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PubMed1999

Staphylococcus aureus adhesion to bone matrix and bone-associated biomaterials.

Staphylococcus aureus is a frequent cause of orthopedic infections in humans. The bacterium expresses several adhesins that facilitate bacterial binding to the bone matrix and to bone implant biomaterials coated with host plasma constituents. The relevant S. aureus adhesins are termed microbial surface components recognizing adhesive matrix molecules (MSCRAMMs) and specific MSCRAMMs are involved in bone and joint infections.

Adhesins, Bacterial↗
PubMed2003

Osseous healing using injectable calcium sulfate-based putty for the delivery of demineralized bone matrix and cancellous bone chips.

Three formulations of injectable calcium sulfate-based putties containing demineralized bone matrix (DBM), 50% DBM/50% cancellous bone (CB) chips, and 30% DBM/70% CB were studied in canines. Four humeral defects per dog were implanted with one of each of the putty formulations while the fourth defect was left untreated. After 6 weeks, the dogs were euthanized. Radiographs and histology showed that the area fraction of new bone in the defects was greater for the three putty formulations than the untreated defects. The area of residual cancellous bone graft remaining in the defects was <10% in both CB putties. Residual calcium sulfate was not apparent in any of the histological sections. We conclude that fast-resorbing calcium sulfate-based putties are effective delivery means of bone graft materials for the successful restoration of bony defects.

Animals↗
PubMed2003

Denaturation of demineralized bone matrix significantly reduces bone formation by guided tissue regeneration.

AIM: To examine in a discriminating capsule model whether denaturation of demineralized bone matrix (DBM) by heating may influence bone formation. MATERIALS AND METHODS: DBM was produced from the long bones of rats. Half the portion of DBM was denatured by heating in distilled water for 20 min at temperatures between 70 degrees C and 90 degrees C. Prior to the study, the destruction of the osteoinductive properties of the DBM was confirmed in three rats following intramuscular implantation. Thirty, 4-month-old, male albino rats of the Wistar strain were used in the study. Following surgical exposure of the mandibular ramus, a hemispherical Teflon capsule (internal diameter = 5.0 mm) was placed, with its open part facing the lateral aspect of the ramus. On one side (test side), the capsule was loosely packed with denatured DBM, while on the contralateral side, serving as control, the capsule was loosely packed with the same amount of non-denatured DBM. After healing periods of 30, 60, and 120 days, groups of 10 animals were killed and 40-70 microm thick undecalcified sections of the capsules were produced. Three sections from each specimen, representing the mid-portion of the capsule, were subjected to histological analysis and computer-assisted planimetric measurements. RESULTS: Increasing amounts of newly formed bone were observed in both test and control capsules during the experimental period. At 4 months, the new bone formed in the control capsules occupied 46.7% of the cross-sectional area of the capsules, while it was only 19.1% in the test capsules (P<0.05). CONCLUSION: Denaturation of DBM by heating significantly reduces bone formation by guided tissue regeneration.

Animals↗
PubMed2001

Human demineralised bone matrix as a bone substitute for reconstruction of cystic defects of the lower jaw.

In a retrospective study validated by a standardized clinical and radiological examination, the bone regeneration in 90 patients with cystic mandibular defects was examined. In 50 patients bony defect reconstructions with human demineralised bone matrix (HDBM) were carried out, while in a comparable group of 40 patients the hollow pockets were left to regenerate bone spontaneously. The bone regeneration after the implantation of human demineralised bone matrix (HDBM) was subjected to a comparative validation. Osteoinductive proteins present in HDBM (bone morphogenetic proteins) can diffuse into the implant seat and induce new bone formation (osteoinduction). A markedly faster and more thorough bone regeneration was demonstrated after the surgical therapy of cystic mandibular lesions with HDBM than without. HDBM also proved to be exceptionally biocompatible.

Journal Article↗
PubMed1999

Histomorphometrical analysis of bone formed in human maxillary sinus floor elevations grafted with OP-1 device, demineralized bone matrix or autogenous bone. Comparison with non-grafted sites in a series of case reports.

Bone morphogenetic proteins have proven to be effective bone inductors in animals and are therefore promising as inductors of bone formation in humans. In the present study we investigated the tissue formed after grafting osteogenic protein 1 on a collagen carrier (OP-1-device) in the human sinus floor elevation procedure. Three patients were grafted with OP-1 device. For comparison 3 groups of 3 patients were included in the study receiving respectively, autogenous bone, human freeze-dried demineralized bone matrix (DBM) or no graft. This last group had a sufficient alveolar bone height for dental implantation. Six months after grafting, at the time of implantation, biopsies were taken from the grafted area and/or the future dental positions. Undecalcified sections were used for histological and histomorphometrical analysis. All grafted sinuses showed an increased osteoid percentage when compared to non-grafted sinuses. Autogenous bone grafts all showed lamellar bone formation. In the DBM grafts mostly woven bone had been formed, predominantly by what appeared to be osteo-conduction. The OP-1 device gave rise to bone formation in 2 of the 3 patients. After 6 months implants could only be placed in 1 out of the 3 patients treated with OP-1 device. This patient showed mature lamellar bone formation, comparable to autogenous bone grafts. In the second patient all bone found was woven and the presence of a high osteoid percentage and large osteocyte lacunae indicated that this was recently-formed bone. Remnants of the collagen carrier were rare and new bone was never found against them, suggesting that this bone was formed by osteo-induction. In the third patient no new bone had been formed. The device had been encapsulated with fibrous tissue and inflammatory reaction was present. We conclude that in the human sinus floor elevation OP-1 has potential bone inductive capacity, but that results in the 3 patients tested with the current OP-1 device are inconsistent.

Adult↗
PubMed1996

Aging effect on inductive capacity of human demineralized bone matrix.

Demineralized bone powder (DBP) prepared from human cortical bone was implanted into subcutaneous pouches of athymic Nu/Nu mice for 28 days. The osteoinductive capacity was evaluated by histomorphometry of the induced cartilage and bone, and by alkaline phosphatase activity in the implant. Very small amounts of new bone and cartilage were found at histological analysis, confirming that human DBP is much less osteoinductive than that from other species. Whereas the morphometric data of the implants from the young and aged donors were not significantly different, the alkaline phosphatase activity was significantly lower in the implants from the old donors than from the younger ones. This difference between the morphometric and biochemical results could reflect the fact that the enzymatic activity is already present in the osteoprogenitor cells. At 28 days, the osteoblastic activity in contact with DBP from the aged group is characterized by a decrease in the enzymatic amount which is not yet visible at the tissue level. This tendency to a decrease in the osteoinductive capacity of bone matrix is an additional aspect of the age-related alterations which occur in bone tissue and could be attributed to modifications of different proteins of the bone matrix, including bone morphogenetic protein.

Aging↗
PubMed1989

Influence of testosterone and dihydrotestosterone on bone-matrix induced endochondral bone formation.

A bone matrix-induced endochondral bone development model has been used to study the effects of androgens on different stages of bone development in castrated young adult rats. Androgen treatment, especially with dihydrotestosterone (DHT) for 7 days, inhibited 35SO4 incorporation by the developing cartilage in the induced plaques. Castrated control animals maintained for 11 days after implantation of bone matrix showed significantly lower calcium levels in the induced implant than was observed earlier in the unoperated controls. DHT treatment for 11 days caused dramatic increases in levels of calcium in the implants. Testosterone had little effect. When androgen treatment was continued for 21 days, while levels of alkaline phosphatase in the implants were unaffected, levels of calcium in the implants were significantly higher than on day 11 for both castrated control and androgen-treated animals. Peak alkaline phosphatase activity (day 10) is known to precede peak calcium mineralizing activity (day 12) in this model and it is also known that calcium levels remain high thereafter. Evaluation of calcium and alkaline phosphatase levels in the proximal tibial metaphyses of castrated control and androgen-treated groups of animals showed no changes after 11 days treatment. Prolonged treatment (21 days) elevated the levels of alkaline phosphatase whereas no change was observed in calcium levels in the tibial metaphyses. These findings demonstrate that androgens stimulate mineralization and that DHT is more active when used for short periods of time and in early stages of bone development in matrix-induced implants.

Alkaline Phosphatase↗
PubMed2003

[An experimental study of demineralized bone matrix to repair bone defects as a scaffold of tissue engineering].

OBJECTIVE: To evaluate application of the sponge of demineralized bone matrix (SDBM) in tissue engineering of bone. METHODS: SDBM was prepared from long bone of rabbits. Bone marrow cells were flushed from the bone shaft of femurs of a two-month-old New Zealand white rabbit. After the cells were cultured for 9 days, the flasks were added into dexamethasone (10(-8) mol/L), beta-glycerophosphate sodium (10 mmol/L) and L-ascorbic acid (50 micrograms/ml). After 5 weeks, the cultured cells were collected and marked by 5-Bromo-2'-dexyouridine (BrdU). The grand sum of cells seeded on a piece of SDBM was about (4-6) x 10(6). The composites of cells and SDBM (tissue engineered chip, TEC) were implanted into muscles and bone defects of radius in rabbits. A standard procedure was applied to make a 10 mm long defect bilaterally in the radius of nine skeletally mature male New Zealand white rabbits. All of the 18 defects were randomly divided into three groups: group I, six defects were grafted by TEC; group II, six defects were grafted with SDBM alone; group III, six defects were empty. RESULTS: The results of radiographic and histological evaluation showed that all of the defects were repaired in group I and group II at 6 weeks, none of the defects was repaired in group III. The results of BrdU staining showed that the staining was positive in group I, but negative in group II. Biomechanical test showed that the compressive ultimate strength (CUS) of new bone in TEC implanted group was comparable with normal radius (P = 0.623) and in SDBM implanted group was significant lower than normal radius (P = 0.038). CONCLUSIONS: The TEC can form cartilage and bone tissue in muscles and repair segmental bone defects. SDBM is a kind of effective natural scaffold in tissue engineering of bone.

Animals↗
PubMed2002

The effect of demineralized bone matrix gel on bone ingrowth and fixation of porous implants.

The presence of demineralized bone matrix (DBM) gel did not enhance or accelerate attachment strength or bone ingrowth and resulted in a significant decrease in implant interface attachment strength at 3 weeks. Hydroxyapatite (HA) coating resulted in significant increases in interface shear strength and bone ingrowth compared with non-HA-coated porous implants at all time periods. The HA-coated implants achieved greater attachment strength and bone ingrowth at earlier time periods and maintained greater attachment strength at long-term periods. The results of this study indicate that in the presence of a good bone-implant interference fit, there is no beneficial effect in applying DBM gel to a porous-coated or HA-coated porous implant surface. The small amount that can be applied and the degree of osteoinductivity of DBM seem to preclude it from having a significant biologic effect.

Animals↗
PubMed1990

Heterotopic new bone formation causes resorption of the inductive bone matrix.

The bone matrix of growing rats was labeled by multiple injections of 3H-proline, and demineralized bone matrix (DBM) was prepared. The DBM was allotransplanted heterotopically into growing rats. New bone formation was induced in and around the implants. The new bone formation was accompanied by a decrease in the content of 3H; 20 and 30 days after implantation, 72% and 46%, respectively, of the activity remained in the implants. Daily injections of indomethacin (2 mg/kg) inhibited calcium uptake by about 20% at 20 and 30 days and inhibited the release of 3H from the DBM to a similar degree. Heterotopic bone induction by DBM is accompanied by matrix resorption, and inhibition of the new bone formation decreases the resorption of DBM.

Animals↗
PubMed1990

Transforming growth factor-beta stimulates bone matrix apposition and bone cell replication in cultured fetal rat calvariae.

Transforming growth factor-beta (TGF beta) stimulates the expression of extracellular matrix proteins and may be a local regulator of bone growth. The aims of this research were to localize the effect of TGF beta on bone matrix formation and to determine if this effect was dependent on increased cell replication, using histomorphometry and autoradiography of bone organ cultures. Half-calvariae of 21-day-old fetal rats were cultured with native or recombinant TGF beta 1 for 24 h and labeled either with [3H]proline for 0-24 or 24-48 h or with [3H]thymidine for the last 6 h of culture. Bones were fixed in glutaraldehyde, embedded in glycol methacrylate, and processed for autoradiography. Bone matrix formation was assessed as the matrix apposition rate per day and the percentage of [3H]proline-labeled bone surface. Cell replication was evaluated based on the number and percentage of [3H]thymidine labeled cells in the osteoblast cell zone, the osteoprogenitor cell zone, and the pericranial fibroblastic periosteum. Both native and recombinant TGF beta at 1-30 ng/ml increased bone matrix formation by 25-40% (P less than 0.05). At 30 ng/ml, TGF beta had a generalized mitogenic effect as cell replication increased by approximately 2-fold in all cell zones of the pericranial periosteum. TGF beta had specific effects on bone cell differentiation. The number of unlabeled cells lining the bone surface increased, and the number of osteoclasts on bone decreased. Inhibition of cell replication by hydroxyurea only partially blocked the stimulatory effect of TGF beta on bone matrix formation, suggesting that TGF beta may have independent effects on cell replication and differentiated bone cell function. In summary, TGF beta had a generalized mitogenic effect on the pericranial periosteum and specific stimulatory and inhibitory effects on bone cell differentiation and function.

Animals↗
PubMed1991

A radiographical and biomechanical study of demineralized bone matrix implanted into a bone defect of rat femurs with and without bone marrow.

Repair of large bone defects represents a challenge to orthopedic surgery since autogenous graft is not available in large amounts. Demineralized bone matrix (DBM) which contains bone morphogenic protein, a potent osteoinductive glycoprotein, and collagen, an osteoconductive matrix, may be an effective substitute for these graft materials. Bone marrow which contains osteoprogenitor cells could potentiate the osteoinductive and osteoconductive properties of demineralized bone matrix. This study tested the ability of demineralized bone matrix with and without bone marrow to bridge large segmental defects, and evaluated the results both radiographically and biomechanically as compared to autogenous (isogeneic) cancellous bone graft. Demineralized bone-matrix segments implanted into a plated femoral segmental defect in rats resulted in firm union in most animals. Bone marrow significantly enhanced bone formation of demineralized bone-matrix implants at an early stage but with time, differences between bone marrow-augmented and bone marrow-deprived demineralized bone implants were no longer demonstrable radiographically and biomechanically. Newly formed bone had about 50% of the strength of the contralateral control bones. Femurs implanted with cancellous bone isografts had similar evidence of absolute union rate, radiographic and mechanical properties as DBM-implanted femurs.

Animals↗
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↗
PubMed1987

Vertebral fusion dynamics in the thoracic and lumbar spine induced by allogenic demineralized bone matrix combined with autogenous bone marrow. An experimental study in rabbits.

The aim of this study was to examine the capability of grafts consisting of demineralized bone matrix combined with autogenous bone marrow to enhance fusion in the thoracic and lumbar spines in rabbits. Thirteen rabbits were operated on, using a posterior approach. A combined intertransverse and interspinous fusion was performed. Fusion was achieved first in the lumbar segments, then in the thoraco-lumbar segments and lastly in the thoracic segments. The best results were achieved in the lumbar spine, where complete fusion occurred in 83%. The combination of demineralized bone matrix and autogenous bone marrow proved to be a reliable transplant material for experimental spinal fusion.

Animals↗
PubMed1991

Analysis of bone morphogenetic protein (BMP) derived from human and bovine bone matrix.

Recently, Bone Morphogenetic Protein (BMP) has attracted the attention of a number of investigators, but its elucidation remains incomplete. At present, the determination of its amino acid sequence, which is necessary for its synthesis, and screening for a carrier that allows BMP to be effective in small amounts are unsolved problems. Bone morphogenetic protein is studied here to clarify its clinical applications. BMP was extracted from human and bovine bone matrix with 4 M guanidine-HCl and purified by liquid chromatography. Acrylamide electrophoresis (SDS-PAGE) and isoelectric focusing (IEF) showed that the purified BMP was homogeneous. We used type I collagen as the carrier in the bioassay. This BMP induced new bone in situ three weeks after implantation in muscle pouches in Wistar rats. The molecular weights of human and bovine bone matrix-derived BMP are 17.0 and 18.0 kDa by SDS-PAGE, and pI values for both are 4.9 by IEF. Human and bovine bone matrix-derived BMP are peptides containing 165 and 163 amino acids, respectively, according to amino acid analysis. The NH2-terminal sequence of bovine bone matrix-derived BMP was obtained from the bovine band, electroblotted onto polyvinylidene difluoride membrane, that corresponded to the final purified fraction. The sequence differs from previously designated BMPs25 and other proteins reported to have similar activity, but the physicochemical characteristics are comparable to the native preparations.

Amino Acid Sequence↗
PubMed1993

Spinal fusion induced by porous hydroxyapatite blocks (HA). Experimental comparative study with HA, demineralized bone matrix and autogenous bone marrow.

Numerous materials have been studied and recently used in clinical practice as synthetic bone biosubstitutes to integrate or even replace autogenous bone grafts. This study consisted of an experimental vertebral fusion model in rabbits using porous hydroxyapatite (HA), both alone and in various combinations with demineralized bone matrix (DBM), as a vehicle for the osteoinductive agent (Bone Morphogenetic Protein-BMP), and with autogenous bone marrow (BM), which supplies BMP-sensitive reticular cells. The animals were divided into two main groups on the basis of the physical form of the biomaterial implanted: HA blocks and HA granules, which in turn included several subgroups based on the composition of the implants (HA, HA+DBM, HA+BM, HA+DBM+BM). There were also control groups with no implant or with bone autografts (ACB). Two months after the operation, analysis of the results (radiograph, fusion stability test on dynamic radiographs, histological evaluation) demonstrated the following: clinical and radiographical results of the group implanted with HA blocks were superimposable on those with autogenous bone grafts, especially in the subgroup treated with DBM and BM, while in the group implanted with granules the results were poor, thus indicating the fundamental influence of the physical state of the biomaterial on bone ingrowth. This was then further confirmed by the histological study in the HA block group, whereas the HA granule group showed a certain fibrous reaction around the granules with poor porous bone ingrowth.

Animals↗
PubMed1987

Bone matrix and marrow versus cancellous bone in rabbit radial defects.

Implants of demineralized bone matrix induce new bone formation. In order to estimate the possible clinical usefulness of this phenomenon, autologous cancellous bone grafts were compared with composite grafts of bone matrix and marrow. Cancellous bone from the tuber ischii of the rabbit was transplanted to a preformed radial defect in the same animal. On the opposite side, a similar defect was filled with a mixture of either allogenous or autogenous bone-matrix particles and autogenous bone marrow. After 25 days, calcium 45 was injected intravenously. Three days later the animals were killed. Standardized segments of the rabbit's forearms, containing the middle of the defect, were cut out, ashed, and analyzed for 45Ca activity. No side difference in 45Ca deposition was found. The callus ash weight of the allogenous matrix-transplanted side was approximately 60% of that of the cancellous bone side. This side difference of ash weights corresponds to the estimated initial mineral content of the cancellous graft. Nontransplanted defects had very low ash weight and 45Ca activity. Thus, in the rabbit, composite grafts of bone matrix and marrow produce a bone yield comparable to that of cancellous bone.

Animals↗
PubMed1988

Irradiation-sterilization of rat bone matrix gelatin.

Bone matrix gelatin induces bone formation in muscle, and when implanted orthotopically it improves bone repair. Co-60 sterilization of bone gelatin impairs the protein-bound induction mechanisms. Gelatin samples nonirradiated or irradiated by 25 or 50 kGy were implanted into a pouch in the abdominal wall of Sprague-Dawley rats, as well as into a 7-mm calvarial defect. Evaluation was done by histologic studies, histomorphometry of orthotopic implants, and determination of alkaline phosphatase in ectopic implants. Gelatin irradiated with 50 kGy was absorbed in the muscle bed without evidence of any specific host reaction. Irradiation of 25 kGy led to histologically confirmed ectopic bone formation, but the wet weight of the explants was only half that of the nonirradiated control samples. Alkaline phosphatase activity was equal in both of these groups. With orthotopic implantation, neither a histologic nor a morphometric effect was seen with 25 kGy. Loss of osteoinduction with 25-kGy irradiation is apparently masked by osteoconductive mechanisms with orthotopic implantation.

Alkaline Phosphatase↗