29 research outputs found
Pre-clinical screening of novel two-photon photopolymerized biomaterials for bone implantation.
No full textWirkung verschiedener Biomaterialien auf Osteoblasten in vitro
No full textKnochenschäden infolge von traumatischen Verletzungen, Tumorresektionen oder operativen Rekonstruktionen erfordern die Anwendung von Knochenersatzmaterialien. Gegenwärtig verfügbare Transplantate, wie die Verwendung von autologen oder allogenen Knochenmaterialen, und Allotransplantaten mit demineralisierter Matrix oder osteogenen Proteinen sind nur limitiert verfügbar. Das Forschungsgebiet der Knochenregenerationsmedizin hat sich aufgrund dieser Notwendigkeit rasch weiterentwickelt und die Entwicklung von Biomaterialien als Knochenersatz stellt dabei einen wichtigen Forschungsbereich dar.
Ziel dieser Diplomarbeit war die Untersuchung von Effekten potentieller Biomaterialen für den Bereich „bone tissue engeneering“. Getestet wurden zwei verschiedene Klassen von abbaubaren Biomaterialien. Einerseits wurden Einflüsse von Gelatine und andererseits von einem synthetischen Copolymer auf den Differenzierungsprozess und der Mineralisationsfähigkeit von Osteoblasten untersucht. Zur Beurteilung dieser Materialien wurden Parameter wie Zellviabilität und knochenspezifische Marker (Alkalische Phosphatase Aktivität und Matrixmineralisation) herangezogen.
Weder das Verfahren zur Gewinnung der Gelatine, noch Unterschiede in physikochemische Parameter wie isoelektrischer Punkt oder Gelierfähigkeit hatten einen Einfluss auf die Alkalische Phosphatase Aktivität MC3T3-E1 Zellen über den getesteten Zeitraum. Aktive Alkalische Phosphatase konnte in sämtlichen Gruppen mittels Färbemethode detektiert werden. Die Fähigkeit extrazelluläre Matrix zu mineralisieren wurde jedoch bei allen Gelatinegruppen gefördert. Physikochemische Parameter und Gewinnungsmethode der Gelatine bewirkten deutliche Unterschiede in der Matrixmineralisation. Sämtliche Beschichtungen, die als Einzelkomponente Gelatine Typ B enthielten oder in einer Mischung mit Gelatine Typ B gefertigt wurden, führten im Vergleich zu den anderen Gelatinebeschichtungen zu einer höheren Matrixmineralisation.
Bei Gegenüberstellung beider Ausgangsmaterialien als Scaffoldmaterial hinsichtlich ihres Effektes auf Zellviabilität und –proliferation, Osteoblastendifferenzierung und Mineralisation extrazellulärer Matrix, zeigte sich in sämtlichen Experimenten, dass LCM6 einen deutlich stärkeren Einfluss auf zelluläre Prozesse ausübt als LCM3. Primären Maus-Osteoblasten konnten auf dem Materialtyp LCM3 adhärieren, proliferieren, differenzieren und schließlich synthetisierte Matrix mineralisieren. In den in vitro Experimenten resultierte für diese Gruppe, trotz verminderter Anzahl an metabolisch aktiven Zellen, eine positive Färbung für das Enzym Alkalische Phosphatase und für die Matrixmineralisation. Im Gegensatz dazu kam es beim Biomaterial LCM6 neben der starken Reduktion in Proliferation und Viabilität zu einer Hemmung der Aktivität der Alkalischen Phosphatase und zu einem nur sehr geringen Anstieg an mineralisierter Matrix über 21 Tage.
Die Materialzusammensetzung, vor allem das Verhältnis der Einzelkomponenten zueinander (Lactid, Caprolacton und Methacrylat) und die damit verbundenen Eigenschaften, nehmen laut den erhaltenen Ergebnissen Einfluss auf Proliferation, den Differenzierungsprozess und auf die Matrixmineralisationsfähigkeit. Im Falle des Biomaterial LCM6 erwiesen sich eine Erhöhung des Lactid-Anteils und ein verringerter Prozentsatz an Methacrylat als nicht günstig für die getesteten Zellfunktionsparameter.Bone loss causing from trauma, surgical tumor removal or reconstructive surgeries requires the usage of bone graft substitutes. Current treatments as autologous or allogenic bone grafts, allografts with demineralized matrix or osteogenic proteins are limited available. Therefore the design of biomaterials with osteogenic, osteoinductive and osteoconductive properties become more and more interesting.
In the present diploma thesis work, the effect of potential biomaterials for bone grafting was tested on osteoblast proliferation and differentiation. Two polymers were investigated. One polymer was the macromolecule gelatin and the other one was a synthetic copolymer (LCM) Mouse calvarial derived osteoblasts or MC3T3-E1 cells were cultured on various gelatine coatings or on different photopolymerized LCM discs. Cells were directed to osteoblast lineage by growing them in differentiation media containing ascorbic acid (50µg/ml) and β-glycerophosphate (5mM). To determine effects on osteoblast proliferation, differentiation and mineralization, assays were performed for cell viability and proliferation (MTS-assay), alkaline phosphatase (ALP) activity and mineralized matrix was visualised with alizarin red.
Independent from the extraction method or gel strength of the tested gelatin coatings, osteoblasts attached on the pre-coated surfaces and showed ALP activity in all groups. There was no significant difference observed between the different investigated gelatin types according to ALP activity. But mineralized matrix production was enhanced in the presence of gelatin compared to the control on tissue culture plastic.
Osteoblast attachment and proliferation was also observed by culturing cells on different synthetic polymer-compositions (LCM3 and LCM6). Even if a reduction in the metabolic activity of osteoblasts on LCM3 and LCM6 was monitored, LCM3, but not LCM6, supported alkaline phosphatase expression and matrix mineralization.
In conclusion, both biomaterials showed supporting effects on osteoblast differentiation in vitro. In the process of bone healing, both, osteoblast and osteoclast play a major role. Therefore further experiments have to be established to determine effects on osteoclast development as well as to determine the potency of these materials for in vivo bone formation
Bone healing around titanium implants in a preclinical model of bile duct ligation‐induced liver injury
No full textOBJECTIVES
Chronic liver disease increases the risk for periodontal disease and osteoporotic fractures, but its impacts on bone regeneration remain unknown. Herein, we studied the impact of liver cirrhosis on peri-implant bone formation.
MATERIAL AND METHODS
A total of 20 male Wistar rats were randomly divided into two groups: one with the common bile duct ligated (BDL) and the respective sham-treated control group (SHAM). After four weeks of disease induction, titanium mini-screws were inserted into the tibia. Successful induction of liver cirrhosis was confirmed by the presence of clinical symptoms. Another four weeks later, peri-implant bone volume per tissue volume (BV/TV) and bone-to-implant contact (BIC) were determined by histomorphometric analysis.
RESULTS
Peri-implant bone formation was not significantly different between the SHAM and BDL groups. In the cortical compartment, the median percentage of peri-implant new bone was 10.1% (95% CI of mean 4.0-35.7) and 22.5% (13.8-30.6) in the SHAM and BDL groups, respectively (p = .26). Consistently, the new bone in direct contact with the implant was 18.1% (0.4-37.8) and 23.3% (9.2-32.8) in SHAM and BDL groups, respectively (p = .38). When measuring the medullary compartment, the new bone area was 7.1% (4.8-10.4) and 10.4% (7.2-13.5) in the SHAM and BDL groups, respectively (p = .17). Medullary new bone in direct contact with the implant was 10.0% (1.2-50.4) and 20.6% (16.8-35.3) in SHAM and BDL groups, respectively, and thus comparable between the two groups (p = .46).
CONCLUSIONS
Bile duct ligation has no significant impact on the early stages of peri-implant bone formation
Impact of a Static Magnetic Field on Early Osseointegration: A Pilot Study in Canines
No full textA static magnetic field generated by neodymium–iron–boron (NdFeB) magnets placed in the inner cavity of dental implants can enhance bone regeneration in rabbits. It is, however, unknown whether static magnetic fields support osseointegration in a canine model. We therefore determined the potential osteogenic effect of implants carrying NdFeB magnets inserted in the tibia of six adult canines in the early stages of osseointegration. Here, we report that after 15 days of healing, magnetic and regular implants showed a high variation with a median new bone-to-implant contact (nBIC) in the cortical (41.3% and 7.3%) and the medullary (28.6% and 44.8%) region, respectively. Consistently, the median new bone volume/tissue volume (nBV/TV) in the cortical (14.9% and 5.4%) and the medullary (22.2% and 22.4%) region were not significantly different. One week of healing only resulted in negligible bone formation. These findings suggest that considering the large variation and the pilot nature of this study, magnetic implants failed to support peri-implant bone formation in a canine model
Active and Passive Mineralization of Bio-Gide® Membranes in Rat Calvaria Defects.
No full textBio-Gide® is a collagen membrane routinely used in guided bone regeneration. Recent studies have shown that this collagen membrane has osteoconductive properties, meaning that it can support the growth of new bone. However, it has also been observed that the collagen membrane has areas of mineralized fibers which can occur spontaneously and independently of osteoblasts. To better understand how this works, we established a model using minced collagen membranes to reduce the active mineralization of intact collagen membranes in favor of passive mineralization. We thus compared the original intact membrane with a minced collagen membrane in a 5 mm calvarial defect model in Sprague Dawley rats. After three weeks of healing, histology and microcomputed tomography (μCT) were performed. Histological analysis confirmed the osteoconductive properties, with new bone growing inside the intact collagen membrane. However, in minced collagen membranes, the osteoconductive properties were restricted to the defect margins. Interestingly, histology revealed large mineralized areas indicating passive mineralization with no signs of bone formation. In the μCT analysis, the intact collagen membranes caused a higher median mineralized volume (1.5 mm3) compared with the minced group (0.4 mm3), but this lacked significance (p = 0.09). The μCT analysis needs to be interpreted carefully, particularly in defects filled with minced membranes, considering that the mineralized tissue may not necessarily be bone but also the result of passive mineralization. Taken together, the findings suggest that Bio-Gide® collagen membranes support bone formation while also exhibiting potential for passive mineralization
Preclinical biological and physicochemical evaluation of two-photon engineered 3D biomimetic copolymer scaffolds for bone healing
No full textA major challenge in orthopedics is the repair of large non-union bone fractures. A promising therapy for this indication is the use of biodegradable bioinspired biomaterials that stabilize the fracture site, relieve pain and initiate bone formation and healing. This study uses a multidisciplinary evaluation strategy to assess immunogenicity, allergenicity, bone responses and physicochemical properties of a novel biomaterial scaffold. Two-photon stereolithography generated personalized custom-built scaffolds with a repeating 3D structure of Schwarz Primitive minimal surface unit cell with a specific pore size of ∼400 μm from three different methacrylated poly(D,L-lactide-co-ε-caprolactone) copolymers with lactide to caprolactone monomer ratios of 16 : 4, 18 : 2 and 9 : 1. Using in vitro and in vivo assays for bone responses, immunological reactions and degradation dynamics, we found that copolymer composition influenced the scaffold physicochemical and biological properties. The scaffolds with the fastest degradation rate correlated with adverse cellular effects and mechanical stiffness correlated with in vitro osteoblast mineralization. The physicochemical properties also correlated with in vivo bone healing and immune responses. Overall these observations provide compelling support for these scaffolds for bone repair and illustrate the effectiveness of a promising multidisciplinary strategy with great potential for the preclinical evaluation of biomaterials
Human versus Rat PRF on Collagen Membranes: A Pilot Study of Mineralization in Rat Calvaria Defect Model.
No full textPlatelet-rich fibrin, the coagulated plasma fraction of blood, is commonly used to support natural healing in clinical applications. The rat calvaria defect is a standardized model to study bone regeneration. It remains, however, unclear if the rat calvaria defect is appropriate to investigate the impact of human PRF (Platelet-Rich Fibrin) on bone regeneration. To this end, we soaked Bio-Gide® collagen membranes in human or rat liquid concentrated PRF before placing them onto 5 mm calvarial defects in Sprague Dawley rats. Three weeks later, histology and micro-computed tomography (μCT) were performed. We observed that the collagen membranes soaked with rat PRF show the characteristic features of new bone and areas of mineralized collagen matrix, indicated by a median mineralized volume of 1.5 mm3 (range: 0.9; 5.3 mm3). Histology revealed new bone growing underneath the membrane and hybrid bone where collagen fibers are embedded in the new bone. Moreover, areas of passive mineralization were observed. The collagen membranes soaked with human PRF, however, were devoid of histological features of new bone formation in the center of the defect; only occasionally, new bone formed at the defect margins. Human PRF (h-PRF) caused a median bone volume of 0.9 mm3 (range: 0.3-3.3 mm3), which was significantly lower than what was observed with rat PRF (r-PRF), with a BV median of 1.2 mm3 (range: 0.3-5.9 mm3). Our findings indicate that the rat calvaria defect model is suitable for assessing the effects of rat PRF on bone formation, but caution is warranted when extrapolating conclusions regarding the efficacy of human PRF
on bone graft remodeling: An in vivo experimental study
No full textOBJECTIVES
Autologous bone is considered the gold standard for grafting, yet it suffers from a tendency to undergo resorption over time. While the exact mechanisms of this resorption remain elusive, osteocytes have been shown to play an important role in stimulating osteoclastic activity through their expression of receptor activator of NF-κB (RANK) ligand (RANKL). The aim of this study was to assess the function of osteocyte-derived RANKL in bone graft remodeling.
MATERIALS AND METHODS
In Tnfsf11fl/fl ;Dmp1-Cre mice without osteocyte-specific RANKL as well as in Dmp1-Cre control mice, 2.6 mm calvarial bone disks were harvested and transplanted into mice with matching genetic backgrounds either subcutaneously or subperiosteally, creating 4 groups in total. Histology and micro-computed tomography of the grafts and the donor regions were performed 28 days after grafting.
RESULTS
Histology revealed marked resorption of subcutaneous control Dmp1-Cre grafts and new bone formation around subperiosteal Dmp1-Cre grafts. In contrast, Tnfsf11fl/fl ;Dmp1-Cre grafts showed effectively neither signs of bone resorption nor formation. Quantitative micro-computed tomography revealed a significant difference in residual graft area between subcutaneous and subperiosteal Dmp1-Cre grafts (p < .01). This difference was not observed between subcutaneous and subperiosteal Tnfsf11fl/fl ;Dmp1-Cre grafts (p = .17). Residual graft volume (p = .08) and thickness (p = .13) did not differ significantly among the groups. Donor area regeneration was comparable between Tnfsf11fl/fl ;Dmp1-Cre and Dmp1-Cre mice and restricted to the defect margins.
CONCLUSIONS
The results suggest an active function of osteocyte-derived RANKL in bone graft remodeling
Acid Dentin Lysate Failed to Modulate Bone Formation in Rat Calvaria Defects
No full textAutogenous tooth roots are increasingly applied as a grafting material in alveolar bone augmentation. Since tooth roots undergo creeping substitution similar to bone grafts, it can be hypothesized that osteoclasts release the growth factors stored in the dentin thereby influencing bone formation. To test this hypothesis, collagen membranes were either soaked in acid dentin lysates (ADL) from extracted porcine teeth or serum–free medium followed by lyophilization. Thereafter, these membranes covered standardized 5-mm-diameter critical-size defects in calvarial bone on rats. After four weeks of healing, micro-computed tomography and histological analyses using undecalcified thin ground sections were performed. Micro-computed tomography of the inner 4.5 mm calvaria defects revealed a median bone defect coverage of 91% (CI: 87–95) in the ADL group and 94% (CI: 65–100) in the control group, without significant differences between the groups (intergroup p > 0.05). Furthermore, bone volume (BV) was similar between ADL group (5.7 mm3, CI: 3.4–7.1) and control group (5.7 mm3, CI: 2.9–9.7). Histomorphometry of the defect area confirmed these findings with bone area values amounting to 2.1 mm2 (CI: 1.2–2.6) in the ADL group and 2.0 mm2 (CI: 1.1–3.0) in the control group. Together, these data suggest that acid dentin lysate lyophilized onto collagen membranes failed to modulate the robust bone formation when placed onto calvarial defects
Regeneration of alveolar bone defects in the experimental pig model: A systematic review and meta-analysis.
No full textOBJECTIVE
Pigs are emerging as a preferred experimental in vivo model for bone regeneration. The study objective was to answer the focused PEO question: in the pig model (P), what is the capacity of experimental alveolar bone defects (E) for spontaneous regeneration in terms of new bone formation (O)?
METHODS
Following PRISMA guidelines, electronic databases were searched for studies reporting experimental bone defects or extraction socket healing in the maxillae or mandibles of pigs. The main inclusion criteria were the presence of a control group of untreated defects/sockets and the assessment of regeneration via 3D tomography [radiographic defect fill (RDF)] or 2D histomorphometry [new bone formation (NBF)]. Random effects meta-analyses were performed for the outcomes RDF and NBF.
RESULTS
Overall, 45 studies were included reporting on alveolar bone defects or extraction sockets, most frequently in the mandibles of minipigs. Based on morphology, defects were broadly classified as 'box-defects' (BD) or 'cylinder-defects' (CD) with a wide range of healing times (10 days to 52 weeks). Meta-analyses revealed pooled estimates (with 95% confidence intervals) of 50% RDF (36.87%-63.15%) and 43.74% NBF (30.47%-57%) in BD, and 44% RDF (16.48%-71.61%) and 39.67% NBF (31.53%-47.81%) in CD, which were similar to estimates of socket-healing [48.74% RDF (40.35%-57.13%) and 38.73% NBF (28.57%-48.89%)]. Heterogeneity in the meta-analysis was high (I2 > 90%).
CONCLUSION
A substantial body of literature revealed a high capacity for spontaneous regeneration in experimental alveolar bone defects of (mini)pigs, which should be considered in future studies of bone regeneration in this animal model
