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PURPOSE: The influence of surface modifications on osseointegration in newly formed bone is not well established. The purpose of this study was to compare osseointegration at acid-etched versus turned implants in newly formed and native bone. METHODS: Supra-alveolar peri-implant defects were created in 8 hound/Labrador mongrel dogs. Titanium implants 10 mm long (2 turned and 1 dual acid-etched) were placed 5 mm into the surgically reduced alveolar crest, creating 5-mm supra-alveolar peri-implant defects. Recombinant human bone morphogenetic protein-2 (rhBMP-2; 0.4 mg) in a collagen carrier was used to induce new bone formation. A macroporous, expanded polytetrafluoroethylene device was used to delineate new bone formation. The animals were euthanized at 8 weeks for histometric analysis of the experimental sites. RESULTS: There were no significant differences in rhBMP-2-induced bone density (mean +/- SD) at acid-etched versus turned implants (20.6% +/- 5.3% vs 23.8% +/- 4.7%; P = .232). However, there was a significant difference in bone-implant contact in favor of the acid-etched implants (12.3% +/- 6.8% vs 7.9% +/- 3.1%; P = .05). Native bone density averaged 63.9% +/- 7.5% and 64.5% +/- 9.0% for acid-etched and turned implants, respectively (P = .641). Nevertheless, bone-implant contact was significantly enhanced at acid-etched versus turned implants (59.7% +/- 11.3% vs 40.7% +/-21.2%; P =.005). CONCLUSIONS: Surface dual acid-etching of titanium implants has a positive effect on osseointegration in newly formed and native bone. Significant differences in bone density do not appear to influence this effect. [ABSTRACT FROM AUTHOR]

Background: Studies using ectopic rodent, orthotopic canine, and non-human primate models show that bone morphogenetic proteins (BMPs) coated onto titanium surfaces induce local bone formation. The objective of this study was to examine the ability of recombinant human BMP-2 (rhBMP-2) coated onto a titanium porous oxide implant surface to stimulate local bone formation including osseointegration and vertical augmentation of the alveolar ridge.Material and Methods: Bilateral, critical-size, 5 mm, supra-alveolar, peri-implant defects were created in 12 young adult Hound Labrador mongrel dogs. Six animals received implants coated with rhBMP-2 at 0.75 or 1.5 mg/ml, and six animals received implants coated with rhBMP-2 at 3.0 mg/ml or uncoated control. Treatments were randomized between jaw quadrants. The mucoperiosteal flaps were advanced, adapted and sutured to submerge the implants for primary intention healing. The animals received fluorescent bone markers at weeks 3, 4, 7 and 8 post-surgery when they were euthanized for histologic evaluation.Results: Jaw quadrants receiving implants coated with rhBMP-2 exhibited gradually regressing swelling that became hard to palpate disguising the contours of the implants. The histologic evaluation showed robust bone formation reaching or exceeding the implant platform. The newly formed bone exhibited characteristics of the adjoining resident Type II bone including cortex formation for sites receiving implants coated with rhBMP-2 at 0.75 or 1.5 mg/ml. Sites receiving implants coated with rhBMP-2 at 3.0 mg/ml exhibited more immature trabecular bone formation, seroma formation and peri-implant bone remodelling resulting in undesirable implant displacement. Control implants exhibited minimal, if any, bone formation. Thus, implants coated with rhBMP-2 at 0.75, 1.5 and 3.0 mg/ml exhibited significant bone formation (height and area) compared with the sham-surgery control averaging (+/-SD) 4.4+/-0.4, 4.2+/-0.7 and 4.2+/-1.2 versus 0.8+/-0.3 mm; and 5.0+/-2.2, 5.6+/-2.2 and 7.4+/-3.5 versus 0.7+/-0.3 mm[2], respectively (p<0.01). All the treatment groups exhibited clinically relevant osseointegration.Conclusions: rhBMP-2 coated onto titanium porous oxide implant surfaces induced clinically relevant local bone formation including vertical augmentation of the alveolar ridge and osseointegration. Higher concentrations/doses were associated with untoward effects. [ABSTRACT FROM AUTHOR]

Background: Studies using ectopic rodent and orthotopic canine models (Type II bone) have shown that titanium porous oxide (TPO) surface implants adsorbed with recombinant human bone morphogenetic protein-2 (rhBMP-2) induce local bone formation including osseointegration. The objective of this study was to evaluate local bone formation and osseointegration at such implants placed into Type IV bone.Material and Methods: rhBMP-2-coated implants were installed into the edentulated posterior maxilla in eight young adult Cynomolgus monkeys: four animals each received three TPO implants adsorbed with rhBMP-2 (2.0 mg/ml) and four animals each received three TPO implants adsorbed with rhBMP-2 (0.2 mg/ml). Contra-lateral jaw quadrants received three TPO implants without rhBMP-2 (control). Treatments were alternated between left and right jaw quadrants. Mucosal flaps were advanced and sutured to submerge the implants. The animals received fluorescent bone markers at weeks 2, 3, 4, and at week 16 when they were euthanized for histologic analysis.Results: Clinical healing was uneventful. Extensive local bone formation was observed in animals receiving implants adsorbed with rhBMP-2 (2.0 mg/ml). The newly formed bone exhibited a specific pinpoint bone-implant contact pattern regardless of rhBMP-2 concentration resulting in significant osseointegration; rhBMP-2 (2.0 mg/ml): 43% and rhBMP-2 (0.2 mg/ml): 37%. Control implants exhibited a thin layer of bone covering a relatively larger portion of the implant threads. Thus, TPO control implants bone exhibited significantly greater bone-implant contact (~75%; p<0.05). There were no statistically significant differences between rhBMP-2-coated and control implants relative to any other parameter including peri-implant and intra-thread bone density.Conclusion: rhBMP-2-coated TPO implants enhanced/accelerated local bone formation in Type IV bone in a dose-dependent fashion in non-human primates resulting in significant osseointegration. rhBMP-2-induced de novo bone formation did not reach the level of osseointegration observed in native resident bone within the 16-week interval. [ABSTRACT FROM AUTHOR]

Background: Conventional oral/maxillofacial implants reach osseointegration over several months during which the titanium fixtures interact with alveolar bone. The objective of this study was to determine if adsorbing recombinant human bone morphogenetic protein-2 (rhBMP-2) onto a titanium porous oxide (TPO) implant surface might enhance or accelerate local bone formation and support osseointegration in a large animal oral/maxillofacial orthotopic model.Material and Methods: Endosseous implants with a TPO surface were installed into the edentulated posterior mandible in eight adult Hound Labrador mongrel dogs. The implant surface had been adsorbed with rhBMP-2 at 0.2 or 4.0 mg/ml. TPO implants without rhBMP-2 served as control. Treatments were randomized between jaw quadrants. Mucosal flaps were advanced and sutured leaving the implants submerged. Clinical and radiographic evaluations were made immediately post-surgery, at day 10 (suture removal), and week 4 and 8 post-surgery. The animals received fluorescent bone markers at week 3, 4, and at week 8 post-surgery, when they were euthanized for histologic analysis.Results: TPO implants coated with rhBMP-2 exhibited dose-dependent bone remodelling including immediate resorption and formation of implant adjacent bone, and early establishment of clinically relevant osseointegration. The resulting bone-implant contact, although clinically respectable, appeared significantly lower for rhBMP-2-coated implants compared with the control [rhBMP-2 (0.2 mg/ml) 43.3+/-10.8%versus 71.7+/-7.8%, p<0.02; rhBMP-2 (4.0 mg/ml) 35.4+/-10.6%versus 68.2+/-11.0%, p<0.03].Conclusions: rhBMP-2 adsorbed onto TPO implant surfaces initiates dose-dependent peri-implant bone re-modelling resulting in the formation of normal, physiologic bone and clinically relevant osseointegration within 8 weeks. [ABSTRACT FROM AUTHOR]

Background: The success rate of rotator cuff repairs is variable. This study was performed to evaluate the ability of recombinant human bone morphogenetic protein-12 (rhBMP-12), administered in several carriers, to accelerate healing in a sheep model of rotator cuff repair.Methods: Local retention of tracer amounts of radiolabeled rhBMP-12, added to non-radiolabeled rhBMP-12 delivered in buffer, hyaluronan paste or sponges, or Type-I or Type-I/III collagen sponges was first evaluated with use of gamma scintigraphy in a pilot study of a rat intramuscular implant model. The rhBMP-12/paste and sponge combinations were then evaluated in eight sheep each with unilateral complete detachment and subsequent double-row reattachment of the infraspinatus tendon to the proximal part of the humerus. Contralateral, normal shoulders from sixteen sheep and shoulders in which a repair had been done without administration of rhBMP-12 in fourteen sheep were also evaluated. The rhBMP-12/Type-I and Type-I/III collagen sponge combinations were each evaluated in eight additional sheep on the basis of superior efficacy. The Type-I/III collagen sponge alone was evaluated in ten sheep to examine the effect of a collagen carrier. Ultrasound imaging was performed at four and eight weeks. Radiographic evaluation, mechanical testing, and biochemical evaluation were performed at eight weeks. Histological evaluation was performed on specimens from the sites of selected repairs following mechanical testing.Results: The sponge carriers had longer local retention of rhBMP-12 than did the buffer or paste carriers in the rat models. All of the sheep shoulder-repair groups demonstrated ultrasound evidence of a gap between the tendon and the humeral insertion. The gap length and the cross-sectional area of the repair tissue decreased with time. The mechanical properties of the repairs treated with rhBMP-12 and hyaluronan paste were similar to those of the untreated repairs. The maximum loads for the rhBMP-12/hyaluronan sponge and rhBMP-12/collagen sponge-treated repairs were 2.1 and 2.7 times greater, respectively, than the loads for the untreated repairs and were 33% and 42% of the value for the normal tendon at eight weeks. The maximum loads for the repairs treated with rhBMP-12 and a Type-I or Type-I/III collagen sponge were 2.1 times greater than those for the repairs treated with the Type-I/III collagen sponge alone. Changes in maximum stiffness followed a similar pattern. Histological evaluation demonstrated accelerated healing of the rhBMP-12-treated repairs compared with the untreated repairs. Bone formation was observed in all repairs, and biochemical measurements were not equivalent to those of normal tendon at eight weeks.Conclusions: Delivery of rhBMP-12 in a collagen or hyaluronan sponge resulted in accelerated healing of acute full-thickness rotator cuff repairs in a sheep model.Clinical Relevance: Delivery of rhBMP-12 in several sponge carriers has the potential to accelerate healing of rotator cuff repairs. Accelerated repair may allow shorter rehabilitation and an earlier return to occupational and recreational activities. [ABSTRACT FROM AUTHOR]

Background: Treatment of segmental bone loss remains a challenge in skeletal repairs. This study was performed to evaluate the efficacy of the use of recombinant bone morphogenetic protein-2 (rhBMP-2) delivered in an injectable calcium phosphate cement (alpha bone substitute material [alpha-BSM]) to bridge critical-sized defects in the rabbit radius.Methods: Unilateral 20-mm mid-diaphyseal defects were created in the radii of thirty-six skeletally mature New Zealand White rabbits. The defects in twelve rabbits each were filled with 0.166 mg/mL rhBMP-2/alpha-BSM cement, 0.033 mg/mL rhBMP-2/alpha-BSM cement, or buffer/alpha-BSM cement. Six rabbits from each group were killed at four weeks, and six were killed at eight weeks. Serial radiographs were made to monitor defect-bridging and residual alpha-BSM carrier. A semiquantitative histological scoring system was used to evaluate defect-bridging. Histomorphometry was used to quantify residual alpha-BSM; trabecular bone area; trabecular bone volume fraction; and cortical length, width, and area.Results: At four weeks, there had been more rapid resorption of alpha-BSM and filling of the defects with trabecular bone in the group treated with 0.166 mg/mL rhBMP-2/alpha-BSM than in the other two groups. Histomorphometry confirmed an increased trabecular area and volume fraction in this group compared with the other two groups. In both rhBMP-2/alpha-BSM-treated groups, the majority of the trabecular bone was formed by a direct process adjacent to the resorbing alpha-BSM. At eight weeks, complete cortical bridging and regeneration of the marrow space were present in all of the defects treated with 0.166 mg/mL rhBMP-2/alpha-BSM. That group also had reduced residual alpha-BSM and trabecular area and volume, compared with the other two groups, at eight weeks as a result of a rapid remodeling process.Conclusions: Treatment of a critical-sized defect in a rabbit radius with 0.166 mg/mL rhBMP-2/alpha-BSM injectable cement can result in bridging with cortical bone and a regenerated bone-marrow space by eight weeks. Site-specific remodeling appears to be responsible for corticalization and marrow regeneration.Clinical Relevance: RhBMP-2 delivered in a calcium phosphate cement may be useful to achieve bridging of critical-sized defects in patients. Its injectable properties may allow minimally invasive use. Delayed percutaneous administration would also be possible when augmentation is desired following an initial surgical procedure or when soft-tissue injuries preclude adequate initial treatment. [ABSTRACT FROM AUTHOR]

OBJECTIVES: The potential of recombinant human bone morphogenetic protein-2 (rhBMP-2) to enhance bone healing following endodontic surgery was tested. The pattern and timing of de novo bone formation and cementum regeneration, and the potential for root resorption and ankylosis to accompany bone formation were evaluated. STUDY DESIGN: Pulpal infections were induced in maxillary and mandibular incisor teeth in young adult Cynomolgus monkeys. The teeth received conventional endodontic treatment immediately followed by surgical root resection. In a randomized split-mouth design, contralateral apical bone defects received rhBMP-2 in absorbable collagen sponge (ACS) carrier or served as sham-surgery controls to provide histological and radiographic evaluations following 1 (mandibular incisors) and 4.5 (maxillary incisors) month(s) postsurgery. RESULTS: At 1 month postsurgery trabecular bone filled the apical bone defects. The newly formed bone appeared considerably more mature and had assumed characteristics of the contiguous resident bone at 4.5 months postsurgery. The resected root tips were almost completely covered by new cementum with a maturing functionally oriented periodontal ligament. Localized inflammatory infiltrates were associated with the filled root canals and extruded root-filling material. Root resorption and ankylosis were not observed. There were no apparent differences in healing patterns between sites implanted with rhBMP-2/ACS and those serving as sham-surgery controls. CONCLUSIONS: Under conditions where the influence of infectious elements and irritation caused by root filling material are minimized, bone formation and cementum regeneration appears rapid following endodontic surgery. rhBMP-2/ACS did not offer an obvious benefit above and beyond that of the native osteogenic potential in this animal model. [ABSTRACT FROM AUTHOR]

Background: While recombinant human bone morphogenetic protein-2 (rhBMP-2) administered in a calcium phosphate cement accelerates osteotomy-site healing in animal models when administered three hours after surgery, definitive fracture treatment is often delayed. The present study evaluated the ability of rhBMP-2, administered in a new particulating calcium phosphate matrix, to accelerate nonhuman primate fibular osteotomy-site healing following treatment at multiple treatment times and concentrations.Methods: The ability of 1.5-mg/mL rhBMP-2/calcium phosphate matrix to accelerate osteotomy-site healing when administered three hours, one day, one week, or two weeks after surgery was first evaluated with use of bilateral proximal and distal fibular osteotomy sites in adult male monkeys. In a second study, the healing of osteotomy sites that had been treated with the administration of calcium phosphate matrix alone and with different concentrations of rhBMP-2/calcium phosphate matrix (0.5 mg/mL, 1.5 mg/mL, or 4.5 mg/mL) seven days after surgery was compared with that of contralateral, untreated osteotomy sites. In a third study, the histologic progression of osteotomy-site healing following treatment with 1.5-mg/mL rhBMP-2/calcium phosphate matrix or calcium phosphate matrix alone, administered three hours or one week after surgery to the osteotomy site, was assessed at multiple time points for as long as twenty-four months after surgery.Results: Radiographs demonstrated increased callus area and more rapid healing in response to 1.5-mg/mL rhBMP-2/calcium phosphate matrix administered over the range of treatment times after surgery as compared with the findings of previous reports on untreated osteotomy sites. Bone formation appeared at the osteotomy sites sooner following treatment at one and two weeks as compared with the findings at the earlier time-points. Scintigraphic imaging at one day and one week after surgery showed prolonged retention of rhBMP-2 at the osteotomy site following an initial burst release. In the second study, radiographic, peripheral quantitative computed tomographic, biomechanical, and microscopic evaluation demonstrated that administration of 1.5 and 4.5-mg/mL rhBMP-2/calcium phosphate matrix one week after surgery accelerated osteotomy-site healing by 40% to 50% compared with the findings in untreated controls. The magnitude of acceleration was less in response to 0.5-mg/mL rhBMP-2/calcium phosphate matrix, and calcium phosphate matrix alone did not accelerate osteotomy-site healing. Histological evaluation indicated that an increased cellular infiltrate and increased direct bone formation contributed to the accelerated osteotomy-site healing following administration of rhBMP-2/calcium phosphate matrix at one week compared with three hours after surgery.Conclusions: A single percutaneous injection of rhBMP-2/calcium phosphate matrix accelerated healing in nonhuman primate fibular osteotomy sites over a wide range of treatment times. Efficacy was optimized in association with the administration of 1.5-mg/mL rhBMP-2/calcium phosphate matrix. Delaying treatment for one week further accelerated healing because of an increase in the number of responding cells and an increase in direct bone formation. [ABSTRACT FROM AUTHOR]

Background: Supraphysiologic bone morphogenetic protein (BMP)-2 concentrations are required to induce spinal fusion. In this study, a BMP-2/BMP-6/activin A chimera (BV-265), optimized for BMP receptor binding, delivered in a recombinant human collagen:CDHA [calcium-deficient hydroxyapatite] porous composite matrix (CM) or bovine collagen:CDHA granule porous composite matrix (PCM), engineered for optimal BV-265 retention and guided tissue repair, was compared with BMP-2 delivered in a bovine absorbable collagen sponge (ACS) wrapped around a MASTERGRAFT Matrix (MM) ceramic-collagen rod (ACS:MM) in a nonhuman primate noninstrumented posterolateral fusion (PLF) model., Methods: In vivo retention of 125I-labeled-BV-265/CM or PCM was compared with 125I-labeled-BMP-2/ACS or BMP-2/buffer in a rat muscle pouch model using scintigraphy. Noninstrumented PLF was performed by implanting CM, BV-265/CM, BV-265/PCM, or BMP-2/ACS:MM across L3-L4 and L5-L6 or L3-L4-L5 decorticated transverse processes in 26 monkeys. Computed tomography (CT) images were acquired at 0, 4, 8, 12, and 24 weeks after surgery, where applicable. Manual palpation, μCT (microcomputed tomography) or nCT (nanocomputed tomography), and histological analysis were performed following euthanasia., Results: Retention of 125I-labeled-BV-265/CM was greater than BV-265/PCM, followed by BMP-2/ACS and BMP-2/buffer. The CM, 0.43 mg/cm3 BMP-2/ACS:MM, and 0.05 mg/cm3 BV-265/CM failed to generate PLFs. The 0.15-mg/cm3 BV-265/CM or 0.075-mg/cm3 BV-265/PCM combinations were partially effective. The 0.25-mg/cm3 BV-265/CM and 0.15 and 0.3-mg/cm3 BV-265/PCM combinations generated successful 2-level PLFs at 12 and 24 weeks., Conclusions: BV-265/CM or PCM can induce fusion in a challenging nonhuman primate noninstrumented PLF model at substantially lower concentrations than BMP-2/ACS:MM., Clinical Relevance: BV-265/CM and PCM represent potential alternatives to induce PLF in humans at substantially lower concentrations than BMP-2/ACS:MM., Competing Interests: Disclosure: Funding was provided by Bioventus LLC, the manufacturer of the BV-0265 chimera. On the Disclosure of Potential Conflicts of Interest forms, which are provided with the online version of the article, one or more of the authors checked “yes” to indicate that the author had a relevant financial relationship in the biomedical arena outside the submitted work (including current and former employment with Bioventus LLC) and “yes” to indicate that the author had a patent and/or copyright, planned, pending, or issued, directly relevant to this work (http://links.lww.com/JBJS/G482)., (Copyright © 2021 by The Journal of Bone and Joint Surgery, Incorporated.)

Background: Synovial membrane-derived factors are implicated in arthritis-related bone changes. The route that synovial factors use to access subchondral bone and the mechanisms responsible for these bone changes remain unclear. A safety study involving intra-articular injection of bone morphogenetic protein-2 (BMP-2)/calcium phosphate matrix (CPM) or CPM addresses these issues., Methods: Knee joints in 21 monkeys were injected with CPM or 1.5 or 4.5 mg/mL BMP-2/CPM and were evaluated at 1 and 8 weeks. Contralateral joints were injected with saline solution. Knee joints in 4 animals each were injected with 1.5 or 4.5 mg/mL BMP-2/CPM. Contralateral joints were injected with corresponding treatments at 8 weeks. Both joints were evaluated at 16 weeks. Harvested joints were evaluated grossly and with histomorphometry. Knee joints in 3 animals were injected with 125I-labeled BMP-2/CPM and evaluated with scintigraphy and autoradiography at 2 weeks to determine BMP-2 distribution., Results: All treatments induced transient synovitis and increased capsular vascularization, observed to anastomose with metaphyseal venous sinusoids, but did not damage articular cartilage. Both treatments induced unanticipated activation of vascular-associated trabecular bone remodeling compartments (BRCs) restricted to injected knees. Bone volume increased in BMP-2/CPM-injected knees at 8 and 16 weeks. Scintigraphy demonstrated metaphyseal 125I-labeled BMP-2 localization restricted to injected knees, confirming local rather than systemic BMP-2 release. Autoradiography demonstrated that BMP-2 diffusion through articular cartilage into the metaphysis was blocked by the tidemark. The lack of marrow activation or de novo bone formation, previously reported following metaphyseal BMP-2/CPM administration, confirmed BMP-2 and synovial-derived factors were not free in the marrow. The 125I-labeled BMP-2/CPM, observed within venous sinusoids of injected knees, confirmed the potential for capsular and metaphyseal venous portal communication., Conclusions: This study identifies a synovitis-induced venous portal circulation between the joint capsule and the metaphysis as an alternative to systemic circulation and local diffusion for synovial membrane-derived factors to reach subchondral bone. This study also identifies vascular-associated BRCs as a mechanism for arthritis-associated subchondral bone changes and provides additional support for their role in physiological trabecular bone remodeling and/or modeling., Clinical Relevance: Inhibition of synovitis and accompanying abnormal vascularization may limit bone changes associated with arthritis., Competing Interests: Disclosure: Funding for this study was provided by Pfizer, Inc. The authors were employees of Pfizer, Inc. at the time the study was performed. On the Disclosure of Potential Conflicts of Interest forms, which are provided with the online version of the article, one or more of the authors checked “yes” to indicate that the author had a relevant financial relationship in the biomedical arena outside the submitted work (including with Pfizer, Inc.) and “yes” to indicate that the author had a patent and/or copyright, planned, pending, or issued, directly relevant to this work (http://links.lww.com/JBJS/G226)., (Copyright © 2020 by The Journal of Bone and Joint Surgery, Incorporated.)

Bone morphogenetic protein (BMP)/carriers approved for orthopedic procedures achieve efficacy superior or equivalent to autograft bone. However, required supraphysiological BMP concentrations have been associated with potential local and systemic adverse events. Suboptimal BMP/receptor binding and rapid BMP release from approved carriers may contribute to these outcomes. To address these issues and improve efficacy, we engineered chimeras with increased receptor binding by substituting BMP-6 and activin A receptor binding domains into BMP-2 and optimized a carrier for chimera retention and tissue ingrowth. BV-265, a BMP-2/BMP-6/activin A chimera, demonstrated increased binding affinity to BMP receptors, including activin-like kinase-2 (ALK2) critical for bone formation in people. BV-265 increased BMP intracellular signaling, osteogenic activity, and expression of bone-related genes in murine and human cells to a greater extent than BMP-2 and was not inhibited by BMP antagonist noggin or gremlin. BV-265 induced larger ectopic bone nodules in rats compared to BMP-2 and was superior to BMP-2, BMP-2/6, and other chimeras in nonhuman primate bone repair models. A composite matrix (CM) containing calcium-deficient hydroxyapatite granules suspended in a macroporous, fenestrated, polymer mesh-reinforced recombinant human type I collagen matrix demonstrated improved BV-265 retention, minimal inflammation, and enhanced handling. BV-265/CM was efficacious in nonhuman primate bone repair models at concentrations ranging from 1 / 10 to 1 / 30 of the BMP-2/absorbable collagen sponge (ACS) concentration approved for clinical use. Initial toxicology studies were negative. These results support evaluations of BV-265/CM as an alternative to BMP-2/ACS in clinical trials for orthopedic conditions requiring augmented healing., (Copyright © 2019 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works.)

Background: Osteoporosis results in a decrease in bone density, bone quality, and strength throughout the skeleton. Despite systemic therapies, the morbidity and mortality that are associated with hip fractures remain a major consequence of osteoporosis., Methods: We used fourteen chronic ovariectomized female cynomolgus monkeys in this study. Six animals received an intraosseous injection of 0.5 mL of 1.5 mg/mL recombinant human bone morphogenetic protein-2/calcium phosphate matrix (rhBMP-2/CPM) into the femoral neck of one femur, and six animals received an intraosseous injection of 0.5 mL of CPM alone into the femoral neck of one femur. The contralateral femur of each of the animals was left untreated. The proximal aspect of each femur was evaluated monthly with use of radiography and at six months with use of peripheral quantitative computed tomography, microcomputed tomography, histological analysis, and mechanical testing. Two additional animals received an intraosseous injection of 0.5 mL of 1.5 mg/mL rhBMP-2/CPM into the femoral neck of one femur. The contralateral femur of each animal was left untreated. Bone formation in the intact specimens from these animals was histologically analyzed at one month in one animal and at three months in the other., Results: Radiographic evaluation over the six-month study period demonstrated an increase in cortical thickness and density in the rhBMP-2/CPM-treated femora as compared to the findings in the untreated contralateral femora or the femora that had been treated with CPM alone. At six months, the rhBMP-2/CPM-treated femora had decreased cortical density and increased cross-sectional area, cortical thickness, trabecular density, and trabecular volume fraction as compared with the contralateral untreated femora and the femora that had received CPM treatment alone, but the differences between the femora that had been treated with CPM alone and the contralateral untreated femora did not reach significance. Increases in bone structure resulted in a 13.7% ± 7.6% (p = 0.032) increase in the maximum bending force at the femoral neck as compared with that at the femoral neck of the contralateral untreated femora. The maximum bending force at the femoral neck was similar between the femora that had been treated with CPM alone and the contralateral untreated femora. De novo and appositional bone formation was present at one month after treatment in the rhBMP-2/CPM-treated femora., Conclusions: This study demonstrates an increase in bone structure and mechanical properties at six months following a single injection of rhBMP-2/CPM into the femoral neck of chronic ovariectomized nonhuman primates.

Background: Transient bone resorption limits the use of recombinant human bone morphogenetic protein-2 (rhBMP-2)/absorbable collagen sponge in metaphyseal bone. The purpose of the present study was to evaluate the efficacy of rhBMP-2/calcium phosphate matrix (CPM) to induce bone formation while limiting transient bone resorption in nonhuman primate core defects., Methods: Metaphyseal core defects were created in eighteen cynomolgus monkeys. rhBMP-2 retention was evaluated in the distal part of the radius. Bone formation was evaluated at eight weeks following treatment with 1.5 or 4.5-mg/mL rhBMP-2/CPM, CPM alone, or no treatment in the distal part of the radius, the proximal part of the tibia, and the proximal part of the femur; at twenty-four weeks following treatment with 1.5-mg/mL rhBMP-2/CPM or CPM alone in the proximal part of the tibia; and at one, two, and four weeks following treatment with 1.5-mg/mL rhBMP-2/CPM or no treatment in the distal part of the radius. Bone resorption was evaluated at four weeks following treatment with 1.5, 2.0, 3.0, and 4.5-mg/mL rhBMP-2/CPM or CPM alone in the distal part of the femur. Evaluations were performed with use of scintigraphy, radiographs, histological analysis, and computed tomography., Results: Seventy-eight percent, 64%, 50%, 35%, and 12% of the rhBMP-2 was retained in the distal part of the radius at one, seven, fourteen, twenty-one, and forty-nine days after surgery. rhBMP-2/CPM increased bone formation within core defects and surrounding trabeculae compared with CPM alone or no treatment at all anatomic locations at eight weeks, and bone formation was ongoing in the rhBMP-2/CPM-treated proximal tibial sites at twenty-four weeks. Bone formation began in the trabeculae surrounding the core defects at one week and was observed adjacent to the resorbing CPM within the core defects and in the surrounding trabecular bone at two and four weeks in the rhBMP-2/CPM-treated distal radial sites. Bone formation was confined to the region immediately surrounding the core defects in the untreated distal radial sites at all time points. Transient bone resorption was only observed in the distal femoral sites treated with 4.5 mg/mL of rhBMP-2/CPM at two weeks., Conclusions: Treatment of nonhuman primate metaphyseal core defects with 1.5 to 3.0-mg/mL rhBMP-2/CPM resulted in bone formation without transient bone resorption., Clinical Relevance: rhBMP-2/CPM may be useful to accelerate healing of metaphyseal bone defects in humans.

Background: Endosseous implants coated with recombinant human bone morphogenetic protein-2 (rhBMP-2) in a laboratory bench setting and air-dried induce relevant bone formation but also resident bone remodeling. Thus, the objective of this study is to evaluate the effect of implants fully or partially coated with rhBMP-2 and vacuum-dried using an industrial process on local bone formation and resident bone remodeling., Methods: Twelve male adult Hound Labrador mongrel dogs were used. Critical-size, supraalveolar, peri-implant defects received titanium porous oxide surface implants coated in their most coronal aspect with rhBMP-2 (coronal-load, six animals), or by immersion of the entire implant in a rhBMP-2 solution (soak-load, six animals) for a total of 30 μg rhBMP-2 per implant. All implants were vacuum-dried. The animals were sacrificed at 8 weeks for histometric evaluation., Results: Clinical healing was unremarkable. Bone formation was not significantly affected by the rhBMP-2 application protocol. New bone height and area averaged (± SE) 3.2 ± 0.5 versus 3.6 ± 0.3 mm, and 2.3 ± 0.5 versus 2.6 ± 0.8 mm(2) for coronal-load and soak-load implants, respectively (P >0.05). The corresponding bone density and bone-implant contact registrations averaged 46.7% ± 5.8% versus 31.6% ± 4.4%, and 28% ± 5.6% versus 36.9% ± 3.4% (P >0.05). In contrast, resident bone remodeling was significantly influenced by the rhBMP-2 application protocol. Peri-implant bone density averaged 72.2% ± 2.1% for coronal-load versus 60.6% ± 4.7% for soak-load implants (P <0.05); the corresponding bone-implant contact averaged 70.7% ± 6.1% versus 47.2% ± 6.0% (P <0.05)., Conclusions: Local application of rhBMP-2 and vacuum-drying using industrial process seems to be a viable technology to manufacture implants that support local bone formation and osseointegration. Coronal-load implants obviate resident bone remodeling without compromising local bone formation.

Background: Implants coated with recombinant human bone morphogenetic protein-2 (rhBMP-2) induce relevant bone formation but also resident bone remodelling., Objectives: To compare the effect of implants fully or partially coated with rhBMP-2 on new bone formation and resident bone remodelling., Materials and Methods: Twelve, male, adult, Hound Labrador mongrel dogs were used. Critical-size, supraalveolar, peri-implant defects received titanium porous oxide surface implants coated in their most coronal aspect with rhBMP-2 (coronal-load/six animals) or by immersion of the entire implant in an rhBMP-2 solution (soak-load/six animals) for a total of 30 mug rhBMP-2/implant. All implants were air-dried. The animals were euthanized at 8 weeks for histometric evaluation., Results: Clinical healing was uneventful. Supraalveolar bone formation was not significantly affected by the rhBMP-2 application protocol. New bone height and area averaged (+/- SE) 3.4 +/- 0.2 versus 3.5 +/- 0.4 mm and 2.6 +/- 0.4 versus 2.5 +/- 0.7 mm(2) for coronal-load and soak-load implants, respectively (p>0.05). The corresponding bone density and bone-implant contact (BIC) recordings averaged 38.0 +/- 3.8%versus 34.4 +/- 5.6% and 25.0 +/- 3.8%versus 31.2 +/- 3.3% (p>0.05). In contrast, resident bone remodelling was significantly influenced by the rhBMP-2 application protocol. Bone density outside the implants threads averaged 74.7 +/- 3.8% and 50.8 +/- 4.1% for coronal-load and soak-load implants, respectively (p<0.05); bone density within the thread area averaged 51.8 +/- 1.2% and 37.8 +/- 2.9%, and BIC 70.1 +/- 6.7% and 43.3 +/- 3.9% (p<0.05)., Conclusion: Local application of rhBMP-2 appears to be a viable technology to support local bone formation and osseointegration. Coronal-load implants obviate resident bone remodelling without compromising new bone formation.

Background: Bone resorption preceding bone formation has been reported following the administration of recombinant human bone morphogenetic protein-2 (rhBMP-2) delivered in an absorbable collagen sponge (ACS) in metaphyseal bone. This study characterizes treatment with rhBMP-2/ACS in metaphyseal bone with use of a nonhuman primate core-defect model., Methods: Unilateral proximal femoral core defects were treated with 360 microg of rhBMP-2/ACS or ACS alone or were left untreated in seven, five, and five adult male cynomolgus monkeys, respectively. Distal femoral core defects in seven of the above animals were treated with 360 microg of rhBMP-2/ACS in one limb and ACS alone in the contralateral limb. Retention of rhBMP-2 in the proximal part of the femora was determined with use of tracer amounts of (125)I-rhBMP-2 imaged with a gamma camera. The distal part of the femora was evaluated with in vivo computed tomography. Computed tomography and histological evaluation were performed on harvested segments in all animals at twenty-four weeks. The histological response in the proximal and distal parts of the femora containing core defects treated with 360 microg of rhBMP-2/ACS in one limb and ACS alone in the contralateral limb was evaluated at one, two, and four weeks in three animals per time point., Results: Approximately 39.9%, 24.2%, 3.4%, and 0.5% of the rhBMP-2 was retained in the proximal part of the femora at one, seven, fourteen, and twenty-one days, respectively. The mineral density and trabecular volume fraction of the core defects treated with rhBMP-2/ACS, those treated with ACS alone, and untreated core defects in the proximal part of the femora were 81%, 54%, and 20%, respectively, and 94%, 36%, and 31%, respectively, of the corresponding region in the contralateral limbs at twenty-four weeks. The mineral density and trabecular volume fraction of the region surrounding the core defects treated with rhBMP-2/ACS, those treated with ACS alone, and untreated core defects were 112%, 105%, and 104%, respectively, and 117%, 108%, and 107%, respectively, of the corresponding region in the contralateral limbs. Treatment with rhBMP-2/ACS increased the size of the proximal and distal core defects compared with treatment with ACS alone. Histological evaluation of the rhBMP-2/ACS-treated limbs demonstrated that bone resorption was initiated at one week in association with osteoclasts and receptor activator of nuclear factor-kappaB ligand-positive stained spindle-shaped cells and peaked at two weeks. Bone formation was observed at two weeks and was ongoing at twenty-four weeks., Conclusions: Treatment of metaphyseal core defects with rhBMP-2/ACS resulted in bone resorption followed by bone formation in nonhuman primates.

Background: Conventional oral/maxillofacial implants reach osseointegration over several months during which the titanium fixtures interact with alveolar bone. The objective of this study was to determine if adsorbing recombinant human bone morphogenetic protein-2 (rhBMP-2) onto a titanium porous oxide (TPO) implant surface might enhance or accelerate local bone formation and support osseointegration in a large animal oral/maxillofacial orthotopic model., Material and Methods: Endosseous implants with a TPO surface were installed into the edentulated posterior mandible in eight adult Hound Labrador mongrel dogs. The implant surface had been adsorbed with rhBMP-2 at 0.2 or 4.0 mg/ml. TPO implants without rhBMP-2 served as control. Treatments were randomized between jaw quadrants. Mucosal flaps were advanced and sutured leaving the implants submerged. Clinical and radiographic evaluations were made immediately post-surgery, at day 10 (suture removal), and week 4 and 8 post-surgery. The animals received fluorescent bone markers at week 3, 4, and at week 8 post-surgery, when they were euthanized for histologic analysis., Results: TPO implants coated with rhBMP-2 exhibited dose-dependent bone remodelling including immediate resorption and formation of implant adjacent bone, and early establishment of clinically relevant osseointegration. The resulting bone-implant contact, although clinically respectable, appeared significantly lower for rhBMP-2-coated implants compared with the control [rhBMP-2 (0.2 mg/ml) 43.3+/-10.8%versus 71.7+/-7.8%, p<0.02; rhBMP-2 (4.0 mg/ml) 35.4+/-10.6%versus 68.2+/-11.0%, p<0.03]., Conclusions: rhBMP-2 adsorbed onto TPO implant surfaces initiates dose-dependent peri-implant bone re-modelling resulting in the formation of normal, physiologic bone and clinically relevant osseointegration within 8 weeks.

Background: Studies using ectopic rodent and orthotopic canine models (Type II bone) have shown that titanium porous oxide (TPO) surface implants adsorbed with recombinant human bone morphogenetic protein-2 (rhBMP-2) induce local bone formation including osseointegration. The objective of this study was to evaluate local bone formation and osseointegration at such implants placed into Type IV bone., Material and Methods: rhBMP-2-coated implants were installed into the edentulated posterior maxilla in eight young adult Cynomolgus monkeys: four animals each received three TPO implants adsorbed with rhBMP-2 (2.0 mg/ml) and four animals each received three TPO implants adsorbed with rhBMP-2 (0.2 mg/ml). Contra-lateral jaw quadrants received three TPO implants without rhBMP-2 (control). Treatments were alternated between left and right jaw quadrants. Mucosal flaps were advanced and sutured to submerge the implants. The animals received fluorescent bone markers at weeks 2, 3, 4, and at week 16 when they were euthanized for histologic analysis., Results: Clinical healing was uneventful. Extensive local bone formation was observed in animals receiving implants adsorbed with rhBMP-2 (2.0 mg/ml). The newly formed bone exhibited a specific pinpoint bone-implant contact pattern regardless of rhBMP-2 concentration resulting in significant osseointegration; rhBMP-2 (2.0 mg/ml): 43% and rhBMP-2 (0.2 mg/ml): 37%. Control implants exhibited a thin layer of bone covering a relatively larger portion of the implant threads. Thus, TPO control implants bone exhibited significantly greater bone-implant contact ( approximately 75%; p<0.05). There were no statistically significant differences between rhBMP-2-coated and control implants relative to any other parameter including peri-implant and intra-thread bone density., Conclusion: rhBMP-2-coated TPO implants enhanced/accelerated local bone formation in Type IV bone in a dose-dependent fashion in non-human primates resulting in significant osseointegration. rhBMP-2-induced de novo bone formation did not reach the level of osseointegration observed in native resident bone within the 16-week interval.

Background: Studies using ectopic rodent, orthotopic canine, and non-human primate models show that bone morphogenetic proteins (BMPs) coated onto titanium surfaces induce local bone formation. The objective of this study was to examine the ability of recombinant human BMP-2 (rhBMP-2) coated onto a titanium porous oxide implant surface to stimulate local bone formation including osseointegration and vertical augmentation of the alveolar ridge., Material and Methods: Bilateral, critical-size, 5 mm, supra-alveolar, peri-implant defects were created in 12 young adult Hound Labrador mongrel dogs. Six animals received implants coated with rhBMP-2 at 0.75 or 1.5 mg/ml, and six animals received implants coated with rhBMP-2 at 3.0 mg/ml or uncoated control. Treatments were randomized between jaw quadrants. The mucoperiosteal flaps were advanced, adapted and sutured to submerge the implants for primary intention healing. The animals received fluorescent bone markers at weeks 3, 4, 7 and 8 post-surgery when they were euthanized for histologic evaluation., Results: Jaw quadrants receiving implants coated with rhBMP-2 exhibited gradually regressing swelling that became hard to palpate disguising the contours of the implants. The histologic evaluation showed robust bone formation reaching or exceeding the implant platform. The newly formed bone exhibited characteristics of the adjoining resident Type II bone including cortex formation for sites receiving implants coated with rhBMP-2 at 0.75 or 1.5 mg/ml. Sites receiving implants coated with rhBMP-2 at 3.0 mg/ml exhibited more immature trabecular bone formation, seroma formation and peri-implant bone remodelling resulting in undesirable implant displacement. Control implants exhibited minimal, if any, bone formation. Thus, implants coated with rhBMP-2 at 0.75, 1.5 and 3.0 mg/ml exhibited significant bone formation (height and area) compared with the sham-surgery control averaging (+/-SD) 4.4+/-0.4, 4.2+/-0.7 and 4.2+/-1.2 versus 0.8+/-0.3 mm; and 5.0+/-2.2, 5.6+/-2.2 and 7.4+/-3.5 versus 0.7+/-0.3 mm(2), respectively (p<0.01). All the treatment groups exhibited clinically relevant osseointegration., Conclusions: rhBMP-2 coated onto titanium porous oxide implant surfaces induced clinically relevant local bone formation including vertical augmentation of the alveolar ridge and osseointegration. Higher concentrations/doses were associated with untoward effects.

Objective: Myostatin is an endogenous negative regulator of muscle growth and a novel target for muscle diseases. We conducted a safety trial of a neutralizing antibody to myostatin, MYO-029, in adult muscular dystrophies (Becker muscular dystrophy, facioscapulohumeral dystrophy, and limb-girdle muscular dystrophy)., Methods: This double-blind, placebo-controlled, multinational, randomized study included 116 subjects divided into sequential dose-escalation cohorts, each receiving MYO-029 or placebo (Cohort 1 at 1 mg/kg; Cohort 2 at 3 mg/kg; Cohort 3 at 10 mg/kg; Cohort 4 at 30 mg/kg). Safety and adverse events were assessed by reported signs and symptoms, as well as by physical examinations, laboratory results, echocardiograms, electrocardiograms, and in subjects with facioscapulohumeral dystrophy, funduscopic and audiometry examinations. Biological activity of MYO-029 was assessed through manual muscle testing, quantitative muscle testing, timed function tests, subject-reported outcomes, magnetic resonance imaging studies, dual-energy radiographic absorptiometry studies, and muscle biopsy., Results: MYO-029 had good safety and tolerability with the exception of cutaneous hypersensitivity at the 10 and 30 mg/kg doses. There were no improvements noted in exploratory end points of muscle strength or function, but the study was not powered to look for efficacy. Importantly, bioactivity of MYO-029 was supported by a trend in a limited number of subjects toward increased muscle size using dual-energy radiographic absorptiometry and muscle histology., Interpretation: This trial supports the hypothesis that systemic administration of myostatin inhibitors provides an adequate safety margin for clinical studies. Further evaluation of more potent myostatin inhibitors for stimulating muscle growth in muscular dystrophy should be considered.

Background: The repulsive guidance molecule (RGM) proteins, originally discovered for their roles in neuronal development, have been recently identified as co-receptors in the bone morphogenetic protein (BMP) signaling pathway. BMPs are members of the TGFbeta superfamily of signaling cytokines, and serve to regulate many aspects of cellular growth and differentiation., Results: Here, we investigate whether RGMa, RGMb, and RGMc play required roles in BMP and TGFbeta signaling in the mouse myoblast C2C12 cell line. These cells are responsive to BMPs and are frequently used to study BMP/TGFbeta signaling pathways. Using siRNA reagents to specifically knock down each RGM protein, we show that the RGM co-receptors are required for significant BMP signaling as reported by two cell-based BMP activity assays: endogenous alkaline phosphatase activity and a luciferase-based BMP reporter assay. Similar cell-based assays using a TGFbeta-induced luciferase reporter show that the RGM co-receptors are not required for TGFbeta signaling. The binding interaction of each RGM co-receptor to each of BMP2 and BMP12 is observed and quantified, and equilibrium dissociation constants in the low nanomolar range are reported., Conclusion: Our results demonstrate that the RGMs play a significant role in BMP signaling and reveal that these molecules cannot functionally compensate for one another.

Background: Alveolar ridge aberrations commonly require bone augmentation procedures for optimal placement of endosseous dental implants. The objective of this study was to evaluate local bone formation following implantation of recombinant human bone morphogenetic protein-2 (rhBMP-2) in an absorbable collagen sponge (ACS) carrier with or without provisions for guided bone regeneration (GBR) as potential treatment modalities for alveolar augmentation., Methods: Surgically induced, large, mandibular alveolar ridge saddle-type defects (2 defects/jaw quadrant) in seven young adult Hound dogs were assigned to receive rhBMP-2/ACS, rhBMP-2/ACS combined with GBR (rhBMP-2/GBR), GBR, and surgery controls. The animals were euthanized at 12 weeks post-surgery when block sections of the defect sites were collected for histologic analysis., Results: Clinical complications included swelling for sites receiving rhBMP-2 and wound failure with exposure of the barrier device for sites receiving GBR (4/6) or rhBMP-2/GBR (3/7). The radiographic evaluation showed substantial bone fill for sites receiving rhBMP-2/ACS, rhBMP-2/GBR, and GBR. In particular, sites receiving rhBMP-2/GBR presented with seroma-like radiolucencies. The surgery control exhibited moderate bone fill. To evaluate the biologic potential of the specific protocols, sites exhibiting wound failure were excluded from the histometric analysis. Sites receiving rhBMP-2/ACS or rhBMP-2/GBR exhibited bone fill averaging 101%. Bone fill averaged 92% and 60%, respectively, for sites receiving GBR and surgery controls. Bone density ranged from 50% to 57% for sites receiving rhBMP-2/ACS, GBR, or surgery controls. Bone density for sites receiving rhBMP-2/GBR averaged 34% largely due to seroma formation encompassing 13% to 97% of the sites., Conclusion: rhBMP-2/ACS appears to be an effective alternative to GBR in the reconstruction of advanced alveolar ridge defects. Combining rhBMP-2/ACS with GBR appears to be of limited value due to the potential for wound failure or persistent seromas.

Objectives: The potential of recombinant human bone morphogenetic protein-2 (rhBMP-2) to enhance bone healing following endodontic surgery was tested. The pattern and timing of de novo bone formation and cementum regeneration, and the potential for root resorption and ankylosis to accompany bone formation were evaluated., Study Design: Pulpal infections were induced in maxillary and mandibular incisor teeth in young adult Cynomolgus monkeys. The teeth received conventional endodontic treatment immediately followed by surgical root resection. In a randomized split-mouth design, contralateral apical bone defects received rhBMP-2 in absorbable collagen sponge (ACS) carrier or served as sham-surgery controls to provide histological and radiographic evaluations following 1 (mandibular incisors) and 4.5 (maxillary incisors) month(s) postsurgery., Results: At 1 month postsurgery trabecular bone filled the apical bone defects. The newly formed bone appeared considerably more mature and had assumed characteristics of the contiguous resident bone at 4.5 months postsurgery. The resected root tips were almost completely covered by new cementum with a maturing functionally oriented periodontal ligament. Localized inflammatory infiltrates were associated with the filled root canals and extruded root-filling material. Root resorption and ankylosis were not observed. There were no apparent differences in healing patterns between sites implanted with rhBMP-2/ACS and those serving as sham-surgery controls., Conclusions: Under conditions where the influence of infectious elements and irritation caused by root filling material are minimized, bone formation and cementum regeneration appears rapid following endodontic surgery. rhBMP-2/ACS did not offer an obvious benefit above and beyond that of the native osteogenic potential in this animal model.

Two major isoforms of the Runx2 gene are expressed by alternative promoter usage: Runx2 type I (Runx2-I) is derived from the proximal promoter (P2), and Runx2 type II (Runx2-II) is produced by the distal promoter (P1). Our previous results indicate that Dlx5 mediates BMP-2-induced Runx2 expression and osteoblast differentiation (Lee, M.-H., Kim, Y-J., Kim, H-J., Park, H-D., Kang, A-R., Kyung, H.-M., Sung, J-H., Wozney, J. M., Kim, H-J., and Ryoo, H-M. (2003) J. Biol. Chem. 278, 34387-34394). However, little is known of the molecular mechanisms by which Dlx5 up-regulates Runx2 expression in BMP-2 signaling. Here, Runx2-II expression was found to be specifically stimulated by BMP-2 treatment or by Dlx5 overexpression. In addition, BMP-2, Dlx5, and Runx2-II were found to be expressed in osteogenic fronts and parietal bones of the developing cranial vault and Runx2-I and Msx2 in the sutural mesenchyme. Furthermore, Runx2 P1 promoter activity was strongly stimulated by Dlx5 overexpression, whereas Runx2 P2 promoter activity was not. Runx2 P1 promoter deletion analysis indicated that the Dlx5-specific response is due to sequences between -756 and -342 bp of the P1 promoter, where three Dlx5-response elements are located. Dlx5 responsiveness to these elements was confirmed by gel mobility shift assay and site-directed mutagenesis. Moreover, Msx2 specifically suppressed the Runx2 P1 promoter, and the responsible region overlaps with that recognized by Dlx5. In summary, Dlx5 specifically transactivates the Runx2 P1 promoter, and its action on the P1 promoter is antagonized by Msx2.

Carriers for bone morphogenetic proteins (BMPs) are used to increase retention of these factors at orthopedic treatment sites for a sufficient period of time to allow regenerative tissue forming cells to migrate to the area of injury and to proliferate and differentiate. Carriers can also serve as a matrix for cell infiltration while maintaining the volume in which repair tissue can form. Carriers have to be biocompatible and are often required to be bioresorbable. Carriers also have to be easily, and cost-effectively, manufactured for large-scale production, conveniently sterilized and have appropriate storage requirements and stability. All of these processes have to be approvable by regulatory agencies. The four major categories of BMP carrier materials include natural polymers, inorganic materials, synthetic polymers, composites of these materials. Autograft or allograft carriers have also used. Carrier configurations range from simple depot delivery systems to more complex systems mimicking the extracellular matrix structure and function. Bone regenerative carriers include depot delivery systems for fracture repair, three-dimensional polymer or ceramic composites for segmental repairs and spine fusion and metal or metal/ceramic composites for augmenting implant integration. Tendon/ligament regenerative carriers range from depot delivery systems to three-dimensional carriers that are either randomly oriented or linearly oriented to improve regenerative tissue alignment. Cartilage regenerative systems generally require three-dimensional matrices and often incorporate cells in addition to factors to augment the repair. Alternative BMP delivery systems include viral vectors, genetically altered cells, conjugated factors and small molecules.

Background: The objective of this study was to evaluate alveolar ridge augmentation following surgical implantation of recombinant human bone morphogenetic protein-2 (rhBMP-2) using two novel space-providing carrier technologies in the baboon (Papio anubis) model., Methods: Standardized alveolar ridge defects ( approximately 15 x 8 x 5 mm) were surgically produced in maxillary and mandibular edentulous areas in four baboons. The defect sites were implanted with rhBMP-2 (0.4 mg/mL) in a tricalcium phosphate/hydroxyapatite/ absorbable collagen sponge composite (TCP/HA/ACS) or calcium phosphate cement (alpha-BSM). Control treatments were TCP/HA/ACS and ?-BSM without rhBMP-2 and sham surgery. Stainless steel pins were placed at the mid-apical and coronal level of the defect sites to provide landmarks for clinical measurements pre- and post-implantation. Impressions were obtained pre- and postimplantation to determine changes in alveolar ridge volume. Radiographic registrations were obtained pre- and post-implantation. Block sections of the defect sites were harvested at week 16 postimplantation and processed for histometric analysis including new bone area and bone density. Statistical comparisons between treatments were made using a mixed effect generalized linear model using least squares estimation., Results: The carrier systems without rhBMP-2 provided a modest ridge augmentation. The addition of rhBMP-2 resulted in an almost 2-fold increase in alveolar ridge width, including a greater percentage of trabecular bone and a higher bone density compared to controls (P < or =0.05) without significant differences between the two rhBMP-2 protocols., Conclusions: TCP/HA/ACS and alphaBSM appear to be suitable carrier technologies for rhBMP-2. Alveolar augmentation procedures using either technology combined with rhBMP-2, rather than stand-alone therapies, may provide clinically relevant augmentation of alveolar ridge defects for placement of endosseous dental implants.

Alkaline phosphatase (ALP) is a widely accepted bone marker. Its expression is stimulated by bone morphogenetic protein (BMP)-2 treatment, the activation of BMP receptors and R-Smads, and the expression of Dlx5 and Runx2. However, how BMP-2 induces ALP expression is not clearly understood. We dissected the murine ALP promoter and found within it a Dlx5-binding cis-acting element by electrophoretic mobility shift assays and site-directed mutagenesis of the element. Dlx5 and the product of its target gene, Runx2, stimulated ALP promoter activity in an additive manner. However, because Dlx5 continued to stimulate ALP expression in Runx2(-/-) cells, the ALP stimulatory activity of Dlx5 is independent of Runx2. We also found that overexpression of Msx2 suppressed the mRNA level and enzyme activity of ALP that were induced by BMP-2 stimulation, and suppressed the Dlx5-stimulated ALP promoter activity by competing with Dlx5 for the cis-acting element in the ALP promoter. Moreover, Msx2 levels are constitutively high in C2C12 myogenic cells but decrease over time after BMP-2 treatment. This may explain why BMP-2 treatment of these cells results in immediate Dlx5 expression yet ALP expression commences only 1-2 days later. In other words, Msx2 in high levels counteracts initially the transcriptional activity of Dlx5 in low levels until a threshold Dlx5:Msx2 ratio is reached to the levels that allow the ALP stimulatory activity of Dlx5 to prevail. Thus, Dlx5 transactivates ALP expression, directly by binding to its cognate response element and/or indirectly by stimulating Runx2 expression, and Msx2 counteracts the direct transactivation of Dlx5.

Bioactive proteins signal host or transplanted cells to form the desired tissue type. Matrix systems are utilized to locally deliver the proteins and to maintain effective protein concentrations. For some indications, a matrix is required to define the physical form of the regenerated tissue. Substantial progress has been made in bone tissue engineering in recent years, based on the results of controlled clinical studies using bone morphogenetic proteins. Ongoing research in this area centers on the design of additional delivery matrices to expand the clinical indications, using synthetic delivery systems that mimic biological qualities of the natural materials currently in use. Although a similar rationale exists for the regeneration of articular cartilage with bioactive factors, advancement in this area has not been as substantial.

Background: Recombinant human bone morphogenetic protein-2 (rhBMP-2) has been shown to induce clinically relevant bone formation for orthopedic, craniofacial, and oral indications. It appears critical, in particular for onlay indications, that the associated carrier technology exhibits structural integrity to offset compressive forces in support of rhBMP-2-induced bone formation. The objective of this study was to evaluate a calcium phosphate (CP) cement, Ceredex, as a candidate carrier for rhBMP-2 in a defect model with limited osteogenic potential., Materials: Bilateral, critical size, 6-mm, supra-alveolar, periodontal defects were created in six, adult, male, Hound Labrador mongrels. Three animals received rhBMP-2/Ceredex (rhBMP-2 at 0.20 and 0.40 mg/ml) in contralateral defect sites (implant volume/defect approximately 1 ml). One defect site in each of the three remaining animals received Ceredex without rhBMP-2 (control). The animals were euthanized at 12 weeks postsurgery for histologic and histometric analysis., Results: Mean induced bone height exceeded 80% of the defect height for supra-alveolar periodontal defects receiving rhBMP-2/Ceredex without major differences between rhBMP-2 concentrations compared with approximately 40% for the control. The newly formed bone, a mixture of lamellar and woven bone in fibrovascular tissue, circumscribed relatively large portions of the residual Ceredex biomaterial. Inflammatory lesions were associated with limited bone formation in some sites. From a periodontal perspective, sites receiving rhBMP-2/Ceredex exhibited increased cementum formation compared with control, but without a functionally oriented periodontal ligament, and increased ankylosis and root resorption. Control sites exhibited early wound failure and exposure, loss of the Ceredex biomaterial, and limited bone formation., Conclusions: The Ceredex CP cement appears a potentially promising carrier technology for rhBMP-2 onlay indications. However, a slow resorption rate may prevent its wider use. This study does not support use of the rhBMP-2/Ceredex combination for periodontal indications.

Background: In recent clinical trials demonstrating the efficacy of recombinant human bone morphogenetic protein-2 (rhBMP-2) for the acceleration of bone-healing, investigators used carriers requiring open surgery for administration. In this study, we used a nonhuman primate fibular osteotomy model to evaluate injectable rhBMP-2/carrier formulations that can be administered in closed fractures., Methods: The fibular osteotomy model was first characterized by evaluating surgically harvested fibular segments containing untreated osteotomy sites (controls) from seventy adult male Cynomolgus monkeys at eight weeks (twenty-four monkeys), ten weeks (thirty-four), twelve weeks (six), and fourteen weeks (six). Fibular segments, from twenty-four animals, in which an osteotomy had not been performed served as normal controls (intact). The contralateral limb of twenty-four of the animals was then used to evaluate the effect of rhBMP-2 administered, three hours after the osteotomy, in eight carrier formulations (buffer, calcium phosphate paste, and hyaluronan gel, hyaluronan paste, and gelatin foam formulated with and without tricalcium phosphate granules). Each carrier was used in three monkeys. At ten weeks, the fibulae with the treated osteotomy sites were harvested and were compared with the contralateral, untreated osteotomized fibulae (paired control). The most promising carrier, calcium phosphate paste (alpha bone substitute material, or alpha-BSM), was then evaluated in eleven additional animals. The outcomes included the findings on radiographs made weekly until the time of fibular harvest, the callus area, the biomechanical properties, and the histologic findings., Results: Radiographic and histologic studies confirmed complete bridging of the control osteotomy sites in most animals by fourteen weeks. The mean torsional stiffness and maximum torque of the control osteotomy sites were 42.7% and 53.7%, 55.2% and 60.4%, 66.7% and 66.4% of the mean torsional stiffness and maximum torque of the intact fibulae at eight, ten, and twelve weeks, respectively, but they were not substantially different from the mean torsional stiffness and maximum torque of the intact fibulae at fourteen weeks (82.3% and 79.8%). In the carrier screening study, outcome measures of healing were more consistently enhanced in the rhBMP-2/alpha-BSM-treated osteotomy sites. In the confirmatory study, the mean callus area, torsional stiffness, and maximum torque were 86%, 72%, and 68% greater in the rhBMP-2/alpha-BSM-treated osteotomy sites than in the paired-control osteotomy sites at ten weeks (p < 0.001). The torsional stiffness and maximum torque in the rhBMP-2/alpha-BSM-treated osteotomy sites were equal to those in the intact fibulae, whereas those parameters in the paired-control osteotomy sites were only 55% and 58%, respectively, of the torsional stiffness and maximum torque of the intact fibulae. Histologic analysis confirmed complete osseous bridging of the rhBMP-2/alpha-BSM-treated osteotomy sites but incomplete bridging of the paired-control osteotomy sites at ten weeks., Conclusions: A single percutaneous injection of rhBMP-2/alpha-BSM accelerates the healing of fibular osteotomy sites in nonhuman primates by approximately 40% compared with the healing of untreated osteotomy sites.

Objectives: Subcutaneous and intramuscular implants of bone morphogenetic protein-12 (BMP-12) have been shown to induce formation of tendon and ligament tissue. BMP-12 induced a new attachment with a distinct fibrocartilaginous zone at the tendon-bone interface in the rat tendon-bone attachment model. Surgical controls showed poor healing and failure to reform the appropriate tendon-bone attachment morphologically. Application of recombinant human BMP-12 (rhBMP-12) to periodontal defects suggests that rhBMP-12 has the potential to support regeneration of the periodontal ligament (PDL). The objective of this pilot study was to evaluate this effect of rhBMP-12 in a tooth replantation model., Methods: Six, young adult, male Hound Labrador mongrel dogs were used. Maxillary and/or mandibular incisor and premolar teeth were extracted and the PDL was either left "intact" or removed by root planing. rhBMP-12 (1.0 mg/ml) or a buffer control was topically applied to teeth with "intact" PDL in contralateral jaw quadrants in each of 3 animals. The teeth were immersed in 1.0 ml of the rhBMP-12 or the buffer solution for 10 min and then replanted. The remaining three animals received rhBMP-12 (1.0 mg/ml) and the buffer control in a similar fashion applied to teeth instrumented to remove the PDL and cementum, and surface demineralized with citric acid. The animals were euthanized at 8 weeks postsurgery and block sections were collected and processed for histopathologic analysis., Results: No dramatic differences were found between teeth receiving topical rhBMP-12 and the buffer control. Application of rhBMP-12 did not have an apparent effect on new cementum and PDL formation in the tooth replantation model. Moreover, application of rhBMP-12 did not increase nor did it decrease the apparent presence and extent of ankylosis along the root surface compared to the control., Conclusions: The observations from this study do not support the use of topical rhBMP-12 to support the reestablishment of the PDL including regeneration of cementum and functionally oriented fibers, and to prevent ankylosis and root resorption following replantation of teeth.

Objectives: Recombinant human bone morphogenetic protein-2 (rhBMP-2) has been shown to stimulate alveolar bone and cementum formation in periodontal defects but not a functionally oriented periodontal ligament (PDL). Subcutaneous and intramuscular implants of BMP-12 have been shown to induce tendon formation and ligament-like tissue. The objective of this study was to evaluate rhBMP-12 for periodontal regeneration, in particular PDL formation., Methods: Six young adult Hound Labrador mongrel dogs were used. Routine supraalveolar periodontal defects were created around the mandibular premolar teeth. Three animals received rhBMP-12(0.04 mg/ml) in an absorbable collagen sponge (ACS) carrier vs. rhBMP-12(0.2 mg/mL)/ACS in contralateral defects. Three animals received rhBMP-12(1.0 mg/ml)/ACS vs. rhBMP-2(0.2 mg/ml)/ACS (total implant volume/defect approximately 1 ml). The animals were euthanized 8 weeks postsurgery and block biopsies were processed for histometric analysis., Results: Bone regeneration appeared increased in sites receiving rhBMP-2/ACS compared to sites receiving rhBMP-12/ACS. Cementum regeneration was similar comparing sites implanted with rhBMP-2/ACS to sites implanted with rhBMP-12/ACS. In contrast, sites receiving rhBMP-12/ACS exhibited a functionally oriented PDL bridging the gap between newly formed bone and cementum whereas this was a rare observation in sites receiving rhBMP-2/ACS. Ankylosis appeared increased in sites receiving rhBMP-2/ACS compared to those receiving rhBMP-12/ACS., Conclusions: The outcomes of this study suggest that rhBMP-12 may have significant effects on regeneration of the PDL. Additional preclinical evaluation is needed to confirm these initial observations prior to clinical application.

Background: Previous studies have shown a limited potential for bone augmentation following guided bone regeneration (GBR) in horizontal alveolar defects. Surgical implantation of recombinant human bone morphogenetic protein-2 (rhBMP-2) in an absorbable collagen sponge carrier (ACS) significantly enhances bone regeneration in such defects; however, sufficient quantities of bone for implant dentistry are not routinely obtained. The objective of this study was to evaluate the potential of rhBMP-2/ACS to enhance GBR using a space-providing, macro-porous expanded polytetrafluoroethylene (ePTFE) device., Methods: Bilateral, critical size, supra-alveolar, peri-implant defects were surgically created in four Hound Labrador mongrel dogs. Two turned and one surface-etched 10-mm titanium dental implant were placed 5 mm into the surgically reduced alveolar ridge creating 5-mm supra-alveolar defects. rhBMP-2/ACS (rhBMP-2 at 0.2 mg/ml) or buffer/ACS was randomly assigned to left and right jaw quadrants in subsequent animals. The space-providing, macro-porous ePTFE device was placed to cover rhBMP-2/ACS and control constructs and dental implants. Gingival flaps were advanced for primary wound closure. The animals were euthanized at 8 weeks postsurgery for histologic and histometric analysis., Results: Bone formation was significantly enhanced in defects receiving rhBMP-2/ACS compared to control. Vertical bone gain averaged (+/- SD) 4.7 +/- 0.3 and 4.8 +/- 0.1 mm, and new bone area 10.3 +/- 2.0 and 8.0 +/- 2.5 mm2 at turned and surface-etched dental implants, respectively. Corresponding values for the control were 1.8 +/- 2.0 and 1.3 +/- 1.3 mm, and 1.8 +/- 1.3 and 1.2 +/- 0.6 mm2. Bone-implant contact in rhBMP-2-induced bone averaged 6.4 +/- 1.4% and 9.6 +/- 7.5% for turned and surface-etched dental implants, respectively (P=0.399). Corresponding values for the control were 14.6 +/- 19.4% and 23.7 +/- 9.7% (P=0.473). Bone-implant contact in resident bone ranged between 43% and 58% without significant differences between dental implant surfaces., Conclusions: rhBMP-2/ACS significantly enhances GBR at turned and surface-etched dental implants. The dental implant surface technology does not appear to substantially influence bone formation.

The purpose of the present study was to determine if recombinant human bone morphogenetic protein-2 (rhBMP-2) enhances bone ingrowth into porous-coated implants and gap healing around the implants. In the presence of a 3-mm gap between the implant and host bone, porous-coated implants were placed bilaterally for four weeks in the proximal humeri of skeletally mature, adult male dogs. In three treatment groups, the test implant was treated with HA/TCP and rhBMP-2 in buffer at a dose of 100 microg/implant (n=5), 400 microg/implant (n=6), or 800 microg/implant (n=5) and placed in the left humerus. In these same animals, an internal control implant was treated only with HA/TCP and buffer and placed in the right humerus. These groups were compared with a previously reported external control group of seven animals in which no growth factor was delivered [J. Orthop. Res. 19 (2001) 85]. The BMP treated implants in the two lower dose groups had significantly more bone ingrowth than the external controls with the greatest effect in the 100 g/implant group (a 3.5-fold increase over the external control, p=0.008). All three dose groups had significantly more bone formation in the 3-mm gap surrounding the BMP treated implants than the external controls with the greatest effect in the 800 microg group (2.9-fold increase, p<0.001). Thus, application of rhBMP-2 to a porous-coated implant stimulated local bone ingrowth and gap healing. The enhancement of bone formation within the implant (bone ingrowth) was inversely related to dose.

Osseointegration [direct bone-implant contact (BIC)] is a primary goal following installation of endosseous dental implants. Such bone contact provides stability for the dental implant over time. The objective of this study was to evaluate bone formation and BIC at long-term, functionally loaded, endosseous dental implants placed into bone induced by recombinant human bone morphogenetic protein-2 (rhBMP-2) in an absorbable collagen sponge (ACS) carrier. Mandibular, saddle-type, alveolar ridge defects (approximately 15 x 10 x 10 mm), two per jaw quadrant, were surgically induced in each of six young adult American fox hounds. The defects were immediately implanted with rhBMP-2/ACS. Two defects per animal additionally received a nonresorbable expanded polytetrafluoroethylene (ePTFE) membrane or a bioresorbable polyglycolide fiber membrane. Healing was allowed to progress for 3 months, when the ePTFE membrane was removed, and machined, threaded, titanium dental implants were installed into the rhBMP-2/ACS induced bone and into the adjacent resident bone. At 4 months of osseointegration, the implants were exposed to receive abutments and prosthetic treatment (two- or three-unit bridges). Some implants were removed for histologic analysis. The remainder of implants were exposed to functional loading for 12 months at which time the animals were killed for histometric analysis. One animal died prematurely due to kidney failure unrelated to the experimental protocol and was not included in the analysis. The 12-month block sections from a second animal were lost in the histological processing. Four sites receiving rhBMP-2/ACS and ePTFE or resorbable membranes experienced wound failure and membrane exposure, and subsequently exhibited limited bone formation. Defects without wound failure filled to contour with the adjacent alveolar bone. The newly formed bone exhibited features of the resident bone with a re-established cortex; however, it commonly included radiolucent areas that resolved over time. Dental implants block biopsied at 4 months exhibited limited, if any, crestal resorption, whereas those exposed to functional loading for 12 months exhibited some crestal resorption. Implants biopsied at 4 months exhibited a mean (+/- SD) BIC of 40.6 +/- 8.2% in rhBMP-2/ACS induced bone vs. 52.7 +/- 11.4% in resident bone. Dental implants exposed to 12 months of functional loading exhibited a mean BIC of 51.7 +/- 7.1% in rhBMP-2/ACS induced bone vs. 74.7 +/- 7.0% in resident bone. There were no significant differences between dental implants placed into rhBMP-2/ACS induced bone and resident bone for any parameter at any observation interval. In conclusion, rhBMP-2/ACS-induced bone allows installation, osseointegration, and long-term functional loading of machined, threaded, titanium dental implants in dogs.

This study evaluated the ability of recombinant human bone morphogenetic protein-2 (rhBMP-2) delivered in an injectable calcium phosphate carrier (alpha-BSM) to accelerate healing in a rabbit ulna osteotomy model compared to untreated surgical controls. Healing was assessed by radiography, histology and biomechanics. Bilateral mid-ulnar osteotomies were created in 16 skeletally mature rabbits. One limb in each animal was injected with either 0.1 mg rhBMP-2/alpha-BSM (BMP) (N=8) or buffer/alpha-BSM (BSM) (N=8). Contralateral osteotomies served as untreated surgical controls (SXCT). Gamma scintigraphy showed 75%, 45% and 5% of the initial 125I-rhBMP-2 dose was retained at the osteotomy site at 3 h, 1 week and 3 weeks. The biological activity of rhBMP-2 (alkaline phosphatase activity from bioassay) extracted from alpha-BSM incubated in vitro up to 30 days at 37 degrees C was unchanged. Radiographs demonstrated complete bridging of the BMP limbs at 4 weeks whereas none of the BSM or SXCT limbs were bridged. Post-mortem peripheral quantitative computed tomography determined mineralized callus area was 62% greater in BMP limbs compared to SXCT limbs. Torsional stiffness and strength were 63% and 103% greater in BMP limbs compared to SXCT limbs. There was no difference in torsional properties between BSM and SXCT limbs. Failure occurred outside the osteotomy in four out of seven of the BMP limbs. All BSM and SXCT limbs failed through the osteotomy. Histology showed bony bridging of the osteotomy and no residual carrier in the BMP limbs. BSM and SXCT groups showed less mature calluses composed of primarily fibrocartilaginous tissue and immature bone in the osteotomy gap. These data indicate rhBMP-2 delivered in alpha-BSM accelerated healing in a rabbit ulna osteotomy model compared to BSM and SXCT groups.

BMP-2 stimulates the expression of three osteogenic master transcription factors: Runx2, Dlx5, and Osterix (Osx). However, the hierarchical regulatory relationships among them are not yet clearly understood. Osx was commonly stimulated in osteogenic and non-osteogenic cells in response to BMP-signaling, as Dlx5 was in our previous report. A cycloheximide experiment indicated that Osx expression by BMP-2 requires new protein synthesis. Even if Osx has been suggested as a downstream target of Runx2, the results of this study indicated that Osx expression was still induced by BMP-2 treatment in Runx2 null cells, but not induced by Runx2 overexpression in myogenic C2C12 cells. Instead, Osx expression by BMP-2 was completely abrogated by the antisense blocking of Dlx5. Depending upon the coincident expression pattern of Osx and Dlx5, and the blocking of Osx expression by the antisense Dlx5, BMP-2-induced Osx expression is mainly mediated not by Runx2, but by Dlx5.

Intramuscular injection of BMP-2 induces ectopic bone formation in vivo. Similarly, BMP-2 treatment blocks myogenic differentiation and induces osteoblastic transdifferentiation of premyoblastic C2C12 cells. Previous reports suggested that BMP-2-stimulated Runx2 expression could play a pivotal role in transdifferentiation. However, increased Runx2 expression by TGF-beta 1 did not support osteoblast differentiation in vitro. These results indicate that the induction of Runx2 is not sufficient to explain the BMP-induced transdifferentiation. We found that Dlx5 is specifically expressed in osteogenic cells, and is specifically induced by BMP-2 or -4 signaling but not by other osteotrophic signals or other TGF-beta superfamily members. Cycloheximide treatment indicated that Dlx5 was immediately induced by BMP signaling, while Runx2 required de novo protein synthesis. In addition, blocking or overexpressing each transcription factor indicated that Dlx5 is an indispensable mediator of BMP-2-induced Runx2 expression but is not involved in TGF-beta 1-induced Runx2 expression. Moreover, TGF-beta 1 opposed BMP-2-induced osteogenic transdifferentiation through Dlx5 suppression by de novo induction of AP-1. Taken together, these results indicate that Dlx5 is an indispensable regulator of BMP-2-induced osteoblast differentiation as well as the counteraction point of the opposing TGF-beta 1 action.

Background: Recombinant human bone morphogenetic protein-2 (rhBMP-2), surgically implanted with a matrix material, has been shown to induce bone formation and enhance fracture repair. The purpose of this investigation was to test the hypothesis that a single, local, percutaneous injection of rhBMP-2 would accelerate fracture-healing in a standard rat femoral fracture model., Methods: Fractures were created, following intramedullary pinning, in the femora of 144 male Sprague-Dawley rats. The animals were divided into three groups of forty-eight each. Six hours after the fracture, one group received an injection of 80 micro g of rhBMP-2 in 25 micro L of buffer vehicle, one received an injection of 25 micro L of buffer vehicle alone, and one did not receive an injection. Twelve animals from each of these three groups were killed at one, two, three, and four weeks after treatment, and the femora were harvested for torsional biomechanical testing. An additional cohort of seventy-two animals, in which a fracture was also created, was divided into the same three treatment groups; six animals from each of these groups was killed at one, two, three, and four weeks; and the femora were processed for qualitative histological analysis., Results: Torsional biomechanical testing indicated that the stiffness of the rhBMP-2-treated fractures was twice that of both control groups at the two, three, and four-week time-points. The strength of the rhBMP-2-treated fractures was 34% greater than that of the buffer-treated controls (p = 0.03) at three weeks and, at four weeks, was 60% and 77% greater than that of the buffer-treated controls and that of the untreated controls, respectively (p < 0.005). At four weeks, the stiffness and strength of the rhBMP-2-treated fractures were equal to those of the intact contralateral femora, whereas the buffer-treated and untreated fractures were significantly weaker than the intact femora. At two and three weeks, large areas of bone formation, typically spanning the fracture, were observed histologically in the rhBMP-2-treated sites. In contrast, the control fractures exhibited primarily soft cartilaginous callus at these time-points. By four weeks, remodeling of the hard callus and recorticalization were observed in the rhBMP-2-treated fracture sites, whereas cartilage and/or soft tissue was still present in the control fracture sites., Conclusions: These data demonstrate that a single, local, percutaneous injection of rhBMP-2 accelerates fracture repair in this rat femoral fracture model. This effect appears to result from a combination of the induction of bone formation at the fracture site and acceleration of the rate at which the fracture callus matures.

Background: Alveolar ridge aberrations commonly compromise optimal dental implant installation. To offset any variance between an aberrant alveolar ridge and prosthetic designs, bone augmentation procedures become necessary. The objective of this study was to evaluate bone formation and osseointegration at alveolar dehiscence defects following augmentation of the defect site with recombinant human bone morphogenetic protein-2 (rhBMP-2) in an absorbable collagen sponge carrier (ACS) at dental implant installation including transmucosal positioning of the dental implant., Methods: Four adult male Cynomolgus monkeys received dental implants in contralateral extraction socket sites with surgically created 6 x 4 mm buccal dehiscence defects following elevation of mucoperiosteal flaps. Contralateral sites received rhBMP-2/ACS (rhBMP-2 at 1.5 mg/ml; 0.1 mg/defect) or served as sham-surgery controls. The flaps were adapted and sutured around the healing abutments leaving the implants in a transmucosal position. The animals were sacrificed at 16 weeks postsurgery and block sections of the implant sites were harvested and prepared for histometric analysis., Results: One dental implant from each treatment group failed to osseointegrate. Another 3 dental implants (sham-surgery controls) failed to osseointegrate with newly-formed bone in the defect area. Thus, 7 of 8 defect sites (4/4 animals) receiving rhBMP-2/ACS compared to 4 of 8 sites (2/4 animals) receiving sham-surgery exhibited evidence of osseointegration with newly formed bone in the defect area. Mean +/- SD defect height amounted to 5.3 +/- 0.2 and 5.4 +/- 0.1 mm for the rhBMP-2/ACS and sham-surgery sites, respectively. Vertical bone gain in rhBMP-2/ACS treated defects (3.9 +/- 0.3 mm) did not differ significantly from that in the sham-surgery control (3.7 +/- 0.4 mm; P > 0.05; paired t-test, N = 4). There were also no significant differences noted for coronal bone-implant contact (3.0 +/- 0.6 versus 3.6 +/- 0.5 mm), and bone-implant contact within the defect site (28.5% +/- 15.1% versus 27.4% +/- 31.7%) and within resident bone (46.9% +/- 26.8% versus 47.8% +/- 39.4%) for the rhBMP-2/ACS and control sites, respectively., Conclusions: The observations in this study point to a substantial native osteogenic potential of the alveolar process that has previously not been explored and show that surgical reentry observations of new bone formation may not necessarily indicate that osseointegration has occurred. Bone formation in control defects was substantially greater than predicted, limiting the value of adding an osteoinductive biologic construct.

Background: Recombinant human bone morphogenetic protein-2 (rhBMP-2) technologies have been shown to significantly support alveolar bone formation. Biomaterial limitations, however, have restricted the biologic potential for onlay indications. The objective of this study was to evaluate regeneration of alveolar bone and periodontal attachment, and biomaterials reaction following surgical implantation of a spaceproviding, bioabsorbable, macroporous, polyglycolic acid-trimethylene carbonate (PGA-TMC) membrane combined with a rhBMP-2 construct in a discriminating onlay defect model., Methods: Routine supraalveolar periodontal defects were created at the mandibular premolar teeth in 9 beagle dogs. Contralateral jaw quadrants in subsequent animals were randomly assigned to receive the domeshaped PGA-TMC (100 to 120 μm pores) membrane with rhBMP-2 (0.2 mg/mL) in a bioresorbable hyaluronan (Hy) carrier or the PGA-TMC membrane with Hy alone (control). The gingival flaps were advanced to submerge the membranes and teeth and sutured. Animals were euthanized at 8 and 24 weeks postsurgery for histologic observations., Results: Jaw quadrants receiving the PGA-TMC membrane alone experienced exposures at various time points throughout the study. Jaw quadrants receiving the PGA-TMC/rhBMP-2 combination remained intact, although one site experienced a late minor exposure. Newly formed alveolar bone approached and became incorporated into the macroporous PGA-TMC membrane in sites receiving rhBMP-2. The PGA-TMC biomaterial was occasionally associated with a limited inflammatory reaction. Residual PGA-TMC could not be observed at 24 weeks postsurgery. Residual Hy could not be observed at any time interval. Regeneration of alveolar bone height (means ± SD) was significantly increased in sites receiving the PGA-TMC/rhBMP 2 combination compared to control (3.8 ± 1.3 versus 0.7 ± 0.5 mm at 8 weeks and 4.6 ± 0.8 versus 2.1 ± 0.4 mm at 24 weeks; P <0.05). Limited cementum regeneration was observed for PGA-TMC/rhBMP-2 and PGA-TMC control sites. Ankylosis compromised regeneration in sites receiving PGA-TMC/rhBMP-2., Conclusions: The bioabsorbable, space-providing, macroporous PGA-TMC membrane appears to be compatible biomaterial for bone augmentation procedures. rhBMP-2 significantly enhances alveolar bone augmentation and soft tissue healing when combined with the PGA-TMC membrane. J Periodontol 2003; 74:635-647., (© 2003 American Academy of Periodontology.)

Background: Surgical implantation of recombinant human bone morphogenetic protein 2 (rhBMP-2) in an absorbable collagen sponge carrier (ACS) significantly enhances bone regeneration in horizontal alveolar defects; however, sufficient quantities of bone for implant dentistry are not routinely obtained., Purpose: The objective of this proof-of-principle study was to evaluate the potential of a space-providing macroporous expanded polytetrafluoroethylene (ePTFE) device to control volume and geometry of rhBMP-2/ACS-induced alveolar bone augmentation., Materials and Methods: Bilateral critical-size supra-alveolar periimplant defects were created in four Hound-Labrador mongrel dogs. Two turned and one surface-etched 10 mm titanium dental implants were placed 5 mm into the surgically reduced alveolar ridge creating 5 mm supra-alveolar defects. rhBMP-2/ACS (0.4 mg rhBMP-2) was placed around the exposed dental implants. Additionally, one jaw quadrant in each animal was randomly assigned to receive the dome-shaped macroporous ePTFE device. Mucoperiosteal flaps were advanced for primary wound closure. The animals were euthanized at 8 weeks post surgery for histometric analysis., Results: The space-providing macroporous ePTFE device defined the volume and geometry of rhBMP-2/ACS-induced bone formation, whereas bone formation at sites receiving rhBMP-2/ACS alone varied considerably. Vertical bone gain at turned dental implants averaged (+/-SD) 4.7 +/-0.2 mm at sites receiving rhBMP-2/ACS and the ePTFE device compared with 3.5 +/-0.9 mm at sites receiving rhBMP-2/ACS only. The corresponding values for rhBMP-2/ACS-induced bone area were 9.6 +/- 0.7 mm2 and 7.5 +/-6.2 mm2. There was a highly significant correlation between induced bone area and the space provided by the ePTFE device (p <.001). There was no difference in induced bone density or bone-implant contact between the two technologies. These observations were consistent with those observed at surface-etched dental implants., Conclusions: The data from this study suggest that a space-providing macroporous ePTFE device defines rhBMP-2/ACS-induced alveolar augmentation to provide adequate bone quantities for implant dentistry. The dental implant surface technology does not appear to substantially influence bone formation.

Background: Recombinant human bone morphogenetic protein-2 (rhBMP-2) in a proper carrier has been shown to induce clinically relevant bone formation for several oral/maxillofacial and periodontal indications and to stimulate regeneration of the periodontal attachment. The objective of this study is to evaluate regeneration of alveolar bone, cementum, periodontal ligament, and associated root resorption and ankylosis following surgical implantation of rhBMP-2 in an absorbable collagen sponge (ACS) or a calcium phosphate putty (alphaBSM) carrier in 3-wall intrabony periodontal defects in the baboon., Methods: rhBMP-2/ACS and rhBMP-2/alphaBSM were implanted in surgically produced, maxillary and mandibular, large size, 3-wall intrabony defects in 4 baboons. Contralateral jaw quadrants were implanted with buffer/ACS, buffer/ alphaBSM, or served as sham-operated surgical controls. Treatments were allocated to left and right, maxillary and mandibular, jaw quadrants following a randomization schedule. Four months following implantation, block biopsies of defect sites were obtained, processed, and subjected to histologic and histometric analysis., Results: Defect sites receiving rhBMP-2/ACS and rhBMP-2/alphaBSM demonstrated significantly greater regeneration than controls. No significant differences were observed between defect sites receiving rhBMP-2/ACS or rhBMP-2/alphaBSM regarding epithelial migration and connective tissue attachment and new bone formation. However, rhBMP-2/ACS supported significantly greater new cementum formation. Ankylosis or root resorption were not observed., Conclusions: The results of this study support the use of rhBMP-2 to enhance periodontal regeneration of intrabony periodontal defects. While this novel technology holds promise, refinement in carrier systems may provide the key to enhancement of the regenerative potential.

Background: Recombinant human bone morphogenetic protein-2 (rhBMP-2) in an absorbable sponge (ACS) carrier is currently being evaluated as candidate therapy for periodontal regeneration. The objective of this study was to characterize, in some detail, tissue reactions following surgical implantation of rhBMP-2/ACS into periodontal defects., Methods: Four young adult, male beagle dogs with surgically induced, bilateral, critical size, supra-alveolar, mandibular premolar defects sequentially received rhBMP-2/ACS (rhBMP-2 at 0.2 mg/ml) in right and left jaw quadrants. After 4 or 8 weeks of healing, experimental teeth with surrounding tissues were harvested and processed for light and transmission electron microscopy., Results: Surgical implantation of rhBMP-2/ACS into large supra-alveolar periodontal defects resulted in a variable tissue response without marked difference between 4- and 8-week observations. New bone, exceeding the volume of the normal alveolar process, had formed within 4 weeks. The regenerated bone tissue consisted of finely trabeculated woven bone. Marrow spaces exhibited a continuous lining of osteoblasts, osteoclasts, and resting cells. The marrow spaces contained numerous large, thin-walled vessels but were almost devoid of collagen fibrils or fibroblasts. Large voids (seromas) encountered in the newly formed bone were free of structured elements except for occasional aggregates of effete erythrocytes. A variety of tissue reactions were observed along the root surface including areas of resorption, areas of hard tissue deposition, and areas without resorptive or appositional activity. Ankylosis was a frequent observation, although areas showing characteristics of a periodontal ligament with a fine layer of acellular fiber cementum and occasional inserting Sharpey's fibers were also observed. Osteoblasts facing the root surface often appeared to be in a highly active state judged by their cuboidal shape, well-developed endoplasmic reticulum and numerous mitochondria, and the presence of an adjacent layer of preosteoblasts. Conspicuous bundles of wide collagen fibrils near the dentin surface as well as within the marrow spaces were considered to represent remnants of the ACS. These fibrils were associated with areas of mineralization as verified by examination of undecalcified specimens., Conclusions: rhBMP-2/ACS elicits a rapid osteoinductive process throughout the implant as well as along and onto the instrumented adjacent root surface. Lamellated trabecular bone was the predominant regenerated tissue. A typical cementum-periodontal ligament-alveolar bone relationship was a rare observation. The great variability in histological tissue response along the instrumented root surface indicates that the stimulus to hard tissue formation resided primarily in the rhBMP-2/ACS implant rather than in the root surface.

Study Design: A literature review was conducted., Objectives: To review the discovery of the bone morphogenetic proteins and describe the bone morphogenetic protein products that will or may be available for clinical use., Summary of Background Data: Bone morphogenetic proteins comprise the osteoinductive component of several tissue engineering products in late-stage development as replacements for autogenous bone graft, and for bone augmentation and repair., Methods: The literature on bone morphogenetic proteins was reviewed., Results: Bone morphogenetic proteins were discovered originally on the basis of their presence in osteoinductive extracts of bone matrix. Molecular cloning of bone morphogenetic proteins demonstrated that they are a family of related differentiation factors, each capable of inducing the formation of new bone tissue when implanted. Two of the molecules in clinical use, recombinant human bone morphogenetic protein-2 and recombinant human bone morphogenetic protein-7 (OP-1) are produced in a biotechnology process using recombinant deoxyribonucleic acid technology that offers unlimited supply and substantial control over purity and reproducible activity. A third material, bovine bone morphogenetic protein extract, is extracted from bone, and contains a mixture of bone morphogenetic protein molecules. Each of these molecules, although osteoinductive in vivo, has different physiologic roles and biologic activities in vivo and in vitro. Successful development of a product for use in spinal fusion involves selecting the osteoinductive molecule, the amount of the bone morphogenetic protein required, and the method of delivery, as well as conducting subsequent preclinical and clinical studies to evaluate its efficacy and safety., Conclusions: On the basis of the data provided in this issue of Spine, some of these bone morphogenetic protein-based products provide for revolutionary therapies in orthopedic practice.