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)

The in vitro culture of porcine bone marrow-derived mesenchymal stem cells (MSCs) was used for the investigation of adult stem cell biology. Isolated porcine MSCs possessed the ability to proliferate extensively in an antioxidants-rich medium containing 5% fetal bovine serum (FBS). Greater than 40 serial MSC passages and 100 cell population doublings have been recorded for some MSC batches. Early and late passage MSCs were defined here as those cultures receiving less than 5 trypsin passages and more than 15 trypsin passages, respectively. Consistent with their robust ability to proliferate, both the early and late passage MSCs expressed the cell-cycle promoting enzyme p34cdc2 kinase. Late MSCs, however, exhibited certain features reminiscent of cellular aging such as actin accumulation, reduced substrate adherence, and increased activity of lysosomal acid beta-galactosidase. Early MSCs retained the multipotentiality capable of chondrogenic, osteogenic, and adipogenic differentiation upon induction in vitro. In contrast, late MSCs were only capable of adipogenic differentiation, which was greatly enhanced at the expense of the osteochondrogenic potential. Along with these changes in multipotentiality, late MSCs expressed decreased levels of the bone morphogenic protein (BMP-7) and reduced activity of alkaline phosphatase. Late MSCs also exhibited attenuated synthesis of the hematopoietic cytokines granulocyte colony-stimulating factor (G-CSF), leukemia inhibitory factor (LIF), and stem cell factor (SCF). The long-term porcine MSC culture, thus, provides a model system to study the molecular interplay between multiple MSC differentiation cascades in the context of cellular aging.
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PMID:Phenotypic changes of adult porcine mesenchymal stem cells induced by prolonged passaging in culture. 1588 Jun 40

To determine how low intensity pulsed ultrasound alters gene expression in rat bone marrow stromal cells and to see if combining this stimulation with BMP-2, cells were pre-cultured for eight days in the presence of 50 microg/ml ascorbic acid and then exposed to either low intensity US or 100 ng/ml BMP-2 or both combined, beginning on the first, third fifth or seventh day of culture so that cells were exposed to the stimuli for one, three, five or seven days. Real time PCR was used to determine the effect of these treatments on gene expression of several genes associated with osteogenesis. The expression of some of the genes (Cbfa-1/Runx2, IGF-receptor, Alk-3, alkaline phosphatase, osteopontin, TGF-beta1, BMP-7) was increased compared to untreated controls. Combination of US and BMP-2 treatment did not lead to synergy of the two stimuli. Cbfa-1 stimulation occurred more quickly with US than with BMP-2. Increases in gene expression were greatest after 3 days exposure to US, with similar results for BMP-2 treatment implying that there may be a time dependence for the stimulus of osteogenic gene expression in stromal cells.
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PMID:Effect of low intensity pulsed ultrasound and BMP-2 on rat bone marrow stromal cell gene expression. 1588 87

Previous studies from this laboratory showed that the action of Osteogenic Protein-1 (OP-1, BMP-7) on osteoblastic cell differentiation could be enhanced by other protein factors, such as Insulin-like Growth Factor (IGF)-I. In the present study, we examined the effects of co-transfection with a combination of the OP-1 and the IGF-I gene on osteoblastic cell differentiation. The results first showed that fetal rat calvaria (FRC) cells transfected with the OP-1 gene under the control of the cytomegalovirus (CMV) promoter showed substantial production of the OP-1 protein. Transfected FRC cells also showed a DNA concentration-dependent increase in alkaline phosphatase (AP) activity, an osteoblastic cell differentiation marker. Von Kossa-positive nodules, a hallmark of bone formation in long-term cultures of bone-derived cells, were also observed in the transfected cells after 26 days in culture, whereas none were observed in control cells. Co-transfection of FRC cells with the combination of the OP-1 and the IGF-I gene resulted in a synergistic stimulation of AP activity. The increase was DNA dose-dependent. The current data show that transfection of OP-1 gene into osteoblastic cells stimulates osteoblastic cell differentiation in vitro. The study further demonstrates the feasibility of employing gene transfer of a second gene in combination with an OP-1 vector to synergistically enhance OP-1 activity.
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PMID:Co-transfection with the osteogenic protein (OP)-1 gene and the insulin-like growth factor (IGF)-I gene enhanced osteoblastic cell differentiation. 1636 66

Both bone morphogenetic proteins (BMPs) and growth factors released from activated platelets both occur at sites of bone regeneration but their functional relationship to regulate the temporal and spatial sequence of cellular events is not well defined. Here we investigated whether supernatants derived from activated platelets can modulate the response of the osteogenic cell line MC3T3-E1 to BMPs, and whether BMPs have an effect on MC3T3-E1 cells stimulated with platelet-released supernatant. Platelet-released supernatant suppressed BMP-2-, BMP-6-, and BMP-7-induced osteogenic differentiation of MC3T3-E1 cells, as indicated by the significant decrease in alkaline phosphatase activity and lower levels of osteocalcin transcripts, whereas BMP-2, BMP-6, and BMP-7 did not modulate migration and proliferation of MC3T3-E1 cells, which were stimulated with platelet-released supernatant. Osteogenic differentiation in response to BMPs was not affected after precultivation of MC3T3-E1 cells with platelet-released supernatant. These data suggest that activated platelets can provide a microenvironment that temporarily suppresses the differentiation of osteogenic cells in the presence of BMPs. Future strategies to stimulate bone regeneration should take the suppression of BMP-induced osteogenic differentiation during the existence of the blood clot into account.
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PMID:Osteogenic differentiation induced by bone morphogenetic proteins can be suppressed by platelet-released supernatant in vitro. 1658 15

Degeneration of the rotator cuff is often associated with inflammation of the subacromial bursa and focal mineralization of the supraspinatus tendon. Portions of the supraspinatus tendon distant from the insertion site could transform into fibrous cartilage, causing rotator-cuff tears owing to mechanical instability. Indirect evidence is presented to link this pathology to ectopic production and secretion of bioactive bone morphogenetic proteins (BMPs) from sites within the subacromial bursa. Surgically removed specimens of subacromial bursa tissue from patients with chronic tears of the rotator cuff were analyzed by immunohistochemistry and reverse transcription-PCR. Bioactive BMP was detected in bursa extracts by a bioassay based on induction of alkaline phosphatase in the osteogenic/myogenic cell line C2C12. Topical and differential expression of BMP-2/4 and BMP-7 mRNA and protein was found in bursa tissue. The bioassay of C2C12 cells revealed amounts of active BMP high enough to induce osteogenic cell types, and blocking BMP with specific antibodies or soluble BMP receptors Alk-3 and Alk-6 abolished the inductive properties of the extract. Sufficient information was gathered to explain how ectopic expression of BMP might induce tissue transformation into ectopic bone/cartilage and, therefore, promote structural degeneration of the rotator cuff. Early surgical removal of the subacromial bursa might present an option to interrupt disease progression.
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PMID:Expression of bioactive bone morphogenetic proteins in the subacromial bursa of patients with chronic degeneration of the rotator cuff. 1671 33

Noggin is a secreted protein that inhibits the binding of bone morphogenetic proteins (BMPs) to their cognate receptor. Its role in human mesenchymal stem cell differentiation has not been well studied. Here, we studied the effect of noggin on human mesenchymal stem cell differentiation induced by inflammatory cytokines (activated T-cell conditioned medium (ACTTCM) or the combination of four T-cell cytokines, TNF-alpha, TGF-beta, IFN-gamma, and IL-17 (TTII)), BMPs, or dexamthasone (DEX). HMSC treated with TTII alone rapidly induced alkaline phosphatase (AlkP) activity. Inclusion of noggin resulted in an additive effect. Noggin acted additively with DEX to induce a significantly higher level of AlkP induction than either noggin or DEX alone. Noggin was examined for its ability to inhibit mineralization in long-term cultures of HMSC stimulated with BMP-2, BMP-6, BMP-7, DEX, or TTII. Surprisingly, noggin alone induced mineralization while it did not inhibit mineralization induced by TTII or BMP-2, BMP-6, or BMP-7. Interestingly, when HMSC were treated with both noggin and DEX they acted synergistically to induce mineralization nearly 3-fold over DEX alone and 30-fold over noggin alone. Reverse transcriptase-polymerase chain reaction (RT-PCR) analysis showed that T-cell cytokines induced noggin, Runx2, BMP-2, and osteocalcin gene expression, while noggin alone induced BMP-2 and osteocalcin gene expression, but not Runx2, although it increased the expression of ActRII, a receptor for BMP-2. These results suggest that in HMSC, the anabolic action of inflammation on bone formation occurs through the induction of noggin, which then induces BMP-2 receptor and BMP-2 leading to the activation of the differentiation process.
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PMID:The role of noggin in human mesenchymal stem cell differentiation. 1713 53

The dental follicle (DF) surrounding the developing tooth germ is an ectomesenchymal tissue composed of various cell populations derived from the cranial neural crest. Human dental follicle cells (HDFC) are believed to contain precursor cells for cementoblasts, periodontal ligament cells, and osteoblasts. Bone morphogenetic proteins (BMPs) produced by Hertwig's epithelial root sheath or present in enamel matrix derivatives (EMD) seem to be involved in the control of DF cell differentiation, but their precise function remains largely unknown. We report the immunolocalization of STRO-1 (a marker of multipotential mesenchymal progenitor cells) and BMP receptors (BMPR) in DF in vivo. In culture, HDFC co-express STRO-1/BMPR and exhibit multilineage properties. Incubation with rhBMP-2 and rhBMP-7 or EMD for 24 h increases the expression of BMP-2 and BMP-7 by HDFC. Long-term stimulation of these cells by rhBMP-2 and/or rhBMP-7 or EMD significantly increases alkaline phosphatase activity (AP) and mineralization. Expression of cementum attachment protein (CAP) and cementum protein-23 (CP-23), two putative cementoblast markers, has been detected in EMD-stimulated whole DF and in cultured HDFC stimulated with EMD or BMP-2 and BMP-7. RhNoggin, a BMP antagonist, abolishes AP activity, mineralization, and CAP/CP-23 expression in HDFC cultures and the expression of BMP-2 and BMP-7 induced by EMD. Phosphorylation of Smad-1 and MAPK is stimulated by EMD or rhBMP-2. However, rhNoggin blocks only Smad-1 phosphorylation under these conditions. Thus, EMD may activate HDFC toward the cementoblastic phenotype, an effect mainly (but not exclusively) involving both exogenous and endogenous BMP-dependent pathways.
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PMID:Human dental follicle cells acquire cementoblast features under stimulation by BMP-2/-7 and enamel matrix derivatives (EMD) in vitro. 1744 52

In this study, we delineate the sequential expression of selected growth factors associated with bone formation in vitro. Mineralization, osteocalcin, and alkaline phosphatase (ALP-2) were measured to monitor the differentiation and maturation of osteoprogenitor cells collected from C57BL mice. Bone-related growth factors, including transforming growth factor beta (TGF-beta), fibroblast growth factor 2 (FGF-2), platelet-derived growth factor (PDGF), insulinlike growth factor (IGF)-1, vascular endothelial growth factor (VEGF), bone morphogenetic protein (BMP)-2, and BMP-7, were selected. Enzyme-linked immunosorbent assay (ELISA) and reverse transcriptase polymerase chain reaction (RT-PCR) were used to measure growth factors at the protein and messenger ribonucleic acid (mRNA) level, respectively. The results found that ALP-2 expression increased progressively over time, whereas mineralization and osteocalcin did not become evident until culture day 14. VEGF and IGF-1 were upregulated early during proliferation. PDGF and TGF-beta mRNA expression was bimodal. FGF-2 and BMP-2 mRNAs were expressed only later in differentiation. FGF-2 mRNA signal levels were highest at day 14 and remained prominent through day 28 of culture. BMP-2 showed a similar profile as FGF-2. BMP-7 was not detectable using RT-PCR or ELISA. Strong correlations existed for the expression patterns between several early-response growth factors (VEGF, TGF-beta, and IGF-1) and were also evident for several late-response growth factors (BMP-2, PDGF, and FGF-2). Differential expression for grouped sets of growth factors occurs during the temporal acquisition of bone-specific markers as osteoprogenitor cell maturation proceeds in vitro.
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PMID:The sequential expression profiles of growth factors from osteoprogenitors [correction of osteroprogenitors] to osteoblasts in vitro. 1752 79

Morselized cancellous allograft bone is frequently used in the reconstruction of bone defects in cases of revision total joint replacement, trauma, spine fusion and treated infection. However, the initial lack of viable bone cells in morselized allograft bone significantly slows the process of graft incorporation compared to autograft bone. This study examined the effects of prolonged local infusion of the growth factors bone morphogenic protein-7 (BMP-7 or OP-1) and fibroblast growth factor-2 (FGF-2 or basic FGF) in the process of allograft incorporation using a rabbit tibial chamber model. New bone formation was evaluated by two indices, the activity of alkaline phosphatase and the level of birefringence. The markers of osteoclast-like cells were also measured. Without the infusion of the growth factors, lower levels of new bone formation were observed in the allograft group, compared to the autograft group. Infusion of growth factors FGF-2 and OP-1, singly or in combination, for 4 weeks, diminished this difference. The numbers of osteoclast-like cells were much higher in the allograft group before the growth factors were delivered. The infusion of FGF, singly, diminished this difference. However, the infusion of OP-1 or the combination of FGF and OP-1 did not decrease the number of osteoclast-like cells to a level comparable to autograft only. Local infusion of growth factors appears to be a useful adjunct to promote the incorporation of allograft bone in vivo.
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PMID:Modulation of allograft incorporation by continuous infusion of growth factors over a prolonged duration in vivo. 1761 98

LIM mineralization protein-1 (LMP-1) is a novel intracellular osteoinductive protein that has been shown to induce bone formation both in vitro and in vivo. LMP-1 contains an N-terminal PDZ domain and three C-terminal LIM domains. In this study, we investigated whether a truncated form of human LMP-1 (hLMP-1[t]), lacking the three C-terminal LIM domains, triggers the differentiation of pluripotent myoblastic C(2)C(12) cells to the osteoblast lineage. C(2)C(12) cells were transiently transduced with Ad5-hLMP-1(t)-green fluorescent protein or viral vector control. The expression of hLMP-1(t) RNA and the truncated protein were examined. The results showed that hLMP-1(t) blocked myotube formation in C(2)C(12) cultures and significantly enhanced the alkaline phosphatase (ALP) activity. In addition, the expressions of ALP, osteocalcin, and bone morphogenetic protein (BMP)-2 and BMP-7 genes were also increased. The induction of these key osteogenic markers suggests that hLMP-1(t) can trigger the pluripotent myoblastic C2C12 cells to differentiate into osteoblastic lineage, thus extending our previous observation that LMP-1 and LMP-1(t) enhances the osteoblastic phenotype in cultures of cells already committed to the osteoblastic lineage. Therefore, C(2)C(12) cells are an appropriate model system for the examination of LMP-1 induction of the osteoblastic phenotype and the study of mechanisms of LMP-1 action.
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PMID:Truncated human LMP-1 triggers differentiation of C2C12 cells to an osteoblastic phenotype in vitro. 1780 64


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