epidermal-growth-factor and Leukemia--Promyelocytic--Acute

P-selectin is an integral membrane glycoprotein on stimulated platelets and endothelial cells that serves as a receptor for leukocytes. To estimate the density of P-selectin in membranes necessary to support adhesion, we incorporated purified P-selectin at varying concentrations into phospholipid bilayers that encapsulated glass microspheres. Maximal binding of these lipospheres to HL60 cells, a P-selectin ligand-expressing cell line, was approached at a P-selectin density of about 100 molecules per microns 2; half-maximal binding was observed at about 50 to 60 molecules per microns 2. Compatible results were obtained with P-selectin expressed on Chinese hamster ovary cells. The P-selectin density on stimulated platelets was estimated to be 150 to 200 molecules/microns 2. To identify the domains of P-selectin required for HL60 cell binding, chimeras of P-selectin and L-selectin were stably expressed in Chinese hamster ovary cells and clones that expressed the chimeras at the estimated physiologic density were selected. Chimeras containing the P-selectin lectin and epidermal growth factor (EGF) domains or the lectin, EGF, and short consensus repeats bound HL60 cells equivalently, but a chimera containing the P-selectin lectin domain alone bound HL60 cells much less well. These results indicate that at a physiologically relevant P-selectin density on membrane surfaces, the lectin, and EGF domains of P-selectin are together required for optimal leukocyte binding.

Topics: Animals; Blood Platelets; Cell Adhesion; Cell Adhesion Molecules; CHO Cells; Cricetinae; Epidermal Growth Factor; Glass; Humans; L-Selectin; Lectins; Leukemia, Promyelocytic, Acute; Leukocytes; Lipid Bilayers; Liposomes; Microspheres; P-Selectin; Platelet Membrane Glycoproteins; Recombinant Fusion Proteins; Structure-Activity Relationship; Transfection

We studied 18 patients with acute promyelocytic leukaemia and 13 with relapsed APL. We found a significantly elevated EGF in acute leukaemia, especially in APL, being 418.59 +/- 19.2 micrograms in the 24-h urine that was much higher than that of the normal controls. When eight APL patients achieved complete remission by RA treatment, the EGF value decreased to 149.9 +/- 27.3 micrograms in the 24-h urine near to normal. In 13 patients with relapsed APL, EGF rose to 446.9 +/- 82.6 micrograms in the 24-h urine. Most interestingly, this elevated EGF could be detected before the relapse by 5 +/- 0.84 months in seven out of eight APL with relapse. We suggest that the unaccountably elevated EGF during remission period may be an indicator of the occurrence of relapse.

Topics: Acute Disease; Adult; Aged; Base Sequence; Biomarkers, Tumor; Cell Differentiation; Epidermal Growth Factor; Female; Humans; Immunologic Factors; Leukemia, Myeloid; Leukemia, Promyelocytic, Acute; Male; Middle Aged; Molecular Sequence Data; Neoplasm Proteins; Neoplasm Recurrence, Local; Neoplasm, Residual; Oncogene Proteins, Fusion; Precursor Cell Lymphoblastic Leukemia-Lymphoma; Remission Induction; Tretinoin

Mitogenic stimulation of quiescent cells not only triggers the cell division cycle but also induces an increase in cell volume, associated with an activation of cellular metabolism. It is therefore likely that genes encoding enzymes and other proteins involved in energy metabolism and biosynthetic pathways represent a major class of mitogen-induced genes. In the present study, we investigated in the non-established human fibroblast line WI-38 the induction by mitogens of 17 genes whose products play a role in different metabolic processes. We show that these genes fall into 4 different categories, i.e. non-induced genes, immediate early (IE) primary genes, delayed early (DE) secondary genes and late genes reaching peak levels in S-phase. In addition, we have analysed the regulation of these genes during normal cell cycle progression, using HL-60 cells separated by counterflow elutriation. A clear cell cycle regulation was seen with those genes that are induced in S-phase, i.e. thymidine kinase, thymidylate synthase and dihydrofolate reductase. In addition, two DE genes showed a cell cycle dependent expression. Ornithine decarboxylase mRNA increased around mid-G1, reaching maximum levels in S/G2, while hexokinase mRNA expression was highest in early G1. In contrast, the expression of other DE and IE genes did not fluctuate during the cell cycle, a result that was confirmed with elutriated WI-38 and serum-stimulated HL-60 cells. These observations suggest that G0-->S and G1-->S transition are distinct processes, exhibiting characteristic programmes of gene regulation, and merging around S-phase entry.

Topics: Base Sequence; Cell Cycle; Cell Line; Culture Media; Cycloheximide; DNA Primers; Epidermal Growth Factor; Gene Expression; Gene Expression Regulation; Gene Expression Regulation, Enzymologic; Genes, Immediate-Early; Hexokinase; Humans; Leukemia, Promyelocytic, Acute; Lung; Models, Biological; Molecular Sequence Data; Ornithine Decarboxylase; Platelet-Derived Growth Factor; Polymerase Chain Reaction; RNA; RNA, Messenger; Tetrahydrofolate Dehydrogenase; Thymidine Kinase; Thymidylate Synthase; Tumor Cells, Cultured

A human promyelocytic cell line, HL-60, treated with activin/EDF was found to differentiate into monocyte/macrophage-like cells. This was shown not only by morphology but by the loss of myeloperoxidase granules and the appearance of nonspecific esterase. Dose-dependent inhibition of the differentiation by follistatin, an activin-binding protein, confirmed that it was indeed caused by activin. Thus, activin/EDF exerts its effect on hematopoietic cells not only on erythroid differentiation but also on at least a part of myeloid cell differentiation.

Topics: Activins; Binding Sites; Cell Differentiation; Cytoplasmic Granules; Epidermal Growth Factor; Follistatin; Glycoproteins; Histocytochemistry; Humans; Inhibins; Leukemia, Promyelocytic, Acute; Peroxidase; Tumor Cells, Cultured

A major peak of tyrosine protein kinase activity was partially purified from a Triton X100 extract of HL-60. This preparation submitted to high pressure gel filtration was eluted at a volume corresponding to a mass of 35/40 kD. This activity was insensitive to EGF and insulin. Autoradiographs of the preparations incubated with [gamma P32]-ATP and separated by electrophoresis do not give any evidence that autophosphorylation occurs for that particular tyrosine protein kinase. Furthermore, we failed to immunoprecipitate the enzyme with a specific antiphosphotyrosine antibody and anti v-src antibody. All the data presented herein suggest that this enzyme has not been previously purified.

Topics: Adenosine Triphosphate; Autoradiography; Binding, Competitive; Chromatography, High Pressure Liquid; Electrophoresis, Agar Gel; Epidermal Growth Factor; Guanosine Triphosphate; Humans; Immunosorbent Techniques; Insulin; Leukemia, Promyelocytic, Acute; Molecular Weight; Octoxynol; Phosphorylation; Polyethylene Glycols; Protein-Tyrosine Kinases; Substrate Specificity; Tumor Cells, Cultured