The bone morphogenetic protein family: multifunctional cellular regulators in the embryo and adult

Originally identified as the active components within osteoinductive extracts derived from bone, the BMPs now are known to include a large family of proteins within the TGF-β superfamily of growth and differentiation factors. Members of the BMP family have been determined to be key signaling molecules in embryogenesis, in species ranging from Drosophila to humans. They are involved in delivering positional information, the development of hard tissues (both bones and teeth), as well as soft tissue types. When implanted into adult animals, several of the BMPs have been shown to initiate the complex cellular process resulting in the induction of bone through both the endochondral and intramembranous bone formation pathways. Preclinical studies have shown the ability of these factors to induce bone and heal large bony defects in a variety of models relevant to clinical problems in orthopedics, as well as the oral and maxillofacial dental areas. For example, implantation of recombinant human BMP-2 (rhBMP-2) has been shown to augment the alveolar ridge in several animal models, and when placed in a periodontal environment to restore not only new bone, but also the attachment tissues. Results from ongoing clinical studies support the ability of rhBMP-2 implants to induce physiologic bone.     

Effects of carrier release kinetics on bone morphogenetic protein-2-induced periodontal regeneration in vivo

BACKGROUND: Bone morphogenetic proteins (BMPs) have shown considerable promise as a therapeutic agent to enhance periodontal regeneration although the optimal characteristics of a suitable release system are not known. AIM: The aim of this study was to compare the effects of slow and fast degrading gelatin carriers on BMP-2-induced periodontal healing. METHOD: Recombinant human bone morphogenetic protein-2 (rhBMP-2) was incorporated into gelatin and subsequently differentially cross-linked to produce slow and fast release carrier systems. Release kinetics were confirmed in vitro, by measuring release of 125I-growth hormone from similar gelatin plugs. Effects of BMP were evaluated in surgically created rat periodontal fenestration defects which were processed for histology 10 days post-operatively. The rats were divided into 4 groups and the control defects were treated with either slow or fast degrading gelatin (CONs or CONf respectively), whilst test groups were treated with 1.25 microg rhBMP-2 in the slow or fast degrading gelatin (BMPs or BMPf respectively). RESULTS: BMPf greatly increased bone formation compared with the control (CONf) (1.67 +/- 0.65 versus 0.34 +/- 0.11 x 10(-4) m2), but no significant differences were observed with BMPs and CONs. In contrast, new cementum formation was significantly greater in the BMPs group compared with all other groups (p<0.05). CONCLUSION: Release kinetics of BMP may have important effects on the outcome of BMP-induced periodontal regeneration. New bone formation may be affected by rapid-release kinetics although further investigation is necessary to confirm this. In contrast, new cementum formation is promoted by slow release of BMP.     

骨形成蛋白家族与牵引成骨术

牵引成骨术(DO)中新骨的形成具有胚胎骨的发生和正常骨折愈合的某些特征,骨形成蛋白家族(BMPs)在其中扮演了重要角色。本文就BMPs与DO的生物学联系、作用调节机制、外源性BMPs的应用等方面作一综述。     

Factors that modulate the effects of bone morphogenetic protein-induced periodontal regeneration: a critical review

The healing process initiated by a single molecular species of bone morphogenetic protein (BMP) such as BMP-2 or BMP-7 sets in motion a cascade of cellular events resulting in differentiation of progenitor cells into phenotypes involved in periodontal regeneration. For example, animal studies show that a single dose of recombinant human (rh) BMP-2 increases the rate of normal intramembranous bone formation and enhanced cementum formation during periodontal wound healing. However, the optimal effects of BMPs are modulated by a range of factors that need careful evaluation in clinical studies. These factors include the influence of root conditioning, occlusal loading, BMP dose, and the release characteristics of the carrier as well as the suitability of the model to evaluate the efficacy of BMPs. Each of these factors may affect the rate of BMP-induced osteogenesis and cementogenesis and subsequent periodontal ligament (PDL) formation during the early and late stages of periodontal wound healing. Although BMP-2 initiates stem cells along an osteogenic pathway, the dose may have to be of sufficient concentration to ensure other growth and differentiation factors do not redirect or retard the osteogenic potential of the cell. Understanding when to manipulate the cell's differentiation pathway with the application of single or multiple doses of BMPs at the appropriate concentration is required to optimize the effect of BMPs in periodontal wound healing. Therefore, different release profiles from the same carrier may be particularly important in tissues with mixed cell populations such as in the periodontium, where similar tissues like bone and cementum grow at different rates. Furthermore, treatment of intrabony defects with BMPs are likely to not only require appropriate temporal release of the BMP(s), but also a carrier that can serve as a template for new tissue formation providing space maintenance and supporting the mucoperiosteal flap. Many of these issues have not been adequately addressed from a periodontal standpoint; therefore the purpose of this review is to clarify our current understanding of the factors that are likely to modulate the effects of BMP-induced periodontal regeneration. Moreover, assessing the importance of these factors is essential prior to conducting expensive human clinical trials.     

Recent outcomes and perspectives of the application of bone morphogenetic proteins in implant dentistry

Background: Since the discovery of bone morphogenetic proteins (BMPs), the number of related studies has increased substantially, and more recent outcomes have cast encouraging perspectives on their use in reconstructive surgery. Purpose: The aim of the present review was to summarize the present knowledge about the use of BMPs in conjunction with dental implants based on the literature. Materials and Methods: Scientific articles dealing with the use of growth factors and bone healing with or without dental implants were searched for on MEDLINE and critically scrutinized. Results: Thirty‐nine scientific reports formed the base for the present review. Whereas the osteoinductive capability of BMPs is well documented, studies on their effects in implant dentistry are still incipient. Preclinical and clinical studies did not show outstandingly good outcomes of the application of BMPs compared with conventional treatments or controls. Conclusions: The number of studies in the field of dental implantology in which BMPs have been used is still too small for establishing clinical protocols of their use in order to improve a recipient bone bed prior to implant placement or to enhance the integration process of an implant.     

Effect of a fibrin-fibronectin sealing system as a carrier for recombinant human bone morphogenetic protein-4 on bone formation in rat calvarial defects

BACKGROUND: Bone morphogenetic proteins (BMPs) have been shown to play an important role in bone formation during development and wound healing. Despite there being good prospects for BMP applications, an ideal carrier system for BMPs has yet to be determined. The purpose of this study was to evaluate the possibility of a fibrin-fibronectin sealing system (FFSS) as a carrier for recombinant human BMP-4 (rhBMP-4) and to evaluate the genuine osteoconductive potential of the FFSS in a rat calvarial defect model. METHODS: An 8-mm, calvarial, critical-size osteotomy defect was created in each of 30 male Sprague-Dawley rats. Three groups of 10 animals each received rhBMP-4 (0.025 mg/ml) in the FFSS, FFSS control, or sham-surgery control. The groups were evaluated using histologic and histometric parameters following 2- and 8-week healing intervals (five animals per group per healing interval). RESULTS: Surgical implantation of rhBMP-4/FFSS resulted in enhanced local bone formation at 2 and 8 weeks. New bone formation was also evident in the FFSS control; however, the amount of defect closure, new bone area, and bone density was significantly greater in the rhBMP-4/FFSS group (P < 0.05). At 8 weeks, the quantity of the new bone was greater than that observed at 2 weeks, and the specimens showed a more advanced stage of remodeling and consolidation in both groups (P < 0.05). Only very limited bone formation was observed in the sham-surgery control. CONCLUSION: The results of the present study indicated that the FFSS has osteoconductive potential and may be employed as a carrier for BMPs.     

Effects of bone morphogenetic protein-6 on periodontal wound healing in a fenestration defect of rats

BACKGROUND: Bone morphogenetic proteins (BMPs) may play significant roles in bone formation. The ability of BMP-6 to promote wound healing has been chosen as the subject of this investigation. In this study, a synthetic rat BMP-6 polypeptide was applied to a periodontal fenestration defect in rats to elucidate the effects of BMP-6 on periodontal wound healing. MATERIAL AND METHODS: Following surgery to create a bony window on the buccal aspects of mandibular molar roots, 24 male Sprague Dawley rats were divided into four groups according to BMP application (0, 1, 3 and 10 microg, respectively). Animals were killed after 28 days and the mandible taken for histological examination. Histometric measurements were performed on sections selected from three levels (coronal, middle and apical levels; with 240 microm apart from the central) of the defect. New bone and cementum formation (including area and thickness) were analyzed and compared. RESULTS: In general, minimal new bone was observed on the surgically created defects in the non-BMP group, whereas a complete osseous healing occurred in all BMP-6 treated animals. New bone formation (both in area and thickness) was significantly influenced by both the dosage and the examining level, whereas new cementum formation was affected by dosage only. An increase in bone and cementum formation was noted in all three BMP groups when compared with the control group at all examined levels. Among the BMP groups, greatest new bone and cementum formation were noted in the 3 microg group. New cementum thickness increased on the cementum surfaces of the defects compared with the dentinal surfaces in all study groups. CONCLUSION: An increase in new bone and cementum formation was noted after applying a synthetic BMP-6 polypeptide to a periodontal fenestration defect in rats. Therefore, we suggest that BMP-6 may play a certain role in periodontal regeneration.     

Periodontal regeneration: potential role of bone morphogenetic proteins

Initiation of osteogenesis and cementogenesis is a problem central to periodontal regeneration. A major advance in the understanding of bone formation has been the identification of an entirely new family of protein initiators, the bone morphogenetic proteins, that regulate cartilage and bone differentiation in vivo . The purification, genetic cloning and expression of recombinant human bone morphogenetic proteins (BMPs) have laid the foundation for the cellular and molecular dissection of bone development and regeneration. The striking evolutionary conservation of the BMP genes indicates that they are critical in the normal development and function of animals. In addition to postfetal osteogenesis, the BMPs may play multiple roles in embryonic development and organogenesis, including skeletogenesis and the development of craniofacial and dental tissues. The availability of recombinant human BMPs provides several challenges and opportunities to gain insights into the mechanisms regulating the regeneration of bone and cementum for optimal outcome in the periodontal patient.     

Histomorphometric analysis of rat alveolar wound healing with hydroxyapatite alone or associated to BMPs

Several materials and techniques have been proposed to improve alveolar wound healing and decrease loss of bone height and thickness that normally follow dental extraction. The objective of this research was the histologic analysis of bone morphogenetic proteins implanted into dental alveoli of rats after extraction. A total of 45 adult male Wistar rats were divided into three groups of 15 animals each: control (no treatment), implanted with pure hydroxyapatite (HA, 3 mg) and implanted with hydroxyapatite plus bone morphogenetic proteins (HA/BMPs, 3 mg). Five animals from each group were sacrificed at 7, 21 and 42 days after extraction for the histometric analyses of the osteoconductive potential of hydroxyapatite associated or not with BMPs. After dissection, fixation, decalcification and serial microtomy of 6-micron thick sections, the samples were stained with hematoxylin-eosin for histologic and histometric analyses. Both HA and HA/BMPs caused a delay in wound healing compared to control animals, evaluated by the percentage of bone tissue in the alveoli. The treatment with HA/BMPs had the greatest delay at 21 days, even though it produced values similar to the control group at 42 days. The materials did not improve alveolar repair in the normal period of wound healing and the association of HA/BMPs did not have osteoconductive properties with granulated hydroxyapatite as the vehicle.     

Identification of bone morphogenetic proteins 2 and 4 in commercial demineralized freeze-dried bone allograft preparations: pilot study

Background: Demineralized freeze‐dried bone allografts (DFDBAs) have been proposed as a useful adjunct in periodontal therapy to induce periodontal regeneration through the induction of new bone formation. The presence of bone morphogenetic proteins (BMPs) within the demineralized matrix has been proposed as a possible mechanism through which DFDBA may exert its biologic effect. However, in recent years, the predicability of results using DFDBA has been variable and has led to its use being questioned. One reason for the variability in tissue response may be attributed to differences in the processing of DFDBA, which may lead to loss of activity of any bioactive substances within the DFDBA matrix. Therefore, the purpose of this investigation was to determine whether there are detectable levels of bone morphogenetic proteins in commercial DFDBA preparations. Methods: A single preparation of DFDBA was obtained from three commercial sources. Each preparation was studied in triplicate. Proteins within the DFDBA samples were first extracted with 4M guanidinium HCI for seven days at 40 degrees Celsius and the residue was further extracted with 4M guanidinium HCL/EDTA for seven days at 40 degrees celsius. Two anti‐human BMP‐2 and ‐4 antibodies were used for the detection of the presence of BMP's in the extracts. Results: Neither BMP‐2 nor BMP‐4 was detected in any of the extracts. When recombinant human BMP‐2 and ‐4 were added throughout the extraction process of DFDBA extraction, not only were intact proteins detected but smaller molecular weight fragments were also noted in the extract. Conclusions: These results indicate that all of the DFDBA samples tested had no detectable amounts of BMP‐2 and ‐4. In addition, an unknown substance present in the DFDBA may be responsible for degradation of whatever BMPs might be present.     

Periodontal tissue regeneration by combined applications of recombinant human osteogenic protein-1 and bone morphogenetic protein-2. A pilot study in Chacma baboons (Papio ursinus)

Native and recombinant human bone morphogenetic/osteogenic proteins (BMPs/ OPs) singly initiate bone induction in vivo. The finding of synchronous but spatially different BMPs/OPs expression during periodontal tissue morphogenesis suggests novel therapeutic approaches using morphogen combinations based on recapitulation of embryonic development. Twelve furcation defects prepared in the first and second mandibular molars of three adult baboons (Papio ursinus) were used to assess whether qualitative histological aspects of periodontal tissue regeneration could be enhanced and tissue morphogenesis modified by combined or single applications of recombinant hOP-1 and hBMP-2. Doses of BMPs/OPs were 100 microg of each protein per 1 g of insoluble collagenous bone matrix as carrier. Approximately 200 mg of carrier matrix was used per furcation defect. Undecalcified sections cut for histological analysis 60 d after healing of hOP-1-treated specimens showed substantial cementogenesis with scattered remnants of the collagenous carrier. hBMP-2 applied alone induced greater amounts of mineralized bone and osteoid when compared to hOP-1 alone or to combined morphogen applications. Combined applications of hOP-1 and hBMP-2 did not enhance alveolar bone regeneration or new attachment formation over and above the single applications of the morphogens. The results of this study, which is the first to attempt to address the structure-activity relationship amongst BMP/OP family members, indicate that tissue morphogenesis induced by hOP-1 and hBMP-2 is qualitatively different when the morphogens are applied singly, with hOP-1 inducing substantial cementogenesis. hBMP-2 treated defects, on the other hand, showed limited cementum formation but a temporal enhancement of alveolar bone regeneration and remodelling. The demonstration of therapeutic mosaicism in periodontal regeneration will require extensive testing of ratios and doses of recombinant morphogen combinations for optimal tissue engineering in clinical contexts.     

Effects of alendronate on bone healing after tooth extraction in rats

Oral Diseases (2010) 16 , 674–685 Objectives: Tooth extraction has been identified as an important risk factor for bisphosphonate‐induced osteonecrosis of the jaw. Therefore, the main goal of this study was to determine the effects of alendronate on healing of the extraction socket and on interdental alveolar bone after tooth extraction in rats. Materials and methods: Animals were injected subcutaneously with vehicle or alendronate for 3–4 weeks before the first mandibular molar was extracted and these treatments were continued during post‐extraction periods of 10, 21, 35 and 70 days. Mandibles were processed to evaluate healing of the extraction socket and adjacent alveolar bone by assessing bone formation, bone resorption and vascularity by histomorphometric techniques. Results: Alendronate decreased new woven bone formation, blood vessel area, perimeter and number in the extraction socket at 10 days postextraction, but not at later time points. Furthermore, alendronate‐treated rats had increased interdental alveolar bone volume and height only at 10 days postextraction. In addition, a 2.5‐fold increase in the percentage of empty osteocyte lacunae was found in alveolar bone of alendronate‐treated rats only at 10 days postextraction. Conclusions: Alendronate transiently decreases bone formation and vascularity in the extraction socket and delays the removal of interdental alveolar bone after tooth extraction in rats.     

Alveolar wound healing after implantation with a pool of commercially available bovine bone morphogenetic proteins (BMPs): a histometric study in rats

The capacity of a commercially available pool of bovine bone morphogenetic proteins (BMPs) to stimulate osteogenesis in the rat alveolar healing was investigated by histometric analysis. Male rats were anesthetized and had their upper incisor extracted. A pool of purified bovine BMPs adsorbed to microgranular resorbable hydroxyapatite was agglutinated with bovine collagen and saline before implantation into the alveolar socket. The implanted and control rats (n=30 per group) were sacrificed 1 to 9 weeks postoperatively, the hemi-maxillae were decalcified, processed for paraffin embedding and semi-serial longitudinal sections were obtained and stained with hematoxylin and eosin. The volume fraction of alveolar healing components was estimated by a differential point-counting method in histologic images. The results showed that in both, control and implanted rats, the alveolar healing followed the histologic pattern usually described in the literature. Quantitative data confirmed that the BMPs mixture did not stimulate new bone formation in the alveolar socket of implanted rats. These results suggest that the pool of BMPs adsorbed to hydroxyapatite and agglutinated with bovine collagen did not warrant incorporation of the osteoinductive proteins to a slow-absorption system that would allow a BMPs release rate compatible to that of new bone formation, and thus more adequate to osteoinduction.     

A review of mechanisms by which low-intensity pulsed ultrasound affects bone regeneration

低强度脉冲超声（LIPUS）是临床常见的物理治疗手段之一,可用于促进骨折愈合以及治疗陈旧性骨不连。局部血管、神经和骨组织密切相关、互相影响,是骨组织再生的重要影响因素。近年来越来越多的数据表明LIPUS不仅能作用于成骨细胞、破骨细胞、间充质干细胞等发挥促成骨效应,还可通过其对血管、神经的作用对骨组织愈合与再生产生一定的积极影响。本文从LIPUS对骨组织的直接作用和LIPUS通过促进血管及神经再生对骨组织的间接作用2个方面,就LIPUS在骨组织再生方面相关分子机制的最新研究进展作一综述,为LIPUS治疗骨相关疾病提供新的思路。     

Bone growth factors in maxillofacial skeletal reconstruction

A literature review was performed to survey the available information on the potential of bone growth factors in skeletal reconstruction in the maxillofacial area. The aim of this review was to characterize the biological and developmental nature of the growth factors considered, their molecular level of activity and their osteogenic potential in craniofacial bone repair and reconstruction. A total of 231 references were selected for evaluation by the content of the abstracts. All growth factors considered have a fundamental role in growth and development. In postnatal skeletal regeneration, PDGF plays an important role in inducing proliferation of undifferentiated mesenchymal cells. It is an important mediator for bone healing and remodelling during trauma and infection. It can enhance bone regeneration in conjunction with other growth factors but is unlikely to provide entirely osteogenic properties itself. IGFs have an important role in general growth and maintenance of the body skeleton. The effect of local application of IGFs alone in craniofacial skeletal defects has not yet shown a clear potential for enhancement of bone regeneration in the reported dosages. The combination of IGF-I with PDGF has been effective in promoting bone regeneration in dentoalveolar defects around implants or after periodontal bone loss. TGFbeta alone in skeletal reconstruction appears to be associated with uncertain results. The presence of committed cells is required for enhancement of bone formation by TGFbeta. It has a biphasic effect, which suppresses proliferation and osteoblastic differentiation at high concentrations. BMPs, BMP2, BMP4 and BMP7 in particular, appear to be the most effective growth factors in terms of osteogenesis and osseous defect repair. Efficacy of BMPs for defect repair is strongly dependent on the type of carrier and has been subject to unknown factors in clinical feasibility trials resulting in ambiguous results. The current lack of clinical data may prolong the period until this factor is introduced into routine clinical application. PRP is supposed to increase proliferation of undifferentiated mesenchymal cells and to enhance angiogenesis. There is little scientific evidence about the benefit of PRP in skeletal reconstructive and preprosthetic surgery yet and it is unlikely that peri-implant bone healing or regeneration of local bone into alloplastic material by the application of PRP alone will be significantly enhanced.     

The Influence of PRP on Early Bone Formation in Membrane Protected Defects. A Histological and Histomorphometric Study in the Rabbit Calvaria

Background: Platelet rich plasma (PRP) has been proposed to be a useful adjunct to bone grafting. Purpose: The aim of the present study was to assess new bone formation in bone regeneration procedures using platelet rich plasma (PRP) alone or in combination with autogenous bone. Materials and Methods: Four surgically created, monocortical defects 5 mm in diameter in the calvariae of 15 New Zealand rabbits were grafted with a coagulum‐filled control, PRP, particulated autogenous bone alone (A), or combined with PRP (A‐PRP). Results: Mean platelet concentration of 1,761,930 ± 680,200/µl was achieved (5.30 ± 2.63 × fold of baseline). Animals were sacrificed 1, 2, and 4 weeks later. Histomorphometric analysis showed no statistical difference for total new bone formation at any time point, however, a detailed analysis revealed a statistically significant higher percentage of lamellar bone than woven bone for the autogenous bone group at 2 weeks; all other groups demonstrated equal percentages of either bone type. At 4 weeks, all groups revealed a statistically greater component of lamellar bone over woven bone. Graft resorption rate was similar for both A and A‐PRP. PRP platelet concentration was significantly positively correlated with TGF‐beta1 but not with PDGF‐AB. Conclusions: Within the limits of the chosen animal model, this study demonstrated that PRP during early healing, whether alone or mixed with autogenous bone, did not lead to greater bone remodelling, as compared to coagulum. In contrast, autogenous bone alone demonstrated accelerated bone remodelling at 2 weeks.     

An evaluation of hydroxyapatite and biphasic calcium phosphate in combination with Pluronic F127 and BMP on bone repair

Calcium phosphates like hydroxyapatite (HA), beta-tricalcium phosphate (beta-TCP), and their mixtures (biphasic calcium phosphates; BCP) are used clinically to repair bone defects. These materials can be difficult to handle and have no inherent biological activity. Handling properties of other bone substitute materials have been improved by combining them with an inert carrier such as Pluronic F-127 (Pluronic, BASF, Mt. Olive, NJ), while the addition of bone morphogenetic proteins (BMP) with such implants has also been shown to enhance bone repair. This study assessed the impact of adding Pluronic and BMPs to an HA (C-Graft) or a BCP (80/20 HA/beta-TCP ratio; Algisorb) implant's ability of promote bony repair in the rabbit calvarial defect model.Twenty-five New Zealand white rabbits were divided into 5 groups of 5 animals each. Bilateral calvarial defects were made in the parietal bones of each animal. HA or BCP alone or combined with Pluronic and/or BMP were implanted into the defect sites. Animals were euthanized at 6 weeks, postoperatively. Bone regeneration was evaluated quantitatively by histomorphometry.The amount of bone regeneration, which occurred in defects containing HA and BCP, was similar over the time period studied. Incorporating Pluronic increased handling and moldability without compromising osteoconductivity of either calcium phosphate. The addition of BMP significantly increased the amount of new bone formed with all calcium phosphates studied (P < 0.05). These results suggest that Pluronic can be added to calcium phosphates to enhance handling and moldability without any negative effects on their biocompatibility and that healing can be enhanced with the incorporation of BMPs.