concanavalin-a and Osteosarcoma

Confluent cultures of rat bone cells synthesize several forms of secreted phosphoprotein 1 (SPP-1, osteopontin), the major phosphorylated forms of which migrate at 55 and 44 kDa on 15% cross-linked SDS-PAGE gels and correspond to the transformation-associated proteins pp 69 and pp 62. A clonal rat calvarial cell line (RCA 11), which expressed the highest level of SPP-1, produced only the 55 kDa form of the phosphorylated protein, whereas normal rat calvarial cells enriched in osteoblastic cells (RC IV cells) produced mostly the 55 kDa form, with small amounts of the 44 kDa form. In contrast, a 44 kDa form was the major [32PO4]-labelled SPP-1 synthesized by a rat osteocarcoma cell line (ROS 17/2.8 cells) with lesser amounts of the 55 kDa SPP-1. When [35S]methionine was used to measure protein synthesis, only the 55 kDa SPP-1 could be clearly detected in confluent cultures of each cell population, indicating that the 55 kDa SPP-1 is the prominent form of SPP-1 synthesized by each cell population. Following treatment of the normal rat bone cells for 24 h with osteotropic hormones (vit D3, PTH and RA), growth factors (PDGF, EGF, TGF-beta), a tumor promoter (TPA) and a plant lectin (Con A), the 55 kDa [35S]methionine labelled SPP-1 was increased 1.7-8.3-fold. Similar, but generally lower responses were observed in the clonal RCA 11 cell line, whereas the ROS 17/2.8 cells were more refractory, showing only a strong response to vit D3. In general, vit D3 produced the strongest stimulation in all populations with TGF-beta producing a good response in the non-transformed cells and RA in the RC IV cells. In contrast, PTH was inhibitory in both RCA 11 and ROS 17/2.8 cells. In most, but not all, cases the alteration in SPP-1 synthesis reflected similar changes in SPP-1 mRNA and in the intensity of the [32PO4]-labelled 55 kDa SPP-1. Collectively, these studies demonstrate that bone cells produce several forms of SPP-1 which are differentially regulated in normal and transformed cells through both transcriptional and posttranscriptional mechanisms.

Topics: Animals; Calcitriol; Cell Line; Cell Line, Transformed; Cells, Cultured; Concanavalin A; Epidermal Growth Factor; Growth Substances; Nucleic Acid Hybridization; Osteopontin; Osteosarcoma; Parathyroid Hormone; Platelet-Derived Growth Factor; Precipitin Tests; RNA, Messenger; Sialoglycoproteins; Tetradecanoylphorbol Acetate; Transcription, Genetic; Transforming Growth Factor beta; Tretinoin

Carbohydrate moieties of cell surface glycoproteins with an external orientation play a role in hormone recognition and/or transmembrane signal transmission. We have examined the effect of various lectins, which interact with specific cell surface glycosyl residues, and of tunicamycin, an antibiotic that inhibits glycosylation of proteins, on the adenosine 3',5'-cyclic monophosphate (cAMP) response to parathyroid hormone (PTH) in confluent cultured osteoblast-like rat osteosarcoma cells (UMR-106) and opossum kidney cells (OK cells). Incubation of both cell lines with wheat germ lectin (WGL), but not with concanavalin A, succinylated wheat germ, ricin, or soybean lectins, markedly reduced the PTH-induced cAMP production, whereas the stimulation obtained with forskolin, a compound that acts directly on the adenylate cyclase enzyme, was not affected. In contrast, tunicamycin did not cause any decrease in the cAMP response to PTH. These results indicate that the masking of sialic acid residue by WGL considerably blunted PTH-stimulated cAMP production in cultured osteoblast-like and kidney cells. An 80% inhibition of glycosylation of cell surface proteins did not appear to affect the response to PTH. Thus the functional role of this carbohydrate moiety in the PTH receptor remains to be determined.

Topics: Acetylglucosamine; Animals; Colforsin; Concanavalin A; Cyclic AMP; Glycosylation; Kidney; Lectins; Opossums; Osteosarcoma; Parathyroid Hormone; Peptide Fragments; Plant Lectins; Rats; Ricin; Soybean Proteins; Tumor Cells, Cultured; Tunicamycin; Wheat Germ Agglutinins

Cultures of normal diploid fibroblasts and of a human osteosarcoma cell line (MG-63) are shown to be able to produce a factor which promotes the growth of B cell hybridomas (hybridoma growth factor, HGF). The induction is stimulated by treatment of the cells with interleukin 1 (IL 1) (alpha or beta) or polyriboinosinic-polyribocytidylic acid [poly(rI).poly(rC)]. Combined treatment with cycloheximide and actinomycin D also stimulates production and enhances production induced by IL 1 or poly(rI).poly(rC). Extremely small doses of IL 1 (0.1 units/ml) are active as inducer of HGF. Also, under optimal conditions the yield of HGF can attain as much as 10(4) units/ml. Tumor necrosis factor (TNF-alpha), which otherwise shares various properties with IL 1, is a weak inducer of HGF. Although there is a superficial resemblance between induction of HGF and that of interferon-beta, the two activities are serologically distinct and conditions for their induction are quite different. In fact, conditions for induction of HGF are indistinguishable from those described for the induction of the mRNA of the so-called 26-kDa protein (also known as interferon-beta 2). Finally, the HGF derived from IL 1- or poly(rI).poly(rC)-treated fibroblasts is serologically not distinguishable from that produced by mitogen-stimulated peripheral blood leukocytes.

Topics: Animals; B-Lymphocytes; Biological Products; Cell Line; Cells, Cultured; Concanavalin A; Cricetinae; Cricetulus; Cycloheximide; Cytokines; Dactinomycin; Female; Fibroblasts; Growth Substances; Humans; Hybridomas; Interferon Type I; Interleukin-1; Kinetics; L Cells; Leukocytes; Mice; Osteosarcoma; Ovary; Poly I-C; Skin

Type V collagen from FBJ virus-induced osteosarcoma of mice has a high content of saccharide as has been noted for type V collagens from different sources. In the present study, this collagen was found to contain significant amounts of mannose and hexosamine. Three alpha chains of this collagen were electrophoretically separated and cleaved by cyanogen bromide. The cyanogen bromide peptides, following their separation by SDS-polyacrylamide gel electrophoresis, were transferred onto nitrocellulose paper and stained with concanavalin A. Several bands derived only from alpha 3(V) stained positively, but this was inhibited by the presence of alpha-methylmannoside. Thus, at least one of these three chains appears to have an asparagine-linked oligosaccharide.

Topics: Animals; Asparagine; Collagen; Concanavalin A; Cyanogen Bromide; Electrophoresis, Polyacrylamide Gel; Hexosamines; Mannose; Osteosarcoma; Peptide Fragments; Rats

The concanavalin A-induced agglutination of cultivated cells of human osteosarcoma Sa-4 was studied in the process of their reversion caused by high polar compounds, dimethylsulphoxide (DMSO) and dimethylformamide (DMFA). Primarily lectin induced an expressed ability to agglutination in Sa-4 cells. In the process of tumour cell reversion under the effect of chemical inducers their agglutination decreases. After the DMSO and DMFA removal the osteosarcoma cells return to their initial state showing a high agglutination. Other high-polar compounds (N-methylformamide and dimethylacetamide) induced no reversion of Sa-4 cells and no agglutination of them as well.

Topics: Acetamides; Agglutination; Concanavalin A; Culture Media; Dimethyl Sulfoxide; Dimethylformamide; Drug Interactions; Formamides; Humans; In Vitro Techniques; Osteosarcoma

The lectin (PHA and Con A) induced agglutination of a human tumour cell line (lung adenocarcinoma and osteosarcoma) was estimated by the spectrophotometric method. A decrease in the optic density for 1 min (delta D) of cell suspension and the time (t0) necessary for a complete sedimentation of cells (at delta D = 0) were used as quantitative indicators of agglutination. An increase in the concentration of tumour cells and lectins resulted in an increase of delta D and a decrease of t0. The results of spectrophotometry were correlated with the microscopic study data for tumour cells agglutination.

Topics: Adenocarcinoma; Agglutination; Cell Aggregation; Cell Count; Cells, Cultured; Concanavalin A; Humans; Lectins; Lung Neoplasms; Neoplasms; Osteosarcoma; Phytohemagglutinins; Spectrophotometry

The monoclonal antibody 791T /36, prepared against a human osteogenic sarcoma cell line, 791T , reacts with a variety of human tumours and also mitogen-stimulated PBMN cells. The target antigen as expressed upon 791T cells is a monomeric plasma membrane-associated glycoprotein with an apparent Mr of 72000. By quantitative flow cytofluorimetry, approx. 10(5) antibody molecules bound per cell to T-lymphoblasts induced with PHA or Con A, whereas only a few thousand antibody molecules bound per cell to unstimulated cells, so that the antigen may be classified as a lymphocyte activation antigen. On lymphoblasts, the 791T /36 again reacted with a protein with an apparent Mr of 72000. This antigen therefore has a dual role as a tumour marker and lymphocyte activation antigen which may be implicated in the regulation of cell proliferation.

Topics: Antibodies, Monoclonal; Antigen-Antibody Complex; Antigens, Neoplasm; Antigens, Surface; Cell Line; Cells, Cultured; Concanavalin A; Flow Cytometry; Humans; Lymphocyte Activation; Neoplasms; Osteosarcoma; T-Lymphocytes

The effects of isoprinosine (ISO) on the immune responses (Con A-induced lymphocyte proliferation, monocyte chemotactic responsiveness, and "natural killer" cytotoxicity) of normal hamsters and hamsters with human osteosarcoma (OS) were investigated. Human osteosarcoma was induced in newborn inbred hamsters (LHX/SsLAK) after induction of tolerance in utero. In vitro, ISO increased Con A-induced proliferation of peripheral blood lymphocytes (PBL) from normal hamsters by 23.4-48.9% and from OS-bearing hamsters by 58.1-107.4% over controls (Con A alone). When ISO was administered in vivo by intraperitoneal injection. Con A-induced proliferation of PBL from both normal and OS-bearing recipients in vitro was increased by 50-55% at 1, 3 and 5 days after injection. The chemotactic responsiveness of monocytes from OS-bearing hamsters was also significantly increased (59.1-97.4%) at 1, 3 and 5 days after injection of ISO. Natural killer cytotoxicity was augmented at 1, 3 and 5 days after injection of ISO by 31.7-83.6% in normal hamsters and 54.6-184% in OS-bearing hamsters. These results indicate that ISO can produce a generalized enhancement of immune function in hamsters with OS.

Topics: Adjuvants, Immunologic; Animals; Chemotaxis, Leukocyte; Concanavalin A; Cricetinae; Cytotoxicity, Immunologic; Disease Models, Animal; Female; Humans; In Vitro Techniques; Inosine; Inosine Pranobex; Lymphocyte Activation; Osteosarcoma; Pregnancy; Sarcoma, Experimental

Peripheral lymphocytes from patients with osteosarcoma receiving interferon (IF) as adjuvant therapy were tested in vitro for response to various mitogens. Prolonged parenteral administration of IF preparations caused no major change of the mitogen responses. The ability of IF, when added in vitro, to inhibit lymphocyte response to mitogens was not altered to any major extent by in vivo administration of IF.

Topics: Adult; Aged; Cells, Cultured; Concanavalin A; Female; Humans; Interferons; Leukocyte Count; Leukocytes; Lymphocyte Activation; Male; Middle Aged; Mitogens; Osteosarcoma; Phytohemagglutinins; Pokeweed Mitogens; Time Factors

Topics: Adenoviridae; Animals; Antibody Formation; Bromodeoxyuridine; Cells, Cultured; Concanavalin A; Culture Media; Female; Fibrosarcoma; Guinea Pigs; Immunodiffusion; Inclusion Bodies, Viral; Injections, Subcutaneous; Liposarcoma; Methylcholanthrene; Osteosarcoma; Sarcoma, Experimental; Thymidine; Tritium

Blocking factors are small polypeptide molecules that may appear in the serum of patients with cancer. These factors block the transformation of lymphocytes in culture to nonspecific mitogens such as phytohemagglutinin or concanavalin A and, therefore, may reflect changes in the immunocompetence of the patient. Blocking factors were monitored during the clinical course of thirty-five patients with cancer. These factors did not develop in patients with response to therapy whereas they did develop in patients without response. A third group of patients without response to therapy after a previous remission showed an absence of lymphocyte responsiveness in culture that was not due to blocking factors, suggesting that immune clone consumption had occurred. Dermal responsiveness to tumor antigen correlated with a favorable clinical course and was usually absent when serum blocking factors were present.

Topics: Antibody Formation; Antigens, Neoplasm; Carcinoma, Squamous Cell; Concanavalin A; Humans; Immunotherapy; Lectins; Leiomyosarcoma; Lymphocyte Activation; Lymphocytes; Melanoma; Neoplasms; Osteosarcoma; Peptides; Pharyngeal Neoplasms; Rhabdomyosarcoma; Skin Tests

Topics: Adsorption; Agglutination; Binding Sites; Cell Fractionation; Cell Membrane; Cell Transformation, Neoplastic; Centrifugation, Density Gradient; Concanavalin A; Methods; Neoplasms, Experimental; Osteosarcoma; Peritoneal Neoplasms; Polyomavirus