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)

Vasculotropin/vascular endothelial growth factor (VAS/VEGF) is an angiogenic growth factor whose biological activity seems to be restricted in vitro to vascular endothelial cells. We describe here that fetal bovine osteoblasts (OB) bind VAS/VEGF but do not proliferate upon its addition. However VAS/VEGF induces migration, PTH-dependent cAMP accumulation and alkaline phosphatase increase when added to OB. The maximal effects reach levels comparable to that obtained with bone morphogenetic protein 2 (BMP-2), although the VAS/VEGF concentrations required are at least 100 fold lower. Our results suggest that VAS/VEGF could be an important regulator of osteoblastic differentiation.
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PMID:Vasculotropin/vascular endothelial growth factor induces differentiation in cultured osteoblasts. 812 39

The 165-amino acid form of vascular endothelial growth factor (VEGF165) is a mitogen for vascular endothelial cells and a potent angiogenic factor. Expression of a chimeric receptor containing the extracellular domain of the flk-1 receptor fused to the transmembrane and intracellular domains of the human c-fms receptor in NIH-3T3 cells, resulted in the appearance of high affinity binding sites for 125I-VEGF165 on transfected cells. The binding of 125I-VEGF165 to the flk-1/fms chimeric receptor of the transfected cells as well as the VEGF165-induced autophosphorylation of the chimeric receptors were inhibited in the presence of low concentrations of heparin (1-10 micrograms/ml). In contrast, similar concentrations of heparin potentiated the binding of 125I-VEGF165 to the endogenous VEGF receptors of the transfected cells, indicating that to some extent, the effect of heparin on 125I-VEGF165 binding is receptor type-dependent. A soluble fusion protein containing the extracellular domain of flk-1 fused to alkaline phosphatase (flk-1/SEAP) was used to study the effects of heparin on the binding of 125I-VEGF165 to flk-1 in a cell-free environment. The fusion protein specifically inhibited VEGF165-induced proliferation of vascular endothelial cells, but bound 125I-VEGF165 inefficiently in the absence of heparin. Addition of low concentrations of heparin or heparan sulfate (0.1-1 microgram/ml) resulted in a strong potentiation of 125I-VEGF165 binding, whereas higher heparin or heparan sulfate concentrations inhibited the binding. The effect of heparin on the binding of 125I-VEGF165 to flk-1/SEAP could not be mimicked by desulfated heparin or by chondroitin sulfate. Both bFGF and aFGF inhibited the binding when low concentrations of heparin were added to the binding reaction. However, higher concentrations of heparin abolished the inhibition, indicating that the inhibition is probably caused by competition for available heparin. Taken as a whole, these results indicate that heparin-like molecules regulate the binding of VEGF165 to its receptors in complex ways which depend on the heparin binding properties of VEGF165, on the specific VEGF receptor type involved, and on the amount and composition of heparin-like molecules that are present on the cell surface of VEGF receptor containing cells.
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PMID:Heparin modulates the interaction of VEGF165 with soluble and cell associated flk-1 receptors. 817 51

Fetal liver kinase-1 (Flk-1) is a transmembrane tyrosine kinase that was identified in endothelial cells and populations of cells enriched in hematopoietic progenitors. To characterize the interaction of Flk-1 with potential ligands the receptor extracellular domain was genetically fused to an alkaline phosphatase (AP) tag. A soluble ligand for Flk-1 was identified in the supernatants of numerous mesenchymal cell lines by co-immunoprecipitation with the Flk1-AP fusion protein. This polypeptide was shown by N-terminal sequencing to be vascular endothelial growth factor (VEGF). Receptor-AP fusion proteins can thus be used to identify soluble ligands as well as transmembrane ligands, and this approach is therefore likely to be widely applicable to many types of orphan receptor. The Flk1-AP soluble receptor was also found to bind to cell surfaces, showing two apparent classes of binding site with different affinities. This interaction could be reconstructed by introducing a VEGF expression plasmid into cells. These results indicate that VEGF presented at the cell surface can bind to the Flk-1 receptor, and could mediate a direct cell-cell interaction. The Flk1-AP fusion protein was also found to bind heparin, implying that ligand binding by the Flk-1 receptor may involve a three way interaction between the Flk-1 receptor, VEGF, and heparin-like cell surface proteoglycans.
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PMID:Interactions between the Flk-1 receptor, vascular endothelial growth factor, and cell surface proteoglycan identified with a soluble receptor reagent. 852 58

Throughout development of the vertebrate retina, progenitor cells are multipotential, producing a variety of distinctive cell types. Little is known of the molecular mechanisms directing the determination of cell fate. We have examined retinal progenitor cells for expression of receptor tyrosine kinases in an attempt to define receptors that could allow a progenitor to respond to its environment. We found that the receptor tyrosine kinase Flk-1, previously shown to be expressed in endothelial cells, is also expressed in neural progenitor cells of the mouse retina. Flk-1 RNA expression in the retinal progenitors commences with the onset of neuronal differentiation and persists throughout retinal neurogenesis. Flk-1 RNA and protein levels in the retina vary temporally during development, as shown by in situ hybridization and Western blot analysis. Patterns of beta-galactosidase expression in mice containing the lacZ gene in place of the Flk-1 gene are consistent with Flk-1 being expressed in retinal progenitors. In addition, we show that the ligand of Flk-1, vascular endothelial growth factor (VEGF), is expressed in the developing retina by differentiated cells and that a chimeric ligand of VEGF fused to alkaline phosphatase binds to proliferating retinal progenitors. Furthermore, the neural retina-derived Flk-1 protein kinase is activated by VEGF in vitro. Thus, the Flk-1 receptor protein kinase is expressed on the surface of neural progenitors in mouse retina and may play a critical role in neurogenesis as well as in vasculogenesis.
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PMID:Flk-1, a receptor for vascular endothelial growth factor (VEGF), is expressed by retinal progenitor cells. 881 91

Human osteoblast-like cells (HOB) produce vascular endothelial growth factor (VEGF), the steady state level of which is stimulated by 1,25-dihydroxyvitamin D3 [1,25-(OH)2D3]. As osteoblasts and endothelial cells are proximally located in skeletal tissue, we investigated the anabolic effects of 1,25-(OH)2D3 and VEGF on HOB cocultured with endothelial cells. When HOB with high alkaline phosphatase (Al-P) activity and human umbilical vein endothelial cells (HUVEC) with little activity were cultured together, Al-P activity increased, accompanied by an increase in cell number. When HOB and HUVEC were cultured separately, 1,25-(OH)2D3 did not directly stimulate [3H]thymidine incorporation into HUVEC, but stimulated it in the presence of HOB. VEGF did not directly stimulate the Al-P activity of HOB but stimulated it in the presence of HUVEC. The conditioned medium of HOB stimulated the proliferation of HUVEC, and this was partially blocked by anti-VEGF antibody. Conversely, the conditioned medium of HUVEC increased Al-P activity and [3H]thymidine incorporation into HOB, and this was partially blocked by antiinsulin-like growth factor I antibody and BQ-123, a specific antagonist of the endothelin-1 (ET-1) receptor. 1,25-(OH)2D3 stimulated the release of VEGF and ET-1 from HOB and HUVEC, respectively. Furthermore, the 1,25-(OH)2D3-induced release of VEGF was enhanced in HOB cocultured with HUVEC. A quantitative reverse transcription-PCR study revealed that genes for VEGF receptors (Flt-1 and KDR) were expressed in HUVEC, but not in HOB, and that 1,25-(OH)2D3 increased the levels of expression of VEGF receptor genes in endothelial cells only when cocultured with HOB. In summary, we demonstrated that 1,25-(OH)2D3 exerts an anabolic effect on osteoblasts by enhancing their production of VEGF, which stimulates its receptors on endothelial cells, followed by increased production of osteotropic growth factors, such as insulin-like growth factor I and ET-1. These in vitro findings suggest that the VEGF/VEGF receptor system may be involved in both bone formation and bone remodeling in vivo.
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PMID:Anabolic effects of 1,25-dihydroxyvitamin D3 on osteoblasts are enhanced by vascular endothelial growth factor produced by osteoblasts and by growth factors produced by endothelial cells. 920 40

A gene therapy strategy involving direct myocardial administration of an adenovirus (Ad) vector encoding the vascular endothelial growth factor 121 cDNA (Ad(GV)VEGF121.10) has been shown to be capable of "biological revascularization" of ischemic myocardium in an established porcine model [Mack, C.A. (1998). J. Thorac. Cardiovasc. Surg. 115, 168-177]. The present study evaluates the local and systemic safety of this therapy in this porcine ischemia model and in normal mice. Myocardial ischemia was induced in Yorkshire swine with an ameroid constrictor 21 days prior to vector administration. Ad(GV)VEGF121.10 (10(9) or 10(10) PFU), Ad5 wild type (10(9) PFU), AdNull (control vector with no transgene; 10(9) PFU), saline, or no injection (naive) was administered in 10 sites in the ischemic, circumflex distribution of the myocardium. Toxicity was assessed by survival, serial echocardiography, blood analyses, and myocardial and liver histology at 3 and 28 days after vector administration. All pigs survived to sacrifice, except for one animal in the Ad(GV)VEGF121.10 (10(10) PFU) group, which died as a result of oversedation. Echocardiograms of Ad(GV)VEGF121.10-treated pigs demonstrated no differences in pericardial effusion, mitral valve regurgitation, or regional wall motion compared with control pigs. Intramyocardial administration of Ad(GV)VEGF121.10 included only minimal myocardial inflammation and necrosis, and no hepatic inflammation or necrosis. Only a mild elevation of the white blood cell count was encountered on day 3, which was transient and self-limited in the Ad(GV)VEGF121.10 group as compared with the saline-treated animals. As a measure of inadvertent intravascular administration of vector, normal C57/BL6 mice received intravenous Ad(GV)VEGF121.10 (10(4), 10(6), 5 x 10(7), or 10(9) PFU), AdNull (5 x 10(7) or 10(9) PFU), or saline. Toxicity was assessed by survival, blood analyses, and organ histology at 3 and 7 days after vector administration. A separate group of C57/BL6 mice received intravenous AdmVEGF164 (Ad vector encoding the murine VEGF164 cDNA), Ad(GV)VEGF121.10, AdNull (10(8) PFU each group), or saline to assess duration of expression and safety of a homologous transgene. All mice survived to sacrifice except for 40% of the mice in the highest (10(9) PFU; a dose more than 10(3)-fold higher by body weight than the efficacious dose in pigs) Ad(GV)VEGF121.10 dose group, which died on days 5-6 after vector administration. The only differences seen in the blood analyses between treated and control mice were in the very high Ad(GV)VEGF121.10 dose group (10(9) PFU), which demonstrated an anemia as well as an increase in alkaline phosphatase when compared with all other treatment groups. Hepatic VEGF levels by ELISA in AdmVEGF164-treated mice did not persist beyond 14 days after vector administration, suggesting that persistent expression of a homologous VEGF gene transferred with an Ad vector is not a significant safety risk. Although this is not a chronic toxicity study, these data demonstrate the safety of direct myocardial administration of Ad(GV)VEGF121.10, and support the potential use of this strategy to treat human myocardial ischemia.
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PMID:Safety of direct myocardial administration of an adenovirus vector encoding vascular endothelial growth factor 121. 1036 64

Osteogenic protein-1 (OP-1 or BMP-7) stimulates new bone formation in vivo and induces cell proliferation and differentiation of osteoblasts in vitro. In the present study, we examined effects of OP-1 on the expression of vascular endothelial growth factor (VEGF) in primary cultures of fetal rat calvaria (FRC) cells. OP-1 increased the steady-state level of VEGF mRNA by about 3-fold in an OP-1 concentration- and time-dependent manner. The increase in VEGF mRNA level depended on transcription and was sensitive to cell replication. The VEGF mRNA stability was unaffected. The mRNA levels for both types of VEGF receptors, Flk-1 and Flt-1 were low but detectable in FRC cells by RT-PCR and were not changed by OP-1. Inhibition of VEGF synthesis and function by antisense oligonucleotide and by suramin, respectively arrested the OP-1-induced alkaline phosphatase activity and mineralized bone nodule formation. Together with published studies of VEGF on vascular endothelial cells which are usually found in close proximity to osteoblastic cells in vivo, these results suggest that VEGF participates in the OP-1-induced osteogenesis by taking part in bone cell differentiation and by promoting angiogenesis at the site of bone formation.
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PMID:Osteogenic protein-1 increases gene expression of vascular endothelial growth factor in primary cultures of fetal rat calvaria cells. 1045 59

To investigate the relationship between angiogenic growth factors and endothelial enzyme activity in capillaries after injury of rat cardiomyocytes caused by X irradiation, 7-week-old male Wistar rats were anesthetized with pentobarbitone and their hearts irradiated (X rays, 20 Gy) through a hole in the lead casing in which they were enclosed. The hearts were excised at 1 h, 1 week and 3 weeks after irradiation. Left ventricular cross sections were stained for capillary enzymes by double staining for two endothelial enzymes, alkaline phosphatase (AP) and dipeptidylpeptidase IV (DPP), immunohistochemically stained for basic fibroblast growth factor (Fgf, also known as bFgf) and vascular endothelial growth factor (Vegf), and stained for nick end-labeling of DNA by the TUNEL method. Staining for distribution of AP in the arteriolar portion was reduced at both 1 and 3 weeks after irradiation with 20 Gy, but staining for DPP in the venular portion was unchanged, suggesting a close relationship between growth factors and injury of the arteriolar capillary portion. Fgf and Vegf proteins were present within the cytoplasm of the cardiomyocytes, or around capillaries, 1 h, 1 week and 3 weeks after irradiation. Many TUNEL-stained cardiomyocyte nuclei were observed at 1 h, but they had decreased markedly at 1 week and had almost disappeared by 3 weeks after irradiation. Thus Fgf and Vegf were induced concomitantly with the decrease in the staining for endothelial AP by 20 Gy X irradiation, which also caused microeffects as indicated by TUNEL staining of many nuclei at 1 h postirradiation.
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PMID:Induction of growth factors in rat cardiac tissue by X irradiation. 1079 Feb 75

The well-described detrimental effects of ionizing radiation on the regeneration of bone within a fracture site include decreased osteocyte number, suppressed osteoblast activity, and diminished vascularity. However, the biologic mechanisms underlying osteoradionecrosis and the impaired fracture healing of irradiated bone remain undefined. Ionizing radiation may decrease successful osseous repair by altering cytokine expression profiles resulting from or leading to a change in the osteoblastic differentiation state. These changes may, in turn, cause alterations in osteoblast proliferation and extracellular matrix formation. The purpose of this study was to investigate the effects of ionizing radiation on the proliferation, maturation, and cytokine production of MC3T3-E1 osteoblast-like cells in vitro. Specifically, the authors examined the effects of varying doses of ionizing radiation (0, 40, 400, and 800 cGy) on the expression of transforming growth factor-beta1 (TGF-beta1), vascular endothelial growth factor (VEGF), and alkaline phosphatase. In addition, the authors studied the effects of ionizing radiation on MC3T3-E1 cellular proliferation and the ability of conditioned media obtained from control and irradiated cells to regulate the proliferation of bovine aortic endothelial cells. Finally, the authors evaluated the effects of adenovirus-mediated TGF-beta1 gene therapy in an effort to "rescue" irradiated osteoblasts. The exposure of osteoblast-like cells to ionizing radiation resulted in dose-dependent decreases in cellular proliferation and promoted cellular differentiation (i.e., increased alkaline phosphatase production). Additionally, ionizing radiation caused dose-dependent decreases in total TGF-beta1 and VEGF protein production. Decreases in total TGF-beta1 production were due to a decrease in TGF-beta1 production per cell. In contrast, decreased total VEGF production was secondary to decreases in cellular proliferation, because the cellular production of VEGF by irradiated osteoblasts was moderately increased when VEGF production was corrected for cell number. Additionally, in contrast to control cells (i.e., nonirradiated), conditioned media obtained from irradiated osteoblasts failed to stimulate the proliferation of bovine aortic endothelial cells. Finally, transfection of control and irradiated cells with a replication-deficient TGF-beta1 adenovirus before irradiation resulted in an increase in cellular production of TGF-beta1 protein and VEGF. Interestingly, this intervention did not alter the effects of irradiation on cellular proliferation, which implies that alterations in TGF-beta1 expression do not underlie the deficiencies noted in cellular proliferation. The authors hypothesize that ionizing radiation-induced alterations in the cytokine profiles and differentiation states of osteoblasts may provide insights into the cellular mechanisms underlying osteoradionecrosis and impaired fracture healing.
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PMID:The effects of ionizing radiation on osteoblast-like cells in vitro. 1103 76

A new biocompatible glass, which is composed of CaO, P2O5, SiO2, and Al2O3 (abbreviated CPSA) and is characterized by higher elasticity than previous bioglass products, was molded into fibers with a diameter of 9 microm. With CPSA fibers, two geometrically different structures, balls and bundles (each 20 mg in weight), were prepared, combined with 2.2 microg of rhBMP-2 (a gift from Yamanouchi Co., Japan) and implanted subcutaneously into rats. The histology showed remarkably higher bone formation in the ball-CPSA/BMP at 2 and 4 weeks than in the bundle-CPSA/BMP. The ball-CPSA/BMP showed 10 times higher alkaline phosphatase (ALP) activity at the second week and 5 times higher osteocalcin content at the fourth week than the bundle-CSPA/BMP. Vascular development in the implants was evaluated by mRNA expression of Flt-1 and KDR, two receptors for vascular endothelial growth factor (VEGF). Both receptors showed higher expression in the case of the ball, while they were not detected in the bundle. It is concluded that the BMP-induced bone formation depends highly upon the porous vasculature-inducing geometry of the matrix, which can be constructed with the new CPSA fibers.
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PMID:Geometric effect of matrix upon cell differentiation: BMP-induced osteogenesis using a new bioglass with a feasible structure. 1113 71


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