bluuebunny/crossref_metadata_2025 huggingface dataset:
Residual amounts of glycoprotein Ib concomitant with near-absence of glycoprotein IX in platelets of Bernard-Soulier patients
Authors: Drouin J , McGregor JL , Parmentier S , Izaguirre CA , Clemetson KJ
Published: October 2019

A study of the Bernard-Soulier syndrome in two unrelated families using different polyclonal antibodies in a sensitive immunoblot assay showed residual amounts of platelet membrane glycoprotein (GP) lb in the eight homozygotes, as well as the near-absence of GPlb beta and GPIX. The eight heterozygotes studied showed a double band pattern for GPlb and about half the normal level of GPlb beta and GPIX. Therefore, we conclude that the Bernard-Soulier syndrome is heterogeneous and is probably not due to gene deletions.
Recombinant granulocyte-macrophage colony-stimulating factor after autologous bone marrow transplantation for relapsed non-Hodgkin's lymphoma: blood and bone marrow progenitor growth studies. A phase II Eastern Cooperative Oncology Group Trial
Authors: Lazarus HM , Andersen J , Chen MG , Variakojis D , Mansour EG , Oette D , Arce CA , Oken MM , Gerson SL
Published: October 2019

Sixteen patients with relapsed non-Hodgkin's lymphoma underwent autologous bone marrow transplantation and infusion of recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF). Treatment consisted of involved-field radiotherapy, cyclophosphamide 60 mg/kg/d intravenously (IV) for 2 days, and fractionated total body irradiation (1,200 cGy). Autologous bone marrow was thawed and infused IV, followed 3 hours later by the first infusion of IV rhGM-CSF 11 micrograms/kg/d over 4 hours. Infusions of rhGM-CSF were continued daily until either both neutrophil count exceeded 1,500/microL and platelet count exceeded 50,000/microL, or until 30 days after marrow re- infusion. Toxicities encountered were mild and included fever, chills, hypertension, alopecia, rash, diarrhea, stomatitis, myalgias, and synovial (knee) effusions. Neutrophil recovery greater than 500/microL occurred a median of 14 days (range, 9 to 30 days) after marrow infusion, significantly earlier than in a comparable group of historic controls who recovered counts at a median time of 20 days (range, 12 to 51 days) (P = .00002). Median time to self-sustaining platelet counts greater than 20,000/microL was 23.5 days (range, 12 to 100 days), comparable with the historic group (P = .38). One bacteremia (central venous catheter exit site infection with Staphylococcus epidermidis) and one local infection (Giardia lamblia in stool) occurred. Patients received a median of 11.4 (range, 4.4 to 20.2) x 10(4) colony-forming unit granulocyte-macrophage (CFU-GM) progenitors per kg. Stem cell progenitors CFU-GM, CFU-granulocyte, erythroid, monocyte, megakaryocyte (CFU-GEMM), and burst-forming unit-erythroid (BFU-E) were detected in the bone marrow as early as 7 days after marrow re-infusion, and increased in proportion to peripheral blood counts, but by 30 to 60 days still remained much lower than before transplant. Neutrophils transiently decreased in 13 of 16 patients (median decrease, 42%) within 24 to 72 hours of discontinuing rhGM-CSF infusions. These data suggest that rhGM-CSF therapy enhances neutrophil recovery by forcing stem cells to produce mature elements at an enhanced rate but may not affect marrow stem cell and early progenitor population sizes.
Clonal development and karyotype evolution during leukemogenesis of BCR/ABL transgenic mice
Authors: Voncken JW , Morris C , Pattengale P , Dennert G , Kikly C , Groffen J , Heisterkamp N
Published: October 2019

The Philadelphia (Ph) translocation is responsible for the generation of the chimeric BCR/ABL oncogene. The Ph chromosome constitutes the earliest detectable chromosome abnormality in chronic myelogenous leukemia and is also found in acute lymphoblastic leukemia. Mice transgenic for a P190 BCR/ABL-producing DNA construct develop lymphoblastic leukemia/lymphoma and provide an opportunity to study early stages of the disease as well as progression. In this study, we have karyotyped the bone marrow of 10 19-day-old BCR/ABL P190 transgenic mice from a line that reproducibly develops leukemia/lymphoma. Leukemic cells from 17 terminally ill transgenic founders and progeny were also karyotyped as well as bone marrow transplant recipients of leukemic donor marrow. Karyotypically visible aberrations were absent from the early stages of BCR/ABL P190-generated leukemia and normal metaphases could be found even in the terminal stages of the disease. A high frequency of aneuploidy was found in advanced leukemia, with a marked preference for the gain of mouse chromosomes 12, 14, or 17. These results point to a primary role for BCR/ABL in leukemogenesis and suggest a destabilizing effect of the BCR/ABL gene on the regulation of cell division.
Autologous bone marrow transplantation in high-risk remission B-lineage acute lymphoblastic leukemia using a cocktail of three monoclonal antibodies (BA-1/CD24, BA-2/CD9, and BA-3/CD10) plus complement and 4- hydroperoxycyclophosphamide for ex vivo bone marrow purging
Authors: Uckun FM , Kersey JH , Haake R , Weisdorf D , Ramsay NK
Published: October 2019

Fourteen patients with high-risk B-lineage acute lymphoblastic leukemia (ALL) in complete remission underwent autologous bone marrow transplantation (BMT) using a combined immunochemopurging protocol. A monoclonal antibody (MoAb) cocktail of BA-1, BA-2, and BA-3 plus rabbit complement (C′) plus 4-hydroperoxycyclophosphamide (4-HC) was used to eliminate residual occult leukemia cells from autografts. All patients were conditioned with single-dose total body irradiation (TBI) followed by high-dose Ara-C. All 14 patients engrafted at a median of 24 days (range, 12 to 36 days). Three patients are alive and disease free at 3.5 years, 3.9 years, and 4.1 years post-BMT. The Kaplan-Meiser estimate and standard error of the probability of sustained remission was 23% +/- 12% at 3.5 years post-BMT with a mean relapse-free interval of 1.4 +/- 0.4 years. The disease-free survival (DFS) at 3.5 years was 21% +/- 11%, with a mean DFS time of 1.3 +/- 0.4 years. A novel and quantitative minimal residual disease (MRD) detection assay, which combines fluorescence-activated multiparameter flow cytometry and cell sorting with leukemic progenitor cell (LPC) colony assays, was used to analyze remission BM samples from B-lineage ALL patients for residual LPC, and to evaluate the efficacy of ex vivo BM purging. Notably, the minimal residual leukemia burden before BMT, as measured by the percentage of B-lineage LPC in the pre-BMT remission BM samples, indicated the outcome of the BMT. The median value for the minimal residual leukemia burden before BMT was 0.0035% (35 LPC/10(6) mononuclear cells). The Kaplan-Meier estimates and standard errors of the probability of remaining in remission after BMT were 43% +/- 19% for patients whose BM samples contained less than or equal to 0.0035% LPC and 0% +/- 0% for patients whose BM samples contained greater than 0.0035% B-lineage LPC (P less than .05). In contrast to the minimal residual leukemia burden measured by the described MRD assay system, the percentage of blasts or TdT+ cells in the remission BM samples did not correlate with the probability of relapse. The applied purging protocol showed variable success in destroying target B-lineage LPC populations contaminating the autografts. While in some cases purging was highly effective, eliminating up to greater than or equal to 4 logs of residual B-lineage LPC, in other cases only 0.1 to 0.2 logs of B- lineage LPC were purged.(ABSTRACT TRUNCATED AT 400 WORDS).
Monoclonal antibodies specific for low-affinity interleukin-3 (IL-3) binding protein AIC2A: evidence that AIC2A is a component of a high- affinity IL-3 receptor
Authors: Ogorochi T , Hara T , Wang HM , Maruyama K , Miyajima A
Published: October 2019

Mouse interleukin-3 (IL-3) binds to its receptor with high and low affinities. Using anti-Aic2 antibody, two distinct cDNAs (AIC2A and AIC2B) were isolated. The AIC2A gene encodes a protein of 120 Kd that binds IL-3 with low affinity, whereas the AIC2B gene encodes a protein that is 91% identical to AIC2A at the amino acid level, but which does not bind IL-3. To study the structure of the functional high-affinity IL-3 receptor (IL-3R), we generated specific monoclonal antibodies against the AIC2A protein. We produced a soluble AIC2A protein by inserting a termination codon at the beginning of the transmembrane domain of the AIC2A cDNA. Soluble AIC2A protein expressed in COS7 cells was purified to homogeneity and three anti-AIC2A monoclonal antibody- producing hybridomas (3D1, 3D4, and 9D3) were obtained from a rat immunized with the purified soluble AIC2A protein. The antibodies were specific for the AIC2A protein and did not bind to the AIC2B protein. Using chimeric receptors between AIC2A and AIC2B, recognition sites of the antibodies were mapped. The antibodies immunoprecipitated a 120-Kd protein from IL-3-dependent PT18 cells. The N-terminal sequence of the 120-Kd protein was consistent with the predicted processing site of the signal sequence of the AIC2A protein. Staining of IL-3-dependent and IL- 3-independent cell lines with the 9D3 antibody were consistent with the IL-3 binding. The 9D3 antibody inhibited both the high-affinity IL-3 binding and the low-affinity binding, as well as IL-3-dependent proliferation. These results indicate that the AIC2A protein is a binding component of a high-affinity IL-3R.
Kinetics of the appearance of Fc epsilon RI-bearing cells in interleukin-3-dependent mouse bone marrow cultures: correlation with histamine content and mast cell maturation
Authors: Rottem M , Barbieri S , Kinet JP , Metcalfe DD
Published: October 2019

While it is known that mast cells arise from pluripotential hematopoietic cells and express their mature phenotypes in tissues, the sequence of events in maturation is incompletely understood. To study early mast cells, we sorted cells from interleukin-3 (IL-3)-dependent mouse bone marrow cultures on the basis of Fc epsilon RI and examined their morphology, histamine content, and growth characteristics. Flow cytometric analysis and sort showed that the Fc epsilon RI-bearing (Fc epsilon RI+) cells increased from 0% on day 0 to 90% by day 21 and that the total number of Fc epsilon RI+ cells increased from 0 at the start of culture to 3.75 x 10(5) cells by day 21 from an initial population of 1 x 10(5) cells. The dissociation rate of 125I-labeled IgE from early cultured cells resembled the dissociation rate of mouse IgE from mature murine mast cells. Mean fluorescence intensity increased over time, reflecting an increase in IgE receptor density. Fc epsilon RI+ cells were also positive for Fc gamma RII/III. Morphologic studies showed gradual acquisition of metachromatic granules in the Fc epsilon RI+ cells, which was paralleled by an increase in histamine content. Sorted Fc epsilon RI+ cells, when placed in liquid suspension culture, gave rise to pure mast cell populations. Fc epsilon RI+ cells sorted at day 3 and cultured in agarose with IL-3 gave rise to 4,800 small and 150 medium-size mast cell colony-forming units per 10(6) cells, while Fc epsilon RI- cells gave rise to 23 medium-size and 49 large mast cell colony-forming units per 10(6) cells. Fc epsilon RI+ cells grown in granulocyte-macrophage colony-stimulating factor (CSF) or macrophage- CSF did not give rise to colony-forming units. These results show that Fc epsilon RI+ cells have proliferative potential, but that there also is a population of mast cell progenitor cells that have not yet expressed Fc epsilon RI, and such individual progenitor cells have greater potential for proliferation than cells that express Fc epsilon RI.
Cytokine regulation of colony-stimulating factor (CSF) production in cultured human synovial fibroblasts. II. Similarities and differences in the control of interleukin-1 induction of granulocyte-macrophage CSF and granulocyte-CSF production
Authors: Hamilton JA , Piccoli DS , Cebon J , Layton JE , Rathanaswani P , McColl SR , Leizer T
Published: October 2019

Synovial fibroblasts are likely to be a significant source of granulocyte-macrophage colony-stimulating factor (GM-CSF) and granulocyte-CSF (G-CSF), which could be crucial to the pathogenesis of rheumatoid arthritis. Using specific enzyme-linked immunosorbent assays (ELISAs) and Northern analysis, GM-CSF and G-CSF expression were followed in human synovial fibroblast-like cells in response to a number of agents, either alone or in the presence of an optimal stimulatory concentration of interleukin-1 (IL-1). For both CSFs, interferon-gamma (100 U/mL) did not increase their levels but dramatically suppressed the stimulatory action of IL-1, while basic fibroblast growth factor (10(-8) mol/L), although nonstimulatory by itself, potentiated IL-1 action. The glucocorticoid, dexamethasone (10(- 7) mol/L), inhibited IL-1-stimulated CSF production. However, evidence was obtained for noncoordinated CSF regulation. Cyclooxygenase inhibitors potentiated the action of IL-1 on GM-CSF synthesis but suppressed G-CSF synthesis, suggesting that endogenous cyclooxygenase products can have opposite effects in modulating the levels of each CSF. Also, the lymphokine, IL-4 (250 pmol/L), slightly inhibited GM-CSF formation in the presence of IL-1 but elevated the G-CSF levels in these cultures without having an effect by itself. Transforming growth factor beta (less than or equal to 20 ng/mL) did not modulate levels of either CSF. Mesenchymal cell production of both GM-CSF and G-CSF is generally viewed as being under coordinate control; our findings suggest that their synthesis in IL-1-stimulated human synoviocytes can be modulated by a number of agents, in some cases with divergent actions depending on which CSF is examined.
Disturbed platelet aggregation to collagen associated with an antibody against an 85- to 90-Kd platelet glycoprotein in a patient with prolonged bleeding time
Authors: Deckmyn H , Van Houtte E , Vermylen J
Published: October 2019

We studied a 5-year-old girl presenting with a markedly prolonged bleeding time. Her platelets were refractory to collagen stimulation, but the response to other agonists was normal. There were no coagulation abnormalities as measured by standard tests. Two- dimensional electrophoresis showed no abnormalities of the patient's platelet membrane glycoproteins. When the patient's plasma or purified plasma IgG was mixed with normal platelets, collagen-induced platelet aggregation was blocked. Western blotting showed the presence of an antibody in the patient's plasma directed against a protein of molecular weight 85 to 90 Kd under both reducing and nonreducing conditions. This protein comigrated with glycoprotein (GP) IV immunoprecipitated by OKM5 from 125I-labeled platelets. Immunoprecipitation of 125I-labeled normal platelet glycoproteins with the patient's IgGs also yielded an 85- to 90-Kd protein that migrated on the diagonal following nonreduced/reduced two-dimensional sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Despite similarities in electrophoretic behavior, the antigen was not demonstrated to be GPIV, since purified GPIV was not recognized by the antibody.
A Factor in Lungs of Anaphylactically Shocked Guinea Pigs Which Can Induce Eosinophilia in Normal Animals
Authors: SAMTER MAX , KOFOED MARTHA A. , PIEPER WILLIAM
Published: October 2019

1 . Absolute eosinophil counts of normal guinea pigs were obtained before, and twenty-four hours after, intraperitoneal implantation of lung slices from normal and sensitized guinea pigs, and of lung and other tissues—intestines, uterus, gall bladder, liver, and skin—from guinea pigs which died in anaphylactic shock. Changes in the number of circulating eosinophils of normal recipients were also determined after transfer of lung obtained from the same sensitized donor animal before and after reinjection of the specific antigen, and of lungs from shocked animals incubated at 56 C. for 40 minutes.

2. Transfer of lung tissue from normal and sensitized (but not reinjected) guinea pigs into the peritoneal cavity of normal animals produces a significant, but limited, increase in the number of circulating eosinophils of the recipients.

3. Transfer of lung tissue of guinea pigs which died in anaphylactic shock into the peritoneal cavity of normal recipients produces a marked transitory eosinophilia.

4. Removal and transfer of one lung of sensitized guinea pigs, compared with transfer of the other lung after reinjection of the specific antigen demonstrates that the eosinotactic factor develops immediately after anaphylactic shock.

5. Transfer of intestines, uterus, gall bladder, and liver (compared with lung) of shocked guinea pigs into the peritoneal cavity of normal animals produces a small, but significant, transitory increase in the number of circulating eosinophils in the recipient; transfer of skin in a limited number of experiments appears to depress the peripheral eosinophil count of the recipients.

6. Transfer of lung tissue from shocked donors incubated at 58 C. for 40 minutes produces a less marked eosinophilia in normal recipients than transfer of lung tissue kept at room temperature for the same period.
The Finding of the L. E. (Lupus Erythematosus) Cells in Smears of Untreated, Freshly Drawn Peripheral Blood
Authors: CHOMET BERNHARD , KIRSHEN MARTIN M. , SCHAEFER GERSHON , MUDRIK POLA
Published: October 2019

A case of lupus erythematosus is reported in which L.E. cells were found in the smears of freshly drawn peripheral blood.
Studies on the Mechanical Fragility of Erythrocytes
Authors: GOLDBLOOM RICHARD B. , FISCHER ENID , REINHOLD JOHN , HSIA DAVID YI-YUNG
Published: October 2019

1. The mechanical fragilities of incubated and nonincubated erythrocytes of normal newborn infants, children and adults have been determined through the use of a simplified rotator.

2. The mean mechanical fragility of nonincubated erythrocytes was 7.1 per cent for newborns, 4.0 per cent for children and adults; the mean mechanical fragility of incubated erythrocytes was 13.4 per cent for newborns, 10.5 per cent for children and 10.8 per cent for adults.

3. The possible relationship of increased mechanical fragility of erythrocytes in the newborn to increased hemolysis in the neonatal period is suggested.

4. Possible causes of variation in the determination of mechanical fragility of incubated erythrocytes are discussed.
Antifibrinolytic Activity in Plasma after Exposure to Cold and after Chymotrypsin Administration
Authors: JENSEN H. , CHAMOVITZ DAVID L. , VOLKRINGER EVELYN , GRAY ELIZABETH , GOSNEY JAMES E. , KOCHOLATY WALTER
Published: October 2019

Exposure of rats and of human subjects to cold was associated with an elevation of the antifibrinolytic activity of blood. Administration of chymotrypsin to rats and rabbits caused a decrease in the antifibrinolytic activity of blood The possible significance of these and earlier observations on the fibrinolysis antifibrinolysis system in blood is discussed.
The Adrenal Cortex and Hemolysis
Authors: FELDMAN J. D. , RACHMILEWITZ M. , STEIN O. , STEIN Y.
Published: October 2019

The intravenous administration of phenylhydrazine caused a significantly greater destruction of red cells in adrenalectomized rats than in controls. Cortisone given to adrenalectomized rats prevented this increased destruction and maintained red cells at a level comparable to controls. ACTH almost completely prevented phenylhydrazine anemia in intact rats and the red cell level was significantly higher than that in intact rats which did not receive ACTH.
Electron Microscopic Studies of the Action of Thrombin on Blood Platelets
Authors: DE ROBERTIS E. , PASEYRO P. , REISSIG MAGDALENA
Published: October 2019

The action of thrombin on blood platelets has been studied under the electron microscope. In control experiments with preparations treated with normal plasma or Tyrode fluid, the normal structure of platelets in their different forms of extension and flattening on the film has been observed.

Thrombin (Parke-Davis Co.) in concentrations of 25 and 50 units per ml. has a lytic effect on platelets. The number of platelets affected by this treatment increases in the period of 5 to 30 minutes.

Disintegration proceeds in the hyalomere from the periphery to the center with the formation of long fibrillar processes constituted by microvesicles. These processes resemble myelin figures. The granulomere turns into an amorphous dense body which also undergoes lysis. The final residue of disintegration is constituted by microvesicular material and dense amorphous bodies.

The purified bovine thrombin of Seegers has exactly the same action. The lytic effect is observed with concentrations of thrombin as low as 2.5 units per ml. The number of platelets undergoing disintegration increases proportionally to the logarithm of the thrombin concentration. The probable role of thrombin and the lysis of platelets in normal blood clotting is discussed.
Hemophilia-like Disease Following Pregnancy
Authors: FRICK PAUL G.
Published: October 2019

1. A hemophilia-like disease acquired during the first pregnancy of a young woman, is described.

2. The clotting abnormality was caused by a circulating anticoagulant which, in all probability, inhibited the antihemophilic globulin.

3. The anticoagulant was demonstrated in the patient's second child during his first two and one-half months of life.

4. The transplacental transfer of the anticoagulant and the analogy of this condition to hemophilia (resistant to therapy) following iso-immunization against antihemophilic globulin, strongly suggest that an immunologic mechanism accounts for the development of this abnormality.

5. The patient recovered eighteen months after her second delivery and sixteen months after x-ray sterilization. The possible relation between endocrine factors and recovery is discussed.
Bone Marrow Aspiration in the Monkey (Macacus Rhesus)
Authors: COHEN BERNARD STANLEY
Published: October 2019

1. A simple method suitable for repeated bone marrow aspirations in the unanesthetized monkey has been described.

2. Data have been presented as to the myelogram of 25 normal Macacus rhesus monkeys.
Inhibition of human B-cell lymphoma growth by CD40 stimulation
Authors: Funakoshi S , Longo DL , Beckwith M , Conley DK , Tsarfaty G , Tsarfaty I , Armitage RJ , Fanslow WC , Spriggs MK , Murphy WJ
Published: October 2019

CD40 is a molecule present on B lymphocyte lineage cells that is important in B-cell differentiation and activation. Signaling through CD40 has been shown to exert costimulatory signals on normal B cells resulting in proliferative and differentiation responses. Examination of several B-cell lymphomas showed cell-surface expression of the CD40 molecule. Incubation of these lymphomas with anti-CD40 antibodies resulted in significant growth inhibition in vitro. Cross-linking of the CD40 antibodies resulted in even greater inhibition of proliferation. A recombinant soluble human CD40 ligand was also shown to inhibit lymphoma proliferation. When various human B-cell lymphomas were transferred into mice with severe combined immune deficiency, the treatment of the mice with anti-CD40 antibodies resulted in significant increases in survival showing that anti-CD40 is efficacious after in vivo administration. Thus, CD40 stimulation by either the antibody or soluble ligand directly inhibits human B-cell lymphoma growth and therefore, may be of significant clinical use in their treatment.
Hemoglobin enhances the production of tissue factor by endothelial cells in response to bacterial endotoxin
Authors: Roth RI
Published: October 2019

Human endothelial cells respond to bacterial endotoxin (lipopolysaccharide [LPS]) with changes that transform the endothelium into a surface with prominent procoagulant properties. Production of tissue factor (TF) in response to LPS is a major alteration that favors coagulation. Biologic activities of LPS have previously been shown to be enhanced by the presence of hemoglobin. Therefore, the ability of human hemoglobin (Hb) to modulate TF production by cultured human umbilical vein endothelial cells (HUVEC) was investigated. Cell-free Hb (10 mg/mL), either purified native (HbAo) or chemically cross-linked (alpha alpha Hb), was incubated with LPS (0.1 microgram/mL), and the mixtures then were added to HUVEC in culture. TF activity was quantified with a clotting assay and TF protein was measured with an enzyme-linked immunosorbent assay. Hb preparations greatly enhanced the production of TF activity (11- to 25-fold greater than TF produced by HUVEC alone) compared with minimal TF activity generated by LPS alone (only twofold greater than HUVEC alone). The enhancement of LPS-induced TF activity was Hb concentration-dependent over a range of 1 to 100 mg/mL. Cross-linked alpha alpha Hb also greatly enhanced the production of TF protein compared with TF protein generated by LPS alone (12-fold greater v 3.5-fold greater than HUVEC alone, respectively). The enhancement of LPS-induced TF protein was Hb concentration-dependent over a range of 0.1 to 2 mg/mL. Enhancement of TF activity by Hb required new protein synthesis. These results show that human Hb can augment the ability of LPS to induce endothelial cell TF and suggest that hemolysis associated with disseminated intravascular coagulation during sepsis may further stimulate coagulation. In addition, these results suggest a potential mechanism for generalized thrombosis in animals that has been associated with the infusion of cell-free Hb for resuscitation.
Functional and binding studies of the roles of prothrombin and beta 2- glycoprotein I in the expression of lupus anticoagulant activity
Authors: Permpikul P , Rao LV , Rapaport SI
Published: October 2019

We present functional and binding data relevant to the reported roles for prothrombin and beta 2-glycoprotein I (beta 2GPI) in the expression of lupus anticoagulant activity. In a purified system containing human prothrombin, Xa, Va, and a rate-limiting concentration of phosphatidylserine (PS)/phosphatidylcholine (PC) vesicles, the preliminary incubation of vesicles with protein A separated IgG preparations from 10 lupus anticoagulant plasmas, calcium, and prothrombin enhanced the inhibitory effect of all IgG preparations upon thrombin generation. Experiments in a purified factor X activation system provided supporting data that a similar preliminary incubation with prothrombin enhanced the inhibitory effect of many of the IgG preparations upon factor X activation. However, we could not obtain unequivocal evidence that prothrombin was an obligatory cofactor for lupus anticoagulant IgG to inhibit procoagulant phospholipid function, because lupus anticoagulant IgG separated by protein A chromatography contained traces of prothrombin. The binding of many IgG preparations to immobilized PS was enhanced by prothrombin when calcium ions were present. beta 2GPI enhanced binding of many of the IgG preparations to immobilized PS both in the presence and absence of calcium, yet beta 2GPI failed to enhance the ability of the IgG preparations to inhibit phospholipid function in purified prothrombin and factor X assays. Moreover, the IgG preparations prolonged the dilute Russell's viper venom time (dRVVT) of beta 2GPI-depleted normal plasma. Nine of 10 IgG preparations bound to prothrombin on Western blots in the absence of calcium and phospholipid, whereas no preparation bound to beta 2GPI. Passage of five citrated lupus anticoagulant plasmas through a prothrombin affinity column in the absence of added calcium and phospholipid removed most of the activity prolonging the dRVVT of normal plasma, and IgG in the pass-through plasma no longer bound to PS in the presence of prothrombin and calcium ions. IgG in prothrombin column eluates had strikingly enhanced specific lupus anticoagulant activity and also specific PS binding activity in the presence of prothrombin and calcium ions. Thus, lupus anticoagulant plasmas were shown to contain IgG binding to prothrombin, in the absence of calcium ions and phospholipid, which could also, in the presence of calcium ions and prothrombin, bind to PS and express lupus anticoagulant activity.
2-chlorodeoxyadenosine induces durable remissions and prolonged suppression of CD4+ lymphocyte counts in patients with hairy cell leukemia
Authors: Seymour JF , Kurzrock R , Freireich EJ , Estey EH
Published: October 2019

A number of effective treatments are available for patients with hairy cell leukemia (HCL). 2-Chlorodeoxyadenosine (2-CdA) induces more than 80% complete responses, but is associated with profound suppression of CD4+ lymphocyte counts. However, the duration of each is uncertain. We have analyzed a previously reported cohort of 40 patients who had responded to 2-CdA. Eight patients (20%) have relapsed at a median of 16 months (range, 3 to 23 months). The remaining 32 patients were observed for a median of 30 months (range, 7 to 43 months). No patients have died. At 3 years, the actuarial disease-free survival rate is 77% (95% confidence interval, 70% to 84%). The median CD4+ lymphocyte count before therapy was 743/microL (range, 58 to 2,201/microL). The median CD4+ nadir after treatment was 139/microL (range, 25 to 580/microL). There was a single opportunistic infection and no second malignancies observed. Although there was evidence of some improvement in CD4+ lymphocyte counts on sequential testing, CD4+ counts remained significantly lower than baseline (P < .0001) at a median of 23 months after therapy (median, 237/microL; range, 25 to 514/microL), and were also lower than baseline (P < .002) in those patients with more than 1 year of follow-up (median, 27 months; range, 13 to 42 months). The median time to reach an absolute CD4+ lymphocyte count of 365/microL, the lower limit of the normal range, was 40 months. Although responses to 2-CdA are durable in the majority of patients with HCL, the uncertain long-term consequences of the observed CD4+ lymphocytopenia suggest caution in the broad application of this therapy.
Adhesion of sickle cells to vascular endothelium is critically dependent on changes in density and shape of the cells
Authors: Kaul DK , Chen D , Zhan J
Published: October 2019

Experiments were performed to explore whether adhesion differences among different density sickle (SS) cells are mainly caused by deformability characteristics attendant to density variations, which could affect their surface contacts with the endothelium, or caused by any changes in their membrane surface (eg, receptors). Also, what is the contribution of morphologic attributes of SS cells in adhesive and obstructive events? To resolve these issues, controlled modifications of cell density were performed using Nystatin and sucrose. This allowed elevation of the mean corpuscular hemoglobin concentration (MCHC) of light-density SS2 discocytes to that of SS4 native dense cells, as well as its normalization in both native and artificially dehydrated cells. Hemodynamic and adhesive characteristics of these individual SS cell populations and their defined mixtures were investigated in the ex vivo mesocecum vasculature. Dehydrated (high MCHC) SS2 discocytes, alone or with SS2 control cells, caused a higher peripheral resistance but no persistent microvascular blockage. Nevertheless, dehydrated SS2 discocytes resulted in a significantly decreased adhesion in venules (adhesion sites). Rehydration of these high MCHC discocytes completely restored their adhesivity, similar to that of SS2 control cells. In contrast, irreversibly sickled cell (ISC)-rich SS4 dense cell class (dense discocytes and 66% to 72% ISCs) was less adherent than dehydrated SS2 discocytes, but caused a persistent blockage of small diameter postcapillary venules when infused with SS2 control or dehydrated discocytes. In the areas of adhesion, SS4 dense discocytes outnumbered ISCs by 4 to 1, demonstrating that ISCs, because of their shape characteristics, are minimally adherent but play a distinct role in postcapillary obstruction. When MCHC (density) of SS4 cells was decreased by rehydration, it resulted in an almost complete reversal of their hemodynamic and adhesive behavior, confirming a profound influence of cell density. These findings demonstrate for the first time that, under shear flow, adhesion of different density SS cells to the endothelium is dependent on the inverse of cell density rather than any changes in their adhesion potential. Finally, the trapping of ISC- rich native dense cells, but not of dehydrated discocytes, in the areas of adhesion shows a distinct contribution of ISC in adhesion-initiated vasoocclusion.
Flt-3 Ligand Synergizes With Granulocyte Colony-Stimulating Factor to Increase Neutrophil Numbers and to Mobilize Peripheral Blood Stem Cells With Long-Term Repopulating Potential
Authors: Molineux Graham , McCrea Clay , Yan Xiao Qiang , Kerzic Patrick , McNiece Ian
Published: October 2019

Flt-3 ligand (FL) shares many features with stem cell factor (SCF), a widely documented cofactor for peripheral blood progenitor cell (PBPC) mobilization. We investigated the mobilization of PBPCs by FL in combination with granulocyte colony-stimulating factor (G-CSF). As a single agent, FL was a relatively modest mobilizer of PBPCs, resulting in 360 granulocyte/macrophage colony-forming cells (GM-CFCs)/mL blood (control, 155 GM-CFCs/mL blood) and no advantage in leukocyte recovery when these PBPCs were transplanted to irradiated recipient mice. G-CSF, on the other hand, mobilized over 20,000 GM-CFCs/mL blood, and the combination of G-CSF + FL resulted in over 100,000 GM-CFCs/mL blood. The combination of G-CSF + FL stimulated increased levels of monocytes and basophils in the peripheral blood. The performance of the mobilized PBPC product in irradiated hosts correlated with progenitor numbers resulting in long-term engraftment in association with accelerated short-term recovery of both leukocytes and platelets. These data demonstrate the potential of FL to synergize with G-CSF to mobilize PBPCs with both short- and long-term engraftment potential. The effect is similar to the synergistic interaction of G-CSF and SCF on PBPC mobilization. The use of FL as opposed to SCF may elicit a different spectrum of toxicities including lymphoid proliferation effects, in contrast to the mast cell degranulation effects of SCF. Clinical studies of FL are needed to evaluate its usefulness in man.
Activation of Human T Cells by Major Histocompatability Complex Class II Expressing Neutrophils: Proliferation in the Presence of Superantigen, But Not Tetanus Toxoid
Authors: Fanger Neil A. , Liu Chunlei , Guyre Paul M. , Wardwell Kathleen , O'Neil Jerome , Guo Tai L. , Christian Todd P. , Mudzinski Stanley P. , Gosselin Edmund J.
Published: October 2019

The primary function of polymorphonuclear neutrophils (PMN) in the immune response appears to be acute phagocytic clearance of foreign pathogens and release of inflammatory mediators. Consistent with their assumed lack of major histocompatibility complex (MHC) class II expression, PMN have not been considered to play a role in antigen presentation and T-cell activation. However, recent reports have shown that human PMN can express MHC class II molecules both in vitro and in vivo after stimulation with either granulocyte-macrophage colony-stimulating factor (GM-CSF ) or interferon-γ (IFN-γ). Thus, under appropriate conditions, PMN could play a significant role in immune regulation, including T-cell activation. In this report, we demonstrate that human class II–expressing PMN can serve as accessory cells in superantigen (SAg)-mediated T-cell activation. This accessory activity for SAg presentation was present only after induction of MHC class II expression, and was especially pronounced following culture of PMN with GM-CSF plus IFN-γ, which acted synergistically to induce MHC class II molecules on PMN. Moreover, the level of MHC class II expression and the magnitude of SAg-induced T-cell responses were found to be highly correlated and distinctly donor dependent, with PMN from some donors repeatedly showing fivefold higher responses than PMN from other donors. On the other hand, culture of PMN with GM-CSF plus IFN-γ under conditions that resulted in optimal MHC class II expression did not enable them to function as antigen-presenting cells for either intact tetanus toxoid (TT) or for a TT peptide. These results delineate a new pathway for T-cell activation by SAg that may play an important role in the severity of SAg-induced inflammatory responses. They also identify a donor-specific polymorphism for induction of PMN MHC class II expression which may be of significance for therapies involving GM-CSF and IFN-γ.
Characterization of Interleukin-10 Receptor Expression on B-Cell Chronic Lymphocytic Leukemia Cells
Authors: Jurlander Jesper , Lai Chun-Fai , Tan Jimmy , Chou Chuan-Chu , Geisler Christian H. , Schriber Jeffrey , Blumenson Leslie E. , Narula Satwant K. , Baumann Heinz , Caligiuri Michael A.
Published: October 2019

B-cell chronic lymphocytic leukemia (B-CLL) cells accumulate in vivo in the G0/G1 phase of the cell cycle, suggesting that their malignant expansion is due, at least in part, to a delay in cell death. However, the cellular or molecular factors responsible for a delay in B-CLL cell death are unknown. B-CLL cells do express receptors for interferon-α (IFN-α) and IFN-γ, and activation of both has been shown to promote B-CLL survival in vitro by preventing apoptosis. The interleukin-10 (IL-10) receptor is another member of the IFN receptor family, but its ligand, IL-10, has been reported to induce apoptosis in B-CLL cells. In the current study, we undertook a biochemical analysis of IL-10 receptor expression on freshly isolated B-CLL cells and characterized the functional responsiveness of IL-10 binding to its constitutively expressed receptor. We show that B-CLL cells bind IL-10 with significant specificity and express between 47 and 127 IL-10 receptor sites per cell, with a dissociation constant in the range of 168 to 426 × 10−12 mol/L. Ligand binding and activation of the IL-10 receptor expressed on B-CLL cells results in the phosphorylation of signal transducer and activator of transcription 1 (STAT1) and STAT3 proteins. This pattern of STAT protein phosphorylation is identical to IL-10 receptor activation on normal cells and similar to IFN-α (STAT1 and STAT3) and IFN-γ (STAT1) receptor activation in CLL. Further, in consecutive samples of fresh blood obtained from patients with B-CLL cells, the addition of IL-10 inhibited B-CLL proliferation, enhanced B-CLL differentiation, but did not induce apoptosis. Indeed, IL-10, like IFN-γ, was able to significantly reduce the amount of B-CLL cell death caused by hydrocortisone-induced apoptosis. We conclude that cytokines, which signal through the interferon family of receptors, have comparable functional effects on B-CLL cells.
Erythropoietin Induces Tyrosine Phosphorylation of the Interleukin-3 Receptor β Subunit (βIL3 ) and Recruitment of Stat5 to Possible Stat5-Docking Sites in βIL3
Authors: Chin Hiroshi , Wakao Hiroshi , Miyajima Atsushi , Kamiyama Ryuichi , Miyasaka Nobuyuki , Miura Osamu
Published: October 2019

The receptors for erythropoietin (Epo) and interleukin-3 (IL-3) both induce the ligand-dependent activation of the Jak2 tyrosine kinase. Activated Jak2 then phosphorylates these receptors and thereby recruits various signaling molecules containing the Src homology (SH)-2 domain, including Stat5, to the tyrosine phosphorylated receptors. In the present study, we demonstrate that Epo stimulation induces unidirectional cross-phosphorylation of the IL-3 receptor β subunit (βIL3) on tyrosines and its rapid and transient association with Stat5 in murine IL-3–dependent cell lines engineered to express the Epo receptor (EpoR). Using cell lines expressing various EpoR mutants, it was demonstrated that the Epo-induced tyrosine phosphorylation of βIL3 is dependent on the membrane-proximal EpoR cytoplasmic region involved in the activation of Jak2, but not on the extracellular and transmembrane regions or on the carboxy-terminal 145 amino acid region containing all the intracellular tyrosine residues. It was also shown that IL-3 induces rapid and dose-dependent association of Jak2 with βIL3. However, Epo failed to induce any detectable association of βIL3 with Jak2 or the EpoR. The present study also demonstrates that in IL-3–stimulated cells, an ovine Stat5 mutant harboring a substitution of Tyr694 to Phe, which abolishes the tyrosine phosphorylation required for activation, fails to dimerize with endogenous Stat5, shows sustained binding with tyrosine-phosphorylated βIL3, and inhibits the tyrosine phosphorylation of endogenous Stat5. These results suggest that βIL3 may have Stat5 docking sites, similar to those found in the EpoR, that facilitate the activation of Stat5 by Jak2 and raise the possibility that Epo may cross-activate or transmodulate the IL-3 receptor signaling pathways.
The Carbohydrate Moiety of Factor V Modulates Inactivation by Activated Protein C
Authors: Fernández José A. , Hackeng Tilman M. , Kojima Kazuhisa , Griffin John H.
Published: October 2019

An important risk factor for thrombosis is the polymorphism R506Q in factor V that causes resistance of factor Va to proteolytic inactivation by activated protein C (APC). To study the potential influence of the carbohydrate moieties of factor Va on its inactivation by APC, factor V was subjected to mild deglycosylation (neuraminidase plus N-glycanase) under nondenaturing conditions. The APC resistance ratio values (ratio of activated partial thromboplastin time [APTT] clotting times with and without APC) of the treated factor V were increased (2.4 to 3.4) as measured in APTT assays. O-glycanase treatment of factor V did not change the APC resistance ratio. The procoagulant activity of factor V as well as its activation by thrombin was not affected by mild deglycosylation. Treatment of factor V with neuraminidase and N-glycanase mainly altered the electrophoretic mobility of the factor Va heavy chain, whereas treatment with O-glycanase changed the mobility of the connecting region. This suggests that the removal of the N-linked carbohydrates from the heavy chain of factor Va, which is the substrate for APC, is responsible for the increase in susceptibility to inactivation by APC. Thus, variability in carbohydrate could account for some of the known variability in APC resistance ratios, including the presence of borderline or low APC resistance ratios among patients who lack the R506Q mutation.
Persistence of Antibody to Human Parvovirus B19 After Allogeneic Bone Marrow Transplantation: Role of Prior Recipient Immunity
Authors: Ang H.A. , Apperley J.F. , Ward K.N.
Published: October 2019

Human parvovirus B19 (B19) IgG was studied retrospectively in 66 allogeneic bone marrow transplantation (BMT) patients using an enzyme-linked immunosorbent assay. Recipient and donor sera had been stored pre-BMT together with sequential sera thereafter. Approximately half of donors and recipients had anti-B19 IgG pre-BMT and thus the relative contributions of donor and recipient immunity to antibody production after transplantation could be assessed. For each patient, a serum taken 2 to 3 years after BMT was also tested and the results show that persistence of B19 antibody depends on prior recipient (P = .0003) but not on donor immunity (P = .8). The findings were similar in both sibling and (VUD) BMT volunteer unrelated donor patients. Analysis of sequential post-BMT sera from 41 of the patients, for whom appropriately timed samples were available, showed primary B19 infection in 3 seronegative individuals, whereas 5 others who were seropositive before BMT underwent recurrent infection. Sequential results from the remaining 33 patients without recent B19 infection showed no evidence for donor antibody transfer and confirmed that antibody persistence depends on prior recipient immunity. B19 IgG levels decreased variably with time and some patients eventually became seronegative. It is concluded that this long-term persistence of B19 antibody post-BMT is most probably due to the existence of long-lived recipient plasma cells.
Tumor Necrosis Factor-α– and Interleukin-6–Triggered Mast Cell Development From Mouse Spleen Cells
Authors: Hu Zhi-Qing , Kobayashi Kazuo , Zenda Naoko , Shimamura Tadakatsu
Published: October 2019

We previously showed that interleukin-3 (IL-3) alone is not sufficient, although it is essential for murine mucosal-type mast cell development and that prostaglandin E (PGE) and interferon-γ (IFN-γ) are critical for survival or differentiation of mast cell precursors. We also confirmed that IL-4 is a key inhibitor for mast cell precursors despite being a growth factor of mast cells. In the present work, mouse spleen cells were cultured with recombinant (r) IL-1β, rIL-5, rIL-6, rIL-9, granulocyte-macrophage colony-stimulating factor (GM-CSF ), stem cell factor (SCF ), tumor transforming growth factor-β (TGF-β), or tumor necrosis factor-α (TNF-α) in the presence of endogenous IL-3. After 12 days of culture, mast cell development was induced by rIL-6 and rTNF-α. rIL-1β, rIL-5, rGM-CSF, rTGF-β and even the mast cell growth factors, rIL-9 and rSCF, failed to induce mast cell development. However, unlike IL-9 and SCF, IL-6 and TNF-α did not promote the growth of mast cells already developed. Macrophage may be one of the responsive cells of IL-6 and TNF-α in the cultures, because removal of macrophages greatly reduced the mast cell development induced by the cytokines. The actions of TNF-α and IL-6 were inhibited by indomethacin, an inhibitor for prostaglandin synthesis, and by neutralizing anti–IFN-γ and anti–IL-3 antibodies. rIL-4, when added at the start of the culture, also inhibited mast cell development induced by rIL-6 and rTNF-α. Nevertheless, neutralizing anti–IL-6 and anti–TNF-α antibodies did not suppress mast cell development induced by PGE and IFN-γ. TNF-α and IL-6 enhanced IFN-γ production, but suppressed IL-4 production in the cultures. Mast cell numbers induced were inversely and directly proportional to IL-4 and IFN-γ levels, respectively. These results indicate that inflammatory mediators as triggers are important for mast cell development, although they are not the mast cell growth factors.
Differential Susceptibility to HIV-GP120–Sensitized Apoptosis in CD4+ T-Cell Clones With Different T-Helper Phenotypes: Role of CD95/CD95L Interactions
Authors: Accornero Paola , Radrizzani Marina , Delia Domenico , Gerosa Franca , Kurrle Roland , Colombo Mario P.
Published: October 2019

The susceptibility of Th1 and Th2 cell clones to apoptosis following HIV-gp120/CD4 cross-linking and TCR activation was investigated. We show that only Th1 clones are susceptible to HIV-gp120-sensitized apoptosis, although both types of clones express similar levels of CD4 and bind similar amounts of recombinant gp120. Both types of clones, however, undergo apoptosis induced by CD95 cross-linking with agonistic monoclonal antibody (MoAb). Apoptosis induced by gp120 in the Th1 clones is inhibited by either an antiCD95 neutralizing MoAb or an anti-CD95L neutralizing MoAb as well as by a specific interleukin-1β converting enzyme (ICE) inhibitor. When triggered to apoptosis by gp120, Th1 but not Th2 clones express both cell-associated and soluble CD95L. The CD95L produced by Th1 clones induces cell death, inhibitable by anti-CD95 neutralizing MoAb, of CD95 positive Jurkat cells. These data suggest that, like activation-induced apoptosis, HIV-gp120 sensitized apoptosis in Th1 clones occurs via CD95/CD95L interaction and that lack or insufficient production of CD95L is responsible, at least in part, for the resistance of Th2 clones to such apoptosis.