How to Get Them off?-Assessment of Innovative Techniques for Generation and Detachment of Mature Osteoclasts for Biomaterial Resorption Studies.

The fusion process of mononuclear monocytes into multinuclear osteoclasts in vitro is an essential process for the study of osteoclastic resorption of biomaterials. Thereby biomaterials offer many influencing factors such as sample shape, material composition, and surface topography, which can have a decisive influence on the fusion and thus the entire investigation. For the specific investigation of resorption, it can therefore be advantageous to skip the fusion on samples and use mature, predifferentiated osteoclasts directly. However, most conventional detachment methods (cell scraper, accutase), lead to a poor survival rate of osteoclasts or to a loss of function of the cells after their reseeding. In the present study different conventional and novel methods of detachment in combination with different culture surfaces were investigated to obtain optimal osteoclast differentiation, yield, and vitality rates without loss of function. The innovative method-using thermoresponsive surfaces for cultivation and detachment-was found to be best suited. This is in particular due to its ability to maintain osteoclast activity, as proven by TRAP 5b-, CTSK-activity and resorption pits on dentin discs and decellularized osteoblast-derived matrix plates. In conclusion, it is shown, that osteoclasts can be predifferentiated on cell culture dishes and transferred to a reference biomaterial under preservation of osteoclastic resorption activity, providing biomaterial researchers with a novel tool for material characterization.

Strontium ions promote in vitro human bone marrow stromal cell proliferation and differentiation in calcium-lacking media.

In order to investigate the influence of calcium and strontium ion concentration on human bone marrow stromal cells and their differentiation to osteoblasts, different cell culture media have been used. Even though this study does not contain a bone substitute material, the reason for this study was the decrease of cation concentration by many biomaterials, due to induced apatite precipitation. As a consequence, the reduced calcium ion concentration is known to affect osteoblastic development. Therefore, the main focus was put on the question, whether an increased strontium concentration (in the range of mM) might be suitable to compensate the lack of calcium ions. The effect of solely strontium ions-with only calcium in the media resulting from fetal calf serum-was investigated. Commercially available calcium-free medium (modified α-MEM) was tested in comparison with media with varied calcium ion concentrations (0.9, 1.8, and 3.6 mM), or strontium ion concentration (0.4, 0.9, 1.8, and 3.6 mM). In case of calcium, higher concentrations cause increased proliferation, while differentiation was shifted to earlier points of time. Differentiation was increased by solely strontium ions only at 0.4-0.9 mM, while proliferation was highest for 0.9-1.8 mM. From these results, it can be concluded that strontium is able to compensate a lack of calcium to a certain degree. Thus, in contrast to calcium ion release, a strontium ion release from bone substitute materials might be applicable for stimulation of bone regeneration without influencing the media saturation.

Periosteal microcirculatory reactions in a zoledronate-induced osteonecrosis model of the jaw in rats.

OBJECTIVES: Nitrogen-containing bisphosphonates induce osteonecrosis mostly in the jaw and less frequently in other bones. Because of the crucial role of periosteal perfusion in bone repair, we investigated zoledronate-induced microcirculatory reactions in the mandibular periosteum in comparison with those in the tibia in a clinically relevant model of bisphosphonate-induced medication-related osteonecrosis of the jaw (MRONJ). MATERIALS AND METHODS: Sprague-Dawley rats were treated with zoledronate (ZOL; 80 i.v. µg/kg/week over 8 weeks) or saline vehicle. The first two right mandibular molar teeth were extracted after 3 weeks. Various systemic and local (periosteal) microcirculatory inflammatory parameters were examined by intravital videomicroscopy after 9 weeks. RESULTS: Gingival healing disorders (∼100%) and MRONJ developed in 70% of ZOL-treated cases but not after saline (shown by micro-CT). ZOL induced significantly higher degrees of periosteal leukocyte rolling and adhesion in the mandibular postcapillary venules (at both extraction and intact sites) than at the tibia. Leukocyte NADPH-oxidase activity was reduced; leukocyte CD11b and plasma TNF-alpha levels were unchanged. CONCLUSION: Chronic ZOL treatment causes a distinct microcirculatory inflammatory reaction in the mandibular periosteum but not in the tibia. The local reaction in the absence of augmented systemic leukocyte inflammatory activity suggests that topically different, endothelium-specific changes may play a critical role in the pathogenesis of MRONJ. CLINICAL RELEVANCE: This model permits for the first time to explore the microvascular processes in the mandibular periosteum after chronic ZOL treatment. This approach may contribute to a better understanding of the pathomechanism and the development of strategies to counteract bisphosphonate-induced side effects.

A novel method for in vivo visualization of the microcirculation of the mandibular periosteum in rats.

OBJECTIVE: The periosteum plays an important role in bone physiology, but observation of its microcirculation is greatly limited by methodological constraints at certain anatomical locations. This study was conducted to develop a microsurgical procedure which provides access to the mandibular periosteum in rats. METHODS: Comparisons of the microcirculatory characteristics with those of the tibial periosteum were performed to confirm the functional integrity of the microvasculature. The mandibular periosteum was reached between the facial muscles and the anterior surface of the superficial masseter muscle at the external surface of the mandibular corpus; the tibial periosteum was prepared by dissecting the covering muscles at the anteromedial surface. Intravital fluorescence microscopy was used to assess the leukocyte-endothelial interactions and the RBCV in the tibial and mandibular periosteum. Both structures were also visualized through OPS and fluorescence CLSM. RESULTS: The microcirculatory variables in the mandibular periosteum proved similar to those in the tibia, indicating that no microcirculatory failure resulted from the exposure technique. CONCLUSION: This novel surgical approach provides simple access to the mandibular periosteum of the rat, offering an excellent opportunity for investigations of microcirculatory manifestations of dentoalveolar and maxillofacial diseases.

Osteogenic stimulation of osteoprogenitors by putamen ovi peptides and hyaluronic acid.

Eggshell peptides (EP) majorly contribute to rapid bone building in chicks, wherefore this paper investigated their potential for stimulating osteogenesis in vitro. In this study, the effects of EP, also called putamen ovi peptides and a combination of hyaluronic acid with EP in cell culture medium were tested towards proliferation, differentiation, gene expression and mineralization of bovine osteoprogenitors and primary human osteoblasts. The influence of EP at concentrations of 0.005 g/L, 0.5 g/L and 0.5 g/L with 0.25% hyaluronic acid was analyzed using immunocytochemical staining of bone-specific matrix proteins, namely collagen type I, osteonectin, osteopontin and osteocalcin, to prove osteoblastic differentiation. Additionally, Richardson-staining was performed. All tests revealed a superior osteoblastic differentiation with EP at 0.5 g/L after 5 days of cultivation. Hyaluronic acid alone showed controversial results and partially constrained osteoblastic differentiation in combination with EP to a level as low as for pure EP at 0.005 g/L. Of particular interest is the osteoblast-typical mineralization, as an important indicator of bone formation, which was measured indirectly via the calcium concentration after cultivation over 4 weeks. The mineralization showed an increase by a factor of 286 during the cultivation of primary human osteoblasts with hyaluronic acid and EP. Meanwhile, cell cultures treated with EP (0.5 g/L) only showed an 80-fold increase in calcium concentration.The influence of EP (0.5 g/L) on primary human osteoblasts was investigated by gene expression after 2 weeks of cultivation. Microarray and qRT-PCR analysis showed a strongly increased expression of main important genes in bone formation, bone regeneration and the physiological bone remodelling processes. Namely, BMP 2, osteopontin and the matrix metalloproteinases 1 and 9, were present during in vitro osteoprogenitor culture with EP. By explicitly underlining the potential of eggshell peptides for stimulating osteogenesis, as well as emphasizing complex and controversial interaction with hyaluronan, this manuscript is relevant for developing new functionalized biomaterials for bone regeneration.

Effects of Gamma Irradiation and Supercritical Carbon Dioxide Sterilization on Methacrylated Gelatin/Hyaluronan Hydrogels.

Biopolymer hydrogels have become an important group of biomaterials in experimental and clinical use. However, unlike metallic or mineral materials, they are quite sensitive to sterilization. The aim of this study was to compare the effects of gamma irradiation and supercritical carbon dioxide (scCO2) treatment on the physicochemical properties of different hyaluronan (HA)- and/or gelatin (GEL)-based hydrogels and the cellular response of human bone marrow-derived mesenchymal stem cells (hBMSC). Hydrogels were photo-polymerized from methacrylated HA, methacrylated GEL, or a mixture of GEL/HA. The composition and sterilization methods altered the dissolution behavior of the biopolymeric hydrogels. There were no significant differences in methacrylated GEL release but increased methacrylated HA degradation of gamma-irradiated samples. Pore size/form remained unchanged, while gamma irradiation decreased the elastic modulus from about 29 kPa to 19 kPa compared to aseptic samples. HBMSC proliferated and increased alkaline phosphatase activity (ALP) particularly in aseptic and gamma-irradiated methacrylated GEL/HA hydrogels alike, while scCO2 treatment had a negative effect on both proliferation and osteogenic differentiation. Thus, gamma-irradiated methacrylated GEL/HA hydrogels are a promising base for multi-component bone substitute materials.

A review on the recent progress, opportunities, and challenges of 4D printing and bioprinting in regenerative medicine.

Four-dimensional (4 D) printing is a novel emerging technology, which can be defined as the ability of 3 D printed materials to change their form and functions. The term 'time' is added to 3 D printing as the fourth dimension, in which materials can respond to a stimulus after finishing the manufacturing process. 4 D printing provides more versatility in terms of size, shape, and structure after printing the construct. Complex material programmability, multi-material printing, and precise structure design are the essential requirements of 4 D printing systems. The utilization of stimuli-responsive polymers has increasingly taken the place of cell traction force-dependent methods and manual folding, offering a more advanced technique to affect a construct's adjusted shape transformation. The present review highlights the concept of 4 D printing and the responsive bioinks used in 4 D printing, such as water-responsive, pH-responsive, thermo-responsive, and light-responsive materials used in tissue regeneration. Cell traction force methods are described as well. Finally, this paper aims to introduce the limitations and future trends of 4 D printing in biomedical applications based on selected key references from the last decade.

Ketoprofen-Based Polymer-Drug Nanoparticles Provide Anti-Inflammatory Properties to HA/Collagen Hydrogels.

Current limitations of wound dressings for treating chronic wounds require the development of novel approaches. One of these is the immune-centered approach, which aims to restore the pro-regenerative and anti-inflammatory properties of macrophages. Under inflammatory conditions, ketoprofen nanoparticles (KT NPs) can reduce pro-inflammatory markers of macrophages and increase anti-inflammatory cytokines. To assess their suitability as part of wound dressings, these NPs were combined with hyaluronan (HA)/collagen-based hydro- (HGs) and cryogels (CGs). Different HA and NP concentrations and loading techniques for NP incorporation were used. The NP release, gel morphology, and mechanical properties were studied. Generally, colonialization of the gels with macrophages resulted in high cell viability and proliferation. Furthermore, direct contact of the NPs to the cells reduced the level of nitric oxide (NO). The formation of multinucleated cells on the gels was low and further decreased by the NPs. For the HGs that produced the highest reduction in NO, extended ELISA studies showed reduced levels of the pro-inflammatory markers PGE2, IL-12 p40, TNF-α, and IL-6. Thus, HA/collagen-based gels containing KT NPs may represent a novel therapeutic approach for treating chronic wounds. Whether effects observed in vitro translate into a favorable profile on skin regeneration in vivo will require rigorous testing.

Covalent linkage of sulfated hyaluronan to the collagen scaffold Mucograft® enhances scaffold stability and reduces proinflammatory macrophage activation in vivo.

Sulfated glycosaminoglycans (sGAG) show interaction with biological mediator proteins. Although collagen-based biomaterials are widely used in clinics, their combination with high-sulfated hyaluronan (sHA3) is unexplored. This study aims to functionalize a collagen-based scaffold (Mucograft®) with sHA3 via electrostatic (sHA3/PBS) or covalent binding to collagen fibrils (sHA3+EDC/NHS). Crosslinking without sHA3 was used as a control (EDC/NHS Ctrl). The properties of the sHA3-functionalized materials were characterized. In vitro growth factor and cytokine release after culturing with liquid platelet-rich fibrin was performed by means of ELISA. The cellular reaction to the biomaterials was analyzed in a subcutaneous rat model. The study revealed that covalent linking of sHA3 to collagen allowed only a marginal release of sHA3 over 28 days in contrast to electrostatically bound sHA3. sHA3+EDC/NHS scaffolds showed reduced vascular endothelial growth factor (VEGF), transforming growth factor beta 1 (TGF-ß1) and enhanced interleukin-8 (IL-8) and epithelial growth factor (EGF) release in vitro compared to the other scaffolds. Both sHA3/PBS and EDC/NHS Ctrl scaffolds showed a high proinflammatory reaction (M1: CD-68+/CCR7+) and induced multinucleated giant cell (MNGC) formation in vivo. Only sHA3+EDC/NHS scaffolds reduced the proinflammatory macrophage M1 response and did not induce MNGC formation during the 30 days. SHA3+EDC/NHS scaffolds had a stable structure in vivo and showed sufficient integration into the implantation region after 30 days, whereas EDC/NHS Ctrl scaffolds underwent marked disintegration and lost their initial structure. In summary, functionalized collagen (sHA3+EDC/NHS) modulates the inflammatory response and is a promising biomaterial as a stable scaffold for full-thickness skin regeneration in the future.

3D Plotting of Silica/Collagen Xerogel Granules in an Alginate Matrix for Tissue-Engineered Bone Implants.

Today, materials designed for bone regeneration are requested to be degradable and resorbable, bioactive, porous, and osteoconductive, as well as to be an active player in the bone-remodeling process. Multiphasic silica/collagen Xerogels were shown, earlier, to meet these requirements. The aim of the present study was to use these excellent material properties of silica/collagen Xerogels and to process them by additive manufacturing, in this case 3D plotting, to generate implants matching patient specific shapes of fractures or lesions. The concept is to have Xerogel granules as active major components embedded, to a large proportion, in a matrix that binds the granules in the scaffold. By using viscoelastic alginate as matrix, pastes of Xerogel granules were processed via 3D plotting. Moreover, alginate concentration was shown to be the key to a high content of irregularly shaped Xerogel granules embedded in a minimum of matrix phase. Both the alginate matrix and Xerogel granules were also shown to influence viscoelastic behavior of the paste, as well as the dimensionally stability of the scaffolds. In conclusion, 3D plotting of Xerogel granules was successfully established by using viscoelastic properties of alginate as matrix phase.

Strontium substitution of gelatin modified calcium hydrogen phosphates as porous hard tissue substitutes.

Aiming at the generation of a high strontium-containing degradable bone substitute, the exchange of calcium with strontium in gelatin-modified brushite was investigated. The ion substitution showed two mineral groups, the high-calcium containing minerals with a maximum measured molar Ca/Sr ratio of 80%/20% (mass ratio 63%/37%) and the high-strontium containing ones with a maximum measured molar Ca/Sr ratio of 21%/79% (mass ratio 10%/90%). In contrast to the high-strontium mineral phases, a high mass loss was observed for the calcium-based minerals during incubation in cell culture medium (alpha-MEM), but also an increase in strength owing to dissolution and re-precipitation. This resulted for the former in a decrease of cation concentration (Ca + Sr) in the medium, while the pH value decreased and the phosphate ion concentration rose significantly. The latter group of materials, the high-strontium containing ones, showed only a moderate change in mass and a decrease in strength, but the Ca + Sr concentration remained permanently above the initial calcium concentration in the medium. This might be advantageous for a future planned application by supporting bone regeneration on the cellular level.

Chemically Modified Biomimetic Carbon-Coated Iron Nanoparticles for Stent Coatings: In Vitro Cytocompatibility and In Vivo Structural Changes in Human Atherosclerotic Plaques.

Atherosclerosis, a systematic degenerative disease related to the buildup of plaques in human vessels, remains the major cause of morbidity in the field of cardiovascular health problems, which are the number one cause of death globally. Novel atheroprotective HDL-mimicking chemically modified carbon-coated iron nanoparticles (Fe@C NPs) were produced by gas-phase synthesis and modified with organic functional groups of a lipophilic nature. Modified and non-modified Fe@C NPs, immobilized with polycaprolactone on stainless steel, showed high cytocompatibility in human endothelial cell culture. Furthermore, after ex vivo treatment of native atherosclerotic plaques obtained during open carotid endarterectomy surgery, Fe@C NPs penetrated the inner structures and caused structural changes of atherosclerotic plaques, depending on the period of implantation in Wistar rats, serving as a natural bioreactor. The high biocompatibility of the Fe@C NPs shows great potential in the treatment of atherosclerosis disease as an active substance of stent coatings to prevent restenosis and the formation of atherosclerotic plaques.

Organically modified hydroxyapatite (ormoHAP) nanospheres stimulate the differentiation of osteoblast and osteoclast precursors: a co-culture study.

Isolated nanospheres consisting of organically modified hydroxyapatite (ormoHAP), prepared by an electric field-assisted ion double migration process, were embedded in foamed gelatin to form a composite scaffold. Degradation rates have been demonstrated to correlate with the crosslinking degree (40%, 80%) as well as with the mineral content of the scaffolds (0%, 20%, 40%). A human co-culture model of osteoblasts and osteoclasts, derived from bone marrow stromal cells and monocytes, respectively, without external addition of the factors RANKL and M-CSF, was run for up to 42 d in order to characterize the action of the ormoHAP-gelatin scaffolds on the co-culture. Examination was performed by quantitative biochemical methods (DNA, LDH, ALP, TRAP5b), gene expression analysis (ALP, BSP II, RANKL, IL-6, VTNR, CTSK, TRAP, OSCAR, CALCR) and confocal laser scanning microscopy (cell nuclei, actin, CD68, TRAP). Results confirm that ormoHAP embedded in the gelatin matrix enhanced TRAP 5b activity. As a feedback, ALP activity and gene expression of BSP II of osteoblasts increased. Finally, a sequence of cell cross-talk actions is suggested, which can explain the behavior of the formed vital co-culture and moreover the influence of the presence and concentration of ormoHAP.

Biomaterial based treatment of osteoclastic/osteoblastic cell imbalance - Gelatin-modified calcium/strontium phosphates.

Osteoporotic bone represents - particularly in case of fractures - difficult conditions for its regeneration. In the present study, the focus was put on a degradable bone substitute material of gelatin-modified calcium and strontium phosphates facing the special demands of osteoporotic bone. The release of strontium ions from the material ought to stimulate osteoblastogenesis either direct by ion release or indirect after material resorption by increased presence and activity of osteoclasts, which subsequently stimulate osteoblasts. A new porous material was produced from calcium phosphate, strontium phosphate and a mixed phase of calcium/strontium phosphate precipitated in presence of gelatin. Initially, ion release was analyzed in standard­calcium containing (2.0 mM) and low-calcium (0.4 mM) minimum essential medium. The cultivation of human peripheral blood mononuclear cells next to the material led to formation of osteoclast-like cells, able to migrate, fuse, and differentiate. Especially, the mixed gelatin-modified calcium/strontium phosphate allowed osteoclastogenesis as proven morphologically and by real-time quantitative polymerase chain reaction (RT-qPCR). It was precisely this material that led to the best osteoblastic reaction of human bone marrow stromal cells cultured on the material. The investigations of the bone substitute material indicate active involvement in the balance of cells of the bone morphogenetic unit.

Odontoblast-like differentiation and mineral formation of pulpsphere derived cells on human root canal dentin in vitro.

BACKGROUND: The revitalization or regeneration of the dental pulp is a preferable goal in current endodontic research. In this study, human dental pulp cell (DPC) spheres were applied to human root canal samples to evaluate their potential adoption for physiological tissue-like regeneration of the dental root canal by odontoblastic differentiation as well as cell-induced mineral formation. METHODS: DPC were cultivated into three-dimensional cell spheres and seeded on human root canal specimens. The evaluation of sphere formation, tissue-like behavior and differentiation as well as mineral formation of the cells was carried out with the aid of optical light microscopy, immunohistochemical staining and scanning electron microscopy (SEM). RESULTS: Spheres and cells migrated out of the spheres showed an intense cell-cell- and cell-dentin-contact with the formation of extra cellular matrix. In addition, the ingrowth of cell processes into dentinal tubules and the interaction of cell processes with the tubule walls were detected by SEM-imaging. Immunohistochemical staining of the odontoblast specific matrix proteins, dentin matrix protein-1, and dentin sialoprotein revealed an odontoblast-like cell differentiation in contact with the dentin surface. This differentiation was confirmed by SEM-imaging of cells with an odontoblast specific phenotype and cell induced mineral formation. CONCLUSIONS: The results of the present study reveal the high potential of pulp cells organized in spheres for dental tissue engineering. The odontoblast-like differentiation and the cell induced mineral formation display the possibility of a complete or partial "dentinal filling" of the root canal and the opportunity to combine this method with other current strategies.

Cell-based bone reconstruction therapies--cell sources.

Over the last few decades, reconstructive surgery has shifted from a resection-oriented approach toward strategies focusing on repair and regeneration of tissues. As the main aim of maxillofacial reconstruction has been the restoration of bone form and function, surgeons used artificial tissue substitutes in the early decades of bone reconstruction. These artificial materials significantly improved the ability of surgeons to restore the form and, to some extent, the function of defective bones. Despite the fact that every artificial material has specific disadvantages, the use of biomaterials is a common treatment option in clinical practice even today. Due to the more detailed understanding that exists concerning transplantation of cells and tissues, autogenous grafts are the second mainstay in clinical practice. However, the main disadvantage of using autogenous grafts is donor site morbidity and donor shortage. Research is currently in progress into the use of cell-based approaches in reconstructive surgery, since cells are the driving elements for all repair and regeneration processes. Various cell populations have been reported on in the relevant literature. These cells can be classified according to differentiation capacity and the tissue from which they originated. In this review, unrestricted cells, multipotential progenitor cells, determined cells, and genetically modified cells are described systematically, and their advantages as well as limitations are discussed. (More than 50 references.)

Cell-based bone reconstruction therapies-principles of clinical approaches.

Cell-based bone tissue engineering is a rapidly evolving therapy option in bone reconstruction strategies. Some cell-driven approaches, especially the biophysical stimulation of the host cell population surrounded by the bone defect, are common treatment methods in maxillofacial surgery. Others, such as autologous cell implantation, have now gained acceptance for clinical trials. More advanced or complex therapeutical options (extracorporeal tissue engineering, stem cell use, genetic engineering) have been tested in preclinical investigations but have not reached the level of clinical use. Two different aspects are of special relevance in cell-based bone reconstruction therapies. The source of cells used to regenerate bone (discussed in detail in a complementary review in this issue of The International Journal of Oral and Maxillofacial Implants) as well as the principal approach of a cell-driven bone regeneration therapy influence the outcome of such engineering strategies. All of the cell-driven repair strategies are under intensive investigation in an effort to provide surgeons with a limitless supply of tissue for bone repair and reconstruction in future procedures. An overview of the basic biological aspects as well as the inherent constraints of different cell-based approaches are given in this paper.

Development and evaluation of an internet-based blended-learning module in biomedicine for university applicants--Education as a challenge for the future.

BACKGROUND: Biomedical science, especially biomaterials, is an expanding field in medicine. Universities are being challenged to gain the best students for a later academic career. Pre-university assessment of pupils has become crucial to reach this aim. Blended learning is an emerging paradigm for science education even though it has not yet been rigorously assessed, especially in the pupil/undergraduate situation. The aim of the study was to develop and preliminarily test a blended-learning system in biomedicine for university applicants. METHODS: An internet-based blended-learning module in material science was developed in close collaboration between a university (Biomaterials Department, Dresden TU), a German Gymnasium and an internationally oriented medical college (IMC®, Münster). Forty pre-university students were taught by this learning module composed of school education and internet-based knowledge transfer and involved in the evaluation of the utility of this learning tool. Finally, the students took first-year university examinations in order to evaluate the success of this kind of education. RESULTS: The internet-based blended-learning module as a combination of e-learning tutorials and live online lectures which was applied in phase 3 of this study was developed on the basis of the findings of both pre-university studies. The results of the learning behavior regarding the number of invokes and the dwell time of the individual pages of the pre-university learning material, the results of the online evaluation and the results of the pre-phase examination were successively used to optimize the next phase. At the end of the pre-university learning, seven of eight participants were able to pass the first-year university examination followed by nationally accepted credit award. CONCLUSION: Internet-based blended-learning module proved to be suitable to prepare students for biomedical university education while also giving them the possibility to assess their qualifications for studying biomedicine and subsequent scientific careers. Moreover, the module can help universities to find the best students.

Gelatine modified monetite as a bone substitute material: An in vitro assessment of bone biocompatibility.

Calcium phosphate phases are increasingly used for bone tissue substitution, and the load bearing properties of these inherently brittle biomaterials are increased by inclusion of organic components. Monetite prepared using mineralization of gelatine pre-structured through phosphate leads to a significantly increased biaxial strength and indirect tensile strength compared to gelatine-free monetite. Besides the mechanical properties, degradation in physiological solutions and osteoblast and osteoclast cell response were investigated. Human bone marrow stromal cells (hBMSCs) showed considerably higher proliferation rates on the gelatine modified monetite than on polystyrene reference material in calcium-free as well as standard cell culture medium (α-MEM). Osteogenic differentiation on the material was comparable to polystyrene in both medium types. Osteoclast-like cells derived from monocytes were able to actively resorb the biomaterial. Osteoblastic differentiation and perhaps even more important the cellular resorption of the biomaterial indicate that it can be actively involved in the bone remodeling process. Thus the behavior of osteoblasts and osteoclasts as well as the adequate degradation and mechanical properties are strong indicators for bone biocompatibility, although in vivo studies are still required to prove this. STATEMENT OF SIGNIFICANCE: New and unique? A low temperature precipitationprocessforcalcium anhydrous hydrogen phosphateallows for the first time to produce monolithic compact composites of monetite and gelatine. The composite is degradable and resorbable. To prove that, the question arises: what is bone biocompatibility? The reaction of both mayor cell types of bone represents this biocompatibility. Therefore, human bone marrow stromal cells were seeded revealing the materials pro-osteogenic properties. Monocyte cultivation, becoming recently focus of interest, revealed the capability of the biomaterial to be actively resorbed by derived osteoclast-like cells. Not new but necessary ismechanical characterization, which is often only investigated as uniaxial property. Here, a biaxial method is applied, to characterize the materials properties closer to its application loads.

Osteoblast alignment, elongation and migration on grooved polystyrene surfaces patterned by Langmuir-Blodgett lithography.

Topographically patterned surfaces are known to influence cellular behavior in a controllable manner. However, the relatively large surface areas (several cm2) required for many biomaterial applications are beyond the practical limits of traditional lithography. Langmuir-Blodgett lithography, a recently developed method, was used to fabricate regularly spaced grooves of different depths (50 and 150 nm) with a periodicity of 500 nm over several square centimeter on silicon surfaces. These topographies were transferred into polystyrene surfaces by means of nanoimprinting. Primary osteoblasts were cultured on the patterned polymer surfaces. They were observed to align, elongate and migrate parallel to the grooves. The combination of Langmuir-Blodgett lithography with nanoimprinting enables the fabrication of large, nanostructured surface areas on a wide spectrum of different biomaterials. Osteoblasts show a significant anisotropic behavior to these surfaces, which can enhance cell settlement on the surface or be used to direct tissue generation on the biomaterial interface.