Effects of porcine bone marrow-derived platelet-rich plasma on bone marrow-derived mesenchymal stem cells and endothelial progenitor cells.

This study investigated the abundance of pro-regenerative growth factors in bone marrow-derived platelet-rich plasma (BM-PRP) and their effects on bone marrow-derived mesenchymal stem cells (BM-MSC) and bone marrow-derived endothelial progenitor cells (BM-EPC). Four 4-5 months-old domestic pigs were included, and each underwent bone marrow aspiration from its humerus bones and processed into bone marrow aspiration concentrate (BMAC) samples. The plasma and cellular portions of BMAC were subsequently separated and collected. The concentration of growth factors including BMP-2, PDGF-BB, TGF-ß1 and VEGF in the plasma portion was measured and compared between BM-PRP and bone marrow-derived platelet-poor plasma (BM-PPP). It was found that platelet count was significantly higher in BM-PRP than in BM-PPP, but the concentration of above-mentioned growth factors was not significantly different between BM-PRP and BM-PPP. As most existing literature has indicated the regenerative potency of PRP, this study focused on assessing the effect of BM-PRP treatment on BM-MSC and BM-EPC proliferation, osteogenic differentiation and angiogenesis capacity by comparing samples with 2.5% BM-PRP treatment and samples without BM-PRP treatment (control). In response to BM-PRP treatment, the cellular doubling time increased with culturing time and was significantly shorter in the BM-PRP-treated samples than in control samples. For osteogenic differentiation, BM-PRP-treated BM-MSCs demonstrated a time-dependent increase in alkaline phosphatase (ALP) activity and expression levels of osteogenic differentiation markers. For the expression of angiogenic genes, none of the differences reached statistical significance despite a tendency of stronger expression at day 18 in BM-PRP-treated BM-EPCs. In conclusion, this in vitro study suggests that most BMP-2, PDGF-BB, TGF-ß1 and VEGF-A contained in BM-PRP are not platelet-released and BM-PRP may have some stimulation (less than 1-fold) for MSC, EPC proliferation and MSC osteogenic differentiation.

Reliability and accuracy of assessing temporary anchorage device-tooth root contact with cone-beam computed tomography.

INTRODUCTION: This study was aimed at investigating the reliability and accuracy of cone-beam computed tomography (CBCT) diagnosis of contact between a temporary anchorage device (TAD) and tooth root and assessing any effect produced by metal brackets, imaging software program, and image segmentation or color enhancement tools. METHODS: Eighteen fresh pig mandibles were used. TADs (Vector, 1.4 × 8 mm) were placed at the buccal intermolar alveolar bone on both sides of the mandibles. With soft tissue kept intact, each mandible underwent CBCT scans (voxel size, 400 µm) before and after placing TADs, and after placing metal brackets on involved molars. Alveolar bone specimens containing the TADs were then exposed to microcomputed tomography (microCT) scans (voxel size, 27 µm) after TAD removal. Two independent raters, blinded of image identity, diagnosed TAD-root contact using ImageJ (National Institutes of Health and the Laboratory for Optical and Computational Instrumentation, University of Wisconsin, Madison, Wis) for microCT; Dolphin (Dolphin Imaging and Management Solutions, Chatsworth, Calif) and Anatomage software programs (Anatomage, Santa Clara, Calif) for CBCT images. Intrarater and interrater reliability and diagnostic accuracy were statistically assessed using Cohen kappa and McNemar tests. RESULTS: Intrarater and interrater reliability of TAD-root contact diagnoses were perfect for microCT diagnoses (κ = 1), generally moderate to good (κ >0.5) for CBCT diagnoses except for the use of color enhancement tools (κ <0.25). For diagnostic accuracy, there was generally a low agreement (κ <0.45) between CBCT and microCT (gold standard). The percent accuracy ranged from 68.1% to 79.2% and was not different among raters, bracket presence/absence, or software choices (chi-square tests, P >0.05). Overall, diagnostic sensitivity was above 80%, whereas specificity was below 55%. CONCLUSIONS: Despite good reliability, diagnoses of TAD-root contact using 400 µm voxel size CBCT imaging tend to be inaccurate, with a likelihood of high false-positive diagnoses.

Enhancement of bone marrow aspirate concentrate with local self-healing corticotomies.

Bone marrow aspirate concentrate (BMAC) is a potentially useful biological product for bone regeneration. This study investigated whether BMAC can be enriched by local minor corticotomies. Five 4-month-old domestic pigs were used with each pig undergoing two minor corticotomies at one randomly-selected tibia. Two weeks after the operation, bone marrow was aspirated from both tibiae and processed into BMAC samples. The amount of mesenchymal stem cells (MSCs) and the concentration of several regenerative growth factors contained in BMAC, as well as the proliferative and osteogenic differentiation capacity of MSCs, were compared between the corticotomy and the control sides. Another four weeks later, healing of the corticotomies was evaluated by radiographic and histological methods. The results demonstrated that BMAC from the corticotomy side contained significantly more MSCs than the control side. MSCs from the corticotomy side also proliferated significantly faster and tended to have stronger osteogenic differentiation than those from the control side. In contrast, the protein concentration of TGF-ß, BMP-2 and PDGF contained in BMAC was only minimally changed by the corticotomies. The corticotomies in all pigs healed uneventfully, showing complete obliteration of the corticotomy gaps on CT images. Comparison between the two sides showed that the corticotomy side had thicker and denser cortical bone and more abundant osteogenic cell differentiation than the control side. These findings suggest that the quantity and proliferative/osteogenic differentiation capacity of MSCs contained in local BMAC can be enhanced by minor corticotomies, and spontaneous healing of the corticotomy can be completed within 6 weeks of the operation.

Xenogeneic mesenchymal stem cell transplantation for mandibular defect regeneration.

BACKGROUND: It is commonly accepted that xenogeneic stem cell transplantation for tissue engineering is faced with host immune rejection. Using a rat critical-size mandibular defect model, this study examined whether the immune rejection can be evaded by diminishing T-cell immunity. METHODS: To examine donor cell survival and host immune reaction, pig bone marrow-derived mesenchymal stem cells (BM-MSCs) were labeled with CM-DiI, loaded onto gelatin sponge (5 × 106 cells/scaffold), and transplanted into 5-mm mandibular defects of immunocompetent and T cell-deficient athymic rats. To examine the effects of xenogeneic BM-MSCs on bone regeneration, athymic rats undergone the same surgeries were terminated at post-operative weeks 1, 3, and 6. Control rats underwent the same jaw surgery without BM-MSC transplantation. RESULTS: The density of CM-DiI-labeled BM-MSCs decreased with time in both strains of rats. Although it was substantially higher in athymic rats than in immunocompetent rats at post-operative day 1, by day 3-7 the density became comparable between the two strains of rats. Apoptosis reflected by cleaved Caspase-3 staining was low in both strains. Stronger infiltration of neutrophils, macrophages, B cells and CD8+ T cells was found in MSC-treated animals. In athymic rats, infiltration of neutrophils and macrophages was strong, but it occurred later than that in immunocompetent rats. While bone volume fraction significantly increased with time (P < .001), no difference was found between MSC-treated and control groups. CONCLUSIONS: Even in hosts with deficient T-cell immunity, xenogeneic BM-MSC transplantation into mandibular critical-sized defects still faces challenges from host innate immunity, which compromises their regenerative efficacy.

The Effect of Parathyroid Hormone Analogues When Added to Mineralized Bone Xenografts.

Implants can be a treatment option when there is sufficient quantity and quality of bone to provide support for long-term success. In the reconstruction of defects, autogenous bone remains the gold standard for its osteogenic and compatibility properties. However, the disadvantage of secondary surgery and the associated donor site morbidity prompts researchers to develop the ideal bone substitute for optimum bone reconstruction. Parathyroid hormone (PTH1-34) has provided a new option for improvement in bone regeneration. This study used a pig model to evaluate the effectiveness of parathyroid hormone when added to a xenograft, Bio-Oss, in reconstructing mandible defects. Six domestic pigs were used to create 3 posterior mandibular defects measuring 2 × 1-cm bilaterally with a total of 36 defects to simulate tooth extraction sites in humans. The defects were grafted in random order and divided into 3 groups as follows: control (no graft), Bio-Oss without PTH, and Bio-Oss with PTH. Defects were assessed with cone beam computerized tomography (CBCT), micro computerized tomography (microCT), nanoindentation, and histology. Results showed that adding PTH1-34 significantly enhanced the graft construct. CBCT showed a significant increase in the degree of bone mineralization. Nanoindentation showed increased hardness of regenerated bone and accelerated bone mineralization with PTH. MicroCT analysis revealed a trend toward higher bone regeneration and mineralization. The histological analysis showed a positive trend of the increase in cortical bone thickness and mineral apposition rate. In conclusion, the local addition of PTH1-34 to a xenograft has shown promising results to enhance bone regeneration in the reconstruction of mandibular defects.

Enhance Mandibular Symphyseal Surface Bone Growth with Autologous Mesenchymal Stem Cell Sheets: An Animal Study.

INTRODUCTION: The size and shape of the chin strongly influence facial profile and harmony. The current correction of chin deficiency mostly relies on genioplasty surgery involving osteotomy. To avoid osteotomy, one possible alternative is to enhance bone growth at the mental protuberance area with cell sheet transplantation. This study was undertaken to evaluate the efficacy of this approach in a pig model. MATERIALS AND METHODS: Five 4-month-old pigs were included for mandibular bone marrow aspiration and MSC isolation. Triple-layer MSC sheets were then fabricated and utilized using culture-expanded MSCs. Four weeks after bone marrow aspiration, subperiosteal pockets were created on the labial symphyseal surface, followed by transplantation of autogenous MSC sheets to one randomly chosen side with the other side (control) receiving no transplantation. Six weeks after the surgery, the pigs were euthanized and the specimens from both sides were collected for computed tomography (CT) and histological and immunohistochemical analysis. Measurements between the experimental and control sides were compared using paired t tests. RESULTS: MSC sheet fabrication and transplantation were reliably conducted. The labial cortical bone thickness increased significantly with MSC sheet transplantation by an average of 2 mm (p = 0.0001). The average measurements of mineral apposition rate and cell proliferation at the cell sheet side tended to be higher than the control side although the differences did not reach statistical significance (p = 0.1-0.2). Tissue mineral density measurements from CT images and bone volume fraction (BV/TV) measurements from histologic images were identical between the two sides (p > 0.5). CONCLUSION: These data provide a proof of concept that autologous MSC sheets may be transplanted to the subperiosteal region of the mandibular symphysis to stimulate local surface bone growth. NO LEVEL ASSIGNED: This journal requires that authors assign a level of evidence to each submission to which Evidence-Based Medicine rankings are applicable. This excludes Review Articles, Book Reviews, and manuscripts that concern Basic Science, Animal Studies, Cadaver Studies, and Experimental Studies. For a full description of these Evidence-Based Medicine ratings, please refer to the Table of Contents or the online Instructions to Authors www.springer.com/00266.

The usefulness of cone-beam computed tomography gray values for alveolar bone linear measurements.

OBJECTIVES: To test a proof-of-concept that the accuracy and reliability of alveolar bone height measurements from orthodontic grade (large field-of-view [FOV], large voxel-size) cone-beam computed tomography (CBCT) images may be improved by using pixel gray values. MATERIALS AND METHODS: Twenty fresh cadaver pig heads underwent CBCT scans (17 × 23 cm FOV, 0.4-mm voxel size). Buccal alveolar bone heights of maxillary first molars were measured using the conventional vision-based (VB) and the proposed gray value-assisted (GVA) methods. The GVA methods entailed localization of landmarks through observation of gray value pattern changes across tissue boundaries followed by mathematical calculation of distances between landmark pixels. Interrater reliability and accuracy of CBCT measurements made by all methods were statistically analyzed by comparing with physical measurements (gold standards). RESULTS: The interrater reliability of CBCT measurements made by GVA methods was comparable to physical measurements but higher than those made by the VB method. The GVA (bend-down pattern) method yielded average measurements similar to physical measurements, while those obtained by the VB and the GVA (straight pattern) methods were significantly larger (repeated measures analysis of variance, P < .001). The GVA (bend-down pattern) method also produced significantly more measurements within one voxel size of physical measurements than did the VB and GVA (straight pattern) methods (Chi-square tests, P < .017). CONCLUSIONS: These data confirm a concept that local gray value change patterns may be used to improve the accuracy and reliability of alveolar bone height measurement from large FOV and large voxel-size CBCT images.

Similarities and differences between porcine mandibular and limb bone marrow mesenchymal stem cells.

OBJECTIVE: Research has shown promise of using bone marrow mesenchymal stem cells (BMSCs) for craniofacial bone regeneration; yet little is known about the differences of BMSCs from limb and craniofacial bones. This study compared pig mandibular and tibia BMSCs for their in vitro proliferation, osteogenic differentiation properties and gene expression. DESIGN: Bone marrow was aspirated from the tibia and mandible of 3-4 month-old pigs (n=4), followed by BMSC isolation, culture-expansion and characterization by flow cytometry. Proliferation rates were assessed using population doubling times. Osteogenic differentiation was evaluated by alkaline phosphatase activity. Affymetrix porcine microarray was used to compare gene expressions of tibial and mandibular BMSCs, followed by real-time RT-PCR evaluation of certain genes. RESULTS: Our results showed that BMSCs from both locations expressed MSC markers but not hematopoietic markers. The proliferation and osteogenic differentiation potential of mandibular BMSCs were significantly stronger than those of tibial BMSCs. Microarray analysis identified 404 highly abundant genes, out of which 334 genes were matched between the two locations and annotated into the same functional groups including osteogenesis and angiogenesis, while 70 genes were mismatched and annotated into different functional groups. In addition, 48 genes were differentially expressed by at least 1.5-fold difference between the two locations, including higher expression of cranial neural crest-related gene BMP-4 in mandibular BMSCs, which was confirmed by real-time RT-PCR. CONCLUSIONS: Altogether, these data indicate that despite strong similarities in gene expression between mandibular and tibial BMSCs, mandibular BMSCs express some genes differently than tibial BMSCs and have a phenotypic profile that may make them advantageous for craniofacial bone regeneration.

Comparison of cone-beam computed tomography with multislice computed tomography in detection of small osseous condylar defects.

INTRODUCTION: Previous studies have indicated that orthodontic-grade cone-beam computed tomography (CBCT) images are limited when displaying small defects at the mandibular condyles for diagnosis. In this study, we investigated whether this limitation was inherent to CBCT by comparing CBCT with multislice computed tomography (CT), and whether image segmentation and color mapping could overcome this limitation. METHODS: Nine fresh pig heads (18 condyles, 36 medial and lateral condylar regions) were used. Small osseous defects (diameter and depth, 1.5 mm) were created at the medial and lateral regions of the condyles shown by gutta percha markers. After the overlying soft tissues were restored, the pig heads underwent orthodontic-grade CBCT scans (0.4-mm voxel size; i-CAT; Imaging Sciences International, Hatfield, Pa) and medical-grade CT scans (0.625-mm voxel size; LightSpeed; GE, Little Chalfont, Buckinghamshire, United Kingdom). Subsequently, 2 calibrated and blinded raters diagnosed the defect numbers in each condylar region from CBCT and CT images using Dolphin 3D software (Patterson Supply, St Paul, Minn) without image segmentation, and then 1 week later with the proprietary image segmentation and color mapping tools of Dolphin 3D. Condylar polyvinyl siloxane impressions were collected and evaluated by the same raters to obtain physical diagnoses. Rediagnoses were made on randomly selected subsamples to assess reliability. Using the physical diagnoses as references, the accuracy of imaging diagnosis was assessed and statistically compared among the varied imaging and analysis methods. RESULTS: Image diagnoses of all imaging and analysis methods showed good or excellent intrarater and interrater reliability values, except for those of the segmented CBCT images, which were substantially lower. The numbers of overdiagnoses and underdiagnoses per condylar region were not significantly different among the varied imaging and analysis methods (Wilcoxon tests, P >0.05), but classification functions demonstrated substantially lower sensitivity and accuracy with CBCT than with CT. Logistic regression also showed that CT had a significantly higher probability (odds ratio, 2.4) than CBCT in reaching the correct diagnosis, whereas use of the image segmentation and color mapping tool proprietary to Dolphin 3D did not improve the diagnostic accuracy from CBCT images. CONCLUSIONS: Even at a lower voxel size than medical CT images, orthodontic-grade CBCT images of mandibular condyles may be inherently less reliable and less accurate for the diagnosis of small condylar defects.

Reconstructing jaw defects with MSCs and PLGA-encapsulated growth factors.

Cell and growth factor-based tissue engineering has shown great potentials for skeletal regeneration. This study tested its feasibility in reconstructing large mandibular defects and compared the efficacy of varied construction materials and sealing methods. Bilateral mandibular critical-size (5-cm(3)) defects were created on six 4-month-old domestic pigs, and grafted with ß-tricalcium phosphate (ßTCP) only (Group-A), ßTCP with autologous bone marrow-derived mesenchymal stem cells (BM-MSCs) (Group-B), and ßTCP with BM-MSCs and biodegradable poly(lactic-co-glycolic acid) (PLGA) microspheres containing bone morphogenetic protein-2 (BMP-2) and vascular endothelial growth factor (VEGF) (Group-C). The buccal sides of Groups-B/-C were either sealed by fibrin sealant or by a biodegradable PLGA barrier membrane before soft-tissue closure. Computed tomography (CT), microCT and histology analyses were performed 12 weeks postoperatively. In vitro data demonstrated that BM-MSCs, with MSC properties confirmed, remained vital after integration with ßTCP; and PLGA microspheres exhibited an initial burst followed by slow and continuous release of growth factors over a period of 28 days. In vivo data demonstrated that Group-B/-C sites had significantly greater gap obliteration, higher tissue mineral densities and more residual ßTCP granules (p<0.05, Kruskal-Wallis tests). Qualitatively, Group-B/-C defect sites had started remodeling while Group-A sites were mainly forming new bone to bridge the gaps. Furthermore, ßTCP degradation was not mediated by macrophages or osteoclasts, and was significantly slowed down by sealing the defects with barrier membrane. Combined, these data present a promising formulation composed of ßTCP granules, autologous MSCs, controlled-release growth factors and biodegradable PLGA barrier membrane for the reconstruction of critical-size mandibular defects.

Establishing a critical-size mandibular defect model in growing pigs: characterization of spontaneous healing.

PURPOSE: A large animal model is desired for preclinical studies aimed at reconstructing severe mandibular skeletal defects using tissue engineering techniques. To identify the size and location requirements for a mandibular critical-size bone defect in growing pigs, the present study investigated the spontaneous healing of surgically created mandibular defects. MATERIALS AND METHODS: Six 4-month-old domestic pigs were used. In pigs 1 and 2, a 3-, 5-, or 7-cm(3) subperiosteal mandibular defect was created. In pigs 3 to 6, 3- to 5-cm(3) bilateral defects were randomly created at the anterior (apical to the molars) and posterior (mandibular angle) mandibular regions. Spontaneous healing of these defects was assessed by serial computed tomography scans (postoperative week 1, 6, and 12) and histologic analyses. RESULTS: In pigs 1 and 2, regardless of defect size, the anterior, but not posterior, defects had largely healed. Systematic analyses of pigs 3 to 6 revealed, first, the extent of defect regeneration from spontaneous healing was significantly less in the posterior than in the anterior defects, with about two thirds and one third of the original defect volume remaining, respectively. Second, histologically, the posterior defects had considerably less regeneration and more evident tapering of the new bone than did the anterior defects. Finally, the buccal periosteum had completely regenerated in the anterior defects, but had only partially done so in the posterior defects. Also, the buccal surface contour was moderately concave in the anterior defects, but it was severely concave in the posterior defects. CONCLUSIONS: Despite robust spontaneous healing of mandibular defects in growing pigs, 5-cm(3) defects in the mandibular angle region without buccal periosteum would be a reasonable critical-size defect model relevant to mandibular defects in adolescent humans.

Evaluation of cone-beam computed tomography in the diagnosis of simulated small osseous defects in the mandibular condyle.

INTRODUCTION: In this study, we investigated the impact of defect size and scan voxel size on the accuracy of cone-beam computed tomography (CBCT) diagnoses of simulated condylar defects and assessed the value of orthodontic CBCT images typically scanned at lower settings (0.4-mm voxel size and full-size field of view) in diagnosing condylar erosion defects. METHODS: Cylindrical holes simulating condylar defects with varied diameters (≤2, 2-3, and >3 mm) and depths (≤2 and >2 mm) were created in 22 fresh pig mandibular condyles, with defect number and size per condyle and quadrant randomly determined. With the soft tissues repositioned, 2 CBCT scans (voxel sizes, 0.4 and 0.2 mm) of the pig heads were obtained from an i-CAT unit (Imaging Science International, Hatfield, Pa). Reconstructed CBCT data were analyzed independently by 2 calibrated, blinded raters using Dolphin-3D (Dolphin Imaging and Management Solutions, Chatsworth, Calif) for defect identification and localization and defect diameter and depth measurements, which were compared with physical diagnoses obtained from polyvinyl siloxane impressions. RESULTS: Identification and localization of simulated defects demonstrated moderate interrater reliability and excellent specificity and sensitivity, except for extremely small defects (both diameter and depth ≤2 mm) viewed with 0.4-mm scans, which had a significantly lower sensitivity (67.3%). Geometric measurements of simulated defects demonstrated good but not excellent interrater reliability and submillimeter inaccuracy for all defects. Receiver operating characteristic analyses demonstrated that the overall accuracy of diagnosing simulated condylar defects based on CBCT geometric measurements was fair and good for the 0.4-mm and 0.2-mm voxel-size scans, respectively. With the prevalence of condylar erosion defects in the patients considered, the positive predictive values of diagnoses based on 0.5-mm size (diameter or depth) cutoff points were near 15% and 50% for asymptomatic and symptomatic temporomandibular joints, respectively; the negative predictive values were near 95% and 90%, respectively. CONCLUSIONS: When using orthodontic CBCT images for diagnosing condylar osseous defects, extremely small (<2 mm) defects can be difficult to detect; caution is also needed for the diagnostic accuracy of positive diagnoses, especially those from asymptomatic temporomandibular joints.

Scaffold-based delivery of autologous mesenchymal stem cells for mandibular distraction osteogenesis: preliminary studies in a porcine model.

PURPOSE: Bone regeneration through distraction osteogenesis (DO) is promising but remarkably slow. To accelerate it, autologous mesenchymal stem cells have been directly injected to the distraction site in a few recent studies. Compared to direct injection, a scaffold-based method can provide earlier cell delivery with potentially better controlled cell distribution and retention. This pilot project investigated a scaffold-based cell-delivery approach in a porcine mandibular DO model. MATERIALS AND METHODS: Eleven adolescent domestic pigs were used for two major sets of studies. The in-vitro set established methodologies to: aspirate bone marrow from the tibia; isolate, characterize and expand bone marrow-derived mesenchymal stem cells (BM-MSCs); enhance BM-MSC osteogenic differentiation using FGF-2; and confirm cell integration with a gelatin-based Gelfoam scaffold. The in-vivo set transplanted autologous stem cells into the mandibular distraction sites using Gelfoam scaffolds; completed a standard DO-course and assessed bone regeneration by macroscopic, radiographic and histological methods. Repeated-measure ANOVAs and t-tests were used for statistical analyses. RESULTS: From aspirated bone marrow, multi-potent, heterogeneous BM-MSCs purified from hematopoietic stem cell contamination were obtained. FGF-2 significantly enhanced pig BM-MSC osteogenic differentiation and proliferation, with 5 ng/ml determined as the optimal dosage. Pig BM-MSCs integrated readily with Gelfoam and maintained viability and proliferative ability. After integration with Gelfoam scaffolds, 2.4-5.8×10(7) autologous BM-MSCs (undifferentiated or differentiated) were transplanted to each experimental DO site. Among 8 evaluable DO sites included in the final analyses, the experimental DO sites demonstrated less interfragmentary mobility, more advanced gap obliteration, higher mineral content and faster mineral apposition than the control sites, and all transplanted scaffolds were completely degraded. CONCLUSION: It is technically feasible and biologically sound to deliver autologous BM-MSCs to the distraction site immediately after osteotomy using a Gelfoam scaffold to enhance mandibular DO.

Bone ingrowth and initial stability of titanium and porous tantalum dental implants: a pilot canine study.

PURPOSE: To investigate if a dental implant system with a midsection covered by 3-dimensionally porous tantalum material would exhibit stability comparable with a traditional threaded titanium alloy implant system and whether bone would grow into the porous section. METHODS: Three experimental and 3 control implants were placed in the individual mandibles of 8 dogs. Resonance frequency analysis assessed implant stability at 0, 2, 4, 8, and 12 weeks of healing. Histomorphometric and backscattered scanning electron microscopic analyses examined the presence of bone ingrowth into the experimental implant's porous section and bone-to-implant contact along the titanium surfaces of both implants. RESULTS: Implant stability did not significantly differ during 0 to 12 weeks of healing. Progressive tissue mineralization developed inside porous sections from weeks 2 to 12. Porous implants exhibited a combination of progressive osseointegration along their titanium surfaces and bone ingrowth inside their porous tantalum sections. CONCLUSIONS: Cortical and apical implant threads, combined with the porous section, were able to stabilize the experimental implant to the same degree as the fully threaded control implant.

Factors affecting the accuracy of buccal alveolar bone height measurements from cone-beam computed tomography images.

INTRODUCTION: The reasons for inaccuracies in alveolar bone measurement from cone-beam computed tomography (CBCT) images might be multifactorial. In this study, we investigated the impact of software, the presence or absence of soft tissues, the voxel size of the scan, and the regions in the jaws on buccal alveolar bone height measurements in pigs at an age equivalent to human adolescents. METHODS: Marker holes, apical to the maxillary and mandibular molar roots, and mesiodistal molar occlusal reference grooves were created in 6 fresh pig heads (12 for each jaw), followed by CBCT scans at 0.4-mm and 0.2-mm voxel sizes under soft-tissue presence and soft-tissue absence conditions. Subsequently, buccolingual sections bisecting the marker holes were cut, from which the physical alveolar bone height and thickness were measured. One blinded rater, using Dolphin (version 11.5 Premium; Dolphin Imaging, Chatsworth, Calif) and OsiriX (version 3.9; www.osirix-viewer.com) software, independently collected alveolar bone height measurements from the CBCT images. Differences between the CBCT and the physical measurements were calculated. The mean differences and the limit of agreement (LOA, ±1.96 SD) for every jaw, voxel-size, soft-tissue, and software condition were depicted. Each measurement was then assessed for clinical inaccuracy by using 2 levels of criteria (absolute differences between CBCT and physical measurements ≥1 mm, or absolute differences between CBCT and physical measurements ≥0.5 mm), and the interactions between soft-tissue and voxel-size factors for every jaw and software condition were assessed by chi-square tests. RESULTS: Overall, the mean differences between the CBCT and the physical measurements for every jaw, voxel-size, soft-tissue, and software condition were near 0. With all other conditions kept equal, the accuracy of the maxillary CBCT measurements was inferior (larger limit of agreement ranges and higher frequencies of clinical inaccuracy) to the mandibular measurements. The physical thickness of the maxillary alveolar crestal bone was less than 1 mm and significantly thinner than the mandibular counterparts. For every jaw and software condition, the accuracy of measurements from the 0.2-mm soft-tissue presence CBCT images was consistently superior (smaller limit of agreement ranges and lower frequencies of clinical inaccuracy) to those from the 0.4-mm soft-tissue presence, the 0.4-mm soft-tissue absence, and the 0.2-mm soft-tissue absence images; all showed similar accuracies. Qualitatively, the soft-tissue absence images demonstrated much brighter enamel and alveolar bone surface contours than did the soft-tissue presence images. CONCLUSIONS: At an adolescent age, the buccal alveolar bone height measured from the maxillary molar region based on 0.4-mm voxel-size CBCT images can have relatively large and frequently inaccurate measurements, possibly due to its thinness. By using 0.2-mm voxel-size scans, measurement accuracy might be improved, but only when the overlying facial and gingival tissues are kept intact.

Alveolar ridge reduction after tooth extraction in adolescents: an animal study.

OBJECTIVE: The mechanism for tooth extraction induced residual alveolar ridge reduction (RRR) during adolescence is poorly understood. This study investigated the alveolar bone morphology, growth, resorption and functional loading at normal and extraction sites using an adolescent pig model. DESIGN: Sixteen 3-month-old pigs were divided into two groups - immediate post-extraction (IE) and 6-week post-extraction (SE). The IE group received an extraction of one deciduous mandibular molar, immediately followed by a final experiment to record masseter muscle EMGs and strains from the buccal surface of the extraction and contralateral non-extraction sites during function (mastication). The SE group was given the same tooth extraction, then kept for 6 weeks before the same final functional recording as the IE group. Both groups also received baseline (pre-extraction) EMGs and fluorescent vital stains 10 and 3 days before the final functional recording. Immediately after the final functional recording, animals were euthanized and alveolar bone specimens from extraction and contralateral non-extraction sites were collected and used to analyse alveolar bone morphology, apposition and resorption based on fluorescent and hematoxylin and eosin stained histological sections. RESULTS: At control sites (IE-extraction, IE-non-extraction and SE-non-extraction), the alveolar ridges grew gingivally and buccally. Bone formation characterized the buccal surface and lingual bundle bone, whereas resorption characterized the lingual surface and buccal bundle bone. The SE-extraction sites showed three major alterations: convergence of the buccal and lingual gingival crests, loss of apposition on the lingual bundle bone, and decelerated growth at the entire buccal surface. These alterations likely resulted from redirected crestal growth as part of the socket healing process, loss of tongue pressure to the lingual side of the teeth which normally provides mechanical stimulation for dental arch expansion, and masticatory underloading during the initial post-extraction period, respectively. CONCLUSIONS: These data indicate that the initial phase of RRR in adolescents is a product of modified growth, not resorption, possibly because of decreased mechanical stimulation at the extraction site.

Increased variability of bone tissue mineral density resulting from estrogen deficiency influences creep behavior in a rat vertebral body.

Progressive vertebral deformation increases the fracture risk of a vertebral body in the postmenopausal patient. Many studies have observed that bone can demonstrate creep behavior, defined as continued time-dependent deformation even when mechanical loading is held constant. Creep is a characteristic of viscoelastic behavior, which is common in biological materials. We hypothesized that estrogen deficiency-dependent alteration of the mineral distribution of bone at the tissue level could influence the progressive postmenopausal vertebral deformity that is observed as the creep response at the organ level. The objective of this study was thus to examine whether the creep behavior of vertebral bone is changed by estrogen deficiency, and to determine which bone property parameters are responsible for the creep response of vertebral bone at physiological loading levels using an ovariectomized (OVX) rat model. Correlations of creep parameters with bone mineral density (BMD), tissue mineral density (TMD) and architectural parameters of both OVX and sham surgery vertebral bone were tested. As the vertebral creep was not fully recovered during the post-creep unloading period, there was substantial residual displacement for both the sham and OVX groups. A strong positive correlation between loading creep and residual displacement was found (r=0.868, p<0.001). Of the various parameters studied, TMD variability was the parameter that best predicted the creep behavior of the OVX group (p<0.038). The current results indicated that creep caused progressive, permanent reduction in vertebral height for both the sham and OVX groups. In addition, estrogen deficiency-induced active bone remodeling increased variability of trabecular TMD in the OVX group. Taken together, these results suggest that increased variability of trabecular TMD resulting from high bone turnover influences creep behavior of the OVX vertebrae.

Mechanical strain at alveolar bone and circummaxillary sutures during acute rapid palatal expansion.

INTRODUCTION: Palatal expansion can potentially affect alveolar bone and circummaxillary sutures. In this study, we characterized their mechanical strain during acute expansion. METHODS: Eight 3- and 6-month-old fresh pig heads received acute palatal expansion with hyrax expanders. Strain gauges were used to measure strain at the buccal alveolar bone of anchor and adjacent nonanchor teeth, and at maxillary-premaxillary, maxillary-zygomatic, and zygomatic-temporal sutures during expansion. Intermolar width changes were measured from dental casts. RESULTS: Intermolar width increased less than expander activation, and the midpalatal sutures were only opened slightly. Alveolar bone strain increased linearly with expander activation and decayed by 20% to 30% during postactivation intervals. Compressive strain at anchor-tooth alveolar bone locations was directed occlusally and apically, related to tooth tipping, and significantly higher than that at nonanchor tooth locations. With expander activation, suture strains increased monotonically and tended to plateau. Suture strain magnitude was generally similar to physiologic (masticatory) strains reported in the literature. The dominant strain polarity was compression at the maxillary-zygomatic and zygomatic-temporal sutures, but there was tension at the maxillary-premaxillary suture. CONCLUSIONS: In these pigs, palatal expansion can cause significant occlusal-apical compression at buccal alveolar bone and physiologic-level strains at circummaxillary sutures.

Effect of bone thickness on alveolar bone-height measurements from cone-beam computed tomography images.

INTRODUCTION: Cone-beam computed tomography (CBCT) has been used to assess alveolar bone changes after rapid palatal expansion. The purpose of this study was to investigate the accuracy of alveolar bone-height measurements from CBCT images with varied bone thicknesses and imaging resolutions. METHODS: Eleven maxillary specimens from 6-month-old pigs were measured for alveolar bone height (distance between drilled reference holes and alveolar crests) at 6 locations with a digital caliper, followed by CBCT scanning at 0.4-mm and 0.25-mm voxel sizes. Buccal alveolar bone of these locations was then reduced approximately by 0.5 to 1.5 mm, followed by CBCT rescanning with the same voxel sizes. The CBCT images were measured by using 3-dimensional software to determine alveolar bone height and thickness in buccolingual slices by independent, blinded raters. The specimens were subsequently cut into buccolingual sections at reference-hole levels, and direct bone height and thickness were measured from these sections. Intrarater and interrater repeatability and the differences between CBCT and direct measurements were assessed. RESULTS: Excellent intrarater (intraclass correlations, r = 0.89-0.98) and good interrater (r = 0.64-0.90) repeatability values were found for alveolar bone-height measurements from the CBCT images. Before alveolar bone reduction, the thickness was much greater than the CBCT voxel size (0.4 mm), and bone-height measurements from the CBCT images were 0.5 to 1 mm more than the direct measurements (paired t tests, P <0.017 at most locations). After bone reduction, the thickness at the subcrest 1-mm level was near or below the CBCT voxel size (0.4 mm), and bone-height measurements from the CBCT images were 0.9 to 1.2 mm less than the direct measurements (paired t tests, P <0.017 at most locations). These measurement inaccuracies were substantially improved by decreasing the CBCT voxel size to 0.25 mm. CONCLUSIONS: Alveolar bone-height measurements from conventional clinical 0.4-mm voxel size CBCT images might overestimate alveolar bone-height loss associated with rapid palatal expansion.

Molecular variations related to the regional differences in periosteal growth at the mandibular ramus.

Periosteal growth at human mandibular ramus is characterized by bone apposition at the posterior border and resorption at the anterior border. Molecular control of this regional variation is unclear. This study examined the expression of several molecules involved in bone apposition/resorption at these regions in vivo and in vitro. By using growing pigs as a model, the periosteal growth was assessed at the mandibular ramus by vital staining and histological observations. In parallel, periosteal tissues were harvested and pulverized for RNA and protein extraction. Periosteal cells were also isolated, expanded in osteogenic media, and subjected to a single dose of dynamic tensile strain (0, 5, or 10% magnitude at 0.5 Hz) to examine their responses to mechanical loading. Real-time RT-PCR and Western blot analyses were used to examine mRNA and protein expression from periosteal tissues and cultured cells. Histological observation confirmed an anterior-resorption/posterior-apposition pattern in the pig mandibular ramus. Both in vivo tissue and in vitro cells demonstrated greater mRNA expression of receptor activator of NF-κB ligand (RANKL)/osteoprotegerin (OPG) ratio and bone morphogenetic protein 2 (BMP2) at the anterior region, while OPG expression at the anterior region was lower than the posterior region. In response to the application of a single dose of dynamic tensile strain, cultured periosteal cells appeared to change the expression profile of osteogenic markers but not that of RANKL/OPG and BMP2. These findings suggest that the unique regional variation of periosteal activity at the mandibular ramus is regulated by a differential expression of RANKL/OPG ratio (likely through differential induction of OPG) and BMP2.