Abstract

Stroke and traumatic brain injury are among the leading causes of death in the world. Until now, there are no effective treatments available. Current pharmaceutical treatments have limited benefits to repair the damaged tissue. Brain tissue engineering is a promising strategy to help brain regeneration after the damage induced by stroke or traumatic brain injury. In this thesis, our work focused on designing and evaluating appropriate silk fibroin-based hydrogels combined with stem cells therapy for brain tissue regeneration. The work initially started from looking for appropriate silk fibroin-based hydrogel substrates which can support the viability and neural differentiation of pluripotent cells. Mouse embryonic stem cells (mESC) were used as a model. Different processing procedures of silk fibroin-based hydrogel substrates were prepared by chemical genipin crosslinking and physical sonication crosslinking. The viability and neural differentiation of pluripotent cells on these hydrogel substrates were evaluated, using tissue culture plates (TCP) as control. Different crosslinking processes were found to modulate the neural differentiation of pluripotent cells. Chemical genipin crosslinked hydrogel substrates could inhibit the neural differentiation of mESC compared to control TCP, while the physical sonication crosslinked hydrogel substrates could support the neural differentiation as TCP. According to the results obtained in the first stage, the physically sonication-crosslinked 3D silk fibroin hydrogel was produced to encapsulate human neural stem cells (hNSC). In order to improve the hNSC attachment and neuronal differentiation, the isoleucine-lysine-valine-alanine-valine (IKVAV) peptide derived from laminin was covalently conjugated to the silk fibroin. The viability and neural differentiation of hNSC were evaluated in the unmodified and IKVAV peptide modified silk fibroin hydrogels. We found that the IKVAV peptide modified silk fibroin hydrogel could increase the viability, proliferation and neuronal differentiation of hNSC. Furthermore, the angiogenesis potential of sonication-induced 3D silk fibroin unmodified and modified with IKVAV and a scramble peptide VVIAK (as control) were evaluated in a human outgrowth endothelial cells (OEC) mono-culture system and a co-culture system in which OEC were cultured with human bone marrow mesenchymal stem cells (BM-MSC). Both the silk fibroin unmodified and modified with IKVAV peptide could not induce angiogenesis in the mono-culture system under the VEGF condition. However, in the co-culture system, we found that unmodified, IKVAV-modified and VVIAK-modified silk fibroin hydrogels all could support angiogenesis. Furthermore, there were no significant differences among unmodified, IKVAV modified and VVIAK modified silk fibroin hydrogels influencing on angiogenesis structure and gene expression related to angiogenesis. The thesis will introduce the detailed work in three different chapters (from chapter 3 to chapter 5) respectively.