methylcellulose and Neuroblastoma

Cell hybrids (BIM) were produced between human neuroblastoma cells (IMR-32) and thymidine auxotrophs (B3T) of rat nerve-like cells (B103) in order to obtain cell lines undergoing stable neuronal differentiation. BIM cells exhibited the growth properties of partial transformation: 1) When the cell growth reached a plateau, BIM cells ceased to proliferate and expressed a differentiated phenotype. The shape of the cells changed from flat to round and they extended neurites. 2) When cultured in methylcellulose, BIM cells formed colonies, indicating that BIM cells have the ability of anchorage-independent growth. By Southern blot analysis, BIM cells had both human and rat types of N-myc genes. The human N-myc genes were amplified, but the extent of the amplification was lower in BIM cells than that in the parental cell line IMR-32. The rat N-myc gene was detected at a similar level in BIM, B3T, B103, and rat fibroblastic cells 3Y1. Therefore, the decrease in amplification of human N-myc genes may be involved in the properties of partial reverse-transformation in BIM cells. When treated with various drugs such as db-cAMP, forskolin, and cAMP with isobutyl-methylxanthine, BIM cells expressed a nerve-like phenotype. These findings indicate that cell hybridization yielded partial normalization of transformed nerve-like cells.

Topics: 1-Methyl-3-isobutylxanthine; Animals; Axons; Bucladesine; Calcimycin; Cell Cycle; Cell Transformation, Neoplastic; Colforsin; Cyclic AMP; Fibroblasts; Humans; Hybrid Cells; Methylcellulose; Neuroblastoma; Neurons; Phenotype; Rats; Thymidine; Tretinoin; Tumor Cells, Cultured

The effects of recombinant products of granulocyte colony-stimulating factors (G-CSF), granulocyte/macrophage CSF (GM-CSF), human interleukin-3 (IL-3), and interleukin-1 (IL-1) were studied using purified target cell populations from patients undergoing peripheral blood stem cell transplantation after myeloablative therapy. Cells were subjected to combined purification procedures including negative selection with a panel of monoclonal antibodies (CD2, 3, 5, 10, and 20). The purified cells were enriched for HLA-DR+ (51% to 71%) and My-10+ (CD34; 37% to 54%) and had a plating efficiency of up to 20%. In the liquid-suspension limiting dilution clonal assay (LDA), purified progenitors responded directly to IL-3 by proliferation with single-hit kinetics. The ability of GM-CSF to support progenitor growth was inferior to that of IL-3, and the cells were virtually unresponsive when cultured with G-CSF, supporting the notion that these blood-derived progenitors belong to a primitive population of hematopoietic progenitor cells. The results obtained in simultaneous methycellulose cultures (MC) showed the same trend and provided additional information on the ability of GM-CSF and IL-3 to support erythroid progenitor growth. The combination of IL-3 plus G-CSF, but not IL-3 plus GM-CSF, resulted in a synergistic increase in colony number. IL-1 alpha increased both the size and number of colonies when added to IL-3 or G-CSF. Study of this enriched progenitor cell population in MC and LDA represents an excellent model for the investigation of myeloid and erythroid differentiation and for evaluating the influence of various cytokines on human hematopoiesis.

Topics: Antigens, CD; Antigens, CD34; Antigens, Differentiation; Cell Division; Cell Separation; Cells, Cultured; Child; Colony-Stimulating Factors; Granulocyte Colony-Stimulating Factor; Granulocyte-Macrophage Colony-Stimulating Factor; Growth Substances; Hematopoietic Stem Cells; HLA-DR Antigens; Humans; Interleukin-1; Interleukin-3; Kinetics; Leukemia; Lymphoma, Non-Hodgkin; Methylcellulose; Neuroblastoma; Recombinant Proteins

An in vitro methylcellulose technique was used in an attempt to culture neuroblastoma cells from 25 bone marrows from eight children with neuroblastoma. Colonies appeared within 5 days in histologically positive bone marrows. Light microscopy, linearity study, and marker study provided evidence for the neuroblastoma origin of the colonies. These colonies could be distinguished from other colonies under the inverted microscope because of its distinct feature. In one case, the characteristic morphology of neuroblastoma was shown in 3 days of culture, while histological evidence is absent. The diagnosis of neuroblastoma was confirmed by subsequent catecholamine determination. All histologically negative specimens formed no colonies, while all positive specimens formed more than three colonies. Potential application of this culturing technique for monitoring of bone marrow involvement and differential diagnosis in children with neuroblastoma is presented.

Topics: Bone Marrow; Child; Clone Cells; Culture Media; Culture Techniques; Diagnosis, Differential; Hematopoietic Stem Cells; Humans; Methylcellulose; Neoplasm Proteins; Neoplastic Stem Cells; Neuroblastoma; Phosphopyruvate Hydratase; Prognosis

Therapeutic efficacy and toxicity were evaluated in 28 children with acute lymphoblastic leukemia, in ten with acute nonlymphoblastic leukemia (ANLL), and in 13 with metastatic neuroblastoma. All were refractory to standard chemotherapeutic agents and 25 were refractory to an investigational drug. The initial dose was 12 mg/m2/day and was based on an established maximal dose tolerated in adults. This dose was found to be intolerable in 5 of 5 children with leukemia. Similarly an initial dose of 9 mg/m2/day was intolerable in 4 of 5 patients with leukemia. The starting dose in the next 28 children with leukemia or neuroblastoma was 3 mg/m2. This drug was gradually increased to the highest tolerated dose by 3-mg/m2 increments. Fifteen children with acute lymphoblastic leukemia, 3 children with ANLL, and 2 children with neuroblastoma received the drug daily. Seven patients with ANLL and 7 patients with neuroblastoma received the drug biweekly. Seventeen patients with acute lymphoblastic leukemia, 6 patients with ANLL, and 5 patients with neuroblastoma had an adequate trial of the drug. An adequate trial was defined as a minimum of 5 weeks of therapy unless progressive disease developed. Side effects of the drug were striking and included fever, hypotension, myalgia, bone pain, arthralgia, arthritis, abdominal pain, liver toxicity, thrombocytopenia, and neurotoxicity. No complete remission occurred although interferon levels above 100 units/ml were induced in nearly 50% of the patients.

Topics: Acute Disease; Adolescent; Carboxymethylcellulose Sodium; Child; Child, Preschool; Drug Evaluation; Humans; Interferon Inducers; Interferons; Leukemia; Methylcellulose; Neuroblastoma; Poly I-C; Polylysine

A Phase II study of poly(I,C)-LC was performed in 28 children and adolescents with acute lymphoblastic leukemia (ALL), 10 with acute nonlymphoblastic leukemia (ANLL), and 13 with metastatic neuroblastoma. All were refractory to standard chemotherapeutic agents and 25 to an investigational drug. Initial doses of 12 mg/m2 and 9 mg/m2 were intolerable. However, 9 mg/m2 was tolerable in the majority of patients when the drug was started at 3 mg/m2 and increased by 3 mg/m2 increments. Fifteen children with ALL, three with ANLL, and two with neuroblastoma received the drug daily. Seven patients with ANLL and seven children with neuroblastoma received the drug biweekly. Twenty-eight patients received an adequate trial, which was defined as a minimum of 5 weeks at the maximal tolerated dose, unless there was progressive disease at the maximal tolerated dose. Side effects of the drug were striking, and included fever, hypotension, myalgia, bone pain, arthralgia, arthritis, abdominal pain, liver toxicity, thrombocytopenia, and neurotoxicity. No complete remissions occurred in spite of interferon levels above 100 U in nearly 50% of patients.

Topics: Acute Disease; Carboxymethylcellulose Sodium; Child; Drug Evaluation; Female; Humans; Interferon Inducers; Interferons; Leukemia; Leukemia, Lymphoid; Male; Methylcellulose; Neuroblastoma; Poly I-C; Polylysine

The addition of 1% (w/v) carboxymethyl cellulose to the culture medium induces the formation of neurites of clone N18 neuroblastoma cells even in the presence of normal (5-10%) serum supplement concentrations, which rivals that previously observed by growth in low 0.1% serum. Heavy metal ions associated with the carboxymethyl cellulose were responsible for small increases in the sizes of cell bodies during treatment. Pretreatment of the PC12 pheochromocytoma line of neuroblasts with carboxymethyl cellulose for 1 day prior to their stimulation with nerve growth factor resulted in an acceleration in the rate, but not extent, of neurite outgrowth.

Topics: Animals; Carboxymethylcellulose Sodium; Cell Differentiation; Cell Line; Copper; Drug Interactions; Ferrous Compounds; Methylcellulose; Mice; Neuroblastoma; Neurons; Pheochromocytoma; Zinc