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Query: UNIPROT:P06889 (Mol)
 630,302 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Human mesenchymal stem cells (hMSCs) are an attractive tissue engineering avenue for the repair and regeneration of bone. In this study we detail the in vivo performance of a novel electrospun polycaprolactone scaffold incorporating the glycosaminoglycan heparan sulfate (HS) as a carrier for hMSC. HS is a multifunctional regulator of many key growth factors expressed endogenously during bone wound repair, and we have found it to be a potent stimulator of proliferation in hMSCs. To assess the potential of the scaffolds to support hMSC function in vivo, hMSCs pre-committed to the osteogenic lineage (human osteoprogenitor cells) were seeded onto the scaffolds and implanted subcutaneously into the dorsum of nude rats. After 6 weeks the scaffolds were retrieved and examined by histological methods. Implanted human cells were identified using a human nuclei-specific antibody. The host response to the implants was characterized by ED1 and ED2 antibody staining for monocytes/macrophages and mature tissue macrophages, respectively. It was found that the survival of the implanted human cells was affected by the host response to the implant regardless of the presence of HS, highlighting the importance of controlling the host response to tissue engineering devices.
J Mol Histol 2007 Oct
PMID:The in vivo assessment of a novel scaffold containing heparan sulfate for tissue engineering with human mesenchymal stem cells. 1769 76

Bone marrow-derived mesenchymal stem cells consist of a developmentally heterogeneous population of cells obtained from colony forming progenitors. As these colonies express the alpha-1 integrin (CD49a), here we single-cell FACS sorted CD49a+ cells from bone marrow in order to create clones and then compared their colony forming efficiency and multilineage differentiation capacity to the unsorted cells. Following selection, 40% of the sorted CD49a+ cells formed colonies, whereas parental cells failed to form colonies following limited dilution plating at 1 cell/well. Following ex vivo expansion, clones shared a similar morphology to the parental cell line, and also demonstrated enhanced proliferation. Further analysis by flow cytometry using a panel of multilineage markers demonstrated that the CD49a+ clones had enhanced expression of CD90 and CD105 compared to unsorted cells. Culturing cells in adipogenic, osteogenic or chondrogenic medium for 7, 10 and 15 days respectively and then analysing them by quantitative PCR demonstrated that CD49a+ clones readily underwent multlineage differentiation into fat, bone and cartilage compared to unsorted cells. These results thus support the use of CD49a selection for the enrichment of mesenchymal stem cells, and describes a strategy for selecting the most multipotential cells from a heterogeneous pool of bone marrow mononuclear stem cells.
J Mol Histol 2007 Oct
PMID:Selection using the alpha-1 integrin (CD49a) enhances the multipotentiality of the mesenchymal stem cell population from heterogeneous bone marrow stromal cells. 1769 77

Ror2 receptor plays a key role in bone formation, but its signaling pathway is not completely understood. We demonstrate that Ror2 homodimerizes at the cell surface, and that dimerization can be induced by a bivalent antibody. Antibody-mediated dimerization causes receptor autophosphorylation and induces functional consequences of its signaling, including osteogenesis in mesenchymal stem cells and bone formation in organ culture. We further show that Ror2 associates with and phosphorylates 14-3-3beta scaffold protein. Endogenous Ror2 binds 14-3-3beta in U2OS osteosarcoma cells, and purified intracellular domain of Ror2 interacts with 14-3-3beta in vitro. 14-3-3beta Is tyrosine phosphorylated in U2OS cells, and this phosphorylation is inhibited by down-regulating Ror2 and enhanced by overexpressing the kinase. Purified Ror2 phosphorylates 14-3-3beta in vitro, confirming 14-3-3beta as the first identified Ror2 substrate. Down-regulating 14-3-3beta potentiates osteoblastogenesis in mesenchymal stem cells and increases bone formation in calvarial cultures, indicating that 14-3-3beta exerts a negative effect on osteogenesis. This raises a possibility that Ror2 induces osteogenic differentiation, at least in part, through a release of the 14-3-3beta-mediated inhibition. Our research forms a foundation for several new areas of investigation, including the molecular regulation of 14-3-3 by tyrosine phosphorylation and the role of this scaffold in osteogenesis.
Mol Endocrinol 2007 Dec
PMID:Homodimerization of Ror2 tyrosine kinase receptor induces 14-3-3(beta) phosphorylation and promotes osteoblast differentiation and bone formation. 1771 73

This paper explores the potential therapeutic role of the naturally occurring sugar heparan sulfate (HS) for the augmentation of bone repair. Scaffolds comprising fibrin glue loaded with 5 microg of embryonically derived HS were assessed, firstly as a release-reservoir, and secondly as a scaffold to stimulate bone regeneration in a critical size rat cranial defect. We show HS-loaded scaffolds have a uniform distribution of HS, which was readily released with a typical burst phase, quickly followed by a prolonged delivery lasting several days. Importantly, the released HS contributed to improved wound healing over a 3-month period as determined by microcomputed tomography (microCT) scanning, histology, histomorphometry, and PCR for osteogenic markers. In all cases, only minimal healing was observed after 1 and 3 months in the absence of HS. In contrast, marked healing was observed by 3 months following HS treatment, with nearly full closure of the defect site. PCR analysis showed significant increases in the gene expression of the osteogenic markers Runx2, alkaline phosphatase, and osteopontin in the heparin sulfate group compared with controls. These results further emphasize the important role HS plays in augmenting wound healing, and its successful delivery in a hydrogel provides a novel alternative to autologous bone graft and growth factor-based therapies.
J Mol Histol 2007 Oct
PMID:Sustained release and osteogenic potential of heparan sulfate-doped fibrin glue scaffolds within a rat cranial model. 1784 24

Runt-related transcription factor Runx2 regulates osteogenic phenotype commitment and attenuates osteoblast growth. Runx2 levels are cell cycle regulated and maximal in the G1 phase of proliferating osteoblasts and during quiescence. The Wnt/Lrp5-Frizzled/beta-catenin/Lef-Tcf signaling cascade also controls progression along the osteogenic lineage with a net anabolic effect that promotes bone formation. However, Lef1 opposes the osteoblast maturation promoting activity of Runx2. Here we examined whether Lef1 controls Runx2 expression during the cell cycle or onset of quiescence in osteoblasts. We inhibited Lef1 expression using short hairpin (sh) RNA interference in stably transfected MC3T3-E1 cells. In asynchronously growing osteoblasts, expression of Lef1 shRNA diminishes Lef1 protein levels, but does not affect Runx2 levels. Cells arrested in different cell cycle stages using mimosine (late G1), hydroxyurea or aphidicolin (S phase) or nocodazole (mitosis) exhibit expected reductions in Runx2 protein levels despite reductions in Lef1. Serum deprived MC3T3-E1 cells normally upregulate Runx2 protein regardless of Lef1 deficiency, although loss of Lef1 reduces cyclin A and increases cyclin D1 expression upon serum withdrawal. Thus, Runx2 protein levels during the cell cycle and onset of quiescence are regulated independently of Lef1, one of the major transcriptional inducers of Wnt signaling in proliferating cells.
J Mol Histol 2007 Oct
PMID:Cell cycle related modulations in Runx2 protein levels are independent of lymphocyte enhancer-binding factor 1 (Lef1) in proliferating osteoblasts. 1788 13

Reparative dentin has a wide variety of manifestations ranging from a regular, tubular form to an irregular, atubular form. However, the characteristics of reparative dentin have not been clarified. This study hypothesized that the level of bone sialoprotein (BSP) expression will increase if the newly formed reparative dentin is bone-like but the dentin sialophosphoprotein (DSPP) level will decrease. In order to test this hypothesis, the expression of BSP and DSP was examined by immunohistochemistry and the expression of BSP was measured by in situ hybridization in an animal model. The pulps of 12 maxillary right first molars from twelve male rats were exposed and capped with MTA. In addition, in order to understand the role of transforming growth factor-beta 1 (TGF-beta1) during reparative dentinogenesis, the expression of BSP and DSPP mRNA was analyzed by RT-PCR in a human dental pulp cell culture, and the transforming growth factor-beta 1 receptors (TbetaRI) and Smad 2/3 were examined by immunofluorescence in an animal model. DSP was expressed in the normal odontoblasts and odontoblast-like cells of the reparative dentin. Interestingly, BSP was strongly expressed in the odontoblast-like cells of reparative dentin. The level of the TbetaRI and Smad 2/3 proteins was higher in the reparative dentin than in the normal dentin. TGF-beta1 up-regulated BSP in the human pulp cell cultures. This suggests that reparative dentin has both dentinogenic and osteogenic characteristics that are mediated by TGF-beta1.
J Mol Histol 2008 Apr
PMID:Influence of TGF-beta1 on the expression of BSP, DSP, TGF-beta1 receptor I and Smad proteins during reparative dentinogenesis. 1792 79

Na+-Ca2+ exchanger (NCX) transports Ca2+ coupled with Na+ across the plasma membrane in a bi-directional mode. Ca2+ flux via NCX mediates osteogenic processes, such as formation of extracellular matrix proteins and bone nodules. However, it is not clearly understood how the NCX regulates cellular Ca2+ movements in osteogenic processes. In this study, the role of NCX in modulating Ca2+ content of intracellular stores ([Ca2+]ER) was investigated by measuring intracellular Ca2+ activity in isolated rat osteoblasts. Removal of extracellular Na+ elicited a transient increase of intracellular Ca2+ concentration ([Ca2+]i). Pretreatment of antisense oligodeoxynucleotide (AS) against NCX depressed this transient Ca2+ rise and raised the basal level of [Ca2+]i. In AS-pretreated cells, the expression and activity of alkaline phosphatase (ALP), an osteogenic marker, were decreased. However, the cell viability was not affected by AS-pretreatment. Suppression of NCX activity by the AS-pretreatment decreased ATP-activated Ca2+ release from intracellular stores and significantly enhanced Ca2+ influx via store operated calcium influx (SOCI), compared to those of S-pretreated or control cells. These results strongly suggest that NCX has a regulatory role in cellular Ca2+ pathways in osteoblasts by modulating intracellular Ca2+ content.
Exp Mol Med 2007 Aug 31
PMID:Na+-Ca2+ exchanger modulates Ca2+ content in intracellular Ca2+ stores in rat osteoblasts. 1793 33

Bone marrow (BM) derived mesenchymal stem cells (MSC) are pluripotent cells which can differentiate into osteogenic, adipogenic and other lineages. In spite of the broad interest, the information about the changes in BM cell composition, in particularly about the variation of MSC number and their properties in relation to the age of the donor is still controversial. The aim of this study was to investigate the age associated changes in variations of BM cell composition, phenotype and differentiation capacities of MSC using a rat model. Cell populations were characterized by flow cytometry using light scattering parameters, DNA content and a set of monoclonal antibodies. Single cell analysis was performed by conventional fluorescent microscopy. In vitro culture of MSC was established and their phenotype and capability for in vitro differentiation into osteogenic and adipogenic cells was shown. Age related changes in tibiae and femurs, amount of BM tissue, BM cell composition, proportions of separated MSC and yield of MSC in 2 weeks of in vitro culture were found. At the same time, neither change in phenotype no in differentiation capacities of MSC was registered. Age-related changes of the number of MSC should be taken into account whenever MSC are intended to be used for investigations.
Mol Cells 2007 Oct 31
PMID:A number of bone marrow mesenchymal stem cells but neither phenotype nor differentiation capacities changes with age of rats. 1797 79

MMPs are endopeptidases that play a pivotal role in ECM turnover. RECK is a single membrane-anchored MMP-regulator. Here, we evaluated the temporal and spatial expression of MMP-2, MMP-9, and RECK during alveolar bone regeneration. The maxillary central incisor of Wistar rats was extracted and the animals were killed at 1, 3, 7, 10, 14, 21, 28, and 42 days post-operatively (n = 3/period). The hemimaxillae were collected, demineralized and embedded in paraffin. Immunohistochemical analysis was performed by the immunoperoxidase technique with polyclonal antibodies. On day 1, polymorphonuclear cells in the blood clot presented mild immunolabeling for MMPs. During bone remodeling, osteoblasts facing new bone showed positive staining for gelatinases and RECK in all experimental periods. MMPs were also found in the connective tissue and endothelial cells. Our results show for the first time that inactive and/or active forms of MMP-2, MMP-9 and RECK are differentially expressed by osteogenic and connective cells during several events of alveolar bone regeneration. This may be important for the replacement of the blood clot by connective tissue, and in the formation, maturation and remodeling of new bone.
J Mol Histol 2008 Apr
PMID:Expression of matrix metalloproteinases-2 and -9 and RECK during alveolar bone regeneration in rat. 1798 94

In this study, we established an in vitro model of osteogenic-inductive differentiation of rat bone marrow mesenchymal stem cells (BMSCs) to determine the mechanisms and relative gene function underlying BMSCs osteogenesis. Osteoplastic differentiation of the third generation BMSCs was induced with the alpha-minimal essential medium containing beta-glyceraldehyde-3-phosphate, L: -ascorbic acid, dexamethasone and 1,25-2(OH)2 vitamin D3 prior to applying gene chip technology (also called microarray technology) for global gene expression screening. Real-time quantitative PCR (Real-time PCR) was used to determine the temporal profile of mRNA expression of regulated genes during osteogenic differentiation of BMSCs. A bioinformatic analysis was utilized to determine the functional significance of the identified osteogenic-related genes. Purkinje cell protein 4 (Pcp4) mRNA expression was identified by the gene chip screening as being up-regulated during osteoplastic differentiation of BMSCs. Real-time PCR analysis confirmed the increased expression of Pcp4 mRNA expression during osteoplastic differentiation of BMSCs with an upward trend that peaked at day 14. The bioinformatic analysis identified Pcp4 as a gene involved in the deposition of calcium and the modulation of CaM-dependent protein kinase. Thus, we hypothesize that Pcp4 osteoplastic differentiation of BMSCs is mediated in part via Pcp4-induced calcium deposition to form mineral nodules and modulation of certain signal transduction pathways of BMPs.
Mol Cell Biochem 2008 Feb
PMID:Expression of Pcp4 gene during osteogenic differentiation of bone marrow mesenchymal stem cells in vitro. 1800 38


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