[Scaffold-based Bone Tissue Engineering]. / Gerüstträgerbasiertes Knochen-Tissue-Engineering.

Tissue engineering provides the possibility of regenerating damaged or lost osseous structures without the need for permanent implants. Within this context, biodegradable and bioresorbable scaffolds can provide structural and biomechanical stability until the body's own tissue can take over their function. Additive biomanufacturing makes it possible to design the scaffold's architectural characteristics to specifically guide tissue formation and regeneration. Its nano-, micro-, and macro-architectural properties can be tailored to ensure vascularization, oxygenation, nutrient supply, waste exchange, and eventually ossification not only in its periphery but also in its center, which is not in direct contact with osteogenic elements of the surrounding healthy tissue. In this article we provide an overview about our conceptual design and process of the clinical translation of scaffold-based bone tissue engineering applications.

Tissue engineered periodontal products.

Attainment of periodontal regeneration is a significant clinical goal in the management of advanced periodontal defects arising from periodontitis. Over the past 30 years numerous techniques and materials have been introduced and evaluated clinically and have included guided tissue regeneration, bone grafting materials, growth and other biological factors and gene therapy. With the exception of gene therapy, all have undergone evaluation in humans. All of the products have shown efficacy in promoting periodontal regeneration in animal models but the results in humans remain variable and equivocal concerning attaining complete biological regeneration of damaged periodontal structures. In the early 2000s, the concept of tissue engineering was proposed as a new paradigm for periodontal regeneration based on molecular and cell biology. At this time, tissue engineering was a new and emerging field. Now, 14 years later we revisit the concept of tissue engineering for the periodontium and assess how far we have come, where we are currently situated and what needs to be done in the future to make this concept a reality. In this review, we cover some of the precursor products, which led to our current position in periodontal tissue engineering. The basic concepts of tissue engineering with special emphasis on periodontal tissue engineering products is discussed including the use of mesenchymal stem cells in bioscaffolds and the emerging field of cell sheet technology. Finally, we look into the future to consider what CAD/CAM technology and nanotechnology will have to offer.

Biomimetic tubular nanofiber mesh and platelet rich plasma-mediated delivery of BMP-7 for large bone defect regeneration.

There is a growing need for successful bone tissue engineering strategies and advanced biomaterials that mimic the structure and function of native tissues carry great promise. Successful bone repair approaches may include an osteoconductive scaffold, osteoinductive growth factors, cells with an osteogenic potential and capacity for graft vascularisation. To increase osteoinductivity of biomaterials, the local combination and delivery of growth factors has been developed. In the present study we investigated the osteogenic effects of calcium phosphate (CaP)-coated nanofiber mesh tube-mediated delivery of BMP-7 from a PRP matrix for the regeneration of critical sized segmental bone defects in a small animal model. Bilateral full-thickness diaphyseal segmental defects were created in twelve male Lewis rats and nanofiber mesh tubes were placed around the defect. Defects received either treatment with a CaP-coated nanofiber mesh tube (n = 6), an un-coated nanofiber mesh tube (n=6) a CaP-coated nanofiber mesh tube with PRP (n=6) or a CaP-coated nanofiber mesh tube in combination with 5 µg BMP-7 and PRP (n = 6). After 12 weeks, bone volume and biomechanical properties were evaluated using radiography, microCT, biomechanical testing and histology. The results demonstrated significantly higher biomechanical properties and bone volume for the BMP group compared to the control groups. These results were supported by the histological evaluations, where BMP group showed the highest rate of bone regeneration within the defect. In conclusion, BMP-7 delivery via PRP enhanced functional bone defect regeneration, and together these data support the use of BMP-7 in the treatment of critical sized defects.

Dynamic compression improves biosynthesis of human zonal chondrocytes from osteoarthritis patients.

OBJECTIVE: We hypothesize that chondrocytes from distinct zones of articular cartilage respond differently to compressive loading, and that zonal chondrocytes from osteoarthritis (OA) patients can benefit from optimized compressive stimulation. Therefore, we aimed to determine the transcriptional response of superficial (S) and middle/deep (MD) zone chondrocytes to varying dynamic compressive strain and loading duration. To confirm effects of compressive stimulation on overall matrix production, we subjected zonal chondrocytes to compression for 2 weeks. DESIGN: Human S and MD chondrocytes from osteoarthritic joints were encapsulated in 2% alginate, pre-cultured, and subjected to compression with varying dynamic strain (5, 15, 50% at 1 Hz) and loading duration (1, 3, 12 h). Temporal changes in cartilage-specific, zonal, and dedifferentiation genes following compression were evaluated using quantitative real-time reverse transcriptase polymerase chain reaction (qRT-PCR). The benefits of long-term compression (50% strain, 3 h/day, for 2 weeks) were assessed by measuring construct glycosaminoglycan (GAG) content and compressive moduli, as well as immunostaining. RESULTS: Compressive stimulation significantly induced aggrecan (ACAN), COL2A1, COL1A1, proteoglycan 4 (PRG4), and COL10A1 gene expression after 2 h of unloading, in a zone-dependent manner (P < 0.05). ACAN and PRG4 mRNA levels depended on strain and load duration, with 50% and 3 h loading resulting in highest levels (P < 0.05). Long-term compression increased collagen type II and ACAN immunostaining and total GAG (P < 0.05), but only S constructs showed more PRG4 stain, retained more GAG (P < 0.01), and developed higher compressive moduli than non-loaded controls. CONCLUSIONS: The biosynthetic activity of zonal chondrocytes from osteoarthritis joints can be enhanced with selected compression regimes, indicating the potential for cartilage tissue engineering applications.

Comparative study of depth-dependent characteristics of equine and human osteochondral tissue from the medial and lateral femoral condyles.

Articular cartilage defects are common after joint injuries. When left untreated, the biomechanical protective function of cartilage is gradually lost, making the joint more susceptible to further damage, causing progressive loss of joint function and eventually osteoarthritis (OA). In the process of translating promising tissue-engineering cartilage repair approaches from bench to bedside, pre-clinical animal models including mice, rabbits, goats, and horses, are widely used. The equine species is becoming an increasingly popular model for the in vivo evaluation of regenerative orthopaedic approaches. As there is also an increasing body of evidence suggesting that successful lasting tissue reconstruction requires an implant that mimics natural tissue organization, it is imperative that depth-dependent characteristics of equine osteochondral tissue are known, to assess to what extent they resemble those in humans. Therefore, osteochondral cores (4-8 mm) were obtained from the medial and lateral femoral condyles of equine and human donors. Cores were processed for histology and for biochemical quantification of DNA, glycosaminoglycan (GAG) and collagen content. Equine and human osteochondral tissues possess similar geometrical (thickness) and organizational (GAG, collagen and DNA distribution with depth) features. These comparable trends further underscore the validity of the equine model for the evaluation of regenerative approaches for articular cartilage.

[Bone tissue engineering. Reconstruction of critical sized segmental bone defects in the ovine tibia]. / Knochen-Tissue-Engineering. Rekonstruktion segmentaler Knochendefekte kritischer Größe in der Schafstibia.

Well-established therapies for bone defects are restricted to bone grafts which face significant disadvantages (limited availability, donor site morbidity, insufficient integration). Therefore, the objective was to develop an alternative approach investigating the regenerative potential of medical grade polycaprolactone-tricalcium phosphate (mPCL-TCP) and silk-hydroxyapatite (silk-HA) scaffolds.Critical sized ovine tibial defects were created and stabilized. Defects were left untreated, reconstructed with autologous bone grafts (ABG) and mPCL-TCP or silk-HA scaffolds. Animals were observed for 12 weeks. X-ray analysis, torsion testing and quantitative computed tomography (CT) analyses were performed. Radiological analysis confirmed the critical nature of the defects. Full defect bridging occurred in the autograft and partial bridging in the mPCL-TCP group. Only little bone formation was observed with silk-HA scaffolds. Biomechanical testing revealed a higher torsional moment/stiffness (p < 0.05) and CT analysis a significantly higher amount of bone formation for the ABG group when compared to the silk-HA group. No significant difference was determined between the ABG and mPCL-TCP groups. The results of this study suggest that mPCL-TCP scaffolds combined can serve as an alternative to autologous bone grafting in long bone defect regeneration. The combination of mPCL-TCP with osteogenic cells or growth factors represents an attractive means to further enhance bone formation.

Water as the often neglected medium at the interface between materials and biology.

Despite its apparent simplicity, water behaves in a complex manner and is fundamental in controlling many physical, chemical and biological processes. The molecular mechanisms underlying interaction of water with materials, particularly polymer networks such as hydrogels, have received much attention in the research community. Despite this, a large gulf still exists in applying what is known to rationalize how the molecular organization of water on and within these materials impacts biological processes. In this perspective, we outline the importance of water in biomaterials science as a whole and give indications for future research directions towards emergence of a complete picture of water, materials and biology.

CD73 and CD29 concurrently mediate the mechanically induced decrease of migratory capacity of mesenchymal stromal cells.

e assumption that mesenchymal stromal cell (MSC)-based-therapies are capable of augmenting physiological regeneration processes has fostered intensive basic and clinical research activities. However, to achieve sustained therapeutic success in vivo, not only the biological, but also the mechanical microenvironment of MSCs during these regeneration processes needs to be taken into account. This is especially important for e.g., bone fracture repair, since MSCs present at the fracture site undergo significant biomechanical stimulation. This study has therefore investigated cellular characteristics and the functional behaviour of MSCs in response to mechanical loading. Our results demonstrated a reduced expression of MSC surface markers CD73 (ecto-5'-nucleotidase) and CD29 (integrin ß1) after loading. On the functional level, loading led to a reduced migration of MSCs. Both effects persisted for a week after the removal of the loading stimulus. Specific inhibition of CD73/CD29 demonstrated their substrate dependent involvement in MSC migration after loading. These results were supported by scanning electron microscopy images and phalloidin staining of actin filaments displaying less cell spreading, lamellipodia formation and actin accumulations. Moreover, focal adhesion kinase and Src-family kinases were identified as candidate downstream targets of CD73/CD29 that might contribute to the mechanically induced decrease in MSC migration. These results suggest that MSC migration is controlled by CD73/CD29, which in turn are regulated by mechanical stimulation of cells. We therefore speculate that MSCs migrate into the fracture site, become mechanically entrapped, and thereby accumulate to fulfil their regenerative functions.

Targeted 2D histology and ultrastructural bone analysis based on 3D microCT anatomical locations.

Histological processing of mineralised tissue (e.g. bone) allows examining the anatomy of cells and tissues as well as the material properties of the tissue. However, resin-embedding offers limited control over the specimen position for cutting. Moreover, specific anatomical planes (coronal, sagittal) or defined landmarks are often missed with standard microtome sectioning. Here we describe a method to precisely locate a specific anatomical 2D plane or any anatomical feature of interest (e.g. bone lesions, newly formed bone, etc.) using 3D micro computed tomography (microCT), and to expose it using controlled-angle microtome cutting. The resulting sections and corresponding specimen's block surface offer correlative information of the same anatomical location, which can then be analysed using multiscale imaging. Moreover, this method can be combined with immunohistochemistry (IHC) to further identify any component of the bone microenvironment (cells, extracellular matrix, proteins, etc.) and guide subsequent in-depth analysis. Overall, this method allows to:•Cut your specimens in a consistent position and precise manner using microCT-based controlled-angle microtome sectioning.•Locate and expose a specific anatomical plane (coronal, sagittal plane) or any other anatomical landmarks of interest based on microCT.•Identify any cell or tissue markers based on IHC to guide further in-depth examination of those regions of interest.

Formalin fixation affects equilibrium partitioning of an ionic contrast agent-microcomputed tomography (EPIC-µCT) imaging of osteochondral samples.

OBJECTIVE: Equilibrium Partitioning of an Ionic Contrast agent with microcomputed tomography (EPIC-µCT) is a non-invasive technique to quantify and visualize the three-dimensional distribution of glycosaminoglycans (GAGs) in fresh cartilage tissue. However, it is unclear whether this technique is applicable to already fixed tissues. Therefore, this study aimed at investigating whether formalin fixation of bovine cartilage affects X-ray attenuation, and thus the interpretation of EPIC-µCT data. DESIGN: Osteochondral samples (n=24) were incubated with ioxaglate, an ionic contrast agent, for 22h prior to µCT scanning. The samples were scanned in both formalin-fixed and fresh conditions. GAG content was measured using a biochemical assay and normalized to wet weight, dry weight, and water content to determine potential reasons for differences in X-ray attenuation. RESULTS: The expected zonal distribution of contrast agent/GAGs was observed for both fixed and fresh cartilage specimens. However, despite no significant differences in GAG concentrations or physical properties between fixed and fresh samples, the average attenuation levels of formalin-fixed cartilage were 14.3% lower than in fresh samples. CONCLUSIONS: EPIC-µCT is useful for three-dimensional visualization of GAGs in formalin-fixed cartilage. However, a significant reduction in X-ray attenuation for fixed (compared to fresh) cartilage must be taken into account and adjusted for accordingly when quantifying GAG concentrations using EPIC-µCT.

Human and mouse bones physiologically integrate in a humanized mouse model while maintaining species-specific ultrastructure.

Humanized mouse models are increasingly studied to recapitulate human-like bone physiology. While human and mouse bone architectures differ in multiple scales, the extent to which chimeric human-mouse bone physiologically interacts and structurally integrates remains unknown. Here, we identify that humanized bone is formed by a mosaic of human and mouse collagen, structurally integrated within the same bone organ, as shown by immunohistochemistry. Combining this with materials science techniques, we investigate the extracellular matrix of specific human and mouse collagen regions. We show that human-like osteocyte lacunar-canalicular network is retained within human collagen regions and is distinct to that of mouse tissue. This multiscale analysis shows that human and mouse tissues physiologically integrate into a single, functional bone tissue while maintaining their species-specific ultrastructural differences. These results offer an original method to validate and advance tissue-engineered human-like bone in chimeric animal models, which grow to be eloquent tools in biomedical research.

Cancer-associated fibroblasts of the prostate promote a compliant and more invasive phenotype in benign prostate epithelial cells.

Reciprocal interactions between prostate epithelial cells and their adjacent stromal microenvironment not only are essential for tissue homeostasis but also play a key role in tumor development and progression. Malignant transformation is associated with the formation of a reactive stroma where cancer-associated fibroblasts (CAFs) induce matrix remodeling and thereby provide atypical biochemical and biomechanical signals to epithelial cells. Previous work has been focused on the cellular and molecular phenotype as well as on matrix stiffness and remodeling, providing potential targets for cancer therapeutics. So far, biomechanical changes in CAFs and adjacent epithelial cells of the prostate have not been explored. Here, we compared the mechanical properties of primary prostatic CAFs and patient-matched non-malignant prostate tissue fibroblasts (NPFs) using atomic force microscopy (AFM) and real-time deformability cytometry (RT-FDC). It was found that CAFs exhibit an increased apparent Young's modulus, coinciding with an altered architecture of the cytoskeleton compared with NPFs. In contrast, co-cultures of benign prostate epithelial (BPH-1) cells with CAFs resulted in a decreased stiffness of the epithelial cells, as well as an elongated morphological phenotype, when compared with co-cultures with NPFs. Moreover, the presence of CAFs increased proliferation and invasion of epithelial cells, features typically associated with tumor progression. Altogether, this study provides novel insights into the mechanical interactions between epithelial cells with the malignant prostate microenvironment, which could potentially be explored for new diagnostic approaches.

Characterisation and evaluation of the regenerative capacity of Stro-4+ enriched bone marrow mesenchymal stromal cells using bovine extracellular matrix hydrogel and a novel biocompatible melt electro-written medical-grade polycaprolactone scaffold.

Many skeletal tissue regenerative strategies centre around the multifunctional properties of bone marrow derived stromal cells (BMSC) or mesenchymal stem/stromal cells (MSC)/bone marrow derived skeletal stem cells (SSC). Specific identification of these particular stem cells has been inconclusive. However, enriching these heterogeneous bone marrow cell populations with characterised skeletal progenitor markers has been a contributing factor in successful skeletal bone regeneration and repair strategies. In the current studies we have isolated, characterised and enriched ovine bone marrow mesenchymal stromal cells (oBMSCs) using a specific antibody, Stro-4, examined their multipotential differentiation capacity and, in translational studies combined Stro-4+ oBMSCs with a bovine extracellular matrix (bECM) hydrogel and a biocompatible melt electro-written medical-grade polycaprolactone scaffold, and tested their bone regenerative capacity in a small in vivo, highly vascularised, chick chorioallantoic membrane (CAM) model and a preclinical, critical-sized ovine segmental tibial defect model. Proliferation rates and CFU-F formation were similar between unselected and Stro-4+ oBMSCs. Col1A1, Col2A1, mSOX-9, PPARG gene expression were upregulated in respective osteogenic, chondrogenic and adipogenic culture conditions compared to basal conditions with no significant difference between Stro-4+ and unselected oBMSCs. In contrast, proteoglycan expression, alkaline phosphatase activity and adipogenesis were significantly upregulated in the Stro-4+ cells. Furthermore, with extended cultures, the oBMSCs had a predisposition to maintain a strong chondrogenic phenotype. In the CAM model Stro-4+ oBMSCs/bECM hydrogel was able to induce bone formation at a femur fracture site compared to bECM hydrogel and control blank defect alone. Translational studies in a critical-sized ovine tibial defect showed autograft samples contained significantly more bone, (4250.63 mm3, SD = 1485.57) than blank (1045.29 mm3, SD = 219.68) ECM-hydrogel (1152.58 mm3, SD = 191.95) and Stro-4+/ECM-hydrogel (1127.95 mm3, SD = 166.44) groups. Stro-4+ oBMSCs demonstrated a potential to aid bone repair in vitro and in a small in vivo bone defect model using select scaffolds. However, critically, translation to a large related preclinical model demonstrated the complexities of bringing small scale reported stem-cell material therapies to a clinically relevant model and thus facilitate progression to the clinic.

Periodontal Tissue Engineering with a Multiphasic Construct and Cell Sheets.

This study reports on scaffold-based periodontal tissue engineering in a large preclinical animal model. A biphasic scaffold consisting of bone and periodontal ligament compartments manufactured by melt and solution electrospinning, respectively, was used for the delivery of in vitro matured cell sheets from 3 sources: gingival cells (GCs), bone marrow-derived mesenchymal stromal cells (Bm-MSCs), and periodontal ligament cells (PDLCs). The construct featured a 3-dimensional fibrous bone compartment with macroscopic pore size, while the periodontal compartment consisted of a flexible porous membrane for cell sheet delivery. The regenerative performance of the constructs was radiographically and histologically assessed in surgically created periodontal defects in sheep following 5 and 10 wk of healing. Histologic observation demonstrated that the constructs maintained their shape and volume throughout the entirety of the in vivo study and were well integrated with the surrounding tissue. There was also excellent tissue integration between the bone and periodontal ligament compartments as well as the tooth root interface, enabling the attachment of periodontal ligament fibers into newly formed cementum and bone. Bone coverage along the root surface increased between weeks 5 and 10 in the Bm-MSC and PDLC groups. At week 10, the micro-computed tomography results showed that the PDLC group had greater bone fill as compared with the empty scaffold, while the GC group had less bone than the 3 other groups (control, Bm-MSC, and PDLC). Periodontal regeneration, as measured by histologically verified new bone and cementum formation with obliquely inserted periodontal ligament fibers, increased between 5 and 10 wk for the empty, Bm-MSC, and PDLC groups, while the GC group was inferior to the Bm-MSC and PDLC groups at 10 wk. This study demonstrates that periodontal regeneration can be achieved via the utilization of a multiphasic construct, with Bm-MSCs and PDLCs obtaining superior results as compared with GC-derived cell sheets.

SpheroidSim-Preliminary evaluation of a new computational tool to predict the influence of cell cycle time and phase fraction on spheroid growth.

BACKGROUND: There is a relative paucity of research that integrates materials science and bioengineering with computational simulations to decipher the intricate processes promoting cancer progression. Therefore, a first-generation computational model, SpheroidSim, was developed that includes a biological data set derived from a bioengineered spheroid model to obtain a quantitative description of cell kinetics. RESULTS: SpheroidSim is a 3D agent-based model simulating the growth of multicellular cancer spheroids. Cell cycle time and phases mathematically motivated the population growth. SpheroidSim simulated the growth dynamics of multiple spheroids by individually defining a collection of specific phenotypic traits and characteristics for each cell. Experimental data derived from a hydrogel-based spheroid model were fit to the predictions providing insight into the influence of cell cycle time (CCT) and cell phase fraction (CPF) on the cell population. A comparison of the number of active cells predicted for each analysis showed that the value and method used to define CCT had a greater effect on the predicted cell population than CPF. The model predictions were similar to the experimental results for the number of cells, with the predicted total number of cells varying by 8% and 12%, respectively, compared to the experimental data. CONCLUSIONS: SpheroidSim is a first step in developing a biologically based predictive tool capable of revealing fundamental elements in cancer cell physiology. This computational model may be applied to study the effect of the microenvironment on spheroid growth and other cancer cell types that demonstrate a similar multicellular clustering behavior as the population develops. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:1335-1343, 2018.

A humanised tissue-engineered bone model allows species-specific breast cancer-related bone metastasis in vivo.

Bone metastases frequently occur in the advanced stages of breast cancer. At this stage, the disease is deemed incurable. To date, the mechanisms of breast cancer-related metastasis to bone are poorly understood. This may be attributed to the lack of appropriate animal models to investigate the complex cancer cell-bone interactions. In this study, two established tissue-engineered bone constructs (TEBCs) were applied to a breast cancer-related metastasis model. A cylindrical medical-grade polycaprolactone-tricalcium phosphate scaffold produced by fused deposition modelling (scaffold 1) was compared with a tubular calcium phosphate-coated polycaprolactone scaffold fabricated by solution electrospinning (scaffold 2) for their potential to generate ectopic humanised bone in NOD/SCID mice. While scaffold 1 was found not suitable to generate a sufficient amount of ectopic bone tissue due to poor ectopic integration, scaffold 2 showed excellent integration into the host tissue, leading to bone formation. To mimic breast cancer cell colonisation to the bone, MDA-MB-231, SUM1315, and MDA-MB-231BO breast cancer cells were cultured in polyethylene glycol-based hydrogels and implanted adjacent to the TEBCs. Histological analysis indicated that the breast cancer cells induced an osteoclastic reaction in the TEBCs, demonstrating analogies to breast cancer-related bone metastasis seen in patients.

Vitrification as a prospect for cryopreservation of tissue-engineered constructs.

Cryopreservation plays a significant function in tissue banking and will presume yet larger value when more and more tissue-engineered products will routinely enter the clinical arena. The most common concept underlying tissue engineering is to combine a scaffold (cellular solids) or matrix (hydrogels) with living cells to form a tissue-engineered construct (TEC) to promote the repair and regeneration of tissues. The scaffold and matrix are expected to support cell colonization, migration, growth and differentiation, and to guide the development of the required tissue. The promises of tissue engineering, however, depend on the ability to physically distribute the products to patients in need. For this reason, the ability to cryogenically preserve not only cells, but also TECs, and one day even whole laboratory-produced organs, may be indispensable. Cryopreservation can be achieved by conventional freezing and vitrification (ice-free cryopreservation). In this publication we try to define the needs versus the desires of vitrifying TECs, with particular emphasis on the cryoprotectant properties, suitable materials and morphology. It is concluded that the formation of ice, through both direct and indirect effects, is probably fundamental to these difficulties, and this is why vitrification seems to be the most promising modality of cryopreservation.

Comparison of human alveolar osteoblasts cultured on polymer-ceramic composite scaffolds and tissue culture plates.

The effects of medical grade polycaprolactone-tricalcium phosphate (mPCL-TCP) (80:20) scaffolds on primary human alveolar osteoblasts (AOs) were compared with standard tissue-culture plates. Of the seeded AOs, 70% adhered to and proliferated on the scaffold surface and within open and interconnected pores; they formed multi-layered sheets and collagen fibers with uniform distribution within 28 days. Elevation of alkaline phosphatase activity occurred in scaffold-cell constructs independent of osteogenic induction. AO proliferation rate increased and significant decrease in calcium concentration of the medium for both scaffolds and plates under induction conditions were seen. mPCL-TCP scaffolds significantly influenced the AO expression pattern of osterix and osteocalcin (OCN). Osteogenic induction down-regulated OCN at both RNA and protein level on scaffolds (3D) by day 7, and up-regulated OCN in cell-culture plates (2D) by day 14, but OCN levels on scaffolds were higher than on cell-culture plates. Immunocytochemical signals for type I collagen, osteopontin and osteocalcin were detected at the outer parts of scaffold-cell constructs. More mineral nodules were found in induced than in non-induced constructs. Only induced 2D cultures showed nodule formation. mPCL-TCP scaffolds appear to stimulate osteogenesis in vitro by activating a cellular response in AO's to form mineralized tissue. There is a fundamental difference between culturing AOs on 2D and 3D environments that should be considered when studying osteogenesis in vitro.

Scaffold-cell bone engineering in a validated preclinical animal model: precursors vs differentiated cell source.

The properties of osteoblasts (OBs) isolated from the axial skeleton (tOBs) differ from OBs of the orofacial skeleton (mOBs) due to the different embryological origins of the bones. The aim of the study was to assess and compare the regenerative potential of allogenic bone marrow-derived mesenchymal progenitor cells with allogenic tOBs and allogenic mOBs in combination with a mPCL-TCP scaffold in critical-sized segmental bone defects in sheep tibiae. After 6 months, the tibiae were explanted and underwent biomechanical testing, micro-computed tomography (microCT) and histological and immunohistochemical analyses. Allogenic MPCs demonstrated a trend towards a better outcome in biomechanical testing and the mean values of newly formed bone. Biomechanical, microCT and histological analysis showed no significant differences in the bone regeneration potential of tOBs and mOBs in our in vitro study, as well as in the bone regeneration potential of different cell types in vivo. Copyright © 2015 John Wiley & Sons, Ltd.