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At least 19 recordsLinked to original sources
PubMed1992

Influence of short-term aluminum exposure on demineralized bone matrix induced bone formation.

The effects of aluminum exposure on bone formation employing the demineralized bone matrix (DBM) induced bone development model were studied using 4-week-old Sprague-Dawley rats injected with a saline (control) or an aluminum chloride (experimental) solution. After 2 weeks of aluminum treatment, 20-mg portions of rat DBM were implanted subcutaneously on each side in the thoracic region of the control and experimental rats. Animals were killed 7, 12, or 21 days after implantation of the DBM and the developing plaques removed. No morphological, histochemical, or biochemical differences were apparent between plaques from day 7 control and experimental rats. Plaques from day 12 control and experimental rats exhibited cartilage formation and alkaline phosphatase activity localized in osteochondrogenic cells, chondrocytes, osteoblasts, and extracellular matrix. Unlike the plaques from control rats that contained many osteoblastic mineralizing fronts, the plaques from the 12-day experimental group had a preponderance of cartilaginous tissue, no evidence of mineralization, increased levels of alkaline phosphatase activity, and a reduced calcium content. Plaques developing for 21 days in control animals demonstrated extensive new bone formation and bone marrow development, while those in the experimental rats demonstrated unmineralized osteoid-like matrix with poorly developed bone marrow. Alkaline phosphatase activity of the plaques continued to remain high on day 21 for the control and experimental groups. Calcium levels were significantly reduced in the experimental group. These biochemical changes correlated with histochemical reductions in bone calcification.(ABSTRACT TRUNCATED AT 250 WORDS)

Alkaline Phosphatase↗
PubMed1992

Effect of a pulsing electromagnetic field on demineralized bone-matrix-induced bone formation in a bony defect in the premaxilla of rats.

A 2-mm non-healing bony defect was prepared in the premaxilla of male Wistar rats weighing about 180 g as a simulation of an alveolar cleft, for determination of whether a pulsing electromagnetic field (PEMF) could promote regeneration of bone induced by demineralized bone matrix (DBM). The defect was either treated with 7 mg DBM or was left as a non-grafted control. The rats were exposed to a PEMF with a frequency of 100 Hz, a 10-ms-wide burst with 100 microseconds-wide quasi-rectangular pulses, repeating at 15 Hz, and magnetic field strength of 1.5-1.8 G. Alkaline phosphatase activity increased significantly from day 7 in the DBM-graft-plus-PEMF group and from day 10 in the DBM-graft group, reaching a maximum on day 14. A greater-than-two-fold rise in alkaline phosphatase activity and a three-fold rise in the amount of 45Ca incorporation in the DBM-graft-plus-PEMF group were attained compared with those of the DBM-graft group. The DBM-graft-plus-PEMF group produced more bone with almost complete osseous bridging in the defect sites than did the group treated with DBM only on day 35. The findings indicate that PEMF had an enhancing effect on the bone-inductive properties of the DBM through the stimulation of osteoblast differentiation induced by DBM.

Alkaline Phosphatase↗
PubMed1992

Bone morphogenetic protein induces bone in the squirrel monkey, but bone matrix does not.

Demineralized bone matrix (DBM) reproducibly induces extraskeletal bone formation in rodents, but its effects in dogs and primates are negative or uncertain. In previous studies on the squirrel monkey, DBM did not induce bone, although the same implants were effective in nude rats. In the present study, the DBM was augmented with recombinant human bone morphogenetic protein-2 (BMP-2). Bone was formed in 10 of 12 monkeys, as verified by histology and calcium content. However, in 4 monkeys, the induced bone mass appeared smaller than the original implant. DBM controls induced microscopic amounts of bone in 2 out of 10 monkeys. In the nude rats, all DBM controls and augmented implants induced bone. The difficulties in achieving bone induction in higher animals may be overcome, at least partially, by using a higher concentration of the inductive protein than is present in DBM.

Animals↗
PubMed1979

Effect of preimplantation treatment on the bone-forming potential of decalcified allogeneic and xenogeneic bone-matrix implants.

Bone-forming property of 0.6 M HCl decalcified (a) allogeneic bone matrix preserved in 70% alcohol, (b) allogeneic bone matrix preserved in anaesthetic ether, (c) allogeneic 'Ossein' provided by the Leather Research Institute, Madras, and (d) xenogeneic bone-matrix preserved in alcohol was studied by fitting the implants in surgically created complete circumferential osteo-periosteal gaps in the ulna of rabbits. Bone formation was assessed radiologically, macroscopically, histologically, and by tetracycline fluorescence up to 16 postimplantation weeks. Successful bridging of the gap by new bone formation was observed in 75% of (a) and 28.6% of (d) preserved up to 2 weeks. Ether-preserved implants did not induce bone formation. The 'Ossein' implants remained as inert material neither invaded by host cells nor inducing any bone formation. The xenogeneic implants exhibited local immune response which was probably responsible for poor osteogeneic response. Bone forming quality of bone-matrix implants appears to be influenced by the chemical treatment during preparation and preservation, host cellular response and immune reaction invoked by the implant.

Animals↗
PubMed1979

Detection of collagen degradation products from subcutaneously implanted organic bone matrix.

Demineralized bovine bone powder was reduced with NaB3H4 to label the collagen crosslinks with tritium. The powder was enclosed in small nylon mesh pouches and implanted subcutaneously into rats for 3 weeks. Histological examinations revealed that multinuclear giant cells accumulated around the bone matrix, some in Howship's lacunae. Collagenous peptides containing intermolecular crosslinks were detected in the urea-soluble fraction extracted from the implant. Two crosslink-containing peptides were isolated from a dialyzable fraction: one contained dihydroxylysinonorleucine and the other hydroxylysinonorleucine. Both peptides had molecular weights of approximately 1000 estimated from the elution positions of gel filtration chromatography; and both had similar quantitative compositions of amino acids. There were no homologous peptides detected in a control experiment of the reduced bone matrix which was incubated in vitro with buffered saline for 1 week at 37 degrees C.

Animals↗
PubMed1992

Factors influencing synthesis and mineralization of bone matrix from fetal bovine bone cells grown in vitro.

This study of the in vitro synthesis and mineralization of bovine bone demonstrates that sheets of mineralized matrix can be produced consistently within 18-24 days of cell isolation. Mineralization surpasses that achieved by other systems with other species: The deposition of mineral extends beyond nodules to form branching trabeculae and then solid wafers of bone. Comparison of the fetal age of the bone source, enzyme digestion methods, seeding density, culture surface, nutritive media, and concentration of fetal calf serum and other additives, including insulin and ascorbic acid, has yielded a set of optimal culture conditions. In the presence of ascorbic acid and beta-glycerol phosphate, insulin has a dose-dependent effect on the morphology of the mineralized bone matrix produced. Quantitative analysis shows that in these cultures calcium accumulates most rapidly between days 6 and 10 after the introduction of mineralization medium but that mineral accretion continues throughout 14-16 days of culture. Alkaline phosphatase levels rise up to 200-fold, concomitant with a rapid increase in the number of cells per culture during the early mineralization phases; both fall as mineralization proceeds. This system has been used to study the induction of mRNA of type I collagen, alkaline phosphatase, and several noncollagenous bone proteins during the course of mineralization. Because of the degree of mineralization achieved with this system, it has many potential applications.

Alkaline Phosphatase↗
PubMed1992

Ultrastructural observation of calcification preceding new bone formation induced by demineralized bone matrix gelatin.

Demineralized bone matrix gelatin (BMG) was implanted into the skeletal muscle of Sprague-Dawley rats, and the resulting ultrastructural changes of the BMG were examined 3, 5, 7, 10, 15 and 20 days later. Most of the implanted BMG became calcified 7-20 days after implantation. Calcification ('acellular mineral deposition') was first observed as needle-shaped crystalline deposits in the BMG matrix on day 7 after implantation, which gradually increased in size and fused with one another in some deposits. They appeared to be divided into small partitions consisting of denatured collagen fibers unlike those of noncalcified BMG. Some deposits had electron-lucent areas in their center adjacent to well-calcified peripheral areas, and the central area contained many collagen-like fibers and spherical vacuoles. Osteoblast-like cells were not present around these calcified deposits 7 days after implantation. After that, new bone formation was often seen near the area of acellular mineral deposition, and the fused calcified deposits remained until day 15 after implantation. The noncalcified BMG was mainly absorbed by macrophages, and the sites of acellular mineral deposition were absorbed by multinucleated cells resembling osteoclasts which were considered to be activated by the implants. Acellular mineral deposition probably started as calcium and phosphate deposits on some materials in the BMG matrix, and by heterogenic nucleation without osteoblasts or matrix vesicles, inducing bone formation. Thus the BMG may be not only a carrier of bone morphogenetic protein, gradually supplying it to the surrounding tissue but also a storage site for minerals that are indispensable for bone formation.(ABSTRACT TRUNCATED AT 250 WORDS)

Alkaline Phosphatase↗
PubMed1992

In situ hybridization of bone matrix proteins in undecalcified adult rat bone sections.

We have developed a method for in situ hybridization of adult bone tissue utilizing undecalcified sections and have used it to histologically examine the mRNA expression of non-collagenous bone matrix proteins such as osteocalcin (bone Gla protein, BGP), matrix Gla protein (MGP), and osteopontin in adult rats. Expression was compared with that in bone tissues of newborn rats. In the adult bone tissue, osteocalcin mRNA was strongly expressed in periosteal and endosteal cuboidal osteoblasts but not in primary spongiosa near the growth plate. Osteopontin mRNA was strongly expressed in cells present on the bone resorption surface, osteocytes, and hypertrophic chondrocytes, but not in cuboidal osteoblasts on the formation surface. Osteopontin and osteocalcin mRNAs were expressed independently and the distribution of cells expressing osteopontin mRNA corresponded with acid phosphatase-positive mononuclear cells and osteoclasts. Expression of MGP mRNA was noted only in hypertrophic chondrocytes. In newborn rat bone tissues, expression of osteocalcin mRNA was much weaker than in adult rat bone tissues. These results clearly indicate the differential expression of mRNAs of non-collagenous bone matrix proteins in adult rat bone tissues.

Animals↗
PubMed1992

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

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.

Alkaline Phosphatase↗
PubMed1990

[Vertebral trabecular bone in various age groups and in osteoporosis-- morphometry and bone matrix biochemistry].

Vertebral trabecular bone was analysed by morphometry and bone matrix biochemistry. Trabecular bone volume (TBV) and mean trabecular plate thickness (MTPT) decreased with age. TBV was significantly correlated with MTPT and mean trabecular plate density (MTPD). The individual structure of trabecular bone could be described by both MTPT and MTPD together, but changes of these parameters, that were pathognomonic for osteopenia, were not found. By measuring TBV 3 cases of severe osteopenia were identified (TBV less than 2s of controls); 2 of them showed matrix abnormalities so far not described. In one case (a 67 year old woman without risk factors for osteoporosis) an abnormal high content of type III collagen was found, in the other case (a 44 year old woman with acromegaly) bone matrix analysis atypically revealed a significant fraction of type II collagen. Further studies will be needed to assess the pathogenetic or diagnostic importance of these new findings.

Adult↗
PubMed1992

Osteoclast recruitment in response to human bone matrix is age related.

The effect of bone matrix age on the recruitment and differentiation of osteoclast precursors was studied using the chick chorioallantoic membrane (CAM) implant system. Devitalized mineralized bone particles (75-250 microns) were prepared from human femoral cortical bone obtained postmortem from 8 men (age range: 18-72 years). The particles were implanted onto the CAM and 8 days later implants were harvested and processed for light microscopic, morphometric or immunohistochemical analysis. Histomorphometric analysis was performed on samples representing each donor age. The analysis was grouped into three categories consisting of bone from young adults (18-20 years), adults (34-53 years) and aged individuals (67 years and older). Total osteoclast number, osteoclast number per bone particle, cell area, cell size, number of nuclei per cell profile, nucleocytoplasmic ratio, and the presence of a distinctive osteoclast antigen defined by monoclonal anti-body 121F were determined. Bone matrix from older individuals, and therefore the oldest age group (67 years and older), elicited significantly fewer multinucleated cells when compared to bone matrix from younger donors. The number of nuclei per cell profile was highest in the adult population (34-53 years), and there was a continuous increase in cell area with aging. As a consequence, the nucleocytoplasmic ratio decreased from the youngest to the oldest age group. These findings indicate that, relative to factors that affect the recruitment and differentiation of osteoclast precursor cells, bone matrix of older individuals is changed in quality and/or quantity compared to bone matrix from younger individuals. It is hypothesized that this decline in osteoclast formation in response to older bone matrix may contribute to the impaired bone remodeling associated with aging.

Adolescent↗
PubMed1992

Demineralized bone matrix polydioxanone composite as a substitute for bone graft: a comparative study in rats.

Demineralized bone matrix (DBM) has been successfully used as a substitute for bone grafting. Autogenous bone grafts (ABG) may cause donor site morbidity and undergo significant resorption. DBM may overcome these problems but is mechanically unstable when originally placed. We explored using a slowly resorbable template, polydioxanone (PDS), in combination with DBM and compared it to ABG in a rat 9 x 9 mm cranial defect model. After both 1 and 3 months, histologically and biochemically well-formed bone was present in ABG/PDS and DBM/PDS-treated defects, but not in control defects (PDS alone). Mechanical push-out tests using a servohydraulic testing frame were conducted. Maximum load before failure of DBM/PDS increased from 65% at 1 month to 100% of that of intact skull at 3 months. In contrast, ABG/PDS was 50% as strong as DBM/PDS and not significantly stronger than PDS alone. ABG/DBM did not significantly increase in strength from 1 to 3 months. We conclude that DBM/PDS is better than ABG/PDS in treating cranial defects in the rat model, and that an absorbable osteoinductive bone substitute with superior mechanical advantage is possible without the disadvantages of ABG.

Animals↗
PubMed1975

Transmembrane bone matrix gelatin-induced differentiation of bone.

In response to chemically-defined bone matrix gelatin (BMG) inside a diffusion chamber implanted in a muscle pouch, mesenchymal cells migrate directionally, aggregate and differentiate into new bone, on the outside of the chamber. BMG diffuses through double membranes 275 to 300 mum in thickness. The inner membrane of pore size is 0.025 mum and the outer membrane of pore size is 0.45 mum. The inner membrane is 1/20 the pore size and the combination is twice the thickness of membranes previously reported to transfer osteoinductive activity of living cells. Autoradiographs show 35S-cysteine-labelled BMG produces very high trans-membrane grain counts while 3H-proline labelled BMG produces very low transmembrane grain counts. Electron micrographs demonstrate that gelatin-derived, uranyl-acetate-stained fine granules interspersed with ruthenium red-staining coarse granules, diffuse through the membrane of 0.025 mum pore size from the inside out. Solitary pale-staining collagen fibrils, possibly formed in interstitial fluid by renaturation of BMG are found in the interior of the chamber and in the interior of the outer 0.45 mum but not the inner 0.025 mum pore size membrane. Densely-stained new bone collagen fiber bundles cover the outer membrane, fill the 0.45 mum subsurface pores for a depth of 0.20 to 30 mum, and thereby attach the new cartilage and bone deposits to the outer surface of the chamber. BMG powders solubilize rapidly in diffusion chambers and produce high yields of new bone. The relationship between denatured collagen and renatured gelatin fibrils in the process of transfer of the bone morphogen from BMG to mesenchymal cell receptors is an intriguing subject for further investigation.

Animals↗
PubMed1992

Compositional analysis of the collagenous bone matrix. A study on adult normal and osteopenic bone tissue.

The collagenous constituents of mature bone of 30 individuals 22-93 years of age were studied by post-mortem morphological and biochemical analysis. Morphometric evaluation of the second lumbar vertebral body revealed striking interindividual differences in bone mass, mean trabecular density and mean trabecular thickness. Collagen extracted from vertebral trabecular bone by limited pepsin digestion consisted mainly of collagen I (92%) and collagen V (8%). Immunohistochemistry revealed a distinct distribution of these two collagen types within the bone matrix. The degree of lysyl hydroxylation of the alpha 2(I) collagen chain correlated inversely with the trabecular bone volume (TBV) and with the mean trabecular plate density. This correlation was statistically significant for the entire study group as well as for the female and male subgroups. Within the female subgroup, the lysyl hydroxylation/TBV ratio was higher in postmenopausal than in premenopausal women and was highest in women with established osteoporosis. No significant correlation was found between the level of lysyl hydroxylation and the age of the patients. The alpha 1(I) collagen chain showed a nearly constant degree of lysyl hydroxylation in all 30 samples. The results provide convincing evidence that morphometric changes associated with osteopenia in adult bone are accompanied by an altered level of lysyl hydroxylation of the alpha 2(I)-chain of collagen I. The biochemical alterations observed may be responsible for the deposition of a deficient bone matrix in osteopenic conditions.

Adult↗
PubMed1979

Proteochondroitin sulfate synthesized in cartilages induced in vivo and in vitro by bone matrix gelatin.

Implanted allogeneic demineralized bone matrix gelatin induced sequential development of cartilage and bone in the recipient rat muscle tissue. Proteoglycans of the implants labeled in vivo with [35S]sulfate at different stages of development were analyzed by sucrose density gradient centrifugation. The major proteoglycan synthesized in day-5 implant, just prior to onset of chondrogenesis, was a dermatan sulfate-containing proteoglycan with relatively slow sedimentation rate. Additionally, a small amount of a faster sedimenting component could be detected. The faster sedimenting proteoglycan, in which chondroitin 4-sulfate accounted for 85% of total radioactivity, became predominant in day-10 sample when cartilage formation was maximal. By day 30, when cartilage had been replaced by newly formed bone, the synthesis of this faster sedimenting component had ceased. A similar, if not identical, proteoglycan was found to be a major one synthesized by the in vitro-induced cartilage. This proteoglycan was smaller in overall size and shorter in length of its chondroitin sulfate chains than a major proteoglycan component obtained from neonatal rat epiphyseal cartilage. Concurrent with these changes in proteoglycan type, there appeared to be a change in collagen type, since type II collagen, in addition to type I collagen, was synthesized in day-10 implant. These results indicate that the proteoglycan can be used as a molecular marker for chondrogenesis by bone matrix gelatin.

Abdominal Muscles↗
PubMed1979

Identification of organic phosphorus covalently bound to collagen and non-collagenous proteins of chicken-bone matrix. The presence of O-phosphoserine and O-phosphothreonine in non-collagenous proteins, and their absence from phosporylated collagen.

Non-collagenous phosphoproteins, almost all of which can be extracted in EDTA at neutral pH in the presence of proteinase inhibitors, are identified in the matrix of chicken bone, and are therefore not covalently bound to collagen. Similarly, all the peptides containing gamma-carboxyglutamic acid are present in the EDTA extract and none in the insoluble residue, confirming that none is covalently linked to chicken bone collagen. However, organic phosphorus is also found to be present in chicken bone collagen, principally in the alpha2-chains. Of the total protein-bound organic phosphorus present in chicken bone matrix, approx. 80% is associated with the non-collagenous proteins and 20% with collagen. The soluble non-collagenous proteins contain both O-phosphoserine and O-phosphothreonine and these account for essentially of their organic phosphorus content. In contrast, collagen contains neither O-phosphoserine nor O-phosphothreonine. Indeed, no phosphorylated hydroxy amino acid, phosphoamidated amino acid or phosphorylated sugar could be identified in purified components of collagen, which contain approximately four to five atoms of organic phosphorus per molecule of collagen. Peptides containing organic phosphorus were isolated from partial acid hydrolysates and enzymic digests of purified collagen components, which contain an as-yet-unidentified cationic amino acid. These data, the very high concentrations of glutamic acid in the phosphorylated peptides, and the pH-stability of the organic phosphorus moiety in intact collagen chains strongly suggest that at least part of the organic phosphorus in collagen is present as phosphorylated glutamic acid. This would indicate that the two major chemically different protein fractions in chicken bone matrix that contain organic phosphorus may represent two distinct metabolic pools of organic phosphorus under separate biological control.

1-Carboxyglutamic Acid↗
PubMedDate not supplied

Diffusion of bone morphogenetic activity from the residue of collagenase digested bone matrix gelatin through interstitial fluid.

Bone morphogenetic activity is transmitted from the residue of a collagenase digest of bone matrix gelatin not only across cellulose acetate membranes but also through an interstitial fluid filled duplex diffusion chamber (a distance 300 + 2,000 mum). Collagenolysis enhances dissociation of the bone morphogenetic property of bone matrix and dissemination among mesenchymal cells proliferating in the host bed surrounding the diffusion chamber. The bone morphogenetic response is associated with secretion of interstitial fluid, enzymes, and fibrin as well as formation of new collagen fibrils beaded with coarse ruthenium red granules in the pores of the cellulose acetate membrane. Membranes with pore sizes too small to accommodate either new collagen fibrils or mesenchymal cell microvilli do not transmit the morphogenetic response.

Animals↗
PubMed1992

Implants of heterologous demineralized bone matrix for induction of posterior spinal fusion in rats.

The authors tested the osteoinductive capacity of powdered heterologous (bovine) demineralized bone matrix in rats. The first part of the study concerned a monolateral posterior spinal implant after decortication of three vertebrae, using as a control area the animal's contralateral side, in which neither bone graft nor any other material were placed. In another group of rats, a comparative evaluation was made of powdered heterologous demineralized bone matrix and fresh autologous bone. In the same animal, autologous bone was implanted to realize a thoracic posterior fusion and demineralized bone matrix, to induce a posterior fusion in the lumbar area. All data obtained suggested a good osteoinductive activity of heterologous powdered demineralized bone matrix. The two posterior spinal fusions done in the same animal with heterologous demineralized bone matrix or autologous bone, respectively, had similar callus development and required the same time for formation.

Animals↗