Thermoresponsive magnetic colloidal gels via surface-initiated polymerisation from functional microparticles.

Novel magnetothermally responsive core-shell microparticles have been synthesized. The aqueous suspensions of these particles exhibit fast thermoreversible fluid-to-gel transitions and retain good magnetic properties. Rheological measurements demonstrated that the viscoelasticity of the prepared particle gels can be tuned, enabling these gels to have the mechanical properties that should facilitate their applications as 3D cell scaffolds for in vitro expansion of cells. Also, it was found that the responsive particles could be used in repeated heating-cooling cycles without marked changes in gel elasticity. PrestoBlue viability assays of 3T3 fibroblasts and human mesenchymal stem cells cultured within the colloidal gel showed that the cells remained viable and proliferated, with significant increases in cell numbers over extended culture times. Confocal microscopy images of 3T3 cells cultured within the colloidal gel demonstrated that cells adhered, spread and retained their normal morphologies during proliferation. Furthermore, magnetic separation allowed efficient recovery of cells after their expansion in vitro without need for enzyme-mediated release steps. Trypsin-free cell passages were performed allowing multiple growth, separation and reloading of cells within the colloidal gels. Overall, the results suggest this colloidal gel has potential as a 3D scaffold for in vitro expansion of a variety of cell types and for enzyme free cell harvesting.

Cell and protein compatible 3D bioprinting of mechanically strong constructs for bone repair.

Rapid prototyping of bone tissue engineering constructs often utilizes elevated temperatures, organic solvents and/or UV light for materials processing. These harsh conditions may prevent the incorporation of cells and therapeutic proteins in the fabrication processes. Here we developed a method for using bioprinting to produce constructs from a thermoresponsive microparticulate material based on poly(lactic-co-glycolic acid) at ambient conditions. These constructs could be engineered with yield stresses of up to 1.22 MPa and Young's moduli of up to 57.3 MPa which are within the range of properties of human cancellous bone. Further study showed that protein-releasing microspheres could be incorporated into the bioprinted constructs. The release of the model protein lysozyme from bioprinted constructs was sustainted for a period of 15 days and a high degree of protein activity could be measured up to day 9. This work suggests that bioprinting is a viable route to the production of mechanically strong constructs for bone repair under mild conditions which allow the inclusion of viable cells and active proteins.

Tissue engineered bone using select growth factors: A comprehensive review of animal studies and clinical translation studies in man.

There is a growing socio-economic need for effective strategies to repair damaged bone resulting from disease, trauma and surgical intervention. Bone tissue engineering has received substantial investment over the last few decades as a result. A multitude of studies have sought to examine the efficacy of multiple growth factors, delivery systems and biomaterials within in vivo animal models for the repair of critical-sized bone defects. Defect repair requires recapitulation of in vivo signalling cascades, including osteogenesis, chondrogenesis and angiogenesis, in an orchestrated spatiotemporal manner. Strategies to drive parallel, synergistic and consecutive signalling of factors including BMP-2, BMP-7/OP-1, FGF, PDGF, PTH, PTHrP, TGF-ß3, VEGF and Wnts have demonstrated improved bone healing within animal models. Enhanced bone repair has also been demonstrated in the clinic following European Medicines Agency and Food and Drug Administration approval of BMP-2, BMP-7/OP-1, PDGF, PTH and PTHrP. The current review assesses the in vivo and clinical data surrounding the application of growth factors for bone regeneration. This review has examined data published between 1965 and 2013. All bone tissue engineering studies investigating in vivo response of the growth factors listed above, or combinations thereof, utilising animal models or human trials were included. All studies were compiled from PubMed-NCBI using search terms including 'growth factor name', 'in vivo', 'model/animal', 'human', and 'bone tissue engineering'. Focus is drawn to the in vivo success of osteoinductive growth factors incorporated within material implants both in animals and humans, and identifies the unmet challenges within the skeletal regenerative area.

Surgical delivery of drug releasing poly(lactic-co-glycolic acid)/poly(ethylene glycol) paste with in vivo effects against glioblastoma.

INTRODUCTION: The median survival of patients with glioblastoma multiforme (astrocytoma grade 4) remains less than 18 months despite radical surgery, radiotherapy and systemic chemotherapy. Surgical implantation of chemotherapy eluting wafers into the resection cavity has been shown to improve length of survival but the current licensed therapy has several drawbacks. This paper investigates in vivo efficacy of a novel drug eluting paste in glioblastoma. METHODS: Poly(lactic-co-glycolic acid)/poly(ethylene glycol) (PLGA/PEG) self-sintering paste was loaded with the chemotherapeutic agent etoposide and delivered surgically into partially resected tumours in a flank murine glioblastoma xenograft model. RESULTS: Surgical delivery of the paste was successful and practical, with no toxicity or surgical morbidity to the animals. The paste was retained in the tumour cavity, and preliminary results suggest a useful antitumour and antiangiogenic effect, particularly at higher doses. Bioluminescent imaging was not affected significantly by the presence of the paste in the tumour. CONCLUSIONS: Chemotherapy loaded PLGA/PEG paste seems to be a promising technology capable of delivering active drugs into partially resected tumours. The preliminary results of this study suggest efficacy with no toxicity and will lead to larger scale efficacy studies in orthotopic glioblastoma models.

Evaluation of skeletal tissue repair, part 2: enhancement of skeletal tissue repair through dual-growth-factor-releasing hydrogels within an ex vivo chick femur defect model.

There is an unmet need for improved, effective tissue engineering strategies to replace or repair bone damaged through disease or injury. Recent research has focused on developing biomaterial scaffolds capable of spatially and temporally releasing combinations of bioactive growth factors, rather than individual molecules, to recapitulate repair pathways present in vivo. We have developed an ex vivo embryonic chick femur critical size defect model and applied the model in the study of novel extracellular matrix (ECM) hydrogel scaffolds containing spatio-temporal combinatorial growth factor-releasing microparticles and skeletal stem cells for bone regeneration. Alginate/bovine bone ECM (bECM) hydrogels combined with poly(d,l-lactic-co-glycolic acid) (PDLLGA)/triblock copolymer (10-30% PDLLGA-PEG-PLDLGA) microparticles releasing dual combinations of vascular endothelial growth factor (VEGF), chondrogenic transforming growth factor beta 3 (TGF-ß3) and the bone morphogenetic protein BMP2, with human adult Stro-1+bone marrow stromal cells (HBMSCs), were placed into 2mm central segmental defects in embryonic day 11 chick femurs and organotypically cultured. Hydrogels loaded with VEGF combinations induced host cell migration and type I collagen deposition. Combinations of TGF-ß3/BMP2, particularly with Stro-1+HBMSCs, induced significant formation of structured bone matrix, evidenced by increased Sirius red-stained matrix together with collagen expression demonstrating birefringent alignment within hydrogels. This study demonstrates the successful use of the chick femur organotypic culture system as a high-throughput test model for scaffold/cell/growth factor therapies in regenerative medicine. Temporal release of dual growth factors, combined with enriched Stro-1+HBMSCs, improved the formation of a highly structured bone matrix compared to single release modalities. These studies highlight the potential of a unique alginate/bECM hydrogel dual growth factor release platform for bone repair.

A biodegradable antibiotic-impregnated scaffold to prevent osteomyelitis in a contaminated in vivo bone defect model.

Open fractures are at risk of serious infection and, if infected, require several surgical interventions and courses of systemic antibiotics. We investigated a new injectable formulation that simultaneously hardens in vivo to form a porous scaffold for bone repair and delivers antibiotics at high concentrations to the local site of infection. Duration of antimicrobial activity against Staphylococcus aureus was determined using the serial plate transfer test. Ultimate compressive strength and porosity of the material was measured with and without antibiotics. The material was evaluated in vivo in an ovine medial femoral condyle defect model contaminated with S. aureus. Sheep were sacrificed at either 2 or 13 weeks and the defect and surrounding bone assessed using micro-computed tomography and histology. Antimicrobial activity in vitro persisted for 19-21 days. Sheep with antibiotic-free material and bacteria became infected, while those with antibiotic-containing material and bacteria did not. Similarly, new bone growth was seen in uninoculated animals with plain polymer, and in those with antibiotic polymer with bacteria, but not in sheep with plain polymer and bacteria. The antibiotic-impregnated scaffolds were effective in preventing S. aureus infections whilst supporting bone growth and repair. If translated into clinical practice, this approach might reduce the need for systemic antibiotics.

Evaluation of skeletal tissue repair, part 1: assessment of novel growth-factor-releasing hydrogels in an ex vivo chick femur defect model.

Current clinical treatments for skeletal conditions resulting in large-scale bone loss include autograft or allograft, both of which have limited effectiveness. In seeking to address bone regeneration, several tissue engineering strategies have come to the fore, including the development of growth factor releasing technologies and appropriate animal models to evaluate repair. Ex vivo models represent a promising alternative to simple in vitro systems or complex, ethically challenging in vivo models. We have developed an ex vivo culture system of whole embryonic chick femora, adapted in this study as a critical size defect model to investigate the effects of novel bone extracellular matrix (bECM) hydrogel scaffolds containing spatio-temporal growth factor-releasing microparticles and skeletal stem cells on bone regeneration, to develop a viable alternative treatment for skeletal degeneration. Alginate/bECM hydrogels combined with poly (d,l-lactic-co-glycolic acid) (PDLLGA)/triblock copolymer (10-30% PDLLGA-PEG-PDLLGA) microparticles releasing VEGF, TGF-ß3 or BMP-2 were placed, with human adult Stro-1+ bone marrow stromal cells, into 2mm central segmental defects in embryonic chick femurs. Alginate/bECM hydrogels loaded with HSA/VEGF or HSA/TGF-ß3 demonstrated a cartilage-like phenotype, with minimal collagen I deposition, comparable to HSA-only control hydrogels. The addition of BMP-2 releasing microparticles resulted in enhanced structured bone matrix formation, evidenced by increased Sirius red-stained matrix and collagen expression within hydrogels. This study demonstrates delivery of bioactive growth factors from a novel alginate/bECM hydrogel to augment skeletal tissue formation and the use of an organotypic chick femur defect culture system as a high-throughput test model for scaffold/cell/growth factor therapies for regenerative medicine.

A comparison of polymer and polymer-hydroxyapatite composite tissue engineered scaffolds for use in bone regeneration. An in vitro and in vivo study.

Previous in vitro work demonstrated porous PLA and PLGA both had the mechanical strength and sustained the excellent skeletal stem cell (SSC) growth required of an osteogenic bonegraft substitute, for use in impaction bone grafting. The purpose of this investigation was to assess the effects of the addition of hydroxyapatite (HA) to the scaffolds before clinical translation. PLA, PLA+10% HA, PLGA, and PLGA+10% HA were milled and impacted into discs before undergoing a standardized shear test. Cellular compatibility analysis followed 14 days incubation with human skeletal stems cells (SSC). The best two performing polymers were taken forward for in vivo analysis. SSC seeded polymer discs were implanted subcutaneously in mice. All polymers had superior mechanical shear strength compared with allograft (p < 0.01). Excellent SSC survival was demonstrated on all polymers, but the PLA polymers showed enhanced osteoblastic activity (ALP assay p < 0.01) and collagen-1 formation. In vivo analysis was performed on PLA and PLA+10% HA. MicroCT analysis revealed increased bone formation on the PLA HA (p < 0.01), and excellent neo-vessel formation in both samples. Histology confirmed evidence of de novo bone formation. PLA HA showed both enhanced osteoinductive and osteogenic capacity. This polymer composite has been selected for scaled-up experimentation before clinical translation.

Hydrogels derived from demineralized and decellularized bone extracellular matrix.

The extracellular matrix (ECM) of mammalian tissues has been isolated, decellularized and utilized as a scaffold to facilitate the repair and reconstruction of numerous tissues. Recent studies have suggested that superior function and complex tissue formation occurred when ECM scaffolds were derived from site-specific homologous tissues compared with heterologous tissues. The objectives of the present study were to apply a stringent decellularization process to demineralized bone matrix (DBM), prepared from bovine bone, and to characterize the structure and composition of the resulting ECM materials and DBM itself. Additionally, we sought to produce a soluble form of DBM and ECM which could be induced to form a hydrogel. Current clinical delivery of DBM particles for treatment of bone defects requires incorporation of the particles within a carrier liquid. Differences in osteogenic activity, inflammation and nephrotoxicity have been reported with various carrier liquids. The use of hydrogel forms of DBM or ECM may reduce the need for carrier liquids. DBM and ECM hydrogels exhibited sigmoidal gelation kinetics consistent with a nucleation and growth mechanism, with ECM hydrogels characterized by lower storage moduli than the DBM hydrogels. Enhanced proliferation of mouse primary calvarial cells was achieved on ECM hydrogels, compared with collagen type I and DBM hydrogels. These results show that DBM and ECM hydrogels have distinct structural, mechanical and biological properties and have the potential for clinical delivery without the need for carrier liquids.

Drug delivery to the ear.

Drug delivery to the ear is used to treat conditions of the middle and inner ear such as acute and chronic otitis media, Ménière's disease, sensorineural hearing loss and tinnitus. Drugs used include antibiotics, antifungals, steroids, local anesthetics and neuroprotective agents. A literature review was conducted searching Medline (1966-2012), Embase (1988-2012), the Cochrane Library and Ovid (1966-2012), using search terms 'drug delivery', 'middle ear', 'inner ear' and 'transtympanic'. There are numerous methods of drug delivery to the middle ear, which can be categorized as topical, systemic (intravenous), transtympanic and via the Eustachian tube. Localized treatments to the ear have the advantages of targeted drug delivery allowing higher therapeutic doses and minimizing systemic side effects. The ideal scenario would be a carrier system that could cross the intact tympanic membrane loaded with drugs or biochemical agents for the treatment of middle and inner ear conditions.

Chemical and spatial analysis of protein loaded PLGA microspheres for drug delivery applications.

Polymer microspheres for controlled release of therapeutic protein from within an implantable scaffold were produced and analysed using complimentary techniques to probe the surface and bulk chemistry of the microspheres. Time of Flight - Secondary Ion Mass Spectrometry (ToF-SIMS) surface analysis revealed a thin discontinuous film of polyvinyl alcohol (PVA) surfactant (circa 4.5 nm thick) at the surface which was readily removed under sputtering with C(60). Atomic Force Microscopy (AFM) imaging of microspheres before and after sputtering confirmed that the PVA layer was removed after sputtering revealing poly(lactic-co-glycolic) acid(PLGA). Scanning electron microscopy showed the spheres to be smooth with some shallow and generally circular depressions, often with pores in their central region. The occurrence of the protein at the surface was limited to areas surrounding these surface pores. This surface protein distribution is believed to be related to a burst release of the protein on dissolution. Analysis of the bulk properties of the microspheres by confocal Raman mapping revealed the 3D distribution of the protein showing large voids within the pores. Protein was found to be adsorbed at the interface with the PLGA oil phase following deposition on evaporation of the solvent. Protein was also observed concentrated within pores measuring approximately 2 µm across. The presence of protein in large voids and concentrated pores was further scrutinised by ToF-SIMS of sectioned microspheres. This paper demonstrates that important information for optimisation of such complex bioformulations, including an understanding of the release profile can be revealed by complementary surface and bulk analysis allowing optimisation of the therapeutic effect of such formulations.

Analysis of sintered polymer scaffolds using concomitant synchrotron computed tomography and in situ mechanical testing.

The mechanical behaviour of polymer scaffolds plays a vital role in their successful use in bone tissue engineering. The present study utilised novel sintered polymer scaffolds prepared using temperature-sensitive poly(DL-lactic acid-co-glycolic acid)/poly(ethylene glycol) particles. The microstructure of these scaffolds was monitored under compressive strain by image-guided failure assessment (IGFA), which combined synchrotron radiation computed tomography (SR CT) and in situ micro-compression. Three-dimensional CT data sets of scaffolds subjected to a strain rate of 0.01%/s illustrated particle movement within the scaffolds with no deformation or cracking. When compressed using a higher strain rate of 0.02%/s particle movement was more pronounced and cracks between sintered particles were observed. The results from this study demonstrate that IGFA based on simultaneous SR CT imaging and micro-compression testing is a useful tool for assessing structural and mechanical scaffold properties, leading to further insight into structure-function relationships in scaffolds for bone tissue engineering applications.

Non-local models for the formation of hepatocyte-stellate cell aggregates.

Liver cell aggregates may be grown in vitro by co-culturing hepatocytes with stellate cells. This method results in more rapid aggregation than hepatocyte-only culture, and appears to enhance cell viability and the expression of markers of liver-specific functions. We consider the early stages of aggregate formation, and develop a new mathematical model to investigate two alternative hypotheses (based on evidence in the experimental literature) for the role of stellate cells in promoting aggregate formation. Under Hypothesis 1, each population produces a chemical signal which affects the other, and enhanced aggregation is due to chemotaxis. Hypothesis 2 asserts that the interaction between the two cell types is by direct physical contact: the stellates extend long cellular processes which pull the hepatocytes into the aggregates. Under both hypotheses, hepatocytes are attracted to a chemical they themselves produce, and the cells can experience repulsive forces due to overcrowding. We formulate non-local (integro-partial differential) equations to describe the densities of cells, which are coupled to reaction-diffusion equations for the chemical concentrations. The behaviour of the model under each hypothesis is studied using a combination of linear stability analysis and numerical simulations. Our results show how the initial rate of aggregation depends upon the cell seeding ratio, and how the distribution of cells within aggregates depends on the relative strengths of attraction and repulsion between the cell types. Guided by our results, we suggest experiments which could be performed to distinguish between the two hypotheses.

A mathematical model of liver cell aggregation in vitro.

The behavior of mammalian cells within three-dimensional structures is an area of intense biological research and underpins the efforts of tissue engineers to regenerate human tissues for clinical applications. In the particular case of hepatocytes (liver cells), the formation of spheroidal multicellular aggregates has been shown to improve cell viability and functionality compared to traditional monolayer culture techniques. We propose a simple mathematical model for the early stages of this aggregation process, when cell clusters form on the surface of the extracellular matrix (ECM) layer on which they are seeded. We focus on interactions between the cells and the viscoelastic ECM substrate. Governing equations for the cells, culture medium, and ECM are derived using the principles of mass and momentum balance. The model is then reduced to a system of four partial differential equations, which are investigated analytically and numerically. The model predicts that provided cells are seeded at a suitable density, aggregates with clearly defined boundaries and a spatially uniform cell density on the interior will form. While the mechanical properties of the ECM do not appear to have a significant effect, strong cell-ECM interactions can inhibit, or possibly prevent, the formation of aggregates. The paper concludes with a discussion of our key findings and suggestions for future work.

Efficient assessment of the utility of immortalized Fa2N-4 cells for cytochrome P450 (CYP) induction studies using multiplex quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) and substrate cassette methodologies.

Induction of cytochrome P450 (CYP) 1A2, CYP2B6, and CYP3A4 by 22 prototypical inducers was evaluated in the Fa2N-4 immortalized human hepatic cell line. To facilitate this a duplex one-step quantitative reverse transcriptase-polymerase chain reaction (qRT-PCR) assay for CYP1A2 and CYP3A4 and a substrate cassette allowing simultaneous monitoring of CYP1A2, CYP2B6, CYP2C9, and CYP3A4 activity were developed. CYP1A2 messenger RNA (mRNA) and activity were induced by the prototypical aryl hydrocarbon receptor (AhR) ligand beta-naphthoflavone (E(max) = 217- and 11-fold, respectively, and EC(50) = 8 microM). CYP3A4 mRNA and activity were induced by the prototypical pregnane X receptor (PXR) ligands, rifampicin (E(max) = 36- and 6-fold, respectively, and EC(50) = 4 microM) and phenobarbital (E(max) = 12- and 4-fold, respectively, and EC(50) = 205 microM). No induction of CYP2B6 was detected with several prototypical constitutive androstane receptor (CAR) ligands. A large mRNA-activity E(max) ratio was observed for some time-dependent inhibitors of CYP3A4, whereas EC(50) determinations appeared to be independent of the endpoint. In conclusion, Fa2N-4 cells are a good surrogate for primary human hepatocytes for assessing AhR and PXR-mediated CYP1A2 and CYP3A4 induction, respectively, but not for CAR-mediated CYP2B6 induction. The sensitive and selective methodologies presented in this paper afford maximal data generation and enhanced throughput capability and are readily transferable to primary human hepatocytes or alternate cellular systems.

In situ monitoring of 3D in vitro cell aggregation using an optical imaging system.

Bioreactor systems that maintain cells and tissues in suspension are increasingly popular for culturing 3D constructs to avoid the loss of in vivo cell function associated with traditional 2D culture methods. There is a need for the online monitoring of such systems to provide better understanding and control of the processes involved and to prevent the disruption of these processes caused by offline sampling and endpoint analysis. We describe a system for the imaging and analysis of cell aggregation, over long periods, within a high aspect rotating vessel (HARV). The system exploits side illumination, using an adjustable beam pattern, to restrict the detected light to that scattered by the cell aggregates, thus eliminating the need for the fluorescent labeling of the cells. The in situ aggregation of mammalian cells (MCF-7 breast carcinoma cells) was monitored over an 8 h period and image sequences showing the growth and motion of the aggregates within the bioreactor were obtained. Detailed size and population data have been derived characterizing the development of the aggregates during this time. We show how the number of resolvable aggregates increases to reach a peak and then declines as these aggregates merge. Once formed, remaining aggregates are found to consolidate to form more tightly packed bodies, typically reducing in cross-sectional area by one third. These results provide the basis for the development of an automated feedback system to control the growth of 3D cell cultures for repeatable, reliable, and quality controlled experimentation.

Putting the fizz into chemistry: applications of supercritical carbon dioxide in tissue engineering, drug delivery and synthesis of novel block copolymers.

This paper describes the recent progress at Nottingham towards the exploitation of the unique properties of scCO(2) (supercritical carbon dioxide) for the preparation of polymeric scaffolds for tissue engineering applications and new devices for controlled drug delivery, as well as the synthesis of novel block copolymers by the combination of eROP (enzymatic ring opening polymerization) and controlled polymerization methods for the potential use as drug carriers.

Cell adhesion and mechanical properties of a flexible scaffold for cardiac tissue engineering.

Cardiac tissue engineering is focused on obtaining functional cardiomyocyte constructs to provide an alternative to cellular cardiomyoplasty. Mechanical stimuli have been shown to stimulate protein expression and the differentiation of mammalian cells from contractile tissues. Our aim was to obtain a flexible scaffold which could be used to apply mechanical forces during tissue regeneration. Poly(1,8-octanediol-co-citric acid) (POC) is an elastomer that can be processed into scaffolds for tissue engineering. We investigated the effect of modifying the porosity on the mechanical properties of the POC scaffolds. In addition, the effects of the storage method and strain rate on material integrity were assessed. The maximum elongation of POC porous films varied from 60% to 160% of their original length. A decrease in the porosity caused a rise in this elastic modulus. The attachment of HL-1 cardiomyocytes to POC was assessed on films coated with fibronectin, collagen and laminin. These extracellular matrix proteins promoted cell adhesion in a protein-type- and concentration-dependent manner. Therefore, POC scaffolds can be optimised to meet the mechanical and biological parameters needed for cardiac culture. This porous material has the potential to be used for cardiac tissue engineering as well as for other soft tissue applications.

Supercritical carbon dioxide generated vascular endothelial growth factor encapsulated poly(DL-lactic acid) scaffolds induce angiogenesis in vitro.

The ability to deliver, over time, biologically active vascular endothelial growth factor-165 (VEGF) through tailored designed scaffolds offers tremendous therapeutic opportunities to tissue-engineered therapies. Porous biodegradable poly(DL-lactic) acid (PLA) scaffolds encapsulating VEGF have been generated using supercritical CO2 (scCO2) and the kinetic release and angiogenic activity of these scaffolds examined in vitro and in an ex vivo chick chorioallantoic membrane (CAM) angiogenesis model. After processing through scCO2, VEGF maintained its angiogenic activity as assessed by increased tubule formation of human umbilical vein endothelial cells (HUVEC) cultured on Matrigel (VEGF = 1937 +/- 205 microm; scCO2-VEGF = 2085 +/- 234 microm; control = 1237 +/- 179 microm). VEGF release kinetics from scCO2-VEGF incorporated PLA monolith scaffolds showed a cumulative release of VEGF (2837 +/- 761 rhog/ml) over a 21 day period in culture. In addition, VEGF encapsulated PLA scaffolds increased the blood vessel network in the CAM compared to controls; control, 24.8 +/- 9.6; VEGF/PLA, 44.1 +/- 12.1 (vessels/field). These studies demonstrate that the controlled release of growth factors encapsulated into three-dimensional PLA scaffolds can actively stimulate the rapid development of therapeutic neovascularisation to regenerate or engineer tissues.

The effect of anisotropic architecture on cell and tissue infiltration into tissue engineering scaffolds.

A common phenomenon in tissue engineering is rapid tissue formation on the outer edge of the scaffold which restricts cell penetration and nutrient exchange to the scaffold centre, resulting in a necrotic core. To address this problem, we generated scaffolds with both random and anisotropic open porous architectures to enhance cell and subsequent tissue infiltration throughout the scaffold for applications in bone and cartilage engineering. Hydroxyapatite (HA) and poly(D,L-lactic acid) (P(DL)LA) scaffolds with random open porosity were manufactured, using modified slip-casting and by supercritical fluid processing respectively, and subsequently characterised. An array of porous aligned channels (400 microm) was incorporated into both scaffold types and cell (human osteoblast sarcoma, for HA scaffolds; ovine meniscal fibrochondrocytes, for P(DL)LA scaffolds) and tissue infiltration into these modified scaffolds was assessed in vitro (cell penetration) and in vivo (tissue infiltration; HA scaffolds only). Scaffolds were shown to have an extensive random, open porous structure with an average porosity of 85%. Enhanced cell and tissue penetration was observed both in vitro and in vivo demonstrating that scaffold design alone can influence cell and tissue infiltration into the centre of tissue engineering scaffolds.