Layer-by-Layer Assembled Protein Films And Their Application In The Improvement Of Cytocompatibility

Biodegradable polymer can degrade to natural metabolites, can be easily processed, and its mechanical properties and degradation properties can be adjusted to meet particular needs, so this kind of material has great potential in tissue engineering. Presumably due to its hydrophobicity and lack of appropriate functional groups at the surface, this kind of material exhibits poor cytocompatibility. Cell has the special response to the extracellular matrix, so extracellular matrix can be used as the scaffold for the cell growth. However, most protein's structure will change after they adsorbed on the substrates that results in the loss of its activity, furthermore, most protein need a warm water environment to survive, so it is very important to choose a suitable method to protect the protein activity after they assembled on the multilayers. In our work, we use the layer-by-layer method to obtain the protein films and study the protein films application in the improvement of cytocompatibility.In chapter 2, four different charge density of carrageenan were assembled with PDDA using the layer-by-layer method. The dynamic assembling process was monitored by the quartz crystal microbalance with the dissipation monitoring (QCM-D), the adsorbed carrageenan amount on the PDDA surface depended on the carrageenan charge density. FTIR and water contact angle results showed that the four different charge density of carrageenan can self-assemble with PDDA successfully. Asι-carrageenan having the special helix structure in solution, AFM results showed that the structure can be kept in the multilayers, so using the layer-by-layer method the surface property can be easily controlled. Subsequently, the model protein BSA was chosen to study the protein adsorption behavior on the (PDDA/carageenan)n multilayer surfaces. The adsorption behavior depended on the pH of BSA solution greatly, the parabola adsorption curve occurred on the different charged substrates and the most adsorbed amount happened to the near of IEP point. At the same time, some effects such as the protein concentration, charge density of substrate, the top surface material and the number of (PDDA/carrageenan)n films also have big effects on the protein adsorption behavior. Based on the dissipation factor (ΔD) we can suppose the BSA film structure on the different substrates.Based on the above work about electrostatic layer-by-layer process and the knowledge about protein adsorption behavior on the charged substrate. In chapter 3, the amphoteric nature of the gelatin was exploited and the gelatin was adsorbed to the negatively charged PLLA/PSS and positively charged PLLA/PAH at pH=3.4 and 7.4, respectively. XPS and water contact angle data indicated that the gelatin adsorption at pH = 3.4 resulted in much higher surface coverage by gelatin than at pH = 7.4. All the modified PLLA surfaces became more hydrophilic than the virgin PLLA. Chondrocyte culture was used to test the cell attachment, cell morphology and cell viability on the modified PLLA substrates. The results showed that the PAH and PSS modified PLLA exhibited better cytocompatibility than virgin PLLA, and the incorporation of the gelatin on these modified PLLA substrates further improved their cytocompatibility, with the PLLA/PSS substrate treated with the gelatin at pH = 3.4 being the best, exceeding the chondrocyte compatibility of the tissue culture polystyrene.Later, further studied the gelatin self-assembled behavior, proposed that electrostatic assembly of one species can be realized using gelatin as a polyampholyte. Under suitable conditions where the electrostatic attraction and repulsion were both significant and in balance, linear growth of multilayers driven by electrostatic interactions was sustained over many successive assembly steps, and the maximum amount of adsorption of each layer was reached when the solution pH was around the isoelectric point. Electrostatic and hydrophobic force were the main driving force to obtain the protein multilayers.Subsequently, at pH=3, 5 and 7 of gelatin solution, assembled the gelatin multilayers on the modified PLLA surface, compared with the only single gelatin layer on the PLLA surface, the cytocompatiblity was improved greatly. The XPS, water contact angle, and atomic force microscopy data indicated that the layer thickness, surface hydrophilicity, and surface morphology of the gelatin multilayers assembled strongly depended on the pH at which the layers were deposited. All these modified PLLAs exhibited dramatically improved cytocompatibility compared to the virgin PLLA.In order to better understand the effects of protein films to the cell growth behavior, in chapter 4 cowpea mosaic virus (CPMV), a plant virus with higher order structures and many functional groups on its surface was chosen to self-assemble into protein films. The results showed that the surface coverage of CPMV nanoparticles depended on adsorption time and pH of the virus solution, with more amount of CPMV adsorption occurring at near its isoelectric point. In addition, the adhesion and proliferation of HIH-3T3 fibroblasts can be controlled by the coverage of viral particles. The kind of smart transition from cytophobic to cytophilic on the PDDA/CPMV multilayer surface was achievable by simply choosing the outermost material.