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)

In order to quantify human interleukin-8 (IL-8), which is chemotactic for T cells and basophils as well as neutrophils, we developed an enzyme-linked immunosorbent assay (ELISA). Since binding inhibition tests indicated that three monoclonal antibodies (mAbs; BS-1, WS-4, WS-6) blocked the binding of 125I-labelled IL-8 to neutrophils, we tested an ELISA using these mAbs as primary antibodies, rabbit anti-IL-8 Ab as the secondary antibody, and alkaline phosphatase-labelled goat anti-rabbit Ab as the conjugate. Among the three mAbs tested, WS-4 was the most sensitive with a detection limit of 16 pg/ml. Several other cytokines, including monocyte chemotactic and activating factor (MCAF), which is structurally related to IL-8, showed no cross-reactivity in this system, indicating that this ELISA is specific for IL-8. The coefficients of variation for the intra- and interassays were below 10%. Furthermore, this ELISA also detected natural IL-8 (including both 72 and 77 amino acid forms) produced by cultured human cells and cell lines stimulated with IL-1, suggesting that this system will be useful in the detection of natural IL-8 in various body fluids.
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PMID:A sensitive enzyme-linked immunosorbent assay for human interleukin-8. 159 35

Leukemia inhibitory factor/differentiation-stimulating factor (LIF/D-factor), expression of its mRNA, and possible roles in bone metabolism were studied in murine primary and clonal osteoblast-like cells. Local bone-resorbing factors such as IL-1, TNF alpha, and LPS strongly induced expression of LIF/D-factor mRNA in both clonal MC3T3-E1 cells and primary osteoblast-like cells. Neither parathyroid hormone nor 1 alpha,25-dihydroxyvitamin D3 stimulated expression of LIF/D-factor mRNA. LIF/D-factor per se did not stimulate expression of its own mRNA. Appreciable amounts of LIF/D-factor were detected in synovial fluids from rheumatoid arthritis (RA) patients but not in those with osteoarthritis (OA). Simultaneous treatment with LIF/D-factor, IL-1, and IL-6 at the concentrations found in synovial fluids from RA patients greatly enhanced bone resorption, though these cytokines did not stimulate bone resorption when separately applied. This suggests that LIF/D-factor produced by osteoblasts is in concert with other bone-resorbing cytokines such as IL-1 and IL-6 involved in the bone resorption seen in the joints of RA patients. LIF/D-factor specifically bound to MC3T3-E1 cells with an apparent dissociation constant of 161 pM and 1,100 binding sites/cell. LIF/D-factor dose-dependently suppressed incorporation of [3H]thymidine into MC3T3-E1 cells. In addition, it potentiated the alkaline phosphatase activity induced by retinoic acid, though LIF/D-factor alone had no effect on enzyme activity. These results suggest that LIF/D-factor is involved in not only osteoclastic bone resorption but also osteoblast differentiation in conjugation with other osteotropic factors.
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PMID:Leukemia inhibitory factor/differentiation-stimulating factor (LIF/D-factor): regulation of its production and possible roles in bone metabolism. 161 24

Ascites sarcoma 180 (S180A) is a transplantable tumor maintained in ddY mice. In the tumor-bearing mice, the plasma Ca, Pi and acid phosphatase levels increased and the plasma alkaline phosphatase levels decreased. The elevation of plasma Pi levels is unusual in humoral hypercalcemia of malignancy (HHM). To characterize the pathogenesis of HHM in the animals, the biological activities in the serum-free conditioned media (CM) of S180A cell cultures were examined. The S180A CM stimulated bone resorption dose dependently and showed TGF-like, IL-1-like and mitogenic activity. Unlike parathyroid hormone (PTH), the factor(s) failed to stimulate cAMP production by either UMR 106-01 cells or neonatal mouse calvaria at concentrations that stimulate bone resorption. Also, the factor(s) stimulated proliferation of UMR 106-01 cells concomitant with a slight increase in intracellular calcium levels. These results indicate that S180A cells produce a factor(s) responsible for bone resorption which is apparently different from PTH-like activity.
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PMID:Ascites sarcoma 180, an animal model of humoral hypercalcemia of malignancy, produces a factor(s) exhibiting potent bone-resorbing activity without any parathyroid hormone-like activity. 165 Nov 37

We have compared the adhesion of 51Cr-labeled eosinophils and neutrophils to cultured human umbilical vein endothelial cell (EC) monolayers that have been stimulated with IL-1, TNF, or LPS. Each agent stimulated the adhesion to EC of both eosinophils and neutrophils in a similar dose- and time-dependent manner. F(ab')2 fragments of mAb 1.2B6 (anti-endothelial leukocyte adhesion molecule (ELAM)-1) and mAb 6.5B5 (anti-intercellular adhesion molecule (ICAM)-1) each inhibited partially, and to a similar extent, eosinophil and neutrophil adhesion to EC monolayers prestimulated with TNF (10 ng/ml) for 6 h. Greater inhibition of both eosinophil and neutrophil adhesion was achieved by combining the effects of mAb 1.2B6 with either mAb 6.5B5 or mAb TS1/18 (anti-CD18). These observations indicate that both ELAM-1 and ICAM-1 are involved in the adhesion of eosinophils and neutrophils to EC stimulated with TNF. In order to determine whether these molecules are expressed in vivo during allergen-induced late phase allergic responses in the skin, human skin biopsies were examined at 6 h after Ag or saline challenge with the use of an alkaline phosphatase-staining technique. Both ELAM-1 and ICAM-1 were expressed with greater intensities in Ag-challenged biopsies, suggesting that these molecules may be involved in granulocyte recruitment in vivo. The similarities we have established between mechanisms of eosinophil and neutrophil adhesion to cytokine-stimulated EC suggests that factors other than differential leukocyte-EC adhesion may be responsible for the selective accumulation of eosinophils at sites of allergic inflammation.
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PMID:Endothelial leukocyte adhesion molecule-1 and intercellular adhesion molecule-1 mediate the adhesion of eosinophils to endothelial cells in vitro and are expressed by endothelium in allergic cutaneous inflammation in vivo. 171 Oct 84

Interleukin 6 (IL-6) exerts well-established effects on cells of the immune system as well as on various other cell types. It has been implicated in the control of connective tissue cells in such conditions as rheumatoid arthritis and osteoporosis. We have investigated the effects of recombinant human interleukin-6 (rhIL-6) on human osteoblastlike cells derived from explants of trabecular bone. ROS 17/2.8 cells were used as an additional osteoblastlike cell model system. We were unable to identify any effects of rhIL-6 (5-5000 pg/ml) on the proliferation, alkaline phosphatase activity. osteocalcin production, or release of cytokines or prostaglandins by either osteoblastlike cell model system. Since we have shown previously that these cells release IL-6 in culture, we used a sheep anti-human IL-6 antibody to investigate the possibility that (1) action of added exogenous IL-6 could be masking endogenous production, and (2) endogenous IL-6 may regulate the effects of osteotropic agents on the osteoblastlike cells. Presence of the antibody exerted no detectable effects on 1,25-(OH)2D3-stimulated alkaline phosphatase or on proliferation or TNF production enhanced by IL-1. Thus IL-6 does not appear to be involved in the regulation of osteoblast activity.
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PMID:Human osteoblastlike cells do not respond to interleukin-6. 170 32

Cytokines released at sites of inflammation and infection may alter normal bone remodeling processes resulting in pathologic bone destruction or bone formation. Interleukin 1, an inflammatory mediator, has been shown to stimulate as well as inhibit parameters associated with bone formation. In this study we have examined temporal aspects of the biphasic effects of recombinant interleukin 1 alpha (IL 1 alpha) on the differentiation of osteoprogenitor cells into bone-forming osteoblasts (bone nodules) in vitro. A dose-dependent stimulation of bone formation over a concentration range of 0.5 to 50 U/mL (1.4 x 10(-12) to 1.4 x 10(-10) M) was observed when preconfluent, primary cultures of fetal rat calvaria (RC) cells were pulsed with IL 1 alpha for 72 to 96 hr from the beginning of the culture period. This was correlated with a stimulation of cell proliferation and alkaline phosphatase activity measured during the late log phase of growth. In contrast, continuous exposure to IL 1 alpha or exposure to IL 1 alpha after confluency resulted in inhibition of bone nodule formation and alkaline phosphatase activity. IL 1 alpha-stimulated prostaglandin E2 (PGE2) production until the RC cells became multilayered, but the addition of the cyclooxygenase inhibitor indomethacin had no effect in reducing the IL 1 alpha-mediated stimulation of cell proliferation or bone nodule formation. However, in cultures continuously exposed to IL 1 alpha, added indomethacin partially reduced the inhibition of bone formation, suggesting that prostaglandin production may play a role in the inhibitory effects of IL 1 alpha on bone formation.
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PMID:Temporal sequence of interleukin 1 alpha-mediated stimulation and inhibition of bone formation by isolated fetal rat calvaria cells in vitro. 210 36

Human osteoblast cultures derived as out-growths from trabecular bone released tumor necrosis factor (TNF alpha) upon stimulation of the cells with human recombinant interleukin 1 (IL1; 10(-13)-10(-11) M), human recombinant granulocyte-macrophage colony-stimulating factor (100-1000 U/ml), and bacterial lipopolysaccharide (5-500 ng/ml). The osteotropic hormones 1,25-dihydroxyvitamin D3, PTH, and calcitonin had no effect on TNF production. The TNF released by the osteoblasts was identified as TNF alpha, using a specific anti-TNF alpha monoclonal antibody to neutralize its activity. Immunohistochemical staining of the cells using the same antibody revealed that all of the cells in the cultures were capable of producing TNF alpha, including those that also expressed alkaline phosphatase activity. Immunoreactive protein could be detected in the perinuclear region when cells were cultured in the presence of monensin, suggesting accumulation of newly synthesised protein in the Golgi apparatus. These results suggest that human osteoblasts, which have been shown previously to respond to TNF alpha, can synthesize and release TNF in response to IL1 and granulocyte-macrophage colony-stimulating factor. TNF may, therefore, not only have a pathological role in conditions of chronic inflammation, but also may act as a local paracrine or autocrine regulator of osteoblast function.
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PMID:Production of tumor necrosis factor by human osteoblasts is modulated by other cytokines, but not by osteotropic hormones. 240 45

The effects of interleukin 1, transforming growth factor-beta (coupling factors), prostaglandin E1, and prostaglandin E2 on incorporation of 45Ca2+ and on alkaline phosphatase activity were studied using cultured ROS 17/2.8 cells, one of cell lines derived from rat osteosarcoma. We found that all these factors stimulate both the incorporation of 45Ca2+ and alkaline phosphatase activity of these cells. On the other hand, one of the bone resorption hormones, parathyroid hormone (PTH), suppressed the proliferation of cells and decreased the alkaline phosphatase activity at considerably low concentrations (1 X 10(-12)-1 X 10(-11) M). However, the hormone stimulated the incorporation of 45Ca2+ by these cells in a dose-dependent manner; the maximum stimulation on day 3 was observed at 1 X 10(-7) M and it was approximately 3 times the control value. The data suggest therefore, that the osteoblasts incorporated calcium ions and transported them while bone resorption was occurring. Thus the ROS 17/2.8 cell line appears to be an advantageous experimental system for the study of calcium metabolism of osteoblasts in vitro.
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PMID:[Effects of various factors involved in bone metabolism on 45Ca2+ incorporation and alkaline phosphatase activity of ROS 17/2.8 cells]. 260 4

An immunoenzymatic detection method of in-situ hybridization reactions for interleukin 1 (IL-1) beta was established, using sulphonated probes. As model system we used unstimulated and lipopolysaccharide (LPS) stimulated peripheral blood mononuclear cells (PBMNC). After hybridization, sulphone groups were targeted with a monoclonal antibody, and bound antibody was visualized by the alkaline phosphatase anti-alkaline phosphatase (APAAP) method. In unstimulated PBMNC both the control and the IL-1 beta specific c-DNA probes were negative, whilst a large proportion of LPS treated PBMNC was positive with the sulphonated IL-1 beta plasmid only. This method may be a powerful alternative to radio-isotopic labeling. Since the entire procedure can be performed within one day, it may be applied for routine diagnostic purposes.
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PMID:Immunoenzymatic assessment of IL-1 beta mRNA by in situ hybridization using sulphonated probes. 267 55

Granulated lymphoid cells (CD2+, CD7+, CD38+, NKH1+, CD3-, CD5-, CD4-, CD8-, CD25-) are prominent in human endometrial stroma in the late secretory phase of the menstrual cycle and in early pregnancy, and may play an important role in implantation and placentation. Cell suspensions enriched for granulated lymphoid cells were prepared from first trimester human decidua using a panning technique; cells were labelled with the monoclonal antibody NKH1 and separated by adherence to immunoglobulin-coated plates. The enriched cells were characterized with a panel of monoclonal antibodies using an indirect immuno-alkaline phosphatase method, and subjected to various functional assays. Most cells in the enriched preparations showed the characteristic morphology of granulated lymphocytes in smears stained with toluidine blue or May Grunwald Giemsa. CD45+ cells were obtained up to 98 +/- 1% purity (n = 10) and CD2+ cells were enriched up to 84 +/- 4%. The enriched populations were efficient effectors in a K562 chromium-release assay but showed minimal proliferative response to phytohaemagglutinin, concanavalin A, ionomycin and phorbol 12, 13 dibutyrate (PdBU), interleukin 1 or interleukin 2. The precise lineage and in vivo function of decidual granulated lymphocytes remains to be established.
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PMID:Isolation and functional studies of granulated lymphocytes in first trimester human decidua. 277 61


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