bluuebunny/crossref_metadata_2025 huggingface dataset:
Eosinophilic cytoplasmic inclusions in fetal leukocytes: are Auer bodies a recapitulation of fetal morphology?
Authors: Newburger PE , Novak TJ , McCaffrey RP
Published: October 2019

Among the most striking morphological features of acute nonlymphoblastic leukemias (ANLL) is the occurrence of eosinophilic cytoplasmic inclusions known as Auer rods on Auer bodies. We examined immature myeloid cells from the peripheral blood of 9 human fetuses of 16–19 wk gestation for the presence of such structures. Five of these 9 samples contained cytoplasmic inclusions, which were identical to the Auer rods typically seen in blast cells from patients with ANLL. The incidence of positive cells was low (1–5 cells/10,000 cells surveyed). The inclusions were azurophilic with Wright-Giemsa staining and were cytochemically positive with peroxidase, acid phosphatase, and Sudan black staining. We observed no inclusions in identically prepared control myeloid cells from the bone marrow of 5 patients with acute lymphoblastic leukemia in remission and 3 patients with chronic myelogenous leukemia in stable phase. Nor were they present in peripheral blood myeloid cells of 10 normal adults. Myeloid precursors in long-term bone marrow culture from 2 normal adult donors did not develop the inclusions during 24 hr of incubation with prostaglandin F2 (the abortifacient). These observations suggest that Auer rod formation is an occasional but normal phenomenon in fetal hematopoiesis.
Selective expression of the common acute lymphoblastic leukemia (gp 100) antigen on immature lymphoid cells and their malignant counterparts
Authors: Greaves MF , Hariri G , Newman RA , Sutherland DR , Ritter MA , Ritz J
Published: October 2019

The selectivity of monoclonal antibody J-5 (anti-gp 100, common ALL antigen) for normal and leukemic hemopoietic cells has been investigated. J-5 gave concordant reactions with rabbit anti-cALL, coredistributed on the cell surface, and precipitated a similar if not identical glycoprotein from leukemic and normal tissue. Normal, immature lymphoid cells reactive with J-5 were detected in bone marrow and in fetal thymus. In marrow they were largely coincident with the TdT+ population. J-5 defines a major subgroup of ALL (common ALL) with a favorable prognosis. Of 853 non-ALL acute leukemias investigated, 80 were J-5 positive. These included 14 cases diagnosed as AML, 51 TdT+ blast crises of CGL, and 15 cases diagnosed as “AUL.” Of the 14 J-5+ AML, 13 were subsequently rediagnosed either as cALL (10 cases) or mixed lymphoid-myeloid leukemias (3 cases). One-hundred forty-three cases of mature lymphoid cell leukemia (91 B, 52 T) were investigated with J-5; 3 cases only, of disseminated B lymphoma, were positive, albeit weakly. A higher proportion of follicular lymphomas are, however, J-5 positive when studied in sections of biopsy material. A similar pattern of selective reactivity was observed in a series of leukemia/lymphoma cell lines. These studies emphasize the diagnostic value of monoclonal anti-cALL reagents.
Isolation of erythropoietin-sensitive cells from Friend virus-infected marrow cultures: characteristics of the erythropoietin response
Authors: Bondurant M , Koury M , Krantz SB , Blevins T , Duncan DT
Published: October 2019

Murine erythroid precursor cells, stimulated to proliferate in vitro in the absence of added erythropoietin (EP) by the anemia strain of Friend virus (FVA), will subsequently respond to EP by complete erythrocyte differentiation. If not exposed to EP, the erythroid cells divide for about 120 hr in culture, and they maintain the potential for full differentiation in response to EP added at any time during the period from 72 to 120 hr. Between 96 and 120 hr of culture without added EP, the EP-sensitive erythroid precursor cells that have formed discrete erythroid bursts can be isolated in relatively large numbers from such cultures by plucking with a Pasteur pipette. The addition of EP initiates the final stages of erythroid differentiation, including heme synthesis in 70%-80% of these isolated cells. With respect to homogeneity of the precursor cells, quantity of EP-responsive cells obtainable, and uniformity of EP responsiveness, this system is uniquely favorable for biochemical studies of the late differentiation effects of EP. The overall changes in gene expression accompanying EP- induced terminal differentiation were examined by two-dimensional gel electrophoresis of proteins labeled for a short time with radioactive amino acids. Several new proteins are synthesized in these erythroid cells during terminal differentiation, but the number is a very small percentage of the total number of proteins being made. Thus, in this system, the effect of EP is to initiate expression of a small group of genes, including those for globins, spectrin, and other proteins involved in the final stages of erythroid differentiation.
No stimulative effect of adipocytes on hematopoiesis in long-term human bone marrow cultures
Authors: Touw I , Lowenberg B
Published: October 2019

Long-term cultures of human bone marrow were established for 5–13 wk to study the role of adipocytes in sustaining hematopoiesis. At weekly intervals, the numbers of nucleated cells and granulocyte-macrophage progenitor cells (GM-CFU) in culture were estimated in relation to the numbers of fat-containing cells present in the adherent stroma layer. In these quantifications, the numbers of GM-CFU trapped in the adherent cell layer were considered separately. It was found that the presence of adipocytes did not correlate with more active hematopoiesis. Fat cells appeared at late stages when successful cultures were being exhausted or early in cultures with poor activity. These observations suggest that human marrow continuous hematopoiesis in vitro, unlike hematopoiesis in the analogous murine bone marrow cultures, does not depend on the presence of adipocytes.
Lysis of leukemia and lymphoma cells by autologous and allogeneic interferon-activated blood mononuclear cells
Authors: Oshimi K , Oshimi Y , Motoji T , Kobayashi S , Mizoguchi H
Published: October 2019

Studies were undertaken to determine whether leukemia and lymphoma cells would be lysed by autologous and allogeneic interferon (IFN) activated peripheral blood mononuclear cells (PBMC). PBMC from healthy donors and from patients were cultured with and without 500 U of highly purified human fibroblast IFN/ml for 24 hr, and then their cytotoxic activity was assayed by a 5-hr 51Cr-release test. Of primary tumor cells isolated from patients, the cells of 5 of 15 patients with acute nonlymphocytic leukemia (ANLL), 5 of 9 patients with acute lymphocytic leukemia (ALL), 2 of 3 patients with chronic phase chronic myelogenous leukemia (CML), 2 of 3 patients with blastic phase CML, 1 patient with hairy cell leukemia, and 6 patients with diffuse non-Hodgkin's lymphoma were sensitive to IFN-activated PBMC of healthy donors, whereas the cells of 3 of the ANLL patients, 2 of the ALL patients, and 3 of the lymphoma patients were sensitive to unstimulated PBMC. Of the ANLL cells tested, myeloblasts, promyelocytes, and monoblasts were sensitive to either unstimulated or IFN-activated PBMC. Compared with the ANLL cells, the lymphoma cells were statistically significantly sensitive to activated effector cells (p less than 0.025). On the basis of the unlabeled target competition test and the recovery of cytotoxic cells within the fractions enriched in natural killer (NK) cells, NK cells appeared to mediate the above unstimulated and IFN-boosted cytotoxicity. In experiments using autologous effector-target cells from 11 patients, the addition of 500 U of IFN/ml enhanced the lytic activity of PBMC against autologous lymphoma cells in 1 patient, and higher concentrations of IFN, i.e., 2500 or 3500 U/ml, enhanced their cytotoxic activity against autologous leukemia or lymphoma cells in 4 of 8 patients. These data indicate that IFN-activated allogeneic PBMC are able to lyse both myeloid and lymphoid tumor cells, whereas higher concentrations of IFN are required to enhance lytic activity against autologous tumor cells.
Kinetics of circulating B lymphocytes in human myeloma
Authors: Boccadoro M , Gavarotti P , Fossati G , Massaia M , Pileri A , Durie BG
Published: October 2019

The tritiated thymidine labeling index (LI%) of peripheral B lymphocytes was studied in eight myeloma patients using simultaneous immunofluorescence and autoradiography. The LI% values were low (0.3%- 5.1%), but significantly increased as compared to normal controls. In addition, there was excellent correlation between the LI% values and myeloma disease activity: lowest LI% values were observed in remission patients and the highest at the time of relapse. Simultaneous LI% evaluation of bone marrow myeloma cells in five patients gave concordant results, indicating the same kinetic behavior in both these compartments, particularly in the relapse phase. These data indicate both that circulating B lymphocytes include the neoplastic clone and that these B lymphocytes and bone marrow myeloma cells have similar kinetics.
Tin-protoporphyrin suppression of hyperbilirubinemia in mutant mice with severe hemolytic anemia
Authors: Sassa S , Drummond GS , Bernstein SE , Kappas A
Published: October 2019

Tin-protoporphyrin is a potent competitive inhibitor of heme oxygenase both in vivo in animals and in vitro in isolated enzyme preparations, and when administered to neonatal rats, prevents the development of postnatal hyperbilirubinemia. In this study we examined the effect of the metalloporphyrin on the activity of heme oxygenase in liver, kidney, and spleen, and on the level of bilirubin in plasma in three types of anemic mutant mice with severe hemolytic diseases. We report that the administration of tin-protoporphyrin to anemic mutants homozygous for severe hemolytic disease results in substantial inhibition of heme oxidation in liver, spleen, and kidney and in significant reduction of plasma bilirubin levels. Tin-protoporphyrin thus has the capacity to significantly inhibit in vivo heme degradation and to concurrently diminish plasma bilirubin levels in severe chronic hemolytic disorders.
Chronic lymphocytic leukemia progenitor cells carry the antigens T65, BA-1, and Ia
Authors: Perri RT , Royston I , LeBien TW , Kay NE
Published: October 2019

CLL B cells may be induced to form B-cell colonies in vitro. Colonies formed are monoclonal and appear to reflect the circulating malignant B- cell clone in vitro. Using hybridoma-produced monoclonal antibodies (MAB) and an in vitro B-cell colony assay, we have provided a characterization of the antigenic phenotype of the clonogenic CLL B cell. B-cell colony growth in both patients and normals was not altered by prior incubation with either MAB or complement (C') alone. CLL B- cell colony formation was markedly reduced after treatment with T101 and C', while normal colonies were unaffected (8 +/- 2 versus 107 +/- 10). None of the residual CLL B-cell colonies after T-101 and C' treatment reacted with T-101. However, BA-1 and la reactivity were still seen in residual CLL B-cell colonies following T-101 treatment. In contrast, a similar percentage reduction of B-cell colony growth was seen for both normals and CLL patients following treatment with BA-1 (76% versus 81%) and Q5/13 (89% versus 92%). These studies suggest that the CLL progenitor cell is characterized by the phenotype la+, BA-1+, T- 101+. Better definition of the CLL progenitor cell has potential implications with regards to clinical utilization of MAB in the treatment of CLL.
Individual BFU-E in polycythemia vera produce both erythropoietin dependent and independent progeny
Authors: Cashman J , Henkelman D , Humphries K , Eaves C , Eaves A
Published: October 2019

Erythropoietic progenitors from peripheral blood of normal individuals or patients with polycythemia vera (PV) were cultured in methylcellulose medium containing 2.5 U/ml of erythropoietin (Ep). After 7–9 days, colonies considered to be early stage large bursts were individually removed, resuspended in a small volume of fresh methylcellulose medium, and then divided between 2 dishes. To one of these secondary cultures, sufficient Ep was added to bring the concentration of Ep up to approximately 3 U/ml. To the other was added an equal volume of medium but no Ep. The final concentration of Ep in these cultures was determined to be less than 0.01 U/ml. Nine days later, both types of secondary cultures were scored for the presence of colonies containing 8 or more hemoglobinized erythroblasts. Of 90 primary colonies from 3 normal individuals assessed in this way, 59 gave secondary erythroid colonies in the high Ep cultures, while none gave secondary erythroid colonies in the low Ep cultures. Additional control experiments in which primary colonies from normal individuals were divided into duplicate high Ep cultures showed that on average, the procedure used divided primary colonies equally. Of 109 primary colonies from 5 PV patients that yielded secondary erythroid colonies in the high Ep cultures, 21 yielded no secondary erythroid colonies in the low Ep cultures. The other 88 yielded erythroid colonies in both, but the secondary colonies in the low Ep cultures were consistently smaller in size and significantly fewer in number. Similar results were obtained when primary colonies were generated in cultures to which no Ep was added. These findings indicate that primitive BFU-E in patients with PV can be subdivided into 2 populations: a minor population restricted to the production of erythroid colony-forming cells (Ep- dependent progenitors) that require Ep for their detection, and a major population that is not restricted in this way. In addition, these experiments show that most of the primitive BFU-E that generate Ep- independent progenitors also produce significant numbers of cells that are Ep-dependent.
In Waldenstrom's macroglobulinemia the quantity of detectable circulating monoclonal B lymphocytes correlates with clinical course
Authors: Smith BR , Robert NJ , Ault KA
Published: October 2019

Using a sensitive flow-cytometer-based method of detecting small numbers of morphologically normal monoclonal B lymphocytes, we have investigated the presence and quantity of these cells in the blood of 12 patients with Waldenstrom's macroglobulinemia. All 12 patients, including 6 at the time of asymptomatic presentation, had such circulating monoclonal cells. By comparison, 2 of 7 patients with multiple myeloma had such cells, and prior studies have shown that 80% of patients with non-Hodgkin's lymphoma exhibit blood involvement by these criteria. Enzymatic removal of surface immunoglobulin (lg) with subsequent regeneration by the cells after overnight culture established that the monoclonal surface lg being studied was of intrinsic cell membrane origin and not passively adsorbed serum lg. Serial studies were performed in 7 patients. In the 4 cases where a clinical response to therapy and a decrease in serum IgM level was seen, there was a corresponding decrease in the estimated number of abnormal cells. In the 3 cases where there was neither clinical nor serum M-component evidence of response, the estimated percent abnormal cells likewise increased or remained constant. We conclude that patients with Waldenstrom's macroglobulinemia have a significant number of peripheral blood monoclonal B lymphocytes, even early in their disease, and that for a given patient, serial determinations of the number of these cells as estimated by flow cytometry reflects the clinical activity of the disease.
Monoclonal antibodies to human coagulation factor V and factor Va
Authors: Foster WB , Tucker MM , Katzmann JA , Miller RS , Nesheim ME , Mann KG
Published: October 2019

BALB/c mice were immunized with human factor V. The immunogen was a mixture of procofactor (factor V) and thrombin-activated cofactor (factor Va). Spleen cells were obtained from an immunized animal and fused with NS-1 murine myeloma cells. Hybrid cell cultures were assayed for the production of antibodies to human factor V and factor Va by a solid-phase radioimmunoassay. Factor V and/or factor-Va-specific antibodies were detected in 38 of the 96 cultures assayed. The cells from 10 of these positive cultures were subcloned by limiting dilution and grown as ascites tumors in BALB/c mice. Ascitic fluids were obtained and characterized with respect to their binding interaction with human factor V and factor Va. Three hybridoma cell lines produce monoclonal antibodies that react equally well with factor V and factor Va. Another antibody reacts with both antigens, but the reactivity with factor V is better than with factor Va. An additional two antibodies react with factor Va better than factor V in the radioimmunoassay (RIA). The remaining four antibodies react exclusively with factor V. A previously described murine monoclonal antibody to human factor V (alpha HFV-1) has been used to study the peptides produced during the thrombin-catalyzed activation of human factor V. This antibody binds both factor V and factor Va, releases them at high ionic strength, and has an apparent dissociation constant for factor Va of 3 x 10(-9)M. When human factor V (mol wt 330,000) is activated by thrombin and passed over an alpha HFV-1-Sepharose affinity resin, factor Va binds and subsequently can be eluted. The eluate in 1.2 M NaCl contains two fragments of apparent mol wt 93,000 and 70,000. EDTA, which inactivates factor Va, promotes release of the mol wt 93,000 fragment from factor Va bound to the antibody. Subsequent elution with 1.2 M NaCl releases the mol wt 70,000 fragment. These observations indicate that human factor Va is a two subunit protein and that the epitope for alpha HFV-1 is on the mol wt 70,000 fragment.
Monitoring the entry of new platelets into the circulation after ingestion of aspirin
Authors: Di Minno G , Silver MJ , Murphy S
Published: October 2019

There have been reports of a 24–48-hr delay in the recovery of platelet cyclooxygenase activity and platelet function after the ingestion of aspirin. However, these studies employed a single aggregating agent to stimulate enzymatic or functional activity. We investigated the effects of some pairs of aggregating agents on 14 platelet-rich plasmas (PRP) from normal subjects 2 and 4 hr after ingestion of 650 mg aspirin and daily up to 72 hr. We studied platelet aggregation and secretion with a lumiaggregometer and thromboxane-B2 formation by radioimmunoassay. Aggregation and secretion occurred as early as 4 hr after aspirin ingestion in response to combinations of arachidonic acid with epinephrine, collagen, or adenosine diphosphate (ADP). Thromboxane formation was detected as early as 4 hr after ingestion of aspirin in response to 1 mM arachidonic acid in combination with 1 microgram/ml collagen. Up to 72 hr, there was a linear return of thromboxane formation stimulated by this combination, reflecting the entry of new platelets into the circulation. In vitro experiments with mixtures of aspirin-free and aspirin-treated platelets showed that the combination of collagen and arachidonic acid (AA) could produce full aggregation and secretion when only 2.5% of aspirin-free platelets were present. Use of the combination of collagen plus AA demonstrates the early entry into the circulation of platelets originating from megakaryocytes whose cyclooxygenase has not been completely acetylated.
Modulation of hemoglobin synthesis in K-562(S) cells treated with interferons
Authors: Cioe L , Meo P , Sorrentino V , Rossi GB
Published: October 2019

Treatment of K-562(S) cells with human interferons (HulFN) alpha, beta, or gamma results in a modulation of hemoglobin (Hb) production. When K- 562(S) cells, “induced” with butyric acid or hemin, are given low dosages of all three types of IFN, the percent benzidine-positive cells doubles. Treatment with low doses of IFN causes the acceleration and increased production of those Hb types (as shown by Cellogel analysis) that are already synthesized either in untreated or in “induced” cultures. These results were confirmed by using pure HulFN-beta, and were abrogated, for HulFN-alpha, in presence of its specific antiserum. In contrast, cultures of K-562 cells treated with either inducer and given more than 10(4) IU/ml of alpha- or beta-IFN show a dose-dependent decrease of hemoglobinization in the absence of cell death. The inhibitory effect was reversible upon removal of IFN and culture reseeding in IFN- and inducer-free medium. The significance of both sets of data is strengthened by the pronounced heterogeneity of K-562S cells with respect to their sensitivity to IFN treatment as evaluated by the establishment of the antiviral state. Apparently, one cell out of two is sensitive to IFN, which suggests that the magnitude of IFN effects described here may be larger than it appears to be from the data taken at face value.
The role of sialic acid in the dysfibrinogenemia associated with liver disease: distribution of sialic acid on the constituent chains
Authors: Martinez J , MacDonald KA , Palascak JE
Published: October 2019

To further evaluate the role of sialic acid in the dysfibrinogenemia associated with liver disease, we studied the effect of removal of excess sialic acid residues from the fibrinogen of five patients with liver disease on the thrombin time and fibrin monomer aggregation. Patient fibrinogens containing 1.4–3.4 residues of sialic acid per molecule in excess of normal controls, with thrombin times 12–22 sec longer than normal and with abnormal fibrin monomer aggregation, were stripped of their excess sialic acid by incubation with Vibrio cholerae neuraminidase, followed by rapid removal of the enzyme by antineuraminidase antibody affinity chromatography. These partially desialylated patient fibrinogens, with a normal number of sialic acid residues remaining, exhibited normal thrombin times and normal fibrin monomer aggregation. Sodium dodecylsulfate-polyacrylamide gel electrophoresis (SDS-PAGE) of reduced normal, patient, and partially desialylated patient (sialyl-3H)-fibrinogen exhibited 60% of the radioactivity in the B beta chain and 40% in the gamma chain. There was no radioactivity detectable in the A alpha chain. These studies provide additional evidence that the increased sialic acid content of the acquired dysfibrinogenemia of liver disease is responsible for its functional defect and that the excess sialic acid is distributed on the B beta chain and gamma chains of the fibrinogen.
Human neutrophil dysfunction with giant granules and defective activation of the respiratory burst
Authors: Newburger PE , Robinson JM , Pryzwansky KB , Rosoff PM , Greenberger JS , Tauber AI
Published: October 2019

We describe a patient whose peripheral blood neutrophils and bone marrow precursors (beyond promyelocytes) contained multiple large azurophilic granules. There were also giant granules in eosinophils, basophils, melanocytes, renal tubules, thyroid, and neurones, but not lymphocytes or monocytes. His clinical course included recurrent (ultimately fatal) infections and severe neurologic impairment. Immunofluorescent staining with fluoroscein- and rhodamine-conjugated antisera to primary and secondary granule markers showed virtually all of the granulocyte granules and rare monocyte granules to be fusion products containing both markers. Electron microscopy showed the granules to be large peroxidase-containing lysosomes. Only rare normal primary and secondary granules were present. Superoxide generation in response to opsonized zymosan was 7.3 nmole/min/10(6) cells (control 8.9); but in response to phorbol myristate acetate, only 2.2 (control 9.4). Nitroblue tetrazolium slides showed 3+ dye reduction in response to opsonized zymosan by 90% of granulocytes (control 91%) and to phorbol myristate acetate by 22% (control 99%), with 71% producing only a minimal 1+ response. Cellular contents of myeloperoxidase and beta- glucuronidase were elevated, but the percent release during exocytic degranulation was equivalent to control. Ingestion of complement- opsonized Staphylococcus aureus and zymosan was also normal. Killing of Staphylococcus aureus was 60% at 90-min incubation (control 92%). Granulocyte cyclic adenosine monophosphate (AMP) content was 4 pmole/10(7) cells (control 3.1). In order to determine whether these characteristics derived from the cells' genetic program or their environment, the patient's bone marrow was grown in long-term culture. Granulocytes produced in vitro demonstrated the same morphology, same defect in activation of nitroblue tetrazolium reduction, and same normal cyclic AMP level as those harvested from peripheral blood. These studies describe a new disorder of granulocytes; the structural similarity to, but biochemical differences from, Chediak-Higashi disease indicate the probable heterogeneity of mechanisms for the same morphological abnormality.
Demonstration of a natural antigalactosyl IgG antibody on thalassemic red blood cells
Authors: Galili U , Korkesh A , Kahane I , Rachmilewitz EA
Published: October 2019

A modified antiglobulin test, based on the high affinity between the Fc portion of the red blood cell (RBC) bound IgG and the Fc receptor on the myeloid cell K-562, was utilized for demonstration of immunoglobulins (Ig) on thalassemic RBC. Ig was found on the RBC of 73 out of 80 patients with thalassemia. The immunoglobulins on the thalassemic RBC belonged to the IgG subclass and were autoreactive. Elution studies utilizing various carbohydrates, or by thermal stripping, indicated that at least part of the IgG molecules found on the thalassemic RBC were specifically reactive with terminal galactosyl residues on the RBC membrane. IgG antibodies with similar reactivity were also demonstrated in normal human serum. These natural antigalactosyl IgG antibodies from normal sera could bind to IgG- depleted thalassemic RBC. Thalassemic RBC and normal senescent RBC were previously found to contain reduced amounts of membrane sialic acid (SA). It is suggested that the antigalactosyl IgG antibodies interact with newly exposed galactosyl residues underlying the sialic acid units. Such interaction may lead to the shortened lifespan of thalassemic RBC and may result in sequestration of senescent normal RBC by the reticuloendothelial system.
Properties of NADH-cytochrome-b5 reductase from human neutrophils
Authors: Badwey JA , Tauber AI , Karnovsky ML
Published: October 2019

An NADH-ferricyanide reductase activity of ca. 170 nmole ferricyanide reduced/min/10(7) cells is present in the membrane fraction of human neutrophils. This membrane-bound activity constitutes ca. 85% of the total NADH-ferricyanide reductase activity that is present in these cells. The enzyme(s) readily utilize(s) purified cytochrome-b5 from beef liver as an electron acceptor. No other physiologic electron acceptors tested (e.g., ubiquinone-30, menadione) were active. The specificities of electron donors (e.g., NADH congruent to deamino-NADH much greater than NADPH) and acceptors (e.g., Fe(CN)6–3 greater than 2,6-dichlorophenol-indophenol much greater than O2) for the enzyme(s) in unfractionated membranes, along with action of inhibitors (e.g., ADP, p-chloromercuribenzoate) and the pH optimum, indicate that virtually all of the membrane-bound ferricyanide reductase activity in these cells is NADH-cytochrome-b5 reductase. This reductase, however, is only slightly solubilized (ca. 10%) by a phosphate buffer extraction procedure that is effective with the liver enzyme.
Phenotypic effect of heterozygous alpha and beta 0-thalassemia interaction
Authors: Melis MA , Pirastu M , Galanello R , Furbetta M , Tuveri T , Cao A
Published: October 2019

In this study, we carried out restriction endonuclease mapping in order to characterize the alpha-globin genotype of 10 Sardinian beta 0- thalassemia heterozygotes, all of whom presented with normal red blood cell indices and increased HbA2 levels. In 8 of these subjects, we found the deletion of two alpha-globin genes (-alpha/-alpha), and in the remaining two the deletion of a single alpha-globin gene (- alpha/alpha alpha). In three of these carriers with the (-alpha/-alpha) alpha-globin genotype and in one with the (-alpha/alpha alpha) genotype, we also found the glucose-6-phosphate dehydrogenase (G6PD) defect of the Mediterranean type. On the basis of these findings, we may conclude that the interaction of heterozygous beta 0-thalassemia with alpha-thalassemia, due to the deletion of either one or two alpha- globin genes, may lead to the production of red blood cells with normal indices. The association of the G6PD defect with this thalassemia gene complex may eventually contribute to this effect. We suggest, therefore, that screening programs for heterozygous beta-thalassemia in populations where alpha-thalassemia is also prevalent, should incorporate the determination of HbA2 in the first set of tests.
Kinetics of platelet adhesion and thrombus growth
Authors: Adams GA , Brown SJ , McIntire LV , Eskin SG , Martin RR
Published: October 2019

Epifluorescent microscopy was used to monitor the adhesion of platelets and the growth of platelet aggregates on collagen-coated glass tubes perfused with whole blood. The maximum basal length and width of the aggregate size increased linearly with time, growing symmetrically transverse to the direction of flow and asymmetrically in the plane longitudinal to the direction of flow. Aggregates had elliptical bases, with the major axis parallel to the direction of blood flow. These studies provide an experimental approach to studies of the kinetics of platelet interaction with artificial surfaces and give further support to the concept that blood flow has a major effect on the development of platelet thrombi.
Adenosine and adenosine analogues are more toxic to chronic lymphocytic leukemia than to normal lymphocytes
Authors: Bajaj S , Insel J , Quagliata F , Hirschhorn R , Silber R
Published: October 2019

We compared the effect of adenosine and adenosine analogues on the phytohemagglutinin-induced proliferative response of blood lymphocytes from normal subjects and patients with chronic lymphocytic leukemia. As measured by the inhibition of thymidine or leucine incorporation, adenosine was more toxic to chronic lymphocytic leukemia (CLL) than to normal lymphocytes. This difference was not affected by the removal of adherent cells. The patients' B lymphocytes were more susceptible to adenosine toxicity than normal B lymphocytes. Similar responses were noted in T lymphocytes from both sources. Differential susceptibility was also observed with deoxyadenosine and adenosine analogues, including 5′deoxyadenosine. Uridine rescue from adenosine toxicity was observed for normal and CLL lymphocytes. In the presence of uridine, there was no difference in the residual inhibition of CLL as compared to normal lymphocytes. Intact CLL lymphocytes metabolized 14C-adenosine at a much lower rate than normal lymphocytes. While it appears that the greater toxicity of adenosine to CLL lymphocytes reflects the impaired catabolism of this nucleoside by these cells, evidence is presented that this is not the only mechanism underlying the differential susceptibility. These results may serve as the basis for further pharmacologic investigations of adenosine and adenosine deaminase inhibitors in chronic lymphocytic leukemia.
Comparison of central nervous system prophylaxis with cranial radiation and intrathecal methotrexate versus intrathecal methotrexate alone in acute lymphoblastic leukemia
Authors: Muriel FS , Svarch E , Pavlovsky S , Eppinger-Helft M , Braier J , Vergara B , Garay G , Kvicala R , Divito JM , Failace R , Dibar E , Jimenez E
Published: October 2019

In acute lymphoblastic leukemia (ALL), central nervous system (CNS) prophylaxis with cranial irradiation plus 5 doses of intrathecal methotrexate (i.t. MTX) reduces the incidence of CNS relapse to 7%-15%. However, increased evidence of CNS delayed toxicity started to be recognized as CT scan abnormalities and neuropsychologic alterations, mainly in children. Two questions were analyzed in the present report: (1) Will further doses of i.t. methotrexate and dexamethasone (i.t. MTX- DMT) decrease the incidence of CNS relapse in patients treated early in remission with cranium irradiation plus i.t. MTX-DMT even more? (2) Is i.t. MTX-DMT given during induction and maintenance equally as effective as cranium irradiation plus i.t. MTX-DMT? A randomized study was designed to answer the first question. Incidence of primary CNS relapse in i.t. MTX-DMT-treated patients with a WBC count less than 50,000 was 11% (15 of 135 patients) and was 11% (17 of 150) in the untreated group. In patients with a WBC count greater than 50,000, it was 16% (6/37) in the treated group and 19% (6/31) in the control group. No difference was observed according to treatment in both prognostic groups. Patients in this study were retrospectively compared with a consecutive protocol in which patients received 3 doses of i.t. MTX-DMT alone during induction plus 3 doses weekly during the first month of remission and every 3 mo thereafter. The incidence of primary CNS leukemia at 60 mo in patients with a WBC count less than 50,000 was 20% in the irradiated group and 32% in the group with i.t. MTX-DMT alone. This difference was not significant. However, the relapse-free survival at 60 mo was 26% and 41%, respectively, (p less than 0.0005). The incidence of primary CNS relapse in patients with a WBC count more than 50,000 at 48 mo was 28% in the irradiated group and 42% in the nonirradiated group. The difference was not significant. The duration of complete remission was similar, remaining at 15% and 16% of patients disease-free at 48 mo, respectively. We conclude that (A) after cranial irradiation plus i.t. MTX-DMT X 5, the use of additional doses of i.t. MTX-DMT is not of further benefit in preventing CNS relapse; (B) the use of i.t. MTX-DMT alone compares similarly with cranial irradiation plus i.t. MTX-DMT in the incidence of CNS relapse; and (C) relapse-free survival and survival in patients with a WBC count less than 50,000 were significantly longer in those without cranial irradiation.
Thirty-six human monoclonal immunoglobulins with antibody activity against cytoskeleton proteins, thyroglobulin, and native DNA: immunologic studies and clinical correlations
Authors: Dighiero G , Guilbert B , Fermand JP , Lymberi P , Danon F , Avrameas S
Published: October 2019

Six hundred and twelve monoclonal Ig (MIg) were studied for their antibody activity against the following autoantigens: actin, tubulin, thyroglobulin, myosin, myoglobin, fetuin, albumin, transferrin, and double-stranded DNA (dsDNA). Of these 612 MIg, 36 (i.e., 5.75%) were shown to possess antibody activity. Thirty-two of these 36 (5.22% of the total) were mainly directed against actin. The four others were directed, respectively, against tubulin, myosin, thyroglobulin, and dsDNA. The interaction of the MIg with the respective antigen was demonstrated by immunoenzymatic methods with monospecific antisera and by blotting experiments. Furthermore, this interaction in the 12 cases studied was mediated by the dimeric fragment F(ab')2 of the MIg. The MIg with antitubulin, antithyroglobulin, and anti-dsDNA activities were exclusively inhibited by their homologous antigens. Those with antiactin activity were predominantly inhibited by actin and also by tubulin and thyroglobulin. The one binding to myosin was, for the most part, inhibited by myosin and also significantly by actin and tubulin. Retrospective clinical analysis was possible for 31/36 patients. Twenty- six of 31 had malignant lymphoplasmocytic disorders. The five others were followed for miscellaneous disorders without overt signs of multiple myeloma (MM) or Waldenstrom's macroglobulinemia (WM). The correlation between the antibody activity of the MIg and the clinical features is discussed. These results indicate that a high proportion of MIg possess antibody activity against actin (5.22%). This incidence contrasts sharply with the positive reactions found toward all the other antigens tested: only one each for dsDNA, tubulin, thyroglobulin, and myosin, and none against myoglobin, fetuin, albumin, and transferrin. The significance of these results and the relationship between MIg and natural antibodies are discussed.
Waldenstrom's macroglobulinemia and peripheral neuropathy: a clinical and immunologic study of 25 patients
Authors: Dellagi K , Dupouey P , Brouet JC , Billecocq A , Gomez D , Clauvel JP , Seligmann M
Published: October 2019

We investigated, by indirect immunofluorescence, the binding of monoclonal IgM to human peripheral nerve in 25 patients with Waldenstrom's macroglobulinemia and peripheral neuropathy. In 10 cases (40%), an antibody activity against the myelin sheaths was demonstrated. The reactivity was mediated by the F(ab')2 fragments of the IgM. Prior delipidation of nervous tissue was needed to allow full expression of the target antigen(s). In nine cases, the IgM reacted with both peripheral and central myelin of primates, but not mouse or rabbit nervous tissue. In one case, the IgM reacted only with human peripheral nerve. The patients, whose IgM had an antibody activity to myelin antigen(s), had some distinct hematologic and neurologic features. The peripheral neuropathy always antedated the hematologic symptoms by several years. The serum level of the monoclonal IgM was low in all cases, and overt lymphoid malignancy was frequently absent. In other patients with neuropathy, the monoclonal IgM, which lacked antimyelin antibody activity, displayed either cross-idiotypic antigenic determinants or anti-intermediate filament antibody activity. These results taken together emphasize the heterogeneity of antibody activity of the monoclonal IgM from patients with Waldenstrom's macroglobulinemia and peripheral neuropathy.
Alpha-thalassemia is related to prolonged survival in sickle cell anemia
Authors: Mears JG , Lachman HM , Labie D , Nagel RL
Published: October 2019

We have determined the frequency of deletional alpha-thalassemia in black populations in the USA and Africa that harbor sickle cell anemia. In normals, the frequency of the chromosome bearing a deletion of one of the two normal alpha gene loci, designated (-alpha), ranged from 0.12 to 0.16, and in sickle trait subjects, the frequency ranged from 0.18 to 0.20. By contrast, in sickle cell anemia subjects, the frequency was significantly greater and ranged from 0.22 to 0.33. Analysis demonstrated that the greater frequency in the last group was primarily a result of an increased number of subjects with alpha- thalassemia trait (also called homozygous alpha-thalassemia-2). In addition, the frequency of the (-alpha) chromosome was found to increase progressively with age, supporting the hypothesis that alpha- thalassemia is favorable to the survival of subjects with sickle cell anemia. Thus, individuals who inherit alpha-thalassemia and sickle cell anemia may represent a subgroup of patients with a longer life expectancy.
Cytochemical, functional, and proliferative characteristics of promonocytes and monocytes from patients with monocytic leukemia
Authors: van Furth R , van Zwet TL
Published: October 2019

This article deals with a prospective study on the cytochemical, functional, and proliferative characteristics of promonocytes and bone marrow and peripheral blood monocytes of 20 patients with acute monocytic leukemia and 7 patients with chronic monocytic leukemia. The results show a wide variation in the peroxidase and esterase activities in these cells, whereas the percentages of mononuclear phagocytes with Fc gamma and C3b receptors did not differ appreciably from those in normal individuals. A discriminant analysis of these data and corresponding data from normal individuals showed that a below-normal peroxidase activity of circulating monocytes has predictive value for the presence of monocytic leukemia; a below-normal esterase activity has less, but nevertheless some, predictive value in this respect. An increase in the percentage of circulating monocytes, a decrease in the percentage of Fc gamma or C3b receptors, and a decline in the ability to phagocytose bacteria has no predictive value for the presence of monocytic leukemia. The mean percentage of patients' promonocytes that incorporated 3H-thymidine amounted to 80.9%, which is close to the control value in normal individuals. The mean values for the labeling indices of cultured bone marrow and peripheral blood monocytes are 1.0% and 0.74%, respectively; when 3H-thymidine was added to whole blood, the labeling index of the monocytes amounted to 3.6%. These percentages are only a little higher than those found for monocytes of normal individuals. These results indicate that the majority of the circulating monocytes in acute and chronic monocytic leukemia are not actively dividing or blast cells.
Changes in actin content during induced myeloid maturation of human promyelocytes
Authors: Meyer WH , Howard TH
Published: October 2019

Actin is an important cytoskeletal protein; new actin synthesis occurs during differentiation of many motile cells. To better understand the process of myeloid maturation, the change in actin content during induced maturation of HL-60 human promyelocytic leukemia cells was studied. HL-60 cells induced toward myeloid maturation by a 5-day exposure to dimethylformamide showed an 86% increase in a 43,000 mol wt protein comigrating with rabbit muscle actin on dodecyl sulfate polyacrylamide gels. To further demonstrate that this was an increase in actin content, the total actin content of lysed HL-60 cells was measured by the ability of actin to inhibit DNAase I. Using this assay, actin content of HL-60 cells increased 96% during induced differentiation. The amount of incorporation of 3H-leucine into actin doubled after a 5-day exposure to dimethylformamide, suggesting the increase in actin was due primarily to new synthesis. Total new protein synthesis increased 2–7-fold during differentiation. Additional analysis of polyacrylamide gels showed increased quantities and new synthesis of a high molecular weight protein comigrating with rabbit muscle myosin. This study shows that actin content increases during myeloid maturation. It also demonstrates that the HL-60 cell line is a useful model to study both functional and biochemical events during human myeloid differentiation.
The endotoxin-induced procoagulant of mouse exudate macrophages: a factor-X activator
Authors: Shands JW Jr
Published: October 2019

The properties of mouse macrophage procoagulant induced by endotoxin in vitro were studied by the acceleration of clotting and by chromogenic assays using as substrates human plasma and bovine components, which are not activated by mouse tissue factor. Maximal macrophage procoagulant activity occurred when activated cells were lysed in culture supernatant fluids, suggesting the interaction of cellular and supernatant factors. This procoagulant was clearly able to activate bovine factor X. The procoagulant also appeared to have prothrombinase activity. However, additional experiments suggested that the bulk of this activity was due to the activation of factor X contaminating the prothrombin. The production of the procoagulant was inhibited by warfarin (5 microM). Its activity was inhibited by 1 mM diisopropylfluorophosphate and unaffected by iodoacetamide, indicating that the procoagulant is a serine protease. Macrophage culture supernatants contained factor-VII-like activity. Neither mouse tissue factor nor macrophage culture supernatants alone activated bovine factor X. The two combined served as an efficient factor-X activator. Active supernatant factor (factor-VII-like) was not produced by macrophages cultured in the presence of warfarin, while the production of the macrophage cellular factor was unaffected by the presence of warfarin. I conclude that exudate macrophages cultured in vitro make and secrete factor VII or a factor-VII-like substance into the culture supernatant. When activated macrophages are lysed in this supernatant, the interaction of a cellular factor (? tissue factor) and factor VII gives rise to a factor-X activator.
5-Azacytidine increases gamma-globin synthesis and reduces the proportion of dense cells in patients with sickle cell anemia
Authors: Ley TJ , DeSimone J , Noguchi CT , Turner PH , Schechter AN , Heller P , Nienhuis AW
Published: October 2019

We previously demonstrated that 5-azacytidine can selectively increase gamma-globin synthesis in a patient with beta +-thalassemia, prompting us to treat two patients with sickle cell anemia and two additional patients with beta + thalassemia. 5-Azacytidine (2 mg/kg/day) was continuously infused for 7 days with no apparent clinical toxicity. The gamma/beta-globin biosynthetic ratio increased fourfold to sixfold in the bone marrow cells of each patient after treatment and remained elevated for 7–14 additional days. Hypomethylation of DNA near the gamma-globin genes in bone marrow cells was demonstrated 2 days after beginning the 5-azacytidine infusion. The peripheral blood fetal hemoglobin (HbF) level increased from 6.0% to 13.7% in one patient with sickle cell anemia and from 1.6% to 8.9% in the second. Stractan gradient analyses of peripheral blood from patients with sickle cell anemia revealed a marked decrease in the percentage of dense cells (cells that contain increased amounts of HbS polymer when deoxygenated) following treatment. These observations provide an impetus to investigate the effects of repeated courses of 5-azacytidine in a small group of severely ill patients to determine whether this drug may have a role in the treatment of patients with sickle cell anemia and beta- thalassemia.
Inhibition of erythroid colony formation by autologous bone marrow adherent cells from patients with the anemia of chronic disease
Authors: Roodman GD , Horadam VW , Wright TL
Published: October 2019

To determine the role marrow-adherent cells may play in the anemia of chronic diseases, marrow samples were collected from ten patients with the anemia of chronic disease, seven control patients with cancer but without the anemia of chronic disease, and five normal volunteers. Marrow was either cultured directly or first depleted of adherent cells and then cultured. Plasma clots containing 6 X 10(4) nonadherent marrow cells were cocultured with marrow-adherent cells prepared by incubating 6 X 10(4)-6 X 10(2) unfractionated marrow cells in microtiter plates and removing the nonadherent cells. Adherent cell depletion of marrow from patients with anemia of chronic disease significantly increased erythroid colony formation. Coculture of adherent cells from anemic patients with autologous nonadherent marrow cells inhibited erythroid colony-forming unit (CFU-E) proliferation in patients with the anemia of chronic disease. In contrast, adherent cells from control patients did not affect autologous erythroid colony formation, and adherent cells from normal volunteers stimulated autologous erythroid colony formation. Coculture of adherent cells from anemia patients with nonadherent marrow from control patients failed to inhibit allogeneic erythroid colony formation. Media conditioned by adherent cells from patients with the anemia of chronic disease failed to suppress consistently the formation of allogeneic erythroid colonies. These data suggest that marrow-adherent cells normally stimulate erythropoiesis, but suppress erythroid progenitors, in patients with the anemia of chronic disease and may in part be responsible for their anemia.