Abstract The MC3T3-E1 mouse calvaria-derived cell line has been used to study the role of collagen synthesis in osteoblast differentiation. cells, like several previously characterized culture systems, expressed markers and formed a mineralized extracellular matrix only after exposure ascorbic acid. Mineralization was stimulated further by β-glycerol phosphate. Ultrastructural observations indicated that produced acid-treated cells highly organized contained well-banded fibrils. Expression...

There is significant interest in the development of injectable carriers for cell transplantation to engineer bony tissues. In this study, we hypothesized that adhesion ligands covalently coupled hydrogel would allow one control pre-osteoblast attachment, proliferation, and differentiation. Modification alginate with an RGDcontaining peptide promoted osteoblast spreading, whereas minimal was observed on unmodified hydrogels. Raising ligand density increased a minimum (1.5-15 femtomoles/cm2)...

The extracellular signal–regulated kinase (ERK)–mitogen-activated protein (MAPK) pathway provides a major link between the cell surface and nucleus to control proliferation differentiation. However, its in vivo role skeletal development is unknown. A transgenic approach was used establish for this bone. MAPK stimulation achieved by selective expression of constitutively active MAPK/ERK1 (MEK-SP) osteoblasts accelerated vitro differentiation calvarial cells, as well bone development, whereas...

Abstract Osteoblasts secrete a complex extracellular matrix (ECM) containing collagenous and noncollagenous proteins, bone morphogenetic proteins (BMPs), growth factors. Osteoblast-specific gene expression requires ascorbic acid (AA)-dependent assembly of ECM. Matrix responsiveness an α2β1 integrin-collagen interaction mitogen-activated protein kinase (MAPK) activity, which phosphorylates activates the osteoblast-specific transcription factor Cbfa1. This study examines interactions between...

Treatment of mouse MC3T3-E1 cells with ascorbic acid initiates the formation a collagenous extracellular matrix and synthesis several osteoblast-related proteins. We recently showed that dramatically increases alkaline phosphatase osteocalcin mRNAs this induction is blocked by inhibitors collagen triple-helix (Franceschi Iyer, J Bone Miner Res 7:235). In present study, relationship between osteoblast-specific gene expression explored in greater detail. Kinetic studies revealed increased...

Engineering new bone tissue with cells and a synthetic extracellular matrix (scaffolding) represents approach for the regeneration of mineralized tissues compared transplantation (autografts or allografts). In present work, highly porous poly(L-lactic acid) (PLLA) PLLA/hydroxyapatite (HAP) composite scaffolds were prepared thermally induced phase separation technique. The seeded osteoblastic cultured in vitro. pure PLLA scaffolds, osteoblasts attached primarily on outer surface polymer....

RUNX2 expression in mesenchymal cells induces osteoblast differentiation and bone formation. BMP blocking agents were used to show that RUNX2-dependent transactivation activity both require signaling and, further, enhances the responsiveness of BMPs.BMPs transcription factor are able stimulate BMPs function by activating SMAD proteins other signal transduction pathways many target genes including RUNX2. In contrast, osteoblast-specific gene directly binding enhancer regions genes. this...

Abstract Bone morphogenetic proteins are essential for bone regeneration/fracture healing but can also induce heterotopic ossification (HO). Understanding accessory factors modulating BMP signaling would provide both a means of enhancing BMP-dependent regeneration while preventing HO. This study focuses on the ability collagen receptor, discoidin domain receptor 2 (DDR2), to regulate activity. As will be shown, induction formation by subcutaneous BMP2 implants is severely compromised in Ddr2...

Fibroblast growth factor 2 (FGF-2) is an important regulator of bone formation and osteoblast activity. However, its mechanism action on cells largely unknown. A major route for FGF signaling through the mitogen-activated protein kinase (MAPK) pathway. We showed recently that this pathway activation phosphorylation Cbfa1/Runx2, osteoblast-related transcription (Xiao, G., Jiang, D., Thomas, P., Benson, M. Guan, K., Karsenty, Franceschi, R. T. (2000)<i>J. Biol. Chem.</i> 275, 4453–4459). The...

Extracellular matrix molecules such as type I collagen are required for the adhesion, migration, proliferation, and differentiation of a number cell types including osteoblasts. Matrix components often affect function by interacting with members integrin family surface receptors. Previous work showed that synthesis, induced addition ascorbic acid to cells, precedes is essential expression osteoblast markers induction osteocalcin promoter in murine MC3T3-E1 cells. This later response requires...

AbstractThe Cbfa1/Runx2 transcription factor is essential for osteoblast differentiation. However, levels of Runx2 are often not well correlated with its transcriptional activity suggesting that this must be activated either by covalent modification or through interactions other nuclear components. phosphorylated and the mitogen-activated protein kinase (MAPK) pathway. This pathway stimulated in at least two ways: binding type I collagen to &#102 2 &#103 1 integrins on surface treatment...

An ex vivo gene therapy strategy was used to achieve localized skeletal regeneration in vivo. When an adenovirus vector engineered express bone morphogenetic protein 7 transduced human gingival fibroblasts or rat dermal fibroblasts, these nonosteogenic tissues formed and supported the development of hematopoietic tissue when transplanted into immunocompromised mice. Transduced marrow-containing ossicles 100% transplants after 1–2 weeks (n = 30). Immunostaining with murine human-specific...

Bone regeneration is based on the hypothesis that healthy progenitor cells, either recruited or delivered to an injured site, can ultimately regenerate lost damaged tissue. Three-dimensional porous polymer scaffolds may enhance bone by creating and maintaining a space facilitates cell migration, proliferation, differentiation. As initial step test this possibility, osteogenic cells were cultured fabricated from biodegradable polymers, development these was evaluated. Porous polymers of...

The role of ATF4 (activating transcription factor 4) in osteoblast differentiation and bone formation was recently described using ATF4-deficient mice (Yang, X., Matsuda, K., Bialek, P., Jacquot, S., Masuoka, H. C., Schinke, T., Li, L., Brancorsini, Sassone-Corsi, Townes, T. M., Hanauer, A., Karsenty, G. (2004) Cell 117, 387-398). However, the mechanisms cells are still not clear. In this study, we determined molecular through which activates mouse osteocalcin (Ocn) gene 2 (mOG2) expression...

Bone morphogenetic proteins (BMPs) are well-established agents for inducing orthotopic and ectopic bone formation. However, their clinical usefulness as regenerative may be limited by a short in vivo half-life low specific activity. BMP gene therapy is an alternative route exploiting the bone-inductive activity of this class molecules. To test feasibility approach, we examined osteogenic AdCMV-BMP7, adenovirus containing BMP7 cDNA under control CMV promoter that was constructed using Cre/lox...

Abstract Bone regeneration requires interactions between a number of factors including bone morphogenetic proteins (BMPs), growth factors, and transcriptional regulators such as Runx2/Cbfa1 (Runx2). Because each component may provide unique contribution to the overall osteogenic response, we hypothesized that formation be enhanced by using combinations complimentary factors. As an initial test this concept, BMP2 Runx2 were examined adenovirus-based expression vectors (AdCMV-Runx2,...

Osteocalcin is a hormonally regulated calcium-binding protein made almost exclusively by osteoblasts. In normal cells, osteocalcin expression requires ascorbic acid (AA), an essential cofactor for osteoblast differentiation both in vivo and vitro. To determine the mechanism of this regulation, subclones MC3T3-E1 preosteoblasts were transiently transfected with 1.3 kb mouse gene 2 promoter driving firefly luciferase. AA stimulated luciferase activity 20-fold after 4–5 days. This response was...

Abstract The ability of the hormonally active vitamin D metabolite, lα,25‐dihydroxyvitamin 3 , to affect cell growth, morphology and fibronectin production has been examined using MG‐63 human osteosarcoma line. Hormone treatment reduced growth rate, saturation density [ H]thymidine incorporation. Inhibition was specific for relative other metabolites (lα,25‐dihydroxyvitamin &gt; 25‐dihydroxyvitamin 24R,25‐dihydroxyvitamin ), antagonized by high concentrations serum readily reversed removal...