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Type-H capillary endothelial cells control bone formation during embryogenesis and postnatal growth but few signalling mechanisms underpinning this influence have been characterised. Here, we identify a highly expressed type-H endothelial cell protein, Clec14a, and explore its role in coordinating osteoblast activity. Expression of Clec14a and its ligand, Mmrn2 are high in murine type-H endothelial cells but absent from osteoblasts. Clec14a -/- mice have premature condensation of the type-H vasculature and expanded distribution of osteoblasts and bone matrix, increased long-bone length and bone density indicative of accelerated skeletal development, and enhanced osteoblast maturation. Antibody-mediated blockade of the Clec14a-Mmrn2 interaction recapitulates the Clec14a -/- phenotype. Endothelial cell expression of Clec14a regulates osteoblast maturation and mineralisation activity during postnatal bone development in mice. This finding underscores the importance of type-H capillary control of osteoblast activity in bone formation and identifies a novel mechanism that mediates this vital cellular crosstalk., (© 2024. The Author(s).)

Introduction: Peri-implantitis is an inflammatory process around dental implants that is characterised by bone loss that may jeopardize the long-term survival of osseo integrated dental implants. The aim of this study was to create a surface coating on titanium abutments that possesses cellular adhesion and anti-microbial properties as a post-implant placement strategy for patients at risk of peri-implantitis., Materials and Methodsmethods: Titanium alloy Grade V stubs were coated with gold particles and then subjected to ceramic conversion treatment (CCT) at 620 °C for 3, 8 and 80 h. The surface characteristics and chemistry were assessed using scanning electron microscopy (SEM), energy dispersive spectroscopy (EDS), and X-ray diffraction (XRD) analysis. The leaching profile was investigated by inductively coupled plasma mass spectroscopy (ICP-MS) for all groups after 7, 14 and 28 days in contact with distilled water. A scratch test was conducted to assess the adhesion of the gold coating to the underlying titanium discs. Two bacterial species (Staphylococcus aureus (SA) & Fusobacterium nucleatum (FN)) were used to assess the antibacterial behaviour of the coated discs using a direct attachment assay test. The potential changes in surface chemistry by the bacterial species were investigated by grazing angle XRD., Results: The gold pre-coated titanium discs exhibited good stability of the coating especially after immersion in distilled water and after bacterial colonisation as evident by XRD analysis. Good surface adhesion of the coating was demonstrated for gold treated discs after scratch test analysis, especially titanium, following a 3-hour (3 H) ceramic conversion treatment. All coated discs exhibited significantly improved antimicrobial properties against both tested bacterial species compared to untreated titanium discs., Conclusions: Ceramic conversion treated titanium with a pre-deposited gold layer showed improved antimicrobial properties against both SA and FN species than untreated Ti-C discs. Scratch test analysis showed good adherence properties of the coated discs the oxide layer formed is firmly adherent to the underlying titanium substrate, suggesting that this approach may have clinical efficacy for coating implant abutments., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Advances in bioprinting have enabled the fabrication of complex tissue constructs with high speed and resolution. However, there remains significant structural and biological complexity within tissues that bioprinting is unable to recapitulate. Bone, for example, has a hierarchical organization ranging from the molecular to whole organ level. Current bioprinting techniques and the materials employed have imposed limits on the scale, speed, and resolution that can be achieved, rendering the technique unable to reproduce the structural hierarchies and cell-matrix interactions that are observed in bone. The shift toward biomimetic approaches in bone tissue engineering, where hydrogels provide biophysical and biochemical cues to encapsulated cells, is a promising approach to enhancing the biological function and development of tissues for in vitro modeling. A major focus in bioprinting of bone tissue for in vitro modeling is creating dynamic microenvironmental niches to support, stimulate, and direct the cellular processes for bone formation and remodeling. Hydrogels are ideal materials for imitating the extracellular matrix since they can be engineered to present various cues whilst allowing bioprinting. Here, recent advances in hydrogels and 3D bioprinting toward creating a microenvironmental niche that is conducive to tissue engineering of in vitro models of bone are reviewed., (© 2023 The Authors. Advanced Materials published by Wiley-VCH GmbH.)

Microfluidics have transformed diagnosis and screening in regenerative medicine. Recently, they are showing much promise in biofabrication. However, their adoption is inhibited by costly and drawn-out lithographic processes thus limiting progress. Here, multi-material fibers with complex core-shell geometries with sizes matching those of human arteries and arterioles are fabricated employing versatile microfluidic devices produced using an agile and inexpensive manufacturing pipeline. The pipeline consists of material extrusion additive manufacturing with an innovative continuously varied extrusion (CONVEX) approach to produce microfluidics with complex seamless geometries including, novel variable-width zigzag (V-zigzag) mixers with channel widths ranging from 100-400 µm and hydrodynamic flow-focusing components. The microfluidic systems facilitated rapid mixing of fluids by decelerating the fluids at specific zones to allow for increased diffusion across the interfaces. Better mixing even at high flow rates (100-1000 µL min -1 ) whilst avoiding turbulence led to high cell cytocompatibility (>86%) even when 100 µm nozzles are used. The presented 3D-printed microfluidic system is versatile, simple and efficient, offering a great potential to significantly advance the microfluidic platform in regenerative medicine., (© 2023 The Authors. Advanced Healthcare Materials published by Wiley-VCH GmbH.)

Chronic wounds are a drain on global health services and remain a major area of unmet clinical need. Chronic wounds are characterised by a stable and stubborn bacterial biofilm which hinders innate immune response and delays or prevents wound healing. Bioactive glass (BG) fibres offer a promising novel treatment for chronic wounds by targeting the wound-associated biofilm. In this study, the antimicrobial properties of silver-doped BG fibres were tested against Pseudomonas aeruginosa biofilms, which are commonly found in chronic wound infections. Results showed that BG fibres doped with silver resulted in a 5log10 reduction in biofilm formation whereas silver-free fibres only reduced formation by log10, therefore silver-doped fibres possess stronger antimicrobial effects. Moreover, there appeared to be a synergistic effect between the fibres and the silver as the application of the silver-doped fibres placed directly in contact with the forming biofilm resulted in a higher reduction in biofilm formation compared to treatments either: using the dissolution ions, using BG powder, or when the fibres were placed in an insert above the biofilm, inhibiting physical contact, instead. This suggests that the physical properties of the fibres, as well as silver, influence biofilm formation. Finally, results demonstrated that silver chloride, which is not antimicrobial, forms and the concentrations of antimicrobial silver species, namely silver ions and nanoparticles, reduce over time when fibres are soaked in cell culture media, which partially explains why the silver-doped dissolution ions contained lower antimicrobial activity compared to the fibres. As silver chloride is more likely to form with increased temperature and time, the antimicrobial activity of silver-containing dissolution ions is highly dependent on the length of ageing and storage conditions. Many studies investigate the antimicrobial and cytotoxic properties of biomaterials through their dissolution products. However, instability of antimicrobial silver species due to silver chloride formation and its effect on antimicrobial properties of silver-based biomaterials has not been reported before and could influence past and future dissolution-based assays as results showed that the antimicrobial activity of silver-based dissolution ions can vary greatly depending on post processing steps and can therefore produce misleading data., Competing Interests: The authors declare the following financial interests/personal relationships which may be considered as potential competing interests:Sandeep Shirgill reports financial support was provided by 10.13039/501100000268Biotechnology and Biological Sciences Research Council. Gowsihan Poologasundarampillai reports financial support was provided by 10.13039/501100000266Engineering and Physical Sciences Research Council., (© 2023 The Authors.)

A lack of models that recapitulate the complexity of human bone marrow has hampered mechanistic studies of normal and malignant hematopoiesis and the validation of novel therapies. Here, we describe a step-wise, directed-differentiation protocol in which organoids are generated from induced pluripotent stem cells committed to mesenchymal, endothelial, and hematopoietic lineages. These 3D structures capture key features of human bone marrow-stroma, lumen-forming sinusoids, and myeloid cells including proplatelet-forming megakaryocytes. The organoids supported the engraftment and survival of cells from patients with blood malignancies, including cancer types notoriously difficult to maintain ex vivo. Fibrosis of the organoid occurred following TGFβ stimulation and engraftment with myelofibrosis but not healthy donor-derived cells, validating this platform as a powerful tool for studies of malignant cells and their interactions within a human bone marrow-like milieu. This enabling technology is likely to accelerate the discovery and prioritization of novel targets for bone marrow disorders and blood cancers., Significance: We present a human bone marrow organoid that supports the growth of primary cells from patients with myeloid and lymphoid blood cancers. This model allows for mechanistic studies of blood cancers in the context of their microenvironment and provides a much-needed ex vivo tool for the prioritization of new therapeutics. See related commentary by Derecka and Crispino, p. 263. This article is highlighted in the In This Issue feature, p. 247., (©2022 The Authors; Published by the American Association for Cancer Research.)

Optical trapping enables the real-time manipulation and observation of morphological evolution of individual particles during reaction chemistry. Here, optical trapping was used in combination with Raman spectroscopy to conduct airborne assembly and kinetic experiments. Micro-droplets of alkoxysilane were levitated in air prior to undergoing either acid- or base-catalyzed sol-gel reaction chemistry to form silica particles. The evolution of the reaction was monitored in real-time; Raman and Mie spectroscopies confirmed the in situ formation of silica particles from alkoxysilane droplets as the product of successive hydrolysis and condensation reactions, with faster reaction kinetics in acid catalysis. Hydrolysis and condensation were accompanied by a reduction in droplet volume and silica formation. Two airborne particles undergoing solidification could be assembled into unique 3D structures such as dumb-bell shapes by manipulating a controlled collision. Our results provide a pipeline combining spectroscopy with optical microscopy and nanoscale FIB-SEM imaging to enable chemical and structural insights, with the opportunity to apply this methodology to probe structure formation during reactive inkjet printing., Competing Interests: The authors declare no competing financial interest., (© 2022 The Authors. Published by American Chemical Society.)

Skin has excellent capacity to regenerate, however, in the event of a large injury or burn skin grafts are required to aid wound healing. The regenerative capacity further declines with increasing age and can be further exacerbated with bacterial infection leading to a chronic wound. Engineered skin substitutes can be used to provide a temporary template for the damaged tissue, to prevent/combat bacterial infection and promote healing. In this study, the sol-gel process and electrospinning were combined to fabricate 3D cotton-wool-like sol-gel bioactive glass fibers that mimic the fibrous architecture of skin extracellular matrix (ECM) and deliver metal ions for antibacterial (silver) and therapeutic (calcium and silica species) actions for successful healing of wounds. This study investigated the effects of synthesis and process parameters, in particular sintering temperature on the fiber morphology, the incorporation and distribution of silver and the degradation rate of fibers. Silver nitrate was found to decompose into silver nanoparticles within the glass fibers upon calcination. Furthermore, with increasing calcination temperature the nanoparticles increased in size from 3 nm at 600 °C to ~25 nm at 800 °C. The antibacterial ability of the Ag-doped glass fibers decreased as a function of the glass calcination temperature. The degradation products from the Ag-doped 3D non-woven sol-gel glass fibers were also found to promote fibroblast proliferation thus demonstrating their potential for use in skin regeneration., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2021 Elsevier B.V. All rights reserved.)

We propose a novel image analysis framework to automate analysis of X-ray microtomography images of sintering ceramics and glasses, using open-source toolkits and machine learning. Additive manufacturing (AM) of glasses and ceramics usually requires sintering of green bodies. Sintering causes shrinkage, which presents a challenge for controlling the metrology of the final architecture. Therefore, being able to monitor sintering in 3D over time (termed 4D) is important when developing new porous ceramics or glasses. Synchrotron X-ray tomographic imaging allows in situ, real-time capture of the sintering process at both micro and macro scales using a furnace rig, facilitating 4D quantitative analysis of the process. The proposed image analysis framework is capable of tracking and quantifying the densification of glass or ceramic particles within multiple volumes of interest (VOIs) along with structural changes over time using 4D image data. The framework is demonstrated by 4D quantitative analysis of bioactive glass ICIE16 within a 3D-printed scaffold. Here, densification of glass particles within 3 VOIs were tracked and quantified along with diameter change of struts and interstrut pore size over the 3D image series, delivering new insights on the sintering mechanism of ICIE16 bioactive glass particles in both micro and macro scales., (© 2021 The Authors. Journal of the American Ceramic Society published by Wiley Periodicals LLC on behalf of American Ceramic Society.)

Hydrogels constructed from naturally derived polymers provide an aqueous environment that encourages cell growth, however, mechanical properties are poor and degradation can be difficult to predict. Whilst, synthetic hydrogels exhibit some improved mechanical properties, these materials lack biochemical cues for cells growing and have limited biodegradation. To produce hydrogels that support 3D cell cultures to form tissue mimics, materials must exhibit appropriate biological and mechanical properties. In this study, novel organic-inorganic hybrid hydrogels based on chitosan and silica were prepared using the sol-gel technique. The chemical, physical and biological properties of the hydrogels were assessed. Statistical analysis was performed using One-Way ANOVAs and independent-sample t -tests. Fourier transform infrared spectroscopy showed characteristic absorption bands including amide II, Si-O and Si-O-Si confirming formation of hybrid networks. Oscillatory rheometry was used to characterise the sol to gel transition and viscoelastic behaviour of hydrogels. Furthermore, in vitro degradation revealed both chitosan and silica were released over 21 days. The hydrogels exhibited high loading efficiency as total protein loading was released in a week. There were significant differences between TC 2 G and C 2 G at all-time points ( p < 0.05). The viability of osteoblasts seeded on, and encapsulated within, the hydrogels was >70% over 168 h culture and antimicrobial activity was demonstrated against Pseudomonas aeruginosa and Enterococcus faecalis . The hydrogels developed here offer alternatives for biopolymer hydrogels for biomedical use, including for application in drug/cell delivery and for bone tissue engineering.

An electrospinning technique was used to produce three-dimensional (3D) bioactive glass fibrous scaffolds, in the SiO 2 -CaO sol-gel system, for wound healing applications. Previously, it was thought that 3D cotton wool-like structures could only be produced from sol-gel when the sol contained calcium nitrate, implying that the Ca 2+ and its electronic charge had a significant effect on the structure produced. Here, fibres with a 3D appearance were also electrospun from compositions containing only silica. A polymer binding agent was added to inorganic sol-gel solutions, enabling electrospinning prior to bioactive glass network formation and the polymer was removed by calcination. While the addition of Ca 2+ contributes to the 3D morphology, here we show that other factors, such as relative humidity, play an important role in producing the 3D cotton-wool-like macrostructure of the fibres. A human dermal fibroblast cell line (CD-18CO) was exposed to dissolution products of the samples. Cell proliferation and metabolic activity tests were carried out and a VEGF ELISA showed a significant increase in VEGF production in cells exposed to the bioactive glass samples compared to control in DMEM. A novel SiO 2 -CaO nanofibrous scaffold was created that showed tailorable physical and dissolution properties, the control and composition of these release products are important for directing desirable wound healing interactions.

Purpose: To develop a biomimetic tumor tissue phantom which more closely reflects water diffusion in biological tissue than previously used phantoms, and to evaluate the stability of the phantom and its potential as a tool for validating diffusion-weighted (DW) MRI measurements., Methods: Coaxial-electrospraying was used to generate micron-sized hollow polymer spheres, which mimic cells. The bulk structure was immersed in water, providing a DW-MRI phantom whose apparent diffusion coefficient (ADC) and microstructural properties were evaluated over a period of 10 months. Independent characterization of the phantom's microstructure was performed using scanning electron microscopy (SEM). The repeatability of the construction process was investigated by generating a second phantom, which underwent high resolution synchrotron-CT as well as SEM and MR scans., Results: ADC values were stable (coefficients of variation (CoVs) < 5%), and varied with diffusion time, with average values of 1.44 ± 0.03 µm 2 /ms (Δ = 12 ms) and 1.20 ± 0.05 µm 2 /ms (Δ = 45 ms). Microstructural parameters showed greater variability (CoVs up to 13%), with evidence of bias in sphere size estimates. Similar trends were observed in the second phantom., Conclusion: A novel biomimetic phantom has been developed and shown to be stable over 10 months. It is envisaged that such phantoms will be used for further investigation of microstructural models relevant to characterizing tumor tissue, and may also find application in evaluating acquisition protocols and comparing DW-MRI-derived biomarkers obtained from different scanners at different sites. Magn Reson Med 80:147-158, 2018. © 2017 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited., (© 2017 The Authors Magnetic Resonance in Medicine published by Wiley Periodicals, Inc. on behalf of International Society for Magnetic Resonance in Medicine.)

Antimicrobial silver nanoparticle coatings have attracted interest for reducing prosthetic joint infection. However, few studies report in vivo investigations of the biotransformation of silver nanoparticles within the regenerating tissue and its impact on bone formation. We present a longitudinal investigation of the osseointegration of silver nanoparticle-coated additive manufactured titanium implants in rat tibial defects. Correlative imaging at different time points using nanoscale secondary ion mass spectrometry, transmission electron microscopy (TEM), histomorphometry, and 3D X-ray microcomputed tomography provided quantitative insight from the nano- to macroscales. The quality and quantity of newly formed bone is comparable between the uncoated and silver coated implants. The newly formed bone demonstrates a trabecular morphology with bone being located at the implant surface, and at a distance, at two weeks. Nanoscale elemental mapping of the bone-implant interface showed that silver was present primarily in the osseous tissue and colocalized with sulfur. TEM revealed silver sulfide nanoparticles in the newly regenerated bone, presenting strong evidence that the previously in vitro observed biotransformation of silver to silver sulfide occurs in vivo.

Hierarchically porous biocompatible Mg-Al-Cl-type layered double hydroxide (LDH) composites containing aluminum hydroxide (Alhy) have been prepared using a phase-separation process. The sol-gel synthesis allows for the hierarchical pores of the LDH-Alhy composites to be tuned, leading to a high specific solid surface area per unit volume available for high-molecular-weight protein adsorptions. A linear relationship between the effective surface area, SEFF, and loading capacity of a model protein, bovine serum albumin (BSA), is established following successful control of the structure of the LDH-Alhy composite. The threshold of the mean pore diameter, Dpm, above which BSA is effectively adsorbed on the surface of LDH-Alhy composites, is deduced as 20 nm. In particular, LDH-Alhy composite aerogels obtained via supercritical drying exhibit an extremely high capacity for protein loading (996 mg/g) as a result of a large mean mesopore diameter (>30 nm). The protein loading on LDH-Alhy is >14 times that of a reference LDH material (70 mg/g) prepared via a standard procedure. Importantly, BSA molecules pre-adsorbed on porous composites were successfully released on soaking in ionic solutions (HPO4(2-) and Cl(-) aqueous). The superior capability of the biocompatible LDH materials for loading, encapsulation, and releasing large quantities of proteins was clearly demonstrated.

A correlative imaging methodology was developed to accurately quantify bone formation in the complex lattice structure of additive manufactured implants. Micro computed tomography (μCT) and histomorphometry were combined, integrating the best features from both, while demonstrating the limitations of each imaging modality. This semi-automatic methodology registered each modality using a coarse graining technique to speed the registration of 2D histology sections to high resolution 3D μCT datasets. Once registered, histomorphometric qualitative and quantitative bone descriptors were directly correlated to 3D quantitative bone descriptors, such as bone ingrowth and bone contact. The correlative imaging allowed the significant volumetric shrinkage of histology sections to be quantified for the first time (~15 %). This technique demonstrated the importance of location of the histological section, demonstrating that up to a 30 % offset can be introduced. The results were used to quantitatively demonstrate the effectiveness of 3D printed titanium lattice implants.

Current materials used for bone regeneration are usually bioactive ceramics or glasses. Although they bond to bone, they are brittle. There is a need for new materials that can combine bioactivity with toughness and controlled biodegradation. Sol-gel hybrids have the potential to do this through their nanoscale interpenetrating networks (IPN) of inorganic and organic components. Poly(γ-glutamic acid) (γ-PGA) was introduced into the sol-gel process to produce a hybrid of γ-PGA and bioactive silica. Calcium is an important element for bone regeneration but calcium sources that are used traditionally in the sol-gel process, such as Ca salts, do not allow Ca incorporation into the silicate network during low-temperature processing. The hypothesis for this study was that using calcium methoxyethoxide (CME) as the Ca source would allow Ca incorporation into the silicate component of the hybrid at room temperature. The produced hybrids would have improved mechanical properties and controlled degradation compared with hybrids of calcium chloride (CaCl2 ), in which the Ca is not incorporated into the silicate network. Class II hybrids, with covalent bonds between the inorganic and organic species, were synthesised by using organosilane. Calcium incorporation in both the organic and inorganic IPNs of the hybrid was improved when CME was used. This was clearly observed by using FTIR and solid-state NMR spectroscopy, which showed ionic cross-linking of γ-PGA by Ca and a lower degree of condensation of the Si species compared with the hybrids made with CaCl2 as the Ca source. The ionic cross-linking of γ-PGA by Ca resulted in excellent compressive strength and reduced elastic modulus as measured by compressive testing and nanoindentation, respectively. All hybrids showed bioactivity as hydroxyapatite (HA) was formed after immersion in simulated body fluid (SBF)., (© 2014 The Authors. Published by Wiley-VCH Verlag GmbH & Co. KGaA. This is an open access article under the terms of the Creative Commons Attribution License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited.)

Understanding the distribution of critical elements (e.g. silicon and calcium) within silica-based bone scaffolds synthesized by different methods is central to the optimization of these materials. Time-of-flight secondary ion mass spectrometry (ToF-SIMS) has been used to determine this information due to its very high surface sensitivity and its ability to map all the elements and compounds in the periodic table with high spatial resolution. The SIMS image data can also be combined with depth profiles to construct three-dimensional chemical maps. However, the scaffolds have interconnected pore networks, which are very challenging structures for the SIMS technique. To overcome this problem two experimental methodologies have been developed. The first method involved the use of the focused ion beam technique to obtain clear images of the regions of interest and subsequently mark them by introducing fiducial marks; the samples were then analysed using the ToF-SIMS technique to yield the chemical analyses of the regions of interest. The second method involved impregnating the pores using a suitable reagent so that a flat surface could be achieved, and this was followed by secondary ion mapping and 3D chemical imaging with ToF-SIMS. The samples used in this work were sol-gel 70S30C foam and electrospun fibres and calcium-containing silica/gelatin hybrid scaffolds. The results demonstrate the feasibility of both these experimental methodologies and indicate that these methods can provide an opportunity to compare various artificial bone scaffolds, which will be of help in improving scaffold synthesis and processing routes. The techniques are also transferable to many other types of porous material.

Vaterite particles containing aminopropyl-functionalized silsesquioxane (SiV) were prepared as osteogenic devices for bone regeneration. The Si x V particles (x = 0, 2.6 and 4.9 wt%) were synthesized by reacting a slurry of calcium hydroxide with carbon dioxide gas in the presence of γ-aminopropyltriethoxysilane (APTES), a source of soluble silica which would genetically enhance osteogenesis. The obtained Si 2.6 V and Si 4.9 V particles were monodispersed with a diameter of 1.4 and 1.5 μm, respectively. The Si 2.6 V particles showed spherical morphologies. On the surface of the Si 4.9 V particle small particles were aggregated, resulting in the formation of irregular textures. Transmission electron microscopy of a sectioned Si 2.6 V particle revealed that the vaterite particles were present as lamellae with a length of 5-20 nm and surrounded by silsesquioxane from APTES. Moreover, the vaterite lamellae were relatively orientated to the c face of the unit lattice, where it is known to be highly polarized, compared to pure vaterite, due to the exposure of the uni-ionic plane with positive (Ca 2+ ) or negative (CO 3 2- ) charge. Attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) revealed the co-existence of amorphous calcium carbonate (ACC) in the SiV particles. On contact with physiological pH buffer solution, the vaterite was transiently stabilized and subsequently dissolved and released after the dissolution of silsesquioxane from the particles. This stabilization time was significantly increased with the increase in silicon content. The vaterite was observed in Si 2.6 V particles up to 3 h of soaking, which extended up to 12 h in Si 4.9 V particles. The formation of the particles from the precursor gel was monitored by laser Raman spectroscopy and ATR-FTIR. During the initial 1 to 2 h of the aging step, maturation of ACC into vaterite and condensation of monomeric APTES molecules were found to begin simultaneously. These reactions proceeded up to 7 h of the analysis period. The condensation of hydrolyzed APTES is suggested to occur in the vicinity of growing vaterite, which might play a role in the enclosure of vaterite in silsesquioxanes.

Bioactive glass has high potential for bone regeneration due to its ability to bond to bone and stimulate osteogenesis whilst dissolving in the body. Although three-dimensional (3-D) bioactive glass scaffolds with favorable pore networks can be made from the sol-gel process, compositional and structural evolutions in their porous structures during degradation in vivo, or in vitro, have not been quantified. In this study, bioactive glass scaffolds were put in a simulated body fluid flow environment through a perfusion bioreactor. X-ray microtomography (μCT) was used to non-destructively image the scaffolds at different degradation stages. A new 3-D image processing methodology was developed to quantify the scaffold's pore size, interconnect size and connectivity from μCT images. The accurate measurement of individual interconnect size was made possible by a principal component analysis-based algorithm. During 28 days of dissolution, the modal interconnect size in the scaffold was reduced from 254 to 206 μm due to the deposition of mineral phases. However, the pore size remained unchanged, with a mode of 682 μm. The data presented are important for making bioactive glass scaffolds into clinical products. The technique described for imaging and quantifying scaffold pore structures as a function of degradation time is applicable to most scaffold systems., (Copyright © 2011 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.)

X-ray microtomography (microCT) is a popular tool for imaging scaffolds designed for tissue engineering applications. The ability of synchrotron microCT to monitor tissue response and changes in a bioactive glass scaffold ex vivo were assessed. It was possible to observe the morphology of the bone; soft tissue ingrowth and the calcium distribution within the scaffold. A second aim was to use two newly developed compression rigs, one designed for use inside a laboratory based microCT machine for continual monitoring of the pore structure and crack formation and another designed for use in the synchrotron facility. Both rigs allowed imaging of the failure mechanism while obtaining stress-strain data. Failure mechanisms of the bioactive glass scaffolds were found not to follow classical predictions for the failure of brittle foams. Compression strengths were found to be 4.5-6 MPa while maintaining an interconnected pore network suitable for tissue engineering applications.

The need to shift from tissue replacement to tissue regeneration has led to the development of tissue engineering and in situ tissue regeneration. Both of these strategies often employ the use of scaffolds--templates that allow cells to attach and then guide the new tissue growth. There are many design criteria for an ideal scaffold. These criteria vary depending on the tissue type and location in the body. In any application of a scaffold it is vital to be able to characterise the scaffold before it goes into in vitro testing. In vitro testing allows the cell response to be investigated before its in vivo performance is assessed. A full characterisation of events in vitro and in vivo, in three dimensions (3D), is necessary if a scaffold's performance and effectiveness is to be fully quantified. This paper focuses on porous scaffolds for bone regeneration, suggests appropriate design criteria for a bone regenerating scaffold and then reviews techniques for obtaining the vitally important quantification of its pore structure. The techniques discussed will include newly developed methods of quantifying X-ray microtomography (microCT) images in 3D and for predicting the scaffolds mechanical properties and the likely paths of fluid flow (and hence potential cell migration). The complications in investigating scaffold performance in vitro are then discussed. Finally, the use of microCT for imaging scaffolds for in vivo tests is reviewed.

In tissue engineering, porous scaffolds are often used as three-dimensional (3D) supports for tissue growth. In scaffold design, it is imperative to be able to quantify the pore sizes and more importantly the interconnects between the pores. X-ray micro-computed tomography (microCT) has become a popular tool for obtaining 3D images of scaffold biomaterials, however images are only qualitative. In this work, methods were developed for obtaining pore size distributions for both the macropores and their interconnects. Scaffolds have been developed, by foaming sol-gel derived bioactive glasses, which have the potential to fulfil the criteria for an ideal scaffold for bone tissue engineering. MicroCT images were obtained from scaffolds with different pore structures. The images were thresholded and three algorithms were applied in 3D to identify pores and interconnects and to obtain pore size distributions. The results were validated against mercury intrusion porosimetry and manual 3D image analysis. The microCT data were then meshed such that predictions of permeability as a function of changes in the pore network could be made. Such predictions will be useful for optimising bioreactor conditions for tissue engineering applications. These techniques would be suitable for many other types of scaffolds.