acid-phosphatase and Cell-Transformation--Viral

Topics: Acid Phosphatase; Alkaline Phosphatase; Animals; Cell Transformation, Viral; Oncogenes; Phosphoprotein Phosphatases; Phosphorylation; Phosphoserine; Phosphothreonine; Phosphotyrosine; Retroviridae; Tyrosine

Autosomal-Recessive Osteopetrosis (ARO) comprises a heterogeneous group of bone diseases for which mutations in five genes are known as causative. Most ARO are classified as osteoclast-rich, but recently a subset of osteoclast-poor ARO has been recognized as due to a defect in TNFSF11 (also called RANKL or TRANCE, coding for the RANKL protein), a master gene driving osteoclast differentiation along the RANKL-RANK axis. RANKL and RANK (coded for by the TNFRSF11A gene) also play a role in the immune system, which raises the possibility that defects in this pathway might cause osteopetrosis with immunodeficiency. From a large series of ARO patients we selected a Turkish consanguineous family with two siblings affected by ARO and hypogammaglobulinemia with no defects in known osteopetrosis genes. Sequencing of genes involved in the RANKL downstream pathway identified a homozygous mutation in the TNFRSF11A gene in both siblings. Their monocytes failed to differentiate in vitro into osteoclasts upon exposure to M-CSF and RANKL, in keeping with an osteoclast-intrinsic defect. Immunological analysis showed that their hypogammaglobulinemia was associated with impairment in immunoglobulin-secreting B cells. Investigation of other patients revealed a defect in both TNFRSF11A alleles in six additional, unrelated families. Our results indicate that TNFRSF11A mutations can cause a clinical condition in which severe ARO is associated with an immunoglobulin-production defect.

Topics: Acid Phosphatase; Actins; Agammaglobulinemia; Amino Acid Sequence; Amino Acid Substitution; Argentina; Arginine; Biopsy; Case-Control Studies; Cell Line, Transformed; Cell Proliferation; Cell Transformation, Viral; Cells, Cultured; Cohort Studies; Consanguinity; Cysteine; Dendrites; DNA Mutational Analysis; Female; Genes, Recessive; Herpesvirus 4, Human; Heterozygote; Homozygote; Humans; Ilium; Isoenzymes; Leukocyte Common Antigens; Leukocytes, Mononuclear; Lipopolysaccharides; Macrophage Colony-Stimulating Factor; Male; Models, Immunological; Molecular Sequence Data; Mutation, Missense; Osteoclasts; Osteopetrosis; Osteoprotegerin; Pakistan; Pedigree; Polymorphism, Genetic; Protein Structure, Tertiary; Radiography, Thoracic; RANK Ligand; Receptor Activator of Nuclear Factor-kappa B; Receptors, Vitronectin; Sequence Homology, Amino Acid; Tartrate-Resistant Acid Phosphatase; Turkey

The secreted and intracellular activities of a number of lysosomal hydrolases were higher in 3T3 cells than in SV40-transformed cells. The number of lysosomes and their total volume were also much larger in 3T3 cells and the surface area of their lysosomal membranes was almost twice that of SV3T3 cells. These differences alone were not sufficiently large, however, to account for the disparity seen in activity of some enzymes. Gel electrophoresis showed that a number of protein components present in lysosomal membranes purified from 3T3 cells were absent from SV3T3 membrane preparations. The absence of these components may be correlated with the reduced enzyme activity of SV3T3 cells particularly with respect to beta-glucosidase and acid phosphatase, both of which are normally found associated with lysosomal membranes.

Topics: Acid Phosphatase; Acridine Orange; Animals; beta-Glucosidase; beta-N-Acetylhexosaminidases; Cell Fractionation; Cell Line; Cell Line, Transformed; Cell Transformation, Viral; Cytophotometry; Hydrolases; Intracellular Membranes; Lysosomes; Povidone; Simian virus 40; Staining and Labeling

The hybrid plasmid pK4 containing the early genes of the simian virus SV-40, under the control of the adenovirus type 5 E1a promoter, was introduced into the multipotent embryonal carcinoma (EC) 1003. Expression of the SV-40 oncogenes was observed at the EC cell stage, and this allowed the derivation of immortalized cells corresponding to early stages of differentiation. Among the immortalized mesodermal derivatives obtained, one clone, C1, is committed to the osteogenic pathway. C1 cells have a stable phenotype, synthesize type I collagen, and express alkaline phosphatase activity. Although immortalized and expressing the SV-40 T antigen, the cells continue to be able to differentiate in vivo and in vitro. In vivo, after injection into syngeneic mice, they produce osteosarcomas. In vitro, the cells form nodules and deposit a collagenous matrix that mineralizes, going to hydroxyapatite crystal formation, in the presence of beta-glycerophosphate. This clonal cell line, which originates from an embryonal carcinoma, therefore differentiates into osteogenic cells in vivo and in vitro. This immortalized cell line will be useful in identifying specific molecular markers of the osteogenic pathway, to investigate gene regulation during osteogenesis and to study the ontogeny of osteoblasts.

Topics: Acid Phosphatase; Adenovirus Early Proteins; Alkaline Phosphatase; Animals; Blotting, Southern; Calcification, Physiologic; Cell Differentiation; Cell Line; Cell Transformation, Viral; Clone Cells; Collagen; Cyclic AMP; DNA-Binding Proteins; Electron Probe Microanalysis; Fluorescent Antibody Technique; Genes, Viral; Mice; Microscopy, Electron; Nucleic Acid Hybridization; Oncogene Proteins, Viral; Osteocytes; Plasmids; Promoter Regions, Genetic; Simian virus 40; Teratoma; Viral Structural Proteins; X-Ray Diffraction

Three cell lines from resident macrophages of BALB/c mice and four from activated macrophages of the same strain were isolated by infection with simian virus 40 (SV40). A majority of these cells showed dependency on L cell-conditioned medium (LCM), which is necessary for proliferation of normal macrophages in vitro. Somatic cell hybridization was applied in the study of macrophage growth responsiveness. A macrophage cell line (BR15) with strict dependency on LCM for growth was fused to a Chinese hamster cell line (hs222-16); it was found that dependency on LCM was a dominant trait in the hybrids. Following fusion of a macrophage cell line (BAM3) which grew without LCM to hs222-16, a large number of colonies appeared in the selection medium containing LCM. Four hybrids not requiring LCM for growth were selected in an LCM-free culture, and their hybrid properties were examined. Three out of the four hybrids secreted colony-stimulating factor (CSF) constitutively, whereas the fourth secreted no CSF. The level of acid phosphatase activity in the hybrids was higher than in the parent cells. Two peaks of CSF activity were observed after gel filtration chromatography of conditioned medium: One was eluted at molecular weight of 36,000 and the other at 17,000.

Topics: Acid Phosphatase; Animals; Cell Division; Cell Line; Cell Transformation, Viral; Colony-Stimulating Factors; Cricetinae; Hybrid Cells; Macrophages; Mice; Microscopy, Electron; Simian virus 40

In order to assess the role of Epstein-Barr virus (EBV) in patients with hairy cell leukemia (HCL), we have sought to characterize 1) the ability of EBV to infect and transform hairy leukemic cells in vitro and 2) the phenotypes of cell lines putatively derived from those leukemic cells. Analysis of EBV-induced transformation and the kinetics of Epstein-Barr nuclear antigen (EBNA) induction in leukemic preparations indicated that most leukemic cells were not susceptible to EBV infection but that at least a small subpopulation of leukemic cells could be infected with EBV. Lymphoblastoid cells lines were established after exposure of peripheral blood or splenic cells from HCL patients to B95-8 or QIMR-WIL EBV. Splenic leukemic cell preparations were more sensitive targets for EBV transformation than were peripheral blood cell samples. The newly established cell lines, but not long-established B lines such as Raji, demonstrated high levels of synthesis of p35, (a protein complex expressed abundantly by cells of a subset of HCL patients) and high levels of tartrate-resistant acid phosphatase (an enzyme relatively diagnostic for HCL). Lymphoblastoid lines from one patient with HCL expressed lambda light chains and no kappa chains as did the patient's leukemic cells. Virus expression in these lines showed that HCL-derived lines had spontaneous early antigen (EA) and viral capsid antigen (VCA) expression. Transforming EBV could be rescued from HCL-derived cell lines but not from cord blood-derived lines.

Topics: Acid Phosphatase; Antigens, Neoplasm; Antigens, Surface; Antigens, Viral; Capsid; Cell Line; Cell Transformation, Viral; Electrophoresis, Polyacrylamide Gel; Herpesvirus 4, Human; Humans; Immunoglobulins; Leukemia, Hairy Cell; Neoplasm Proteins; Phenotype

Myocrisin given to mice i.p. causes depression of lysosomal enzyme activity (acid phosphatase, beta-glucuronidase and N-acetyl-beta-D-glucosaminidase) in peritoneal macrophages. If avirulent Semliki Forest virus (SFV) is given i.p. 3 h after the Myocrisin, further depression of lysosomal enzyme activity occurs, a very high titre of virus is produced in these macrophages and the virus becomes lethal, causing 100% mortality. The possible interrelationships between depressed lysosomal activity, high virus titres and the production of a lethal virus infection are discussed.

Topics: Acetylglucosaminidase; Acid Phosphatase; Animals; Ascitic Fluid; Cell Transformation, Viral; Glucuronidase; Gold Sodium Thiomalate; Lysosomes; Macrophages; Mice; Semliki forest virus; Togaviridae Infections; Virulence; Virus Replication

Topics: 4-Nitrophenylphosphatase; Acetylglucosaminidase; Acid Phosphatase; Cell Fractionation; Cell Transformation, Viral; Centrifugation; Electrophoresis; Herpesvirus 4, Human; Humans; Lymphocytes; Lysosomes; Microscopy, Electron

Functionally differentiated chicken macrophages were derived by in vitro differentiation of embryonic yolk sac cells and were characterized by several macrophage-specific cell markers. Uniform, infected, virus-producing cultures were obtained after exposure of these macrophages to avian myoblastosis virus (AMV), avian myelocytomatosis virus (MC29), myeloblastosis-associated virus (MAV-2), and Prague strain of Rous sarcoma virus (PR-B RSV). Both AMV and MC29 induced morphological transformation typical of the in vivo leukemias induced by these virus strains. Analysis of the expression of macrophage-specific markers in these two transformed cell types demonstrated that different markers of the mature macrophage were suppressed by each virus, even though the parental cell immediately preceding the transformation event was a mature macrophage in both cases. Cells infected with PR-B RSV and MAV-2 showed no observable difference from uninfected macrophages in terms of morphological characteristics, growth rate, or expression of the differentiated functions of macrophages. Ths system provides demonstrations of a cell type that produces infectious, transforming RSV but fails to respond by functional alterations induced by the transforming gene, src.

Topics: Acid Phosphatase; Alpharetrovirus; Animals; Cell Adhesion; Cell Transformation, Viral; Chick Embryo; Chickens; Macrophages; Phagocytosis

Topics: 1-Acylglycerophosphocholine O-Acyltransferase; Acid Phosphatase; Animals; Cell Line; Cell Transformation, Viral; Cricetinae; Dengue Virus; Diacylglycerol Cholinephosphotransferase; Enzyme Activation; Glucuronidase; Kidney; Kinetics; Lysophospholipase; Microsomes; Phospholipases; Phospholipases A; Phospholipases A2; Phosphotransferases; Subcellular Fractions

Differences between adult and fetal human fibroblasts have been found in the enzyme patterns of adenosine deaminase, acid phosphatase and lactate dehydrogenase. In each case the pattern seen in adult fibroblast cultures transformed with SV 40 virus resembled that of fetal fibroblasts.

Topics: Acid Phosphatase; Adenosine Deaminase; Adult; Alkaline Phosphatase; Cell Transformation, Viral; Female; Fetus; Fibroblasts; Humans; Infant; Isoenzymes; L-Lactate Dehydrogenase; Pregnancy; Simian virus 40

It is shown that human embryonic cells transformed by Rous sarcoma virus (stable cell line 23) and those transformed by polyoma virus (stable cell line P-2) are morphologically distinguished from the normal human embryonic cells. The mitotic activity of P-2 cells was 51% and the mitotic activity of 23 cells was 48%. While the mitosis activity of human embryo fibroblast was 28%. The duration of the mitosis of P-2 cells was 20 hours and that of 23 cells was 18 hr. The duration of the mitotic cycle of human embryo fibroblast was 18 hr. The G1 periods lasted 6 hours for both the cell lines; the S period of P-2 cells lasted 8 hr and the S period of 23 cells was 6 hr. Both the cell lines had a high content of RNA, DNA, protein bound SH-groups, and a high activity of acid phosphatase, acid RNAase and glucose-6-phosphatase. The content of glycogen, and acidic mucopolysaccharides, the activity of NADPH-tetrazolium reductase, succinic dehydrogenase of both the lines were the same as in normal human cells.

Topics: Acid Phosphatase; Avian Sarcoma Viruses; Cell Cycle; Cell Line; Cell Transformation, Neoplastic; Cell Transformation, Viral; DNA, Neoplasm; Glycogen; Humans; Polyomavirus; RNA, Neoplasm