretreatment of CML progenitors and clonogenic HL-60 and KG-1 cells failed to protect CFU- GM from Ara-C-induced cytotoxicity.
Differences in T cell subsets between men and women with idiopathic thrombocytopenic purpura
Authors: Mylvaganam R , Ahn YS , Harrington WJ , Kim CI , Gratzner HG
Published: October 2019

Idiopathic thrombocytopenic purpura (ITP) is an autoimmune disorder, occurring predominantly in women. We studied by flow cytofluorimetry the T cell subsets in men and women with ITP and compared them with healthy sex-matched volunteers. In healthy controls, women were found to have higher proportions of T helper/inducer (Th/i) and lower T suppressor/cytotoxic (Ts/c) lymphocytes and consequently higher Th/i:Ts/c ratios than men. Accordingly, in clinical surveys, patients and controls should be matched for sex for proper comparisons. In patients with ITP in its active phase, an imbalance in T cell subsets was found in both sexes. The perturbation was more severe in women who had a marked decrease in number and proportion of Th/i lymphocytes and an increase in the proportion of Ts/c lymphocytes, whereas in men only, the proportion of Th/i lymphocytes was decreased. When patients with active disease were compared to those with ITP in remission, the decrease in Th/i subsets still persisted in both sexes but the Ts/c subset in women had returned to normal proportions. Therefore, the immune imbalance in ITP is more marked in women than men; imbalances in both Th/i and Ts/c are present in women while Ts/c appears not to be involved in men.
A longitudinal immunologic evaluation of hemophiliac patients
Authors: deShazo RD , Daul CB , Andes WA , Bozelka BE
Published: October 2019

Over an average span of one year, we performed a prospective clinical and immunologic evaluation of 30 patients with hemophilia. No patient developed life-threatening opportunistic infection or malignancy; however, the immunologic abnormalities and lymphadenopathy initially present in nine patients (lymphadenopathy group) persisted. In addition, five patients, representing 24% of the initial group without lymphadenopathy, developed generalized lymphadenopathy (converter group). One episode of idiopathic thrombocytopenia (ITP) and one episode of staphylococcal sepsis occurred in this “converter” group; one episode of ITP also occurred in the lymphadenopathy group. Sixteen patients remained asymptomatic. At the time of the follow-up evaluation, those differences in mononuclear cell (MNC) percentages and numbers noted initially among the three hemophiliac groups were no longer present. Natural killer cell function alone or in the presence of biologic response modifiers was not different among hemophiliac and control groups. Before developing lymphadenopathy, the converter group of patients had significantly better lymphocyte mitogenic function than did the other two groups of patients with hemophilia. However, lymphocyte mitogenic responses of all groups of patients with hemophilia significantly deteriorated over the course of the study. The abnormal mitogenic responses noted in these patients was explained in part by higher levels of spontaneous suppressor cell activity in mononuclear cell preparations from patients with hemophilia. We conclude that long-term immunologic studies of this patient population requires both quantitative and qualitative evaluations. Our data show that patients with hemophilia have progressive dysfunction of cell- mediated immunity.
Acute mixed lineage leukemia: clinicopathologic correlations and prognostic significance
Authors: Mirro J , Zipf TF , Pui CH , Kitchingman G , Williams D , Melvin S , Murphy SB , Stass S
Published: October 2019

The frequency and clinical significance of acute leukemia displaying both lymphoid and myeloid characteristics was determined in 123 consecutive children using a panel of lineage-associated markers. The leukemic blasts from 18 of 95 children (19%) with the diagnosis of acute lymphoblastic leukemia (ALL) by standard diagnostic criteria expressed myeloid-associated cell surface antigens. Despite immunological evidence of lymphoid differentiation (17 CALLA + and one T cell-associated antigen +) and findings of immunoglobulin gene rearrangement, blasts from these patients reacted with one to five monoclonal antibodies identifying myeloid-associated cell surface antigens (My-1, MCS.2, Mo1, SJ-D1, or 5F1). Dual staining with microsphere-conjugated antibodies and analysis by flow cytometry confirmed that some blasts were simultaneously expressing lymphoid- and myeloid-associated antigens. Conversely, blasts from seven of 28 patients (25%) with acute nonlymphocytic leukemia (ANLL), diagnosed by otherwise standard morphological and cytochemical criteria, expressed lymphoid-associated surface antigens. Dual staining of individual blasts demonstrated simultaneous expression of myeloperoxidase (MPO) (including Auer rods) in association with either T-11, CALLA, or terminal deoxynucleotidyl transferase. Blasts from one patient with ANLL demonstrated T cell receptor gene rearrangement, while blasts from another patient demonstrated characteristics associated with T (T-11), B (CALLA and heavy-chain immunoglobulin gene rearrangement), and myeloid (MPO) lineage. There were no consistent cytogenetic abnormalities, and no patient demonstrated independent leukemic clones. Each patient with typical ALL, except for myeloid-associated antigens, achieved complete remission with conventional induction therapy for ALL. By contrast, three of the seven children with ANLL whose blasts expressed the T-11 surface antigen failed ANLL induction therapy. These three patients subsequently achieved remission with ALL therapy.
The genotype and phenotype of T cell and non-T, non-B acute lymphoblastic leukemia
Authors: Aisenberg AC , Wilkes BM
Published: October 2019

We used Southern blot analysis to investigate the T cell receptor and immunoglobulin (Ig) genes in tumor DNA derived from 13 patients with acute lymphoblastic leukemia (ALL). Rearrangement of both alleles of the T cell receptor beta chain gene was demonstrated in the three individuals with T cell ALL, while the Ig genes remained in germline configuration. In contrast, in nine of ten patients with non-T, non-B ALL (seven common ALL antigen [CALLA]-positive), Ig genes were rearranged while the T cell receptor genes were intact: the noteworthy exception was a CALLA-positive individual in whom one allele of the T cell receptor was rearranged and in whom a minor rearranged Ig band was detected as well. This exception indicates that CALLA-positive non-T, non-B ALL includes proliferations that have undergone an initial genetic step toward T cell differentiation. With probes now available for both the Ig and T cell receptor genes, analysis of genomic tumor DNA is useful in all variants of ALL.
Hepatitis B virus DNA is enriched in polymorphonuclear leukocytes
Authors: Hoar DI , Bowen T , Matheson D , Poon MC
Published: October 2019

DNA hybridization and cell separation techniques were used to determine which blood components contained hepatitis B viral DNA sequences. Free monomer-length hepatitis B virus was found in large amounts in the polymorphonuclear leukocyte cell fraction in two of five HBsAG-positive patients. In these two patients, viral DNA sequences were not detected in the plasma or platelet fraction, whereas the mononuclear cell DNA contained small amounts of a 7.2 kb size unintegrated hepatitis B genome. These studies indicate that the major reservoir of unit-length viral DNA in the asymptomatic hepatitis B carriers studied here was in the polymorphonuclear leukocyte fraction. The basis for the presence of the viral DNA within these cells is presently unknown, but may relate to viral replication within, or phagocytosis of virus by, these cells.
Normal cycling patterns of hematopoietic stem cell subpopulations: an assay using long-term in vivo BrdU infusion
Authors: Pietrzyk ME , Priestley GV , Wolf NS
Published: October 2019

It was found in a long-term bromodeoxyuridine (BrdU) infusion study that two or more different subpopulations of bone marrow stem cells exist in mice. One of these subpopulations appears to be noncycling and forms approximately 10% of eight-day CFU-S. Another one, a subpopulation of slowly cycling bone marrow cells, is represented as 14- day CFU-S. The 14-day CFU-S have a regular increment in the percentage of the subpopulation entering the cycle over time, with a cell generation half-time of 21 days. The cycling status in these experiments was ascertained by in vivo continuous long-term BrdU infusion. An improved method is presented for long-term BrdU infusion with UV killing of cycled cells.
Clinical and biologic features predict a poor prognosis in acute lymphoid leukemias in infants: a Pediatric Oncology Group Study
Authors: Crist W , Pullen J , Boyett J , Falletta J , van Eys J , Borowitz M , Jackson J , Dowell B , Frankel L , Quddus F
Published: October 2019

Analysis of remission induction rates for 1,117 children 18 months to 10 years of age (group 1) and 90 infants less than 18 months of age (group 2) with acute lymphoid leukemia (ALL) and of duration of continuous complete remission (CCR) for 454 in group 1 and 33 in group 2 revealed that infants fared significantly worse in both measures of outcome (P = .03 and P less than .0001). To examine potential reasons for the poor prognosis of affected infants, clinical and biologic features of their ALL were compared. Infants had higher WBC counts (P less than .001), a higher incidence of massive splenomegaly (P less than .001), massive hepatomegaly (P less than .001), more central nervous system (CNS) disease at diagnosis (P less than .01), and lower platelet counts (P less than .001). Also, their blasts were less often PAS+ (P = .02). The incidence of non(T, B, pre-B), T and pre-B immunophenotypes of ALL did not differ significantly between the two groups. However, in patients with non(T, B, pre-B) ALL, the majority (51%) of infants had common ALL antigen (CALLA)-negative blasts, as compared with only 7% in group 1 (P less than .001). Furthermore, infants with non(T, B, pre-B) cell ALL who were less than 12 months of age were almost always CALLA- (18 of 21). The blasts of children from both groups usually expressed Ia-like antigens. These data illustrate that infants with ALL have extensive and bulky disease more often than do older children and are more often affected with a prognostically unfavorable phenotype of acute leukemia (AL) which expresses Ia-like antigens but is more often PAS- and CALLA-. We believe that these clinical and biological differences predict and explain in part the observed poor response to treatment of infants with ALL.
Modulation of human neutrophil effector functions by monoclonal antibodies against surface membrane molecules of 94,000 and 180,000 molecular weight
Authors: King CH , Peck CA , Haimes CS , Kazura JW , Spagnuolo PJ , Sawyer JA , Olds GR , Mahmoud AA
Published: October 2019

Function-related antigens on the neutrophil (PMN) surface were identified using two newly developed PMN-specific mouse monoclonal antibodies. These IgG antibodies, designated Ab 1–14 and Ab 1–15, were selected for detailed study after initial testing revealed their significant inhibition of PMN superoxide generation in response to N- formyl-Met-Leu-Phe (FMLP) (64% for 1–14 and 64% for 1–15; P less than .05). In further experiments, Ab 1–14 augmented PMN adhesion (by 111%; P less than .01) and degranulation (by 52%; P less than .05) in response to FMLP, while Ab 1–15 inhibited these responses by 42% and 29%, respectively (P less than .05). Ab 1–14 reduced PMN chemotaxis in response to FMLP by 37% (P less than .02), and unlike Ab 1–15, Ab 1–14 significantly reduced unstimulated PMN binding of complement-coated sheep red blood cells. Ab 1–14 and Ab 1–15 significantly reduced PMN superoxide production in response to phorbol myristate acetate (PMA) (14% and 23%, respectively; P less than .05). Whereas 1–14 was found to increase PMA-induced cell degranulation significantly (175%), Ab 1–15 did not alter degranulation response to PMA. Immunoprecipitation showed that Ab 1–14 and Ab 1–15 recognized respective surface antigens of 94,000 mol wt and 130,000 to 180,000 mol wt. Our findings suggest that the surface molecules identified by these two monoclonal antibodies play a significant role in neutrophil activation by both FMLP and PMA.
Disparities in estimates of IgG bound to normal platelets
Authors: Blumberg N , Masel D , Stoler M
Published: October 2019

Estimates of the number of IgG molecules bound to normal platelets have ranged from several hundred to several tens of thousands. The lower estimates were generated from direct binding assays and stoichiometric assumptions. The higher values derive from competitive binding assays, in which platelet-associated IgG (PAIgG) is calculated from a standard curve using soluble IgG standards. Using a kinetic-ELISA (enzyme-linked immunosorbent assay) antiglobulin assay, we measured normal platelet IgG to be 21,200 +/- 9,400 molecules per platelet when a competitive assay and soluble IgG standards were used. Direct measurement of bound antiglobulin by kinetic-ELISA and stoichiometric assumptions yielded a measurement of 259 +/- 117 IgG molecules per platelet. Soluble IgG and PAIgG are not comparable in their ability to bind anti-IgG. Disparities in estimates of normal PAIgG are probably due to methodological differences. The estimate most likely to be correct is several hundred IgG or less per normal platelet.
Immunohistochemical localization of membrane and alpha-granule proteins in human megakaryocytes: application to plastic-embedded bone marrow biopsy specimens
Authors: Beckstead JH , Stenberg PE , McEver RP , Shuman MA , Bainton DF
Published: October 2019

Using a new technique for antigen localization, we have demonstrated platelet proteins in megakaryocytes in plastic-embedded biopsy specimens of normal human bone marrow. In a series of 25 specimens, megakaryocytes showed labeling with antibodies to the integral membrane glycoproteins IIIa, IIb, and the IIb-IIIa complex; granule membrane protein 140; and five alpha-granule matrix proteins: thrombospondin, factor VIII-related antigen, beta-thromboglobulin, platelet factor 4, and fibrinogen. The antibodies to the membrane glycoproteins IIIa, IIb, and IIb-IIIa produced diffuse cytoplasmic staining and heavier staining on the plasma membrane, whereas the antibodies to the alpha-granule matrix proteins produced a distinct granular staining within the cytoplasm. Staining for granule membrane protein 140 was also granular in distribution. Rare mononuclear cells consistent with megakaryocyte precursors were labeled with these markers. Other enzyme histochemical and lectin-binding studies showed that the enzyme alpha-naphthyl acetate esterase, the lectin Ulex europaeus I, and the periodic-acid Schiff reaction were consistent, but not specific, markers of megakaryocytes. This immunohistochemical technique should facilitate the examination of qualitative and quantitative changes in megakaryocytes in a variety of physiologic and pathologic processes.
Influence of the calcium-sensitive fluorophore, Quin 2, on platelet function
Authors: Rao GH , Peller JD , Semba CP , White JG
Published: October 2019

Recent investigations using Quin 2, a fluorophore used to monitor cytosolic free calcium shifts, have shown that strong agonists cause a dramatic dose-dependent increase in platelet fluorescence. However, weak agonists stimulated little or no increase in light emission of Quin 2-loaded platelets, suggesting that calcium flux is not involved in activation by these agents. The present study has sought an alternative explanation for the failure of weak stimuli to cause a rise in cytosolic free calcium in platelets containing Quin 2. Conditions used to prepare, wash, load, gel-filter, and evaluate the fluorophore- filled cells were examined for their compatibility with retention of sensitivity to activation by weak agonists. The technique used to measure shifts in cytosolic calcium with Quin 2 requires multiply washed, unstirred platelets. Under these conditions, platelets do not aggregate or secrete in response to weak agonists. Quin 2, at concentrations greater than 40 mumol/L, inhibits the response of platelets to strong agonists, and completely blocks their reaction to weak agonists. Quin 2 inhibition of platelet function appears related to high buffering capacity for free calcium, although other mechanisms cannot be ruled out. This suggestion is supported by the observation that Quin 2-induced blockade can be overcome by membrane modulation, which is a calcium-dependent process. However, since both agonists are weak, significant elevation in cytosolic calcium concurrent with functional restoration could not be demonstrated under the experimental conditions used for monitoring calcium. Thus, the conditions used to prepare platelets for Quin 2 evaluation and Quin 2 itself appear to be responsible for the failure of weak agonists to cause evidence of a calcium shift in fluorophore-loaded cells.
Clinical and hematologic characteristics in acute leukemia with 11q23 translocations
Authors: Kaneko Y , Maseki N , Takasaki N , Sakurai M , Hayashi Y , Nakazawa S , Mori T , Sakurai M , Takeda T , Shikano T
Published: October 2019

We studied the clinical, morphological, and immunologic characteristics of 11 patients with 11q translocation-associated acute leukemia. There were three patients with t(9;11)(p22;q23), one with a variant of the t(9;11), three with t(11;19)(q23;p13), two with t(1;11)(p32;q23), one with t(10;11)(p15;q22or23), and one with t(11;17) (q23;q25). The breakpoints in chromosome 11 clustered in band q23. The morphological feature was FAB-M5 in two patients, FAB-M2 in one, FAB-L1 in six, and lymphoblastic lymphoma in one. The remaining patient underwent morphological changes from FAB-L1 seen at the time of diagnosis to M5b at relapse. Immunologic marker studies in ten patients revealed that one had T cell type; another pre-B cell type; three CALLA- Ia- non-T, non-B type; two CAL-LA- Ia+ non-T, non-B type; two monocytic type (positive Fc-receptor); and the remaining one underwent phenotypic changes from CALLA+ Ia+ non-T, non-B type to monocytic type. The patients were usually young; five were under 1 year and two were 9 and 13 years. Hyperleukocytosis was observed in eight of the ten patients with acute leukemia, and two of the eight died of intracranial hemorrhage within two days of admission, associated with disseminated intravascular coagulation. These findings indicate that leukemia with the 11q23 translocation share certain characteristics in common, irrespective of the recipient chromosome, even though the latter may have some influence on the morphological and immunologic phenotype. Our data provide a hypothesis that multipotent stem cells are involved in the genesis of the 11q translocation-associated leukemia.
Enhanced survival of sickle erythrocytes upon treatment with glyceraldehyde
Authors: Benjamin LJ , Manning JM
Published: October 2019

Glyceraldehyde has been demonstrated to be an antisickling agent in vitro. In the present investigation, chromium-51 red cell studies were used to investigate the life span in vivo of sickle erythrocytes after treatment with glyceraldehyde in vitro. The mean survival (T1/2) of control cells was 5.8 +/- 1.6 days, whereas cells treated with 10 mmol/L or 20 mmol/L glyceraldehyde survived 9.0 +/- 1.4 (P less than .05) and 11.3 +/- 0.8 (P less than .002) days, respectively. The extent of modification by glyceraldehyde was 0.4 to 1.0 lysine residue per hemoglobin tetramer. These studies demonstrate not only a prolongation of the life span of sickle erythrocytes by treatment with glyceraldehyde but also the absence of any deleterious effects that would be revealed by this study.
Cytogenetic and immunophenotypic analysis of cell lines established from patients with T cell leukemia/lymphoma
Authors: Smith SD , Morgan R , Link MP , McFall P , Hecht F
Published: October 2019

Cell lines were established from five patients with T cell malignancies. Two patients had T cell lymphoblastic lymphoma (T-LL), whereas three patients had T cell acute lymphoblastic leukemia (T-ALL). Both T-LL cell lines expressed cell surface antigens characteristic of midthymocytes (Leu 2, 3, 6+). One T-ALL cell line also expressed this immunophenotype, one expressed suppressor/cytotoxic antigens (Leu 2+; Leu 3, 6-), and one expressed antigens of a mature but uncommitted T cell (Leu 4+; Leu 2, 3, 6-). Cytogenetic analysis showed that each cell line had 46 chromosomes with pseudodiploidy. The three T-ALL cell lines had only a few chromosome changes; one cell line had one deletion, another had two deletions, and the third had a translocation and two deletions (including loss of part of 9p). In comparison, both T-LL cell lines had complex chromosome changes, including most notably a rearrangement of band 14q11.2. The immunophenotypes and chromosome breakpoints showed patterns of interlock between the T-LL and T-ALL cell lines because common abnormalities occurred at six distinct chromosome sites. Cell lines with limited and specific chromosomal abnormalities are important because they can provide the basic material for molecular genetic studies that could elucidate the genetic mechanisms involved in neoplasia.
Cellular ras oncogene expression and cell cycle measured by flow cytometry in hematopoietic cell lines
Authors: Andreeff M , Slater DE , Bressler J , Furth ME
Published: October 2019

Human hematopoietic malignancies provide an excellent model for the study of the activity of cellular oncogenes in a context of known defects in cell proliferation and differentiation. A flow cytometric immunofluorescence assay was developed to quantitate the expression of the cellular ras oncogenes in relation to the cell cycle in individual leukemic cells. Specific binding of a monoclonal antibody to the 21-kd protein (p21ras) encoded by the Ha-ras, Ki-ras, and N-ras genes was measured by flow cytometry and confirmed by fluorescence microscopy. P21ras was detected in 416B, a murine hematopoietic precursor cell characterized by a high level of Ki-ras expression, and in the human leukemic cell lines P-12 and KG-1. The presence of p21ras in the cell lines was also shown by immunoprecipitation. Cellular DNA content was determined simultaneously to define cell cycle phases. There was an equal distribution of p21ras in G1, S, and G2M, with considerable heterogeneity of ras gene expression in the G1 compartment. The assay allows oncogene expression to be studied in populations of intact single cells in which cell heterogeneity is maintained, requires very few cells per sample, and directly correlates oncogene expression to cell kinetic data.
Acid phosphatase activity in mononuclear phagocytes and the U937 cell line: monocyte-derived macrophages express tartrate-resistant acid phosphatase
Authors: Snipes RG , Lam KW , Dodd RC , Gray TK , Cohen MS
Published: October 2019

Tartrate-resistant acid phosphatase (TRAcP) is used as a marker for osteoclasts, which are believed to be derived from phagocytic cells or phagocyte stem cell precursors. To further investigate the relationship between monocytic phagocytes and osteoclasts, acid phosphatase (AcP) activity was measured by three different techniques in human peripheral blood monocytes, monocyte-derived macrophages, and the U937 cell line. We found that cytochemistry and gel electrophoresis led to similar results, but that the colorimetric assay was inconsistent. Normal human peripheral monocytes expressed both tartrate-sensitive and -resistant AcP. In culture these cells formed polykaryons and expressed TRAcP activity that was further identified as an isoenzyme associated with bone tissue. In contrast, the U937 cells did not express TRAcP activity as measured by gel electrophoresis. Both U937 cells and monocytes possess material that interferes with interpretation of the colorimetric assay of AcP. The presence of TRAcP in monocyte-derived macrophages further supports the relationship between phagocytic cells and bone osteoclasts.
Diffuse large cell lymphoma in a patient with hairy cell leukemia: immunoglobulin gene analysis reveals separate clonal origins
Authors: Downing JR , Grossi CE , Smedberg CT , Burrows PD
Published: October 2019

A 75-year-old man with hairy cell leukemia (HCL) was found to have an immunoblastic lymphoma of the small bowel. Immunologic and genotypic characterization of these neoplasms revealed both the HCL and the immunoblastic lymphoma to be of the B cell lineage. The HCL expressed the HCL-associated antigens detected by the monoclonal antibodies HC-1 and HC-2, whereas the immunoblastic lymphoma failed to react with these antibodies. In addition, surface immunoglobulin light chains could not be accurately determined for the hairy cells, whereas the immunoblastic lymphoma was shown to express only kappa immunoglobulin light chains. These immunophenotype differences were compatible with either the clonal evolution of the HCL into the immunoblastic lymphoma or a separate clonal origin for these two neoplasms. An analysis of tumor DNA by Southern blot hybridization revealed different heavy-chain and kappa light-chain gene rearrangements in these two malignancies. Thus the occurrence of the large cell lymphoma most likely represents the emergence of a second clonally unrelated B cell malignancy.
Carbonyl iron therapy for iron deficiency anemia
Authors: Gordeuk VR , Brittenham GM , McLaren CE , Hughes MA , Keating LJ
Published: October 2019

To determine if elemental carbonyl iron powder is safe and effective therapy for iron deficiency anemia, 20 nonanemic and 32 anemic volunteers were studied. Single doses of 1,000 to 10,000 mg of carbonyl iron (15 to 150 times the 65 mg of iron in the usual dose of ferrous sulfate) were tolerated by nonanemic volunteers with no evidence of toxicity and only minor gastrointestinal side effects. Anemic volunteers (menstruating women who had previously donated blood) were treated with several regimens providing 1,000 to 3,000 mg of carbonyl iron daily in one to three doses for 8 to 28 days. After 12 weeks anemia was corrected in 29 of 32 patients, and serum ferritin was greater than 12 micrograms/L in 14. Hemoglobin regeneration proceeded at a rate similar to that described for therapy with oral iron salts and parenteral iron dextran. There was no evidence of hematologic, hepatic, or renal toxicity, but mild gastrointestinal side effects occurred in a majority of anemic volunteers. Carbonyl iron is an effective, inexpensive treatment for iron deficiency anemia, is accompanied by tolerable side effects and may have an advantage over therapy with iron salts by substantially reducing or eliminating the risk of iron poisoning in children.
Loss of the largest von Willebrand factor multimers from the plasma of patients with congenital cardiac defects
Authors: Gill JC , Wilson AD , Endres-Brooks J , Montgomery RR
Published: October 2019

We identified a consecutive series of 12 children with noncyanotic congenital cardiac lesions with loss of the largest plasma von Willebrand factor (vWF) multimers determined by SDS-agarose electrophoresis. Seven had previous histories of mucocutaneous hemorrhage; ten had a prolonged bleeding time. Analysis of the factor VIII molecular complex revealed that six patients had reduced vWF measured both immunologically (vW:Ag) and by ristocetin cofactor assay (vW:rist). All had normal or borderline normal factor VIII procoagulant (F VIII) concentrations. Three children had prolonged partial thromboplastin times due to concurrent factor XII deficiency; none had laboratory evidence of intravascular coagulation. Five of the children were restudied after surgical correction of their cardiac lesions. Four had normalization of vWF multimers; the fifth, whose vWF was abnormal postoperatively, had a residual pressure gradient across a previous pulmonary artery banding site. Multimeric abnormalities were not found in the parents of three patients. Thus some patients with noncyanotic congenital heart disease may have an acquired abnormality of vWF that is normalized with correction of the abnormal hemodynamic state.
Effect of acidic and basic isoferritins on in vitro growth of human granulocyte-monocyte progenitors
Authors: Dezza L , Cazzola M , Piacibello W , Arosio P , Levi S , Aglietta M
Published: October 2019

Acidic isoferritins have been previously found to be highly potent inhibitors of hematopoietic progenitors at concentrations of 10(-16) to 10(-18) mol/L, and it has been suggested that acidic isoferritin inhibitory activity plays a role in the regulation of normal hematopoiesis and also in the pathogenesis of leukemia. To characterize the ferritin species that affect the in vitro growth of human colony- forming unit-granulocyte-macrophage (CFU-GM), we tested different preparations of basic (L-subunit-rich) and acidic (H-subunit-rich) isoferritins. Three preparations of human liver (basic) ferritin did not show any effects on CFU-GM growth at concentrations up to 10(-9) mol/L, irrespective of the degree of glycosylation. Acidic isoferritins were purified both from HeLa cells and human heart. HeLa cell ferritin did not affect in vitro colony formation. One of two preparations of human heart ferritin, containing 5% glycosylated ferritin, showed a mean inhibition of 26% +/- 8% of the control at 10(-9) mol/L (P less than .02), whereas the other preparation, which contained no glycosylated ferritin, did not show any effect of CFU-GM growth. A preparation enriched for glycosylated acidic isoferritins from human heart was found to produce a mean inhibition of 32% +/- 11% of the control at 10(-9) mol/L (P less than .01), whereas another one was ineffective. A significant part of the inhibitory activity was removed by preincubation with the monoclonal antibody 2A4 directed against human heart ferritin. The present findings indicate that basic isoferritins, ie, the predominant ferritin type in human blood, have no effect on the growth of human CFU-GM, and this is in keeping with indirect clinical evidence. Inhibition of colony formation may be obtained by some preparations of acidic isoferritins that are rich in H subunits and bind to concanavalin A. The mechanism(s) responsible for this are not clear, but the effective concentrations are higher than those found in human blood both under normal conditions and in leukemia. At present, the physiologic significance of the observed inhibitory activity is uncertain.
Development of a radioimmunoassay for the fibrinogen-derived peptide B beta 1-42
Authors: Weitz JI , Koehn JA , Canfield RE , Landman SL , Friedman R
Published: October 2019

The peptide B beta 1–42 is the initial cleavage product of plasmin- mediated proteolysis of the NH2-terminal region of fibrinogen or fibrin I, while beta 15–42 is the major fragment released by plasmin degradation of fibrin II. Numerous studies have described the measurement of plasma B beta 1–42 levels as an index of plasmin activity. Previous assays were indirect and included quantitation of thrombin-increasable fibrinopeptide B immunoreactivity (TIFPB) or measurement of beta 15–42 with an antiserum (132) which cross-reacted with B beta 1–42. We report on a new antiserum (R142) directed against B beta 1–42 which does not cross-react with beta 15–42 or fibrinopeptide B. Employing this antiserum, a specific assay for B beta 1–42 was developed. This assay was used to measure plasmin-mediated B beta 1–42 release from fibrinogen and its subsequent proteolysis by thrombin. The selectivity constant (Kcat/Km) for thrombin cleavage of the B beta 14–15 bond of B beta 1–42 was 10(5)-fold less than that for proteolysis of the same bond in the intact fibrinogen molecule, thus explaining the stability of this peptide in the presence of thrombin activity in the blood. Similarly, the selectivity constant for plasmin cleavage of the B beta 21–22 bond of B beta 1–42 was 140-fold less than that for proteolysis of the B beta 42–43 bond of fibrinogen, indicating that secondary plasmin attack of the B beta 1–42 molecule is not physiologically important. The specific B beta 1–42 assay provided excellent recovery of peptide added to blood or plasma. Comparison of B beta 1–42 levels with TIFPB values in 37 patient samples yielded good correlation over a wide range of levels (r2 = 0.91). The median plasma B beta 1–42 level in 15 normal individuals was 1.2 pmol/mL. This is similar to the previously reported normal range for TIFPB (1 to 4 pmol/mL) but is higher than the normal level of 0.4 pmol/mL reported with the assay employing antiserum 132. This discrepancy reflects rapid removal of Arg (B beta 42) by plasma carboxypeptidase activity resulting in 50% loss of B beta 1–42 immunoreactivity with antiserum 132 but no loss with R142. To circumvent this problem, we have developed a beta 15–42 antiserum (R154) which, like antiserum 132, cross-reacts with B beta 1–42. However, B beta 1–42/beta 15–42 immunoreactivity with R154 is stable in the presence of carboxypeptidase activity.(ABSTRACT TRUNCATED AT 400 WORDS)
Modulation of in vitro growth of murine myeloid leukemia by an autologous substance immunochemically cross-reactive with insulin and antiinsulin serum
Authors: Vuk-Pavlovic Z , Pavelic K , Vuk-Pavlovic S
Published: October 2019

Murine myeloid leukemia secretes a substance immunochemically cross- reactive with insulin (SICRI) both in vivo and in serum-free media. High SICRI concentrations in peripheral blood of tumorous animals do not affect circulating glucose levels. In culture, DNA synthesis rate per leukemic cell is proportional to cell density and is reduced by antiinsulin serum. Culture medium conditioned by leukemia cells as well as SICRI affinity purified from this medium stimulate DNA synthesis in cultured leukemia cells. It appears that autocrine stimulation of murine myeloid leukemia can be mediated in part by an insulin-related growth factor.
Activation of human granulocytes by arachidonic acid: its use and limitations for investigating granulocyte functions
Authors: Cohen HJ , Chovaniec ME , Takahashi K , Whitin JC
Published: October 2019

The effects of arachidonic acid on human granulocytes (polymorphonuclear neutrophil leukocytes, PMN) were examined with respect to early events associated with activation of the superoxide (O2-)-generating system and its reactivation with other stimuli after the addition of fatty acid-free bovine serum albumin. As with other stimuli, O2- production occurred after a lag that was dependent on the amount of arachidonic acid used. Although the lag, rate, and extent of O2- production were dose dependent, at all concentrations used, the duration of O2- production was four to six minutes. As previously described, when fatty acid-free albumin was added one minute after stimulation of PMN by arachidonic acid, O2- production ceased immediately. The O2(-)-generating system could then be reactivated by the addition of either phorbol myristate acetate (PMA), concanavalin A, or serum-treated zymosan. In previously activated cells, the lag time for reactivation was shorter and the rate of O2- production greater with PMA. PMN membrane potential was depolarized by arachidonic acid. This depolarization was not reversed with the addition of albumin. PMN cytoplasts were also capable of reversible activation by arachidonic acid in a manner identical to whole cells. Divalent cations were found to be necessary for the activation of PMN by arachidonic acid. In the absence of calcium, arachidonic acid caused rapid lysis of the cells, as manifested by the release of lactic acid dehydrogenase. We were unable to measure later events in PMN stimulated by arachidonic acid since even in the presence of divalent cations lactic acid dehydrogenase was released from the cells after five minutes of incubation. We conclude that, in addition to its ability to activate PMN, arachidonic acid is toxic to the cells and cannot be used to study late events in granulocyte activity, and, in the absence of calcium, it may be difficult to interpret its action as a stimulant of the oxidase.
Effect of activated lymphocytes on the regulation of hematopoiesis: enhancement and suppression of in vitro BFU-E growth by T cells stimulated by autologous non-T cells
Authors: Harada M , Nakao S , Kondo K , Odaka K , Ueda M , Shiobara S , Matsue K , Mori T , Matsuda T
Published: October 2019

Autologous mixed lymphocyte culture (AMLR) is an immunologic response with memory and specificity and plays a role in immune regulation. Effects of T cells activated by AMLR were studied in the regulation of in vitro erythropoiesis. AMLR-activated T cells were cocultured with autologous non-T, nonphagocytic peripheral blood mononuclear cells for assaying erythroid progenitor cells (BFU-E). T cells activated for 3 days in AMLR showed significant enhancement of in vitro colony growth by BFU-E. In contrast, activated T cells from day 7 AMLR caused significant suppression of BFU-E growth. Both enhancing and suppressing activities of AMLR-activated T cells were mediated by an la-positive and radiosensitive population within the OKT4+ subset. These observations suggest that AMLR-activated T cells may play a role in the immune-mediated regulation of in vitro erythropoiesis. It is also suggested that heterogeneous T-cell subsets may exert regulatory functions in the regulation of in vitro hematopoiesis.
The cofactor role of protein S in the acceleration of whole blood clot lysis by activated protein C in vitro
Authors: de Fouw NJ , Haverkate F , Bertina RM , Koopman J , van Wijngaarden A , van Hinsbergh VW
Published: October 2019

The effect of purified human activated protein C (APC) and protein S on fibrinolysis was studied by using an in vitro blood clot lysis technique. Blood clots were formed from citrated blood (supplemented with 125I-fibrinogen) by adding thrombin and Ca2+-ions; lysis of the clots was achieved by adding tissue-type plasminogen activator. The release of labeled fibrin degradation products from the clots into the supernatant was followed in time. We clearly demonstrated that APC accelerates whole blood clot lysis in vitro. The effect of APC was completely quenched by antiprotein C IgG, pretreatment of APC with diisopropylfluorophosphate, and preincubation of the blood with antiprotein S IgG. This demonstrates that both the active site of APC and the presence of the cofactor, protein S, are essential for the expression of the profibrinolytic properties. At present, the substrate of APC involved in the regulation of fibrinolysis is not yet known. Analysis of the radiolabeled fibrin degradation products demonstrated that APC had no effect on the fibrin cross-linking capacity of factor XIII.
Deficient induction of leukotriene synthesis in human neutrophils by lipoxygenase-deficient platelets
Authors: Kanaji K , Okuma M , Uchino H
Published: October 2019

The effect of human platelets with deficient lipoxygenase activities on leukotriene B4 (LTB4) synthesis by neutrophils was studied. When arachidonic acid (AA) metabolites obtained from the incubation of washed normal neutrophils and platelets with N- formylmethionylleucylphenylalanine (FMLP), cytochalasin B, and AA were analyzed by reversed-phase high-performance liquid chromatography, the synthesis of 5-lipoxygenase products, including LTB4, was remarkably stimulated by platelets, with their maximal effect at a ratio of platelets to neutrophils of 15:1. However, the use of lipoxygenase- deficient platelets obtained from four patients with myeloproliferative disorders instead of normal platelets showed the deficient production of 5-lipoxygenase-derived products, whereas platelets with normal lipoxygenase activities obtained from MPD patients stimulated the 5- lipoxygenase pathway similarly to the way in which normal platelets did. The addition of 12-hydroperoxyeicosatetraenoic acid (12-HPETE), a labile AA metabolite via the platelet lipoxygenase pathway, could activate the 5-lipoxygenase pathway in neutrophils incubated with FMLP, cytochalasin B and AA, but its stable end product, 12- hydroxyeicosatetraenoic acid, could not. Thus, it is suggested that lipoxygenase-deficient platelets did not sufficiently stimulate LTB4 synthesis during platelet-neutrophil interactions because of defective formation of 12-HPETE. This altered interaction between platelets and neutrophils through the lipoxygenase pathway might result in deficient responses at sites of thrombosis or inflammation in patients with deficient platelet lipoxygenase activities.
Chemoattractant-mediated stimulation of the respiratory burst in human polymorphonuclear leukocytes may require appearance of protein kinase activity in the cells' particulate fraction
Authors: Pike MC , Jakoi L , McPhail LC , Snyderman R
Published: October 2019

Low doses of aliphatic alcohols produce divergent effects on the function of chemoattractant receptors on human polymorphonuclear leukocytes (PMNs) since they enhance chemotaxis but inhibit stimulation of superoxide production by chemoattractants. As such, alcohols can provide useful pharmacologic tools to probe the mechanisms of stimulus- response coupling in leukocytes. A role for protein kinase C has been implicated in the activation of the respiratory burst in PMNs. Although the vast majority of this enzyme activity is located in the cytosolic fraction of unactivated PMNs, protein kinase C activity appears in the particulate fraction of the cells when they are stimulated to produce superoxide by either chemoattractants or by phorbol myristate acetate (PMA). Doses of the alcohols that selectively inhibited stimulation of superoxide production by chemoattractants also inhibited the appearance of protein kinase C activity as well as an undefined protein kinase activity in the particulate fraction of the cells. In contrast, the alcohols did not affect either the ability of PMA to stimulate the production of superoxide in PMNs nor the appearance of protein kinase activity in the cells' particulate fraction. PMA is known to bind and activate protein kinase C directly, thus bypassing receptor-mediated events. These data suggest that alcohols inhibit the stimulation of the respiratory burst by chemoattractants in PMNs by blocking the ability of receptor occupancy to induce the appearance of protein kinase activity in particulate fractions. These results moreover suggest that the appearance of protein kinase activity in the particulate fraction may be required for activation of the respiratory burst in PMNs.
Homozygous hemoglobin Knossos (alpha 2 beta 227(B9) Ala----Ser): a new variety of beta+-thalassemia intermedia associated with delta degree- thalassemia
Authors: Baklouti F , Dorleac E , Morle L , Laselve P , Peyramond D , Aubry M , Godet J , Delaunay J
Published: October 2019

Hb Knossos (beta 27 (B9) Ala----Ser) is a recently discovered hemoglobin variant endowed with beta-thalassemic properties (1,2) We present the first homozygous cases. The propositus, a 19-year-old man is originally from northeast Algeria, but is unrelated to other Algerians who have hemoglobin Knossos. He has a beta+-thalassemia intermedia syndrome, including microcytic, hypochromic anemia, enlargement of the spleen, and an increase in the number of reticulocytes. The reduction of beta-chain synthesis is pronounced (alpha/non alpha:2.76). Whole cells containing Hb Knossos have a dramatically low oxygen affinity (P50:38 mm Hg). The propositus also has homozygous delta degrees-thalassemia. The chromosome carrying these mutations is characterized by the DNA haplotype I.