Enhanced alveolar ridge preservation with hyaluronic acid-enriched allografts: a comparative study of granular allografts with and without hyaluronic acid addition.

PURPOSE: Ridge preservation is essential to restore alveolar ridge volume and to enhance esthetic and functional outcomes for dental implants. The addition of hyaluronic acid to allogeneic bone substitute materials might enhance these outcomes. This clinical study evaluated the efficacy of ridge preservation after tooth extraction using granular allografts with and without hyaluronic acid addition. METHODS: In this retrospective study, 40 patients with compromised extraction sockets were enrolled. Among them, 19 received particulate allogeneic bone substitutes (Allo), 21 received allogeneic bone substitutes with hyaluronic acid (AlloHya). Vertical and horizontal graft stability, graft shrinkage rate, and bone mineral density were assessed using radiographic measurements on CBCT scans conducted before tooth extraction, directly after ridge preservation and after four months. Patients were followed up 12 months post-implantation. RESULTS: Vertical height loss after 4 months was significantly greater in the Allo group (-0.82 ± 0.95 mm) compared to the AlloHya group (-0.19 ± 0.51 mm; p = 0.011). Graft shrinkage rate was 16.9 ± 11.5% (Allo) and 10.3 ± 7.7% (AlloHya) (p = 0.038). After four months, average bone density was significantly higher in the AlloHya compared to the Allo group (p = 0.004). Nearly all implants (39 out of 40) were classified as "Success" according to the ICOI scheme, with no differences in implant quality between the two study groups. CONCLUSIONS: Improved graft stability, reduced resorption, and increased bone density were observed in hyaluronic acid-enriched allografts compared to pure allografts. Adding hyaluronic acid to allogeneic bone grafts significantly enhanced outcomes in ridge preservation.

First Clinical Case Report of a Xenograft-Allograft Combination for Alveolar Ridge Augmentation Using a Bovine Bone Substitute Material with Hyaluronate (Cerabone® Plus) Combined with Allogeneic Bone Granules (Maxgraft®).

BACKGROUND: A patient had lost the first left maxillary incisor in the esthetic zone. METHODS: The defect in the alveolar ridge was reconstructed for an implant-supported restoration using a new xenogeneic bone substitute containing hyaluronate, which was used in combination with allogeneic bone granules. RESULTS: After three years of follow-up, the dental implant was stable and showed no signs of infection. CONCLUSIONS: This is the first case report with a long-term follow-up time of three years of a successful clinical application of a xenograft-allograft combination (cerabone® plus combined with maxgraft®) for alveolar ridge augmentation before dental implantation. Cerabone® plus offers volume stability, provides reliable and efficient structural support of the oral soft tissues in the augmented region (particularly crucial in the aesthetic zone), and preserves the alveolar ridge shape.

Risk Factors for Complications Following Staged Alveolar Ridge Augmentation and Dental Implantation: A Retrospective Evaluation of 151 Cases with Allogeneic and 70 Cases with Autogenous Bone Blocks.

Purpose: the aim of this study was to identify potential risk factors favoring complications by assessing the number and types of complications associated with allogeneic or autogenous bone blocks applied as onlay grafts for alveolar ridge augmentation prior to implantation. Methods: A retrospective chart review on the success of 151 allogeneic and 70 autogenous bone blocks in a cohort of 164 consecutive patients, who were treated over a period of 6 years by the same surgeon, was conducted. Statistical conclusions were based on ROC curves and multiple logistic regression models. Results: Complications were observed more frequently with autogenous bone blocks (14 out of 70 cases; 20%) compared to allogeneic bone blocks (12 out of 151 cases; 7.9%; p = 0.013). However, these complications were minor and did not impact the successful dental rehabilitation. In a multiple logistic regression model, the risk of a complication was increased by the use of an autogenous bone block (OR = 3.2; p = 0.027), smoking (OR = 4.8; p = 0.007), vertical augmentation above a threshold of 2.55 mm (OR = 5.0; p = 0.002), and over-contouring (OR = 15.3; p < 0.001). Conclusions: Overall, the complication rate of ridge augmentations carried out with autogenous or allogeneic bone blocks was low. Despite previous recommendations, over-contouring and a vertical augmentation above a threshold of 2.55 mm should be avoided.

Comparison of allogeneic and autogenous bone grafts for augmentation of alveolar ridge defects-A 12-month retrospective radiographic evaluation.

OBJECTIVES: The aim of this study was to compare three-dimensional alterations following the use of autogenous versus allogeneic onlay grafts for augmentation at single tooth defects. MATERIALS AND METHODS: Alveolar bone width at specific implant sites were assessed using sagittal and cross-sectional CBCT images prior grafting and at three subsequent time points. Twenty-one patients received autogenous bone blocks harvested from the retromolar region and another 21 patients received freeze-dried cancellous allogeneic bone blocks. RESULTS: The vertical and horizontal dimensions did not significantly differ between autogenous and allogeneic bone grafts at any time point. In addition, there were no statistically significant differences in graft remodeling rates between autogenous (mean shrinkage rate after 12 months: 12.5% ± 7.8%) and allogeneic onlay grafts (mean shrinkage rate after 12 months: 14.4% ± 9.8%). CONCLUSIONS: Freeze-dried cancellous allogeneic bone blocks showed equivalent volumetric shrinkage rates as autogenous bone blocks when used for treating circumscribed bone defects classified as Type-II to Type-IV according to the ITI-treatment guide categories. Therefore, it is not necessary to over-contour the alveolar ridge when using allogeneic blocks for treating single tooth defects, but to apply the same procedure as when using autogenous blocks.

Long-lived murine osteocytes are embodied by craniofacial skeleton in young and old animals whereas they decrease in number in postcranial skeletons at older ages.

BACKGROUND: Osteocytes are engaged in life-enduring processes such as bone remodelling, fracture healing or osseointegration of implants. Over age, ossification processes and regenerative capacity can greatly differ in mandible and femur. OBJECTIVE: Mesenchymal stem cells from cranial and postcranial bones are of different embryologic origin. This may be the reason why the regenerative capacity differs between cranial and postcranial bones in old patients. It was hypothesised that different ageing patterns, reflected by osteocyte density, lacunar density and osteoid formation, exist between murine mandibles and femurs. MATERIAL AND METHODS: Mandible and femur of young (4 months) and old (34-36 months old) male C57Bl/6 mice were histologically investigated to determine the number of lacunae occupied with osteocytes. Osteoid formation was revealed by Masson-Goldner staining, and the spatial distribution of BMP-2 synthesis was examined. RESULTS: Over lifetime, the number of lacunae occupied with osteocytes only showed a modest decrease in mandibular bone (old 85.63%/young 91.12%) while greatly diverging in the femur (old 55.99%/young 93.28%). In equal measure, old femur exhibited less osteoid formation and decreased BMP-2 expression. CONCLUSION: Tissue-specific conduct of bone ageing is moulded by osteocytic activities, which was found to vary between postcranial and craniofacial skeleton. The latter harbours long-lived osteocytes also in old animals which assures lifelong bone integrity. Preliminary concurring findings from a human cadaver, also presented in this contribution, provided a rationale for recommending the translatability to humans.

Validation of Next-Generation Sequencing of Entire Mitochondrial Genomes and the Diversity of Mitochondrial DNA Mutations in Oral Squamous Cell Carcinoma.

BACKGROUND: Oral squamous cell carcinoma (OSCC) is mainly caused by smoking and alcohol abuse and shows a five-year survival rate of ~50%. We aimed to explore the variation of somatic mitochondrial DNA (mtDNA) mutations in primary oral tumors, recurrences and metastases. METHODS: We performed an in-depth validation of mtDNA next-generation sequencing (NGS) on an Illumina HiSeq 2500 platform for its application to cancer tissues, with the goal to detect low-level heteroplasmies and to avoid artifacts. Therefore we genotyped the mitochondrial genome (16.6 kb) from 85 tissue samples (tumors, recurrences, resection edges, metastases and blood) collected from 28 prospectively recruited OSCC patients applying both Sanger sequencing and high-coverage NGS (~35,000 reads per base). RESULTS: We observed a strong correlation between Sanger sequencing and NGS in estimating the mixture ratio of heteroplasmies (r = 0.99; p<0.001). Non-synonymous heteroplasmic variants were enriched among cancerous tissues. The proportions of somatic and inherited variants in a given gene region were strongly correlated (r = 0.85; p<0.001). Half of the patients shared mutations between benign and cancerous tissue samples. Low level heteroplasmies (<10%) were more frequent in benign samples compared to tumor samples, where heteroplasmies >10% were predominant. Four out of six patients who developed a local tumor recurrence showed mutations in the recurrence that had also been observed in the primary tumor. Three out of five patients, who had tumor metastases in the lymph nodes of their necks, shared mtDNA mutations between primary tumors and lymph node metastases. The percentage of mutation heteroplasmy increased from the primary tumor to lymph node metastases. CONCLUSIONS: We conclude that Sanger sequencing is valid for heteroplasmy quantification for heteroplasmies ≥10% and that NGS is capable of reliably detecting and quantifying heteroplasmies down to the 1%-level. The finding of shared mutations between primary tumors, recurrences and metastasis indicates a clonal origin of malignant cells in oral cancer.

Life-threatening complications of deep neck space infections.

Odontogenic infections represented a challenge for every craniomaxillofacial surgeon in the pre-antibiotic era, not least due to the frequent association with septic complications and lethal consequences. Nowadays, the incidences of serious infection-related complications are less frequent thanks to the medical progress. Nevertheless, the development of severe infections is still hard to foresee. The following analysis provides an overview of four patients with severe, life-threatening odontogenic deep neck space infections treated at the University Clinic for Oral and Maxillofacial Surgery in Innsbruck and describes the etiology, therapy, and outcome for each patient while analyzing differences between the single cases. Crucial aspects in origin, progression, and treatment of DNIs are highlighted, comparing the results with the international literature.

Biological effects of functionalizing copolymer scaffolds with nanodiamond particles.

Significant evidence has indicated that poly(L-lactide)-co-(ɛ-caprolactone) [(poly(LLA-co-CL)] scaffolds could be one of the suitable candidates for bone tissue engineering. Oxygen-terminated nanodiamond particles (n-DP) were combined with poly(LLA-co-CL) and revealed to be positive for cell growth. In this study, we evaluated the influence of poly(LLA-co-CL) scaffolds modified by n-DP on attachment, proliferation, differentiation of bone marrow stromal cells (BMSCs) in vitro, and on bone formation using a sheep calvarial defect model. BMSCs were seeded on either poly(LLA-co-CL)- or n-DP-coated scaffolds and incubated for 1 h. Scanning electron microscopy (SEM) and fluorescence microscopy were used in addition to protein and DNA measurements to evaluate cellular attachment on the scaffolds. To determine the effect of n-DP on proliferation of BMSCs, cell/scaffold constructs were harvested after 3 days and evaluated by Bicinchoninic Acid (BCA) protein assay and SEM. In addition, the osteogenic differentiation of cells grown for 2 weeks on the various scaffolds and in a dynamic culture condition was evaluated by real-time RT-PCR. Unmodified and modified scaffolds were implanted into the calvaria of six-year-old sheep. The expression of collagen type I (COL I) and bone morphogenetic protein-2 (BMP-2) after 4 weeks as well as the formation of new bone after 12 and 24 weeks were analyzed by immunohistochemistry and histology. Scaffolds modified with n-DP supported increased cell attachment and the mRNA expression of osteopontin (OPN), bone sialoprotein (BSP), and BMP-2 were significantly increased after 2 weeks of culture. The BMSCs had spread well on the various scaffolds investigated after 3 days in the study with no significant difference in cell proliferation. Furthermore, the in vivo data revealed more positive staining of COL I and BMP-2 in relation to the n-DP-coated scaffolds after 4 weeks and presented more bone formation after 12 and 24 weeks. n-DP modification significantly increased cell attachment and differentiation of BMSCs on poly(LLA-co-CL) scaffolds in vitro and enhanced bone formation in vivo.

BMP-2 immobilized on nanocrystalline diamond-coated titanium screws; demonstration of osteoinductive properties in irradiated bone.

BACKGROUND: Irradiation results in impaired bone healing. Thus, osteosynthesis procedures are afflicted with increased failure rates. To improve osseointegration bone morphogenetic protein-2 (BMP-2) immobilized on nanocrystalline diamond (NCD)-coated implant surfaces might be 1 solution. METHODS: By 4 weeks after irradiation of pig's mandible with a dose of 60 Gy a fracture was accomplished. Osteosynthesis was performed either with titanium osteosynthesis screws or NCD-coated screws with immobilized BMP-2. Nonirradiated animals served as control. After 1, 2, 4, and 8 weeks screws were evaluated histologically. Bone biopsies were gained to extract mesenchymal stem or precursor cells (MSCs). RESULTS: MSCs after irradiation demonstrated a behavior comparable to that of unirradiated cells. Consequently, immobilized BMP-2 resulted in an initial increased bone contact ratio (p = .014) but demonstrated no sustainable effect compared with osseointegration in nonirradiated bone (p = .08). CONCLUSION: Immobilized BMP-2 demonstrates an osteoinductive effect in irradiated bone. MSCs as effector cells possess protective mechanisms to overcome the destructive effect of irradiation.

Mesenchymal stem cells show radioresistance in vivo.

Irradiation impacts on the viability and differentiation capacity of tissue-borne mesenchymal stem cells (MSC), which play a pivotal role in bone regeneration. As a consequence of radiotherapy, bones may develop osteoradionecrosis. When irradiating human bone-derived MSC in vitro with increasing doses, the cells' self-renewal capabilities were greatly reduced. Mitotically stalled cells were still capable of differentiating into osteoblasts and pre-adipocytes. As a large animal model comparable to the clinical situation, pig mandibles were subjected to fractionized radiation of 2 χ 9 Gy within 1 week. This treatment mimics that of a standardized clinical treatment regimen of head and neck cancer patients irradiated 30 χ 2 Gy. In the pig model, fractures which had been irradiated, showed delayed osseous healing. When isolating MSC at different time points post-irradiation, no significant changes regarding proliferation capacity and osteogenic differentiation potential became apparent. Therefore, pig mandibles were irradiated with a single dose of either 9 or 18 Gy in vivo, and MSC were isolated immediately afterwards. No significant differences between the untreated and 9 Gy irradiated bone with respect to proliferation and osteogenic differentiation were unveiled. Yet, cells isolated from 18 Gy irradiated specimens exhibited a reduced osteogenic differentiation capacity, and during the first 2 weeks proliferation rates were greatly diminished. Thereafter, cells recovered and showed normal proliferation behaviour. These findings imply that MSC can effectively cope with irradiation up to high doses in vivo. This finding should thus be implemented in future therapeutic concepts to protect regenerating tissue from radiation consequences.

Epidemiologic evidence of barometric pressure changes inducing increased reporting of oral pain.

BACKGROUND AND OBJECTIVE: The daily patient flow at dental emergency departments is subject to fluctuations. The aim of the present study was to investigate whether meteorological parameters were associated with the number of patients reporting acute pain in the oral cavity. METHODS: We performed a retrospective review of all patients presenting with acute oro-dental pain (n=1090) at the dental outpatient clinic in Innsbruck, Austria from January to March 2005. In addition, local weather data were supplied by the Central Institute for Meteorology and Geodynamics Austria. After exponential smoothing of both series of observations, linear regression on the number of patients was performed. RESULTS: Statistical analysis revealed that the acute pain symptoms are not related to the air pressure itself but to the deviation of air pressure. The stronger the daily deviation from the average atmospheric pressure was, the more patients presented with acute pain at the dental emergency department (p=4.2 × 10(-14)). Our findings implicate that the association of air pressure deviation with variation in patient flow at dental clinics is a consequence of barometric pressure-related oral pain. CONCLUSIONS: These findings suggest that changes in barometric pressure were the reason for the observed initiation and/or exacerbation of the various oral pains observed in the emergency room setting. This association has been considered to be rare, with most reports suggesting this phenomenon primarily affects divers and aircraft personnel. Our data suggests otherwise, by providing clear evidence that atmospheric pressure changes commonly influences painful oral conditions.

In vivo investigation on connective tissue healing to polished surfaces with different surface wettability.

OBJECTIVES: Connective tissue in contact to transgingival/-dermal implants presents itself as tight scar formation. Although rough surfaces support the attachment they increase bacterial colonisation as well. In contrast to surface roughness, little is known about the influence of surface wettability on soft-tissue healing in vivo. We therefore investigated the influence of different surface wettabilities on connective tissue healing at polished implant surfaces in vivo. MATERIAL AND METHODS: Three polished experimental groups (titanium, titanium coated with hydrophobic nano-crystalline diamond (H-NCD) and titanium coated with hydrophilic nano-crystalline diamond (O-NCD) were inserted into the subcutaneous connective tissue of the abdominal wall of 24 rats. Animals were sacrificed after 1 and 4 weeks resulting in eight specimen per group per time point. Specimen were subjected to histological evaluation (van Giesson's staining) and immunohistochemistry staining for proliferating cell nuclear antigen (PCNA), fibronectin and tumour necrosis factor-alpha (TNF-α). RESULTS: Histological evaluation revealed dense scar formation at the titanium and H-NCD surfaces. In contrast, the connective tissue was loose at the O-NCD surface with a significantly higher number of cells after 4 weeks. O-NCD demonstrated a strong expression of PCNA and fibronectin but a weak expression of TNF-α. In contrast, the PCNA and fibronectin expression was low at the titanium and H-NCD, with a strong signal of TNF-α at the H-NCD surface. CONCLUSIONS: Hydrophilicity influences the connective tissue healing at polished implant surfaces in vivo positively. The attachment of connective tissue and the number of cells in contact to the surface were increased. Moreover, the inflammatory response is decreased at the hydrophilic surface.

Pullout strength of a biodegradable free form osteosynthesis plate.

The Inion(®) Free Form Plate is a newly designed biodegradable plate. After drilling through the plate and tapping, a biodegradable screw can be inserted, followed by removal of the screw head. As an alternative a countersink screw can be used. Aim of the study was to compare the mechanical properties of the 1.4 mm Free Form Plate with the 2.0 mm conventional shaped plate. Mechanical testing of the plate pullout strength was conducted for the Inion(®) Free Form Plate fixed with an Inion OTPS™ 2.0 × 20 mm Screw. In addition, the failure mode was reported. Overlapping confidence levels were found with regard to the yield load, first peak load and maximum load, when comparing the Free Form Plate and the conventional 4-hole plate. The Free Form Plate fixed with a screw with head and countersink showed the highest stability at maximum load. The results of the mechanical stability testing showed no significant differences between the tested plates. The main failure mode was a failure of the screw shaft. The results of the current investigation imply that the 1.4 mm Free Form Plate could be used as an alternative to the 2.0 mm conventional shaped plate.

The role of oxygen termination of nanocrystalline diamond on immobilisation of BMP-2 and subsequent bone formation.

Medical implants are increasingly often inserted into bone of frail patients, who are advanced in years. Due to age, severe trauma or pathology-related bone changes, osseous healing at the implant site is frequently limited. We were able to demonstrate that coating of endosseous implants with nanocrystalline diamond (NCD) allows stable functionalization by means of physisorption with BMP-2. Strong physisorption was shown to be directly related to the unique properties of NCD, and BMP-2 in its active form interacted strongly when NCD was oxygen-terminated. The binding of the protein was monitored under physiological conditions by single molecule force spectroscopy, and the respective adsorption energies were further substantiated by force-field-calculations. Implant surfaces refined in such a manner yielded enhanced osseointegration in vivo, when inserted into sheep calvaria. Our results further suggest that this technical advancement can be readily applied in clinical therapies with regard to bone healing, since primary human mesenchymal stromal cells strongly activated the expression of osteogenic markers when being cultivated on NCD physisorbed with physiological amounts of BMP-2.

Nanocrystalline diamond--an excellent platform for life science applications.

Nanocrystalline diamond (NCD) has recently been successfully utilized in a variety of life science applications. NCD films are favorable and salubrious substrates for cells during cultivation. Therefore NCD has also been employed in tissue engineering strategies. NCD as reported in this contribution was grown by means of a modified hot-filament chemical vapor deposition technique, which results in less than 3% sp2-hybridization and yields grain sizes of 5-20 nm. After production the NCD surface was rather hydrophobic, however it could be efficiently refined to exhibit more hydrophilic properties. Changing of the surface structure was found to be an efficient means to influence growth and differentiation capacity of a variety of cells. The particular needs for any given cell type has to be proven empirically. Yet flexible features of NCD appear to be superior to plastic surfaces which can be hardly changed in quality. Besides its molecular properties, crystal structural peculiarities of NCD appear to influence cell growth as well. In our attempt to facilitate, highly specialized applications in biomedicine, we recently discovered that growth factors can be tightly bound to NCD by mere physisorption. Hence, combination of surface functionalization together with further options to coat NCD with any kind of three-dimensional structure opens up new avenues for many more applications. In fact, high through-put protein profiling of early disease stages may become possible from serum samples, because proteins bound to NCD can now be efficiently analyzed by MALDI/TOF-MS. Given these results, it is to be presumed that the physical properties and effective electrochemical characteristics of NCD will allow tailoring devices suitable for many more diagnostic as well as therapeutic applications.

Strong binding of bioactive BMP-2 to nanocrystalline diamond by physisorption.

Nano-crystalline diamond (NCD)-coated surfaces were efficiently functionalized with bone morphogenetic protein-2 (BMP-2) by means of physisorption. Due to their randomly oriented texture, NCD-coated surfaces appear to bind complex molecules firmly. Applying various highly sensitive analytical methods, the interaction was found extremely stable. The strength of the experimentally measured adherence between BMP-2 and NCD was further corroborated by theoretical calculations. Oxygen treatment rendered NCD hydrophilic by the appearance of surface oxygen containing groups. This particular NCD surface exhibited even higher binding energies towards BMP-2 than the hydrophobic surface, and this surface was also favoured by cultured cells. Most importantly in this context, bound BMP-2 was found fully active. When cultured on BMP-2-treated NCD, osteosarcoma cells strongly up-regulated alkaline phosphatase, a specific marker for osteogenic differentiation. Hence, this simple method will allow generating highly versatile surfaces with complex biomimetic coatings, essentials for novel medical devices and implants as well as for innovative scaffolds in tissue engineering.