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)

The bone morphogenetic protein (BMP)-2 is a potent osteoinductive signal, inducing bone formation in vivo and osteoblast differentiation from non-osseous cells in vitro. The runt domain-related protein Cbfa1/PEBP2alphaA/AML-3 is a critical component of bone formation in vivo and transcriptional regulator of osteoblast differentiation. To investigate the relationship between the extracellular BMP-2 signal, Cbfa1, and osteogenesis, we examined expression of Cbfa1 and osteoblastic genes during the BMP-2 induced osteogenic transdifferentiation of the myoblastic cell line C2C12. BMP-2 treatment completely blocked myotube formation and transiently induced expression of Cbfa1 and the bone-related homeodomain protein Msx-2 concomitant with loss of the myoblast phenotype. While induction of collagen type I and alkaline phosphatase (AP) expression coincided with Cbfa1 expression, Cbfa1 mRNA was strikingly downregulated at the onset of expression of osteopontin (OPN) and osteocalcin (OCN) genes, reflecting the mature osteoblast phenotype. TGF-beta1 treatment effectively suppressed myogenesis and induced Cbfa1 expression but was insufficient to support osteoblast differentiation reflected by the absence of ALP, OPN, and OCN. We addressed whether induction of Cbfa1 in response to BMP-2 results in the transcriptional activation of the OC promoter which contains three enhancer Cbfa1 elements. Transfection studies show BMP-2 suppresses OC promoter activity in C2C12, but not in osteoblastic ROS 17/2.8 cells. Maximal suppression of OC promoter activity in response to BMP-2 requires sequences in the proximal promoter (up to nt -365) and may occur independent of the three Cbfa sites. Taken together, our results demonstrate a dissociation of Cbfa1 expression from development of the osteoblast phenotype. Our findings suggest that Cbfal may function transiently to divert a committed myoblast to a potentially osteogenic cell. However, other factors induced by BMP-2 appear to be necessary for complete expression of the osteoblast phenotype.
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PMID:Transient upregulation of CBFA1 in response to bone morphogenetic protein-2 and transforming growth factor beta1 in C2C12 myogenic cells coincides with suppression of the myogenic phenotype but is not sufficient for osteoblast differentiation. 1008 30

TAK-778 [(2R,4S)-(-)-N-(4-Diethoxyphosphorylmethylphenyl)-1,2,4,5-tetrahydro-4-methyl-7,8-methylenedioxy-5-oxo-3-benzothiepin-2-carboxamide: mw 505.52], a novel compound promoting osteoblast differentiation, promotes osteogenesis in vitro and enhances bone formation during skeletal repair in vivo. In this study, we further evaluated the effects of TAK-778 on the differentiation of cultured bone marrow stromal cells into osteoblasts in the presence of dexamethasone, paying particular attention to the expression of transcription factors involved in regulating osteoblast differentiation. Treatment of TAK-778 (10(-7)-10(-5) M) for 4 h resulted in an increase in the mRNA expression of Msx2, but not Cbfa1 or Dlx5. This transcriptional alteration preceded the changes in other markers related to the osteoblast phenotype, such as alkaline phosphatase and osteocalcin mRNA. The transfection of Msx2-antisense in the cells caused a significant reduction in the levels of alkaline phosphatase mRNA expression induced by TAK-778. These results suggest that TAK-778 promotes osteoblast differentiation partly through the expression of Msx2, a homeobox-related gene.
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PMID:Enhancement of osteogenesis in vitro by a novel osteoblast differentiation-promoting compound, TAK-778, partly through the expression of Msx2. 1222 24

Recent studies have suggested that regionally differentiated dura mater regulates murine cranial suture fate by providing growth factors to the osteoblasts in the overlying suture complex. To determine if regionally differentiated dura mater is capable of effecting changes in osteoblast gene expression, an in vitro coculture system was established in which osteoblast-enriched cell cultures derived from neonatal rat calvaria were grown in serum-free media in the presence of dural cells derived from posterior frontal (PF) or sagittal (SAG) dural tissues, recapitulating the in situ relation between the underlying dura mater and the osteoblasts in the overlying cranial suture. In this study, the changes in osteoblast gene expression induced by signaling from regional dura mater were examined by analyzing total cellular RNA isolated from osteoblasts cocultured with PF or SAG dural cells. The expression of extracellular matrix molecules (alkaline phosphatase, bone sialoprotein, osteopontin, and osteocalcin) and the transcription factor Msx2 was assessed. Consistent with previous data, the findings demonstrate that osteoblasts cocultured with dural cells undergo changes in gene expression indicative of a more differentiated osteoblast. Additionally, the data suggest that regionally differentiated dura mater isolated from the PF suture enhances the expression of osteogenic genes to a greater extent than SAG suture-derived dural cells. These data support an osteoinductive role for suture-derived dural cells in vitro that may have implications for suture biology in vivo.
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PMID:Regional dura mater differentially regulates osteoblast gene expression. 1282 8

In the aorta, diabetes activates an osteogenic program that includes expression of bone morphogenetic protein-2 (BMP2) and the osteoblast homeoprotein Msx2. To evaluate BMP2-Msx2 signaling in vascular calcification, we studied primary aortic myofibroblasts. These cells express vascular smooth muscle cell (VSMC) markers, respond to BMP2 by up-regulating Msx2, and undergo osteogenic differentiation with BMP2 treatment or transduction with a virus encoding Msx2. The osteoblast factor osterix (Osx) is up-regulated 10-fold by Msx2, but Runx2 mRNA is unchanged; the early osteoblast marker alkaline phosphatase increases 50-fold with mineralized nodule formation enhanced 30-fold. Adipocyte markers are concomitantly suppressed. To better understand Msx2 actions on osteogenesis versus adipogenesis, mechanistic studies were extended to C3H10T1/2 mesenchymal cells. Msx2 enhances osteogenic differentiation in synergy with BMP2. Osteogenic actions depend upon intrinsic Msx2 DNA binding; the gain-of-function variant Msx2(P148H) directs enhanced mineralization, whereas the binding-deficient variant Msx2(T147A) is inactive. Adipogenesis (lipid accumulation, Pparg expression) is inhibited by Msx2. By contrast, suppression of adipogenesis does not require Msx2 DNA binding; inhibition occurs in part via protein-protein interactions with C/EBPalpha that control Pparg transcription. Thus, Msx2 regulates osteogenic versus adipogenic differentiation of aortic myofibroblasts. Myofibroblasts capable of both fates can be diverted to the osteogenic lineage by BMP2-Msx2 signaling and contribute to vascular calcification.
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PMID:MSX2 promotes osteogenesis and suppresses adipogenic differentiation of multipotent mesenchymal progenitors. 1292 29

Bone morphogenetic proteins (BMPs) promote the differentiation of osteoprogenitor cells, and also induce osteogenesis in bone marrow stromal cells (MSC) from rats and mice. However, compared to results with animal models, BMPs are relatively inefficient in inducing human MSC to undergo osteogenesis, and are much less effective in promoting bone formation in human clinical trials. Previous studies indicated that, while human MSC respond to dexamethasone with elevated levels of the osteoblast marker alkaline phosphatase, most isolates of human MSC fail to show alkaline phosphatase induction in response to BMP-2, BMP-4, or BMP-7. Several other genes known to be induced by BMPs are appropriately regulated; thus, human MSC are capable of some BMP-activated signaling. Analysis of the BMP receptors ALK-3 and ALK-6 indicated that, although ALK-6 mRNA was not expressed in human MSC, overexpressing a constitutively active ALK-6 receptor did not induce elevated alkaline phosphatase. Real-time RT-PCR was used to investigate expression of several osteoblast-related transcription factors in MSC after 6 days' exposure to BMP2 or dexamethasone. Msx-2, a transcription factor that has been reported to inhibit differentiation of osteoprogenitor cells, showed 10-fold elevation in BMP-2-treated human MSC, but not in BMP-2-treated rat MSC. Overexpression of Msx-2 in human and rat MSC, however, did not alter alkaline phosphatase levels, which suggests that absence of BMP-stimulated alkaline phosphatase was not caused by the BMP-2-induced increase in Msx-2. Although Runx2 isoforms have been implicated in control of osteoblast differentiation, levels of this transcription factor were unaffected by BMP treatment. Expression of the FKHR transcription factor, which has been reported to regulate alkaline phosphatase transcription in mouse cells, showed a modest increase in response to BMP-2, but a much greater increase in dexamethasone-treated cells. We propose that BMP regulation of the bone/liver/kidney alkaline phosphatase gene is indirect, requiring expression of new transcription factor(s) that behave differently in rodent and human MSC.
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PMID:Different effects of BMP-2 on marrow stromal cells from human and rat bone. 1474 40

Sclerosteosis, a skeletal disorder characterized by high bone mass due to increased osteoblast activity, is caused by loss of the SOST gene product, sclerostin. The localization in bone and the mechanism of action of sclerostin are not yet known, but it has been hypothesized that it may act as a bone morphogenetic protein (BMP) antagonist. We show here that SOST/sclerostin is expressed exclusively by osteocytes in mouse and human bone and inhibits the differentiation and mineralization of murine preosteoblastic cells (KS483). Although sclerostin shares some of the actions of the BMP antagonist noggin, we show here that it also has actions distinctly different from it. In contrast to noggin, sclerostin did not inhibit basal alkaline phosphatase (ALP) activity in KS483 cells, nor did it antagonize BMP-stimulated ALP activity in mouse C2C12 cells. In addition, sclerostin had no effect on BMP-stimulated Smad phosphorylation and direct transcriptional activation of MSX-2 and BMP response element reporter constructs in KS483 cells. Its unique localization and action on osteoblasts suggest that sclerostin may be the previously proposed osteocyte-derived factor that is transported to osteoblasts at the bone surface and inhibits bone formation.
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PMID:Sclerostin is an osteocyte-expressed negative regulator of bone formation, but not a classical BMP antagonist. 1502 46

Mice deficient in the Msx2 gene manifest defects in skull ossification and a marked reduction in bone formation associated with decreases in osteoblast numbers, thus suggesting that Msx2 is involved in bone formation. However, the precise role of Msx2 during osteoblast differentiation is not fully understood. In the present study, we investigated the role of Msx2 in the regulation of osteoblast differentiation in the multipotent mesenchymal cell lines C3H10T1/2 and C2C12 and in murine primary osteoblasts. Introduction of Msx2 induced alkaline phosphatase activity in C3H10T1/2 and C2C12 cells and promoted the calcification of murine primary osteoblasts. This effect of Msx2 was also observed in mesenchymal cells isolated from Runx2-deficient mice. Interestingly the expression of Msx2 was induced by bone morphogenetic protein 2 treatment in Runx2-deficient mesenchymal cells. In contrast, Msx2 diminished peroxisome proliferator-activated receptor gamma (PPARgamma) expression and adipogenesis of the preadipocytic cell line 3T3-F442A. Moreover Msx2 inhibited the transcriptional activity of PPARgamma, CCAAT/enhancer-binding protein beta (C/EBPbeta), and C/EBPdelta and blocked adipocyte differentiation of mesenchymal cells induced by overexpression of PPARgamma, C/EBPalpha, C/EBPbeta, or C/EBPdelta. These data indicate that Msx2 promotes osteoblast differentiation independently of Runx2 and negatively regulates adipocyte differentiation through inhibition of PPARgamma and the C/EBP family.
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PMID:Reciprocal roles of MSX2 in regulation of osteoblast and adipocyte differentiation. 1517 25

Alx4 and Msx2 encode homeodomain-containing transcription factors that show a clear functional overlap. In both mice and humans, loss of function of either gene is associated with ossification defects of the skull vault, although the major effect is on the frontal bones in mice and the parietal bones in humans. This study was undertaken to discover whether Alx4 and Msx2 show a genetic interaction in skull vault ossification, and to test the hypothesis that they interact with the pathway that includes the Fgfr genes, Twist1 and Runx2. We generated Alx4(+/-)/Msx2(+/-) double heterozygous mutant mice, interbred them to produce compound genotypes and analysed the genotype-phenotype relationships. Loss of an increasing number of alleles correlated with an incremental exacerbation of the skull vault defect; loss of Alx4 function had a marginally greater effect than loss of Msx2 and also affected skull thickness. In situ hybridization showed that Alx4 and Msx2 are expressed in the cranial skeletogenic mesenchyme and in the growing calvarial bones. Studies of the coronal suture region at embryonic day (E)16.5 revealed that Alx4 expression was decreased, but not abolished, in Msx2(-/-) mutants, and vice versa; expression of Fgfr2 and Fgfr1, but not Twist1, was reduced in both mutants at the same stage. Runx2 expression was unaffected in the coronal suture; in contrast, expression of the downstream ossification marker Spp1 was delayed. Double homozygous pups showed substantial reduction of alkaline phosphatase expression throughout the mineralized skull vault; they died at birth due to defects of the heart, lungs and diaphragm not previously associated with Alx4 or Msx2. Our observations suggest that Alx4 and Msx2 are partially functionally redundant, acting within a network of transcription factors and signalling events that regulate the rate of osteogenic proliferation and differentiation at a stage after the commitment of mesenchymal stem cells to osteogenesis.
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PMID:Alx4 and Msx2 play phenotypically similar and additive roles in skull vault differentiation. 1519 90

Osteoblast differentiation is controlled by multiple transcription factors, Runx2, AJ18, Osterix, Dlx5 and Msx2. The mechanisms of regulation of AJ18 mRNA expression by the transforming growth factor beta (TGF-beta) superfamily remain poorly understood. However, it is known that BMP-2 induces differentiation of C26 cells into more mature osteoblastic cells. The present study, using Northern blot and real-time reverse transcription polymerase chain reaction analyses, investigated the effects of bone morphogenetic protein-2 (BMP-2) and TGF-beta1 on mRNA expression of AJ18 and Runx2 in a clonal osteoblast precursor cell line ROB-C26 (C26) cultured for 3, 6 or 9 days in the presence or absence of BMP-2. Although mRNA expression of Osterix and bone sialoprotein (BSP) was undetectable in the C26 culture, BMP-2 induced Osterix expression on days 3-9, but not BSP expression. BMP-2 also stimulated significantly Dlx5 expression on days 3-9, Msx2 and matrix Gla protein expressions on days 3 and 6, Runx2, alkaline phosphatase and osteocalcin expressions on days 6 and 9 in the culture. Furthermore, BMP-2 increased significantly Smad5 mRNA in the culture on day 3, indicating BMP-2 involvement in the regulation of Smad5 mRNA expression. In contrast, the inhibitory effects of BMP-2 on AJ18 mRNA expression were significant on days 3-9, indicating that a decrease in AJ18 mRNA expression is essential for the increased osteoblastic differentiation. Furthermore, TGF-beta1 (0, 0.1, 1.0 and 5.0 ng/ml) treatment of C26 cells cultured for 6 days in the presence or absence of BMP-2 for 24h stimulated mRNA levels of AJ18 and Runx2, maximal stimulation occurring principally at 1.0 ng/ml. These observations indicate that the expression of AJ18 and Runx2 mRNAs in C26 cells is under the control of BMP-2 and TGF-beta1, which exert different effects on AJ18 mRNA expression, but are potent stimulators of Runx2 mRNA expression during osteoblast differentiation.
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PMID:Effects of bone morphogenetic protein-2 and transforming growth factor beta1 on gene expression of transcription factors, AJ18 and Runx2 in cultured osteoblastic cells. 1532 53

In diabetic LDLR-/- mice, an ectopic BMP2-Msx2 gene regulatory program is upregulated in association with vascular calcification. We verified the procalcific actions of aortic Msx2 expression in vivo. CMV-Msx2 transgenic (CMV-Msx2Tg(+)) mice expressed 3-fold higher levels of aortic Msx2 than nontransgenic littermates. On high-fat diets, CMV-Msx2Tg(+) mice exhibited marked cardiovascular calcification involving aortic and coronary tunica media. This corresponded to regions of Msx2 immunoreactivity in adjacent adventitial myofibroblasts, suggesting a potential paracrine osteogenic signal. To better understand Msx2-regulated calcification, we studied actions in 10T1/2 cells. We found that conditioned media from Msx2-transduced 10T1/2 cells (Msx2-CM) is both pro-osteogenic and adipostatic; these features are characteristic of Wnt signaling. Msx2-CM stimulated Wnt-dependent TCF/LEF transcription, and Msx2-transduced cells exhibited increased nuclear beta-catenin localization with concomitant alkaline phosphatase induction. Msx2 upregulated Wnt3a and Wnt7a but downregulated expression of the canonical inhibitor Dkk1. Dkk1 treatment reversed osteogenic and adipostatic actions of Msx2. Teriparatide, a PTH1R agonist that inhibits murine vascular calcification, suppressed vascular BMP2-Msx2-Wnt signaling. Analyses of CMV-Msx2Tg(+) mice confirmed that Msx2 suppresses aortic Dkk1 and upregulates vascular Wnts; moreover, TOPGAL(+) (Wnt reporter); CMV-Msx2Tg(+) mice exhibited augmented aortic LacZ expression. Thus, Msx2-expressing cells elaborated an osteogenic milieu that promotes vascular calcification in part via paracrine Wnt signals.
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PMID:Msx2 promotes cardiovascular calcification by activating paracrine Wnt signals. 1584 Dec 9


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