Greater Bone Volume Research Articles

Impacts of permeability and effective diffusivity of porous scaffolds on bone ingrowth: In silico and in vivo analyses

The permeability and the effective diffusivity of a porous scaffold are critical in the bone-ingrowth process. However, design guidelines for porous structures are still lacking due to inadequate understanding of the complex physiological processes involved. In this study, a model integrating the fundamental biological processes of bone regeneration was constructed to investigate the roles of permeability and effective diffusivity in regulating bone deposition in scaffolds. The in silico analysis results were confirmed in vivo by examining bone depositions in three diamond lattice scaffolds manufactured using selective laser melting. The findings show that the scaffolds with better permeability and effective diffusivity had deeper bone ingrowth and greater bone volume. Compared to permeability, effective diffusivity exhibited greater sensitivity to the orientation of porous structures, and bone ingrowth was deeper in the directions with higher effective diffusivity in spite of identical pore size. A 4.8-fold increase in permeability and a 1.6-fold increase in effective diffusivity by changing the porous structure led to a 1.5-fold increase in newly formed bone. The effective diffusivity of the porous scaffold affects the distribution of osteogenic growth factor, which in turn impacts cell migration and bone deposition through chemotaxis effects. Therefore, effective diffusivity may be a more suitable indicator for porous scaffolds because our study shows changes in this parameter determine changes in bone distribution and bone volume.

Bone regeneration using activin A/BMP2 chimera (AB204) with collagen membrane in rats with calvarial defects.

Collagen has long been recognized as an excellent carrier for growth factors, and membrane-type collagen has been widely applied in dentistry for guided bone regeneration. This study was conducted to examine the effects of an activin A/BMP2 chimera (AB204) combined with a collagen membrane (CM) on bone repair in a rat calvarial defect model. A unilateral calvarial defect measuring 5.0 mm was surgically created in 32 Sprague-Dawley rats. The rats were then randomly assigned to 1 of 4 groups, each consisting of 8 animals: control (untreated), CM (treated with a CM only), CM/bone morphogenetic protein 2 (BMP2) (treated with a CM and 1.0 μg of BMP2), and CM/AB204 (treated with a CM and 1.0 μg of AB204). Bone regeneration was evaluated using micro-computed tomography (CT) and histological analysis at 2 and 4 weeks following surgery. Micro-CT analysis revealed that bone formation in the CM/BMP2 and CM/AB204 groups was superior to that observed in the control and CM groups at both 2 and 4 weeks postoperatively. BMP2 induced greater bone regeneration than AB204 at 2 weeks; however, AB204 resulted in a greater bone volume at 4 weeks, achieving the highest values recorded. No significant differences were found between the CM/BMP2 and CM/AB204 groups at either time point (P>0.05). On histological examination, new bone formation was evident in both CM/BMP2 and CM/AB204 groups. Within the limitations of this study, the findings indicate that AB204 may enhance osteogenic potential when used in combination with CM for bone regeneration.

Can growth in captivity alter the calcaneal microanatomy of a wild ungulate?

Reduced mobility associated with captivity induces changes in biomechanical stress on the skeleton of domesticated animals. Due to bone plasticity, bone’s morphology and internal structure can respond to these new biomechanical stresses over individuals’ lifetime. In a context where documenting early process of animal domestication is challenging, this study will test the hypothesis that change in mobility patterns during a wild ungulate’s life will alter the internal structure of its limb bones and provide a proof of concept for the application of this knowledge in Zooarchaeology. Using the calcaneus as a phenotypic marker through qualitative and quantitative 3D microanatomical analyses, we relied on a comparative study across wild boars (Sus scrofa) populations from controlled experimental conditions with different mobility patterns (natural habitat, large pen, and stall) and archaeological specimens collected from middle and late Mesolithic as surrogate for the norm of reaction in European wild boar phenotype before the spread of agriculture and domestic pigs. Results provide evidence for compressive and tensile forces as the main elements affecting the variation in the cortical thickness along the calcaneus. Furthermore, changes in the internal structure of the calcaneus between mobility patterns are observed but their intensity is not directly associated with the degree of mobility restriction and only weakly impacted by the size or weight of the individuals. Despite having greater bone volume, the calcaneus of the Mesolithic wild boars displays a very similar microanatomy compared to the present-day hunted or captive wild boars. These results suggest that calcaneal microanatomy is more affected by population differences than by locomotor variation. For all these reasons, this preliminary study doesn’t support the use of microanatomy of the calcaneus as an indicator of change in locomotor behaviour induced by captivity in the archaeological record.

Impact of Bifidobacterium animalis subsp. lactis HN019 probiotic in the prevention of periodontitis associated with immunosuppression.

This study evaluated the effects of the probiotic Bifidobacterium animalis subsp. lactis HN019 (HN019) in the development of experimental periodontitis (EP) in rats submitted to chemotherapy (5-fluorouracil [5FU]). Eighty male rats were divided into the following groups: control (C); treated with 5FU (60mg/kg at day 30 and 40mg/kg at day 32); treated with probiotic (HN019) (daily, for 44 days, starting at day 0); treatment with 5FU and probiotic (5FU-HN019); only EP (EP) (ligature placed on lower first molars at day 30, maintained for 14 days); EP and treatment with 5FU (EP-5FU); EP and treatment with probiotic (EP-HN019); and EP and treatment with 5FU and probiotic (EP-5FU-HN019). Euthanasia occurred at day 44. Morphometric, histomorphometric, microtomographic, immunohistochemical, immunoenzymatic, and gene expressions analyses were performed. The data obtained were statistically analyzed (p<0.05). The EP-5FU-HN019 group showed less bone and connective tissue loss when compared with EP-5FU group, while EP-HN019 and EP-5FU-HN019 groups had greater bone volume than EP and EP-5FU groups, respectively (p<0.05). A decrease in immunostaining for tartrate-resistant acid phosphatase and RANKL, an increase for osteoprotegerin and lower interleukin-1β levels were observed in EP-5FU-HN019 group, when compared with EP-5FU group (p<0.0001). Probiotic therapy led to an increase in the proportions of B. lactis in the feces (p=0.0018), but not in the biofilm, and reduced the expression of Fusobacterium nucleatum and Prevotella intermedia in the biofilm (p<0.0001). B. lactis HN019 reduced the severity of EP in rats submitted to chemotherapy, modulating immunoinflammatory parameters in periodontal tissues and reducing periodontopathogens expression on biofilm in rats submitted to chemotherapy.

Bone structural, biomechanical, and histomorphometric characteristics of the hindlimb skeleton in the marsh rice rat (Oryzomys palustris).

The rice rat (Oryzomys palustris) is a nonconventional laboratory rodent species used to model some human bone disorders. However, no studies have been conducted to characterize the postcranial skeleton. Therefore, we aimed to investigate age- and gender-related features of the hindlimb skeleton of this species. We used femurs and tibiae from 94 rats of both genders aged 4-28 weeks. Bone mineral content (BMC), volumetric bone mineral density (vBMD), and biomechanical properties were determined in femurs. In addition, bone histomorphometry of tibiae was conducted to assess bone cell activities and bone turnover over time. Bone length, total metaphysis BMC and vBMD, mid-diaphyseal BMC and vBMD, cortical thickness, and cortical area progressively augmented with age. Whereas the increase in these parameters plateaued at age 16-22 weeks in female rats, they continued to rise to age 28 weeks in male rats. Furthermore, bone strength parameters increased with age, with few differences between genders. We also observed a rapid decrease in longitudinal growth between age 4 and 16 weeks. Whereas young rats had a greater bone formation rate and bone turnover, older rice rats had greater bone volume and trabecular thickness, with no differences between genders. (a) Sexual dimorphism in the rice rat becomes grossly evident at age 16 weeks; (b) the age-related increases in bone mass, structural cortical parameters, and in some biomechanical property parameters plateau at an older age in male than in female rats; and (c) bone growth and remodeling significantly decreased with age irrespective of the gender.

Antibacterial Fluorinated Diamond-like Carbon Coating Promotes Osteogenesis—Comparison with Titanium Alloy

Fluorinated diamond-like carbon (F-DLC) coating is biologically safe, provides superior antibacterial properties, and shows promise in preventing postoperative peri-implant infections. However, potential negative effects of this coating on in vivo bone formation and resorption have not been studied. The authors investigated the effects of F-DLC coatings on bone union in beagle dogs. Seventy-two solid columns of titanium alloy were prepared with equally spaced slits. Half of these columns were coated with F-DLC (Group F), and the others were left uncoated as controls (Group C). Columns were implanted in the femurs of beagle dogs, and in vivo bone formation and resorption were assessed 4, 8, and 12 weeks after implantation. In comparison to Group C, Group F showed significantly greater bone volume and trabecular thickness at Week 8 (p &lt; 0.05) and Week 12 (p &lt; 0.005) and significantly lower bone resorption activity, measured by the ratio of osteoclasts to bone surface and of eroded surface to bone surface, at Week 12 (p &lt; 0.05). The F-DLC coating encouraged bone formation in vivo more effectively than uncoated titanium alloy, suggesting that F-DLC will prove to be a useful coating material for antibacterial intraosseous implants.

Microstructural and mechanical recovery of bone in ovariectomized rats: The effects of menaquinone-7

The loss of bone quantity and quality in postmenopausal female patients can be a problem for dental treatment. A sufficient intake of nutrients such as calcium, magnesium, and vitamins D and K is likely correlated with the mechanical properties of bone. In particular, vitamin K2, also called menaquinone (MK), inhibits bone loss in postmenopausal women. Here we demonstrate the microstructural and mechanical properties of bone recovery in ovariectomized (OVX) rats during MK-7 administration. Bilateral ovariectomy and a sham operation were performed on 14-week-old female SPF Wistar rats. MK-4 and -7 were orally administered at 30 mg/kg daily for 12 weeks. The femur was used for the 3-point bending test and microstructural analysis of the cancellous bone by micro-CT, and the mandibular cortical bone for the evaluation of mechanical properties on a nanoscale. Micro-computed tomography revealed irregular trabecular architecture, hollow marrow cavities, and sparse trabecular bone in the femurs of the OVX group. Trabecular bone structure analysis showed that the MK-7 group had greater bone volume per tissue volume (BV/TV) and a higher trabecular number than the OVX group. The bulk-scale 3-point bending test did not allow the mechanical properties between OVX and OVX/MK7 groups to be discerned, yet at the smallest level, the elastic–plastic transition point of the nanoindentation stress–strain curve of the mandibular cortical bone was higher in the MK-7 group than in the OVX group. These findings suggest that MK-7 enables bone microstructural and mechanical recovery in the OVX model.

Acceleration of bone formation using insitu-formed hyaluronan-hydrogel containing bone morphogenetic protein-2 in a mouse critical size bone defect model.

An enzymatic crosslinking strategy using hydrogen peroxide and horseradish peroxidase is receiving increasing attention for application with in situ-formed hydrogels (IFHGs). IFHGs may also be ideal carrier materials for bone repair, although their ability to carry bone morphogenetic protein-2 (BMP2) has yet to be examined. We examined the effectiveness of an IFHG made of hyaluronan (IFHG-HA) containing BMP2 for promoting bone formation in a mouse critical size bone defect model. C57/BL6J mice received a 2-mm femoral critical-sized bone defect before being randomly assigned to one of the following treatment groups (n =6): control (no treatment), IFHG-HA only, PBS with BMP2, and IFHG-HA with BMP2. X-ray radiographs were utilized to track new bone formation, and micro-computed tomography and histological examination were performed on new bone formed at the bone defect site two weeks after surgery. Mice treated with PBS with BMP2 and IFHG-HA with BMP2 had greater bone volume (BV) and bone mineral content (BMC) than those receiving control, and successfully achieved consolidation. Mice treated with IFHG-HA with BMP2 had significantly higher BV and BMC than those treated with PBS with BMP2. IFHG-HA may be an effective carrier for BMP2 to enable delivery for bone defect repair.

Poly(POG)n loaded with recombinant human bone morphogenetic protein-2 accelerates new bone formation in a critical-sized bone defect mouse model

BackgroundDelivery of bone morphogenetic protein-2 (BMP-2) via animal-derived absorbable collagen materials is used for the treatment of large bone defects. However, the administration of bovine proteins to humans is associated with the risk of zoonotic complications. We therefore examined the effect of combining BMP-2 with collagen-like peptides, poly(POG)n, in a critical-sized bone defect mouse model.MethodsA 2-mm critical-sized bone defect was created in the femur of 9-week-old male C57/BL6J mice. Mice were randomly allocated into one of four treatment groups (n = 6 each): control (no treatment), poly(POG)n only, 0.2 μg, or 2.0 μg BMP-2 with poly(POG)n. New bone formation was monitored using soft X-ray radiographs, and bone formation at the bone defect site was examined using micro-computed tomography and histological examination at 4 weeks after surgery.ResultsAdministration of 2.0 μg of BMP-2 with poly(POG)n promoted new bone formation and resulted in greater bone volume and bone mineral content than that observed in the control group and successfully achieved consolidation. In contrast, bone formation in all other groups was scarce.ConclusionsOur findings suggest the potential of BMP-2 with poly(POG)n as a material, free from animal-derived collagen, for the treatment of large bone defects.

Acceleration of bone union by in situ-formed hydrogel containing bone morphogenetic protein-2 in a mouse refractory fracture model

BackgroundAn enzymatic crosslinking strategy using hydrogen peroxide and horseradish peroxidase is receiving increasing attention for application with in situ-formed hydrogels (IFHs). Several studies have reported the application of IFHs in cell delivery and tissue engineering. IFHs may also be ideal carrier materials for bone repair, although their potential as a carrier for bone morphogenetic protein (BMP)-2 has yet to be examined. Here, we examined the effect of an IFH made of hyaluronic acid (IFH-HA) containing BMP-2 in promoting osteogenesis in a mouse refractory fracture model.MethodsImmediately following a fracture procedure, animals either received no treatment (control) or an injection of IFH-HA/PBS or IFH-HA containing 2 μg BMP-2 (IFH-HA/BMP-2) into the fracture site (n = 16, each treatment).ResultsFracture sites injected with IFH-HA/BMP-2 showed significantly greater bone volume, bone mineral content, and bone union compared with sites receiving no treatment or treated with IFH-HA/PBS alone (each n = 10). Gene expression levels of osteogenic markers, Alpl, Bglap, and Osx, were significantly raised in the IFH-HA/BMP-2 group compared to the IFH-HA/PBS and control groups (each n = 6).ConclusionIFH-HA/BMP-2 may contribute to the treatment of refractory fractures.

Effect of DNA/protamine complex paste on bone augmentation of the mandible: A pilot study on dogs

ObjectiveOur previous studies found that a salmon DNA-based scaffold containing protamine promoted bone regeneration of the calvarial defects of rats. The aim of the present pilot study was to examine the influence of the DNA/protamine (DP) complex on bone regeneration of a saddle type, alveolar ridge defects of the dog mandible. DesignAlveolar ridge defects were performed in the mandibles of five adult female beagles. The following three treatment modalities were randomly allocated: (1) the DP complex paste, (2) a beta-tricalcium phosphate (β-TCP), and (3) a blank (control). Healing of bone defects were evaluated by periapical radiography, micro-computed tomography (micro-CT), and histology. ResultsPeriodical radiographic images revealed that a higher percentage of regenerated bone height was consistently achieved in the DP group, as compared with blank controls. All three-dimensional, sagittal, and coronal images of micro-CT showed increased amounts of newly formed bone and a greater bone volume/ tissue volume ratio, as compared with the blank and β-TCP groups. In contrast, there was no significant difference in bone mineral density among the groups. Histological analysis confirmed that the alveolar bone defects were filled with newly formed bone with mature and compact properties in the DP group. ConclusionsThese findings indicate that the DP complexes enhanced regeneration of vertical alveolar bone defects of the dog mandible.

Ingestion of gastrolith mineralized matrix increases bone volume and tissue volume in mouse long bone fracture model

PurposeFracture healing often requires extended convalescence as the bony fragments consolidate into restored viable tissue for load-bearing. Development of interventions to improve healing remains a priority for orthopaedic research. The goal of this study was to evaluate the ability of a naturally occurring matrix of amorphous calcium carbonate to affect fracture healing in an uninstrumented long bone model. MethodsComplete transverse fracture was induced in the fibula of mature mice, followed by daily gavage of crushed gastrolith from crayfish at doses of 0 (control), 1 (1 MG), and 5 (5 MG) mg/kg. At Day 17, bones and sera were harvested. ResultsMorphologically, the 1 MG treated group had greater bone volume (BV), and both 1 MG and 5 MG had greater tissue volume (TV) than control (p < 0.05), as determined by μCT; BV/TV and mineral density did not yield a statistical difference. Histologically, regional variations in mineralized matrix were evident in all specimens, indicating a broad continuum of healing within the callus. Among serum proteins, bone-specific alkaline phosphatase, indicative of active mineralization, was greater in 5 MG than control (p < 0.05). Sclerostin, an inhibitor of osteogenesis, was lower in 5 MG than control (p < 0.05), also suggestive of enhanced healing. ConclusionsAn increase in bone volume, tissue volume and cellular signaling for osteogenesis at 17 days following fibula fracture in this mouse model suggests that gastrolith treatment holds potential for improving fracture healing. Further study at subsequent time points is warranted to determine the extent to which the increase in callus size with gastrolith treatment may accelerate restoration of tissue integrity.

Bone regeneration capacities of alveolar bone mesenchymal stem cells sheet in rabbit calvarial bone defect

Mesenchymal stem cells sheets have been verified as a promising non-scaffold strategy for bone regeneration. Alveolar bone marrow mesenchymal stem cells, derived from neural crest, have the character of easily obtained and strong multi-differential potential. However, the bone regenerative features of alveolar bone marrow mesenchymal stem cells sheets in the craniofacial region remain unclear. The purpose of the present study was to compare the osteogenic differentiation and bone defect repairment characteristics of bone marrow mesenchymal stem cells sheets derived from alveolar bone (alveolar bone marrow mesenchymal stem cells) and iliac bone (Lon-bone marrow mesenchymal stem cells) in vitro and in vivo. Histology character, osteogenic differentiation, and osteogenic gene expression of human alveolar bone marrow mesenchymal stem cells and Lon-bone marrow mesenchymal stem cells were compared in vitro. The cell sheets were implanted in rabbit calvarial defects to evaluate tissue regeneration characteristics. Integrated bioinformatics analysis was used to reveal the specific gene and pathways expression profile of alveolar bone marrow mesenchymal stem cells. Our results showed that alveolar bone marrow mesenchymal stem cells had higher osteogenic differentiation than Lon-bone marrow mesenchymal stem cells. Although no obvious differences were found in the histological structure, fibronectin and integrin β1 expression between them, alveolar-bone marrow mesenchymal stem cells sheet exhibited higher mineral deposition and expression levels of osteogenic marker genes. After being transplanted in the rabbit calvarial defects area, the results showed that greater bone volume and trabecular thickness regeneration were found in bone marrow mesenchymal stem cells sheet group compared to Lon-bone marrow mesenchymal stem cells group at both 4 weeks and 8 weeks. Finally, datasets of bone marrow mesenchymal stem cells versus Lon-bone marrow mesenchymal stem cells, and periodontal ligament mesenchymal stem cells (another neural crest derived mesenchymal stem cells) versus umbilical cord mesenchymal stem cells were analyzed. Total 71 differential genes were identified by overlap between the 2 datasets. Homeobox genes, such as LHX8, MKX, PAX9, MSX, and HOX, were identified as the most significantly changed and would be potential specific genes in neural crest mesenchymal stem cells. In conclusion, the Al-bone marrow mesenchymal stem cells sheet-based tissue regeneration appears to be a promising strategy for craniofacial defect repair in future clinical applications.

Skeletal Site-Specific Response of Jawbones and Long Bones to Surgical Interventions in Rats Treated with Zoledronic Acid.

Bisphosphonates (BPs) have been extensively used for management of bone diseases with pathologically high resorption. Despite the great clinical benefits, a severe complication known as medication-related osteonecrosis of the jaw (MRONJ) has been reported. It is found that most of the reported MRONJ cases were limited in the jawbones/craniofacial bones instead of long bones. The present study aims to investigate the differential bone response to surgical procedures between jawbones and long bones exposed to BPs. Forty-eight skeletal mature Sprague Dawley female rats were administered oncologic dose of zoledronic acid (ZA) or normal saline for 4 weeks and then subjected to tooth extraction on the mandible and maxilla, and a bone defect creation on the femur. After surgical procedures, ZA or saline treatment were continued until sacrifice at week 2, week 4, and week 8, post-operatively. The samples were subjected to micro-computerized tomography (micro-CT) and histological assessment. Osteonecrosis was only found in jawbones in ZA-treated rats. ZA-treated rats showed significantly higher bone mineral density with greater bone volume in all surgical sites than that in the controls. The length of exposure of ZA did not seem to affect trabecular microstructure, and it only showed higher bone volume and BMD with longer healing time which is expected in the healing process.

Abaloparatide, a PTH receptor agonist with homology to PTHrP, enhances callus bridging and biomechanical properties in rats with femoral fracture.

Fractures typically heal via endochondral and intramembranous bone formation, which together form a callus that achieves union and biomechanical recovery. PTHrP, a PTH receptor agonist, plays an important physiological role in fracture healing as an endogenous stimulator of endochondral and intramembranous bone formation. Abaloparatide, a novel systemically-administered osteoanabolic PTH receptor agonist that reduces fracture risk in women with postmenopausal osteoporosis, has 76% homology to PTHrP, suggesting it may have potential to improve fracture healing. To test this hypothesis, ninety-six 12-week-old male rats underwent unilateral internally-stabilized closed mid-diaphyseal femoral fractures and were treated starting the next day with daily s.c. saline (Vehicle) or abaloparatide at 5 or 20 µg/kg/d for 4 or 6 weeks (16 rats/group/time point). Histomorphometry and histology analyses indicated that fracture calluses from the abaloparatide groups exhibited significantly greater total area, higher fluorescence scores indicating more newly-formed bone, and higher fracture bridging scores versus Vehicle controls. Callus bridging score best correlated with callus cartilage score (r = 0.64) and fluorescence score (r = 0.67) at week 4, and callus area correlated with cartilage score (r = 0.60) and fluorescence score (r = 0.89) at Week 6. By micro-CT, calluses from one or both abaloparatide groups had greater bone volume, bone volume fraction, bone mineral content, bone mineral density, and cross-sectional area at both time points versus Vehicle controls. Destructive bending tests indicated greater callus maximum load and stiffness in one or both abaloparatide groups at both time points versus Vehicle controls. These results provide preliminary preclinical evidence for improved fracture healing with systemically-administered abaloparatide. © 2019 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res.

Osteogenic parameters surrounding trabecular tantalum metal implants in osteotomies prepared via osseodensification drilling.

Background Surgical fixation of implants into bone for the correction of bone deformities or defects is a traditional approach for skeletal stabilization. Important measures of efficacy of implants include implant stability and osseointegration—the direct interaction between living bone and an implant. Osseointegration depends on successful implant placement and subsequent bone remodeling. This study utilized osseodensification drilling (OD) in a low bone density model using trabecular metal (TM) implants. Material and Methods Three osteotomy sites, Regular, OD-CW (clockwise), and OD-CCW (counterclockwise), were prepared in each ilium of three female sheep. Drilling was performed at 1100rpm with saline irrigation. Trabecular metal (TM) (Zimmer®, Parsippany, NJ, USA) implants measuring 3.7mm in diameter x 10mm length were placed into respective osteotomies. A three-week period post-surgery was given to allow for healing to take place after which all three sheep were euthanized and the ilia were collected. Samples were prepared, qualitatively and quantitatively analyzed using histology micrographs and image analysis software (ImageJ, NIH, Bethesda, MD). Bone-to-implant contact (BIC) and bone area fraction occupancy (BAFO) were quantified to evaluate the osseointegration parameters. Results All implants exhibit successful bone formation in the peri-implant environment as well as within the open spaces of the trabecular network. Osseointegration within the TM (quantified by %BIC) as a function of drilling technique was more pronounced in OD samples(p>0.05). The %BAFO however shows a significant difference (p=0.036) between the CCW and R samples. Greater bone volume and frequency of bone chips are observed in OD samples. Conclusions The utilization of OD as a design for improved fixation of hardware was supported by increased levels of stability, both primary and secondary. Histological data with OD provided notably different results from those of the regular drilling method. Key words:Osseodensification drilling, trabecular tantalum metal, osteotomies, implants, subtractive drilling.

Exploring bone volume and skeletal weight in the Middle Pleistocene humans from the Sima de los Huesos site (Sierra de Atapuerca, Spain).

Body mass estimation in fossil human species is a crucial topic in paleoanthropology as it yields information about ecologically relevant characteristics. Nevertheless, variables crucial to body mass estimation such as bone volume and skeletal weight have never before been calculated in a fossil human species. The exceptional state of preservation of several fossil human long bones from the Sima de los Huesos (SH) Middle Pleistocene site, in the Sierra de Atapuerca, makes it possible to calculate for the first time the absolute bone volume in five complete long bones (two femora and three humeri) of a fossil human species, an approach not possible in fragmentary or poorly preserved fossils. We have relied on computed tomography scans and 3D reconstructions to calculate bone volume. A sample of 62 complete bones of robust recent humans was also used for comparative purposes. The male SH femora (weight-bearing bones) and humeri (non-weight-bearing bones) have, relative to their size, greater bone volume (volume of bone tissue over total bone volume) than the equivalent bones in our recent human sample. As mass is volume × density, and bone tissue density (as a material) is similar across mammals, we calculate bone mass, and our results show that the SH hominins had on average heavier long bones than extant humans of the same size. From the femoral weight at hand, we have estimated the total skeletal weight in two SH individuals, which is about 36% heavier than in the recent humans of the equivalent body size. Using different methods and skeletal variables, including skeletal weight, to estimate body mass in these two SH humans, we highlight the considerable differences in body mass estimates we obtained, and that the largest body mass estimate is the one based on the skeletal weight. Our results suggest that we cannot assume the same relative proportion of bone volume and bone and skeletal weight characterized the entire genus Homo. Given that skeletal weight has a significant influence on body mass, current body mass estimates of fossil Homo specimens could be systematically underestimated. Thus, the significantly larger bone volume and heavier bones, probably throughout the entire skeleton, of SH humans could have had consequences for many biological parameters in this Pleistocene population and considerable importance for studies focusing on adaptive and ecologically relevant characteristics. Although more recent human samples should be analyzed, in our view, the high skeletal robusticity of the SH sample, including larger bone volume and skeletal weight, is part of their adaptive body type selected for throughout the Pleistocene to support different mechanical and activity regimes and formed under tight genetic control, including control over bone formative and regulatory processes.

Abstract P285: Carotid Sinus Nerve Stimulation Attenuates Alveolar Bone Loss in Rats With Induced Periodontitis.

Introduction: Few studies have focused on the impact of the progression of periodontitis on hypertension. Baroreflex Activation Therapy (BAT) has been used to treat patients with resistant hypertension. Our laboratory has investigated the role of electrical carotid sinus nerve (CSN) stimulation in unanesthetized rats, on local and systemic inflammation. Since periodontitis is a chronic inflammatory disease, the aim of this study was to evaluate the role of the baro- and chemoreflex activation, by means of electrical stimulation of the CSN, on alveolar bone loss in rats with periodontitis. Methods and Results: Under ketamine/xylazine anesthesia Wistar Hannover rats were implanted with electrodes around the CSN and a catheter inserted into the abdominal aorta for blood pressure recording. After 48h periodontitis was induced by the ligation of the bilateral mandibular first molar, followed by the electrical stimulation of the CSN (1,5-4V, 1ms, 30Hz for 10 min); which was continued during the next eight days. At the 8 th day after the induction of periodontitis, the rats were euthanized and the jaws were resected; besides, microtomographic analysis was performed by bi and three-dimensional quantification using micro-computerized tomography. As compared to baseline results the electrical stimulation of the CSN (N=7) promoted a decrease in mean arterial pressure (108± 8 vs. 91± 5 mmHg; p&lt;0.05) and heart rate (374±10 vs. 319±15 bpm; p&lt;0.05). The CSN stimulated rats (N=7) with periodontitis showed greater bone volume and bone surface than the non-stimulated control rats (N=4) with periodontitis, evaluated by three-dimensional analysis (0.5±0.005 vs. 0.7±0.03 mm 3 ; 16.54±0.22 vs. 23.30±0.66 mm 2 , respectively; p&lt;0.05). Moreover, the CSN stimulated rats with periodontitis presented a decrease of the furcation area and interproximal region as compared to the non-stimulated control rats (1.14±0.06 vs. 0.5±0.07 mm; 3.25±0.03 vs. 2,48±0,13 mm, respectively; p&lt;0.05). Conclusions: Overall, these results demonstrate that stimulation of the CSNs promotes a protective effect on alveolar bone loss elicited by periodontitis involving the activation of the baro- and chemoreflex.

Sclerostin Neutralizing Antibody Treatment Enhances Bone Formation but Does Not Rescue Mechanically Induced Delayed Healing

During bone healing, tissue formation processes are governed by mechanical strain. Sost/sclerostin, a key Wnt signaling inhibitor and mechano-sensitive pathway, is downregulated in response to mechanical loading. Sclerostin neutralizing antibody (SclAb) increases bone formation. Nevertheless, it remains unclear whether sclerostin inhibition can rescue bone healing in situations of mechanical instability, which otherwise delay healing. We investigated SclAb's influence on tissue formation in a mouse femoral osteotomy, stabilized with rigid or semirigid external fixation. The different fixations allowed different magnitudes of interfragmentary movement during weight bearing, thereby influencing healing outcome. SclAb or vehicle (veh) was administeredand bone healing was assessed at multiple time points up to day 21 postoperatively by in vivo micro-computed tomography, histomorphometry, biomechanical testing, immunohistochemistry, and gene expression. Our results show that SclAb treatment caused a greater bone volume than veh. However, SclAb could not overcome the characteristic delayed healing of semirigid fixation. Indeed, semirigid fixation resulted in delayed healing with a prolonged endochondral ossification phase characterized by increased cartilage, lower bone volume fraction, and less bony bridging across the osteotomy gap than rigid fixation. In a control setting, SclAb negatively affected later stages of healing under rigid fixation, evidenced by the high degree of endosteal bridging at 21 days in the rigid-SclAb group compared with rigid-veh, indicating delayed fracture callus remodeling and bone marrow reconstitution. Under rigid fixation, Sost and sclerostin expression at the gene and protein level, respectively, were increased in SclAb compared with veh-treated bones, suggesting a negative feedback mechanism. Our results suggest that SclAb could be used to enhance overall bone mass but should be carefully considered in bone healing. SclAb may help to increase bone formation early in the healing process but not during advanced stages of fracture callus remodeling and not to overcome delayed healing in semirigid fixation. © 2018 American Society for Bone and Mineral Research.

Effects of Immobilizations of rhBMP-2 and/or rhPDGF-BB on Titanium Implant Surfaces on Osseointegration and Bone Regeneration

The aim of this study was to examine the effects of immobilizing rhPDGF-BB plus rhBMP-2 on heparinized-Ti implants on in vivo osseointegration and vertical bone regeneration at alveolar ridges. Successful immobilizations of rhPDGF-BB and/or rhBMP-2 onto heparinized-Ti (Hepa/Ti) were confirmed by in vitro analysis, and both growth factors were found to be sustained release. To evaluate bone regeneration, rhPDGF-BB, and/or rhBMP-2-immobilized Hepa/Ti implants were inserted into beagle dogs; implant stability quotients (ISQ), bone mineral densities, bone volumes, osseointegration, and bone formation were assessed by micro CT and histometrically. In vivo study showed that the osseointegration and bone formation were greater in the rhPDGF-BB/rhBMP-2-immobilized Hepa/Ti group than in the rhPDGF-BB-immobilized Hepa/Ti group. The rhPDGF-BB/rhBMP-2 immobilized Hepa/Ti group also showed better implant stability and greater bone volume around defect areas and intra-thread bone density (ITBD) than the rhBMP-2-immobilized Hepa/Ti group. However, no significant differences were observed between these two groups. Through these results, we conclude rhBMP-2 immobilized, heparin-grafted implants appear to offer a suitable delivery system that enhances new bone formation in defect areas around implants. However, we failed to observe the synergetic effects for the rhBMP-2 and rhPDGF-BB combination.