Gene/Protein Disease Symptom Drug Enzyme Compound
Pivot Concepts:   Target Concepts:
Query: EC:3.1.3.1 (alkaline phosphatase)
 47,916 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Endochondral ossification in growth plates proceeds through several consecutive steps of late cartilage differentiation leading to chondrocyte hypertrophy, vascular invasion, and, eventually, to replacement of the tissue by bone. The subchondral vascular system is essential for this process and late chondrocyte differentiation is subject to negative control at several checkpoints. Endothelial cells of subchondral blood vessels not only are the source of vascular invasion accompanying the transition of hypertrophic cartilage to bone but also produce factors overruling autocrine barriers against late chondrocyte differentiation. Here, we have determined that the action of proteases secreted by endothelial cells were sufficient to derepress the production of the hypertrophy-markers collagen X and alkaline phosphatase in arrested populations of chicken chondrocytes. Signalling by thyroid hormones was also necessary but endothelial factors other than proteinases were not. Negative signalling by PTH/PTHrP- or TGF-beta-receptors remained unaffected by the endothelial proteases whereas signalling by FGF-2 did not suppress, but rather activated late chondrocyte differentiation under these conditions. A finely tuned balance between chondrocyte-derived signals repressing cartilage maturation and endothelial signals promoting late differentiation of chondrocytes is essential for normal endochondral ossification during development, growth, and repair of bone. A dysregulation of this balance in permanent joint cartilage also may be responsible for the initiation of pathological cartilage degeneration in joint diseases.
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PMID:Role of the subchondral vascular system in endochondral ossification: endothelial cell-derived proteinases derepress late cartilage differentiation in vitro. 1142 Jan 52

High throughput screening (HTS) of large compound libraries for inhibitors of growth factors raises the requirement for simple yet reliable assays. Fibroblast growth factors (FGFs) play a pivotal role in the multistep pathway of malignant transformation, tumor progression, metastasis, and angiogenesis. FGF-2 (basic FGF) requires a cooperative interaction with heparin or heparan sulfate proteoglycans in order to form functional growth factor-receptor complexes that are essential for receptor binding and activation. We have developed a simple screening system, devised to identify molecules that modulate heparin-FGF-receptor interactions. The system is composed of a heparin matrix, FGF-2, and a FGF receptor-1 protein engineered by genetically fusing the extracellular domain of FGF receptor-1 to alkaline phosphatase (FRAP). The screen is conducted using 96-well plates to which heparin has been covalently attached. FGF-2 is then bound to the plates through heparin-FGF interactions, followed by the addition of FRAP and compounds to be screened for modulation of heparin-FGF, receptor-heparin, and receptor-FGF interactions. The endpoint of the assay is measured enzymatically using the alkaline phosphatase (AP)-catalyzed formation of a chromogenic product, which is directly proportional to the amount of FRAP present on the plates as a heparin-FGF-FRAP ternary complex. Reduced AP values relative to control, as measured by spectrophotometry, indicate inhibition of the formation of an active FGF-receptor-heparin complex. The simple and versatile nature of the assay makes it an attractive HTS system. The screen has identified several potent inhibitors of FGF-2 receptor binding and activation. Furthermore, secondary screening of the HTS-recognized compounds identified several compounds that have the capacity to block growth factor-mediated tumor progression and angiogenesis in vivo.
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PMID:Development of a high throughput screening assay for inhibitors of fibroblast growth factor-receptor-heparin interactions. 1168 13

FGF-2 stimulates bone formation in vitro and in vivo in rats. However, there are limited studies in mice and no data on the mechanism(s) by which FGF-2 induces bone formation. We assessed whether short-term FGF-2 treatment of marrow stromal cells from young mice would increase alkaline phosphatase-positive (ALP), mineralized colony formation and expression of genes important in osteoblast maturation. Short-term treatment with FGF-2 (0.01-1.0 nM) for the first 3 days of a 14- or 21-day culture period increased the number of ALP mineralized colonies in bone marrow stromal cells. FGF-2 (0.1 nM) increased the mRNAs for type 1 collagen: osteocalcin, runt domain/core binding factor, PTH/PTHR receptor, and insulin-like growth factor 1 (IGF-1) at 14 and 21 days. We conclude that short-term FGF-2 treatment enhances osteoblast maturation in vitro. Furthermore, the anabolic effect of FGF-2 may be attributed in part to regulation of IGF-1 in osteoblasts.
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PMID:FGF-2 increases colony formation, PTH receptor, and IGF-1 mRNA in mouse marrow stromal cells. 1177 3

To clarify the effect of recombinant human basic fibroblast growth factor (FGF-2) on the osteoinductive activity of recombinant human bone morphogenetic protein-2 (BMP-2) in vivo, different amounts of FGF-2 (0, 16, 80 and 400 ng, and 2, 10 and 50 micro g: n=10 in each group), BMP-2 (2 micro g) and type I collagen as a carrier were mixed and implanted into rat calf muscles. Three weeks after implantation, compared with the controls, the radiopaque shadows of the implants were increased in the 16, 80 and 400 ng FGF-2-treated groups, but decreased in the 2, 10 and 50 micro g FGF-2-treated groups. In addition, alkaline phosphatase activity was increased in the 16, 80 and 400 ng FGF-2-treated groups but decreased in the 50 micro g FGF-2-treated group. Histological examination revealed increased bone formation in the 16, 80 and 400 ng FGF-2-treated groups. These results show that combined treatment with FGF-2 and BMP-2 has a biphasic effect on osteoinductive activity, i.e. it increases with low doses of FGF-2 and decreases with high doses of FGF-2.
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PMID:The effect of fibroblast growth factor-2 on the osteoinductive activity of recombinant human bone morphogenetic protein-2 in rat muscle. 1222 Oct 14

The cause of progressive dermal sclerosis, proliferation of fibroblasts, and collagen deposition in scleromyxedema is unknown. We analyzed the heparan sulphate proteoglycans (HSPG) in cutaneous nodules from a patient with scleromyxedema in order to ascertain their role in the binding of fibroblast growth factor (FGF) and promoting signaling complex assembly. Total heparan sulphate (HS) was detected with a monoclonal antibody to HSPG on paraffin sections. Binding of FGF to HS was assessed using FGF-2 followed by anti-FGF-2 antibody. The formation of HS-mediated signaling complex was studied using soluble FR1-AP, which contains the extracellular domain of FGF receptor-1 linked to alkaline phosphatase (AP) and monoclonal anti-AP-antibody. Anti FGF-2 and anti-AP antibodies were visualized using the DAKO Envision Plus system. The dermal nodule of scleromyxedema contained ample HS and these bound FGF-2 and FR1-AP. Specificity was confirmed by prior incubation with heparitinase (no staining) and omission of FGF-2 (no staining). Increased amounts of HSPG were present in the dermal nodules of scleromyxedema compared to adjacent normal dermis and these bound FGF-2, immobilized the soluble receptor protein FGFR-1 and, therefore, formed a ternary complex composed of HSPG, FGF-2 and FGFR-1 in vitro. Since this complex resembles the signaling complex formed on live cells, HSPG in the nodules of scleromyxedema are in a configuration that promotes FGF activity.
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PMID:Heparan sulphate proteoglycan in scleromyxedema promotes FGF-2 activity. 1249 27

We examined the effects of basic fibroblast growth factor (FGF-2) on cultured lower molar tooth germ at the differentiative (bell) stage. Although FGF-2 has been detected in odontogenesis, its roles in biological activities, such as cell proliferation, differentiation and extracellular matrix mineralization are unclear. We assayed mRNA levels of the differentiation markers, dentine sialophosphoprotein (DSPP), amelogenin and alkaline phosphatase (ALP) using reverse transcription-polymerase chain reaction (RT-PCR), and histological methods. Tooth germs dissected from 17-day-old embryonic mice were cultured for 4 days with either recombinant human FGF-2 or specific antisense phosphorothioate oligodeoxynucleotide (antisense ODN) for FGF-2. Exogenous FGF-2 decreased the gene expression of differentiation markers in molars at the bell stage. Abrogation of endogenous FGF-2 by antisense ODN increased the gene expression of differentiation markers, and also significantly enhanced enamel and dentine formation. This histological change was recovered by adding exogeneous FGF-2. These findings suggest that FGF-2 at the bell stage regulates cell differentiation and matrix secretion.
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PMID:Fgf-2 regulates enamel and dentine formation in mouse tooth germ. 1295 98

Fibroblast growth factors (FGFs) are involved in stimulation of angiogenesis in tumors and other pathological circumstances. Increased activity of normal skeletal muscles resulting from chronic electrical stimulation is a very potent stimulus for capillary growth but a relationship between the initiation of this angiogenesis and the involvement of autocrine growth factors has yet to be established. Although FGF expression has been reported in muscles stimulated for 3 weeks, capillary growth is underway significantly earlier, beginning around 3 days. The present experiments have therefore studied the possible involvement of basic fibroblast growth factor (FGF-2) in stimulated rat fast skeletal muscles prior to, and coincident with, capillary growth. Muscle contractions were induced via electrodes implanted in the vicinity of the peroneal nerve and maintained for 8h/day for 2, 4 or 7 days. Capillary/fiber ratio (C/F), based on staining of capillary endothelium for alkaline phosphatase, was not changed in either extensor digitorum longus (EDL) or tibialis anterior (TA) after 2 days stimulation, but increased in TA stimulated for 4 days and in both muscles after 7 days. The expression of mRNA for FGF-2, detected by ribonuclease protection assay, was decreased in all stimulated muscles compared with control or contralateral muscles; immunohistochemistry showed FGF-2 gene product in nerves and larger blood vessels but not in capillaries. There was no evidence from immunohistochemistry for up-regulation of receptors flg and bek for FGF-2. The presence of FGF-2, flg and bek in arterioles may indicate a possible role for FGF-2 in the regulation of blood flow since we have previously shown it to be a dilator of small arterioles. However, based on the lack of correlation between changes in capillary density and the expression of mRNA and protein for FGF-2 and its receptors, it is unlikely that it is directly linked with the initiation of angiogenesis resulting from chronic activity in skeletal muscles.
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PMID:Lack of involvement of basic fibroblast growth factor (FGF-2) in capillary growth in skeletal muscles exposed to long-term contractile activity. 1451 78

The osteogenic factors bone morphogenetic protein (BMP-7), platelet-derived growth factor (PDGF)-BB, and fibroblast growth factor (FGF-2) regulate the recruitment of osteoprogenitor cells and their proliferation and differentiation into mature osteoblasts. However, their mechanisms of action on osteoprogenitor cell growth, differentiation, and bone mineralization remain unclear. Here, we tested the hypothesis that these osteogenic agents were capable of regulating osteoblast differentiation and bone formation in vitro. Normal human bone marrow stromal (HBMS) cells were treated with BMP-7 (40 ng ml(-1)), PDGF-BB (20 ng ml(-1)), FGF-2 (20 ng ml(-1)), or FGF-2 plus BMP-7 for 28 days in a serum-containing medium with 10 mM beta-glycerophosphate and 50 microg ml(-1) ascorbic acid. BMP-7 stimulated a morphological change to cuboidal-shaped cells, increased alkaline phosphatase (ALKP) activity, bone sialoprotein (BSP) gene expression, and alizarin red S positive nodule formation. Hydroxyapatite (HA) crystal deposition in the nodules was demonstrated by Fourier transform infrared (FTIR) spectroscopy only in BMP-7- and dexamethasone (DEX)-treated cells. DEX-treated cells appeared elongated and fibroblast-like compared to BMP-7-treated cells. FGF-2 did not stimulate ALKP, and cell morphology was dystrophic. PDGF-BB had little or no effect on ALKP activity and biomineralization. Alizarin Red S staining of cells and calcium assay indicated that BMP-7, DEX, and FGF-2 enhanced calcium mineral deposition, but FTIR spectroscopic analysis demonstrated no formation of HA similar to human bone in control, PDGF-BB-, and FGF-2-treated samples. Thus, FGF-2 stimulated amorphous octacalcium phosphate mineral deposition that failed to mature into HA. Interestingly, FGF-2 abrogated BMP-7-induced ALKP activity and HA formation. Results demonstrate that BMP-7 was competent as a sole factor in the differentiation of human bone marrow stromal cells to bone-forming osteoblasts confirmed by FTIR examination of mineralized matrix. Other growth factors, PDGF, and FGF-2 were incompetent as sole factors, and FGF-2 inhibited BMP-7-stimulated osteoblast differentiation.
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PMID:Differential growth factor control of bone formation through osteoprogenitor differentiation. 1500 88

The objective of this study is to investigate the proliferation and differentiation of stromal cells derived from human adipose tissues cultured on substrates with different surface properties. In addition, a similar investigation was performed on cells proliferated in different concentrations of basic fibroblast growth factor (FGF-2). The culture substrates include several polymer films with different water wettabilities, glass or a cell-culture plate, and that coated with collagen type I or IV, gelatin and FGF-2. The proliferation profiles of cells were influenced by the type of culture substrate and the growth factor concentration. A larger number of proliferated cells was observed for substrates with a water contact angle around 80 degrees, while the cell number was significantly larger for every protein-coated substrate. The rate of cell proliferation became maximal at a FGF-2 concentration of 1000 ng/ml. The FGF-2 concentration used for cell proliferation affected the differentiation profile of cells proliferated. Stromal cells, proliferated in 1 ng/ml FGF-2, were osteogenically differentiated to the strongest and fastest extent among those in other growth factor doses. The alkaline phosphatase (ALP) activity of cells increased with the increased cell number, although the activity per cell was identical, irrespective of the substrate type. The strongest adipogenic differentiation was observed for cells proliferated in 1000 ng/ml FGF-2 and the differentiation induction was maintained for a long time period. No clear dependence of the cell number on adipogenesis was observed. These findings indicate that the proliferation and differentiation of human adipose tissue-derived stromal cells are influenced by the culture substrate and the concentration of FGF-2 used for proliferation.
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PMID:Effect of culture substrate and fibroblast growth factor addition on the proliferation and differentiation of human adipo-stromal cells. 1579 5

We have developed a serum-free medium, designated ESF7, in which leukemia inhibitory factor (LIF) clearly stimulated murine embryonic stem (ES) cell proliferation accompanied by increased expression of nanog and Rex-1 and decreased FGF-5 expression. These effects were dependent on the concentration of LIF. The ES cells maintained in ESF7 medium for more than 2 yr retained an undifferentiated phenotype, as manifested by the expression of the transcription factor Oct-3/4, the stem cell marker SSEA-1, and alkaline phosphatase. Withdrawal of LIF from ESF7 medium resulted in ES cell apoptosis. Addition of serum to ESF7 medium promoted ES cell differentiation. Addition of BMP4 promoted ES cell differentiation into simple epithelial-like cells. In contrast, FGF-2 promoted ES cell differentiation into neuronal and glial-like cells. Under serum-free culture conditions, LIF was sufficient to stimulate cell proliferation, it inhibited cell differentiation, and it maintained self-renewal of ES cells. Because this simple serum-free adherent monoculture system supports the long-term propagation of pluripotent ES cells in vitro, it will allow the elucidation of ES cell responses to growth factors under defined conditions.
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PMID:Leukemia inhibitory factor as an anti-apoptotic mitogen for pluripotent mouse embryonic stem cells in a serum-free medium without feeder cells. 1592 56


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