Development of Organic Solvent Free Fibrillar Scaffolds for Tissue Engineering Applications

Abstract

The increasing use of polymeric biomaterials in the health industries has created many new methods for the processing of polymers into lifesaving products; however, many processes often include the use of toxic organic solvents throughout the manufacturing and fabrication process. The micro-/nano-fibrillar composite (MFC/NFC) technique has the potential to produce tissue scaffolds completely free from organic solvents. The development of tissue scaffolds using this manufacturing method is the main topic of this research. Tissue scaffolds were fabricated using the MFC/NFC concept from poly(L-lactide) (PLA) and poly(ethylene terephthalate) glycol (PETG). These MFC scaffolds were studied alongside, and compared to, the well-known and studied electrospinning technique. Biocompatibility is one of the most important characteristics a scaffold must possess - without it, the best structure or material would not be suitable for use as a scaffold. In this study, cell culture techniques have been used to study mouse osteoblast-like MC3T3-E1 cell and primary rat tenocytes. The MFC/NFC scaffold cytocompatibility was assessed to qualitatively and quantitatively understand the cellscaffold interaction. Live/dead staining provides a means of differentiating between live and dead cells that have been cultured on the scaffolds. It has been found that there is a confluent layer of live cells present on all scaffolds after seven days of culture. To quantitatively assess the cell growth, alamarBlue® assays are conducted using both osteoblasts and tenocytes. The cell numbers for both the PLA and PETG scaffolds fabricated from the MFC/NFC technique, increase at a faster rate than the electrospun scaffolds, over the culture periods of the cells. Sirius red assays are used to detect collagen deposition from the cells. Preliminary experiments show that the collagen levels on the MFC/NFC scaffolds have a tendency to increase over the culture period. Gene expression analysis is studied on the MFC/NFC scaffolds using primary rat tenocytes. Results show that scleraxis and tenomodulin; markers for tenocytic behaviour, increase over the 14 day culture period. The MFC/NFC technique which uses a common polymer manufacturing process, extrusion, has been used to successfully create nanoporous, three-dimensional interconnected scaffolds. The biocompatibility of these scaffolds was initially tested using mouse osteoblasts. The cells on the MFC/NFC scaffolds attached and proliferated better than on the electrospun scaffolds. This led to further tests on the MFC/NFC scaffolds by studying the cell differentiation potential. This study demonstrates that the MFC/NFC technique has the potential to produce completely organic solvent-free scaffolds for bone and tendon regeneration applications.