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Sample records for bone matrix protein

  1. A strategy to quantitate global phosphorylation of bone matrix proteins.

    Science.gov (United States)

    Sroga, Grażyna E; Vashishth, Deepak

    2016-04-15

    Current studies of protein phosphorylation focus primarily on the importance of specific phosphoproteins and their landscapes of phosphorylation in the regulation of different cellular functions. However, global changes in phosphorylation of extracellular matrix phosphoproteins measured "in bulk" are equally important. For example, correct global phosphorylation of different bone matrix proteins is critical to healthy tissue biomineralization. To study changes of bone matrix global phosphorylation, we developed a strategy that combines a procedure for in vitro phosphorylation/dephosphorylation of fully mineralized bone in addition to quantitation of the global phosphorylation levels of bone matrix proteins. For the first time, we show that it is possible to enzymatically phosphorylate/dephosphorylate fully mineralized bone originating from either cadaveric human donors or laboratory animals (mice). Using our strategy, we detected the difference in the global phosphorylation levels of matrix proteins isolated from wild-type and osteopontin knockout mice. We also observed that the global phosphorylation levels of matrix proteins isolated from human cortical bone were lower than those isolated from trabecular bone. The developed strategy has the potential to open new avenues for studies on the global phosphorylation of bone matrix proteins and their role in biomineralization as well for other tissues/cells and protein-based materials.

  2. Induction of Bone Matrix Protein Expression by Native Bone Matrix Proteins in C2C12 Culture

    Institute of Scientific and Technical Information of China (English)

    ZHEN-MING HU; SEAN A. F. PEEL; STEPHEN K. C. HO; GEORGE K. B. SANDOR; CAMERON M. L. CLOKIE

    2009-01-01

    Objective To study the expression of bone matrix protein (BMP) induced by bovine bone morphogenetic proteins (BMPs) in vitro. Methods Type I collagen, osteopontin (OPN), osteonectin (ON), osteocalcin (OC), and bone sialoprotein (BSP) were detected by immunohistochemistry in C2C12 cultured from day 1 to day 28. Results The signaling of bone matrix protein expression became weaker except for type I collagen, OC and BSP after 5 days. Fourteen days after culture, the positive signaling of type I collagen, OPN, ON, OC, and BSP was gradually declined, and could be detected significantly as compared with that of the negative control on day 28. BMP assay showed that the Ikaline phosphatase (ALP) activity was higher in C2C12 culture than in the control during the 14-day culture. Also, total protein and DNA significantly increased during the 14-day culture. High levels of ALP were seen in preosteoblasts and osteoblsts in vivo and in differentiating ostcoblasts in vitro. ALP was well recognized as a marker reflecting osteoblastic activity. Conclusion Native bovine BMP induces conversion of myoblasts into osteoblasts, produces type 1 collagen, and plays significantly role in osteoinduction and bone matrix mineralization of C2C12 in vitro.

  3. The effect of enamel matrix proteins and deproteinized bovine bone mineral on heterotopic bone formation.

    Science.gov (United States)

    Donos, Nikolaos; Kostopoulos, Lambros; Tonetti, Maurizio; Karring, Thorkild; Lang, Niklaus P

    2006-08-01

    To evaluate the osteoinductive potential of deproteinized bovine bone mineral (DBBM) and an enamel matrix derivative (EMD) in the muscle of rats. Sixteen rats were used in this study. The animals were divided in three groups. Group A: a pouch was created in one of the pectoralis profundis muscles of the thorax of the rats and DBBM particles (Bio-Oss) were placed into the pouch. Healing: 60 days. Group B: a small pouch was created on both pectoralis profundis muscles at each side of the thorax midline. In one side, a mixture of EMD (Emdogain) mixed with DBBM was placed into one of the pouches, whereas in the contralateral side of the thorax the pouch was implanted with DBBM mixed with the propylene glycol alginate (PGA--carrier for enamel matrix proteins of EMD). Healing: 60 days. Group C: the same procedure as group B, but with a healing period of 120 days. Qualitative histological analysis of the results was performed. At 60 days, the histological appearance of the DBBM particles implanted alone was similar to that of the particles implanted together with EMD or PGA at both 60 and 120 days. The DBBM particles were encapsulated into a connective tissue stroma and an inflammatory infiltrate. At 120 days, the DBBM particles implanted together with EMD or PGA exhibited the presence of resorption lacunae in some cases. Intramuscular bone formation was not encountered in any group. The implantation of DBBM particles alone, combined with EMD or its carrier (PGA) failed to exhibit extraskeletal, bone-inductive properties.

  4. Dentin extracellular matrix (ECM) proteins: comparison to bone ECM and contribution to dynamics of dentinogenesis.

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    Butler, William T; Brunn, Jan C; Qin, Chunlin

    2003-01-01

    Dentinogenesis involves the initial odontoblastic synthesis of a collagen-rich extracellular matrix (ECM) and predentin that is converted to dentin when the collagen fibrils become mineralized. Since the width of predentin is rather uniform, we postulate that extracellular events regulate dentinogenesis. Similarly, osteogenesis involves an initial unmineralized osteoid that is mineralized and converted to bone. To gain insights into these two processes, we compared ECM proteins in bone with those in dentin, focusing upon the sialic acid (SA)-rich proteins. We observed qualitative similarities between the SA-rich proteins, but distinct differences in the amounts of osteopontin (OPN) and dentin sialoprotein (DSP). OPN, a predominant protein in bone, was found in much smaller amounts in dentin. Conversely, DSP was abundant in dentin ECM, but found sparingly in bone. Molecular cloning experiments indicate that coding sequences for DSP and dentin phosphoprotein (DPP) are found on the same mRNA. We believe that the initial form of the precursor protein DSPP is inactive in influencing the mineralization process and that it must be activated by cleavage of peptide bonds in conserved regions. Thus, unknown proteinases would act on DSPP, possibly at the mineralization front, and liberate active DPP, which plays an initiation and regulatory role in the formation of apatite crystals. This post-translational processing reaction would represent an important control point in dentinogenesis. Recently, we identified uncleaved DSPP in dentin extracts, which should allow us to test portions of our hypothesis.

  5. Comparison of efficacies of different bone substitutes adhered to osteoblasts with and without extracellular matrix proteins

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    Li-Ling Tseng

    2013-12-01

    Conclusion: The results indicated that ECM proteins increased cell attachment to bone substitutes in vitro. The preferential affinity of different bone substitutes to certain ECM proteins was evident. Cerasorb and BoneCeramic had better MG63 human osteosarcoma cell adhesion ability than Bio-Oss and MBCP.

  6. Bone regeneration with osteogenically enhanced mesenchymal stem cells and their extracellular matrix proteins.

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    Clough, Bret H; McCarley, Matthew R; Krause, Ulf; Zeitouni, Suzanne; Froese, Jeremiah J; McNeill, Eoin P; Chaput, Christopher D; Sampson, H Wayne; Gregory, Carl A

    2015-01-01

    Although bone has remarkable regenerative capacity, about 10% of long bone fractures and 25% to 40% of vertebral fusion procedures fail to heal. In such instances, a scaffold is employed to bridge the lesion and accommodate osteoprogenitors. Although synthetic bone scaffolds mimic some of the characteristics of bone matrix, their effectiveness can vary because of biological incompatibility. Herein, we demonstrate that a composite prepared with osteogenically enhanced mesenchymal stem cells (OEhMSCs) and their extracellular matrix (ECM) has an unprecedented capacity for the repair of critical-sized defects of murine femora. Furthermore, OEhMSCs do not cause lymphocyte activation, and ECM/OEhMSC composites retain their in vivo efficacy after cryopreservation. Finally, we show that attachment to the ECM by OEhMSCs stimulates the production of osteogenic and angiogenic factors. These data demonstrate that composites of OEhMSCs and their ECM could be utilized in the place of autologous bone graft for complex orthopedic reconstructions.

  7. Increased serum cartilage oligomeric matrix protein levels and decreased patellar bone mineral density in patients with chondromalacia patellae.

    OpenAIRE

    2002-01-01

    BACKGROUND: Chondromalacia patellae is a potentially disabling disorder characterised by features of patellar cartilage degradation. OBJECTIVE: To evaluate markers of cartilage and bone turnover in patients with chondromalacia patellae. METHODS: 18 patients with chondromalacia patellae were studied. Serum cartilage oligomeric matrix protein (s-COMP) and bone sialoprotein (s-BSP) levels were measured by enzyme linked immunosorbent assay (ELISA) and compared with those of age and sex matched he...

  8. Increased serum cartilage oligomeric matrix protein levels and decreased patellar bone mineral density in patients with chondromalacia patellae.

    OpenAIRE

    Murphy, E; Fitzgerald, O; Saxne, Tore; Bresnihan, B

    2002-01-01

    BACKGROUND: Chondromalacia patellae is a potentially disabling disorder characterised by features of patellar cartilage degradation. OBJECTIVE: To evaluate markers of cartilage and bone turnover in patients with chondromalacia patellae. METHODS: 18 patients with chondromalacia patellae were studied. Serum cartilage oligomeric matrix protein (s-COMP) and bone sialoprotein (s-BSP) levels were measured by enzyme linked immunosorbent assay (ELISA) and compared with those of age and sex matched he...

  9. Skeletal growth after oral administration of demineralized bone matrix.

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    Martínez, J A; Elorriaga, M; Marquínez, M; Larralde, J

    1993-03-01

    Oral administration of bone extracts obtained from bovine demineralized bone matrix to rats has a direct effect on bone metabolism, affecting bone proportions and some markers of bone formation such as bone malate dehydrogenase, serum alkaline phosphatase and serum osteocalcin. Furthermore collagen deposition, bone protein synthesis and nucleic acids content were significantly increased by the treatment.

  10. The bone matrix protein secreted phosphoprotein 24 kD (Spp24): bone metabolism regulator and starting material for biotherapeutic materials.

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    Murray, Samuel S; Wang, Jeffrey C; Duarte, Maria Eugenia Leite; Zhao, Ke-Wei; Tian, Haijun; Francis, Timothy; Brochmann Murray, Elsa J

    2015-05-01

    Secreted phosphoprotein 24 kD (Spp24) is a bone matrix protein that appears to be derived primarily from the liver and delivered to other tissues in a protective complex. A significant role in bone growth and turnover is suggested by genetic studies that associate the gene locus (SPP2) with bone mineral density and bone quality. The function of this protein in the normal bone environment is unknown but clues are given by the fact that Spp24, or proteolytic products of Spp24, bind cytokines of the TGF-β superfamily and also activate intracellular signaling pathways. Several potential biotherapeutics have been engineered from this protein including materials that enhance BMP-induced bone healing and, on the other hand, materials that inhibit BMPs in clinical situations where this is called for such as reducing BMP-induced inflammation and inhibiting tumors dependent on BMP autocrine systems. As understanding of the structure and function of this protein increases, more opportunities for rationally developed therapeutics will become apparent.

  11. Demineralized bone matrix combined bone marrow mesenchymal stem cells, bone morphogenetic protein-2 and transforming growth factor-β3 gene promoted pig cartilage defect repair.

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    Xin Wang

    Full Text Available OBJECTIVES: To investigate whether a combination of demineralized bone matrix (DBM and bone marrow mesenchymal stem cells (BMSCs infected with adenovirus-mediated- bone morphogenetic protein (Ad-BMP-2 and transforming growth factor-β3 (Ad-TGF-β3 promotes the repair of the full-thickness cartilage lesions in pig model. METHODS: BMSCs isolated from pig were cultured and infected with Ad-BMP-2(B group, Ad-TGF-β3 (T group, Ad-BMP-2 + Ad-TGF-β3(BT group, cells infected with empty Ad served as a negative group(N group, the expression of the BMP-2 and TGF-β3 were confirmed by immunofluorescence, PCR, and ELISA, the expression of SOX-9, type II collagen(COL-2A, aggrecan (ACAN in each group were evaluated by real-time PCR at 1w, 2w, 3w, respectively. The chondrogenic differentiation of BMSCs was evaluated by type II collagen at 21d with immunohistochemical staining. The third-passage BMSCs infected with Ad-BMP-2 and Ad-TGF-β3 were suspended and cultured with DBM for 6 days to construct a new type of tissue engineering scaffold to repair full-thickness cartilage lesions in the femur condyles of pig knee, the regenerated tissue was evaluated at 1,2 and 3 months after surgery by gross appearance, H&E, safranin O staining and O'driscoll score. RESULTS: Ad-BMP-2 and Ad-TGF-β3 (BT group infected cells acquired strong type II collagen staining compared with Ad-BMP-2 (B group and Ad-TGF-β3 (T group along. The Ad-BMP-2 and Ad-TGF-β3 infected BMSCs adhered and propagated well in DBM and the new type of tissue engineering scaffold produced hyaline cartilage morphology containing a stronger type II collagen and safranin O staining, the O'driscoll score was higher than other groups. CONCLUSIONS: The DBM compound with Ad-BMP-2 and Ad-TGF-β3 infected BMSCs scaffold has a good biocompatibility and could well induce cartilage regeneration to repair the defects of joint cartilage. This technology may be efficiently employed for cartilage lesions repair in vivo.

  12. The effects of orbital spaceflight on bone histomorphometry and messenger ribonucleic acid levels for bone matrix proteins and skeletal signaling peptides in ovariectomized growing rats

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    Cavolina, J. M.; Evans, G. L.; Harris, S. A.; Zhang, M.; Westerlind, K. C.; Turner, R. T.

    1997-01-01

    A 14-day orbital spaceflight was performed using ovariectomized Fisher 344 rats to determine the combined effects of estrogen deficiency and near weightlessness on tibia radial bone growth and cancellous bone turnover. Twelve ovariectomized rats with established cancellous osteopenia were flown aboard the space shuttle Columbia (STS-62). Thirty ovariectomized rats were housed on earth as ground controls: 12 in animal enclosure modules, 12 in vivarium cages, and 6 killed the day of launch for baseline measurements. An additional 18 ovary-intact rats were housed in vivarium cages as ground controls: 8 rats were killed as baseline controls and the remaining 10 rats were killed 14 days later. Ovariectomy increased periosteal bone formation at the tibia-fibula synostosis; cancellous bone resorption and formation in the secondary spongiosa of the proximal tibial metaphysis; and messenger RNA (mRNA) levels for the prepro-alpha2(1) subunit of type 1 collagen, osteocalcin, transforming growth factor-beta, and insulin-like growth factor I in the contralateral proximal tibial metaphysis and for the collagen subunit in periosteum pooled from tibiae and femora and decreased cancellous bone area. Compared to ovariectomized weight-bearing rats, the flight group experienced decreases in periosteal bone formation, collagen subunit mRNA levels, and cancellous bone area. The flight rats had a small decrease in the cancellous mineral apposition rate, but no change in the calculated bone formation rate. Also, spaceflight had no effect on cancellous osteoblast and osteoclast perimeters or on mRNA levels for bone matrix proteins and signaling peptides. On the other hand, spaceflight resulted in an increase in bone resorption, as ascertained from the diminished retention of a preflight fluorochrome label. This latter finding suggests that osteoclast activity was increased. In a follow-up ground-based experiment, unilateral sciatic neurotomy of ovariectomized rats resulted in cancellous

  13. Increased serum cartilage oligomeric matrix protein levels and decreased patellar bone mineral density in patients with chondromalacia patellae.

    Science.gov (United States)

    Murphy, E; FitzGerald, O; Saxne, T; Bresnihan, B

    2002-11-01

    Chondromalacia patellae is a potentially disabling disorder characterised by features of patellar cartilage degradation. To evaluate markers of cartilage and bone turnover in patients with chondromalacia patellae. 18 patients with chondromalacia patellae were studied. Serum cartilage oligomeric matrix protein (s-COMP) and bone sialoprotein (s-BSP) levels were measured by enzyme linked immunosorbent assay (ELISA) and compared with those of age and sex matched healthy control subjects. Periarticular bone mineral density (BMD) of both knee joints was assessed by dual energy x ray absorptiometry (DXA). s-COMP levels were significantly raised in all patients with chondromalacia patellae compared with healthy control subjects (p=0.0001). s-BSP levels did not differ significantly between the groups (p=0.41). BMD of the patella was significantly reduced in patients with chondromalacia patellae compared with the control subjects (p=0.016). In patients with bilateral chondromalacia patellae, BMD of the patella was lower in the more symptomatic knee joint (p=0.005). Changes in periarticular BMD were localised to the patella and were not present in femoral regions. Neither s-COMP (p=0.18) nor s-BSP (p=0.40) levels correlated with patellar BMD. Increased s-COMP levels, reflecting cartilage degradation, and reduced BMD localised to the patella may represent clinically useful markers in the diagnosis and monitoring of patients with chondromalacia patellae. Measures of cartilage degradation did not correlate with loss of patellar bone density, suggesting dissociated pathophysiological mechanisms.

  14. Increased expression of osteonectin and osteopontin, two bone matrix proteins, in human breast cancer.

    OpenAIRE

    Bellahcène, A.; Castronovo, V

    1995-01-01

    Microcalcifications are a common phenomenon associated with breast cancer and are often the only mammographic sign of a malignant breast disease. Although microcalcifications are not restricted to breast cancer and can be also associated with benign lesions, it is noteworthy that they are composed exclusively of hydroxyapatite in breast carcinoma. Hydroxyapatite is the bone-associated phosphocalcic crystal the deposition of which in bone tissue requires the coordinated expression of several m...

  15. Decellularized bone matrix grafts for calvaria regeneration

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    Lee, Dong Joon; Diachina, Shannon; Lee, Yan Ting; Zhao, Lixing; Zou, Rui; Tang, Na; Han, Han; Chen, Xin; Ko, Ching-Chang

    2016-01-01

    Decellularization is a promising new method to prepare natural matrices for tissue regeneration. Successful decellularization has been reported using various tissues including skin, tendon, and cartilage, though studies using hard tissue such as bone are lacking. In this study, we aimed to define the optimal experimental parameters to decellularize natural bone matrix using 0.5% sodium dodecyl sulfate and 0.1% NH4OH. Then, the effects of decellularized bone matrix on rat mesenchymal stem cell proliferation, osteogenic gene expression, and osteogenic differentiations in a two-dimensional culture system were investigated. Decellularized bone was also evaluated with regard to cytotoxicity, biochemical, and mechanical characteristics in vitro. Evidence of complete decellularization was shown through hematoxylin and eosin staining and DNA measurements. Decellularized bone matrix displayed a cytocompatible property, conserved structure, mechanical strength, and mineral content comparable to natural bone. To study new bone formation, implantation of decellularized bone matrix particles seeded with rat mesenchymal stem cells was conducted using an orthotopic in vivo model. After 3 months post-implantation into a critical-sized defect in rat calvaria, new bone was formed around decellularized bone matrix particles and also merged with new bone between decellularized bone matrix particles. New bone formation was analyzed with micro computed tomography, mineral apposition rate, and histomorphometry. Decellularized bone matrix stimulated mesenchymal stem cell proliferation and osteogenic differentiation in vitro and in vivo, achieving effective bone regeneration and thereby serving as a promising biological bone graft. PMID:28228929

  16. Decellularized bone matrix grafts for calvaria regeneration

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    Dong Joon Lee

    2016-12-01

    Full Text Available Decellularization is a promising new method to prepare natural matrices for tissue regeneration. Successful decellularization has been reported using various tissues including skin, tendon, and cartilage, though studies using hard tissue such as bone are lacking. In this study, we aimed to define the optimal experimental parameters to decellularize natural bone matrix using 0.5% sodium dodecyl sulfate and 0.1% NH4OH. Then, the effects of decellularized bone matrix on rat mesenchymal stem cell proliferation, osteogenic gene expression, and osteogenic differentiations in a two-dimensional culture system were investigated. Decellularized bone was also evaluated with regard to cytotoxicity, biochemical, and mechanical characteristics in vitro. Evidence of complete decellularization was shown through hematoxylin and eosin staining and DNA measurements. Decellularized bone matrix displayed a cytocompatible property, conserved structure, mechanical strength, and mineral content comparable to natural bone. To study new bone formation, implantation of decellularized bone matrix particles seeded with rat mesenchymal stem cells was conducted using an orthotopic in vivo model. After 3 months post-implantation into a critical-sized defect in rat calvaria, new bone was formed around decellularized bone matrix particles and also merged with new bone between decellularized bone matrix particles. New bone formation was analyzed with micro computed tomography, mineral apposition rate, and histomorphometry. Decellularized bone matrix stimulated mesenchymal stem cell proliferation and osteogenic differentiation in vitro and in vivo, achieving effective bone regeneration and thereby serving as a promising biological bone graft.

  17. The NH2-terminal and COOH-terminal fragments of dentin matrix protein 1 (DMP1) localize differently in the compartments of dentin and growth plate of bone.

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    Maciejewska, Izabela; Cowan, Cameron; Svoboda, Kathy; Butler, William T; D'Souza, Rena; Qin, Chunlin

    2009-02-01

    Multiple studies have shown that dentin matrix protein 1 (DMP1) is essential for bone and dentin mineralization. After post-translational proteolytic cleavage, DMP1 exists within the extracellular matrix of bone and dentin as an NH2-terminal fragment, a COOH-terminal fragment, and the proteoglycan form of the NH2-terminal fragment (DMP1-PG). To begin to assess the biological function of each fragment, we evaluated the distribution of both fragments in the rat tooth and bone using antibodies specific to the NH2-terminal and COOH-terminal regions of DMP1 and confocal microscopy. In rat first molar organs, the NH2-terminal fragment localized to predentin, whereas the COOH-terminal fragment was mainly restricted to mineralized dentin. In the growth plate of bone, the NH2-terminal fragment appeared in the proliferation and hypertrophic zones, whereas the COOH-terminal fragment occupied the ossification zone. Forster resonance energy transfer analysis showed colocalization of both fragments of DMP1 in odontoblasts and predentin, as well as hypertrophic chondrocytes within the growth plates of bone. The biochemical analysis of bovine teeth showed that predentin is rich in DMP1-PG, whereas mineralized dentin primarily contains the COOH-terminal fragment. We conclude that the differential patterns of expression of NH2-terminal and COOH-terminal fragments of DMP1 reflect their potentially distinct roles in the biomineralization of dentin and bone matrices.

  18. Osteomimicry of mammary adenocarcinoma cells in vitro; increased expression of bone matrix proteins and proliferation within a 3D collagen environment.

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    Rachel F Cox

    Full Text Available Bone is the most common site of metastasis for breast cancer, however the reasons for this remain unclear. We hypothesise that under certain conditions mammary cells possess osteomimetic capabilities that may allow them to adapt to, and flourish within, the bone microenvironment. Mammary cells are known to calcify within breast tissue and we have recently reported a novel in vitro model of mammary mineralization using murine mammary adenocarcinoma 4T1 cells. In this study, the osteomimetic properties of the mammary adenocarcinoma cell line and the conditions required to induce mineralization were characterized extensively. It was found that exogenous organic phosphate and inorganic phosphate induce mineralization in a dose dependent manner in 4T1 cells. Ascorbic acid and dexamethasone alone have no effect. 4T1 cells also show enhanced mineralization in response to bone morphogenetic protein 2 in the presence of phosphate supplemented media. The expression of several bone matrix proteins were monitored throughout the process of mineralization and increased expression of collagen type 1 and bone sialoprotein were detected, as determined by real-time RT-PCR. In addition, we have shown for the first time that 3D collagen glycosaminoglycan scaffolds, bioengineered to represent the bone microenvironment, are capable of supporting the growth and mineralization of 4T1 adenocarcinoma cells. These 3D scaffolds represent a novel model system for the study of mammary mineralization and bone metastasis. This work demonstrates that mammary cells are capable of osteomimicry, which may ultimately contribute to their ability to preferentially metastasize to, survive within and colonize the bone microenvironment.

  19. Purification of matrix Gla protein from a marine teleost fish, Argyrosomus regius: calcified cartilage and not bone as the primary site of MGP accumulation in fish.

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    Simes, D C; Williamson, M K; Ortiz-Delgado, J B; Viegas, C S B; Price, P A; Cancela, M L

    2003-02-01

    Matrix Gla protein (MGP) belongs to the family of vitamin K-dependent, Gla-containing proteins, and in mammals, birds, and Xenopus, its mRNA was previously detected in extracts of bone, cartilage, and soft tissues (mainly heart and kidney), whereas the protein was found to accumulate mainly in bone. However, at that time, it was not evaluated if this accumulation originated from protein synthesized in cartilage or in bone cells because both coexist in skeletal structures of higher vertebrates and Xenopus. Later reports showed that MGP also accumulated in costal calcified cartilage as well as at sites of heart valves and arterial calcification. Interestingly, MGP was also found to accumulate in vertebra of shark, a cartilaginous fish. However, to date, no information is available on sites of MGP expression or accumulation in teleost fishes, the ancestors of terrestrial vertebrates, who have in their skeleton mineralized structures with both bone and calcified cartilage. To analyze MGP structure and function in bony fish, MGP was acid-extracted from the mineralized matrix of either bone tissue (vertebra) or calcified cartilage (branchial arches) from the bony fish, Argyrosomus regius, separated from the mineral phase by dialysis, and purified by Sephacryl S-100 chromatography. No MGP was recovered from bone tissue, whereas a protein peak corresponding to the MGP position in this type of gel filtration was obtained from an extract of branchial arches, rich in calcified cartilage. MGP was identified by N-terminal amino acid sequence analysis, and the resulting protein sequence was used to design specific oligonucleotides suitable to amplify the corresponding DNA by a mixture of reverse transcription-polymerase chain reaction (RT-PCR) and 5'rapid amplification of cDNA (RACE)-PCR. In parallel, ArBGP (bone Gla protein, osteocalcin) was also identified in the same fish, and its complementary DNA cloned by an identical procedure. Tissue distribution/accumulation was

  20. Extracellular Matrix Proteins, Alkaline Phosphatase and Pyrophosphate as Molecular Determinants of Bone, Tooth, Kidney and Vascular Calcification

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    McKee, Marc D.

    2008-09-01

    Progress in biomineralization research in recent years has identified, characterized and described functions for key noncollagenous extracellular matrix proteins regulating crystal growth in the skeleton and dentition. Some of these same proteins expressed in soft tissues undergoing pathologic calcification also inhibit ectopic crystal growth. In addition to extracellular matrix proteins regulating matrix mineralization, the enzyme tissue-nonspecific alkaline phosphatase—which is highly expressed by cells in mineralized tissues—cleaves pyrophosphate, an anionic small-molecule inhibitor of mineralization. Together with the required mineral ion availability necessary for crystal growth, these molecular determinants appear to function in limiting the spread of pathologic calcification seen in soft tissues such as blood vessels and kidneys. Osteopontin, in particular, is a potent calcification inhibitor that accumulates in mineralized tissues and in calcified deposits during vascular calcification and nephrolithiasis/urolithiasis. Additional research is required to establish the exact temporal sequence in which the molecular determinants of pathologic calcification appear relative to mineral crystal growth in different tissues, and to establish their relationship (if any) to the activation of osteogenic differentiation programs.

  1. Extracellular Matrix Proteins

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    Linda Christian Carrijo-Carvalho

    2012-01-01

    Full Text Available Lipocalin family members have been implicated in development, regeneration, and pathological processes, but their roles are unclear. Interestingly, these proteins are found abundant in the venom of the Lonomia obliqua caterpillar. Lipocalins are β-barrel proteins, which have three conserved motifs in their amino acid sequence. One of these motifs was shown to be a sequence signature involved in cell modulation. The aim of this study is to investigate the effects of a synthetic peptide comprising the lipocalin sequence motif in fibroblasts. This peptide suppressed caspase 3 activity and upregulated Bcl-2 and Ki-67, but did not interfere with GPCR calcium mobilization. Fibroblast responses also involved increased expression of proinflammatory mediators. Increase of extracellular matrix proteins, such as collagen, fibronectin, and tenascin, was observed. Increase in collagen content was also observed in vivo. Results indicate that modulation effects displayed by lipocalins through this sequence motif involve cell survival, extracellular matrix remodeling, and cytokine signaling. Such effects can be related to the lipocalin roles in disease, development, and tissue repair.

  2. Adsorption of enamel matrix proteins to a bovine-derived bone grafting material and its regulation of cell adhesion, proliferation, and differentiation.

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    Miron, Richard J; Bosshardt, Dieter D; Hedbom, Erik; Zhang, Yufeng; Haenni, Beat; Buser, Daniel; Sculean, Anton

    2012-07-01

    The use of various combinations of enamel matrix derivative (EMD) and grafting materials has been shown to promote periodontal wound healing/regeneration. However, the downstream cellular behavior of periodontal ligament (PDL) cells and osteoblasts has not yet been studied. Furthermore, it is unknown to what extent the bleeding during regenerative surgery may influence the adsorption of exogenous proteins to the surface of bone grafting materials and the subsequent cellular behavior. In the present study, the aim is to test EMD adsorption to the surface of natural bone mineral (NBM) particles in the presence of blood and determine the effect of EMD coating to NBM particles on downstream cellular pathways, such as adhesion, proliferation, and differentiation of primary human osteoblasts and PDL cells. NBM particles were precoated in various settings with EMD or human blood and analyzed for protein adsorption patterns via fluorescent imaging and high-resolution immunocytochemistry with an anti-EMD antibody. Cell attachment and cell proliferation were quantified using fluorescent double-stranded DNA-binding dye. Cell differentiation was analyzed using real-time polymerase chain reaction for genes encoding runt-related transcription factor 2, alkaline phosphatase (ALP), osteocalcin (OC), and collagen1α1 (COL1A1), and mineralization was assessed using red dye staining. Analysis of cell attachment and cell proliferation revealed significantly higher osteoblast and PDL cell attachment on EMD-coated surfaces when compared with control and blood-coated surfaces. EMD also stimulated release of growth factors and cytokines, including bone morphogenetic protein 2 and transforming growth factor β1. Moreover, there were significantly higher mRNA levels of osteoblast differentiation markers, including COL1A1, ALP, and OC, in osteoblasts and PDL cells cultured on EMD-coated NBM particles. The present results suggest that 1) EMD enhances osteoblast and PDL cell attachment

  3. Biomimetically Enhanced Demineralized Bone Matrix for Bone Regenerative Applications

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    Sriram eRavindran

    2015-10-01

    Full Text Available Demineralized bone matrix (DBM is one of the most widely used bone graft materials in dentistry. However, the ability of DBM to reliably and predictably induce bone regeneration has always been a cause for concern. The quality of DBM varies greatly depending on several donor dependent factors and also manufacturing techniques. In order to standardize the quality and to enable reliable and predictable bone regeneration, we have generated a biomimetically-enhanced version of DBM (BE-DBM using clinical grade commercial DBM as a control. We have generated the BE-DBM by incorporating a cell-derived pro-osteogenic extracellular matrix (ECM within clinical grade DBM. In the present study, we have characterized the BE-DBM and evaluated its ability to induce osteogenic differentiation of human marrow derived stromal cells (HMSCs with respect to clinical grade commercial DBM. Our results indicate that the BE-DBM contains significantly more pro-osteogenic factors than DBM and enhances HMSC differentiation and mineralized matrix formation in vitro and in vivo. Based on our results, we envision that the BE-DBM has the potential to replace DBM as the bone graft material of choice.

  4. Enhanced hyaline cartilage matrix synthesis in collagen sponge scaffolds by using siRNA to stabilize chondrocytes phenotype cultured with bone morphogenetic protein-2 under hypoxia.

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    Legendre, Florence; Ollitrault, David; Hervieu, Magalie; Baugé, Catherine; Maneix, Laure; Goux, Didier; Chajra, Hanane; Mallein-Gerin, Frédéric; Boumediene, Karim; Galera, Philippe; Demoor, Magali

    2013-07-01

    Cartilage healing by tissue engineering is an alternative strategy to reconstitute functional tissue after trauma or age-related degeneration. However, chondrocytes, the major player in cartilage homeostasis, do not self-regenerate efficiently and lose their phenotype during osteoarthritis. This process is called dedifferentiation and also occurs during the first expansion step of autologous chondrocyte implantation (ACI). To ensure successful ACI therapy, chondrocytes must be differentiated and capable of synthesizing hyaline cartilage matrix molecules. We therefore developed a safe procedure for redifferentiating human chondrocytes by combining appropriate physicochemical factors: hypoxic conditions, collagen scaffolds, chondrogenic factors (bone morphogenetic protein-2 [BMP-2], and insulin-like growth factor I [IGF-I]) and RNA interference targeting the COL1A1 gene. Redifferentiation of dedifferentiated chondrocytes was evaluated using gene/protein analyses to identify the chondrocyte phenotypic profile. In our conditions, under BMP-2 treatment, redifferentiated and metabolically active chondrocytes synthesized a hyaline-like cartilage matrix characterized by type IIB collagen and aggrecan molecules without any sign of hypertrophy or osteogenesis. In contrast, IGF-I increased both specific and noncharacteristic markers (collagens I and X) of chondrocytes. The specific increase in COL2A1 gene expression observed in the BMP-2 treatment was shown to involve the specific enhancer region of COL2A1 that binds the trans-activators Sox9/L-Sox5/Sox6 and Sp1, which are associated with a decrease in the trans-inhibitors of COL2A1, c-Krox, and p65 subunit of NF-kappaB. Our procedure in which BMP-2 treatment under hypoxia is associated with a COL1A1 siRNA, significantly increased the differentiation index of chondrocytes, and should offer the opportunity to develop new ACI-based therapies in humans.

  5. The effects of bone morphogenetic protein-2 and enamel matrix derivative on the bioactivity of mineral trioxide aggregate in MC3T3-E1cells

    Directory of Open Access Journals (Sweden)

    Youngdan Jeong

    2014-08-01

    Full Text Available Objectives The effects of bone morphogenetic protein-2 (BMP-2 and enamel matrix derivative (EMD respectively with mineral trioxide aggregate (MTA on hard tissue regeneration have been investigated in previous studies. This study aimed to compare the osteogenic effects of MTA/BMP-2 and MTA/EMD treatment in MC3T3-E1 cells. Materials and Methods MC3T3-E1 cells were treated with MTA (ProRoot, Dentsply, BMP-2 (R&D Systems, EMD (Emdogain, Straumann separately and MTA/BMP-2 or MTA/EMD combination. Mineralization was evaluated by staining the calcium deposits with alkaline phosphatase (ALP, Sigma-Aldrich and Alizarin red (Sigma-Aldrich. The effects on the osteoblast differentiation were evaluated by the expressions of osteogenic markers, including ALP, bone sialoprotein (BSP, osteocalcin (OCN, osteopontin (OPN and osteonectin (OSN, as determined by reverse-transcription polymerase chain reaction analysis (RT-PCR, AccuPower PCR, Bioneer. Results Mineralization increased in the BMP-2 and MTA/BMP-2 groups and increased to a lesser extent in the MTA/EMD group but appeared to decrease in the MTA-only group based on Alizarin red staining. ALP expression largely decreased in the EMD and MTA/EMD groups based on ALP staining. In the MTA/BMP-2 group, mRNA expression of OPN on day 3 and BSP and OCN on day 7 significantly increased. In the MTA/EMD group, OSN and OCN gene expression significantly increased on day 7, whereas ALP expression decreased on days 3 and 7 (p < 0.05. Conclusions These results suggest the MTA/BMP-2 combination promoted more rapid differentiation in MC3T3-E1 cells than did MTA/EMD during the early mineralization period.

  6. [Actin cytoskeleton organization and spreading of bone marrow stromal cells and cartilage cells during their combined and independent cultivation on different extracellular matrix proteins].

    Science.gov (United States)

    Sakhenberg, E I; Nikolaenko, N S; Pinaev, G P

    2014-01-01

    To clarify the mutual influence of bone marrow stromal cells (BMSCs) and cartilage cells we studied the organization of their actin cytoskeleton and cell spreading on different extracellular matrix proteins--laminin 2/4, collagen type I or fibronectin. It has been shown that the most pronounced difference in morphological characteristics of the cells such as their form, size and actin cytoskeleton organization occur in the case of interaction with fibronectin. So, after separate brief incubation of both cell types on fibronectin, the average area of BMSCs spreading was about 4 times greater than the area of the cartilage cell spreading. However, in the co-culture of these cells in a ratio of 1:1, the average jointed spreading area on fibronctin was nearly 1.5 times less than the theoretically calculated. To determine the nature of exposure of the cells to each other we have studied spreading of these cells in the media conditioned by another cell type. We have found that the area of BMSC's spreading in the medium conditioned by cartilage cells is markedly smaller than the area of spreading of the same cells in the control medium. These data suggest that the cartilage cells secrete factors that reduce BMSC's spreading.

  7. Bone morphogenetic proteins: Periodontal regeneration

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    Subramaniam M Rao

    2013-01-01

    Full Text Available Periodontitis is an infectious inflammatory disease that results in attachment loss and bone loss. Regeneration of the periodontal tissues entails de novo formation of cementum, periodontal ligament, and alveolar bone. Several different approaches are currently being explored to achieve complete, reliable, and reproducible regeneration of periodontal tissues. The therapeutic management of new bone formation is one of the key issues in successful periodontal regeneration. Bone morphogenetic proteins form a unique group of proteins within the transforming growth factor superfamily of genes and have a vital role in the regulation in the bone induction and maintenance. The activity of bone morphogenetic proteins was first identified in the 1960s, but the proteins responsible for bone induction were unknown until the purification and cloning of human bone morphogenetic proteins in the 1980s, because of their osteoinductive potential. Bone morphogenetic proteins have gained a lot of interest as therapeutic agents for treating periodontal defects. A systematic search for data related to the use of bone morphogenetic proteins for the regeneration of periodontal defects was performed to recognize studies on animals and human (PUBMED, MEDLINE, COCHRANE, and Google search. All the studies included showed noticeable regeneration of periodontal tissues with the use of BMP.

  8. The Expression of Bone Morphogenetic Protein 2 and Matrix Metalloproteinase 2 through Retinoic Acid Receptor Beta Induced by All-Trans Retinoic Acid in Cultured ARPE-19 Cells.

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    Zhenya Gao

    Full Text Available All-trans retinoic acid (ATRA plays an important role in ocular development. Previous studies found that retinoic acid could influence the metabolism of scleral remodeling by promoting retinal pigment epithelium (RPE cells to secrete secondary signaling factors. The purpose of this study was to investigate whether retinoic acid affected secretion of bone morphogenetic protein 2 (BMP-2 and matrix metalloproteinase 2 (MMP-2 and to explore the signaling pathway of retinoic acid in cultured acute retinal pigment epithelial 19 (ARPE-19 cells.The effects of ATRA (concentrations from 10-9 to 10-5 mol/l on the expression of retinoic acid receptors (RARs in ARPE-19 cells were examined at the mRNA and protein levels using reverse transcription-polymerase chain reaction (RT-PCR and western blot assay, respectively. The effects of treating ARPE-19 cells with ATRA concentrations ranging from 10-9 to 10-5 mol/l for 24 h and 48 h or with 10-6mol/l ATRA at different times ranging from 6h to 72h were assessed using real-time quantitative PCR (qPCR and enzyme-linked immunosorbent assay (ELISA. The contribution of RARβ-induced activation of ARPE-19 cells was confirmed using LE135, an antagonist of RARβ.RARβ mRNA levels significantly increased in the ARPE-19 cells treated with ATRA for 24h and 48h. These increases in RARβ mRNA levels were dose dependent (at concentrations of 10-9 to 10-5 mol/l with a maximum effect observed at 10-6 mol/l. There were no significant changes in the mRNA levels of RARα and RARγ. Western blot assay revealed that RARβ protein levels were increased significantly in a time-dependent manner in ARPE-19 cells treated with 10-6 mol/l ATRA from 12 h to 72 h, with a marked increase observed at 24 h and 48 h. The upregulation of RARβ and the ATRA-induced secretion in ARPE-19 cells could be inhibited by the RARβ antagonist LE135.ATRA induced upregulation of RARβ in ARPE-19 cells and stimulated these cells to secrete BMP-2 and MMP-2.

  9. Biomineralization of a Self-Assembled Extracellular Matrix for Bone Tissue Engineering

    OpenAIRE

    Meng, Yizhi; Qin, Yi-Xian; DiMasi, Elaine; Ba, Xiaolan; Rafailovich, Miriam; Pernodet, Nadine

    2008-01-01

    Understanding how biomineralization occurs in the extracellular matrix (ECM) of bone cells is crucial to the understanding of bone formation and the development of a successfully engineered bone tissue scaffold. It is still unclear how ECM mechanical properties affect protein-mineral interactions in early stages of bone mineralization. We investigated the longitudinal mineralization properties of MC3T3-E1 cells and the elastic modulus of their ECM using shear modulation force microscopy, sync...

  10. Microspectroscopic evidence of cretaceous bone proteins.

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    Johan Lindgren

    Full Text Available Low concentrations of the structural protein collagen have recently been reported in dinosaur fossils based primarily on mass spectrometric analyses of whole bone extracts. However, direct spectroscopic characterization of isolated fibrous bone tissues, a crucial test of hypotheses of biomolecular preservation over deep time, has not been performed. Here, we demonstrate that endogenous proteinaceous molecules are retained in a humerus from a Late Cretaceous mosasaur (an extinct giant marine lizard. In situ immunofluorescence of demineralized bone extracts shows reactivity to antibodies raised against type I collagen, and amino acid analyses of soluble proteins extracted from the bone exhibit a composition indicative of structural proteins or their breakdown products. These data are corroborated by synchrotron radiation-based infrared microspectroscopic studies demonstrating that amino acid containing matter is located in bone matrix fibrils that express imprints of the characteristic 67 nm D-periodicity typical of collagen. Moreover, the fibrils differ significantly in spectral signature from those of potential modern bacterial contaminants, such as biofilms and collagen-like proteins. Thus, the preservation of primary soft tissues and biomolecules is not limited to large-sized bones buried in fluvial sandstone environments, but also occurs in relatively small-sized skeletal elements deposited in marine sediments.

  11. Comparison of Demineralized Dentin and Demineralized Freeze Dried Bone as Carriers for Enamel Matrix Proteins in a Rat Critical Size Defect

    Science.gov (United States)

    2005-05-01

    usually requires resorption prior to the formation of new bone. At this early two weeks stage it is possible that there was insufficient time for the... resorptive and formative processes to be completed. At eight weeks the mean RIDIT values for all groups were greater than 0.5, 49 indicating that the...non-collageous proteins of rat incisors dentin. Calcified Tissue Research 1978;25:169-178. Finkelman R., Mohan S., Jennings J., Taylor A., Jepsen S

  12. Effect of rhBMP-2 Immobilized Anorganic Bovine Bone Matrix on Bone Regeneration

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    Jung-Bo Huh

    2015-07-01

    Full Text Available Anorganic bovine bone matrix (Bio-Oss® has been used for a long time for bone graft regeneration, but has poor osteoinductive capability. The use of recombinant human bone morphogenetic protein-2 (rhBMP-2 has been suggested to overcome this limitation of Bio-Oss®. In the present study, heparin-mediated rhBMP-2 was combined with Bio-Oss® in animal experiments to investigate bone formation performance; heparin was used to control rhBMP-2 release. Two calvarial defects (8 mm diameter were formed in a white rabbit model and then implanted or not (controls with Bio-Oss® or BMP-2/Bio-Oss®. The Bio-Oss® and BMP-2/Bio-Oss® groups had significantly greater new bone areas (expressed as percentages of augmented areas than the non-implanted controls at four and eight weeks after surgery, and the BMP-2/Bio-Oss® group (16.50 ± 2.87 (n = 6 had significantly greater new bone areas than the Bio-Oss® group (9.43 ± 3.73 (n = 6 at four weeks. These findings suggest that rhBMP-2 treated heparinized Bio-Oss® markedly enhances bone regeneration.

  13. Osteoblast Differentiation and Bone Matrix Formation In Vivo and In Vitro.

    Science.gov (United States)

    Blair, Harry C; Larrouture, Quitterie C; Li, Yanan; Lin, Hang; Beer-Stoltz, Donna; Liu, Li; Tuan, Rocky S; Robinson, Lisa J; Schlesinger, Paul H; Nelson, Deborah J

    2017-06-01

    We review the characteristics of osteoblast differentiation and bone matrix synthesis. Bone in air breathing vertebrates is a specialized tissue that developmentally replaces simpler solid tissues, usually cartilage. Bone is a living organ bounded by a layer of osteoblasts that, because of transport and compartmentalization requirements, produce bone matrix exclusively as an organized tight epithelium. With matrix growth, osteoblasts are reorganized and incorporated into the matrix as living cells, osteocytes, which communicate with each other and surface epithelium by cell processes within canaliculi in the matrix. The osteoblasts secrete the organic matrix, which are dense collagen layers that alternate parallel and orthogonal to the axis of stress loading. Into this matrix is deposited extremely dense hydroxyapatite-based mineral driven by both active and passive transport and pH control. As the matrix matures, hydroxyapatite microcrystals are organized into a sophisticated composite in the collagen layer by nucleation in the protein lattice. Recent studies on differentiating osteoblast precursors revealed a sophisticated proton export network driving mineralization, a gene expression program organized with the compartmentalization of the osteoblast epithelium that produces the mature bone matrix composite, despite varying serum calcium and phosphate. Key issues not well defined include how new osteoblasts are incorporated in the epithelial layer, replacing those incorporated in the accumulating matrix. Development of bone in vitro is the subject of numerous projects using various matrices and mesenchymal stem cell-derived preparations in bioreactors. These preparations reflect the structure of bone to variable extents, and include cells at many different stages of differentiation. Major challenges are production of bone matrix approaching the in vivo density and support for trabecular bone formation. In vitro differentiation is limited by the organization and

  14. Influence of heat stress to matrix on bone formation.

    Science.gov (United States)

    Yoshida, Keiko; Uoshima, Katsumi; Oda, Kimimitsu; Maeda, Takeyasu

    2009-08-01

    It is important to know the etiology of implant failure. It has been reported that heat stress during drilling was one of the causes for failure and the threshold was 47 degrees C. However, clinically, we encounter cases in which overheating does not seem to affect osseointegration eventually. The purpose of this study was to assess histologically the spatio-temporal effect of heat stress on bone formation after overheating the bone matrix. Rat calvarial bone was heated to 37 degrees C, 43 degrees C, 45 degrees C and 48 degrees C for 15 min by a temperature stimulator. Paraffin sections were prepared 1, 3 and 5 weeks after heating and investigated histologically under light microscopy. Hematoxylin and eosin staining, alkaline phosphatase (ALP), osteopontin (OPN), heat shock protein 27 (Hsp27) and heat shock protein 70 (Hsp70) immunohistochemistry and tartrate-resistant acid phosphatase (TRAP) enzyme histochemistry were carried out. The area of dead osteocytes was calculated and statistically analyzed. Apoptotic osteocytes were detected by the terminal deoxynucleotidyltransferase-mediated dUTP nick end-labeling (TUNEL) method. Along with the temperature increase, the area of dead osteocytes increased and regeneration of the periosteal membrane was delayed. Hsps- and TUNEL-positive cells were only seen in the 48 degrees C group. Spatio-temporal changes of TRAP- and ALP-positive cell numbers were observed, while OPN expression was mostly absent. Even after 48 degrees C stimulation, bone formation on the calvarial surface was observed after 5 weeks. Although there was a temperature-dependent delay in bone formation after heat stress, the 48 degrees C heat stress did not obstruct bone formation eventually. This delay was probably caused by slow periosteal membrane regeneration.

  15. Osteogenic Matrix Cell Sheets Facilitate Osteogenesis in Irradiated Rat Bone

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    Yoshinobu Uchihara

    2015-01-01

    Full Text Available Reconstruction of large bone defects after resection of malignant musculoskeletal tumors is a significant challenge in orthopedic surgery. Extracorporeal autogenous irradiated bone grafting is a treatment option for bone reconstruction. However, nonunion often occurs because the osteogenic capacity is lost by irradiation. In the present study, we established an autogenous irradiated bone graft model in the rat femur to assess whether osteogenic matrix cell sheets improve osteogenesis of the irradiated bone. Osteogenic matrix cell sheets were prepared from bone marrow-derived stromal cells and co-transplanted with irradiated bone. X-ray images at 4 weeks after transplantation showed bridging callus formation around the irradiated bone. Micro-computed tomography images at 12 weeks postoperatively showed abundant callus formation in the whole circumference of the irradiated bone. Histology showed bone union between the irradiated bone and host femur. Mechanical testing showed that the failure force at the irradiated bone site was significantly higher than in the control group. Our study indicates that osteogenic matrix cell sheet transplantation might be a powerful method to facilitate osteogenesis in irradiated bones, which may become a treatment option for reconstruction of bone defects after resection of malignant musculoskeletal tumors.

  16. Glycosylation of Dentin Matrix Protein 1 is critical for osteogenesis.

    Science.gov (United States)

    Sun, Yao; Weng, Yuteng; Zhang, Chenyang; Liu, Yi; Kang, Chen; Liu, Zhongshuang; Jing, Bo; Zhang, Qi; Wang, Zuolin

    2015-12-04

    Proteoglycans play important roles in regulating osteogenesis. Dentin matrix protein 1 (DMP1) is a highly expressed bone extracellular matrix protein that regulates both bone development and phosphate metabolism. After glycosylation, an N-terminal fragment of DMP1 protein was identified as a new proteoglycan (DMP1-PG) in bone matrix. In vitro investigations showed that Ser(89) is the key glycosylation site in mouse DMP1. However, the specific role of DMP1 glycosylation is still not understood. In this study, a mutant DMP1 mouse model was developed in which the glycosylation site S(89) was substituted with G(89) (S89G-DMP1). The glycosylation level of DMP1 was down-regulated in the bone matrix of S89G-DMP1 mice. Compared with wild type mice, the long bones of S89G-DMP1 mice showed developmental changes, including the speed of bone remodeling and mineralization, the morphology and activities of osteocytes, and activities of both osteoblasts and osteoclasts. These findings indicate that glycosylation of DMP1 is a key posttranslational modification process during development and that DMP1-PG functions as an indispensable proteoglycan in osteogenesis.

  17. The "love-hate" relationship between osteoclasts and bone matrix.

    Science.gov (United States)

    Rucci, Nadia; Teti, Anna

    2016-01-01

    Osteoclasts are unique cells that destroy the mineralized matrix of the skeleton. There is a "love-hate" relationship between the osteoclasts and the bone matrix, whereby the osteoclast is stimulated by the contact with the matrix but, at the same time, it disrupts the matrix, which, in turn, counteracts this disruption by some of its components. The balance between these concerted events brings about bone resorption to be controlled and to contribute to bone tissue integrity and skeletal health. The matrix components released by osteoclasts are also involved in the local regulation of other bone cells and in the systemic control of organismal homeostasis. Disruption of this regulatory loop causes bone diseases, which may end up with either reduced or increased bone mass, often associated with poor bone quality. Expanding the knowledge on osteoclast-to-matrix interaction could help to counteract these diseases and improve the human bone health. In this article, we will present evidence of the physical, molecular and regulatory relationships between the osteoclasts and the mineralized matrix, discussing the underlying mechanisms as well as their pathologic alterations and potential targeting.

  18. Mimicking the nanostructure of bone matrix to regenerate bone

    Directory of Open Access Journals (Sweden)

    Robert Kane

    2013-11-01

    Full Text Available Key features of bone tissue structure and composition are capable of directing cellular behavior toward the generation of new bone tissue. Bone tissue, as well as materials derived from bone, have a long and successful history of use as bone grafting materials. Recent developments in design and processing of synthetic scaffolding systems has allowed the replication of the bone's desirable biological activity in easy to fabricate polymeric materials with nano-scale features exposed on the surface. The biological response to these new tissue-engineering scaffold materials oftentimes exceeds that seen on scaffolds produced using biological materials.

  19. The classic: Bone morphogenetic protein.

    Science.gov (United States)

    Urist, Marshall R; Strates, Basil S

    2009-12-01

    This Classic Article is a reprint of the original work by Marshall R. Urist and Basil S. Strates, Bone Morphogenetic Protein. An accompanying biographical sketch of Marshall R. Urist, MD is available at DOI 10.1007/s11999-009-1067-4; a second Classic Article is available at DOI 10.1007/s11999-009-1069-2; and a third Classic Article is available at DOI 10.1007/s11999-009-1070-9. The Classic Article is copyright 1971 by Sage Publications Inc. Journals and is reprinted with permission from Urist MR, Strates BS. Bone morphogenetic protein. J Dent Res. 1971;50:1392-1406.

  20. Immobilization of cross linked Col-I–OPN bone matrix protein on aminolysed PCL surfaces enhances initial biocompatibility of human adipogenic mesenchymal stem cells (hADMSC)

    Energy Technology Data Exchange (ETDEWEB)

    Kim, Young-Hee; Jyoti, Md. Anirban; Song, Ho-Yeon, E-mail: songmic@sch.ac.kr

    2014-06-01

    In bone tissue engineering surface modification is considered as one of the important ways of fabricating successful biocompatible material. Addition of biologically active functionality on the surfaces has been tried for improving the overall biocompatibility of the system. In this study poly-ε-caprolactone film surfaces have been modified through aminolysis and immobilization process. Collagen type I (COL-I) and osteopontin (OPN), which play an important role in osteogenesis, was immobilized onto PCL films followed by aminolysis treatment using 1,6-hexanediamine. Characterization of animolysed and immobilized surfaces were done by a number techniques using scanning electron microscopy (SEM), FT-IR, XPS, ninhydrin staining, SDS-PAGE and confocal microscopy and compared between the modified and un-modified surfaces. Results of the successive experiments showed that aminolysis treatment was homogeneously achieved which helped to entrap or immobilize Col-I–OPN proteins on surfaces of PCL film. In vitro studies with human adipogenic mesenchymal stem cells (hADMSC) also confirmed the attachment and proliferation of cells was better in modified PCL surfaces than the unmodified surfaces. SEM, confocal microscopy and MTT assay showed a significant increase in cell spreading, attachment and proliferations on the biofunctionalized surfaces compared to the unmodified PCL surfaces at all-time points indicating the success of surface biofunctionalization.

  1. Autologous bone graft versus demineralized bone matrix in internal fixation of ununited long bones

    Directory of Open Access Journals (Sweden)

    Rubenbauer Bianka

    2009-12-01

    Full Text Available Abstract Background Non-unions are severe complications in orthopaedic trauma care and occur in 10% of all fractures. The golden standard for the treatment of ununited fractures includes open reduction and internal fixation (ORIF as well as augmentation with autologous-bone-grafting. However, there is morbidity associated with the bone-graft donor site and some patients offer limited quantity or quality of autologous-bone graft material. Since allogene bone-grafts are introduced on the market, this comparative study aims to evaluate healing characteristics of ununited bones treated with ORIF combined with either iliac-crest-autologous-bone-grafting (ICABG or demineralized-bone-matrix (DBM. Methods and results From 2000 to 2006 out of sixty-two consecutive patients with non-unions presenting at our Level I Trauma Center, twenty patients had ununited diaphyseal fractures of long bones and were treated by ORIF combined either by ICABG- (n = 10 or DBM-augmentation (n = 10. At the time of index-operation, patients of the DBM-group had a higher level of comorbidity (ASA-value: p = 0.014. Mean duration of follow-up was 56.6 months (ICABG-group and 41.2 months (DBM-group. All patients were clinically and radiographically assessed and adverse effects related to bone grafting were documented. The results showed that two non-unions augmented with ICABG failed osseous healing (20% whereas all non-unions grafted by DBM showed successful consolidation during the first year after the index operation (p = 0.146. No early complications were documented in both groups but two patients of the ICABG-group suffered long-term problems at the donor site (20% (p = 0.146. Pain intensity were comparable in both groups (p = 0.326. However, patients treated with DBM were more satisfied with the surgical procedure (p = 0.031. Conclusion With the use of DBM, the costs for augmentation of the non-union-site are more expensive compared to ICABG (calculated difference: 160

  2. Autologous bone graft versus demineralized bone matrix in internal fixation of ununited long bones.

    Science.gov (United States)

    Pieske, Oliver; Wittmann, Alexandra; Zaspel, Johannes; Löffler, Thomas; Rubenbauer, Bianka; Trentzsch, Heiko; Piltz, Stefan

    2009-12-15

    Non-unions are severe complications in orthopaedic trauma care and occur in 10% of all fractures. The golden standard for the treatment of ununited fractures includes open reduction and internal fixation (ORIF) as well as augmentation with autologous-bone-grafting. However, there is morbidity associated with the bone-graft donor site and some patients offer limited quantity or quality of autologous-bone graft material. Since allogene bone-grafts are introduced on the market, this comparative study aims to evaluate healing characteristics of ununited bones treated with ORIF combined with either iliac-crest-autologous-bone-grafting (ICABG) or demineralized-bone-matrix (DBM). From 2000 to 2006 out of sixty-two consecutive patients with non-unions presenting at our Level I Trauma Center, twenty patients had ununited diaphyseal fractures of long bones and were treated by ORIF combined either by ICABG- (n = 10) or DBM-augmentation (n = 10). At the time of index-operation, patients of the DBM-group had a higher level of comorbidity (ASA-value: p = 0.014). Mean duration of follow-up was 56.6 months (ICABG-group) and 41.2 months (DBM-group). All patients were clinically and radiographically assessed and adverse effects related to bone grafting were documented. The results showed that two non-unions augmented with ICABG failed osseous healing (20%) whereas all non-unions grafted by DBM showed successful consolidation during the first year after the index operation (p = 0.146). No early complications were documented in both groups but two patients of the ICABG-group suffered long-term problems at the donor site (20%) (p = 0.146). Pain intensity were comparable in both groups (p = 0.326). However, patients treated with DBM were more satisfied with the surgical procedure (p = 0.031). With the use of DBM, the costs for augmentation of the non-union-site are more expensive compared to ICABG (calculated difference: 160 euro/case). Nevertheless, this study demonstrated that the

  3. Antibodies to Staphylococcus aureus Bone Sialoprotein-Binding Protein Indicate Infectious Osteomyelitis▿

    OpenAIRE

    Persson, Lena; Johansson, Christian; Rydén, Cecilia

    2009-01-01

    Discrimination of soft tissue infection from osteomyelitis in diabetic foot infections is a common clinical problem. Staphylococcus aureus isolates from patients with osteomyelitis express bone sialoprotein-binding protein (Bbp) that binds the bone matrix protein bone sialoprotein. The serological assay with Bbp discriminated cases of osteomyelitis from soft tissue infections in patients with diabetic foot ulcers.

  4. Matrix Gla protein and osteocalcin: from gene duplication to neofunctionalization.

    Science.gov (United States)

    Cancela, M Leonor; Laizé, Vincent; Conceição, Natércia

    2014-11-01

    Osteocalcin (OC or bone Gla protein, BGP) and matrix Gla protein (MGP) are two members of the growing family of vitamin K-dependent (VKD) proteins. They were the first VKD proteins found not to be involved in coagulation and synthesized outside the liver. Both proteins were isolated from bone although it is now known that only OC is synthesized by bone cells under normal physiological conditions, but since both proteins can bind calcium and hydroxyapatite, they can also accumulate in bone. Both OC and MGP share similar structural features, both in terms of protein domains and gene organization. OC gene is likely to have appeared from MGP through a tandem gene duplication that occurred concomitantly with the appearance of the bony vertebrates. Despite their relatively close relationship and the fact that both can bind calcium and affect mineralization, their functions are not redundant and they also have other unrelated functions. Interestingly, these two proteins appear to have followed quite different evolutionary strategies in order to acquire novel functionalities, with OC following a gene duplication strategy while MGP variability was obtained mostly by the use of multiple promoters and alternative splicing, leading to proteins with additional functional characteristics and alternative gene regulatory pathways. Copyright © 2014 Elsevier Inc. All rights reserved.

  5. HBM Mice Have Altered Bone Matrix Composition and Improved Material Toughness.

    Science.gov (United States)

    Ross, Ryan D; Mashiatulla, Maleeha; Acerbo, Alvin S; Almer, Jonathan D; Miller, Lisa M; Johnson, Mark L; Sumner, D Rick

    2016-10-01

    The G171V mutation in the low-density lipoprotein receptor-related protein 5 (LRP5) leads to a high bone mass (HBM) phenotype. Studies using HBM transgenic mouse models have consistently found increased bone mass and whole-bone strength, but little attention has been paid to the composition of the bone matrix. The current study sought to determine if the cortical bone matrix composition differs in HBM and wild-type mice and to determine how much of the variance in bone material properties is explained by variance in matrix composition. Consistent with previous studies, HBM mice had greater cortical area, moment of inertia, ultimate force, bending stiffness, and energy to failure than wild-type animals. The increased energy to failure was primarily caused by a large increase in post-yield behavior, with no difference in pre-yield behavior. The HBM mice had increased mineral-to-matrix and collagen cross-link ratios, and decreased crystallinity, carbonate, and acid phosphate substitution as measured by Fourier transform infrared microspectroscopy, but no differences in crystal length, intra-fibular strains, and mineral spacing compared to wild-type controls, as measured by X-ray scattering. The largest between genotype difference in material properties was a twofold increase in the modulus of toughness in HBM mice. Step-wise regression analyses showed that the specific matrix compositional parameters most closely associated with material properties varied between the wild-type and HBM genotypes. Although the mechanisms controlling the paradoxical combination of more mineralized yet tougher bone in HBM mice remain to be fully explained, the findings suggest that LRP5 represents a target to not only build bone mass but also to improve bone quality.

  6. Spine fusion using cell matrix composites enriched in bone marrow-derived cells.

    Science.gov (United States)

    Muschler, George F; Nitto, Hironori; Matsukura, Yoichi; Boehm, Cynthia; Valdevit, Antonio; Kambic, Helen; Davros, William; Powell, Kimerly; Easley, Kirk

    2003-02-01

    Bone marrow-derived cells including osteoblastic progenitors can be concentrated rapidly from bone marrow aspirates using the surface of selected implantable matrices for selective cell attachment. Concentration of cells in this way to produce an enriched cellular composite graft improves graft efficacy. The current study was designed to test the hypothesis that the biologic milieu of a bone marrow clot will significantly improve the efficacy of such a graft. An established posterior spinal fusion model and cancellous bone matrix was used to compare an enriched cellular composite bone graft alone, bone matrix plus bone marrow clot, and an enriched bone matrix composite graft plus bone marrow clot. Union score, quantitative computed tomography, and mechanical testing were used to define outcome. The union score for the enriched bone matrix plus bone marrow clot composite was superior to the enriched bone matrix alone and the bone matrix plus bone marrow clot. The enriched bone matrix plus bone marrow clot composite also was superior to the enriched bone matrix alone in fusion volume and in fusion area. These data confirm that the addition of a bone marrow clot to an enriched cell-matrix composite graft results in significant improvement in graft performance. Enriched composite grafts prepared using this strategy provide a rapid, simple, safe, and inexpensive method for intraoperative concentration and delivery of bone marrow-derived cells and connective tissue progenitors that may improve the outcome of bone grafting.

  7. Bone induction by composite of bioerodible polyorthoester and demineralized bone matrix in rats

    DEFF Research Database (Denmark)

    Pinholt, E M; Solheim, E; Bang, G

    1991-01-01

    A composite of a local, sustained, drug-release system, Alzamer bioerodible polyorthoester, and demineralized bone-matrix (DBM) particles implanted in the abdominal muscle of 89 Wistar rats induced cartilage and bone formation at the same rate as DBM when evaluated histologically and by 85Sr uptake...

  8. Bone induction by composite of bioerodible polyorthoester and deminiralized bone matrix in rats

    Energy Technology Data Exchange (ETDEWEB)

    Pinholt, E.M.; Solheim, E. (Institute for Surgical Research, Rikshospitalet, University of Oslo (Norway)); Bang, G. (Department of Oral Pathology and Forensic Odontology, University of Bergen (Norway)); Sudmann, E. (Hagavik Orthopedic Hospital, University of Bergen (Norway))

    1991-01-01

    A composite of a local, sustained, drug-release system, Alzamer bioerodible polyorthoester, and demineralized bone-matrix (DBM) particles implanted in the abdominal muscle of 89 Wistar rats induced cartilage and bone formation at the same rate as DBM when evaluated histologically and by {sup 85}Sr uptake. The composite implant was technically easier to use than DBM alone. (author).

  9. Positive modulator of bone morphogenic protein-2

    Energy Technology Data Exchange (ETDEWEB)

    Zamora, Paul O.; Pena, Louis A.; Lin, Xinhua; Kazuyuki, Takahashi

    2017-06-06

    Compounds of the present invention of formula I and formula II are disclosed in the specification and wherein the compounds are modulators of Bone Morphogenic Protein activity. Compounds are synthetic peptides having a non-growth factor heparin binding region, a linker, and sequences that bind specifically to a receptor for Bone Morphogenic Protein. Uses of compounds of the present invention in the treatment of bone lesions, degenerative joint disease and to enhance bone formation are disclosed.

  10. Positive modulator of bone morphogenic protein-2

    Science.gov (United States)

    Zamora, Paul O [Gaithersburg, MD; Pena, Louis A [Poquott, NY; Lin, Xinhua [Plainview, NY; Takahashi, Kazuyuki [Germantown, MD

    2009-01-27

    Compounds of the present invention of formula I and formula II are disclosed in the specification and wherein the compounds are modulators of Bone Morphogenic Protein activity. Compounds are synthetic peptides having a non-growth factor heparin binding region, a linker, and sequences that bind specifically to a receptor for Bone Morphogenic Protein. Uses of compounds of the present invention in the treatment of bone lesions, degenerative joint disease and to enhance bone formation are disclosed.

  11. Alveolar Ridge Preservation Using Xenogeneic Collagen Matrix and Bone Allograft

    Directory of Open Access Journals (Sweden)

    Andreas O. Parashis

    2014-01-01

    Full Text Available Alveolar ridge preservation (ARP has been shown to prevent postextraction bone loss. The aim of this report is to highlight the clinical, radiographic, and histological outcomes following use of a bilayer xenogeneic collagen matrix (XCM in combination with freeze-dried bone allograft (FDBA for ARP. Nine patients were treated after extraction of 18 teeth. Following minimal flap elevation and atraumatic extraction, sockets were filled with FDBA. The XCM was adapted to cover the defect and 2-3 mm of adjacent bone and flaps were repositioned. Healing was uneventful in all cases, the XCM remained in place, and any matrix exposure was devoid of further complications. Exposed matrix portions were slowly vascularized and replaced by mature keratinized tissue within 2-3 months. Radiographic and clinical assessment indicated adequate volume of bone for implant placement, with all planned implants placed in acceptable positions. When fixed partial dentures were placed, restorations fulfilled aesthetic demands without requiring further augmentation procedures. Histological and immunohistochemical analysis from 9 sites (4 patients indicated normal mucosa with complete incorporation of the matrix and absence of inflammatory response. The XCM + FDBA combination resulted in minimal complications and desirable soft and hard tissue therapeutic outcomes, suggesting the feasibility of this approach for ARP.

  12. Bioinspired Design of Polycaprolactone Composite Nanofibers as Artificial Bone Extracellular Matrix for Bone Regeneration Application.

    Science.gov (United States)

    Gao, Xiang; Song, Jinlin; Zhang, Yancong; Xu, Xiao; Zhang, Siqi; Ji, Ping; Wei, Shicheng

    2016-10-07

    The design and development of functional biomimetic systems for programmed stem cell response is a field of topical interest. To mimic bone extracellular matrix, we present an innovative strategy for constructing drug-loaded composite nanofibrous scaffolds in this study, which could integrate multiple cues from calcium phosphate mineral, bioactive molecule, and highly ordered fiber topography for the control of stem cell fate. Briefly, inspired by mussel adhesion mechanism, a polydopamine (pDA)-templated nanohydroxyapatite (tHA) was synthesized and then surface-functionalized with bone morphogenetic protein-7-derived peptides via catechol chemistry. Afterward, the resulting peptide-loaded tHA (tHA/pep) particles were blended with polycaprolactone (PCL) solution to fabricate electrospun hybrid nanofibers with random and aligned orientation. Our research demonstrated that the bioactivity of grafted peptides was retained in composite nanofibers. Compared to controls, PCL-tHA/pep composite nanofibers showed improved cytocompatibility. Moreover, the incorporated tHA/pep particles in nanofibers could further facilitate osteogenic differentiation potential of human mesenchymal stem cells (hMSCs). More importantly, the aligned PCL-tHA/pep composite nanofibers showed more osteogenic activity than did randomly oriented counterparts, even under nonosteoinductive conditions, indicating excellent performance of biomimetic design in cell fate decision. After in vivo implantation, the PCL-tHA/pep composite nanofibers with highly ordered structure could significantly promote the regeneration of lamellar-like bones in a rat calvarial critical-sized defect. Accordingly, the presented strategy in our work could be applied for a wide range of potential applications in not only bone regeneration application but also pharmaceutical science.

  13. Enamel matrix proteins; old molecules for new applications.

    Science.gov (United States)

    Lyngstadaas, S P; Wohlfahrt, J C; Brookes, S J; Paine, M L; Snead, M L; Reseland, J E

    2009-08-01

    Emdogain (enamel matrix derivative, EMD) is well recognized in periodontology, where it is used as a local adjunct to periodontal surgery to stimulate regeneration of periodontal tissues lost to periodontal disease. The biological effect of EMD is through stimulation of local growth factor secretion and cytokine expression in the treated tissues, inducing a regenerative process that mimics odontogenesis. The major (>95%) component of EMD is Amelogenins (Amel). No other active components have so far been isolated from EMD, and several studies have shown that purified amelogenins can induce the same effect as the complete EMD. Amelogenins comprise a family of highly conserved extracellular matrix proteins derived from one gene. Amelogenin structure and function is evolutionary well conserved, suggesting a profound role in biomineralization and hard tissue formation. A special feature of amelogenins is that under physiological conditions the proteins self-assembles into nanospheres that constitute an extracellular matrix. In the body, this matrix is slowly digested by specific extracellular proteolytic enzymes (matrix metalloproteinase) in a controlled process, releasing bioactive peptides to the surrounding tissues for weeks after application. Based on clinical and experimental observations in periodontology indicating that amelogenins can have a significant positive influence on wound healing, bone formation and root resorption, several new applications for amelogenins have been suggested. New experiments now confirm that amelogenins have potential for being used also in the fields of endodontics, bone regeneration, implantology, traumatology, and wound care.

  14. Ubiquitination of specific mitochondrial matrix proteins.

    Science.gov (United States)

    Lehmann, Gilad; Ziv, Tamar; Braten, Ori; Admon, Arie; Udasin, Ronald G; Ciechanover, Aaron

    2016-06-17

    Several protein quality control systems in bacteria and/or mitochondrial matrix from lower eukaryotes are absent in higher eukaryotes. These are transfer-messenger RNA (tmRNA), The N-end rule ATP-dependent protease ClpAP, and two more ATP-dependent proteases, HslUV and ClpXP (in yeast). The lost proteases resemble the 26S proteasome and the role of tmRNA and the N-end rule in eukaryotic cytosol is performed by the ubiquitin proteasome system (UPS). Therefore, we hypothesized that the UPS might have substituted these systems - at least partially - in the mitochondrial matrix of higher eukaryotes. Using three independent experimental approaches, we demonstrated the presence of ubiquitinated proteins in the matrix of isolated yeast mitochondria. First, we show that isolated mitochondria contain ubiquitin (Ub) conjugates, which remained intact after trypsin digestion. Second, we demonstrate that the mitochondrial soluble fraction contains Ub-conjugates, several of which were identified by mass spectrometry and are localized to the matrix. Third, using immunoaffinity enrichment by specific antibodies recognizing digested ubiquitinated peptides, we identified a group of Ub-modified matrix proteins. The modification was further substantiated by separation on SDS-PAGE and immunoblots. Last, we attempted to identify the ubiquitin ligase(s) involved, and identified Dma1p as a trypsin-resistant protein in our mitochondrial preparations. Taken together, these data suggest a yet undefined role for the UPS in regulation of the mitochondrial matrix proteins.

  15. Ubiquitination of specific mitochondrial matrix proteins

    Energy Technology Data Exchange (ETDEWEB)

    Lehmann, Gilad [The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096 (Israel); Ziv, Tamar [The Smoler Proteomics Center, Faculty of Biology – Technion-Israel Institute of Technology, Haifa, 32000 (Israel); Braten, Ori [The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096 (Israel); Admon, Arie [The Smoler Proteomics Center, Faculty of Biology – Technion-Israel Institute of Technology, Haifa, 32000 (Israel); Udasin, Ronald G. [The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096 (Israel); Ciechanover, Aaron, E-mail: aaroncie@tx.technion.ac.il [The Janet and David Polak Tumor and Vascular Biology Research Center and the Technion Integrated Cancer Center (TICC), The Rappaport Faculty of Medicine and Research Institute, Haifa, 31096 (Israel)

    2016-06-17

    Several protein quality control systems in bacteria and/or mitochondrial matrix from lower eukaryotes are absent in higher eukaryotes. These are transfer-messenger RNA (tmRNA), The N-end rule ATP-dependent protease ClpAP, and two more ATP-dependent proteases, HslUV and ClpXP (in yeast). The lost proteases resemble the 26S proteasome and the role of tmRNA and the N-end rule in eukaryotic cytosol is performed by the ubiquitin proteasome system (UPS). Therefore, we hypothesized that the UPS might have substituted these systems – at least partially – in the mitochondrial matrix of higher eukaryotes. Using three independent experimental approaches, we demonstrated the presence of ubiquitinated proteins in the matrix of isolated yeast mitochondria. First, we show that isolated mitochondria contain ubiquitin (Ub) conjugates, which remained intact after trypsin digestion. Second, we demonstrate that the mitochondrial soluble fraction contains Ub-conjugates, several of which were identified by mass spectrometry and are localized to the matrix. Third, using immunoaffinity enrichment by specific antibodies recognizing digested ubiquitinated peptides, we identified a group of Ub-modified matrix proteins. The modification was further substantiated by separation on SDS-PAGE and immunoblots. Last, we attempted to identify the ubiquitin ligase(s) involved, and identified Dma1p as a trypsin-resistant protein in our mitochondrial preparations. Taken together, these data suggest a yet undefined role for the UPS in regulation of the mitochondrial matrix proteins. -- Highlights: •Mitochondrial matrix contains ubiquitinated proteins. •Ubiquitination occurs most probably in the matrix. •Dma1p is a ubiquitin ligase present in mitochondrial preparations.

  16. How Osteoblasts Sense their Environment: Integrin-Extracellular Matrix Interactions and Mechanical Loading of Bone

    Science.gov (United States)

    Globus, Ruth K.; Dalton, Bonnie (Technical Monitor)

    2002-01-01

    Osteoblasts are the cells responsible for forming and replacing bone throughout life. We know that mechanical stimulation through weight-bearing at I gravity on Earth is needed to maintain healthy bone, and that osteoblasts play a critical role in that process. Over the last 9 years in my laboratory at NASA ARC, we have studied the regulation of osteoblast function by interactions between the extracellular matrix and die cell. Using a cell culture approach, we defined the repertoire of adhesion receptors, called integrins, which are expressed on the osteoblast surface, as well as specific extracellular matrix proteins, which are needed for cellular differentiation and survival. We are now extending these observations to determine if integrin signaling is involved in the skeletal responses to disuse and recovery from disuse using the rodent model of hindlimb unloading by tail suspension. Together, our cell culture and animal studies are providing new insight into the regulation of osteoblast function in bone.

  17. Extracellular matrix proteins and the dynamics of dentin formation.

    Science.gov (United States)

    Butler, William T; Brunn, Jan C; Qin, Chunlin; McKee, Marc D

    2002-01-01

    Dentinogenesis involves controlled reactions that result in conversion of unmineralized predentin to dentin when apatite crystals are formed. This process is dynamic: Maturation events occur within predentin beginning at the proximal layer and progressing to the predentin-dentin (PD) border. One type of controlled reaction is the proteolytic processing of dentin sialophosphoprotein (DSPP) to dentin sialoprotein (DSP) and dentin phosphoprotein (DPP), by cleavage of at least three highly conserved peptide bonds. We postulate that this processing event represents an activation step, resulting in release of DPP, which is active in its effects on formation and growth of apatite crystals. Dentin matrix protein 1 (DPM1), present as a processed fragment (57-kD protein) in bone, is seen in dentin on sodium dodecyl sulfate polyacrylamide gel electrophoresis as one intact protein of 150-200 kD. Anti-57-kD antibodies elicit immunoreactivity in bone, dentin, and cellular cementum. In bone, the reactivity is associated with osteocytes and their cell processes. Similarly, dentin shows reactivity in odontoblasts, predentin, and the odontoblast processes. In summary, the processing of large sialic acid-rich proteins into smaller fragments may be an important part of the controlled conversion of predentin to dentin and osteoid to bone.

  18. REGENERATION OF ARTICULAR CARTILAGE UNDER THE IMPLANTATION OF BONE MATRIX

    Directory of Open Access Journals (Sweden)

    Yuri M. Iryanov, Nikolay A. Kiryanov, Olga V. Dyuriagina , Tatiana Yu. Karaseva, Evgenii A. Karasev

    2015-07-01

    Full Text Available Background: The damage or loss of articular cartilage is costly medical problem. The purpose of this work – morphological analysis of reparative chondrogenesis when implanted in the area of the knee joint cartilage of granulated mineralized bone matrix. Material and Methods: The characteristic features of the knee cartilage regeneration studied experimentally in pubertal Wistar rats after modeling a marginal perforated defect and implantation of granulated mineralized bone matrix obtained according to original technology without heat and demineralizing processing into the injury zone. Results: This biomaterial established to have pronounced chondro- and osteoinductive properties, and to provide prolonged activation of reparative process, accelerated organotypical remodeling and restoration of the articular cartilage injured. Conclusion: The data obtained demonstrate the efficacy of МВМ in clinical practice for the treatment of diseases and injuries of the articular cartilage.

  19. BONE REGENERATION AFTER DEMINERALIZED BONE MATRIX AND CASTOR OIL (RICINUS COMMUNIS) POLYURETHANE IMPLANTATION

    Science.gov (United States)

    Leite, Fábio Renato Manzolli; Ramalho, Lizeti Toledo de Oliveira

    2008-01-01

    Innocuous biocompatible materials have been searched to repair or reconstruct bone defects. Their goal is to restore the function of live or dead tissues. This study compared connective tissue and bone reaction when exposed to demineralized bovine bone matrix and a polyurethane resin derived from castor bean (Ricinus communis). Forty-five rats were assigned to 3 groups of 15 animals (control, bovine bone and polyurethane). A cylindrical defect was created on mandible base and filled with bovine bone matrix and the polyurethane. Control group received no treatment. Analyses were performed after 15, 45 and 60 days (5 animals each). Histological analysis revealed connective tissue tolerance to bovine bone with local inflammatory response similar to that of the control group. After 15 days, all groups demonstrated similar outcomes, with mild inflammatory reaction, probably due to the surgical procedure rather than to the material. In the polymer group, after 60 days, scarce multinucleated cells could still be observed. In general, all groups showed good stability and osteogenic connective tissue with blood vessels into the surgical area. The results suggest biocompatibility of both materials, seen by their integration into rat mandible. Moreover, the polyurethane seems to be an alternative in bone reconstruction and it is an inexhaustible source of biomaterial. PMID:19089203

  20. Physicomechanical properties of the extracellular matrix of a demineralized bone

    Science.gov (United States)

    Kirilova, I. A.; Sharkeev, Yu. P.; Nikolaev, S. V.; Podorozhnaya, V. T.; Uvarkin, P. V.; Ratushnyak, A. S.; Chebodaeva, V. V.

    2016-08-01

    The article describes the results of a study of physicomechanical properties of a demineralized bone matrix of human cancellous and compact bones. A demineralized cancellous bone was shown to have the best characteristics of a porous system for colonization of matrices by cells. The ultimate stress and elasticity modulus of samples of demineralized femoral heads isolated in primary hip replacement was demonstrated to vary in wide ranges. The elasticity modulus ranged from 50 to 250 MPa, and the tensile strength varied from 1.1 to 5.5 MPa. Microhardness measurements by the recovered indentation method were not possible because of the viscoelastic properties of a bone material. To study the piezoelectric properties of samples, a measuring system was developed that comprised a measuring chamber with contact electrodes, a system for controlled sample loading, an amplifier-converter unit, and signal recording and processing software. The measurement results were used to determine the dependence of the signal amplitude on the dynamic deformation characteristics. The findings are discussed in terms of the relationship between the mechanical and electrical properties and the structure of the organic bone component.

  1. Enamel matrix protein derivatives: role in periodontal regeneration

    Directory of Open Access Journals (Sweden)

    Rathva VJ

    2011-12-01

    Full Text Available Vandana J RathvaDepartment of Periodontics, KM Shah Dental College and Hospital, Sumandeep University, Gujarat, IndiaAbstract: The role of regenerative periodontal therapy is the reconstitution of lost periodontal structures, ie, new formation of root cementum, periodontal ligament, and alveolar bone. The outcome of basic research has pointed to the important role of enamel matrix protein derivative (EMD in periodontal wound healing. Histologic results from animal and human studies have shown that treatment with EMD promotes periodontal regeneration. Moreover, clinical studies have indicated that treatment with EMD positively influences periodontal wound healing in humans. The goal of this paper is to review the existing literature on EMD.Keywords: enamel matrix protein derivative, Emdogain®, periodontal regeneration

  2. Mouse basophils reside in extracellular matrix-enriched bone marrow niches which control their motility.

    Science.gov (United States)

    Smaniotto, Salete; Schneider, Elke; Goudin, Nicolas; Bricard-Rignault, Rachel; Machavoine, François; Dardenne, Mireille; Dy, Michel; Savino, Wilson

    2013-01-01

    Basophils co-express FcεRIα and CD49b, the α-2 chain of integrin-type receptor VLA-2 (α2β1), which recognizes type-1 collagen as a major natural ligand. The physiological relevance of this integrin for interactions with extracellular bone marrow matrix remains unknown. Herein, we examined the expression of several receptors of this family by bone marrow-derived basophils sorted either ex-vivo or after culture with IL-3. Having established that both populations display CD49d, CD49e and CD49f (α-4, α-5 and α-6 integrins subunits, respectively), we addressed receptor functions by measuring migration, adhesion, proliferation and survival after interacting with matched natural ligands. Type I collagen, laminin and fibronectin promoted basophil migration/adhesion, the former being the most effective. None of these ligands affected basophil viability and expansion. Interactions between basophils and extracellular matrix are likely to play a role in situ, as supported by confocal 3D cell imaging of femoral bone marrow sections, which revealed basophils exclusively in type-1 collagen-enriched niches that contained likewise laminin and fibronectin. This is the first evidence for a structure/function relationship between basophils and extracellular matrix proteins inside the mouse bone marrow.

  3. Mouse basophils reside in extracellular matrix-enriched bone marrow niches which control their motility.

    Directory of Open Access Journals (Sweden)

    Salete Smaniotto

    Full Text Available Basophils co-express FcεRIα and CD49b, the α-2 chain of integrin-type receptor VLA-2 (α2β1, which recognizes type-1 collagen as a major natural ligand. The physiological relevance of this integrin for interactions with extracellular bone marrow matrix remains unknown. Herein, we examined the expression of several receptors of this family by bone marrow-derived basophils sorted either ex-vivo or after culture with IL-3. Having established that both populations display CD49d, CD49e and CD49f (α-4, α-5 and α-6 integrins subunits, respectively, we addressed receptor functions by measuring migration, adhesion, proliferation and survival after interacting with matched natural ligands. Type I collagen, laminin and fibronectin promoted basophil migration/adhesion, the former being the most effective. None of these ligands affected basophil viability and expansion. Interactions between basophils and extracellular matrix are likely to play a role in situ, as supported by confocal 3D cell imaging of femoral bone marrow sections, which revealed basophils exclusively in type-1 collagen-enriched niches that contained likewise laminin and fibronectin. This is the first evidence for a structure/function relationship between basophils and extracellular matrix proteins inside the mouse bone marrow.

  4. 釉基质蛋白对猪骨髓基质细胞生物学特性的影响%Effects of Enamel Matrix Proteins on the Biological Activity of Porcine Bone Marrow Stromal Cells in Vitro

    Institute of Scientific and Technical Information of China (English)

    宋爱梅; 束蓉; 蒋欣泉; 张秀丽; 刘晓峰; 张文

    2005-01-01

    目的 研究釉基质蛋白(enamel matrix proteins,EMPs)对体外培养的猪骨髓基质细胞(bone marrow stromal cells,BMSCs)生物学特性的影响.方法 抽取猪髂骨骨髓,全血培养法获得骨髓基质细胞.培养液中EMPs的浓度分别为25、50、100、200ug/ml,以不加EMPs为对照.用比色法检测不同浓度EMPs对BMSCs粘附性的影响.通过计数预定视野中伸展的细胞数,计算BMSCs在1h、3.5h、6.5h的伸展率.MTT法测定各组细胞的增殖活性.结果 猪BMSCs在含有EMPs的培养液中生长良好.对照组以及不同浓度EMPs对细胞粘附性的影响无统计学差异.在1h、3.5h、6.5h,各组细胞的伸展率无显著不同.EMPs对BMSCs的促增殖作用呈浓度和时间依赖性,200ug/ml浓度的EMPs从实验的第三天开始显著促进猪BMSCs的增殖.结论 EMPs对体外培养的猪BMSCs的粘附性和伸展率无影响,200ug/ml浓度的EMPs可显著促进猪BMSCs增殖,提示可尝试联合应用EMPs和BMSCs修复牙周组织缺损.

  5. Delivery Systems for Bone Morphogenetic Protein (BMP) for Repair of Battle Incurred Bone Injuries.

    Science.gov (United States)

    1987-11-01

    infections, congenital malformations that fail to heal are eligible for BMP treatment. I (my child/my ward) will be one of 50 patients to be treated with...Fusions in Dogs 6. Craniotomy Defects in Sheep t0 7. Craniotomy Defects in Monkeys 10 8. BMP Delivery System of Bone Matrix Non Collagenous 11 Proteins...effects. The most important and indispensptle substitutes for experiments in human beings are adult mongrel dogs, monkeys, and sheep . Experimental .S

  6. Bone protein “extractomics”: comparing the efficiency of bone protein extractions of Gallus gallus in tandem mass spectrometry, with an eye towards paleoproteomics

    Directory of Open Access Journals (Sweden)

    Elena R. Schroeter

    2016-10-01

    Full Text Available Proteomic studies of bone require specialized extraction protocols to demineralize and solubilize proteins from within the bone matrix. Although various protocols exist for bone protein recovery, little is known about how discrete steps in each protocol affect the subset of the bone proteome recovered by mass spectrometry (MS analyses. Characterizing these different “extractomes” will provide critical data for development of novel and more efficient protein extraction methodologies for fossils. Here, we analyze 22 unique sub-extractions of chicken bone and directly compare individual extraction components for their total protein yield and diversity and coverage of bone proteins identified by MS. We extracted proteins using different combinations and ratios of demineralizing reagents, protein-solubilizing reagents, and post-extraction buffer removal methods, then evaluated tryptic digests from 20 µg aliquots of each fraction by tandem MS/MS on a 12T FT-ICR mass spectrometer. We compared total numbers of peptide spectral matches, peptides, and proteins identified from each fraction, the redundancy of protein identifications between discrete steps of extraction methods, and the sequence coverage obtained for select, abundant proteins. Although both alpha chains of collagen I (the most abundant protein in bone were found in all fractions, other collagenous and non-collagenous proteins (e.g., apolipoprotein, osteonectin, hemoglobin were differentially identified. We found that when a standardized amount of extracted proteins was analyzed, extraction steps that yielded the most protein (by weight from bone were often not the ones that produced the greatest diversity of bone proteins, or the highest degree of protein coverage. Generally, the highest degrees of diversity and coverage were obtained from demineralization fractions, and the proteins found in the subsequent solubilization fractions were highly redundant with those in the previous

  7. Effect of cefazolin loaded bone matrix gelatin on repairing large segmental bone defects and preventing infection

    Institute of Scientific and Technical Information of China (English)

    游洪波; 陈安民

    2004-01-01

    Objective: To explore the possibility of repairing long segmental bone defects and preventing infection with cefazolin loaded bone matrix gelatin (C-BMG). Methods: C-BMG was made from putting cefazolin into BMG by vacuum absorption and lyophilization techniques. The sustaining period of effective drug concentration in vitro and in vivo was detected. The time of inhibiting bacteria, and the drug concentration in local tissues ( bone and muscle) and plasma after implantation of C-BMG were examined by high performance liquid chromatography.Results: The effective inhibition time to staphylococcus aureus of C-BMG was 22 days in vitro; while 14 days in vivo. The cefazolin concentration in local tissues was higher in early stage, and later it kept a stable and low drug release. C-BMG showed an excellent ability to repair segmental long bone defects.Conclusions: C-BMG can gradually release cefazolin with effective drug concentration and has excellent ability to repair segmental bone defects. It can be used to repair segmental long bone defects and prevent infection after operation.

  8. Bone Formation is Affected by Matrix Advanced Glycation End Products (AGEs) In Vivo.

    Science.gov (United States)

    Yang, Xiao; Mostafa, Ahmed Jenan; Appleford, Mark; Sun, Lian-Wen; Wang, Xiaodu

    2016-10-01

    Advanced glycation end products (AGEs) accumulate in bone extracellular matrix as people age. Although previous evidence shows that the accumulation of AGEs in bone matrix may impose significant effects on bone cells, the effect of matrix AGEs on bone formation in vivo is still poorly understood. To address this issue, this study used a unique rat model with autograft implant to investigate the in vivo response of bone formation to matrix AGEs. Fluorochrome biomarkers were sequentially injected into rats to label the dynamic bone formation in the presence of elevated levels of matrix AGEs. After sacrificing animals, dynamic histomorphometry was performed to determine mineral apposition rate (MAR), mineralized surface per bone surface (MS/BS), and bone formation rate (BFR). Finally, nanoindentation tests were performed to assess mechanical properties of newly formed bone tissues. The results showed that MAR, MS/BS, and BFR were significantly reduced in the vicinity of implant cores with high concentration of matrix AGEs, suggesting that bone formation activities by osteoblasts were suppressed in the presence of elevated matrix AGEs. In addition, MAR and BFR were found to be dependent on the surrounding environment of implant cores (i.e., cortical or trabecular tissues). Moreover, MS/BS and BFR were also dependent on how far the implant cores were away from the growth plate. These observations suggest that the effect of matrix AGEs on bone formation is dependent on the biological milieu around the implants. Finally, nanoindentation test results indicated that the indentation modulus and hardness of newly formed bone tissues were not affected by the presence of elevated matrix AGEs. In summary, high concentration of matrix AGEs may slow down the bone formation process in vivo, while imposing little effects on bone mineralization.

  9. Extracellular matrix proteins involved in pseudoislets formation.

    Science.gov (United States)

    Maillard, Elisa; Sencier, Marie-Christine; Langlois, A; Bietiger, William; Krafft, Mp; Pinget, Michel; Sigrist, Séverine

    2009-01-01

    Extracellular matrix proteins are known to mediate, through integrins, cell adhesion and are involved in a number of cellular processes, including insulin expression and secretion in pancreatic islets. We investigated whether expression of some extracellular matrix proteins were implied in islets-like structure formation, named pseudoislets. For this purpose, we cultured the β-cell line, RINm5F, during 1, 3, 5 and 7 days of culture on treated or untreated culture plate to form adherent cells or pseudoislets and analysed insulin, collagen IV, fibronectin, laminin 5 and β1-integrin expression. We observed that insulin expression and secretion were increased during pseudoislets formation. Moreover, we showed by immunohistochemistry an aggregation of insulin secreting cells in the centre of the pseudoislets. Peripheral β-cells of pseudoislets did not express insulin after 7 days of culture. RT-PCR and immunohistochemistry studies showed a transient expression of type IV collagen in pseudoislets for the first 3 days of culture. Study of fibronectin expression indicated that adherent cells expressed more fibronectin than pseudoislets. In contrast, laminin 5 was more expressed in pseudoislets than in adherent cells. Finally, expression of β1-integrin was increased in pseudoislets as compared to adherent cells. In conclusion, laminin 5 and collagen IV might be implicated in pseudoislets formation whereas fibronectin might be involved in cell adhesion. These data suggested that extracellular matrix proteins may enhance the function of pseudoislets.

  10. 'GOING NANO' in the field of osteology: Is bone fragility already determined at the level of bone mineralized matrix?

    Directory of Open Access Journals (Sweden)

    Milovanović Petar

    2016-01-01

    Full Text Available Bone fractures frequently occur in elderly persons even after low-energy trauma. Given that the young individuals would not sustain a fracture under such conditions, it is clear that the main cause of easy bone fracturing originates from the structural and compositional characteristics of the aged bone itself. Observing bone, from macro- to nano-level, shows us the complexity of bone hierarchical organization and reveals various determinants of bone strength. In particular, recent studies focusing on bone at nano-scale revealed distinctive features of the bone matrix that could provide additional explanation for the increased bone fragility in advanced age, independent from age-related effects at other levels of bone hierarchical structure.

  11. Calcification preceding new bone formation induced by demineralized bone matrix gelatin.

    Science.gov (United States)

    Yamashita, K; Takagi, T

    1992-03-01

    Demineralized bone matrix gelatin (BMG) was implanted into the skeletal muscle of Sprague-Dawley (S.D.) rats, and histological changes were examined 3, 5, 7, 10 and 15 days later. Before bone formation, a specific calcification process was found in most of the BMG from day 5 and 7 after implantation. The heterotopic calcified sites were not always consistent with the sites of the alkaline phosphatase activity. It was considered that this calcification progresses without any cellular components, and we distinguished this type of calcification as "acellular mineral deposition" from the calcification which occurs in new bone formation. This "acellular mineral deposition" was first observed as small spherical calcified deposits in the BMG on day 7 after implantation; these deposits then gradually grew and fused with each other. Some multinucleated cells appeared near the site of calcification on day 7 after implantation, but osteoblasts or osteoblast-like cells were scarcely observed around the calcified deposits in BMG until day 7. Vascularization was often observed near the "acellular mineral deposition" and the new bone formation. Fourier transform infrared spectroscopy showed that the calcified deposits in BMG were composed of hydroxyapatite, carbonateapatite and other calcium phosphate components, and that the first two components became prominent with time. It is believed that the "acellular mineral deposition" is due to the deposition of calcium and phosphate into the BMG by a process of heterogenic nucleation that does not involve osteoblasts or matrix vesicles. Bone formation induced by the BMG occurred after the "acellular mineral deposition." The experimental calcification shown in this paper seems a useful model for the study of biocalcification.

  12. Biomineralization of a Self-Assembled Extracellular Matrix for Bone Tissue Engineering

    Science.gov (United States)

    Meng, Yizhi; DiMasi, Elaine; Ba, Xiaolan; Rafailovich, Miriam; Pernodet, Nadine

    2009-01-01

    Understanding how biomineralization occurs in the extracellular matrix (ECM) of bone cells is crucial to the understanding of bone formation and the development of a successfully engineered bone tissue scaffold. It is still unclear how ECM mechanical properties affect protein-mineral interactions in early stages of bone mineralization. We investigated the longitudinal mineralization properties of MC3T3-E1 cells and the elastic modulus of their ECM using shear modulation force microscopy, synchrotron grazing incidence X-ray diffraction (GIXD), scanning electron microscopy, energy dispersive X-ray spectroscopy, and confocal laser scanning microscopy (CLSM). The elastic modulus of the ECM fibers underwent significant changes for the mineralizing cells, which were not observed in the nonmineralizing cells. On substrates conducive to ECM network production, the elastic modulus of mineralizing cells increased at time points corresponding to mineral production, whereas that of the nonmineralizing cells did not vary over time. The presence of hydroxyapatite in mineralizing cells and the absence thereof in the nonmineralizing ones were confirmed by GIXD, and CLSM showed that a restructuring of actin occurred only for mineral-producing cells. These results show that the correct and complete development of the ECM network is required for osteoblasts to mineralize. This in turn requires a suitably prepared synthetic substrate for bone development to succeed in vitro. PMID:18759666

  13. Biomineralization of a Self-Assembled Extracellular Matrix for Bone Tissue Engineering

    Energy Technology Data Exchange (ETDEWEB)

    Yizhi, M.; Yi-Xian, Q; DiMasi, E; Xiaolan, B; Rafailovich, M; Pernodet, N

    2009-01-01

    Understanding how biomineralization occurs in the extracellular matrix (ECM) of bone cells is crucial to the understanding of bone formation and the development of a successfully engineered bone tissue scaffold. It is still unclear how ECM mechanical properties affect protein-mineral interactions in early stages of bone mineralization. We investigated the longitudinal mineralization properties of MC3T3-E1 cells and the elastic modulus of their ECM using shear modulation force microscopy, synchrotron grazing incidence X-ray diffraction (GIXD), scanning electron microscopy, energy dispersive X-ray spectroscopy, and confocal laser scanning microscopy (CLSM). The elastic modulus of the ECM fibers underwent significant changes for the mineralizing cells, which were not observed in the nonmineralizing cells. On substrates conducive to ECM network production, the elastic modulus of mineralizing cells increased at time points corresponding to mineral production, whereas that of the nonmineralizing cells did not vary over time. The presence of hydroxyapatite in mineralizing cells and the absence thereof in the nonmineralizing ones were confirmed by GIXD, and CLSM showed that a restructuring of actin occurred only for mineral-producing cells. These results show that the correct and complete development of the ECM network is required for osteoblasts to mineralize. This in turn requires a suitably prepared synthetic substrate for bone development to succeed in vitro.

  14. Role of Soy Protein on Bone Turnover

    Directory of Open Access Journals (Sweden)

    A Haghighian roudsari

    2004-03-01

    Full Text Available Bone mass loss is one of the commonest menopause symptoms, resulting from cessation of estrogen production. Compounds which have estrogen – like biological activity similar to “Isoflavones” present in plants especially soy, may reduce bone loss in postmenopausal women, because as they are similar in structure to estrogens. This study, therefore, was undertaken to assess the effect of soy protein on bone metabolism biomarkers in postmenopausal women with osteopenia. This “before and after” clinical trial was carried out, on 15 postmenopausal women with osteopenia, between 45 to 64 years of age. The subjects were asked to consume 35 gram/day of soy protein for 12 weeks. Blood and urine samples, were taken at 0, 6 and 12 weeks of the study. Anthropometric measurements and a 2-day dietary recall were done at the beginning of the study, and at the 6 and 12 weeks. The food consumption data were analyzed by “Food Proccessor” software. Repeated measurement analysis was utilized to determine the changes in biochemical indices, anthropometric and dietary data. P-values less than 0.05 were considered as significant. Comparison of weight, BMI, physical activity and dietary intake of subjects during the study did not show any significant differences. Soy protein consumption, showed significant reductions in deoxypyridinoline (biochemical marker of bone resorption and significant increase in total alkaline phosphatase ( biochemical marker of bone formation.There were no significant differences in serum osteocalcin, C- telopeptide, insulin- like growth factor binding protein 3 (IGFBP3, and type-I- collagen telopeptides. Considering the beneficial effects of soy protein consumption on bone metabolism biomarkers, inclusion of this inexpensive and available food item in postmenopausal women diet, may reduce bone loss and could be recommended for the prevention of osteoporosis.

  15. Role of Soy Protein on Bone Turnover

    Directory of Open Access Journals (Sweden)

    A Haghighian roudsari

    2004-11-01

    Full Text Available Bone mass loss is one of the commonest menopause symptoms, resulting from cessation of estrogen production. Compounds which have estrogen – like biological activity similar to “Isoflavones” present in plants especially soy, may reduce bone loss in postmenopausal women, because as they are similar in structure to estrogens. This study, therefore, was undertaken to assess the effect of soy protein on bone metabolism biomarkers in postmenopausal women with osteopenia. This “before and after” clinical trial was carried out, on 15 postmenopausal women with osteopenia, between 45 to 64 years of age. The subjects were asked to consume 35 gram/day of soy protein for 12 weeks. Blood and urine samples, were taken at 0, 6 and 12 weeks of the study. Anthropometric measurements and a 2-day dietary recall were done at the beginning of the study, and at the 6 and 12 weeks. The food consumption data were analyzed by “Food Proccessor” software. Repeated measurement analysis was utilized to determine the changes in biochemical indices, anthropometric and dietary data. P-values less than 0.05 were considered as significant. Comparison of weight, BMI, physical activity and dietary intake of subjects during the study did not show any significant differences. Soy protein consumption, showed significant reductions in deoxypyridinoline (biochemical marker of bone resorption and significant increase in total alkaline phosphatase ( biochemical marker of bone formation.There were no significant differences in serum osteocalcin, C- telopeptide, insulin- like growth factor binding protein 3 (IGFBP3, and type-I- collagen telopeptides. Considering the beneficial effects of soy protein consumption on bone metabolism biomarkers, inclusion of this inexpensive and available food item in postmenopausal women diet, may reduce bone loss and could be recommended for the prevention of osteoporosis.

  16. The application of an enamel matrix protein derivative (Emdogain) in regenerative periodontal therapy: a review.

    NARCIS (Netherlands)

    Sculean, A.; Schwarz, F.; Becker, J.; Brecx, M.

    2007-01-01

    Regenerative periodontal therapy aims at reconstitution of the lost periodontal structures such as new formation of root cementum, periodontal ligament and alveolar bone. Findings from basic research indicate that enamel matrix protein derivative (EMD) has a key role in periodontal wound healing. Hi

  17. A comparative study of fibrous dysplasia and osteofibrous dysplasia with regard to expressions of c-fos and c-jun products and bone matrix proteins: a clinicopathologic review and immunohistochemical study of c-fos, c-jun, type I collagen, osteonectin, osteopontin, and osteocalcin.

    Science.gov (United States)

    Sakamoto, A; Oda, Y; Iwamoto, Y; Tsuneyoshi, M

    1999-12-01

    Fibrous dysplasia and osteofibrous dysplasia are both benign fibro-osseous lesions of the bone and are generally seen during childhood or adolescence. Histologically, the features of these bone lesions sometimes look quite similar, but their precise nature remains controversial. We retrospectively studied clinicopathologic findings in 62 cases of fibrous dysplasia and 20 cases of osteofibrous dysplasia with regard to their anatomic location and histological appearance. From among these cases, the immunohistochemical expressions of c-fos and c-jun proto-oncogene products and bone matrix proteins of type I collagen, osteonectin, osteopontin, and osteocalcin were evaluated in 20 typical fibrous dysplasias and 17 osteofibrous dysplasias using paraffin sections, and these expressions were then assessed semiquantitatively. Microscopically, fibrous dysplasia showed various secondary changes, such as hyalinization, hemorrhage, xanthomatous reaction, and cystic change in 22 of the 62 cases (35%). This was a higher incidence than in osteofibrous dysplasia, in which only 2 of the 20 cases (10%) showed such changes. In the elderly fibrous dysplasia cases, the cellularity of fibroblast-like cells was rather low, and those cases were hyalinized. Almost all of the cases of fibrous dysplasia and osteofibrous dysplasia showed positive expressions of c-fos and c-jun products. The expressions of type I collagen and osteopontin showed no difference between fibrous dysplasia and osteofibrous dysplasia. Immunoreactivity for osteonectin in bone matrix was detected in only 1 case of fibrous dysplasia (1 of 20), whereas it was recognized in 14 of the 17 cases of osteofibrous dysplasia. Furthermore, the immunoreactivity for osteocalcin in bone matrix and fibroblast-like cells was higher in fibrous dysplasia than it was in osteofibrous dysplasia, semiquantitatively. Our immunohistochemical results regarding osteonectin and osteocalcin suggest that the bone matrix of fibrous dysplasia is

  18. Low dose BMP-2 treatment for bone repair using a PEGylated fibrinogen hydrogel matrix.

    Science.gov (United States)

    Ben-David, Dror; Srouji, Samer; Shapira-Schweitzer, Keren; Kossover, Olga; Ivanir, Eran; Kuhn, Gisela; Müller, Ralph; Seliktar, Dror; Livne, Erella

    2013-04-01

    Bone repair strategies utilizing resorbable biomaterial implants aim to stimulate endogenous cells in order to gradually replace the implant with functional repair tissue. These biomaterials should therefore be biodegradable, osteoconductive, osteoinductive, and maintain their integrity until the newly formed host tissue can contribute proper function. In recent years there has been impressive clinical outcomes for this strategy when using osteoconductive hydrogel biomaterials in combination with osteoinductive growth factors such as human recombinant bone morphogenic protein (hrBMP-2). However, the success of hrBMP-2 treatments is not without risks if the factor is delivered too rapidly and at very high doses because of a suboptimal biomaterial. Therefore, the aim of this study was to evaluate the use of a PEGylated fibrinogen (PF) provisional matrix as a delivery system for low-dose hrBMP-2 treatment in a critical size maxillofacial bone defect model. PF is a semi-synthetic hydrogel material that can regulate the release of physiological doses of hrBMP-2 based on its controllable physical properties and biodegradation. hrBMP-2 release from the PF material and hrBMP-2 bioactivity were validated using in vitro assays and a subcutaneous implantation model in rats. Critical size calvarial defects in mice were treated orthotopically with PF containing 8 μg/ml hrBMP-2 to demonstrate the capacity of these bioactive implants to induce enhanced bone formation in as little as 6 weeks. Control defects treated with PF alone or left empty resulted in far less bone formation when compared to the PF/hrBMP-2 treated defects. These results demonstrate the feasibility of using a semi-synthetic biomaterial containing small doses of osteoinductive hrBMP-2 as an effective treatment for maxillofacial bone defects.

  19. Structure and assembly of a paramyxovirus matrix protein.

    Science.gov (United States)

    Battisti, Anthony J; Meng, Geng; Winkler, Dennis C; McGinnes, Lori W; Plevka, Pavel; Steven, Alasdair C; Morrison, Trudy G; Rossmann, Michael G

    2012-08-28

    Many pleomorphic, lipid-enveloped viruses encode matrix proteins that direct their assembly and budding, but the mechanism of this process is unclear. We have combined X-ray crystallography and cryoelectron tomography to show that the matrix protein of Newcastle disease virus, a paramyxovirus and relative of measles virus, forms dimers that assemble into pseudotetrameric arrays that generate the membrane curvature necessary for virus budding. We show that the glycoproteins are anchored in the gaps between the matrix proteins and that the helical nucleocapsids are associated in register with the matrix arrays. About 90% of virions lack matrix arrays, suggesting that, in agreement with previous biological observations, the matrix protein needs to dissociate from the viral membrane during maturation, as is required for fusion and release of the nucleocapsid into the host's cytoplasm. Structure and sequence conservation imply that other paramyxovirus matrix proteins function similarly.

  20. The importance of the SIBLING family of proteins on skeletal mineralisation and bone remodelling.

    Science.gov (United States)

    Staines, Katherine A; MacRae, Vicky E; Farquharson, Colin

    2012-09-01

    The small integrin-binding ligand N-linked glycoprotein (SIBLING) family consists of osteopontin, bone sialoprotein, dentin matrix protein 1, dentin sialophosphoprotein and matrix extracellular phosphoglycoprotein. These proteins share many structural characteristics and are primarily located in bone and dentin. Accumulating evidence has implicated the SIBLING proteins in matrix mineralisation. Therefore, in this review, we discuss the individual role that each of the SIBLING proteins has in this highly orchestrated process. In particular, we emphasise how the nature and extent of their proteolytic processing and post-translational modification affect their functional role. Finally, we describe the likely roles of the SIBLING proteins in clinical disorders of hypophosphataemia and their potential therapeutic use.

  1. Effects of demineralized bone matrix and a 'Ricinus communis' polymer on bone regeneration: a histological study in rabbit calvaria.

    Science.gov (United States)

    Laureano Filho, José R; Andrade, Emanuel S S; Albergaria-Barbosa, José R; Camargo, Igor B; Garcia, Robson R

    2009-09-01

    The aim of the present study was to histologically analyze the effects of bovine and human demineralized bone matrix and a Ricinus communis polymer on the bone regeneration process. Two surgical bone defects were created in rabbit calvaria, one on the right and the other on the left side of the parietal suture. Eighteen rabbits were divided into three groups. In Group I, the experimental defect was treated with bovine demineralized bone matrix, Group II with human demineralized bone matrix, and in Group III, the experimental cavity was treated with polyurethane resin derived from Ricinus communis oil. The control defects were filled with the animals' own blood. The animals were sacrificed after 7 and 15 weeks. Histological analysis revealed that in all groups (control and experimental), bone regeneration increased with time. The least time required for bone regeneration was noted in the control group, with a substantial decrease in the thickness of the defect. All materials proved to be biologically compatible, but polyurethane resorbed more slowly and demonstrated considerably better results than the demineralized bone matrices.

  2. A composite demineralized bone matrix--self assembling peptide scaffold for enhancing cell and growth factor activity in bone marrow.

    Science.gov (United States)

    Hou, Tianyong; Li, Zhiqiang; Luo, Fei; Xie, Zhao; Wu, Xuehui; Xing, Junchao; Dong, Shiwu; Xu, Jianzhong

    2014-07-01

    The need for suitable bone grafts is high; however, there are limitations to all current graft sources, such as limited availability, the invasive harvest procedure, insufficient osteoinductive properties, poor biocompatibility, ethical problems, and degradation properties. The lack of osteoinductive properties is a common problem. As an allogenic bone graft, demineralized bone matrix (DBM) can overcome issues such as limited sources and comorbidities caused by invasive harvest; however, DBM is not sufficiently osteoinductive. Bone marrow has been known to magnify osteoinductive components for bone reconstruction because it contains osteogenic cells and factors. Mesenchymal stem cells (MSCs) derived from bone marrow are the gold standard for cell seeding in tissue-engineered biomaterials for bone repair, and these cells have demonstrated beneficial effects. However, the associated high cost and the complicated procedures limit the use of tissue-engineered bone constructs. To easily enrich more osteogenic cells and factors to DBM by selective cell retention technology, DBM is modified by a nanoscale self-assembling peptide (SAP) to form a composite DBM/SAP scaffold. By decreasing the pore size and increasing the charge interaction, DBM/SAP scaffolds possess a much higher enriching yield for osteogenic cells and factors compared with DBM alone scaffolds. At the same time, SAP can build a cellular microenvironment for cell adhesion, proliferation, and differentiation that promotes bone reconstruction. As a result, a suitable bone graft fabricated by DBM/SAP scaffolds and bone marrow represents a new strategy and product for bone transplantation in the clinic.

  3. A 3D printed nano bone matrix for characterization of breast cancer cell and osteoblast interactions

    Science.gov (United States)

    Zhu, Wei; Castro, Nathan J.; Cui, Haitao; Zhou, Xuan; Boualam, Benchaa; McGrane, Robert; Glazer, Robert I.; Zhang, Lijie Grace

    2016-08-01

    Bone metastasis is one of the most prevalent complications of late-stage breast cancer, in which the native bone matrix components, including osteoblasts, are intimately involved in tumor progression. The development of a successful in vitro model would greatly facilitate understanding the underlying mechanism of breast cancer bone invasion as well as provide a tool for effective discovery of novel therapeutic strategies. In the current study, we fabricated a series of in vitro bone matrices composed of a polyethylene glycol hydrogel and nanocrystalline hydroxyapatite of varying concentrations to mimic the native bone microenvironment for the investigation of breast cancer bone metastasis. A stereolithography-based three-dimensional (3D) printer was used to fabricate the bone matrices with precisely controlled architecture. The interaction between breast cancer cells and osteoblasts was investigated in the optimized bone matrix. Using a Transwell® system to separate the two cell lines, breast cancer cells inhibited osteoblast proliferation, while osteoblasts stimulated breast cancer cell growth, whereas, both cell lines increased IL-8 secretion. Breast cancer cells co-cultured with osteoblasts within the 3D bone matrix formed multi-cellular spheroids in comparison to two-dimensional monolayers. These findings validate the use of our 3D printed bone matrices as an in vitro metastasis model, and highlights their potential for investigating breast cancer bone metastasis.

  4. Age-Related Effects of Advanced Glycation End Products (Ages) in Bone Matrix on Osteoclastic Resorption.

    Science.gov (United States)

    Yang, Xiao; Gandhi, Chintan; Rahman, Md Mizanur; Appleford, Mark; Sun, Lian-Wen; Wang, Xiaodu

    2015-12-01

    Advanced glycation end products (AGEs) accumulate in bone extracellular matrix as people age. Previous studies have shown controversial results regarding the role of in situ AGEs accumulation in osteoclastic resorption. To address this issue, this study cultured human osteoclast cells directly on human cadaveric bone slices from different age groups (young and elderly) to warrant its relevance to in vivo conditions. The cell culture was terminated on the 3rd, 7th, and 10th day, respectively, to assess temporal changes in the number of differentiated osteoclasts, the number and size of osteoclastic resorption pits, the amount of bone resorbed, as well as the amount of matrix AGEs released in the medium by resorption. In addition, the in situ concentration of matrix AGEs at each resorption pit was also estimated based on its AGEs autofluorescent intensity. The results indicated that (1) osteoclastic resorption activities were significantly correlated with the donor age, showing larger but shallower resorption pits on the elderly bone substrates than on the younger ones; (2) osteoclast resorption activities were not significantly dependent on the in situ AGEs concentration in bone matrix, and (3) a correlation was observed between osteoclast activities and the concentration of AGEs released by the resorption. These results suggest that osteoclasts tend to migrate away from initial anchoring sites on elderly bone substrate during resorption compared to younger bone substrates. However, such behavior is not directly related to the in situ concentration of AGEs in bone matrix at the resorption sites.

  5. Extracellular Vesicle-functionalized Decalcified Bone Matrix Scaffolds with Enhanced Pro-angiogenic and Pro-bone Regeneration Activities.

    Science.gov (United States)

    Xie, Hui; Wang, Zhenxing; Zhang, Liming; Lei, Qian; Zhao, Aiqi; Wang, Hongxiang; Li, Qiubai; Cao, Yilin; Jie Zhang, Wen; Chen, Zhichao

    2017-04-03

    Vascularization is crucial for bone regeneration after the transplantation of tissue-engineered bone grafts in the clinical setting. Growing evidence suggests that mesenchymal stem cell (MSC)-derived extracellular vesicles (EVs) are potently pro-angiogenic both in vitro and in vivo. In the current study, we fabricated a novel EV-functionalized scaffold with enhanced pro-angiogenic and pro-bone regeneration activities by coating decalcified bone matrix (DBM) with MSC-derived EVs. EVs were harvested from rat bone marrow-derived MSCs and the pro-angiogenic potential of EVs was investigated in vitro. DBM scaffolds were then coated with EVs, and the modification was verified by scanning electron microscopy and confocal microscopy. Next, the pro-angiogenic and pro-bone regeneration activities of EV-modified scaffolds were evaluated in a subcutaneous bone formation model in nude mice. Micro-computed tomography scanning analysis showed that EV-modified scaffolds with seeded cells enhanced bone formation. Enhanced bone formation was confirmed by histological analysis. Immunohistochemical staining for CD31 proved that EV-modified scaffolds promoted vascularization in the grafts, thereby enhancing bone regeneration. This novel scaffold modification method provides a promising way to promote vascularization, which is essential for bone tissue engineering.

  6. Vitamin D Binding Protein and Bone Health

    Directory of Open Access Journals (Sweden)

    Ishir Bhan

    2014-01-01

    Full Text Available Vitamin D binding protein (DBP is the major carrier protein of 25-hydroxyvitamin D (25(OH D in the circulation, where it may serve roles in maintaining stable levels during times of decreased 25(OH availability and in regulating delivery of 25(OH D to target tissues. Several genetic polymorphisms of DBP have been described that lead to phenotypic changes in the protein that may affect affinity, activity, and concentration. These polymorphisms have been linked with alterations in bone density in several populations. One of the mechanisms by which DBP may alter bone health involves regulating vitamin D bioavailability. DBP-bound vitamin is thought to be relatively unavailable to target tissues, and thus alterations in DBP levels or affinity could lead to changes in vitamin D bioactivity. As a result, functional vitamin D status may differ greatly between individuals with similar total 25(OH D levels. Additionally, DBP may have independent roles on macrophage and osteoclast activation. This review will summarize recent findings about DBP with respect to measures of bone density and health.

  7. Bone matrix calcification during embryonic and postembryonic rat calvarial development assessed by SEM-EDX spectroscopy, XRD, and FTIR spectroscopy.

    Science.gov (United States)

    Henmi, Akiko; Okata, Hiroshi; Anada, Takahisa; Yoshinari, Mariko; Mikami, Yasuto; Suzuki, Osamu; Sasano, Yasuyuki

    2016-01-01

    Bone mineral is constituted of biological hydroxyapatite crystals. In developing bone, the mineral crystal matures and the Ca/P ratio increases. However, how an increase in the Ca/P ratio is involved in maturation of the crystal is not known. The relationships among organic components and mineral changes are also unclear. The study was designed to investigate the process of calcification during rat calvarial bone development. Calcification was evaluated by analyzing the atomic distribution and concentration of Ca, P, and C with scanning electron microscopy (SEM)-energy-dispersive X-ray (EDX) spectroscopy and changes in the crystal structure with X-ray diffraction (XRD) and Fourier transform infrared (FTIR) spectroscopy. Histological analysis showed that rat calvarial bone formation started around embryonic day 16. The areas of Ca and P expanded, matching the region of the developing bone matrix, whereas the area of C became localized around bone. X-ray diffraction and FTIR analysis showed that the amorphous-like structure of the minerals at embryonic day 16 gradually transformed into poorly crystalline hydroxyapatite, whereas the proportion of mineral to protein increased until postnatal week 6. FTIR analysis also showed that crystallization of hydroxyapatite started around embryonic day 20, by which time SEM-EDX spectroscopy showed that the Ca/P ratio had increased and the C/Ca and C/P ratios had decreased significantly. The study suggests that the Ca/P molar ratio increases and the proportion of organic components such as proteins of the bone matrix decreases during the early stage of calcification, whereas crystal maturation continues throughout embryonic and postembryonic bone development.

  8. Bisphosphonate treatment affects trabecular bone apparent modulus through micro-architecture rather than matrix properties.

    Science.gov (United States)

    Day, J S; Ding, M; Bednarz, P; van der Linden, J C; Mashiba, T; Hirano, T; Johnston, C C; Burr, D B; Hvid, I; Sumner, D R; Weinans, H

    2004-05-01

    Bisphosphonates are emerging as an important treatment for osteoporosis. But whether the reduced fracture risk associated with bisphosphonate treatment is due to increased bone mass, improved trabecular architecture and/or increased secondary mineralization of the calcified matrix remains unclear. We examined the effects of bisphosphonates on both the trabecular architecture and matrix properties of canine trabecular bone. Thirty-six beagles were divided into a control group and two treatment groups, one receiving risedronate and the other alendronate at 5-6 times the clinical dose for osteoporosis treatment. After one year, the dogs were killed, and samples from the first lumbar vertebrae were examined using a combination of micro-computed tomography, finite element modeling, and mechanical testing. By combining these methods, we examined the treatment effects on the calcified matrix and trabecular architecture independently. Conventional histomorphometry and microdamage data were obtained from the second and third lumbar vertebrae of the same dogs [Bone 28 (2001) 524]. Bisphosphonate treatment resulted in an increased apparent Young's modulus, decreased bone turnover, increased calcified matrix density, and increased microdamage. We could not detect any change in the effective Young's modulus of the calcified matrix in the bisphosphonate treated groups. The observed increase in apparent Young's modulus was due to increased bone mass and altered trabecular architecture rather than changes in the calcified matrix modulus. We hypothesize that the expected increase in the Young's modulus of the calcified matrix due to the increased calcified matrix density was counteracted by the accumulation of microdamage.

  9. Titanium implants induce expression of matrix metalloproteinases in bone during osseointegration

    National Research Council Canada - National Science Library

    Shubayev, Veronica I; Brånemark, Rickard; Steinauer, Joanne; Myers, Robert R

    2004-01-01

    ...) bond at the titanium-bone interface. In this study, we used a rodent femur model of intramedullary osseointegration to analyze the changes in immunoreactivity of ECM-controlling matrix metalloproteinases (MMPs...

  10. 3D printed nanocomposite matrix for the study of breast cancer bone metastasis.

    Science.gov (United States)

    Zhu, Wei; Holmes, Benjamin; Glazer, Robert I; Zhang, Lijie Grace

    2016-01-01

    Bone is one of the most common metastatic sites of breast cancer, but the underlying mechanisms remain unclear, in part due to an absence of advanced platforms for cancer culture and study that mimic the bone microenvironment. In the present study, we integrated a novel stereolithography-based 3D printer and a unique 3D printed nano-ink consisting of hydroxyapatite nanoparticles suspended in hydrogel to create a biomimetic bone-specific environment for evaluating breast cancer bone invasion. Breast cancer cells cultured in a geometrically optimized matrix exhibited spheroid morphology and migratory characteristics. Co-culture of tumor cells with bone marrow mesenchymal stem cells increased the formation of spheroid clusters. The 3D matrix also allowed for higher drug resistance of breast cancer cells than 2D culture. These results validate that our 3D bone matrix can mimic tumor bone microenvironments, suggesting that it can serve as a tool for studying metastasis and assessing drug sensitivity. From the Clinical Editor: Cancer remains a major cause of mortality for patients in the clinical setting. For breast cancer, bone is one of the most common metastatic sites. In this intriguing article, the authors developed a bone-like environment using 3D printing technology to investigate the underlying biology of bone metastasis. Their results would also allow a new model for other researchers who work on cancer to use.

  11. Nanometeric hydroxyapatite in situ formed in demineralized bone matrix by electrochemical technique

    Institute of Scientific and Technical Information of China (English)

    CHEN Ji-da; WANG Yuan-ling; CAI Hao-xi; CAO Yin

    2001-01-01

    @@ To prepare biomimic bone materials, some thick meshes of demineralized bone matrix (DBM), which are the interest work of collagen fibrils, have been prepared in normal way, and calcium phosphate of nanometeric apatite-li.ke was introduced into these mesh works with aqueous calcium and phosphate salt solution by means of the electrochemical technique at room temperature.

  12. Evaluation of heterotopic bone formation induced by squalane and bone morphogenetic protein composite.

    Science.gov (United States)

    Kawakami, T; Kawai, T; Takei, N; Kise, T; Eda, S; Urist, M R

    1997-04-01

    Bone morphogenetic protein is an important molecule whose bioactivity depends on the carrier. Squalane is used in the formulation of various kinds of cosmetics because it is easily emulsified and has the property of spreading well. Thus, squalane might be effective as a bone morphogenetic protein delivery system. As a test for this possibility, gelatin capsules containing squalane and bone morphogenetic protein (bovine derived partially purified) composite were implanted under the hind-quarter perimuscular membrane of ddY mice. Control capsules containing only bone morphogenetic protein were used for controls. The implants were radiographically and histologically examined at 1 to 4 weeks after the operation. According to the radiographic analysis, squalane and bone morphogenetic protein composite and bone morphogenetic protein only control specimens formed widespread heterotopic bone tissues. The amount of heterotopic bone formation in the composite experimental specimens was approximately 40% greater than that in the controls. Histologic examination of experimental and control specimens revealed varying amounts of perichondral ossification by 2 weeks. By 3 and 4 weeks, the bone deposits were colonized by hematopoietic bone marrow. Squalane was effective for the slow local release of bone morphogenetic protein. Furthermore, the squalane and bone morphogenetic protein composite was a reliable osteoinductive biomaterial.

  13. Exercise Alters Mineral and Matrix Composition in the Absence of Adding New Bone

    OpenAIRE

    Kohn, David H.; Nadder D Sahar; Wallace, Joseph M; Golcuk, Kurtulus; Morris, Michael D.

    2008-01-01

    The mechanical properties of bone are dictated by its amount, distribution and ‘quality’. The composition of the mineral and matrix phases is integral to defining ‘bone quality’. Exercise can potentially increase resistance to fracture, yet the effects of exercise on skeletal fragility, and how alterations in fragility are modulated by the amount, distribution and composition of bone, are unknown. In this investigation, the effects of exercise on the size, composition, mechanical properties a...

  14. A comparative analysis of viral matrix proteins using disorder predictors

    Directory of Open Access Journals (Sweden)

    Dunker A Keith

    2008-10-01

    Full Text Available Abstract Background A previous study (Goh G.K.-M., Dunker A.K., Uversky V.N. (2008 Protein intrinsic disorder toolbox for comparative analysis of viral proteins. BMC Genomics. 9 (Suppl. 2, S4 revealed that HIV matrix protein p17 possesses especially high levels of predicted intrinsic disorder (PID. In this study, we analyzed the PID patterns in matrix proteins of viruses related and unrelated to HIV-1. Results Both SIVmac and HIV-1 p17 proteins were predicted by PONDR VLXT to be highly disordered with subtle differences containing 50% and 60% disordered residues, respectively. SIVmac is very closely related to HIV-2. A specific region that is predicted to be disordered in HIV-1 is missing in SIVmac. The distributions of PID patterns seem to differ in SIVmac and HIV-1 p17 proteins. A high level of PID for the matrix does not seem to be mandatory for retroviruses, since Equine Infectious Anemia Virus (EIAV, an HIV cousin, has been predicted to have low PID level for the matrix; i.e. its matrix protein p15 contains only 21% PID residues. Surprisingly, the PID percentage and the pattern of predicted disorder distribution for p15 resemble those of the influenza matrix protein M1 (25%. Conclusion Our data might have important implications in the search for HIV vaccines since disorder in the matrix protein might provide a mechanism for immune evasion.

  15. Bone graft substitutes and bone morphogenetic proteins for osteoporotic fractures: What is the evidence?

    NARCIS (Netherlands)

    E.M.M. van Lieshout (Esther); V. Alt (Volker)

    2016-01-01

    textabstractDespite improvements in implants and surgical techniques, osteoporotic fractures remain challenging to treat. Among other major risk factors, decreased expression of morphogenetic proteins has been identified for impaired fracture healing in osteoporosis. Bone grafts or bone graft

  16. Bone graft substitutes and bone morphogenetic proteins for osteoporotic fractures: What is the evidence?

    NARCIS (Netherlands)

    E.M.M. van Lieshout (Esther); V. Alt (Volker)

    2016-01-01

    textabstractDespite improvements in implants and surgical techniques, osteoporotic fractures remain challenging to treat. Among other major risk factors, decreased expression of morphogenetic proteins has been identified for impaired fracture healing in osteoporosis. Bone grafts or bone graft substi

  17. Erythropoietin Modulates the Structure of Bone Morphogenetic Protein 2–Engineered Cranial Bone

    OpenAIRE

    Sun, Hongli; Jung, Younghun; Shiozawa, Yusuke; Taichman, Russell S.; Krebsbach, Paul H.

    2012-01-01

    The ideally engineered bone should have similar structural and functional properties to the native tissue. Although structural integrity is critical for functional bone regeneration, we know less about modulating the structural properties of the engineered bone elicited by bone morphogenetic protein (BMP) than efficacy and safety. Erythropoietin (Epo), a primary erythropoietic hormone, has been used to augment blood transfusion in orthopedic surgery. However, the effects of Epo on bone regene...

  18. An Overview of the Medical Applications of Marine Skeletal Matrix Proteins

    Directory of Open Access Journals (Sweden)

    M. Azizur Rahman

    2016-09-01

    Full Text Available In recent years, the medicinal potential of marine organisms has attracted increasing attention. This is due to their immense diversity and adaptation to unique ecological niches that has led to vast physiological and biochemical diversification. Among these organisms, marine calcifiers are an abundant source of novel proteins and chemical entities that can be used for drug discovery. Studies of the skeletal organic matrix proteins of marine calcifiers have focused on biomedical applications such as the identification of growth inducing proteins that can be used for bone regeneration, for example, 2/4 bone morphogenic proteins (BMP. Although a few reports on the functions of proteins derived from marine calcifiers can be found in the literature, marine calcifiers themselves remain an untapped source of proteins for the development of innovative pharmaceuticals. Following an overview of the current knowledge of skeletal organic matrix proteins from marine calcifiers, this review will focus on various aspects of marine skeletal protein research including sources, biosynthesis, structures, and possible strategies for chemical or physical modification. Special attention will be given to potential medical applications and recent discoveries of skeletal proteins and polysaccharides with biologically appealing characteristics. In addition, I will introduce an effective protocol for sample preparation and protein purification that includes isolation technology for biopolymers (of both soluble and insoluble organic matrices from coralline algae. These algae are a widespread but poorly studied group of shallow marine calcifiers that have great potential for marine drug discovery.

  19. An Overview of the Medical Applications of Marine Skeletal Matrix Proteins

    Science.gov (United States)

    Rahman, M. Azizur

    2016-01-01

    In recent years, the medicinal potential of marine organisms has attracted increasing attention. This is due to their immense diversity and adaptation to unique ecological niches that has led to vast physiological and biochemical diversification. Among these organisms, marine calcifiers are an abundant source of novel proteins and chemical entities that can be used for drug discovery. Studies of the skeletal organic matrix proteins of marine calcifiers have focused on biomedical applications such as the identification of growth inducing proteins that can be used for bone regeneration, for example, 2/4 bone morphogenic proteins (BMP). Although a few reports on the functions of proteins derived from marine calcifiers can be found in the literature, marine calcifiers themselves remain an untapped source of proteins for the development of innovative pharmaceuticals. Following an overview of the current knowledge of skeletal organic matrix proteins from marine calcifiers, this review will focus on various aspects of marine skeletal protein research including sources, biosynthesis, structures, and possible strategies for chemical or physical modification. Special attention will be given to potential medical applications and recent discoveries of skeletal proteins and polysaccharides with biologically appealing characteristics. In addition, I will introduce an effective protocol for sample preparation and protein purification that includes isolation technology for biopolymers (of both soluble and insoluble organic matrices) from coralline algae. These algae are a widespread but poorly studied group of shallow marine calcifiers that have great potential for marine drug discovery. PMID:27626432

  20. An Overview of the Medical Applications of Marine Skeletal Matrix Proteins.

    Science.gov (United States)

    Rahman, M Azizur

    2016-09-12

    In recent years, the medicinal potential of marine organisms has attracted increasing attention. This is due to their immense diversity and adaptation to unique ecological niches that has led to vast physiological and biochemical diversification. Among these organisms, marine calcifiers are an abundant source of novel proteins and chemical entities that can be used for drug discovery. Studies of the skeletal organic matrix proteins of marine calcifiers have focused on biomedical applications such as the identification of growth inducing proteins that can be used for bone regeneration, for example, 2/4 bone morphogenic proteins (BMP). Although a few reports on the functions of proteins derived from marine calcifiers can be found in the literature, marine calcifiers themselves remain an untapped source of proteins for the development of innovative pharmaceuticals. Following an overview of the current knowledge of skeletal organic matrix proteins from marine calcifiers, this review will focus on various aspects of marine skeletal protein research including sources, biosynthesis, structures, and possible strategies for chemical or physical modification. Special attention will be given to potential medical applications and recent discoveries of skeletal proteins and polysaccharides with biologically appealing characteristics. In addition, I will introduce an effective protocol for sample preparation and protein purification that includes isolation technology for biopolymers (of both soluble and insoluble organic matrices) from coralline algae. These algae are a widespread but poorly studied group of shallow marine calcifiers that have great potential for marine drug discovery.

  1. Gray-Level Co-occurrence Matrix Bone Fracture Detection

    Directory of Open Access Journals (Sweden)

    Hum Y. Chai

    2011-01-01

    Full Text Available Problem statement: Currently doctors in orthopedic wards inspect the bone x-ray images according to their experience and knowledge in bone fracture analysis. Manual examination of x-rays has multitude drawbacks. The process is time-consuming and subjective. Approach: Since detection of fractures is an important orthopedics and radiologic problem and therefore a Computer Aided Detection(CAD system should be developed to improve the scenario. In this study, a fracture detection CAD based on GLCM recognition could improve the current manual inspection of x-ray images system. The GLCM for fracture and non-fracture bone is computed and analysis is made. Features of Homogeneity, contrast, energy, correlation are calculated to classify the fractured bone. Results: 30 images of femur fractures have been tested, the result shows that the CAD system can differentiate the x-ray bone into fractured and nonfractured femur. The accuracy obtained from the system is 86.67. Conclusion: The CAD system is proved to be effective in classifying the digital radiograph of bone fracture. However the accuracy rate is not perfect, the performance of this system can be further improved using multiple features of GLCM and future works can be done on classifying the bone into different degree of fracture specifically.

  2. Evaluation of bone morphogenic proteins in periodontal practice.

    Science.gov (United States)

    Kaur, Supreet; Grover, Vishakha; Kaur, Harkiran; Malhotra, Ranjan

    2016-01-01

    Forty years ago Marshal R. Urist discovered a substance in bone matrix that had inductive properties for the development of bone and cartilage, until date, at least 20 bone morphogenetic proteins (BMPs) have been identified, some of which have been shown in vitro to stimulate the process of stem cell differentiation into osteoblasts in human and animal models. The purpose of this paper is to give a brief overview of BMPs and to review critically the clinical data currently available on the use of BMPs in various periodontal applications. The literature on BMPs was reviewed. A comprehensive search was designed. The articles were independently screened for eligibility. Articles with authentic controls and proper randomization and pertaining specifically to their role in periodontal applications were included. The available literature was analyzed and compiled. The analysis indicates BMPs to be a promising, as well as an effective novel approach to reconstruct and engineer the periodontal apparatus. Here, we represent several articles, as well as recent texts that make up a special and an in-depth review on the subject. On the basis of the data provided in the studies that were reviewed BMPs provide revolutionary therapies in periodontal practice.

  3. Use of bone morphogenetic proteins in mesenchymal stemcell stimulation of cartilage and bone repair

    Institute of Scientific and Technical Information of China (English)

    2016-01-01

    The extracellular matrix-associated bone morphogeneticproteins (BMPs) govern a plethora of biological processes.The BMPs are members of the transforming growthfactor-β protein superfamily, and they actively participateto kidney development, digit and limb formation,angiogenesis, tissue fibrosis and tumor development.Since their discovery, they have attracted attentionfor their fascinating perspectives in the regenerativemedicine and tissue engineering fields. BMPs havebeen employed in many preclinical and clinical studiesexploring their chondrogenic or osteoinductive potentialin several animal model defects and in human diseases.During years of research in particular two BMPs, BMP2and BMP7 have gained the podium for their use inthe treatment of various cartilage and bone defects.In particular they have been recently approved foremployment in non-union fractures as adjunct therapies.On the other hand, thanks to their potentialities inbiomedical applications, there is a growing interest instudying the biology of mesenchymal stem cell (MSC),the rules underneath their differentiation abilities, andto test their true abilities in tissue engineering. In fact,the specific differentiation of MSCs into targeted celltypelineages for transplantation is a primary goal of theregenerative medicine. This review provides an overviewon the current knowledge of BMP roles and signaling inMSC biology and differentiation capacities. In particularthe article focuses on the potential clinical use of BMPsand MSCs concomitantly, in cartilage and bone tissuerepair.

  4. Demineralized Bone Matrix Scaffolds Modified by CBD-SDF-1α Promote Bone Regeneration via Recruiting Endogenous Stem Cells.

    Science.gov (United States)

    Shi, Jiajia; Sun, Jie; Zhang, Wen; Liang, Hui; Shi, Qin; Li, Xiaoran; Chen, Yanyan; Zhuang, Yan; Dai, Jianwu

    2016-10-07

    The reconstruction of bone usually depends on substitute transplantation, which has drawbacks including the limited bone substitutes available, comorbidity, immune rejection, and limited endogenous bone regeneration. Here, we constructed a functionalized bone substitute by combining application of the demineralized bone matrix (DBM) and collagen-binding stromal-cell-derived factor-1α (CBD-SDF-1α). DBM was a poriferous and biodegradable bone substitute, derived from bovine bone and consisting mainly of collagen. CBD-SDF-1α could bind to collagen and be controllably released from the DBM to mobilize stem cells. In a rat femur defect model, CBD-SDF-1α-modified DBM scaffolds could efficiently mobilize CD34(+) and c-kit(+) endogenous stem cells homing to the injured site at 3 days after implantation. According to the data from micro-CT, CBD-SDF-1α-modified DBM scaffolds could help the bone defects rejoin with mineralization accumulated and bone volume expanded. Interestingly, osteoprotegerin (OPG) and osteopontin (OPN) were highly expressed in CBD-SDF-1α group at an early time after implantation, while osteocalcin (OCN) was more expanded. H&E and Masson's trichrome staining showed that the CBD-SDF-1α-modified DBM scaffold group had more osteoblasts and that the bone defect rejoined earlier. The ultimate strength of the regenerated bone was investigated by three-point bending, showing that the CBD-SDF-1α group had superior strength. In conclusion, CBD-SDF-1α-modified DBM scaffolds could promote bone regeneration by recruiting endogenous stem cells.

  5. OPTICAL DENSITY OF CORTICAL BONE MATRIX IS DIMINISHED IN EXPERIMENTALLY INDUCED OSTEOPOROSIS

    Directory of Open Access Journals (Sweden)

    Jovan Janić

    2016-06-01

    Full Text Available Osteoporosis is characterized by low bone mineral density (BMD and abnormalities in bone structural and material properties, with unexplained low trauma fractures. The aim of the present study was to quantify the optical density of cortical bone matrix in rats with experimentally induced osteoporosis by ovariectomy. The experimental group was divided in two equal subgroups, the first sacrificed in the third month after ovariectomy and second sacrificed in the fifth month after ovariectomy. After decalcification, on routinely stained histopathologic sections optical density (OD, standard deviation of OD, mode OD, minimal and maximal OD of cortical bone matrix were estimated. Mean optical density and mode optical density of cortical bone were statistically higher in the control than in the experimental group (p<0.05. Maximal optical density of cortical bone was significantly lower in rats three months after ovariectomy than in other groups. Obtained results indicate that in experimentally induced osteoporosis the optical density of cortical bone matrix is diminished, similarly to low bone mineral density.

  6. Human Bone Matrix Changes During Deep Saturation Dives

    Science.gov (United States)

    2008-08-08

    agreement notwithstanding, much remains unknown about its pathogenesis, prevention, and treatment . DON is currently disqualifying for U.S. Navy divers...recourse for symptomatic treatment is surgical joint replacement.7 The principal mechanism of bone injury is generally accepted to be bubble formation...urine concentrations of Ntx have been demonstrated in bone diseases such as osteoporosis, primary hyperthyroidism , and Paget’s disease. Also

  7. Analysis of bone protein and mineral composition in bone disease using synchrotron infrared microspectroscopy

    Science.gov (United States)

    Miller, Lisa M.; Hamerman, David; Chance, Mark R.; Carlson, Cathy S.

    1999-10-01

    by lower mineral/protein ratios and higher acid phosphate content. This immature bone may also be a symptom of slower bone formation rates related to estrogen deficiency.

  8. Reconstitution of bone-like matrix in osteogenically differentiated mesenchymal stem cell–collagen constructs: A three-dimensional in vitro model to study hematopoietic stem cell niche

    Directory of Open Access Journals (Sweden)

    WY Lai

    2013-10-01

    Full Text Available Mesenchymal stem/stromal cells (MSCs and osteoblasts are important niche cells for hematopoietic stem cells (HSCs in bone marrow osteoblastic niche. Here, we aim to partially reconstitute the bone marrow HSC niche in vitro using collagen microencapsulation for investigation of the interactions between HSCs and MSCs. Mouse MSCs (mMSCs microencapsulated in collagen were osteogenically differentiated to derive a bone-like matrix consisting of osteocalcin, osteopontin, and calcium deposits and secreted bone morphogenic protein 2 (BMP2. Decellularized bone-like matrix was seeded with fluorescence-labeled human MSCs and HSCs. Comparing with pure collagen scaffold, significantly more HSCs and HSC–MSC pairs per unit area were found in the decellularized bone-like matrix. Moreover, incubation with excess neutralizing antibody of BMP2 resulted in a significantly higher number of HSC per unit area than that without in the decellularized matrix. This work suggests that the osteogenic differentiated MSC–collagen microsphere is a valuable three-dimensional in vitro model to elucidate cell–cell and cell–matrix interactions in HSC niche.

  9. Cross-talk between bone morphogenetic proteins and inflammatory pathways.

    Science.gov (United States)

    van der Kraan, Peter M; Davidson, Esmeralda N Blaney

    2015-11-23

    Pro-inflammatory cytokines and bone morphogenetic proteins are generally studied separately and considered to be elements of different worlds, immunology and developmental biology. Varas and colleagues report that these factors show cross-talk in rheumatoid arthritis synoviocytes. They show that pro-inflammatory cytokines not only stimulate the production of bone morphogenetic proteins but that these endogenously produced bone morphogenetic proteins interfere with the effects of pro-inflammatory cytokines on synoviocytes.

  10. PTH(1-84) Administration in Hypoparathyroidism Transiently Reduces Bone Matrix Mineralization.

    Science.gov (United States)

    Misof, Barbara M; Roschger, Paul; Dempster, David W; Zhou, Hua; Bilezikian, John P; Klaushofer, Klaus; Rubin, Mishaela R

    2016-01-01

    Patients with hypoparathyroidism have low circulating parathyroid (PTH) levels and higher cancellous bone volume and trabecular thickness. Treatment with PTH(1-84) was shown to increase abnormally low bone remodeling dynamics. In this work, we studied the effect of 1-year or 2-year PTH(1-84) treatment on cancellous and cortical bone mineralization density distribution (Cn.BMDD and Ct.BMDD) based on quantitative backscattered electron imaging (qBEI) in paired transiliac bone biopsy samples. The study cohort comprised 30 adult hypoparathyroid patients (14 treated for 1 year; 16 treated for 2 years). At baseline, Cn.BMDD was shifted to higher mineralization densities in both treatment groups (average degree of mineralization Cn.CaMean +3.9% and +2.7%, p mineralizing surface) was predictive for Cn.BMDD outcomes in the 1-year PTH(1-84) group, but not in the 2-year PTH(1-84) group. Our findings suggest higher baseline bone matrix mineralization consistent with the decreased bone turnover in hypoparathyroidism. PTH(1-84) treatment caused differential effects dependent on treatment duration that were consistent with the histomorphometric bone formation outcomes. The greater increase in bone formation during the first year of treatment was associated with a decrease in bone matrix mineralization, suggesting that PTH(1-84) exposure to the hypoparathyroid skeleton has the greatest effects on BMDD early in treatment.

  11. Differential regulation of dentin matrix protein 1 expression during odontogenesis.

    Science.gov (United States)

    Lu, Yongbo; Zhang, Shubin; Xie, Yixia; Pi, Yuli; Feng, Jian Q

    2005-01-01

    Dentin matrix protein 1 (DMP1) is highly expressed in mineralized tooth and bone. Both in vitro and in vivo data show that DMP1 is critical for mineralization and tooth morphogenesis (growth and development). In this study, we studied Dmp1 gene regulation. The in vitro transient transfection assay identified two important DNA fragments, the 2.4- and 9.6-kb promoter regions. We next generated and analyzed transgenic mice bearing the beta-galactosidase (lacZ) reporter gene driven by the 2.4- or 9.6-kb promoter with the complete 4-kb intron 1. The 9.6-kb Dmp1-lacZ mice conferred a DMP1 expression pattern in odontoblasts identical to that in the endogenous Dmp1 gene. This is reflected by lacZ expression in Dmp1-lacZ knock-in mice during all stages of odontogenesis. In contrast, the 2.4-kb Dmp1-lacZ mice display activity in odontoblast cells only at the early stage of odontogenesis. Thus, we propose that different transcription factors regulate early or later cis-regulatory domains of the Dmp1 promoter, which gives rise to the unique spatial and temporal expression pattern of Dmp1 gene at different stages of tooth development. 2005 S. Karger AG, Basel

  12. Nipah virus matrix protein: expert hacker of cellular machines.

    Science.gov (United States)

    Watkinson, Ruth E; Lee, Benhur

    2016-08-01

    Nipah virus (NiV, Henipavirus) is a highly lethal emergent zoonotic paramyxovirus responsible for repeated human outbreaks of encephalitis in South East Asia. There are no approved vaccines or treatments, thus improved understanding of NiV biology is imperative. NiV matrix protein recruits a plethora of cellular machinery to scaffold and coordinate virion budding. Intriguingly, matrix also hijacks cellular trafficking and ubiquitination pathways to facilitate transient nuclear localization. While the biological significance of matrix nuclear localization for an otherwise cytoplasmic virus remains enigmatic, the molecular details have begun to be characterized, and are conserved among matrix proteins from divergent paramyxoviruses. Matrix protein appropriation of cellular machinery will be discussed in terms of its early nuclear targeting and later role in virion assembly.

  13. THE MEASUREMENT OF BONE QUALITY USING GRAY LEVEL CO-OCCURRENCE MATRIX TEXTURAL FEATURES.

    Science.gov (United States)

    Shirvaikar, Mukul; Huang, Ning; Dong, Xuanliang Neil

    2016-10-01

    In this paper, statistical methods for the estimation of bone quality to predict the risk of fracture are reported. Bone mineral density and bone architecture properties are the main contributors of bone quality. Dual-energy X-ray Absorptiometry (DXA) is the traditional clinical measurement technique for bone mineral density, but does not include architectural information to enhance the prediction of bone fragility. Other modalities are not practical due to cost and access considerations. This study investigates statistical parameters based on the Gray Level Co-occurrence Matrix (GLCM) extracted from two-dimensional projection images and explores links with architectural properties and bone mechanics. Data analysis was conducted on Micro-CT images of 13 trabecular bones (with an in-plane spatial resolution of about 50μm). Ground truth data for bone volume fraction (BV/TV), bone strength and modulus were available based on complex 3D analysis and mechanical tests. Correlation between the statistical parameters and biomechanical test results was studied using regression analysis. The results showed Cluster-Shade was strongly correlated with the microarchitecture of the trabecular bone and related to mechanical properties. Once the principle thesis of utilizing second-order statistics is established, it can be extended to other modalities, providing cost and convenience advantages for patients and doctors.

  14. Aluminum and iron can be deposited in the calcified matrix of bone exostoses.

    Science.gov (United States)

    Chappard, Daniel; Mabilleau, Guillaume; Moukoko, Didier; Henric, Nicolas; Steiger, Vincent; Le Nay, Patrick; Frin, Jean-Marie; De Bodman, Charlotte

    2015-11-01

    Exostosis (or osteochondroma) is the most common benign bone tumor encountered in children and adults. Exostoses may occur as solitary or multiple tumors (in the autosomal syndromes of hereditary multiple exostoses). Exostoses are composed of cortical and medullary bone covered by an overlying hyaline cartilage cap. We have searched iron (Fe) and aluminum (Al) in the matrix of cortical and trabecular bone of 30 patients with exostosis. Al(3+) and Fe(3+) are two cations which can substitute calcium in the hydroxyapatite crystals of the bone matrix. The bone samples were removed surgically and were studied undecalcified. Perls' Prussian blue staining (for Fe) and solochrome azurine B (for Al) were used on the histological sections of the tumors. Al(3+) was detected histochemically in 21/30 patients as linear bands deposited by the osteoblasts. Fe(3+) was detected in 10 out of these 21 patients as linear bands in the same locations. Fe(3+) and Al(3+) were not identified in the bone matrix of a control group of 20 osteoporotic patients. Energy X-ray Dispersive Spectrometry failed to identify Fe and Al in bone of these tumors due to the low sensitivity of the method. Wavelength Dispersive Spectrometry identified them but the concentrations were very low. Histochemistry appears a very sensitive method for Fe(3+) and Al(3+) in bone.The presence of these two metals in the exostoses advocates for a disturbed metabolism of osteoblasts which can deposit these metals into the bone matrix, similar to which is observed in case of hemochromatosis with Fe(3+).

  15. Cartilage oligomeric matrix protein in patients with juvenile idiopathic arthritis

    DEFF Research Database (Denmark)

    Bjørnhart, Birgitte; Juul, Anders; Nielsen, Susan

    2009-01-01

    Cartilage oligomeric matrix protein (COMP) has been identified as a prognostic marker of progressive joint destruction in rheumatoid arthritis. In this population based study we evaluated associations between plasma concentrations of COMP, disease activity, and growth velocity in patients...

  16. Nuclear variants of bone morphogenetic proteins

    Directory of Open Access Journals (Sweden)

    Meinhart Christopher A

    2010-03-01

    Full Text Available Abstract Background Bone morphogenetic proteins (BMPs contribute to many different aspects of development including mesoderm formation, heart development, neurogenesis, skeletal development, and axis formation. They have previously been recognized only as secreted growth factors, but the present study detected Bmp2, Bmp4, and Gdf5/CDMP1 in the nuclei of cultured cells using immunocytochemistry and immunoblotting of nuclear extracts. Results In all three proteins, a bipartite nuclear localization signal (NLS was found to overlap the site at which the proproteins are cleaved to release the mature growth factors from the propeptides. Mutational analyses indicated that the nuclear variants of these three proteins are produced by initiating translation from downstream alternative start codons. The resulting proteins lack N-terminal signal peptides and are therefore translated in the cytoplasm rather than the endoplasmic reticulum, thus avoiding proteolytic processing in the secretory pathway. Instead, the uncleaved proteins (designated nBmp2, nBmp4, and nGdf5 containing the intact NLSs are translocated to the nucleus. Immunostaining of endogenous nBmp2 in cultured cells demonstrated that the amount of nBmp2 as well as its nuclear/cytoplasmic distribution differs between cells that are in M-phase versus other phases of the cell cycle. Conclusions The observation that nBmp2 localization varies throughout the cell cycle, as well as the conservation of a nuclear localization mechanism among three different BMP family members, suggests that these novel nuclear variants of BMP family proteins play an important functional role in the cell.

  17. Nanoporous Structure of Bone Matrix at Osteoporosis from Data of Atomic Force Microscopy and IR Spectroscopy

    Directory of Open Access Journals (Sweden)

    A. A. Gaidash

    2011-01-01

    Full Text Available It was found that in an osteoporotic bone the fraction of nanosized pores decreases, the mineral phase amorphizes, hydrated shells around mineralized particles of the bone matrix thicken, and adhesion forces increase. This contributes to the formation of water clusters similar to bulk water clusters compared to the healthy bone tissue and leads to the accumulation of more viscous liquid with increased intermolecular interaction forces in the pores of the bone matrix. Given this, the rates of chemical reactions proceeding in the water phase of ultrathin channels of general parts of collagen fibrils decrease. Ultimately, nanopores of collagen-apatite interfaces lose, to a certain extent, the capability of catalyzing the hydroxyapatite crystallization.

  18. Histologic and clinical evaluation of an allogeneic bone matrix for the treatment of periodontal osseous defects.

    Science.gov (United States)

    Mellonig, James T

    2006-12-01

    The objective of this study was to evaluate the potential of an allogeneic bone matrix (Grafton, Osteotech) to regenerate new bone, new cementum, and a new periodontal ligament around teeth previously contaminated by bacterial plaque. Four patients with chronic advanced periodontitis and who were scheduled for full-mouth extraction were enrolled in the study. One patient dropped out from the study before any therapy began. One tooth with an intraosseous defect in each patient was selected for treatment. Measurements of probing depth, gingival recession, and clinical attachment level were made. After flap reflection, a root notch was placed at the apical level of calculus, the root was debrided, and allogenic bone matrix was inserted into the defect. After 6 months of healing, the teeth were removed en bloc and evaluated histologically for a new attachment apparatus. Two of the three teeth demonstrated regeneration of new bone, cementum, and periodontal ligament.

  19. Bisphosphonate treatment affects trabecular bone apparent modulus through micro-architecture rather than matrix properties

    DEFF Research Database (Denmark)

    Ding, Ming

    2004-01-01

    and trabecular architecture independently. Conventional histomorphometry and microdamage data were obtained from the second and third lumbar vertebrae of the same dogs [Bone 28 (2001) 524]. Bisphosphonate treatment resulted in an increased apparent Young's modulus, decreased bone turnover, increased calcified......Bisphosphonates are emerging as an important treatment for osteoporosis. But whether the reduced fracture risk associated with bisphosphonate treatment is due to increased bone mass, improved trabecular architecture and/or increased secondary mineralization of the calcified matrix remains unclear....... We examined the effects of bisphosphonates on both the trabecular architecture and matrix properties of canine trabecular bone. Thirty-six beagles were divided into a control group and two treatment groups, one receiving risedronate and the other alendronate at 5-6 times the clinical dose...

  20. Multifunctional Bone Morphogenetic Protein System in Endocrinology

    Directory of Open Access Journals (Sweden)

    Otsuka,Fumio

    2013-04-01

    Full Text Available New biological activities of bone morphogenetic proteins (BMPs in the endocrine system have recently been revealed. The BMP system is composed of approximately 30 ligands and preferential combinations of type I and type II receptors. The BMP system not only induces bone formation but also plays unique tissue-specific roles in various organs. For instance, the ovarian BMP system is a physiological inhibitor of luteinization in growing ovarian follicles. In the ovary, the expression of oocyte-derived BMP-15 is critical for female reproduction. In the pituitary, BMP-4 is a key player for initial development of the anterior pituitary, while it is also functionally involved in some differentiated pituitary tumors, including prolactinoma and Cushingʼs disease. In the adrenal glands, BMP-6 and BMP-4 modulate aldosterone and catecholamine production, respectively, which contributes to a functional interaction between the cortex and medulla. In the present review, recent advances in BMP biology in the field of endocrinology are described and the possibility for clinical application of BMP activity is discussed.

  1. Relation Between Bone Matrix Proteins and Monocyte Chemoattractant Protein-1 in Renal Small Artery in Diabetic Nephropathy Rats%糖尿病肾病大鼠肾小动脉骨基质蛋白表达及其与MCP-1的关系

    Institute of Scientific and Technical Information of China (English)

    赵安菊; 黄颂敏; 欧三桃; 刘芳; 陈泽君; 胡章学; 唐万欣

    2012-01-01

    of bone matrix proteins and monocyte chemoattractant protein-1 ((MCP-1) in the renal arteriole of diabetic nephropathy CDN) rats and analyze their correlations and roles in diabetic nephropathy. Methods Adult Sprague-Dawley male rats were used to establish the animal model of diabetic nephropathy induced by peritoneal injection of 55 mg/kg of streptozocin. Calcium deposit around the renal arteriole was observed by alizarin red staining. The protein and mRNA levels of core-bind factor alpha 1 (cbfα1), bone morphogenetic protein 2 (BMP-2) and matrix Gla protein (MGP) in renal arteriole of DN rats were detected by immunohistochemistry, in-situ hybridization and real-time PCR. The biochemical indices were detected by routine test. Results ①Blood glucose and Urine protein of 24 h were significantly increased in the renal arteriole of DN rats vs. the control rats (P<0. 05),serum creatinine (SCr) and phosphorus were significantly increased from 12 weeks. ② Little deposit of calcium salt was observed in the renal arteriole of DN rats at the 4th week and a large amount of deposit was observed at 24th week, but no calcium deposit was observed in control rats. ③Cbfal and BMP-2 expressions were significantly increased in the renal arteriole of DN rats from 4 to 24 weeks vs. the control rats. MGP mRNA expression in the renal arteriole of DN rats was significantly decreased from 4 to 24 weeks. MCP-1 expression was obviously upregulated in the renal arteriole of DN rats at 24th week vs. that at 4th and 12th week. No MCP-1 expression was observed in the renal arterioles of control rats. ④MCP-1 were positively correlated with the expression of cbfα1 and BMP-2. Conclusion bone matrix proteins has already expressed in renal arteriole before the formation of vascular calcification. MCP-1 can affect the expression of cbfα1,BMP-2 ;Cbfα1. BMP-2, MGP and MCP-1 may be involved in the formation of vascular lesions of DN.

  2. Hybrid Matrix Grafts to Favor Tissue Regeneration in Rabbit Femur Bone Lesions

    Science.gov (United States)

    Goy, Dante Pascual; Gorosito, Emmanuel; Costa, Hermes S; Mortarino, Pablo; Pedemonte, Noelia Acosta; Toledo, Javier; Mansur, Herman S; Pereira, Marivalda M; Battaglino, Ricardo; Feldman, Sara

    2012-01-01

    At present, typical approaches employed to repair fractures and other bone lesions tend to use matrix grafts to promote tissue regeneration. These grafts act as templates, which promote cellular adhesion, growth and proliferation, osteoconduction, and even osteoinduction, which commonly results in de novo osteogenesis. The present work aimed to study the bone-repairing ability of hybrid matrixes (HM) prepared with polyvinyl alcohol (PVA) and bioactive glass in an experimental rabbit model. The HM were prepared by combining 30% bioactive glass (nominal composition of 58% SiO2 -33 % CaO - 9% P2O5) and 70% PVA. New Zealand rabbits were randomly divided into the control group (C group) and two groups with bone lesions, in which one received a matrix implant HM (Implant group), while the other did not (no Implant group). Clinical monitoring showed no altered parameters from either the Implant or the no Implant groups as compared to the control group, for the variables of diet grades, day and night temperatures and hemograms. In the Implant group, radiologic and tomographic studies showed implanted areas with clean edges in femoral non-articular direction, and radio-dense images that suggest incipient integration. Minimum signs of phlogosis could be observed, whereas no signs of rejection at this imaging level could be identified. Histological analysis showed evidence of osteo-integration, with the formation of a trabecular bone within the implant. Together, these results show that implants of hybrid matrixes of bioactive glass are capable of promoting bone regeneration. PMID:22848334

  3. Ultrastructural studies on the origin and structure of matrix vesicles in bone of young rats.

    Science.gov (United States)

    Ornoy, A; Atkin, I; Levy, J

    1980-01-01

    Tibiae of young rats were examined by scanning electron microscopy (SEM) and transmission electron microscopy (TEM) in order to study the origin and structure of matrix vesicles in periosteal bone. SEM studies have shown that periosteal osteoblasts have elongated processes with globular structures of 0.1 micron in diameter attached to the cell surface and processes. Similar structures were found to cover the inner surface of osteoblastic lacunae. The SEM studies have further shown that in the periosteal surface (forming bone) the above-described globules once mineralized, aggregated to form larger, nonhomogeneous mineralized spherules in which, by proper treatment with NaOCl, hydroxyapatite crystals could be exposed. Endosteal osteoblasts had fewer processes, devoid of the globular structures. Similarly, osteocytic and osteoclastic processes, although elongated and numberous, were not covered by the globular structures. In the matrix, collagen fibers of forming bone were randomly orientated, while in the deeper areas of bone they formed bundles with a longitudinal orientation. TEM studies have shown that the structures found on the osteoblastic surface and in the matrix are membrane-bound matrix vesicles which seem to be formed by budding from cell processes. Preformed membrane-bound vesicles were also observed by TEM inside sections of osteoblastic processes. These vesicles resembled the extracellular matrix vesicles in size and shape, thus giving the impression that at least some of the matrix vesicles are preformed cellular structures. While comparing SEM with TEM, it can be conducted that in bone, as in cartilage, matrix vesicles which probably serve as the initial locus of calcification, are formed directly by osteoblasts.

  4. Titanium implants induce expression of matrix metalloproteinases in bone during osseointegration.

    Science.gov (United States)

    Shubayev, Veronica I; Brånemark, Rickard; Steinauer, Joanne; Myers, Robert R

    2004-01-01

    Implanted pure titanium fixtures are able to completely integrate with bone, in part because of the formation of a strong extracellular matrix (ECM) bond at the titanium-bone interface. In this study, we used a rodent femur model of intramedullary osseointegration to analyze the changes in immunoreactivity of ECM-controlling matrix metalloproteinases (MMPs), tissue inhibitor of metalloproteinase-3 (TIMP-3), and tumor necrosis factor alpha (TNF-alpha) during osseointegration. We observed dramatic increases in MMP-2, MMP-9, MMP-7, TIMP-3, and TNF-alpha in osteocytes, osteoclasts, haversian canals, and the interface matrix in bone ipsilateral to the titanium implant. An increase in TIMP-3, MMP-9, and MMP-7 in hypertrophied chondrocytes and the vascular component of the epiphysial growth plate was also observed in experimental bone. These findings were not seen in contralateral or sham-operated bone, where the titanium fixtures were threaded into the femur and immediately removed. Our data link titanium-induced bone remodeling to changes in expression and distribution of MMPs.

  5. Modifications in Bone Matrix of Estrogen-Deficient Rats Treated with Intermittent PTH

    Directory of Open Access Journals (Sweden)

    Rafael Pacheco-Costa

    2015-01-01

    Full Text Available Bone matrix dictates strength, elasticity, and stiffness to the bone. Intermittent parathyroid hormone (iPTH, a bone-forming treatment, is widely used as a therapy for osteoporosis. We investigate whether low doses of intermittent PTH (1-34 change the profile of organic components in the bone matrix after 30 days of treatment. Forty 6-month-old female Wistar rats underwent ovariectomy and after 3 months received low doses of iPTH administered for 30 days: daily at 0.3 µg/kg/day (PTH03 or 5 µg/kg/day (PTH5; or 3 times per week at 0.25 µg/kg/day (PTH025. After euthanasia, distal femora were processed for bone histomorphometry, histochemistry for collagen and glycosaminoglycans, biochemical quantification of sulfated glycosaminoglycans, and hyaluronan by ELISA and TUNEL staining. Whole tibiae were used to estimate the bone mineral density (BMD. Histomorphometric analysis showed that PTH5 increased cancellous bone volume by 6% over vehicle-treated rats. In addition, PTH5 and PTH03 increased cortical thickness by 21% and 20%, respectively. Tibial BMD increased in PTH5-treated rats and this group exhibited lower levels of chondroitin sulfate; on the other hand, hyaluronan expression was increased. Hormonal administration in the PTH5 group led to decreased collagen maturity. Further, TUNEL-positive osteocytes were decreased in the cortical compartment of PTH5 whereas administration of PTH025 increased the osteocyte death. Our findings suggest that daily injections of PTH at low doses alter the pattern of organic components from the bone matrix, favoring the increase of bone mass.

  6. The requirement of matrix ATP for the import of precursor proteins into the mitochondrial matrix and intermembrane space

    NARCIS (Netherlands)

    Stuart, Rosemary A.; Gruhler, Albrecht; Klei, Ida van der; Guiard, Bernard; Koll, Hans; Neupert, Walter

    1994-01-01

    The role of ATP in the matrix for the import of precursor proteins into the various mitochondrial subcompartments was investigated by studying protein translocation at experimentally defined ATP levels. Proteins targeted to the matrix were neither imported or processed when matrix ATP was depleted.

  7. Recent advances in the study of zebrafish extracellular matrix proteins.

    Science.gov (United States)

    Jessen, Jason R

    2015-05-01

    The zebrafish extracellular matrix (ECM) is a dynamic and pleomorphic structure consisting of numerous proteins that together regulate a variety of cellular and morphogenetic events beginning as early as gastrulation. The zebrafish genome encodes a similar complement of ECM proteins as found in other vertebrate organisms including glycoproteins, fibrous proteins, proteoglycans, glycosaminoglycans, and interacting or modifying proteins such as integrins and matrix metalloproteinases. As a genetic model system combined with its amenability to high-resolution microscopic imaging, the zebrafish allows interrogation of ECM protein structure and function in both the embryo and adult. Accumulating data have identified important roles for zebrafish ECM proteins in processes as diverse as cell polarity, migration, tissue mechanics, organ laterality, muscle contraction, and regeneration. In this review, I highlight recently published data on these topics that demonstrate how the ECM proteins fibronectin, laminin, and collagen contribute to zebrafish development and adult homeostasis.

  8. Vitamin d binding protein and bone health

    National Research Council Canada - National Science Library

    Bhan, Ishir

    2014-01-01

    .... These polymorphisms have been linked with alterations in bone density in several populations. One of the mechanisms by which DBP may alter bone health involves regulating vitamin D bioavailability...

  9. Implants of polyanionic collagen matrix in bone defects of ovariectomized rats.

    Science.gov (United States)

    Cunha, Marcelo Rodrigues; Santos, Arnaldo Rodrigues; Goissis, Gilberto; Genari, Selma C

    2008-03-01

    In recent years, there has been a great interest in the development of biomaterials that could be used in the repair of bone defects. Collagen matrix (CM) has the advantage that it can be modified chemically to improve its mechanical properties. The aim of the present study was to evaluate the effect of three-dimensional membranes of native or anionic (submitted to alkaline treatment for 48 or 96 h) collagen matrix on the consolidation of osteoporosis bone fractures resulting from the gonadal hormone alterations caused by ovariectomy in rats subjected to hormone replacement therapy. The animals received the implants 4 months after ovariectomy and were sacrificed 8 weeks after implantation of the membranes into 4-mm wide bone defects created in the distal third of the femur with a surgical bur. Macroscopic analysis revealed the absence of pathological alterations in the implanted areas, suggesting that the material was biocompatible. Microscopic analysis showed a lower amount of bone ingrowth in the areas receiving the native membrane compared to the bone defects filled with the anionic membranes. In ovariectomized animals receiving anionic membranes, a delay in bone regeneration was observed mainly in animals not subjected to hormone replacement therapy. We conclude that anionic membranes treated with alkaline solution for 48 and 96 h presented better results in terms of bone ingrowth.

  10. Ultra-structural defects cause low bone matrix stiffness despite high mineralization in osteogenesis imperfecta mice☆

    Science.gov (United States)

    Vanleene, Maximilien; Porter, Alexandra; Guillot, Pascale-Valerie; Boyde, Alan; Oyen, Michelle; Shefelbine, Sandra

    2012-01-01

    Bone is a complex material with a hierarchical multi-scale organization from the molecule to the organ scale. The genetic bone disease, osteogenesis imperfecta, is primarily caused by mutations in the collagen type I genes, resulting in bone fragility. Because the basis of the disease is molecular with ramifications at the whole bone level, it provides a platform for investigating the relationship between structure, composition, and mechanics throughout the hierarchy. Prior studies have individually shown that OI leads to: 1. increased bone mineralization, 2. decreased elastic modulus, and 3. smaller apatite crystal size. However, these have not been studied together and the mechanism for how mineral structure influences tissue mechanics has not been identified. This lack of understanding inhibits the development of more accurate models and therapies. To address this research gap, we used a mouse model of the disease (oim) to measure these outcomes together in order to propose an underlying mechanism for the changes in properties. Our main finding was that despite increased mineralization, oim bones have lower stiffness that may result from the poorly organized mineral matrix with significantly smaller, highly packed and disoriented apatite crystals. Using a composite framework, we interpret the lower oim bone matrix elasticity observed as the result of a change in the aspect ratio of apatite crystals and a disruption of the crystal connectivity. PMID:22449447

  11. Novel approaches to bone grafting: porosity, bone morphogenetic proteins, stem cells, and the periosteum.

    Science.gov (United States)

    Petrochenko, Peter; Narayan, Roger J

    2010-01-01

    The disadvantages involving the use of a patient's own bone as graft material have led surgeons to search for alternative materials. In this review, several characteristics of a successful bone graft material are discussed. In addition, novel synthetic materials and natural bone graft materials are being considered. Various factors can determine the success of a bone graft substitute. For example, design considerations such as porosity, pore shape, and interconnection play significant roles in determining graft performance. The effective delivery of bone morphogenetic proteins and the ability to restore vascularization also play significant roles in determining the success of a bone graft material. Among current approaches, shorter bone morphogenetic protein sequences, more efficient delivery methods, and periosteal graft supplements have shown significant promise for use in autograft substitutes or autograft extenders.

  12. Utility of tricalcium phosphate and osteogenic matrix cellsheet constructs for bone defect reconstruction

    Institute of Scientific and Technical Information of China (English)

    2015-01-01

    AIM To determine the effects of transplanting osteogenicmatrix cell sheets and beta-tricalcium phosphate(TCP) constructs on bone formation in bone defects.METHODS: Osteogenic matrix cell sheets were preparedfrom bone marrow stromal cells (BMSCs), anda porous TCP ceramic was used as a scaffold. Threeexperimental groups were prepared, comprised of TCPscaffolds (1) seeded with BMSCs; (2) wrapped withosteogenic matrix cell sheets; or (3) both. Constructswere implanted into a femoral defect model in rats andbone growth was evaluated by radiography, histology,biochemistry, and mechanical testing after 8 wk.RESULTS: In bone defects, constructs implanted withcell sheets showed callus formation with segmental or continuous bone formation at 8 wk, in contrast toTCP seeded with BMSCs, which resulted in bone nonunion.Wrapping TCP constructs with osteogenic matrixcell sheets increased their osteogenic potential andresulting bone formation, compared with conventionalbone tissue engineering TCP scaffolds seeded withBMSCs. The compressive stiffness (mean ± SD) valueswere 225.0 ± 95.7, 30.0 ± 11.5, and 26.3 ± 10.6MPa for BMSC/TCP/Sheet constructs with continuousbone formation, BMSC/TCP/Sheet constructs withsegmental bone formation, and BMSC/TCP constructs,respectively. The compressive stiffness of BMSC/TCP/Sheet constructs with continuous bone formation wassignificantly higher than those with segmental boneformation and BMSC/TCP constructs.CONCLUSION: This technique is an improvementover current methods, such as TCP substitution, andis useful for hard tissue reconstruction and inducingearlier bone union in defects.

  13. Relationships between serum osteoprotegerin, matrix metalloproteinase-2 levels and bone metabolism in postmenopausal women

    Institute of Scientific and Technical Information of China (English)

    DAI Yi; SHEN Lin

    2007-01-01

    Background Serum osteoprotegerin (OPG) and matrix metalloproteinase-2 (MMP-2) have been shown to play a role in bone metabolism by degrading the bone matrix. The present study was undertaken to compare OPG and MMP-2 with bone mineral density and three markers (alkaline phosphatase (AKP), calcium and phosphorus) in postmenopausal women in Wuhan.Methods Serum OPG, MMP-2, and AKP of 78 Chinese postmenopausal women aged 48 to 65 were measured using enzyme-linked immunosorbent assay (ELISA). Bone mineral density was measured with dual energy X-ray absorptiometry (DEXA), and serum calcium and phosphorus were measured by auto biochemical analysis.Results Serum OPG and MMP-2 concentrations were significantly higher in postmenopausal women with osteoporosis ((127.6±6.3) ng/L; (1388±121) μg/L)) than those in age-matched normal controls ((72.3±2.4) ng/L; (1126±141) μg/L,P<0.01). Negative relationships were found between serum OPG, MMP-2 levels and bone mineral density in osteoporotic women. Adjusted by age and body mass index (BMI), the correlation of MMP-2 with bone mineral density of the neck of the femur disappeared. In osteoporotic women, negative correlations between OPG, MMP-2 levels and serum calcium were found (r=-0.216; r=-0.269, P<0.05), but positive correlations between OPG and serum AKP, serum phosphorus (r=0.235; r=0.124, P<0.05).Conclusions Significant correlations exist between serum OPG, MMP-2 levels and bone metabolism in high bone turnover of postmenopausal osteoporotic women. The concentrations of serum OPG and MMP-2 increase possibly as a concomitant event in the high bone turnover state, such as postmenopausal osteoporosis. Therefore serum OPG and MMP-2 could be used as indicators for the bone metabolism in postmenopausal osteoporotic women.

  14. The Bone Morphogenetic Protein 1/Tolloid-like Metalloproteinases

    Science.gov (United States)

    Hopkins, Delana R.; Keles, Sunduz; Greenspan, Daniel S.

    2009-01-01

    A decade ago, bone morphogenetic protein 1 (BMP1) was shown to provide the activity necessary for proteolytic removal of the C-propeptides of procollagens I–III: precursors of the major fibrillar collagens. Subsequent studies have shown BMP1 to be the prototype of a small group of extracellular metalloproteinases that play manifold roles in regulating formation of the extracellular matrix (ECM). Soon after initial cloning of BMP1, genetic studies showed the related Drosophila proteinase Tolloid (TLD) to be necessary for formation of the dorsal-ventral axis in early embryogenesis. It is now clear that the BMP1/TLD-like proteinases, conserved in species ranging from Drosophila to humans, act in dorsal-ventral patterning via activation of transforming growth factor β (TGFβ)-like proteins BMP2, BMP4 (vertebrates) and decapentaplegic (arthropods). More recently, it has become apparent that the BMP1/TLD-like proteinases are key activators of a broader subset of the TGFβ superfamily of proteins, with implications that these proteinases may be key in orchestrating formation of ECM with growth factor activation and BMP signaling in morphogenetic processes. PMID:17560775

  15. Protein structure estimation from NMR data by matrix completion.

    Science.gov (United States)

    Li, Zhicheng; Li, Yang; Lei, Qiang; Zhao, Qing

    2017-02-06

    Knowledge of protein structures is very important to understand their corresponding physical and chemical properties. Nuclear Magnetic Resonance (NMR) spectroscopy is one of the main methods to measure protein structure. In this paper, we propose a two-stage approach to calculate the structure of a protein from a highly incomplete distance matrix, where most data are obtained from NMR. We first randomly "guess" a small part of unobservable distances by utilizing the triangle inequality, which is crucial for the second stage. Then we use matrix completion to calculate the protein structure from the obtained incomplete distance matrix. We apply the accelerated proximal gradient algorithm to solve the corresponding optimization problem. Furthermore, the recovery error of our method is analyzed, and its efficiency is demonstrated by several practical examples.

  16. Aluminum and bone: Review of new clinical circumstances associated with Al(3+) deposition in the calcified matrix of bone.

    Science.gov (United States)

    Chappard, D; Bizot, P; Mabilleau, G; Hubert, L

    2016-06-01

    Several decades ago, aluminum encephalopathy associated with osteomalacia has been recognized as the major complication of chronic renal failure in dialyzed patients. Removal of aluminum from the dialysate has led to a disappearance of the disease. However, aluminum deposit occurs in the hydroxyapatite of the bone matrix in some clinical circumstances that are presented in this review. We have encountered aluminum in bone in patients with an increased intestinal permeability (coeliac disease), or in the case of prolonged administration of aluminum anti-acid drugs. A colocalisation of aluminum with iron was also noted in cases of hemochromatosis and sickle cell anemia. Aluminium was also identified in a series of patients with exostosis, a frequent benign bone tumor. Corrosion of prosthetic implants composed of grade V titanium (TA6V is an alloy containing 6% aluminum and 4% vanadium) was also observed in a series of hip or knee revisions. Aluminum can be identified in undecalcified bone matrix stained by solochrome azurine, a highly specific stain allowing the detection of 0.03 atomic %. Colocalization of aluminum and iron does not seem to be the fruit of chance but the cellular and molecular mechanisms are still poorly understood. Histochemistry is superior to spectroscopic analyses (EDS and WDS in scanning electron microscopy).

  17. The effect of enamel matrix derivative (Emdogain) on bone formation: a systematic review.

    NARCIS (Netherlands)

    Rathe, F.; Junker, R.; Chesnutt, B.C.; Jansen, J.A.

    2009-01-01

    This systematic review focused on the question, if and to what extent enamel matrix derivative (Emdogain) [EMD]) promotes the regeneration of bone. The influence of combinations with other biomaterials was additionally evaluated. Twenty histomorphometric studies were included in this systematic revi

  18. Changes in chemical composition of bone matrix in ovariectomized (OVX) rats detected by Raman spectroscopy and multivariate analysis

    Science.gov (United States)

    Oshima, Yusuke; Iimura, Tadahiro; Saitou, Takashi; Imamura, Takeshi

    2015-02-01

    Osteoporosis is a major bone disease that connotes the risk of fragility fractures resulting from alterations to bone quantity and/or quality to mechanical competence. Bone strength arises from both bone quantity and quality. Assessment of bone quality and bone quantity is important for prediction of fracture risk. In spite of the two factors contribute to maintain the bone strength, only one factor, bone mineral density is used to determine the bone strength in the current diagnosis of osteoporosis. On the other hand, there is no practical method to measure chemical composition of bone tissue including hydroxyapatite and collagen non-invasively. Raman spectroscopy is a powerful technique to analyze chemical composition and material properties of bone matrix non-invasively. Here we demonstrated Raman spectroscopic analysis of the bone matrix in osteoporosis model rat. Ovariectomized (OVX) rat was made and the decalcified sections of tibias were analyzed by a Raman microscope. In the results, Raman bands of typical collagen appeared in the obtained spectra. Although the typical mineral bands at 960 cm-1 (Phosphate) was absent due to decalcified processing, we found that Raman peak intensities of amide I and C-C stretching bands were significantly different between OVX and sham-operated specimens. These differences on the Raman spectra were statistically compared by multivariate analyses, principal component analysis (PCA) and liner discrimination analysis (LDA). Our analyses suggest that amide I and C-C stretching bands can be related to stability of bone matrix which reflects bone quality.

  19. Castor oil polymer induces bone formation with high matrix metalloproteinase-2 expression.

    Science.gov (United States)

    Saran, Wallace Rocha; Chierice, Gilberto Orivaldo; da Silva, Raquel Assed Bezerra; de Queiroz, Alexandra Mussolino; Paula-Silva, Francisco Wanderley Garcia; da Silva, Léa Assed Bezerra

    2014-02-01

    The aim of this study was to evaluate the modulation of matrix metalloproteinase-2 (MMP-2) and -9 (MMP-9) expression in newly formed bone tissue at the interface between implants derived from castor oil (Ricinus communis) polymer and the tibia medullary canal. Forty-four rabbits were assigned to either Group 1 (n = 12; control) or Group 2 (n = 30), which had the tibial medullary canals reamed bilaterally and filled with polymer. CT scans showed no space between the material surface and the bone at the implant/bone marrow interface, and the density of the tissues at this interface was similar to the density measured of other regions of the bone. At 90 days postimplantation, the interface with the polymer presented a thick layer of newly formed bone tissue rich in osteocytes. This tissue exhibited ongoing maturation at 120 and 150 days postimplantation. Overall, bone remodeling process was accompanied by positive modulation of MMP-2 and low MMP-9 expression. Differently, in control group, the internal surface close to the medullary canal was lined by osteoblasts, followed by a bone tissue zone with few lacunae filled with osteocytes. Maturation of the tissue of the medullary internal surface occurred in the inner region, with the bone being nonlamellar.

  20. Chemical modification of extracellular matrix by cold atmospheric plasma-generated reactive species affects chondrogenesis and bone formation.

    Science.gov (United States)

    Eisenhauer, Peter; Chernets, Natalie; Song, You; Dobrynin, Danil; Pleshko, Nancy; Steinbeck, Marla J; Freeman, Theresa A

    2016-09-01

    The goal of this study was to investigate whether cold plasma generated by dielectric barrier discharge (DBD) modifies extracellular matrices (ECM) to influence chondrogenesis and endochondral ossification. Replacement of cartilage by bone during endochondral ossification is essential in fetal skeletal development, bone growth and fracture healing. Regulation of this process by the ECM occurs through matrix remodelling, involving a variety of cell attachment molecules and growth factors, which influence cell morphology and protein expression. The commercially available ECM, Matrigel, was treated with microsecond or nanosecond pulsed (μsp or nsp, respectively) DBD frequencies conditions at the equivalent frequencies (1 kHz) or power (~1 W). Recombinant human bone morphogenetic protein-2 was added and the mixture subcutaneously injected into mice to simulate ectopic endochondral ossification. Two weeks later, the masses were extracted and analysed by microcomputed tomography. A significant increase in bone formation was observed in Matrigel treated with μsp DBD compared with control, while a significant decrease in bone formation was observed for both nsp treatments. Histological and immunohistochemical analysis showed Matrigel treated with μsp plasma increased the number of invading cells, the amount of vascular endothelial growth factor and chondrogenesis while the opposite was true for Matrigel treated with nsp plasma. In support of the in vivo Matrigel study, 10 T1/2 cells cultured in vitro on μsp DBD-treated type I collagen showed increased expression of adhesion proteins and activation of survival pathways, which decreased with nsp plasma treatments. These results indicate DBD modification of ECM can influence cellular behaviours to accelerate or inhibit chondrogenesis and endochondral ossification. Copyright © 2016 John Wiley & Sons, Ltd. Copyright © 2016 John Wiley & Sons, Ltd.

  1. Cartilage oligomeric matrix protein enhances the vascularization of acellular nerves

    Directory of Open Access Journals (Sweden)

    Wei-ling Cui

    2016-01-01

    Full Text Available Vascularization of acellular nerves has been shown to contribute to nerve bridging. In this study, we used a 10-mm sciatic nerve defect model in rats to determine whether cartilage oligomeric matrix protein enhances the vascularization of injured acellular nerves. The rat nerve defects were treated with acellular nerve grafting (control group alone or acellular nerve grafting combined with intraperitoneal injection of cartilage oligomeric matrix protein (experimental group. As shown through two-dimensional imaging, the vessels began to invade into the acellular nerve graft from both anastomotic ends at day 7 post-operation, and gradually covered the entire graft at day 21. The vascular density, vascular area, and the velocity of revascularization in the experimental group were all higher than those in the control group. These results indicate that cartilage oligomeric matrix protein enhances the vascularization of acellular nerves.

  2. Cartilage oligomeric matrix protein enhances the vascularization of acellular nerves

    Institute of Scientific and Technical Information of China (English)

    Wei-ling Cui; Long-hai Qiu; Jia-yan Lian; Jia-chun Li; Jun Hu; Xiao-lin Liu

    2016-01-01

    Vascularization of acellular nerves has been shown to contribute to nerve bridging. In this study, we used a 10-mm sciatic nerve defect model in rats to determine whether cartilage oligomeric matrix protein enhances the vascularization of injured acellular nerves. The rat nerve defects were treated with acellular nerve grafting (control group) alone or acellular nerve grafting combined with intraperitoneal injection of cartilage oligomeric matrix protein (experimental group). As shown through two-dimensional imaging, the vessels began to invade into the acellular nerve graft from both anastomotic ends at day 7 post-operation, and gradually covered the entire graft at day 21. The vascular density, vascular area, and the velocity of revascularization in the experimental group were all higher than those in the control group. These results indicate that cartilage oligomeric matrix protein enhances the vascularization of acellular nerves.

  3. Different matrix evaluation for the bone regeneration of rats' femours using time domain optical coherence tomography

    Science.gov (United States)

    Rusu, Laura-Cristina; Negrutiu, Meda Lavinia; Sinescu, Cosmin; Hoinoiu, Bogdan; Zaharia, Cristian; Ardelean, Lavinia; Duma, Virgil-Florin; Podoleanu, Adrian G.

    2014-01-01

    The osteoconductive materials are important in bone regeneration procedures. Three dimensional (3D) reconstructions were obtained from the analysis. The aim of this study is to investigate the interface between the femur rat bone and the new bone that is obtained using a method of tissue engineering that is based on two artificial matrixes inserted in previously artificially induced defects. For this study, under strict supervision 20 rats were used in conformity with ethical procedures. In all the femurs a round defect was induced by drilling with a 1 mm spherical Co-Cr surgical drill. The matrixes used were IngeniOss (for ten samples) and 4Bone(for the other ten samples). These materials were inserted into the induced defects. The femurs were investigated at 1 month, after the surgical procedures. The interfaces were examined using Time Domain (TD) Optical Coherence Tomography (OCT) combined with Confocal Microscopy (CM). The scanning procedure is similar to that used in any CM, where the fast scanning is en-face (line rate) and the scanning in depth is much slower (at the frame rate). The optical configuration uses two single mode directional couplers with a superluminiscent diode as the source centered at 1300 nm. The results showed open interfaces due to the insufficient healing process, as well as closed interfaces due to a new bone formation inside the defect. The conclusion of this study is that TD-OCT can act as a valuable tool in the investigation of the interface between the old bone and the one that has been newly created due to the osteoinductive process. The TD-OCT has proven a valuable tool for the non-invasive evaluation of the matrix bone interfaces.

  4. Exercise alters mineral and matrix composition in the absence of adding new bone.

    Science.gov (United States)

    Kohn, David H; Sahar, Nadder D; Wallace, Joseph M; Golcuk, Kurtulus; Morris, Michael D

    2009-01-01

    The mechanical properties of bone are dictated by its amount, distribution and 'quality'. The composition of the mineral and matrix phases is integral to defining 'bone quality'. Exercise can potentially increase resistance to fracture, yet the effects of exercise on skeletal fragility, and how alterations in fragility are modulated by the amount, distribution and composition of bone, are unknown. In this investigation, the effects of exercise on the size, composition, mechanical properties and damage resistance of bones from mice of various ages, background strains and genetic makeup were assessed, as a means of testing the hypothesis that mechanical loading can improve skeletal fragility via compositional alterations. C57BL/6 mice (4-month-old males) ran on a treadmill for 21 days. Tibiae from exercised and control mice were analyzed for cross-sectional geometry, mechanical properties, microdamage and composition. Exercise significantly increased strength without increasing cross-sectional properties, suggesting that mechanical stimulation led to changes in the bone matrix, and these changes led to the improvements in mechanical properties. Consistent with this interpretation, the mineral/matrix ratio was significantly increased in exercised bones. The number of fatigue-induced microcracks was significantly lower in exercised bones, providing evidence that exercise modulates fatigue resistance. The ratio of nonreducible/reducible cross-links mirrored the damage data. Similar trends (exercise induced increases in mechanical properties without increases in cross-sectional properties, but with compositional changes) were also observed in 2-month-old biglycan-deficient and wild-type mice bred on a C57BL/6x129 genetic background. Copyright 2008 S. Karger AG, Basel.

  5. Identification of Proteins with Potential Osteogenic Activity Present in the Water-Soluble Matrix Proteins from Crassostrea gigas Nacre Using a Proteomic Approach

    Directory of Open Access Journals (Sweden)

    Daniel V. Oliveira

    2012-01-01

    Full Text Available Nacre, when implanted in vivo in bones of dogs, sheep, mice, and humans, induces a biological response that includes integration and osteogenic activity on the host tissue that seems to be activated by a set of proteins present in the nacre water-soluble matrix (WSM. We describe here an experimental approach that can accurately identify the proteins present in the WSM of shell mollusk nacre. Four proteins (three gigasin-2 isoforms and a cystatin A2 were for the first time identified in WSM of Crassostrea gigas nacre using 2DE and LC-MS/MS for protein identification. These proteins are thought to be involved in bone remodeling processes and could be responsible for the biocompatibility shown between bone and nacre grafts. These results represent a contribution to the study of shell biomineralization process and opens new perspectives for the development of new nacre biomaterials for orthopedic applications.

  6. [Mineral phase and protein matrix status of rat bony tissue after a flight on the Kosmos-1129 biosatellite].

    Science.gov (United States)

    Prokhonchukov, A A; Desiatnichenko, K S; Tigranian, R A; Komissarova, N A

    1982-01-01

    The major parameters of the mineral component and protein matrix of bones were investigated in 30 rats flown onboard Cosmos-1129. Postflight, the content of calcium decreased by 7.8%, that of phosphorus diminished by 11.8%, the Ca/P ratio increased by 5.9%, the content of collagen diminished by 14.7% and that of non-collagenous proteins by 45.7% and the content of sialic and hexuronic acids increased by 36.2% and 14.6%, respectively, as compared to the vivarium controls. The paper discusses the role of EDTA-and HCl-protein extracts, soluble and poorly soluble calcium fractions, protein-Ca-phosphate complex, sialic and hexuronic acids in the mechanism of calcium binding by the bone organic matrix.

  7. Squalane as a possible carrier of bone morphogenetic protein.

    Science.gov (United States)

    Kawakami, T; Uji, H; Antoh, M; Hasegawa, H; Kise, T; Eda, S

    1993-07-01

    Gelatin capsules containing squalane partially purified bone morphogenetic protein (BMP) complex were placed on the perimuscular membrane of rats. Two kinds of control, gelatin capsules containing only BMP and those bearing squalane only, were used. The embedded areas were histopathologically examined at 3 and 6 wk after the operation. The observations revealed that the squalane/BMP complex elicited wide heterotopic bone formation with bone marrow tissue, suggesting that squalane is a possible carrier of BMP for clinical applications.

  8. A Novel MALDI Matrix for Analyzing Peptides and Proteins: Paraffin Wax Immobilized Matrix

    Institute of Scientific and Technical Information of China (English)

    WEI Yuanlong; MEI Yuan; XU Zhe; WANG Cuihong; GUO Yinlong; DU Yiping; ZHANG Weibing

    2009-01-01

    A new kind of MALDI matrix, termed paraffin wax immobilized matrix, was used to study peptide mixtures and proteins. During the preparation process, the paraffin wax was heated and coated on the stainless-steel target plate, and then 2,5-dihydrobenzoic acid (DHB) was deposited on the paraffin layer and stainless-steel target plate to obtain different kinds of matrix spots. The morphology of matrices on different supports and peptide-matrix co-crystallization were observed by a high resolution digital-video microscopy system. Peptide mixtures and bovine serum albumin (BSA) digests were used to investigate the performance of the immobilized matrices on the paraffin target. The MALDI-FTMS analysis results also showed that the detection sensitivity of matrices immobilized in the paraffin sample support was better than that on other sample supports.

  9. Histologic evaluation of an allogeneic mineralized bone matrix in the treatment of periodontal osseous defects.

    Science.gov (United States)

    Koylass, Jana-Marie; Valderrama, Pilar; Mellonig, James T

    2012-08-01

    The aim of this study was to evaluate the potential of an allogeneic bone matrix to regenerate new bone, cementum, and periodontal ligament around a previously diseased root surface. Four patients with severe chronic periodontitis and teeth with hopeless periodontal or restorative prognoses participated in this study. One tooth with a severe intraosseous defect was selected per patient. At baseline, measurements of probing depth, gingival recession, and clinical attachment level were obtained. Following flap reflection, a root notch was placed at the apical extent of the calculus; the root was debrided, and the allogeneic bone graft material was placed into the defect. After a minimum of 6 months of healing, the teeth were removed en bloc and prepared for histologic examination. Two of four teeth demonstrated regeneration of new bone, cementum, and periodontal ligament. One tooth healed by new connective tissue attachment, and another by junctional epthelium.

  10. The history and histology of bone morphogenetic protein.

    Science.gov (United States)

    Murray, Samuel S; Brochmann Murray, Elsa J; Wang, Jeffrey C; Duarte, Maria Eugenia Leite

    2016-07-01

    Bone morphogenetic proteins are a group of structurally related proteins within the TGF-β superfamily of proteins with a diverse repertoire of functions in embryonic and adult organisms. As is apparent from the name, the members first characterized participate in bone growth, development, and remodeling. The "morphogenic" activity per se is defined as the induction of a recapitulation of endochondral bone formation by appropriate stem cells. The regenerative capacity of bone has been recognized since ancient times. The mechanism, applications, and conceptual basis of bone transplantation, bone implantation, ectopic bone formation, and exogenously induced bone formation have been studied by many investigators for more than a century. This review examines the efforts to characterize this activity in the European and American literature over approximately the last century. Because of the inherently complex nature of the process induced by these molecules (inflammation, stem cell proliferation, cartilage differentiation, replacement of cartilage with bone) it is important to evaluate previous investigations through a histological perspective. The cellular basis of the contemporary bioassay for BMP activity is illustrated and discussed from the histological point of view.

  11. Congenital Bone Fractures in Spinal Muscular Atrophy: Functional Role for SMN Protein in Bone Remodeling

    Science.gov (United States)

    Shanmugarajan, Srinivasan; Swoboda, Kathryn J.; Iannaccone, Susan T.; Ries, William L.; Maria, Bernard L.; Reddy, Sakamuri V.

    2009-01-01

    Spinal muscular atrophy is the second most common fatal childhood disorder. Core clinical features include muscle weakness caused by degenerating lower motor neurons and a high incidence of bone fractures and hypercalcemia. Fractures further compromise quality of life by progression of joint contractures or additional loss of motor function. Recent observations suggest that bone disease in spinal muscular atrophy may not be attributed entirely to lower motor neuron degeneration. The presence of the spinal muscular atrophy disease-determining survival motor neuron gene (SMN), SMN expression, and differential splicing in bone-resorbing osteoclasts was recently discovered. Its ubiquitous expression and the differential expression of splice variants suggest that SMN has specific roles in bone cell function. SMN protein also interacts with osteoclast stimulatory factor. Mouse models of human spinal muscular atrophy disease suggest a potential role of SMN protein in skeletal development. Dual energy x-ray absorptiometry analysis demonstrated a substantial decrease in total bone area and poorly developed caudal vertebra in the mouse model. These mice also had pelvic bone fractures. Studies delineating SMN signaling mechanisms and gene transcription in a cell-specific manner will provide important molecular insights into the pathogenesis of bone disease in children with spinal muscular atrophy. Moreover, understanding bone remodeling in spinal muscular atrophy may lead to novel therapeutic approaches to enhance skeletal health and quality of life. This article reviews the skeletal complications associated with spinal muscular atrophy and describes a functional role for SMN protein in osteoclast development and bone resorption activity. PMID:17761651

  12. The Structure and Function of Non-Collagenous Bone Proteins

    Science.gov (United States)

    Hook, Magnus; McQuillan, David J.

    1997-01-01

    The research done under the cooperative research agreement for the project titled 'The structure and function of non-collagenous bone proteins' represented the first phase of an ongoing program to define the structural and functional relationships of the principal noncollagenous proteins in bone. An ultimate goal of this research is to enable design and execution of useful pharmacological compounds that will have a beneficial effect in treatment of osteoporosis, both land-based and induced by long-duration space travel. The goals of the now complete first phase were as follows: 1. Establish and/or develop powerful recombinant protein expression systems; 2. Develop and refine isolation and purification of recombinant proteins; 3. Express wild-type non-collagenous bone proteins; 4. Express site-specific mutant proteins and domains of wild-type proteins to enhance likelihood of crystal formation for subsequent solution of structure.

  13. Histological and radiographic evaluations of demineralized bone matrix and coralline hydroxyapatite in the rabbit tibia.

    Science.gov (United States)

    Zhukauskas, Rasa; Dodds, Robert A; Hartill, Caroline; Arola, Travis; Cobb, Ronald R; Fox, Casey

    2010-03-01

    Complex fractures resulting in bone loss or impaired fracture healing remain problematic in trauma and orthopedic surgeries. Many bone graft substitutes have been developed and are commercially available. These products differ in their osteoconductive and osteoinductive properties. Differential enhancement of these properties may optimize the performance of these products for various orthopedic and craniofacial applications. The use of bone graft substitutes offers the ability to lessen the possible morbidity of the harvest site in autografts. The objective of the present study was to compare the ability of two bone graft substitutes, BioSet RT, an allograft demineralized bone matrix formulation, and ProOsteon 500R, a coralline hydroxyapatite, in a rabbit critical tibial defect model. BioSet RT and ProOsteon 500R were implanted into a unicortical proximal metaphyseal tibial defect and evaluated for new bone formation. Samples were analyzed radiographically and histologically at 1 day, 6 weeks, 12 weeks, and 24 weeks post surgery. Both materials were biocompatible and demonstrated significant bone growth and remodeling. At 12 weeks, the BioSet RT implanted sites demonstrated significantly more defect closure and bone remodeling as determined by radiographic analyses with 10 out of 14 defects being completely healed versus 1 out of 14 being completely healed in the ProOsteon 500R implanted sites. At 24 weeks, both materials demonstrated complete closure of the defect as determined histologically. There were no statistical differences in radiographic scores between the two implanted materials. However, there was an observable trend that the BioSet RT material generated higher histological and radiographic scores, although not statistically significant. This study provides evidence that both BioSet RT and ProOsteon 500R are biocompatible and able to induce new bone formation as measured in this rabbit model. In addition, this in vivo study demonstrates the ability of

  14. Bone Morphogenetic Protein-2 Nonviral Gene Therapy in a Goat Iliac Crest Model for Bone Formation

    NARCIS (Netherlands)

    Loozen, Loek D.; van der Helm, Yvonne J. M.; Oner, F. Cumhur; Dhert, W.J.A.; Kruyt, Moyo C.; Alblas, Jacqueline

    2015-01-01

    Treatment and reconstruction of large bone defects, delayed unions, and nonunions is challenging and has resulted in an ongoing search for novel tissue-engineered therapies. Bone morphogenetic protein-2 (BMP-2) gene therapy is a promising strategy to provide sustained production of BMP-2 locally. Al

  15. The effect of enamel matrix derivative (Emdogain) on bone formation: a systematic review.

    Science.gov (United States)

    Rathe, Florian; Junker, Rüdiger; Chesnutt, Betsy M; Jansen, John A

    2009-09-01

    This systematic review focused on the question, if and to what extent enamel matrix derivative (Emdogain) [EMD]) promotes the regeneration of bone. The influence of combinations with other biomaterials was additionally evaluated. Twenty histomorphometric studies were included in this systematic review. Main results of the reviewed articles were (i) guide tissue regeneration (GTR) of infrabony defects seems to result in a higher degree of bone regeneration compared to treatment with EMD; (ii) combined therapy (GTR + EMD) of infrabony defects might not lead to better results than GTR therapy alone; (iii) there seems to be no additional benefit of combined therapy (GTR + EMD) in furcation defects over GTR therapy alone; (iv) EMD seems to lead to more bone regeneration of infrabony defects compared to open flap debridement; (v) however, EMD application might result in more bone formation when applied in supporting defects compared to nonsupporting defects; and (vi) EMD does not seem to promote external jaw/parietal bone formation in the titanium capsule model. The results of one study that suggest that EMD increases the initial growth of trabecular bone around endosseous implants by new bone induction need to be confirmed by additional research.

  16. Erythropoietin modulates the structure of bone morphogenetic protein 2-engineered cranial bone.

    Science.gov (United States)

    Sun, Hongli; Jung, Younghun; Shiozawa, Yusuke; Taichman, Russell S; Krebsbach, Paul H

    2012-10-01

    The ideally engineered bone should have similar structural and functional properties to the native tissue. Although structural integrity is critical for functional bone regeneration, we know less about modulating the structural properties of the engineered bone elicited by bone morphogenetic protein (BMP) than efficacy and safety. Erythropoietin (Epo), a primary erythropoietic hormone, has been used to augment blood transfusion in orthopedic surgery. However, the effects of Epo on bone regeneration are not well known. Here, we determined the role of Epo in BMP2-induced bone regeneration using a cranial defect model. Epo administration improved the quality of BMP2-induced bone and more closely resembled natural cranial bone with a higher bone volume (BV) fraction and lower marrow fraction when compared with BMP2 treatment alone. Epo increased red blood cells (RBCs) in peripheral blood and also increased hematopoietic and mesenchymal stem cell (MSC) populations in bone marrow. Consistent with our previous work, Epo increased osteoclastogenesis both in vitro and in vivo. Results from a metatarsal organ culture assay suggested that Epo-promoted osteoclastogenesis contributed to angiogenesis because angiogenesis was blunted when osteoclastogenesis was blocked by alendronate (ALN) or osteoprotegerin (OPG). Earlier calcification of BMP2-induced temporary chondroid tissue was observed in the Epo+BMP group compared to BMP2 alone. We conclude that Epo significantly enhanced the outcomes of BMP2-induced cranial bone regeneration in part through its actions on osteoclastogenesis and angiogenesis.

  17. Tendon Reattachment to Bone in an Ovine Tendon Defect Model of Retraction Using Allogenic and Xenogenic Demineralised Bone Matrix Incorporated with Mesenchymal Stem Cells

    Science.gov (United States)

    2016-01-01

    Background Tendon-bone healing following rotator cuff repairs is mainly impaired by poor tissue quality. Demineralised bone matrix promotes healing of the tendon-bone interface but its role in the treatment of tendon tears with retraction has not been investigated. We hypothesized that cortical demineralised bone matrix used with minimally manipulated mesenchymal stem cells will result in improved function and restoration of the tendon-bone interface with no difference between xenogenic and allogenic scaffolds. Materials and Methods In an ovine model, the patellar tendon was detached from the tibial tuberosity and a complete distal tendon transverse defect measuring 1 cm was created. Suture anchors were used to reattach the tendon and xenogenic demineralised bone matrix + minimally manipulated mesenchymal stem cells (n = 5), or allogenic demineralised bone matrix + minimally manipulated mesenchymal stem cells (n = 5) were used to bridge the defect. Graft incorporation into the tendon and its effect on regeneration of the enthesis was assessed using histomorphometry. Force plate analysis was used to assess functional recovery. Results Compared to the xenograft, the allograft was associated with significantly higher functional weight bearing at 6 (P = 0.047), 9 (P = 0.028), and 12 weeks (P = 0.009). In the allogenic group this was accompanied by greater remodeling of the demineralised bone matrix into tendon-like tissue in the region of the defect (p = 0.015), and a more direct type of enthesis characterized by significantly more fibrocartilage (p = 0.039). No failures of tendon-bone healing were noted in either group. Conclusion Demineralised bone matrix used with minimally manipulated mesenchymal stem cells promotes healing of the tendon-bone interface in an ovine model of acute tendon retraction, with superior mechanical and histological results associated with use of an allograft. PMID:27606597

  18. Demineralized Bone Matrix (DBM) as a Bone Void Filler in Lumbar Interbody Fusion: A Prospective Pilot Study of Simultaneous DBM and Autologous Bone Grafts

    Science.gov (United States)

    Kim, Bum-Joon; Kim, Se-Hoon; Lee, Haebin; Lee, Seung-Hwan; Kim, Won-Hyung; Jin, Sung-Won

    2017-01-01

    Objective Solid bone fusion is an essential process in spinal stabilization surgery. Recently, as several minimally invasive spinal surgeries have developed, a need of artificial bone substitutes such as demineralized bone matrix (DBM), has arisen. We investigated the in vivo bone growth rate of DBM as a bone void filler compared to a local autologous bone grafts. Methods From April 2014 to August 2015, 20 patients with a one or two-level spinal stenosis were included. A posterior lumbar interbody fusion using two cages and pedicle screw fixation was performed for every patient, and each cage was packed with autologous local bone and DBM. Clinical outcomes were assessed using the Numeric Rating Scale (NRS) of leg pain and back pain and the Korean Oswestry Disability Index (K-ODI). Clinical outcome parameters and range of motion (ROM) of the operated level were collected preoperatively and at 3 months, 6 months, and 1 year postoperatively. Computed tomography was performed 1 year after fusion surgery and bone growth of the autologous bone grafts and DBM were analyzed by ImageJ software. Results Eighteen patients completed 1 year of follow-up, including 10 men and 8 women, and the mean age was 56.4 (32–71). The operated level ranged from L3/4 to L5/S1. Eleven patients had single level and 7 patients had two-level repairs. The mean back pain NRS improved from 4.61 to 2.78 (p=0.003) and the leg pain NRS improved from 6.89 to 2.39 (posteoporosis. PMID:28264244

  19. Novel silk protein barrier membranes for guided bone regeneration.

    Science.gov (United States)

    Smeets, Ralf; Knabe, Christine; Kolk, Andreas; Rheinnecker, Michael; Gröbe, Alexander; Heiland, Max; Zehbe, Rolf; Sachse, Manuela; Große-Siestrup, Christian; Wöltje, Michael; Hanken, Henning

    2016-10-12

    This study assesses the biocompatibility of novel silk protein membranes with and without modification, and evaluates their effect on facilitating bone formation and defect repair in guided bone regeneration. Two calvarian bone defects 12 mm in diameter were created in each of a total of 38 rabbits. Four different types of membranes, (silk-, hydroxyapatite-modified silk-, β-TCP-modified silk- and commonly clinically used collagen-membranes) were implanted to cover one of the two defects in each animal. Histologic analysis did not show any adverse tissue reactions in any of the defect sites indicating good biocompatibility of all silk protein membranes. Histomorphometric and histologic evaluation revealed that collagen and β-TCP modified silk membranes supported bone formation (collagen: bone area fraction p = 0.025; significant; β-TCP modified silk membranes bone area fraction: p = 0.24, not significant), guided bone regeneration and defect bridging. The bone, which had formed in defects covered by β-TCP modified silk membranes, displayed a more advanced stage of bone tissue maturation with restoration of the original calvarial bone microarchitecture when compared to the bone which had formed in defects, for which any of the other test membranes were used. Micro-CT analysis did not reveal any differences in the amount of bone formation between defects with and without membranes. In contrast to the collagen membranes, β-TCP modified silk membranes were visible in all cases and may therefore be advantageous for further supporting bone formation beyond 10 weeks and preventing soft tissue ingrowth from the periphery. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 2016.

  20. Mouse Basophils Reside in Extracellular Matrix-Enriched Bone Marrow Niches Which Control Their Motility

    OpenAIRE

    Salete Smaniotto; Elke Schneider; Nicolas Goudin; Rachel Bricard-Rignault; François Machavoine; Mireille Dardenne; Michel Dy; Wilson Savino

    2013-01-01

    Basophils co-express FcεRIα and CD49b, the α-2 chain of integrin-type receptor VLA-2 (α2β1), which recognizes type-1 collagen as a major natural ligand. The physiological relevance of this integrin for interactions with extracellular bone marrow matrix remains unknown. Herein, we examined the expression of several receptors of this family by bone marrow-derived basophils sorted either ex-vivo or after culture with IL-3. Having established that both populations display CD49d, CD49e and CD49f (...

  1. Dietary protein: an essential nutrient for bone health.

    Science.gov (United States)

    Bonjour, Jean-Philippe

    2005-12-01

    Nutrition plays a major role in the development and maintenance of bone structures resistant to usual mechanical loadings. In addition to calcium in the presence of an adequate vitamin D supply, proteins represent a key nutrient for bone health, and thereby in the prevention of osteoporosis. In sharp opposition to experimental and clinical evidence, it has been alleged that proteins, particularly those from animal sources, might be deleterious for bone health by inducing chronic metabolic acidosis which in turn would be responsible for increased calciuria and accelerated mineral dissolution. This claim is based on an hypothesis that artificially assembles various notions, including in vitro observations on the physical-chemical property of apatite crystal, short term human studies on the calciuric response to increased protein intakes, as well as retrospective inter-ethnic comparisons on the prevalence of hip fractures. The main purpose of this review is to analyze the evidence that refutes a relation of causality between the elements of this putative patho-physiological "cascade" that purports that animal proteins are causally associated with an increased incidence of osteoporotic fractures. In contrast, many experimental and clinical published data concur to indicate that low protein intake negatively affects bone health. Thus, selective deficiency in dietary proteins causes marked deterioration in bone mass, micro architecture and strength, the hallmark of osteoporosis. In the elderly, low protein intakes are often observed in patients with hip fracture. In these patients intervention study after orthopedic management demonstrates that protein supplementation as given in the form of casein, attenuates post-fracture bone loss, increases muscles strength, reduces medical complications and hospital stay. In agreement with both experimental and clinical intervention studies, large prospective epidemiologic observations indicate that relatively high protein intakes

  2. Matrix Gla Protein polymorphisms are associated with coronary artery calcification

    Science.gov (United States)

    Matrix Gla Protein (MGP) is a key regulator of vascular calcification. Genetic variation at the MGP locus could modulate the development of coronary artery calcification (CAC). We examined the cross-sectional association between MGP SNPs [rs1800802 (T-138C), rs1800801 (G-7A),and rs4236 (Ala102Thr)...

  3. Cartilage oligomeric matrix protein specific antibodies are pathogenic

    DEFF Research Database (Denmark)

    Geng, Hui; Nandakumar, Kutty Selva; Pramhed, Anna

    2012-01-01

    ABSTRACT: INTRODUCTION: Cartilage oligomeric matrix protein (COMP) is a major non-collagenous component of cartilage. Earlier, we developed a new mouse model for rheumatoid arthritis using COMP. This study was undertaken to investigate the epitope specificity and immunopathogenicity of COMP...

  4. Mixed-matrix membrane adsorbers for protein separation

    NARCIS (Netherlands)

    Avramescu, Maria-Elena; Borneman, Zandrie; Wessling, Matthias

    2003-01-01

    The separation of two similarly sized proteins, bovine serum albumin (BSA) and bovine hemoglobin (Hb) was carried out using a new type of ion-exchange mixed-matrix adsorber membranes. The adsorber membranes were prepared by incorporation of various types of Lewatit ion-exchange resins into an ethyle

  5. The application of an enamel matrix protein derivative (Emdogain) in regenerative periodontal therapy: a review.

    Science.gov (United States)

    Sculean, Anton; Schwarz, Frank; Becker, Jurgen; Brecx, Michel

    2007-01-01

    Regenerative periodontal therapy aims at reconstitution of the lost periodontal structures such as new formation of root cementum, periodontal ligament and alveolar bone. Findings from basic research indicate that enamel matrix protein derivative (EMD) has a key role in periodontal wound healing. Histological results from animal and human studies have shown that treatment with EMD promotes periodontal regeneration. Moreover, clinical studies have indicated that treatment with EMD positively influences periodontal wound healing in humans. This review aims to present an overview of evidence-based clinical indications for regenerative therapy with EMD.

  6. Gallium nitrate increases type I collagen and fibronectin mRNA and collagen protein levels in bone and fibroblast cells.

    Science.gov (United States)

    Bockman, R S; Guidon, P T; Pan, L C; Salvatori, R; Kawaguchi, A

    1993-08-01

    Gallium is a Group IIIa transitional element with therapeutic efficacy in the treatment of metabolic bone disorders. Previously described antiresorptive effects of gallium on osteoclasts are not sufficient to account for the full range of effects of gallium on bone structure and metabolism. We have recently shown that gallium nitrate inhibits osteocalcin gene expression and the synthesis of osteocalcin protein, an osteoblast-specific bone matrix protein that is thought to serve as a signal to trigger osteoclastic resorption. Here we present evidence for an additional mechanism by which gallium may function to augment bone mass by altering matrix protein synthesis by osteoblastic and fibroblastic cells. Rat calvarial explants exposed to gallium nitrate for 48 h showed increased incorporation of 3H-proline into hydroxyproline and collagenase digestible protein. In addition, gallium treatment increased steady-state mRNA levels for fibronectin and type I procollagen chains in primary rat calvarial osteoblast-enriched cultures, the ROS 17/2.8 osteoblastic osteosarcoma line, and nontransformed human dermal fibroblasts. These findings suggest that the exposure of mesenchymally-derived cells to gallium results in an altered pattern of matrix protein synthesis that would favor increased bone formation.

  7. Effect of enamel matrix derivative (Emdogain) on bone defects in rabbit tibias.

    Science.gov (United States)

    Cornelini, Roberto; Scarano, Antonio; Piattelli, Maurizio; Andreana, Sebastiano; Covani, Ugo; Quaranta, Alessandro; Piattelli, Adriano

    2004-01-01

    The aim of this study was to assess the effect of an enamel matrix derivative (Emdogain, Biora, AB, Malmö, Sweden) on bone healing. Ten New Zealand rabbits, weighing about 2.5 kg, were used. One 8-mm bone defect was created in each tibia. The defect on the right leg was filled with Emdogain, whereas the defect on the opposite leg was left unfilled as control. A total of 20 defects were created. Five rabbits each were killed at 4 and 8 weeks with an overdose of Tanax. Block sections containing the defects were retrieved and the specimens processed for light microscopy examination. The slides were stained with acid and basic fuchsin and toluidine blue. Histologically, no differences were noted in both groups at each observation period; in the test group, remnants of the implanted Emdogain were not present at 4 weeks. Newly formed bone was detectable in both groups at all observation times. At 8 weeks, both groups showed mature bone, and in the test group the material implanted was not visible. No inflammatory cells were visible in both groups. In conclusion, our results indicate that Emdogain implanted in bone defects is fully resorbed after 4 to 8 weeks and does not adversely affect bone formation.

  8. Development of biomimetic nanocomposites as bone extracellular matrix for human osteoblastic cells.

    Science.gov (United States)

    Bhowmick, Arundhati; Mitra, Tapas; Gnanamani, Arumugam; Das, Manas; Kundu, Patit Paban

    2016-05-05

    Here, we have developed biomimetic nanocomposites containing chitosan, poly(vinyl alcohol) and nano-hydroxyapatite-zinc oxide as bone extracellular matrix for human osteoblastic cells and characterized by Fourier transform infrared spectroscopy, powder X-ray diffraction. Scanning electron microscopy images revealed interconnected macroporous structures. Moreover, in this study, the problem related to fabricating a porous composite with good mechanical strength has been resolved by incorporating 5wt% of nano-hydroxyapatite-zinc oxide into chitosan-poly(vinyl alcohol) matrix; the present composite showed high tensile strength (20.25MPa) while maintaining appreciable porosity (65.25%). These values are similar to human cancellous bone. These nanocomposites also showed superior water uptake, antimicrobial and biodegradable properties than the previously reported results. Compatibility with human blood and pH was observed, indicating nontoxicity of these materials to the human body. Moreover, proliferation of osteoblastic MG-63 cells onto the nanocomposites was also observed without having any negative effect.

  9. Novel identification of matrix proteins involved in calcitic biomineralization.

    Science.gov (United States)

    Rose-Martel, Megan; Smiley, Sandy; Hincke, Maxwell T

    2015-02-26

    Calcitic biomineralization is essential for otoconia formation in vertebrates. This process is characterized by protein-crystal interactions that modulate crystal growth on an extracellular matrix. An excellent model for the study of calcitic biomineralization is the avian eggshell, the fastest known biomineralization process. The objective of this study is to identify and characterize matrix proteins associated with the eggshell mammillary cones, which are hypothesized to regulate the earliest stage of eggshell calcification. Mammillary cones were isolated from 2 models, fertilized and unfertilized, and the released proteins were identified by RP-nanoLC and ES-MS/MS proteomics. Proteomics analysis identified 49 proteins associated with the eggshell membrane fibers and, importantly, 18 mammillary cone-specific proteins with an additional 18 proteins identified as enriched in the mammillary cones. Among the most promising candidates for modulating protein-crystal interactions were extracellular matrix proteins, including ABI family member 3 (NESH) binding protein (ABI3BP), tiarin-like, hyaluronan and proteoglycan link protein 3 (HAPLN3), collagen alpha-1(X), collagen alpha-1(II) and fibronectin, in addition to the calcium binding proteins calumenin, EGF-like repeats and discoidin 1-like domains 3 (EDIL3), nucleobindin-2 and SPARC. In conclusion, we identified several cone-resident proteins that are candidates to regulate initiation of eggshell calcification. Further study of these proteins will determine their roles in modulating calcitic biomineralization and lead to insight into the process of otoconia formation/regeneration. Biomineralization is essential for the development of hard tissues in vertebrates, which includes both calcium phosphate and calcium carbonate structures. Calcitic mineralization by calcium carbonate is an important process in the formation of otoconia, which are gravity receptor organs located in the inner ear and are responsible for balance

  10. Effects of matrix metalloproteinase-1 on the myogenic differentiation of bone marrow-derived mesenchymal stem cells in vitro

    Energy Technology Data Exchange (ETDEWEB)

    Zheng, Zhenyang [Department of Neurology, The First Affiliated Hospital, Sun Yat-sen University, No. 58, Zhongshan 2nd Road, Guangzhou 510080, Guangdong Province (China); Leng, Yan [Department of Rehabilitation, The First Affiliated Hospital, Sun Yat-sen University, No. 58, Zhongshan 2nd Road, Guangzhou 510080, Guangdong Province (China); Zhou, Chen; Ma, Zhenyu; Zhong, Zhigang [Department of Neurology, The First Affiliated Hospital, Sun Yat-sen University, No. 58, Zhongshan 2nd Road, Guangzhou 510080, Guangdong Province (China); Shi, Xing-Ming [Institute of Molecular Medicine and Genetics, Georgia Health Sciences University, Augusta, GA 30912 (United States); Zhang, Weixi, E-mail: weixizhang@qq.com [Department of Neurology, The First Affiliated Hospital, Sun Yat-sen University, No. 58, Zhongshan 2nd Road, Guangzhou 510080, Guangdong Province (China)

    2012-11-16

    Highlights: Black-Right-Pointing-Pointer MMP-1 is a member of the zinc-dependent endopeptidase family. Black-Right-Pointing-Pointer MMP-1 has no cytotoxic effects on BMSCs. Black-Right-Pointing-Pointer MMP-1 can promote the myogenic differentiation of BMSCs. Black-Right-Pointing-Pointer MyoD and desmin were chosen as myogenic markers in this study. -- Abstract: Matrix metalloproteinase-1 (MMP-1) is a member of the family of zinc-dependent endopeptidases that are capable of degrading extracellular matrix (ECM) and certain non-matrix proteins. It has been shown that MMP-1 can enhance muscle regeneration by improving the differentiation and migration of myoblasts. However, it is still not known whether MMP-1 can promote the myogenesis of bone marrow-derived mesenchymal stem cells (BMSCs). To address this question, we isolated BMSCs from C57BL/6J mice and investigated the effects of MMP-1 on their proliferation and myogenic differentiation. Our results showed that MMP-1 treatment, which had no cytotoxic effects on BMSCs, increased the mRNA and protein levels of MyoD and desmin in a dose-dependent manner, indicating that MMP-1 promoted myogenic differentiation of BMSCs in vitro. These results suggest that BMSCs may have a therapeutic potential for treating muscular disorders.

  11. Demineralized bone matrix and human cancellous bone enhance fixation of titanium implants

    DEFF Research Database (Denmark)

    Babiker, Hassan; Ding, Ming; Overgaard, Søren

    Denmark, DenmarkAbstractReplacement of extensive local bone loss especially in revision joint arthroplasty and spine fusion is a significant clinical challenge. Allograft and autograft have been considered as gold standard for bone replacement. However, there are several disadvantages such as donor site...... from human tissue were included (IsoTis OrthoBiologics, Inc. USA). Both materials are commercially available. Titanium alloy implants (Biomet Inc.) of 10 mm in length and 10 mm in diameter were inserted bilaterally into the femoral condyles of 8 skeletally mature sheep. Thus four implants...... with a concentric gap of 2 mm were implanted in each sheep. The gap was filled with: DBM; DBM/CB with ratio of 1/3; DBM/allograft with ratio of 1/3; or allograft (Gold standard), respectively. Standardised surgical procedure was used1. At sacrifice, 6 weeks after surgery, both distal femurs were harvested...

  12. Effect of bone graft density on in vitro cell behavior with enamel matrix derivative.

    Science.gov (United States)

    Miron, Richard J; Caluseru, Oana M; Guillemette, Vincent; Zhang, Yufeng; Buser, Daniel; Chandad, Fatiha; Sculean, Anton

    2015-09-01

    Bone replacement grafting materials play an important role in regenerative dentistry. Despite a large array of tested bone-grafting materials, little information is available comparing the effects of bone graft density on in vitro cell behavior. Therefore, the aim of the present study is to compare the effects of cells seeded on bone grafts at low and high density in vitro for osteoblast adhesion, proliferation, and differentiation. The response of osteoblasts to the presence of a growth factor (enamel matrix derivative, (EMD)) in combination with low (8 mg per well) or high (100 mg per well) bone grafts (BG; natural bone mineral, Bio-Oss®) density, was studied and compared for osteoblast cell adhesion, proliferation, and differentiation as assessed by real-time PCR. Standard tissue culture plastic was used as a control with and without EMD. The present study demonstrates that in vitro testing of bone-grafting materials is largely influenced by bone graft seeding density. Osteoblast adhesion was up to 50 % lower when cells were seeded on high-density BG when compared to low-density BG and control tissue culture plastic. Furthermore, proliferation was affected in a similar manner whereby cell proliferation on high-density BG (100 mg/well) was significantly increased when compared to that on low-density BG (8 mg/well). In contrast, cell differentiation was significantly increased on high-density BG as assessed by real-time PCR for markers collagen 1 (Col 1), alkaline phosphatase (ALP), and osteocalcin (OC) as well as alizarin red staining. The effects of EMD on osteoblast adhesion, proliferation, and differentiation further demonstrated that the bone graft seeding density largely controls in vitro results. EMD significantly increased cell attachment only on high-density BG, whereas EMD was able to further stimulate cell proliferation and differentiation of osteoblasts on control culture plastic and low-density BG when compared to high-density BG. The results

  13. Fibular Allograft and Demineralized Bone Matrix for the Treatment of Slipped Capital Femoral Epiphysis.

    Science.gov (United States)

    Murray, Travis; Morscher, Melanie A; Krahe, Amy M; Adamczyk, Mark J; Weiner, Dennis S

    2016-05-01

    Previous studies documented the use of fibular allograft in the treatment of slipped capital femoral epiphysis (SCFE) with bone graft epiphysiodesis (BGE). This study describes the results of using a 10-mm diameter premilled fibular allograft packed with demineralized bone matrix placed across the physis in an open surgical approach under image intensification. A review identified 45 cases of BGE using fibular allograft and demineralized bone matrix in 34 patients with a diagnosis of SCFE performed by a single surgeon during an 8-year period. Thirty-four cases (25 patients) had at least 1 year of follow-up and were included in the study. Medical records were reviewed for complications, subsequent surgeries, and time to physeal closure. Of the 34 cases included, there were no cases of acute chondrolysis. Complications included 1 case of bone graft extrusion that required surgical replacement and 1 re-slip requiring surgical stabilization. Five cases of avascular necrosis (AVN) were encountered (1 unstable slip with total head AVN, and 4 stable slips with 3 total head and 1 partial head AVN). In 1 patient, small loose bony fragments were noted on postoperative radiographs that appeared outside of the articular surface of the hip and were asymptomatic. Two patients encountered wound healing issues that resolved with appropriate wound care. In light of the occurrence of AVN in stable cases, BGE with autogenous corticocancellous graft is preferable to BGE with autologous fibular graft for the treatment of SCFE. [Orthopedics. 2016; 39(3):e519-e525.].

  14. Effect of HIP/ribosomal protein L29 deficiency on mineral properties of murine bones and teeth.

    Science.gov (United States)

    Sloofman, Laura G; Verdelis, Kostas; Spevak, Lyudmila; Zayzafoon, Majd; Yamauchi, Mistuo; Opdenaker, Lynn M; Farach-Carson, Mary C; Boskey, Adele L; Kirn-Safran, Catherine B

    2010-07-01

    Mice lacking HIP/RPL29, a component of the ribosomal machinery, display increased bone fragility. To understand the effect of sub-efficient protein synthetic rates on mineralized tissue quality, we performed dynamic and static histomorphometry and examined the mineral properties of both bones and teeth in HIP/RPL29 knock-out mice using Fourier transform infrared imaging (FTIRI). While loss of HIP/RPL29 consistently reduced total bone size, decreased mineral apposition rates were not significant, indicating that short stature is not primarily due to impaired osteoblast function. Interestingly, our microspectroscopic studies showed that a significant decrease in collagen crosslinking during maturation of HIP/RPL29-null bone precedes an overall enhancement in the relative extent of mineralization of both trabecular and cortical adult bones. This report provides strong genetic evidence that ribosomal insufficiency induces subtle organic matrix deficiencies which elevates calcification. Consistent with the HIP/RPL29-null bone phenotype, HIP/RPL29-deficient teeth also showed reduced geometric properties accompanied with relative increased mineral densities of both dentin and enamel. Increased mineralization associated with enhanced tissue fragility related to imperfection in organic phase microstructure evokes defects seen in matrix protein-related bone and tooth diseases. Thus, HIP/RPL29 mice constitute a new genetic model for studying the contribution of global protein synthesis in the establishment of organic and inorganic phases in mineral tissues.

  15. Osteoinductivity assay of the variability of repeated extractions of bone morphogenetic proteins from bovine bone at different times

    Institute of Scientific and Technical Information of China (English)

    HU Zhen-ming 胡侦明; Sean AF Peel; Cameron ML Clokie

    2004-01-01

    Objective:To observe the activity of repeated extracts of bone matrix and the production of purified bone morphogenetic proteins (BMPs).Methods: BMPs were extracted 1- 4 times from fresh bovine cortical bone by the modified Urist's method, with each collected precipitate separated and lyophilized as partially purified BMPs. Another fresh bovine bone was extracted three times and the precipitates were mixed and lyophilized. Meanwhile, the alkaline phosphatase (ALP)activity was measured by an in vitro assay employing cultured C2C12 mouse myoblast cells through the osteoinductivity of bovine BMPs extracted four times at days 1, 4, 7, and 14, and the correlation between BMPs quantities and costing during extraction processes was analyzed.Results:The purified and the cost showed a positive correlation(r=0.969).To separate and lyophilize each collected precipitate as partially purified BMPs raised the cost,and mixed precipitates also cost much.ALPactivities of 1st and mixed extractions of BMPs were shown to be highly osteoinductive and keep a significantly high level(P<0.05-0.01)4 days after culturing compared with the 2nd,3rd and 4th extractions,especially the control group.However,the more times the extraction ws done,the less activity of BMPs was shown and more costing was.The x-ray and histological analysis also showed that the 1st extraction of BMPs induced more ossicles and new bone formation.Conclusions:The results indicated that BMPs enhanced the abilities of osteoinductiviyt in C2C12 culture in vitro.The first extraction of BMPsfrom bone is fitfull,4th extractions are unnecessary for they cost more and waste more time,say nothing of mixed extractions.

  16. The Effect of Bioceramic Composite Extracellular Matrixes Used to Repair Bone Deficiency on Relevant Blood Biochemical Indices

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    At the base of experimental animal model construction of bone defect in New Zealand rabbit, the promoting repair effect of bioactive ceramics on bone defect as well as its machanism was studied through testing body mineral elements, enzymes related to bone morphogenetic proteins and some biochemical indexes. Refering to some documents, materials of TCP, CHA and HA were combined and TCP/BMP/ TCP-β1 and CHA/BMP/ TCP-β1, HA/BMP/ TCP-β1 composite materials were made. All kinds of them were implanted into the radial defect site of rabbit, respectively. The chosen blood indexes (Ca, P, ALP, GGT, AST, ALT, TPA, BUN and Cr) were tested by colorimetry, speed rate and bromocresol green testing methods. No abnormal effects were found in any animal after operation. Serum concentrations of Ca, P and ALP were increased with the length of time in all groups of the three kinds of composite material, mixed material and pure materials. The increases in composite material groups were more significant ( P <0.05). Comparison of the three kinds of material showed TCP > CHA > HA. There was a tendency of increased TPA and decreased BUN with the length of time. There was no significant difference between the composite material groups and pure material group (P >0.05). The three kinds of bioactive ceramics composed of extracellular matrix could increase the serum concentrations of Ca and P and activity of ALP after being implanted into defect bone and showed some repairing capacity. This provided a new area of machanism study of bone defect repair by biomaterials.

  17. Evolution of the vertebrate bone matrix: an expression analysis of the network forming collagen paralogues in amphibian osteoblasts.

    Science.gov (United States)

    Aldea, Daniel; Hanna, Patricia; Munoz, David; Espinoza, Javier; Torrejon, Marcela; Sachs, Laurent; Buisine, Nicolas; Oulion, Silvan; Escriva, Hector; Marcellini, Sylvain

    2013-09-01

    The emergence of vertebrates is closely associated to the evolution of mineralized bone tissue. However, the molecular basis underlying the origin and subsequent diversification of the skeletal mineralized matrix is still poorly understood. One efficient way to tackle this issue is to compare the expression, between vertebrate species, of osteoblastic genes coding for bone matrix proteins. In this work, we have focused on the evolution of the network forming collagen family which contains the Col8a1, Col8a2, and Col10a1 genes. Both phylogeny and synteny reveal that these three paralogues are vertebrate-specific and derive from two independent duplications in the vertebrate lineage. To shed light on the evolution of this family, we have analyzed the osteoblastic expression of the network forming collagens in endochondral and intramembraneous skeletal elements of the amphibian Xenopus tropicalis. Remarkably, we find that amphibian osteoblasts express Col10a1, a gene strongly expressed in osteoblasts in actinopterygians but not in amniotes. In addition, while Col8a1 is known to be robustly expressed in mammalian osteoblasts, the expression levels of its amphibian orthologue are dramatically reduced. Our work reveals that while a skeletal expression of network forming collagen members is widespread throughout vertebrates, osteoblasts from divergent vertebrate lineages express different combinations of network forming collagen paralogues.

  18. Endochondral ossification for enhancing bone regeneration: converging native extracellular matrix biomaterials and developmental engineering in vivo.

    Science.gov (United States)

    Dennis, S Connor; Berkland, Cory J; Bonewald, Lynda F; Detamore, Michael S

    2015-06-01

    Autologous bone grafting (ABG) remains entrenched as the gold standard of treatment in bone regenerative surgery. Consequently, many marginally successful bone tissue engineering strategies have focused on mimicking portions of ABG's "ideal" osteoconductive, osteoinductive, and osteogenic composition resembling the late reparative stage extracellular matrix (ECM) in bone fracture repair, also known as the "hard" or "bony" callus. An alternative, less common approach that has emerged in the last decade harnesses endochondral (EC) ossification through developmental engineering principles, which acknowledges that the molecular and cellular mechanisms involved in developmental skeletogenesis, specifically EC ossification, are closely paralleled during native bone healing. EC ossification naturally occurs during the majority of bone fractures and, thus, can potentially be utilized to enhance bone regeneration for nearly any orthopedic indication, especially in avascular critical-sized defects where hypoxic conditions favor initial chondrogenesis instead of direct intramembranous ossification. The body's native EC ossification response, however, is not capable of regenerating critical-sized defects without intervention. We propose that an underexplored potential exists to regenerate bone through the native EC ossification response by utilizing strategies which mimic the initial inflammatory or fibrocartilaginous ECM (i.e., "pro-" or "soft" callus) observed in the early reparative stage of bone fracture repair. To date, the majority of strategies utilizing this approach rely on clinically burdensome in vitro cell expansion protocols. This review will focus on the confluence of two evolving areas, (1) native ECM biomaterials and (2) developmental engineering, which will attempt to overcome the technical, business, and regulatory challenges that persist in the area of bone regeneration. Significant attention will be given to native "raw" materials and ECM-based designs that

  19. An Osteoconductive Antibiotic Bone Eluting Putty with a Custom Polymer Matrix

    Directory of Open Access Journals (Sweden)

    John Curley

    2016-06-01

    Full Text Available With the rising tide of antibiotic resistant bacteria, extending the longevity of the current antibiotic arsenal is becoming a necessity. Developing local, controlled release antibiotic strategies, particularly for difficult to penetrate tissues such as bone, may prove to be a better alternative. Previous efforts to develop an osteoconductive local antibiotic release device for bone were created as solid molded composites; however, intimate contact with host bone was found to be critical to support host bone regrowth; thus, an osteocondconductive antibiotic releasing bone void filling putty was developed. Furthermore, a controlled releasing polymer matrix was refined using pendant-functionalized diols to provide tailorable pharmacokinetics. In vitro pharmacokinetic and bioactivity profiles were compared for a putty formulation with an analogous composition as its molded counterpart as well as four new pendant-functionalized polymers. A best-fit analysis of polymer composition in either small cylindrical disks or larger spheres revealed that the new pendant-functionalized polymers appear to release vancomycin via both diffusion and erosion regardless of the geometry of the putty. In silico simulations, a valuable technique for diffusion mediated controlled release models, will be used to confirm and optimize this property.

  20. Nucleation of apatite crystals in vitro by self-assembled dentin matrix protein 1

    Science.gov (United States)

    He, Gen; Dahl, Tom; Veis, Arthur; George, Anne

    2003-08-01

    Bones and teeth are biocomposites that require controlled mineral deposition during their self-assembly to form tissues with unique mechanical properties. Acidic extracellular matrix proteins play a pivotal role during biomineral formation. However, the mechanisms of protein-mediated mineral initiation are far from understood. Here we report that dentin matrix protein 1 (DMP1), an acidic protein, can nucleate the formation of hydroxyapatite in vitro in a multistep process that begins by DMP1 binding calcium ions and initiating mineral deposition. The nucleated amorphous calcium phosphate precipitates ripen and nanocrystals form. Subsequently, these expand and coalesce into microscale crystals elongated in the c-axis direction. Characterization of the functional domains in DMP1 demonstrated that intermolecular assembly of acidic clusters into a β-sheet template was essential for the observed mineral nucleation. Protein-mediated initiation of nanocrystals, as discussed here, might provide a new methodology for constructing nanoscale composites by self-assembly of polypeptides with tailor-made peptide sequences.

  1. Stimulatory effect of puerarin on bone formation through co-activation of nitric oxide and bone morphogenetic protein-2/mitogen-activated protein kinases pathways in mice

    Institute of Scientific and Technical Information of China (English)

    SHEU Shiow-yunn; TSAI Chia-chung; SUN Jui-sheng; CHEN Ming-hong; LIU Man-hai; SUN Man-ger

    2012-01-01

    Background Estrogen deficiency results in loss of bone mass.Phytoestrogens are plant-derived non-steroidal compounds with estrogen-like activity that bind to estrogen receptors.The main aim of this study was to investigate the effect of the phytoestrogen puerarin on adult mouse osteoblasts.Methods Osteoblast cells were harvested from 8-month old female imprinting control region (ICR) mice.The effects of puerarin stimulation on the proliferation,differentiation and maturation of osteoblasts were examined.The production of nitric oxide (NO) and the expression of bone morphogenetic protein-2 (BMP-2),SMAD4,mitogen-activated protein kinases (MAPK),core binding factor α1/runt-related transcription factor 2 (Cbfa1/Runx2),osteoprotegerin (OPG),and receptor activator of NF-kB ligand (RANKL) genes were analyzed.The activation of signal pathways was further confirmed by specific pathway inhibitors.Results The osteoblast viability reached its maximum at 10-8 mol/L puerarin.At this concentration,puerarin increases the proliferation and matrix mineralization of osteoblasts and promotes NO synthesis.With 10-8 mol/L puerarin treatment,BMP-2,SMAD4,Cbfa1/Runx2,and OPG gene expression were up-regulated,while the RANKL gene expression is down-regulated.Concurrent treatment involving the (bone morphogenetic protein) BMP antagonist Noggin or the NOS inhibitor L-NAME diminishes puerarin induced cell proliferation,Alkaline phosphatase (ALP) activity,NO production,as well as the BMP-2,SMAD4,Cbfa1/Runx2,OPG,and RANKL gene expression.Conclusions In this in vitro study,we demonstrate that puerarin is a bone anabolic agent that exerts its osteogenic effects through the induction of BMP-2 and NO synthesis,subsequently regulating Cbfa1/Runx2,OPG,and RANKL gene expression.This effect may contribute to its induction of osteoblast proliferation and differentiation,resulting in bone formation.

  2. Biological conduits combining bone marrow mesenchymal stem cells and extracellular matrix to treat long-segment sciatic nerve defects

    Directory of Open Access Journals (Sweden)

    Yang Wang

    2015-01-01

    Full Text Available The transplantation of polylactic glycolic acid conduits combining bone marrow mesenchymal stem cells and extracellular matrix gel for the repair of sciatic nerve injury is effective in some respects, but few data comparing the biomechanical factors related to the sciatic nerve are available. In the present study, rabbit models of 10-mm sciatic nerve defects were prepared. The rabbit models were repaired with autologous nerve, a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells, or a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel. After 24 weeks, mechanical testing was performed to determine the stress relaxation and creep parameters. Following sciatic nerve injury, the magnitudes of the stress decrease and strain increase at 7,200 seconds were largest in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group, followed by the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group, and then the autologous nerve group. Hematoxylin-eosin staining demonstrated that compared with the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells group and the autologous nerve group, a more complete sciatic nerve regeneration was found, including good myelination, regularly arranged nerve fibers, and a completely degraded and resorbed conduit, in the polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel group. These results indicate that bridging 10-mm sciatic nerve defects with a polylactic glycolic acid conduit + bone marrow mesenchymal stem cells + extracellular matrix gel construct increases the stress relaxation under a constant strain, reducing anastomotic tension. Large elongations under a constant physiological load can limit the anastomotic opening and shift, which is beneficial for the regeneration and functional reconstruction of sciatic nerve. Better

  3. Bone morphogenetic protein-2-encapsulated grafted-poly-lactic acid-polycaprolactone nanoparticles promote bone repair.

    Science.gov (United States)

    Xu, Xiaojun; Yang, Jun; Ding, Lifeng; Li, Jianjun

    2015-01-01

    The aim of this study is to test the efficacy of a novel tissue-engineered bone in repairing bone defects, using poly-lactic-acid-polycaprolactone (PLA-PCL) scaffolding seeded with PEG-bone morphogenetic protein-2 (BMP-2)-transfected rBMSCs (rabbit bone marrow stromal cells). The rBMSCs were transfected with PEG/BMP-2 or liposome/BMP-2, and then implanted into a PLA-PCL tissue-engineered bone. The protein level of BMP-2 was assessed by Western blot analysis and immunohistochemistry. ELISA was used to measure the amount of BMP-2 secreted in the culture media. The mRNA level of BMP-2 and osteocalcin was assayed quantitatively by real-time PCR. The middle portion of the bilateral radius in New Zealand rabbits was excised and implanted with tissue-engineered bone, and the modified areas were monitored by X-ray, hematoxylin-eosin staining, and immunohistochemistry staining of BMP-2. PEG-BMP-2 nanoparticles (NPs) and BMP-2-loaded PEG-PLA-PCL tissue-engineered bones were successfully constructed. The novel PEG-PLA-PCL NPs/DNA complex was a superior option for transfecting BMP-2 in rBMSCs compared to normal liposomes Moreover, the mRNA level of osteocalcin and alkaline phosphatase activity was also elevated upon transfection of BMP-2-encapsulated NPs. In vivo implants with BMP-2-carried tissue-engineered bone exhibited dramatic augmentation of BMP-2 and effective bone formation in the rabbit ectopic model. The PEG-PLA-PCL NPs/BMP-2 complex had an advantageous effect on bone repair, which provided an important theoretic basis for potential clinical treatments.

  4. Oxidized alginate hydrogels for bone morphogenetic protein-2 delivery in long bone defects.

    Science.gov (United States)

    Priddy, Lauren B; Chaudhuri, Ovijit; Stevens, Hazel Y; Krishnan, Laxminarayanan; Uhrig, Brent A; Willett, Nick J; Guldberg, Robert E

    2014-10-01

    Autograft treatment of large bone defects and fracture non-unions is complicated by limited tissue availability and donor site morbidity. Polymeric biomaterials such as alginate hydrogels provide an attractive tissue engineering alternative due to their biocompatibility, injectability, and tunable degradation rates. Irradiated RGD-alginate hydrogels have been used to deliver proteins such as bone morphogenetic protein-2 (BMP-2), to promote bone regeneration and restoration of function in a critically sized rat femoral defect model. However, slow degradation of irradiated alginate hydrogels may impede integration and remodeling of the regenerated bone to its native architecture. Oxidation of alginate has been used to promote degradation of alginate matrices. The objective of this study was to evaluate the effects of alginate oxidation on BMP-2 release and bone regeneration. We hypothesized that oxidized-irradiated alginate hydrogels would elicit an accelerated release of BMP-2, but degrade faster in vivo, facilitating the formation of higher quality, more mature bone compared to irradiated alginate. Indeed, oxidation of irradiated alginate did accelerate in vitro BMP-2 release. Notably, the BMP-2 retained within both constructs was bioactive at 26days, as observed by induction of alkaline phosphatase activity and positive Alizarin Red S staining of MC3T3-E1 cells. From the in vivo study, robust bone regeneration was observed in both groups through 12weeks by radiography, micro-computed tomography analyses, and biomechanical testing. Bone mineral density was significantly greater for the oxidized-irradiated alginate group at 8weeks. Histological analyses of bone defects revealed enhanced degradation of oxidized-irradiated alginate and suggested the presence of more mature bone after 12weeks of healing.

  5. Impact of extracellular matrix derived from osteoarthritis subchondral bone osteoblasts on osteocytes: role of integrinβ1 and focal adhesion kinase signaling cues.

    Science.gov (United States)

    Prasadam, Indira; Farnaghi, Saba; Feng, Jian Q; Gu, Wenyi; Perry, Samuel; Crawford, Ross; Xiao, Yin

    2013-10-09

    Our recent study indicated that subchondral bone pathogenesis in osteoarthritis (OA) is associated with osteocyte morphology and phenotypic abnormalities. However, the mechanism underlying this abnormality needs to be identified. In this study we investigated the effect of extracellular matrix (ECM) produced from normal and OA bone on osteocytic cells function. De-cellularized matrices, resembling the bone provisional ECM secreted from primary human subchondral bone osteoblasts (SBOs) of normal and OA patients were used as a model to study the effect on osteocytic cells. Osteocytic cells (MLOY4 osteocyte cell line) cultured on normal and OA derived ECMs were analyzed by confocal microscopy, scanning electron microscopy (SEM), cell attachment assays, zymography, apoptosis assays, qRT-PCR and western blotting. The role of integrinβ1 and focal adhesion kinase (FAK) signaling pathways during these interactions were monitored using appropriate blocking antibodies. The ECM produced by OA SBOs contained less mineral content, showed altered organization of matrix proteins and matrix structure compared with the matrices produced by normal SBOs. Culture of osteocytic cells on these defective OA ECM resulted in a decrease of integrinβ1 expression and the de-activation of FAK cell signaling pathway, which subsequently affected the initial osteocytic cell's attachment and functions including morphological abnormalities of cytoskeletal structures, focal adhesions, increased apoptosis, altered osteocyte specific gene expression and increased Matrix metalloproteinases (MMP-2) and -9 expression. This study provides new insights in understanding how altered OA bone matrix can lead to the abnormal osteocyte phenotypic changes, which is typical in OA pathogenesis.

  6. Hyperunstable matrix proteins in the byssus of Mytilus galloprovincialis.

    Science.gov (United States)

    Sagert, Jason; Waite, J Herbert

    2009-07-01

    The marine mussel Mytilus galloprovincialis is tethered to rocks in the intertidal zone by a holdfast known as the byssus. Functioning as a shock absorber, the byssus is composed of threads, the primary molecular components of which are collagen-containing proteins (preCOLs) that largely dictate the higher order self-assembly and mechanical properties of byssal threads. The threads contain additional matrix components that separate and perhaps lubricate the collagenous microfibrils during deformation in tension. In this study, the thread matrix proteins (TMPs), a glycine-, tyrosine- and asparagine-rich protein family, were shown to possess unique repeated sequence motifs, significant transcriptional heterogeneity and were distributed throughout the byssal thread. Deamidation was shown to occur at a significant rate in a recombinant TMP and in the byssal thread as a function of time. Furthermore, charge heterogeneity presumably due to deamidation was observed in TMPs extracted from threads. The TMPs were localized to the preCOL-containing secretory granules in the collagen gland of the foot and are assumed to provide a viscoelastic matrix around the collagenous fibers in byssal threads.

  7. Biofilm-specific extracellular matrix proteins of nontypeable Haemophilus influenzae.

    Science.gov (United States)

    Wu, Siva; Baum, Marc M; Kerwin, James; Guerrero, Debbie; Webster, Simon; Schaudinn, Christoph; VanderVelde, David; Webster, Paul

    2014-12-01

    Nontypeable Haemophilus influenzae (NTHi), a human respiratory tract pathogen, can form colony biofilms in vitro. Bacterial cells and the amorphous extracellular matrix (ECM) constituting the biofilm can be separated using sonication. The ECM from 24- and 96-h NTHi biofilms contained polysaccharides and proteinaceous components as detected by nuclear magnetic resonance (NMR) and Fourier transform infrared spectroscopy (FTIR) spectroscopy. More conventional chemical assays on the biofilm ECM confirmed the presence of these components and also DNA. Proteomics revealed eighteen proteins present in biofilm ECM that were not detected in planktonic bacteria. One ECM protein was unique to 24-h biofilms, two were found only in 96-h biofilms, and fifteen were present in the ECM of both 24- and 96-h NTHi biofilms. All proteins identified were either associated with bacterial membranes or cytoplasmic proteins. Immunocytochemistry showed two of the identified proteins, a DNA-directed RNA polymerase and the outer membrane protein OMP P2, associated with bacteria and biofilm ECM. Identification of biofilm-specific proteins present in immature biofilms is an important step in understanding the in vitro process of NTHi biofilm formation. The presence of a cytoplasmic protein and a membrane protein in the biofilm ECM of immature NTHi biofilms suggests that bacterial cell lysis may be a feature of early biofilm formation.

  8. STUDY ON NUCLEAR MATRIX PROTEINS FROM HUMAN BREAST CARCINOMA

    Institute of Scientific and Technical Information of China (English)

    HE Qian; ZHANG Shu-qun; CHU Yong-lie; JIA Xiao-li; JIANG Jian-tao

    2009-01-01

    Objective To investigate the marker protein of human breast carcinoma from nuclear matrix proteins (NMPs).Methods NMPs were injected subcutaneously into rabbit to get antiserum, which was used to detect the NMPs specificity for breast carcinoma.Results There was an apparent positive band (100kD) in the NMPs of breast carcinoma, which did not exist in normal breast and other tumors that were detected.Conclusion One or one group of 100kD NMPs were found to be related to human breast carcinoma, which may be involved in the carcinogenesis and development of human breast carcinoma and valuable for breast carcinoma diagnosis.

  9. Unconventional Secretion of Ebola Virus Matrix Protein VP40

    OpenAIRE

    Reynard, Olivier; Reid, St. Patrick; Page, Audrey; Mateo, Mathieu; Alazard-Dany, Nathalie; Raoul, Hervé; Basler, Christopher F.; Volchkov, Viktor E.

    2011-01-01

    The Ebola virus matrix protein VP40 plays an essential role in virus assembly and budding. In this study we reveal that transient VP40 expression results in the release into the culture medium of substantial amounts of soluble monomeric VP40 in addition to the release of virus-like particles containing an oligomeric form of this protein as previously described. We show that VP40 secretion is endoplasmic reticulum/Golgi–independent and is not associated with cell death. Soluble VP40 was observ...

  10. Effect of allogenic freeze-dried demineralized bone matrix on guided tissue regeneration in dogs.

    Science.gov (United States)

    Caplanis, N; Lee, M B; Zimmerman, G J; Selvig, K A; Wikesjö, U M

    1998-08-01

    This randomized, split-mouth study was designed to evaluate the adjunctive effect of allogenic, freeze-dried, demineralized bone matrix (DBM) to guided tissue regeneration (GTR). Contralateral fenestration defects (6 x 4 mm) were created 6 mm apical to the buccal alveolar crest on maxillary canine teeth in 6 beagle dogs. DBM was implanted into one randomly selected fenestration defect. Expanded polytetrafluoroethylene (ePTFE) membranes were used to provide bilateral GTR. Tissue blocks including defects with overlying membranes and soft tissues were harvested following a four-week healing interval and prepared for histometric analysis. Differences between GTR+DBM and GTR defects were evaluated using a paired t-test (N = 6). DBM was discernible in all GTR+DBM defects with limited, if any, evidence of bone metabolic activity. Rather, the DBM particles appeared solidified within a dense connective tissue matrix, often in close contact to the instrumented root. There were no statistically significant differences between the GTR+DBM versus the GTR condition for any histometric parameter examined. Fenestration defect height averaged 3.7+/-0.3 and 3.9+/-0.3 mm, total bone regeneration 0.8+/-0.6 and 1.5+/-0.8 mm, and total cementum regeneration 2.0+/-1.3 and 1.6+/-1.7 mm for GTR+DBM and GTR defects, respectively. The histologic and histometric observations, in concert, suggest that allogenic freeze-dried DBM has no adjunctive effect to GTR in periodontal fenestration defects over a four-week healing interval. The critical findings were 1) the DBM particles remained, embedded in dense connective tissue without evidence of bone metabolic activity; and 2) limited and similar amounts of bone and cementum regeneration were observed for both the GTR+DBM and GTR defects.

  11. Two-Stage Revision Anterior Cruciate Ligament Reconstruction: Bone Grafting Technique Using an Allograft Bone Matrix.

    Science.gov (United States)

    Chahla, Jorge; Dean, Chase S; Cram, Tyler R; Civitarese, David; O'Brien, Luke; Moulton, Samuel G; LaPrade, Robert F

    2016-02-01

    Outcomes of primary anterior cruciate ligament (ACL) reconstruction have been reported to be far superior to those of revision reconstruction. However, as the incidence of ACL reconstruction is rapidly increasing, so is the number of failures. The subsequent need for revision ACL reconstruction is estimated to occur in up to 13,000 patients each year in the United States. Revision ACL reconstruction can be performed in one or two stages. A two-stage approach is recommended in cases of improper placement of the original tunnels or in cases of unacceptable tunnel enlargement. The aim of this study was to describe the technique for allograft ACL tunnel bone grafting in patients requiring a two-stage revision ACL reconstruction.

  12. Pooled-matrix protein interaction screens using Barcode Fusion Genetics.

    Science.gov (United States)

    Yachie, Nozomu; Petsalaki, Evangelia; Mellor, Joseph C; Weile, Jochen; Jacob, Yves; Verby, Marta; Ozturk, Sedide B; Li, Siyang; Cote, Atina G; Mosca, Roberto; Knapp, Jennifer J; Ko, Minjeong; Yu, Analyn; Gebbia, Marinella; Sahni, Nidhi; Yi, Song; Tyagi, Tanya; Sheykhkarimli, Dayag; Roth, Jonathan F; Wong, Cassandra; Musa, Louai; Snider, Jamie; Liu, Yi-Chun; Yu, Haiyuan; Braun, Pascal; Stagljar, Igor; Hao, Tong; Calderwood, Michael A; Pelletier, Laurence; Aloy, Patrick; Hill, David E; Vidal, Marc; Roth, Frederick P

    2016-04-22

    High-throughput binary protein interaction mapping is continuing to extend our understanding of cellular function and disease mechanisms. However, we remain one or two orders of magnitude away from a complete interaction map for humans and other major model organisms. Completion will require screening at substantially larger scales with many complementary assays, requiring further efficiency gains in proteome-scale interaction mapping. Here, we report Barcode Fusion Genetics-Yeast Two-Hybrid (BFG-Y2H), by which a full matrix of protein pairs can be screened in a single multiplexed strain pool. BFG-Y2H uses Cre recombination to fuse DNA barcodes from distinct plasmids, generating chimeric protein-pair barcodes that can be quantified via next-generation sequencing. We applied BFG-Y2H to four different matrices ranging in scale from ~25 K to 2.5 M protein pairs. The results show that BFG-Y2H increases the efficiency of protein matrix screening, with quality that is on par with state-of-the-art Y2H methods.

  13. Biomimetic tendon extracellular matrix composite gradient scaffold enhances ligament-to-bone junction reconstruction.

    Science.gov (United States)

    Liu, Huanhuan; Yang, Long; Zhang, Erchen; Zhang, Rui; Cai, Dandan; Zhu, Shouan; Ran, Jisheng; Bunpetch, Varitsara; Cai, Youzhi; Heng, Boon Chin; Hu, Yejun; Dai, Xuesong; Chen, Xiao; Ouyang, Hongwei

    2017-07-01

    Management of ligament/tendon-to-bone-junction healing remains a formidable challenge in the field of orthopedic medicine to date, due to deficient vascularity and multi-tissue transitional structure of the junction. Numerous strategies have been employed to improve ligament-bone junction healing, including delivery of stem cells, bioactive factors, and synthetic materials, but these methods are often inadequate at recapitulating the complex structure-function relationships at native tissue interfaces. Here, we developed an easily-fabricated and effective biomimetic composite to promote the regeneration of ligament-bone junction by physically modifying the tendon extracellular matrix (ECM) into a Random-Aligned-Random composite using ultrasound treatment. The differentiation potential of rabbit bone marrow stromal cells on the modified ECM were examined in vitro. The results demonstrated that the modified ECM enhanced expression of chondrogenesis and osteogenesis-associated epigenetic genes (Jmjd1c, Kdm6b), transcription factor genes (Sox9, Runx2) and extracellular matrix genes (Col2a1, Ocn), resulting in higher osteoinductivity than the untreated tendon ECM in vitro. In the rabbit anterior cruciate ligament (ACL) reconstruction model in vivo, micro-computed tomography (Micro-CT) and histological analysis showed that the modified Random-Aligned-Random composite scaffold enhanced bone and fibrocartilage formation at the interface, more efficaciously than the unmodified tendon ECM. Therefore, these results demonstrated that the biomimetic Random-Aligned-Random composite could be a promising scaffold for ligament/tendon-bone junction repair. The native transitional region consists of several distinct yet contiguous tissue regions, composed of soft tissue, non-calcified fibrocartilage, calcified fibrocartilage, and bone. A stratified graft whose phases are interconnected with each other is essential for supporting the formation of functionally continuous multi

  14. Pigment epithelium derived factor suppresses expression of Sost/Sclerostin by osteocytes: implication for its role in bone matrix mineralization.

    Science.gov (United States)

    Li, Feng; Song, Na; Tombran-Tink, Joyce; Niyibizi, Christopher

    2015-06-01

    Mutations in Serpinf1 gene which encodes pigment epithelium derived factor (PEDF) lead to osteogenesis imperfecta type VI whose hallmark is defective mineralization. Mechanisms by which PEDF regulates matrix mineralization remain unknown. We examined effect of exogenous PEDF on expression of osteoblastic and osteocytic related genes and proteins in mineralizing osteoblast culture. Mineralizing human osteoblasts supplemented with exogenous PEDF for 14 days deposited 47% more mineral than cells cultured without PEDF. Analysis of selected gene expression by cells in mineralizing cultures supplemented with exogenous PEDF showed reduction in expression of Sclerostin (Sost) by 70%, matrix extracellular phosphoglycoprotein (MEPE) by 75% and dentin matrix protein (DMP-1) by 20% at day 14 of culture. Phosphate-regulating gene with homologies to endopeptidases on the X chromosome (PHEX) expression was not affected. Western blotting and immunoprecipitation showed that sclerostin and MEPE synthesis by osteocytes were reduced by 50% and 60% respectively in mineralizing osteoblasts containing exogenous PEDF. Primary osteocytes exposed to PEDF also reduced synthesis of Sost/sclerostin by 50% within 24 h. For osteoblastic genes, Bone sialoprotein (BSP) was expressed at 75% higher by day 7 in cultures containing exogenous PEDF while Col1A1 expression remained high at all-time points. Total beta-catenin was increased in mineralizing osteoblastic cells suggesting increased Wnt activity. Taken together, the data indicate that PEDF suppressed expression of factors that inhibit mineralization while enhancing those that promote mineralization. The findings also suggest that PEDF may regulate Sost expression by osteocytes leading to enhanced osteoblastic differentiation and increased matrix mineralization.

  15. Two essential peritrophic matrix proteins mediate matrix barrier functions in the insect midgut.

    Science.gov (United States)

    Agrawal, Sinu; Kelkenberg, Marco; Begum, Khurshida; Steinfeld, Lea; Williams, Clay E; Kramer, Karl J; Beeman, Richard W; Park, Yoonseong; Muthukrishnan, Subbaratnam; Merzendorfer, Hans

    2014-06-01

    The peritrophic matrix (PM) in the midgut of insects consists primarily of chitin and proteins and is thought to support digestion and provide protection from abrasive food particles and enteric pathogens. We examined the physiological roles of 11 putative peritrophic matrix protein (PMP) genes of the red flour beetle, Tribolium castaneum (TcPMPs). TcPMP genes are differentially expressed along the length of the midgut epithelium of feeding larvae. RNAi of individual PMP genes revealed no abnormal developmental phenotypes for 9 of the 11 TcPMPs. However, RNAi for two PMP genes, TcPMP3 and TcPMP5-B, resulted in depletion of the fat body, growth arrest, molting defects and mortality. In situ permeability assays after oral administration of different-sized FITC-dextran beads demonstrated that the exclusion size of the larval peritrophic matrix (PM) decreases progressively from >2 MDa to RNAi for TcPMP3 and TcPMP5-B, these dextrans penetrated the epithelium of the median midgut, indicating loss of structural integrity and barrier function of the larval PM. In contrast, RNAi for TcPMP5-B, but not RNAi for TcPMP3, resulted in breakdown of impermeability to 4 and 40 kDa dextrans in the PM of the posterior midgut. These results suggest that specific PMPs are involved in the regulation of PM permeability, and that a gradient of barrier function is essential for survival and fat body maintenance.

  16. Molecular events in matrix protein metabolism in the aging kidney.

    Science.gov (United States)

    Sataranatarajan, Kavithalakshmi; Feliers, Denis; Mariappan, Meenalakshmi M; Lee, Hak Joo; Lee, Myung Ja; Day, Robert T; Yalamanchili, Hima Bindu; Choudhury, Goutam G; Barnes, Jeffrey L; Van Remmen, Holly; Richardson, Arlan; Kasinath, Balakuntalam S

    2012-12-01

    We explored molecular events associated with aging-induced matrix changes in the kidney. C57BL6 mice were studied in youth, middle age, and old age. Albuminuria and serum cystatin C level (an index of glomerular filtration) increased with aging. Renal hypertrophy was evident in middle-aged and old mice and was associated with glomerulomegaly and increase in mesangial fraction occupied by extracellular matrix. Content of collagen types I and III and fibronectin was increased with aging; increment in their mRNA varied with the phase of aging. The content of ZEB1 and ZEB2, collagen type I transcription inhibitors, and their binding to the collagen type Iα2 promoter by ChIP assay also showed age-phase-specific changes. Lack of increase in mRNA and data from polysome assay suggested decreased degradation as a potential mechanism for kidney collagen type I accumulation in the middle-aged mice. These changes occurred with increment in TGFβ mRNA and protein and activation of its SMAD3 pathway; SMAD3 binding to the collagen type Iα2 promoter was also increased. TGFβ-regulated microRNAs (miRs) exhibited selective regulation. The renal cortical content of miR-21 and miR-200c, but not miR-192, miR-200a, or miR-200b, was increased with aging. Increased miR-21 and miR-200c contents were associated with reduced expression of their targets, Sprouty-1 and ZEB2, respectively. These data show that aging is associated with complex molecular events in the kidney that are already evident in the middle age and progress to old age. Age-phase-specific regulation of matrix protein synthesis occurs and involves matrix protein-specific transcriptional and post-transcriptional mechanisms. © 2012 The Authors Aging Cell © 2012 Blackwell Publishing Ltd/Anatomical Society of Great Britain and Ireland.

  17. Cloning of matrix Gla protein in a marine cartilaginous fish, Prionace glauca: preferential protein accumulation in skeletal and vascular systems.

    Science.gov (United States)

    Ortiz-Delgado, J B; Simes, D C; Viegas, C S B; Schaff, B J; Sarasquete, C; Cancela, M L

    2006-07-01

    Matrix Gla protein (MGP) belongs to the family of vitamin K dependent, Gla containing proteins and, in mammals, birds and Xenopus, its mRNA has been previously detected in bone, cartilage and soft tissue extracts, while the accumulation of the protein was found mainly in calcified tissues. More recently, the MGP gene expression was also studied in marine teleost fish where it was found to be associated with chondrocytes, smooth muscle and endothelial cells. To date no information is available on the sites of MGP expression or accumulation in cartilaginous fishes that diverged from osteichthyans, a group that includes mammals, over 400 million years ago. The main objectives of this work were to study the sites of MGP gene expression and protein accumulation by means of in situ hybridization and immunohistochemistry. MGP mRNA and protein were localized as expected not only in cartilage from branchial arches and vertebra but also in the endothelia of the vascular system as well as in the tubular renal endothelium. The accumulation of MGP in non mineralized soft tissues was unexpected and suggests differences in localization or regulation of this protein in shark soft tissues compared to tetrapods and teleosts. Our results also corroborate the hypothesis that in Prionace glauca, as previously shown in mammals, the MGP protein probably also acts as a calcification inhibitor, protecting soft tissues from abnormal and ectopic calcification.

  18. Quantitative (31)P NMR spectroscopy and (1)H MRI measurements of bone mineral and matrix density differentiate metabolic bone diseases in rat models.

    Science.gov (United States)

    Cao, Haihui; Nazarian, Ara; Ackerman, Jerome L; Snyder, Brian D; Rosenberg, Andrew E; Nazarian, Rosalynn M; Hrovat, Mirko I; Dai, Guangping; Mintzopoulos, Dionyssios; Wu, Yaotang

    2010-06-01

    In this study, bone mineral density (BMD) of normal (CON), ovariectomized (OVX), and partially nephrectomized (NFR) rats was measured by (31)P NMR spectroscopy; bone matrix density was measured by (1)H water- and fat-suppressed projection imaging (WASPI); and the extent of bone mineralization (EBM) was obtained by the ratio of BMD/bone matrix density. The capability of these MR methods to distinguish the bone composition of the CON, OVX, and NFR groups was evaluated against chemical analysis (gravimetry). For cortical bone specimens, BMD of the CON and OVX groups was not significantly different; BMD of the NFR group was 22.1% (by (31)P NMR) and 17.5% (by gravimetry) lower than CON. For trabecular bone specimens, BMD of the OVX group was 40.5% (by (31)P NMR) and 24.6% (by gravimetry) lower than CON; BMD of the NFR group was 26.8% (by (31)P NMR) and 21.5% (by gravimetry) lower than CON. No significant change of cortical bone matrix density between CON and OVX was observed by WASPI or gravimetry; NFR cortical bone matrix density was 10.3% (by WASPI) and 13.9% (by gravimetry) lower than CON. OVX trabecular bone matrix density was 38.0% (by WASPI) and 30.8% (by gravimetry) lower than CON, while no significant change in NFR trabecular bone matrix density was observed by either method. The EBMs of OVX cortical and trabecular specimens were slightly higher than CON but not significantly different from CON. Importantly, EBMs of NFR cortical and trabecular specimens were 12.4% and 26.3% lower than CON by (31)P NMR/WASPI, respectively, and 4.0% and 11.9% lower by gravimetry. Histopathology showed evidence of osteoporosis in the OVX group and severe secondary hyperparathyroidism (renal osteodystrophy) in the NFR group. These results demonstrate that the combined (31)P NMR/WASPI method is capable of discerning the difference in EBM between animals with osteoporosis and those with impaired bone mineralization.

  19. The molecular elasticity of the extracellular matrix protein tenascin

    Science.gov (United States)

    Oberhauser, Andres F.; Marszalek, Piotr E.; Erickson, Harold P.; Fernandez, Julio M.

    1998-05-01

    Extracellular matrix proteins are thought to provide a rigid mechanical anchor that supports and guides migrating and rolling cells. Here we examine the mechanical properties of the extracellular matrix protein tenascin by using atomic-force-microscopy techniques. Our results indicate that tenascin is an elastic protein. Single molecules of tenascin could be stretched to several times their resting length. Force-extension curves showed a saw-tooth pattern, with peaks of force at 137pN. These peaks were ~25nm apart. Similar results have been obtained by study of titin. We also found similar results by studying recombinant tenascin fragments encompassing the 15 fibronectin type III domains of tenascin. This indicates that the extensibility of tenascin may be due to the stretch-induced unfolding of its fibronectin type III domains. Refolding of tenascin after stretching, observed when the force was reduced to near zero, showed a double-exponential recovery with time constants of 42 domains refolded per second and 0.5 domains per second. The former speed of refolding is more than twice as fast as any previously reported speed of refolding of a fibronectin type III domain,. We suggest that the extensibility of the modular fibronectin type III region may be important in allowing tenascin-ligand bonds to persist over long extensions. These properties of fibronectin type III modules may be of widespread use in extracellular proteins containing such domain,.

  20. Distance matrix-based approach to protein structure prediction.

    Science.gov (United States)

    Kloczkowski, Andrzej; Jernigan, Robert L; Wu, Zhijun; Song, Guang; Yang, Lei; Kolinski, Andrzej; Pokarowski, Piotr

    2009-03-01

    Much structural information is encoded in the internal distances; a distance matrix-based approach can be used to predict protein structure and dynamics, and for structural refinement. Our approach is based on the square distance matrix D = [r(ij)(2)] containing all square distances between residues in proteins. This distance matrix contains more information than the contact matrix C, that has elements of either 0 or 1 depending on whether the distance r (ij) is greater or less than a cutoff value r (cutoff). We have performed spectral decomposition of the distance matrices D = sigma lambda(k)V(k)V(kT), in terms of eigenvalues lambda kappa and the corresponding eigenvectors v kappa and found that it contains at most five nonzero terms. A dominant eigenvector is proportional to r (2)--the square distance of points from the center of mass, with the next three being the principal components of the system of points. By predicting r (2) from the sequence we can approximate a distance matrix of a protein with an expected RMSD value of about 7.3 A, and by combining it with the prediction of the first principal component we can improve this approximation to 4.0 A. We can also explain the role of hydrophobic interactions for the protein structure, because r is highly correlated with the hydrophobic profile of the sequence. Moreover, r is highly correlated with several sequence profiles which are useful in protein structure prediction, such as contact number, the residue-wise contact order (RWCO) or mean square fluctuations (i.e. crystallographic temperature factors). We have also shown that the next three components are related to spatial directionality of the secondary structure elements, and they may be also predicted from the sequence, improving overall structure prediction. We have also shown that the large number of available HIV-1 protease structures provides a remarkable sampling of conformations, which can be viewed as direct structural information about the

  1. Salicylic Acid-Based Polymers for Guided Bone Regeneration Using Bone Morphogenetic Protein-2.

    Science.gov (United States)

    Subramanian, Sangeeta; Mitchell, Ashley; Yu, Weiling; Snyder, Sabrina; Uhrich, Kathryn; O'Connor, J Patrick

    2015-07-01

    Bone morphogenetic protein-2 (BMP-2) is used clinically to promote spinal fusion, treat complex tibia fractures, and to promote bone formation in craniomaxillofacial surgery. Excessive bone formation at sites where BMP-2 has been applied is an established complication and one that could be corrected by guided tissue regeneration methods. In this study, anti-inflammatory polymers containing salicylic acid [salicylic acid-based poly(anhydride-ester), SAPAE] were electrospun with polycaprolactone (PCL) to create thin flexible matrices for use as guided bone regeneration membranes. SAPAE polymers hydrolyze to release salicylic acid, which is a nonsteroidal anti-inflammatory drug. PCL was used to enhance the mechanical integrity of the matrices. Two different SAPAE-containing membranes were produced and compared: fast-degrading (FD-SAPAE) and slow-degrading (SD-SAPAE) membranes that release salicylic acid at a faster and slower rate, respectively. Rat femur defects were treated with BMP-2 and wrapped with FD-SAPAE, SD-SAPAE, or PCL membrane or were left unwrapped. The effects of different membranes on bone formation within and outside of the femur defects were measured by histomorphometry and microcomputed tomography. Bone formation within the defect was not affected by membrane wrapping at BMP-2 doses of 12 μg or more. In contrast, the FD-SAPAE membrane significantly reduced bone formation outside the defect compared with all other treatments. The rapid release of salicylic acid from the FD-SAPAE membrane suggests that localized salicylic acid treatment during the first few days of BMP-2 treatment can limit ectopic bone formation. The data support development of SAPAE polymer membranes for guided bone regeneration applications as well as barriers to excessive bone formation.

  2. ANA deficiency enhances bone morphogenetic protein-induced ectopic bone formation via transcriptional events.

    Science.gov (United States)

    Miyai, Kentaro; Yoneda, Mitsuhiro; Hasegawa, Urara; Toita, Sayaka; Izu, Yayoi; Hemmi, Hiroaki; Hayata, Tadayoshi; Ezura, Yoichi; Mizutani, Shuki; Miyazono, Kohei; Akiyoshi, Kazunari; Yamamoto, Tadashi; Noda, Masaki

    2009-04-17

    Ectopic bone formation after joint replacement or brain injury in humans is a serious complication that causes immobility of joints and severe pain. However, mechanisms underlying such ectopic bone formation are not fully understood. Bone morphogenetic protein (BMPs) are defined as inducers of ectopic bone formation, and they are regulated by several types of inhibitors. ANA is an antiproliferative molecule that belongs to Tob/BTG family, but its activity in bone metabolism has not been known. Here, we examined the role of ANA on ectopic bone formation activity of BMP. In ANA-deficient and wild-type mice, BMP2 was implanted to induce ectopic bone formation in muscle. ANA deficiency increased mass of newly formed bone in vivo compared with wild-type based on 3D-muCT analyses. ANA mRNA was expressed in bone in vivo as well as in osteoblastic cells in vitro. Such ANA mRNA levels were increased by BMP2 treatment in MC3T3-E1 osteoblastic cells. Overexpression of ANA suppressed BMP-induced expression of luciferase reporter gene linked to BMP response elements in these cells. Conversely, ANA mRNA knockdown by small interference RNA enhanced the BMP-dependent BMP response element reporter expression. It also enhanced BMP-induced osteoblastic differentiation in muscle-derived C2C12 cells. Immunoprecipitation assay indicated that ANA interacts with Smad8. Thus, ANA is a suppressor of ectopic bone formation induced by BMP, and this inhibitory ANA activity is a part of the negative feedback regulation of BMP function.

  3. [Glycation of extracellular matrix proteins and its role in atherosclerosis].

    Science.gov (United States)

    Kuzan, Aleksandra; Chwiłkowska, Agnieszka; Kobielarz, Magdalena; Pezowicz, Celina; Gamian, Andrzej

    2012-10-29

    Glycation consists in formation of advanced glycation end-products (AGE) during non-enzymatic reaction between reducing sugars and proteins, lipids or nucleic acids. This review is focused mainly on glycation of collagen and its role in acceleration of vascular disease. Collagen is an extracellular matrix protein characterized by unique structure forming fibrils with great anti-tensile and anti-breaking strength. The protein builds the connective tissue and is responsible for biomechanical properties of blood vessels. It is reported that higher content of glycated collagen correlates with lower elasticity and greater toughness of the vessel walls and, as a consequence, a faster rate of atherosclerosis development. Numerous mechanisms connected with AGE formation are involved in atherogenesis, among others: receptor-mediated production of free radicals, triggering an inflammatory process, activation of leukocytes and thrombocytes, facilitation of LDL binding, change in level of growth factors, adhesion molecules, MMP and some other proteins' expression. The coverages allow the development of therapeutic strategies to prevent or slow down the pathological processes connected with glycation of collagen and other proteins in the artery wall. The main strategies are based on limitation of exogenous AGE, consumption of products which contain rutin, treatment with drugs which inhibit AGE formation, such as pyridoxamine, and chemicals which are able to cleave already formed AGE protein-protein crosslinks, such as ALT-711.

  4. Healing of periodontal defects treated with enamel matrix proteins and root surface conditioning--an experimental study in dogs.

    Science.gov (United States)

    Sakallioğlu, Umur; Açikgöz, Gökhan; Ayas, Bülent; Kirtiloğlu, Tuğrul; Sakallioğlu, Eser

    2004-05-01

    Application of enamel matrix proteins has been introduced as an alternative method for periodontal regenerative therapy. It is claimed that this approach provides periodontal regeneration by a biological approach, i.e. creating a matrix on the root surfaces that promotes cementum, periodontal ligament (PDL) and alveolar bone regeneration, thus mimicking the events occurring during tooth development. Although there have been numerous in vitro and in vivo studies demonstrating periodontal regeneration, acellular cementum formation and clinical outcomes via enamel matrix proteins usage, their effects on the healing pattern of soft and hard periodontal tissues are not well-established and compared with root conditioning alone. In the present study, the effects of Emdogain (Biora, Malmö, Sweden), an enamel matrix derivative mainly composed of enamel matrix proteins (test), on periodontal wound healing were evaluated and compared with root surface conditioning (performed with 36% orthophosphoric acid) alone (control) histopathologically and histomorphometrically by means of the soft and hard tissue profile of periodontium. An experimental periodontitis model performed at premolar teeth of four dogs were used in the study and the healing pattern of periodontal tissues was evaluated at days 7, 14, 21, 28 (one dog at each day), respectively. At day 7, soft tissue attachment evaluated by means of connective tissue and/or epithelial attachment to the root surfaces revealed higher connective tissue attachment rate in the test group and the amount of new connective tissue proliferation in the test group was significantly greater than the control group (p0.05). A firm attachment of acellular cementum to the root dentin with functional organization of its collagen fibers was noted, and, the accumulation and organization of cellular cementum in the control group was more irregular than the cellular cementum formed in the test group. The amount of new bone was 2.41+/-0.75 mm in

  5. Extracellular matrix-inspired growth factor delivery systems for bone regeneration

    Energy Technology Data Exchange (ETDEWEB)

    Martino, Mikaël M. [Osaka Univ. (Japan). Immunology Frontier Research Center; Briquez, Priscilla S. [Ecole Polytechnique Federale de Lausanne (Switzerland). Inst. of Bioengineering; Maruyama, Kenta [Osaka Univ. (Japan). Immunology Frontier Research Center; Hubbell, Jeffrey A. [Ecole Polytechnique Federale de Lausanne (Switzerland). Inst. of Bioengineering; Univ. of Chicago, IL (United States). Inst. for Molecular Engineering; Argonne National Lab. (ANL), Argonne, IL (United States)

    2015-04-17

    Growth factors are very promising molecules to enhance bone regeneration. However, their translation to clinical use has been seriously limited, facing issues related to safety and cost-effectiveness. These problems derive from the vastly supra-physiological doses of growth factor used without optimized delivery systems. Therefore, these issues have motivated the development of new delivery systems allowing better control of the spatio-temporal release and signaling of growth factors. Because the extracellular matrix (ECM) naturally plays a fundamental role in coordinating growth factor activity in vivo, a number of novel delivery systems have been inspired by the growth factor regulatory function of the ECM. After introducing the role of growth factors during the bone regeneration process, this review exposes different issues that growth factor-based therapies have encountered in the clinic and highlights recent delivery approaches based on the natural interaction between growth factor and the ECM.

  6. Biomineralization of bone: a fresh view of the roles of non-collagenous proteins

    Science.gov (United States)

    Gorski, Jeffrey Paul

    2015-01-01

    The impact of genetics has dramatically affected our understanding of the functions of non-collagenous proteins. Specifically, mutations and knockouts have defined their cellular spectrum of actions. However, the biochemical mechanisms mediated by non-collagenous proteins in biomineralization remain elusive. It is likely that this understanding will require more focused functional testing at the protein, cell, and tissue level. Although initially viewed as rather redundant and static acidic calcium binding proteins, it is now clear that non-collagenous proteins in mineralizing tissues represent diverse entities capable of forming multiple protein-protein nteractions which act in positive and negative ways to regulate the process of bone mineralization. Several new examples from the author’s laboratory are provided which illustrate this theme including an apparent activating effect of hydroxyapatite crystals on metalloproteinases. This review emphasizes the view that secreted non-collagenous proteins in mineralizing bone actively participate in the mineralization process and ultimately control where and how much mineral crystal is deposited, as well as determining the quality and biomechanical properties of the mineralized matrix produced. PMID:21622198

  7. Effect of the “protein diet” and bone tissue.

    Science.gov (United States)

    Nascimento da Silva, Zoraide; Azevedo de Jesuz, Vanessa; De Salvo Castro, Eduardo; Soares da Costa, Carlos Alberto; Teles Boaventura, Gilson; Blondet de Azeredo, Vilma

    2014-01-01

    The aim of this study is to evaluate the effect of the hyperproteic diet consumption on bone tissue. The study was conducted during sixty days. Twenty eight Wistar albinus rats, adults, originated from Laboratory of Experimental Nutrition were divided in four groups: (n = 7); Control 1 (C1), Control 2 (C2), Hyperproteic 1 (HP1) e Hyperproteic 2 (HP2). The C2 and HP2 groups were submitted to 30% of food restriction. The hyperproteic diet was based on the Atkins diet and prepared to simulate the protein diet. At the end of the study the animals were anesthetized to performer bone densitometry analyses by DEXA and blood and tissue collection. Serum and bone minerals analyses were conducted by colorimetric methods in automated equipment. The total bone mineral density (BMD) of the pelvis and the spine of the food restriction groups (HP2 e C2) were lower (p hyperproteic groups (HP1 e HP2). It was observed similar effect on the osteocalcin level, that presented lower (p hyperproteic groups. The insulin level was lower only in HP2 and serum calcium of the HP1 and HP2 groups was lower than C1. The protein diet promotes significant bone change on femur and in the hormones levels related to bone synthesis and maintenance of this tissue.

  8. Contribution of matrix vesicles and alkaline phosphatase to ectopic bone formation

    Directory of Open Access Journals (Sweden)

    Ciancaglini P.

    2006-01-01

    Full Text Available Endochondral calcification involves the participation of matrix vesicles (MVs, but it remains unclear whether calcification ectopically induced by implants of demineralized bone matrix also proceeds via MVs. Ectopic bone formation was induced by implanting rat demineralized diaphyseal bone matrix into the dorsal subcutaneous tissue of Wistar rats and was examined histologically and biochemically. Budding of MVs from chondrocytes was observed to serve as nucleation sites for mineralization during induced ectopic osteogenesis, presenting a diameter with Gaussian distribution with a median of 306 ± 103 nm. While the role of tissue-nonspecific alkaline phosphatase (TNAP during mineralization involves hydrolysis of inorganic pyrophosphate (PPi, it is unclear how the microenvironment of MV may affect the ability of TNAP to hydrolyze the variety of substrates present at sites of mineralization. We show that the implants contain high levels of TNAP capable of hydrolyzing p-nitrophenylphosphate (pNPP, ATP and PPi. The catalytic properties of glycosyl phosphatidylinositol-anchored, polidocanol-solubilized and phosphatidylinositol-specific phospholipase C-released TNAP were compared using pNPP, ATP and PPi as substrates. While the enzymatic efficiency (k cat/Km remained comparable between polidocanol-solubilized and membrane-bound TNAP for all three substrates, the k cat/Km for the phosphatidylinositol-specific phospholipase C-solubilized enzyme increased approximately 108-, 56-, and 556-fold for pNPP, ATP and PPi, respectively, compared to the membrane-bound enzyme. Our data are consistent with the involvement of MVs during ectopic calcification and also suggest that the location of TNAP on the membrane of MVs may play a role in determining substrate selectivity in this micro-compartment.

  9. The effect of enamel matrix derivative (Emdogain®) on gene expression profiles of human primary alveolar bone cells.

    Science.gov (United States)

    Yan, X Z; Rathe, F; Gilissen, C; van der Zande, M; Veltman, J; Junker, R; Yang, F; Jansen, J A; Walboomers, X F

    2014-06-01

    Emdogain® is frequently used in regenerative periodontal treatment. Understanding its effect on gene expression of bone cells would enable new products and pathways promoting bone formation to be established. The aim of the study was to analyse the effect of Emdogain® on expression profiles of human-derived bone cells with the help of the micro-array, and subsequent validation. Bone was harvested from non-smoking patients during dental implant surgery. After outgrowth, cells were cultured until subconfluence, treated for 24 h with either Emdogain® (100 µg/ml) or control medium, and subsequently RNA was isolated and micro-array was performed. The most important genes demonstrated by micro-array data were confirmed by qPCR and ELISA tests. Emdogain tipped the balance between genes expressed for bone formation and bone resorption towards a more anabolic effect, by interaction of the PGE2 pathway and inhibition of IL-7 production. In addition the results of the present study indicate that Emdogain possibly has an effect on gene expression for extracellular matrix formation of human bone cells, in particular on bone matrix formation and on proliferation and differentiation. With the micro-array and the subsequent validation, the genes possibly involved in Emdogain action on bone cells were identified. These results can contribute to establishing new products and pathways promoting bone formation.

  10. In vivo study of extracellular matrix coating enhancing fixation of the pedicle screw-bone's interface

    Institute of Scientific and Technical Information of China (English)

    LIU Guo-min; ZHANG Xing-yi; XU Chuan-jie; ZHU Xiao-min; WANG Jun; LIU Yi

    2011-01-01

    Background Based on in vivo research on the effect of the coating of the extracellular matrix composition of pedicle screws on the conduction and induction of bone formation in young sheep,the aim of this study was to investigate the application of coated pedicle screws in sheep with scoliosis whose spines are under constant development.Methods Four groups of pedicle screws were randomly implanted into bilateral L2-L5 pedicles of 2.5- to 3-month-old sheep.A static experiment was performed on one side and a loading test was performed on the other side by implanting connecting rods at the L2-L3 and L4-L5 segments.The changes in the force on the coated screws and the combination of the surface of the coated screws with the surrounding bone in the growth process of young sheep's spines with aging were observed.After 3 months,the lumbar vertebrae with the screws were removed and examined by micro-CT,histological,and biomechanical analyses.Results Under nonloading conditions,there is bone formation around the surfaces of coated screws.The bone forming on the surface of collagen/chondroitin sulfate/hydroxyapatite coating of pedicle screws is the most,the one of the collagen / chondrcitin sulfate coating and hydroxyapatite coating is followed,and no significant difference between the two groups.In terms of the trabecular bone morphology parameters of the region of interest around the surface of the pedicle screws,such as bone mineral content,bone mineral density,tissue mineral content,tissue bone mineral density,bone volume fraction,and connection density,those associated with collagen/chondroitin sulfate/hydroxyapatite coatings are largest and those unassociated with coatings are smallest.Under nonloading conditions,the pullout strength of the collagen/chondroitin sulfate/hydroxyapatite-coated screws was largest,and that of the uncoated screws was minimal (P <0.01).Under loading conditions,the maximum pullout strength of each group of pedicle screws was less than that

  11. Stromal cell-derived factor-1 potentiates bone morphogenetic protein-2 induced bone formation.

    Science.gov (United States)

    Higashino, Kosaku; Viggeswarapu, Manjula; Bargouti, Maggie; Liu, Hui; Titus, Louisa; Boden, Scott D

    2011-02-01

    The mechanisms driving bone marrow stem cell mobilization are poorly understood. A recent murine study found that circulating bone marrow-derived osteoprogenitor cells (MOPCs) were recruited to the site of recombinant human bone morphogenetic protein-2 (BMP-2)-induced bone formation. Stromal cell-derived factor-1α (SDF-1α) and its cellular receptor CXCR4 have been shown to mediate the homing of stem cells to injured tissues. We hypothesized that chemokines, such as SDF-1, are also involved with mobilization of bone marrow cells. The CD45(-) fraction is a major source of MOPCs. In this report we determined that the addition of BMP-2 or SDF-1 to collagen implants increased the number of MOPCs in the peripheral blood. BMP-2-induced mobilization was blocked by CXCR4 antibody, confirming the role of SDF-1 in mobilization. We determined for the first time that addition of SDF-1 to implants containing BMP-2 enhances mobilization, homing of MOPCs to the implant, and ectopic bone formation induced by suboptimal BMP-2 doses. These results suggest that SDF-1 increases the number of osteoprogenitor cells that are mobilized from the bone marrow and then home to the implant. Thus, addition of SDF-1 to BMP-2 may improve the efficiency of BMPs in vivo, making their routine use for orthopaedic applications more affordable and available to more patients.

  12. The role of Matrix Gla Protein in ossification and recovery of the avian growth plate

    Directory of Open Access Journals (Sweden)

    Harel eDan

    2012-07-01

    Full Text Available ECM mineralization is an essential physiologic process in bone, teeth, and hypertrophic cartilage. Matrix Gla Protein (MGP, an inhibitor of mineralization, is expressed by chondrocytes and vascular smooth muscle cells to inhibit calcification of those soft tissues.Tibial Dyschondroplasia (TD, a skeletal abnormality apparent as a plug of non-vascularized, non-mineralized, white opaque cartilage in the tibial growth plate of avian species can serve as a good model for studying process and genes involved in matrix mineralization and calcification. In this work, we studied the involvement of MGP in the development of TD, as well as in the processes of spontaneous and induced recovery from this syndrome. First, we found that during normal bone development, MGP is expressed in specific time and locations, starting from wide spread expression in the yet un-ossified diaphysis during embryonic development, to specific expression in hypertrophic chondrocytes adjacent to the chondro-osseous junction and the secondary ossification center just prior to calcification. In addition, we show that MGP is not expressed in the impaired TD lesion, however when the lesion begins to heal, it strongly express MGP prior to its calcification. Moreover, we show that when calcification is inhibited, a gap is formed between the expression zones of MGP and BMP2 and that this gap is closed during the healing process. To conclude, we suggest that MGP, directly or through interaction with BMP2, plays a role as ossification regulator, rather then simple inhibitor that acts prior to ossification.

  13. Time domain optical coherence tomography investigation of bone matrix interface in rat femurs

    Science.gov (United States)

    Rusu, Laura-Cristina; Negruá¹±iu, Meda-Lavinia; Sinescu, Cosmin; Hoinoiu, Bogdan; Topala, Florin-Ionel; Duma, Virgil-Florin; Rominu, Mihai; Podoleanu, Adrian G.

    2013-08-01

    The materials used to fabricate scaffolds for tissue engineering are derived from synthetic polymers, mainly from the polyester family, or from natural materials (e.g., collagen and chitosan). The mechanical properties and the structural properties of these materials can be tailored by adjusting the molecular weight, the crystalline state, and the ratio of monomers in the copolymers. Quality control and adjustment of the scaffold manufacturing process are essential to achieve high standard scaffolds. Most scaffolds are made from highly crystalline polymers, which inevitably result in their opaque appearance. Their 3-D opaque structure prevents the observation of internal uneven surface structures of the scaffolds under normal optical instruments, such as the traditional light microscope. The inability to easily monitor the inner structure of scaffolds as well as the interface with the old bone poses a major challenge for tissue engineering: it impedes the precise control and adjustment of the parameters that affect the cell growth in response to various mimicked culture conditions. The aim of this paper is to investigate the interface between the femur rat bone and the new bone that is obtained using a method of tissue engineering that is based on different artificial matrixes inserted in previously artificially induced defects. For this study, 15 rats were used in conformity with ethical procedures. In all the femurs a round defect was induced by drilling with a 1 mm spherical Co-Cr surgical drill. The matrixes used were Bioss and 4bone. These materials were inserted into the induced defects. The femurs were investigated at 1 week, 1 month, 2 month and three month after the surgical procedures. The interfaces were examined using Time Domain (TD) Optical Coherence Tomography (OCT) combined with Confocal Microscopy (CM). The optical configuration uses two single mode directional couplers with a superluminiscent diode as the source centered at 1300 nm. The scanning

  14. Implications of combined ovariectomy and glucocorticoid (dexamethasone) treatment on mineral, microarchitectural, biomechanical and matrix properties of rat bone.

    Science.gov (United States)

    Govindarajan, Parameswari; Khassawna, Thaqif; Kampschulte, Marian; Böcker, Wolfgang; Huerter, Britta; Dürselen, Lutz; Faulenbach, Miriam; Heiss, Christian

    2013-12-01

    Osteoporosis is one of the deleterious side effects of long-term glucocorticoid therapy. Since the condition is particularly aggressive in postmenopausal women who are on steroid therapy, in this study we have attempted to analyse the combined effect of glucocorticoid (dexamethasone) treatment and cessation of oestrogen on rat bone. The dual aim was to generate osteoporotic bone status in a short time scale and to characterise the combination of glucocorticoid-postmenopausal osteoporotic conditions. Sprague Dawley rats (N = 42) were grouped randomly into three groups: untreated control, sham-operated and ovariectomized-steroid (OVX-Steroid) rats. Control animals were euthanized with no treatment [Month 0 (M0)], while sham and OVX-Steroid rats were monitored up to 1 month (M1) and 3 months (M3) post laparotomy/post OVX-Steroid treatment. Histology, dual-energy X-ray absorptiometry (DXA), micro-computed tomography (micro-CT), and biomechanical and mRNA expression analysis of collagenous, non-collagenous matrix proteins and osteoclast markers were examined. The study indicated enhanced osteoclastogenesis and significantly lower bone mineral density (BMD) in the OVX-Steroid rats with Z-scores below -2.5, reduced torsional strength, reduced bone volume (BV/TV%), significantly enhanced trabecular separation (Tb.S), and less trabecular number (Tb.N) compared with sham rats. Osteoclast markers, cathepsin K and MMP 9 were upregulated along with Col1α1 and biglycan with no significant expression variation in fibronectin, MMP 14, LRP-5, Car II and TNC. These results show higher bone turnover with enhanced bone resorption accompanied with reduced torsional strength in OVX-Steroid rats; and these changes were attained within a short timeframe. This could be a useful model which mimics human postmenopausal osteoporosis that is associated with steroid therapy and could prove of value both in disease diagnosis and for testing generating and testing biological agents which could

  15. Great increase in bone 66 kDa protein and osteocalcin at later stages with healing rat fractures: effect of zinc treatment.

    Science.gov (United States)

    Igarashi, Aki; Yamaguchi, Masayoshi

    2003-02-01

    Fracture healing has been demonstrated to increase production of bone growth factors, and this elevation has been shown to be enhanced by zinc treatment. Moreover, the effect of zinc treatment on production of bone osteocalcin, which is a kind of Ca2+-binding protein localized in bone matrix, at the later stages with bone fracture was investigated. Rats were sacrificed 7 (earlier stage) or 21 (later stage) days after fracture of femoral diaphysis. Femoral-diaphyseal tissues with fracture healing were cultured in a serum-free medium for 24 h. Many proteins in the bone tissues were released into the medium. Bone protein production was markedly elevated 21 days after bone fracture as compared with that of 7 days. A approximately 66 kDa protein molecule, a major protein component which was produced by the diaphyseal tissues during fracture healing, was predominantly increased at the later stages with fracture healing. Bone osteocalcin production was significantly increased during fracture healing. This increase was enhanced at the later stages with fracture healing. The presence of zinc acexamate (10(-4) M) in culture medium caused a significant increase in bone protein and osteocalcin production at 7 or 21 days after bone fracture. The effect of zinc acexamate in increasing bone total protein and osteocalcin production was remarkable at the later stages with fracture healing. Moreover, zinc treatment caused a significant increase in alkaline phosphatase activity, deoxyribonucleic acid (DNA) and calcium content in the femoral-diaphyseal tissues of the later stages with fracture healing in vitro. The present study demonstrates that bone protein production is markedly increased at the later stages with fracture healing, and that zinc treatment can enhance production of bone protein components including osteocalcin in vitro. Zinc treatment may stimulate the healing of femoral fracture at earlier and later stages.

  16. Carbon nanotube interaction with extracellular matrix proteins producing scaffolds for tissue engineering

    Directory of Open Access Journals (Sweden)

    Tonelli FM

    2012-08-01

    Full Text Available Fernanda MP Tonelli,1 Anderson K Santos,1 Katia N Gomes,2 Eudes Lorençon,2 Silvia Guatimosim,3 Luiz O Ladeira,2 Rodrigo R Resende11Cell Signaling and Nanobiotechnology Laboratory, Department of Biochemistry and Immunology, Federal University of Minas Gerais, Belo Horizonte, Brazil; 2Nanomaterials Laboratory, Department of Physics, Federal University of Minas Gerais, Belo Horizonte, Brazil; 3Intracellular Cardiomiocyte Signaling Laboratory, Department of Physiology and Biophysics, Federal University of Minas Gerais, Belo Horizonte, BrazilAbstract: In recent years, significant progress has been made in organ transplantation, surgical reconstruction, and the use of artificial prostheses to treat the loss or failure of an organ or bone tissue. In recent years, considerable attention has been given to carbon nanotubes and collagen composite materials and their applications in the field of tissue engineering due to their minimal foreign-body reactions, an intrinsic antibacterial nature, biocompatibility, biodegradability, and the ability to be molded into various geometries and forms such as porous structures, suitable for cell ingrowth, proliferation, and differentiation. Recently, grafted collagen and some other natural and synthetic polymers with carbon nanotubes have been incorporated to increase the mechanical strength of these composites. Carbon nanotube composites are thus emerging as potential materials for artificial bone and bone regeneration in tissue engineering.Keywords: carbon nanotubes, tissue engineering, extracellular matrix proteins, collagen, hyaluronic acid, stem cells

  17. Physical distribution and characteristics of meat & bone meal protein

    Science.gov (United States)

    Meat & bone meal (MBM) is a high-protein commodity produced by the rendering of fat from unmarketable animal tissue. Concerns related to bovine spongiform encephalopathy have progressively restricted MBM’s conventional use as a feed ingredient. Consequently, significant attention has focused on th...

  18. Estrogen modulates the mRNA levels for cancellous bone protein of ovariectomized rats.

    Science.gov (United States)

    Salih, M A; Liu, C C; Arjmandi, B H; Kalu, D N

    1993-12-01

    This study was undertaken to examine the effects of ovariectomy and 17 beta-estradiol (E2) on the gene expression of type 1 collagen, osteocalcin and the protooncogen, c-myc, in cancellous bone. Female Sprague-Dawley rats, aged 95 days, were divided into 4 groups. Group 1 was sham operated and Groups 2-4 were ovariectomized. Groups 3 and 4 received daily injections of 160 ng and 1600 ng E2/kg body weight, respectively. Groups 1 and 2 received the solvent vehicle. All animals were sacrificed after 14 days. The femurs were dissected out and cancellous bone scraped from the distal metaphysis. RNA was isolated from the cancellous bone, immobilized on filters or size-fractionated by agarose gel electrophoresis and adsorbed on filters which were then hybridized with specific cDNA probes. Ovariectomy resulted in a significant increase in the mRNAs of type 1 collagen, osteocalcin and c-myc. The increase was suppressed in animals that received 17 beta-estradiol injections. In addition, ovariectomy caused the expected decrease in cancellous bone in the proximal tibia and increased osteoclast and osteoblast numbers. The ovariectomy-induced changes were prevented by 17 beta-estradiol administration. These findings suggest that the lack of ovarian hormones shortly after ovariectomy up-regulates and estrogen administration down-regulates the expression of important cancellous bone matrix proteins as well as the protooncogen, c-myc.

  19. The density of collagen fiber in alveolus mandibular bone of rabbit after augmentation with powder demineralized bone matrix post incisivus extraction

    Directory of Open Access Journals (Sweden)

    Regina TC. Tandelilin

    2006-06-01

    Full Text Available The bone defect due to tooth extraction contributes the most cases reported in the aspects of oral surgery. The defect can be preventively managed by adding powder bone matrix intended for augmentation which eventually induces the formation of new bones. This hard tissue wound healing is preceded by the presence of collagen fibers. The aim of this study was to determine the density of collagen fiber in the alveolus mandibular bone of rabbit which was augmented using powder demineralized bone matrix (DBM post incisivus extraction. Twenty four male rabbits aged 2.5–3 months weighed 900–1,100 grams were randomly divided into two groups. The treated rabbits were augmented with DBM after the incisivus extraction on mandible. The mucosa was then sutured. On the other hand, the controlled rabbits received similar treatments with those of the treated rabbits except there was no augmentation of DBM. Decapitation of treated and controlled rabbits was made on day 5, 7, 10, and 14 days post surgery, each with three rabbits. Mandibles were cut, decalcified, and imbedded in paraffin block. The staining was done using Mallory. Significant differences in the density of collagen were noted on day 10 and 14 post surgery, indicating that powder demineralized bone matrix successfully induced the stimulation of collagen.

  20. Expression of genes encoding extracellular matrix proteins: a macroarray study.

    Science.gov (United States)

    Futyma, Konrad; Miotła, Paweł; Różyńska, Krystyna; Zdunek, Małgorzata; Semczuk, Andrzej; Rechberger, Tomasz; Wojcierowski, Jacek

    2014-12-01

    Endometrial cancer (EC) is one of the most common gynecological malignancies in Poland, with well-established risk factors. Genetic instability and molecular alterations responsible for endometrial carcinogenesis have been systematically investigated. The aim of the present study was to investigate, by means of cDNA macroarrays, the expression profiles of genes encoding extracellular matrix (ECM) proteins in ECs. Tissue specimens were collected during surgical procedures from 40 patients with EC, and control tissue was collected from 9 patients with uterine leiomyomas. RNA was isolated and RT-PCR with radioisotope-labeled cDNA was performed. The levels of ECM protein gene expression in normal endometrial tissues were compared to the expression of these genes in EC specimens. Statistically significant differences in gene expression, stratified by clinical stage of the ECs, were detected for aggrecan, vitronectin, tenascin R, nidogen and two collagen proteins: type VIII chain α1 and type XI chain α2. All of these proteins were overexpressed in stage III endometrial carcinomas compared to levels in stage I and II uterine neoplasms. In conclusion, increased expression of genes encoding ECM proteins may play an important role in facilitating accelerated disease progression of human ECs.

  1. Aberrant expression of nuclear matrix proteins during HMBA-induced differentiation of gastric cancer cells

    Institute of Scientific and Technical Information of China (English)

    2010-01-01

    AIM: To investigate the aberrant expression of nuclear matrix proteins in human gastric cancer cells before and after hexamethylene bisacetamide (HMBA) treatment.METHODS: Proteomics analysis of differential nuclear matrix proteins was performed by two dimensional electrophoresis polyacrylamide gel electrophoresis and matrix-assisted laser desorption/ionization time-of-flight mass spectrometry.The expression levels of three nuclear matrix proteins were further confirmed by Western blotting and their location...

  2. In vitro cartilage tissue engineering using cancellous bone matrix gelatin as a biodegradable scaffold.

    Science.gov (United States)

    Yang, Bo; Yin, Zhanhai; Cao, Junling; Shi, Zhongli; Zhang, Zengtie; Song, Hongxing; Liu, Fuqiang; Caterson, Bruce

    2010-08-01

    In this study, we constructed tissue-engineered cartilage using allogeneic cancellous bone matrix gelatin (BMG) as a scaffold. Allogeneic BMG was prepared by sequential defatting, demineralization and denaturation. Isolated rabbit chondrocytes were seeded onto allogeneic cancellous BMG, and cell-BMG constructs were harvested after 1, 3 and 6 weeks for evaluation by hematoxylin and eosin staining for overall morphology, toluidine blue for extracellular matrix (ECM) proteoglycans, immunohistochemical staining for collagen type II and a transmission electron microscope for examining cellular microstructure on BMG. The prepared BMG was highly porous with mechanical strength adjustable by duration of demineralization and was easily trimmed for tissue repair. Cancellous BMG showed favorable porosity for cell habitation and metabolism material exchange with larger pore sizes (100-500 microm) than in cortical BMG (5-15 microm), allowing cell penetration. Cancellous BMG also showed good biocompatibility, which supported chondrocyte proliferation and sustained their differentiated phenotype in culture for up to 6 weeks. Rich and evenly distributed cartilage ECM proteoglycans and collagen type II were observed around chondrocytes on the surface and inside the pores throughout the cancellous BMG. Considering the large supply of banked bone allografts and relatively convenient preparation, our study suggests that allogeneic cancellous BMG is a promising scaffold for cartilage tissue engineering.

  3. In vitro cartilage tissue engineering using cancellous bone matrix gelatin as a biodegradable scaffold

    Energy Technology Data Exchange (ETDEWEB)

    Yang Bo; Yin Zhanhai; Cao Junling; Shi Zhongli; Zhang Zengtie; Liu Fuqiang [College of Medicine, Xi' an Jiaotong University, Yanta West Road, No 76, Yanta District, Xi' an, Shaanxi Province 710061 (China); Song Hongxing [Department of Orthopedics, Xuanwu Hospital, Capital Medical University, Beijing 100053 (China); Caterson, Bruce, E-mail: caojl@mail.xjtu.edu.c [Connective Tissue Biology Laboratories, Cardiff School of Biosciences, Cardiff University, Biomedical Building, Museum Avenue, Cardiff, CF10 3US (United Kingdom)

    2010-08-01

    In this study, we constructed tissue-engineered cartilage using allogeneic cancellous bone matrix gelatin (BMG) as a scaffold. Allogeneic BMG was prepared by sequential defatting, demineralization and denaturation. Isolated rabbit chondrocytes were seeded onto allogeneic cancellous BMG, and cell-BMG constructs were harvested after 1, 3 and 6 weeks for evaluation by hematoxylin and eosin staining for overall morphology, toluidine blue for extracellular matrix (ECM) proteoglycans, immunohistochemical staining for collagen type II and a transmission electron microscope for examining cellular microstructure on BMG. The prepared BMG was highly porous with mechanical strength adjustable by duration of demineralization and was easily trimmed for tissue repair. Cancellous BMG showed favorable porosity for cell habitation and metabolism material exchange with larger pore sizes (100-500 {mu}m) than in cortical BMG (5-15 {mu}m), allowing cell penetration. Cancellous BMG also showed good biocompatibility, which supported chondrocyte proliferation and sustained their differentiated phenotype in culture for up to 6 weeks. Rich and evenly distributed cartilage ECM proteoglycans and collagen type II were observed around chondrocytes on the surface and inside the pores throughout the cancellous BMG. Considering the large supply of banked bone allografts and relatively convenient preparation, our study suggests that allogeneic cancellous BMG is a promising scaffold for cartilage tissue engineering.

  4. Demineralized bone matrix as a bone tissue engineering scaffold material%骨组织工程支架材料:脱矿骨基质

    Institute of Scientific and Technical Information of China (English)

    陈海霞; 谢志刚

    2014-01-01

    BACKGROUND:Demineralized bone matrix as bone tissue engineering biological scaffold material is more researched currently, which has osteoinductive and osteoconductive. OBTECTIVE:To summarize the development of demineralized bone matrix as bone tissue engineering scaffold material and to prospect its trend. METHODS:The relative literatures addressing demineralized bone matrix as bone tissue engineering scaffold material published between January 1965 and May 2013 in PubMed database, Chinese Biomedical Database, Wanfang Database and FMJS database were searched by the first author. The key words were“demineralized bone matrix, scaffold material, growth factor, cells, drugs”in English and Chinese. According to the inclusion criteria, repetitive researches were excluded, and final y 34 articles were included. RESULTS AND CONCLUSION:Scaffold material is the key composition of tissue engineered bone, and only demineralized bone matrix has both osteoinductive and osteoconductive properties which cannot only provide space for bone repair, but also can be combined with bioactive factors, living cells, antibiotics in vitro to construct bone graft thereby promoting healing of bone defects. However, this technique is also facing some problems to be solved such as the ratio of demineralized bone matrix and various substances, disinfection, preservation of osteogenetic activity and antigenicity elimination. Sufficiently understanding demineralized bone matrix as bone tissue engineering scaffold can provide a theoretical basis for its clinical service.%背景:脱矿骨基质是目前研究较多的具备骨诱导及骨引导的生物支架材料之一。目的:总结脱矿骨基质作为骨组织工程支架材料的研究进展,并展望其发展趋势。  方法:由第一作者检索1965年1月至2013年5月PubMed数据库、中国生物医学数据库、万方数据库及FMJS数据库有关脱矿骨基质及其作为骨组织工程支架材料的相关文

  5. Enhanced Bone Morphogenetic Protein-2-Induced Ectopic and Orthotopic Bone Formation by Intermittent Parathyroid Hormone (1-34) Administration

    NARCIS (Netherlands)

    Kempen, Diederik H. R.; Lu, Lichun; Hefferan, Theresa E.; Creemers, Laura B.; Heijink, Andras; Maran, Avudaiappan; Dhert, Wouter J. A.; Yaszemski, Michael J.

    2010-01-01

    Bone morphogenetic proteins (BMPs) play a central role in local bone regeneration strategies, whereas the anabolic features of parathyroid hormone (PTH) are particularly appealing for the systemic treatment of generalized bone loss. The aim of the current study was to investigate whether local BMP-2

  6. A comprehensive clinical review of recombinant human bone morphogenetic protein-2 (INFUSE® Bone Graft)

    Science.gov (United States)

    Peckham, Steven M.; Badura, Jeffrey M.

    2007-01-01

    The combination of recombinant human bone morphogenetic protein-2 (rhBMP-2) on an absorbable collagen sponge (ACS) carrier has been shown to induce bone formation in a number of preclinical and clinical investigations. In 2002, rhBMP-2/ACS at a 1.5-mg/cc concentration (INFUSE® Bone Graft, Medtronic Spinal and Biologics, Memphis, TN) was FDA-approved as an autograft replacement for certain interbody spinal fusion procedures. In 2004, INFUSE® Bone Graft was approved for open tibial fractures with an intermedullary (IM) nail fixation. Most recently, in March 2007, INFUSE® Bone Graft was approved as an alternative to autogenous bone grafts for sinus augmentations, and for localised alveolar ridge augmentations for defects associated with extraction sockets. The culmination of extensive preclinical and clinical research and three FDA approvals makes rhBMP-2 one of the most studied, published and significant advances in orthopaedics. This review article summarises a number of clinical findings of rhBMP-2/ACS, including the FDA-approved investigational device exemption (IDE) studies used in gaining the aforementioned approvals. PMID:17639384

  7. Effect of the "protein diet" and bone tissue

    Directory of Open Access Journals (Sweden)

    Zoraide Nascimento da Silva

    2014-01-01

    Full Text Available The aim of this study is to evaluate the effect of the hyperproteic diet consumption on bone tissue. Methods: The study was conducted during sixty days. Twenty eight Wistar albinus rats, adults, originated from Laboratory of Experimental Nutrition were divided in four groups: (n = 7; Control 1 (C1, Control 2 (C2, Hyperproteic 1 (HP1 e Hyperproteic 2 (HP2. The C2 and HP2 groups were submitted to 30% of food restriction. The hyperproteic diet was based on the Atkins diet and prepared to simulate the protein diet. At the end of the study the animals were anesthetized to performer bone densitometry analyses by DEXA and blood and tissue collection. Serum and bone minerals analyses were conducted by colorimetric methods in automated equipment. Results: The total bone mineral density (BMD of the pelvis and the spine of the food restriction groups (HP2 e C2 were lower (p < 0.05 than C1 e HP1 groups. While the femur BMD of the HP2 was lower (p < 0.05 related to others groups. It had been observed reduction (p < 0.05 in the medium point of the width of femur diaphysis and in bone calcium level in the hyperproteic groups (HP1 e HP2. It was observed similar effect on the osteocalcin level, that presented lower (p < 0.05 in the hyperproteic groups. The insulin level was lower only in HP2 and serum calcium of the HP1 and HP2 groups was lower than C1. Conclusion: The protein diet promotes significant bone change on femur and in the hormones levels related to bone synthesis and maintenance of this tissue.

  8. Ridge preservation with acellular dermal matrix and anorganic bone matrix cell-binding peptide P-15 after tooth extraction in humans. A histologic and morphometric study

    Directory of Open Access Journals (Sweden)

    Arthur B. Novaes Jr

    2012-06-01

    Full Text Available Aim: The aim of this study was to analyze by histomorphometric parameters the use of acellular dermal matrix (ADM with or without anorganic bovine bone matrix (ABM / synthetic cell-binding peptide P-15 in the formation of bone in human alveoli. Materials and methods: Eighteen patients in need of extraction of maxillary anterior teeth were selected and randomly assigned to the test group (ADM plus ABM/P-15 or the control group (ADM only. Histomorphometric measurements and histological analysis were recorded about 6 months after ridge preservation procedures in ten patients. The amount of newly formed bone, the most recently formed bone, fibrous tissue plus marrow spaces and remaining graft particles were measured and analyzed. Results: At 6 months, the new bone area parameter and the percentage of fibrous tissue plus marrow space areas showed higher values to the control group, and statistically significant differences when compared with the test group (p=0.03. Conclusion: The ADM acted as a membrane. The association of ABM/P-15 with ADM resulted in new bone formation within the alveoli, but the results were not considered relevant when used in this indication.

  9. Platelet activation by extracellular matrix proteins in haemostasis and thrombosis.

    Science.gov (United States)

    Watson, Steve P

    2009-01-01

    The prevention of excessive blood loss to avoid fatal haemorrhage is a pivotal process for all organisms possessing a circulatory system. Increased circulating blood volume and pressure, as required in larger animals, make this process all the more important and challenging. It is essential to have a powerful and rapid system to detect damage and generate an effective seal, and which is also exquisitely regulated to prevent unwanted, excessive or systemic activation so as to avoid blockage of vessels. Thus, a highly specialised and efficient haemostatic system has evolved that consists of cellular (platelets) and protein (coagulation factors) components. Importantly, this is able to support haemostasis in both the low shear environment of the venous system and the high shear environment of the arterial system. Endothelial cells, lining the entire circulation system, play a crucial role in the delicate balance between activation and inhibition of the haemostatic system. An intact and healthy endothelium supports blood flow by preventing attachment of cells and proteins which is required for initiation of coagulation and platelet activation. Endothelial cells produce and release the two powerful soluble inhibitors of platelet activation, nitric oxide and prostacyclin, and express high levels of CD39 which rapidly metabolises the major platelet feedback agonist, ADP. This antithrombotic environment however can rapidly change following activation or removal of endothelial cells through injury or rupture of atherosclerotic plaques. Loss of endothelial cells exposes the subendothelial extracellular matrix which creates strong signals for activation of the haemostatic system including powerful platelet adhesion and activation. Quantitative and qualitative changes in the composition of the subendothelial extracellular matrix influence these prothrombotic characteristics with life threatening thrombotic and bleeding complications, as illustrated by formation of

  10. Human breast cancer cell-mediated bone collagen degradation requires plasminogen activation and matrix metalloproteinase activity

    Directory of Open Access Journals (Sweden)

    Hill Peter A

    2005-02-01

    Full Text Available Abstract Background Breast cancer cells frequently metastasize to the skeleton and induce extensive bone destruction. Cancer cells produce proteinases, including matrix metalloproteinases (MMPs and the plasminogen activator system (PAS which promote invasion of extracellular matrices, but whether these proteinases degrade bone matrix is unclear. To characterize the role that breast cancer cell proteinases play in bone degradation we compared the effects of three human breast cancer cell lines, MDA-MB-231, ZR-75-1 and MCF-7 with those of a normal breast epithelial cell line, HME. The cell lines were cultured atop radiolabelled matrices of either mineralized or non-mineralized bone or type I collagen, the principal organic constituent of bone. Results The 3 breast cancer cell lines all produced significant degradation of the 3 collagenous extracellular matrices (ECMs whilst the normal breast cell line was without effect. Breast cancer cells displayed an absolute requirement for serum to dissolve collagen. Degradation of collagen was abolished in plasminogen-depleted serum and could be restored by the addition of exogenous plasminogen. Localization of plasmin activity to the cell surface was critical for the degradation process as aprotinin, but not α2 antiplasmin, prevented collagen dissolution. During ECM degradation breast cancer cell lines expressed urokinase-type plasminogen activator (u-PA and uPA receptor, and MMPs-1, -3, -9,-13, and -14. The normal breast epithelial cell line expressed low levels of MMPs-1, and -3, uPA and uPA receptor. Inhibitors of both the PAS (aprotinin and PA inhibitor-1 and MMPs (CT1166 and tisue inhibitor of metalloproteinase blocked collagen degradation, demonstrating the requirement of both plasminogen activation and MMP activity for degradation. The activation of MMP-13 in human breast cancer cells was prevented by plasminogen activator inhibitor-1 but not by tissue inhibitor of metalloproteinase-1, suggesting

  11. Bone morphogenetic proteins: from structure to clinical use

    Directory of Open Access Journals (Sweden)

    Granjeiro J.M.

    2005-01-01

    Full Text Available Bone morphogenetic proteins (BMPs are multi-functional growth factors belonging to the transforming growth factor ß superfamily. Family members are expressed during limb development, endochondral ossification, early fracture, and cartilage repair. The activity of BMPs was first identified in the 1960s but the proteins responsible for bone induction were unknown until the purification and cloning of human BMPs in the 1980s. To date, about 15 BMP family members have been identified and characterized. The signal triggered by BMPs is transduced through serine/threonine kinase receptors, type I and II subtypes. Three type I receptors have been shown to bind BMP ligands, namely: type IA and IB BMP receptors and type IA activin receptors. BMPs seem to be involved in the regulation of cell proliferation, survival, differentiation and apoptosis, but their hallmark is their ability to induce bone, cartilage, ligament, and tendon formation at both heterotopic and orthotopic sites. This suggests that, in the future, they may play a major role in the treatment of bone diseases. Several animal studies have illustrated the potential of BMPs to enhance spinal fusion, repair critical-size defects, accelerate union, and heal articular cartilage lesions. Difficulties in producing and purifying BMPs from bone tissue have prompted the attempts made by several laboratories, including ours, to express these proteins in the recombinant form in heterologous systems. This review focuses on BMP structure, molecular mechanisms of action and significance and potential applications in medical, dental and veterinary practice for the treatment of cartilage and bone-related diseases.

  12. Comparative experiment of four different materials as carriers of Bone morphogenetic protein to repair long bone defect

    Institute of Scientific and Technical Information of China (English)

    WEI Kuan-hai; PEI Guo-xian; YANG Run-gong

    2001-01-01

    @@ OBJECTIVE To investigate the effects of four different materials as carriers of bone morphogenetic protein (BMP) to repair long bone defect. METHODS 12 mm radius bone defects were made. They were divided into 4 groups in random and repaired respectively with the vascular muscle flap combined with FS/BMP (group A), vascular muscle flap/BMP (group B), bloodless muscle flap/BMP (group C) and autolyzed antigen-extracted allogeneic bone (AAA)/BMP (group D).Their abilities of bone forming to repair bone defects were observed.

  13. Influence of mineral phase in mineralization of a biocomposite containing chitosan, demineralized bone matrix and bone ash—in vitro study

    Indian Academy of Sciences (India)

    Krithiga Gunasekaran; Santhosh Kumar Baskar; Divya Sapphire Mohan; Thotapalli P Sastry

    2014-05-01

    A resorbable composite which acts as a active barrier in guided bone regeneration was fabricated using chitosan, demineralized bone matrix and bone ash. Its potential to form bone like apatite in simulated body fluid was assessed in this study. The mechanical strength of these composites was correlated with bone ash ratios and composites with better tensile strength were studied for their acellular bioactivity by incubating in simulated body fluid for 21 days. Composites without bone ash did not show acellular bioactivity which was confirmed by thermogravimetric analysis. In case of biocomposites with bone ash, there was an increase in residual weight indicating the mineralization of the composite. The composite containing bone ash has shown the peaks related to phosphate vibrations in its Fourier-transform infrared spectrum. Scanning micrographs revealed formation of apatite like crystals on its surface. Ca/P ratio was found to be 1.7 which is nearer to that of natural bone. Thus, prepared composites can be used as resorbable biocomposite in maxillofacial and oral defects.

  14. Promotion of hepatic differentiation of bone marrow mesenchymal stem cells on decellularized cell-deposited extracellular matrix.

    Science.gov (United States)

    He, Hongliang; Liu, Xiaozhen; Peng, Liang; Gao, Zhiliang; Ye, Yun; Su, Yujie; Zhao, Qiyi; Wang, Ke; Gong, Yihong; He, Fan

    2013-01-01

    Interactions between stem cells and extracellular matrix (ECM) are requisite for inducing lineage-specific differentiation and maintaining biological functions of mesenchymal stem cells by providing a composite set of chemical and structural signals. Here we investigated if cell-deposited ECM mimicked in vivo liver's stem cell microenvironment and facilitated hepatogenic maturation. Decellularization process preserved the fibrillar microstructure and a mix of matrix proteins in cell-deposited ECM, such as type I collagen, type III collagen, fibronectin, and laminin that were identical to those found in native liver. Compared with the cells on tissue culture polystyrene (TCPS), bone marrow mesenchymal stem cells (BM-MSCs) cultured on cell-deposited ECM showed a spindle-like shape, a robust proliferative capacity, and a suppressed level of intracellular reactive oxygen species, accompanied with upregulation of two superoxide dismutases. Hepatocyte-like cells differentiated from BM-MSCs on ECM were determined with a more intensive staining of glycogen storage, an elevated level of urea biosynthesis, and higher expressions of hepatocyte-specific genes in contrast to those on TCPS. These results demonstrate that cell-deposited ECM can be an effective method to facilitate hepatic maturation of BM-MSCs and promote stem-cell-based liver regenerative medicine.

  15. Promotion of Hepatic Differentiation of Bone Marrow Mesenchymal Stem Cells on Decellularized Cell-Deposited Extracellular Matrix

    Directory of Open Access Journals (Sweden)

    Hongliang He

    2013-01-01

    Full Text Available Interactions between stem cells and extracellular matrix (ECM are requisite for inducing lineage-specific differentiation and maintaining biological functions of mesenchymal stem cells by providing a composite set of chemical and structural signals. Here we investigated if cell-deposited ECM mimicked in vivo liver's stem cell microenvironment and facilitated hepatogenic maturation. Decellularization process preserved the fibrillar microstructure and a mix of matrix proteins in cell-deposited ECM, such as type I collagen, type III collagen, fibronectin, and laminin that were identical to those found in native liver. Compared with the cells on tissue culture polystyrene (TCPS, bone marrow mesenchymal stem cells (BM-MSCs cultured on cell-deposited ECM showed a spindle-like shape, a robust proliferative capacity, and a suppressed level of intracellular reactive oxygen species, accompanied with upregulation of two superoxide dismutases. Hepatocyte-like cells differentiated from BM-MSCs on ECM were determined with a more intensive staining of glycogen storage, an elevated level of urea biosynthesis, and higher expressions of hepatocyte-specific genes in contrast to those on TCPS. These results demonstrate that cell-deposited ECM can be an effective method to facilitate hepatic maturation of BM-MSCs and promote stem-cell-based liver regenerative medicine.

  16. PROTEIN OF MEAT AND BONE MEAL FOR PIGS

    Directory of Open Access Journals (Sweden)

    Patieva S. V.

    2015-09-01

    Full Text Available The modern requirements of intergovernmental standards to the quality and safety of livestock produce provide for the use of highly productive animals capable under small expenses to produce more the high quality produce. In particular, at the formation of meat productivity at pigs the great significance has an achievement of optimal digestion and assimilability of consumed fodder means. In the connection, the study of digestion of meat and bone meal from slaughterhouse wastes of cattle (MCM and poultry (MCBM presents the scientific interest. In the fodder experience on the growing pigs with the fistula of iliac intestines there was investigated the digestion of two types of meat and bone meal from slaughterhouse wastes of cattle (MCM and poultry (MKBM. The iliac accessibility of amino acids of meat and bone meal found itself too low: 49,3 % - 69,3 %. The accessibility of general protein reliably did not differ from the average accessibility on main amino acids - 61,5 %. To count the real iliac accessibility of raw protein and amino acids of meat and bone meal there was determined an endogenous emission of these substances on the casein diet. The real iliac accessibility of protein and individual amino acids did not leave the limits in 73% on МCM and 69% - on МCBМ. The accessibility of lysine, leucine and isoleucine MCBM is reliably higher than the same in MCM (P

  17. An Autologous Bone Marrow Mesenchymal Stem Cell–Derived Extracellular Matrix Scaffold Applied with Bone Marrow Stimulation for Cartilage Repair

    Science.gov (United States)

    Tang, Cheng; Jin, Chengzhe; Du, Xiaotao; Yan, Chao; Min, Byoung-Hyun; Xu, Yan

    2014-01-01

    Purpose: It is well known that implanting a bioactive scaffold into a cartilage defect site can enhance cartilage repair after bone marrow stimulation (BMS). However, most of the current scaffolds are derived from xenogenous tissue and/or artificial polymers. The implantation of these scaffolds adds risks of pathogen transmission, undesirable inflammation, and other immunological reactions, as well as ethical issues in clinical practice. The current study was undertaken to evaluate the effectiveness of implanting autologous bone marrow mesenchymal stem cell–derived extracellular matrix (aBMSC-dECM) scaffolds after BMS for cartilage repair. Methods: Full osteochondral defects were performed on the trochlear groove of both knees in 24 rabbits. One group underwent BMS only in the right knee (the BMS group), and the other group was treated by implantation of the aBMSC-dECM scaffold after BMS in the left knee (the aBMSC-dECM scaffold group). Results: Better repair of cartilage defects was observed in the aBMSC-dECM scaffold group than in the BMS group according to gross observation, histological assessments, immunohistochemistry, and chemical assay. The glycosaminoglycan and DNA content, the distribution of proteoglycan, and the distribution and arrangement of type II and I collagen fibers in the repaired tissue in the aBMSC-dECM scaffold group at 12 weeks after surgery were similar to that surrounding normal hyaline cartilage. Conclusions: Implanting aBMSC-dECM scaffolds can enhance the therapeutic effect of BMS on articular cartilage repair, and this combination treatment is a potential method for successful articular cartilage repair. PMID:24666429

  18. NMR structure of the myristylated feline immunodeficiency virus matrix protein.

    Science.gov (United States)

    Brown, Lola A; Cox, Cassiah; Baptiste, Janae; Summers, Holly; Button, Ryan; Bahlow, Kennedy; Spurrier, Vaughn; Kyser, Jenna; Luttge, Benjamin G; Kuo, Lillian; Freed, Eric O; Summers, Michael F

    2015-04-30

    Membrane targeting by the Gag proteins of the human immunodeficiency viruses (HIV types-1 and -2) is mediated by Gag's N-terminally myristylated matrix (MA) domain and is dependent on cellular phosphatidylinositol-4,5-bisphosphate [PI(4,5)P2]. To determine if other lentiviruses employ a similar membrane targeting mechanism, we initiated studies of the feline immunodeficiency virus (FIV), a widespread feline pathogen with potential utility for development of human therapeutics. Bacterial co-translational myristylation was facilitated by mutation of two amino acids near the amino-terminus of the protein (Q5A/G6S; myrMAQ5A/G6S). These substitutions did not affect virus assembly or release from transfected cells. NMR studies revealed that the myristyl group is buried within a hydrophobic pocket in a manner that is structurally similar to that observed for the myristylated HIV-1 protein. Comparisons with a recent crystal structure of the unmyristylated FIV protein [myr(-)MA] indicate that only small changes in helix orientation are required to accommodate the sequestered myr group. Depletion of PI(4,5)P2 from the plasma membrane of FIV-infected CRFK cells inhibited production of FIV particles, indicating that, like HIV, FIV hijacks the PI(4,5)P2 cellular signaling system to direct intracellular Gag trafficking during virus assembly.

  19. NMR Structure of the Myristylated Feline Immunodeficiency Virus Matrix Protein

    Directory of Open Access Journals (Sweden)

    Lola A. Brown

    2015-04-01

    Full Text Available Membrane targeting by the Gag proteins of the human immunodeficiency viruses (HIV types-1 and -2 is mediated by Gag’s N-terminally myristylated matrix (MA domain and is dependent on cellular phosphatidylinositol-4,5-bisphosphate [PI(4,5P2]. To determine if other lentiviruses employ a similar membrane targeting mechanism, we initiated studies of the feline immunodeficiency virus (FIV, a widespread feline pathogen with potential utility for development of human therapeutics. Bacterial co-translational myristylation was facilitated by mutation of two amino acids near the amino-terminus of the protein (Q5A/G6S; myrMAQ5A/G6S. These substitutions did not affect virus assembly or release from transfected cells. NMR studies revealed that the myristyl group is buried within a hydrophobic pocket in a manner that is structurally similar to that observed for the myristylated HIV-1 protein. Comparisons with a recent crystal structure of the unmyristylated FIV protein [myr(-MA] indicate that only small changes in helix orientation are required to accommodate the sequestered myr group. Depletion of PI(4,5P2 from the plasma membrane of FIV-infected CRFK cells inhibited production of FIV particles, indicating that, like HIV, FIV hijacks the PI(4,5P2 cellular signaling system to direct intracellular Gag trafficking during virus assembly.

  20. Whey Protein Concentrate Hydrolysate Prevents Bone Loss in Ovariectomized Rats.

    Science.gov (United States)

    Kim, Jonggun; Kim, Hyung Kwan; Kim, Saehun; Imm, Ji-Young; Whang, Kwang-Youn

    2015-12-01

    Milk is known as a safe food and contains easily absorbable minerals and proteins, including whey protein, which has demonstrated antiosteoporotic effects on ovariectomized rats. This study evaluated the antiosteoporotic effect of whey protein concentrate hydrolysate (WPCH) digested with fungal protease and whey protein concentrate (WPC). Two experiments were conducted to determine (1) efficacy of WPCH and WPC and (2) dose-dependent impact of WPCH in ovariectomized rats (10 weeks old). In Experiment I, ovariectomized rats (n=45) were allotted into three dietary treatments of 10 g/kg diet of WPC, 10 g/kg diet of WPCH, and a control diet. In Experiment II, ovariectomized rats (n=60) were fed four different diets (0, 10, 20, and 40 g/kg of WPCH). In both experiments, sham-operated rats (n=15) were also fed a control diet containing the same amount of amino acids and minerals as dietary treatments. After 6 weeks, dietary WPCH prevented loss of bone, physical properties, mineral density, and mineral content, and improved breaking strength of femurs, with similar effect to WPC. The bone resorption enzyme activity (tartrate resistance acid phosphatase) in tibia epiphysis decreased in response to WPCH supplementation, while bone formation enzyme activity (alkaline phosphatase) was unaffected by ovariectomy and dietary treatment. Bone properties and strength increased as the dietary WPCH level increased (10 and 20 g/kg), but there was no difference between the 20 and 40 g/kg treatment. WPCH and WPC supplementation ameliorated bone loss induced by ovariectomy in rats.

  1. Novel biocompatible polymeric blends for bone regeneration: Material and matrix design and development

    Science.gov (United States)

    Deng, Meng

    The first part of the work presented in this dissertation is focused on the design and development of novel miscible and biocompatible polyphosphazene-polyester blends as candidate materials for scaffold-based bone tissue engineering applications. Biodegradable polyesters such as poly(lactide-co-glycolide) (PLAGA) are among the most widely used polymeric materials for bone tissue engineering. However, acidic degradation products resulting from the bulk degradation mechanism often lead to catastrophic failure of the structure integrity, and adversely affect biocompatibility both in vitro and in vivo. One promising approach to circumvent these limitations is to blend PLAGA with other macromolecules that can buffer the acidic degradation products with a controlled degradation rate. Biodegradable polyphosphazenes (PPHOS), a new class of biomedical materials, have proved to be superior candidate materials to achieve this objective due to their unique buffering degradation products. A highly practical blending approach was adopted to develop novel biocompatible, miscible blends of these two polymers. In order to achieve this miscibility, a series of amino acid ester, alkoxy, aryloxy, and dipeptide substituted PPHOS were synthesized to promote hydrogen bonding interactions with PLAGA. Five mixed-substituent PPHOS compositions were designed and blended with PLAGA at different weight ratios producing candidate blends via a mutual solvent method. Preliminary characterization identified two specific side groups namely glycylglycine dipeptide and phenylphenoxy that resulted in improved blend miscibility and enhanced in vitro osteocompatibility. These findings led to the synthesis of a mixed-substituent polyphosphazene poly[(glycine ethyl glycinato)1(phenylphenoxy)1phosphazene] (PNGEGPhPh) for blending with PLAGA. Two dipeptide-based blends having weight ratios of PNGEGPhPh to PLAGA namely 25:75 (Matrix1) and 50:50 (Matrix2) were fabricated. Both of the blends were

  2. The effect of bone injury on extracellular matrix vesicle proliferation and mineral formation.

    Science.gov (United States)

    Sela, J; Schwartz, Z; Amir, D; Swain, L D; Boyan, B D

    1992-05-01

    Removal of tibial bone marrow in rats is followed by primary bone formation, resorption and marrow restitution. The first week of healing is characterized by partially calcified trabeculae. After 2 weeks, a higher degree of calcification and partial resorption are observed. The third week is characterized by massive resorption of the trabeculae, which are replaced in the fourth week by new bone marrow tissue. This model was used to study primary calcification. Transmission electron micrographs of the young bone revealed osteoblasts, matrix vesicles and calcified fronts. The different vesicular types were defined as 'empty', 'amorphous', 'crystal', and 'rupture'. The vesicles were studied on days 3, 6, 8, 12, 14, 18, 21, 23 and 28 after injury. The mean diameters of most vesicles ranged between 100.3 and 121.9 nm, and their mean distance from the calcified front was less than 976.6 nm. Vesicular density, calculated as number per 10 m2, increased on the eighth day and decreased from the fourteenth day onwards. Highest diameter values were recorded on the sixth day, and decreased onward. Vesicular distance from the calcified front decreased continuously. Distribution of vesicle number, diameter, and distance in each class showed that numbers of empty and amorphous vesicles decreased and of crystal and rupture increased throughout the experiment. Distances from the calcified front and vesicular diameters varied as follows: 'rupture', 'crystal', amorphous', and 'empty', the 'rupture' type being the closest to the front and of the largest diameter. The results confirm the hypothesis that the cell is responsible for the secretion of electron lucent vesicles that accumulate Ca and Pi to form amorphous calcium phosphate complexes that convert to hydroxyapatite. Crystal growth is followed by membrane rupture.

  3. The role of lysyl oxidase, the extracellular matrix and the pre-metastatic niche in bone metastasis

    Directory of Open Access Journals (Sweden)

    Alison Gartland

    2016-09-01

    Full Text Available Most deaths from solid cancers occur as a result of secondary metastasis to distant sites. Bone is the most frequent metastatic site for many cancer types and can account for up to 80% of cancer-related deaths in certain tumours. The progression from a discrete solid primary tumour to devastating and painful bone metastases is a complex process involving multiple cell types and steps. There is increasing evidence that modulation of the extracellular matrix plays an important role in the lethal transition from a primary to disseminated metastatic bone tumour. This review provides an overview of the current understanding on the role of role of lysyl oxidase, the extracellular matrix and the pre-metastatic niche in bone metastasis

  4. The role of lysyl oxidase, the extracellular matrix and the pre-metastatic niche in bone metastasis

    DEFF Research Database (Denmark)

    Gartland, Alison; Erler, Janine Terra; Cox, Thomas Robert

    2016-01-01

    and painful bone metastases is a complex process involving multiple cell types and steps. There is increasing evidence that modulation of the extracellular matrix plays an important role in the lethal transition from a primary to disseminated metastatic bone tumour. This review provides an overview......Most deaths from solid cancers occur as a result of secondary metastasis to distant sites. Bone is the most frequent metastatic site for many cancer types and can account for up to 80% of cancer-related deaths in certain tumours. The progression from a discrete solid primary tumour to devastating...... of the current understanding on the role of role of lysyl oxidase, the extracellular matrix and the pre-metastatic niche in bone metastasis....

  5. Biological conduits combining bone marrow mesenchymal stem cells and extracellular matrix to treat long-segment sciatic nerve defects

    Institute of Scientific and Technical Information of China (English)

    Yang Wang; Zheng-wei Li; Min Luo; Ya-jun Li; Ke-qiang Zhang

    2015-01-01

    The transplantation of polylactic glycolic acid conduits combining bone marrow mesenchymal stem cells and extracellular matrix gel for the repair of sciatic nerve injury is effective in some re-spects, but few data comparing the biomechanical factors related to the sciatic nerve are available. In the present study, rabbit models of 10-mm sciatic nerve defects were prepared. The rabbit models were repaired with autologous nerve, a polylactic glycolic acid conduit+bone marrow mesenchymal stem cells, or a polylactic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel. After 24 weeks, mechanical testing was performed to determine the stress relaxation and creep parameters. Following sciatic nerve injury, the magnitudes of the stress decrease and strain increase at 7,200 seconds were largest in the polylactic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel group, followed by the polylactic glycolic acid conduit+bone marrow mesenchymal stem cells group, and then the autologous nerve group. Hematoxylin-eosin staining demonstrated that compared with the poly-lactic glycolic acid conduit+bone marrow mesenchymal stem cells group and the autologous nerve group, a more complete sciatic nerve regeneration was found, including good myelination, regularly arranged nerve ifbers, and a completely degraded and resorbed conduit, in the polylac-tic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel group. These results indicate that bridging 10-mm sciatic nerve defects with a polylactic glycolic acid conduit+bone marrow mesenchymal stem cells+extracellular matrix gel construct increases the stress relaxation under a constant strain, reducing anastomotic tension. Large elongations under a constant physiological load can limit the anastomotic opening and shift, which is ben-eifcial for the regeneration and functional reconstruction of sciatic nerve. Better regeneration was found with the

  6. Expression of 16 Nitrogenase Proteins within the Plant Mitochondrial Matrix

    Science.gov (United States)

    Allen, Robert S.; Tilbrook, Kimberley; Warden, Andrew C.; Campbell, Peter C.; Rolland, Vivien; Singh, Surinder P.; Wood, Craig C.

    2017-01-01

    The industrial production and use of nitrogenous fertilizer involves significant environmental and economic costs. Strategies to reduce fertilizer dependency are required to address the world's increasing demand for sustainable food, fibers, and biofuels. Biological nitrogen fixation, a process unique to diazatrophic bacteria, is catalyzed by the nitrogenase complex, and reconstituting this function in plant cells is an ambitious biotechnological strategy to reduce fertilizer use. Here we establish that the full array of biosynthetic and catalytic nitrogenase (Nif) proteins from the diazotroph Klebsiella pneumoniae can be individually expressed as mitochondrial targeting peptide (MTP)-Nif fusions in Nicotiana benthamiana. We show that these are correctly targeted to the plant mitochondrial matrix, a subcellular location with biochemical and genetic characteristics potentially supportive of nitrogenase function. Although Nif proteins B, D, E, F, H, J, K, M, N, Q, S, U, V, X, Y, and Z were all detectable by Western blot analysis, the NifD catalytic component was the least abundant. To address this problem, a translational fusion between NifD and NifK was designed based on the crystal structure of the nitrogenase MoFe protein heterodimer. This fusion protein enabled equimolar NifD:NifK stoichiometry and improved NifD expression levels in plants. Finally, four MTP-Nif fusion proteins (B, S, H, Y) were successfully co-expressed, demonstrating that multiple components of nitrogenase can be targeted to plant mitochondria. These results establish the feasibility of reconstituting the complete componentry for nitrogenase in plant cells, within an intracellular environment that could support the conversion of nitrogen gas into ammonia. PMID:28316608

  7. Expression of 16 Nitrogenase Proteins within the Plant Mitochondrial Matrix.

    Science.gov (United States)

    Allen, Robert S; Tilbrook, Kimberley; Warden, Andrew C; Campbell, Peter C; Rolland, Vivien; Singh, Surinder P; Wood, Craig C

    2017-01-01

    The industrial production and use of nitrogenous fertilizer involves significant environmental and economic costs. Strategies to reduce fertilizer dependency are required to address the world's increasing demand for sustainable food, fibers, and biofuels. Biological nitrogen fixation, a process unique to diazatrophic bacteria, is catalyzed by the nitrogenase complex, and reconstituting this function in plant cells is an ambitious biotechnological strategy to reduce fertilizer use. Here we establish that the full array of biosynthetic and catalytic nitrogenase (Nif) proteins from the diazotroph Klebsiella pneumoniae can be individually expressed as mitochondrial targeting peptide (MTP)-Nif fusions in Nicotiana benthamiana. We show that these are correctly targeted to the plant mitochondrial matrix, a subcellular location with biochemical and genetic characteristics potentially supportive of nitrogenase function. Although Nif proteins B, D, E, F, H, J, K, M, N, Q, S, U, V, X, Y, and Z were all detectable by Western blot analysis, the NifD catalytic component was the least abundant. To address this problem, a translational fusion between NifD and NifK was designed based on the crystal structure of the nitrogenase MoFe protein heterodimer. This fusion protein enabled equimolar NifD:NifK stoichiometry and improved NifD expression levels in plants. Finally, four MTP-Nif fusion proteins (B, S, H, Y) were successfully co-expressed, demonstrating that multiple components of nitrogenase can be targeted to plant mitochondria. These results establish the feasibility of reconstituting the complete componentry for nitrogenase in plant cells, within an intracellular environment that could support the conversion of nitrogen gas into ammonia.

  8. Multi-protein delivery by nanodiamonds promotes bone formation.

    Science.gov (United States)

    Moore, L; Gatica, M; Kim, H; Osawa, E; Ho, D

    2013-11-01

    Bone morphogenetic proteins (BMPs) are well-studied regulators of cartilage and bone development that have been Food and Drug Administration (FDA)-approved for the promotion of bone formation in certain procedures. BMPs are seeing more use in oral and maxillofacial surgeries because of recent FDA approval of InFUSE(®) for sinus augmentation and localized alveolar ridge augmentation. However, the utility of BMPs in medical and dental applications is limited by the delivery method. Currently, BMPs are delivered to the surgical site by the implantation of bulky collagen sponges. Here we evaluate the potential of detonation nanodiamonds (NDs) as a delivery vehicle for BMP-2 and basic fibroblast growth factor (bFGF). Nanodiamonds are biocompatible, 4- to 5-nm carbon nanoparticles that have previously been used to deliver a wide variety of molecules, including proteins and peptides. We find that both BMP-2 and bFGF are readily loaded onto NDs by physisorption, forming a stable colloidal solution, and are triggered to release in slightly acidic conditions. Simultaneous delivery of BMP-2 and bFGF by ND induces differentiation and proliferation in osteoblast progenitor cells. Overall, we find that NDs provide an effective injectable alternative for the delivery of BMP-2 and bFGF to promote bone formation.

  9. Heterotopic bone formation in the musculus latissimus dorsi of sheep using β-tricalcium phosphate scaffolds: evaluation of an extended prefabrication time on bone formation and matrix degeneration.

    Science.gov (United States)

    Spalthoff, S; Jehn, P; Zimmerer, R; Möllmann, U; Gellrich, N-C; Kokemueller, H

    2015-06-01

    We previously generated viable heterotopic bone in living animals and found that 3 months of intrinsic vascularization improved bone formation and matrix degeneration. In this study, we varied the pre-vascularization time to determine its effects on the kinetics of bone formation and ceramic degradation. Two 25-mm-long cylindrical β-tricalcium phosphate scaffolds were filled intraoperatively with autogenous iliac crest bone marrow and implanted in the latissimus dorsi muscle in six sheep. To examine the effect of axial perfusion, one scaffold was surgically implanted with (group C) or without (group D) a central vascular bundle. All animals were sacrificed 6 months postoperatively and histomorphometric measurements were compared to previous results. All implanted scaffolds exhibited ectopic bone growth. However, bone growth was not significantly different between the 3-month (group A, 0.191±0.097 vs. group C, 0.237±0.075; P=0.345) and 6-month (group B, 0.303±0.105 vs. group D, 0.365±0.258; P=0.549) pre-vascularization durations, regardless of vessel supply; early differences between surgically and extrinsically vascularized constructs disappeared after 6 months. Here, we describe a reliable procedure for generating ectopic bone in vivo. A 3-month pre-vascularization duration appears sufficient and ceramic degradation proceeds in accordance with bone generation, supporting the hypothesis of cell-mediated resorption.

  10. Porcine bladder acellular matrix (ACM): protein expression, mechanical properties.

    Science.gov (United States)

    Farhat, Walid A; Chen, Jun; Haig, Jennifer; Antoon, Roula; Litman, Jessica; Sherman, Christopher; Derwin, Kathleen; Yeger, Herman

    2008-06-01

    Experimentally, porcine bladder acellular matrix (ACM) that mimics extracellular matrix has excellent potential as a bladder substitute. Herein we investigated the spatial localization and expression of different key cellular and extracellular proteins in the ACM; furthermore, we evaluated the inherent mechanical properties of the resultant ACM prior to implantation. Using a proprietary decellularization method, the DNA contents in both ACM and normal bladder were measured; in addition we used immunohistochemistry and western blots to quantify and localize the different cellular and extracellular components, and finally the mechanical testing was performed using a uniaxial mechanical testing machine. The mean DNA content in the ACM was significantly lower in the ACM compared to the bladder. Furthermore, the immunohistochemical and western blot analyses showed that collagen I and IV were preserved in the ACM, but possibly denatured collagen III in the ACM. Furthermore, elastin, laminin and fibronectin were mildly reduced in the ACM. Although the ACM did not exhibit nucleated cells, residual cellular components (actin, myosin, vimentin and others) were still present. There was, on the other hand, no significant difference in the mean stiffness between the ACM and the bladder. Although our decellularization method is effective in removing nuclear material from the bladder while maintaining its inherent mechanical properties, further work is mandatory to determine whether these residual DNA and cellular remnants would lead to any immune reaction, or if the mechanical properties of the ACM are preserved upon implantation and cellularization.

  11. Porcine bladder acellular matrix (ACM): protein expression, mechanical properties

    Energy Technology Data Exchange (ETDEWEB)

    Farhat, Walid A [Department of Surgery, Division of Urology, University of Toronto and Hospital for Sick Children, Toronto, ON M5G 1X8 (Canada); Chen Jun; Haig, Jennifer; Antoon, Roula; Litman, Jessica; Yeger, Herman [Department of Developmental and Stem Cell Biology, Research Institute, Hospital for Sick Children, Toronto, ON M5G 1X8 (Canada); Sherman, Christopher [Department of Anatomic Pathology, Sunnybrook and Women' s College Health Sciences Centre, Toronto, ON (Canada); Derwin, Kathleen [Department of Biomedical Engineering, Lerner Research Institute and Orthopaedic Research Center, Cleveland Clinic Foundation, Cleveland, OH (United States)], E-mail: walid.farhat@sickkids.ca

    2008-06-01

    Experimentally, porcine bladder acellular matrix (ACM) that mimics extracellular matrix has excellent potential as a bladder substitute. Herein we investigated the spatial localization and expression of different key cellular and extracellular proteins in the ACM; furthermore, we evaluated the inherent mechanical properties of the resultant ACM prior to implantation. Using a proprietary decellularization method, the DNA contents in both ACM and normal bladder were measured; in addition we used immunohistochemistry and western blots to quantify and localize the different cellular and extracellular components, and finally the mechanical testing was performed using a uniaxial mechanical testing machine. The mean DNA content in the ACM was significantly lower in the ACM compared to the bladder. Furthermore, the immunohistochemical and western blot analyses showed that collagen I and IV were preserved in the ACM, but possibly denatured collagen III in the ACM. Furthermore, elastin, laminin and fibronectin were mildly reduced in the ACM. Although the ACM did not exhibit nucleated cells, residual cellular components (actin, myosin, vimentin and others) were still present. There was, on the other hand, no significant difference in the mean stiffness between the ACM and the bladder. Although our decellularization method is effective in removing nuclear material from the bladder while maintaining its inherent mechanical properties, further work is mandatory to determine whether these residual DNA and cellular remnants would lead to any immune reaction, or if the mechanical properties of the ACM are preserved upon implantation and cellularization.

  12. Preparation of Extracellular Matrix Protein Fibers for Brillouin Spectroscopy.

    Science.gov (United States)

    Edginton, Ryan S; Mattana, Sara; Caponi, Silvia; Fioretto, Daniele; Green, Ellen; Winlove, C Peter; Palombo, Francesca

    2016-09-15

    Brillouin spectroscopy is an emerging technique in the biomedical field. It probes the mechanical properties of a sample through the interaction of visible light with thermally induced acoustic waves or phonons propagating at a speed of a few km/sec. Information on the elasticity and structure of the material is obtained in a nondestructive contactless manner, hence opening the way to in vivo applications and potential diagnosis of pathology. This work describes the application of Brillouin spectroscopy to the study of biomechanics in elastin and trypsin-digested type I collagen fibers of the extracellular matrix. Fibrous proteins of the extracellular matrix are the building blocks of biological tissues and investigating their mechanical and physical behavior is key to establishing structure-function relationships in normal tissues and the changes which occur in disease. The procedures of sample preparation followed by measurement of Brillouin spectra using a reflective substrate are presented together with details of the optical system and methods of spectral data analysis.

  13. Expression of uncarboxylated matrix Gla protein in ankylosing spondylitis and its significance

    Directory of Open Access Journals (Sweden)

    Han-qing HUANG

    2013-07-01

    Full Text Available Objective To investigate the serum level of uncarboxylated matrix Gla protein (ucMGP in ankylosing spondylitis (AS patients, and to evaluate its diagnostic value and the relation of ucMGP to inflammation and ossification process in AS. Methods Eight-two AS patients and 76 healthy controls were enrolled in this randomized controlled study. The clinical indices (age, gender, course of disease, disease activity, changes in radiographic studies, and indices of bone metabolism or inflammation, including erythrocyte sedimentation rate (ESR, C-reactive protein (CRP, osteocalcin (OC, and bone-specific alkaline phosphatase (BALP were evaluated or measured. The disease activity was assessed by Bath Ankylosing Spondylitis Disease Activity Index (BASDAI, and changes in radiographic pictures were evaluated according to the modified Stoke AS Spine Score (mSASSS, and serum level of ucMGP was measured by a competitive ELISA. The relationship between ucMGP and clinical indexes, radiographic scoring, indices in bone metabolism or inflammation was estimated by SPSS software, and the diagnostic value of ucMGP was analyzed by receiver operator characteristic (ROC curve. Results The levels of ESR and CRP in AS patients were higher than those in healthy controls, but the serum ucMGP was lower (2958±654nmol/L compared with healthy controls (4551±1036nmol/L, P0, r=-0.715, P1, r=-0.741, P10, r=-0.776, P<0.01; mSASSS <10, r=-0.297, P=0.028. Conclusion Serum ucMGP may serve as a diagnostic biomarker of AS and progression index of ossification, especially in late stage of AS.

  14. Cartilage oligomeric matrix protein specific antibodies are pathogenic

    DEFF Research Database (Denmark)

    Geng, Hui; Nandakumar, Kutty Selva; Pramhed, Anna;

    2012-01-01

    ABSTRACT: INTRODUCTION: Cartilage oligomeric matrix protein (COMP) is a major non-collagenous component of cartilage. Earlier, we developed a new mouse model for rheumatoid arthritis using COMP. This study was undertaken to investigate the epitope specificity and immunopathogenicity of COMP...... and the pathogenicity of mAbs was investigated by passive transfer experiments. RESULTS: B cell immunodominant epitopes were localized within 4 antigenic domains of the COMP but with preferential response to the epidermal growth factor (EGF)-like domain. Some of our anti-COMP mAbs showed interactions with the native...... form of COMP, which is present in cartilage and synovium. Passive transfer of COMP-specific mAbs enhanced arthritis when co-administrated with a sub-arthritogenic dose of a mAb specific to collagen type II. Interestingly, we found that a combination of 5 COMP mAbs was capable of inducing arthritis...

  15. Characterization of canine platelet adhesion to extracellular matrix proteins.

    Science.gov (United States)

    Pelagalli, Alessandra; Pero, Maria Elena; Mastellone, Vincenzo; Cestaro, Anna; Signoriello, Simona; Lombardi, Pietro; Avallone, Luigi

    2011-07-01

    Canine platelets have been extensively studied but little is known about specific aspects such as adhesion. Platelet adhesion is a critical step during haemostasis and thrombosis as well as during inflammatory and immunopathogenic responses. The aim of this study was to evaluate the adhesive properties of canine platelets using fibrinogen and collagen as substrates immobilized on plates. Adhesion was monitored for 120 min and the effect of adenosine 5'-diphosphate (ADP) was assayed. The results showed that canine platelets displayed good adhesion activity that was significantly time-dependent. Moreover, ADP was able to enhance platelet adhesion in a dose-dependent manner. The findings aid knowledge of the adhesion process and suggest a specific role of surface platelet receptors in mediating the interaction with extracellular matrix proteins.

  16. Calcium Sulfate with Stearic Acid as an Encouraging Carrier for Reindeer Bone Protein Extract

    Directory of Open Access Journals (Sweden)

    Pekka Jalovaara

    2011-07-01

    Full Text Available Various bone proteins and growth factors in specific concentrations are required for bone formation. If the body cannot produce sufficient quantities of these factors, bone trauma can be healed with an implant that includes the required factors in a carrier. This study was designed to evaluate various calcium salt candidates that can be used as carrier with reindeer bone protein extract to induce ectopic bone formation in the muscle pouch model of mouse. The bone protein extract was either impregnated into the disc form of carrier or mixed with carrier powder before implantation. The radiographic analysis indicated increased bone formation in all of the active groups containing the bone protein extract compared to the controls within 21 days follow-up. The highest bone formation was seen in the group with calcium sulfate with stearic acid where new bone and calcified cartilage were clearly visible. The greatest bone formation occurred in the groups that had bone protein extract readily available. This indicates that the bone forming factors in sufficient concentrations are required at the early stage of bone formation. The calcium sulfate with stearic acid was the most suitable and effective carrier for reindeer bone protein extract.

  17. Glycation of extracellular matrix proteins impairs migration of immune cells.

    Science.gov (United States)

    Haucke, Elisa; Navarrete-Santos, Alexander; Simm, Andreas; Silber, Rolf-Edgar; Hofmann, Britt

    2014-01-01

    The immune response during aging and diabetes is disturbed and may be due to the altered migration of immune cells in an aged tissue. Our study should prove the hypothesis that age and diabetes-related advanced glycation end products (AGEs) have an impact on the migration and adhesion of human T-cells. To achieve our purpose, we used in vitro AGE-modified proteins (soluble albumin and fibronectin [FN]), as well as human collagen obtained from bypass graft. A Boyden chamber was used to study cell migration. Migrated Jurkat T-cells were analyzed by flow cytometry and cell adhesion by crystal violet staining. Actin polymerization was determined by phalloidin-Alexa-fluor 488-labeled antibody and fluorescence microscopy. We found that significantly fewer cells (50%, p = 0.003) migrated through methylglyoxal modified FN. The attachment to FN in the presence of AGE-bovine serum albumin (BSA) was also reduced (p < 0.05). In ex vivo experiments, isolated collagen from human vein graft material negatively affected the migration of the cells depending on the grade of AGE modification of the collagen. Collagen with a low AGE level reduced the cell migration by 30%, and collagen with a high AGE level by 60%. Interaction of the cells with an AGE-modified matrix, but not with soluble AGEs like BSA-AGE per se, was responsible for a disturbed migration. The reduced migration was accompanied by an impaired actin polymerization. We conclude that AGEs-modified matrix protein inhibits cell migration and adhesion of Jurkat T-cells.

  18. Harmine promotes osteoblast differentiation through bone morphogenetic protein signaling

    Energy Technology Data Exchange (ETDEWEB)

    Yonezawa, Takayuki [Department of Nutriproteomics, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Lee, Ji-Won [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Hibino, Ayaka; Asai, Midori [Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Hojo, Hironori [Center for Disease Biology and Integrative Medicine, Faculty of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Cha, Byung-Yoon [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Teruya, Toshiaki [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Faculty of Education, University of the Ryukyus, 1 Senbaru, Nishihara, Okinawa 903-0213 (Japan); Nagai, Kazuo [Research Institute for Biological Functions, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Department of Biological Chemistry, College of Bioscience and Biotechnology, Chubu University, 1200 Matsumoto, Kasugai, Aichi 487-8501 (Japan); Chung, Ung-Il [Center for Disease Biology and Integrative Medicine, Faculty of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Yagasaki, Kazumi [Department of Nutriproteomics, Graduate School of Medicine, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-0033 (Japan); Division of Applied Biological Chemistry, Institute of Agriculture, Tokyo Noko University, 3-5-8 Saiwai, Fuchu, Tokyo 183-8509 (Japan); and others

    2011-06-03

    Highlights: {yields} Harmine promotes the activity and mRNA expression of ALP. {yields} Harmine enhances the expressions of osteocalcin mRNA and protein. {yields} Harmine induces osteoblastic mineralization. {yields} Harmine upregulates the mRNA expressions of BMPs, Runx2 and Osterix. {yields} BMP signaling pathways are involved in the actions of harmine. -- Abstract: Bone mass is regulated by osteoblast-mediated bone formation and osteoclast-mediated bone resorption. We previously reported that harmine, a {beta}-carboline alkaloid, inhibits osteoclast differentiation and bone resorption in vitro and in vivo. In this study, we investigated the effects of harmine on osteoblast proliferation, differentiation and mineralization. Harmine promoted alkaline phosphatase (ALP) activity in MC3T3-E1 cells without affecting their proliferation. Harmine also increased the mRNA expressions of the osteoblast marker genes ALP and Osteocalcin. Furthermore, the mineralization of MC3T3-E1 cells was enhanced by treatment with harmine. Harmine also induced osteoblast differentiation in primary calvarial osteoblasts and mesenchymal stem cell line C3H10T1/2 cells. Structure-activity relationship studies using harmine-related {beta}-carboline alkaloids revealed that the C3-C4 double bond and 7-hydroxy or 7-methoxy group of harmine were important for its osteogenic activity. The bone morphogenetic protein (BMP) antagonist noggin and its receptor kinase inhibitors dorsomorphin and LDN-193189 attenuated harmine-promoted ALP activity. In addition, harmine increased the mRNA expressions of Bmp-2, Bmp-4, Bmp-6, Bmp-7 and its target gene Id1. Harmine also enhanced the mRNA expressions of Runx2 and Osterix, which are key transcription factors in osteoblast differentiation. Furthermore, BMP-responsive and Runx2-responsive reporters were activated by harmine treatment. Taken together, these results indicate that harmine enhances osteoblast differentiation probably by inducing the expressions of

  19. Osteoblast Differentiation and Bone: Relevant proteins, regulatory processes and the vascular connection

    OpenAIRE

    2012-01-01

    textabstractBone is a highly specialized form of connective tissue present in most vertebrate animals as part of the endoskeleton. Structurally speaking, bone is mainly constituted by an organic extracellular matrix (ECM) hardened by deposited mineral. The blending between the organic and inorganic parts originates two main types of osseous tissue. The outer part of the tissue, the cortex, is hard compact bone and surrounds the inner trabecular bone, a spongy-like structure. In terms of funct...

  20. Cartilage damage involving extrusion of mineralisable matrix from the articular calcified cartilage and subchondral bone

    Directory of Open Access Journals (Sweden)

    A Boyde

    2011-05-01

    Full Text Available Arthropathy of the distal articular surfaces of the third metacarpal (Mc3 and metatarsal (Mt3 bones in the Thoroughbred racehorse (Tb is a natural model of repetitive overload arthrosis. We describe a novel pathology that affects the articular calcified cartilage (ACC and subchondral bone (SCB and which is associated with hyaline articular cartilage degeneration.Parasagittal slices cut from the palmar quadrant of the distal condyles of the left Mc3/Mt3 of 39 trained Tbs euthanased for welfare reasons were imaged by point projection microradiography, and backscattered electron (BSE scanning electron microscopy (SEM, light microscopy, and confocal scanning light microscopy. Mechanical properties were studied by nanoindentation. Data on the horses' training and racing career were also collected.Highly mineralised projections were observed extending from cracks in the ACC mineralising front into the hyaline articular cartilage (HAC up to two-thirds the thickness of the HAC, and were associated with focal HAC surface fibrillation directly overlying their site. Nanoindentation identified this extruded matrix to be stiffer than any other mineralised phase in the specimen by a factor of two. The presence of projections was associated with a higher cartilage Mankin histology score (P < 0.02 and increased amounts of gross cartilage loss pathologically on the condyle (P < 0.02. Presence of projections was not significantly associated with: total number of racing seasons, age of horse, amount of earnings, number of days in training, total distance galloped in career, or presence of wear lines.

  1. Ornamenting 3D printed scaffolds with cell-laid extracellular matrix for bone tissue regeneration.

    Science.gov (United States)

    Pati, Falguni; Song, Tae-Ha; Rijal, Girdhari; Jang, Jinah; Kim, Sung Won; Cho, Dong-Woo

    2015-01-01

    3D printing technique is the most sophisticated technique to produce scaffolds with tailorable physical properties. But, these scaffolds often suffer from limited biological functionality as they are typically made from synthetic materials. Cell-laid mineralized ECM was shown to be potential for improving the cellular responses and drive osteogenesis of stem cells. Here, we intend to improve the biological functionality of 3D-printed synthetic scaffolds by ornamenting them with cell-laid mineralized extracellular matrix (ECM) that mimics a bony microenvironment. We developed bone graft substitutes by using 3D printed scaffolds made from a composite of polycaprolactone (PCL), poly(lactic-co-glycolic acid) (PLGA), and β-tricalcium phosphate (β-TCP) and mineralized ECM laid by human nasal inferior turbinate tissue-derived mesenchymal stromal cells (hTMSCs). A rotary flask bioreactor was used to culture hTMSCs on the scaffolds to foster formation of mineralized ECM. A freeze/thaw cycle in hypotonic buffer was used to efficiently decellularize (97% DNA reduction) the ECM-ornamented scaffolds while preserving its main organic and inorganic components. The ECM-ornamented 3D printed scaffolds supported osteoblastic differentiation of newly-seeded hTMSCs by upregulating four typical osteoblastic genes (4-fold higher RUNX2; 3-fold higher ALP; 4-fold higher osteocalcin; and 4-fold higher osteopontin) and increasing calcium deposition compared to bare 3D printed scaffolds. In vivo, in ectopic and orthotopic models in rats, ECM-ornamented scaffolds induced greater bone formation than that of bare scaffolds. These results suggest a valuable method to produce ECM-ornamented 3D printed scaffolds as off-the-shelf bone graft substitutes that combine tunable physical properties with physiological presentation of biological signals.

  2. The diversity of shell matrix proteins: genome-wide investigation of the pearl oyster, Pinctada fucata.

    Science.gov (United States)

    Miyamoto, Hiroshi; Endo, Hirotoshi; Hashimoto, Naoki; Limura, Kurin; Isowa, Yukinobu; Kinoshita, Shigeharu; Kotaki, Tomohiro; Masaoka, Tetsuji; Miki, Takumi; Nakayama, Seiji; Nogawa, Chihiro; Notazawa, Atsuto; Ohmori, Fumito; Sarashina, Isao; Suzuki, Michio; Takagi, Ryousuke; Takahashi, Jun; Takeuchi, Takeshi; Yokoo, Naoki; Satoh, Nori; Toyohara, Haruhiko; Miyashita, Tomoyuki; Wada, Hiroshi; Samata, Tetsuro; Endo, Kazuyoshi; Nagasawa, Hiromichi; Asakawa, Shuichi; Watabe, Shugo

    2013-10-01

    In molluscs, shell matrix proteins are associated with biomineralization, a biologically controlled process that involves nucleation and growth of calcium carbonate crystals. Identification and characterization of shell matrix proteins are important for better understanding of the adaptive radiation of a large variety of molluscs. We searched the draft genome sequence of the pearl oyster Pinctada fucata and annotated 30 different kinds of shell matrix proteins. Of these, we could identified Perlucin, ependymin-related protein and SPARC as common genes shared by bivalves and gastropods; however, most gastropod shell matrix proteins were not found in the P. fucata genome. Glycinerich proteins were conserved in the genus Pinctada. Another important finding with regard to these annotated genes was that numerous shell matrix proteins are encoded by more than one gene; e.g., three ACCBP-like proteins, three CaLPs, five chitin synthase-like proteins, two N16 proteins (pearlins), 10 N19 proteins, two nacreins, four Pifs, nine shematrins, two prismalin-14 proteins, and 21 tyrosinases. This diversity of shell matrix proteins may be implicated in the morphological diversity of mollusc shells. The annotated genes reported here can be searched in P. fucata gene models version 1.1 and genome assembly version 1.0 ( http://marinegenomics.oist.jp/pinctada_fucata ). These genes should provide a useful resource for studies of the genetic basis of biomineralization and evaluation of the role of shell matrix proteins as an evolutionary toolkit among the molluscs.

  3. The effect of stromelysin-1 (MMP-3) on non-collagenous extracellular matrix proteins of demineralized dentin and the adhesive properties of restorative resins.

    Science.gov (United States)

    Boukpessi, T; Menashi, S; Camoin, L; Tencate, J M; Goldberg, M; Chaussain-Miller, C

    2008-11-01

    Dentin non-collagenous matrix components (NCPs) are structural proteins involved in the formation, the architecture and the mineralization of the extracellular matrix (ECM). We investigated here how recombinant metalloproteinase stromelysin-1, also termed MMP-3, initiates the release of ECM molecules from artificially demineralized human dentin. Analysis of the supernatants by Western blotting reveals that MMP-3 extracts PGs (decorin, biglycan), and also a series of phosphorylated proteins: dentin sialoprotein (DSP), osteopontin (OPN), bone sialoprotein (BSP) and MEPE, but neither dentin matrix protein-1 (DMP1), another member of the SIBLING family, nor osteocalcin (OC), a non-phosphorylated matrix molecule. After treatment of dentin surfaces by MMP-3, scanning electron microscope (SEM) examination of resin replica shows an increased penetration of the resin into the dentin tubules when compared to surfaces only treated by demineralizing solutions. This preclinical investigation suggests that MMP-3 may be used to improve the adhesive properties of restorative materials.

  4. Using bone marrow matrix to analyze meprobamate for forensic toxicological purposes.

    Science.gov (United States)

    Bévalot, F; Gustin, M P; Cartiser, N; Gaillard, Y; Le Meur, C; Fanton, L; Guitton, J; Malicier, D

    2013-09-01

    Bone marrow (BM) analysis is of forensic interest for postmortem toxicological investigations where blood samples are unavailable or unusable. Due to the lack of studies, it remains difficult to interpret concentrations of xenobiotics measured in this matrix. Based on a statistical approach published previously to interpret meprobamate concentrations in bile and vitreous humor, we propose here a diagnostic test for interpretation of BM meprobamate concentrations from analysis of 99 sets of autopsy data. The mean age was 48 years (range 18-80 years, one unknown) for males and 50 years (range 19-80 years, one unknown) for females, with a male/female ratio at 0.768. A BM concentration threshold of 11.3 μg/g was found to be statistically equivalent to that of a blood meprobamate concentration threshold of 50 μg/ml in distinguishing overdose from therapeutic use. The intrinsic qualities of this diagnostic test were good with sensitivity of 0.82 and specificity of 0.92. Compared to previous tests published with the same objective on vitreous humor and bile, this study shows that BM is a useful alternative matrix to reveal meprobamate overdose when blood, vitreous humor, and bile are not available or unusable.

  5. Matrix metalloproteinase-9 and cell division in neuroblastoma cells and bone marrow macrophages.

    Science.gov (United States)

    Sans-Fons, M Gloria; Sole, Sonia; Sanfeliu, Coral; Planas, Anna M

    2010-12-01

    Matrix metalloproteinases (MMPs) degrade the extracellular matrix and carry out key functions in cell development, cancer, injury, and regeneration. In addition to its well recognized extracellular action, functional intracellular MMP activity under certain conditions is supported by increasing evidence. In this study, we observed higher gelatinase activity by in situ zymography and increased MMP-9 immunoreactivity in human neuroblastoma cells and in bone marrow macrophages undergoing mitosis compared with resting cells. We studied the pattern of immunoreactivity at the different stages of cell division by confocal microscopy. Immunostaining with different monoclonal antibodies against MMP-9 revealed a precise, dynamic, and well orchestrated localization of MMP-9 at the different stages of cell division. The cellular distribution of MMP-9 staining was studied in relation to that of microtubules. The spatial pattern of MMP-9 immunoreactivity suggested some participation in both the reorganization of the nuclear content and the process of chromatid segmentation. We then used several MMP-9 inhibitors to find out whether MMP-9 might be involved in the cell cycle. These drugs impaired the entry of cells into mitosis, as revealed by flow cytometry, and reduced cell culture growth. In addition, the silencing of MMP-9 expression with small interfering RNA also reduced cell growth. Taken together, these results suggest that intracellular MMP-9 is involved in the process of cell division in neuroblastoma cells and in primary cultures of macrophages.

  6. Protein expression profile in the differentiation of rat bone marrow stromal cells into Schwann cell-like cells

    Institute of Scientific and Technical Information of China (English)

    LI WenTing; SUN HuaLin; XU ZengLu; DING Fei; GU XiaoSong

    2009-01-01

    During the last decade, increasing evidence suggested that bone marrow stromal cells (MSCs) have the potential to differentiate into neural lineages. Many studies have reported that MSCs showed morpho-logical changes and expressed a limited number of neural proteins under experimental conditions. However, no proteomic studies on MSCs differentiated into Schwann cell-like cells have been reported. In this study, we isolated MSCs from adult Sprague-Dawley rat femur and tibia bone marrows and in-duced the cells in vitro under specific conditions. By using two-dimensional gel electrophoresis (2-DE), we compared the protein profiles of MSCs before and after induced differentiation. We obtained 792 protein spots in the protein profile by 2-DE, and found that 74 spots changed significantly before and after the differentiation using PDQuest software, with 43 up-regulated and 31 down-regulated. We ana-lyzed these 74 spots by a matrix assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF-MS) and by database searching, and found that they could be grouped into various classes, including cytoskeleton and structure proteins, growth factors, metabolic proteins, chaperone proteins, receptor proteins, cell cycle proteins, calcium binding proteins, and other proteins. These proteins also include neural and glial proteins, such as BDNF, CNTF and GFAP. The results may provide valuable proteomic information about the differentiation of MSCs into Schwann cell-like cells.

  7. Protein expression profile in the differentiation of rat bone marrow stromal cells into Schwann cell-like cells

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    During the last decade,increasing evidence suggested that bone marrow stromal cells(MSCs) have the potential to differentiate into neural lineages.Many studies have reported that MSCs showed morphological changes and expressed a limited number of neural proteins under experimental conditions.However,no proteomic studies on MSCs differentiated into Schwann cell-like cells have been reported.In this study,we isolated MSCs from adult Sprague-Dawley rat femur and tibia bone marrows and induced the cells in vitro under specific conditions.By using two-dimensional gel electrophoresis(2-DE),we compared the protein profiles of MSCs before and after induced differentiation.We obtained 792 protein spots in the protein profile by 2-DE,and found that 74 spots changed significantly before and after the differentiation using PDQuest software,with 43 up-regulated and 31 down-regulated.We analyzed these 74 spots by a matrix assisted laser desorption ionization-time of flight mass spectrometry(MALDI-TOF-MS) and by database searching,and found that they could be grouped into various classes,including cytoskeleton and structure proteins,growth factors,metabolic proteins,chaperone proteins,receptor proteins,cell cycle proteins,calcium binding proteins,and other proteins.These proteins also include neural and glial proteins,such as BDNF,CNTF and GFAP.The results may provide valuable proteomic information about the differentiation of MSCs into Schwann cell-like cells.

  8. Fabrication method, structure, mechanical, and biological properties of decellularized extracellular matrix for replacement of wide bone tissue defects.

    Science.gov (United States)

    Anisimova, N Y; Kiselevsky, M V; Sukhorukova, I V; Shvindina, N V; Shtansky, D V

    2015-09-01

    The present paper was focused on the development of a new method of decellularized extracellular matrix (DECM) fabrication via a chemical treatment of a native bone tissue. Particular attention was paid to the influence of chemical treatment on the mechanical properties of native bones, sterility, and biological performance in vivo using the syngeneic heterotopic and orthotopic implantation models. The obtained data indicated that after a chemical decellularization treatment in 4% aqueous sodium chlorite, no noticeable signs of the erosion of compact cortical bone surface or destruction of trabeculae of spongy bone in spinal channel were observed. The histological studies showed that the chemical treatment resulted in the decellularization of both bone and cartilage tissues. The DECM samples demonstrated no signs of chemical and biological degradation in vivo. Thorough structural characterization revealed that after decellularization, the mineral frame retained its integrity with the organic phase; however clotting and destruction of organic molecules and fibers were observed. FTIR studies revealed several structural changes associated with the destruction of organic molecules, although all organic components typical of intact bone were preserved. The decellularization-induced structural changes in the collagen constituent resulted changed the deformation under compression mechanism: from the major fracture by crack propagation throughout the sample to the predominantly brittle fracture. Although the mechanical properties of radius bones subjected to decellularization were observed to degrade, the mechanical properties of ulna bones in compression and humerus bones in bending remained unchanged. The compressive strength of both the intact and decellularized ulna bones was 125-130 MPa and the flexural strength of humerus bones was 156 and 145 MPa for the intact and decellularized samples, respectively. These results open new avenues for the use of DECM samples as

  9. Histological analysis of cells and matrix mineralization of new bone tissue induced in rabbit femur bones by Mg-Zr based biodegradable implants.

    Science.gov (United States)

    Ragamouni, Sravanthi; Kumar, Jerald Mahesh; Mushahary, Dolly; Nemani, Harishankar; Pande, Gopal

    2013-09-01

    The biological efficacy of bone inducing implant materials in situ can be assessed effectively by performing histological analysis. We studied the peri-implant bone regeneration around two types of biodegradable magnesium-zirconium alloys, Mg-5Zr and Mg-Zr-2Sr, using histological, histochemical and immunohistochemical methods in the femur of New Zealand White strain rabbits. Our study includes three animal groups: (a) Mg-5Zr, (b) Mg-Zr-2Sr and (c) control. In each group three animals were used and in groups 'a' and 'b' the respective alloys were implanted in cavities made at the distal ends of the femur; control animals were left without implants to observe natural bone healing. Qualitative assessment of the cellularity and matrix mineralization events of the newly formed bone tissue was done at three months after implantation by histological methods in methyl methacrylate embedded tissue without decalcifying the bone. Quantitative mineral content and density of the new bone (NB) were evaluated by the statistical analysis of dual energy X-ray absorptiometry (DXA) data obtained from three animals in each experimental group. Based on our analysis we conclude that Mg-Zr-2Sr alloy showed better osseointegration of the newly formed bone with the implant surface. Our methodology of studying peri-implant osteoinduction of degradable implants using low temperature methyl methacrylate embedding resin can be useful as a general method for determining the bio-efficacy of implant materials. Copyright © 2013 Elsevier GmbH. All rights reserved.

  10. The Effect of an Enamel Matrix Derivative (Emdogain Combined with Bone Ceramic on Bone Formation in Mandibular Defects: A Histomorphometric and Immunohistochemical Study in the Canine

    Directory of Open Access Journals (Sweden)

    Reza Birang

    2012-01-01

    Full Text Available Background. The purpose of this study was to evaluate the combination of an enamel matrix derivative (EMD and an osteoconductive bone ceramic (BC in improving bone regeneration. Materials and Methods. Four cylindrical cavities (6×6mm were prepared bilaterally in the mandible in three dogs. The defects were randomly assigned to four different treatments—filled with EMD/BC and covered with a nonresorbable membrane, filled with EMD/BC without membrane, membrane coverage only, or control (left untreated—and healed for 2, 4, or 6 weeks. Harvested specimens were prepared for histologic, histomorphometric, and immunohistochemical analyses. Results. Sites treated with EMD/BC with or without membrane showed more total bone formation and lamellar bone formation than membrane-only and control defects. There were no statistically significant differences in total bone formation between EMD/BC with or without membrane. Conclusion. EMD with BC might improve bone formation in osseous defects more than membrane coverage alone; the use of a membrane had no significant additive effect on total bone formation.

  11. Photoencapsulation of bone morphogenetic protein-2 and periosteal progenitor cells improve tendon graft healing in a bone tunnel.

    Science.gov (United States)

    Chen, Chih-Hwa; Liu, Hsia-Wei; Tsai, Ching-Lin; Yu, Chung-Ming; Lin, I-Hsuan; Hsiue, Ging-Ho

    2008-03-01

    Tissue-engineered solutions for promoting the tendon graft incorporation within the bone tunnel appear to be promising. To determine the feasibility that conjugation of hyaluronic acid-tethered bone morphogenetic protein-2 can be used to stimulate periosteal progenitor cells direct fibrocartilagenous attachment and new bone formation in an extra-articular tendon-bone healing model. Controlled laboratory study. A total of 42 mature New Zealand White rabbits were used. The long digitorum extensor tendon was transplanted into a bone tunnel of the proximal tibia. The tendon was pulled through a drill hole in the proximal tibia and attached to the medial aspect of the tibia. Photopolymerizable hydrogel based on poly (ethylene glycol) diacrylate with hyaluronic acid-tethered bone morphogenetic protein-2 was injected and photogelated in a bone tunnel. Histological and biomechanical examination of the tendon-bone interface was evaluated at postoperative weeks 3 and 6. Histological analysis showed an interface fibrocartilage and new bone formed by photoencapsulation of bone morphogenetic protein-2 and periosteal progenitor cells at 6 weeks. Biomechanical testing revealed higher maximum pullout strength and stiffness in experimental groups with a statistically significant difference at 3 and 6 weeks after tendon transplantation. The healing tendon-bone interface undergoes a gradual remodeling process; it appears that photoencapsulation of bone morphogenetic protein-2 and periosteal progenitor cells possesses a powerful inductive ability between the tendon and the bone to incorporate the healing in a rabbit model. Novel technologies, such as those described in this study, including photopolymerization and tissue engineering, may provide minimally invasive therapeutic procedures via arthroscopy to enhance biological healing after reconstruction of the anterior cruciate ligament.

  12. The tent pole splint: a bone-supported stereolithographic surgical splint for the soft tissue matrix expansion graft procedure.

    Science.gov (United States)

    Cillo, Joseph E; Theodotou, Nicholas; Samuels, Marc; Krajekian, Joseph

    2010-06-01

    This report details the use of computer-aided planning and intraoperative stereolithographic direct-bone-contact surgical splints for the accurate extraoral placement of dental implants in the soft tissue matrix expansion (tent pole) graft of the severely resorbed mandible.

  13. Stimulation of Bone Healing by Sustained Bone Morphogenetic Protein 2 (BMP-2 Delivery

    Directory of Open Access Journals (Sweden)

    Mirja Faßbender

    2014-05-01

    Full Text Available The aim of the study was to investigate the effect of a sustained release of bone morphogenetic protein2 (BMP-2 incorporated in a polymeric implant coating on bone healing. In vitro analysis revealed a sustained, but incomplete BMP-2 release until Day 42. For the in vivo study, the rat tibia osteotomy was stabilized either with control or BMP-2 coated wires, and the healing progress was followed by micro computed tomography (µCT, biomechanical testing and histology at Days 10, 28, 42 and 84. MicroCT showed an accelerated formation of mineralized callus, as well as remodeling and an increase of mineralized/total callus volume (p = 0.021 at Day 42 in the BMP-2 group compared to the control. Histology revealed an increased callus mineralization at Days 42 and 84 (p = 0.006 with reduced cartilage at Day 84 (p = 0.004 in the BMP-2 group. Biomechanical stiffness was significantly higher in the BMP-2 group (p = 0.045 at Day 42. In summary, bone healing was enhanced after sustained BMP-2 application compared to the control. Using the same drug delivery system, but a burst release of BMP-2, a previous published study showed a similar positive effect on bone healing. Distinct differences in the healing outcome might be explained due to the different BMP release kinetics and dosages. However, further studies are necessary to adapt the optimal release profiles to physiological mechanisms.

  14. Bone marrow derived cell-seeded extracellular matrix: A novel biomaterial in the field of wound management

    Directory of Open Access Journals (Sweden)

    V. Remya

    2014-11-01

    Full Text Available Aim: Extensive or irreversible damage to the skin often requires additional skin substitutes for reconstruction. Biomaterials have become critical components in the development of effective new medical therapies for wound care. Materials and Methods: In the present study, a cell matrix construct (bone marrow-derived cells (BMdc seeded extracellular matrix [ECM] was used as a biological substitute for the repair of full-thickness skin wound. ECM was developed by decellularizing fish swim bladder (FSB. Goat bone marrow-derived cells (G-BMdc were seeded over this decellularized matrix. Efficacy of this cell matrix construct in wound repair was tested by implanting it over 20 mm2 × 20 mm2 size fullthickness skin wound created over the dorsum of rat. The study was conducted in 16 clinically healthy adult rats of either sex. The animals were randomly divided into 2 equal groups of 8 animals each. In Group I, animal’s wounds were repaired with a cellular FSB matrix. In Group II, wounds were repaired with G-BMdc seeded a cellular FSB matrix. Immune response and efficacy of healing were analyzed. Results: Quality of healing and immuno tolerance to the biological substitute was significantly better in Group II than Group I. Conclusion: Seeding with BMdc increases the wound healing potency and modulates the immune response to a significantly negligible level. The BMdc seeded acellular FSB matrix was found to be a novel biomaterial for wound management.

  15. The association of matrix Gla protein isomers with calcification in capsules surrounding silicone breast implants.

    Science.gov (United States)

    Hunter, Larry W; Lieske, John C; Tran, Nho V; Miller, Virginia M

    2011-11-01

    Implanted silicone medical prostheses induce a dynamic sequence of histologic events in adjacent tissue resulting in the formation of a fibrotic peri-prosthetic capsule. In some cases, capsular calcification occurs, requiring surgical intervention. In this study we investigated capsules from silicone gel-filled breast prostheses to test the hypothesis that this calcification might be regulated by the small vitamin K-dependent protein, matrix Gla protein (MGP), a potent inhibitor of arterial calcification, or by Fetuin-A, a hepatocyte-derived glycoprotein also implicated as a regulator of pathologic calcification. Immunolocalization studies of explanted capsular tissue, using conformation-specific antibodies, identified the mineralization-protective γ-carboxylated MGP isomer (cMGP) within cells of uncalcified capsules, whereas the non-functional undercarboxylated isomer (uMGP) was typically absent. Both were upregulated in calcific capsules and co-localized with mineral plaque and adjacent fibers. Synovial-like metaplasia was present in one uncalcified capsule in which MGP species were differentially localized within the pseudosynovium. Fetuin-A was localized to cells within uncalcified capsules and to mineral deposits within calcific capsules. The osteoinductive cytokine bone morphogenic protein-2 localized to collagen fibers in uncalcified capsules. These findings demonstrate that MGP, in its vitamin K-activated conformer, may represent a pharmacological target to sustain the health of the peri-prosthetic tissue which encapsulates silicone breast implants as well as other implanted silicone medical devices.

  16. Regulation of matrix metalloproteinase-9 protein expression by 1α, 25-(OH)₂D₃ during osteoclast differentiation.

    Science.gov (United States)

    Gu, Jian-Hong; Tong, Xi-Shuai; Chen, Guo-Hong; Liu, Xue-Zhong; Bian, Jian-Chun; Yuan, Yan; Liu, Zong-Ping

    2014-01-01

    To investigate 1α,25-(OH)₂D₃ regulation of matrix metalloproteinase-9 (MMP-9) protein expression during osteoclast formation and differentiation, receptor activator of nuclear factor kB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) were administered to induce the differentiation of RAW264.7 cells into osteoclasts. The cells were incubated with different concentrations of 1α,25-(OH)₂D₃ during culturing, and cell proliferation was measured using the methylthiazol tetrazolium method. Osteoclast formation was confirmed using tartrate-resistant acid phosphatase (TRAP) staining and assessing bone lacunar resorption. MMP-9 protein expression levels were measured with Western blotting. We showed that 1α,25-(OH)₂D₃ inhibited RAW264.7 cell proliferation induced by RANKL and M-CSF, increased the numbers of TRAP-positive osteoclasts and their nuclei, enhanced osteoclast bone resorption, and promoted MMP-9 protein expression in a concentration-dependent manner. These findings indicate that 1α,25-(OH)₂D₃ administered at a physiological relevant concentration promoted osteoclast formation and could regulate osteoclast bone metabolism by increasing MMP-9 protein expression during osteoclast differentiation.

  17. Biological assessment of a calcium silicate incorporated hydroxyapatite-gelatin nanocomposite: a comparison to decellularized bone matrix.

    Science.gov (United States)

    Lee, Dong Joon; Padilla, Ricardo; Zhang, He; Hu, Wei-Shou; Ko, Ching-Chang

    2014-01-01

    Our laboratory utilized biomimicry to develop a synthetic bone scaffold based on hydroxyapatite-gelatin-calcium silicate (HGCS). Here, we evaluated the potential of HGCS scaffold in bone formation in vivo using the rat calvarial critical-sized defect (CSD). Twelve Sprague-Dawley rats were randomized to four groups: control (defect only), decellularized bone matrix (DECBM), and HGCS with and without multipotent adult progenitor cells (MAPCs). DECBM was prepared by removing all the cells using SDS and NH4OH. After 12 weeks, the CSD specimens were harvested to evaluate radiographical, histological, and histomorphometrical outcomes. The in vitro osteogenic effects of the materials were studied by focal adhesion, MTS, and alizarin red. Micro-CT analysis indicated that the DECBM and the HGCS scaffold groups developed greater radiopaque areas than the other groups. Bone regeneration, assessed using histological analysis and fluorochrome labeling, was the highest in the HGCS scaffold seeded with MAPCs. The DECBM group showed limited osteoinductivity, causing a gap between the implant and host tissue. The group grafted with HGCS+MAPCs resulting in twice as much new bone formation seems to indicate a role for effective bone regeneration. In conclusion, the novel HGCS scaffold could improve bone regeneration and is a promising carrier for stem cell-mediated bone regeneration.

  18. Biomechanical and radiographic comparison of demineralized bone matrix, and a coralline hydroxyapatite in a rabbit spinal fusion model.

    Science.gov (United States)

    Dodds, Robert A; York-Ely, Amanda M; Zhukauskas, Rasa; Arola, Travis; Howell, John; Hartill, Caroline; Cobb, Ronald R; Fox, Casey

    2010-09-01

    The use of bone grafts is an essential component in spinal fusion. Autologous bone has been shown to result in long-term stable arthrodesis between spinal motion segments. However, autograft can be associated with significant morbidity and a limited supply. Alternatives, such as allogeneic demineralized bone matrix (DBM), are a potential source and supplement to autograft bone. The current study compares the ability of a DBM product (BioSet RT) and a coralline hydroxyapatite (Pro Osteon 500R), for inducing spinal fusion in a rabbit model. BioSet RT, alone or in combination with autograft, and Pro Osteon 500R were implanted in the posterior lateral inter-transverse process region of the rabbit spine. The spines were evaluated at 18 weeks for fusion of the L4-L5 transverse processes using a total of 33 skeletally mature male rabbits; 4 naïve animals were also included in the study. Samples were evaluated radiographically, histologically, by palpation, and through mechanical strength testing. Radiographical, histological, and palpation measurements demonstrated the ability of BioSet RT to induce new bone formation and bridging fusion comparable to autograft. This material performed well alone or in combination with autograft material. Despite significantly higher biomechanical testing results, minimal bone formation and fusion was recorded for the Pro Osteon 500R-treated group. This in vivo study demonstrates the ability of BioSet RT to induce new bone formation, and there was a clear relationship between bridging bone and mechanical strength.

  19. Biological Assessment of a Calcium Silicate Incorporated Hydroxyapatite-Gelatin Nanocomposite: A Comparison to Decellularized Bone Matrix

    Directory of Open Access Journals (Sweden)

    Dong Joon Lee

    2014-01-01

    Full Text Available Our laboratory utilized biomimicry to develop a synthetic bone scaffold based on hydroxyapatite-gelatin-calcium silicate (HGCS. Here, we evaluated the potential of HGCS scaffold in bone formation in vivo using the rat calvarial critical-sized defect (CSD. Twelve Sprague-Dawley rats were randomized to four groups: control (defect only, decellularized bone matrix (DECBM, and HGCS with and without multipotent adult progenitor cells (MAPCs. DECBM was prepared by removing all the cells using SDS and NH4OH. After 12 weeks, the CSD specimens were harvested to evaluate radiographical, histological, and histomorphometrical outcomes. The in vitro osteogenic effects of the materials were studied by focal adhesion, MTS, and alizarin red. Micro-CT analysis indicated that the DECBM and the HGCS scaffold groups developed greater radiopaque areas than the other groups. Bone regeneration, assessed using histological analysis and fluorochrome labeling, was the highest in the HGCS scaffold seeded with MAPCs. The DECBM group showed limited osteoinductivity, causing a gap between the implant and host tissue. The group grafted with HGCS+MAPCs resulting in twice as much new bone formation seems to indicate a role for effective bone regeneration. In conclusion, the novel HGCS scaffold could improve bone regeneration and is a promising carrier for stem cell-mediated bone regeneration.

  20. The comparative effectiveness of demineralized bone matrix, beta-tricalcium phosphate, and bovine-derived anorganic bone matrix on inflammation and bone formation using a paired calvarial defect model in rats

    Directory of Open Access Journals (Sweden)

    Khoshzaban A

    2011-09-01

    Full Text Available Ahad Khoshzaban1,2,3, Shahram Mehrzad1, Vida Tavakoli2, Saeed Heidari Keshel2, Gholam Reza Behrouzi2, Maryam Bashtar2 1Iranian Tissue Bank Research and Preparation Center, Imam Khomeini Hospital Complex, 2Stem Cells Preparation Unit, Eye Research Center, Farabi Hospital, Tehran University of Medical Science, 3Dental Bio Material Department, Tehran University of Medical Science, Faculty of Dentistry, Tehran, Iran Background: In this study, the effectiveness of Iranian Tissue Bank–produced demineralized bone matrix (ITB-DBM, beta-tricalcium phosphate (ßTCP, and Bio-Oss® (Geistlich Pharma AG, Wolhusen, Switzerland were evaluated and compared with double controls. The main goal was to measure the amount of new bone formation in the center of defects created in rat calvaria. Another goal was to compare the controls and evaluate the effects of each treatment material on their adjacent untreated (control defects. Methods: In this study, 40 male Wistar rats were selected and divided into four groups, In each group, there were ten rats with two defects in their calvarias; one of them is considered as control and the other one was treated with ITB-DBM (group 1, BIO-OSS (group2, and ßTCP (group 3, respectively. But in group 4, both defects were considered as control. The amount of inflammation and new bone formation were evaluated at 4 and 10 weeks. In the first group, one defect was filled with ITB-DBM; in the second group, one defect was filled with Bio-Oss; in the third group, one defect was filled with ßTCP; and in the fourth group, both defects were left unfilled. Zeiss microscope (Carl Zeiss AG, Oberkochen, Germany and Image Tool® (version 3.0; University of Texas Health Science Center at San Antonio, San Antonio, TX software were used for evaluation. SPSS Statistics (IBM Corp, Somers, NY was used for statistical analysis. Results: Maximum bone formation at 4 and 10 weeks were observed in the ITB-DBM group (46.960% ± 4.366%, 94.970% ± 0

  1. Identification of calprotectin, a calcium binding leukocyte protein, in human dental calculus matrix.

    Science.gov (United States)

    Kido, J; Nishikawa, S; Ishida, H; Yamashita, K; Kitamura, S; Kohri, K; Nagata, T

    1997-05-01

    Calprotectin is a calcium binding protein produced by leukocytes, macrophages and epithelial cells, and its levels in several tissues increase during infections and in many inflamed areas, suggesting that it may be an indicator of inflammatory activity. Osteopontin is a prominent phosphorylated glycoprotein in bone matrix, having calcium binding capacity. Recently, it has been reported that calprotectin and osteopontin are present in urinary stones (pathological mineralized masses in the body), and that these proteins may be involved in their formation. Dental calculus formed by mineralization of dental plaque is an inflammatory factor which may contribute to periodontal disease. It contains many organic components involved in mineralization. We recently found osteopontin molecules in human dental calculus and suggested that the components of its matrix may be similar to those of urinary stones. In this study, we investigated the presence of calprotectin in human dental calculus by immunohistochemical and immunoblotting analyses using a specific antibody for calprotectin. After fixation and demineralization of dental calculi adhered to tooth roots, sections embedded in paraffin were immunoreacted with the antibody for calprotectin and positive immunostaining for calprotectin was observed. Dental calculus proteins were then extracted with EDTA and separated by electrophoresis on 15% polyacrylamide gels. By immunoblotting analysis, 3 or 4 bands were observed at 11, 14.5, 22-25, 28 or 36.5 kDa and these patterns corresponded to those of calprotectin subunits. When non-immune rabbit serum was used instead of calprotectin-specific antibody as a negative control, no immunoreactivity was observed. These findings indicate that calprotectin is associated not only with antibacterial action but also with calcium binding capacity during dental calculus formation.

  2. Altered distribution of extracellular matrix proteins in the periodontal ligament of periostin-deficient mice.

    Science.gov (United States)

    Tabata, Chihiro; Hongo, Hiromi; Sasaki, Muneteru; Hasegawa, Tomoka; de Freitas, Paulo Henrique Luiz; Yamada, Tamaki; Yamamoto, Tomomaya; Suzuki, Reiko; Yamamoto, Tsuneyuki; Oda, Kimimitsu; Li, Minqi; Kudo, Akira; Iida, Junichiro; Amizuka, Norio

    2014-06-01

    Verifying whether periostin affects the distribution of type I collagen, fibronectin and tenascin C in the periodontal ligament (PDL) is important to contribute to a more thorough understanding of that protein's functions. In this study, we have histologically examined incisor PDL of mandibles in 20 week-old male wild-type and periostin-deficient (periostin-/-) mice, by means of type I collagen, fibronectin, tenascin C, proliferating cell nuclear antigen, matrix metallo-proteinase (MMP)-1 and F4/80-positive monocyte/macrophage immunostaining, transmission electron microscopy and quantitative analysis of cell proliferation. Wild-type PDL featured well-arranged layers of collagen bundles intertwined with PDL cells, whose longitudinal axis ran parallel to the collagen fibers. However, cells in the periostin-/- PDL were irregularly distributed among collagen fibrils, which were also haphazardly arranged. Type I collagen and fibronectin reactivity was seen throughout the wild-type PDL, while in the periostin-/- PDL, only focal, uneven staining for these proteins could be seen. Similarly, tenascin C staining was evenly distributed in the wild-type PDL, but hardly seen in the periostin-/- PDL. MMP-1 immunoreactivity was uniformly distributed in the wild-type PDL, but only dotted staining could be discerned in the periostin-/- PDL. F4/80-positive monocyte/macrophages were found midway between tooth- and bone-related regions in the wild-type PDL, a pattern that could not be observed in the periostin-/- PDL. In summary, periostin deficiency may not only cause PDL collagen fibril disorganization, but could also affect the distribution of other major extracellular matrix proteins such as fibronectin and tenascin C.

  3. Nanofibrous yet injectable polycaprolactone-collagen bone tissue scaffold with osteoprogenitor cells and controlled release of bone morphogenetic protein-2

    Energy Technology Data Exchange (ETDEWEB)

    Subramanian, Gayathri; Bialorucki, Callan [Department of Bioengineering, College of Engineering, University of Toledo, Toledo, OH 43606 (United States); Yildirim-Ayan, Eda, E-mail: eda.yildirimayan@utoledo.edu [Department of Bioengineering, College of Engineering, University of Toledo, Toledo, OH 43606 (United States); Department of Orthopaedic Surgery, University of Toledo Medical Center, Toledo, OH 43614 (United States)

    2015-06-01

    In this work, we developed a nanofibrous, yet injectable orthobiologic tissue scaffold that is capable of hosting osteoprogenitor cells and controlling kinetic release profile of the encapsulated pro-osteogenic factor without diminishing its bioactivity over 21 days. This innovative injectable scaffold was synthesized by incorporating electrospun and subsequently O{sub 2} plasma-functionalized polycaprolactone (PCL) nanofibers within the collagen type-I solution along with MC3T3-E1 cells (pre-osteoblasts) and bone morphogenetic protein-2 (BMP2). Through changing the PCL nanofiber concentration within the injectable scaffolds, we were able to tailor the mechanical strength, protein retention capacity, bioactivity preservation, and osteoinductive potential of the scaffolds. The nanofibrous internal structure of the scaffold allowed us to use a low dose of BMP2 (200 ng/ml) to achieve osteoblastic differentiation in in vitro culture. The osteogenesis capacity of the injectable scaffolds were evaluated though measuring MC3T3-E1 cell proliferation, ALP activity, matrix mineralization, and early- and late-osteoblast specific gene expression profiles over 21 days. The results demonstrated that the nanofibrous injectable scaffold provides not only an osteoinductive environment for osteoprogenitor cells to differentiate, but also a suitable biomechanical and biochemical environment to act as a reservoir for osteogenic factors with controlled release profile. - Highlights: • Injectable nanofibrous scaffold with osteoprogenitor cells and BMP2 was synthesized. • PCL nanofiber concentration within collagen scaffold affected the BMP2 retention and bioactivity. • Optimal PCL concentration was identified for mechanical stability, injectability, and osteogenic activity. • Scaffolds exhibited long-term osteoinductive capacity for bone repair and regeneration.

  4. Role of osteogenic protein-1/bone morphogenetic protein-7 in spinal fusion

    Directory of Open Access Journals (Sweden)

    Justin Munns

    2009-10-01

    Full Text Available Justin Munns, Daniel K Park, Kern SinghDepartment of Orthopedic Surgery, Rush University Medical Center, Chicago, Illinois, USAAbstract: Osteogenic protein-1 (OP-1, also known as bone morphogenetic protein-7 (BMP-7, is a protein in the TGF-β family of cellular proteins that has shown potential for application in patients undergoing spinal fusion due to its proven osteoinductive effects, particularly in patients with spondylolisthesis. OP-1 initiates numerous processes at the cellular level, acting on mesenchymal stem cells (MSCs, osteoblasts, and osteoclasts to stimulate bone growth. Animal studies of OP-1 have provided strong evidence for the ability of OP-1 to initiate ossification in posterolateral arthrodesis. Promising findings in early clinical trials with OP-1 prompted FDA approval for use in long bone nonunions in 2001 and subsequently for revision posterolateral arthrodesis in 2004 under a conditional Humanitarian Device Exemption. Larger clinical trials have recently shown no notable safety concerns or increases in adverse events associated with OP-1. However, a recent clinical trial has not conclusively demonstrated the noninferiority of OP-1 compared to autograft in revision posterolateral arthrodesis. The future of OP-1 application in patients with spondylolisthesis thus remains uncertain with the recent rejection of Premarket Approval (PMA status by the FDA (April 2009. Further investigation of its treatment success and immunological consequences appears warranted to establish FDA approval for its use in its current form.Keywords: osteogenic protein-1, bone morphogenetic protein-7, spinal fusion

  5. Protein and amino acid quality of meat and bone meal.

    Science.gov (United States)

    Parsons, C M; Castanon, F; Han, Y

    1997-02-01

    The in vivo protein quality of 14 meat and bone meals (MBM) was evaluated in three chick growth assays and a 48-h excreta collection assay using conventional and cecectomized roosters. In addition, in vitro evaluation of protein quality was assessed using pepsin N digestibility (0.2, 0.002, or 0.0002% pepsin), KOH protein solubility, and multi-enzyme pH change. Crude protein, lysine, and SAA in the MBM varied from 48 to 56, 2.32 to 3.01, and 1.0 to 2.13%, respectively. Protein efficiency ratio (weight gain:protein intake) estimated from feeding chicks diets containing 9% protein from a MBM ranged from 0.61 to 2.89 and averaged 1.78. Lysine bioavailability determined by slope-ratio chick assay ranged from 43 to 89%. True amino acid digestibility and TMEn values determined in cecectomized roosters were generally lower (P < 0.05) than those determined in conventional roosters. True digestibility of amino acids (percentage) also varied among MBM, with the mean (and range) for lysine, methionine, and cystine in cecectomized birds being 81 (73 to 88), 85 (77 to 91), and 58% (37 to 72%), respectively. Pepsin N digestibility values determined using 0.002 or 0.0002% pepsin were positively correlated (P < 0.05) with lysine digestibility. Pepsin N digestibility determined using 0.2% pepsin, KOH protein solubility, and multi-enzyme pH change were not significantly correlated with in vivo protein quality. Ash content was negatively correlated (-0.80, P < 0.05) with protein efficiency ratio. These results indicated that there is substantial variation in protein quality among commercial MBM and that pepsin N digestibility and ash content are correlated with some in vivo protein quality measurements.

  6. Enhanced healing of rabbit segmental radius defects with surface-coated calcium phosphate cement/bone morphogenetic protein-2 scaffolds

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Yi; Hou, Juan; Yin, ManLi [Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237 (China); Wang, Jing, E-mail: biomatwj@163.com [Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237 (China); Liu, ChangSheng, E-mail: csliu@sh163.net [Engineering Research Center for Biomedical Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237 (China); The State Key Laboratory of Bioreactor Engineering, East China University of Science and Technology, Shanghai 200237 (China); Key Laboratory for Ultrafine Materials of Ministry of Education, East China University of Science and Technology, Shanghai 200237 (China)

    2014-11-01

    Large osseous defects remain a difficult clinical problem in orthopedic surgery owing to the limited effective therapeutic options, and bone morphogenetic protein-2 (BMP-2) is useful for its potent osteoinductive properties in bone regeneration. Here we build a strategy to achieve prolonged duration time and help inducting new bone formation by using water-soluble polymers as a protective film. In this study, calcium phosphate cement (CPC) scaffolds were prepared as the matrix and combined with sodium carboxymethyl cellulose (CMC-Na), hydroxypropylmethyl cellulose (HPMC), and polyvinyl alcohol (PVA) respectively to protect from the digestion of rhBMP-2. After being implanted in the mouse thigh muscles, the surface-modified composite scaffolds evidently induced ectopic bone formation. In addition, we further evaluated the in vivo effects of surface-modified scaffolds in a rabbit radius critical defect by radiography, three dimensional micro-computed tomographic (μCT) imaging, synchrotron radiation-based micro-computed tomographic (SRμCT) imaging, histological analysis, and biomechanical measurement. The HPMC-modified CPC scaffold was regarded as the best combination for segmental bone regeneration in rabbit radius. - Highlights: • A simple surface-coating method was used to fabricate composite scaffolds. • Growth factor was protected from rapid depletion via superficial coating. • Significant promotion of bone regeneration was achieved. • HPMC-modification displayed optimal effect of bone regeneration.

  7. Calcium Phosphate Scaffolds Combined with Bone Morphogenetic Proteins or Mesenchymal Stem Cells in Bone Tissue Engineering

    Directory of Open Access Journals (Sweden)

    Han Sun

    2015-01-01

    Full Text Available Objective: The purpose of this study was to review the current status of calcium phosphate (CaP scaffolds combined with bone morphogenetic proteins (BMPs or mesenchymal stem cells (MSCs in the field of bone tissue engineering (BTE. Date Sources: Data cited in this review were obtained primarily from PubMed and Medline in publications from 1979 to 2014, with highly regarded older publications also included. The terms BTE, CaP, BMPs, and MSC were used for the literature search. Study Selection: Reviews focused on relevant aspects and original articles reporting in vitro and/or in vivo results concerning the efficiency of CaP/BMPs or CaP/MSCs composites were retrieved, reviewed, analyzed, and summarized. Results: An ideal BTE product contains three elements: Scaffold, growth factors, and stem cells. CaP-based scaffolds are popular because of their outstanding biocompatibility, bioactivity, and osteoconductivity. However, they lack stiffness and osteoinductivity. To solve this problem, composite scaffolds of CaP with BMPs have been developed. New bone formation by CaP/BMP composites can reach levels similar to those of autografts. CaP scaffolds are compatible with MSCs and CaP/MSC composites exhibit excellent osteogenesis and stiffness. In addition, a CaP/MSC/BMP scaffold can repair bone defects more effectively than an autograft. Conclusions: Novel BTE products possess remarkable osteoconduction and osteoinduction capacities, and exhibit balanced degradation with osteogenesis. Further work should yield safe, viable, and efficient materials for the repair of bone lesions.

  8. Calcium Phosphate Scaffolds Combined with Bone Morphogenetic Proteins or Mesenchymal Stem Cells in Bone Tissue Engineering

    Institute of Scientific and Technical Information of China (English)

    Han Sun; Hui-Lin Yang

    2015-01-01

    Objective:The purpose of this study was to review the current status of calcium phosphate (CaP) scaffolds combined with bone morphogenetic proteins (BMPs) or mesenchymal stem cells (MSCs) in the field of bone tissue engineering (BTE).Date Sources:Data cited in this review were obtained primarily from PubMed and Medline in publications from 1979 to 2014,with highly regarded older publications also included.The terms BTE,CaP,BMPs,and MSC were used for the literature search.Study Selection:Reviews focused on relevant aspects and original articles reporting in vitro and/or in vivo results concerning the efficiency of CaP/BMPs or CaP/MSCs composites were retrieved,reviewed,analyzed,and summarized.Results:An ideal BTE product contains three elements:Scaffold,growth factors,and stem cells.CaP-based scaffolds are popular because of their outstanding biocompatibility,bioactivity,and osteoconductivity.However,they lack stiffness and osteoinductivity.To solve this problem,composite scaffolds of CaP with BMPs have been developed.New bone formation by CaP/BMP composites can reach levels similar to those of autografts.CaP scaffolds are compatible with MSCs and CaP/MSC composites exhibit excellent osteogenesis and stiffness.In addition,a CaP/MSC/BMP scaffold can repair bone defects more effectively than an autograft.Conclusions:Novel BTE products possess remarkable osteoconduction and osteoinduction capacities,and exhibit balanced degradation with osteogenesis.Further work should yield safe,viable,and efficient materials for the repair of bone lesions.

  9. The effect of statins in colorectal cancer is mediated through the bone morphogenetic protein pathway

    NARCIS (Netherlands)

    Kodach, Liudmila L.; Bleuming, Sylvia A.; Peppelenbosch, Maikel P.; Hommes, Daniel W.; Van Den Brink, Gus R.; Hardwick, James C. H.

    2007-01-01

    Background & Aims: Epidemiological evidence suggests that statins prevent colorectal cancer (CRC), but the biological mechanism remains obscure. Statins induce bone morphogenetic protein (BMP) expression in bone cells. We have previously shown that BMPs act as tumor suppressors in CRC. We

  10. Electron cryotomography of measles virus reveals how matrix protein coats the ribonucleocapsid within intact virions.

    Science.gov (United States)

    Liljeroos, Lassi; Huiskonen, Juha T; Ora, Ari; Susi, Petri; Butcher, Sarah J

    2011-11-01

    Measles virus is a highly infectious, enveloped, pleomorphic virus. We combined electron cryotomography with subvolume averaging and immunosorbent electron microscopy to characterize the 3D ultrastructure of the virion. We show that the matrix protein forms helices coating the helical ribonucleocapsid rather than coating the inner leaflet of the membrane, as previously thought. The ribonucleocapsid is folded into tight bundles through matrix-matrix interactions. The implications for virus assembly are that the matrix already tightly interacts with the ribonucleocapsid in the cytoplasm, providing a structural basis for the previously observed regulation of RNA transcription by the matrix protein. Next, the matrix-covered ribonucleocapsids are transported to the plasma membrane, where the matrix interacts with the envelope glycoproteins during budding. These results are relevant to the nucleocapsid organization and budding of other paramyxoviruses, where isolated matrix has been observed to form helices.

  11. Disposable polymeric cryogel bioreactor matrix for therapeutic protein production.

    Science.gov (United States)

    Jain, Era; Kumar, Ashok

    2013-05-01

    Low cost and high efficiency make disposable bioreactors feasible for small-scale therapeutic development and initial clinical trials. We have developed a cryogel-based disposable bioreactor matrix, which has been used for production of protein therapeutics such as urokinase and monoclonal antibodies (mAbs). The protocol discusses the application of a cryogel bioreactor for mAb production. Cryogels composed of either polyacrylamide (PAAm) coupled to gelatin or semi-interpenetrating PAAm-chitosan are synthesized by free-radical polymerization at -12 °C. Hybridoma cells are immobilized over the cryogel bioreactor and incubated for 48 h. Medium is circulated thereafter at 0.2 ml min(-1) and bioreactors can be run continuously for 60 d. The cryogel-based packed-bed bioreactor can be formulated as a monolith or as beads; it also has an efficiency four times what can be obtained using a tissue-culture flask, a high surface-to-volume ratio and effective nutrient transport. After incubation, the bioreactor setup will take about 60 min using a pre-prepared sterilized cryogel.

  12. Protein and mineral characterisation of rendered meat and bone meal.

    Science.gov (United States)

    Buckley, M; Penkman, K E H; Wess, T J; Reaney, S; Collins, M J

    2012-10-01

    We report the characterisation of meat and bone meal (MBM) standards (Set B-EFPRA) derived from cattle, sheep, pig and chicken, each rendered at four different temperatures (133, 137, 141 and 145 °C). The standards, prepared for an EU programme STRATFEED (to develop new methodologies for the detection and quantification of illegal addition of mammalian tissues in feeding stuffs), have been widely circulated and used to assess a range of methods for identification of the species composition of MBM. The overall state of mineral alteration and protein preservation as a function of temperature was monitored using small angle X-ray diffraction (SAXS), amino acid composition and racemization analyses. Progressive increases in protein damage and mineral alteration in chicken and cattle standards was observed. In the case of sheep and pig, there was greater damage to the proteins and alteration of the minerals at the lowest treatment temperature (133 °C), suggesting that the thermal treatments must have been compromised in some way. This problem has probably impacted upon the numerous studies which tested methods against these heat treatments. We use protein mass spectrometric methods to explore if thermostable proteins could be used to identify rendered MBM. In more thermally altered samples, so-called 'thermostable' proteins such as osteocalcin which has been proposed as a ideal target to speciate MBM were no longer detectable, but the structural protein type I collagen could be used to differentiate all four species, even in the most thermally altered samples.

  13. Clotting protein - An extracellular matrix (ECM) protein involved in crustacean hematopoiesis.

    Science.gov (United States)

    Junkunlo, Kingkamon; Söderhäll, Kenneth; Söderhäll, Irene

    2017-09-21

    Hematopoietic progenitor cells in crustaceans are organized in lobule-like structures surrounded by different types of cells and extracellular matrix (ECM) protein in a Hematopoietic tissue (HPT). Here we show that the clotting protein (CP) is part of the ECM in HPT and is secreted during HPT cell culture. The formation of a filamentous network of CP was observed in HPT cell culture. A high amount of CP protein was detected at the surfaces of undifferentiated cells (round-shaped) compared with migrating cells (spindle shaped). Co-localization of the CP protein and TGase activity was observed on the cell surface and filamentous network between cells. A role for CP together with collagen was revealed in a 3D culture in which a collagen-I matrix was immobilized with CP or supplemented with CP. The results showed possible functions of CP, collagen, TGase and cytokine Ast1 in the regulation of HPT progenitor cell behavior. This is the first study to provide insight into the role of CP, which probably not only participates in clot formation but also functions as an ECM component protein controlling hematopoietic stem cell behavior. Copyright © 2017. Published by Elsevier Ltd.

  14. Purification of capping protein using the capping protein binding site of CARMIL as an affinity matrix.

    Science.gov (United States)

    Remmert, Kirsten; Uruno, Takehito; Hammer, John A

    2009-10-01

    Capping protein (CP) is a ubiquitously expressed, heterodimeric actin binding protein that is essential for normal actin dynamics in cells. The existing methods for purifying native CP from tissues and recombinant CP from bacteria are time-consuming processes that involve numerous conventional chromatographic steps and functional assays to achieve a homogeneous preparation of the protein. Here, we report the rapid purification of Acanthamoeba CP from amoeba extracts and recombinant mouse CP from E. coli extracts using as an affinity matrix GST-fusion proteins containing the CP binding site from Acanthamoeba CARMIL and mouse CARMIL-1, respectively. This improved method for CP purification should facilitate the in vitro analysis of CP structure, function, and regulation.

  15. Compound soft regenerated skull material for repairing dog skull defects using bone morphogenetic protein as an inductor and nanohydroxyapatite as a scaffold

    Institute of Scientific and Technical Information of China (English)

    Zhidong Shi; Mingwang Liu; Zhongzong Qin; Qinmei Wang; Ying Guo; Haiyong He; Zhonghe Yu

    2008-01-01

    BACKGROUND: In previous studies of skull defects and regeneration, bone morphogenetic protein as an inductor and nanohydroxyapatite as a scaffold have been cocultured with osteoblasts.OBJECTIVE: To verify the characteristics of the new skull regenerated material after compound soft regenerated skull material implantatiom.DESIGN, TIME AND SETTING: The self-control and inter-group control animal experiment was perfurmed at the Sun Yat-sen University, China from February to July 2007.MATERIALS: Twenty-tour healthy adult dogs of both genders weighing 15-20 kg were used in this study. Nanohydroxyapatite as a scaffold was cocultured with osteoblasts. Using demineralized canine bone matrix as a carrier, recombinant human bone morphogenetic protein-2 was employed to prepare compound soft regenerated skull material. Self-designed compound soft regenerated skull material was implanted in models of skull defects.METHODS: Animals were randomly assigned into two groups, Group A (n = 16) and Group B (n = 8).Bilateral 2.5-cm-diameter full-thickness parietal skull defects were made in all animals. In Group A, the right side was reconstructed with calcium alginate gel, osteoblasts, and nanomcter bone meal composite;the left side was reconstructed with calcium alginate gel, osteoblasts, nanometer bone meal and recombinant human bone morphogenetic protein-2 composite. In Group B, the right side was kept as a simple skull detect, and the left side was reconstructed with calcium alginate gel, osteoblasts, nanometer bone meal and recombinant human bone morphogenetic protein-2 composite.MAIN OUTCOME MEASURES: Bone regeneration and histopathological changes at the site of the skull defect were observed with an optical microscope and a scanning electron microscope after surgery.The ability to form bone was measured by alizarin red S staining. In vitro cultured osteoblasts were observed for morphology.RESULTS: One month following surgery, newly formed bone trabeculae mostly covered the

  16. Nell1-deficient mice have reduced expression of extracellular matrix proteins causing cranial and vertebral defects

    Energy Technology Data Exchange (ETDEWEB)

    Desai, Jayashree [University of Tennessee, Knoxville (UTK) & Oak Ridge National Laboratory (ORNL); Shannon, Mark E. [Applied Biosystems; Johnson, Mahlon D. [University of Tennessee Graduate School of Medicine; Ruff, David W. [Applied Biosystems; Hughes, Lori A [ORNL; Kerley, Marilyn K [ORNL; Carpenter, D A [ORNL; Johnson, Dabney K [ORNL; Rinchik, Eugene M. [University of Tennessee, Knoxville (UTK) & Oak Ridge National Laboratory (ORNL); Culiat, Cymbeline T [ORNL

    2006-01-01

    The mammalian Nell1 gene encodes a protein kinase C-b1 (PKC-b1) binding protein that belongs to a new class of cell-signaling molecules controlling cell growth and differentiation. Over-expression of Nell1 in the developing cranial sutures in both human and mouse induces craniosynostosis, the premature fusion of the growing cranial bone fronts. Here, we report the generation, positional cloning and characterization of Nell16R, a recessive, neonatal-lethal point mutation in the mouse Nell1 gene, induced by N-ethyl-N-nitrosourea. Nell16R has a T!A base change that converts a codon for cysteine into a premature stop codon [Cys(502)Ter], resulting in severe truncation of the predicted protein product and marked reduction in steady-state levels of the transcript. In addition to the expected alteration of cranial morphology, Nell16R mutants manifest skeletal defects in the vertebral column and ribcage, revealing a hitherto undefined role for Nell1 in signal transduction in endochondral ossification. Real-time quantitative reverse transcription-PCR assays of 219 genes showed an association between the loss of Nell1 function and reduced expression of genes for extracellular matrix (ECM) proteins critical for chondrogenesis and osteogenesis. Several affected genes are involved in the human cartilage disorder Ehlers-Danlos Syndrome and other disorders associated with spinal curvature anomalies. Nell16R mutant mice are a new tool for elucidating basic mechanisms in osteoblast and chrondrocyte differentiation in the developing skull and vertebral column and understanding how perturbations in the production of ECM proteins can lead to anomalies in these structures.

  17. Molecular cloning and characterization of lustrin A, a matrix protein from shell and pearl nacre of Haliotis rufescens.

    Science.gov (United States)

    Shen, X; Belcher, A M; Hansma, P K; Stucky, G D; Morse, D E

    1997-12-19

    A specialized extracellular matrix of proteins and polysaccharides controls the morphology and packing of calcium carbonate crystals and becomes occluded within the mineralized composite during formation of the molluscan shell and pearl. We have cloned and characterized the cDNA coding for Lustrin A, a newly described matrix protein from the nacreous layer of the shell and pearl produced by the abalone, Haliotis rufescens, a marine gastropod mollusc. The full-length cDNA is 4,439 base pairs (bp) long and contains an open reading frame coding for 1,428 amino acids. The deduced amino acid sequence reveals a highly modular structure with a high proportion of Ser (16%), Pro (14%), Gly (13%), and Cys (9%). The protein contains ten highly conserved cysteine-rich domains interspersed by eight proline-rich domains; a glycine- and serine-rich domain lies between the two cysteine-rich domains nearest the C terminus, and these are followed by a basic domain and a C-terminal domain that is highly similar to known protease inhibitors. The glycine- and serine-rich domain and at least one of the proline-rich domains show sequence similarity to proteins of two extracellular matrix superfamilies (one of which also is involved in the mineralized matrixes of bone, dentin, and avian eggshell). The arrangement of alternating cysteine-rich domains and proline-rich domains is strikingly similar to that found in frustulins, the proteins that are integral to the silicified cell wall of diatoms. Its modular structure suggests that Lustrin A is a multifunctional protein, whereas the occurrence of related sequences suggest it is a member of a multiprotein family.

  18. Extracellular matrix mineralization in periodontal tissues: Noncollagenous matrix proteins, enzymes, and relationship to hypophosphatasia and X-linked hypophosphatemia.

    Science.gov (United States)

    McKee, Marc D; Hoac, Betty; Addison, William N; Barros, Nilana M T; Millán, José L; Chaussain, Catherine

    2013-10-01

    As broadly demonstrated for the formation of a functional skeleton, proper mineralization of periodontal alveolar bone and teeth - where calcium phosphate crystals are deposited and grow within an extracellular matrix - is essential for dental function. Mineralization defects in tooth dentin and cementum of the periodontium invariably lead to a weak (soft or brittle) dentition in which teeth become loose and prone to infection and are lost prematurely. Mineralization of the extremities of periodontal ligament fibers (Sharpey's fibers) where they insert into tooth cementum and alveolar bone is also essential for the function of the tooth-suspensory apparatus in occlusion and mastication. Molecular determinants of mineralization in these tissues include mineral ion concentrations (phosphate and calcium), pyrophosphate, small integrin-binding ligand N-linked glycoproteins and matrix vesicles. Amongst the enzymes important in regulating these mineralization determinants, two are discussed at length here, with clinical examples given, namely tissue-nonspecific alkaline phosphatase and phosphate-regulating gene with homologies to endopeptidases on the X chromosome. Inactivating mutations in these enzymes in humans and in mouse models lead to the soft bones and teeth characteristic of hypophosphatasia and X-linked hypophosphatemia, respectively, where the levels of local and systemic circulating mineralization determinants are perturbed. In X-linked hypophosphatemia, in addition to renal phosphate wasting causing low circulating phosphate levels, phosphorylated mineralization-regulating small integrin-binding ligand N-linked glycoproteins, such as matrix extracellular phosphoglycoprotein and osteopontin, and the phosphorylated peptides proteolytically released from them, such as the acidic serine- and aspartate-rich-motif peptide, may accumulate locally to impair mineralization in this disease.

  19. Mitochondrial unfolded protein response controls matrix pre-RNA processing and translation.

    Science.gov (United States)

    Münch, Christian; Harper, J Wade

    2016-06-30

    The mitochondrial matrix is unique in that it must integrate the folding and assembly of proteins derived from the nuclear and mitochondrial genomes. In Caenorhabditis elegans, the mitochondrial unfolded protein response (UPRmt) senses matrix protein misfolding and induces a program of nuclear gene expression, including mitochondrial chaperonins, to promote mitochondrial proteostasis. While misfolded mitochondrial-matrix-localized ornithine transcarbamylase induces chaperonin expression, our understanding of mammalian UPRmt is rudimentary, reflecting a lack of acute triggers for UPRmt activation. This limitation has prevented analysis of the cellular responses to matrix protein misfolding and the effects of UPRmt on mitochondrial translation to control protein folding loads. Here we combine pharmacological inhibitors of matrix-localized HSP90/TRAP1 (ref. 8) or LON protease, which promote chaperonin expression, with global transcriptional and proteomic analysis to reveal an extensive and acute response of human cells to UPRmt. This response encompasses widespread induction of nuclear genes, including matrix-localized proteins involved in folding, pre-RNA processing and translation. Functional studies revealed rapid but reversible translation inhibition in mitochondria occurring concurrently with defects in pre-RNA processing caused by transcriptional repression and LON-dependent turnover of the mitochondrial pre-RNA processing nuclease MRPP3 (ref. 10). This study reveals that acute mitochondrial protein folding stress activates both increased chaperone availability within the matrix and reduced matrix-localized protein synthesis through translational inhibition, and provides a framework for further dissection of mammalian UPRmt.

  20. Alteration of nuclear matrix-intermediate filament system and differential expression of nuclear matrix proteins during human hepatocarcinoma cell differentiation

    Institute of Scientific and Technical Information of China (English)

    Jian Tang; Jing-Wen Niu; Dong-Hui Xu; Zhi-Xing Li; Qi-Fu Li; Jin-An Chen

    2007-01-01

    AIM:To investigate the association between the configurational and compositional changes of nuclear matrix and the differentiation of carcinoma cells.METHODS: Cells cultured with or without 5 × 10-3mmol/L of hexamethylene bisacetamide (HMBA) on Nickel grids were treated by selective extraction and prepared for whole mount observation under electron microscopy. The samples were examined under transmission electron microscope. Nuclear matrix proteins were selectively extracted and subjected to subcellular proteomics study. The protein expression patterns were analyzed by PDQuest software. Spots of differentially expressed nuclear matrix proteins were excised and subjected to in situ digestion with trypsin.The peptides were analyzed by matrix-assisted laserdesorption/ionization time of flight mass spectrometry (MALDI-TOF-MS). Data were submitted for database searching using Mascot tool (www. Matrixscience.com).RESULTS: The nuclear matrix (NM) and intermediate filament (IF) in SMMC-7721 hepatocarcinoma cells were found relatively sparse and arranged irregularly.The nuclear lamina was non-uniform, and two kinds of filaments were not tightly connected. After induction for differentiation by HMBA, the NM-IF filaments were concentrated and distributed uniformly. The heterogeneous population of filaments, including highly branched utrathin filaments could also be seen in the regular meshwork. The connection between the two kinds of filaments and the relatively thin, condensed and sharply demarcated lamina composed of intermediatesized filaments was relatively fastened. Meanwhile, 21NM proteins changed remarkably during SMMC-7721cell differentiation. Four proteins, I.e. Mutant Pyst1,hypothetical protein, nucleophosmin1, and LBP were downregulated, whereas four other proteins, eIF6, p44subunit, β-tubulin, and SIN3B were upregulated with the last one, SR2/ASF found only in the differentiated SMMC-7721 cells.CONCLUSION: The induced differentiation of SMMC-7721cells by HMBA is

  1. Adhesion of the human pathogen Sporothrix schenckii to several extracellular matrix proteins

    Directory of Open Access Journals (Sweden)

    O.C. Lima

    1999-05-01

    Full Text Available The pathogenic fungus Sporothrix schenckii is the causative agent of sporotrichosis. This subcutaneous mycosis may disseminate in immunocompromised individuals and also affect several internal organs and tissues, most commonly the bone, joints and lung. Since adhesion is the first step involved with the dissemination of pathogens in the host, we have studied the interaction between S. schenckii and several extracellular matrix (ECM proteins. The binding of two morphological phases of S. schenckii, yeast cells and conidia, to immobilized type II collagen, laminin, fibronectin, fibrinogen and thrombospondin was investigated. Poly (2-hydroxyethyl methacrylate (poly-HEMA was used as the negative control. Cell adhesion was assessed by ELISA with a rabbit anti-S. schenckii antiserum. The results indicate that both morphological phases of this fungus can bind significantly to type II collagen, fibronectin and laminin in comparison to the binding observed with BSA (used as blocking agent. The adhesion rate observed with the ECM proteins (type II collagen, fibronectin and laminin was statistically significant (P<0.05 when compared to the adhesion obtained with BSA. No significant binding of conidia was observed to either fibrinogen or thrombospondin, but yeast cells did bind to the fibrinogen. Our results indicate that S. schenckii can bind to fibronectin, laminin and type II collagen and also show differences in binding capacity according to the morphological form of the fungus.

  2. Comparison between heparin-conjugated fibrin and collagen sponge as bone morphogenetic protein-2 carriers for bone regeneration

    OpenAIRE

    Yang, Hee Seok; La, Wan-Geun; Cho, Yong-Min; Shin, Wangsoo; Yeo, Guw-Dong; Kim, Byung-Soo

    2012-01-01

    Bone morphogenetic protein-2 (BMP-2) is used to promote bone regeneration. However, the bone regeneration ability of BMP-2 relies heavily on the delivery vehicle. Previously, we have developed heparin-conjugated fibrin (HCF), a vehicle for long-term delivery of BMP-2 and demonstrated that long-term delivery of BMP-2 enhanced its osteogenic efficacy as compared to short-term delivery at an equivalent dose. The aim of this study was to compare the bone-forming ability of the BMP-2 delivered by ...

  3. A Novel High Mechanical Property PLGA Composite Matrix Loaded with Nanodiamond-Phospholipid Compound for Bone Tissue Engineering.

    Science.gov (United States)

    Zhang, Fan; Song, Qingxin; Huang, Xuan; Li, Fengning; Wang, Kun; Tang, Yixing; Hou, Canglong; Shen, Hongxing

    2016-01-20

    A potential bone tissue engineering material was produced from a biodegradable polymer, poly(lactic-co-glycolic acid) (PLGA), loaded with nanodiamond phospholipid compound (NDPC) via physical mixing. On the basis of hydrophobic effects and physical absorption, we modified the original hydrophilic surface of the nanodiamond (NDs) with phospholipids to be amphipathic, forming a typical core-shell structure. The ND-phospholipid weight ratio was optimized to generate sample NDPC50 (i.e., ND-phospholipid weight ratio of 100:50), and NDPC50 was able to be dispersed in a PLGA matrix at up to 20 wt %. Compared to a pure PLGA matrix, the introduction of 10 wt % of NDPC (i.e., sample NDPC50-PF10) resulted in a significant improvement in the material's mechanical and surface properties, including a decrease in the water contact angle from 80 to 55°, an approximately 100% increase in the Young's modulus, and an approximate 550% increase in hardness, thus closely resembling that of human cortical bone. As a novel matrix supporting human osteoblast (hFOB1.19) growth, NDPC50-PFs with different amounts of NDPC50 demonstrated no negative effects on cell proliferation and osteogenic differentiation. Furthermore, we focused on the behaviors of NDPC-PFs implanted into mice for 8 weeks and found that NDPC-PFs induced acceptable immune response and can reduce the rapid biodegradation of PLGA matrix. Our results represent the first in vivo research on ND (or NDPC) as nanofillers in a polymer matrix for bone tissue engineering. The high mechanical properties, good in vitro and in vivo biocompatibility, and increased mineralization capability suggest that biodegradable PLGA composite matrices loaded with NDPC may potentially be useful for a variety of biomedical applications, especially bone tissue engineering.

  4. Effect of Extracellular Matrix Membrane on Bone Formation in a Rabbit Tibial Defect Model

    Directory of Open Access Journals (Sweden)

    Jin Wook Hwang

    2016-01-01

    Full Text Available Absorbable extracellular matrix (ECM membrane has recently been used as a barrier membrane (BM in guided tissue regeneration (GTR and guided bone regeneration (GBR. Absorbable BMs are mostly based on collagen, which is more biocompatible than synthetic materials. However, implanted absorbable BMs can be rapidly degraded by enzymes in vivo. In a previous study, to delay degradation time, collagen fibers were treated with cross-linking agents. These compounds prevented the enzymatic degradation of BMs. However, cross-linked BMs can exhibit delayed tissue integration. In addition, the remaining cross-linker could induce inflammation. Here, we attempted to overcome these problems using a natural ECM membrane. The membrane consisted of freshly harvested porcine pericardium that was stripped from cells and immunoreagents by a cleaning process. Acellular porcine pericardium (APP showed a bilayer structure with a smooth upper surface and a significantly coarser bottom layer. APP is an ECM with a thin layer (0.18–0.35 mm but with excellent mechanical properties. Tensile strength of APP was 14.15±2.24 MPa. In in vivo experiments, APP was transplanted into rabbit tibia. The biocompatible material was retained for up to 3 months without the need for cross-linking. Therefore, we conclude that APP could support osteogenesis as a BM for up to 3 months.

  5. Osteoblast Differentiation and Bone: Relevant proteins, regulatory processes and the vascular connection

    NARCIS (Netherlands)

    R.D.A.M. Alves (Rodrigo)

    2012-01-01

    textabstractBone is a highly specialized form of connective tissue present in most vertebrate animals as part of the endoskeleton. Structurally speaking, bone is mainly constituted by an organic extracellular matrix (ECM) hardened by deposited mineral. The blending between the organic and inorganic

  6. Cloning of Mouse Enamel Matrix Serine Proteinase Encoding Mature Protein

    Institute of Scientific and Technical Information of China (English)

    MU Ya-bing; SUN Hong-chen; ZHANG Ze-bing; OUYANG Jie

    2003-01-01

    Objective: To clone cDNA of enamel matrix serine proteinase (EMSP1) encoding mature protein from mouse dental germs. Methods: Total RNA was isolated from developing incisors and molars of 7 days mouse pups and reverse-transcribed into cDNA. Two pairs of specific primers was designed to obtain the desired gene by Touchdown PCR and Nested PCR. The segment was inserted into Vector pMD-18T, and recombined vectors was transformed into E.coli JM109.The positive clone was chose and analysed by restriction endonuclease mapping and DNA sequencing. Results:700 bp of cDNA of mouse EMSP1 was sueccessfully cloned from mouse tooth germs tissue. The sequence was consistent with that displayed in PubMed. Conclusion:The mouse EMSP1 cDNA encoding mature protein is obtained for further study.%目的:克隆小鼠牙胚组织中釉基质丝氨酸蛋白酶(EMSP1)成熟肽编码区基因.方法:提取出生后7 d昆明种小白鼠切牙、磨牙牙胚总RNA,逆转录为cDNA,设计两对特异性引物,采用Touchdown PCR 和嵌套PCR方法,扩增出小鼠EMSP1起始密码子至终止密码子基因片段.将目的基因连入载体pMD-18T,转化入大肠杆菌JM109,通过蓝白筛选,挑选阳性克隆培养扩增,纯化重组质粒进行限制性酶切和核苷酸序列分析鉴定.结果:限制性酶切图谱和核苷酸序列分析均表明所克隆cDNA为小鼠700 bp的EMSP1成熟肽基因编码.结论:成功地克隆了小鼠编码EMSP1成熟肽基因片段.

  7. Photolytic Cross-Linking to Probe Protein-Protein and Protein-Matrix Interactions in Lyophilized Powders.

    Science.gov (United States)

    Iyer, Lavanya K; Moorthy, Balakrishnan S; Topp, Elizabeth M

    2015-09-08

    Protein structure and local environment in lyophilized formulations were probed using high-resolution solid-state photolytic cross-linking with mass spectrometric analysis (ssPC-MS). In order to characterize structure and microenvironment, protein-protein, protein-excipient, and protein-water interactions in lyophilized powders were identified. Myoglobin (Mb) was derivatized in solution with the heterobifunctional probe succinimidyl 4,4'-azipentanoate (SDA) and the structural integrity of the labeled protein (Mb-SDA) confirmed using CD spectroscopy and liquid chromatography/mass spectrometry (LC-MS). Mb-SDA was then formulated with and without excipients (raffinose, guanidine hydrochloride (Gdn HCl)) and lyophilized. The freeze-dried powder was irradiated with ultraviolet light at 365 nm for 30 min to produce cross-linked adducts that were analyzed at the intact protein level and after trypsin digestion. SDA-labeling produced Mb carrying up to five labels, as detected by LC-MS. Following lyophilization and irradiation, cross-linked peptide-peptide, peptide-water, and peptide-raffinose adducts were detected. The exposure of Mb side chains to the matrix was quantified based on the number of different peptide-peptide, peptide-water, and peptide-excipient adducts detected. In the absence of excipients, peptide-peptide adducts involving the CD, DE, and EF loops and helix H were common. In the raffinose formulation, peptide-peptide adducts were more distributed throughout the molecule. The Gdn HCl formulation showed more protein-protein and protein-water adducts than the other formulations, consistent with protein unfolding and increased matrix interactions. The results demonstrate that ssPC-MS can be used to distinguish excipient effects and characterize the local protein environment in lyophilized formulations with high resolution.

  8. Demineralized bone matrix and human cancellous bone enhance fixation of porous-coated titanium implants in sheep

    DEFF Research Database (Denmark)

    Babiker, Hassan; Ding, Ming; Overgaard, Søren

    2016-01-01

    Allogenic bone graft has been considered the gold standard in connection with bone graft material in revision joint arthroplasty. However, the lack of osteogenic potential and the risk of disease transmission are clinical challenges. The use of osteoinductive materials, such as demineralized bone...... of DBM alone, DBM with CB, or allograft on the fixation of porous-coated titanium implants. DBM100 and CB produced from human tissue were included. Both materials are commercially available. DBM granules are placed in pure DBM and do not contain any other carrier. Titanium alloy implants, 10 mm long × 10...... (gold standard), respectively. A standardized surgical procedure was used. At sacrifice 6 weeks after implantation, both distal femurs were harvested. The implant fixation was evaluated by mechanical push-out testing to test shear mechanical properties between implant and the host bone...

  9. Actin-binding protein coronin 1A controls osteoclastic bone resorption by regulating lysosomal secretion of cathepsin K

    Science.gov (United States)

    Ohmae, Saori; Noma, Naruto; Toyomoto, Masayasu; Shinohara, Masahiro; Takeiri, Masatoshi; Fuji, Hiroaki; Takemoto, Kenji; Iwaisako, Keiko; Fujita, Tomoko; Takeda, Norihiko; Kawatani, Makoto; Aoyama, Mineyoshi; Hagiwara, Masatoshi; Ishihama, Yasushi; Asagiri, Masataka

    2017-01-01

    Osteoclasts degrade bone matrix proteins via the secretion of lysosomal enzymes. However, the precise mechanisms by which lysosomal components are transported and fused to the bone-apposed plasma membrane, termed ruffled border membrane, remain elusive. Here, we identified coronin 1A as a negative regulator of exocytotic release of cathepsin K, one of the most important bone-degrading enzymes in osteoclasts. The modulation of coronin 1A expression did not alter osteoclast differentiation and extracellular acidification, but strongly affected the secretion of cathepsin K and osteoclast bone-resorption activity, suggesting the coronin 1A-mediated regulation of lysosomal trafficking and protease exocytosis. Further analyses suggested that coronin 1A prevented the lipidation-mediated sorting of the autophagy-related protein LC3 to the ruffled border and attenuated lysosome–plasma membrane fusion. In this process, the interactions between coronin 1A and actin were crucial. Collectively, our findings indicate that coronin 1A is a pivotal component that regulates lysosomal fusion and the secretion pathway in osteoclast-lineage cells and may provide a novel therapeutic target for bone diseases. PMID:28300073

  10. When size matters: differences in demineralized bone matrix particles affect collagen structure, mesenchymal stem cell behavior, and osteogenic potential.

    Science.gov (United States)

    Dozza, B; Lesci, I G; Duchi, S; Della Bella, E; Martini, L; Salamanna, F; Falconi, M; Cinotti, S; Fini, M; Lucarelli, E; Donati, D

    2017-04-01

    Demineralized bone matrix (DBM) is a natural, collagen-based, osteoinductive biomaterial. Nevertheless, there are conflicting reports on the efficacy of this product. The purpose of this study was to evaluate whether DBM collagen structure is affected by particle size and can influence DBM cytocompatibility and osteoinductivity. Sheep cortical bone was ground and particles were divided in three fractions with different sizes, defined as large (L, 1-2 mm), medium (M, 0.5-1 mm), and small (S, structure, with DBM-M being altered but not as much as DBM-S. DBM-M displayed a preferable trend in almost all biological characteristics tested, although all DBM particles revealed an optimal cytocompatibility. Subcutaneous implantation of DBM particles into immunocompromised mice resulted in bone induction only for DBM-M. When sheep MSC were seeded onto particles before implantation, all DBM particles were able to induce new bone formation with the best incidence for DBM-M and DBM-S. In conclusion, the collagen alteration in DBM-M is likely the best condition to promote bone induction in vivo. Furthermore, the choice of 0.5-1 mm particles may enable to obtain more efficient and consistent results among different research groups in bone tissue-engineering applications. © 2017 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 105A: 1019-1033, 2017.

  11. Effects of casein, whey and soy proteins on volumetric bone density and bone strength in immunocompromised piglets

    DEFF Research Database (Denmark)

    Budek, Alicja Zofia; Bjørnvad, Charlotte; Mølgaard, Christian

    2007-01-01

    assigned to a formula based on either casein (n=11), whey (n=11) or soy (n=10) as the protein source (each 55 g/L), and equal amounts of fat, carbohydrates, calcium and phosphorus. Results & Conclusion: Despite efforts to sustain immuno-protection (sow serum and antibiotic injections), some piglets became......Summary:Background and aims: Bone-promoting effect of different proteins in early life, under immunocompromised conditions, is unknown. We investigated effects of milk- and plantderived proteins on bone development in immunocompromised piglets. Methods: Newborn, colostrum-deprived piglets were...... sick and were early euthanised. After 6 days, bone density (peripheral quantitative computed tomography), bone mechanical strength (three-point bending test) and serum insulin-like growth factor-I (sIGF-I) (immunoassay) were measured in the surviving piglets (casein n=5, whey n=9, soy n=5)....

  12. Relative and combined effects of ethanol and protein deficiency on bone manganese and copper.

    Science.gov (United States)

    González-Pérez, José M; González-Reimers, Emilio; DeLaVega-Prieto, María José; Durán-Castellón, María del Carmen; Viña-Rodríguez, José; Galindo-Martín, Luis; Alvisa-Negrín, Julio; Santolaria-Fernández, Francisco

    2012-06-01

    Both manganese and copper may affect bone synthesis. Bone content of both metals can be altered in alcoholics, although controversy exists regarding this matter. To analyse the relative and combined effects of ethanol and a low protein diet on bone copper and manganese, and their relationships with bone structure and metabolism, including trabecular bone mass (TBM), osteoid area (OA), osteocalcin (OCN), insulin-like growth factor-1 (IGF-1), parathyroid hormone (PTH), urinary hydroxyproline (uHP) and vitamin D. Adult male Sprague-Dawley rats were divided into four groups. The control rats received a 18% protein-containing diet; a second group, an isocaloric, 2% protein-containing diet; a third one, an isocaloric, 36% ethanol-containing diet and a fourth, an isocaloric diet containing 2% protein and 36% ethanol. After sacrifice, TBM and OA were histomorphometrically assessed; bone and serum manganese and copper were determined by atomic absorption spectrophotometry, and serum OCN, IGF-1, PTH, uHP and vitamin D by radioimmunoassay. Ethanol-fed rats showed decreased TBM and bone manganese. Significant relationships existed between bone manganese and TBM, serum IGF-1 and OCN. Ethanol leads to a decrease in bone manganese, related to decreased bone mass and bone synthesis. No alterations were found in bone copper.

  13. Combination of absorbable mesh and demineralized bone matrix in orbital wall fracture for preventing herniation of orbit.

    Science.gov (United States)

    Tak, Kyoung Seok; Jung, Min Su; Lee, Byeong Ho; Kim, Joo Hyun; Ahn, Duk Kyun; Jeong, Hii Sun; Park, Young Kyu; Suh, In Suck

    2014-07-01

    After restoration of orbit wall fracture, preventing sequelae is important. An absorbable mesh is commonly used in orbit wall fracture, yet it has limitation due to orbit sagging when bony defect is larger than the moderate size (1 × 1 cm2). In this study, the authors present a satisfactory result in treating orbit wall fracture larger than the moderate size with a combination of absorbable mesh and demineralized bone matrix.From 2009 to 2012, 63 patients with bony defect larger than the moderate size, who were treated with a combination of absorbable mesh and demineralized bone matrix, were reviewed retrospectively. The site of bony defect, size, and applied amount of demineralized bone matrix were reviewed, and a 2-year follow-up was done. Facial computed tomography scans were checked preoperative, immediate postoperative, and 2-year postoperative.Among the 63 patients, there were 52 men and 11 women. Mean age was 33.3 years. The most common cause was blunt blow (35 cases); mean defect size was 13.36 × 12.82 mm2 in inferior wall fracture and 20.69 × 14.41 mm2 in medial wall fracture. There was no complication except for 3 cases of infraorbital nerve hypoesthesia. A 2-year follow-up computed tomography showed that the surgical site preserved bony formation without herniation. In treating moderate-sized bony defect in orbit wall fracture, absorbable mesh and demineralized bone matrix can maintain structural stability through good bony formation even after degradation of absorbable mesh.

  14. Scaffolds for bone regeneration made of hydroxyapatite microspheres in a collagen matrix

    Energy Technology Data Exchange (ETDEWEB)

    Cholas, Rahmatullah, E-mail: rahmat.cholas@gmail.com; Kunjalukkal Padmanabhan, Sanosh, E-mail: sanosh2001@gmail.com; Gervaso, Francesca; Udayan, Gayatri; Monaco, Graziana; Sannino, Alessandro; Licciulli, Antonio

    2016-06-01

    Biomimetic scaffolds with a structural and chemical composition similar to native bone tissue may be promising for bone tissue regeneration. In the present work hydroxyapatite mesoporous microspheres (mHA) were incorporated into collagen scaffolds containing an ordered interconnected macroporosity. The mHA were obtained by spray drying of a nano hydroxyapatite slurry prepared by the precipitation technique. X-ray diffraction (XRD) analysis revealed that the microspheres were composed only of hydroxyapatite (HA) phase, and energy-dispersive x-ray spectroscopy (EDS) analysis revealed the Ca/P ratio to be 1.69 which is near the value for pure HA. The obtained microspheres had an average diameter of 6 μm, a specific surface area of 40 m{sup 2}/g as measured by Brunauer-Emmett-Teller (BET) analysis, and Barrett-Joyner-Halenda (BJH) analysis showed a mesoporous structure with an average pore diameter of 16 nm. Collagen/HA-microsphere (Col/mHA) composite scaffolds were prepared by freeze-drying followed by dehydrothermal crosslinking. SEM observations of Col/mHA scaffolds revealed HA microspheres embedded within a porous collagen matrix with a pore size ranging from a few microns up to 200 μm, which was also confirmed by histological staining of sections of paraffin embedded scaffolds. The compressive modulus of the composite scaffold at low and high strain values was 1.7 and 2.8 times, respectively, that of pure collagen scaffolds. Cell proliferation measured by the MTT assay showed more than a 3-fold increase in cell number within the scaffolds after 15 days of culture for both pure collagen scaffolds and Col/mHA composite scaffolds. Attractive properties of this composite scaffold include the potential to load the microspheres for drug delivery and the controllability of the pore structure at various length scales. - Highlights: • Mesoporous hydroxyapatite microsphere(mHA) synthesized by spray drying method • Porous collagen/mHA composite scaffold made by freeze

  15. Vitamin D and calcium supplementation for three years in postmenopausal osteoporosis significantly alters bone mineral and organic matrix quality.

    Science.gov (United States)

    Paschalis, E P; Gamsjaeger, S; Hassler, N; Fahrleitner-Pammer, A; Dobnig, H; Stepan, J J; Pavo, I; Eriksen, E F; Klaushofer, K

    2017-02-01

    Prospective, controlled clinical trials in postmenopausal osteoporosis typically compare effects of an active drug with placebo in addition to vitamin D and calcium supplementation in both treatment arms. While clinical benefits are documented, the effect of this supplementation in the placebo arm and in clinical practice on bone material composition properties is unknown. The purpose of the present study was to evaluate these bone quality indices (specifically mineral/matrix, nanoporosity, glycosaminoglycan content, mineral maturity/crystallinity, and pyridinoline content) in patients that either received long-term vitamin D (400-1200IU) and calcium (1.0-1.5g) supplementation, or did not. We have analyzed by Raman microspectroscopy the bone forming trabecular surfaces of iliac crest in pre-treatment samples of a teriparatide study and the endpoint biopsies of the control arm obtained from the HORIZON trial. In general, the mineral/matrix ratio and the glycosaminoglycan (GAG) content was higher while nanoporosity, (a surrogate for tissue water content), the mineral maturity/crystallinity (MMC) and the pyridinoline (Pyd) content was lower in patients without long-term supplementation. Moreover, all indices were significantly dependent on tissue age. In conclusion, vitamin D and calcium supplementation is associated with altered mineral and organic matrix properties.

  16. Combined intervention of dietary soybean proteins and swim training: effects on bone metabolism in ovariectomized rats.

    Science.gov (United States)

    Figard, Hélène; Mougin, Fabienne; Gaume, Vincent; Berthelot, Alain

    2006-01-01

    Soybean proteins, a rich source of isoflavones, taken immediately after an ovariectomy prevent bone loss in rats. Exercise-induced stimuli are essential for bone growth. Few studies exist about the combined effects of swim training and soybean protein supplementation on bone metabolism. So, the purpose of this study was to investigate, in 48 female Sprague-Dawley rats (12 weeks old) the effects of an 8-week swim-training regimen (1 h/day, 5 days/week) and dietary soybean proteins (200 g/kg diet) on bone metabolism. Rats were randomly assigned to four groups: (1) ovariectomized fed with a semisynthetic control diet; (2) ovariectomized fed with a soybean protein-enriched semisynthetic diet; (3) ovariectomized trained to exercise and fed with control diet; (4) ovariectomized trained to exercise and fed with a soybean protein diet. Following the treatment period, body weight gain was identical in the four groups. Soybean protein supplementation increased bone calcium content, and reduced plasma osteocalcin values, without significant modification of calcium balance and net calcium absorption. Swim training enhanced plasma and bone calcium content and calcium balance and net calcium absorption. It did not modify either plasma osteocalcin values or urinary deoxypyridinoline excretion. Both exercise and soybean protein intake increased plasma on bone calcium without modifying net calcium absorption or bone markers. In conclusion, we demonstrated, in ovariectomized rats, that swimming exercise and dietary supplementation with soy proteins do not have synergistic effects on calcium metabolism and bone markers.

  17. Influence of heating and cyclic tension on the induction of heat shock proteins and bone-related proteins by MC3T3-E1 cells.

    Science.gov (United States)

    Chung, Eunna; Sampson, Alana Cherrell; Rylander, Marissa Nichole

    2014-01-01

    Stress conditioning (e.g., thermal, shear, and tensile stress) of bone cells has been shown to enhance healing. However, prior studies have not investigated whether combined stress could synergistically promote bone regeneration. This study explored the impact of combined thermal and tensile stress on the induction of heat shock proteins (HSPs) and bone-related proteins by a murine preosteoblast cell line (MC3T3-E1). Cells were exposed to thermal stress using a water bath (44°C for 4 or 8 minutes) with postheating incubation (37°C for 4 hours) followed by exposure to cyclic strain (equibiaxial 3%, 0.2 Hz, cycle of 10-second tensile stress followed by 10-second rest). Combined thermal stress and tensile stress induced mRNA expression of HSP27 (1.41 relative fold induction (RFI) compared to sham-treated control), HSP70 (5.55 RFI), and osteopontin (1.44 RFI) but suppressed matrix metalloproteinase-9 (0.6 RFI) compared to the control. Combined thermal and tensile stress increased vascular endothelial growth factor (VEGF) secretion into the culture supernatant (1.54-fold increase compared to the control). Therefore, combined thermal and mechanical stress preconditioning can enhance HSP induction and influence protein expression important for bone tissue healing.

  18. No effect of bone morphogenetic protein-7 (OP-1) on the incorporation of impacted bone grafts in a realistic acetabular model.

    NARCIS (Netherlands)

    Buma, P.; Arts, J.J.C.; Gardeniers, J.W.M.; Verdonschot, N.J.J.; Schreurs, B.W.

    2008-01-01

    Bone morphogenetic proteins (BMPs) accelerate bone repair in experimental and clinical conditions. Impacted Morsellized Cancellous Bone grafts (MCB) are successfully used to reconstruct bone defects after failed hip implants. The main question in this study was if BMP-7 (OP-1) mixed with MCB could a

  19. Quantitative sandwich ELISA for determination of traces of hazelnut (Corylus avellana) protein in complex food matrixes.

    Science.gov (United States)

    Holzhauser, T; Vieths, S

    1999-10-01

    A hazelnut-specific sandwich-type ELISA based on polyclonal antisera was developed for detection of hidden hazelnut protein residues in complex food matrixes. In the absence of a food matrix, extractable protein from different native and toasted hazelnuts was detected at rates of 94 +/- 13 and 96 +/- 7% applying standards prepared from native and toasted hazelnuts, respectively. From complex food matrixes, 0.001-10% of hazelnut was recovered between 67 and 132%, in average by 106 +/- 17%. Depending on the food matrix, hazelnut protein could be detected down to the ppb (ng/g) level. Intraassay precision was hazelnut >/= 0.001% and interassay precision was hazelnut >/= 0.01%. In 12 of 28 commercial food products without labeling or declaration of hazelnut components, between 2 and 421 ppm of hazelnut protein was detected, demonstrating a remarkable presence of potentially allergenic hazelnut protein "hidden" in commercial food products.

  20. Structural and contractile proteins. Part E: Extracellular matrix

    Energy Technology Data Exchange (ETDEWEB)

    Cunningham, L.W.

    1987-01-01

    This book contains 20 papers. Some of the titles are: Characterization of Pro-..cap alpha..2(I) Collagen Gene Mutation by Nuclease S/sub 1/ Mapping: Analysis of Cytoplasmic and Nuclear Messenger RNA in Fibroblasts from Patients with Type I Osteogenesis Imperfecta; R-Loop Analysis of Procollagen Messenger RNA from the Assessment of Human Collagen Mutations; and Bone Glycoproteins.

  1. Cartilage tissue engineering of nasal septal chondrocyte-macroaggregates in human demineralized bone matrix.

    Science.gov (United States)

    Liese, Juliane; Marzahn, Ulrike; El Sayed, Karym; Pruss, Axel; Haisch, Andreas; Stoelzel, Katharina

    2013-06-01

    Tissue Engineering is an important method for generating cartilage tissue with isolated autologous cells and the support of biomaterials. In contrast to various gel-like biomaterials, human demineralized bone matrix (DBM) guarantees some biomechanical stability for an application in biomechanically loaded regions. The present study combined for the first time the method of seeding chondrocyte-macroaggregates in DBM for the purpose of cartilage tissue engineering. After isolating human nasal chondrocytes and creating a three-dimensional macroaggregate arrangement, the DBM was cultivated in vitro with the macroaggregates. The interaction of the cells within the DBM was analyzed with respect to cell differentiation and the inhibitory effects of chondrocyte proliferation. In contrast to chondrocyte-macroaggregates in the cell-DBM constructs, morphologically modified cells expressing type I collagen dominated. The redifferentiation of chondrocytes, characterized by the expression of type II collagen, was only found in low amounts in the cell-DBM constructs. Furthermore, caspase 3, a marker for apoptosis, was detected in the chondrocyte-DBM constructs. In another experimental setting, the vitality of chondrocytes as related to culture time and the amount of DBM was analyzed with the BrdU assay. Higher amounts of DBM tended to result in significantly higher proliferation rates of the cells within the first 48 h. After 96 h, the vitality decreased in a dose-dependent fashion. In conclusion, this study provides the proof of concept of chondrocyte-macroaggregates with DBM as an interesting method for the tissue engineering of cartilage. The as-yet insufficient redifferentiation of the chondrocytes and the sporadic initiation of apoptosis will require further investigations.

  2. Combined role of type IX collagen and cartilage oligomeric matrix protein in cartilage matrix assembly: Cartilage oligomeric matrix protein counteracts type IX collagen-induced limitation of cartilage collagen fibril growth in mouse chondrocyte cultures

    NARCIS (Netherlands)

    Blumbach, K.; Bastiaansen-Jenniskens, Y.M.; Groot, J. de; Paulsson, M.; Osch, G.J.V.M. van; Zaucke, F.

    2009-01-01

    Objective. Defects in the assembly and composition of cartilage extracellular matrix are likely to result in impaired matrix integrity and increased susceptibility to cartilage degeneration. The aim of this study was to determine the functional interaction of the collagen fibril-associated proteins

  3. Combined role of type IX collagen and cartilage oligomeric matrix protein in cartilage matrix assembly: Cartilage oligomeric matrix protein counteracts type IX collagen-induced limitation of cartilage collagen fibril growth in mouse chondrocyte cultures

    NARCIS (Netherlands)

    Blumbach, K.; Bastiaansen-Jenniskens, Y.M.; Groot, J. de; Paulsson, M.; Osch, G.J.V.M. van; Zaucke, F.

    2009-01-01

    Objective. Defects in the assembly and composition of cartilage extracellular matrix are likely to result in impaired matrix integrity and increased susceptibility to cartilage degeneration. The aim of this study was to determine the functional interaction of the collagen fibril-associated proteins

  4. Use of a graft of demineralized bone matrix along with TGF-β1 leads to an early bone repair in dogs.

    Science.gov (United States)

    Servin-Trujillo, Miguel Angel; Reyes-Esparza, Jorge Alberto; Garrido-Fariña, German; Flores-Gazca, Enrique; Osuna-Martinez, Ulises; Rodriguez-Fragoso, Lourdes

    2011-09-01

    Tibia fractures are common in small animal practice. Over the past decade, improvements to animal internal fracture fixation have been developed. TGF-β1 has been shown to be crucial in the development, induction and repair of bone. In present study, we investigate the effect of local application of a graft demineralized bone matrix (DBM) along with TGF-β1 in a model of open osteotomy induced experimentally in dogs. Tibia fracture was brought about by using an open osteotomy model in young male dogs. Fracture repair was evaluated by a histological and biochemical analysis. Collagen content, proteolytic activity and urokinase-type plasminogen activator (uPA) expression were analyzed at the end of the study. Radiographic analysis, alkaline phosphatase and hematological evaluation were performed weekly. At the fifth week, there was an improvement and restoration of bone architecture in animals treated with a graft containing TGF-β1 (5 ng/ml) compared with the control and graft groups, as was evidenced by the presence of an early formation of wide callus and bone regeneration. In addition, local application of TGF-β1 led to an increase in collagen and proteolytic activity. More immunopositive osteoclast and mesenchymal cells were found in bone tissue from animals treated with TGF-β1 as compared with the control group. No changes in alkaline phosphatase, hematological and clinical parameters were observed. This study shows that the combined use of DBM along with TGF-β1 is able to improve and accelerate the bone repair.

  5. Sequence preservation of osteocalcin protein and mitochondrial DNA in bison bones older than 55 ka

    Science.gov (United States)

    Nielsen-Marsh, Christina M.; Ostrom, Peggy H.; Gandhi, Hasand; Shapiro, Beth; Cooper, Alan; Hauschka, Peter V.; Collins, Matthew J.

    2002-12-01

    We report the first complete sequences of the protein osteocalcin from small amounts (20 mg) of two bison bone (Bison priscus) dated to older than 55.6 ka and older than 58.9 ka. Osteocalcin was purified using new gravity columns (never exposed to protein) followed by microbore reversed-phase high-performance liquid chromatography. Sequencing of osteocalcin employed two methods of matrix-assisted laser desorption ionization mass spectrometry (MALDI-MS): peptide mass mapping (PMM) and post-source decay (PSD). The PMM shows that ancient and modern bison osteocalcin have the same mass to charge (m/z) distribution, indicating an identical protein sequence and absence of diagenetic products. This was confirmed by PSD of the m/z 2066 tryptic peptide (residues 1 19); the mass spectra from ancient and modern peptides were identical. The 129 mass unit difference in the molecular ion between cow (Bos taurus) and bison is caused by a single amino-acid substitution between the taxa (Trp in cow is replaced by Gly in bison at residue 5). Bison mitochondrial control region DNA sequences were obtained from the older than 55.6 ka fossil. These results suggest that DNA and protein sequences can be used to directly investigate molecular phylogenies over a considerable time period, the absolute limit of which is yet to be determined.

  6. The effect of SDF-1α on low dose BMP-2 mediated bone regeneration by release from heparinized mineralized collagen type I matrix scaffolds in a murine critical size bone defect model.

    Science.gov (United States)

    Zwingenberger, Stefan; Langanke, Robert; Vater, Corina; Lee, Geoffrey; Niederlohmann, Eik; Sensenschmidt, Markus; Jacobi, Angela; Bernhardt, Ricardo; Muders, Michael; Rammelt, Stefan; Knaack, Sven; Gelinsky, Michael; Günther, Klaus-Peter; Goodman, Stuart B; Stiehler, Maik

    2016-09-01

    The treatment of critical size bone defects represents a challenge. The growth factor bone morphogenetic protein 2 (BMP-2) is clinically established but has potentially adverse effects when used at high doses. The aim of this study was to evaluate if stromal derived factor-1 alpha (SDF-1α) and BMP-2 released from heparinized mineralized collagen type I matrix (MCM) scaffolds have a cumulative effect on bone regeneration. MCM scaffolds were functionalized with heparin, loaded with BMP-2 and/or SDF-1α and implanted into a murine critical size femoral bone defect (control group, low dose BMP-2 group, low dose BMP-2 + SDF-1α group, and high dose BMP-2 group). After 6 weeks, both the low dose BMP-2 + SDF-1α group (5.8 ± 0.6 mm³, p = 0.0479) and the high dose BMP-2 group (6.5 ± 0.7 mm³, p = 0.008) had a significantly increased regenerated bone volume compared to the control group (4.2 ± 0.5 mm³). There was a higher healing score in the low dose BMP-2 + SDF-1α group (median grade 8; Q1-Q3 7-9; p = 0.0357) than in the low dose BMP-2 group (7; Q1-Q3 5-9) histologically. This study showed that release of BMP-2 and SDF-1α from heparinized MCM scaffolds allows for the reduction of the applied BMP-2 concentration since SDF-1α seems to enhance the osteoinductive potential of BMP-2. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 104A: 2126-2134, 2016.

  7. 人纳米脱钙骨基质复合物的理化性质和安全性%Physicochemical property and safety of nanometer human demineralized bone matrix composite

    Institute of Scientific and Technical Information of China (English)

    房雷; 陈雄生; 黄凯; 周盛源; 朱巍; 王辉; 邵将; 贾连顺

    2013-01-01

    背景:脱钙骨基质和骨形态发生蛋白已被证实具有良好的骨诱导性,但有关纳米脱钙骨基质的研究较少,其理化性质和生物安全性尚不明确。  目的:在前期实验制备人纳米脱钙骨基质的基础上加载重组人骨形态发生蛋白2,分析人纳米脱钙骨基质复合重组人骨形态发生蛋白2的理化性质及生物安全性。  方法:采用改良Urist法制备人脱钙骨基质,并进行纳米化处理,再将骨形态发生蛋白2与其按特定比例混合,冻干塑型行以下实验:①热源实验:将材料浸提液经耳静脉注入兔体内。②毒性实验:将材料浸提液与生理盐水分别经尾静脉注入小白鼠体内。③植入实验:在兔两侧后肢肌肉内分别植入实验材料和β-磷酸三钙。  结果与结论:冻干塑型后,纳米人脱钙骨基质材料表面致密,孔隙直径100-400μm,孔隙分布欠均匀,孔隙率小于30%,以碳、氧和氮为主要元素组成。人纳米脱钙骨基质复合重组人骨形态发生蛋白2材料无热源效应,注射后未见兔体温有明显波动。急性全身毒性实验结果表明人纳米脱钙骨基质复合重组人骨形态发生蛋白2材料符合国家相关规定,注射后未见小鼠出现明显毒性反应。人纳米脱钙骨基质复合重组人骨形态发生蛋白2材料植入兔体内的炎症反应明显轻于β-磷酸三钙植入后的反应。结果表明人纳米脱钙骨基质复合重组人骨形态发生蛋白2是一种无毒、组织相容性好、生物利用度高、炎症反应轻的纳米同种异体骨移植替代物。%BACKGROUND: Demineralized bone matrix and bone morphogenetic protein have been shown to have good bone induction, but less studies concerned nanometer demineralized bone matrix. Its physical and chemical properties and biological security are not yet clear. OBJECTIVE:On the basis of preparing the nanometer human demineralized bone

  8. Extracellular matrix production by nucleus pulposus and bone marrow stem cells in response to altered oxygen and glucose microenvironments.

    Science.gov (United States)

    Naqvi, Syeda M; Buckley, Conor T

    2015-12-01

    Bone marrow (BM) stem cells may be an ideal source of cells for intervertebral disc (IVD) regeneration. However, the harsh biochemical microenvironment of the IVD may significantly influence the biological and metabolic vitality of injected stem cells and impair their repair potential. This study investigated the viability and production of key matrix proteins by nucleus pulposus (NP) and BM stem cells cultured in the typical biochemical microenvironment of the IVD consisting of altered oxygen and glucose concentrations. Culture-expanded NP cells and BM stem cells were encapsulated in 1.5% alginate and ionically crosslinked to form cylindrical hydrogel constructs. Hydrogel constructs were maintained under different glucose concentrations (1, 5 and 25 mM) and external oxygen concentrations (5 and 20%). Cell viability was measured using the Live/Dead® assay and the production of sulphated glycosaminoglycans (sGAG), and collagen was quantified biochemically and histologically. For BM stem cells, IVD-like micro-environmental conditions (5 mM glucose and 5% oxygen) increased the accumulation of sGAG and collagen. In contrast, low glucose conditions (1 mM glucose) combined with 5% external oxygen concentration promoted cell death, inhibiting proliferation and the accumulation of sGAG and collagen. NP-encapsulated alginate constructs were relatively insensitive to oxygen concentration or glucose condition in that they accumulated similar amounts of sGAG under all conditions. Under IVD-like microenvironmental conditions, NP cells were found to have a lower glucose consumption rate compared with BM cells and may in fact be more suitable to adapt and sustain the harsh microenvironmental conditions. Considering the highly specialised microenvironment of the central NP, these results indicate that IVD-like concentrations of low glucose and low oxygen are critical and influential for the survival and biological behaviour of stem cells. Such findings may promote and accelerate

  9. Effect of Emdogain enamel matrix derivative and BMP-2 on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells.

    Science.gov (United States)

    Fawzy El-Sayed, Karim M; Dörfer, Christof; Ungefroren, Hendrick; Kassem, Neemat; Wiltfang, Jörg; Paris, Sebastian

    2014-07-01

    The objective of this study was to evaluate the effect of Emdogain (Enamel Matrix Derivative, EMD) and Bone Morphogenetic Protein-2 (BMP-2), either solely or in combination, on the gene expression and mineralized nodule formation of alveolar bone proper-derived stem/progenitor cells. Stem/progenitor cells were isolated from human alveolar bone proper, magnetically sorted using STRO-1 antibodies, characterized flowcytometrically for their surface markers' expression, and examined for colony formation and multilineage differentiation potential. Subsequently, cells were treated over three weeks with 100 μg/ml Emdogain (EMD-Group), or 100 ng/ml BMP-2 (BMP-Group), or a combination of 100 ng/ml BMP-2 and 100 μg/ml Emdogain (BMP/EMD-Group). Unstimulated stem/progenitor cells (MACS(+)-Group) and osteoblasts (OB-Group) served as controls. Osteogenic gene expression was analyzed using RTq-PCR after 1, 2 and 3 weeks (N = 3/group). Mineralized nodule formation was evaluated by Alizarin-Red staining. BMP and EMD up-regulated the osteogenic gene expression. The BMP Group showed significantly higher expression of Collagen-I, III, and V, Alkaline phosphatase and Osteonectin compared to MACS(+)- and OB-Group (p Emdogain and BMP-2 up-regulate the osteogenic gene expression of stem/progenitor cells. The combination of BMP-2 and Emdogain showed no additive effect and would not be recommended for a combined clinical stimulation.

  10. Guaifenesin stone matrix proteomics: a protocol for identifying proteins critical to stone formation.

    Science.gov (United States)

    Kolbach-Mandel, A M; Mandel, N S; Cohen, S R; Kleinman, J G; Ahmed, F; Mandel, I C; Wesson, J A

    2017-04-01

    Drug-related kidney stones are a diagnostic problem, since they contain a large matrix (protein) fraction and are frequently incorrectly identified as matrix stones. A urine proteomics study patient produced a guaifenesin stone during her participation, allowing us to both correctly diagnose her disease and identify proteins critical to this drug stone-forming process. The patient provided three random midday urine samples for proteomics studies; one of which contained stone-like sediment with two distinct fractions. These solids were characterized with optical microscopy and Fourier transform infrared spectroscopy. Immunoblotting and quantitative mass spectrometry were used to quantitatively identify the proteins in urine and stone matrix. Infrared spectroscopy showed that the sediment was 60 % protein and 40 % guaifenesin and its metabolite guaiacol. Of the 156 distinct proteins identified in the proteomic studies, 49 were identified in the two stone-components with approximately 50 % of those proteins also found in this patient's urine. Many proteins observed in this drug-related stone have also been reported in proteomic matrix studies of uric acid and calcium containing stones. More importantly, nine proteins were highly enriched and highly abundant in the stone matrix and 8 were reciprocally depleted in urine, suggesting a critical role for these proteins in guaifenesin stone formation. Accurate stone analysis is critical to proper diagnosis and treatment of kidney stones. Many matrix proteins were common to all stone types, but likely not related to disease mechanism. This protocol defined a small set of proteins that were likely critical to guaifenesin stone formation based on their high enrichment and high abundance in stone matrix, and it should be applied to all stone types.

  11. Growth of thin films of low molecular weight proteins by matrix assisted pulsed laser evaporation (MAPLE)

    DEFF Research Database (Denmark)

    Matei, Andreea; Schou, Jørgen; Constantinescu, C.;

    2011-01-01

    Thin films of lysozyme and myoglobin grown by matrix assisted pulsed laser evaporation (MAPLE) from a water ice matrix have been investigated. The deposition rate of these two low molecular weight proteins (lysozyme: 14307 amu and myoglobin: 17083 amu) exhibits a maximum of about 1–2 ng/cm2 per...

  12. Biomimetic Mineralization on a Macroporous Cellulose-Based Matrix for Bone Regeneration

    OpenAIRE

    2013-01-01

    The aim of this study is to investigate the biomimetic mineralization on a cellulose-based porous matrix with an improved biological profile. The cellulose matrix was precalcified using three methods: (i) cellulose samples were treated with a solution of calcium chloride and diammonium hydrogen phosphate; (ii) the carboxymethylated cellulose matrix was stored in a saturated calcium hydroxide solution; (iii) the cellulose matrix was mixed with a calcium silicate solution in order to introduce ...

  13. Extracellular matrix proteins: A positive feedback loop in lung fibrosis?

    NARCIS (Netherlands)

    Blaauboer, M.E.; Boeijen, F.R.; Emson, C.L.; Turner, S.M.; Zandieh-Doulabi, B.; Hanemaaijer, R.; Smit, T.H.; Stoop, R.; Everts, V.

    2014-01-01

    Lung fibrosis is characterized by excessive deposition of extracellular matrix. This not only affects tissue architecture and function, but it also influences fibroblast behavior and thus disease progression. Here we describe the expression of elastin, type V collagen and tenascin C during the

  14. Matrix Gla Protein polymorphism, but not concentrations, is associated with radiographic hand osteoarthritis

    Science.gov (United States)

    Objective. Factors associated with mineralization and osteophyte formation in osteoarthritis (OA) are incompletely understood. Genetic polymorphisms of matrix Gla protein (MGP), a mineralization inhibitor, have been associated clinically with conditions of abnormal calcification. We therefore evalua...

  15. Detection of HOCl-mediated protein oxidation products in the extracellular matrix of human atherosclerotic plaques

    DEFF Research Database (Denmark)

    Woods, Alan A; Linton, Stuart M; Davies, Michael Jonathan

    2003-01-01

    for 83-96% of the total oxidized protein side-chain products detected in these plaques. Oxidation of matrix components extracted from healthy artery tissue, and model proteins, with reagent HOCl is shown to give rise to a similar pattern of products to those detected in advanced human lesions......Oxidation is believed to play a role in atherosclerosis. Oxidized lipids, sterols and proteins have been detected in early, intermediate and advanced human lesions at elevated levels. The spectrum of oxidized side-chain products detected on proteins from homogenates of advanced human lesions has...... by activated monocytes (and possibly macrophages) and is a highly basic protein, it would be expected to associate with polyanions such as the glycosaminoglycans of the extracellular matrix, and might result in damage being localized at such sites. In this study proteins extracted from extracellular matrix...

  16. Interplay of matrix stiffness and protein tethering in stem cell differentiation

    Science.gov (United States)

    Wen, Jessica H.; Vincent, Ludovic G.; Fuhrmann, Alexander; Choi, Yu Suk; Hribar, Kolin C.; Taylor-Weiner, Hermes; Chen, Shaochen; Engler, Adam J.

    2014-10-01

    Stem cells regulate their fate by binding to, and contracting against, the extracellular matrix. Recently, it has been proposed that in addition to matrix stiffness and ligand type, the degree of coupling of fibrous protein to the surface of the underlying substrate, that is, tethering and matrix porosity, also regulates stem cell differentiation. By modulating substrate porosity without altering stiffness in polyacrylamide gels, we show that varying substrate porosity did not significantly change protein tethering, substrate deformations, or the osteogenic and adipogenic differentiation of human adipose-derived stromal cells and marrow-derived mesenchymal stromal cells. Varying protein-substrate linker density up to 50-fold changed tethering, but did not affect osteogenesis, adipogenesis, surface-protein unfolding or underlying substrate deformations. Differentiation was also unaffected by the absence of protein tethering. Our findings imply that the stiffness of planar matrices regulates stem cell differentiation independently of protein tethering and porosity.

  17. Structural and functional features of a collagen-binding matrix protein from the mussel byssus.

    Science.gov (United States)

    Suhre, Michael H; Gertz, Melanie; Steegborn, Clemens; Scheibel, Thomas

    2014-02-26

    Blue mussels adhere to surfaces by the byssus, a holdfast structure composed of individual threads representing a collagen fibre reinforced composite. Here, we present the crystal structure and function of one of its matrix proteins, the proximal thread matrix protein 1, which is present in the proximal section of the byssus. The structure reveals two von Willebrand factor type A domains linked by a two-β-stranded linker yielding a novel structural arrangement. In vitro, the protein binds heterologous collagens with high affinity and affects collagen assembly, morphology and arrangement of its fibrils. By providing charged surface clusters as well as insufficiently coordinated metal ions, the proximal thread matrix protein 1 might interconnect other byssal proteins and thereby contribute to the integrity of the byssal threads in vivo. Moreover, the protein could be used for adjusting the mechanical properties of collagen materials, a function likely important in the natural byssus.

  18. Cartilage oligomeric matrix protein in patients with juvenile idiopathic arthritis: relation to growth and disease activity

    DEFF Research Database (Denmark)

    Bjørnhart, Birgitte; Juul, Anders; Nielsen, Susan

    2009-01-01

    OBJECTIVE: Cartilage oligomeric matrix protein (COMP) has been identified as a prognostic marker of progressive joint destruction in rheumatoid arthritis. In this population based study we evaluated associations between plasma concentrations of COMP, disease activity, and growth velocity in patie......OBJECTIVE: Cartilage oligomeric matrix protein (COMP) has been identified as a prognostic marker of progressive joint destruction in rheumatoid arthritis. In this population based study we evaluated associations between plasma concentrations of COMP, disease activity, and growth velocity...

  19. Altered expression of nuclear matrix proteins in etoposide induced apoptosis in HL-60 cells

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The events of cell death and the expression of nuclear matrix protein(NMP)have been investigated in a promyelocytic leukemic cell line HL-60 induced with etoposide.By means of TUNEL assay,the nuclei displayed a characteristic morphology change,and the amount of apoptotic cells increased early and reached maximun about 39% after treatment with etoposide for 2 h.Nucleosomal DNA fragmentation was observed after treatment for 4 h.The morphological change of HL-60 cells,thus,occurred earlier than the appearance of DNA ladder.Total nuclear matrix proteins were analyzed by 2-dimensional gel electrophoresis.Differential expression of 59 nuclear matrix proteins was found in 4 h etoposide treated cells.Western blotting was then performed on three nuclear matrix acssociated proteins,PML,HSC70 and NuMA.The expression of the suppressor PML protein and heat shock protein HSC70 were significantly upregulated after etoposide treatment,while NuMA,a nuclear mitotic apparatus protein,was down regulated.These results demonstrate that significant biochemical alterations in nuclear matrix proteins take place during the apoptotic process.

  20. 弹状病毒基质蛋白(Matrix protein)的研究进展%Advances in research on rhabdovirus matrix protein

    Institute of Scientific and Technical Information of China (English)

    杨振慧; 葛均青; 龚晖; 林天龙

    2011-01-01

    Rhabdovirus contains a linear, negative-sense and single-stranded RNA genome. It encodes 5 structural proteins, including nucleoprotein (N), phosphoprotein (P), matrix protein (M), glycoprotein (G) and RNA polymerase (L). The matrix protein takes multi-function in the virus infection cycle, including participating in virus budding and assembly, inhibiting protein nucleocytoplasmic transport, and regulating host gene transcription and inducing cell apoptosis. Recent advances in the research on the matrix protein are reviewed.%弹状病毒的基因组由1段线性单股不分节的负链RNA组成,主要编码5个结构蛋白,分别为核蛋白(Nucleoprotein,N)、磷蛋白(Phosphoprotein,P)、基质蛋白(Mattix protein,M)、糖蛋白(Glycoprotein,G)和RNA聚合酶(RNA polymerase,L).其中,基质蛋白(M)是一种多功能性蛋白,在病毒的侵染过程中参与病毒粒子的出芽、装配,抑制核质运输,调控宿主的基因转录,诱导细胞凋亡等.本文综述M蛋白的研究进展.

  1. Novel Approaches to Bone Grafting: Porosity, Bone Morphogenetic Proteins, Stem Cells, and the Periosteum

    OpenAIRE

    Petrochenko, Peter; Narayan, Roger J.

    2010-01-01

    The disadvantages involving the use of a patient’s own bone as graft material have led surgeons to search for alternative materials. In this review, several characteristics of a successful bone graft material are discussed. In addition, novel synthetic materials and natural bone graft materials are being considered. Various factors can determine the success of a bone graft substitute. For example, design considerations such as porosity, pore shape, and interconnection play significant roles i...

  2. Predicting protein-ligand affinity with a random matrix framework

    OpenAIRE

    Lee, Alpha Albert; Brenner, MP; Colwell, Lucy Jane

    2016-01-01

    Rapid determination of whether a candidate compound will bind to a particular target receptor remains a stumbling block in drug discovery. We use an approach inspired by random matrix theory to decompose the known ligand set of a target in terms of orthogonal "signals" of salient chemical features, and distinguish these from the much larger set of ligand chemical features that are not relevant for binding to that particular target receptor. After removing the noise caused by finite sampling, ...

  3. [Electron transfer between globular proteins. Evaluation of a matrix element].

    Science.gov (United States)

    Lakhno, V D; Chuev, G N; Ustinin, M N

    1998-01-01

    The dependence of the matrix element of the probability of interprotein electron transfer on the mutual orientation of the donor and acceptor centers and the distance between them was calculated. The calculations were made under the assumption that electron transfer proceeds mainly by a collective excitation of polaron nature, like a solvated electron state. The results obtained are consistent with experimental data and indicate the nonexponential behavior of this dependence in the case when the distance transfer is less than 20 A.

  4. Stroma and extracellular matrix proteins in canine tumours

    OpenAIRE

    2004-01-01

    In this thesis, studies on temporal and spatial changes in stromal cells and extracellular matrix (ECM) molecules in canine gastrointestinal (GIT) tumours and canine transmissible venereal (CTVT) tumours are described. The mechanisms involved in the phenotypic transformation of fibroblasts to myofibroblasts, and ECM changes were investigated. We found that the myofibroblast is the most common stromal cell in canine GIT epithelial tumours and most likely originated from pre-existing fibroblast...

  5. Genomic analysis of a spontaneous model of breast cancer metastasis to bone reveals a role for the extracellular matrix.

    Science.gov (United States)

    Eckhardt, Bedrich L; Parker, Belinda S; van Laar, Ryan K; Restall, Christina M; Natoli, Anthony L; Tavaria, Michael D; Stanley, Kym L; Sloan, Erica K; Moseley, Jane M; Anderson, Robin L

    2005-01-01

    A clinically relevant model of spontaneous breast cancer metastasis to multiple sites, including bone, was characterized and used to identify genes involved in metastatic progression. The metastatic potential of several genetically related tumor lines was assayed using a novel real-time quantitative RT-PCR assay of tumor burden. Based on this assay, the tumor lines were categorized as nonmetastatic (67NR), weakly metastatic to lymph node (168FARN) or lung (66cl4), or highly metastatic to lymph node, lung, and bone (4T1.2 and 4T1.13). In vitro assays that mimic stages of metastasis showed that highly metastatic tumors lines were more adhesive, invasive, and migratory than the less metastatic lines. To identify metastasis-related genes in this model, each metastatic tumor was array profiled against the nonmetastatic 67NR using 15,000 mouse cDNA arrays. A significant proportion of genes relating to the extracellular matrix had elevated expression in highly metastatic tumors. The role of one of these genes, POEM, was further investigated in the model. In situ hybridization showed that POEM expression was specific to the tumor epithelium of highly metastatic tumors. Decreased POEM expression in 4T1.2 tumors significantly inhibited spontaneous metastasis to the lung, bone, and kidney. Taken together, our data support a role for the extracellular matrix in metastatic progression and describe, for the first time, a role for POEM in this process.

  6. Self-assembled composite matrix in a hierarchical 3-D scaffold for bone tissue engineering

    DEFF Research Database (Denmark)

    Chen, Muwan; Le, Dang Quang Svend; Baatrup, Anette

    2011-01-01

    It is of high clinical relevance in bone tissue engineering that scaffolds promote a high seeding efficiency of cells capable of osteogenic differentiation, such as human bone marrow-derived mesenchymal stem cells (hMSCs). We evaluated the effects of a novel polycaprolactone (PCL) scaffold on h...

  7. Collagen osteoid-like model allows kinetic gene expression studies of non-collagenous proteins in relation with mineral development to understand bone biomineralization.

    Science.gov (United States)

    Silvent, Jérémie; Nassif, Nadine; Helary, Christophe; Azaïs, Thierry; Sire, Jean-Yves; Guille, Marie Madeleine Giraud

    2013-01-01

    Among persisting questions on bone calcification, a major one is the link between protein expression and mineral deposition. A cell culture system is here proposed opening new integrative studies on biomineralization, improving our knowledge on the role played by non-collagenous proteins in bone. This experimental in vitro model consisted in human primary osteoblasts cultured for 60 days at the surface of a 3D collagen scaffold mimicking an osteoid matrix. Various techniques were used to analyze the results at the cellular and molecular level (adhesion and viability tests, histology and electron microscopy, RT- and qPCR) and to characterize the mineral phase (histological staining, EDX, ATG, SAED and RMN). On long term cultures human bone cells seeded on the osteoid-like matrix displayed a clear osteoblast phenotype as revealed by the osteoblast-like morphology, expression of specific protein such as alkaline phosphatase and expression of eight genes classically considered as osteoblast markers, including BGLAP, COL1A1, and BMP2. Von Kossa and alizarine red allowed us to identify divalent calcium ions at the surface of the matrix, EDX revealed the correct Ca/P ratio, and SAED showed the apatite crystal diffraction pattern. In addition RMN led to the conclusion that contaminant phases were absent and that the hydration state of the mineral was similar to fresh bone. A temporal correlation was established between quantified gene expression of DMP1 and IBSP, and the presence of hydroxyapatite, confirming the contribution of these proteins to the mineralization process. In parallel a difference was observed in the expression pattern of SPP1 and BGLAP, which questioned their attributed role in the literature. The present model opens new experimental possibilities to study spatio-temporal relations between bone cells, dense collagen scaffolds, NCPs and hydroxyapatite mineral deposition. It also emphasizes the importance of high collagen density environment in bone cell

  8. Gradual downhill running improves age-related skeletal muscle and bone weakness: implication of autophagy and bone morphogenetic proteins.

    Science.gov (United States)

    Kim, Jeong-Seok; Lee, Young-Hee; Yi, Ho-Keun

    2016-12-01

    What is the central question of this study? Exercise training by running has an effect on age-related muscle and bone wasting that improves physical activity and quality of life in the elderly. However, the effect of downhill running on age-related muscle and bone wasting, and its mechanisms, are unclear. What is the main finding and its importance? Gradual downhill running can improve skeletal muscle growth and bone formation by enhancing autophagy and bone morphogenetic protein signalling in aged rats. Therefore, downhill running exercise might be a practical intervention to improve skeletal muscle and bone protection in the elderly. Recent evidence suggests that autophagy and the bone morphogenetic protein (BMP) signalling pathway regulate skeletal muscle growth and bone formation in aged rats. However, the effect of downhill running on muscle growth and bone formation is not well understood. Thus, we investigated the effect of downhill and uphill running on age-related muscle and bone weakness. Young and late middle-aged rats were randomly assigned to control groups (young, YC; and late middle-aged, LMC) and two types of running training groups (late middle-aged downhill, LMD; and late middle-aged uphill, LMU). Training was progressively carried out on a treadmill at a speed of 21 m min(-1) with a slope of +10 deg for uphill training versus 16 m min(-1) with a slope of -16 deg for downhill training, both for 60 min day(-1) , 5 days week(-1) for 8 weeks. Downhill and uphill training increased autophagy-related protein 5, microtubule-associated protein light chain, Beclin-1 and p62 proteins in aged rats. In addition, superoxide dismutase, haem oxygenase-1 and the BMP signalling pathway were elevated. Phosphorylation of mammalian target of rapamycin and myogenic differentiation were increased significantly in the LMD and LMU groups. Consequently, in the femur, BMP-2, BMP-7 and autophagy molecules were highly expressed in the LMD and LMU groups. These

  9. Bone Formation in a Rat Tibial Defect Model Using Carboxymethyl Cellulose/BioC/Bone Morphogenic Protein-2 Hybrid Materials

    Directory of Open Access Journals (Sweden)

    Sang-Heon Song

    2014-01-01

    Full Text Available The objective of this study was to assess whether carboxymethyl cellulose- (CMC- based hydrogel containing BioC (biphasic calcium phosphate (BCP; tricalcium phosphate (TCP : hydroxyapatite (Hap = 70 : 30 and bone morphogenic protein-2 (BMP-2 led to greater bone formation than CMC-based hydrogel containing BioC without BMP-2. In order to demonstrate bone formation at 4 and 8 weeks, plain radiographs, microcomputed tomography (micro-CT evaluation, and histological studies were performed after implantation of all hybrid materials on an 8 mm defect of the right tibia in rats. The plain radiographs and micro-CT analyses revealed that CMC/BioC/BMP-2 (0.5 mg led to much greater mineralization at 4 and 8 weeks than did CMC/BioC or CMC/Bio/BMP-2 (0.1 mg. Likewise, bone formation and bone remodeling studies revealed that CMC/BioC/BMP-2 (0.5 mg led to a significantly greater amount of bone formation and bone remodeling at 4 and 8 weeks than did CMC/BioC or CMC/BioC/BMP-2 (0.1 mg. Histological studies revealed that mineralized bone tissue was present around the whole circumference of the defect site with CMC/BioC/BMP-2 (0.5 mg but not with CMC/BioC or CMC/BioC/BMP-2 (0.1 mg at 4 and 8 weeks. These results suggest that CMC/BioC/BMP-2 hybrid materials induced greater bone formation than CMC/BioC hybrid materials. Thus, CMC/BioC/BMP-2 hybrid materials may be used as an injectable substrate to regenerate bone defects.

  10. Kinetics of protein-protein complex coacervation and biphasic release of salbutamol sulfate from coacervate matrix.

    Science.gov (United States)

    Tiwari, Ananya; Bindal, Sonal; Bohidar, H B

    2009-01-12

    Turbidimetric titration was used to initiate associative intermolecular interactions between a pair of protein molecules, gelatin-A and gelatin-B, having complementary charges that led to pH-induced liquid-liquid phase separation and the formation of complex coacervate. The stoichiometric binding ratio was found to be [gelatin-A]/[gelatin-B]=3:2. The size of soluble intermolecular aggregates present in the supernatant exhibited interesting time-dependent coacervation because of residual electrostatic interactions. Dynamic light scattering and turbidity studies provided a systematic account of coacervation behavior. Rheology studies attributed the softening of the coacervate matrix to the presence of encapsulated salbutamol sulfate. The in vitro drug release kinetics was probed in simulated gastric fluid medium at physiological temperature (37 degrees C), which showed biphasic behavior. The initial release kinetics exhibited an exponential growth to saturation behavior, followed by a slower logarithmic release process.

  11. Effects of bone morphogenetic protein-2 on bone cells in primary culture: immunohistochemical and electronmicroscopical studies

    Energy Technology Data Exchange (ETDEWEB)

    Schmitz, I.; Prochnow, N.; Mueller, K.M. [Berufsgenossenschaftliche Kliniken Bergmannsheil, Bochum (Germany). Inst. fuer Pathologie; Wiemann, M.; Schirrmacher, K.; Bingmann, D. [Essen Univ. (Germany). Inst. fuer Physiologie; Sebald, W. [Wuerzburg Univ. (Germany). Inst. fuer Physiologische Chemie II

    2001-02-01

    Bone morphogenetic protein 2 (BMP-2), among other morphogenetic effects on non osseous tissues, promotes bone formation in vivo. Therefore, BMP-2 may accelerate the integration of osseous implants. Although the effects of BMPs on cell proliferation have been studied extensively in vivo or in cell lines, little is published about effects on bone cells in primary cultures, especially on cell differentiation. As such information is a prerequisite to understand and to control effects of BMPs on cells at the surface of implant materials, the present experiments aimed to describe effects of BMP-2 on primary cultures derived from calvarial fragments of neonatal rats. The cells were stimulated with 50 nM BMP-2 added to the nutrient medium for 3 or 6 days. Light- and electronmicroscopical studies showed that cells in the sprouting zones were larger and more often spindle shaped. Stimulated cells had more nucleoli than control cells and the endoplasmic reticulum was widened. They retained properties of typical bone cells: An immunhistochemical analysis showed that stimulated cells increased the activity of alkaline phosphatase, they secreted collagen type I and to a minor extent collagen type III. In BMP-2 treated cells the pattern of cells stained for actin, desmin and vimentin hardly changed whereas extracellular fibronectin appeared to be less cross-linked in BMP-2 treated cultures. The distribution and labeling strength of osteocalcin, a specific marker protein of bone cells did not change markedly. After exposure to BMP-2 cells tended to detach from the cover slips. Electron microscopy showed a reduced number of cell processes possibly facilitating the detachment and/or mobility. Stimulated cells contained an increased number of lamellar bodies which may reflect an increased synthesis and/or membrane turnover. Staining of non-osseous cells with anti-CD68-or anti-myeloid antibodies revealed that the small percentage of these cells regularly occurring in primary cultures

  12. Detecting Protein-Protein Interactions with a Novel Matrix-Based Protein Sequence Representation and Support Vector Machines

    Directory of Open Access Journals (Sweden)

    Zhu-Hong You

    2015-01-01

    Full Text Available Proteins and their interactions lie at the heart of most underlying biological processes. Consequently, correct detection of protein-protein interactions (PPIs is of fundamental importance to understand the molecular mechanisms in biological systems. Although the convenience brought by high-throughput experiment in technological advances makes it possible to detect a large amount of PPIs, the data generated through these methods is unreliable and may not be completely inclusive of all possible PPIs. Targeting at this problem, this study develops a novel computational approach to effectively detect the protein interactions. This approach is proposed based on a novel matrix-based representation of protein sequence combined with the algorithm of support vector machine (SVM, which fully considers the sequence order and dipeptide information of the protein primary sequence. When performed on yeast PPIs datasets, the proposed method can reach 90.06% prediction accuracy with 94.37% specificity at the sensitivity of 85.74%, indicating that this predictor is a useful tool to predict PPIs. Achieved results also demonstrate that our approach can be a helpful supplement for the interactions that have been detected experimentally.

  13. Effect of enamel matrix derivative and parathyroid hormone on bone formation in standardized osseous defects: an experimental study in minipigs

    DEFF Research Database (Denmark)

    Jensen, Simon S; Chen, B; Bornstein, Michael M;

    2011-01-01

    or directly on the bone-forming cells. In addition, it is not known if the presentation of PTH by adding the amino acid sequence Arg-Gly-Asp (RGD) is essential for its osteopromotive effect. Local delivery of a bioactive substance at the right time and in the right concentration often constitutes a major...... challenge. Polyethylene glycol-based hydrogel (PEG) is a degradable vehicle developed for delivery of bioactive proteins. To enhance the mechanical stability of the PEG-bioactive substance complex, an osteoconductive bone substitute material is often needed....

  14. Development and Characterization of a Bioinspired Bone Matrix with Aligned Nanocrystalline Hydroxyapatite on Collagen Nanofibers

    Directory of Open Access Journals (Sweden)

    Hsi-Chin Wu

    2016-03-01

    Full Text Available Various kinds of three-dimensional (3D scaffolds have been designed to mimic the biological spontaneous bone formation characteristics by providing a suitable microenvironment for osteogenesis. In view of this, a natural bone-liked composite scaffold, which was combined with inorganic (hydroxyapatite, Hap and organic (type I collagen, Col phases, has been developed through a self-assembly process. This 3D porous scaffold consisting of a c-axis of Hap nanocrystals (nHap aligning along Col fibrils arrangement is similar to natural bone architecture. A significant increase in mechanical strength and elastic modulus of nHap/Col scaffold is achieved through biomimetic mineralization process when compared with simple mixture of collagen and hydroxyapatite method. It is suggested that the self-organization of Hap and Col produced in vivo could also be achieved in vitro. The oriented nHap/Col composite not only possesses bone-like microstructure and adequate mechanical properties but also enhances the regeneration and reorganization abilities of bone tissue. These results demonstrated that biomimetic nHap/Col can be successfully reconstructed as a bone graft substitute in bone tissue engineering.

  15. Influence of select extracellular matrix proteins on mesenchymal stem cell osteogenic commitment in three-dimensional contexts.

    Science.gov (United States)

    Becerra-Bayona, Silvia; Guiza-Arguello, Viviana; Qu, Xin; Munoz-Pinto, Dany J; Hahn, Mariah S

    2012-12-01

    Growth factors have been shown to be powerful mediators of mesenchymal stem cell (MSC) osteogenic differentiation. However, their use in tissue engineered scaffolds not only can be costly but also can induce undesired responses in surrounding tissues. Thus, the ability to specifically promote MSC osteogenic differentiation in the absence of exogenous growth factors via the manipulation of scaffold material properties would be beneficial. The current work examines the influence of select extracellular matrix (ECM) proteins on MSC osteogenesis toward the goal of developing scaffolds with intrinsically osteoinductive properties. Fibrinogen (FG), fibronectin (FN) and laminin-1 (LN) were chosen for evaluation due to their known roles in bone morphogenesis or bone fracture healing. These proteins were conjugated into poly(ethylene glycol) diacrylate (PEGDA) hydrogels and their effects on encapsulated 10T½ MSCs were evaluated. Specifically, following 1week of culture, mid-term markers of various MSC lineages were examined in order to assess the strength and specificity of the observed osteogenic responses. PEG-LN gels demonstrated increased levels of the osteogenic transcription factor osterix relative to day 0 levels. In addition, PEG-FG and PEG-LN gels were associated with increased deposition of bone ECM protein osteocalcin relative to PEG-FN gels and day 0. Importantly, the osteogenic response associated with FG and LN appeared to be specific in that markers for chondrocytic, smooth muscle cell and adipocytic lineages were not similarly elevated relative to day 0 in these gels. To gain insight into the integrin dynamics underlying the observed differentiation results, initial integrin adhesion and temporal alterations in cell integrin profiles were evaluated. The associated results suggest that α(2), α(v) and α(6) integrin subunits may play key roles in integrin-mediated osteogenesis.

  16. Successful treatment of a humeral capitulum osteonecrosis with bone morphogenetic protein-7 combined with autologous bone grafting.

    Science.gov (United States)

    Marsell, Richard; Hailer, Nils P

    2014-08-01

    We present the case of a 27-year-old female with subcortical osteonecrosis of the humeral capitulum. Percutaneous retrograde drilling of the lesion and application of recombinant human bone morphogenetic protein (BMP)-7 were combined with autologous bone grafting. At follow-up the patient was almost pain-free, had normalized her range of motion, and radiography showed consolidation of the lesion without any heterotopic bone formation. By timing surgery prior to subchondral collapse, biomechanical stability of the subchondral bone was maintained. To our knowledge, this is the first report on the treatment of an osteonecrosis in this location with a BMP, and this strategy could potentially be applied in other locations with juxta-articular osteonecrosis.

  17. Mixed matrix membrane adsorbers for protein and blood purification

    NARCIS (Netherlands)

    Saiful,

    2007-01-01

    Biotechnology and bio-manufacturing markets are continuously growing, generating new sources of many valuable healthcare and life science products including therapeutic proteins and polysaccharides, monoclonals, vaccines, diagnostics, pharmaceutical chemicals and enzymes. These bioproducts have to b

  18. Towards a matrix mechanics framework for dynamic protein network.

    Science.gov (United States)

    Bhattacharya, Sanjoy K

    2010-06-01

    Protein-protein interaction networks are currently visualized by software generated interaction webs based upon static experimental data. Current state is limited to static, mostly non-compartmental network and non time resolved protein interactions. A satisfactory mathematical foundation for particle interactions within a viscous liquid state (situation within the cytoplasm) does not exist nor do current computer programs enable building dynamic interaction networks for time resolved interactions. Building mathematical foundation for intracellular protein interactions can be achieved in two increments (a) trigger and capture the dynamic molecular changes for a select subset of proteins using several model systems and high throughput time resolved proteomics and, (b) use this information to build the mathematical foundation and computational algorithm for a compartmentalized and dynamic protein interaction network. Such a foundation is expected to provide benefit in at least two spheres: (a) understanding physiology enabling explanation of phenomenon such as incomplete penetrance in genetic disorders and (b) enabling several fold increase in biopharmaceutical production using impure starting materials.

  19. New collagen matrix to avoid the reduction of keratinized tissue during guided bone regeneration in postextraction sites.

    Science.gov (United States)

    De Santis, Daniele; Cucchi, Alessandro; de Gemmis, Antonio; Nocini Pier, Francesco

    2012-05-01

    For decades, there has been an ongoing controversy regarding the need for an "adequate" width of keratinized gingiva/mucosa to preserve periodontal and implant health. Today, the presence of a certain width of keratinized tissue is recommended for achieving long-term periodontal and implant success, and therefore, a new collagen matrix has been developed to enhance the width of keratinized gingiva/mucosa. During postextraction socket preservation, guided bone regeneration techniques require complete coverage of the barrier membrane to reduce the risk of infection, occasionally causing a reduction of the width of keratinized tissue. Using the new collagen matrix, it is possible to leave the membrane intentionally uncovered, without suturing the surgical flap above it, to avoid the reduction of such tissue.

  20. Conjugation of extracellular matrix proteins to basal lamina analogs enhances keratinocyte attachment.

    Science.gov (United States)

    Bush, Katie A; Downing, Brett R; Walsh, Sarah E; Pins, George D

    2007-02-01

    The dermal-epidermal junction of skin contains extracellular matrix proteins that are involved in initiating and controlling keratinocyte signaling events such as attachment, proliferation, and terminal differentiation. To characterize the relationship between extracellular matrix proteins and keratinocyte attachment, a biomimetic design approach was used to precisely tailor the surface of basal lamina analogs with biochemistries that emulate the native biochemical composition found at the dermal-epidermal junction. A high-throughput screening device was developed by our laboratory that allows for the simultaneous investigation of the conjugation of individual extracellular matrix proteins (e.g. collagen type I, collagen type IV, laminin, or fibronectin) as well as their effect on keratinocyte attachment, on the surface of an implantable collagen membrane. Fluorescence microscopy coupled with quantitative digital image analyses indicated that the extracellular matrix proteins adsorbed to the collagen-GAG membranes in a dose-dependent manner. To determine the relationship between extracellular matrix protein signaling cues and keratinocyte attachment, cells were seeded on protein-conjugated collagen-GAG membranes and a tetrazolium-based colorimetric assay was used to quantify viable keratinocyte attachment. Our results indicate that keratinocyte attachment was significantly enhanced on the surfaces of collagen membranes that were conjugated with fibronectin and type IV collagen. These findings define a set of design parameters that will enhance keratinocyte binding efficiency on the surface of collagen membranes and ultimately improve the rate of epithelialization for dermal equivalents.

  1. Biomimetic Mineralization on a Macroporous Cellulose-Based Matrix for Bone Regeneration

    Directory of Open Access Journals (Sweden)

    Odeta Petrauskaite

    2013-01-01

    Full Text Available The aim of this study is to investigate the biomimetic mineralization on a cellulose-based porous matrix with an improved biological profile. The cellulose matrix was precalcified using three methods: (i cellulose samples were treated with a solution of calcium chloride and diammonium hydrogen phosphate; (ii the carboxymethylated cellulose matrix was stored in a saturated calcium hydroxide solution; (iii the cellulose matrix was mixed with a calcium silicate solution in order to introduce silanol groups and to combine them with calcium ions. All the methods resulted in a mineralization of the cellulose surfaces after immersion in a simulated body fluid solution. Over a period of 14 days, the matrix was completely covered with hydroxyapatite crystals. Hydroxyapatite formation depended on functional groups on the matrix surface as well as on the precalcification method. The largest hydroxyapatite crystals were obtained on the carboxymethylated cellulose matrix treated with calcium hydroxide solution. The porous cellulose matrix was not cytotoxic, allowing the adhesion and proliferation of human osteoblastic cells. Comparatively, improved cell adhesion and growth rate were achieved on the mineralized cellulose matrices.

  2. Localized ridge defect augmentation using human pericardium membrane and demineralized bone matrix

    Directory of Open Access Journals (Sweden)

    Arun Kumar Vidyadharan

    2014-01-01

    Conclusion and Clinical Implications: The results suggested that HP Allograft membrane may be a suitable component for augmentation of localized alveolar ridge defects in conjunction with DBM with bone chips.

  3. Efficacy comparison of Accell Evo3 and Grafton demineralized bone matrix putties against autologous bone in a rat posterolateral spine fusion model.

    Science.gov (United States)

    Brecevich, Antonio T; Kiely, Paul D; Yoon, B Victor; Nguyen, Joseph T; Cammisa, Frank P; Abjornson, Celeste

    2017-06-01

    Spinal fusion procedures are intended to stabilize the spinal column for a multitude of disorders including abnormal curvature, traumatic instability, degenerative instability, and damage from infections or tumors. As an aid in the bone healing response, bone graft materials are used to bridge joints for arthrodesis and promote unions in pseudoarthrosis. Currently, the gold standard for stabilizing fusion masses in spinal procedures involves using the osteogenic, osteoinductive, and osteoconductive properties of autologous iliac crest corticocancellous bone. However, considerable morbidity is associated with harvesting the autologous graft. Donor site complications including infection, large hematomas, and pain have been reported at rates as high as 50% (Boden and Jeffrey, 1995). Biologically, the rate of bone repair dictates the rate at which the fusion mass will unite under autologous graft conditions. The purpose of this study is to compare the quality and rate of fusion between Accell Evo3 and Grafton demineralized bone matrix (DBM), with the gold standard iliac crest bone graft (ICBG) as the control, in athymic rat posterolateral fusion. This study was a randomized, controlled study in a laboratory setting at the Hospital for Special Surgery in New York City. Blinded observations were made, which created an assessment of outcomes for successful fusions between each method. Forty-eight (48) athymic rats were used in this study and underwent posterolateral lumbar fusion. They were assessed at either 3 weeks or 9 weeks to see the rate and efficacy of fusion. Outcome measures will be the efficacy of the different bone grafts and their success rates of fusion in the rats. A comparison of the quality and rate of fusion between Accell Evo3® (DBM A) and Grafton (DBM B), with the gold standard iliac crest bone graft (ICBG) as the control, was performed using the established posterolateral intertransverse process on an athymic rat model. Materials were evaluated for

  4. INFLUENCE OF THE SKELETON HIERARCHICAL ORGANIZATION ON ELECTRONIC STATE OF IONS IN BONE MATRIX

    Directory of Open Access Journals (Sweden)

    A. S. Avrunin

    2016-01-01

    Full Text Available The authors suggested the 3D-superlattice (3DSL model to describe the effect of coplanar assembly of the hydroxyapatite (HA nanocrystallites on local electronic state of ions in mineralized bone. This model is based on the main structural and functional relationships between adjacent levels of the hierarchical organization of bone tissue. In the framework of the 3DSL model the authors predicted the distinct assembly-to-crystal red shift of the unoccupied electronic states located near the bottom of the conduction band in HA and dependence of this shift on the ratio of the thickness of the hydrated layer to the crystallite size. To check these predictions the experimental X-ray absorption studies of native bone are performed near the Ca2р1/2,3/2-, P2р1/2,3/2- и O1s edges. Comparison of the measured spectra with the known spectra of the reference compounds has confirmed appearance of the distinct assembly-to-crystal red shift. The observed effect is the ground for development of new diagnostic methods for bone status and imaging changes in the local electronic structure of bone tissue by using ultrasoft X-ray absorption spectroscopy and measuring the assembly-tocrystal shifts. The experimental data analysis proved the applicability of the 3DSL model for better understanding of the hierarchical organization of bone at nanolevel.

  5. Chronic cyclophosphamide exposure alters the profile of rat sperm nuclear matrix proteins.

    Science.gov (United States)

    Codrington, Alexis M; Hales, Barbara F; Robaire, Bernard

    2007-08-01

    Chronic exposure of male rats to the alkylating agent cyclophosphamide, a well-known male-mediated developmental toxicant, alters gene expression in male germ cells as well as in early preimplantation embryos sired by cyclophosphamide-exposed males. Sperm DNA is organized by the nuclear matrix into loop-domains in a sequence-specific manner. In somatic cells, loop-domain organization is involved in gene regulation. Various structural and functional components of the nuclear matrix are targets for chemotherapeutic agents. Consequently, we hypothesized that cyclophosphamide treatment would alter the expression of sperm nuclear matrix proteins. Adult male rats were treated for 4 wk with saline or cyclophosphamide (6.0 mg kg(-1) day(-1)), and the nuclear matrix was extracted from cauda epididymal sperm. Proteins were analyzed by two-dimensional gel electrophoresis. Identified proteins within the nuclear matrix proteome were mainly involved in cell structure, transcription, translation, DNA binding, protein processing, signal transduction, metabolism, cell defense, or detoxification. Interestingly, cyclophosphamide selectively induced numerous changes in cell defense and detoxification proteins, most notably, in all known forms of the antioxidant enzyme glutathione peroxidase 4, in addition to an uncharacterized 54-kDa form; an overall increase in glutathione peroxidase 4 immunoreactivity was observed in the nuclear matrix extracts from cyclophosphamide-exposed spermatozoa. An increase in glutathione peroxidase 4 expression suggests a role for this enzyme in maintaining nuclear matrix stability and function. These results led us to propose that a change in composition of the nuclear matrix in response to drug exposure was a factor in altered sperm function and embryo development.

  6. Effects of Bone Morphogenic Proteins on Engineered Cartilage

    Science.gov (United States)

    Gooch, Keith, J.; Blunk, Torsten; Courter, Donald L.; Sieminski, Alisha; Vunjak-Novakovic, Gordana; Freed, Lisa E.

    2007-01-01

    A report describes experiments on the effects of bone morphogenic proteins (BMPs) on engineered cartilage grown in vitro. In the experiments, bovine calf articular chondrocytes were seeded onto biodegradable polyglycolic acid scaffolds and cultured in, variously, a control medium or a medium supplemented with BMP-2, BMP-12, or BMP-13 in various concentrations. Under all conditions investigated, cell-polymer constructs cultivated for 4 weeks macroscopically and histologically resembled native cartilage. At a concentration of 100 ng/mL, BMP-2, BMP-12, or BMP-13 caused (1) total masses of the constructs to exceed those of the controls by 121, 80, or 62 percent, respectively; (2) weight percentages of glycosaminoglycans in the constructs to increase by 27, 18, or 15, respectively; and (3) total collagen contents of the constructs to decrease to 63, 89, or 83 percent of the control values, respectively. BMP-2, but not BMP-12 or BMP-13, promoted chondrocyte hypertrophy. These observations were interpreted as suggesting that the three BMPs increase the growth rates and modulate the compositions of engineered cartilage. It was also concluded that in vitro engineered cartilage is a suitable system for studying effects of BMPs on chondrogenesis in a well-defined environment.

  7. The effect of bone morphogenetic protein-2 on osteosarcoma metastasis

    Science.gov (United States)

    Gill, Jonathan; Connolly, Patrick; Roth, Michael; Chung, So Hak; Zhang, Wendong; Piperdi, Sajida; Hoang, Bang; Yang, Rui; Guzik, Hillary; Gorlick, Richard; Geller, David S.

    2017-01-01

    Purpose Bone Morphogenetic Protein-2 (BMP-2) may offer the potential to enhance allograft-host osseous union in limb-salvage surgery following osteosarcoma resection. However, there is concern regarding the effect of locally applied BMP-2 on tumor recurrence and metastasis. The purpose of this project was to evaluate the effect of exogenous BMP-2 on osteosarcoma migration and invasion across a panel of tumor cell lines in vitro and to characterize the effect of BMP-2 on pulmonary osteosarcoma metastasis within a xenograft model. Experimental design The effect of BMP-2 on in vitro tumor growth and development was assessed across multiple standard and patient-derived xenograft osteosarcoma cell lines. Tumor migration capacity, invasion, and cell proliferation were characterized. In addition, the effect on metastasis was measured using a xenograft model following tail-vein injection. The effect of exogenous BMP-2 on the development of metastases was measured following both single and multiple BMP-2 administrations. Results There was no significant difference in migration capacity, invasion, or cell proliferation between the BMP-2 treated and the untreated osteosarcoma cell lines. There was no significant difference in pulmonary metastases between either the single-dose or multi-dose BMP-2 treated animals and the untreated control animals. Conclusions In the model systems tested, the addition of BMP-2 does not increase osteosarcoma proliferation, migration, invasion, or metastasis to the lungs. PMID:28264040

  8. The effect of bone morphogenetic protein-2 on osteosarcoma metastasis.

    Science.gov (United States)

    Gill, Jonathan; Connolly, Patrick; Roth, Michael; Chung, So Hak; Zhang, Wendong; Piperdi, Sajida; Hoang, Bang; Yang, Rui; Guzik, Hillary; Morris, Jonathan; Gorlick, Richard; Geller, David S

    2017-01-01

    Bone Morphogenetic Protein-2 (BMP-2) may offer the potential to enhance allograft-host osseous union in limb-salvage surgery following osteosarcoma resection. However, there is concern regarding the effect of locally applied BMP-2 on tumor recurrence and metastasis. The purpose of this project was to evaluate the effect of exogenous BMP-2 on osteosarcoma migration and invasion across a panel of tumor cell lines in vitro and to characterize the effect of BMP-2 on pulmonary osteosarcoma metastasis within a xenograft model. The effect of BMP-2 on in vitro tumor growth and development was assessed across multiple standard and patient-derived xenograft osteosarcoma cell lines. Tumor migration capacity, invasion, and cell proliferation were characterized. In addition, the effect on metastasis was measured using a xenograft model following tail-vein injection. The effect of exogenous BMP-2 on the development of metastases was measured following both single and multiple BMP-2 administrations. There was no significant difference in migration capacity, invasion, or cell proliferation between the BMP-2 treated and the untreated osteosarcoma cell lines. There was no significant difference in pulmonary metastases between either the single-dose or multi-dose BMP-2 treated animals and the untreated control animals. In the model systems tested, the addition of BMP-2 does not increase osteosarcoma proliferation, migration, invasion, or metastasis to the lungs.

  9. Bone morphogenetic proteins in multiple sclerosis: Role in neuroinflammation.

    Science.gov (United States)

    Eixarch, Herena; Calvo-Barreiro, Laura; Montalban, Xavier; Espejo, Carmen

    2017-02-27

    Bone morphogenetic proteins (BMPs) are growth factors that represent the largest subgroup of signalling ligands of the transforming growth factor beta (TGF-β) superfamily. Their participation in the proliferation, survival and cell fate of several cell types and their involvement in many pathological conditions are now well known. BMP expression is altered in multiple sclerosis (MS) patients, suggesting that BMPs have a role in the pathogenesis of this disease. MS is a demyelinating and neurodegenerative autoimmune disorder of the central nervous system (CNS). MS is a complex pathological condition in which genetic, epigenetic and environmental factors converge, although its aetiology remains elusive. Multifunctional molecules, such as BMPs, are extremely interesting in the field of MS because they are involved in the regulation of several adult tissues, including the CNS and the immune system. In this review, we discuss the extensive data available regarding the role of BMP signalling in neuronal progenitor/stem cell fate and focus on the participation and expression of BMPs in CNS demyelination. Additionally, we provide an overview of the involvement of BMPs as modulators of the immune system, as this subject has not been thoroughly explored even though it is of great interest in autoimmune disorders. Moreover, we describe the data on BMP signalling in autoimmunity and inflammatory diseases, including MS and its experimental models. Thus, we aim to provide an integrated view of the putative role of BMPs in MS pathogenesis and to open the field for the further development of alternative therapeutic strategies for MS patients.

  10. Predicting protein-ligand affinity with a random matrix framework.

    Science.gov (United States)

    Lee, Alpha A; Brenner, Michael P; Colwell, Lucy J

    2016-11-29

    Rapid determination of whether a candidate compound will bind to a particular target receptor remains a stumbling block in drug discovery. We use an approach inspired by random matrix theory to decompose the known ligand set of a target in terms of orthogonal "signals" of salient chemical features, and distinguish these from the much larger set of ligand chemical features that are not relevant for binding to that particular target receptor. After removing the noise caused by finite sampling, we show that the similarity of an unknown ligand to the remaining, cleaned chemical features is a robust predictor of ligand-target affinity, performing as well or better than any algorithm in the published literature. We interpret our algorithm as deriving a model for the binding energy between a target receptor and the set of known ligands, where the underlying binding energy model is related to the classic Ising model in statistical physics.

  11. Capacidade da matriz extracelular da medula óssea de induzir proliferação de células mielóides in vitro no modelo de desnutrição protéica em camundongos Capacity of the extracellular matrix of the bone marrow to induce proliferation of myeloid cells in vitro in model of protein malnutrition in mice

    Directory of Open Access Journals (Sweden)

    Cidônia de Lourdes Vituri

    2008-09-01

    Full Text Available Este trabalho tem por objetivo verificar se a matriz extracelular (MEC obtida da medula óssea de camundongos com desnutrição protéica energética sustenta a sobrevivência, se induz proliferação de células mielóides, bem como avaliar a capacidade desta MEC de interagir com citocinas hematopoiéticas in vitro. Camundongos machos "Swiss" foram submetidos à desnutrição protéica (4% de caseína até que perdessem 20% do peso inicial e o grupo-controle foi mantido com uma dieta contendo 14% de caseína. A medula óssea foi extraída com tampão PBS suplementado com 1 mg de aprotinina/mL. Os ensaios de proliferação foram realizados com a linhagem mielóide FDC-P1, pelo método colorimétrico de redução do MTT. A MEC obtida tanto do grupo-controle como do desnutrido induziu proliferação celular in vitro. Os ensaios de interação foram realizados com IL-3 e GM-CSF na concentração de 10 ρg e 500 ρg/mL, que demonstraram efeito sinérgico e efeito regulatório, respectivamente. A MEC obtida de animais do grupo desnutrido quando submetida ao ensaio de ligação ao GM-CSF mostrou maior proliferação celular do que a MEC obtida de animais do grupo-controle (pThe aim of this study was to verify the capacity of the extracellular matrix (ECM obtained from bone marrow of malnourished mice to sustain survival and to induce the proliferation of myeloid cells. We also verified the capacity of the tests to interact with in vitro hematopoietic cytokines. Male "Swiss" mice were submitted to protein malnutrition with a diet content of '4% casein until they lost 20% of the original weight, while the group-control was kept with a diet content of 14% of casein. The bone marrow was extracted with 1.0 mg of aprotinin/mL in PBS. The proliferation tests were carried out with myeloid cell line FDCP-1, by the colorimetric method of reduction of the MTT. The obtained ECM from nourished and undernourished mice induced cellular proliferation invitro. Tests

  12. Subcutaneous administration of insulin-like growth factor (IGF)-II/IGF binding protein-2 complex stimulates bone formation and prevents loss of bone mineral density in a rat model of disuse osteoporosis

    Science.gov (United States)

    Conover, Cheryl A.; Johnstone, Edward W.; Turner, Russell T.; Evans, Glenda L.; John Ballard, F. John; Doran, Patrick M.; Khosla, Sundeep

    2002-01-01

    Elevated serum levels of insulin-like growth factor binding protein-2 (IGFBP-2) and a precursor form of IGF-II are associated with marked increases in bone formation and skeletal mass in patients with hepatitis C-associated osteosclerosis. In vitro studies indicate that IGF-II in complex with IGFBP-2 has high affinity for bone matrix and is able to stimulate osteoblast proliferation. The purpose of this study was to determine the ability of the IGF-II/IGFBP-2 complex to increase bone mass in vivo. Osteopenia of the femur was induced by unilateral sciatic neurectomy in rats. At the time of surgery, 14-day osmotic minipumps containing vehicle or 2 microg IGF-II+9 microg IGFBP-2/100g body weight/day were implanted subcutaneously in the neck. Bone mineral density (BMD) measurements were taken the day of surgery and 14 days later using a PIXImus small animal densitometer. Neurectomy of the right hindlimb resulted in a 9% decrease in right femur BMD (PBone histomorphometry indicated increases in endocortical and cancellous bone formation rates and in trabecular thickness. These results demonstrate that short-term administration of the IGF-II/IGFBP-2 complex can prevent loss of BMD associated with disuse osteoporosis and stimulate bone formation in adult rats. Furthermore, they provide proof of concept for a novel anabolic approach to increasing bone mass in humans with osteoporosis.

  13. Effects of extracellular matrix proteins on macrophage differentiation, growth, and function: comparison of liquid and agar culture systems

    Science.gov (United States)

    Armstrong, J. W.; Chapes, S. K.; Spooner, B. S. (Principal Investigator)

    1994-01-01

    Both spaceflight and skeletal unloading suppress the haematopoietic differentiation of macrophages (Sonnenfeld et al., Aviat. Space Environ. Med., 61:648-653, 1990; Armstrong et al., J. Appl. Physiol., 75:2734-2739, 1993). The mechanism behind this reduction in haematopoiesis has yet to be elucidated. However, changes in bone marrow extracellular matrix (ECM) may be involved. To further understand the role of ECM products in macrophage differentiation, we have performed experiments evaluating the effects of fibronectin, laminin, collagen type I, and collagen type IV on macrophage development and function. Bone marrow-derived macrophages cultured on four different ECM substrates in liquid culture medium showed less growth than those cultured on plastic. Significant morphological differences were seen on each of the substrates used. Phenotypically and functionally, as measured by class II major histocompatibility molecule (MHCII) expression, MAC-2 expression, and the secretion of interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-alpha), these macrophages were similar. In contrast, bone marrow-derived macrophages cultured in suspension, using agar, showed no difference in growth when exposed to ECM proteins. However, IL-6 and TNF-alpha secretion was affected by fibronectin, laminin, collagen type I, and collagen type IV in a concentration-dependent manner. We conclude that the ECM products fibronectin, laminin, collagen type I, and collagen type IV have profound effects on macrophage development and function. Additionally, we suggest that an ECM-supplemented agar culture system provides an environment more analogous to in vivo bone marrow than does a traditional liquid culture system.

  14. Import of peroxisomal matrix proteins in the yeast Hansenula polymorpha

    NARCIS (Netherlands)

    Gunkel, Katja

    2005-01-01

    Archaea, prokaryotes and eukaryotes form the three kingdoms of life. The smallest unit of life, which can exist independently, is a cell. Archaea and prokaryotes have a relatively very simple architecture. The cytoplasm (cellulars pace), containing all metabolites, proteins and genetic material (DNA

  15. The influence of dietary crude protein intake on bone and mineral metabolism in sheep

    Directory of Open Access Journals (Sweden)

    T.S. Brand

    1999-07-01

    Full Text Available Increased dietary protein consumption is thought to cause calciuresis, a negative calcium balance and increased bone loss that may result in skeletal deformities and fracture. To explore this hypothesis, 40 approximately 100-day-old meat-type Merino ram lambs were fed, for 6 months, diets with an increasing crude protein (CP content (114, 142, 171 and 190 g/kg DM but approximately on an iso-nutrient basis with regard to metabolisable energy, calcium and phosphorus. Increased protein consumption modestly (NS enhanced calciuresis and resulted in significant (P < 0.01 limb skewness. This could not, however, be ascribed to osteopaenic bones, and compared with animals consuming lower protein rations, the bone mineral density (BMD and vertebral trabecular bone volume of animals fed high protein diets were significantly increased: theBMDof thoracic vertebrae was positively related to the CP intake (r=0.62; P < 0.001. In animals consuming higher protein diets, skeletal radiology and quantitative bone histology revealed no evidence of increased bone turnover as would be expected in animals that are in negative calcium balance. No relationship existed between limb skewness and the growth rate of lambs. However, the ratio of Ca:P in the forelimb (r = -0.98, vertebrae (r = -0.72 and rib (r = -0.42 was found to be inversely correlated with increased protein intake and resulted from an increase in the phosphorus content of bone, while the amount of bone calcium was unaffected. We conclude that qualitative micro-architectural abnormalities, and not mere bone loss, may underlie the skeletal deformities induced by increased protein consumption in sheep.

  16. Osteocalcin and matrix GLA protein in developing teleost teeth: identification of sites of mRNA and protein accumulation at single cell resolution.

    Science.gov (United States)

    Ortiz-Delgado, J B; Simes, D C; Gavaia, P; Sarasquete, C; Cancela, M L

    2005-08-01

    In this study, the tissue distribution and accumulation of osteocalcin or bone Gla protein (BGP) and matrix Gla protein (MGP) were determined during tooth development in a teleost fish, Argyrosomus regius. In this species, the presence of BGP and MGP mRNA in teeth was revealed by in situ hybridization. mRNA for BGP was detected in the odontoblasts as well as in its cytoplasmic processes emerging through dentinal tubules, while mRNA for MGP was expressed in the enamel portion within the apical portion of the elongated cell bodies of enameloblasts, adjacent to the root of the teeth as well as in cells within the pulpal space. Immunolocalization of BGP and MGP demonstrated that these proteins accumulate mainly in the mineralized dentin or in enameloblastic processes, confirming in situ hybridization results. In this study, we examined for the first time the localization of both BGP and MGP gene expression and protein accumulation within the different regions of the vertebrate tooth. We clearly demonstrated that although the overall pattern of BGP and MGP gene expression and protein accumulation in A. regius teeth was in general agreement to what is known for other vertebrates such as rats or rodents, our study provided novel information and highlighted some species-differences between fish and higher vertebrates.

  17. The effect of statins in colorectal cancer is mediated through the bone morphogenetic protein pathway

    NARCIS (Netherlands)

    Kodach, Liudmila L.; Bleuming, Sylvia A.; Peppelenbosch, Maikel P.; Hommes, Daniel W.; Van Den Brink, Gus R.; Hardwick, James C. H.

    2007-01-01

    Background & Aims: Epidemiological evidence suggests that statins prevent colorectal cancer (CRC), but the biological mechanism remains obscure. Statins induce bone morphogenetic protein (BMP) expression in bone cells. We have previously shown that BMPs act as tumor suppressors in CRC. We hypothesiz

  18. Differential expression pattern of extracellular matrix molecules during chondrogenesis of mesenchymal stem cells from bone marrow and adipose tissue

    DEFF Research Database (Denmark)

    Mehlhorn, A T; Niemeyer, P; Kaiser, S;

    2006-01-01

    Adipose-derived adult stem cells (ADASCs) or bone marrow-derived mesenchymal stem cells (BMSCs) are considered as alternative cell sources for cell-based cartilage repair due to their ability to produce cartilage-specific matrix. This article addresses the differential expression pattern of extra......Adipose-derived adult stem cells (ADASCs) or bone marrow-derived mesenchymal stem cells (BMSCs) are considered as alternative cell sources for cell-based cartilage repair due to their ability to produce cartilage-specific matrix. This article addresses the differential expression pattern...... mRNA was monitored via reverse transcriptase-polymerase chain reaction (RT-PCR). Corresponding ECM synthesis was demonstrated using immunohistochemistry. After chondroinduction, expression of collagen type II, type X, COMP and aggrecan mRNA was 3-15-fold higher than in ADASCs. The type IIA splicing...... form of alpha(1)-procollagen type II was expressed in both populations, and the type IIB splicing form was exclusively detected in BMSCs. In response to TGF-beta, collagen type II and X were secreted more strongly by BMSCs than by ADASCs. BMSCs express a more mature phenotype than ADASCs after...

  19. Moderate cyclic tensile strain alters the assembly of cartilage extracellular matrix proteins in vitro.

    Science.gov (United States)

    Bleuel, Judith; Zaucke, Frank; Brüggemann, Gert-Peter; Heilig, Juliane; Wolter, Marie-Louise; Hamann, Nina; Firner, Sara; Niehoff, Anja

    2015-06-01

    Mechanical loading influences the structural and mechanical properties of articular cartilage. The cartilage matrix protein collagen II essentially determines the tensile properties of the tissue and is adapted in response to loading. The collagen II network is stabilized by the collagen II-binding cartilage oligomeric matrix protein (COMP), collagen IX, and matrilin-3. However, the effect of mechanical loading on these extracellular matrix proteins is not yet understood. Therefore, the aim of this study was to investigate if and how chondrocytes assemble the extracellular matrix proteins collagen II, COMP, collagen IX, and matrilin-3 in response to mechanical loading. Primary murine chondrocytes were applied to cyclic tensile strain (6%, 0.5 Hz, 30 min per day at three consecutive days). The localization of collagen II, COMP, collagen IX, and matrilin-3 in loaded and unloaded cells was determined by immunofluorescence staining. The messenger ribo nucleic acid (mRNA) expression levels and synthesis of the proteins were analyzed using reverse transcription-polymerase chain reaction (RT-PCR) and western blots. Immunofluorescence staining demonstrated that the pattern of collagen II distribution was altered by loading. In loaded chondrocytes, collagen II containing fibrils appeared thicker and strongly co-stained for COMP and collagen IX, whereas the collagen network from unloaded cells was more diffuse and showed minor costaining. Further, the applied load led to a higher amount of COMP in the matrix, determined by western blot analysis. Our results show that moderate cyclic tensile strain altered the assembly of the extracellular collagen network. However, changes in protein amount were only observed for COMP, but not for collagen II, collagen IX, or matrilin-3. The data suggest that the adaptation to mechanical loading is not always the result of changes in RNA and/or protein expression but might also be the result of changes in matrix assembly and structure.

  20. Conformal nanopatterning of extracellular matrix proteins onto topographically complex surfaces.

    Science.gov (United States)

    Sun, Yan; Jallerat, Quentin; Szymanski, John M; Feinberg, Adam W

    2015-02-01

    Our Patterning on Topography (PoT) printing technique enables fibronectin, laminin and other proteins to be applied to biomaterial surfaces in complex geometries that are inaccessible using traditional soft lithography techniques. Engineering combinatorial surfaces that integrate topographical and biochemical micropatterns enhances control of the biotic-abiotic interface. Here, we used this method to understand cardiomyocyte response to competing physical and chemical cues in the microenvironment.

  1. Human keratinocytes synthesize and secrete the extracellular matrix protein, thrombospondin.

    Science.gov (United States)

    Wikner, N E; Dixit, V M; Frazier, W A; Clark, R A

    1987-02-01

    Thrombospondin (TSP) a glycoprotein originally identified as the endogenous lectin of platelets, is also synthesized by fibroblasts, endothelial cells, pneumocytes, smooth muscle cells, and macrophages. Thrombospondin is subdivided into functional domains which bind specifically to heparin, fibronectin, collagen, and to specific cellular receptors. It is found within the basement membranes of kidney, lung, smooth muscle, and skin. Thus TSP may serve as an important link between cells and matrices. Thrombospondin also has been reported at the epidermal-dermal junction. We wished to determine whether human keratinocytes synthesize and secrete TSP. Pure human keratinocytes were grown in defined medium without fibroblast feeder layers. Immunofluorescent staining with either rabbit polyclonal or mouse monoclonal antibodies to human platelet TSP yielded specific granular staining within the cytoplasm of keratinocytes. Culture media and cellular lysates were harvested from cultures metabolically labeled with [35S]methionine. Trichloroacetic acid precipitation, sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and autoradiography revealed a major labeled band comigrating with purified platelet TSP in both the media and the cellular lysates. Immunoprecipitation with either the polyclonal or the monoclonal anti-TSP antibodies followed by SDS-PAGE and autoradiography identified this band as TSP. Thus keratinocytes in culture synthesize and secrete TSP. Thrombospondin may play an important role in epidermal interactions with extracellular matrix.

  2. Serum protein fractionation using supported molecular matrix electrophoresis.

    Science.gov (United States)

    Dong, Weijie; Matsuno, Yu-ki; Kameyama, Akihiko

    2013-08-01

    Supported molecular matrix electrophoresis (SMME), in which a hydrophilic polymer such as PVA serves as a support within a porous PVDF membrane, was recently developed. This method is similar to cellulose acetate membrane electrophoresis but differs in the compatibility to glycan analysis of the separated bands. In this report, we describe the first instance of the application of SMME to human serum fractionation, and demonstrate the differences with serum fractionation by cellulose acetate membrane electrophoresis. The SMME membrane exhibited almost no EOF during electrophoresis, unlike the cellulose acetate membrane, but afforded comparative results for serum fractionation. The visualization of each fraction was achieved by conventional staining with dye such as Direct Blue-71, and objective quantification was obtained by densitometry after inducing membrane transparency with 1-nonene. Immunostaining was also achieved. Moreover, mass spectrometric analysis of both N-linked and O-linked glycans from the separated bands was demonstrated. Serum fractionation and glycan profiling of each fraction using SMME will enable novel insights into the relationships between various glycosylation profiles and disease states.

  3. Degenerated human intervertebral discs contain autoantibodies against extracellular matrix proteins

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    S Capossela

    2014-04-01

    Full Text Available Degeneration of intervertebral discs (IVDs is associated with back pain and elevated levels of inflammatory cells. It has been hypothesised that discogenic pain is a direct result of vascular and neural ingrowth along annulus fissures, which may expose the avascular nucleus pulposus (NP to the systemic circulation and induce an autoimmune reaction. In this study, we confirmed our previous observation of antibodies in human degenerated and post-traumatic IVDs cultured in vitro. We hypothesised that the presence of antibodies was due to an autoimmune reaction against specific proteins of the disc. Furthermore we identified antigens which possibly trigger an autoimmune response in degenerative disc diseases. We demonstrated that degenerated and post-traumatic IVDs contain IgG antibodies against typical extracellular proteins of the disc, particularly proteins of the NP. We identified IgGs against collagen type II and aggrecan, confirming an autoimmune reaction against the normally immune privileged NP. We also found specific IgGs against collagens types I and V, but not against collagen type III. In conclusion, this study confirmed the association between disc degeneration and autoimmunity, and may open the avenue for future studies on developing prognostic, diagnostic and therapy-monitoring markers for degenerative disc diseases.

  4. Degenerated human intervertebral discs contain autoantibodies against extracellular matrix proteins.

    Science.gov (United States)

    Capossela, S; Schläfli, P; Bertolo, A; Janner, T; Stadler, B M; Pötzel, T; Baur, M; Stoyanov, J V

    2014-04-04

    Degeneration of intervertebral discs (IVDs) is associated with back pain and elevated levels of inflammatory cells. It has been hypothesised that discogenic pain is a direct result of vascular and neural ingrowth along annulus fissures, which may expose the avascular nucleus pulposus (NP) to the systemic circulation and induce an autoimmune reaction. In this study, we confirmed our previous observation of antibodies in human degenerated and post-traumatic IVDs cultured in vitro. We hypothesised that the presence of antibodies was due to an autoimmune reaction against specific proteins of the disc. Furthermore we identified antigens which possibly trigger an autoimmune response in degenerative disc diseases. We demonstrated that degenerated and post-traumatic IVDs contain IgG antibodies against typical extracellular proteins of the disc, particularly proteins of the NP. We identified IgGs against collagen type II and aggrecan, confirming an autoimmune reaction against the normally immune privileged NP. We also found specific IgGs against collagens types I and V, but not against collagen type III. In conclusion, this study confirmed the association between disc degeneration and autoimmunity, and may open the avenue for future studies on developing prognostic, diagnostic and therapy-monitoring markers for degenerative disc diseases.

  5. Bone morphogenetic protein signaling protects against cerulein-induced pancreatic fibrosis.

    Directory of Open Access Journals (Sweden)

    Xuxia Gao

    Full Text Available Bone morphogenetic proteins (BMPs have an anti-fibrogenic function in the kidney, lung, and liver. However, their role in chronic pancreatitis (CP is unknown. The aim of this study was to define the anti-fibrogenic role of BMP signaling in the pancreas in vivo under CP induction. Mice with a deletion of BMP type II receptor (BMPR2(+/- were used in this study in comparison with wild-type mice. CP was induced by repetitive cerulein injection intraperitoneally for 4 weeks, and the severity of CP was evaluated. Pancreatic stellate cells (PSCs were isolated from the mice and treated with BMP2 and TGF-β in vitro, and extracellular matrix protein (ECM production was measured. Smad and mitogen-activated protein kinase (MAPK signaling was also evaluated. BMPR2(+/- mice revealed a greater pancreatic fibrosis, PSC activation and leukocyte infiltration after CP induction compared to wild-type mice (P<0.05. Under CP induction, phospho (pSmad1/5/8 was elevated in wild-type mice and this effect was abolished in BMPR2(+/- mice; pSmad2 and pp38(MAPK were further enhanced in BMPR2(+/- mice compared to wild-type mice (P<0.05. In vitro, BMP2 inhibited TGF-β-induced ECM protein fibronectin production in wild-type PSCs; this effect was abolished in BMPR2(+/- PSCs (P<0.05. In BMPR2(+/- PSCs, pSmad1/5/8 level was barely detectable upon BMP2 stimulation, while pSmad2 level was further enhanced by TGF-β stimulation, compared to wild-type PSCs (P<0.05. BMPR2/Smad1/5/8 signaling plays a protective role against cerulein-induced pancreatic fibrosis by inhibiting Smad2 and p38(MAPK signaling pathways.

  6. Histologic analysis of a novel extracellullar matrix membrane for guided bone regeneration: an experimental study in rabbits.

    Science.gov (United States)

    Al-Asfour, Adel; Tengvall, Pentti; Andersson, Lars; Dahlin, Christer

    2013-01-01

    This experiment was conducted to study the histologic feasibility and biologic impact of Ti02 impregnation of an extracellullar matrix (ECM) membrane in guided bone regeneration. Eighteen adult New Zealand White rabbits were used. Bilateral bone defects were created in edentulous areas of the maxilla. The defects were filled with demineralized freeze-dried bone (DFDB). ECM was randomly pretreated with a suspension containing saline and 3 mg Ti02 granules. A regular ECM membrane served as a control on the contralateral side. Healing periods were 2, 4, and 8 weeks. Histologic and histomorphometric analyses were performed. The parameters assessed were (1) zone of inflammatory cells adjacent to ECM membrane, (2) presence of cellullar ingrowth into ECM, and (3) presence of Ti02 particles within the ECM barrier membrane. In general, no adverse reactions toward both groups of ECM membranes could be noted. The Ti02 particles remained within the ECM after 8 weeks of healing, making histologic detection of ECM easy. Histomorphometric analysis revealed low numbers of inflammatory cells adjacent to the ECM surface and adequate preservation and integration of the barrier. Contrasting Ti02 particles impregnated into the ECM membrane can be a very useful tool for the detection of similar biologic materials in in vivo models.

  7. Poly(ADP-ribosyl)ation of proteins associated with nuclear matrix in rat testis

    Energy Technology Data Exchange (ETDEWEB)

    Quesada, P.; Atorino, L.; Faraone-Mennella, M.R.; Farina, B. [Naples Univ. (Italy); Caiafa, P. [Rome Univ. (Italy)

    1995-12-31

    We have previously demonstrated that a significant percentage of poly(ADPR)polymerase is present, as a tightly-bound form, at the third level of chromatin organization defined by chromosomal loops and nuclear matrix. The present work is focused on the study of poly(ADP-ribosyl)ation of proteins present in these nuclear subfractions. It has been shown that, due to the action of poly(ADPR) polymerase, the ADP-ribose moiety of [{sup 14}C]NAD is transferred to both loosely-bound and tightly-bound chromosomal proteins, which in consequence are modified by chain polymers of ADP-ribose of different lengths. Moreover, histone-like proteins seem to be ADP-ribosylated in chromosomal loops and nuclear matrix associated regions of DNA loops (MARS). A hypothesis can be put forward that the ADP-ribosylation system is functionally related to the nuclear processes, actively coordinated by the nuclear matrix. (author). 34 refs, 4 figs.

  8. Vibrio cholerae utilizes direct sRNA regulation in expression of a biofilm matrix protein.

    Directory of Open Access Journals (Sweden)

    Tianyan Song

    Full Text Available Vibrio cholerae biofilms contain exopolysaccharide and three matrix proteins RbmA, RbmC and Bap1. While much is known about exopolysaccharide regulation, little is known about the mechanisms by which the matrix protein components of biofilms are regulated. VrrA is a conserved, 140-nt sRNA of V. cholerae, whose expression is controlled by sigma factor σE. In this study, we demonstrate that VrrA negatively regulates rbmC translation by pairing to the 5' untranslated region of the rbmC transcript and that this regulation is not stringently dependent on the RNA chaperone protein Hfq. These results point to VrrA as a molecular link between the σE-regulon and biofilm formation in V. cholerae. In addition, VrrA represents the first example of direct regulation of sRNA on biofilm matrix component, by-passing global master regulators.

  9. Novel proteins identified in the insoluble byssal matrix of the freshwater zebra mussel.

    Science.gov (United States)

    Gantayet, Arpita; Rees, David J; Sone, Eli D

    2014-04-01

    The freshwater zebra mussel, Dreissena polymorpha, is an invasive, biofouling species that adheres to a variety of substrates underwater, using a proteinaceous anchor called the byssus. The byssus consists of a number of threads with adhesive plaques at the tips. It contains the unusual amino acid 3, 4-dihydroxyphenylalanine (DOPA), which is believed to play an important role in adhesion, in addition to providing structural integrity to the byssus through cross-linking. Extensive DOPA cross-linking, however, renders the zebra mussel byssus highly resistant to protein extraction, and therefore limits byssal protein identification. We report here on the identification of seven novel byssal proteins in the insoluble byssal matrix following protein extraction from induced, freshly secreted byssal threads with minimal cross-linking. These proteins were identified by LC-MS/MS analysis of tryptic digests of the matrix proteins by spectrum matching against a zebra mussel cDNA library of genes unique to the mussel foot, the organ that secretes the byssus. All seven proteins were present in both the plaque and thread. Comparisons of the protein sequences revealed common features of zebra mussel byssal proteins, and several recurring sequence motifs. Although their sequences are unique, many of the proteins display similarities to marine mussel byssal proteins, as well as to adhesive and structural proteins from other species. The large expansion of the byssal proteome reported here represents an important step towards understanding zebra mussel adhesion.

  10. Expression Patterns of Extracellular Matrix Proteins during Posterior Commissure Development

    Science.gov (United States)

    Stanic, Karen; Saldivia, Natalia; Förstera, Benjamín; Torrejón, Marcela; Montecinos, Hernán; Caprile, Teresa

    2016-01-01

    Extracellular matrix (ECM) molecules are pivotal for central nervous system (CNS) development, facilitating cell migration, axonal growth, myelination, dendritic spine formation, and synaptic plasticity, among other processes. During axon guidance, the ECM not only acts as a permissive or non-permissive substrate for navigating axons, but also modulates the effects of classical guidance cues, such as netrin or Eph/ephrin family members. Despite being highly important, little is known about the expression of ECM molecules during CNS development. Therefore, this study assessed the molecular expression patterns of tenascin, HNK-1, laminin, fibronectin, perlecan, decorin, and osteopontin along chick embryo prosomere 1 during posterior commissure development. The posterior commissure is the first transversal axonal tract of the embryonic vertebrate brain. Located in the dorso-caudal portion of prosomere 1, posterior commissure axons primarily arise from the neurons of basal pretectal nuclei that run dorsally to the roof plate midline, where some turn toward the ipsilateral side. Expressional analysis of ECM molecules in this area these revealed to be highly arranged, and molecule interactions with axon fascicles suggested involvement in processes other than structural support. In particular, tenascin and the HNK-1 epitope extended in ventro-dorsal columns and enclosed axons during navigation to the roof plate. Laminin and osteopontin were expressed in the midline, very close to axons that at this point must decide between extending to the contralateral side or turning to the ipsilateral side. Finally, fibronectin, decorin, and perlecan appeared unrelated to axonal pathfinding in this region and were instead restricted to the external limiting membrane. In summary, the present report provides evidence for an intricate expression of different extracellular molecules that may cooperate in guiding posterior commissure axons. PMID:27733818

  11. Expression Patterns of Extracellular Matrix Proteins during Posterior Commissure Development

    Directory of Open Access Journals (Sweden)

    Karen Stanic

    2016-09-01

    Full Text Available Extracellular matrix (ECM molecules are pivotal for central nervous system development, facilitating cell migration, axonal growth, myelination, dendritic spine formation, and synaptic plasticity, among other processes. During axonal guidance, the ECM not only acts as a permissive or non-permissive substrate for navigating axons, but also modulates the effects of classical guidance cues, such as netrin or Eph/ephrin family members. Despite being highly important, little is known about the expression of ECM molecules during central nervous system development. Therefore, this study assessed the molecular expression patterns of tenascin, HNK-1, laminin, fibronectin, perlecan, decorin, and osteopontin along chick embryo prosomere 1 during posterior commissure development. The posterior commissure is the first transversal axonal tract of the embryonic vertebrate brain. Located in the dorso-caudal portion of prosomere 1, posterior commissure axons primarily arise from the neurons of basal pretectal nuclei that run dorsally to the roof plate midline, where some turn towards the ipsilateral side. Expressional analysis of ECM molecules in this area these revealed to be highly arranged, and molecule interactions with axon fascicles suggested involvement in processes other than structural support. In particular, tenascin and the HNK-1 epitope extended in ventro-dorsal columns and enclosed axons during navigation to the roof plate. Laminin and osteopontin were expressed in the midline, very close to axons that at this point must decide between extending to the contralateral side or turning to the ipsilateral side. Finally, fibronectin, decorin, and perlecan appeared unrelated to axonal pathfinding in this region and were instead restricted to the external limiting membrane. In summary, the present report provides evidence for an intricate expression of different extracellular molecules that may cooperate in guiding posterior commissure axons.

  12. Altering adsorbed proteins or cellular gene expression in bone-metastatic cancer cells affects PTHrP and Gli2 without altering cell growth

    Directory of Open Access Journals (Sweden)

    Jonathan M. Page

    2015-09-01

    Full Text Available The contents of this data in brief are related to the article titled “Matrix Rigidity Regulates the Transition of Tumor Cells to a Bone-Destructive Phenotype through Integrin β3 and TGF-β Receptor Type II”. In this DIB we will present our supplemental data investigating Integrin expression, attachment of cells to various adhesion molecules, and changes in gene expression in multiple cancer cell lines. Since the interactions of Integrins with adsorbed matrix proteins are thought to affect the ability of cancer cells to interact with their underlying substrates, we examined the expression of Integrin β1, β3, and β5 in response to matrix rigidity. We found that only Iβ3 increased with increasing substrate modulus. While it was shown that fibronectin greatly affects the expression of tumor-produced factors associated with bone destruction (parathyroid hormone-related protein, PTHrP, and Gli2, poly-l-lysine, vitronectin and type I collagen were also analyzed as potential matrix proteins. Each of the proteins was independently adsorbed on both rigid and compliant polyurethane films which were subsequently used to culture cancer cells. Poly-l-lysine, vitronectin and type I collagen all had negligible effects on PTHrP or Gli2 expression, but fibronectin was shown to have a dose dependent effect. Finally, altering the expression of Iβ3 demonstrated that it is required for tumor cells to respond to the rigidity of the matrix, but does not affect other cell growth or viability. Together these data support the data presented in our manuscript to show that the rigidity of bone drives Integrinβ3/TGF-β crosstalk, leading to increased expression of Gli2 and PTHrP.

  13. A proteomic study of protein variation between osteopenic and age-matched control bone tissue.

    Science.gov (United States)

    Chaput, Christopher D; Dangott, Lawrence J; Rahm, Mark D; Hitt, Kirby D; Stewart, Donald S; Wayne Sampson, H

    2012-05-01

    The focus of this study was to identify changes in protein expression within the bone tissue environment between osteopenic and control bone tissue of human femoral neck patients with osteoarthritis. Femoral necks were compared from osteopenic patients and age-matched controls. A new method of bone protein extraction was developed to provide a swift, clear view of the bone proteome. Relative changes in protein expression between control and osteopenic samples were quantified using difference gel electrophoresis (DIGE) technology after affinity chromatographic depletion of albumin and IgG. The proteins that were determined to be differentially expressed were identified using standard liquid chromatography mass spectrometry (LC/MS/MS) and database searching techniques. In order to rule out blood contamination, blood from age-matched osteoporotic, osteopenic and controls were analyzed in a similar manner. Image analysis of the DIGE gels indicated that 145 spots in the osteopenic bone samples changed at least ± 1.5-fold from the control samples (P proteins were identified by LC/MS/MS. Of the proteins that increased in the osteopenic femurs, two were especially significant: carbonic anhydrase I and phosphoglycerate kinase 1. Apolipoprotein A-I was the most prominent protein that significantly decreased in the osteopenic femurs. The blood samples revealed no significant differences between groups for any of these proteins. In conclusion, carbonic anhydrase I, phosphoglycerate kinase 1 and apolipoprotein A-I appeared to be the most significant variations of proteins in patients with osteopenia and osteoarthritis.

  14. Uncovering Molecular Bases Underlying Bone Morphogenetic Protein Receptor Inhibitor Selectivity.

    Directory of Open Access Journals (Sweden)

    Abdelaziz Alsamarah

    Full Text Available Abnormal alteration of bone morphogenetic protein (BMP signaling is implicated in many types of diseases including cancer and heterotopic ossifications. Hence, small molecules targeting BMP type I receptors (BMPRI to interrupt BMP signaling are believed to be an effective approach to treat these diseases. However, lack of understanding of the molecular determinants responsible for the binding selectivity of current BMP inhibitors has been a big hindrance to the development of BMP inhibitors for clinical use. To address this issue, we carried out in silico experiments to test whether computational methods can reproduce and explain the high selectivity of a small molecule BMP inhibitor DMH1 on BMPRI kinase ALK2 vs. the closely related TGF-β type I receptor kinase ALK5 and vascular endothelial growth factor receptor type 2 (VEGFR2 tyrosine kinase. We found that, while the rigid docking method used here gave nearly identical binding affinity scores among the three kinases; free energy perturbation coupled with Hamiltonian replica-exchange molecular dynamics (FEP/H-REMD simulations reproduced the absolute binding free energies in excellent agreement with experimental data. Furthermore, the binding poses identified by FEP/H-REMD led to a quantitative analysis of physical/chemical determinants governing DMH1 selectivity. The current work illustrates that small changes in the binding site residue type (e.g. pre-hinge region in ALK2 vs. ALK5 or side chain orientation (e.g. Tyr219 in caALK2 vs. wtALK2, as well as a subtle structural modification on the ligand (e.g. DMH1 vs. LDN193189 will cause distinct binding profiles and selectivity among BMP inhibitors. Therefore, the current computational approach represents a new way of investigating BMP inhibitors. Our results provide critical information for designing exclusively selective BMP inhibitors for the development of effective pharmacotherapy for diseases caused by aberrant BMP signaling.

  15. Demineralized Bone Matrix Add-On for Acceleration of Bone Healing in Atypical Subtrochanteric Femoral Fracture: A Consecutive Case-Control Study

    Directory of Open Access Journals (Sweden)

    Noratep Kulachote

    2016-01-01

    Full Text Available Background. Delayed union and nonunion are common complications in atypical femoral fractures (AFFs despite having good fracture fixation. Demineralized bone matrix (DBM is a successfully proven method for enhancing fracture healing of the long bone fracture and nonunion and should be used in AFFs. This study aimed to compare the outcome after subtrochanteric AFFs (ST-AFFs fixation with and without DBM. Materials and Methods. A prospective study was conducted on 9 ST-AFFs patients using DBM (DBM group during 2013-2014 and compared with a retrospective consecutive case series of ST-AFFs patients treated without DBM (2010–2012 (NDBM group, 9 patients. All patients were treated with the same standard guideline and followed up until fractures completely united. Postoperative outcomes were then compared. Results. DBM group showed a significant shorter healing time than NDBM group (28.1 ± 14.4 versus 57.9 ± 36.8 weeks, p=0.04. Delayed union was found in 4 patients (44% in DBM group compared with 7 patients (78% in NDBM group (p>0.05. No statistical difference of nonunion was demonstrated between both groups (DBM = 1 and NDBM = 2, p>0.05. Neither postoperative infection nor severe local tissue reaction was found. Conclusions. DBM is safe and effective for accelerating the fracture healing in ST-AFFx and possibly reduces nonunion after fracture fixation. Trial registration number is TCTR20151021001.

  16. Bone

    Science.gov (United States)

    Helmberger, Thomas K.; Hoffmann, Ralf-Thorsten

    The typical clinical signs in bone tumours are pain, destruction and destabilization, immobilization, neurologic deficits, and finally functional impairment. Primary malignant bone tumours are a rare entity, accounting for about 0.2% of all malignancies. Also benign primary bone tumours are in total rare and mostly asymptomatic. The most common symptomatic benign bone tumour is osteoid osteoma with an incidence of 1:2000.

  17. Protein malnutrition attenuates bone anabolic response to PTH in female rats.

    Science.gov (United States)

    Ammann, P; Zacchetti, G; Gasser, J A; Lavet, C; Rizzoli, R

    2015-02-01

    PTH is indicated for the treatment of severe osteoporosis. Elderly osteoporotic patients frequently suffer from protein malnutrition, which may contribute to bone loss. It is unknown whether this malnutrition may affect the response to PTH. Therefore, the aim of the present study was to assess whether an isocaloric low-protein (LP) diet may influence the bone anabolic response to intermittent PTH in 6-month-old female rats. Six-month-old female rats were either pair fed an isocaloric LP diet (2.5% casein) or a normal-protein (NP) diet (15% casein) for 2 weeks. The rats continued on their respective diet while being treated with 5- or 40-μg/kg recombinant human PTH amino-terminal fragment 1-34 (PTH-[1-34]) daily, or with vehicle for 4 weeks. At the end of this period, areal bone mineral density, bone mineral content, microstructure, and bone strength in axial compression of proximal tibia or 3-point bending for midshaft tibia tests were measured. Blood was collected for the determination of IGF-I and osteocalcin. After 4 weeks of PTH-(1-34), the dose-dependent increase of proximal tibia bone mineral density, trabecular microstructure variables, and bone strength was attenuated in rats fed a LP diet as compared with rats on a NP intake. At the level of midshaft tibia cortical bone, PTH-(1-34) exerted an anabolic effect only in the NP but not in the LP diet group. Protein malnutrition was associated with lower IGF-I levels. Protein malnutrition attenuates the bone anabolic effects of PTH-(1-34) in rats. These results suggest that a sufficient protein intake should be recommended for osteoporotic patients undergoing PTH therapy.

  18. Bone morphogenetic protein-induced heterotopic bone formation: What have we learned from the history of a half century?

    Directory of Open Access Journals (Sweden)

    Takenobu Katagiri, PhD

    2015-05-01

    Full Text Available Bone morphogenetic protein (BMP was originally discovered by Marshall Urist a half century ago following the observation of a unique activity that induced heterotopic bone formation in skeletal muscle tissue. The molecular mechanisms underlying the induction of heterotopic bone formation in skeletal muscle by BMPs were elucidated through the purification and molecular cloning of BMPs and identification of their functional receptors and downstream effectors, as well as from genetic disorders related to BMP activity. BMPs are important regulators of not only skeletal development and regeneration but also the homeostasis of normal skeletal muscle mass. There is still much to learn about the physiology and pathology at the interface of BMPs and skeletal muscle.

  19. Surgical Outcomes of Anterior Cervical Fusion Using Deminaralized Bone Matrix as Stand-Alone Graft Material: Single Arm, Pilot Study.

    Science.gov (United States)

    Chung, Ho-Jung; Hur, Jung-Woo; Ryu, Kyeong-Sik; Kim, Jin-Sung; Seong, Ji-Hoon

    2016-09-01

    To investigate the safety and efficacy of demineralized bone matrix (DBM) as a bone graft substitute for anterior cervical discectomy and fusion (ACDF) surgery. Twenty consecutive patients treated with ACDF using stand-alone polyestheretherketone (PEEK) cages (Zero-P) with DBM(CGDBM100) were prospectively evaluated with a minimum of 6 months of follow-up. Radiologic efficacy was evaluated with a 6-point scoring method for osseous fusion using plain radiograph and computed tomogrpahy scans. Clinical efficacy was evaluated using the visual analogue scale (VAS), Owestry disability index (ODI), and short-form health questionnaire-36. The safety of the bone graft substitute was assessed with vital sign monitoring and a survey measuring complications at each follow-up visit. There were significant improvements in VAS and ODI scores at a mean 6-month follow-up. Six months after surgery, solid fusion was achieved in all patients. Mean score on the 6-point scoring system was 5.1, and bony formation was found to score at least 4 points in all patients. There was no case with implant-related complications such as cage failure or migration, and no complications associated with the use of CGDBM100. ACDF using CGDBM100 demonstrated good clinical and radiologic outcomes. The fusion rate was comparable with the published results of traditional ACDF. Therefore, the results of this study suggest that the use of a PEEK cage packed with DBM for ACDF is a safe and effective alternative to the gold standard of autologous iliac bone graft.

  20. Increase in bone protein components with healing rat fractures: enhancement by zinc treatment.

    Science.gov (United States)

    Igarashi, A; Yamaguchi, M

    1999-12-01

    The alteration in bone components in the femoral-diaphyseal tissues with fracture healing was investigated. Rats were sacrificed 7 and 14 days after the femoral fracture. Protein content in the femoral-diaphyseal tissues was markedly elevated by fracture healing. Analysis with sodium dodecyl sulfate-polyacrylamide gel electrophoresis showed that many protein molecules were induced in the diaphyseal tissues with fracture healing. Moreover, when the femoral-diaphyseal tissues with fracture healing were cultured for 24 and 48 h in a serum-free medium, many proteins in the bone tissues were released into the medium. Also, the culture of the diaphyseal tissues with fracture healing caused a significant increase in bone alkaline phosphatase activity and deoxyribonucleic acid (DNA) content. Meanwhile, the presence of zinc acexamate (10-5 and 10-4 M), a stimulator of bone formation, in a culture medium induced a significant elevation of protein content and alkaline phosphatase activity in the diaphyseal tissues with fracture healing. Such an effect was completely abolished by the presence of cycloheximide (10-6 M), an inhibitor of protein synthesis. The present study suggests that fracture healing induces a newly synthesized bone protein component including stimulatory factor(s) for bone formation. Zinc supplementation may stimulate the healing of femoral fracture.

  1. The incorporation of extracellular matrix proteins in protein polymer hydrogels to improve encapsulated beta-cell function.

    Science.gov (United States)

    Beenken-Rothkopf, Liese N; Karfeld-Sulzer, Lindsay S; Davis, Nicolynn E; Forster, Ryan; Barron, Annelise E; Fontaine, Magali J

    2013-01-01

    Biomaterial encapsulation of islets has been proposed to improve the long-term success of islet transplantation by recreating a suitable microenvironment and enhancing cell-matrix interactions that affect cellular function. Protein polymer hydrogels previously showed promise as a biocompatible scaffold by maintaining high cell viability. Here, enzymatically-crosslinked protein polymers were used to investigate the effects of varying scaffold properties and of introducing ECM proteins on the viability and function of encapsulated MIN6 β-cells. Chemical and mechanical properties of the hydrogel were modified by altering the protein concentrations while collagen IV, fibronectin, and laminin were incorporated to reestablish cell-matrix interactions lost during cell isolation. Rheology indicated all hydrogels formed quickly, resulting in robust, elastic hydrogels with Young's moduli similar to soft tissue. All hydrogels tested supported both high MIN6 β-cell viability and function and have the potential to serve as an encapsulation platform for islet cell delivery in vivo.

  2. Effect of acemannan, an extracted polysaccharide from Aloe vera, on BMSCs proliferation, differentiation, extracellular matrix synthesis, mineralization, and bone formation in a tooth extraction model.

    Science.gov (United States)

    Boonyagul, Sani; Banlunara, Wijit; Sangvanich, Polkit; Thunyakitpisal, Pasutha

    2014-07-01

    Aloe vera is a traditional wound healing medicine. We hypothesized acemannan, a polysaccharide extracted from Aloe vera gel, could affect bone formation. Primary rat bone marrow stromal cells (BMSCs) were treated with various concentrations of acemannan. New DNA synthesis, VEGF, BMP-2, alkaline phosphatase activity, bone sialoprotein, osteopontin expression, and mineralization were determined by [(3)H] thymidine incorporation assay, ELISA, biochemical assay, western blotting, and Alizarin Red staining, respectively. In an animal study, mandibular right incisors of male Sprague-Dawley rats were extracted and an acemannan treated sponge was placed in the socket. After 1, 2, and 4 weeks, the mandibles were dissected. Bone formation was evaluated by dual-energy X-ray absorptiometry and histopathological examination. The in vitro results revealed acemannan significantly increased BMSC proliferation, VEGF, BMP-2, alkaline phosphatase activity, bone sialoprotein and osteopontin expression, and mineralization. In-vivo results showed acemannan-treated groups had higher bone mineral density and faster bone healing compared with untreated controls. A substantial ingrowth of bone trabeculae was observed in acemannan-treated groups. These data suggest acemannan could function as a bioactive molecule inducing bone formation by stimulating BMSCs proliferation, differentiation into osteoblasts, and extracellular matrix synthesis. Acemannan could be a candidate natural biomaterial for bone regeneration.

  3. Parathyroid Hormone-Related Protein, Its Regulation of Cartilage and Bone Development, and Role in Treating Bone Diseases.

    Science.gov (United States)

    Martin, T John

    2016-07-01

    Although parathyroid hormone-related protein (PTHrP) was discovered as a cancer-derived hormone, it has been revealed as an important paracrine/autocrine regulator in many tissues, where its effects are context dependent. Thus its location and action in the vasculature explained decades-long observations that injection of PTH into animals rapidly lowered blood pressure by producing vasodilatation. Its roles have been specified in development and maturity in cartilage and bone as a crucial regulator of endochondral bone formation and bone remodeling, respectively. Although it shares actions with parathyroid hormone (PTH) through the use of their common receptor, PTHR1, PTHrP has other actions mediated by regions within the molecule beyond the amino-terminal sequence that resembles PTH, including the ability to promote placental transfer of calcium from mother to fetus. A striking feature of the physiology of PTHrP is that it possesses structural features that equip it to be transported in and out of the nucleus, and makes use of a specific nuclear import mechanism to do so. Evidence from mouse genetic experiments shows that PTHrP generated locally in bone is essential for normal bone remodeling. Whereas the main physiological function of PTH is the hormonal regulation of calcium metabolism, locally generated PTHrP is the important physiological mediator of bone remodeling postnatally. Thus the use of intermittent injection of PTH as an anabolic therapy for bone appears to be a pharmacological application of the physiological function of PTHrP. There is much current interest in the possibility of developing PTHrP analogs that might enhance the therapeutic anabolic effects. Copyright © 2016 the American Physiological Society.

  4. Recombinant human bone morphogenetic protein-2 suspended in fibrin glue enhances bone formation during distraction osteogenesis in rabbits

    Science.gov (United States)

    Li, Yunfeng; Li, Rui; Hu, Jing; Song, Donghui; Jiang, Xiaowen

    2016-01-01

    Introduction Bone morphogenetic protein-2 (BMP-2) has high potential for bone formation, but its in vivo effects are unpredictable due to the short life time. This study was designed to evaluate the effects of recombinant human (rh) BMP-2 suspended in fibrin on bone formation during distraction osteogenesis (DO) in rabbits. Material and methods The in vitro release kinetics of rhBMP-2 suspended in fibrin was tested using an enzyme-linked immunosorbent assay. Unilateral tibial lengthening for 10 mm was achieved in 48 rabbits. At the completion of osteodistraction, vehicle, fibrin, rhBMP-2 or rhBMP-2 suspended in fibrin (rhBMP-2 + fibrin) was injected into the center of the lengthened gap, with 12 animals in each group. Eight weeks later, the distracted callus was examined by histology, micro-CT and biomechanical testing. Radiographs of the distracted tibiae were taken at both 4 and 8 weeks after drug treatment. Results It was found that fibrin prolonged the life span of rhBMP-2 in vitro with sustained release during 17 days. The rhBMP-2 + fibrin treated animals showed the best results in bone mineral density, bone volume fraction, cortical bone thickness by micro-CT evaluation and mechanical properties by the three-point bending test when compared to the other groups (p < 0.05). In histological images, rhBMP-2 + fibrin treatment showed increased callus formation and better gap bridging compared to the other groups. Conclusions The results of this study suggest that fibrin holds promise to be a good carrier of rhBMP-2, and rhBMP-2 suspended in fibrin showed a stronger promoting effect on bone formation during DO in rabbits. PMID:27279839

  5. Low protein intake magnifies detrimental effects of ovariectomy and vitamin D on bone.

    Science.gov (United States)

    Marotte, Clarisa; Gonzales Chaves, Macarena M S; Pellegrini, Gretel G; Friedman, Silvia M; Lifshitz, Fima; Mandalunis, Patricia; Zeni, Susana N

    2013-08-01

    Protein-induced changes in bone and calcium homeostasis could potentially be greater in the elderly and in women at risk for osteoporosis. We hypothesize that a low protein intake would magnify the negative changes in bone metabolism seen in vitamin D (vitD) insufficiency and/or estrogen deficiency. The present study was undertaken to better understand how a low protein diet along with vitD insufficiency could affect bone metabolism using a rodent ovariectomized (OVX) model. Rats (n = 60) underwent ovariectomy (OVX) or sham operation. The first 15 days after surgery, all rats were fed a standard rodent diet. Thereafter, rats (n = 10/group) were fed a low protein diet (LP; 2.5 %) or a control diet (NP; 12.5 %) with 100 IU% vitD (+D; cholecalciferol) or without vitD (-D) for 45 days. The groups were as follows: SHAM + NP + D (control); SHAM + LP + D; SHAM + LP - D; OVX + NP + D; OVX + LP + D; OVX + LP - D. Body weight (BW) of control and OVX + NP + D groups increased while those feeding the LP diet, independently of vitD feedings, decreased (p bone mineral content, proximal tibia bone mineral density, bone volume and trabecular number levels decreased as follows: SHAM + NP + D (controls) > SHAM + LP + D > OVX + NP + D > SHAM + LP - D > OVX + LP + D > OVX + LP - D (p bone mass and magnified the detrimental effects of vitD and/or estrogen deficiencies.

  6. Effects of ionizing radiation on proteins in demineralized, lyophilized or frozen human bone

    Energy Technology Data Exchange (ETDEWEB)

    Antebi, Uri; Mathor, Monica B., E-mail: uri@usp.br, E-mail: mathor@ipen.br [Instituto de Pesquisas Energeticas e Nucleares (IPEN/CNEN-SP), Sao Paulo, SP (Brazil); Guimaraes, Rodrigo P., E-mail: clinicaguimaraes@gmail.com [Santa Casa de Sao Paulo (FCM/SCSP), Sao Paulo, SP (Brazil). Faculdade de Ciencias Medicas

    2015-07-01

    The aim is the study of the application of ionizing radiation (gamma and electron) as sterilizing agents at doses of 15 kGy, 25 kGy and 50 kGy, the demineralized bone tissue frozen and freeze-dried for use in transplants. Five human femoral diaphysis of different donors demineralized bone tissues were preserved as lyophilized or frozen at - 80 deg C. The samples were divided into non-irradiated groups (control) and irradiated by gamma rays or electron beam. The bone proteins were extracted and used to determine the concentrations of total protein, BMP 2 and 7. It was observed a decrease in total protein concentrations, and BMP 2 and 7. The decrease in total protein concentrations, as compared to respective control groups was significant in the lyophilized and frozen samples irradiated at a dose of 50 kGy gamma radiation and beam electrons with greater than 30% reduction. The significant decrease in the levels of BMP 2 and 7 were also observed in higher doses and especially by electron beam. The reductions in the concentrations of total protein and osteoinductive proteins (BMP 2 and 7), were related to the radiation dose, i.e., increase with higher doses of ionizing radiation type and the type of preservation of the bones. The largest reductions in concentrations were observed in bone irradiated by electron beam and at a dose of 50 kGy. But this type of radiation and this high dose are not usual practice for the sterilization of bone tissue. Keywords: demineralized bone tissue, ionizing radiation, Tissue Bank, BMP 2, BMP 7, bone proteins. (author)

  7. Calcinosis in juvenile dermatomyositis : a possible role for the vitamin K-dependent protein matrix Gla protein

    NARCIS (Netherlands)

    Van Summeren, M. J. H.; Spliet, W. G. M.; Van Royen-Kerkhof, A.; Vermeer, C.; Lilien, M.; Kuis, W.; Schurgers, L. J.

    2008-01-01

    Objectives. The aims of the present study were to investigate whether the calcification inhibitor matrix Gla protein (MGP) is expressed in muscle biopsies of patients with juvenile dermatomyositis (JDM), and whether different forms of MGP are differentially expressed in JDM patients with and without

  8. Distribution of cytoskeletal proteins, integrins, leukocyte adhesion molecules and extracellular matrix proteins in plastic-embedded human and rat kidneys

    NARCIS (Netherlands)

    van Goor, H; Coers, W; van der Horst, MLC; Suurmeijer, AJH

    2001-01-01

    OBJECTIVE: To study the distribution of cytoskeletal proteins (actin, alpha -actinin, vinculin, beta -tubulin, keratin, vimentin, desmin), adhesion molecules for cell-matrix interations (very later antigens [VLA1-6], beta1, beta2 [CD18], vitronectin receptor [alphav beta3], CD 11b), leukocyte adhesi

  9. The use of beta-tricalcium phosphate and bovine bone matrix in the guided tissue regeneration treatment of deep infra-bony defects

    OpenAIRE

    Luković Natalija; Zelić Obrad; Čakić Saša; Petrović Vanja

    2009-01-01

    Introduction. The primary goal of bone regeneration procedures with application of various regenerative biologic agents and biomaterials is to facilitate the formation of periodontal tissues lost as a result of periodontitis. Objective. The aim of the study was to compare clinical outcome of the guided tissue regeneration (GTR) treatment with the use of β-tricalcium phosphate and with bovine bone matrix in human deep intra-osseous defects. Methods. Twenty-one systemically healthy subjects wit...

  10. Promotion of bone growth by dietary soy protein isolate: Comparision with dietary casein, whey hydrolysate and rice protein isolate in growing female rats

    Science.gov (United States)

    The effects of different dietary protein sources(casein (CAS), soy protein isolate (SPI), whey protein hydrolysate (WPH) and rice protein isolate (RPI)) on bone were studied in intact growing female rats and in ovarectomized (OVX) rats showing sex steroid deficiency-induced bone loss. In addition, S...

  11. Histologic and histomorphometric evaluation of osteogenesis induced by octacalcium phosphate (OCP combined with bone matrix gelatin (BMG in rat skull defects

    Directory of Open Access Journals (Sweden)

    Sargolzaei F.

    2005-05-01

    Full Text Available Statement of Problem: Several methods are used to enhance bone repair and new bone formation, and bone matrix gelatin (BMG is recently introduced. Purpose: The purpose of this histologic and histomorphometric study was to assess the osteogenic potential and the quantity of new trabecular bone formation after implantation of OCP and BMG alone and in combination into the cranial defects in rat. Materials and Methods: In this experimental study, 100 young male Sprague Dawley rats (5-6 weeks age and 120-150gr weight were divided into four groups randomly. A full thickness standard trephine defect 5mm in diameter was made in the rat’s parietal bone, and 5mg of OCP, BMG alone and in combination were implanted into the defects. No OCP and BMG particles were implanted in control group which was otherwise treated identically. On the 5th, 7th, 14th, 21st and 56th days after implantation, the rats were killed and bone samples collected. After processing the samples by routine histological procedures, 5µm thick sections of bone were cut and stained with Haematoxyline & Eosin (H&E and Alcian Blue and studied histologically and histomorphometrically using light microscope and eyepiece graticule. The amount of newly formed bone was quantitatively measured by the use of histomorphometric methods. Data were analyzed with SAS statistical package using ANOVA and Duncan tests. Results: In the experimental groups, the new bone formation was initiated from the margin of defects during 5-14 days after implantation. During 14-21 days after implantation, bone marrow cavities and bone marrow tissues in newly formed bone were seen. By the end of the study, the newly formed bone increased and was relatively matured and almost all of the implanted materials were absorbed. In control group, at the end of the study, a few clusters of new bone were seen near to the defect margins and host bone. The histomorphometric analysis indicated statistical significant differences in

  12. Matrix elasticity of void-forming hydrogels controls transplanted-stem-cell-mediated bone formation

    Science.gov (United States)

    Huebsch, Nathaniel; Lippens, Evi; Lee, Kangwon; Mehta, Manav; Koshy, Sandeep T.; Darnell, Max C.; Desai, Rajiv M.; Madl, Christopher M.; Xu, Maria; Zhao, Xuanhe; Chaudhuri, Ovijit; Verbeke, Catia; Kim, Woo Seob; Alim, Karen; Mammoto, Akiko; Ingber, Donald E.; Duda, Georg N.; Mooney, David J.

    2015-12-01

    The effectiveness of stem cell therapies has been hampered by cell death and limited control over fate. These problems can be partially circumvented by using macroporous biomaterials that improve the survival of transplanted stem cells and provide molecular cues to direct cell phenotype. Stem cell behaviour can also be controlled in vitro by manipulating the elasticity of both porous and non-porous materials, yet translation to therapeutic processes in vivo remains elusive. Here, by developing injectable, void-forming hydrogels that decouple pore formation from elasticity, we show that mesenchymal stem cell (MSC) osteogenesis in vitro, and cell deployment in vitro and in vivo, can be controlled by modifying, respectively, the hydrogel’s elastic modulus or its chemistry. When the hydrogels were used to transplant MSCs, the hydrogel’s elasticity regulated bone regeneration, with optimal bone formation at 60 kPa. Our findings show that biophysical cues can be harnessed to direct therapeutic stem cell behaviours in situ.

  13. Adhesion protein networks reveal functions proximal and distal to cell-matrix contacts.

    Science.gov (United States)

    Byron, Adam; Frame, Margaret C

    2016-04-01

    Cell adhesion to the extracellular matrix is generally mediated by integrin receptors, which bind to intracellular adhesion proteins that form multi-molecular scaffolding and signalling complexes. The networks of proteins, and their interactions, are dynamic, mechanosensitive and extremely complex. Recent efforts to characterise adhesions using a variety of technologies, including imaging, proteomics and bioinformatics, have provided new insights into their composition, organisation and how they are regulated, and have also begun to reveal unexpected roles for so-called adhesion proteins in other cellular compartments (for example, the nucleus or centrosomes) in diseases such as cancer. We believe this is opening a new chapter on understanding the wider functions of adhesion proteins, both proximal and distal to cell-matrix contacts.

  14. LRPPRC is a mitochondrial matrix protein that is conserved in metazoans

    Energy Technology Data Exchange (ETDEWEB)

    Sterky, Fredrik H. [Department of Laboratory Medicine, Karolinska Institutet, Retzius vaeg 8, SE-171 77 Stockholm (Sweden); Ruzzenente, Benedetta [Max-Planck-Institut fuer Biologie des Alterns, Gleueler Str. 50a, D-50931 Cologne (Germany); Department of Biology, University of Padova, Via Ugo Bassi 58B, I-35127 Padova (Italy); Gustafsson, Claes M. [Max-Planck-Institut fuer Biologie des Alterns, Gleueler Str. 50a, D-50931 Cologne (Germany); Department of Biochemistry and Cell Biology, University of Gothenburg, P.O. Box 440, SE-405 30 Gothenburg (Sweden); Samuelsson, Tore [Department of Biochemistry and Cell Biology, University of Gothenburg, P.O. Box 440, SE-405 30 Gothenburg (Sweden); Larsson, Nils-Goeran, E-mail: larsson@age.mpg.de [Max-Planck-Institut fuer Biologie des Alterns, Gleueler Str. 50a, D-50931 Cologne (Germany)

    2010-08-06

    Research highlights: {yields} LRPPRC orthologs are restricted to metazoans. {yields} LRPPRC is imported to the mitochondrial matrix. {yields} No evidence of nuclear isoform. -- Abstract: LRPPRC (also called LRP130) is an RNA-binding pentatricopeptide repeat protein. LRPPRC has been recognized as a mitochondrial protein, but has also been shown to regulate nuclear gene transcription and to bind specific RNA molecules in both the nucleus and the cytoplasm. We here present a bioinformatic analysis of the LRPPRC primary sequence, which reveals that orthologs to the LRPPRC gene are restricted to metazoan cells and that all of the corresponding proteins contain mitochondrial targeting signals. To address the subcellular localization further, we have carefully analyzed LRPPRC in mammalian cells and identified a single isoform that is exclusively localized to mitochondria. The LRPPRC protein is imported to the mitochondrial matrix and its mitochondrial targeting sequence is cleaved upon entry.

  15. Human epidermal keratinocyte cell response on integrin-specific artificial extracellular matrix proteins.

    Science.gov (United States)

    Tjin, Monica Suryana; Chua, Alvin Wen Choong; Ma, Dong Rui; Lee, Seng Teik; Fong, Eileen

    2014-08-01

    Cell-matrix interactions play critical roles in regulating cellular behavior in wound repair and regeneration of the human skin. In particular, human skin keratinocytes express several key integrins such as alpha5beta1, alpha3beta1, and alpha2beta1 for binding to the extracellular matrix (ECM) present in the basement membrane in uninjured skin. To mimic these key integrin-ECM interactions, artificial ECM (aECM) proteins containing functional domains derived from laminin 5, type IV collagen, fibronectin, and elastin are prepared. Human skin keratinocyte cell responses on the aECM proteins are specific to the cell-binding domain present in each construct. Keratinocyte attachment to the aECM protein substrates is also mediated by specific integrin-material interactions. In addition, the aECM proteins are able to support the proliferation of keratinocyte stem cells, demonstrating their promise for use in skin tissue engineering.

  16. Matrix protein 2 of influenza A virus blocks autophagosome fusion with lysosomes

    DEFF Research Database (Denmark)

    Gannagé, Monique; Dormann, Dorothee; Albrecht, Randy

    2009-01-01

    Influenza A virus is an important human pathogen causing significant morbidity and mortality every year and threatening the human population with epidemics and pandemics. Therefore, it is important to understand the biology of this virus to develop strategies to control its pathogenicity. Here, we...... demonstrate that influenza A virus inhibits macroautophagy, a cellular process known to be manipulated by diverse pathogens. Influenza A virus infection causes accumulation of autophagosomes by blocking their fusion with lysosomes, and one viral protein, matrix protein 2, is necessary and sufficient...... for this inhibition of autophagosome degradation. Macroautophagy inhibition by matrix protein 2 compromises survival of influenza virus-infected cells but does not influence viral replication. We propose that influenza A virus, which also encodes proapoptotic proteins, is able to determine the death of its host cell...

  17. Bone Morphogenetic Proteins stimulate mammary fibroblasts to promote mammary carcinoma cell invasion.

    Directory of Open Access Journals (Sweden)

    Philip Owens

    Full Text Available Bone Morphogenetic Proteins (BMPs are secreted cytokines that are part of the Transforming Growth Factor β (TGFβ superfamily. BMPs have been shown to be highly expressed in human breast cancers, and loss of BMP signaling in mammary carcinomas has been shown to accelerate metastases. Interestingly, other work has indicated that stimulation of dermal fibroblasts with BMP can enhance secretion of pro-tumorigenic factors. Furthermore, treatment of carcinoma-associated fibroblasts (CAFs derived from a mouse prostate carcinoma with BMP4 was shown to stimulate angiogenesis. We sought to determine the effect of BMP treatment on mammary fibroblasts. A large number of secreted pro-inflammatory cytokines and matrix-metallo proteases (MMPs were found to be upregulated in response to BMP4 treatment. Fibroblasts that were stimulated with BMP4 were found to enhance mammary carcinoma cell invasion, and these effects were inhibited by a BMP receptor kinase antagonist. Treatment with BMP in turn elevated pro-tumorigenic secreted factors such as IL-6 and MMP-3. These experiments demonstrate that BMP may stimulate tumor progression within the tumor microenvironment.

  18. High Protein Intake Improves Insulin Sensitivity but Exacerbates Bone Resorption in Immobility (WISE Study)

    Science.gov (United States)

    Heer, Martina; Smith, Scott M.; Frings-Meuthen, Petra; Zwart, Sara R.; Baecker, Natalie

    2012-01-01

    Inactivity, like bed rest (BR), causes insulin resistance (IR) and bone loss even in healthy subjects. High protein intake seems to mitigate this IR but might exacerbate bone loss. We hypothesized that high protein intake (animal:vegetable protein ratio: 60:40), isocaloric, compared to the control group plus high potassium intake would prevent IR without affecting bone turnover. After a 20-day ambulatory adaptation to controlled confinement and diet, 16 women participated in a 60-day, 6 deg head-down-tilt BR and were assigned randomly to one of the two groups. Control subjects (CON, n=8) received 1g/kg body mass/d dietary protein. Nutrition subjects (NUT, n=8) received 1.45g/kg body mass/d dietary protein plus 7.2g branched chain amino acids per day during BR. All subjects received 1670 kcal/d. Bed rest decreased glucose disposal by 35% (pprotein intake prevented insulin resistance, but exacerbated bed rest induced increase in bone resorption markers C-telopeptide (> 30%) and Ntelopeptide (>20%) (both: pprotein intake. We conclude from these results that high protein intake might positively affect glucose tolerance, but might also foster bone loss. Further long-duration studies are mandatory before high protein intake for diabetic patients, who have an increased fracture risk, might be recommended.

  19. Preparation of bioconjugates by solid-phase conjugation to ion exchange matrix-adsorbed carrier proteins

    DEFF Research Database (Denmark)

    Houen, G.; Olsen, D.T.; Hansen, P.R.;

    2003-01-01

    A solid-phase conjugation method utilizing carrier protein bound to an ion exchange matrix was developed. Ovalbumin was adsorbed to an anion exchange matrix using a batch procedure, and the immobilized protein was then derivatized with iodoacetic acid N-hydroxysuccinimid ester. The activated......, and immunization experiments with the eluted conjugates showed that the more substituted conjugates gave rise to the highest titers of glutathione antibodies. Direct immunization with the conjugates adsorbed to the ion exchange matrix was possible and gave rise to high titers of glutathione antibodies. Conjugates...... of ovalbumin and various peptides were prepared in a similar manner and used for production of peptide antisera by direct immunization with the conjugates bound to the ion exchanger. Advantages of the method are its solid-phase nature, allowing fast and efficient reactions and intermediate washings...

  20. Extra-cellular matrix proteins induce matrix metalloproteinase-1 (MMP-1 activity and increase airway smooth muscle contraction in asthma.

    Directory of Open Access Journals (Sweden)

    Natasha K Rogers

    Full Text Available Airway remodelling describes the histopathological changes leading to fixed airway obstruction in patients with asthma and includes extra-cellular matrix (ECM deposition. Matrix metalloproteinase-1 (MMP-1 is present in remodelled airways but its relationship with ECM proteins and the resulting functional consequences are unknown. We used airway smooth muscle cells (ASM and bronchial biopsies from control donors and patients with asthma to examine the regulation of MMP-1 by ECM in ASM cells and the effect of MMP-1 on ASM contraction. Collagen-I and tenascin-C induced MMP-1 protein expression, which for tenascin-C, was greater in asthma derived ASM cells. Tenascin-C induced MMP-1 expression was dependent on ERK1/2, JNK and p38 MAPK activation and attenuated by function blocking antibodies against the β1 and β3 integrin subunits. Tenascin-C and MMP-1 were not expressed in normal airways but co-localised in the ASM bundles and reticular basement membrane of patients with asthma. Further, ECM from asthma derived ASM cells stimulated MMP-1 expression to a greater degree than ECM from normal ASM. Bradykinin induced contraction of ASM cells seeded in 3D collagen gels was reduced by the MMP inhibitor ilomastat and by siRNA knockdown of MMP-1. In summary, the induction of MMP-1 in ASM cells by tenascin-C occurs in part via integrin mediated MAPK signalling. MMP-1 and tenascin-C are co-localised in the smooth muscle bundles of patients with asthma where this interaction may contribute to enhanced airway contraction. Our findings suggest that ECM changes in airway remodelling via MMP-1 could contribute to an environment promoting greater airway narrowing in response to broncho-constrictor stimuli and worsening asthma symptoms.

  1. The association of Matrix Gla protein isomers with calcification in capsules surrounding silicone breast implants

    OpenAIRE

    Larry W. Hunter; Lieske, John C.; Tran, Nho V.; Miller, Virginia M.

    2011-01-01

    Implanted silicone medical prostheses induce a dynamic sequence of histologic events in adjacent tissue resulting in the formation of a fibrotic peri-prosthetic capsule. In some cases, capsular calcification occurs, requiring surgical intervention. In this study we investigated capsules from silicone gel-filled breast prostheses to test the hypothesis that this calcification might be regulated by the small vitamin K-dependent protein, matrix Gla protein (MGP), a potent inhibitor of arterial c...

  2. Effects of resistance training and protein supplementation on bone turnover in young adult women

    Directory of Open Access Journals (Sweden)

    Sinning Wayne E

    2005-08-01

    Full Text Available Abstract Background The strength of aging bone depends on the balance between the resorption and formation phases of the remodeling process. The purpose of this study was to examine the interaction of two factors with the potential to exert opposing influences on bone turnover, resistance exercise training and high dietary protein intake. It was hypothesized that resistance training by young, healthy, untrained women with protein intakes near recommended levels (0.8 g·kg-1·d-1 would promote bone formation and/or inhibit bone resorption, and that subsequent supplementation to provide 2.4 g protein·kg-1·d-1 would reverse these effects. Methods Bone formation was assessed with serum bone-specific alkaline phosphatase (BAP and osteocalcin (OC, and bone resorption with urinary calcium and deoxypyridinoline (DPD. Biochemical, strength, anthropometric, dietary, and physical activity data were obtained from 24 healthy, untrained, eumenorrheic women (18–29y at baseline, after eight weeks of resistance training (3 d·wk-1, ~1 hr·d-1; 3 sets, 6–10 repetitions, 13 exercises, 75–85% maximum voluntary contraction, and after 12 weeks of resistance training and 10 days of protein/placebo supplementation. Subjects were randomized (double-blind to either a high protein (HP or training control (TC group and, during the final 10 days, consumed either enough purified whey protein to bring daily protein intake to 2.4 g·kg-1·d-1, or an equivalent dose of isoenergetic, carbohydrate placebo. Results Strength, lean tissue mass, and DPD increased significantly in both groups over time, while percent body fat and BAP decreased (repeated measures ANOVA, p ≤ 0.05, Bonferroni correction. No significant changes were observed for serum OC or urinary calcium, and no significant group (TC, HP × time (baseline, week 8, week 12 interactions emerged for any of the biochemical measures. Conclusion (1 Twelve weeks of high-intensity resistance training did not appear to

  3. Extracellular matrix proteins modulate asthmatic airway smooth muscle cell proliferation via an autocrine mechanism

    NARCIS (Netherlands)

    Johnson, Peter R A; Burgess, Janette K; Underwood, P Anne; Au, Wendy; Poniris, Maree H; Tamm, Michael; Ge, Qi; Roth, Michael; Black, Judith L

    2004-01-01

    BACKGROUND: Airway remodeling is a key feature of persistent asthma and includes alterations in the extracellular matrix protein profile around the airway smooth muscle (ASM) and hyperplasia of the ASM. We have previously shown that nonasthmatic ASM cells in culture produce a range of extracellular

  4. Changes of β3 Integrins and Extracellular Matrix Proteins in the Endometrium of Unexplained Infertility

    Institute of Scientific and Technical Information of China (English)

    王化丽; 曲陆荣; 何丽霞; 张颐

    1999-01-01

    The purpose of this study was to investigate changes of β3 integrins and extracellular matrix proteins including fibronectin (FN) , laminin (LN) and collagen type Ⅳ (CL type Ⅳ) on the endometrium of secretory phase from 31 fertile women (fertility group)and 34 women with unexplained infertility (infertility group) by a histochemical method. The results were as follows : In glandular epithelium, β3 integrin appeared in the mid secretory phase and continued to late secretory phase in the fertility group, but was not expressed during the secretory phase in the infertility group.Extracellular matrix proteins from the fertility group were expressed more strongly in mid secretory phase than that in the early secretory phase, and were weakest in the late secretory phase. Compared with the fertility group, the levels of extracellular matrix proteins in the infertility group were elevated in the secretory phase. In conclusion: our current study demonstrate that fie integrin and extracellular matrix proteins are expressed at different levels in the endometrium during the menstrual cycle. They are involved in endometrial changes during the menstrual cycle and during the implantation of the blastocyst. Their unusual expression result in the failure of implantation.

  5. Aligning protein sequence and analysing substitution pattern using a class-specific matrix

    Indian Academy of Sciences (India)

    Hai Song Xu; Wen Ke Ren; Xiao Hui Liu; Xiao Qin Li

    2010-06-01

    Aligning protein sequences using a score matrix has became a routine but valuable method in modern biological research. However, alignment in the ‘twilight zone’ remains an open issue. It is feasible and necessary to construct a new score matrix as more protein structures are resolved. Three structural class-specific score matrices (all-alpha, all-beta and alpha/beta) were constructed based on the structure alignment of low identity proteins of the corresponding structural classes. The class-specific score matrices were significantly better than a structure-derived matrix (HSDM) and three other generalized matrices (BLOSUM30, BLOSUM60 and Gonnet250) in alignment performance tests. The optimized gap penalties presented here also promote alignment performance. The results indicate that different protein classes have distinct amino acid substitution patterns, and an amino acid score matrix should be constructed based on different structural classes. The class-specific score matrices could also be used in profile construction to improve homology detection.

  6. Vitamin K-dependent carboxylation of matrix gla protein influences the risk of calciphylaxis

    Science.gov (United States)

    Matrix Gla protein (MGP) is a potent inhibitor of vascular calcification. The ability of MGP to inhibit calcification requires the activity of a vitamin K-dependent enzyme, which mediates MGP carboxylation. We investigated how MGP carboxylation influences the risk of calciphylaxis in adult patients ...

  7. BIBF1120 inhibits fibroblasts proliferation and production of the extracellular matrix protein fibulin-1

    NARCIS (Netherlands)

    Burgess, Janette; Munk, Lizzie; Jaffar, Jade; Black, Judith; Oliver, Brian

    2015-01-01

    Introduction: In patients with idiopathic pulmonary fibrosis (IPF) transforming growth factorbeta 1 (TGFβ1) induces excessive extracellular matrix (ECM) protein deposition leading to fibrosis. Our recent studies have shown that the glycoprotein fibulin-1 is increased in serum and lung tissue from pa

  8. Inflammatory Bowel Disease in a Rodent Model Alters Osteocyte Protein Levels Controlling Bone Turnover.

    Science.gov (United States)

    Metzger, Corinne E; Narayanan, Anand; Zawieja, David C; Bloomfield, Susan A

    2017-04-01

    Bone loss is a common comorbidity of inflammatory bowel disease (IBD), leading to elevated fracture risk in these patients. Inflammatory factors associated with IBD cause increased bone resorption and decreased bone formation with multiple factors implicated as instigators of these alterations. In this project, we examined the influence of IBD on osteocyte proteins in male rats (2 months old) divided into two groups: induced gut inflammation via 2,4,6-trinitrobenzenesulfonic acid (TNBS) enema, and vehicle control. We examined the prevalence of two pro-inflammatory cytokines, tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6), an anti-inflammatory cytokine, interleukin-10 (IL-10), the anabolic factor insulin-like growth factor-I (IGF-I), osteoclastogenesis regulators RANKL and OPG, and the bone formation inhibitor sclerostin in osteocytes in three bone compartments 4 weeks after initiation of gut inflammation. Histomorphometry of the proximal tibia and fourth lumbar vertebra revealed lower bone volume, lower bone formation rate (BFR), lower osteoid surface (OS), and higher osteoclast surface (Oc.S) with TNBS. Tibial mid-shaft periosteal BFR was also lower with TNBS. Immunohistochemical staining of the distal femur demonstrated that %TNF-α(+) , %IL-6(+) , %RANKL(+) , and %OPG(+) osteocytes were elevated in cancellous bone in TNBS animals compared to vehicle. These changes were coincident with increased bone resorption. With regression analysis, %RANKL(+) osteocytes statistically predicted the increase in cancellous Oc.S (R(2)  = 0.565). Increased %sclerostin(+) osteocytes observed in the TNBS treatment predicted declines in cancellous OS (R(2)  = 0.581) as well as BFR in cancellous and cortical bone (R(2)  = 0.674, R(2)  = 0.908, respectively). Contrary to our hypothesis, %IGF-I(+) osteocytes increased in TNBS animals. In conclusion, the IBD model produced a systemic inflammation that altered the regulatory protein profile in osteocytes that

  9. A comparative study of the proliferation and osteogenic differentiation of human periodontal ligament cells cultured on β-TCP ceramics and demineralized bone matrix with or without osteogenic inducers in vitro.

    Science.gov (United States)

    An, Shaofeng; Gao, Yan; Huang, Xiangya; Ling, Junqi; Liu, Zhaohui; Xiao, Yin

    2015-05-01

    The repair of bone defects that result from periodontal diseases remains a clinical challenge for periodontal therapy. β-tricalcium phosphate (β-TCP) ceramics are biodegradable inorganic bone substitutes with inorganic components that are similar to those of bone. Demineralized bone matrix (DBM) is an acid-extracted organic matrix derived from bone sources that consists of the collagen and matrix proteins of bone. A few studies have documented the effects of DBM on the proliferation and osteogenic differentiation of human periodontal ligament cells (hPDLCs). The aim of the present study was to investigate the effects of inorganic and organic elements of bone on the proliferation and osteogenic differentiation of hPDLCs using three-dimensional porous β-TCP ceramics and DBM with or without osteogenic inducers. Primary hPDLCs were isolated from human periodontal ligaments. The proliferation of the hPDLCs on the scaffolds in the growth culture medium was examined using a Cell-Counting kit-8 (CCK-8) and scanning electron microscopy (SEM). Alkaline phosphatase (ALP) activity and the osteogenic differentiation of the hPDLCs cultured on the β-TCP ceramics and DBM were examined in both the growth culture medium and osteogenic culture medium. Specific osteogenic differentiation markers were examined using reverse transcription-quantitative polymerase chain reaction (RT-qPCR). SEM images revealed that the cells on the β-TCP were spindle-shaped and much more spread out compared with the cells on the DBM surfaces. There were no significant differences observed in cell proliferation between the β-TCP ceramics and the DBM scaffolds. Compared with the cells that were cultured on β-TCP ceramics, the ALP activity, as well as the Runx2 and osteocalcin (OCN) mRNA levels in the hPDLCs cultured on DBM were significantly enhanced both in the growth culture medium and the osteogenic culture medium. The organic elements of bone may exhibit greater osteogenic differentiation effects

  10. Effect of nano-hydroxyapatite/collagen composite and bone morphogenetic protein-2 on lumbar intertransverse fusion in rabbits

    Institute of Scientific and Technical Information of China (English)

    孙天胜; 关凯; 时述山; 朱兵; 郑永军; 崔福斋; 张伟; 廖素三

    2004-01-01

    Objective: To investigate the effect of nano-hydroxyapatite/collagen (nHA/collagen) composite as a graft extender and enhancer when combined with recombinant human bone morphogenetic protein-2 (rhBMP-2) on lumbar intertransverse fusion in rabbits.Methods: Sixty-four adult female New Zealand white rabbits, aged 1 year and weighing 3.5-4.5 kg, underwent similar posterolateral intertransverse process arthrodesis and were randomly divided into 4 groups based on different grafts: autogenous cancellous bone alone (ACB group), nHA/collagen alone (HAC group), half autogenous cancellous bone and half nHA/collagen (ACB+HAC group) and nHA/collagen combined with rhBMP-2 (HAC+BMP group). The fusion masses were analyzed by manual palpation, radiography, biomechanical testing and histological examination. Results: Fusion was observed in 4 cases in the 6th week and in 5 cases in the 10th week after surgery in ACB group. No case showed fusion in HAC group. In ACB+HAC group, there was fusion in 3 cases in the 6th week and in 4 cases in the 10th week after surgery. In HAC+BMP group, fusion in 1 case was found in the 4th week, in 5 cases in the 6th week and in 6 cases in the 10th week after surgery. It suggested that ACB, ACB+HAC and HAC+BMP groups showed similar fusion ratio and mechanical strength in the 6th and 10th week after surgery. According to the microstructure analysis of the samples, nHA/collagen had no negative effect when implanted together with ilium autograft. In HAC+BMP group, new bone-like tissue was observed in the 2nd week postoperatively, and nearly all of the implanted composites were replaced by mature bone matrix and new bones in 10th week postoperatively.Conclusions: The nHA/collagen, especially combined with rhBMP-2, is a promising bone substitute, for it has quick biodegradation, fine bone-bending ability, and high osteoconductivity on posterolateral spinal fusion in rabbits.

  11. Altered profiles of nuclear matrix proteins during the differentiation of human gastric mucous adenocarcinoma MGc80-3 cells

    Institute of Scientific and Technical Information of China (English)

    Chun-Hong Zhao; Qi-Fu Li

    2005-01-01

    AIM: To find and identify specific nuclear matrix proteins associated with proliferation and differentiation of carcinoma cells, which will be potential markers for cancer diagnosis and targets in cancer therapy.METHODS: Nuclear matrix proteins were selectively extracted from MGc80-3 cells treated with or without hexamethylamine bisacetamide (HMBA), and subjected to 2-D gel electrophoresis. The resulted protein patterns were analyzed by Melanie software. Spots of nuclear matrix proteins differentially expressed were excised and subjected to in situ digestion with trypsin. Peptide masses were obtained by matrix-assisted laser-desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS) analysis and submitted for database searching using Mascot tool.RESULTS: The MGc80-3 cells were induced into differentiation by HMBA. There were 22 protein spots which changed remarkably in the nuclear matrix, from differentiation of MGc80-3 cells compared to control.Eleven of which were identified. Seven proteins -actin, prohibitin, porin 31HL, heterogeneous nuclear ribonucleoprotein A2/B1, vimentin, ATP synthase, and heatshock protein 60 were downregulated, whereas three proteins - heat shock protein gp96, heat shock protein 90-beta, and valosin-containing protein were upregulated,and the oxygen-regulated protein was only found in the differentiated MGc80-3 cells.CONCLUSION: The induced differentiation of carcinoma cells is accompanied by the changes of nuclear matrix proteins. Further characterization of those proteins will show the mechanism of cellular proliferation and differentiation, as well as cancer differentiation.

  12. Mechanical Loading Improves Tendon-Bone Healing in a Rabbit Anterior Cruciate Ligament Reconstruction Model by Promoting Proliferation and Matrix Formation of Mesenchymal Stem Cells and Tendon Cells

    Directory of Open Access Journals (Sweden)

    Fanglong Song

    2017-02-01

    Full Text Available Background/Aims: This study investigated the effect of mechanical stress on tendon-bone healing in a rabbit anterior cruciate ligament (ACL reconstruction model as well as cell proliferation and matrix formation in co-culture of bone-marrow mesenchymal stem cells (BMSCs and tendon cells (TCs. Methods: The effect of continuous passive motion (CPM therapy on tendon-bone healing in a rabbit ACL reconstruction model was evaluated by histological analysis, biomechanical testing and gene expressions at the tendon-bone interface. Furthermore, the effect of mechanical stretch on cell proliferation and matrix synthesis in BMSC/TC co-culture was also examined. Results: Postoperative CPM therapy significantly enhanced tendon-bone healing, as evidenced by increased amount of fibrocartilage, elevated ultimate load to failure levels, and up-regulated gene expressions of Collagen I, alkaline phosphatase, osteopontin, Tenascin C and tenomodulin at the tendon-bone junction. In addition, BMSC/TC co-culture treated with mechanical stretch showed a higher rate of cell proliferation and enhanced expressions of Collagen I, Collagen III, alkaline phosphatase, osteopontin, Tenascin C and tenomodulin than that of controls. Conclusion: These results demonstrated that proliferation and differentiation of local precursor cells could be enhanced by mechanical stimulation, which results in enhanced regenerative potential of BMSCs and TCs in tendon-bone healing.

  13. Fos/AP-1 proteins in bone and the immune system.

    Science.gov (United States)

    Wagner, Erwin F; Eferl, Robert

    2005-12-01

    The skeleton and the immune system share a variety of different cytokines and transcription factors, thereby mutually influencing each other. These interactions are not confined to the bone marrow cavity where bone cells and hematopoietic cells exist in proximity but also occur at locations that are target sites for inflammatory bone diseases. The newly established research area termed 'osteoimmunology' attempts to unravel these skeletal/immunological relationships. Studies towards a molecular understanding of inflammatory bone diseases from an immunological as well as a bone-centered perspective have been very successful and led to the identification of several signaling pathways that are causally involved in inflammatory bone loss. Induction of receptor activator of nuclear factor (NF)-kappaB ligand (RANKL) signals by activated T cells and subsequent activation of the key transcription factors Fos/activator protein-1 (AP-1), NF-kappaB, and NF for activation of T cells c1 (NFATc1) are in the center of the signaling networks leading to osteoclast-mediated bone loss. Conversely, nature has employed the interferon system to antagonize excessive osteoclast differentiation, although this counteracting activity appears to be overruled under pathological conditions. Here, we focus on Fos/AP-1 functions in osteoimmunology, because this osteoclastogenic transcription factor plays a central role in inflammatory bone loss by regulating genes like NFATc1 as well as the interferon system. We also attempt to put potential therapeutic strategies for inflammatory bone diseases in perspective.

  14. The role of the BH3-only protein Noxa in bone homeostasis.

    Science.gov (United States)

    Idrus, Erik; Nakashima, Tomoki; Wang, Ling; Hayashi, Mikihito; Okamoto, Kazuo; Kodama, Tatsuhiko; Tanaka, Nobuyuki; Taniguchi, Tadatsugu; Takayanagi, Hiroshi

    2011-07-08

    Bone homeostasis is maintained by a dynamic balance between bone resorption by osteoclasts and bone formation by osteoblasts. Since excessive osteoclast activity is implicated in pathological bone resorption, understanding the mechanism underlying osteoclast differentiation, function and survival is of both scientific and clinical importance. Osteoclasts are monocyte/macrophage lineage cells with a short life span that undergo rapid apoptosis, the rate of which critically determines the level of bone resorption in vivo. However, the molecular basis of rapid osteoclast apoptosis remains obscure. Here we report the role of a BH3-only protein, Noxa (encoded by the Pmaip1 gene), in bone homeostasis using Noxa-deficient mice. Among the Bcl-2 family members, Noxa was selectively induced during osteoclastogenesis. Mice lacking Noxa exhibit a severe osteoporotic phenotype due to an increased number of osteoclasts. Noxa deficiency did not have any effect on the number of osteoclast precursor cells or the expression of osteoclast-specific genes, but led to a prolonged survival of osteoclasts. Furthermore, adenovirus-mediated Noxa overexpression remarkably reduced bone loss in a model of inflammation-induced bone destruction. This study reveals Noxa to be a crucial regulator of osteoclast apoptosis, and may provide a molecular basis for a new therapeutic approach to bone diseases.

  15. Effects of cell-attachment and extracellular matrix on bone formation in vivo in collagen-hydroxyapatite scaffolds.

    Directory of Open Access Journals (Sweden)

    Max M Villa

    Full Text Available Cell-based tissue engineering can be used to replace missing or damaged bone, but the optimal methods for delivering therapeutic cells to a bony defect have not yet been established. Using transgenic reporter cells as a donor source, two different collagen-hydroxyapatite (HA scaffolds, and a critical-size calvarial defect model, we investigated the effect of a cell-attachment period prior to implantation, with or without an extracellular matrix-based seeding suspension, on cell engraftment and osteogenesis. When quantitatively compared, the in-house scaffold implanted immediately had a higher mean radiopacity than in-house scaffolds incubated overnight. Both scaffold types implanted immediately had significantly higher area fractions of donor cells, while the in-house collagen-HA scaffolds implanted immediately had higher area fractions of the mineralization label compared with groups incubated overnight. When the cell loading was compared in vitro for each delivery method using the in-house scaffold, immediate loading led to higher numbers of delivered cells. Immediate loading may be preferable in order to ensure robust bone formation in vivo. The use of a secondary ECM carrier improved the distribution of donor cells only when a pre-attachment period was applied. These results have improved our understanding of cell delivery to bony defects in the context of in vivo outcomes.

  16. Clinical evaluation of a combined regenerative technique with enamel matrix derivative, bone grafts, and guided tissue regeneration.

    Science.gov (United States)

    Harris, Randall J; Harris, Laura E; Harris, Christopher R; Harris, Anne J

    2007-04-01

    The goal of this study was to evaluate the clinical changes obtained when intra-bony defects were treated with an enamel matrix derivative (EMD), a bone graft, and guided tissue regeneration. Fifty patients with a periodontal defect not associated with a furcation and with an attachment loss of at least 7.0 mm were included in this study. Full-thickness flaps were reflected, the roots were planed, EMD was applied, a demineralized freeze-dried bone allograft combined with EMD was placed, a bioabsorbable membrane was placed, and more EMD was applied. The defect areas were then sutured. At a mean of 5.3 months after treatment, there was a mean increase in recession of 0.7 mm, a mean reduction in probing depth of 5.7 mm, and a mean gain in attachment level of 5.0 mm. In this study there was more recession in smokers than in nonsmokers and in defects associated with anterior teeth. Additionally, the deeper defects (those with greater probing depths and attachment level loss) had the greatest reductions in probing depth and gains in attachment level. Based on this study, this technique proved itself to be an effective method to improve the clinical situation when treating periodontal defects not involving furcations.

  17. Associations of total, dairy, and meat protein with markers for bone turnover in healthy, prepubertal boys

    DEFF Research Database (Denmark)

    Budek, Alicja Zofia; Hoppe, Camilla; Michaelsen, Kim Fleischer

    2007-01-01

    We previously reported that high intake of milk, but not meat, equal in protein content, increased serum insulin-like growth factor-I (sIGF-I) in prepubertal boys. sIGF-I plays a key role in bone metabolism. Therefore, the aim of this cross-sectional study was to investigate associations of total...... focus on differential effects of dairy and meat protein on bone health during growth.......We previously reported that high intake of milk, but not meat, equal in protein content, increased serum insulin-like growth factor-I (sIGF-I) in prepubertal boys. sIGF-I plays a key role in bone metabolism. Therefore, the aim of this cross-sectional study was to investigate associations of total......, dairy, and meat protein intake with markers for bone turnover and sIGF-I in prepubertal, healthy boys (n ¼ 81). We measured bone turnover (enzyme-linked immunoassay) in serum osteocalcin (sOC), bone-specific alkaline phosphatase (sBAP), and C-terminal telopeptide of collagen type-I (sCTX); dietary...

  18. Controlled protein delivery from electrospun non-wovens: novel combination of protein crystals and a biodegradable release matrix.

    Science.gov (United States)

    Puhl, Sebastian; Li, Linhao; Meinel, Lorenz; Germershaus, Oliver

    2014-07-07

    Poly-ε-caprolactone (PCL) is an excellent polymer for electrospinning and matrix-controlled drug delivery combining optimal processability and good biocompatibility. Electrospinning of proteins has been shown to be challenging via the use of organic solvents, frequently resulting in protein unfolding or aggregation. Encapsulation of protein crystals represents an attractive but largely unexplored alternative to established protein encapsulation techniques because of increased thermodynamic stability and improved solvent resistance of the crystalline state. We herein explore the electrospinning of protein crystal suspensions and establish basic design principles for this novel type of protein delivery system. PCL was deployed as a matrix, and lysozyme was used as a crystallizing model protein. By rational combination of lysozyme crystals 0.7 or 2.1 μm in diameter and a PCL fiber diameter between 1.6 and 10 μm, release within the first 24 h could be varied between approximately 10 and 100%. Lysozyme loading of PCL microfibers between 0.5 and 5% was achieved without affecting processability. While relative release was unaffected by loading percentage, the amount of lysozyme released could be tailored. PCL was blended with poly(ethylene glycol) and poly(lactic-co-glycolic acid) to further modify the release rate. Under optimized conditions, an almost constant lysozyme release over 11 weeks was achieved.

  19. Comparative proteomic analysis of extracellular matrix proteins secreted by hypertrophic scar with normal skin fibroblasts

    Directory of Open Access Journals (Sweden)

    Li Ma

    2014-04-01

    Full Text Available The formation of hypertrophic scars (HSs is a fibroproliferative disorder of abnormal wound healing. HSs usually characterize excessive proliferation of fibroblasts, abnormal deposition of extracellular matrix (ECM during wound healing, associated with cosmetic, functional, and psychological problems. Owing to the role of ECM proteins in scar formation, we comparatively analyzed matrix proteins secreted by normal skin fibroblasts (NSFs and HS fibroblasts (HSFs. The acetone-extracted secreted proteins were separated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE, and identified by mass spectrometry (MS. Based on Go annotation of MS data, the profiling of ECM proteins was established and scar-related proteins have been screened out. The functions of several ECM proteins identified by MS have been discussed, such as collagens I, VI, XII, fibronectin, decorin, lumican, and protein procollagen C endopeptidase enhancer 1 (PCPE-1. Among them, the MS result of PCPE-1 was supported by Western blotting that PCPE-1 from HSFs were significantly upregulated than that from NSFs. It is suggested that PCPE-1 could be a potential target for scar treatment. The exploration of scar related proteins may provide new perspectives on understanding the mechanism of scar formation and open a new way to scar treatment and prevention.

  20. Combined effects of porous hydroxyapatite and demineralized bone matrix on bone induction: in vitro and in vivo study using a nude rat model

    Energy Technology Data Exchange (ETDEWEB)

    Lee, Jae Hyup; Lee, Kyung-Mee; Baek, Hae-Ri; Jang, Soo-Jeong; Lee, Ji-Ho [Department of Orthopedic Surgery, Seoul National University School of Medicine, SMG-SNU Boramae Medical Center, Seoul 156-707 (Korea, Republic of); Ryu, Hyun-Seung, E-mail: spinelee@snu.ac.kr [Research and Development Center, CGBio Inc., Seong-Nam 462-120 (Korea, Republic of)

    2011-02-15

    Hydroxyapatite (HA) is an osteoconductive material used as a bone graft extender and demineralized bone matrix (DBM) has been used as a source of osteoinductive factors. A combination of DBM and HA is expected to create a composite with both osteoconductive and osteoinductive properties. This study examined the effect of a combination of DBM and HA on osteogenesis both in vitro and in vivo using an athymic nude rat abdominal muscle pouch model, and evaluated the possibility of HA as a carrier of DBM. Alkaline phosphatase (ALP) staining, ALP assay and measurements of the mRNA expression of ALP and Runx2 by RT-PCR were performed by transplanting human mesenchymal stem cells onto a plate. Five athymic nude rats each were assigned to one of two experimental groups (DBM/HA putty and only HA, i.e. 15 pouches per group). The muscle pouches were filled with DBM/HA putty or only HA. Radiographs were obtained at weeks 4 and 8, postoperatively. The animals were sacrificed at week 8 postoperatively and high resolution microCT was used to confirm the newly formed mineralized tissue. Each pouch was fixed, embedded, sectioned and processed for hematoxylin and eosin staining. The ALP value of the DBM/HA putty was higher than those of HA and control (p < 0.05, each). The expression of ALP mRNA appeared higher on the DBM/HA putty than on HA and control. MicroCT and histology examinations of the DBM/HA putty demonstrated the presence of newly generated mineralized tissues but there was no mineralized tissue in the HA cases. In conclusion, the DBM/HA putty indicated osteoblastic differentiation in vitro and showed ectopic mineralized tissue formation in the rat abdominal pouch model. These findings indicate that the DBM/HA putty can retain its oteoinductivity and HA can be used as a carrier of DBM.

  1. Proteomic analysis of a pleistocene mammoth femur reveals more than one hundred ancient bone proteins

    DEFF Research Database (Denmark)

    Cappellini, Enrico; Jensen, Lars Juhl; Szklarczyk, Damian Milosz

    2012-01-01

    We used high-sensitivity, high-resolution tandem mass spectrometry to shotgun sequence ancient protein remains extracted from a 43 000 year old woolly mammoth (Mammuthus primigenius) bone preserved in the Siberian permafrost. For the first time, 126 unique protein accessions, mostly low-abundance...

  2. Extraction and Purification of Matrix Protein from the Nacre of Pearl Oyster Pinctada fucata

    Institute of Scientific and Technical Information of China (English)

    MA Caiping; ZHANG Cen; NIE Yancheng; XIE Liping; ZHANG Rongqing

    2005-01-01

    A soluble matrix protein P14 with an apparent molecular mass of 14.5 kDa was isolated from fragmented nacre of pearl oysters (Pinctada fucata) treated with 10% NaOH solution to investigate the nacre matrix proteins and their effect on the CaCO3 crystal. The protein was characterized by gel exclusion chromatography and reversed-phase high performance liquid chromatography after demineralization by 10% acetic acid. The X-ray diffraction pattern of P14 crystals indicates that P14 plays an important role in nacre biomineralization. P14 can induce aragonite formation, stimulate CaCO3 crystal formation, and accelerate aragonite precipitation. Heating of the acid insoluble nacre residue, which was named conchiolin, in 10% sodium dodecyl sulfate solution supplemented with 10% β-mercaptoethanol solution for 10-20 min at about 100(C gave two other soluble proteins having molecular masses of 19.4 kDa and 25.0 kDa. The present study suggests that these two proteins are linked to the insoluble organic matrix by disulfide bridges because the extraction yield increases when β-mercaptoethanol is added to the medium.

  3. Adhesion properties of Lactobacillus rhamnosus mucus-binding factor to mucin and extracellular matrix proteins.

    Science.gov (United States)

    Nishiyama, Keita; Nakamata, Koichi; Ueno, Shintaro; Terao, Akari; Aryantini, Ni Putu Desy; Sujaya, I Nengah; Fukuda, Kenji; Urashima, Tadasu; Yamamoto, Yuji; Mukai, Takao

    2015-01-01

    We previously described potential probiotic Lactobacillus rhamnosus strains, isolated from fermented mare milk produced in Sumbawa Island, Indonesia, which showed high adhesion to porcine colonic mucin (PCM) and extracellular matrix (ECM) proteins. Recently, mucus-binding factor (MBF) was found in the GG strain of L. rhamnosus as a mucin-binding protein. In this study, we assessed the ability of recombinant MBF protein from the FSMM22 strain, one of the isolates of L. rhamnosus from fermented Sumbawa mare milk, to adhere to PCM and ECM proteins by overlay dot blot and Biacore assays. MBF bound to PCM, laminin, collagen IV, and fibronectin with submicromolar dissociation constants. Adhesion of the FSMM22 mbf mutant strain to PCM and ECM proteins was significantly less than that of the wild-type strain. Collectively, these results suggested that MBF contribute to L. rhamnosus host colonization via mucin and ECM protein binding.

  4. Infant formula promotes bone grow