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)

Fibroblast growth factors (FGF) play a critical role in bone growth and development affecting both chondrogenesis and osteogenesis. During the process of intramembranous ossification, which leads to the formation of the flat bones of the skull, unregulated FGF signaling can produce premature suture closure or craniosynostosis and other craniofacial deformities. Indeed, many human craniosynostosis disorders have been linked to activating mutations in FGF receptors (FGFR) 1 and 2, but the precise effects of FGF on the proliferation, maturation and differentiation of the target osteoblastic cells are still unclear. In this report, we studied the effects of FGF treatment on primary murine calvarial osteoblast, and on OB1, a newly established osteoblastic cell line. We show that FGF signaling has a dual effect on osteoblast proliferation and differentiation. FGFs activate the endogenous FGFRs leading to the formation of a Grb2/FRS2/Shp2 complex and activation of MAP kinase. However, immature osteoblasts respond to FGF treatment with increased proliferation, whereas in differentiating cells FGF does not induce DNA synthesis but causes apoptosis. When either primary or OB1 osteoblasts are induced to differentiate, FGF signaling inhibits expression of alkaline phosphatase, and blocks mineralization. To study the effect of craniosynostosis-linked mutations in osteoblasts, we introduced FGFR2 carrying either the C342Y (Crouzon syndrome) or the S252W (Apert syndrome) mutation in OB1 cells. Both mutations inhibited differentiation, while dramatically inducing apoptosis. Furthermore, we could also show that overexpression of FGF2 in transgenic mice leads to increased apoptosis in their calvaria. These data provide the first biochemical analysis of FGF signaling in osteoblasts, and show that FGF can act as a cell death inducer with distinct effects in proliferating and differentiating osteoblasts.
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PMID:Signaling by fibroblast growth factors (FGF) and fibroblast growth factor receptor 2 (FGFR2)-activating mutations blocks mineralization and induces apoptosis in osteoblasts. 1085 Oct 26

Mutations in fibroblast growth factor receptor 2 (FGFR2), a transmembrane receptor expressed in suture mesenchyme, osteogenic fronts, and dura, have been implicated in the etiopathogenesis of craniosynostosis syndromes. The C278F- and P253R-FGFR2 mutations result in Crouzon and Apert syndromes, respectively. The dura mater plays a critical role in the formation and maintenance of cranial sutures. However, its role in syndromic craniosynostosis remains unclear. This study examines the influence of FGFR2 mutations in dural cells on osteoblast proliferation and differentiation. Primary cultures of dural cells and osteoblasts were established, and adenoviral-FGFR2 constructs were prepared by subcloning mutant (C278F and P253R) FGFR2 constructs into adenovirus. Dural cells were infected with adenovirus, and dural protein expression was confirmed by immunostaining. Infected dural cells were cocultured with osteoblasts using a transwell system for 7 days. Dural cells infected with null adenovirus served as the negative control. In separate cultures, osteoblasts were directly infected with the adenoviral-FGFR2 constructs. Osteoblast proliferation was analyzed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, and differentiation was analyzed by alkaline phosphatase assay, histochemical staining, and gene expression studies. Osteoblasts directly infected with the Crouzon (C278F-FGFR2) mutation demonstrated an increase in cell proliferation (P < 0.05). Osteoblasts directly infected with the Apert (P253R-FGFR2) mutation demonstrated an increase in alkaline phosphatase activity. In coculture experiments, osteoblasts cocultured with Crouzon-transformed dural cells demonstrated increased cell proliferation (P < 0.05), whereas osteoblasts cocultured with Apert-transformed dural cells showed an increase in alkaline phosphatase activity (P < 0.05). In addition, osteogenic gene expression (alkaline phosphatase, osteopontin, and runx2) were up-regulated in osteoblasts cocultured with Apert-expressing dural cells. These experiments suggest that FGFR2 mutations in dural cells alter normal dural signaling. Apert mutations promote osteodifferentiation, whereas Crouzon mutations result in enhanced cell proliferation. These mutations may induce craniosynostosis in part through the influence of mutation-induced constitutive signaling in the dura, with subsequent enhancement of dural-mediated osteogenesis.
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PMID:Dura in the pathogenesis of syndromic craniosynostosis: fibroblast growth factor receptor 2 mutations in dural cells promote osteogenic proliferation and differentiation of osteoblasts. 2048 51

We recently reported that cranial bones of Fgfr2(C342Y/+) craniosynostotic mice are diminished in density when compared to those of wild type mice, and that cranial bone cells isolated from the mutant mice exhibit inhibited late stage osteoblast differentiation. To provide further support for the idea that craniosynostosis-associated Fgfr mutations lead to cell autonomous defects in osteoblast differentiation and mineralized tissue formation, here we tested bone marrow stromal cells isolated from Fgfr2(C342Y/+) mice for their ability to differentiate into osteoblasts. Additionally, to determine if the low bone mass phenotype of Crouzon syndrome includes the appendicular skeleton, long bones were assessed by micro CT. Fgfr2(C342Y/+) cells showed increased osteoblastic gene expression during early osteoblastic differentiation but decreased expression of alkaline phosphatase mRNA and enzyme activity, and decreased mineralization during later stages of differentiation, when cultured under 2D in vitro conditions. Cells isolated from Fgfr2(C342Y/+) mice also formed less bone when allowed to differentiate in a 3D matrix in vivo. Cortical bone parameters were diminished in long bones of Fgfr2(C342Y/+) mice. These results demonstrate that marrow stromal cells of Fgfr2(C342Y/+) mice have an autonomous defect in osteoblast differentiation and bone mineralization, and that the Fgfr2(C342Y) mutation influences both the axial and appendicular skeletons.
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PMID:Craniosynostosis-associated Fgfr2(C342Y) mutant bone marrow stromal cells exhibit cell autonomous abnormalities in osteoblast differentiation and bone formation. 2376 37

The purposes of this study were to find a novel mutation of FGFR2 in Korean Crouzon syndrome patients and to identify the functional consequences of this mutation. The samples consisted of 16 Crouzon patients. Peripheral venous blood was collected from the patients. FGFR2 mutation screening was performed by direct PCR sequencing of all exons and part of the introns. Restriction fragment length polymorphism (RFLP) analysis was performed to confirm the novel mutation. For functional studies, we performed luciferase assay for Runx2 transcriptional activity, real-time PCR for the bone markers (osteocalcin and alkaline phosphatase), and Western blot for phosphorylated FGFR2 and ERK1/2-MAPK protein. Among 16 patients, 10 showed FGFR2 mutations that had already been reported elsewhere. A novel FGFR2 mutation associated with tyrosine kinase II (TK-II) domain, L617F, was found in one Crouzon syndrome patient by direct PCR sequencing. Presence of this mutation was confirmed using RFLP analysis. Runx2 transcriptional activity and expression of osteocalcin and alkaline phosphatase significantly increased in L617F-transfected cells compared to wild-type cells. FGFR2 autophosphorylation in L617F-transfected cells increased in 1% serum, but ERK1/2-MAPK protein was not activated. The FGFR2-L617F mutation associated with the TK domain is potentially related to premature suture closure in Crouzon syndrome patient.
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PMID:A novel FGFR2 mutation in tyrosine kinase II domain, L617F, in Crouzon syndrome. 2391 23

Fibroblast growth factor receptor 2 (FGFR2) signaling is critical for proper craniofacial development. A gain-of-function mutation in the 2c splice variant of the receptor's gene is associated with Crouzon syndrome, which is characterized by craniosynostosis, the premature fusion of one or more of the cranial vault sutures, leading to craniofacial maldevelopment. Insight into the molecular mechanism of craniosynostosis has identified the ERK-MAPK signaling cascade as a critical regulator of suture patency. The aim of this study is to investigate the role of FGFR2c-induced ERK-MAPK activation in the regulation of coronal suture development. Loss-of-function and gain-of-function Fgfr2c mutant mice have overlapping phenotypes, including coronal synostosis and craniofacial dysmorphia. In vivo analysis of coronal sutures in loss-of-function and gain-of-function models demonstrated fundamentally different pathogenesis underlying coronal suture synostosis. Calvarial osteoblasts from gain-of-function mice demonstrated enhanced osteoblastic function and maturation with concomitant increase in ERK-MAPK activation. In vitro inhibition with the ERK protein inhibitor U0126 mitigated ERK protein activation levels with a concomitant reduction in alkaline phosphatase activity. This study identifies FGFR2c-mediated ERK-MAPK signaling as a key mediator of craniofacial growth and coronal suture development. Furthermore, our results solve the apparent paradox between loss-of-function and gain-of-function FGFR2c mutants with respect to coronal suture synostosis.
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PMID:FGFR2c-mediated ERK-MAPK activity regulates coronal suture development. 2703 31

Craniosynostosis is the premature fusion of cranial bones. The goal of this study was to determine if delivery of recombinant tissue nonspecific alkaline phosphatase (TNAP) could prevent or diminish the severity of craniosynostosis in a C57BL/6 FGFR2C342Y/+ model of neonatal onset craniosynostosis or a BALB/c FGFR2C342Y/+ model of postnatal onset craniosynostosis. Mice were injected with a lentivirus encoding a mineral targeted form of TNAP immediately after birth. Cranial bone fusion as well as cranial bone volume, mineral content and density were assessed by micro CT. Craniofacial shape was measured with calipers. Alkaline phosphatase, alanine amino transferase (ALT) and aspartate amino transferase (AST) activity levels were measured in serum. Neonatal delivery of TNAP diminished craniosynostosis severity from 94% suture obliteration in vehicle treated mice to 67% suture obliteration in treated mice, p<0.02) and the incidence of malocclusion from 82.4% to 34.7% (p<0.03), with no effect on cranial bone in C57BL/6 FGFR2C342Y/+ mice. In contrast, treatment with TNAP increased cranial bone volume (p< 0.01), density (p< 0.01) and mineral content (p< 0.01) as compared to vehicle treated controls, but had no effect on craniosynostosis or malocclusion in BALB/c FGFR2C342Y/+ mice. These results indicate that postnatal recombinant TNAP enzyme therapy diminishes craniosynostosis severity in the C57BL/6 FGFR2C342Y/+ neonatal onset mouse model of Crouzon syndrome, and that effects of exogenous TNAP are genetic background dependent.
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PMID:Viral delivery of tissue nonspecific alkaline phosphatase diminishes craniosynostosis in one of two FGFR2C342Y/+ mouse models of Crouzon syndrome. 3247 62