Many of the current tissue-engineering scaffold-based strategies have suffered from limited cell depth viability when cultured in vitro, with viable cells existing within the outer 250-500 /im from the fluid-scaffold interface. This is prim arily believed to be due to the lack of nutrient delivery into and waste removal from the inner regions of the scaffold construct. Other issues associated with scaffolds involve poor seeding efficiencies and limited cell penetration resulting in heterogenous cellular distributions.

This work focused on developing a novel hydroxyapatite multi-domain porous scaffold architecture {i.e. a scaffold providing a discrete domain for cell occupancy and a separate domain for nutrient delivery) with the specific objectives of em bodying in one scaffold the structures required to optimise cell seeding, cell proliferation and m igration and potentially to facilitate vascularisation once implanted in vivo. It has been dem onstrated th a t incorporating unidirectional macrochannels into a porous scaffold m aterial significantly enhances initial cell seeding distribution, while m aintaining relatively high seeding efficiencies. This work has also dem onstrated th a t for seeding of three-dimensional porous scaffolds, appropriately matching the seeding volume to the saturation capacity of the scaffold is the optimum approach in terms of promoting homogeneous cell seeding while m aintaining relatively high seeding efficiencies.

Through in vitro static culturing it has been shown th a t providing a discrete domain for nutrient diffusion and metabolic waste removal is insufficient to maintain cell viability throughout the entire scaffold depth. Finite element analysis incorporating measured effective diffusion coefficients, cellular oxygen consumption rates and cellular distributions demonstrate that the formation of this necrotic core cannot be solely due to oxygen diffusion limitations and is possibly due to other diffusive limited species concentrations such as glucose or metabolic waste products. Therefore, in order to overcome these diffusion based limitations it has been shown th at dynamic rotational culturing of multi-domain porous scaffolds can maintain uniform cell viability throughout the scaffold depth with increasing culturing time and increase the rate and extent of cell proliferation (~ 2 fold) compared to static culturing.

This work provides significant insight into current tissue engineering strategies and raises important questions regarding scaffold architecture and short-term conditioning of scaffolds in vitro prior to in vivo implantation.