Extent Of Bone Formation Research Articles

Effect of Collagen Plug and Bone Graft on Bone Formation in Rabbit Extraction Model

1. Purpose : This study aimed to investigate the differences in bone regeneration capacity following extraction between a control group without biomaterial placement and experimental groups where biomaterials were inserted (including xenograft, synthetic bone-containing collagen plugs, and collagen plugs).2. Materials and Methods : Bilateral extraction of maxillary first premolars or second premolars was performed, dividing the rabbits into four groups [① Group 1 :no bone graft or collagen plug inserted, ② Group 2: collagen plug inserted after extraction, ③ Group 3: collagen plug containing synthetic bone inserted after extraction, ④ Group 4: xenograft material inserted after extraction]. After euthanized the rabbit, Four and eight weeks postoperatively, the extent of bone formation were evaluated histologically and radiographically.3 Results : The degree of new bone formation using histological analysis showed higher results in both the control groups at weeks 4 and 8, although not statistically significant. Across all groups, higher bone formation was observed at 8 weeks compared to 4 weeks. In radiological analysis using micro CT, Bone Mineral Density (BMD) and Bone Volume/Tissue Volume(BV/TV) were highest in Group 4. Trabecular bone thickness(Tb.Th) was highest in the control group, but not statistically significant.4. Conclusion : In all groups, the degree of new bone formation at 4 and 8 weeks did not show statistically significant differences histologically and radiologically.

The effects of alendronate on the suppression of bone resorption and the promotion of cartilage formation in the human mosaicplasty donor site: A randomized, double-blind, placebo-controlled prospective study

BackgroundWe previously reported that early alendronate administration accelerated bone formation and improved the quality of repaired cartilage in the donor site in rabbits. To investigate whether alendronate administration has effects in humans similar to those observed in rabbits. MethodsThe study cohort included 35 patients over the age of 12-years old who underwent mosaicplasty without osteoporotic therapy from March 2011 to October 2012. The donor sites were medial or lateral in the patellofemoral joint. Placebo (P) or Bonalon containing 35 mg of alendronate (A) was administered orally every week for 8 weeks. The cohort comprised 15 male and 20 female, including 14 right and 21 left knees. The mean age at the time of surgery was 57.1 years. Bone formation was examined using computer tomography and lateral knee radiography, and cartilage formation was examined using magnetic resonance imaging (MRI), second-look assessment, and intraoperative acoustic evaluation. The clinical outcomes were assessed using the Japanese Orthopaedic Association knee score and visual analog scale (VAS). Bone and cartilage formation in the donor site and clinical outcomes were assessed at 3, 6, and 12 months after mosaicplasty. ResultsThe ratio of TRAP-5b in group A was significantly smaller than that in group P at 2 and 8 weeks after mosaicplasty. The extent of bone formation in the donor sites in group A was significantly greater than that in group P at 3 and 6 months after mosaicplasty. Cartilage formation did not differ significantly between the two groups as determined by MRI, macroscopic assessment, and intraoperative acoustic evaluation. Clinical outcomes did not differ significantly between the two groups, and no negative clinical outcomes were observed. ConclusionEarly alendronate administration accelerated bone formation but not cartilage formation in the mosaicplasty donor site in humans.

Partial Bone Formation in Additive Manufactured Porous Implants Reduces Predicted Stress and Danger of Fatigue Failure

New porous implant designs made possible by additive manufacturing allow for increased osseointegration, potentially improving implant performance and longevity for patients that require massive bone implants. The aim of this study was to evaluate how implantation and the strain distribution in the implant affect the pattern of bone ingrowth and how changes in tissue density within the pores alter the stresses in implants. The hypothesis was that porous metal implants are susceptible to fatigue failure, and that this reduces as osteointegration occurs. A phenomenological, finite element analysis (FEA) bone remodelling model was used to predict partial bone formation for two porous (pore sizes of 700 μm and 1500 μm), laser sintered Ti6Al4V implants in an ovine condylar defect model, and was compared and verified against in vivo, histology results. The FEA models predicted partial bone formation within the porous implants, but over-estimated the amount of bone-surface area compared to histology results. The stress and strain in the implant and adjacent tissues were assessed before, during bone remodelling, and at equilibrium. Results showed that partial bone formation improves the stress distribution locally by reducing stress concentrations for both pore sizes, by at least 20%. This improves the long-term fatigue resistance for the larger pore implant, as excessively high stress is reduced to safer levels (86% of fatigue strength) as bone forms. The stress distribution only changed slightly in regions without bone growth. As the extent of bone formation into extensively porous bone implants depends on the level of stress shielding, the design of the implant and stiffness have significant influence on bone integration and need to be considered carefully to ensure the safety of implants with substantial porous regions. To our knowledge this is the first time that the effect of bone formation on stress distribution within a porous implant has been described and characterised.

The Induction of Bone Formation: The Translation Enigma.

A paradigmatic shift in the way of thinking is what bone tissue engineering science requires to decrypt the translation conundrum from animal models into human. The deductive work of Urist (1965), who discerned the principle of bone induction from the pioneering works of Senn, Huggins, Lacroix, Levander, and other bone regenerative scientists, provided the basis that has assisted future bone tissue regenerative scientists to extend the bone tissue engineering field and its potential uses for bone regenerative medicine in humans. However, major challenges remain that are preventing the formation of bone by induction clinically. Growing experimental evidence is indicating that bone inductive studies are non-translatable from animal models into a clinical environment. This is preventing bone tissue engineering from reaching the next phase in development. Countless studies are trying to discern how the formation of bone by induction functions mechanistically, so as to try and solve this enigmatic problem. However, are the correct questions being asked? Why do bone inductive animal studies not translate into humans? Why do bone induction principles not yield the same extent of bone formation as an autogenous bone graft? What are bone tissue engineering scientists missing? By critically re-assessing the past and present discoveries of the bone induction field, this review article attempts to re-discover the field of bone formation by induction, identifying some key features that may have been missed. These include a detailed library of all proteins in bones and their arrangement in the 3D superstructure of the bone together with some other important criteria not considered by tissue engineering scientists. The review therefore not only re-iterates possible avenues of research that need to be re-explored but also seeks to guide present and future scientists in how they assess their own research in light of experimental design and results. By addressing these issues bone formation by induction without autografts might finally become clinically viable.

Do we need retarded delivery of bone growth factors in facial bone repair? An experimental study in rats.

The aim of the present study was to evaluate the effect of different dosages of retarded vs. rapid release of bone morphogenic protein 2 (BMP2) at different recipient sites. Porous composite poly(D,L-lactic acid) (PDLLA)/CaCO3 scaffolds were loaded with three different dosages of rhBMP2 (24 µg, 48 µg and 96 µg) and implanted, together with blank controls, both into non-healing defects of the mandibles and into the gluteal muscles of 24 adult male Wistar rats. After 26 weeks, bone formation and expression of bone specific markers [alkaline phosphatase (AP) and Runx2] were evaluated by histomorphometry and immunohistochemistry. Results showed that the mode of delivery had no quantitative effect on bone formation in mandibular sites. Expression of AP and Runx2 showed significant differences among the three dosage groups. There were significant correlations between the expression of both AP and Runx2 as well as the extent of bone formation, with both retarded and rapid release of rhBMP2. In ectopic sites, retarded release significantly enhanced bone formation in the low and medium dosage groups, compared to rapid release. Expression of AP was significantly higher and Runx2 significantly lower in ectopic sites, compared to mandibular sites. Significant correlations between the expression of bone specific markers and bone formation occurred only in the retarded delivery groups, but not in the rapid release groups. Within the limitations of the experimental model, it was concluded that retarded delivery of BMP2 was effective, preferably in sites with low or non-existing pristine osteogenic activity. Expression of bone specific markers indicated that osteogenic pathways might be different in mandibular vs. ectopic sites.

Glucose Uptake and Runx2 Synergize to Orchestrate Osteoblast Differentiation and Bone Formation

The synthesis of type I collagen, the main component of bone matrix, precedes the expression of Runx2, the earliest determinant of osteoblast differentiation. We hypothesized that the energetic needs of osteoblasts might explain this apparent paradox. We show here that glucose, the main nutrient of osteoblasts, is transported in these cells through Glut1, whose expression precedes that of Runx2. Glucose uptake favors osteoblast differentiation by suppressing the AMPK-dependent proteasomal degradation of Runx2 and promotes bone formation by inhibiting another function of AMPK. While RUNX2 cannot induce osteoblast differentiation when glucose uptake is compromised, raising blood glucose levels restores collagen synthesis in Runx2-null osteoblasts and initiates bone formation in Runx2-deficient embryos. Moreover, RUNX2 favors Glut1 expression, and this feedforward regulation between RUNX2 and Glut1 determines the onset of osteoblast differentiation during development and the extent of bone formation throughout life. These results reveal an unexpected intricacy between bone and glucose metabolism.

Histological Evaluation of Degradable Guided Bone Regeneration Membranes Prepared from Poly(trimethylene carbonate) and Biphasic Calcium Phosphate Composites

SummaryIn oral and maxillofacial surgery, guided bone regeneration using barrier membranes is an important strategy to treat bone defects. The currently used barrier membranes have important disadvantages. Barrier membranes prepared from resorbable poly(trimethylene carbonate) (PTMC) performed as well as collagen barrier membranes. We hypothesized that composite membranes prepared from surface‐eroding PTMC and osteoinductive biphasic calcium phosphate (BCP) would enhance bone formation even further. Bicortical critical size defects in the mandibular angle of rats were covered on both sides with the membranes. After 2, 4, and 12 weeks the extent of bone formation in the defects and the soft tissue reaction towards the membranes was examined histologically. At 2 and 4 weeks, the formation of new bone was observed in defects covered with PTMC, PTMC‐BCP and Biogide collagen membranes. At 12 weeks, bone defects that were covered with PTMC membranes and control Biogide collagen membranes were fully filled with new formed bone. However, at this time point, defects covered with PTMC‐BCP composite membranes had not led to new bone in the defects. Instead a significant tissue reaction, likely to remaining BCP particles, was observed.

Effects of the systemic administration of alendronate on bone formation in a porous hydroxyapatite/collagen composite and resorption by osteoclasts in a bone defect model in rabbits

Several bisphosphonates are now available for the treatment of osteoporosis. Porous hydroxyapatite/collagen (HA/Col) composite is an osteoconductive bone substitute which is resorbed by osteoclasts. The effects of the bisphosphonate alendronate on the formation of bone in porous HA/Col and its resorption by osteoclasts were evaluated using a rabbit model. Porous HA/Col cylinders measuring 6 mm in diameter and 8 mm in length, with a pore size of 100 μm to 500 μm and 95% porosity, were inserted into a defect produced in the lateral femoral condyles of 72 rabbits. The rabbits were divided into four groups based on the protocol of alendronate administration: the control group did not receive any alendronate, the pre group had alendronate treatment for three weeks prior to the implantation of the HA/Col, the post group had alendronate treatment following implantation until euthanasia, and the pre+post group had continuous alendronate treatment from three weeks prior to surgery until euthanasia. All rabbits were injected intravenously with either saline or alendronate (7.5 μg/kg) once a week. Each group had 18 rabbits, six in each group being killed at three, six and 12 weeks post-operatively. Alendronate administration suppressed the resorption of the implants. Additionally, the mineral densities of newly formed bone in the alendronate-treated groups were lower than those in the control group at 12 weeks post-operatively. Interestingly, the number of osteoclasts attached to the implant correlated with the extent of bone formation at three weeks. In conclusion, the systemic administration of alendronate in our rabbit model at a dose-for-weight equivalent to the clinical dose used in the treatment of osteoporosis in Japan affected the mineral density and remodelling of bone tissue in implanted porous HA/Col composites.

Multiscale osteointegration as a new paradigm for the design of calcium phosphate scaffolds for bone regeneration

The role of macropore size (>100 μm) and geometry in synthetic scaffolds for bone regeneration has been studied extensively, but successful translation to the clinic has been slow. Significantly less attention has been given to porosity at the microscale (0.5–10 μm). While some have shown that microporosity in calcium phosphate (CaP)-based scaffolds can improve rate and extent of bone formation in macropores, none has explored microporosity as an additional and important space for bone ingrowth. Here we show osteointegration of biphasic calcium phosphate (BCP) scaffolds at both the macro and micro length scales. Bone, osteoid, and osteogenic cells fill micropores in scaffold rods and osteocytes are embedded in mineralized matrix in micropores, without the addition of growth factors. This work further highlights the importance of considering design parameters at the microscale and demonstrates the possibility for a bone–scaffold composite with no “dead space.” Embedded osteocytes distributed throughout microporous rods may form a mechanosensory network, which would not be possible in scaffolds without microporosity. Multiscale osteointegration has the potential to greatly improve overall performance of these scaffolds through an improvement of mechanical properties, load transfer, and stability in the long and short term, and represents a new paradigm for scaffold design.

Beneficial Role of Periosteum in Distraction Osteogenesis of Mandible: Its Preservation Prevents the External Bone Resorption

Distraction osteogenesis (DO) is a surgical process of new bone generation through the gradual extension of two segments of existing bone. DO is applied for maxillofacial surgeries to manage defects in mandibular continuity. Vertical DO with an oral device is often employed to augment the alveolar bone height for better implant anchorage for esthetic purposes or functional prosthetic requirements. To determine how the periosteum affects the vertical DO in mandibular reconstruction, we extracted the teeth and resected the alveolar parts of the mandible on both sides of dogs, along with removal of the surrounding periosteum in the right, but not left side. Three months later, box-shaped bone segments (vectors) were prepared from the resected alveolar part, and the segments were vertically elongated using a distraction device on both sides at 0.9 mm/day for one week. The extent of bone formation after distraction was determined with micro-focused computed tomography and by measuring incorporation of tetracycline and calcein with confocal laser scanning microscopy. During the initial two months after distraction, new bone formation was observed more prominently in the left side than in the right side of mandible with the periosteum. However, this difference was less clear during the bone-remodeling period. One notable change was the reduced height of the alveolar part of the right-side mandible, a sign of external bone resorption, observed in two out of three dogs at 6-month post-consolidation. These findings suggest that preservation of periosteum prevents the external bone resorption during the vertical DO of mandible.

Repair of osteochondral defects with biodegradable hydrogel composites encapsulating marrow mesenchymal stem cells in a rabbit model

This work investigated the delivery of marrow mesenchymal stem cells (MSCs), with or without the growth factor transforming growth factor-β1 (TGF-β1), from biodegradable hydrogel composites on the repair of osteochondral defects in a rabbit model. Three formulations of oligo(poly(ethylene glycol) fumarate) (OPF) hydrogel composites containing gelatin microparticles (GMPs) and MSCs were implanted in osteochondral defects, including (i) OPF/GMP hydrogel composites; (ii) OPF/GMP hydrogel composites encapsulating MSCs; and (iii) OPF hydrogel composites containing TGF-β1-loaded GMPs and MSCs. At 12weeks, the quality of new tissue formed in chondral and subchondral regions of defects was evaluated based on subjective and quantitative histological analysis. OPF hydrogel composites were partially degraded and the defects were filled with newly formed tissue at 12weeks with no sign of persistent inflammation. With the implantation of scaffolds alone, newly formed chondral tissue had an appearance of hyaline cartilage with zonal organization and intense staining for glycosaminoglycans, while in the subchondral region hypertrophic cartilage with some extent of bone formation was often observed. The addition of MSCs, especially with TGF-β1-loaded GMPs, facilitated subchondral bone formation, as evidenced by more trabecular bone appearance. However, the delivery of MSCs with or without TGF-β1 at the dosage investigated did not improve cartilage morphology. While OPF-based hydrogel composites supported osteochondral tissue generation, further investigations are necessary to elucidate the effects of MSC seeding density and differentiation stage on new tissue formation and regeneration.

RhBMP-2 Modulation of Gene Expression in Infected Segmental Bone Defects

The osteoinductive capability of BMPs appears diminished in the setting of acute infection. We applied rhBMP-2 to a segmental defect in a rat femur and measured the expression of key bone formation genes in the presence of acute infection. Types I and II collagen, osteocalcin, and BMP Type II receptor mRNA expression were characterized in 72 Sprague-Dawley rats, which received either bovine collagen carrier with 200 mug rhBMP-2 plus Staphylococcus aureus, carrier with bacteria only, carrier with rhBMP-2 only, or carrier alone. Six animals from each group were euthanized at 1, 2, and 4 weeks. Total RNA was isolated and extracted, and mRNA was determined by real-time comparative quantitative PCR. Infected defects had little expression of collagen I and II and osteocalcin mRNAs, while BMP receptor II expression with infection was greater than carrier-only controls at weeks 2 and 4. Notably, all four genes were upregulated in infected defects in the presence of rhBMP-2. Thus, in a clinical setting with a high risk of infection and nonunion, such as a compound fracture with bone loss, rhBMP-2 may increase the rate and extent of bone formation. Even if infection does occur, rhBMP-2 may allow a quicker overall recovery time.

Enhanced ectopic bone formation using a combination of plasmid DNA impregnation into 3-D scaffold and bioreactor perfusion culture

The objective of this study is to enhance in vivo ectopic bone formation by combination of plasmid DNA impregnation into three-dimensional (3-D) cell scaffolds and a developed in vitro culture method. Gelatin was cationized by introducing spermine (Sm) to the carboxyl groups for complexation with the plasmid DNA. As the MSC scaffold, collagen sponge reinforced by incorporation of poly(glycolic acid) (PGA) fibers was used. A complex of the cationized gelatin and plasmid DNA of BMP-2 was impregnated into the scaffold. MCS were seeded into each scaffold and cultured by a static and perfusion methods. When MSC were cultured in the PGA-reinforced collagen sponge, the level of BMP-2 expression was significantly enhanced by the perfusion culture compared with static method. When the osteoinduction activity of the PGA-reinforced collagen sponges seeded with PBS, MSC, naked plasmid DNA-BMP-2, cationized gelatin-plasmid DNA-BMP-2 complex, and transfected MSC by static and perfusion method, were studied following the implantation into the back subcutis of rats in terms of histological and biochemical examinations, homogeneous bone formation was histologically observed throughout the sponges seeded with cationized gelatin-plasmid DNA of BMP-2 complex and transfected MSC by perfusion method, although the extent of bone formation was higher for the later one. The level of alkaline phosphatase activity and osteocalcin content at the implanted sites of sponges seeded with transfected MSC by perfusion method were significantly high compared with those seeded with other agents. We conclude that combination of plasmid DNA-impregnated PGA-reinforced collagen sponge and the perfusion method was promising to promote the in vitro gene expression for MSC and in vivo ectopic bone formation.

Enhanced osteoinduction by controlled release of bone morphogenetic protein-2 from biodegradable sponge composed of gelatin and β-tricalcium phosphate

Biodegradable gelatin sponges at different contents of β-tricalcium phosphate ( β-TCP) were fabricated to allow bone morphogenetic protein (BMP)-2 to incorporate into them. The in vivo osteoinduction activity of the sponges incorporating BMP-2 was investigated, while their in vivo profile of BMP-2 release was evaluated. The sponges prepared had an interconnected pore structure with an average pore size of 200 μm, irrespective of the β-TCP content. The in vivo release test revealed that BMP-2 was released in vivo at a similar time profile, irrespective of the β-TCP content. The in vivo time period of BMP-2 retention was longer than 28 days. When the osteoinduction activity of gelatin or gelatin– β-TCP sponges incorporating BMP-2 was studied following the implantation into the back subcutis of rats in terms of histological and biochemical examinations, homogeneous bone formation was histologically observed throughout the sponges, although the extent of bone formation was higher in the sponges with the lower contents of β-TCP. On the other hand, the level of alkaline phosphatase activity and osteocalcin content at the implanted sites of sponges decreased with an increase in the content of β-TCP. The gelatin sponge exhibited significantly higher osteoinduction activity than that of any gelatin– β-TCP sponge, although every sponge with or without β-TCP showed a similar in vivo profile of BMP-2 release. In addition, the in vitro collagenase digestion experiments revealed that the gelatin– β-TCP sponge collapsed easier than the gelatin sponge without β-TCP incorporation. These results suggest that the maintenance of the intrasponge space necessary for the osteoinduction is one factor contributing to the osteoinduction extent of BMP-2-incorporating sponges.

Interbody cages: be careful what you pack

Interbody cages: be careful what you pack

Effects of guided bone regeneration with non-resorbable and bioabsorbable barrier membranes on osseointegration around hydroxyapatite-coated and uncoated threaded titanium dental implants placed into a surgically-created dehiscence type defect in rabbit tibia: A pilot study.

The purpose of this study was to evaluate the effects of guided bone regeneration (GBR) with either a non-resorbable (ePTFE) or bioabsorbable barrier membrane (RSLT) on osseointegration and extent of bone formation around hydroxyapatite-coated (HA) and uncoated threaded titanium (Ti) dental implants placed into surgically-created dehiscence type defects in rabbit tibia. A dehiscence type bone defect, approximately 3 mm in width and height was surgically created on the outer surface of the bone in each tibia of 9 rabbits. For the conventional group, either a HA or a Ti implant was then placed at this site. After the same procedure was performed as in the conventional group, the implant site of the GBR group was covered with either an ePTFE or a RSLT. After 4 months, the rabbits were sacrificed. Specimens were prepared and examined histometrically. It was found that the mean percentage of osseointegration tended to increase in HA compared to Ti implants, both with and without membranes. There was a tendency that the extent of newly regenerated bone was higher in the GBR group than that in the conventional group.

Alveolar bone formation at dental implant dehiscence defects following guided bone regeneration and xenogeneic freeze-dried demineralized bone matrix.

The present study evaluated rate and extent of alveolar bone formation in dental implant dehiscence defects following guided bone regeneration (GBR) and implantation of xenogeneic freeze-dried demineralized bone matrix (xDBM). A total of 16 titanium plasma-sprayed (TPS) and 16 hydroxyapatite-coated (HA) titanium cylinder implants were inserted in 4 mongrel dogs following extraction of the mandibular premolar teeth. Four implant sites per jaw quadrant (2 TPS and 2 HA implant sites) were prepared into extraction sockets in each dog. Buccal alveolar bone was removed to create 3 x 5 mm dehiscence defects. Two jaw quadrants in separate animals received GBR, GBR + xDBM, xDBM (control), or gingival flap surgery alone (GFS; control). Thus, four conditions were available for each implant type (TPS or HA): GBR, GBR + xDBM; xDBM and GFS. The animals received fluorescent bone labels to allow observations of rate and extent of bone formation. Animals were sacrificed at 12 weeks postsurgery and block sections were harvested for histologic analysis. There were no apparent histologic differences between TPS and HA implant defects. GBR and GBR + xDBM resulted in almost complete bone closure of the dental implant dehiscence defect. Rate of bone formation appeared higher following GBR alone. Extent of bone formation appeared somewhat greater following GBR + xDBM; however, delayed. xDBM alone did not adequately resolve the bony defect. In conclusion, GBR results in rapid, clinically relevant bone closure of dental implant dehiscence defects. Adjunctive implantation of xDBM does not appear to significantly improve the healing response in the model used.

Single-Colony Derived Strains of Human Marrow Stromal Fibroblasts Form Bone After Transplantation In Vivo

Populations of marrow stromal fibroblasts (MSFs) can differentiate into functional osteoblasts and form bone in vivo. It is not known, however, what proportion of MSF precursor cells, colony forming units-fibroblast (CFU-Fs), have osteogenic potential. In the present study, analysis of bone formation in vivo by single-colony derived strains of human marrow stromal fibroblasts (HMSFs) has been performed for the first time. Each strain originated from an individual CFU-F and underwent four passages in vitro prior to subcutaneous implantation into immunodeficient mice within vehicles containing hydroxyapatite-tricalcium phosphate ceramic. Multicolony derived HMSF strains were also transplanted to serve as positive controls. After 8 weeks, abundant bone formation was found in the transplants of all multicolony derived HMSF strains, whereas 20 out of 34 (58.8%) single-colony derived strains from four donors formed bone. Immunostaining with antibody directed against human osteonectin and in situ hybridization for human-specific alu sequences demonstrated that cells forming new bone were of human origin and were vital for at least 45 weeks post-transplantation. Both the incidence of bone-forming colonies and the extent of bone formation by single-colony derived HMSF strains were increased by cultivation with dexamethasone and ascorbic acid phosphate. Other factors, including type of transplantation vehicle, morphology, size, and structure of the original HMSF colonies showed no obvious correlation with the incidence or extent of bone formation. Hematopoietic tissue within the newly formed bone was developed in the transplants exhibiting exuberant bone formation. These results provide evidence that individual human CFU-Fs have osteogenic potential and yet differ from each other with respect to their osteogenic capacity.

Promotion of the osteogenetic activity of recombinant human bone morphogenetic protein by prostaglandin E1.

We investigated the promotive effect of prostaglandin (PG) E1 on the osteogenetic activity of bone morphogenetic protein (BMP) using porous chemically synthesized hydroxyapatite ceramic (HAP) pellets as the carrier. After treating the pellets with recombinant human (rh) BMP-2 with or without PGE1, both of which were used at two different concentrations, they were inserted beneath the cranial periosteum of a rabbit. The degree of osteogenesis and osteoconductivity was then examined histopathologically as well as by means of an image-analyzing procedure. Results showed that there was extensive bone formation around the pellets as early as 3 weeks after insertion in the group, which received pellets treated with a relatively large amount of rhBMP alone as well as in the group receiving pellets treated with a small amount of rhBMP combined with PGE1. In the later group, the extent of bone formation was dose-dependent. Subsequent osteogenesis within the pores of the pellets in these groups slowly progressed over time, and by 9 weeks after the insertion, most of the pellet pores had filled with newly generated bone. In addition, the groups which received pellets treated with PGE1 alone also showed osteogenesis as demonstrated by osteoconduction, which was not observed in the group that received the phosphate buffered saline (PBS) treated pellets. The group that received pellets treated with a small amount of rhBMP plus a high concentration of PGE1 exhibited significantly greater bone induction than the group which received the pellets treated with a small amount rhBMP alone, and there was no statistically significant difference between the former group and the group that received a high rhBMP dose. These results clearly indicated that PGE1 has a strong and dose-dependent promotive effect on the osteogenetic activity of rhBMP and that it also promotes osteoconduction, even when used alone. This suggests that enormous reductions can be made in the amount of rhBMP administered when PGE1 is used in conjunction with porous HAP. This should prove to be very advantageous and practical in the clinical application of these materials.

Comparison of the distribution of 3H-alendronate and 3H-Etidronate in rat and mouse bones

Alendronate and etidronate are bisphosphonates used clinically to treat diseases associated with increased bone resorption. Etidronate is less potent and was reported to cause osteomalacia. This study examines if differences in distribution of alendronate and etidronate in the skeleton can explain differences in efficacy and in effects on mineralization between the two drugs. Eight-day old rat pups were injected s.c. with 3H-alendronate or 3H-etidronate both at either 1.3 mumol/kg or at their respective pharmacological effective doses in the growing rat of 0.12 mumol/kg for alendronate and 72.8 mumol/kg for etidronate. Twelve hours after administration at 1.3 mumol/kg both drugs showed a three- to fourfold higher localization on osteoclast vs. osteoblast surface. At the pharmacologically effective doses, 3H-alendronate labeled eightfold more osteoclast surface than osteoblast surface. In contrast, 3H-etidronate labeled approximately equal fractions of osteoclast and osteoblast surface. When similar doses of 3H-etidronate and 3H-alendronate (0.24 mumol/kg 3H-etidronate vs. 0.20 mumol/kg 3H-alendronate; 1.5 mumol/kg 3H-etidronate vs. 1.2 mumol/kg 3H-alendronate; and 14.6 mumol/kg 3H-etidronate vs. 12.0 mumol/kg 3 H-alendronate) were injected intravenously into adult mice at similar specific activities, 3H-etidronate labeled 1.5-2.5 times more osteoclast surface than 3-H-alendronate, but 3 to 15 times more osteoblast surface. Consequently, the ratio between the fraction of labeled osteoclast surface and the fraction of labeled osteoblast surface ranged for 3H-alendronate from 9 to 24, whereas for 3H-etidronate the range was from 4 to 7, due to more extensive labeling of osteoblast surface by 3H-etidronate. In a third experiment, we confirmed in adult mice the previous observation made in rat pups that normal bone formation occurs over alendronate-covered bone surfaces, and found that it occurred over etidronate-covered surfaces as well. Forty nine days after s.c. administration of alendronate at 0.12 mumol/kg or etidronate at 1.3 mumol/kg or 55.3 mumol/kg into adult mice bone formed over drug label. The distance from incorporated label to bone surface for both drugs (12.7 microns for alendronate and 8.7 and 9.2 microns for etidronate) was similar to wall width (defined by cement line) in controls (10.6 microns). In conclusion, alendronate, especially at pharmacologically active doses, shows higher uptake on resorption vs. formation surfaces than etidronate. The extent of bone formation on surfaces containing alendronate or etidronate is similar and is comparable to the "wall width" in controls.