agar and Leukemia--Myeloid

The in vitro cloning of haemopoietic precursors is a rapidly growing field. The data reviewed above and the current practical applications of the techniques can be expected to increase quite rapidly in the next decade. Despite the technical problems of tissue culture and the special problems associated with culturing human cells, it is clear that these procedures can be effectively applied at the clinical level. The value of the data obtained will vary directly with the quality of the culture techniques. Any centre undertaking these techniques must be prepared to properly equip the culture laboratory, appoint a full-time staff member for the work and maintain a constant surveillance of the quality of the culture work.

Topics: Agar; Anemia, Aplastic; Cells, Cultured; Colony-Forming Units Assay; Colony-Stimulating Factors; Culture Media; Granulocytes; Hematopoietic Stem Cells; Humans; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Monocytes; Myeloproliferative Disorders; Polycythemia Vera; Preleukemia; Primary Myelofibrosis; Time Factors

Haematopoietic stem cells are capable of developing colonies of differentiated granulocytes in a semi-stable agar environment under corresponding experimental conditions. Number and size of aggregations developing from a single stem cell in each case called "in culture Colony Forming Unit (CFU-c)" enable conclusions to be made about the functional ability of the cultivated human or animal bone marrow. The method is applied in investigating the kinetics in the haematopoietic system and, in addition, it is used for diagnostics, therapy and control of the course in haematological diseases as well as to check the proliferous ability of cryopreserved bone marrow cells. As to the standardization of the procedure the instability of the active principle of the foetal calf serum as well as the essential colony stimulating factor represent limiting items at present.

Topics: Agar; Animals; B-Lymphocytes; Bone Marrow Cells; Cell Differentiation; Cell Division; Cells, Cultured; Chick Embryo; Colony-Stimulating Factors; Culture Techniques; Hematopoietic Stem Cells; Humans; Leukemia, Myeloid; Methylcellulose; Mice; T-Lymphocytes; Thymus Gland

Topics: Agar; Cell Transformation, Neoplastic; Cells, Cultured; Chromosomes, Human, 21-22 and Y; Hematopoietic Stem Cells; Humans; Leukemia, Myeloid; Time Factors

Topics: Agar; Animals; Antigens; Blood; Bone Marrow Cells; Cell Division; Clone Cells; Culture Techniques; Humans; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Leukocytes; Macrophages; Methods; Mice

Topics: Agar; Bone Marrow; Bone Marrow Cells; Cell Separation; Cells, Cultured; Centrifugation; Humans; Leukemia, Myeloid; Leukocytes; Monocytes

Topics: Acetamides; Acute Disease; Adult; Agar; Anti-Bacterial Agents; Bone Marrow Transplantation; Drug Resistance; Humans; Immunocompromised Host; Leukemia, Myeloid; Linezolid; Male; Microbial Sensitivity Tests; Oxazolidinones; Staphylococcal Infections; Staphylococcus epidermidis; Staphylococcus haemolyticus; Vancomycin

Cord plasma contains colony-stimulating activity (CSA) which stimulates the in vitro clonal growth of neutrophils, eosinophils, macrophages, erythrocytes, and persisting mast cells in semisolid cultures. Analysis of day 35 colonies in agar cultures was found to be a suitable means of demonstrating this activity and discriminating between it and granulocyte-macrophage colony-stimulating factor (GM-CSF). Serum (10%) from patients with acute and chronic myeloid leukemia (AML and CML) was added to normal human bone marrow cultures to search for similar activity in these patient's serum. Although the number of colonies on day 12 (predominantly neutrophils and macrophages) was not significantly different from the number of colonies in cultures containing normal serum, the number of colonies increased 500% in cultures containing CML serum on day 35. Serum from patients with AML during regeneration also stimulated an increased number of colonies on day 35. Although both eosinophil and mast cell colonies were still present on day 35, only mast cell colonies persisted for 150 days. On day 35, cultures containing 10% CML serum contained predominantly eosinophil colonies (84%), whereas cultures containing AML serum contained predominantly mast cell colonies (76%). Although serum contains various CSFs, the specific factor which stimulates persisting mast cell colonies may be the human equivalent of murine persisting (P) cell-stimulating factor (Multi-CSF).

Topics: Agar; Cells, Cultured; Colony-Forming Units Assay; Colony-Stimulating Factors; Fetal Blood; Granulocytes; Hematopoietic Stem Cells; Humans; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Macrophages

The density distribution in Percoll gradients of clonogenic cells forming colonies in diffusion chambers (CFU-D) or in agar culture (CFU-GM) was studied in chronic myeloid leukaemia (CML). The density distribution of CFU-GM in CML was homogeneous with peaks within 1.058-1.061 g/ml, which is slightly lower than normal. CFU-D, on the other hand, showed heterogeneous distributions both in CML and in normal controls. Two separable populations of CFU-D were recognized, one with the same or lower density than CFU-GM that formed almost exclusively neutrophilic colonies in diffusion chambers, and a second population concentrating in the density range 1.068-1.075 g/ml which primarily formed macrophage colonies. The second population contained colony-forming cells derived from both Fc-receptor positive and Fc-receptor negative precursor cells, suggesting that at least some colonies in diffusion chambers arise from Fc-receptor positive granulopoietic cells of intermediate maturity and/or monocytes. The concentration of CFU-D in peripheral blood was increased 40- to 100-fold in 2 patients currently off treatment who had increased WBC counts.

Topics: Agar; Bone Marrow; Cell Count; Cell Separation; Cells, Cultured; Centrifugation, Density Gradient; Culture Techniques; Diffusion; Humans; Leukemia, Myeloid; Neoplastic Stem Cells; Receptors, Fc; Stem Cells

Marrow or peripheral blood cells from 28 patients with acute myeloid leukemia (AML) or chronic myeloid leukemia in blastic crisis (CML-BC) were studied in both liquid and agar cultures. The proliferation and maturation of these cells were followed for 15-20 days in liquid culture with or without the addition of human placenta conditioned medium (HPCM) and/or retinoic acid (RA). In nine patients (group 1), cells underwent both proliferation and maturation, i.e., the percentage of peroxidase-positive cells (PO), phagocytic cells, and mature forms increased. For the remaining 19 patients (group 2), no proliferation was observed. However, 11 of these leukemic cell samples showed maturation (group 2A), while the eight others remained immature (group 2B). In agar culture, cell samples from group 1 showed cluster growth, group 2 no growth. Maturation without proliferation was observed for group-1 liquid cultures not containing HPCM and those containing HPCM and RA. The viability rapidly decreased in liquid cultures with only addition of RA. HPCM and RA showed no effect on group-2 cell cultures.

Topics: Adult; Agar; Aged; Bone Marrow; Cell Division; Cells, Cultured; Child; Culture Media; Culture Techniques; Follow-Up Studies; Humans; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Middle Aged; Phagocytosis; Placenta

The production of basophils in semisolid agar cultures form normal and chronic myeloid leukemia (CML) committed granulocyte-macrophage precursors was investigated using an original whole-dish staining technique with toluidine blue which produces a specific metachromasia in basophils. As additional proof of the basophilic nature of metachromatic cells, their degranulation after challenge with C5a anaphylotoxin and Synachten was observed. Our studies show that few basophils are produced in cultures form normal bone marrow. CML CFUc produce more basophils, their number being roughly correlated with the degree of basophilia. We observed only clusters composed by a pure basophilic population, while larger aggregates in which basophils could be detected were composed also by other granulocytic cells.

Topics: Agar; Anaphylatoxins; Basophils; Bone Marrow Cells; Cosyntropin; Culture Techniques; Cytoplasmic Granules; Humans; Leukemia, Myeloid; Staining and Labeling; Tolonium Chloride

T lymphocytic colony formation by peripheral lymphocytes separated by discontinuous albumin gradient centrifugation was evaluated in 8 patients with Philadelphia (Ph1)-positive chronic myeloid leukemia (CML). Colonies were obtained using a liquid-on-agar culture system recently introduced (PHA overlayer-leukocyte feeder layer assay) which has been shown to be simple and reliable. The pattern of colony growth in CML and in normal controls was similar, the peak ranging from the 4th to the 6th day. Also the morphological aspects of colonies did not differ in the two groups. Cells recovered from CML lymphocytic colonies were shown to belong to T cell lineage, as they are able to form spontaneous E-rosettes and to respond to mitogenic stimulation in vitro. In contrast, cells recovered from all other cultured fractions failed to display these properties. Cytogenetic analysis showed that T colony cells were Ph1-negative whereas the chromosome anomaly was found in nonlymphoid colonies of the same patients, thus suggesting a nonclonal origin of T lymphocytes in CML.

Topics: Agar; Cell Transformation, Neoplastic; Cells, Cultured; Centrifugation, Density Gradient; Chromosomes, Human, 21-22 and Y; Concanavalin A; Humans; Leukemia, Myeloid; Phytohemagglutinins; Pokeweed Mitogens; Rosette Formation; T-Lymphocytes

Human leukaemic cell specimens were obtained from patients and directly plated into soft agar (t = 0) or cultured for 1 week in liquid phase and then plated in soft agar. Growth for 1 week in liquid phase allowed the clonal growth in agar of leukaemic specimens which were unable to clone at t = 0. Clonal growth after liquid culture consisted of the usual leukaemic type of cluster-colonies, growth of a new type of 'syncytial' cell colony or a mixture of colony types. In addition, marrow from a patient with acute lymphocytic leukaemia produced normal-appearing colonies after 1 week of growth in liquid phase. These studies suggest a similarity in the growth requirements of some leukaemic cells and normal CFUd cells.

Topics: Adult; Agar; Aged; Cell Division; Cells, Cultured; Clone Cells; Culture Media; Female; Humans; Kinetics; Leukemia; Leukemia, Erythroblastic, Acute; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Leukocytes; Macrophages; Male; Middle Aged

Six patients with Philadelphia positive chronic myeloid leukaemia (CML) were treated with single high doses of busulphan. The action of busulphan on the in vivo kinetics of circulating progenitor cells (colony forming cells) was measured using an agar culture system which involved scoring of total colonies and clusters at 7 days and of granulocyte, monocyte and eosinophil colonies at 14 days. High dose busulphan was found to be effective in suppressing circulating granulocyte, monocyte and eosinophil progenitor cells. The effect of busulphan on progenitor cells was rapid and their levels fell by at least 85% within five days. By contrast, the white blood cells fell by only 9% and the platelets fell by 10% over this time. Subsequently, the white cell count and platelet count fell to near normal levels. The progenitor cell levels began to rise again at a mean of 35 days following busulphan treatment and the white blood cells at a mean of 39 days in four patients. One patient remained in haematological remission for six months following 100 mg of busulphan with less than 1 progenitor per 5 x 10(5) peripheral blood nucleated cells. One patient in myeloblastic transformation had a previously not described culture pattern consisting of a high cluster to colony ratio at 7 days and of an increased number of predominantly eosinophilic colonies at 14 days. There was no significant fall in progenitor cell levels following busulphan and this patient died.

Topics: Administration, Oral; Adult; Agar; Aged; Busulfan; Chromosomes, Human, 21-22 and Y; Culture Media; Dose-Response Relationship, Drug; Eosinophils; Female; Granulocytes; Hematopoietic Stem Cells; Humans; Leukemia, Myeloid; Male; Middle Aged; Monocytes

Topics: Agar; Cell Aggregation; Cell Line; Colony-Forming Units Assay; Colony-Stimulating Factors; Culture Media; Humans; Leukemia, Myeloid; Methylcellulose; Time Factors

Topics: Adult; Agar; Cell Division; Cells, Cultured; Colony-Stimulating Factors; Cytological Techniques; Hematopoiesis; Hematopoietic Stem Cells; Humans; Leukemia; Leukemia, Myeloid

Using a specific stain and electron microscopy, small numbers of mast cells were detected in human bone marrow cultures. However, they were not detected in agar cultures containing murine bone marrow cells. In bone marrow cultures from three patients with acute myeloid leukemia the number of mast cells was elevated.

Topics: Agar; Animals; Bone Marrow Cells; Cell Transformation, Neoplastic; Cells, Cultured; Cytoplasmic Granules; Humans; Leukemia, Myeloid; Mast Cells; Mice; Mice, Inbred BALB C; Mice, Inbred C57BL; Staining and Labeling

Infection of long-term BDF1 marrow cultures with Friend leukaemia virus complex (FLV) induced transformed cells with myelomonocytic characteristics, which were isolated only 14 days after the viral infection. Criteria for transformation were growth in suspension cultures and high plating efficiency in agar. The lymphatic leukaemia virus (LLV) replicates in these suspension cultures, but the spleen focus-forming virus (SFFV) component of the FLV complex has not been detected. Injection of the transformed cells into syngeneic neonatal or adult mice leads to the development of leukaemia which can be demonstrated to be of donor origin by the presence of two metacentric marker chromosomes which are also seen in the cultured cells.

Topics: Agar; Animals; Bone Marrow; Cell Division; Cell Line; Cell Transformation, Neoplastic; Cell Transformation, Viral; Colony-Forming Units Assay; Friend murine leukemia virus; Genetic Markers; Leukemia, Experimental; Leukemia, Myeloid; Mice; Neoplasm Transplantation; Time Factors; Transplantation, Isogeneic

Four children with chronic myeloproliferative disorders (three with Philadelphia [Ph1] chromosome-positive chronic myelogenous leukemia [CML] were studied with soft agar culture at diagnosis (before therapy) in an attempt to define abnormalities in granulopoiesis. The three patients with CML had elevated peripheral blood golony-forming cells (CFCs) and/or normal or decreased bone marrow CFCs (in those studied). Colony-stimulating activity (CSA) was markedly decreased or absent at diagnosis in all three. Maturation of myeloid cells eithin the colonies in agar was normal, indicating that no block in myeloid maturation was present. These findings are in general agreement with results previously reported in untreated adults with Ph1 chromosome-positive CML and further define the similarity with the adult form of the disease. One Ph1 chromosome-negative patient with a clinically similar chronic myeloproliferative disorder was studied and had similarly elevated peripheral blood CFCs. She had normal CSA with a similarly high WBC count. This finding was unexpected and suggests that, unlike the patients with CML, her monocytes were capable of elaboration CSA. This difference might prove helpful in the classification of this type of disorder in cases where the Ph1 chromosome abnormality is not present.

Topics: Adolescent; Agar; Bone Marrow; Cell Division; Child; Child, Preschool; Chromosomes, Human, 21-22 and Y; Chronic Disease; Clone Cells; Culture Media; Female; Granulocytes; Hematopoiesis; Humans; Leukemia, Myeloid; Leukocytes; Male; Myeloproliferative Disorders

Bone marrow cells from BALB/c mice with myeloid leukemia, lymphosarcoma, erythroblastosis, or mammary tumor produce small clusters in semisolid agar cultures in the absence of specific colony-stimulating factor. This spontaneous growth is observed only when high cell numbers (5 x 10 5 cells/ml) are plated. The phenomenon was encountered only when mice had an elevated number of mature or immature granulocytes in the peripheral blood. Removal of the adherent cells from the bone marrow did not abolish spontaneous growth, indicating that this colony-stimulating factor independency is not due to a high number of colony-stimulating cells in the bone marrow cells. This excluded the possibility that the spontaneous growth was due to a high endogenous stimulating activity of the bone marrow from tumor-bearing mice.

Topics: Agar; Animals; Bone Marrow; Bone Marrow Cells; Cell Division; Clone Cells; Granulocytes; Leukemia, Myeloid; Leukocyte Count; Lymphoma, Non-Hodgkin; Mammary Neoplasms, Experimental; Mice; Mice, Inbred BALB C; Neoplasms, Experimental; Rauscher Virus; Sarcoma, Experimental

Topics: Agar; Bone Marrow Cells; Cell Division; Cell Transformation, Neoplastic; Cells, Cultured; Clone Cells; Humans; In Vitro Techniques; Leukemia, Monocytic, Acute; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Leukocytes

Topics: Adult; Agar; Blood Platelets; Bone Marrow; Bone Marrow Cells; Cell Division; Cell Fractionation; Cell Separation; Clone Cells; Culture Media; Female; Hematopoiesis; Hemoglobins; Humans; Leukemia, Myeloid; Leukocyte Count; Leukocytes; Male; Middle Aged

Topics: Agar; Animals; Cell Division; Centrifugation, Density Gradient; Clone Cells; Cross Reactions; Hematopoietic Stem Cells; Humans; Immune Adherence Reaction; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Leukocytes; Monocytes; Rabbits

Topics: Agar; Cell Count; Cell Division; Centrifugation, Density Gradient; Clone Cells; Hematopoietic Stem Cells; Humans; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Lymphocyte Activation; Myeloproliferative Disorders; Thymidine; Tritium

Topics: Acid Phosphatase; Agar; Alkaline Phosphatase; Amylases; Animals; Cattle; Chickens; Chromatography, Gel; Deoxyribonucleases; Diffusion; Egg White; Gels; Intestines; Kinetics; Leukemia, Myeloid; Macaca; Male; Methods; Muramidase; Pancreas; Plants; Polysaccharides; Ribonucleases; Semen; Swine; Triticum

Topics: Agar; Electrophoresis; Erythrocytes; Freezing; Gels; Hemolysis; Humans; Isoenzymes; L-Lactate Dehydrogenase; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Leukocytes; Lymphocytes; Myeloproliferative Disorders; Primary Myelofibrosis; Staining and Labeling

Topics: Agar; Animals; Bone Marrow; Bone Marrow Cells; Cerebrospinal Fluid; Culture Techniques; Humans; Leukemia; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Mice; Retroviridae; Sepsis

Topics: Agar; Carcinoma; Electrophoresis; Gels; Half-Life; Hodgkin Disease; Humans; Isoenzymes; L-Lactate Dehydrogenase; Leukemia, Myeloid; Leukemia, Myeloid, Acute; Leukocytes; Lymphatic Diseases; Lymphoma; Neoplasms; Osteosclerosis; Plasmacytoma; Sarcoma; Tissue Extracts

An obligately anaerobic, gram-negative microorganism identified as a Sphaerophorus species was recovered from the fecal material of two cancer (chronic myelogenous leukemia and idiopathic thrombocythemia) patients receiving cobalt radiation therapy. The organism, isolated on sheep blood-agar, exhibited extreme pleomorphism (rods, filaments, and spheroids) and was a major component of the anaerobic fecal microflora. In one patient the numbers of Sphaerophorus species (designated as isolate 6-13-68), Bacteroides species, and Clostridium perfringens declined after irradiation; however, they were stable in this same patient after a second therapeutic dose of radiation. The numbers of anaerobes in the other patient remained fairly consistent after radiation. The biochemical and morphological characteristics and carbohydrate fermentation reactions of isolate 6-13-68 most closely resembled those of Sphaerophorus ridiculosis.

Topics: Agar; Animals; Bacteroides; Blood; Clostridium perfringens; Cobalt Isotopes; Culture Media; Feces; Fusobacterium; Humans; Intestine, Large; Leukemia, Myeloid; Sheep; Thrombocythemia, Essential

Topics: Adolescent; Adult; Agar; Anemia, Aplastic; Antibodies; Antibodies, Heterophile; Antigens; Blood Protein Electrophoresis; Blood Sedimentation; Chemical Precipitation; Female; Humans; Immune Sera; Immunoelectrophoresis; Leukemia, Myeloid; Leukocytes; Lupus Erythematosus, Systemic; Male; Serositis