Salem, Aliasger K.
(2002)
An injectable degradable porous polymer scaffold for tissue engineering and drug delivery.
PhD thesis, University of Nottingham.
Abstract
Cell transplantation on biodegradable scaffolds is an established approach in tissue engineering to the problem of the regeneration of diseased or damaged tissues. As cells grow and organise themselves, they secrete their own extracellular matrix, while the polymer degrades into natural metabolites resulting in eventual natural tissue replacement. Polymeric materials used for these scaffolds must satisfy a number of requirements. These include defined cell-interactive properties, porosity, biodegradability, mechanical and controlled release properties.
To date, scaffolds have been designed to conform to these requirements. However, the need to perform defined three-dimensional structures requires prior knowledge of the dimensions of the defect or cavity to be filled. Furthermore the general use of toxic solvents in the processing of these scaffolds prevents the incorporation of biological agents and cells during fabrication. Therefore, poor transportation of cells through the scaffolds can result in low cell seeding efficiencies. Finally such scaffolds require an invasive operation for transplantation of the material.
In contrast a number of injectable materials have been proposed and investigated. The transformation from liquid pre-cursor to gel in such systems can, however, require cell harmful trigger signals such as UV exposure or pH changes. Furthermore, these injectable gels lack a porous structure preventing effective cell migration and restricting tissue formation and vascularisation tothe barrier of diffusion for signalling and nutrient molecules.
The work in this thesis presents a scaffold that is both injectable and conforms to the requirements of water-insoluble porous scaffolds. This starts with the synthesis of a biotinylated poly (lactic acid)-poly (ethylene glycol) (PLA-PEG) copolymer. The polymer is degradable, protein resistant and cell interactive when used in conjunction with biotinylated cell adhesive peptides. The biotin unit tethered to the PEG-PLA also provides the polymer with self-assembling properties when used in conjunction with avidin.
In contrast to alternative injectable materials, the scaffold presented in this thesis is porous. This porosity is necessary for tissue ingrowth and vascularization. Therefore, before progressing on to the manufacture of the scaffold, a systematic study of two cell types involved in vascularisation was carried out over defined pore features. These studies revealed that cell behaviour over pore features was related to cell type, cell density and pore size. This had significant implications for the injectable scaffold in development because proposed advantages were delivery of a variety of cell types, controlled porous structure, and efficient cell seeding.
Microparticles were then manufactured from the PLA-PEG-biotin using a single emulsion manufacturing process. Surface Plasmon Resonance (SPR) confirmed that these microparticles would bind efficiently to avidin. The condition for optimum self-assembling of particles was then determined using aggregation studies. These studies showed that a critical quantity of avidin was required for microparticles to aggregate together. The ability to aggregate particles of different sizes leads to the potential for controlling scaffold porosity. Rheological testing showed that the scaffold's mechanical properties could be tailored to that of the tissue in which regeneration is required.
The self-assembly of microparticles was also demonstrated to form complex three-dimensional scaffolds without the use of toxic solvents. Scaffolds prepared in simulated tissues maintained shape upon injection. Scaffolds were then self-assembled with cells entrapped within them. Cell viability within the self-assembling scaffolds was confirmed by Alamar Blue assays. In vivo studies have demonstrated that cell-scaffold composites permit tissue ingrowth and thus readily undergo vascularisation.
The novel molecular-interaction mechanism of self-assembly of these scaffolds differentiates this material from other injectable systems. The formation of porous scaffolds within a cavity or a soft-tissue could be a pre-requisite for tissue remodelling using new cell sources that are dependent on vascularisation and tissue ingrowth. The basic component of the scaffold is a biodegradable microparticle that presents a protein resistant surface with biotinylated moieties. Therefore, standard controlled release technologies and biotin-avidin mediated surface engineering can be combined with the self-assembly to form biomimetic scaffolds that stimulate integrin-mediated cell adhesion and then release growth factors.
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