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)

Bone morphogenetic protein-2 (BMP-2), a member of transforming growth factor-beta superfamily, inhibits the terminal differentiation of C2C12 myoblasts and changes their differentiation pathway into cells expressing osteoblast phenotypes such as alkaline phosphatase (ALP) activity and osteocalcin production (Katagiri et al., 1994, J. Cell Biol. 127, 1755-1766). Two type I receptors for BMP-2 (BMPR-IA and BMPR-IB) have been cloned, but the role of the respective receptors in signal transduction is not clear. In the present study, we examined the signal transduction of BMP-2 in C2C12 cells using constitutively activated mutant BMPR-IA and BMPR-IB. C2C12 cells expressed BMPR-IA and BMPR-II mRNAs, but not BMPR-IB mRNA at detectable levels in Northern blotting. When mutated BMPR-IA and BMPR-IB were transiently transfected into C2C12 cells, both BMPR-IA and BMPR-IB similarly induced ALP activity in the absence of BMP-2. We also established subclonal cell lines of C2C12 cells by stably transfecting mutated BMPR-IB. When the mutated BMPR-IB-transfected cells were cultured in medium with low serum (differentiation medium) without BMP-2, the cells differentiated into ALP-positive mononuclear cells and not into myosin heavy chain-positive myotubes. These mutated BMPR-IB-transfected cells expressed ALP activity and osteocalcin mRNA in a time-dependent manner, but neither muscle creatine kinase nor myogenin mRNAs. These results indicate that the mutated BMP-2 type I receptors can constitutively transduce BMP-2 signals in the absence of the ligand in C2C12 cells.
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PMID:Constitutively active BMP type I receptors transduce BMP-2 signals without the ligand in C2C12 myoblasts. 929 60

To examine the role of bone morphogenetic protein (BMP) signaling in chondrocytes during endochondral ossification, the dominant negative (DN) forms of BMP receptors were introduced into immature and mature chondrocytes isolated from lower and upper portions of chick embryo sternum, respectively. We found that control sternal chondrocyte populations expressed type IA, IB, and II BMP receptors as well as BMP-4 and -7. Expression of a DN-type II BMP receptor (termed DN-BMPR-II) in immature lower sternal (LS) chondrocytes led to a loss of differentiated functions; compared with control cells, the DN-BMPR- II-expressing LS chondrocytes proliferated more rapidly, acquired a fibroblastic morphology, showed little expression of type II collagen and aggrecan genes, and upregulated type I collagen gene expression. Expression of DN-BMPR-II in mature hypertrophic upper sternal (US) chondrocytes caused similar effects. In addition, the DN-BMPR-II-expressing US cells exhibited little alkaline phosphatase activity and type X collagen gene expression, while the control US cells produced both alkaline phosphatase and type X collagen. Both DN-BMPR-II-expressing US and LS chondrocytes failed to respond to treatment with BMP-2 . When we examined the effects of DN forms of types IA and IB BMP receptors, we found that DN-BMPR-IA had little effect, while DN-BMPR-IB had similar but weaker effects compared with those of DN-BMPR-II. We conclude that BMP signaling, particularly that mediated by the type II BMP receptor, is required for maintenance of the differentiated phenotype, control of cell proliferation, and expression of hypertrophic phenotype.
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PMID:Bone morphogenetic protein signaling is required for maintenance of differentiated phenotype, control of proliferation, and hypertrophy in chondrocytes. 944 16

Cartilage-derived morphogenetic proteins-1 and -2 (CDMP-1 and CDMP-2) are members of the bone morphogenetic protein (BMP) family, which play important roles in embryonic skeletal development. We studied the biological activities of recombinant CDMP-1 and CDMP-2 in chondrogenic and osteogenic differentiation and investigated their binding properties to type I and type II serine/threonine kinase receptors. In vivo, CDMP-1 and CDMP-2 were capable of inducing dose-dependently de novo cartilage and bone formation in an ectopic implantation assay. In vitro studies using primary chondrocyte cultures showed that both CDMP-1 and CDMP-2 stimulated equally de novo synthesis of proteoglycan aggrecan in a concentration-dependent manner. This activity was equipotent when compared with osteogenic protein-1 (OP-1). In contrast, CDMPs were less stimulatory than OP-1 in osteogenic differentiation as evaluated by alkaline phosphatase activity and expression levels of bone markers in ATDC5, ROB-C26, and MC3T3-E1 cells. CDMP-2 was the least osteogenic in these assays. Receptor binding studies of CDMP-1 and CDMP-2 revealed that both have affinity for the BMP receptor type IB (BMPR-IB) and BMPR-II, and weakly for BMPR-IA. Moreover, using a promoter/reporter construct, transcriptional activation signal was transduced by BMPR-IB in the presence of BMPR-II upon CDMP-1 and CDMP-2 binding. Our data show that distinct members of the BMP family differentially regulate the progression in the osteogenic lineage, and this may be due to their selective affinity for specific receptor complexes.
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PMID:Cartilage-derived morphogenetic proteins and osteogenic protein-1 differentially regulate osteogenesis. 952 38

Osteoblasts enzymatically isolated from newborn rat calvariae show various phenotypes including formation of mineralized bone nodules in culture. We investigated the temporal changes in osteoblast phenotype in these cells up to day 20 in culture. These cells formed unmineralized nodules by day 5. Mineralization was observed at the center of nodules by day 10, and nodules became larger on day 15. The nodules were surrounded by numerous alkaline phosphatase (ALP)-positive cells. ALP activity gradually increased by day 20. Parathyroid hormone (PTH) responsiveness increased with time in culture. Osteoblasts produced no osteocalcin by day 10, but its synthesis was detected from day 15. These cells expressed substantial levels of ALP and PTH/PTHrP receptor mRNAs as early as day 5 in culture, but very weak expression of osteocalcin mRNA on day 5. The levels of expression of these transcripts increased with time in culture. In situ hybridization demonstrated that PTH/PTHrP receptor and osteocalcin mRNAs were strongly expressed in nodules, but the former appeared much earlier than the latter. BMP-2 and BMP-4 mRNAs also appeared in the cells forming nodules. Immunohistochemical analysis demonstrated that cells expressing either BMP-2/4 or their receptors (BMPR-IA, BMPR-IB, and BMPR-II) preferentially appeared in nodules. These observations suggested that BMPs play an important role in the formation of mineralized bone nodules in an autocrine and/or paracrine fashion in these cells. The present study confirmed that osteoblasts enzymatically isolated from newborn rat calvariae are a useful tool for studying the differentiation process of osteoblasts.
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PMID:Changes in osteoblast phenotype during differentiation of enzymatically isolated rat calvaria cells. 960 Jul 81

Osteogenic protein-1 (OP-1 or bone morphogenetic protein-7 [BMP-7]) stimulates osteoblast differentiation in vitro and induces bone formation in vivo. BMPs exert their effects through complex formation with a heterodimeric receptor composed of a type I and a type II polypeptide. In the present study, mRNAs for three BMP subtype I receptors (ActR-I, BMPR-IA, and BMPR-IB) and one BMPR-II receptor were detected by Northern analysis in two human osteosarcoma cell lines (SaOS-2 and TE85) and in the primary cultures of fetal rat calvaria (FRC) cells. OP-1 affected the steady-state mRNA levels of these receptors differently among these cell types. To study the role of each receptor type in OP-1 action in FRC cells, receptor synthesis was inhibited by antisense oligonucleotides. Inhibition of receptor synthesis was confirmed by immunoprecipitation of radiolabeled cellular proteins with specific antibodies. The osteogenic action of OP-1 was measured by alkaline phosphatase (ALP) activity and mineralized bone nodule formation in FRC cells. Results showed that inhibition of synthesis of a single subtype I receptor alone did not affect significantly the OP-1-stimulated ALP activity. Inhibition of BMPR-II synthesis reduced the OP-1-stimulated ALP activity by about 50%. Inhibition of synthesis of any one of the type I receptor plus the BMPR-II receptor did not reduce the OP-1-stimulated ALP activity significantly beyond that observed by inhibition of BMPR-II alone. Under these conditions, nodule formation was affected similarly, thus supporting the observations made with the ALP measurements. The present results suggest that the ActR-I, BMPR-IA, and BMPR-IB receptors and the BMPR-II receptor are expressed and functional for OP-1 in FRC cells and that regulation of synthesis of these receptors may be a mechanism by which a specific cell type responds to OP-1. The turnover rate of these receptor proteins might be relatively long and another type II receptor(s) for OP-1 might be functional in FRC cells.
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PMID:Inhibition of BMP receptor synthesis by antisense oligonucleotides attenuates OP-1 action in primary cultures of fetal rat calvaria cells. 984 5

Bone morphogenetic protein (BMP)-6 is a member of the transforming growth factor (TGF)-(&bgr;) superfamily, and is most similar to BMP-5, osteogenic protein (OP)-1/BMP-7, and OP-2/BMP-8. In the present study, we characterized the endogenous BMP-6 signaling pathway during osteoblast differentiation. BMP-6 strongly induced alkaline phosphatase (ALP) activity in cells of osteoblast lineage, including C2C12 cells, MC3T3-E1 cells, and ROB-C26 cells. The profile of binding of BMP-6 to type I and type II receptors was similar to that of OP-1/BMP-7 in C2C12 cells and MC3T3-E1 cells; BMP-6 strongly bound to activin receptor-like kinase (ALK)-2 (also termed ActR-I), together with type II receptors, i.e. BMP type II receptor (BMPR-II) and activin type II receptor (ActR-II). In addition, BMP-6 weakly bound to BMPR-IA (ALK-3), to which BMP-2 also bound. In contrast, binding of BMP-6 to BMPR-IB (ALK-6), and less efficiently to ALK-2 and BMPR-IA, together with BMPR-II was detected in ROB-C26 cells. Intracellular signalling was further studied using C2C12 and MC3T3-E1 cells. Among the receptor-regulated Smads activated by BMP receptors, BMP-6 strongly induced phosphorylation and nuclear accumulation of Smad5, and less efficiently those of Smad1. However, Smad8 was constitutively phosphorylated, and no further phosphorylation or nuclear accumulation of Smad8 by BMP-6 was observed. These findings indicate that in the process of differentiation to osteoblasts, BMP-6 binds to ALK-2 as well as other type I receptors, and transduces signals mainly through Smad5 and possibly through Smad1.
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PMID:Characterization of bone morphogenetic protein-6 signaling pathways in osteoblast differentiation. 1050

Osteogenic Protein-1 (OP-1, BMP-7), a member of the bone morphogenetic protein family, stimulates synthesis of biochemical markers characteristic of the osteoblastic and chondrocytic phenotypes and induces new bone formation. Interleukin-6 (IL-6), a cytokine produced by a wide variety of cells, appears to interact with other factors producing different biological effects. In the present study, we showed that OP-1 action in fetal rat calvaria (FRC) cells was enhanced by the combination of IL-6 and the soluble receptor IL-6sR. OP-1 alone induced alkaline phosphatase (AP) activity by 4- to 5-fold above the control. Exogenous IL-6 soluble receptor (IL-6sR) synergistically stimulated the OP-1-induced AP activity and mineralized bone nodule formation by an additional 3-fold. The stimulation was IL-6sR concentration-dependent. The combination of IL-6 and IL-6sR synergistically stimulated OP-1 action by an additional 6- to 7-fold. BMPR-II receptor mRNA expression in FRC cells treated with OP-1 and IL-6 plus IL-6sR was stimulated further, while BMPR-IA, -IB, and ActR-I expressions were not affected. The intracellular signaling molecules Smad2 and Smad5 mRNA expressions were not changed under these conditions. The expression of selected BMP family members (BMP-3, -4, and -6) was altered in FRC cells treated with OP-1 in combination with IL-6 and IL-6sR. The combination of IL-6 and IL-6sR reduced the OP-1-stimulated BMP-3 mRNA levels and enhanced the suppressive effect of OP-1 on BMP-4 and -6 mRNA expressions. In conclusion, the present results demonstrate that exogenous IL-6 and IL-6sR synergistically stimulate OP-1 action in primary cultures of rat osteoblastic cells. One possible mechanism of synergy involves differential regulation of the effects of OP-1 on the expression of the type II BMP receptor and several other BMPs.
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PMID:Osteogenic protein-1 and interleukin-6 with its soluble receptor synergistically stimulate rat osteoblastic cell differentiation. 1185 48

In this study, we examine the role of bone morphogenetic protein (BMP) signaling during differentiation of the murine preosteoblastic KS483 cell line, which formed alkaline phosphatase (ALP)-positive and mineralized nodules during a 3 week culture period. Semiquantitative reverse transcription-polymerase chain reaction (RT-PCR) demonstrated the presence of various BMPs (BMP-2, -3, -4, -6, -7, and -8A and -8B), BMP type I and II receptors (ALK2, ALK3, ALK4, BMPR-II, and ActR-IIA and -IIB), BMP antagonists (DAN, gremlin, chordin, cerberus, noggin, and tsg), and Smads 1-8. mRNA expression of these genes did not change during differentiation, except for BMP-3, BMP-8a, and noggin. BMP-3 increased gradually, particularly in the matrix formation phase; BMP-8a was induced from the onset of matrix maturation and mineralization, in parallel to the expression of osteocalcin; and noggin tended to decline during the mineralization phase. Treatment of KS483 cells with the BMP antagonists noggin or soluble truncated BMPR-IA, either continuously or during distinct periods of osteoblast differentiation; that is, matrix formation or matrix maturation and mineralization phase, decreased ALP-positive and mineralized nodule area independent of the phase of osteoblast differentiation. Notably, the antagonists inhibited mineralization of already existing nodules. Similarly, BMP-4 stimulated differentiation not only at the beginning of the culture period, but also at late stages of differentiation. These data indicate that autocrine BMP signaling is involved in KS483 osteoblastic differentiation not only during the early phase of differentiation, but also during matrix maturation and mineralization. The different expression patterns of components of BMP signaling in the KS483 cells suggest distinct functions of individual BMPs during osteoblast differentiation. In summary, our data suggest that BMP activity is required not only for initiation of osteoblast differentiation and further development of early osteoblasts, but is also involved in late-stage osteoblast differentiation and matrix mineralization.
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PMID:Differentiation of murine preosteoblastic KS483 cells depends on autocrine bone morphogenetic protein signaling during all phases of osteoblast formation. 1253 59

Osteogenic protein-1 (OP-1, also called BMP-7), a member of the BMP family and the TGF-beta superfamily, induces formation of new bone and cartilage, but also regulates a wide array of processes. In the present study, the expression of several characteristic biochemical markers of ligaments, such as Six1, Scleraxis, aggrecan, and type I collagen in primary cultures of adult rat medial collateral ligament (MCL) cells was determined. The effects of OP-1 on cell proliferation and on gene expression were subsequently examined. OP-1 stimulated cell proliferation, alkaline phosphatase (AP) activity, and the steady-state mRNA levels of the transcription factor Runx2/Cbfa1 in a dose- and time-dependent manner. The mRNA levels of type I collagen only increased slightly, but the activity of the cloned collagen promoter increased by 2-fold in transiently transfected MCL cells. OP-1 also stimulated aggrecan mRNA expression. The mRNA levels of Six1 and Scleraxis were not detectably altered by OP-1. In control cultures, the steady-state mRNA levels of ActR-I, BMPR-IA, BMPR-IB, and BMPR-II increased as a function of time in culture. The mRNA levels of BMP-1 and -4 increased significantly after 12 days, but those of BMP-2 and -6 did not change. The GDF-1, -3, -5, -6, and -8 mRNA levels in the control cultures also increased as a function of time. OP-1 treatment stimulated mRNA expression of BMPR-IA and BMPR-II, but had little effect on ActR-I and BMPR-IB mRNA expression. OP-1 lowered the BMP-1, -2, and -6 mRNA levels without changing the BMP-4 mRNA level. OP-1 treatment also reduced the mRNA levels of GDFs detected. In summary, the present study demonstrated that OP-1 stimulated cell proliferation and mRNA expression of several biochemical markers in this ligament cell culture model and established the spatial and temporal appearance of several members of the TGF-beta superfamily.
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PMID:Effects of osteogenic protein-1 (OP-1, BMP-7) on gene expression in cultured medial collateral ligament cells. 1458 33

Cartilage-derived morphogenetic protein-1, -2, and -3 (CDMP-1, -2, and -3) are members of the bone morphogenetic protein (BMP) family and have been shown to exhibit a variety of biological activities. In the present study, effects of these CDMPs on the temporal and spatial expression of genes in the pluripotent mesenchymal cell line C2C12 were examined. Cells cultured in the presence of CDMPs lost the characteristic elongated shape of myoblasts. At the molecular level, CDMP treatment did not change the mRNA expression of MyoD, aggrecan, Six1, and tendin. Scleraxis mRNA level was reduced by CDMP treatment. CDMP-1 and -3, but not CDMP-2, stimulated expression of osteogenic markers, such as alkaline phosphatase (AP), osteocalcin (OC), BSP, and type I collagen, in a dose- and time-dependent manner. With few exceptions, the three CDMPs changed, with different potencies, the expression profile of different members of the BMP family in a similar temporal pattern. Except at the late phase of treatment, CDMP treatment did not change the expression of ActR-IA, BMPR-IA, BMPR-IB, BMPR-II, and ALK-7 mRNAs. Based on the current data, the CDMPs appear to be able to stimulate the C2C12 cells to differentiate into the osteoblast pathway.
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PMID:Cartilage-derived morphogenetic proteins induce osteogenic gene expression in the C2C12 mesenchymal cell line. 1575 75


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