Structural and biological evaluation of new chitosan membrane for dural closure
Keywords:dura mater substitute, chitin, chitosan, degradation, mechanical properties, cell culture
Objective: to study the structural and biological evaluation of chitin-chitosan based membrane for dura mater replacement.
Materials and methods. Chitosan-based films were made out of 3% solution of chitosan for the research. We used 200, 500 and 700 kDa chitosan (deacetylation rate 80–90 %) to produce chitin-chitosan membrane by using solvent evaporation method. For enhancing mechanical properties and reducing the degradation, chitin particles were added to the chitosan solution. Chitosan and chitin ratio was 80/20. The chitin/chitosan solution in Petri dishes was dried out during 3 days at room temperature.
To obtain information about the structure of membrane surface and cross-section scanning, electron microscopy was performed.
Hydrolytic degradation was studied by pouring into SBF solution. To determine the rate of enzymatic degradation, trypsin solution was used. To determine the mass loss percentage, we measured the sample weight after 7, 14, 21, 30 and 60 days after being in the appropriate solutions.
Relative elongation and strength were measured by digital dynamometer to study membranes mechanical properties such as the strength and elasticity. MLO-A5 cells were used to assess biocompatibility of new materials.
Results. Macroscopic view of obtained samples has shown their relative transparency with impregnation of chitin particle that elevated over the membrane surface without any diversity between different chitosan molecular weight samples.
Due to scanning electron microscopy, principal diversity between the samples of different molecular weight has being seen: rough pore surface at 200 and 500 kDa and flat with minimal roughness surface of 700 kDa membranes. Cross-section of 500 and 700 kDa membranes are dense with no pores, but 200 kDa membrane are sponge like and it can be prediction for fluid sorption and cell migration during healing process.
Chitin-chitosan membranes are biocompatible and degrade in aqueous and enzymatic solutions. Due to polysaccharide nature of chitosan and chitin, enzymatic degradation has shown higher trend compare to the hydrolytic ones. 200 kDa membrane degrades faster with final mass loss 83.2 % and completely due to porous structure that allows fluid sorption.
Membrane mechanical parameters strongly depend on their structure. 200 kDa membrane has shown 2-fold higher elongation compared to 500 kDa and 3-fold — compared to 700 kDa ones. The compensation of mechanical forces ensured by porous structure is better than in dense ones. Tensile strength was in 2-fold better in 200 kDa membranes than in 500 and 700 kDa ones.
Cell culture experiment has shown the better adhesion at the 3rd day for 200 kDa membrane and minimal cell adhesion for 700 kDa membrane, probably due to smooth surface. The reduction rate between all samples and PCT control differ a lot, except for 200 kDa membrane that has the same proliferation rate as TCP.
Conclusion. Chitin-chitosan membranes, made from different molecular weight chitosan, are transparent and has appropriate structure for being used as a dura mater substitute. They are biocompatible and degrade in aqueous and enzymatic solutions. Due to porous structure, excellent mechanical properties as well as better cell adhesion and proliferation, 200 kDa chitosan membrane is more applicable for neurosurgical issues.
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