Leukemic Sera Research Articles

Early syncytium formation by bovine leukemia virus.

Bovine leukemia virus (BLV) from either persistently infected bat cells or fetal lamb kidney cells induced rapid syncytium formation in F81 indicator cells. Distinct syncytia were seen within 2 h after inoculation of cells with highly concentrated (500-fold) cell-free BLV preparations and within 4 to 8 h when unconcentrated cell-free BLV preparations were used. Indicator cell densities of 1 x 10(5) to 2 x 10(5) were optimal for rapid and maximal syncytium formation. Pretreatment of BLV with reference BLV leukemic serum and antiserum prepared against purified BLV significantly inhibited (95%) syncytium formation. Reference bovine viral diarrhea virus serum, foamy-like bovine syncytial virus serum, and control serum had little effect (17% inhibition). Antiserum to BLV gp51 inhibited syncytium formation by greater than 96%, whereas antiserum to BLV p24 reduced syncytium activity to a much lesser extent (38% inhibition). Treatment of BLV with beta-propiolactone (0.005 to 0.05%) had little or no effect upon syncytium-forming activity, whereas UV irradiation (15 ergs/mm(2) per s for 30 min) reduced, but did not completely destroy, the fusion activity. However, both beta-propiolactone and UV irradiation drastically reduced the replication potential of BLV, as demonstrated by the lack of p24 expression in the inoculated cells. Concentrations of cycloheximide, cytosine arabinoside, tunicamycin, and 2-deoxy-D-glucose which effectively blocked cellular macromolecular synthesis did not significantly inhibit syncytium formation. These latter results suggested that de novo protein and DNA synthesis as well as protein glycosylation were not required for early syncytium formation. Thus, these experiments demonstrated that replication of BLV by the indicator cells was not essential for cell fusion.

ANTIBODY AND COMPLEMENT-MEDIATED CYTOTOXICITY OF NUCLEATED CELLS INHIBITION BY THE SERA OF PATIENTS WITH ACUTE LYMPHOBLASTIC LEUKEMIA

SUMMARY Optimal conditions for the killing of normal human lymphocytes by IgG antibody and autologous normal human serum were established. When serum from a patient with acute lymphoblastic leukemia was used in place of the normal serum, the rate and extent of killing was markedly reduced. Levels of all complement components were equivalent in the normal and leukemic sera. Furthermore, if the patient's serum was mixed with normal serum, killing was inhibited at concentrations of the patient's serum of 25% or more. A purified preparation of a C3 inhibitor, previously demonstrated to occur in these patients, had a similar effect when mixed with normal serum. In addition, sensitized target cells incubated with this inhibitor and normal serum remained viable after washing and reexposure to fresh serum. If, however, the inhibitor was added to the sensitized cells in the absence of serum and then removed, killing proceeded normally on the subsequent addition of normal serum. Finally, the inhibitor was able to be eluted from sensitized target cells exposed to the inhibitor in the presence of serum. It would appear, therefore, that the inhibitor fixes to a sensitized cell in the presence of complement yielding a target cell that is resistant to the cytotoxic activity of complement. The fact that the inhibitor may also be eluted from cells obtained from the bone marrow of patients with acute lymphoblastic leukemia when the tumor burden is high suggests that a similar process may also occur in vivo.

Radioimmunoassay for urinary lysozyme in human serum from leukemic patients.

We present a radioimmunoassay for lysozyme in human serum, based upon human lysozyme isolated from the urine of leukemic patients and antiserum prepared against this lysozyme in the goat. In the separation step, a second antibody is used. By properly adjusting the concentrations of unlabeled and 125I-labeled lysozyme and of the antibodies, maximal precision (SD, 0.04 mg/litre) was obtained in the range 0.00 to 2.00 mg/litre. In 20 normal volunteers the lysozyme concentration was 4.6 +/- 0.8 mg/litre (mean +/- SD), in 13 patients with monocytic leukemia 34.4 +/- 8.6 mg/litre. Correlation with lysoplate determinations was excellent in leukemic sera (r = 0.97) but was poor in normal sera (r = 0.35), possibly owing to the existence of isoenzymes.

Identification and behavior of the precipitating BLV antibodies in sera of leukotic cattle.

Precipitating bovine leukemia virus antibodies isolated from bovine leukemic sera by ion-exchange chromatography, gel filtration and affinity chromatography were identified as immunoglobulins of the IgG1 subclass and IgA class. They proved to be different with regard to molecular size and electrical charge. Immunoglobulins IgG2 and IgM as well as low-molecular components ranging in the microglobulin regions (less than 4S) failed to precipitate BLV antigens. Individual bovine leukemic sera showed differing precipitating activities against p24 and gp69 antigens. Precipitating monospecific gp69 antibodies represented exclusively the IgG1 subclass. Some unidentified antigens in disrupted BLV preparations showed serological relationship to p24 and gp69, respectively.

Decreased microviscosity of membrane lipids in leukemic cells: Two possible mechanisms

Steady-state fluorescence polarization studies with the fluorescent lipid probe 1,6-diphenyl 1,3,5-hexatriene were done to determine the degree of microviscosity of cellular membrane lipids and serum lipoproteins in human normal donors and leukemic patients. The results show a marked decrease in microviscosity of cellular membrane lipids in both intact lymphocytes and isolated cellular plasma membranes obtained from leukemic patients in clinical relapse as compared to intact lymphocytes and isolated cellular plasma membranes obtained from normal donors and leukemic patients in complete clinical remission. Concomitant to these dynamic changes in cellular membrane lipids, the degree of microviscosity of lipids in the blood serum of leukemic patients in clinical relapse is markedly reduced as compared to serum obtained from normal donors and leukemic patients in complete clinical remission. Moreover, an in vitro incubation of leukemic lymphocytes with normal low density lipoproteins results in an increased microviscosity of cellular membrane lipids. In addition to the interrelation between cellular membrane lipids and serum lipoproteins, plasma membrane vesicles with a high degree of lipid microviscosity were isolated from the blood serum and pleural effusion of leukemic patients in clinical relapse. Such membrane vesicles could not be detected in normal serum. Therefore, we suggest that the two major mechanisms associated with the decreased microviscosity of membrane lipids in human leukemic cells are an abnormal exchange in lipids between the leukemic cell surface membrane and leukemic serum lipoproteins and an exfoliation of plasma membrane vesicles with a high degree of microviscosity from the cell surface of leukemic cells.

Kinetic features of a lymphoid population in the blastic crisis of chronic granulocytic leukaemia

Kinetic and chromosome studies were performed on the primitive cells from a patient in the blastic phase of chronic granulocytic leukaemia who had two distinct cell populations. Lymphoblastic cells predominated and stained with fluorescent labelled anti-acute lymphoblastic leukaemic sera. There was variation in the proliferative activity of the myeloid population but the lymphoid cells remained essentially quiescent. A Ph 1 chromosome was identified in the myeloid but not found in the lymphoid cells. The findings suggest the presence of a population unlike those of acute lymphoblastic leukaemia with limited metabolic capacity.

Microbial-like antigens in human leukemia.

Antigens present in leukemic sera and organ extracts, but absent from non-leukemic sera and tissues, were detected by examining their reactions with anti-microbial antisera by means of counter-immunoelectrophoresis. Out of 110 different antisera used, 12 antisera, those produced against Micropolyspora, Coccidioides, Salmonella, Mycoplasma, Mycobacterium simiae, Absidia, pseudorabies virus and adenovirus, proved to react regularly with the leukemic sera. Absorption studies point to an immunological specificity of these cross-antigenicities. Interpretations and implications of these discoveries are discussed in the paper.

Antibodies in human sera to oncorna virus-like proteins from normal or leukemia marrow cell cultures.

Some human marrows in culture release particles with oncornavirus-like properties. This study was designed to examine the immunological properties of similar particles in human marrow culture supernates. Leukemic and nonleukemic marrows were cultured for 5-7 days in the presence of [14C]uridine and [3H]leucine or [3H]glucosamine. Labeled supernatant components banding in sucrose gradient densities of 1.20-1.24 g/ml were used as antigen in a double antibody immunoprecipitation assay. The assay was validated by end point titrations and competition with unlabeled antigen; purified myeloma proteins were used as negative controls. Cross-reactivity with mammalian oncornaviruses, as judged by competitive inhibition of precipitation by these viruses, was slight and at the border of the sensitivity of the method. Precipitated antigens analyzed by SDS polyacrylamide gel electrophoresis contained three distinct polypeptides of about 70,000, 45,000 and 30,000 mol wt; these comigrated with the gp 70, pg 45, and p 30 of a murine leukemia virus. Similar polypeptides were obtained from both leukemic and nonleukemic marrow culture supernates. As determined by the radioimmunoprecipitation assay, 32 of 45 leukemic sera (71%), 36 of 45 normal sera (80%), 15 of 19 sera from family contacts of leukemic patients (79%), 14 of 21 cord blood specimens (67%), and 21 of 23 sera (91%) from patients with systemic lupus erythematosus had detectable antibody activity.

Lymphocyte transformation in leukemic serum

Lymphocyte transformation in leukemic serum

Studies on mouse Moloney virus induced tumours: II. Detection of p30 in the serum of mice with Moloney leukaemia by in vitro blocking of complement dependent antibody mediated cytotoxicity.

Sera from Balb/c mice bearing Moloney leukaemia block complement dependent antibody mediated cytotoxicity of an antiserum prepared in rats against syngeneic Moloney virus induced lymphomata when either spleen cells from mice bearing Moloney leukaemia (M) or an in vitro line of Moloney virus transformed cells (MSC) are used as targets. This antiserum has been shown to recognize p30, the major internal virion protein, as a cytotoxic target on these cells. Viral particles were identified by electron microscopic examination of pelleted material obtained from leukaemic sera after high speed centrifugation. However, removal of virus did not affect the capacity of the leukaemic sera to absorb cytotoxicity of rat ILR-3 for MSC targets, and only depressed somewhat its ability to absorb activity of the same antisera against M targets. Virus-free leukaemic sera also blocks complement dependent antibody mediated cytotoxicity of an antiserum prepared in goats against the gs3 determinant of p30. This indicates that the material in leukaemic sera responsible for the in vitro block of antibody mediated cytotoxicity was p30. A lesser degree of block was observed with sera obtained from normal Balb/c mice, but the nature of material responsible is as yet undefined.

THE INHIBITORY EFFECT OF LEUKAEMIA ASSOCIATED ANTIGEN AND LEUKAEMIC SERUM ON THE LEUKOCYTE MIGRATION OF CHILDREN WITH ACUTE LEUKAEMIA IN REMISSION

AllergyVolume 29, Issue 4 p. 288-296 THE INHIBITORY EFFECT OF LEUKAEMIA ASSOCIATED ANTIGEN AND LEUKAEMIC SERUM ON THE LEUKOCYTE MIGRATION OF CHILDREN WITH ACUTE LEUKAEMIA IN REMISSION R. SZIGETI, Corresponding Author R. SZIGETI From the Second Department of Paediatrics, Semmelweis University Medical School and Oncopathological Research Institute, Budapest, Hungary.Dr. R. Szigeti, 2nd Dept. of Paediatrics, Semmelweis Univ. Med. School, 1094 Budapest, Tüizoltó u. 7–9, HungarySearch for more papers by this authorT. RÉVÉSZ, T. RÉVÉSZ From the Second Department of Paediatrics, Semmelweis University Medical School and Oncopathological Research Institute, Budapest, Hungary.Search for more papers by this authorÉVA GERÖ-FERENCZ, ÉVA GERÖ-FERENCZ From the Second Department of Paediatrics, Semmelweis University Medical School and Oncopathological Research Institute, Budapest, Hungary.Search for more papers by this author R. SZIGETI, Corresponding Author R. SZIGETI From the Second Department of Paediatrics, Semmelweis University Medical School and Oncopathological Research Institute, Budapest, Hungary.Dr. R. Szigeti, 2nd Dept. of Paediatrics, Semmelweis Univ. Med. School, 1094 Budapest, Tüizoltó u. 7–9, HungarySearch for more papers by this authorT. RÉVÉSZ, T. RÉVÉSZ From the Second Department of Paediatrics, Semmelweis University Medical School and Oncopathological Research Institute, Budapest, Hungary.Search for more papers by this authorÉVA GERÖ-FERENCZ, ÉVA GERÖ-FERENCZ From the Second Department of Paediatrics, Semmelweis University Medical School and Oncopathological Research Institute, Budapest, Hungary.Search for more papers by this author First published: August 1974 https://doi.org/10.1111/j.1398-9995.1974.tb01656.xCitations: 4AboutPDF ToolsRequest permissionExport citationAdd to favoritesTrack citation ShareShare Give accessShare full text accessShare full-text accessPlease review our Terms and Conditions of Use and check box below to share full-text version of article.I have read and accept the Wiley Online Library Terms and Conditions of UseShareable LinkUse the link below to share a full-text version of this article with your friends and colleagues. Learn more.Copy URL Citing Literature Volume29, Issue4August 1974Pages 288-296 RelatedInformation

Leukemic Serum Inhibition of Phytohemagglutinin-Induced Transformation

Abstract The influence of leukemic serum on phytohemagglutinin-P (PHA-P)-induced lymphocyte transformation was studied. Twenty of 22 pretreatment sera collected at diagnosis inhibited the transformation of normal lymphocytes stimulated with PHA-P by more than 50%. In these experiments with PHA-P, kinetic studies of transformation demonstrated that leukemic serum inhibited rather than delayed transformation. The inhibition of transformation by PHA-P was assumed to be due to binding of PHA by components of leukemic serum because the inhibition could be overcome by an increase in the PHA concentration. A biologic assay of the PHA content of lysates of lymphocytes preincubated with PHA and leukemic serum further substantiated the PHA-binding mechanism of leukemic serum. Whether the differences observed in the binding capacity of normal and leukemic serum are qualitative or quantitative remains to be determined.

Influence of chemotherapeutic agents on the antibacterial activity of normal and leukemic sera.

The influence of various chemotherapeutic agents on the antibacterial activity of normal and leukemic sera was tested by bacteriolytic and bactericidal assays with use of 2 rough strains of Escherichia coli, one sensitive to the antlbacterlal activity of human serum and another resistant to such action. With a “low” dose of drugs, daunorublcine, mechlorethamine, and methotrexate slightly inhibited early stages of bacteriolytic activity of normal human serum against E. coli Lilly. With a "high" dose of drugs, mechlorethamine, daunorubicine, and cytarabine definitely enhanced early stages of bacteriolytic activity. Enhancement of bactericidal activity of normal human serum against E. coli Lilly was achieved by admixture with 1,3-bis(2-chloroethyl)-l-nitrosourea (SCN.U, carmustine) fluorouracil, methotrexate, vincrisUne, cytarabine, and prednisone. There was no enhancement of antibacterial activity by any drug against E. coli resistant to the action of normal human serum. Preincubation of E. coli Lilly with various drugs indicated that their enhancing activity was not expressed when the bacterial envelope was intact. Sera of patients receiving cyclophosphamid e intravenously did not differ in antibacterial activity from the samples drawn before the injection. Apparently certain chemotherapeutic agents influence antibacterial activity of the serum in vitro against some microorganisms. This fact should be considered when defense mechanismsof patients with leukemia or cancer are investigated.

Pseudomonas bacteremia: pharmacologic and other bases for failure of treatment with gentamicin.

The species Pseudomonas aeruginosa is sensitive to gentamicin, but among 21 patients treated for pseudomonas bacteremia, only six were cured and 15 (71%) died. Seven patients with a rapidly fatal underlying disease died; five of eight considered to have ultimately fatal diseases died, and three of six patients with nonfatal primary diseases died. The dose was not predictive of the level in serum, and most patients received a suboptimal dose. The concentration in the serum was directly related to eradication of bacteremia and survival of the patient. The protective level in serum corroborated the in-vitro sensitivity tests. Gentamicin had a slow maximal bactericidal rate on Pseudomonas. Leukemic leukocytes in leukemic serum were less effective than normal leukocytes in normal serum in killing P. aeruginosa. The high fatality rate from pseudomonas bacteremia results from inadequate treatment, resistance of the organism to killing, and defective defense mechanisms in the patients. Proper use of gentamicin should cure patients if they do not have fatal underlying diseases.

Detection of human leukaemia associated antigens in leukaemic serum and normal embryos.

THE occurrence of tumours or leukaemia in experimental animals is frequently associated with the appearance of “new” antigens perhaps indicative of gene activation or mutation, or infection with oncogenic virus. The reliable detection of such antigens in man would provide an immunological signal of incipient tumour formation and might permit early or more precise diagnosis. The study of human tumour antigens might also be expected to yield information about the causes and natural history of human cancer as well as suggest improved means of treatment. This communication is concerned with the detection of one group of “new” antigens, those associated with human leukaemia.

Acrylamide gel electrophoretic abnormalities in chronic lymphatic leukemic sera.

Using disk acrylamide gel electrophoresis, an abnormal pattern was found in all sera from patients with chronic lymphatic leukemia but in none of the sera from normal persons or patients with lymphosarcoma. The abnormality is the presence of an intense band (X) cathodal to the usual pretransferrin band (P) of normal sera. The P bands were either absent or very weak in all sera of the patients with chronic lymphatic leukemia.

Effect of splenic X-irradiation on bone marrow function: experimental studies of a serum mitotic inhibitor in rabbit and in patients with chronic myelocytic leukaemia.

Summary. The effect of a selective splenic irradiation on bone marrow function was studied in rabbits given a single splenic exposure with 300 r. of X‐rays. Serum collected shortly thereafter decreased mitotic activity in suspensions of normal rabbit bone marrow. This in vitro inhibitory effect was only transiently demonstrable in the serum appearing 2 hours after irradiation and disappearing 3 hours later.Five patients with chronic myelocytic leukaemia in relapse receiving splenic radiotherapy in large single doses of 200–400 r. (surface dose) were studied along these lines. Serum collected 3 hours after X‐ray treatment was tested with suspensions of normal bone marrow and was found to decrease erythroblastic mitotic activity significantly below the control values observed with serum collected before irradiation. The magnitude of this in vitro effect was to some extent related to the spleen size and the radiation dose. No differences were observed between bone marrow suspensions incubated with pre‐irradiation leukaemic serum and normal control serum.

Control of DNA duplication in rat chloroleukaemia by means of the granulocytic chalone

It is known that the mitotic activity of the normal granulocytic progenitor cells is inhibited by a tissue-specific granulocytic chalone which has Ve Vo =2·73±0·035 on Sephadex G- 75. Using 3 H-thymidine and 14 C-formate and a liquid scintillation counter together with autoradiography, it has now been shown that this same chalone also exists in chloroleukaemic cells and that it is capable of inhibiting their mitotic activity. However, the leukaemic cells are not as sensitive as are the normal progenitor cells. It has been found that the chalone concentration in a chloroleukaemic cell is 1 10 – 1 40 of that in a normal mature granulocyte and that the leukaemic cells probably also synthesize less chalone. The chalone concentration in the leukaemic serum is higher than that in normal serum. This is evidently due to a high rate of chalone loss from the leukaemic cells combined with the large numbers of these cells that are present in the body. In spite of the reduced chalone concentration within the cells the mitotic rate of the chloroleukaemic cells is lower than that of the normal progenitor cells. Their high rate of cell production is due solely to their ‘delayed maturation’. From evidence given here, and from other evidence, it is clear that whether the granulocytic chalone is derived from normal or leukaemic cells, it is tissue-specific since it has no action on the mitotic activity of erythrocytic progenitor cells. Its action is fully reversible since it causes a delay rather than a permanent inhibition of the cell proliferation rate. Although the chalone causes a strong inhibition of DNA synthesis in both normal and leukaemic cells, it inhibits RNA synthesis only in normal cells. The chloroleukaemic cells utilize exogenous thymidine less effectively and exogenous formate more effectively than do normal cells. The main results and conclusions given here regarding chloroleukaemia in rats are essentially similar to those obtained with the V× 2 epidermal tumour of rabbits and with melanomata of both mice and hamsters.

Antilymphocytic leukemic serum and chemotherapy in the treatment of murine leukemia.

Antiserum against L 1210 murine leukemia was prepared in rabbits and goats. Murine antibody titers against such antisera were lower than against normal foreign serum, suggesting an immunosuppressive effect. Additional suppression of antibody formation was obtained by the concomitant use of cyclophosphamide. The antiserum had an equally cytotoxic effect against normal and leukemic lymphocytes in vitro. The antileukemic effect in vivo was demonstrated by a statistically significant but slight prolongation of survival time of leukemia-bearing mice. The results of therapy with the antileukemic serum were enhanced in some experiments by the incorporation of cyclophoshamide and 6-mercaptopurine into the treatment schedule.

Chieco-Bianchi L,+CHIECOAABIANCHI L: [Cytoxic activity in vitro of normal and leukemic mouse antileukocytic serum

These experiments were designed to evaluate the specificity of cytotoxic antibodies for normal and leukemic leukocytes from mice with different genotypes. C57BL and C3Hf/Gs mice were immunized with allogeneic splenic cells from C3Hf/Gs, C57BL and (C3Hf/Gs x CBA T6T6) F1 donors. The animals were injected intraperitoneally once a week for 6–7 weeks and serum was obtained 7 days after the last injection. The cytotoxic effect of the sera on leukocytes was evaluated using the Terasaki test, which is based on direct phase contrast observation of refraction modification in damaged cellular elements. By using donors and recipients with dissimilar H-2 locus, an increase in the cytotoxic index was observed which was related to the number of immunizing injections. Values were fairly uniform for the different combinations tested. The maximum antibody titer was obtained 7 days after the last injection with notable decrease after 14 and 30 days. Cross tests between antisera and leukocytes with the same or different H-2 allele have demonstrated a high specificity of the isoantisera for the different H-2 locus alleles (k, b). In another group of experiments, the effect of isoantisera on leukocytes from animals with generalized leukemia was tested. The leukemia was induced by inoculation at birth with either Graffi or Gross virus. In the 12 cases studied, the cytotoxic effect on leukemic cells was always less than on normal cells of the same strain. Finally, a group of experiments was carried out using leukemic anti-leukocyte sera obtained by immunizing C57BL animals with cells derived from 2 Graffi virus-induced leukemias grafted into (C3Hf/Gs x CBA T6T6) F1 mice. These sera presented maximum cytotoxic index values on leukocytes of (C3Hf/Gs x CBA T6T6) F1 animals with Graffi virus-induced leukemia. Lower values were observed for normal leukocytes of the same strain. (C3Hf/Gs x CBA T6T6) F1 cells originating from Gross virus-induced leukemia were less sensitive to these sera, not only in comparison with Graffi virus-induced leukemia leukocytes but with normal (C3Hf/Gs x CBA T6T6) F1 leukocytes as well. It was also possible to demonstrate positivity in the cytotoxic response with C57BL leukemic leukocytes (Graffi virus-induced). These data seem to indicate that a reduction in the histocompatibility antigens of the leukemic leukocyte occurs contemporaneously with the appearance of tumor-specific antigens, and both phenomena are most likely related to neoplastic transformation. The following hypotheses are discussed as possible explanations of these phenomena: the possibility of true antigen deletion; the spreading out of membrane antigenic receptors as a consequence of increased cellular volume; the appearance of the TL antigen in the leukemic cells causing a partial loss of H-2 antigens.