sodium-dodecyl-sulfate and Leukemia--Myeloid

The effect of sodium dodecyl sulfate (SDS), an anionic amphiphilic detergent, on the function of human neutrophils and of the human promyelocytic leukemia cell line HL-60 was investigated. SDS modulated the respiratory burst in human neutrophils and HL-60 cells which were stimulated with phorbol 12-myristate 13-acetate (PMA). In concentrations above 1 X 10(-6) M it also caused release of lysosomal enzymes (beta-D-glucuronidase, myeloperoxidase and lysozyme) from neutrophils. Our results demonstrate that SDS at concentrations 1 X 10(-6) M-1 X 10(-4) M strongly affect properties of human phagocytic cells.

Topics: Adjuvants, Immunologic; Cell Division; Glucuronidase; Humans; Leukemia, Myeloid; Lysosomes; Neutrophils; Peroxidase; Respiratory Burst; Sodium Dodecyl Sulfate; Superoxides; Tetradecanoylphorbol Acetate; Thymidine; Tumor Cells, Cultured

Many functions of polymorphonuclear leukocytes (PMNs) appear to alter and be affected by changes in the intracellular and/or extracellular acid-base milieu, suggesting that carbonic anhydrase (CA) may be important. Although small amounts of CA activity in PMNs have been reported, it has not been characterized fully. We therefore studied isolated mature circulating human PMNs and cultured HL-60 cells, an undifferentiated myelopoietic cell line, and compared these to human red cells (RBCs) for CA activity. Activity and sulfonamide inhibition were measured by a modified micromethod assay. Isoenzyme and total CA concentrations were determined by radioimmunoassay for human isozyme CA I, differential inhibition by MK-927, inhibition by 0.2% sodium dodecyl sulfate (SDS), and quantitative sulfonamide binding. Total CA activity (units/10(6) cells) was 0.04 in PMNs, 0.06 in HL-60 cells, and 0.62 in RBCs. Human PMNs have a total CA concentration of 1.3 microM, of which 0.9 microM is CA I and the remainder is CA II. Total loss of CA activity with 100 microM ethoxzolamide and 0.2% SDS ruled out significant CA III or CA IV activity. Subcellular fractionation of PMNs revealed that all CA activity was cytosolic. The absence of CA activity in mitochondrial and microsomal membrane fractions argues against any mitochondrial CA V or membrane-bound CA IV contribution to total CA activity. Neutrophils contain both CA I and II isozymes in roughly the same proportion as RBCs but at much lower concentrations, suggesting that in the course of maturation the CA content of neutrophils is regulated differently from that in erythrocytes.

Topics: Carbonic Anhydrase Inhibitors; Carbonic Anhydrases; Chromatography, High Pressure Liquid; Electrophoresis, Polyacrylamide Gel; Erythrocytes; Ethoxzolamide; Humans; Immunoblotting; Isoenzymes; Leukemia, Myeloid; Neutrophils; Radioimmunoassay; Sodium Dodecyl Sulfate; Sulfonamides; Thiophenes; Tumor Cells, Cultured

In this work granulocytes from normal human donors and patients suffering from chronic myeloid leukemia (CML) were externally labeled with 125Iodine, using the Iodogen method. 125Iodine labeled Concanavalin A binding proteins (CBP) and detergent-resistant proteins (DRP) were isolated from the cell lysates and characterized by one- and two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis (1D- and 2D-SDS-PAGE). Autoradiographs of the 2D-gels of DRP show seven proteins with Mr 118,000 (spot 1 a), Mr 112,000 (spot 1b), Mr 78,000-85,000 (spot 2), Mr 85,000 (spot 4), Mr 52,000 (spot 3, 3 a and 3 b). Of this set, spot 1 b, 2 and 4 are also present in the autoradiographs of 2D-gels of CBP and, hence, may be considered to be transmembrane components. Spot 4 is expressed more intensely in the normal granulocytes while spots 3 a and 3 b are mainly expressed on the leukemic granulocytes.

Topics: Autoradiography; Cell Fractionation; Cytoskeleton; Detergents; Electrophoresis, Polyacrylamide Gel; Granulocytes; Humans; Iodine Radioisotopes; Leukemia, Myeloid; Membrane Proteins; Molecular Weight; Peptide Mapping; Receptors, Concanavalin A; Sodium Dodecyl Sulfate; Surface-Active Agents

Complement receptor activity for cell bound C3b and C3d was detected on plasma membrane fragments prepared by nitrogen cavitation from cultured human lymphoid cells. The activity of the membrane fragments reflected the activity of the whole cells in that cells which did not form rosettes (P3J and RPMI 4098) resulted in inactive membranes and cells with high rosette formation (NC37 and Raji) yielded highly active membrane fragments. Two test systems were devised to detect these receptor activities, namely a rosette inhibition and a hemagglutination assay. Solubilization of C3 receptors was accomplished by extraction of active plasma membrane fragments with 2 MKBr. Dissociation and reassociation experiments suggest C3b and C3d receptors to be highly complex molecular structures. It appears that these complement receptors on plasma membranes rely on both protein and lipid moieties for the expression of their activity.

Topics: Binding Sites; Burkitt Lymphoma; Butanols; Cell Fractionation; Cell Line; Cell Membrane; Cells, Cultured; Complement C3; Complement System Proteins; Deoxycholic Acid; Electrophoresis, Polyacrylamide Gel; Erythrocytes; Ethanol; Hemagglutination Tests; HEPES; Humans; Immune Adherence Reaction; Leukemia, Myeloid; Lipids; Lymphocytes; Neoplasm Proteins; Polyethylene Glycols; Sodium Dodecyl Sulfate; Sucrose

Topics: Amino Acids; Aminocaproates; Animals; Blood Proteins; Centrifugation, Zonal; Chromatography, Gel; Chromatography, Ion Exchange; Cytoplasmic Granules; Dextrans; DNA; Electrophoresis, Polyacrylamide Gel; Humans; Immune Sera; Immunodiffusion; Immunoelectrophoresis; Leukemia, Myeloid; Leukocytes; Molecular Weight; Muramidase; Polysaccharides; Rabbits; RNA; Sodium Dodecyl Sulfate; Ultrafiltration

Topics: Animals; Antigens, Neoplasm; Burkitt Lymphoma; Cell Line; Chromatography, Affinity; Concanavalin A; Electrophoresis, Polyacrylamide Gel; Humans; Immune Sera; Iodine Radioisotopes; Leukemia; Leukemia, Myeloid; Mercaptoethanol; Molecular Weight; Papain; Rabbits; Sodium Dodecyl Sulfate; Urea

Topics: Acrylamides; Burkitt Lymphoma; Cell Nucleus; Cell Transformation, Neoplastic; Cells, Cultured; DNA, Neoplasm; Electrophoresis; Genetic Code; Humans; In Vitro Techniques; Lectins; Leukemia; Leukemia, Lymphoid; Leukemia, Monocytic, Acute; Leukemia, Myeloid; Lymphocytes; Neoplasm Proteins; Nucleoproteins; Sodium Dodecyl Sulfate; Thymidine; Tritium

Topics: Amino Acids; Binding Sites; Blood Proteins; Carbohydrates; Carrier Proteins; Centrifugation, Density Gradient; Chromatography, Affinity; Chromatography, Gel; Electrophoresis, Disc; Electrophoresis, Polyacrylamide Gel; Humans; Leukemia, Myeloid; Leukocytes; Molecular Weight; Polysaccharides; Protein Binding; Sodium Dodecyl Sulfate; Spectrophotometry; Ultracentrifugation; Vibration; Vitamin B 12