bluuebunny/crossref_metadata_2025 huggingface dataset:
Platelet-collagen interaction: inhibition by ristocetin and enhancement by von Willebrand factor-platelet binding
Authors: LaDuca FM , Bettigole RE , Bell WR , Robson EB
Published: October 2019

The contribution of von Willebrand factor (vWF)-platelet binding to platelet-collagen interaction was examined in vitro. The binding of vWF to platelets was mediated and regulated by ristocetin. Subthreshold concentrations of ristocetin (less than or equal to 1 mg/mL), insufficient to cause ristocetin-induced platelet aggregation (RIPA), were added to platelet-rich plasma (PRP) prior to the addition of collagen. The collagen-induced platelet aggregation (CIPA) was modified by ristocetin and the degree of alteration was dependent on the ristocetin concentration. Response as a function of ristocetin concentration was designated the Collagen-Platelet Aggregation Response (CoI-PAR). In normal PRP the CoI-PAR was a progressive inhibition followed by decreasing inhibition and then an enhanced response. The enhanced response occurred over a narrow range of ristocetin concentrations (0.8 to 1.0 mg/mL). In the absence of vWF (severe von Willebrand's disease, Type I, vWF less than 1%) the CoI-PAR was a progressive, eventually complete inhibition with no enhanced response (with ristocetin concentrations up to 3.0 mg/mL). With addition of vWF to this PRP an enhanced response was observed at a ristocetin concentration inversely proportional to the vWF level. PRP from a patient with severe Hemophilia A showed a response within the normal range. Subthreshold ristocetin did not cause plasma protein precipitation or platelet release of 3H-serotonin, nor induce micro platelet aggregate formation. Digestion of platelet membrane glycoproteins (GP(s] with chymotrypsin demonstrated that upon removal of GPI, RIPA was absent, CIPA retained and the CoI-PAR was progressive inhibition, with no enhancement. With removal of GPs I, II, and III, RIPA, CIPA, and the CoI-PAR were absent. A dose-response 125I-vWF- platelet binding occurred with increasing ristocetin concentrations which was unchanged by the addition of collagen. These results demonstrated that ristocetin-platelet association inhibited CIPA, and vWF-platelet binding enhanced platelet-collagen adhesion and platelet aggregation. The in vitro-enhanced CIPA represents a vWF-dependent aggregation of sufficient magnitude to overcome the inhibitory effect of ristocetin. These studies demonstrate an influential interaction of ristocetin, vWF, and collagen with the platelet membrane and imply an important hemostatic contribution of vWF-platelet binding in platelet- collagen interaction.
Cytotoxicity of interleukin 2-activated lymphocytes for leukemia and lymphoma cells
Authors: Oshimi K , Oshimi Y , Akutsu M , Takei Y , Saito H , Okada M , Mizoguchi H
Published: October 2019

Studies were undertaken to determine whether leukemia and lymphoma cells would be lysed by autologous and allogeneic lymphokine-activated killer (LAK) cells. Peripheral blood mononuclear cells (PBMC) from patients and normal donors were cultured for five days, 2 weeks, and 4 weeks with medium containing 2,500 units of recombinant interleukin 2 (IL-2) per mL, and their cytotoxicity was assayed by a five-hour 51Cr- release test. Of primary tumors isolated from patients with acute nonlymphoblastic leukemia, acute lymphoblastic leukemia, and non- Hodgkin's lymphoma, tumors of 37 out of 40 patients tested were shown to be susceptible to normal donors' LAK, and tumors of 18 of 20 patients tested were shown to be susceptible to autologous LAK. LAK cultured for longer periods showed a tendency to have lower cytotoxicity. LAK had also low, but significant, levels of cytotoxicity for nonmalignant target cells. Because PBMC expanded in IL-2-containing medium consisted mainly of OKT3-positive pan T cells, OKT8-positive suppressor/cytotoxic cells, and Leu-11-positive natural killer (NK) cells, and treatment with OKT3 and Leu-11 monoclonal antibodies (mAb) reduced LAK activity for autologous and allogeneic tumor cells, both T and NK cells appeared to be effector cells for LAK activity. Mechanisms of target-cell recognition in the LAK system seem to be different from those in alloreactive cytotoxic T lymphocytes (CTL) based on the results that, while cytotoxicity of alloreactive CTL was inhibited by the treatment of effector cells with mAb, OKT3, and OKT8, and by the treatment of target cells with a mAb that reacts with HLA class I antigen, LAK activity was not inhibited by the above treatment. When chromosomes of IL-2-expanded PBMC in nine patients and two normal individuals were analyzed, PBMC from one patient showed chromosomes of clonal abnormalities, and PBMC from five donors showed those of nonclonal abnormalities.
Acute lymphoblastic leukemia in infants: evidence for B cell origin of disease by use of monoclonal antibody phenotyping
Authors: Dinndorf PA , Reaman GH
Published: October 2019

Since the prognosis of infants with acute lymphoblastic leukemia (ALL) is so poor, it has been suggested that these leukemias may not be lymphoid in origin, but may originate from stem cell, myeloid, or megakaryocytic progenitors. Alternately it has been hypothesized that these leukemias originate in lymphoid cells at the earliest stages of B cell development. Another possibility is that these leukemias may be of more than one lineage. Therefore we examined leukemic blasts from 12 infants with ALL using monoclonal antibodies to myeloid and lymphoid differentiation antigens. The majority of specimens expressed HLA/DR and reacted with B4 (CD19) but failed to react with stem cell, myeloid, megakaryocytic, or T cell associated antibodies. These results support the speculation that the majority of these leukemias arise in cells at the earliest stages of B cell commitment, and are not of a myeloid or biphenotypic nature.
Variables influencing the timing of marrow transplantation in patients with chronic myelogenous leukemia
Authors: Segel GB , Simon W , Lichtman MA
Published: October 2019

The prognosis for patients with chronic myelogenous leukemia (CML) has improved only for patients who can receive marrow transplantation from a histocompatible sibling. The timing of the marrow transplant is made difficult by the high peritransplant mortality of 20% to 35% and a group of patients with a prolonged chronic phase of CML, which can be identified on the basis of prognostic indexes (age, percent blood myeloblasts, spleen size, and platelet count). We have developed a mathematic model and computer program that consider age, prognostic index, and projected survival rate by transplantation to balance the risk of peritransplant mortality against the risk of delaying the transplantation of patients with Philadelphia chromosome-positive CML. The computation assesses the risk of delaying transplantation; it does not offer the option of avoiding transplantation, since long-term survival ultimately requires transplantation. Three prognostic groups were considered as described by Sokal and co-workers (Blood 63:789, 1984) (I, best; II, intermediate; III, worst prognosis). The computation used the projected survival rates of transplantation from the Seattle experience and from the International Bone Marrow Transplant Registry. As an example of the model's utility, we have determined the ratio of the calculated life expectancy to the normal life expectancy for hypothetical patients up to 50 years of age in each of the three prognostic categories. A value of 20% is used for patients who successfully receive transplants after the onset of the accelerated phase. The analysis allows assessment of the risk of delaying transplantation for a finite time in patients with CML. The importance of the method rests in its consideration of multiple variables, including the peritransplant mortality, transplant projected survival before and upon entering the accelerated phase, age, prognostic group, and other risk factors. The program permits a change in these parameters as new information or advances in treatment occur. This analysis does not replace the diagnostic deliberations of the clinician. Rather, it provides a numeric framework for prognosis based on the currently available data. The physician in conjunction with the patient, not the algorithm, makes the decisions of whether and when to transplant.
Frequent and extensive deletion during the 9,22 translocation in CML
Authors: Popenoe DW , Schaefer-Rego K , Mears JG , Bank A , Leibowitz D
Published: October 2019

Chromosomal translocation is one mechanism by which cellular oncogenes may be activated during tumorigenesis. The translocation of the abl oncogene to the Philadelphia chromosome in chronic myelogenous leukemia (CML) results in a new RNA transcript that fuses sequence from chromosome 22 to sequence from the abl oncogene. This RNA presumably codes for a new abl-related protein product found in CML, the activity of which is different from the normal abl protein. The molecular structure of the translocation varies from patient to patient, and the individual variation in RNA transcript and protein product remains to be defined. This report describes the frequent occurrence of chromosomal deletion within the 9q+ chromosome during these translocations. The location of the deletions suggests that some mechanism maintains the chromosomal breakpoint on the Philadelphia chromosome within a limited region. These deletions complicate the interpretation of Southern blots as a means of detecting the translocation.
A prospective study of the value of bone marrow erythroid progenitor cultures in polycythemia
Authors: Lemoine F , Najman A , Baillou C , Stachowiak J , Boffa G , Aegerter P , Douay L , Laporte JP , Gorin NC , Duhamel G
Published: October 2019

The diagnostic value in polycythemia of the presence of endogenous erythroid colonies derived from bone marrow cells (EECs) was assessed in a prospective study on 108 patients referred for polycythemia (Hb greater than g/dL in men, greater than 16 g/dL in women) with normal plasma volume by comparison with the standard criteria, the bone marrow grade, and the serum erythropoietin (Epo) level. Total red cell volume (TRCV) was high (greater than 36 mL/kg in men, 32 mL/kg in women) in 87 cases (group A) and slightly increased in 21 cases (group B). Standard criteria were applicable in 63 of 108 cases (57%); 46 were PV and 17 were secondary polycythemia (SP). Standard criteria were nonapplicable in 45 cases. EECs were present in 65 cases (60%) with a ratio of EEC/Epo-stimulated colonies of 39.5% +/- 18% (extremes 10% to 80%). EECs were noted in 43 of 46 polycythemia vera (PV) and 0 of 17 SP. Among the 45 unclassified cases, EECs were noted in 22: 18 of 29 cases from group A (10 with 2 major and 1 minor criteria; 8 with 2 major criteria) and 4 of 16 cases from group B (with variable standard criteria, 2 belonging to a PV family). In group A, there was a positive significant correlation between EECs and the presence of two major and 1 minor criteria (P less than .0001). In group B, there was a positive significant correlation between EECs and the presence of at least 1 major criterion and 2 minor criteria or a family background (P less than .0001). The unclassified polycythemias with EECs in the bone marrow are characterized by a bone marrow grade and a mean serum Epo level not different from that of patients with PV and an active course of the disease. The unclassified polycythemias without EECs in the bone marrow are a heterogeneous group corresponding in some cases to SPs of unknown origin (slightly increased bone marrow grade and/or high serum Epo level), and in others cases to spurious polycythemias (normal bone marrow grade and/or normal Epo level). In conclusion, EECs were of great value in differentiating PV from SP (P less than .001), and in allowing the diagnosis of PV in the absence of all the standard criteria even when TRCV was slightly increased. In our study, EEC improved the classification of polycythemia by 22%. The recommended diagnostic steps for the evaluation of polycythemia must be reconsidered.
Use of surface marker analysis to predict outcome of adult acute myeloblastic leukemia
Authors: Griffin JD , Davis R , Nelson DA , Davey FR , Mayer RJ , Schiffer C , McIntyre OR , Bloomfield CD
Published: October 2019

In order to investigate the clinical significance of surface antigen analysis in acute myeloblastic leukemia (AML), the blasts from 196 patients with AML were analyzed prospectively with a panel of 16 monoclonal antibodies. The antibodies were selected to identify differentiation-associated antigens of either the myeloid lineage (MY9, PM-81, AML-2–23, MY7, MCS-1, MY8, Mo1, MY1, MY4, Mo2), T cell lineage (T101, T11), B cell lineage (B1, B4) or multiple lineages [J5 (CALLA), HLA-DR]. Independent morphological review and classification by French- American-British (FAB) criteria was performed in 161 of the 196 cases. One or more myeloid surface antigens were detected on the blasts of 195 cases, while B and T cell markers were detected on 0% to 2% of cases. When both blood and marrow samples were studied on the same patient, very few differences were noted between the antigenic profiles of the paired specimens. The frequency of expression of individual myeloid antigens ranged from 91% (PM-81) to 29% (Mo2). Expression of individual antigens was found to correlate significantly with several clinical parameters including FAB classification, cytochemical staining for alpha naphthyl acetate esterase, leukocyte count, and the presence of extramedullary disease at presentation. Two myeloid antigens (MY4 and MY7) predicted for a low rate of complete remission (CR) to standard induction chemotherapy. MY4+ cases (37% of the total population) had a CR rate of 53%, while M4- cases had a CR rate of 69% (P = .03). MY7+ cases (57% of the total population) had a CR rate of 55% while MY7- cases had a CR rate of 73% (P = .01). Neither MY4 nor MY7 antigen expression was correlated with patient age. Paired combinations of antigens were also examined. The [MY4- MY7-] phenotype was exhibited by 32% of all cases and was associated with an 82% CR rate while the CR rate of all other cases was 54% (P = .001). The expression of three antigens (HLA-DR, MY8, Mo1) was associated with a decreased continuous complete remission (P less than .05, median follow-up time of 19 months). Expression of MY8 antigen was also associated with decreased survival (P = .03). These results confirm earlier reports of antigenic heterogeneity in AML, and indicate that immunologically defined subgroups of AML patients which are of potential clinical significance can be identified.
Acute myelomonocytic leukemia with abnormal eosinophils and inv(16) or t(16;16) has a favorable prognosis
Authors: Larson RA , Williams SF , Le Beau MM , Bitter MA , Vardiman JW , Rowley JD
Published: October 2019

Inv(16)(p13q22) and t(16;16)(p13;q22) are recurring chromosomal rearrangements which juxtapose the metallothionein gene cluster at 16q22 with other DNA sequences from 16p13. We have studied 20 men and 13 women who had acute nonlymphocytic leukemia; 27 patients had an inv(16) and six patients had a t(16;16). Eight patients also had trisomy 22, and four had trisomy 8. All but two patients had the unique morphologic features of acute myelomonocytic leukemia with abnormal eosinophils (M4Eo). In one patient with M4 leukemia, abnormal eosinophils were not observed in the marrow. A second patient had acute monocytic leukemia, plus abnormal eosinophils. Eosinophils constituted 1% to 46% (median, 6%) of the bone marrow cells, and in all but a single patient, the eosinophils exhibited distinctly abnormal morphology. Twenty-five patients have had a complete remission (78% of treated patients). Nine patients have remained in remission longer than 24 months. No patient had symptoms of central nervous system (CNS) disease at diagnosis, and none had CNS leukemic mass lesions at any time. Treatment with high-dose cytarabine may have provided prophylactic CNS therapy. Four additional patients with chromosomal rearrangements involving a breakpoint at 16q22 but not at 16p13 have had different morphological features and different clinical courses. Thus, the juxtaposition of genes at 16p13 and 16q22, which occurs both in the inv(16) and the t(16;16), results in a specific subset of acute nonlymphocytic leukemia that has a favorable prognosis.
Leukemia diagnosis and testing of complement-fixing antibodies for bone marrow purging in acute lymphoid leukemia
Authors: Campana D , Janossy G
Published: October 2019

In this paper a microplate method is described for diagnosing acute leukemia and for investigating the reactivity of monoclonal antibodies (MoAbs) against membrane antigens in combination with rabbit or murine antibodies to nuclear terminal transferase (TdT). The speed of this method facilitates the investigation of fresh leukemic cells from individual patients and assesses the cytolytic efficacy of the relevant MoAbs in the presence of complement (C'). Lymphoblasts (TdT+) are mixed in equal proportions with known numbers of “inert” cells, eg, RBC or nonleukemic bone marrow (BM). Following incubation with MoAbs and C' the ratio of residual TdT+ cells and inert cells is determined on cytospin preparations. Initially, percentages of TdT+ cells are counted in a unit volume of 5,000 inert cells, followed by the scanning of greater than 2 X 10(4) inert cells on entire slides. With this method more than 4 log cytoreduction of TdT + cells is detected. The method is also applicable for studying the cytolysis of malignant B cells by using mostly monoclonal lg expression rather than TdT for the identification of residual B cells. Ten representative patients selected from a group of greater than 100 are reported. In some cases cytoreduction of greater than 4 log with no identifiable residual TdT + cells is achieved by a single C'-fixing MoAb: anti-CD10 (RFAL3) in common acute lymphoid leukemia (ALL) and anti-CD7 (RFT2) in T cell ALL (T-ALL). Other cases require cocktails of anti-CD10, anti-CD19, and anti-CD24 in common ALL or anti-CD7 and anti-CD8 in T-ALL. In T-ALL a few TdT + cells remain that exhibit the features of normal TdT + BM cells (CD7-, HLA-DR+). This is particularly noticeable when patients are studied in partial remission or if nonleukemic BM is used as a source of inert cells. The methods described here contribute to establishing a range of MoAbs (ie, of IgM class) and techniques for efficient purging and to comparing the efficacy of “clean-up,” in remission, of common ALL, T-ALL, and B cell malignancies.
Proteolysis of platelet glycoprotein Ib by plasmin is facilitated by plasmin lysine-binding regions
Authors: Adelman B , Michelson AD , Greenberg J , Handin RI
Published: October 2019

We have characterized the effects of plasmin on glycoprotein Ib (GpIb), a platelet membrane receptor for von Willebrand factor (vWF), and on glycocalicin, a fragment of the alpha chain of GpIb that contains the vWF-binding region. The addition of 4.5 X 10(-7) mol/L plasmin to washed platelets caused a time-dependent decrease in ristocetin- induced, vWF-dependent platelet agglutination. epsilon-Aminocaproic acid (EACA) inhibited plasmin release of glycocalicin-related antigen from washed platelets and preserved vWF-dependent platelet agglutination, thus indicating that the lysine-binding sites on plasmin facilitated its degradation of GpIb. To demonstrate a direct interaction between plasmin and the vWF-binding region of GpIb we incubated purified glycocalicin with plasmin. Plasmin degraded the glycocalicin into two small carbohydrate-poor peptides and into a larger carbohydrate-rich fragment. EACA was able to inhibit plasmin- mediated degra dation of glycocalicin in a concentration-dependent fashion. These studies indicated that plasmin degradation of GpIb was due to a direct interaction between plasmin and GpIb and that this effect was mediated by the lysine-binding region of the plasmin molecule.
Denaturant-induced stimulation of the beta-migrating plasminogen activator inhibitor in endothelial cells and serum
Authors: Erickson LA , Hekman CM , Loskutoff DJ
Published: October 2019

Cultured bovine aortic endothelial cells and human serum contain plasminogen activator inhibitors (PAIs) that are immunologically related. In the present study, the electrophoretic mobilities, molecular weights (mol wt), and activities of these PAIs were compared. When fractionated by agarose zone electrophoresis, both PAIs migrated with beta mobility as compared with the mobilities of human plasma/serum proteins. Two-dimensional electrophoretic analysis, employing agarose zone electrophoresis in the first dimension and sodium dodecyl sulfate-polyacrylamide gel electrophoresis in the second dimension, indicated that these beta-PAIs comigrated, both having a mol wt of approximately 50,000. The activity of the PAI in endothelial cell- conditioned medium is enhanced severalfold by treatment with either sodium dodecyl sulfate or guanidine. In preliminary experiments, we were unable to stimulate the PAI activity of undiluted serum by similar treatments. However, the PAI activities in both diluted serum and gel- filtered or electrophoretically fractionated serum were enhanced by treatment with these denaturants. The gel filtration studies also revealed that serum contains multiple forms of the beta-PAI. These forms may represent polymeric PAI and/or complexes between the PAI and other serum components. These findings indicate that the primary PAIs in bovine endothelial cells and human serum are not only immunologically related but are also biochemically similar.
Beta zero-thalassemia in association with a gamma-globin gene quadruplication
Authors: Yang KG , Liu JZ , Kutlar F , Kutlar A , Altay C , Gurgey A , Huisman TH
Published: October 2019

We have studied the hematology, hemoglobin composition, and globin gene arrangements in one young Turkish boy with a beta zero-thalassemia homozygosity and in 11 of his relatives. Evidence is presented that the chromosome with the beta zero-thalassemia determinant carries a gamma- globin gene quadruplication, perhaps in a -G gamma-G gamma-G gamma-A gamma-gene arrangement. The eight gamma-globin genes in this patient produced G gamma and A gamma chains in a 95 to 5 ratio, and nearly 99% of the patient's hemoglobin was of the fetal type. The clinical condition resembled that of a thalassemia intermedia. HbF levels in eight beta-thalassemia heterozygotes varied between 0.5 and 4.2% and the percentages of G gamma in this HbF averaged at 87% or 95%; this level is to some extent related to the haplotype of the normal chromosome. All subjects carried four alpha-globin genes; a new BglII polymorphism was observed within the psi alpha-globin gene.
Studies on B lymphoid tumors treated with monoclonal anti-idiotype antibodies: correlation with clinical responses
Authors: Lowder JN , Meeker TC , Campbell M , Garcia CF , Gralow J , Miller RA , Warnke R , Levy R
Published: October 2019

Monoclonal anti-idiotype antibodies can be made which are exquisitely specific for B lymphocytic malignancies. We have conducted a clinical trial in which some patients' tumors regressed after infusion of such antibodies. Here, we evaluated characteristics of the antibodies, the tumors, and the patients to determine which features best correlated with the clinical response. Neither the isotype of the murine antibodies, nor their avidity were predictive of clinical outcome. The specific epitope to which the antibodies bound was characterized by immunochemical techniques. Reactivity with a heavy-light chain combinatorial determinant correlated somewhat with clinical effect. Variations in the characteristics of the individual tumors such as antigen sites per cell and ability to modulate the surface immunoglobulin were not predictive of response. In one patient with prolymphocytic leukemia the anti-idiotype antibody had a direct antiproliferative effect on tumor cells in vitro. This patient's tumor response was explainable by such a direct mechanism. In the other patients, who had lymphomas, therapeutic outcome correlated with the number of host nontumor cells infiltrating the tumor. The vast majority of these nontumor cells were mature T lymphocytes of the Leu 4, Leu 3 (T3, T4) phenotype. Thus, a preexistent host-tumor interaction seems to be important in the in vivo effect of anti-idiotype antibodies in B cell tumors.
Activation of plasminogen by single-chain urokinase or by two-chain urokinase--a demonstration that single-chain urokinase has a low catalytic activity (pro-urokinase)
Authors: Pannell R , Gurewich V
Published: October 2019

Single-chain urokinase (SC-UK) has an intrinsic amidolytic activity, as measured with synthetic substrate (Kabi S-2444; pyro-Glu-Gly-Arg- pNitroanalide), which was found to be 0.1% to 0.2% that of its plasmin- activated derivative, two-chain UK (TC-UK). A study of the reaction of SC-UK with plasminogen is complicated by the effect of the reaction product, plasmin, on both reactants. The resultant generation of TC-UK and Lys-plasminogen produces secondary reactions which greatly augment plasminogen activation. To confine enzymatic activity to the primary reaction, after pretreatment to eliminate trace TC-UK contaminants, SC- UK was incubated with Glu- or Lys-plasminogen in the presence of aprotinin (500 KIU/mL) to inhibit generated plasmin and dansyl-glutamyl- glycyl-arginyl-chloromethylketone (20 mumol/L), which irreversibly inhibited TC-UK but not SC-UK. Analysis by reducing sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) showed a plasminogen-activating activity for SC-UK that was approximately 0.4% that of TC-UK. Both SC-UK and TC-UK preferentially activated Lys- plasminogen over Glu-plasminogen. Similarly, Glu-plasminogen activation was augmented by lysine or soluble fibrin. The ratio of the reaction rates of SC-UK and TC-UK were comparable for Glu- and Lys-plasminogen. It is concluded that there is a major difference in the catalytic activities of SC-UK and TC-UK against plasminogen that is comparable to that against synthetic substrate.
Terminal deoxynucleotidyl transferase expression can precede T cell receptor beta chain and gamma chain rearrangement in T cell acute lymphoblastic leukemia
Authors: Greenberg JM , Kersey JH
Published: October 2019

The nuclear enzyme terminal deoxynucleotidyl transferase (TdT) is thought to contribute to the diversity of certain immunoglobulin and T cell receptor gene rearrangements through the addition of random nucleotides at their variable (V)-joining (J) region junctions. An acute lymphoblastic leukemia (ALL) with an immature T cell phenotype (CD7+, CD5+, CD1+/-, CD2+/-, CD3-, CD4-, CD8-) was found to be TdT+ with germline immunoglobulin heavy chain, T cell receptor beta chain, and T cell gamma chain genes. The data indicate that TdT expression can precede T gamma and T beta rearrangement during T lymphoid ontogeny consistent with its proposed association with the T cell receptor rearrangement process. Southern analysis of certain cases of T-ALL may not result in the detection of a monoclonal population of cells.
Platelet aggregation in whole blood from patients with Glanzmann's thrombasthenia
Authors: Burgess-Wilson ME , Cockbill SR , Johnston GI , Heptinstall S
Published: October 2019

We examined platelet aggregation in platelet-rich plasma (PRP) and in whole blood from two patients with Glanzmann's thrombasthenia. In PRP, aggregation was measured by monitoring the changes in light absorbance that occurred in response to aggregating agents; to measure platelet aggregation in whole blood, we used a platelet counting technique. In PRP, the patients' platelets showed defective aggregation in response to ADP, adrenaline, arachidonic acid (AA), and collagen, but normal agglutination occurred in response to ristocetin. In whole blood, however, platelet aggregation in response to the aggregating agents appeared to be either very similar to that which occurred in blood from normal subjects or only slightly reduced. There was a reduced response to all concentrations of ADP and to low concentrations of collagen but a normal response to all concentrations of adrenaline, AA, and higher concentrations of collagen. Conversely, there seemed to be an increased agglutination response to ristocetin. The abnormality in our two patients with Glanzmann's thrombasthenia probably lies in the inability of their platelets to form large, macroscopic aggregates rather than in platelet aggregation per se.
Purification and properties of bacterially synthesized human granulocyte-macrophage colony stimulating factor
Authors: Burgess AW , Begley CG , Johnson GR , Lopez AF , Williamson DJ , Mermod JJ , Simpson RJ , Schmitz A , DeLamarter JF
Published: October 2019

Human granulocyte-macrophage colony stimulating factor (GM-CSF) has been synthesized in high yield using a temperature inducible plasmid in Escherichia coli. The human GM-CSF is readily isolated from the bacterial proteins because of its differential solubility and chromatographic properties. The bacterially synthesized form of the human GM-CSF contains an extra methionine residue at position 1, but otherwise it is identical to the polypeptide predicted from the cDNA sequence. The specific activity of 2.9 X 10(7) units/mg of protein for purified bacterially synthesized human GM-CSF indicates that despite the lack of glycosylation, the molecule is substantially in its native conformation. This molecule stimulated the same number and type of both seven- and 14-day human bone marrow colonies as the CSF alpha preparation from human placental conditioned medium. Human GM-CSF had no activity on murine bone marrow or murine leukemic cells. There was no detectable, direct stimulation of adult human erythroid burst forming units (BFU-E) by the bacterially synthesized human GM-CSF. Although impure preparations containing native human GM-CSF (eg, human placental conditioned medium) stimulated the formation of mixed colonies, even in the presence of erythropoietin, the bacterially synthesized human GM-CSF failed to stimulate the formation of mixed colonies from adult human bone marrow cells. The bacterially synthesized human GM-CSF increased N-formyl-methionyl-leucyl- phenylalanine (FMLP)-induced superoxide production and lysozyme secretion. Antibody-dependent cytotoxicity and phagocytosis by human neutrophils was stimulated by the bacterially synthesized human GM-CSF and eosinophils were also activated in the antibody-dependent cytotoxicity assay.
Divalent cation-dependent and independent surface expression of thrombospondin on thrombin-stimulated human platelets
Authors: Aiken ML , Ginsberg MH , Plow EF
Published: October 2019

Thrombospondin (TSP), a platelet alpha-granule protein, becomes expressed on the surface of thrombin-stimulated platelets. The surface expression of this protein occurs through two distinct mechanisms. At low platelet concentrations (1 X 10(8)/mL), a divalent ion-dependent, low-capacity mechanism predominates. At higher cell concentrations, a divalent ion-dependent, higher capacity mechanism prevails that can account for greater than 90% of all the TSP surface expression measured. This mechanism requires the presence of both calcium and magnesium (Ca + Mg). The dependence of the divalent ion-dependent surface expression on platelet concentration suggests that release of the molecule from the cell followed by its binding to the cell surface mediates this component of the endogenous TSP-platelet interaction. These data are consistent with a two-receptor model for the platelet- surface expression of the endogenous TSP pool.
Chlorpropamide-induced pure white cell aplasia
Authors: Levitt LJ
Published: October 2019

We investigated the mechanism for isolated agranulocytosis and marrow pure white cell aplasia in an elderly man receiving 0.5 to 1.0 g per day of chlorpropamide (Chl) without other toxic drug exposure or overt systemic illness. Patient marrow revealed an absence of recognizable granulocytic precursors; megakaryocytes and erythroid precursors were normal. The WBC count was 1800/mm3 on admission with only 2% neutrophils; the absolute neutrophil count first exceeded 500/mm3 on the 17th day following cessation of Chl. A serum Chl level on admission was 100 micrograms/mL (acute phase, AP); no Chl was detected in serum (convalescent phase, CP) assessed on the 22nd hospital day. Antineutrophil antibodies were not detected, and T cell depletion failed to augment patient in vitro granulopoiesis. Patient AP serum produced potent complement-mediated inhibition (87% +/- 7%) of autologous granulocyte progenitors (CFU-GM) with minimal inhibition of erythroid (11% +/- 5%) or multipotent (5% +/- 4%) progenitor cells. Selective inhibition by patient AP serum of CFU-GM (74% +/- 11%) was also seen against two allogeneic marrows. Patient CP serum no longer inhibited (6% +/- 4%) autologous CFU-GM. Addition of Chl (5 to 120 micrograms/mL) to CP serum but not to control serum resulted in potent drug concentration-dependent complement-mediated inhibition of autologous and allogeneic CFU-GM. Inhibition of CFU-GM in the presence of Chl was no longer demonstrable following immunoabsorbent removal of IgG from patient serum. Patient serum in the presence of Chl had limited activity against morphologically recognizable marrow granulocytic precursors in a microimmunofluorescence assay. These results are most consistent with the development of Chl-dependent, selective antibody-mediated immune inhibition of granulopoiesis.
Murine malaria decreases hemopoietic stem cells
Authors: Silverman PH , Schooley JC , Mahlmann LJ
Published: October 2019

The causes of anemia and immunosuppression, major outcomes of malaria, are not well established. This study was undertaken to investigate whether erythropoietin (EP) production is adequate and whether the hemopoietic stem cells (CFU-S) were affected during the course of infection. Groups of female Balb/c mice infected with Plasmodium vinckei vinckei, Plasmodium berghei, or Plasmodium chabaudi adami were exposed to five hours of simulated altitude equivalent to 22,000 ft. Plasma samples were collected for EP bioassay and radioimmunoassay (RIA). Using radioiron incorporation as an index of erythropoiesis, differences in response to infection with different species of plasmodia were observed. In general, decreases in erythropoietic activity were observed in bone marrow and spleen as the infection progressed and continued to be depressed after apparent resolution of a nonlethal infection with P. chabaudi adami. Marrow from infected and control femurs were tested for CFU-S content using the spleen colony assay. The cellularity and CFU-S content of the femoral marrow decrease as the parasitemia increases. All three species of plasmodia stimulate EP production during peak parasitemias, indicating that adequate amounts of EP are available to the erythron during malarial infection. Depletion of CFU-S and probable lack of compensatory turnover of CFU-S may contribute to the disease characteristics of malaria.
Epidemiological investigation of the prevalence of von Willebrand's disease
Authors: Rodeghiero F , Castaman G , Dini E
Published: October 2019

To evaluate the prevalence of von Willebrand's disease (vWd) we carried out an epidemiological investigation among school children of the Veneto region in northern Italy. A total of 1,218 of 1,281 possible children participated in the study. They were 11 to 14 years of age, and all attended secondary schools in two distinct small areas, 70 km apart, between which there is no social contact. A blood sample was taken from each subject for determination of the blood group and von Willebrand factor (vWf) level (measured as ristocetin cofactor and expressed in IU/dL after calibration of the internal pool against an international standard), and the parents were given a questionnaire concerning hemorrhagic symptoms in the members of the family in the last three generations. Separate normal ranges were calculated for blood group O and non-O subjects (1,166 children and 289 adults) with a nonparametric method because the distribution curves of the reference values did not fit the gaussian distribution. Diagnoses of vWd were considered only for children who had low vWf levels and were members of a family with a convincing bleeding history (case of “probable vWd”). A final diagnosis was assigned if, in addition to these criteria, at least one other family member on the side with hemorrhagic history had a low vWf level. Of the 1,218 children examined, ten were classified as having vWd (0.82%). Taking into account the 90% confidence interval for the lower limit of the normal range, this figure could range from 7 (0.57%) to 14 (1.15%). All these subjects were mildly to moderately affected and presented features of heterozygous classic vWd (type I). Affected subjects were distributed evenly in the two areas examined. Our results suggest that the prevalence of vWd might be much higher than previously reported and that a different screening approach might be of use for patients with mild bleeding diathesis.
The role of glutathione reductase in maintaining human granulocyte function and sensitivity to exogenous H2O2
Authors: Cohen HJ , Tape EH , Novak J , Chovaniec ME , Liegey P , Whitin JC
Published: October 2019

Human granulocytes (polymorphonuclear leukocytes, PMN) produce H2O2 and other reactive oxygen species while undergoing phagocytosis. To examine the role of the glutathione cycle in metabolizing H2O2, we incubated PMN with 1,3-bis (2-chloroethyl) nitrosourea (BCNU). Incubation of PMN with BCNU results in a dose-dependent inhibition of PMN glutathione reductase (GRED), with 50% inhibition occurring at approximately 2 micrograms/mL BCNU. PMN hexose monophosphate shunt activity stimulated with an exogenous H2O2-generating system was inhibited only when the GRED activity was reduced to less than 30% of control. BCNU-treated cells contained lower levels of reduced sulfhydryls and reduced glutathione, which decreased even more in the presence of an exogenous H2O2-generating system. The effect of BCNU and exogenous H2O2 on various aspects of phagocytosis were examined. Exposure of BCNU-treated PMN to an H2O2-generating system resulted in an inhibition of chemotactic peptide-induced shape changes and degranulation. The ability of BCNU-treated cells to produce O2- was diminished only when the PMN were incubated with an H2O2-generating system in the presence of cyanide. Ingestion of opsonized bacteria by BCNU-treated PMN was unaffected by incubation in an H2O2-generating system even in the presence of cyanide. We conclude that PMN GRED is inhibited by BCNU, the ability of PMN to metabolize H2O2 is affected only when GRED is reduced more than 70%, this inhibition affects the glutathione content of these cells, and some, but not all of the phagocytic functions of GRED-inhibited PMN are inhibited after exposure to an H2O2-generating system.
Antibodies to granulocyte precursors in selective myeloid hypoplasia and other suspected autoimmune neutropenias: use of HL-60 cells as targets
Authors: Currie MS , Weinberg JB , Rustagi PK , Logue GL
Published: October 2019

Patients with syndromes of autoantibody-mediated hematocytopenias may manifest signs of increased cell destruction and/or decreased cell production, depending on the maturity of the target cell and the effects of antibody binding. The purpose of this study was to use a cultured human cell line of hematopoietic origin for in vitro assays of antibody binding to overcome the relative inaccessibility of natural human marrow progenitor cells. This report describes the detection, using radioiodinated staphylococcal protein A (SPA), of antibodies binding to a human promyelocytic cell line (HL-60) in sera from three patients with chronic idiopathic granulocytic hypoplasia (“pure white cell aplasia,” PWCA) and 22 patients with other syndromes of suspected immune neutropenia. Bone marrow from patients with increased IgG binding to HL-60 cells showed less than 15% granulocytic lineage cellularity in 11 of 17 cases. In vitro differentiation of HL-60 cells by retinoic acid resulted in increased IgG binding for sera that had shown increased IgG binding to mature granulocytes but not undifferentiated HL-60 cells; in contrast, for sera with antibodies to untreated HL-60 cells and for normal serum, in vitro differentiation had little effect on IgG binding. Antibodies eluted from mature granulocytes were similar to the parent serum regarding the ratio of IgG binding to mature cells v HL-60 cells. No sera from 19 patients with febrile transfusion reactions showed increased IgG binding to HL- 60 cells in the absence of increased IgG binding to mature granulocytes, although two sera had antibodies to both cell types. The use of HL-60 cells as targets may permit measurement of serum antibodies associated with granulocytic hypoplasia. In combination with assays to detect antibody binding to mature granulocytes, these techniques may discriminate among autoantibody specificities for antigens that are gained, conserved, or lost during myeloid maturation.
Leukemic blast cell colony formation in semisolid culture with erythropoietin: a case report of acute poorly differentiated erythroid leukemia
Authors: Tomonaga M , Jinnai I , Tagawa M , Amenomori T , Nishino K , Yao E , Nonaka H , Kuriyama K , Yoshida Y , Matsuo T
Published: October 2019

The bone marrow of a patient with acute undifferentiated leukemia developed unique colonies after a 14-day culture in erythropoietin (EPO)-containing methylcellulose. The colonies consisted of 20 to 200 nonhemoglobinized large blast cells. Cytogenetic analysis of single colonies revealed hypotetraploid karyotypes with several marker chromosomes that were identical to those found in directly sampled bone marrow. The concurrently formed erythroid bursts showed only normal karyotypes. No leukemic colony formation was observed in other culture systems with either colony-stimulating activity (CSA) or phytohemagglutinin-stimulated leukocyte-conditioned medium (PHA-LCM). The leukemic colonies exhibited a complete EPO-dose dependency similar to that of the patient's normal BFU-E. Although cytochemical and immunologic marker studies of the bone marrow cells failed to clarify the cell lineage of the leukemic cells with extraordinarily large cell size, ultrastructural study revealed erythroid differentiation such as siderosome formation in the cytoplasm and ferritin particles in the rhophecytosis invaginations. These findings indicate that the patient had poorly differentiated erythroid leukemia and that some of the clonogenic cells might respond to EPO in vitro. Corresponding to this biological feature, the leukemic cells were markedly decreased in number in response to repeated RBC transfusions, and partial remission was obtained. These observations suggest that erythroid leukemia distinct from erythroleukemia (M6) with a myeloblastic component, can develop as a minor entity of human acute leukemia.
Characterization of monoclonal IgG cryoglobulins: fine-structural and morphological analysis
Authors: Podell DN , Packman CH , Maniloff J , Abraham GN
Published: October 2019

The morphology of the amorphous, gelatinous, and crystalline varieties of monoclonal IgG cryoglobulins was analyzed by light and transmission and scanning electron microscopy. Each cryoglobulin had a characteristic fine structure that correlated with its gross morphology. Transmission electron microscopy showed that the amorphous precipitates were random and disorganized molecular clumps. In contrast, cryogels were thin-walled, well-organized, and hydrated strawlike clusters, whereas cryocrystals formed tightly compacted, highly structured molecular clusters. Crystals that formed in blood produced rouleaux, and analysis by scanning electron microscopy indicated that the crystals could form thick-walled, branching, macromolecular nets that could physically trap cells. The morphological properties provided visual impressions by which cryoglobulins could cause clinical disease secondary to vascular occlusion produced by self- associated IgG cryoglobulin molecules.
Control of hemopoiesis by a bone marrow stromal cell clone: lipopolysaccharide- and interleukin-1-inducible production of colony- stimulating factors
Authors: Rennick D , Yang G , Gemmell L , Lee F
Published: October 2019

A stromal cell line, GY30, was cloned from mouse bone marrow adherent cell layers. In culture, GY30 cells sustain the production of granulocyte-macrophage progenitor cells (GM-CFU) but fail to support the survival of pluripotential stem cells (CFU-S). GY30 cells secrete two growth factor activities distinct from interleukin-3 (IL-3), IL-2, and macrophage colony-stimulating factor (M-CSF) but functionally similar to GM-CSF and G-CSF. The production of both CSFs is increased 70- to 200-fold by treating GY30 cells with lipopolysaccharide or IL-1. RNA blot analysis reveals the presence of GM-CSF and G-CSF transcripts and demonstrates that IL-1 regulates the production of both factors at the mRNA level. Further, these studies show that the GM-CSF secreted by GY30 cells is structurally similar to the GM-CSF produced by activated T cells.
Lack of association between abnormalities of the chromosome 9 short arm and either "lymphomatous" features or T cell phenotype in childhood acute lymphocytic leukemia
Authors: Carroll AJ , Castleberry RP , Crist WM
Published: October 2019

In childhood acute lymphocytic leukemia (ALL), abnormalities in the short (p) arm of chromosome 9, particularly those leading to the loss of material in the p21–p22 region, may be associated with bulky disease at diagnosis (so-called “lymphomatous” ALL) and a T cell immunophenotype. To assess these associations further, we reviewed the clinical and laboratory data for 100 consecutively evaluated children with ALL who had successful cytogenetic studies. From analysis of clinical and laboratory features, 8 of the 100 patients were classified as having lymphomatous ALL. Seven of the 100 patients had aberrations involving the 9p arm or were missing an entire chromosome 9. The prevalence rate of 9p abnormalities in cases of lymphomatous ALL was not significantly different from that in cases without lymphomatous features (1 of 8 v 6 of 92, P = .62). Moreover, all seven patients with 9p abnormalities had the common ALL phenotype. These data suggest that although 9p abnormalities in childhood ALL occur frequently, there is no consistent association with either the occurrence of lymphomatous clinical features or the presence of T cell disease.