Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound   Target Concepts: Gene/Protein Disease Symptom Drug Enzyme Compound

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Query: EC:3.1.3.1 (alkaline phosphatase)
47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

We present a method to detect and enumerate individual interferon (IFN)-producing human lymphocytes. The assay is based on the ELISA-plaque assay developed by Sedgwick and Holt (J. Exp. Med. (1983) 157, 2178; J. Immunol. Methods (1986) 87, 37). Mitogen-stimulated T cells are seeded in anti-IFN-gamma-coated wells. After a 16 h incubation period, the cells are removed. Subsequently a rabbit anti-IFN-gamma-antiserum followed by goat anti-rabbit antiserum conjugated to alkaline phosphatase are used to detect the IFN-gamma spots. Application of the spot-ELISA in combination with the conventional ELISA reveals the amount of IFN-gamma produced per cell. The spot-ELISA is a highly sensitive, easy to perform and rapid assay. Provided specific antisera are available, this method is suitable to detect production of other lymphokines at the single-cell level. To our knowledge, this is the first report of a single, well-defined T cell product measurement by the spot-ELISA.
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PMID:Enumeration of IFN-gamma-producing human lymphocytes by spot-ELISA. A method to detect lymphokine-producing lymphocytes at the single-cell level. 313 88

Rat placental cells (RPCs) derived from the chorioallantoic placenta of day-12 Holtzman rats were tested for the expression of class I and class II RT I histocompatibility antigens, transferrin receptors, intermediate filaments, and alkaline phosphatase. The binding of mouse monoclonal antibodies to those antigens by RPCs was compared with the binding of the same reagents to rat placental cells in situ. RPCs expressed low levels of class I antigens and failed to express detectable levels of class II antigens. RPCs resisted up-regulation of expression of class I antigens by interferon-gamma, and did not express class II antigens following exposure to medium containing interferon. Transferrin receptors; cytokeratin intermediate filaments, and alkaline phosphatase were universally expressed by RPCs. Taken together with the patterns of expression of the same antigens by rat placental cells in situ, the results suggest that RPCs comprise labyrinthine trophoblast cells. Those cells may provide a valuable new approach for studying the structures and functions of trophoblast cells in vitro.
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PMID:Expression of histocompatibility antigens, transferrin receptors, intermediate filaments, and alkaline phosphatase by in vitro cultured rat placental cells and rat placental cells in situ. 313 46

Using cultured human osteoblast-like cells, we studied the effects of tumor necrosis factor (TNF) and recombinant human gamma-interferon (gamma-IFN) on osteoblast growth and function, and demonstrated that TNF stimulated bone cell proliferation and prostaglandin production while inhibiting 1,25-(OH)2D3-stimulated alkaline phosphatase activity and osteocalcin release. In contrast, gamma-IFN inhibited proliferation and stimulated alkaline phosphatase activity of the cells, while inhibiting 1,25-(OH)2D3-stimulated osteocalcin production and having variable effects on the release of prostaglandins, depending on the presence of other factors. Our results suggest that TNF and gamma-IFN can act directly on bone-forming cells to affect both their proliferation and their differentiated function, and that changes in the ability of cells to produce these factors in disease states may contribute to alterations in the integrity of connective tissue matrices.
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PMID:Actions of recombinant human gamma-interferon and tumor necrosis factor alpha on the proliferation and osteoblastic characteristics of human trabecular bone cells in vitro. 314 69

The 2- and 8-azido trimer 5'-triphosphate photoprobes of 2-5A have been enzymatically synthesized from [gamma-32P]2-azidoATP and [alpha-32P]8-azidoATP by 2-5A synthetase from rabbit reticulocyte lysates. Identification and structural determination of the 2- and 8-azido adenylate trimer 5'-triphosphates were accomplished by enzymatic hydrolyses with T2 RNase, snake venom phosphodiesterase, and bacterial alkaline phosphatase. Hydrolysis products were identified by HPLC and PEI-cellulose TLC analyses. The 8-azido photoprobe of 2-5A displaces p3A4[32P]pCp from RNase L with affinity equivalent to p3A3 (IC50 = 2 X 10(-9) M in radiobinding assays). The 8-azido photoprobe also activates RNase L to hydrolyze poly(U) [32P]pCp 50% at 7 X 10(-9) M in core-cellulose assays. The 2- and 8-azido photoprobes and authentic p3A3 activate RNase L to cleave 28S and 18S rRNA to specific cleavage products at 10(-9) M in rRNA cleavage assays. The nucleotide binding site(s) of RNase L and/or other 2-5A binding proteins in extracts of interferon-treated L929 cells were investigated by photoaffinity labeling. Dramatically different photolabeling patterns were observed with the 2- and 8-azido photoprobes. The [gamma-32P]2-azido adenylate trimer 5'-triphosphate photolabels only one polypeptide with a molecular weight of 185,000 as determined by SDS gel electrophoresis, whereas the [alpha-32P]8-azido adenylate trimer 5'-triphosphate covalently photolabels six polypeptides with molecular weights of 46,000, 63,000, 80,000, 89,000, 109,000, and 158,000. Evidence that the photolabeling by 2- and 8-azido 2-5A photoprobes was highly specific for the p3A3 allosteric binding site was obtained as follows.(ABSTRACT TRUNCATED AT 250 WORDS)
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PMID:2- and 8-azido photoaffinity probes. 1. Enzymatic synthesis, characterization, and biological properties of 2- and 8-azido photoprobes of 2-5A and photolabeling of 2-5A binding proteins. 324 13

Cloned MC3T3-E1 cells which have retained several osteoblast-like characteristics were derived from newborn mouse calvaria. In order to elucidate the function of osteoblasts, the effects of 1,25(OH)2D3, interleukin (IL)-1 beta, IL-3, interferon(INF)-gamma and epidermal growth factor(EGF) on the activity of alkaline phosphatase(Al-P'ase), DNA synthesis and the production of prostaglandin E2(PGE2) in MC3T3-E1 cells were studied. The influence of cyclosporin A(CSA), a potent immunosuppressive agent, was also studied. The following results were obtained: 1. 1,25(OH)2D3 increased the incorporation of [45Ca]Cl2 into matrix and accelerated the calcification of MC3T3-E1 cells. 2. Al-P'ase activity and the incorporation of [3H]-thymidine into MC3T3-E1 cells were increased by 1,25(OH)2D3 but decreased by IL-1 beta, INF-gamma, IL-3 and EGF. 3. IL-1 beta increased and INF-gamma decreased PG-E2 production by MC3T3-E1 cells. 4. CSA decreased either Al-P'ase activity or incorporation of [3H]-thymidine, and increased PG-E2 production in MC3T3-E1 cells. CSA which was simultaneously incubated with these various cell growth factors, showed a similar effect to that of CSA alone. These results suggest that cytokines produced from immune cells, could affect osteoblasts besides that of calcium regulating hormones like parathyroid hormone and 1,25(OH)2D3, implying a probability for the participation of immunocompetent cells in the regulation of bone metabolism.
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PMID:[The effects of various cell growth factors and cyclosporin A, an immunosuppressive agent, on cloned osteoblastic cell line, MC3T3-E1 cells]. 326 92

The effect of interleukin-1 beta, the major component of osteoclast-activating factor (OAF), on bone formation by fetal rat osteoblast-rich cells was investigated. An in vitro culture system developed by Ecarot-Charrier et al. (1983) and Bellows et al. (1986) was utilized in which osteoblasts form mineralized nodules which closely resemble woven bone. Continuous exposure of cultures to homogenous IL-1 beta resulted in potent inhibition of mineralized nodule formation, which was half maximal at 0.1 U/ml (7.5 X 10(-13) M). Bone formation may thus be considerably more sensitive to IL-1 beta than is bone resorption (half maximal at 3.8 X 10(-11) M). Inhibition of bone formation occurred when cultures were exposed to IL-1 beta at both early and late time periods and was unaffected by the presence of indomethacin or by the osteoclast inhibitors calcitonin and gamma-interferon. Instead, IL-1 beta exerted multiple inhibitory effects on osteoblast functions, including inhibition of collagen and noncollagen protein synthesis, alkaline phosphatase expression, and cell replication. On a dose response basis, the inhibition of protein synthesis correlated most closely with inhibition of bone formation. IL-1 beta is clearly inhibitory rather than stimulatory for bone formation as assessed in this system and is therefore unlikely to function as a coupling factor linking the processes of bone resorption and bone formation.
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PMID:Interleukin-1 beta is a potent inhibitor of bone formation in vitro. 350 84

We studied changes in peripheral blood and bone marrow biopsy specimens obtained before, during, and after recombinant alpha 2b-interferon (IFN-alpha 2b) therapy in 25 patients with hairy cell leukemia. During therapy, only 1 patient showed no improvement in at least one of the parameters monitored. Granulocytopenia, thrombocytopenia, and monocytopenia resolved in 19/20, 14/15, and 17/18 patients, respectively. In 18/21 patients with Hb less than 12g/dl before treatment, the anemia became less severe. Hairy cells disappeared or decreased in numbers in the peripheral blood in all patients. In the bone marrow, numbers of hairy cells decreased and numbers of granulocytic, erythroid, and megakaryocytic cells increased usually within 3-6 months after the start of therapy. In no patient were hairy cells ever completely absent from the bone marrow. After cessation of IFN-alpha, the median Hb value, WBC, and platelet counts changed little for up to 12 months, but the absolute neutrophil count and absolute monocyte count decreased. Hairy cells reappeared in the peripheral blood of three patients. In the bone marrow the percentage of hairy cells increased, whereas the percentage of granulocytic and erythroid cells decreased. Neutrophil alkaline phosphatase (NAP) scores were abnormally high in 18/18 patients studied prior to IFN-alpha, but became normal in 17 of these during therapy and were normal in seven first studied during therapy. The median NAP score doubled by 3 months after cessation of therapy and was abnormal in 17/19 patients followed for 6 months. NAP score may be useful in predicting changes in the bone marrow in patients treated with IFN-alpha. We did not find any parameter in the pretherapy specimens that would have allowed us to predict individual response.
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PMID:Changes in peripheral blood and bone marrow specimens during and after alpha 2b-interferon therapy for hairy cell leukemia. 366 60

We constructed a new vector containing the promoter and the signal sequence of E. coli phoA gene, the structural gene for the periplasmic alkaline phosphatase. One of the most useful characteristics of this vector is the unique HindIII restriction site located just at the end of the phoA signal sequence. This restriction site was generated by oligonucleotide-directed site-specific mutagenesis without changing the amino acid sequence of the signal peptide. Any kind of foreign structural gene can be easily inserted into the HindIII site by using synthetic oligonucleotides to construct a hybrid gene which has neither an extra sequence nor a deletion between the phoA signal sequence and the foreign structural gene. Human alpha-interferon gene was inserted into this HindIII site. When this hybrid gene was expressed under the control of the phoA promoter region, a low but significant activity was recovered in the cold water wash of the cells after an osmotic shock procedure.
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PMID:Secretion of human interferon-alpha induced by using secretion vectors containing a promoter and signal sequence of alkaline phosphatase gene of Escherichia coli. 392 9

Three patients with pulmonary metastases of osteosarcoma underwent treatment with intravenous or intramuscular administration of human leukocyte interferon. In 2 cases, the size of the metastasized tumor mass diminished temporarily six or eight months after interferon treatment, and serum alkaline phosphatase levels decreased to normal. In 1 case, the tumor cell from the pleural exudate 1 could be isolated and cultivated in vitro. When 2,000 IU of interferon was added to the tumor cell culture, marked inhibition of tumor cell growth resulted. In the other case, interferon had no effect on the metastasized lung tumor.
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PMID:Effect of human leukocyte interferon on the metastatic lung tumor of osteosarcoma: case reports. 615 71

Leukocyte interferon was given by i.m. injection as adjuvant therapy to 9 patients with osteosarcoma. The dose was 3 X 10(6) standard units daily for one month, and then 3 times a week for the next 17 months. Blood samples were drawn at intervals for a number of routine tests during the 18-month course of interferon administration and during the subsequent 18 months. On withdrawal of the interferon treatment, the mean Hb concentration rose significantly and the mean ESR fell significantly. There was no significant change in the leukocyte and platelet counts or in the alkaline phosphatase, alanine and aspartate aminotransferase or plasma protein levels.
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PMID:Effect of long-term treatment with human leukocyte interferon on various laboratory parameters. 615 78


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