Cornelia Ganz

Evaluation of injectable silica-embedded nanohydroxyapatite bone substitute in a rat tibia defect model

In clinical practice, vertebral compression fractures occur after trauma and osteoporosis. Kyphoplasty is a minimally invasive procedure using bone filler material for the treatment of such fractures. A full synthetic injectable bone substitute (SIBS) was manufactured by means of spray drying. The aim of this study was to characterize the SIBS and to analyze the remodelling process during degradation of the biomaterial and new bone formation after implantation. SIBS is an aqueous suspension of donut-like microparticles. These microparticles consist of nanocrystallites of synthetic hydroxyapatite embedded in amorphous silica gel. After implantation of SIBS in a proximal tibial diaphyseal defect in 52 rats, grafts were harvested for subsequent analysis on different days. Newly formed bone originating from endosteum was observed on day 6. Hematomas in the medullary space and cortical wounds disappeared on day 12. The wound region was completely replaced by a composite of newly formed cancellous bone, extracellular matrix, and SIBS. At day 63 the cortical defect was fully healed by bone, while newly formed bone in the medullary space almost disappeared and was replaced with bone marrow. In conclusion, SIBS demonstrated a unique structure with osteoinductive and bioresorbable properties, which induced fast bone regeneration. Therefore, a clinical application of SIBS for kyphoplasty is promising.

In vivo and in vitro investigations of a nanostructured coating material – a preclinical study

Immediate loading of dental implants is only possible if a firm bone-implant anchorage at early stages is developed. This implies early and high bone apposition onto the implant surface. A nanostructured coating material based on an osseoinductive bone grafting is investigated in relation to the osseointegration at early stages. The goal is to transmit the structure (silica matrix with embedded hydroxyapatite) and the properties of the bone grafting into a coating material. The bone grafting substitute offers an osseoinductive potential caused by an exchange of the silica matrix in vivo accompanied by vascularization. X-ray diffraction and transmission electron microscopy analysis show that the coating material consists of a high porous silica matrix with embedded nanocrystalline hydroxyapatite with the same morphology as human hydroxyapatite. An in vitro investigation shows the early interaction between coating and human blood. Energy-dispersive X-ray analysis showed that the silica matrix was replaced by an organic matrix within a few minutes. Uncoated and coated titanium implants were inserted into the femora of New Zealand White rabbits. The bone-to-implant contact (BIC) was measured after 2, 4, and 6 weeks. The BIC of the coated implants was increased significantly at 2 and 4 weeks. After 6 weeks, the BIC was decreased to the level of the control group. A histological analysis revealed high bone apposition on the coated implant surface after 2 and 4 weeks. Osteoblastic and osteoclastic activities on the coating material indicated that the coating participates in the bone-remodeling process. The nanostructure of the coating material led to an exchange of the silica matrix by an autologous, organic matrix without delamination of the coating. This is the key issue in understanding initial bone formation on a coated surface.

Hydrogel-embedded nanocrystalline hydroxyapatite granules (elastic blocks) based on a cross-linked polyvinylpyrrolidone as bone grafting substitute in a rat tibia model

Purpose The aim of this study was to examine the in vivo characteristics and levels of integration and degradation of a ready-to-use bone grafting block with elastic properties (elastic block) for the use in surgery. Materials and methods Thirty-six male Wistar rats underwent surgical creation of a well-defined bone defect in the tibia. All created defects – one per animal – were filled with an unsintered nanocrystalline hydroxyapatite embedded either with a non-cross-linked hydrogel carrier (CONT, n=18) or a cross-linked hydrogel carrier (elastic block [EB], n=18) based on polyvinylpyrrolidone (PVP) and silica sol, respectively. The animals were killed after 12 (n=12), 21 (n=12) and 63 days (n=12). The bone formation and defect healing were quantified by histomorphometric measurements made in paraffin sections. Additionally, immunohistochemical (tartrate-resistant acid phosphatase [TRAP] and alkaline phosphatase [aP]), antibody-based examinations (CD68) and energy-dispersive x-ray scattering measurements of silica atom concentration were carried out. Results A larger remaining bone defect area overall was observed in EB after 12 days and 21 days. After 63 days, similar areas of remaining bone defects were found. The amount of the remaining carrier material in EB overall was higher at all times. In CONT no residual carrier material was found at 12 days and later. CD68 analyses showed significantly lower level of CD68-positive marked cells after 21 days in CONT, and nonsignificant differences at 12 and 63 days, respectively. Additionally, a significantly higher level of aP-positive marked cells was observed in CONT after 12 days. Later on, the levels of aP-positive marked cells were slightly higher in EB (21 and 63 days). Furthermore, no significant differences regarding the level of TRAP-positive marked cells in each group were observed. Conclusion The bone substitute (EB) with the cross-linked PVP-based hydrogel carrier leads at the beginning to a higher amount of remaining carrier material and remaining bone substitute. This delayed degradation is supposed to be the reason for the observed lower level of bone remodeling and is caused by the irradiation changes (cross links) in the structure in PVP.

Comparison of Bone Substitutes in a Tibia Defect Model in Wistar-Rats

Various bone graft substitutes were used in clinical practise in the treatment of bone defects after trauma or osteoporosis. Many synthetic biomaterials were developed in recent years primarily based on hydroxyapatite (HA). NanoBone® is a nanocrystalline hydroxyapatite (HA) embedded in a porous matrix of silica (SiO2). The ratio of HA:SiO2 varied between 76:24 (wt%; NanoBone®) and 61:39 (wt%; Nanobone® S). The two bone substitutes NB and NB S and a natural bovine bone substitute Bio-Oss® (BO) were evaluated by means of implantation in the tibia of the rat. The aim of this study was to analyze the remodelling process and to measure new bone formation and degradation after implantation of these biomaterials. A tibia defect model was used for all investigations with testing periods of 12, 21 and 84 days. (n=5 for each time point). The results showed, that all bone grafts were well accepted by the host tissue without inflammatory reactions. In comparison to the biomaterial BO, NanoBone® and NanoBone® S were quickly degraded, whereas autologous proteins were incorporated into nanopores. New bone formation was statistically higher in NanoBone® S compared to Bio-Oss® in defect area after 84 days implantation. The presence of osteoclasts in tissue sections were demonstrated by TRAP- and ED1-immunohistology.

How to Enhance Osseointegration – Roughness, Hydrophilicity or Bioactive Coating

The apposition of bone at early stages is critical for rapid loading and therefore there is much effort in improving the implant surfaces for a rapid osseointegration. The aim of this study is to investigate the effect of roughness, hydrophilicity and coating on osseointegration. Machined (smooth), sand-blasted (rough), hydrophilic and coated implants were tested in vivo for 2, 4 and 6 weeks. The hydrophilic surfaces were obtained by atmospheric oxygen plasma treatment of machined and sand-blasted implants. The coating is obtained by a spin-spray-process using sol-gel-technique. SEM and TEM investigations revealed that the coating consists of a nanoporous silica matrix with embedded synthetic, nanocrystalline hydroxyapatite. Histological polished sections were manufactured and the bone-to-implant-contact was calculated. The difference between smooth and rough implants was marginal and not significant. There were no statistical differences between hydrophilic and control implants, whereas the BIC of the hydrophilic surfaces was lower by trend. All coated implants offered an increased bone to implant-contact. However, the BIC was decreasing at 6 weeks due to the missing of mechanical stress and a faster bone metabolism in rabbits. The coating offers a new opportunity to enhance the osseointegration and therefore an earlier implant loading.

Bone Substitutes as a Drug Delivery of Antibiotics

The aim of the present study was the in vitro investigation of a synthetic bone graft substitute loaded with individual antibiotics for the treatment of osteomyelitis and infectious bone disease. The elution of gentamicin, an aminoglycoside antibiotic, from the NanoBone® products NanoBone® S granules (NBG) and lyophilized NanoBone® (NBP) putty was tested over a period of one week. An indirect photometrically-based detection system was used to measure the released antibiotic concentration. Both materials showed very different release behaviour. After one day lyophilized NanoBone® putty delivered 100% of the gentamicin value, whereas NanoBone® S granules released one-fifth of the used gentamicin level.

Elastic Blocks: Hydrogel-Embedded Granules as Bone Grafting Substitutes

NanoBone® Block is the alternative to autogenous bone blocks and offers doctors a rapid, simple operating procedure in combination with a low risk of complications. Aim of this work was to develop a bone grafting block with elastic properties for dental and orthopedic surgery which is ready to use. An easy handling has to be connected with quick regeneration. Therefore, NB granules have been combined with an elastic hydrogel carrier based on Polyvinylpyrrolidone (PVP) and silica sol. Mechanical properties were optimized for an enhanced handling by cross linking and simultaneous sterilization of the PVP with gamma radiation. Cross linked PVP has the capability of swelling in water without being solved. To approve biocompatibility of adjusted material an in vivo study was analyzed using a standardized bone defect model in rat tibiae. Defect was filled with elastic bone grafting material. After 21 and 63 days rats were sacrificed and tibias analyzed. Histomorphological analysis showed an increased bone formation but even a decelerated resorption of elastic biomaterial. Quantitative compositional analysis showed a decrease in silicon content of granules as a process of matrix change.

Nanostructured Bone Grafting Substitutes Versus Autologous Cancellous Bone – An Animal Experiment in Sheep

In an In vivo study the full synthetic bone substitute NanoBone® S (NBS) was analyzed using a standardized bone defect (6 x 12 x 24mm) model in the ovine tibial metaphysis. The defect on the left side was filled with NBS granules and on the right side, autologous bone, harvested from the hip of the same animal, was inserted. After six, 12 and 26 weeks sheep were sacrificed and the tibiae analyzed. Quantitative histomorphological analysis after six weeks showed a resorption of biomaterial from over 60 to 24 percent. In contrast the bone formation after 6, and 12 weeks revealed an osteoneogenesis of 19%, and 34%, respectively. Hematoxylin and eosin sections demonstrated multinucleated giant cells on the surface of the biomaterial and resorption lacunae, indicating osteoclastic resorptive activity.

Controlled Self-Coating of Implant Surfaces with Autologous Molecules

mplant fixation is correlated with direct bone apposition on the implant surface. In a former study it was reported that a new coating material enhances the bone-to-implant-contact in comparison to machined and rough surfaces. This study is aimed at clarifying the effect of an enhanced bone-to-implant-contact that is induced by a new coating material. The coating is produced by spin and spray coating and consists of a silica matrix, in which nanocrystalline hydroxyapatite is embedded. The coating material exhibits a high porosity in the micrometer and nanometer scale. Coated implants were inserted subcutaneously in Wistar Rats. The specimens were excised after 6 and 12 days. EDX and SEM analysis showed a reduction of the silica amount within 6 days. In accordance to former results of a bone grafting material with the same structure, this matrix change is responsible for an initial bone formation at early stages.

Bone Grafting Putty – Animal Experiments and Clinical Applications

The aim of the described study was to generate and evaluate a putty-like bone graft substitute ready to use for dental and orthopedic surgery. According to the asking of clinicians the new material should avoid the necessity of mixing blood and bone graft during the surgical process. Therefor the granulous material NanoBone® has been combined with a hydrogel based on Polyvinylpyrrolidone (PVP) and tested in standardized rat tibia defect over a period of 12 weeks and evaluated histologically. The results showed no limitations of the granulate characteristics in matrix change and hence a high level of vascularization and bone formation. An example for dental application shows the outcome in the case of socket preservation.