Identification of bone morphogenetic proteins 2 and 4 in commercial demineralized freeze-dried bone allograft preparations : pilot study

BACKGROUND: Demineralized freeze-dried bone allografts (DFDBAs) have been proposed as a useful adjunct in periodontal therapy to induce periodontal regeneration through the induction of new bone formation. The presence of bone morphogenetic proteins (BMPs) within the demineralized matrix has been proposed as a possible mechanism through which DFDBA may exert its biologic effect. However, in recent years, the predictability of results using DFDBA has been variable and has led to its use being questioned. One reason for the variability in tissue response may be attributed to differences in the processing of DFDBA, which may lead to loss of activity of any bioactive substances within the DFDBA matrix. Therefore, the purpose of this investigation was to determine whether there are detectable levels of bone morphogenetic proteins in commercial DFDBA preparations. METHODS: A single preparation of DFDBA was obtained from three commercial sources. Each preparation was studied in triplicate. Proteins within the DFDBA samples were first extracted with 4M guanidinium HCI for seven days at 40 degrees celsius and the residue was further extracted with 4M guanidinium HCL/EDTA for seven days at 40 degrees celsius. Two anti-human BMP-2 and -4 antibodies were used for the detection of the presence of BMP's in the extracts. RESULTS: Neither BMP-2 nor BMP-4 was detected in any of the extracts. When recombinant human BMP-2 and -4 were added throughout the extraction process of DFDBA extraction, not only were intact proteins detected but smaller molecular weight fragments were also noted in the extract. CONCLUSIONS: These results indicate that all of the DFDBA samples tested had no detectable amounts of BMP-2 and -4. In addition, an unknown substance present in the DFDBA may be responsible for degradation of whatever BMPs might be present.

Alternative modes of entrepreneurial opportunity exploitation : the case on inter-organizational project

While entrepreneurship research has taken firm formation to be the predominant mode of opportunity exploitation, entrepreneurship can take place through many other types of organizational arrangements. In the present article, we consider one such alternative arrangement, namely the formation of inter-organizational projects (IOPs). We propose a multi-level contingency model that suggests that uncertainty both at the level of the firm and at the level of the environment makes the exploitation of opportunities through IOPs more likely. The model is tested by telephone survey data collected amongst a panel of 1725 SMEs and longitudinal industry data. Our findings provide strong support for the industry-level part of the model, but interestingly, only partial support for the firm level part of the model. This indicates that the effects of uncertainty need to be dissected into different levels of analysis to understand the conditions under which alternative modes of opportunity exploitation can be a prominent entrepreneurial alternative to new firm formation.

Effects of scaffold architecture on cranial bone healing

Scaffolds for bone tissue engineering should be designed to optimize cell migration, enhance new bone formation and give mechanical support. In the present study, we used polycaprolactone-tricalciumphosphate (PCL/TCP) scaffolds with two different fibre lay down patterns which were coated with hydroxyapatite and gelatine as an approach for optimizing bone regeneration in a critical sized calvarial defect. After 12 weeks bone regeneration was quantified using microCT analysis, biomechanical testing and histological evaluation. Notably, the experimental groups containing coated scaffolds showed lower bone formation and lower biomechanical properties within the defect compared to the uncoated scaffolds. Surprisingly, the different lay down pattern of the fibres resulted in different bone formation and biomechanical properties; namely 0/60/120° scaffolds revealed lower bone formation and biomechanical properties compared to the 0/90° scaffolds in all the experimental groups. The different architecture of the scaffold fibres may have an effect on nutrition supply as well as the attachment of the newly formed matrix to the scaffold. Therefore, future bone regeneration strategies utilising scaffolds should consider scaffold architecture as an important factor during the scaffold optimisation stages in order to move closer to a clinical application.

Turnover in entrepreneurial teams : are spousal teams different?

Entrepreneurial teams are increasingly prevalent. An important stage in new venture formation involves the recruitment and retention of team members, characterised by changes in the membership of the ET. ET turnover involves members joining or leaving the team, which could have positive or negative effects on the team and the firm. Spousal Entrepreneurial Teams (SET) are a frequently occurring, yet unique type of ET, with very distinct characteristics and dynamics. In an SET turnover is likely to be have different meanings and effects than in Other Entrepreneurial Teams. This research asks: What influences members joining and leaving entrepreneurial teams? And are SET different? Using data from 561 young firms over X years, we find significant differences between SET and NSET. Specifically, we find X.

The relationship between airborne small ions and particles in urban environments

Ions play an important role in affecting climate and particle formation in the atmosphere. Small ions rapidly attach to particles in the air and, therefore, studies have shown that they are suppressed in polluted environments. Urban environments, in particular, are dominated by motor vehicle emissions and, since motor vehicles are a source of both particles and small ions, the relationship between these two parameters is not well known. In order to gain a better understanding of this relationship, an intensive campaign was undertaken where particles and small ions of both signs were monitored over two week periods at each of three sites A, B and C that were affected to varying degrees by vehicle emissions. Site A was close to a major road and reported the highest particle number and lowest small ion concentrations. Precursors from motor vehicle emissions gave rise to clear particle formation events on five days and, on each day this was accompanied by a suppression of small ions. Observations at Site B, which was located within the urban airshed, though not adjacent to motor traffic, showed particle enhancement but no formation events. Site C was a clean site, away from urban sources. This site reported the lowest particle number and highest small ion concentration. The positive small ion concentration was 10% to 40% higher than the corresponding negative value at all sites. These results confirm previous findings that there is a clear inverse relationship between small ions and particles in urban environments dominated by motor vehicle emissions.

Alternative modes of entrepreneurial opportunity exploitation

While entrepreneurship research has taken firm formation to be the predominant mode of opportunity exploitation, entrepreneurship can take place through many other types of organizational arrangements. In the present article, we consider one such alternative arrangement, namely the formation of inter-organizational projects (IOPs). We propose a multi-level contingency model that suggests that uncertainty both at the level of the firm and at the level of the environment makes the exploitation of opportunities through IOPs more likely. The model is tested by telephone survey data collected amongst a panel of 1725 SMEs and longitudinal industry data. Our findings provide strong support for the industry-level part of the model, but interestingly, only partial support for the firm level part of the model. This indicates that the effects of uncertainty need to be dissected into different levels of analysis to understand the conditions under which alternative modes of opportunity exploitation can be a prominent entrepreneurial alternative to new firm formation.

The influence of cellular source on periodontal regeneration using calcium phosphate coated polycaprolactone scaffold supported cell sheets

Cell-based therapy is considered a promising approach to achieving predictable periodontal regeneration. In this study, the regenerative potential of cell sheets derived from different parts of the periodontium (gingival connective tissue, alveolar bone and periodontal ligament) were investigated in an athymic rat periodontal defect model. Periodontal ligament (PDLC), alveolar bone (ABC) and gingival margin-derived cells (GMC) were obtained from human donors. The osteogenic potential of the primary cultures was demonstrated in vitro. Cell sheets supported by a calcium phosphate coated melt electrospun polycaprolactone (CaP-PCL) scaffold were transplanted to denuded root surfaces in surgically created periodontal defects, and allowed to heal for 1 and 4 weeks. The CaP-PCL scaffold alone was able to promote alveolar bone formation within the defect after 4 weeks. The addition of ABC and PDLC sheets resulted in significant periodontal attachment formation. The GMC sheets did not promote periodontal regeneration on the root surface and inhibited bone formation within the CaP-PCL scaffold. In conclusion, the combination of either PDLC or ABC sheets with a CaP-PCL scaffold could promote periodontal regeneration, but ABC sheets were not as effective as PDLC sheets in promoting new attachment formation.

Controlling whole blood activation and resultant clot properties on various material surfaces : a possible therapeutic approach for enhancing bone healing

Injured bone initiates the healing process by forming a blood clot at the damaged site. However, in severe damage, synthetic bone implants are used to provide structural integrity and restore the healing process. The implant unavoidably comes into direct contact with whole blood, leading to a blood clot formation on its surface. Despite this, most research in bone tissue engineering virtually ignores the important role of a blood clot in supporting healing. Surface chemistry of a biomaterial is a crucial property in mediating blood-biomaterials interactions, and hence the formation of the resultant blood clot. Surfaces presenting mixtures of functional groups carboxyl (–COOH) and methyl (–CH3) have been shown to enhance platelet response and coagulation activation, leading to the formation of fibrin fibres. In addition, it has been shown that varying the compositions of these functional groups and the length of alkyl groups further modulate the immune complement response. In this study, we hypothesised that a biomaterial surface with mixture of –COOH/–CH3(methyl), –CH2CH3 (ethyl) or –(CH2)3CH3 (butyl) groups at different ratios would modulate blood coagulation and complement activation, and eventually tailor the structural and functional properties of the blood clot formed on the surface, which subsequently impacts new bone formation. Firstly, we synthesised a series of materials composed of acrylic acid (AA), and methyl (MMA), ethyl (EMA) or butyl methacrylates (BMA) at different ratios and coated on the inner surfaces of incubation vials. Our surface analysis showed that the amount of –COOH groups on the surface coatings was lower than the ratios of AA prepared in the materials even though the surface content of –COOH groups increased with increasing in AA ratios. It was indicated that the surface hydrophobicity increased with increasing alkyl chain length: –CH 3 > –CH2CH3 > –(CH2)3CH3, and decreased with increasing –COOH groups. No significant differences in surface hydrophobicity was found on surfaces with –CH3 and –CH2CH3 groups in the presence of –COOH groups. The material coating was as smooth as uncoated glass and without any major flaws. The average roughness of material-coated surface (3.99 ± 0.54 nm) was slightly higher than that of uncoated glass surface (2.22 ± 0.29 nm). However, no significant differences in surface average roughness was found among surfaces with the same functionalities at different –COOH ratios nor among surfaces with different alkyl groups but the same –COOH ratios. These suggested that the surface functional groups and their compositions had a combined effect on modulating surface hydrophobicity but not surface roughness. The second part of our study was to investigate the effect of surface functional groups and their compositions on blood cascade activation and structural properties of the formed clots. It was found that surfaces with –COOH/–(CH2)3CH3 induced a faster coagulation activation than those with –COOH/–CH3 and –CH2CH3, regardless of the –COOH ratios. An increase in –COOH ratios on –COOH/–CH3 and –CH2CH3 surfaces decreased the rate of activation. Moreover, all material-coated surfaces markedly reduced the complement activation compared to uncoated glass surfaces, and the pattern of complement activation was entirely similar to that of surface-induced coagulation, suggesting there is an interaction between two cascades. The clots formed on material-coated surfaces had thicker fibrin with a tighter network at the exterior when compared to uncoated glass surfaces. Compared to the clot exteriors, thicker fibrins with a loose network were found in clot interiors. Coated surfaces resulted in more rigid clots with a significantly slower fibrinolysis after 1 h of lysis when compared to uncoated glass surfaces. Significant differences in fibrinolysis after 1 h of lysis among clots on material-coated surfaces correlated well with the differences in fibrin thickness and density at clot exterior. In addition, more growth factors were released during clot formation than during clot lysis. From an intact clot, there was a correlation between the amount of PDGF-AB release and fibrin density. Highest amount of PDGF-AB was released from clots formed on surfaces with 40% –COOH/60% –CH 3 (i.e. 65MMA). During clot lysis, the release of PDGF-AB also correlated with the fibrinolytic rate while the release of TGF-â1 was influenced by the fibrin thickness. This suggested that different clot structures led to different release profiles of growth factors in clot intact and degrading stages. We further validated whether the clots formed on material-coatings provide the microenvironment for improved bone healing by using a rabbit femoral defect model. In this pilot study, the implantation of clots formed on 65MMA coatings significantly increased new bone formation with enhanced chondrogenesis, osteoblasts activity and vascularisation, but decreased inflammatory macrophage number at the defects after 4 weeks when compared to commercial bone grafts ChronOSTM â-TCP granules. Empty defects were observed when blood clot formation was inhibited. In summary, our study demonstrated that surface functional groups and their relative ratios on material coatings synergistically modulate activation of blood cascades, resultant fibrin architecture, rigidity, susceptibility to fibrinolysis as well as growth factor release of the formed clots, which ultimately alter the healing microenvironment of injured bones.

Osteogenic differentiation of bone marrow MSCs by β-tricalcium phosphate stimulating macrophages via BMP2 signalling pathway

Immune reactions play important roles in determining the in vivo fate of bone substitute materials, either in new bone formation or inflammatory fibrous tissue encapsulation. The paradigm for the development of bone substitute materials has been shifted from inert to immunomodulatory materials, emphasizing the importance of immune cells in the material evaluation. Macrophages, the major effector cells in the immune reaction to implants, are indispensable for osteogenesis and their heterogeneity and plasticity render macrophages a primer target for immune system modulation. However, there are very few reports about the effects of macrophages on biomaterial-regulated osteogenesis. In this study, we used b-tricalcium phosphate (b-TCP) as a model biomaterial to investigate the role of macrophages on the material stimulated osteogenesis. The macrophage phenotype switched to M2 extreme in response to b-TCP extracts, which was related to the activation of calcium-sensing receptor (CaSR) pathway. Bone morphogenetic protein 2 (BMP2) was also significantly upregulated by the b-TCP stimulation, indicating that macrophage may participate in the b-TCP stimulated osteogenesis. Interestingly, when macrophageconditioned b-TCP extracts were applied to bone marrow mesenchymal stem cells (BMSCs), the osteogenic differentiation of BMSCs was significantly enhanced, indicating the important role of macrophages in biomaterial-induced osteogenesis. These findings provided valuable insights into the mechanism of material-stimulated osteogenesis, and a strategy to optimize the evaluation system for the in vitro osteogenesis capacity of bone substitute materials.

A biomimetic extracellular matrix for cartilage tissue engineering centered on photocurable gelatin, hyaluronic acid and chondroitin sulfate

The development of hydrogels tailored for cartilage tissue engineering has been a research and clinical goal for over a decade. Directing cells towards a chondrogenic phenotype and promoting new matrix formation are significant challenges that must be overcome for the successful application of hydrogels in cartilage tissue therapies. Gelatin-methacrylamide (Gel-MA) hydrogels have shown promise for the repair of some tissues, but they have not been extensively investigated for cartilage tissue engineering. We encapsulated human chondrocytes in gel-MA based hydrogels, and show that with the incorporation of small quantities of photo-crosslinkable hyaluronic acid methacrylate (HA-MA), and to a lesser extent chondroitin sulfate methacrylate (CS-MA), chondrogenesis and mechanical properties can be enhanced. The addition of HA-MA to Gel-MA constructs resulted in more rounded cell morphologies, enhanced chondrogenesis as assessed by gene expression and immunofluorescence, and increased quantity and distribution of the newly synthesised ECM throughout the construct. Consequently, while the compressive moduli of control Gel-MA constructs increased by 26 kPa after 8 weeks culture, constructs with HA-MA and CS-MA increased by 96 kPa. The enhanced chondrogenic differentiation, distribution of ECM, and improved mechanical properties make these materials potential candidates for cartilage tissue engineering applications.

Angiogenesis as a biomarker and target in cancer chemoprevention

Angiogenesis, the formation of new blood vessels from existing vasculature, is essential to the late stages of carcinogenesis, allowing tumours to grow beyond 1-2 mm in diameter, invade surrounding tissue, and metastasise. However, more than two decades ago, angiogenesis that preceded neoplastic transformation was seen. Indeed, it can be detected in inflammatory and infectious diseases that increase the risk of developing cancer. Recent advances in fluorescence endoscopy and histological assessment suggest that, for certain cancers, the degree of new blood-vessel formation may differ between the early and late stages of carcinogenic progression. The association between angiogenesis and cancer occurrence, and ease of detection of this process in accessible tissues early in carcinogenesis, mean that angiogenesis fulfils the criteria for a biomarker of the effectiveness of chemopreventive intervention. There is also some evidence that biochemical assays of angiogenic growth factors may after similar potential as surrogate biomarkers. Many natural and synthetic chemopreventive agents in development or in clinical use inhibit new vessel formation in vivo. Validation of angiogenesis as a biomarker for the effectiveness of chemoprevention should further the advancement of some chemopreventive agents.

The role of biological extracellular matrix scaffolds in vascularized three-dimensional tissue growth in vivo

An in vivo murine vascularized chamber model has been shown to generate spontaneous angiogenesis and new tissue formation. This experiment aimed to assess the effects of common biological scaffolds on tissue growth in this model. Either laminin-1, type I collagen, fibrin glue, hyaluronan, or sea sponge was inserted into silicone chambers containing the epigastric artery and vein, one end was sealed with adipose tissue and the other with bone wax, then incubated subcutaneously. After 2, 4, or 6 weeks, tissue from chambers containing collagen I, fibrin glue, hyaluronan, or no added scaffold (control) had small amounts of vascularized connective tissue. Chambers containing sea sponge had moderate connective tissue growth together with a mild "foreign body" inflammatory response. Chambers containing laminin-1, at a concentration 10-fold lower than its concentration in Matrigel™, resulted in a moderate adipogenic response. In summary, (1) biological hydrogels are resorbed and gradually replaced by vascularized connective tissue; (2) sponge-like matrices with large pores support connective tissue growth within the pores and become encapsulated with granulation tissue; (3) laminin-containing scaffolds facilitate adipogenesis. It is concluded that the nature and chemical composition of the scaffold exerts a significant influence on the amount and type of tissue generated in this in vivo chamber model.

A comparative study of Sr-incorporated mesoporous bioactive glass scaffolds for regeneration of osteopenic bone defects

SUMMARY: Recently, the use of the pharmacological agent strontium ranelate has come to prominence for the treatment of osteoporosis. While much investigation is focused on preventing disease progression, here we fabricate strontium-containing scaffolds and show that they enhance bone defect healing in the femurs of rats induced by ovariectomy. INTRODUCTION: Recently, the use of the pharmacological agent strontium ranelate has come to prominence for the treatment of osteoporosis due to its ability to prevent bone loss in osteoporotic patients. Although much emphasis has been placed on using pharmacological agents for the prevention of disease, much less attention has been placed on the construction of biomaterials following osteoporotic-related fracture. The aim of the present study was to incorporate bioactive strontium (Sr) trace element into mesoporous bioactive glass (MBG) scaffolds and to investigate their in vivo efficacy for bone defect healing in the femurs of rats induced by ovariectomy. METHODS: In total, 30 animals were divided into five groups as follows: (1) empty defect (control), (2) empty defects with estrogen replacement therapy, (3) defects filled with MBG scaffolds alone, (4) defects filled with MBG + estrogen replacement therapy, and (5) defects filled with strontium-incorporated mesopore-bioglass (Sr-MBG) scaffolds. RESULTS: The two groups demonstrating the highest levels of new bone formation were the defects treated with MBG + estrogen replacement therapy and the defects receiving Sr-MBG scaffolds as assessed by μ-CT and histological analysis. Furthermore, Sr scaffolds had a reduced number of tartrate-resistant acid phosphatase-positive cells when compared to other modalities. CONCLUSION: The results from the present study demonstrate that the local release of Sr from bone scaffolds may improve fracture repair. Future large animal models are necessary to investigate the future relationship of Sr incorporation into biomaterials.

Structure and assembly of immature HIV

The major structural components of HIV are synthesized as a 55-kDa polyprotein, Gag. Particle formation is driven by the self-assembly of Gag into a curved hexameric lattice, the structure of which is poorly understood. We used cryoelectron tomography and contrast-transfer-function corrected subtomogram averaging to study the structure of the assembled immature Gag lattice to approximate to 17-angstrom resolution. Gag is arranged in the immature virus as a single, continuous, but incomplete hexameric lattice whose curvature is mediated without a requirement for pentameric defects. The resolution of the structure allows positioning of individual protein domains. High-resolution crystal structures were fitted into the reconstruction to locate protein-protein interfaces involved in Gag assembly, and to identify the structural transformations associated with virus maturation. The results of this study suggest a concept for the formation of nonsymmetrical enveloped viruses of variable sizes.

Lindquist and Pytel

The foremost event in the international architecture calendar is the Venice International Architecture Biennale. In 2012, Creative Directors Gerard Reinmuth and Anthony Burke with TOKO Concept Design, led the Australian Pavilion exhibition, entitled FORMATIONS: New Practices in Australian Architecture. The exhibition focus was to explore and celebrate “the nature of innovative configurations of architectural practice in Australia today and the desire for a renewed form of architectural agency which drives them”. The Australian Pavilion exhibition purposely chose to highlight the actions and processes behind contemporary architectural practice, focusing not on ‘starchitecture’ projects but those far reaching and socially-engaged “practitioners who are making a substantial and consequential impact in the field and well beyond it”. FORMATIONS had two overarching themes: (1) to stimulate critical disciplinary commentary on a range of new types of Australian practices and their potentialities and (2) exciting a public audience with a spatially dynamic and thought provoking exhibition of new forms of architectural practice, their spatial consequences and transformative potentials. Six projects were displayed in the Australian Pavilion in Venice, with the printed catalogue showcasing 33 ground-breaking examples of Australian practitioners addressing internationally relevant issues in their practice. Lindquist and Pytels collaborative practice is programmed between the demands of academia and commercial fashion practice. Their interests lie in exploring the relationship between the body, new materiality and its application within different facts of design production. The creative practice is underpinned by scholarly theory such as Heidegger’s "nearness and revealing" (1927-1954), Simondon’s "transduction theory" (1989) and the Burke's "sublime" (1757). Outcomes feedback into academic studio programs, scholarly research and material development for commercial, installation and speculative design production.