cytochalasin-b and Leukemia--Lymphoid

We have previously demonstrated that the murine monoclonal antibody T101 induces antigenic modulation when infused into patients with chronic lymphocytic leukemia and cutaneous T-cell lymphoma. In this paper, we extend our studies of T101-induced modulation and compare it to T101-induced capping. We found that, in contrast to antigenic modulation, capping occurred only in the presence of secondary anti-mouse IgG antisera and was altered by drugs that affect the cellular cytoskeleton or energy metabolism. F(ab')2 fragments of T101 induced antigenic modulation with kinetics similar to those of intact T101, but Fab-induced modulation proceeded more slowly and required the continual presence of Fab throughout the incubation. Experiments with radioiodinated T101 demonstrated that initial internalization of the antibody is followed by rapid efflux of intact, immunoreactive T101 from the cells. These data indicate important differences between capping and modulation and suggest that these two phenomena proceed by different mechanisms. More importantly, the data have implications for the potential therapeutic use of monoclonal antibody immunoconjugates.

Topics: Antibodies, Monoclonal; Antigen-Antibody Reactions; Antigens, Differentiation, B-Lymphocyte; Colchicine; Cytochalasin B; Cytoskeleton; Endocytosis; Immunologic Capping; Kinetics; Leukemia, Lymphoid; Membrane Glycoproteins

Peripheral blood lymphocytes from eight patients with chronic lymphocytic leukemia (CLL) were cultured with Epstein Barr virus (EBV) and cytochalasin B. All eight cytochalasin B cultures had analyzable metaphases whereas only six of the EBV cultures were successful. Furthermore, the number of abnormal metaphases and the mitotic indices were greater in the cytochalasin B cultures than in the EBV cultures. Trisomy 12, alone or in combination with other abnormalities, was the most frequent cytogenetic finding. Structural abnormalities of chromosomes #6 and #14 were also found. Cytochalasin B appears to be an effective mitogen for demonstrating abnormal metaphases in patients with CLL.

Topics: Aged; Cells, Cultured; Chromosome Aberrations; Cytochalasin B; Female; Herpesvirus 4, Human; Humans; Karyotyping; Leukemia, Lymphoid; Lymphocyte Activation; Lymphocytes; Male; Middle Aged; Mitogens

The human acute promyelocytic leukemia cell line HL-60 is induced by retinoic acid (RA) and N,N-dimethylformamide (DMF) to differentiate into cells having many of the functional and morphologic characteristics of mature granulocytes. With normal human phagocytic cells there is both superoxide anion (O2-) production and chemotaxis in response to chemoattractants such as N-formyl-methionyl-leucyl-phenylalanine (FMLP). We have now found that although HL-60 cells induced with RA alone produce O2- in response to 12-0-tetradecanoyl-phorbol-13-acetate (TPA) they are deficient in FMLP-stimulated O2- production and chemotaxis. In contrast, HL-60 induced either with DMF or with a combination of 10 nmol/L RA and a T cell-derived lymphokine, differentiation-inducing activity (DIA), produce O2- and exhibit chemotaxis in response to FMLP. The basis for these results appears to be the concentration of cell surface chemotactic peptide receptors. Thus, untreated HL-60 and HL-60 induced with either RA alone or DIA alone do not have measurable levels of FMLP receptors, whereas HL-60 induced with a combination of RA and DIA has 5,400 receptors per cell. HL-60 induced with RA and DIA plus 1 mumol/L dexamethasone have 25,000 receptors per cell and have greater chemotactic activity than HL-60 induced with the combination of RA and DIA. Thus, differentiation of HL-60 to cells with many properties of normal phagocytes can be induced in vitro by physiologic substances.

Topics: Cell Adhesion; Cell Differentiation; Cell Line; Chemotaxis; Cytochalasin B; Dimethylformamide; Humans; Kinetics; Leukemia, Lymphoid; Lymphokines; N-Formylmethionine Leucyl-Phenylalanine; Phagocytes; Receptors, Formyl Peptide; Receptors, Immunologic; Superoxides; Tetradecanoylphorbol Acetate; Tretinoin

It is widely accepted that the neoplastic B cells from patients with chronic lymphocytic leukemia (CLL) respond poorly to common mitogens. The fungal metabolite cytochalasin B (0.5 micrograms/ml) is a weak mitogen for normal lymphocytes. However, when peripheral blood lymphocytes from 19 patients with CLL of B cell origin (B-CLL) were cultured with 0.5 micrograms cytochalasin B/ml, significant new DNA synthesis ( [14C]thymidine incorporation) occurred in 18. Stimulation indices with cytochalasin B varied widely (range = 1.9-28.2, mean +/- SD = 10.6 +/- 7.5; delta cpm range = 1,157-153,818; n = 26) but in 11 cases exceeded those seen with concanavalin A (Con A), phytohemagglutinin, or pokeweed mitogen. In all 11, the mitogenic response to cytochalasin B exceeded that to pokeweed mitogen, which is believed to be a T cell-dependent B cell mitogen. In three cases, the responses to cytochalasin B were 8.6, 3.5, and 2.3 times greater than those to Con A. As with other mitogens, the DNA synthetic response to cytochalasin B was time and dose dependent. Peak thymidine incorporation occurred at 72-88 h and declined thereafter. Significant mitogenic effects were observed with 0.1-5 micrograms cytochalasin B/ml with a peak at 0.5-2 micrograms/ml. Stimulated DNA synthesis was abolished by 1 mM hydroxyurea. Cells from two patients with B-CLL were separated by rosetting with sheep erythrocytes (E). Depletion of E-rosette-positive cells from the CLL cell population abolished the response to Con A but did not affect the response to cytochalasin B. Cytochalasin B is a potent mitogen for B-CLL cells and may be useful in cytogenetic studies of this often indolent neoplasm.

Topics: Cell Transformation, Neoplastic; Cytochalasin B; Cytochalasins; Dimethyl Sulfoxide; DNA; Dose-Response Relationship, Immunologic; Female; Humans; Karyotyping; Kinetics; Leukemia, Lymphoid; Lymphocyte Activation; Male; Mitogens; Rosette Formation

Pre-B cells from the bone marrow of children with acute lymphoblastic leukaemia (ALL) survived up to 144 hr after the completion of treatment and divided in culture with maximum cell numbers at 24 hr. There was no rise in B cell number and no evidence of differentiation from pre-B to B cells. Binucleated pre-B cells in cultures containing cytochalasin B confirmed that pre-B cell division was occurring. Cycloheximide reduced cell numbers in culture but bromodeoxyuridine did not. Pre-B cell numbers also increased in culture of morphologically normal marrows from treated and untreated patients with solid tumours, and probably in normal marrows from patients with non-malignant diseases.

Topics: Adolescent; B-Lymphocytes; Bromodeoxyuridine; Cell Division; Cell Survival; Cells, Cultured; Child; Child, Preschool; Colchicine; Cycloheximide; Cytochalasin B; Hematopoietic Stem Cells; Humans; Leukemia, Lymphoid

A factor on human lymphocytes has been identified as factor B of the alternative pathway. Lymphocytes can replace factor B in the fluid phase formation of C3 convertase with cobra venom factor (CVF). This lymphocyte activity is inhibited by specific anti-human factor B, and it is shown by Burkitt lymphoma cell lines cultured in the absence of any factor B source. After the reaction with CVF all the C3-converting activity is found in the cell supernatant, and the same cells can undergo several successive cycles of 'activation' by CVF. Factor B is distinct from the C3b receptor, and its presence could not be detected antigenically on the lymphocyte membrane. It may be secreted by the cells, but the reaction was not affected by sodium azide or cytochalasin B. No detectable factor B activity was found in the culture medium of cells grown in the absence of CVF.

Topics: Animals; Burkitt Lymphoma; Cell Line; Cells, Cultured; Complement C3; Complement System Proteins; Culture Media; Cytochalasin B; Edetic Acid; Humans; Immune Sera; Leukemia, Lymphoid; Lymphocytes; Peptide Hydrolases; Phospholipases; Properdin; Snakes; Specimen Handling; Thymus Extracts; Trypsin; Venoms

The characteristics of human lymphocyte motility and its relationship to the redistribution of surface membrane antigens (capping) are poorly defined. Since chronic lymphocytic leukemia (CLL) cells cap poorly when compared with normal human lymphocytes, this study was undertaken to compare the motility of these two cell types. A modification of the Boyden chamber system was employed to quantify lymphocyte motility by placing lymphocyte suspensions on 8-mum convoluted-pore nitrocellulose filters and measuring the depth of migration of the cells into the filter at 37 degrees C. After 3 hr of incubation, CLL cells migrated significantly less into the filter than normal cells. Incubation in the presence of sodium azide or at 4 degrees C abolished all motility, indicating the active nature of the process. The relative motility of individual CLL patients' cells correlated best with the proportion of abnormal cells present as determined by surface receptor assays. The possibility that decreased cell motility in CLL was a reflection of enrichment by a "bone marrow-derived" (B cell) population was eliminated by the finding that normal B cells purified by gradient separation of rosetted cells migrated faster than normal T cells and considerably faster than CLL cells. Motility of normal and CLL lymphocytes was decreased by cytochalasin B and increased by colchicine, vincristine, and vinblastine. Thus, human lymphocyte motility appears to be dependent on microfilament integrity but not to require the colchicine-sensitive cytoskeleton. The decreased motility of CLL cells is the result of an intrinsic cell abnormality, but this finding cannot fully explain the decreased capping, since in human lymphocytes the latter is not prevented by an inhibitor of motility.

Topics: Adult; Azides; B-Lymphocytes; Cell Movement; Cell Separation; Colchicine; Cytochalasin B; Humans; Incubators; Leukemia, Lymphoid; Lymphocytes; Middle Aged; T-Lymphocytes; Temperature; Time Factors

In health up to 6% of human lymphocytes will form rosettes with homologous group O rhesus-negative erythrocytes (H rosettes). Increased numbers of H rosettes were found in T-cell-proliferative diseases-- namely, infectious mononucleosis, Sézary's syndrome, and T-cell leukaemia. H-rosette formation is thus a marker for a subpopulation of T lymphocytes, and this easily performed test of T-cell activation may have clinical value in characterising changes in blood lymphocyte populations in disease.

Topics: ABO Blood-Group System; Colchicine; Cytochalasin B; Humans; Immune Adherence Reaction; Infectious Mononucleosis; Isoantibodies; Leukemia, Lymphoid; Lymphatic Diseases; Lymphocyte Activation; Lymphocyte Culture Test, Mixed; T-Lymphocytes; Vinblastine

The phenomenon of redistribution of surface membrane immunoglobulin (Ig) components (capping) has been well described in mouse lymphoid cells. The characteristics of this process in human lymphocytes are less clear. This study characterizes the phenomenon of surface membrane Ig redistribution of normal and chronic lymphocytic leukemia (CLL) lymphocytes with the use of fluoroscein-labeled anti-Ig sera. Normal lymphocytes underwent rapid cap formation after incubation with anti-Ig serum in the cold and subsequent rewarming. The morphology was characteristic with aggregation over the pole of the cell opposite the nucleus and over the uropod when present. The process was energy dependent but independent of protein synthesis, and could be inhibited by vincristine, vinblastine, and colchicine but not by cytochalasin B. CLL cells, on the other hand, though showing fluorescent complex aggregation on the surface, rarely demonstrated unidirectional movement of these aggregates to form a cap. Cap formation in these cells could not be stimulated by supplementing the energy source or protein concentration of the medium nor by adding glutamic acid which could partially reverse the vincristine and vinblastine inhibition of normal capping. The failure of agents which inhibit motility to inhibit capping of the normal lymphocytes suggests that active locomotion is not a direct prerequisite for capping. The results also suggest the involvement of microtubules in normal capping and the possibility that abnormal membrane structure or microtubular function could explain the failure of CLL cells to behave normally in this regard. The role of this cellular defect in the immune deficiencies exhibited by many patients with CLL, however, is not established.

Topics: Animals; Antibodies, Anti-Idiotypic; Azides; Cattle; Cell Membrane; Cell Survival; Colchicine; Culture Media; Cyanides; Cytochalasin B; Depression, Chemical; Fluoresceins; Fluorescent Antibody Technique; Glutamates; Humans; Immunoglobulins; Leukemia, Lymphoid; Lymphocytes; Microscopy, Fluorescence; Puromycin; Thiocyanates; Time Factors; Vinca Alkaloids

Topics: Agammaglobulinemia; Antibodies, Anti-Idiotypic; Antigen-Antibody Complex; Azides; Binding Sites, Antibody; Cell Membrane; Cytochalasin B; Epitopes; Fluorescent Antibody Technique; Humans; Immunoglobulin G; Immunoglobulins; Leukemia, Lymphoid; Lymphocyte Activation; Lymphocytes; Pinocytosis; Pronase; Sarcoidosis; Sodium; Trypsin

Topics: Acridines; Alkaloids; Animals; Antineoplastic Agents; Cell Adhesion; Cell Division; Cell Line; Cell Nucleus; Cells, Cultured; Cyclic AMP; Cytochalasin B; Female; Golgi Apparatus; Humans; Leukemia, Experimental; Leukemia, Lymphoid; Melanoma; Mice; Microscopy, Electron; Mitochondria; Mitosis; Simian virus 40; Time Factors; Tumor Virus Infections; Uterine Cervical Neoplasms