The shape of a bone scraper: an in vitro pilot study using porcine bone chips.

Bone scrapers are commonly used to harvest autologous bone in oral and implant surgery. The angle of the cutting blade is a variable that distinguishes bone scrapers. In the present study, the impact of the angle of the cutting blade on the in vitro characteristics of harvested bone was determined. Bone scrapers with blade angles of 15°, 25°, 35°, 45°, and 55° were used to harvest porcine cortical mandibular bone. The number and characteristics of the cells that grew out from the bone chips were examined. The data showed that, independent of the angle of the cutting blade, viable cells were barely detectable in fresh bone grafts. However, cells with a fibroblastic morphology appeared within 1 week in the culture dishes. After 21 days, the number of cells did not differ significantly between the five preparations. Moreover, cells responded to incubation with bone morphogenetic protein 7 (BMP-7) with an increased alkaline phosphatase activity, irrespective of the preparation. The data suggest that bone scrapers with different cutting angles produce bone chips with comparable in vitro characteristics.

Human bone chips release of sclerostin and FGF-23 into the culture medium: an in vitro pilot study

Abstract OBJECTIVE: Signaling molecules derived from osteocytes have been proposed as a mechanism by which autografts contribute to bone regeneration. However, there have been no studies that determined the role of osteocytes in bone grafts. MATERIAL AND METHOD: Herein, it was examined whether bone chips and demineralized bone matrix release sclerostin and FGF-23, both of which are highly expressed by osteocytes. RESULTS: Bone grafts from seven donors were placed in culture medium. Immunoassay showed that bone chips released sclerostin (median 1.0 ng/ml) and FGF-23 (median 9.8 relative units/ml) within the first day, with declining levels overtime. Demineralized bone matrix also released detectable amounts of sclerostin into culture medium, while FGF-23 remained close to the detection limit. In vitro expanded isolated bone cells failed to release detectable amounts of sclerostin and FGF-23. CONCLUSION: These results suggest that autografts but also demineralized bone matrix can release signaling molecules that are characteristically produced by osteocytes. © 2014 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd. KEYWORDS: FGF-23; autologous bone; bone grafts; demineralized bone matrix; osteocytes; sclerostin

Antiseptic solutions modulate the paracrine-like activity of bone chips: differential impact of chlorhexidine and sodium hypochlorite

AIM: Chemical decontamination increases the availability of bone grafts; however, it is unclear whether antiseptic processing changes the biological activity of bone. MATERIALS AND METHODS: Bone chips were incubated with 4 different antiseptic solutions including (1) povidone-iodine (0.5%), (2) chlorhexidine diguluconate (0.2%), (3) hydrogen peroxide (1%) and (4) sodium hypochlorite (0.25%). After 10 minutes of incubation, changes in the capacity of the bone-conditioned medium to modulate gene expression of gingival fibroblasts was investigated. RESULTS: Conditioned medium obtained from freshly prepared bone chips increased the expression of TGF-β target genes interleukin 11 (IL11), proteoglycan4 (PRG4), NADPH oxidase 4 (NOX4), and decreased the expression of adrenomedullin (ADM), and pentraxin 3 (PTX3) in gingival fibroblasts. Incubation of bone chips with 0.2% chlorhexidine, followed by vigorously washing resulted in a bone-conditioned medium with even higher expression of IL11, PRG4, and NOX4. These findings were also found with a decrease in cell viability and an activation of apoptosis signaling. Chlorhexidine alone, at low concentrations, increased IL11, PRG4 and NOX4 expression, independent of the TGF-β receptor I kinase activity. In contrast, 0.25% sodium hypochlorite almost entirely abolished the activity of bone-conditioned medium, while the other two antiseptic solutions, 1% hydrogen peroxide and 0.5% povidone-iodine, had relatively no impact, respectively. CONCLUSION: These in vitro findings demonstrate that incubation of bone chips with chlorhexidine differentially affects the activity of the respective bone-conditioned medium compared to the other antiseptic solutions. The data further suggest that the main effects are caused by chlorhexidine remaining in the bone-conditioned medium after repeated washing of the bone chips. This article is protected by copyright. All rights reserved. This article is protected by copyright. All rights reserved. KEYWORDS: Autografts; TGF-β; antiseptic solution; bone; bone conditioned medium; bone supernatant; chlorhexidine; hydrogen peroxide; povidone-iodine; sodium hypochlorite

Diagnostic microchip to assay 3D colony-growth potential of captured circulating tumor cells

Microfluidic technology has been successfully applied to isolate very rare tumor-derived epithelial cells (circulating tumor cells, CTCs) from blood with relatively high yield and purity, opening up exciting prospects for early detection of cancer. However, a major limitation of state-of-the-art CTC-chips is their inability to characterize the behavior and function of captured CTCs, for example to obtain information on proliferative and invasive properties or, ultimately, tumor re-initiating potential. Although CTCs can be efficiently immunostained with markers reporting phenotype or fate (e.g. apoptosis, proliferation), it has not yet been possible to reliably grow captured CTCs over long periods of time and at single cell level. It is challenging to remove CTCs from a microchip after capture, therefore such analyses should ideally be performed directly on-chip. To address this challenge, we merged CTC capture with three-dimensional (3D) tumor cell culture on the same microfluidic platform. PC3 prostate cancer cells were isolated from spiked blood on a transparent PDMS CTC-chip, encapsulated on-chip in a biomimetic hydrogel matrix (QGel™) that was formed in situ, and their clonal 3D spheroid growth potential was assessed by microscopy over one week in culture. The possibility to clonally expand a subset of captured CTCs in a near-physiological in vitro model adds an important element to the expanding CTC-chip toolbox that ultimately should improve prediction of treatment responses and disease progression.

Effect of two different bioabsorbable collagen membranes on guided bone regeneration: a comparative histomorphometric study in the dog mandible

BACKGROUND: This study compared bone regeneration following guided bone regeneration with two bioabsorbable collagen membranes in saddle-type bone defects in dog mandibles. METHODS: Three standardized defects were created, filled with bone chips and deproteinized bovine bone mineral (DBBM), and covered by three different methods: control = no membrane; test 1 = collagen membrane; and test 2 = cross-linked collagen membrane (CCM). Each side of the mandible was allocated to one of two healing periods (8 or 16 weeks). The histomorphometric analysis assessed the percentage of bone, soft tissue, and DBBM in the regenerate; the absolute area in square millimeters of the bone regenerate; and the distance in millimeters from the bottom of the defect to the highest point of the regenerate. RESULTS: In the 8-week healing group, two dehiscences occurred with CCM. After 8 weeks, all treatment modalities showed no significant differences in the percentage of bone regenerate. After 16 weeks, the percentage of bone had increased for all treatment modalities without significant differences. For all groups, the defect fill height increased between weeks 8 and 16. The CCM group showed a statistically significant (P = 0.0202) increase over time and the highest value of all treatment modalities after 16 weeks of healing, CONCLUSIONS: The CCM showed a limited beneficial effect on bone regeneration in membrane-protected defects in dog mandibles when healing was uneventful. The observed premature membrane exposures resulted in severely compromised amounts of bone regenerate. This increased complication rate with CCM requires a more detailed preclinical and clinical examination before any clinical recommendations can be made.

Evaluation of a novel biphasic calcium phosphate in standardized bone defects: a histologic and histomorphometric study in the mandibles of minipigs

OBJECTIVE: A novel biphasic calcium phosphate (CaP) granulate consisting of hydroxyapatite (HA) and beta-tricalciumphosphate (TCP) was compared with pure HA and pure TCP and with autograft as positive control. MATERIALS AND METHODS: Four standardized bone defects were prepared in both mandibular angles of 16 minipigs and grafted with autogenous bone chips, HA, HA/TCP (60% : 40%), or TCP. Histologic and histomorphometric analysis of bone formation and graft degradation followed healing periods of 2, 4, 8, and 24 weeks. RESULTS: 2 weeks: more bone formation in defects filled with autograft than with the three CaP materials (P<0.05). 4 weeks: bone formation differed significantly (P<0.05) between all four materials (autograft>TCP>HA/TCP>HA). 8 weeks: more bone formation in defects with autograft and TCP than with HA/TCP (P<0.05), and HA/TCP had more bone formation than HA (P<0.05). 24 weeks: no difference in bone formation between the groups. Autograft and TCP resorbed quickly and almost completely over 8 weeks, whereas HA/TCP and HA showed limited degradation over 24 weeks. CONCLUSION: All defects healed with mature lamellar bone and intimate contact between bone and the remaining graft material. The rate of bone formation corresponded to the content of TCP in the CaP materials.

Performance of dental implants after staged sinus floor elevation procedures: 5-year results of a prospective study in partially edentulous patients

OBJECTIVES: The aim of this prospective study was to evaluate the 5-year performance and success rate of titanium screw-type implants with the titanium plasma spray (TPS) or the sand-blasted, large grit, acid-etched (SLA) surface inserted in a two-stage sinus floor elevation (SFE) procedure in the posterior maxilla. MATERIAL AND METHODS: A total of 59 delayed SFEs were performed in 56 patients between January 1997 and December 2001, using a composite graft with autogenous bone chips combined with deproteinized bovine bone mineral (DBBM) or synthetic porous beta-tricalcium phosphate (beta-TCP). After a healing period averaging 7.75 months, 111 dental implants were inserted. After an additional 8-14-week healing period, all implants were functionally loaded with cemented crowns or fixed partial dentures. The patients were recalled at 12 and 60 months for clinical and radiographic examination. RESULTS: One patient developed an acute infection in the right maxillary sinus after SFE and did not undergo implant therapy. Two of the 111 inserted implants had to be removed because of a developing atypical facial pain, and 11 implants were lost to follow-up and were considered drop-outs. The remaining 98 implants showed favorable clinical and radiographic findings at the 5-year examination. The peri-implant soft tissues were stable over time; the mean probing depths and mean attachment levels did not change during the follow-up period. The measurement of the bone crest levels (DIB values) indicated stability as well. Based on strict success criteria, all 98 implants were considered successfully integrated, resulting in a 5-year success rate of 98% (for TPS implants 89%, for SLA implants 100%). CONCLUSION: This prospective study assessing the performance of dental implants inserted after SFE demonstrated that titanium implants can achieve and maintain successful tissue integration with high predictability for at least 5 years of follow-up in carefully selected patients.

Early implant placement following single-tooth extraction in the esthetic zone: biologic rationale and surgical procedures

Early implant placement is one treatment option for implant therapy following single-tooth extraction in the anterior maxilla. The surgical technique presented here is characterized by tooth extraction without flap elevation, a 4- to 8-week soft tissue healing period, implant placement in a correct three-dimensional position, simultaneous contour augmentation on the facial aspect with guided bone regeneration using a bioabsorbable collagen membrane combined with autogenous bone chips and a low-substitution bone filler, and tension-free primary wound closure. The surgical step-by-step procedure is presented with a case report. In addition, the biologic rationale is discussed.

A feasibility study evaluating an in situ formed synthetic biodegradable membrane for guided bone regeneration in dogs

PURPOSE: The aim was (1) to evaluate the soft-tissue reaction of a synthetic polyethylene glycol (PEG) hydrogel used as a barrier membrane for guided bone regeneration (GBR) compared with a collagen membrane and (2) to test whether or not the application of this in situ formed membrane will result in a similar amount of bone regeneration as the use of a collagen membrane. MATERIAL AND METHODS: Tooth extraction and preparation of osseous defects were performed in the mandibles of 11 beagle dogs. After 3 months, 44 cylindrical implants were placed within healed dehiscence-type bone defects resulting in approximately 6 mm exposed implant surface. The following four treatment modalities were randomly allocated: PEG+autogenous bone chips, PEG+hydroxyapatite (HA)/tricalcium phosphate (TCP) granules, bioresorbable collagen membrane+autogenous bone chips and autogenous bone chips without a membrane. After 2 and 6 months, six and five dogs were sacrificed, respectively. A semi-quantitative evaluation of the local tolerance and a histomorphometric analysis were performed. For statistical analysis, repeated measures analysis of variance (ANOVA) and subsequent pairwise Student's t-test were applied (P<0.05). RESULTS: No local adverse effects in association with the PEG compared with the collagen membrane was observed clinically and histologically at any time-point. Healing was uneventful and all implants were histologically integrated. Four out of 22 PEG membrane sites revealed a soft-tissue dehiscence after 1-2 weeks that subsequently healed uneventful. Histomorphometric measurement of the vertical bone gain showed after 2 months values between 31% and 45% and after 6 months between 31% and 38%. Bone-to-implant contact (BIC) within the former defect area was similarly high in all groups ranging from 71% to 82% after 2 months and 49% to 91% after 6 months. However, with regard to all evaluated parameters, the PEG and the collagen membranes did not show any statistically significant difference compared with sites treated with autogenous bone without a membrane. CONCLUSION: The in situ forming synthetic membrane made of PEG was safely used in the present study, revealing no biologically significant abnormal soft-tissue reaction and demonstrated similar amounts of newly formed bone for defects treated with the PEG membrane compared with defects treated with a standard collagen membrane.

Contribution of genome-wide significant single-nucleotide polymorphisms and antiretroviral therapy to dyslipidemia in HIV-infected individuals: a longitudinal study

BACKGROUND: HIV-infected individuals have an increased risk of myocardial infarction. Antiretroviral therapy (ART) is regarded as a major determinant of dyslipidemia in HIV-infected individuals. Previous genetic studies have been limited by the validity of the single-nucleotide polymorphisms (SNPs) interrogated and by cross-sectional design. Recent genome-wide association studies have reliably associated common SNPs to dyslipidemia in the general population. METHODS AND RESULTS: We validated the contribution of 42 SNPs (33 identified in genome-wide association studies and 9 previously reported SNPs not included in genome-wide association study chips) and of longitudinally measured key nongenetic variables (ART, underlying conditions, sex, age, ethnicity, and HIV disease parameters) to dyslipidemia in 745 HIV-infected study participants (n=34 565 lipid measurements; median follow-up, 7.6 years). The relative impact of SNPs and ART to lipid variation in the study population and their cumulative influence on sustained dyslipidemia at the level of the individual were calculated. SNPs were associated with lipid changes consistent with genome-wide association study estimates. SNPs explained up to 7.6% (non-high-density lipoprotein cholesterol), 6.2% (high-density lipoprotein cholesterol), and 6.8% (triglycerides) of lipid variation; ART explained 3.9% (non-high-density lipoprotein cholesterol), 1.5% (high-density lipoprotein cholesterol), and 6.2% (triglycerides). An individual with the most dyslipidemic antiretroviral and genetic background had an approximately 3- to 5-fold increased risk of sustained dyslipidemia compared with an individual with the least dyslipidemic therapy and genetic background. CONCLUSIONS: In the HIV-infected population treated with ART, the weight of the contribution of common SNPs and ART to dyslipidemia was similar. When selecting an ART regimen, genetic information should be considered in addition to the dyslipidemic effects of ART agents.

Bone-Conditioned Medium Inhibits Osteogenic and Adipogenic Differentiation of Mesenchymal Cells In Vitro

BACKGROUND AND PURPOSE Autografts are used for bone reconstruction in regenerative medicine including oral and maxillofacial surgery. Bone grafts release paracrine signals that can reach mesenchymal cells at defect sites. The impact of the paracrine signals on osteogenic, adipogenic, and chondrogenic differentiation of mesenchymal cells has remained unclear. MATERIAL AND METHODS Osteogenesis, adipogenesis, and chondrogenesis were studied with murine ST2 osteoblast progenitors, 3T3-L1 preadipocytes, and ATDC5 prechondrogenic cells, respectively. Primary periodontal fibroblasts from the gingiva, from the periodontal ligament, and from bone were also included in the analysis. Cells were exposed to bone-conditioned medium (BCM) that was prepared from porcine cortical bone chips. RESULTS BCM inhibited osteogenic and adipogenic differentiation of ST2 and 3T3-L1 cells, respectively, as shown by histological staining and gene expression. No substantial changes in the expression of chondrogenic genes were observed in ATDC5 cells. Primary periodontal fibroblasts also showed a robust decrease in alkaline phosphatase and peroxisome proliferator-activated receptor gamma (PPARγ) expression when exposed to BCM. BCM also increased collagen type 10 expression. Pharmacologic blocking of transforming growth factor (TGF)-β receptor type I kinase with SB431542 and the smad-3 inhibitor SIS3 at least partially reversed the effect of BCM on PPARγ and collagen type 10 expression. In support of BCM having TGF-β activity, the respective target genes were increasingly expressed in periodontal fibroblasts. CONCLUSIONS The present work is a pioneer study on the paracrine activity of bone grafts. The findings suggest that cortical bone chips release soluble signals that can modulate differentiation of mesenchymal cells in vitro at least partially involving TGF-β signaling.

Nanomaterials and the human lung: what is known and what must be deciphered to realise their potential advantages?

Due to the constant expansion within the nanotechnology industry in the last decade, nanomaterials are omnipresent in society today. Nanotechnology-based products have numerous different applications ranging from electronic (e.g., advanced memory chips) to industrial (e.g., coatings or composites) to biomedical (e.g., drug delivery systems, diagnostics). Although these new nanomaterials can be found in many "everyday" products, their effects on the human body have still to be investigated in order to identify not only their risk, but also their potential benefits towards human health. Since the lung is commonly thought to be the main portal of entry into the human body for nanomaterials released within the environment, this review will attempt to summarise the current knowledge and understanding of how nanomaterials interact with the respiratory tract. Furthermore, the advantages and disadvantages of different experimental model systems that are commonly used to study this exposure route to the human body will be discussed.

Proteomic analysis of porcine bone-conditioned medium.

PURPOSE Autografts are considered to support bone regeneration. Paracrine factors released from cortical bone might contribute to the overall process of graft consolidation. The aim of this study was to characterize the paracrine factors by means of proteomic analysis. MATERIALS AND METHODS Bone-conditioned medium (BCM) was prepared from fresh bone chips of porcine mandibles and subjected to proteomic analysis. Proteins were categorized and clustered using the bioinformatic tools UNIPROT and PANTHER, respectively. RESULTS Proteomic analysis showed that BCM contains more than 150 proteins, of which 43 were categorized into "secreted" and "extracellular matrix." Growth factors that are not only detectable in BCM, but potentially also target cellular processes involved in bone regeneration, eg, pleiotrophin, galectin-1, transforming growth factor beta (TGF-β)-induced gene (TGFBI), lactotransferrin, insulin-like growth factor (IGF)-binding protein 5, latency-associated peptide forming a complex with TGF-β1, and TGF-β2, were discovered. CONCLUSION The present results demonstrate that cortical bone chips release a large spectrum of proteins with the possibility of modulating cellular aspects of bone regeneration. The data provide the basis for future studies to understand how these paracrine factors may contribute to the complex process of graft consolidation.

Long-term stability of contour augmentation in the esthetic zone: histologic and histomorphometric evaluation of 12 human biopsies 14 to 80 months after augmentation.

BACKGROUND Contour augmentation around early-placed implants (Type 2 placement) using autogenous bone chips combined with deproteinized bovine bone mineral (DBBM) and a collagen barrier membrane has been documented to predictably provide esthetically satisfactory clinical outcomes. In addition, recent data from cone beam computed tomography studies have shown the augmented volume to be stable long-term. However, no human histologic data are available to document the tissue reactions to this bone augmentation procedure. METHODS Over an 8-year period, 12 biopsies were harvested 14 to 80 months after implant placement with simultaneous contour augmentation in 10 patients. The biopsies were subjected to histologic and histomorphometric analysis. RESULTS The biopsies consisted of 32.0% ± 9.6% DBBM particles and 40.6% ± 14.6% mature bone. 70.3% ± 14.5% of the DBBM particle surfaces were covered with bone. On the remaining surface, multinucleated giant cells with varying intensity of tartrate-resistant acid phosphatase staining were regularly present. No signs of inflammation were visible, and no tendency toward a decreasing volume fraction of DBBM over time was observed. CONCLUSIONS The present study confirms previous findings that osseointegrated DBBM particles do not tend to undergo substitution over time. This low substitution rate may be the reason behind the clinically and radiographically documented long-term stability of contour augmentation using a combination of autogenous bone chips, DBBM particles, and a collagen membrane.

Technical complications and failures of zirconia-based prostheses supported by implants followed up to 7 years: a case series.

PURPOSE To evaluate technical complications and failures of zirconia-based fixed prostheses supported by implants. MATERIALS AND METHODS Consecutive patients received zirconia-based single crowns (SCs) and fixed dental prostheses (FDPs) on implants in a private clinical setting between 2005 and 2010. One dentist performed all surgical and prosthetic procedures, and one master technician performed and coordinated all laboratory procedures. One-piece computer-aided design/ computer-assisted manufacture technology was used to fabricate abutments and frameworks, which were directly connected at the implant level, where possible. All patients were involved in a recall maintenance program and were finally reviewed in 2012. Data on framework fractures, chipping of veneering ceramics, and other technical complications were recorded. The primary endpoint was failure of the prostheses, ie, the need for a complete remake. A life table analysis was calculated. RESULTS A total of 289 implants supported 193 zirconia-based prostheses (120 SCs and 73 FDPs) in 127 patients (51 men, 76 women; average age: 62.5 ± 13.4 years) who were reviewed in 2012. Twenty-five (13%) prostheses were cemented on 44 zirconia abutments and 168 (87%) prostheses were screw-retained directly at the implant level. Fracture of 3 frameworks (1 SC, 2 FDPs) was recorded, and significant chipping resulted in the remake of 3 prostheses (1 SC, 2 FDPs). The 7-year cumulative survival rate was 96.4% ± 1.99%. Minor complications comprised 5 loose screws (these were retightened), small chips associated with 3 prostheses (these were polished), and dislodgement of 3 prostheses (these were recemented). Overall, 176 prostheses remained free of technical problems. CONCLUSIONS Zirconia-based prostheses screwed directly to implants are clinically successful in the short and medium term.