15 research outputs found
3D bioprinting for potential use in nasal cartilage reconstruction
3D printing is an additive manufacturing technique that is rapidly gaining traction in health and medical applications. This technique could potentially benefit plastic and reconstructive surgeries by fabricating patient-specific tissue replacements with tissue-like functions and mechanical properties. One specific example in the field of plastic and constructive surgery is nose reconstruction. Current gold standard for nasal reconstruction after rhinectomy or severe trauma involves a three stage surgery that requires a minimum of three and maximum of seven operations to achieve an acceptable result. The surgical procedure require transposition of autologous cartilage grafts in conjunction with coverage using an autologous skin flap. Harvest of autologous rib cartilage requires a major additional procedure which creates donor site morbidity. Additionally, major nasal reconstruction also requires sculpting autologous cartilages to form a cartilage framework, which is complex, highly-skill demanding and time-consuming.
These drawbacks of the current approach for nasal reconstruction are some of the reasons why facial plastic and reconstructive surgeons are interested in the application of tissue engineering and 3D printing for reconstructive surgeries. To address these clinical challenges, the aim of the work presented in this thesis was to fabricate a personalised 3D bioprinted composite scaffold for nasal reconstruction mimicking the mechanical properties and architecture of nasal cartilage. The composite consists of biodegradable thermoplastic polycaprolactone (PCL) to provide structural support, and cell-laden thermoresponsive and UV crosslinkable gelatin methacrylate (GelMA) to act as a cell carrier.
We first investigated the appropriate cell source to use for cartilage tissue engineering and 3D bioprinting. Primary sheep articular chondrocytes (ShCh) and sheep bone marrow derived Mesenchymal Stem Cells (ShMSCs) were isolated, expanded and differentiated; followed by an assessment of the effects of the 3D printing process on cell viability and functionality. From these studies it was observed that ShCh were easier to isolate and expand than ShMSCs because less steps are required and the doubling time is 50% shorter. Additionally, 80% of the ShCh survived the printing process compared to a 50% of the ShMSCs, suggesting that chondrocytes were able to tolerate higher stress caused by the 3D printing process.
PCL and poly (lactic-co-glycolic acid) (PLGA) scaffolds were printed and seeded with chondrocytes post-printing. The printing process and the 3D printed structures of these polymers were characterised before and after printing by measuring their molecular weight, thermal and mechanical properties. It was found that the printing process reduced the molecular weight of PLGA by 50% percent due to thermal degradation. Consequently, its glass transition temperature and youngâs modulus decreased post printing. On the contrary, PCLâs molecular weight remain unchanged after printing. Characterisation of the chondrocytes showed that whilst both scaffold materials supported cell attachment the ECM secreted deformed the PLGA whilst the PCL scaffolds were unaffected. Due to superior mechanical properties PCL was selected to 3D print the personalised nose scaffolds. Additional studies on the 3D printed scaffolds showed that controlling the surface pores of scaffolds was important for cell infiltration and proliferation Scaffolds with larger surface pores were 3D printed and these resulted in increased cell seeding and proliferation demonstrated by DNA quantification.
Moreover, the printing process of the cell carrier GelMA was optimised by utilising its thermoresponsive properties. A rheological study of three different concentrations of GelMA was performed in order to identify the most suitable for bioprinting. GelMA 15% and 20% at 15 °C and 18 °C respectively were found the appropriate ones. Finally, multi-material 3D bioprinting of PCL and chondrocyte-laden GelMA was utilised for making cartilage constructs. The 3D bioprinted constructs showed neocartilage formation and similar mechanical properties to nasal alar cartilage after a 50-day culture period. Neocartilage formation was evidenced by the presence of glycosaminoglycans and collagen type II after cultivation. The findings in this thesis therefore support the feasibility of using 3D bioprinted composite constructs for nasal reconstruction
Synthesis of methacrylate-terminated block copolymers with reduced transesterification by controlled ring-opening polymerization
This work presents a robust method to achieve the synthesis of low molecular weight polyesters 26 via ring-opening polymerization (ROP) initiated by 2-hydroxyethyl-methacrylate (HEMA) 27 when using triazabicyclodecene (TBD) as catalyst. The effect that the HEMA:TBD ratio has 28 upon the final reaction rate and final polymer molecular architecture is discussed. The optimum 29 HEMA:TBD ratio and reaction conditions required to minimize competing transesterification 30 reactions were determined, in order to synthesize successfully the target ROP macromonomer 31 species containing only a single 2-methacryloyloxyethyl end-group. Additionally, to confirm 32 the terminal end-group fidelity of the product macromonomers and confirm TBD utility for 33 block copolymer manufacture, a small series of di-block polyesters were synthesized using 34 TBD and shown to exhibit good control over the final polymer structure whilst negating the 35 side transesterification reactions, irrespective of the monomers used
A Reactive Prodrug Ink Formulation Strategy for Inkjet 3D Printing of Controlled Release Dosage Forms and Implants
We propose a strategy for creating tuneable 3D printed drug delivery devices. 3D printing offers the opportunity for improved compliance and patient treatment outcomes through personalisation, but bottlenecks include finding formulations that provide a choice of drug loading and release rate, are tuneable and avoid the need for surgical removal. Our solution is to exploit 3D inkjet printing freedoms. We use a reactive prodrug that can polymerize into drug-attached macromolecules during 3D printing, and by tuning the hydrophilicity we can facilitate or hinder hydrolysis, which in turn controls the drug release. To demonstrate this approach, we attach ibuprofen to 2-hydroxyethyl acrylate through a cleavable ester bond, formulate it for inkjet 3D printing, and then print to produce a solid dosage form. This allows a much higher loading than is usually achievable-in our case up to 58 wt%. Of equal importance, the 3D inkjet printing freedoms mean that our drug delivery device is highly tuneable: by selection of spacer monomers to adjust the hydrophilicity; through geometry; by spatially varying the components. Consequently, we create bespoke, hierarchical release systems, from the molecular to macro. This approach represents a new paradigm for the formulation of printable inks for drug-loaded medical devices
Characterisation of the surface structure of 3D printed scaffolds for cell infiltration and surgical suturing
3D printing is of great interest for tissue engineering scaffolds due to the ability to form complex geometries and control internal structures, including porosity and pore size. The porous structure of scaffolds plays an important role in cell ingrowth and nutrition infusion. Although the internal porosity and pore size of 3D printed scaffolds have been frequently studied, the surface porosity and pore size, which are critical for cell infiltration and mass transport, have not been investigated. The surface geometry can differ considerably from the internal scaffold structure depending on the 3D printing process. It is vital to be able to control the surface geometry of scaffolds as well as the internal structure to fabricate optimal architectures. This work presents a method to control the surface porosity and pore size of 3D printed scaffolds. Six scaffold designs have been printed with surface porosities ranging from 3% to 21%. We have characterised the overall scaffold porosity and surface porosity using optical microscopy and microCT. It has been found that surface porosity has a significant impact on cell infiltration and proliferation. In addition, the porosity of the surface has been found to have an effect on mechanical properties and on the forces required to penetrate the scaffold with a surgical suturing needle. To the authors' knowledge, this study is the first to investigate the surface geometry of extrusion-based 3D printed scaffolds and demonstrates the importance of surface geometry in cell infiltration and clinical manipulation
Correction to âBespoke 3D-Printed Polydrug Implants Created via Microstructural Control of Oligomersâ
The chemical structure of the drug trandolapril has been corrected in Figure 4c. The conclusions of the work have not been affected by this correction. (Figure present)
Bespoke 3D-Printed Polydrug Implants Created via Microstructural Control of Oligomers
Controlling the microstructure of materials by means of phase separation is a versatile tool for optimizing material properties. Phase separation has been exploited to fabricate intricate microstructures in many fields including cell biology, tissue engineering, optics, and electronics. The aim of this study was to use phase separation to tailor the spatial location of drugs and thereby generate release profiles of drug payload over periods ranging from 1 week to months by exploiting different mechanisms: polymer degradation, polymer diluent dissolution, and control of microstructure. To achieve this, we used drop-on-demand inkjet three-dimensional (3D) printing. We predicted the microstructure resulting from phase separation using high-throughput screening combined with a model based on the Flory-Huggins interaction parameter and were able to show that drug release from 3D-printed objects can be predicted from observations based on single drops of mixtures. We demonstrated for the first time that inkjet 3D printing yields controllable phase separation using picoliter droplets of blended photoreactive oligomers/monomers. This new understanding gives us hierarchical compositional control, from droplet to device, allowing release to be "dialled up"without manipulation of device geometry. We exemplify this approach by fabricating a biodegradable, long-term, multiactive drug delivery subdermal implant ("polyimplant") for combination therapy and personalized treatment of coronary heart disease. This is an important advance for implants that need to be delivered by cannula, where the shape is highly constrained and thus the usual geometrical freedoms associated with 3D printing cannot be easily exploited, which brings a hitherto unseen level of understanding to emergent material properties of 3D printing
Poly (glycerol adipate) (PGA) backbone modifications with a library of functional diols: Chemical and physical effects
Enzymatically synthesised poly(glycerol adipate) (PGA) has shown a palette of key desirable properties required for a biomaterial to be considered a âversatile polymeric toolâ in the field of drug delivery. PGA and its variations can self-assemble into nanoparticles (NPs) and interact at different levels with small active molecules. PGA derivatives are usually obtained by functionalising the glyceryl side hydroxyl group present along the main polymer scaffold. However, if the synthetic pathways are not finely tuned, the self-assembling ability of these new polymeric modifications might be hampered by the poor amphiphilic balance. For this reason, we have designed a straightforward one-pot synthetic modification, using a small library of diols in combination with glycerol, aimed at altering the backbone of the polymer without affecting the hydrophilic glyceryl portion. The diols introduce additional functionality into the backbone of PGA alongside the secondary hydroxyl group already present. We have investigated how extra functionalities along the polymer backbone alter the final polymer reactivity as well the chemical and biological properties of the nanoparticles. In addition, with the intent to further improve the green credentials of the enzymatic synthesis, a solvent derived from renewable resources, (2-methyl tetrahydrofuran, 2-MeTHF) was employed for the synthesis of all the PGA-variants as a replacement for the more traditionally used and fossil-based tetrahydrofuran (THF). In vitro assays carried out to evaluate the potential of these novel materials for drug delivery applications demonstrated very low cytotoxicity characteristic against NIH 3T3 model cell line
Glycerol-based sustainably sourced resin for volumetric printing
Volumetric Additive Manufacturing (VAM) represents a revolutionary advancement in the field of Additive Manufacturing, as it allows for the creation of objects in a single, cohesive process, rather than in a layer-by-layer approach. This innovative technique offers unparalleled design freedom and significantly reduces printing times. A current limitation of VAM is the availability of suitable resins with the required photoreactive chemistry and from sustainable sources. To support the application of this technology, we have developed a sustainable resin based on polyglycerol, a bioderived (e.g., vegetable origin), colourless, and easily functionisable oligomer produced from glycerol. To transform polyglycerol-6 into an acrylate photo-printable resin we adopted a simple, one-step, and scalable synthesis route. Polyglycerol-6-acrylate fulfils all the necessary criteria for volumetric printing (transparency, photo-reactivity, viscosity) and was successfully used to print a variety of models with intricate geometries and good resolution. The waste resin was found to be reusable with minimal performance issues, improving resin utilisation and minimising waste material. Furthermore, by incorporating dopants such as poly(glycerol) adipate acrylate (PGA-A) and 10,12-pentacosadyinoic acid (PCDA), we demonstrated the ability to print objects with a diverse range of functionalities, including temperature sensing probes and a polyester excipient, highlighting the potential applications of these new resins
Nanoscale coatings for ultralow dose BMP-2-driven regeneration of critical-sized bone defects
While new biomaterials for regenerative therapies are being reported in the literature, clinical translation is slow. Some existing regenerative approaches rely on high doses of growth factors, such as bone morphogenetic protein-2 (BMP-2) in bone regeneration, which can cause serious side effects. An ultralow-dose growth factor technology is described yielding high bioactivity based on a simple polymer, poly(ethyl acrylate) (PEA), and report mechanisms to drive stem cell differentiation and bone regeneration in a critical-sized murine defect model with translation to a clinical veterinary setting. This material-based technology triggers spontaneous fibronectin organization and stimulates growth factor signalling, enabling synergistic integrin and BMP-2 receptor activation in mesenchymal stem cells. To translate this technology, for the first time, plasma-polymerized PEA is used on 2D and 3D substrates to enhance cell signalling in vitro, showing the complete healing of a critical sized bone injury in mice in vivo. Efficacy is demonstrated in a MĂŒnsterlĂ€nder dog with a nonhealing humerus fracture, establishing the clinical translation of advanced ultralow-dose growth factor treatment
3D bioprinting for potential use in nasal cartilage reconstruction
3D printing is an additive manufacturing technique that is rapidly gaining traction in health and medical applications. This technique could potentially benefit plastic and reconstructive surgeries by fabricating patient-specific tissue replacements with tissue-like functions and mechanical properties. One specific example in the field of plastic and constructive surgery is nose reconstruction. Current gold standard for nasal reconstruction after rhinectomy or severe trauma involves a three stage surgery that requires a minimum of three and maximum of seven operations to achieve an acceptable result. The surgical procedure require transposition of autologous cartilage grafts in conjunction with coverage using an autologous skin flap. Harvest of autologous rib cartilage requires a major additional procedure which creates donor site morbidity. Additionally, major nasal reconstruction also requires sculpting autologous cartilages to form a cartilage framework, which is complex, highly-skill demanding and time-consuming.
These drawbacks of the current approach for nasal reconstruction are some of the reasons why facial plastic and reconstructive surgeons are interested in the application of tissue engineering and 3D printing for reconstructive surgeries. To address these clinical challenges, the aim of the work presented in this thesis was to fabricate a personalised 3D bioprinted composite scaffold for nasal reconstruction mimicking the mechanical properties and architecture of nasal cartilage. The composite consists of biodegradable thermoplastic polycaprolactone (PCL) to provide structural support, and cell-laden thermoresponsive and UV crosslinkable gelatin methacrylate (GelMA) to act as a cell carrier.
We first investigated the appropriate cell source to use for cartilage tissue engineering and 3D bioprinting. Primary sheep articular chondrocytes (ShCh) and sheep bone marrow derived Mesenchymal Stem Cells (ShMSCs) were isolated, expanded and differentiated; followed by an assessment of the effects of the 3D printing process on cell viability and functionality. From these studies it was observed that ShCh were easier to isolate and expand than ShMSCs because less steps are required and the doubling time is 50% shorter. Additionally, 80% of the ShCh survived the printing process compared to a 50% of the ShMSCs, suggesting that chondrocytes were able to tolerate higher stress caused by the 3D printing process.
PCL and poly (lactic-co-glycolic acid) (PLGA) scaffolds were printed and seeded with chondrocytes post-printing. The printing process and the 3D printed structures of these polymers were characterised before and after printing by measuring their molecular weight, thermal and mechanical properties. It was found that the printing process reduced the molecular weight of PLGA by 50% percent due to thermal degradation. Consequently, its glass transition temperature and youngâs modulus decreased post printing. On the contrary, PCLâs molecular weight remain unchanged after printing. Characterisation of the chondrocytes showed that whilst both scaffold materials supported cell attachment the ECM secreted deformed the PLGA whilst the PCL scaffolds were unaffected. Due to superior mechanical properties PCL was selected to 3D print the personalised nose scaffolds. Additional studies on the 3D printed scaffolds showed that controlling the surface pores of scaffolds was important for cell infiltration and proliferation Scaffolds with larger surface pores were 3D printed and these resulted in increased cell seeding and proliferation demonstrated by DNA quantification.
Moreover, the printing process of the cell carrier GelMA was optimised by utilising its thermoresponsive properties. A rheological study of three different concentrations of GelMA was performed in order to identify the most suitable for bioprinting. GelMA 15% and 20% at 15 °C and 18 °C respectively were found the appropriate ones. Finally, multi-material 3D bioprinting of PCL and chondrocyte-laden GelMA was utilised for making cartilage constructs. The 3D bioprinted constructs showed neocartilage formation and similar mechanical properties to nasal alar cartilage after a 50-day culture period. Neocartilage formation was evidenced by the presence of glycosaminoglycans and collagen type II after cultivation. The findings in this thesis therefore support the feasibility of using 3D bioprinted composite constructs for nasal reconstruction