Isolation of selected populations from heterogeneous cell mixtures and retrieval of the captured population of interest for regenerative medicine and diagnostics applications is one of the challenges that may be addressed by microfluidics. An affinity adhesion strategy was tested using the tetrapeptides RGDS (arg-gly-asp-ser), REDV (arg-glu-asp-val) and VAPG (val-ala-pro-gly) to modify an alginate hydrogel surface layer to selectively adhere fibroblast (FB), endothelial (EC) and smooth muscle cell (SMC) populations, respectively, of the non-myocyte cardiac cell fraction. Incorporation of peptides into sodium alginate gel surface coatings demonstrated a preferential, seeding density-dependent adhesion relationship on alginate-RGDS when tested with a cardiomyocyte-depleted cell suspension in both static culture and in microfluidic devices. Seeding density-dependent attachment was seen with close to 100% release of viable cells from coated surfaces upon application of ethylenediaminetetraacetic acid (EDTA). Further work will optimize the system with REDV and VAPG to capture ECs and SMCs.

Isolation of selected populations from heterogeneous cell mixtures and retrieval of the captured population of interest for regenerative medicine and diagnostics applications is one of the challenges that may be addressed by microfluidics. An affinity adhesion strategy was tested using the tetrapeptides RGDS (arg-gly-asp-ser), REDV (arg-glu-asp-val) and VAPG (val-ala-pro-gly) to modify an alginate hydrogel surface layer to selectively adhere fibroblast (FB), endothelial (EC) and smooth muscle cell (SMC) populations, respectively, of the non-myocyte cardiac cell fraction. Incorporation of peptides into sodium alginate gel surface coatings demonstrated a preferential, seeding density-dependent adhesion relationship on alginate-RGDS when tested with a cardiomyocyte-depleted cell suspension in both static culture and in microfluidic devices. Seeding density-dependent attachment was seen with close to 100% release of viable cells from coated surfaces upon application of ethylenediaminetetraacetic acid (EDTA). Further work will optimize the system with REDV and VAPG to capture ECs and SMCs.

Tissue engineering has the potential to create new tissue and organs from cultured cells for transplantation. Biodegradable and biocompatible scaffolds play a vital role in the transfer of the cultured cells to a new tissue. Various scaffolds for soft tissue engineering have been developed, however there is not any structure totally mimicking the natural extracellular matrix (ECM), ready to use. In this study biodegradable and biocompatible scaffolds were developed from natural polymers by tissue engineering approach and tested in vitro. Scaffolds (SCAF) were prepared with freeze drying and composed of chitosan, gelatin and dermatan sulfate. Polymer solutions were treated with different stirring rates (500 rpm and 2000 rpm), freezing temperatures (-20 &deg / C and -80 &deg / C) and molding (cylindrical mold and petri dish) to achieve porous structure in order to provide sufficient space for cell growth and extracellular matrix production. Among the prepared scaffolds at different conditions, the scaffolds prepared at 500 rpm and frozen at -80 &deg / C, (SCAF-1), was chosen for further studies. These scaffolds achieved 0.512 MPa tensile strength, with 9.165 MPa tension modulus and 3.428 MPa compression modulus. Besides in lysozyme containing degradation medium they conserved their integrity and lost about 30 % of their initial weight in 30 days period. Mechanical and enzymatic degradation tests showed that scaffolds have physical integrity for the tissue engineering applications. To mimic the natural tissue and enhance cell growth, biologically active arginine &amp / #8211 / glycine - aspartic acid - serine (RGDS) peptides and platelet derived growth factor-BB (PDGF-BB) were immobilized on the SCAF-1. Fibroblast cells were seeded on the scaffolds containing RGDS, (SCAF-1-RGDS), and PDGF-BB, (SCAF-1-RGDS-PDGF), and incubated in media either free of serum or containing serum. Scaffolds immobilized with RGDS and PDGF-BB had the highest attached cell number by the day 15. Florescence microscopy studies also indicated that RGDS and RGDS-PDGF modified scaffolds were more suitable than controls, (SCAF-1), for cell growth and proliferation. According to scanning electron microscopy (SEM) results, modified scaffolds demonstrated better cell morphology and attachment of cells. Based on the obtained results, it can be concluded that RGDS-PDGF immobilized chitosan-gelatin-dermatan sulfate systems have a great potential to be used as a scaffold for soft tissue engineering applications.

The central nervous system (CNS) is susceptible to several disorders that can affect the structure or function of the brain or spinal cord, such as stroke and spinal cord injury (SCI). CNS disorders are complex, frequently causing failure of cognitive, motor and sensory functions. Unfortunately, there are only a few care alternatives for patients with CNS disorders, due to the limited capacity of the CNS to spontaneously regenerate; what expresses the need to develop innovative solutions, such as scaffolds that also could act as drug delivery systems to promote tissue and functional repairs in the CNS. To achieve this goal, three main projects were developed in this thesis. In the first project, a novel drug releasing duraplasty that can be applied as part of decompressive craniectomy (DC) was designed and tested. While DC can significantly reduce the risk of death, this procedure does not reverse the stroke damage. Thus, biosynthesized cellulose (BC) was used to produce a new duraplasty loaded with growth factors. The in vivo animal studies revealed that our duraplasty had excellent biocompatibility when implanted onto rodents’ brains. In the second project, BC tubes were prepared and nerve growth factor was incorporated into the tubes to be used as potential nerve guides to assist with the reconstitution of nerve tissues across SCI lesion. Physical and mechanical properties of the drug delivery systems produced were evaluated and compared to the neural native tissue. In addition, cell cultures demonstrated that growth factors released from both drug delivery systems were bioactive for over 7 days. In the third project, linear and 2-branched peptides were synthesized as potential bioactive molecules to improve tissue regeneration. These peptides, containing the RGDS sequence, were synthesized through Solid Phase Peptide Synthesis and characterized by mass spectrometry, high-performance liquid chromatography, and their conformational structures were analyzed by an energy minimized 3D model. In summary, this thesis explores the use of BC as drug releasing systems, which are promising and clinically relevant strategies to enhance nerve regeneration for many patients facing physical, mental and financial strains due to stroke, SCI or other difficult-to-cure injuries to the CNS.