Spatial and temporal localization of FGF-2 and VEGF in healing tooth extraction sockets in a rabbit model.

PURPOSE: The purpose of this study was to test the hypothesis that spatial and temporal localization of growth factors FGF-2 and VEGF in a rabbit tooth extraction socket model correlate with the histologic events of healing. MATERIALS AND METHODS: Twenty-four male New Zealand white rabbits divided into 8 groups of 3 were used in the study. Incisor teeth were extracted from both jaws and the healing extraction socket with surrounding jaw bone was harvested at 48 hours, 4 days, 1, 2, 4, 8, 12, and 16 weeks. Tissues were fixed, decalcified, and processed for hematoxylin-eosin and immunohistochemical staining. The sections were stained to detect FGF-2 and VEGF. The stained sections were then imaged and an automated computer program was used to detect the brown diaminobenzidine stain that represented the growth factors of interest. Data was obtained in the form of percentage area and intensity of stain and analyzed using the analysis of variance (ANOVA - Tukey Kramer and Scheffe's post-test). RESULTS: Spatial and temporal differences in localization of FGF-2 and VEGF were observed across all time frames in both jaws. Statistically significant differences in percentage area and intensity of brown diaminobenzidine stain were seen temporally between FGF-2 and VEGF (P < .05). CONCLUSION: The results of this study showed positive correlation of histologic events to spatial and temporal localization of FGF-2 and VEGF in a rabbit tooth extraction model.

Prototype testing for a new bioabsorbable Le Fort III distraction device: a pilot study.

PURPOSE: The goal of the study was to design and test prototypes for a new, internal resorbable Le Fort III distractor in the minipig model. MATERIALS AND METHODS: Le Fort III osteotomies were performed on 3 juvenile minipigs, followed by implantation of the bioabsorbable distractors. After a 5-day latency period, the distraction progressed at a rate of 1 mm/day in 2 equal increments for 12 days. This was followed by a 6-week consolidation phase. Cephalometric radiographs were taken postoperatively, on completion of distraction and after the consolidation phase, to evaluate the amount of distraction that was obtained. Necropsy was performed at the completion of each experiment. RESULTS: The first minipig was sacrificed at the end of the distraction period because of device failure. On necropsy, it was noted that the anterior footplate on either side had failed. The second minipig was sacrificed during the latency period because of severe pneumonia. On necropsy, it was noted that none of the footplates had failed but that the distraction barrels on either side had become dislodged from the anterior bioabsorbable footplates. The third minipig was sacrificed at the end of the distraction period because of device failure. On necropsy, it was noted that the posterior bioabsorbable footplates on either side were fractured across the posterior screw lines. CONCLUSIONS: The bioabsorbable devices evaluated in this study could not withstand the forces generated during distraction osteogenesis in the minipig model. Experience with the prototypes may help in designing internal bioresorbable Le Fort III distraction devices.

Biomechanical evaluation of the pins of a mandibular external distractor.

PURPOSE: Our goal was to establish whether the pins of an external distractor were capable of overcoming tissue resistance to distraction. MATERIALS AND METHODS: The study was carried out in 2 parts. The first part of the study determined the bending rigidity of the distractors pins. To accomplish this, the distractor was installed on the mandible of a stereolithographic model using 4 regular 2.0-mm steel pins. An osteotomy was not performed. The distractor was activated using a torque gauge, and the bending rigidity of the pins was recorded. The second part of the study determined the tissue resistance to mandibular distraction using fresh cadavers. Six cadavers were divided into 2 groups to determine tissue resistance to angular and linear distraction, respectively. The devices used to measure tissue resistance were a modified external multiplanar distractor and a torque gauge. RESULTS: In the anteroposterior direction, the tissue resistance to linear distraction clearly exceeded the bending rigidity of the pins for the first 7.5 mm of activation. After this, the opposite was true. In the vertical direction, the tissue resistance clearly exceeded the bending rigidity of the pins for the first 8.0 mm of activation. After this, the opposite was true. For the first 15.0 degrees of angular distraction in the sagittal plane, the tissue resistance was almost identical to the bending rigidity of the pins. After this, the tissue resistance significantly exceeded the bending rigidity of the pins. For the first 7.0 degrees of angular distraction in the transverse plane, the tissue resistance was marginally greater than the bending rigidity of the pins. After this, the bending rigidity of the pins increased significantly. CONCLUSION: The 2.0-mm steel pins used in most external multiplanar distractors are not capable of overcoming the tissue resistance to linear or angular distraction.

Spatial and temporal localization of secretory IgA in healing tooth extraction sockets in a rabbit model.

PURPOSE: The purpose of this study was to look at the spatial and temporal localization of secretory IgA in healing tooth extraction sockets in a rabbit model. MATERIALS AND METHODS: Twenty-four male New Zealand White rabbits were used in the study. Incisor teeth were extracted from both jaws, and the healing extraction socket with the surrounding jaw bone was harvested at 48 hours, 4 days, and 1, 2, 4, 8, 12, and 16 weeks. Tissues were fixed, decalcified, and processed for hematoxylin and eosin and immunohistochemical staining. The sections were stained to detect secretory IgA. The stained sections were then imaged, and an automated computer program was used to detect the brown 3,3'-diaminobenzidine stain that represented the secretory IgA. The data were obtained in the form of percentage area and intensity of stain and analyzed using analysis of variance (Tukey-Kramer and Scheffé's tests). RESULTS: Spatial and temporal differences in localization of secretory IgA were observed across time frames in both jaws. CONCLUSION: The results of this study showed definite trends in the spatial and temporal localization of secretory IgA in healing tooth extraction sockets in a rabbit model.

Effect of biomaterial properties on bone healing in a rabbit tooth extraction socket model.

In this work we sought to understand the effect of biomaterial properties upon healing bone tissue. We hypothesized that a hydrophilic polymer gel implanted into a bone tissue defect would impede the healing process owing to the biomaterial's prevention of protein adsorption and thus cell adhesion. To test this hypothesis, healing bone was investigated within a rabbit incisor extraction socket, a subcritical size bone defect that resists significant soft tissue invasion by virtue of its conformity. After removal of the incisor teeth, one tooth socket was left as an empty control, one was filled with crosslinked polymer networks formed from the hydrophobic polymer poly(propylene fumarate) (PPF), and one was filled with a hydrogel formed from the hydrophilic oligomer oligo(poly(ethylene glycol) fumarate) (OPF). At five different times (4 days as well as 1, 2, 4, and 8 weeks), jaw bone specimens containing the tooth sockets were removed. We analyzed bone healing by histomorphometrical analysis of hematoxylin and eosin stained sections as well as immunohistochemically stained sections. The proposed hypothesis, that a hydrophilic material would hinder bone healing, was supported by the histomorphometrical results. In addition, the immunohistochemical results reflect molecular signaling indicative of the early invasion of platelets, the vascularization of wound-healing tissue, the differentiation of migrating progenitor cells, and the formation and remodeling of bone tissue. Finally, the results emphasize the need to consider biomaterial properties and their differing effects upon endogenous growth factors, and thus bone healing, during the development of tissue engineering devices.

Spatial and temporal localization of transforming growth factor-beta1, bone morphogenetic protein-2, and platelet-derived growth factor-A in healing tooth extraction sockets in a rabbit model.

PURPOSE: The purpose of this study was to test the hypothesis that spatial and temporal localization of growth factors transforming growth factor (TGF)-beta1, bone morphogenetic protein (BMP)-2, and platelet derived growth factor (PDGF)-A in a rabbit tooth extraction model correlate with the histologic events contributing toward healing. MATERIALS AND METHODS: Twenty-four male New Zealand White rabbits were used in the study. Incisor teeth were extracted from both jaws, and the healing extraction socket with the surrounding jaw bone was harvested at 48 hours, 4 days, and 1, 2, 4, 8, 12, and 16 weeks. Tissues were fixed, decalcified, and processed for hematoxylin and eosin and immunohistochemical staining. The sections were stained to detect TGF-beta1, BMP-2, and PDGF-A. Stained sections were then imaged, and an automated computer program was used to detect the brown 3,3'-diaminobenzidine regions representing the location of growth factors of interest. The data were collected in terms of percentage area and intensity of stain, and an analysis of variance was conducted (Tukey-Kramer and Scheffe's test) for statistical comparison between different time points, jaws, and growth factors. These results were also compared with hematoxylin and eosin-stained histologic specimens obtained at similar time points. RESULTS: Spatial and temporal differences in localization of TGF-beta1, BMP-2, and PDGF-A were observed across all time frames in both jaws. Statistically significant differences in percentage area and intensity of brown diaminobenzidine stain were detected temporally between TGF-beta1, BMP-2, and PDGF-A (P < or =.0001). CONCLUSION: The results of this study showed positive correlation between histologic events and the spatial and temporal localization of TGF-beta1, BMP-2, and PDGF-A in a rabbit tooth extraction model.

Automated selection of DAB-labeled tissue for immunohistochemical quantification.

The increased use of immunohistochemistry (IHC) in both clinical and basic research settings has led to the development of techniques for acquiring quantitative information from immunostains. Staining correlates with absolute protein levels and has been investigated as a clinical tool for patient diagnosis and prognosis. For these reasons, automated imaging methods have been developed in an attempt to standardize IHC analysis. We propose a novel imaging technique in which brightfield images of diaminobenzidene (DAB)-labeled antigens are converted to normalized blue images, allowing automated identification of positively stained tissue. A statistical analysis compared our method with seven previously published imaging techniques by measuring each one's agreement with manual analysis by two observers. Eighteen DAB-stained images showing a range of protein levels were used. Accuracy was assessed by calculating the percentage of pixels misclassified using each technique compared with a manual standard. Bland-Altman analysis was then used to show the extent to which misclassification affected staining quantification. Many of the techniques were inconsistent in classifying DAB staining due to background interference, but our method was statistically the most accurate and consistent across all staining levels.

Aesthetic and functional outcomes using osseous or soft-tissue free flaps.

The authors compared functional and aesthetic outcomes following reconstruction of the posterior mandible (i.e., unilateral defects distal to the mental foramen) with free flaps containing vascularized bone (VBFs) or only soft-tissue (STFs). Fifty-eight reconstructions of the posterior mandible were performed with STFs (n=32) or VBFs (n=26) and were available for review. All patients (n=58) completed a functional outcome questionnaire containing questions about diet, oral competence, pain, trismus, speech, cosmesis, social reintegration, and return to work. Computer-assisted image analysis was performed comparing facial proportions of patients with STFs (n=10), VBFs (n=11), or unoperated controls (n=10). VBFs yielded statistically significantly superior functional scores compared to STFs in 5/12 categories. Normal scores were found for diet in 31 percent (10/32) vs. 21 percent (3/26) ( p=0.006); oral competence in 56 percent (18/32) vs. 35 percent (9/26) ( p=0.05); and speech in 47 percent (15/32) vs. 30 percent (7/26) ( p=0.026). VBF patients returned to public dining (72 percent, 23/32) more frequently than STF patients (35 percent, 9/26), ( p=0.003). VBFs yielded superior results for midline symmetry with a mean mandible deviation of 3.3+/-2.3 mm vs. 5.8+/-3.6 mm (STFs), and 1.2+/-0.7 mm (controls, p=0.001). An objective method for the evaluation of aesthetic and functional outcomes in posterior mandible reconstruction was developed. This method demonstrated that posterior mandible reconstruction with VBFs is associated with superior results in speech, diet, and midline symmetry. However, rigorous analyses of oropharyngeal soft-tissue defects and the radiation therapy protocols utilized, were not performed in this study. Without these additional analyses, one cannot definitively conclude that VBFs are superior to STFs.