Absorbable Collagen Sponge Research Articles

Mesenchymal Stem Cell Exosomes Enhance Posterolateral Spinal Fusion in a Rat Model.

Spinal fusion, a common surgery performed for degenerative lumbar conditions, often uses recombinant human bone morphogenetic protein 2 (rhBMP-2) that is associated with adverse effects. Mesenchymal stromal/stem cells (MSCs) and their extracellular vesicles (EVs), particularly exosomes, have demonstrated efficacy in bone and cartilage repair. However, the efficacy of MSC exosomes in spinal fusion remains to be ascertained. This study investigates the fusion efficacy of MSC exosomes delivered via an absorbable collagen sponge packed in a poly Ɛ-caprolactone tricalcium phosphate (PCL-TCP) scaffold in a rat posterolateral spinal fusion model. Herein, it is shown that a single implantation of exosome-supplemented collagen sponge packed in PCL-TCP scaffold enhanced spinal fusion and improved mechanical stability by inducing bone formation and bridging between the transverse processes, as evidenced by significant improvements in fusion score and rate, bone structural parameters, histology, stiffness, and range of motion. This study demonstrates for the first time that MSC exosomes promote bone formation to enhance spinal fusion and mechanical stability in a rat model, supporting its translational potential for application in spinal fusion.

A Mechanistic and Preclinical Assessment of BioRestore Bioactive Glass as a Synthetic Bone Graft Extender and Substitute for Osteoinduction and Spine Fusion.

Preclinical animal study. Evaluate the osteoinductivity and bone regenerative capacity of BioRestore bioactive glass. BioRestore is a Food and Drug Administration (FDA)-approved bone void filler that has not yet been evaluated as a bone graft extender or substitute for spine fusion. In vitro and in vivo methods were used to compare BioRestore with other biomaterials for the capacity to promote osteodifferentiation and spinal fusion. The materials evaluated (1) absorbable collagen sponge (ACS), (2) allograft, (3) BioRestore, (4) Human Demineralized Bone Matrix (DBM), and (5) MasterGraft. For in vitro studies, rat bone marrow-derived stem cells (BMSC) were cultured on the materials in either standard or osteogenic media (SM, OM), followed by quantification of osteogenic marker genes ( Runx2, Osx, Alpl, Bglap, Spp1 ) and alkaline phosphatase (ALP) activity. Sixty female Fischer rats underwent L4-5 posterolateral fusion (PLF) with placement of 1 of 5 implants: (1) ICBG from syngeneic rats; (2) ICBG+BioRestore; (3) BioRestore alone; (4) ICBG+Allograft; or (5) ICBG+MasterGraft. Spines were harvested 8 weeks postoperatively and evaluated for bone formation and fusion via radiography, blinded manual palpation, microCT, and histology. After culture for 1 week, BioRestore promoted similar expression levels of Runx2 and Osx to cells grown on DBM. At the 2-week timepoint, the relative ALP activity for BioRestore-OM was significantly higher ( P <0.001) than that of ACS-OM and DBM-OM ( P <0.01) and statistically equivalent to cells grown on allograft-OM. In vivo, radiographic and microCT evaluation showed some degree of bridging bone formation in all groups tested, with the exception of BioRestore alone, which did not produce successful fusions. This study demonstrates the capacity of BioRestore to promote osteoinductivity in vitro. In vivo, BioRestore performed similarly to commercially available bone graft extender materials but was incapable of producing fusion as a bone graft substitute. Level V.

CALCIUM SULFATE/HYDROXYAPATITE-MEDIATED CONTROLLED CO-DELIVERY OF BMP-2 AND ZOLEDRONIC ACID ENHANCES SPINAL FUSION

Although bone morphogenetic protein 2 (BMP-2) has been FDA-approved for spinal fusion for decades, its disadvantages of promoting osteoclast-based bone resorption and suboptimal carrier (absorbable collagen sponge) leading to premature release of the protein limit its clinical applications. Our recent study showed an excellent effect on bone regeneration when BMP-2 and zoledronic acid (ZA) were co-delivered based on a calcium sulphate/hydroxyapatite (CaS/HA) scaffold in a rat critical-size femoral defect model. Therefore, the aim of this study was to evaluate whether local application of BMP-2 and ZA released from a CaS/HA scaffold is favorable for spinal fusion. We hypothesized that CaS/HA mediated controlled co-delivery of rhBMP-2 and ZA could show an improved effect in spinal fusion over BMP-2 alone. 120, 8-week-old male Wistar rats (protocol no. 25-5131/474/38) were randomly divided into six groups in this study (CaS/HA, CaS/HA + BMP-2, CaS/HA + systemic ZA, CaS/HA + local ZA, CaS/HA + BMP-2 + systemic ZA, CaS/HA + BMP-2 + local ZA). A posterolateral spinal fusion at L4 to L5 was performed bilaterally by implanting group-dependent scaffolds. At 3 weeks and 6 weeks, 10 animals per group were euthanized for µCT, histological staining, or mechanical testing. µCT and histological results showed that the CaS/HA + BMP-2 + local ZA group significantly promoted bone regeneration than other treated groups. Biomechanical testing showed breaking force in CaS/HA + BMP + local ZA group was significantly higher than other groups at 6 weeks. In conclusion, the CaS/HA-based biomaterial functionalized with bioactive molecules rhBMP-2 and ZA enhanced bone formation and concomitant spinal fusion outcomeAcknowledgements: Many thanks to Ulrike Heide, Anna-Maria Placht (assistance with surgeries) as well as Suzanne Manthey &amp; Annett Wenke (histology).

Posterior open wound healing in immediate implant placement using reactive soft tissue versus absorbable collagen sponge: a retrospective cohort study

The soft and hard tissue healing of open wounds in immediate implant placement are yet to be explored. The aim of this study was to compare the clinical outcomes of open wound healing using reactive soft tissue (RST) and absorbable collagen sponge (ACS). Forty implants placed immediately in posterior sockets were included; autologous RST was used in 20 and ACS substitute was used in 20. Soft tissue healing was primarily assessed through a novel scoring system and the evaluation of gingival recession. The horizontal bone width (HBW) and interproximal marginal bone level (MBL) were measured on radiographs to observe the hard tissue healing. No significant difference in total soft tissue healing score was observed at 2 weeks postoperatively. Notably, the ACS group showed better tissue colour (P = 0.016) but worse fibrous repair (P = 0.043) scores than the RST group. Gingival recession levels were comparable in the two groups, both before tooth extraction and after placement of the restoration. Regarding hard tissue, HBW and MBL changes showed no intergroup differences. Within the limitations of this study, both RST and ACS seemed effective for open wound closure, achieving ideal soft and hard tissue healing in immediate implant placement.

Application of Recombinant Human Bone Morphogenetic Protein-2 (RhBMP-2) in the Reconstruction of Edentulous Posterior Maxilla: Clinical Protocol, Histological Analysis, and Long-Term Implant Success and Survival Rates

Background: This study evaluated the efficacy of recombinant human bone morphogenetic protein-2 (rhBMP-2) in inducing de novo bone formation during the augmentation of the severely resorbed posterior maxilla for dental implant placement. Methods: Composite grafts consisting of rhBMP-2/absorbable collagen sponge (ACS) mixed with deproteinized bovine bone mineral (DBBM) Bio-Oss in a 1:1 ratio were used to reconstruct severely resorbed posterior maxillae in 13 patients (five men, eight women; age range: 50 to 66 years). Three patients were treated with sinus floor augmentation alone, and 10 patients with a combination of sinus floor augmentation and guided bone regeneration. A total of 71 dental implants were used in this study. The residual bone height below the maxillary sinus ranged from 0.5 to 3.0 mm. Clinical, radiological, and histomorphometric analyses were performed to assess the outcomes of bone augmentation. Results: The healing period was uneventful in all patients. The mean alveolar ridge height increased by 14 mm (range, 10–18 mm) and the mean alveolar width increased by 6 mm (range, 4–7 mm). Histopathological analysis revealed that newly formed bone was detectable throughout the implantation sites of sinus biopsies. The analysis of vascularization of the implant bed revealed large numbers of high-lumen vessels, in addition to moderate numbers of smaller blood vessels within the connective tissue. The histomorphometrical analysis of the tissue distribution showed that the amount of newly formed bone was 20.39 ± 4.95%, the amount of the remaining bone substitute was 41.85 ± 11.97%, and connective tissue was 37.76 ± 8.82%. The overall dental implant success and survival rates were 100% and remained unchanged at a follow-up period of 3 to 12 years. Conclusions: The results of this study demonstrated that a composite graft of rhBMP-2/ACS and DBBM can result in predictable reconstruction of a large bone volume of the maxilla for dental implant placement and functional loading.

RhBMP6 in autologous blood coagulum is a preferred osteoinductive device to rhBMP2 on bovine collagen sponge in the rat ectopic bone formation assay

Osteoinductive BMPs require a suitable delivery system for treating various pathological conditions of the spine and segmental bone defects. INFUSE, the only commercially available BMP-based osteoinductive device, consisting of rhBMP2 on bovine absorbable collagen sponge (ACS) showed major disadvantages due to serious side effects. A novel osteoinductive device, OSTEOGROW, comprised of rhBMP6 dispersed within autologous blood coagulum (ABC) is a promising therapy for bone regeneration, subjected to several clinical trials for diaphysial bone repair and spinal fusion. In the present study, we have examined the release dynamics showing that the ABC carrier provided a slower, more steady BMP release in comparison to the ACS. Rat subcutaneous assay was employed to evaluate cellular events and the time course of ectopic osteogenesis. The host cellular response to osteoinductive implants was evaluated by flow cytometry, while dynamics of bone formation and maintenance in time were evaluated by histology, immunohistochemistry and micro CT analyses. Flow cytometry revealed that the recruitment of lymphoid cell populations was significantly higher in rhBMP6/ABC implants, while rhBMP2/ACS implants recruited more myeloid populations. Furthermore, rhBMP6/ABC implants more efficiently attracted early and committed progenitor cells. Dynamics of bone formation induced by rhBMP2/ACS was characterized by a delayed endochondral ossification process in comparison to rhBMP6/ABC implants. Besides, rhBMP6/ABC implants induced more ectopic bone volume in all observed time points in comparison to rhBMP2/ACS implants. These results indicate that OSTEOGROW was superior to INFUSE due to ABC's advantages as a carrier and rhBMP6 superior efficacy in inducing bone.

Effects of rhBMP-2 with various carriers on maxillofacial bone regeneration through computed tomography evaluation

BackgroundrhBMP-2 is regarded as the most potent osteoinductive growth factor, and it has been used in the oral cavity with different carriers. The purpose of this study is to evaluate the bone-regenerative effect of rhBMP-2 delivered with different carrier systems through three-dimensional cone beam computed tomography analysis.MethodA total of 112 patients underwent oral surgery with rhBMP-2 application (Group 1, n = 53) or without rhBMP-2 application (Group 2, n = 59). Group 1 was divided into 3 groups according to carriers, rhBMP-2 with allograft (Group 1–1, n = 34), rhBMP-2 with xenograft (Group 1–2, n = 5), and rhBMP-2 with absorbable collagen sponge (Group 1–3, n = 14). Cone beam computed tomography scans were taken before surgery (T0) 6 months after surgery (T1). The volume of defects was measured through the three-dimensional image analysis tool.ResultsThe average bone regeneration rate of Group 1 was significantly greater than that of Group 2. Within Group 1, the group that used allograft as a carrier (Group 1–1) showed significantly higher bone regeneration rates than the group that used absorbable collagen sponge as a carrier (Group 1–3).ConclusionThe use of rhBMP-2 after oral surgery results in a superior bone regeneration rate compared to not using rhBMP-2, and its efficacy depends on the carriers it is used with. Allograft affects bone regeneration more than absorbable collagen sponge when it is carried with rhBMP-2. Therefore, the appropriate use of rhBMP-2 with suitable bone grafting materials is useful for promoting postoperative bone regeneration in oral surgery.

Recombinant Human Bone Morphogenetic Protein-2 as Bone Additive and its Relation with the Dental Implant Dimensions and Stability: Split-Mouth Randomized Clinical Trial

Background: The bone morphogenetic protein belongs to transforming growth factor beta (TGF-β), and it is regarded as one of the biological factors that play major roles in the process of osteogenesis. Objective: To measure the effect of recombinant human bone morphogenetic protein-2 with an absorbable collagen sponge carrier (rhBMP-2/ACS) on secondary implant stability and study the relation of other parameters like implant receipt jaws, implant diameter, and implant length with implant stability. Methods: Ten participants were enrolled in the study after a selective diagnosis. Forty-seven implants were classified into two groups: the study group included twenty-three dental implants with an additive of rhBMP-2/ACS, and the control group included twenty-four dental implants without an additive. Each patient received at least two implants on each ipsilateral side. The primary implant stability was measured at the surgical phase, and the secondary implant stability was recorded after 16–24 weeks by using the Resonance Frequency Analysis device. Results: There was a weak but not significant correlation between implant dimensions and stability, except in the control group. Concerning the ISQ relation to the jaw, the mandible showed a significant increase in primary stability for the study group compared to that in the control group, but no statistical relation was recorded. Conclusion: The mandible had higher ISO values for primary stability than the maxilla in both groups, with a non-significant connection. The maxilla in both groups had improved secondary stability, whereas the mandible had decreased. (clinicaltrials.gov-NCT05719181).

Pilot Study on Percutaneous Delivery of Recombinant Human Bone Morphongenetic Protein-2 Augments Fusion in a Nicotine-impaired Rabbit Fusion Model.

A nicotine-impaired spinal fusion rabbit model. To examine whether controlled delivery of morselized absorbable collagen sponge recombinant human bone morphogenetic protein-2 (rhBMP2) in a delayed manner postsurgery would allow for improved bone healing. The current delivery method of rhBMP-2 during surgery causes a burst of rhBMP-2, which is not sustained. Given that bone morphogenetic protein 2 (BMP-2) expression peaks later in the fusion process, there may be the benefit of delivery of rhBMP-2 later in the healing process. Sixteen male 1-year-old rabbits underwent a posterolateral spinal fusion with iliac crest bone graft at L5-L6 while being given nicotine to prevent spinal fusion as previously published. Eight were controls, whereas 8 had morselized rhBMP-2 (4.2 mg) injected at the fusion site at 4 weeks postoperatively. Histologic, radiologic, and palpation examinations were performed at 12 weeks to determine fusion status and the volume of bone formed. Hematoxylin and eosin stains were used for histology. A Student t test was used to compare the computed tomography scan measured volume of bone created between the control cohort (CC) and rhBMP-2 delayed delivery cohort (BMP-DDC). Of the total, 7/8 rabbits in the BMP-DDC and 5/8 rabbits in the CC formed definitive fusion with a positive palpation examination, bridging bone between transverse processes on computed tomography scan, and an x-ray showing fusion. Histologic analysis revealed newly remodeled bone within the BMP-DDC. There was an increased average volume of bone formed within the BMP-DDC versus the CC (22.6 ± 13.1 vs 11.1 ± 3.6 cm 3 , P = 0.04). Our study shows that injectable morselized absorbable collagen sponge/rhBMP-2 can create twice as much bone within a nicotine-impaired rabbit spine fusion model when delivered 4 weeks out from the time of surgery.

Systemic Diclofenac Sodium Reduces Postoperative rhBMP-2 Induced Neuroinflammation: A Rodent Model Study.

This is a basic science, animal research study. This study aims to explore, in rodent models, the effectiveness of systemic nonsteroidal anti-inflammatory drugs in reducing recombinant human bone morphogenetic protein-2 (rhBMP-2) induced neuroinflammation. rhBMP-2 is increasingly used to augment fusion in lumbar interbody fusion surgeries, although it can cause complications including postoperative radiculitis. Eighteen 8-week-old Sprague-Dawley rats underwent Hargreaves testing to measure the baseline thermal withdrawal threshold before undergoing surgical intervention. The L5 nerve root was exposed and wrapped with an Absorbable Collagen Sponge containing rhBMP-2. Rats were randomized into 3 groups: (1) Low dose (LD), (2) high dose (HD) diclofenac sodium, and (3) saline, receiving daily injection treatment. Hargreaves testing was performed postoperatively on days 5 and 7. Seroma volumes were measured by aspiration and the nerve root was then harvested for hematoxylin and eosin, immunohistochemistry, Luxol Fast Blue staining, and real-time quantitative polymerase chain reaction. The Student t test was used to evaluate the statistical significance among groups. The intervention groups showed reduced seroma volume, and a general reduction of inflammatory markers (MMP12, MAPK6, GFAP, CD68, and IL18) compared with controls, with the reduction in MMP12 being statistically significant ( P = 0.02). Hematoxylin and eosin and immunohistochemistry of the nerve roots showed the highest macrophage density in the saline controls and the lowest in the HD group. Luxol Fast Blue staining showed the greatest extent of demyelination in the LD and saline groups. Lastly, Hargreaves testing, a functional measure of neuroinflammation, of the HD group demonstrated a minimal change in thermal withdrawal latency. In contrast, the thermal withdrawal latency of the LD and saline groups showed a statistically significant decrease of 35.2% and 28.0%, respectively ( P < 0.05). This is the first proof-of-concept study indicating that diclofenac sodium is effective in alleviating rhBMP-2-induced neuroinflammation. This can potentially impact the clinical management of rhBMP-2-induced radiculitis. It also presents a viable rodent model for evaluating the effectiveness of analgesics in reducing rhBMP-2-induced inflammation.

Can a low dosage of recombinant human bone morphogenetic protein-2 loaded on collagen sponge induce ectopic bone?

Recombinant human bone morphogenetic protein-2 (rhBMP-2) is one of the growth factors that may induce the formation of new bone. The aim was to determine the efficacy of low doses of rhBMP-2 for bone regeneration using a collagen sponge as a carrier. Three doses of rhBMP-2 (1.167, 0.117, and 0.039 mg/mL) were combined with an absorbable collagen sponge (ACS) as a delivery vehicle. The rhBMP-2/ACS implants were placed in the subcutaneous tissues of rat backs. X-ray microcomputed tomography (micro-CT) and histological analysis were used to evaluate bone formation. The samples treated with 1.167 mg/mL of rhBMP-2 showed greater bone formation than the samples treated with 0.117 mg/mL of rhBMP-2 four weeks after surgery. However, there was no evidence of bone formation in the samples that were treated with 0.039 mg/mL of rhBMP-2. It was found that rhBMP-2 was osteogenic even at one-tenth of its manufacturer's recommended concentration (1.167 mg/mL), indicating its potential for clinical use at lower concentrations.

Combination of a surgical template and a collagen strip for guiding sinus floor elevation in the oblique sinus floor: A technical note

Sinus floor elevation (SFE) by transcrestal approach has been proven to be a predictable and minimally invasive treatment that augments posterior maxilla with insufficient bone height, allowing the prosthetic rehabilitation of this area with dental implants. However, precise and sufficient elevation of the Schneiderian membrane without perforation is challenging through this blind technique especially in the presence of anatomical restrictions. This note describes a novel technique combining a surgical template and an absorbable collagen sponge (ACS) strip for transcrestal SFE in the oblique sinus floor. A surgical template was used to locate the oblique sinus floor and a collagen strip was placed to orient membrane elevation, meanwhile, protect the sinus membrane. Within the limits of present observation, this technique may increase the manipuility while reducing the risk of complications.

BIOMIMETIC HEMATOMA: NOVEL CARRIER DELIVERS EXTREMELY LOW-DOSE RHBMP-2 FOR HIGHLY EFFECTIVE HEALING OF LARGE BONE DEFECTS IN GOATS

Our previous rat study demonstrated an ex vivo-created “Biomimetic Hematoma” (BH) that mimics the intrinsic structural properties of normal fracture hematoma, consistently and efficiently enhanced the healing of large bone defects at extremely low doses of rhBMP-2 (0.33 μg). The aim of this study was to determine if an extremely low dose of rhBMP-2 delivered within BH can efficiently heal large bone defects in goats.Goat 2.5 cm tibial defects were stabilized with circular fixators, and divided into groups (n=2-3): 2.1 mg rhBMP-2 delivered on an absorbable collagen sponge (ACS); 52.5 μg rhBMP-2 delivered within BH; and an empty group. BH was created using autologous blood with a mixture of calcium and thrombin at specific concentrations. Healing was monitored with X-rays. After 8 weeks, femurs were assessed using microCT.Using 2.1 mg on ACS was sufficient to heal 2.5 cm bone defects. Empty defects resulted in a nonunion after 8 weeks. Radiographic evaluation showed earlier and more robust callus formation with 97.5 % (52.5 μg) less of rhBMP-2 delivered within the BH, and all tibias were fully bridged at 3 weeks. The bone mineral density was significantly higher in defects treated with BH than with ACS. Defects in the BH group had smaller amounts of intramedullary and cortical trabeculation compared to the ACS group, indicating advanced remodeling.The results confirm that the delivery of rhBMP-2 within the BH was much more efficient than on an ACS. Not only did the large bone defects heal consistently with a 40x lower dose of rhBMP-2, but the quality of the defect regeneration was also superior in the BH group. These findings should significantly influence how rhBMP-2 is delivered clinically to maximize the regenerative capacity of bone healing while minimizing the dose required, thereby reducing the risk of adverse effects.

Segmental defect healing in the presence or absence of recombinant human BMP2: Novel insights from a rat model.

This study defined and compared the course of native, impaired and growth factor-stimulated bone regeneration in a rat femoral defect model. A mid-diaphyseal defect with rigid internal fixation was surgically created in the right femur of male Fischer rats and serially analyzed over 36 weeks. Native bone regeneration was modeled using a sub-critical, 1 mm size defect, which healed uneventfully. Critical size defects of 5 mm were used to analyze impaired bone regeneration. In a third group, the 5 mm defects were filled with 11 µg of recombinant human bone morphogenetic protein 2 (rhBMP2) impregnated onto an absorbable collagen sponge, modeling its clinical use. Native bone regeneration was characterized by endochondral ossification with progressive remodeling to ultimately resemble intact femora. An endochondral response was also observed under conditions of impaired bone regeneration, but by week 8 medullary capping occurred with fibrofatty consolidation of the tissue within the defect, resembling an atrophic non-union. rhBMP2 treatment was associated with prolonged inflammatory cytokine expression and rapid intramembranous bone formation occurring with reduced expression of cartilage-associated collagens. Between weeks 4 and 36, rhBMP2-treated bones demonstrated decreased trabecular number and increased trabecular separation, which resulted in inferior mechanical properties compared with bones that healed naturally. Clinical Significance: Recombinant human bone morphogenetic protein 2 (rhBMP2) is used clinically to promote healing of long bones. Our data suggest that it drives intramembraneous ossification producing an inferior regenerate that deteriorates with time. Clinical outcomes would be improved by technologies favoring endochondral regenerative ossification.

Bone Morphogenetic Protein 2 Promotes Bone Formation in Bone Defects in Which Bone Remodeling Is Suppressed by Long-Term and High-Dose Zoledronic Acid.

The use of long-term and high-dose bisphosphate is associated with severely suppressed bone turnover and the delayed union of fractures. However, therapeutic methods to overcome the negative effects of bisphosphonate use are lacking. Bone morphogenetic proteins (BMPs) are powerful osteoinductive proteins. The development of the delivery system using BMP has been verified to have an excellent effect on fracture healing and the enhancement of osteointegration. We hypothesized that BMPs had similar effects as autografts in patients with decreased bone healing potential due to long-term bisphosphonate treatment. Forty rats were divided into the following four groups depending upon the materials implanted into the femoral defect after ten weeks of bisphosphonate (zoledronic acid) injections: Group I: absorbable collagen sponge (control); group II: demineralized freeze-dried bone graft; group III: autogenous bone graft; and group IV: rhBMP-2 with an absorbable collagen sponge. Radiographic union, micro-computed tomography (CT) analysis, manual palpation, and histologic analysis were evaluated. The radiographic union rate, manual union rate, and micro-CT bone volume in groups III and IV were significantly higher than those in groups I and II. Groups III and IV showed similar results to each other. Although the amount of immature bone in the BMP-treated group was large, the effect was similar to that of autografts in the bone defect model in which bone turnover was severely reduced by bisphosphonate treatment. BMP might be a good substitute for autografts in patients with decreased bone healing potential due to long-term bisphosphonate treatment.

The optimal dosage of hyaluronic acid for bone regeneration in rat calvarial defects.

Hyaluronic acid (HA) affects angiogenesis and promotes the migration and differentiation of mesenchymal cells, thereby activating the osteogenic ability of osteoblasts. Although studies on the action of HA during bone regeneration are being actively conducted, the optimal dose of HA required for bone regeneration remains unclear. Therefore, the purpose of this study was to elucidate the most effective HA dose for bone formation using a rat critical-size defect model. Thirty rats were randomly divided into 5 groups, with 6 rats in each group. An absorbable collagen sponge soaked with HA or saline was used to fill an 8-mm defect, which was then covered with a collagen membrane. Different treatments were performed for each group as follows: (1) saline control, (2) 1 mg/mL HA, (3) 25 mg/mL HA, (4) 50 mg/mL HA, or (5) 75 mg/mL HA. After a healing period of 4 weeks, micro-computed tomography and histological analysis were performed. The obtained values were analyzed using analysis of variance and the Tukey test (P<0.05). At week 4, the 75 mg/mL HA group had the highest bone volume/total volume ratio, new bone, and bone fill among the 5 groups, and these values were significantly different from those observed in the control group (P<0.01) and 1 mg/mL HA group (P<0.001). More active bone formation was observed in the higher-dose HA groups (25 mg/mL, 50 mg/mL, and 75 mg/mL HA), which included a large amount of woven bone. The 75 mg/mL HA group showed better bone formation than the other groups (1, 25, and 50 mg/mL HA and control).

Carrier systems for bone morphogenetic proteins: An overview of biomaterials used for dentoalveolar and maxillofacial bone regeneration.

Different types of biomaterials have been used to fabricate carriers to deliver bone morphogenetic proteins (BMPs) in both dentoalveolar and maxillofacial bone regeneration procedures. Despite that absorbable collagen sponge (ACS) is considered the gold standard for BMP delivery, there is still some concerns regarding its use mainly due to its poor mechanical properties. To overcome this, novel systems are being developed, however, due to the wide variety of biomaterial combination, the heterogeneous assessment of newly formed tissue, and the intended clinical applications, there is still no consensus regarding which is more efficient in a particular clinical scenario. The combination of two or more biomaterials in different topological configurations has allowed specific controlled-release patterns for BMPs, improving their biological and mechanical properties compared with classical single-material carriers. However, more basic research is needed. Since the BMPs can be used in multiple clinical scenarios having different biological and mechanical needs, novel carriers should be developed in a context-specific manner. Thus, the purpose of this review is to gather current knowledge about biomaterials used to fabricate delivery systems for BMPs in both dentoalveolar and maxillofacial contexts. Aspects related with the biological, physical and mechanical characteristics of each biomaterial are also presented and discussed.

8. Controlling the rate of presentation of BMP-2 from a highly structured composite for interbody fusion

<h3>BACKGROUND CONTEXT</h3> Clinical outcomes for patients with intervertebral disc injury or disease could be enhanced by a new generation of regenerative matrices that create tissue environments to control angiogenesis, chondrogenesis and bone formation. Regenerative matrices are a new class of controlled release biomaterials that use surface chemistry, materials architecture, and drug delivery kinetics to create optimised conditions for cell interactions that result in high quality tissue formation. A novel biodegradable polymer matrix that releases bone morphogenetic protein 2 (BMP-2) over a period of one month with a low Cmax and sustained presentation of active protein is investigated in this study. <h3>PURPOSE</h3> BMP-2 release kinetics and protein activity from a new regenerative matrix was quantified to develop an implant with 1-month release of the drug within a macroporous matrix. A rabbit posterolateral (PLF) vertebral fusion model was performed to assess bone formation and fusion outcomes as a function of dose and source of BMP-2 (E coli and CHO cells). <h3>STUDY DESIGN/SETTING</h3> The study was designed to investigate the effect of various biomaterial design factors and manufacturing methods on the rate of release of BMP-2, the activity of BMP-2 and the extent of in vivo bone formation. <h3>PATIENT SAMPLE</h3> This study did not involve patients. <h3>OUTCOME MEASURES</h3> Outcome measures were the mechanical properties of the regenerative matrix, the architecture and porosity of the matrix, BMP-2 release kinetics, BMP-2 activity as a function of release time and bone formation. <h3>METHODS</h3> BMP-2 activity was tested using two in vitro cell-based assays. BMP-2 solutions and regenerative matrix extracts were tested for their ability to induce alkaline phosphatase (ALP) activity in W-20-17 stromal cells using a modified version of the ASTM standard protocol for in vitro biological activity of recombinant human BMP-2 (F2131-02). Cells were exposed to serial dilutions of BMP-2 for 24 h, assayed for ALP activity and EC50s calculated allowing the relative potency to NIBSC WHO BMP-2 standard to be calculated. BMP-2 release was assessed with an in vitro assay of ALP activity in the mouse muscle myoblast cell line C2C12. Formulations were suspended above a C2C12 monolayer in an insert with a porous (8 µm) PET membrane. Following 3-4 days of exposure, the inserts containing LDGraft were transferred to fresh cells for a further 3-4 days. Cells exposed to LDGraft in this manner were assayed for ALP activity (normalised to protein content) once the inserts had been transferred to fresh cells. Fusion rate and new bone formation were compared to INFUSE in a single-level lumbar (L4-L5) posterolateral (PLF) vertebral fusion model in the New Zealand White rabbit based on standard protocols (Boden et al, 1995, ASTM F3207). Pre-formed implants (3 cc) of INFUSE (rhBMP-2 and absorbable collagen sponge) were implanted on decorticated dorsal surfaces of the transverse processes of L4 and L5 adjacent to the laminae. Fusion was assessed by radiograph at 6 weeks and by manual palpation, radiography and microCT at 12 weeks. <h3>RESULTS</h3> BMP-2 activity was retained over one month of release with the addition of protectants in the formulation and precise control of manufacturing processes. The matrix had a porosity of >70% with a high proportion of macropores (>200 micron diameter). Low doses of E. coli-produced BMP-2 induced fusion in the PLF model. <h3>CONCLUSIONS</h3> A one-month releasing BMP-2 formulation is described. This new regenerative matrix had internal architecture designed to host angiogenesis. Manufacturing and design challenges can be overcome to retain high biologic activity throughout the release period. <h3>FDA DEVICE/DRUG STATUS</h3> BMP-2, not approved for this indication.

P7. Percutaneous delivery of recombinant human bone morphongenetic protein-2 augments fusion in a nicotine impaired rabbit fusion model

<h3>BACKGROUND CONTEXT</h3> Several experiments have shown that delivery of BMP-2 in a delayed fashion from initial surgery may be superior for optimal bone growth. Studies of posterolateral spinal fusion in rabbits have shown that BMP-2 expression physiologically peaks from 4-6 weeks out from surgery. The hypothesis of this experiment was that delayed delivery of ACS/BMP-2 would augment bone growth in a rabbit spine fusion model. By percutaneously delivering ACS/BMP-2 we would be able to deliver osteoinductive material at an optimal time point away from the typical time of surgery application. <h3>PURPOSE</h3> To determine if controlled delivery of morselized absorbable collagen sponge (ACS) Rh-BMP2 in a delayed manner postsurgery would allow for increased fusion mass in a rabbit spine fusion model. <h3>STUDY DESIGN/SETTING</h3> A nicotine-impaired spinal fusion rabbit model. <h3>PATIENT SAMPLE</h3> Sixteen male one-year-old rabbits. <h3>OUTCOME MEASURES</h3> Histologic, radiologic (radiographic, CT scans) and palpation examinations were performed to determine fusion status and the volume of bone formed. <h3>METHODS</h3> Sixteen male one-year-old rabbits underwent a posterolateral spinal fusion with iliac crest bone graft at L5-L6 while being given nicotine to prevent spinal fusion as previously published. Eight were controls while 8 had morselized RhBMP-2 (4.2mg) injected at the fusion site at 4 weeks postoperatively. At 12 weeks, the rabbits were sacrificed. Histologic, radiologic (radiographic, CT scans) and palpation examinations were performed to determine fusion status and the volume of bone formed. Hematoxylin and eosin (H&E) stain were used for histology. There were 4 of 8 rabbits in the control cohort with bilateral fusion and 6 of 8 rabbits in the BMP-DDC that had bilateral fusion based on CT scan analysis. A student's t-test was used to compare the CT scan measured volume of bone created between control (CC) and RhBMP-2 delayed delivery cohorts (BMP-DDC). <h3>RESULTS</h3> Seven of 8 rabbits in the BMP-DDC and 5 of 8 rabbits in the CC formed definitive fusion with a positive palpation exam, bridging bone between transverse processes on CT scan and an x-ray showing fusion. Histologic analysis revealed new remodeled bone within the BMP-DDC. There was increased average volume of bone formed within the BMP-DDC vs the CC (22.6±13.1 vs 11.1±3.6 cm^3, p=0.04) <h3>CONCLUSIONS</h3> Our study shows that injectable morselized ACS/RhBMP-2 can create twice as much bone within a nicotine-impaired rabbit spine fusion model when delivered 4 weeks out from time of surgery. <h3>FDA DEVICE/DRUG STATUS</h3> This abstract does not discuss or include any applicable devices or drugs.

P9. Sex differences in response to recombinant human bone morphogenetic protein-2 (rhBMP-2) in a rat posterolateral fusion spine model

<h3>BACKGROUND CONTEXT</h3> Estrogen modulates bone activity largely through inhibition of osteoclast bone resorption and bone morphogenetic protein-2 (BMP-2) signaling pathways in osteoblasts. Minimal data exist concerning potential sex-dependent differences in BMP-2-mediated bone regeneration. Here, we aimed to quantify sex-dependent differences in the bone healing response to recombinant human BMP-2 (rhBMP-2) treatment in a rat posterolateral spinal fusion model. <h3>METHODS</h3> A total of 48 female and male Sprague-Dawley rats (N=24/group), underwent L4-L5 posterolateral fusion with bilateral placement of absorbable collagen sponges, loaded with 5 µg of rhBMP-2 (10 µg/animal). At 8 weeks postoperatively, 10 specimens of each sex were tested in flexion-extension with quantification of range of motion (ROM) and stiffness. The remaining specimens were evaluated for new bone growth and fusion via radiography, blinded manual palpation and microcomputed tomography (microCT). Laboratory microCT quantified bone microarchitecture and synchrotron microCT examined bone microstructure. On histology, the number of adipocytes within a standardized field of view was counted for both male and female fusion masses. <h3>RESULTS</h3> Manual palpation scores differed significantly between sexes, with mean fusion scores of 2.4±0.4 in females vs 3.1±0.6 in males, p < 0.001. Biomechanical stiffness did not differ between sexes, but ROM was significantly greater and more variable for females vs males (3.7°±5.6° vs 0.27°±0.15°,p < 0.005, respectively. Laboratory microCT showed significantly smaller volumes of fusion masses in females vs males (262±87 mm³ vs 732±238 mm³, respectively, p < 0.001) but significantly higher bone volume fraction (0.27±0.08 vs 0.12±0.05, respectively, p < 0.001). Mean trabecular thickness was not different, but trabecular number was significantly greater in females (3.1±0.5 mm-1vs 1.5±0.4 mm-1, respectively, p<0.001). Synchrotron microCT showed fine bone structures developing in both sexes at the 8-week time point. Adipocyte density was found to be significantly higher in male fusion masses than in female fusion masses. <h3>CONCLUSIONS</h3> This study demonstrates that sex-based factors may influence both the quantity and quality of bone formation in patients receiving rhBMP-2 for a variety of orthopedic applications. We found that male rats had significantly higher fusion scores, greater fusion-mass volume, and lower ratio of new bone volume to total volume compared to female rats. Male and female mean trabecular thicknesses were equal, but the female fusion masses were more densely filled with trabeculae (higher trabecular number) than those of males. MicroCT results suggested that the male trabeculae were more rod-like and the female more plate-like, although the methods that were used are generally considered less accurate for trabecular bone, which has a high degree of concavity. The incomplete fusions on three female rats, evidenced by high ROM and small gaps on microCT, suggest that females may have a more variable response to rhBMP-2. Investigation into whether these sex-based differences are specific to rhBMP-2-induced bone formation or are more general to bone regeneration/healing is prudent. <h3>FDA DEVICE/DRUG STATUS</h3> This abstract does not discuss or include any applicable devices or drugs.