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PubMed1990

Bone transplantation.

According to one principle of surgery, the transplantation of vital tissue is the best method of reconstructing a defect. Because of absent immunologic reactions, high osteogenic potency, and preserved stability, transplantation of autogenous bone shows the best results. Necrosis of transplanted bone, leading inevitably to absorption and remodeling of the graft, can be avoided if microsurgically vascularized autogenous bone segments are transferred. Disadvantages are the low availability and the necessity of additional operations. As an alternative, deep-frozen allogeneic bone is used. However, this kind of bone shows delayed incorporation based on cellular and humoral immune reactions, and it is also installed into the host bed after overcoming the immune barrier. The risk of microbiological contamination or transmission of unrecognized germs such as HIV is a cause of great expense in bone banking techniques. If one succeeds in reducing (a) the immunologic defense reaction and (b) the risk of infection by sterilization or disinfection without damaging the osteoinductive proteins of bone matrix, the rate of complications can be lowered. Demineralized bone matrix can be used if biomechanical stability is not required. Its ability to induce osteogenesis without a major immune reaction or the risk of transmitting diseases justifies its clinical application. Further intensive research in these areas is unavoidable.

Bone Transplantation↗
PubMed1993

Apatite formation on three kinds of bioactive material at an early stage in vivo: a comparative study by transmission electron microscopy.

Apatite formation on the surface of three kinds of bioactive material at an early stage after implantation in bone was studied using transmission electron microscopy (TEM). The materials were apatite- and wollastonite-containing glass-ceramic (A-W GC) as a surface-active glass-ceramic, dense sintered hydroxyapatite (HA) as a surface-active ceramic, and dense sintered beta-tricalcium phosphate (beta-TCP) as a resorbable ceramic. Particles of these materials, ranging from 100-300 microns in diameter, were implanted into rat tibiae, and specimens were prepared at 3, 7, 10, and 14 days after implantation. For A-W GC, dissolution of the glassy and probably wollastonite phase was observed in the surface region on and after the third day, and a collagen-free thin apatite layer on the surface of the material was evident on and after the seventh day. This apatite layer was observed before the mineralization of the surrounding bone matrix and was sometimes evident even where the material bordered on the bone marrow. On and after the tenth day, the surrounding bone matrix calcified and A-W GC-bone bonding through an apatite layer was completed. For HA, a mineralized collagen-free layer was observed on the surface of the ceramic on and after the tenth day. This layer was always present near calcifying bone and it was difficult to distinguish from immature bone. For beta-TCP, such a surface mineralized layer was rarely evident, even just before bone-ceramic contact, and finally the bone bonded to beta-TCP directly. Cell-mediated degradation of beta-TCP was frequently observed.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed2006

Calvarial osteoclasts express a higher level of tartrate-resistant acid phosphatase than long bone osteoclasts and activation does not depend on cathepsin K or L activity.

Bone resorption by osteoclasts depends on the activity of various proteolytic enzymes, in particular those belonging to the group of cysteine proteinases. Next to these enzymes, tartrate-resistant acid phosphatase (TRAP) is considered to participate in this process. TRAP is synthesized as an inactive proenzyme, and in vitro studies have shown its activation by cysteine proteinases. In the present study, the possible involvement of the latter enzyme class in the in vivo modulation of TRAP was investigated using mice deficient for cathepsin K and/or L and in bones that express a high (long bone) or low (calvaria) level of cysteine proteinase activity. The results demonstrated, in mice lacking cathepsin K but not in those deficient for cathepsin L, significantly higher levels of TRAP activity in long bone. This higher activity was due to a higher number of osteoclasts. Next, we found considerable differences in TRAP activity between calvarial and long bones. Calvarial bones contained a 25-fold higher level of activity than long bones. This difference was seen in all mice, irrespective of genotype. Osteoclasts isolated from the two types of bone revealed that calvarial osteoclasts expressed higher enzyme activity as well as a higher level of mRNA for the enzyme. Analysis of TRAP-deficient mice revealed higher levels of nondigested bone matrix components in and around calvarial osteoclasts than in long bone osteoclasts. Finally, inhibition of cysteine proteinase activity by specific inhibitors resulted in increased TRAP activity. Our data suggest that neither cathepsin K nor L is essential in activating TRAP. The findings also point to functional differences between osteoclasts from different bone sites in terms of participation of TRAP in degradation of bone matrix. We propose that the higher level of TRAP activity in calvarial osteoclasts compared to that in long bone cells may partially compensate for the lower cysteine proteinase activity found in calvarial osteoclasts and TRAP may contribute to the degradation of noncollagenous proteins during the digestion of this type of bone.

Acid Phosphatase↗
PubMed1996

Human osteosarcoma (OST) induces mouse reactive bone formation in xenograft system.

To distinguish the origin of bone-forming cells in the osteosarcoma (OST) tumor inoculated into nude mice, we have developed a novel in situ hybridization technique. The system used digoxygenin (DIG) labeled DNA probes that encoded human specific repetitive gene, Alu, and mouse specific repetitive gene, mouse L1 (m-L1). The chondrogenic and osteogenic cells in the tumor had strongly positive signals for m-L1 probe without any signals for Alu probe. The expression of bone matrix proteins was also examined by in situ hybridization. The bone-forming cells were positive for mRNAs of mouse osteonectin, osteopontin, and osteocalcin relating to calcification during bone formation, while these were negative for human mRNAs of these bone matrix proteins. The OST cells in the tumor expressed the human bone morphogenetic proteins (BMPs) mRNAs by RT-PCR. These data indicated that the mouse cells, not the human sarcoma cells, are responsible for cartilage and bone formation in the OST tumor inoculated into nude mice, and we speculated that BMPs, at least in part, could play an important role in this ossification.

Animals↗
PubMed2004

Are recombinant human bone morphogenetic protein-7 and tobramycin compatible? An experiment in rats.

OBJECTIVES: To evaluate the effects of local antibiotics on bone morphogenetic protein-induced new bone formation in vivo. DESIGN: In the research laboratory, inactive collagenous bone matrix was reconstituted with 1 microg of recombinant human bone morphogenetic protein-7 and implanted subcutaneously in the thorax bilaterally in 30 male Long-Evans rats. INTERVENTION: In group A (n = 2), the inactive collagenous bone matrix alone was implanted, bilaterally, and one of these pellets treated with either 500 microg tobramycin in aqueous solution or 3 tobramycin-impregnated polymethyl methacrylate beads. In group B (n = 4), the reconstituted pellets were not treated with tobramycin. In group C (n = 8), 1 reconstituted pellet in each rat was treated with 500 microg tobramycin in aqueous solution. In group D (n = 8), 3 tobramycin beads were placed in contact with 1 of the 2 reconstituted pellets in each rat. In group E (n = 8), 3 tobramycin beads were placed on the dorsal surface of 4 of the rats. All rats were killed on day 11. MAIN OUTCOME MEASUREMENT: Bone formation was evaluated by alkaline phosphatase assay and histology. Tobramycin elution from the beads after day 11 was measured by placing the explanted beads into a phosphate buffer solution to incubate for 24 hours. RESULTS: There was no difference in the alkaline phosphatase activity between the tobramycin treated and untreated implants. Histologic evaluation of the implants revealed areas of robust new bone formation in both the tobramycin treated and untreated implants. CONCLUSIONS: The results by both alkaline phosphatase assay and histologic evaluation in this rat model indicate that there is no inhibition of recombinant human bone morphogenetic protein-7-induced new bone formation by locally applied tobramycin. Recombinant human bone morphogenetic protein-7 is osteoinductive in the presence of locally applied tobramycin. A composite osteogenic device containing both tobramycin and recombinant human bone morphogenetic protein-7 may be developed that can simultaneously induce bone healing and decrease the risk for infection.

Administration, Topical↗
PubMed2006

Tensile behavior of cortical bone: dependence of organic matrix material properties on bone mineral content.

A porous composite model is developed to analyze the tensile mechanical properties of cortical bone. The effects of microporosity (volksman's canals, osteocyte lacunae) on the mechanical properties of bone tissue are taken into account. A simple shear lag theory, wherein tensile loads are transferred between overlapped mineral platelets by shearing of the organic matrix, is used to model the reinforcement provided by mineral platelets. It is assumed that the organic matrix is elastic in tension and elastic-perfectly plastic in shear until it fails. When organic matrix shear stresses at the ends of mineral platelets reach their yield values, the stress-strain curve of bone tissue starts to deviate from linear behavior. This is referred as the microscopic yield point. At the point where the stress-strain behavior of bone shows a sharp curvature, the organic phase reaches its shear yield stress value over the entire platelet. This is referred as the macroscopic yield point. It is assumed that after macroscopic yield, mineral platelets cannot contribute to the load bearing capacity of bone and that the mechanical behavior of cortical bone tissue is determined by the organic phase only. Bone fails when the principal stress of the organic matrix is reached. By assuming that mechanical properties of the organic matrix are dependent on bone mineral content below the macroscopic yield point, the model is used to predict the entire tensile mechanical behavior of cortical bone for different mineral contents. It is found that decreased shear yield stresses and organic matrix elastic moduli are required to explain the mechanical behavior of bones with lowered mineral contents. Under these conditions, the predicted values (elastic modulus, 0.002 yield stress and strain, and ultimate stress and strain) are within 15% of experimental data.

Biomechanical Phenomena↗
PubMed1997

Stage-specific expression patterns of alkaline phosphatase during development of the first arch skeleton in inbred C57BL/6 mouse embryos.

Timing and pattern of expression of alkaline phosphatase was examined during early differentiation of the 1st arch skeleton in inbred C57BL/6 mice. Embryos were recovered between 10 and 18 d of gestation and staged using a detailed staging table of craniofacial development prior to histochemical examination. Expression of alkaline phosphatase is initiated at stage 20.2 in the plasma membrane of mesenchymal cells in the distal region of the first arch. Expression is strongest in osteoid (unmineralised bone matrix) and presumptive periosteum at stage 21.32. Mineralisation begins at stage E23. Expression is present in the mineralised bone matrix. Secondary cartilages form in the condylar and angular processes by stage M24. The cartilaginous cells and surrounding cells in the processes are all alkaline phosphatase-positive and surrounded by the common periosteum, suggesting that progenitor cells of the processes, dentary ramus and secondary cartilages all originate from a common pool. Nonhypertrophied chondrocytes of Meckel's cartilage express alkaline phosphatase at stage M23. Expression in these chondrocytes is preceded by the expression in their adjacent perichondrium. This is true of chondrocytes in all other cranial cartilages examined. 3-D reconstruction of expression in Meckel's cartilage also revealed that the chondrocytes of Meckel's cartilage which express alkaline phosphatase and the matrix of which undergoes mineralisation are those surrounded by the alkaline phosphatase-positive dentary ramus. By stage 25, coincident with mineralisation in the distal section of Meckel's cartilage, most chondrocytes are strongly positive. The perichondria of malleus and incus cartilages express alkaline phosphatase at stage M24. Nonhypertrophied chondrocytes along these perichondria also express alkaline phosphatase. Superficial and deep cells in the dental laminae of incisor and 1st molar teeth become alkaline phosphatase-positive at the bud stage, stages 21.16 and 21.32, respectively. Dental papillae are negative until stage M24 when alkaline phosphatase expression begins in the dental papillae and follicles of the incisor teeth and the dental follicles of the 1st molar teeth. The dental papillae of the 1st molar teeth express alkaline phosphatase at stage 25. Expression in the dental papillae and follicles appears to coincide with cellular differentiation of follicle from papilla. The presumptive squamosal, ectotympanic and gonial membrane bones, lingual oral epithelial cells connected to the dental laminae of the incisor teeth, hair follicle papillae and sheath and surrounding dermis all express alkaline phosphatase in a stage-specific manner.

Alkaline Phosphatase↗
PubMed1989

Ectopic induction of cartilage and bone by water-soluble proteins from bovine bone using a collagenous delivery vehicle.

A controlled-release delivery vehicle for water-soluble osteogenic proteins from demineralized bone matrix was constructed using purified type I collagen. The water-soluble proteins were isolated from a 4 M GdnHCl extract of bone matrix. Although the water-soluble proteins were capable of inducing cartilage formation in vitro, they were incapable of inducing cartilage or bone in vivo when implanted intramuscularly into mice in the absence of an appropriate delivery vehicle. The collagen-based delivery vehicle alone was also incapable of inducing osteogenesis in vivo. However, when the water-soluble proteins were incorporated into the delivery vehicle, the combination was capable of inducing cartilage and bone 76% of the time. These results demonstrate that it is possible to formulate a controlled-release delivery vehicles for soluble bioactive factors which upon release interact with local responsive cells.

Animals↗
PubMed1992

The effect of tissue type plasminogen activator (tPA) on osteoclastic resorption in embryonic mouse long bone explants: a possible role for the growth factor domain of tPA.

Osteoblasts produce proteolytic enzymes and their production is regulated by osteotropic agents. It has been suggested that these proteases play a role in bone resorption by removing the superficial collagenous layer from the bone matrix and indirectly inducing migration of osteoclast precursors towards the bone matrix. We examined the effect of the plasminogen activator tPA on osteoclastic resorption using 17-day-old mouse embryonic long bone explants representing different stages of osteoclast development, that is, radii containing already mature osteoclasts and metacarpals containing no mature osteoclasts but only osteoclast precursors/progenitors which are still confined to the periosteum. Tissue type PA stimulated osteoclastic resorption (measured as 45Ca-release) in 17-day-old fetal metacarpals but not in radii of the same animal. Blocking the enzymatic activity of tPA did not inhibit its effect on osteoclastic resorption. Plasmin, the direct product of PA enzymatic activity, did not induce osteoclastic resorption. However, a tPA-mutant missing the growth-factor-like domain of the molecule, failed to stimulate 45Ca-release from the metacarpals. In addition, in both systems tPA and transforming growth factor alpha had similar effects on osteoclastic resorption. The finding that tPA stimulated 45Ca-release only in the metacarpals suggests that tPA has an effect on osteoclast formation rather than on the activity of already mature osteoclasts. Under the experimental conditions used this effect seems to be mediated by the growth factor domain of tPA rather than by the enzymatic activity of the molecule.

Amino Acid Chloromethyl Ketones↗
PubMed1991

In vivo induction of bone by recombinant human transforming growth factor beta 1.

A single application of recombinant human transforming growth factor beta 1 (rhTGF-beta 1) adjacent to cartilage was found to induce bone formation in rabbit ear full-thickness skin wounds. At doses that optimally promote soft tissue healing, 25-100 ng rhTGF-beta 1 per wound caused osseous tissue formation starting 21 days after wounding to reach a peak incidence and area of bone formation at day 42. Bone formation was followed by active remodeling, resulting in lower incidence and area of bone formation at days 56 and 70. The early phase of bone formation was located overlying the cartilage and involved perichondrial cells that appeared to differentiate directly into osteoblasts forming bone matrix without a cartilage precursor. Cartilage was replaced with bone at later time points. rhTGF-beta 1 was able to increase the ratio of osteoblasts to osteoclasts lining the trabecular surface of bone and thus increase the net amount of bone formation. The present studies suggest a potential therapeutic role for rhTGF-beta 1 in hard tissue repair.

Animals↗
PubMed2005

DBM induced ectopic bone formation in the rat: the importance of surface area.

Demineralized bone matrix (DBM) has been shown to induce ectopic endochondral bone formation, when intramuscularly implanted in rats. In earlier studies we have found a variation in bone formation capacity of this DBM. This might be due to the properties of the DBM itself, but the use of DBM blocks could be of influence as well. Therefore, this study was designed to investigate whether increasing the surface area of the DBM by morsellizing, influences the bone formation capacity. In view of this, DBM implants and morsellized DBM (MDBM) implants were placed intramuscularly in a rat model. At six weeks the implants were retrieved and evaluated by histology and histomorphometry. The results demonstrated that significant amounts of newly formed bone were present in some DBM as well as some MDBM implants while in others no, or very little new bone was found. Histomorphometric analysis showed an average bone formation of 2.6% in DBM implants and an average of 1.9% in MDBM implants. Still, the amount of bone formation was limited compared with previous studies. It is concluded that enlargement of the surface area by morsellizing DBM implants is not an important factor in bone forming capacity.

Animals↗
PubMed1998

Enhancement of fracture healing with autogenous and allogeneic bone grafts.

The factors contributing to a delayed union or nonunion are many. In general they may be divided into three major categories: deficiencies in vascularity and angiogenesis, deficiencies in the robustness of the chondroosseous response, and deficiencies in stability, strain, or physical continuity. Frequently, deficiencies in more than one category are present, thus complicating the approach to therapy. For a bone grafts to enhance fracture healing, it must provide or stimulate that which is deficient. Autogenous fresh cancellous and cortical bone most frequently are used, but other common grafts include allogeneic frozen, freeze dried, or processed allogeneic cortical, corticocancellous and cancellous grafts, and demineralized bone matrix. These grafts have varying capacities to provide active bone formation, to induce bone formation by cells of the surrounding soft tissue, and to serve as a substrate for bone formation. However, the graft cannot exert its biologic activity in isolation, dependent as it is on the surrounding environment for cells to respond to its signals and, in some cases, for blood supply. The mechanical environment of the graft site is also important. Successful graft incorporation requires that an appropriate match must be made between the biologic activity of a bone graft, the condition of the perigraft environment, and the mechanical environment. The task of the clinician who performs a bone grafting procedure for the enhancement of fracture healing is to choose the right graft or combination of grafts for the biologic and mechanical environment into which the graft will be placed.

Bone Matrix↗
PubMed1981

Abnormalities in the biosynthesis of cartilage and bone proteoglycans in experimental diabetes.

Proteoglycans synthesized in developing cartilage and bone were investigated in control and streptozotocin-induced (65 mg/kg, i.v.) diabetic rats. Ten days after streptozotocin injection, animals were implanted subcutaneously with demineralized bone matrix particles. This system induces formation of cartilage and bone on days 7 and 14, respectively. Two hours before they were killed, animals were injected with 35SO4 and the labeled proteoglycans were extracted from the explants and metaphyses by either a direct associative extraction (0.5 M GuCl2) or a direct dissociative extraction (4.0 M GuCl2). These procedures extract 80-90% of the total counts incorporated. To characterize the proteoglycans, extracts were subjected to cesium chloride density gradient centrifugation and molecular sieve chromatography. These data showed that (1) there is less proteoglycan made in diabetic bone; (2) the proteoglycan aggregate is of a smaller molecular weight in bone than in cartilage; (3) 10% of the proteoglycan synthesized in diabetic bone was in the form of aggregates compared with 48% of the control bone; (4) aggregates did form in the diabetic cartilage, and their molecular weight was smaller than in normal cartilage. This investigation shows that proteoglycans, structurally important macromolecules of cartilage and bone, are altered in experimental diabetes. This metabolic abnormality may be an important factor contributing to decreased bone formation observed in diabetes.

Animals↗
PubMed1977

A histological demonstration of nerves in subchondral bone.

Several different staining procedures were carried out on decalcified histological sections from human femoral heads to demonstrate the nerves in subchondral bone. The femoral heads were obtained at surgery from patients with fractures of the femoral neck or osteoarthritic hip joints. The Bodian technique was found to be the most suitable. Serial sections were used in order to disclose the various sources of error. It was not possible to demonstrate nerves in the bone matrix, but they were easily seen in the subchondral bone marrow, after related to the vessels. A comparison of the fracture and osteoarthritic cases revealed an obvious difference; more nerves were seen in osteoarthritis. The method described is considered suitable for further study of the nerves in osteoarthritic femoral heads.

Bone Matrix↗
PubMed1999

Effect of cytokines on in vitro bone resorption by cells isolated from giant cell tumor of bone.

OBJECTIVE: To investigate the effect of cytokines on in vitro bone resorption by cells isolated from giant cell tumor of bone. METHODS: Mononuclear stromal cells and multinucleated giant cells (MGC) were isolated from 11 cases of giant cell tumor of bone (GCT) and their bone resorption capability in an in vitro cell-bone resorption model were tested. Expressions of some cytokines were detected by immunohistochemistry, Western blotting analysis and Enzyme-linked immunoabsorbent assay (ELISA) in the GCT. RESULTS: The results showed that MGCs of GCT had capability to resorb bone matrix directly. Fibroblast-like stromal cells (FC) could not only resorb bone matrix directly, but also secret unknown factors to facilitate bone resorption of MGC. Exogenous TNF-alpha could significantly increase the bone resorption by both kinds of stromal cells, while exogenous IL-1 did not. Expression rate of M-CSF and level of TNF-alpha in GCT were higher than in osteosarcoma and normal serum. CONCLUSIONS: The characteristic bone resorption behavior of GCT might be caused by its three major cell components. The M-CSF and TNF-alpha could promote their bone resorption capability.

Animals↗
PubMed2005

[Septic complication in hip and knee arthroplasty].

The authors present the management of acute infections affecting joint alloarthroplasty. Study consist 23 patients (10 male 13 female. Age between 48-84 years). Consisting of surgical elimination of inflammated soft tissue, and granulation surrounding the joint endoprothesis, local antibiotic application and suction drenage instalation. In cases of chronic infection associated with loosening of the implant and bone-matrix destruction, dealloplasty is indicated with precise removal of bone - cement followed by, local antibiotic administration, and changes affecting the bone-matrix should be approached as posttraumatic osteitis with all consequences, related to it.

Aged↗
PubMed1992

An active neutral metalloproteinase bound to the insoluble collagen in the mineralized phase matrix of adult rat calvaria.

Two kinds of proteinases were found in the mineralized phase matrix of 24-week-old rat calvaria by means of enzymography using gelatin as a substrate. One proteinase was a neutral thiol 58kD proteinase as shown in a previous paper [2]. The other was a neutral metalloproteinase that had a molecular mass of 56kD and was detected only when calcium (Ca) ions were added to the incubation buffer. It is believed that the 56kD proteinase is bound to the insoluble collagen of the bone matrix, as it is solubilized by 4 M guanidine HC1 solution from the insoluble collagen fraction, when prepared by removing extractable proteins of the mineralized phase matrix. The insoluble collagen fraction could also be solubilized and prepared as gelatin by heating at 65 degrees C for 5 minutes. The gelatin was then incubated at 37 degrees C without further treatment and became degraded without an activation of 4-aminophenylmercuric acetate (APMA). This nonactivated degradation was enhanced by adding Ca ions. These results suggest that the 56kD metalloproteinase bound to the insoluble collagen of bone matrix is in an active form and may participate in the rapid degradation of collagen during bone resorption. As partially purified 56kD metalloproteinase degraded cartilage type proteoglycan, but not type I, IV, and V collagens, it is possibly related to the degradation of proteoglycans before it binds to collagen fibers during bone formation.

Animals↗
PubMed2005

The use of guided tissue regeneration principles in endodontic surgery for induced chronic periodontic-endodontic lesions: a clinical, radiographic, and histologic evaluation.

BACKGROUND: Chronic periodontic-endodontic lesions are not uncommon in clinical practice and their regenerative capacity has long been questioned. However, there are no published studies investigating the application of guided tissue regeneration techniques in combination with endodontic surgery using an induced perio-endo defect model. This study evaluated the clinical, radiographic, and histologic outcomes of three surgical procedures used to treat induced perio-endo lesions. METHODS: Pulpal necrosis was induced in foxhounds along with surgical removal of radicular buccal bone. After 4 weeks, chronic lesions were clinically and radiographically assessed. Treatment surgery consisted of apicoectomy, root canal instrumentation, and retrofilling with mineral trioxide aggregate. Teeth were then assigned to one of the following treatment groups: open flap debridement only (OFD), OFD with bioabsorbable porcine-derived collagen membrane (BG), or OFD with BG and anorganic bovine bone matrix (BO/BG). Clinical parameters and standardized radiographs were assessed at defect creation; treatment surgery; and at 1, 2, 4, and 6 months. Animals were sacrificed at 6 months and specimens prepared for histometric analysis. RESULTS: Clinical and radiographic conditions improved during the study period. Mean epithelial attachment was similar between all groups. Mean connective tissue attachment for groups OFD, BG, and BO/BG was 3.79 mm, 2.63 mm, and 1.75 mm, respectively, and mean radicular bone height was 2.16 mm, 3.24 mm, and 3.45 mm, respectively. Statistically significant increases in the amount of new cementum were observed in groups BG and BO/BG when compared with OFD (P <0.05). CONCLUSIONS: Treatment of combined induced perio-endo lesions using bioabsorbable collagen membranes alone or in combination with anorganic bovine bone matrix resulted in increased amounts of bone, periodontal ligament, and significant increases in the amount of new cementum when compared to open flap debridement in a canine model.

Absorbable Implants↗