Bone Formation In Scaffolds Research Articles

Dynamics of actinotrichia, fibrous collagen structures in zebrafish fin tissues, unveiled by novel fluorescent probes

Abstract Collagen fibers provide physical support to animal tissues by orienting in the correct position and at optimal density. Actinotrichia are thick collagen fibers that are present at the tips of fish fins and serve as scaffolds for bone formation. The arrangement and density of actinotrichia must be constantly maintained with a high degree of regularity to form spatial patterns in the fin bones, but the mechanisms of this process are largely unknown. To address this issue, we first identified two fluorescent probes that can stain actinotrichia clearly in vivo. Using these probes and time-lapse observation of actinotrichia synthesized at different growth stages, we revealed the following previously unknown dynamics of actinotrichia. (i) Actinotrichia do not stay stationary at the place where they are produced; instead, they move towards the dorsal area during the notochord bending and (ii) move towards the distal tip during the fin growth. (iii) Actinotrichia elongate asymmetrically as new collagen is added at the proximal side. (iv) Density is maintained by the insertion of new actinotrichia. (v) Actinotrichia are selectively degraded by osteoclasts. These findings suggest that the regular arrangement of actinotrichia is the outcome of multiple dynamic processes.

Effect of Mechanical Loading on Bone Regeneration in HA/β-TCP/SF Scaffolds Prepared by Low-Temperature 3D Printing In Vivo.

It has been well demonstrated that a dynamic culture environment improves tissue-engineered bone formation in vitro, but little is known about how cyclical mechanical loading induced bone formation in scaffolds in situ. To mimic the organic and inorganic components and multilevel structure of a bony microenvironment, hydroxyapatite/β tricalcium phosphate/silk fibroin(HA/β-TCP/SF) composite scaffolds with macro- and micropores were fabricated in this study. The mechanical properties and structure of the scaffolds were adjusted based on the ratio of organic and inorganic components and three-dimensional (3D) printing parameters. Dynamic sinusoidal loading with different frequencies was applied to the composite scaffold. Mouse bone precursor cells MC3T3-E1 were seeded on the scaffolds, and the cell compatibility of the scaffolds was investigated by MTT, SEM, and HE. The effect of the loading on bone formation in the scaffold in situ was investigated in a rabbit tibia defect model. The scaffold showed viscoelasticity and hysteresis under dynamic sinusoidal loading with different frequencies. With an increase in HA/β-TCP, the stress and modulus of the scaffolds increased. MTT, SEM, and HE results showed that MC3T3-E1 cells could adhere and proliferate on the composite scaffolds. After loading in vivo, the quantity of newly formed bone and the bone volume fraction increased. Micro-CT, undecalcified Van Gieson (VG) staining, and fluorescent double-labeling results suggested that appropriate cyclical mechanical loading at frequencies of 1 and 10 Hz had positive effects on bone formation in situ and it may play a role in clinical bone defect repair.

Orsellinic acid-loaded chitosan nanoparticles in gelatin/nanohydroxyapatite scaffolds for bone formation in vitro

AimOrsellinic acid (2,4-Dimethoxy-6-methylbenzoic acid) (OA) is a hydrophobic polyphenolic compound with therapeutic potential, but its impact on actuating osteogenesis remains unknown. The bioavailability of OA is hampered by its hydrophobic nature. This study aimed to fabricate nano-drug delivery system-based scaffolds for OA and test its potential for osteogenesis in vitro. Materials and methodsOA was loaded into chitosan nanoparticles (nCS + OA) using the ionic gelation technique at different concentrations. nCS + OA were incorporated onto the scaffolds containing gelatin (Gel) and nanohydroxyapatite (nHAp) by the lyophilization method. Biocomposite scaffolds were examined for their physicochemical and material characteristic properties. The effect of OA in the scaffolds for osteoblast differentiation was determined by alizarin red and von Kossa staining at the cellular level and by reverse transcriptase-qPCR and western blot analysis at the molecular level. Key findingsThe scaffolds showed excellent physiochemical and material characteristics and remained cyto-friendly to mouse mesenchymal stem cells (mMSCs, C3H10T1/2). The release of OA from Gel/nHAp/nCS scaffolds enhanced the differentiation of mMSCs towards osteoblasts, as observed through cellular and molecular studies. Moreover, the osteogenic potential of OA was mediated by the activation of FAK and ERK signaling pathways through integrins. SignificanceThe inclusion of OA into Gel/nHAp/nCS biocomposite scaffolds at 80 μM concentration promoted osteoblast differentiation via cell adhesion mediated signaling, compared with that shown by Gel/nHAp/nCS alone. Overall, this study identified the potential therapeutic OA containing Gel/nHAp/nCS scaffolds, accelerating its potential for clinical application towards bone regeneration.

Three-dimensional Printing of Biomimetic Titanium Mimicking Trabecular Bone Induces Human Mesenchymal Stem Cell Proliferation: An In-vitro Analysis.

In vitro analysis. The aim of this study was to assess the effect of three-dimensional (3D) printing of porous titanium on human mesenchymal stem cell (hMSC) adhesion, proliferation, and osteogenic differentiation. A proprietary implant using three-dimensional porous titanium (3D-pTi) that mimics trabecu-lar bone structure, roughness, porosity, and modulus of elasticity was created (Ti-LIFE technology™, Spineart SA Switzerland). Such implants may possess osteoinductive properties augmenting fusion in addition to their structural advantages. However, the ability of 3D-pTi to affect in vitro cellular proliferation and osteogenic differentiation remains undefined. Disks of 3D-pTi with a porosity of 70% to 75% and pore size of 0.9 mm were produced using additive manufacturing technology. 2D Ti6Al4V (2D-Ti) and 2D polyetheretherketone (2D-PEEK) disks were prepared using standard manufacturing process. Tissue culture plastic (TCP) served as the control surface. All discs were characterized using 2D-micros-copy, scanning electron microscopy (SEM), and x-ray micro-computed tomography. Forty thousand hMSCs were seeded on the disks and TCP and cultured for 42 days. hMSC morphology was assessed using environmental SEM and confocal imaging following phalloidin staining. hMSC proliferation was evaluated using DNA fluorescent assay. hMSC differentiation was assessed using RT-qPCR for genes involved in hMSC osteogenic differentiation and biochemical assays were performed for alkaline phosphatase activity (ALP) and calcium content. 3D-pTi lead to a higher cell number as compared to 2D-Ti and 2D-PEEK at D21, D28 and D42. ALP activity of hMSCs seeded into 3D-pTi scaffolds was as high as or higher than that of hMSCs seeded onto TCP controls over all time points and consistently higher than that of hMSCs seeded onto 2D-Ti scaffolds. However, when ALP activity was normalized to protein content, no statistical differences were found between all scaffolds tested and TCP controls. 3D-pTi provides a scaffold for bone formation that structurally mimics cancellous bone and improves hMSC adhesion and proliferation compared to 2D-Ti and PEEK.

Macrophage-Mediated Bone Formation in Scaffolds Modified With MSC-Derived Extracellular Matrix Is Dependent on the Migration Inhibitory Factor Signaling Pathway.

The positive role of macrophages in the osteogenesis of mesenchymal stem cells (MSCs) has been a recent research focus. On the other hand, MSCs could carefully regulate the paracrine molecules derived from macrophages. Human umbilical cord mesenchymal stem cells (hucMSCs) can reduce the secretion of inflammatory factors from macrophages to improve injury healing. hucMSC-derived extracellular matrix (hucMSC-ECM) has the similar effect to hucMSCs, which could combat the inflammatory response of macrophages. Additionally, MSC-derived extracellular matrix also enhanced bone regeneration by inhibiting osteoclastic differentiation of monocyte/macrophage lineage. However, whether hucMSC-ECM could improve bone formation by guiding macrophage-induced osteogenic differentiation of MSCs is unknown. Here, we present decalcified bone scaffolds modified by hucMSC-derived extracellular matrix (DBM-ECM), which maintained multiple soluble cytokines from hucMSCs, including macrophage migration inhibitory factor (MIF). Compared with DBM, the DBM-ECM scaffolds induced bone formation in an improved heterotopic ossification model of severe combined immunodeficiency (SCID) mice in a macrophage-dependent manner. Macrophages cocultured with DBM-ECM expressed four osteoinductive cytokines (BMP2, FGF2, TGFβ3 and OSM), which were screened out by RNA sequencing and measured by qPCR and western blot. The conditioned medium from macrophages cocultured with DBM-ECM improved the osteogenic differentiation of hBMSCs. Furthermore, DBM-ECM activated CD74/CD44 (the typical MIF receptors) signal transduction in macrophages, including phosphorylation of P38 and dephosphorylation of c-jun. On the other side, the inhibitory effects of the DBM-ECM scaffolds with a deficient of MIF on osteogenesis in vitro and in vivo revealed that macrophage-mediated osteogenesis depended on MIF/CD74 signal transduction. The results of this study indicate that the coordinated crosstalk of macrophages and MSCs plays a key role on bone regeneration, with an emphasis on hucMSC-ECM constructing a macrophage-derived osteoinductive microenvironment.

A machine learning-based multiscale model to predict bone formation in scaffolds.

Computational modeling methods combined with non-invasive imaging technologies have exhibited great potential and unique opportunities to model new bone formation in scaffold tissue engineering, offering an effective alternate and viable complement to laborious and time-consuming in vivo studies. However, existing numerical approaches are still highly demanding computationally in such multiscale problems. To tackle this challenge, we propose a machine learning (ML)-based approach to predict bone ingrowth outcomes in bulk tissue scaffolds. The proposed in silico procedure is developed by correlating with a dedicated longitudinal (12-month) animal study on scaffold treatment of a major segmental defect in sheep tibia. Comparison of the ML-based time-dependent prediction of bone ingrowth with the conventional multilevel finite element (FE2) model demonstrates satisfactory accuracy and efficiency. The ML-based modeling approach provides an effective means for predicting in vivo bone tissue regeneration in a subject-specific scaffolding system.

Time-lapsed imaging of nanocomposite scaffolds reveals increased bone formation in dynamic compression bioreactors

Progress in bone scaffold development relies on cost-intensive and hardly scalable animal studies. In contrast to in vivo, in vitro studies are often conducted in the absence of dynamic compression. Here, we present an in vitro dynamic compression bioreactor approach to monitor bone formation in scaffolds under cyclic loading. A biopolymer was processed into mechanically competent bone scaffolds that incorporate a high-volume content of ultrasonically treated hydroxyapatite or a mixture with barium titanate nanoparticles. After seeding with human bone marrow stromal cells, time-lapsed imaging of scaffolds in bioreactors revealed increased bone formation in hydroxyapatite scaffolds under cyclic loading. This stimulatory effect was even more pronounced in scaffolds containing a mixture of barium titanate and hydroxyapatite and corroborated by immunohistological staining. Therefore, by combining mechanical loading and time-lapsed imaging, this in vitro bioreactor strategy may potentially accelerate development of engineered bone scaffolds and reduce the use of animals for experimentation.

Therapeutic Potential of Orsellinic Acid-Loaded Chitosan Nanoparticles in Gelatin/Nanohydroxyapatite Scaffolds for Bone Formation

Therapeutic Potential of Orsellinic Acid-Loaded Chitosan Nanoparticles in Gelatin/Nanohydroxyapatite Scaffolds for Bone Formation

Effects of concurrent use of royal jelly with hydroxyapatite on bone healing in rabbit model: radiological and histopathological evaluation

BackgroundBone grafts have been used to enhance bone fracture healing in orthopedic surgery. Bone grafts enhance bone healing either by mechanical support or acting as a scaffold for bone formation. Fresh autograft is the most effective biomaterial because it is histocompatible with less complication about transmissible disease. Hydroxyapatite is a well-established material for bone repair and very comparable to natural apatite providing a strong biomechanical interlock with host tissue. Royal jelly is the principal food for the honeybee queen. This biomaterial has been demonstrated to have several pharmacological activities, such as antiallergic, antitumor and anti-inflammatory effects. ObjectivesThis study was design to evaluate the effect of concurrent using of Royal jelly with hydroxyapatite on bone healing in rabbit model. Methods15 adult rabbits weighting approximately 2 kg had been used. They were divided into three groups randomly. In first group (N = 5) mid radius bone defect created and left empty. The second group (N = 5) filled with hydroxy apatite alone and the last group (N = 5) filled with royal jelly and hydroxy apatite combination. Radiological evaluation performed on days14th, 28th and 42nd after operation. Histopathological evaluation was done on 56th postoperative day. ResultsRadiological evaluation showed significant superior bone healing in hydroxyapatite and hydroxyapatite-Royal jelly groups in comparison to control group. Control group was the inferior group between three groups. There were not any significant differences between three groups in histopathological group. ConclusionIn conclusion our study showed the best results with using the hydroxyl apatite and Royal-jelly group because they provide not only scaffold for bone healing but also do, they provide some osteoinduction materials for bone healing.

The Cocktail Effect of BMP-2 and TGF-β1 Loaded in Visible Light-Cured Glycol Chitosan Hydrogels for the Enhancement of Bone Formation in a Rat Tibial Defect Model.

Bone tissue engineering scaffolds offer the merits of minimal invasion as well as localized and controlled biomolecule release to targeted sites. In this study, we prepared injectable hydrogel systems based on visible light-cured glycol chitosan (GC) hydrogels containing bone morphogenetic protein-2 (BMP-2) and/or transforming growth factor-beta1 (TGF-β1) as scaffolds for bone formation in vitro and in vivo. The hydrogels were characterized by storage modulus, scanning electron microscopy (SEM) and swelling ratio analyses. The developed hydrogel systems showed controlled releases of growth factors in a sustained manner for 30 days. In vitro and in vivo studies revealed that growth factor-loaded GC hydrogels have no cytotoxicity against MC3T3-E1 osteoblast cell line, improved mRNA expressions of alkaline phosphatase (ALP), type I collagen (COL 1) and osteocalcin (OCN), and increased bone volume (BV) and bone mineral density (BMD) in tibia defect sites. Moreover, GC hydrogel containing BMP-2 (10 ng) and TGF-β1 (10 ng) (GC/BMP-2/TGF-β1-10 ng) showed greater bone formation abilities than that containing BMP-2 (5 ng) and TGF-β1 (5 ng) (GC/BMP-2/TGF-β1-5 ng) in vitro and in vivo. Consequently, the injectable GC/BMP-2/TGF-β1-10 ng hydrogel may have clinical potential for dental or orthopedic applications.

Effect of Alendronate with β - TCP Bone Substitute in Surgical Therapy of Periodontal Intra-Osseous Defects: A Randomized Controlled Clinical Trial.

Alendronate (ALN), an aminobisphosphonate, inhibits osteoclastic bone resorption and also stimulates osteogenesis. Beta-Tricalcium Phosphate (β-TCP) is an osteoconductive graft material which provides a scaffold for bone formation and also a widely used drug delivery vehicle for growth factors and antibiotics. Drug delivery vehicles, like β-TCP, improve the potency of the drugs by specific local site delivery of the drug, optimal release characteristics and easy handling. The aim of the this study was to evaluate the bone formation potential of 400μg ALN delivered in β-TCP in the treatment of periodontal intra-osseous defects. Thirty patients with periodontal defects were randomly assigned to 400μg ALN + β-TCP + Saline (test) group and β-TCP + Saline (active-control) group. Clinical parameters like Clinical Attachment Level (CAL) gain, Probing Depth (PD) reduction, post-operative Gingival Recession (GR) were assessed from the baseline, 3 months and 6 months recordings. Radiographic parameters like Linear Bone Growth (LBG), Percentage Bone Fill (%BF), and change in alveolar crest height (ACH) were assessed from baseline and 6 months radiographs. Mean measurements in the ALN test group for CAL gain (3.4 ± 0.74 mm), PD reduction (4.33 ± 0.82 mm), LBG (2.88 ± 0.88 mm), and %BF (51.98 ± 15.84%) were significantly greater with a p-value <0.05 compared to the mean measurements of CAL gain (2.20 ± 0.86 mm), PD reduction (3.20 ± 1.15 mm), LBG (1.70 ± 0.39 mm), and %BF (30.35 ± 6.88%) of the control group. There was mild alveolar crestal apposition (0.32 ± 0.68 mm) in the ALN test group and mild alveolar crestal resorption (-0.24 ± 0.40 mm) in the control group. 400μg ALN combined with β-TCP bone graft material was effective in improving soft tissue parameters, inhibiting alveolar crestal resorption and enhancing bone formation, compared to β-TCP alone.

Bilayered scaffolds colonized with dental pulp stem cells for osteochondral tissue engineering application

The development of osteochondral tissue engineered interfaces would be a novel treatment for traumatic injuries and aging associated diseases that affect joints [1]. Dental Pulp Stem Cells (DPSC) are a promising alternative source in cell-based tissue engineering for the easily recruitment, the self-renewal and differentiation potential, being able to differentiate toward chondrogenic and osteogenic lineage [2]. The aim of this study was to develop scaffold that would support an osteochon- dral interface. DPSCs, isolated from dental pulp by selection with c-Kit-CD34 and STRO1 antibodies, were either cultured on fibroin and collagen scaffolds and differ- entiated into osteogenic cells or encapsulated in alginate and differentiated into chondrogenic cells. After 3 weeks of culture, the differentiation of the cell-scaffold con- structs was analyzed by histology and immunofluorescence. The cell seeded scaffolds were then brought together (fibroin+alginate and collagen+alginate) in order to form a combination that would provide a supportive scaffold for bone formation as well as a soft matrix for cartilage growth. The resulting single constructs were cultured for 2 weeks in a osteochondral media, developed by combining some of the growth factors and nutrients of osteoand chondrogenic media. The constructs were then characterized for expression of bone and cartilage specific markers to evaluate the capability of the cells to not dedifferentiate in the new cultured conditions. We demonstrated that both collagen and fibroin appear to be excellent scaffold for cells differentiated into osteogenic lineage, although they have different structural characteristics. Similarly, the alginate has allowed cells to produce ECM-rich in collagen II and aggrecan during chondrogenic differentiation. Moreover DPSC in the bilay- ered constructs remain viable and differentiated even in the osteochondral medium. Thus the integrated fibroin-alginate or collagen-alginate proved to be functional in vitro and may lead to the development of new strategies for osteochondral repair and regeneration.

Enhanced bone regeneration in rat calvarial defects implanted with surface-modified and BMP-loaded bioactive glass (13-93) scaffolds

The repair of large bone defects, such as segmental defects in the long bones of the limbs, is a challenging clinical problem. Our recent work has shown the ability to create porous scaffolds of silicate 13-93 bioactive glass by robocasting which have compressive strengths comparable to human cortical bone. The objective of this study was to evaluate the capacity of those strong porous scaffolds with a grid-like microstructure (porosity=50%; filament width=330μm; pore width=300μm) to regenerate bone in a rat calvarial defect model. Six weeks post-implantation, the amount of new bone formed within the implants was evaluated using histomorphometric analysis. The amount of new bone formed in implants composed of the as-fabricated scaffolds was 32% of the available pore space (area). Pretreating the as-fabricated scaffolds in an aqueous phosphate solution for 1, 3 and 6days to convert a surface layer to hydroxyapatite prior to implantation enhanced new bone formation to 46%, 57% and 45%, respectively. New bone formation in scaffolds pretreated for 1, 3 and 6days and loaded with bone morphogenetic protein-2 (BMP-2) (1μg per defect) was 65%, 61% and 64%, respectively. The results show that converting a surface layer of the glass to hydroxyapatite or loading the surface-treated scaffolds with BMP-2 can significantly improve the capacity of 13-93 bioactive glass scaffolds to regenerate bone in an osseous defect. Based on their mechanical properties evaluated previously and their capacity to regenerate bone found in this study, these 13-93 bioactive glass scaffolds, pretreated or loaded with BMP-2, are promising in structural bone repair.

Evaluation of a Thiolated Chitosan Scaffold for Local Delivery of BMP-2 for Osteogenic Differentiation and Ectopic Bone Formation

Thiolated chitosan (Thio-CS) is a well-established pharmaceutical excipient for drug delivery. However, its use as a scaffold for bone formation has not been investigated. The aim of this study was to evaluate the potential of Thio-CS in bone morphogenetic protein-2 (BMP-2) delivery and bone formation. In vitro study showed that BMP-2 interacted with the Thio-CS and did not affect the swelling behavior. The release kinetics of BMP-2 from the Thio-CS was slightly delayed (70%) within 7 days compared with that from collagen gel (Col-gel, 85%), which is widely used in BMP-2 delivery. The BMP-2 released from Thio-CS increased osteoblastic cell differentiation but did not show any cytotoxicity until 21 days. Analysis of the in vivo ectopic bone formation at 4 weeks of posttransplantation showed that use of Thio-CS for BMP-2 delivery induced more bone formation to a greater extent (1.8 fold) than that of Col-gel. However, bone mineral density in both bones was equivalent, regardless of Thio-CS or Col-gel carrier. Taken together, Thio-CS system might be useful for delivering osteogenic protein BMP-2 and present a promising bone regeneration strategy.

Bone scaffold architecture modulates the development of mineralized bone matrix by human embryonic stem cells

Decellularized bone has been widely used as a scaffold for bone formation, due to its similarity to the native bone matrix and excellent osteoinductive and biomechanical properties. We have previously shown that human mesenchymal and embryonic stem cells form functional bone matrix on such scaffolds, without the use of growth factors. In this study, we focused on differences in bone matrix that exist even among identical harvesting sites, and the effects of the matrix architecture and mineral content on bone formation by human embryonic stem cells (hESC). Mesenchymal progenitors derived from hESCs were cultured for 5 weeks in decellularized bone scaffolds with three different densities: low (0.281 ± 0.018 mg/mm3), medium (0.434 ± 0.015 mg/mm3) and high (0.618 ± 0.027 mg/mm3). The medium-density group yielded highest densities of cells and newly assembled bone matrix, presumably due to the best balance between the transport of nutrients and metabolites to and from the cells, space for cell infiltration, surface for cell attachment and the mechanical strength of the scaffolds, all of which depend on the scaffold density. Bone mineral was beneficial for the higher expression of bone markers in cultured cells and more robust accumulation of the new bone matrix.

Bone formation using β-tricalcium phosphate/carboxymethyl-chitin composite scaffold in rat calvarial defects

The aim of the present study was to evaluate the effects of β-tricalcium phosphate/carboxymethyl-chitin material (β-TCP/CM-chitin) on bone formation in rat calvarial defects. Eighteen animals surgically received 2 calvarial defects (5 mm) bilaterally in each parietal bone. β-TCP/CM-chitin was implanted in one side of each defect, and the contralateral side of the defect was left empty as a control. The animals were humanely killed at 4, 8, and 12 weeks after surgery for histologic evaluation. New bone formation in the β-TCP/CM-chitin group was significantly greater than that in the control group throughout the healing periods (P < .05). β-TCP/CM-chitin was remarkably resorbed 12 weeks after surgery. These results indicate that β-TCP/CM-chitin is useful as a scaffold for bone formation.

Critical illness-related bone loss is associated with osteoclastic and angiogenic abnormalities

Critically ill patients are at increased risk of fractures during rehabilitation, and can experience impaired healing of traumatic and surgical bone fractures. In addition, markers of bone resorption are markedly increased in critically ill patients, while markers of bone formation are decreased. In the current study, we have directly investigated the effect of critical illness on bone metabolism and repair. In a human in vitro model of critical illness, Fluorescence-activated cell sorting (FACS) analysis revealed an increase in circulating CD14+/CD11b+ osteoclast precursors in critically ill patient peripheral blood compared to healthy controls. In addition, the formation of osteoclasts was increased in patient peripheral blood mononuclear cell (PBMC) cultures compared to healthy controls, both in the presence and absence of osteoclastogenic factors receptor activator of NF-κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF). Culturing PBMCs with 10% critically ill patient serum further increased osteoclast formation and activity in patient PBMCs only, and neutralization studies revealed that immunoglobulin G (IgG) antibody signaling through the immunoreceptor Fc receptor common γ chain III (FcRγIII) played an important role. When analyzing bone formation, no differences in osteogenic differentiation were observed using human periosteal-derived cells (hPDCs) treated with patient serum in vitro, but a decrease in the expression of vascular endothelial growth factor receptor 1 (VEGF-R1) suggested impaired vascularization. This was confirmed using serum-treated hPDCs implanted onto calcium phosphate scaffolds in a murine in vivo model of bone formation, where decreased vascularization and increased osteoclast activity led to a decrease in bone formation in scaffolds with patient serum-treated hPDCs. Together, these findings may help to define novel therapeutic targets to prevent bone loss and optimize fracture healing in critically ill patients.

Prediction of spatio-temporal bone formation in scaffold by diffusion equation

Developing a successful bone tissue engineering strategy entails translation of experimental findings to clinical needs. A major leap forward toward this goal is developing a quantitative tool to predict spatial and temporal bone formation in scaffold. We hypothesized that bone formation in scaffold follows diffusion phenomenon. Subsequently, we developed an analytical formulation for bone formation, which had only three unknown parameters: C, the final bone volume fraction, α , the so-called scaffold osteoconduction coefficient, and h, the so-called peri-scaffold osteoinduction coefficient. The three parameters were estimated by identifying the model with in vivo data of polymeric scaffolds implanted in the femoral condyle of rats. In vivo data were obtained by longitudinal micro-CT scanning of the animals. Having identified the three parameters, we used the model to predict the course of bone formation in two previously published in vivo studies. We found the predicted values to be consistent with the experimental ones. Bone formation into a scaffold can then adequately be described through diffusion phenomenon. This model allowed us to spatially and temporally predict the outcome of tissue engineering scaffolds with only 3 physically relevant parameters.

The synergetic bone-forming effects of combinations of growth factors expressed by adenovirus vectors on chitosan/collagen scaffolds

Supporting and enhancing the regeneration and stability of alveolar bone at dental implant sites is necessary to improve implant integration and stability. In this study chitosan/collagen scaffolds combined with adenoviruses expressing either bone morphogenetic protein 7 (BMP-7), platelet-derived growth factor B (PDGF-B), or a combination of both BMP-7 and PDGF-B, were prepared by a freeze–drying method. The cytotoxicity of the resulting biomaterials, and their potential effects on cell proliferation and differentiation towards an osteoblast phenotype, were studied in an initial in vitro study using human periodontal ligament cells (HPLCs). These in vitro results indicate that the scaffold materials are not cytotoxic, and moreover that scaffolds expressing PDGF-B show a higher proliferation rate, while scaffolds expressing BMP-7 show a stronger differentiation towards the osteoblast phenotype. In a subsequent in vivo study, scaffolds were implanted into surgically induced defects on both sides of the dog mandible. Bone formation, which was evaluated histomorphometrically at 4 and 8 weeks and after sacrificing the animals at 12 weeks, revealed a significant increase in bone formation in scaffolds expressing only BMP-7 and also the combined growth factors, with the highest increase occurring for the scaffold expressing both BMP-7 and PDGF-B. This study demonstrates the promising potential of biomaterial expression of combinations of growth factors such as BMP-7 and PDGF-B for bone regeneration in tissue engineering applications.

In vivo performance of selective electron beam‐melted Ti‐6Al‐4V structures

Highly porous titanium structures are widely used for maxillofacial and orthopedic surgery because of their excellent mechanical properties similar to those of human bone and their facilitation of bone ingrowth. In contrast to common methods, the generation of porous titaniumproducts by selective electron beam melting (SEBM), an additive manufacturing technology, overcomes difficulties concerning the extreme chemical affinity of liquid titanium to atmospheric gases which consequently leads to strongly reduced ductility of the metal. The purpose of this study was to assess the suitability of a smooth compact and a porous Ti-6Al-4V structure directly produced by the SEBM process as scaffolds for bone formation. SEBM-processed titanium implants were placed into defects in the frontal skull of 15 domestic pigs. To evaluate the direct contact between bone and implant surfaces and to assess the ingrowth of osseous tissue into the porous structure, microradiographs and histomorphometric analyses were performed 14, 30, and 60 days after surgery. Bone ingrowth increased significantly during the period of this study. After 14 days the most outer regions of the implants were already filled with newly formed bone tissue (around 14%). After 30 days the bone volume inside the implants reached almost 30% and after 60 days abundant bone formation inside the implants attained 46%. During the study only scarce bone-implant contact was found around all implants, which did not exceed 9% around compact specimens and 6% around porous specimens after 60 days. This work demonstrates that highly porous titanium implants with excellent interconnectivity manufactured using the SEBM method are suitable scaffolds for bone ingrowth. This technique is a good candidate for orthopedic and maxillofacial applications.