The surface of solvent cast chitosan membranes was modified using a two-step procedure. Oxygen plasma treatment was used at the first activation step followed by vinyl monomer graft polymerization. Two monomers were used in order to compare the influence of different functional groups on cell adhesion and proliferation; acrylic acid (AA) was used to introduce carboxyl groups and vinyl sulfonic acid (VSA) was used as a source of sulfonic groups. The surface chemistry/energy changes were characterized by means of X-ray photoelectron spectroscopy (XPS), Fourier transform infrared spectroscopy (FTIR-ATR), and contact angle measurements. Additionally, alterations in the surface morphology were investigated by scanning electron microscopy (SEM). XPS analyses confirmed the polymer grafting on the surface; an S 2s peak appears in the VSA survey spectrum and an O-CLO peak emerges in the C 1s high resolution spectrum after AA grafting. Moreover, contact angle measurements showed an increment in the values of the surface energy polar and Lewis base components for all treated samples, confirming the introduction of additional polar groups by the modification processes. FTIR-ATR spectra showed no significant difference between treated and original materials. These results confirmed that only the very top (a few angstroms) surface layer, but not the bulk of the material, was modified. The effect of modification on the adhesion and proliferation of osteoblast-like cells was studied on a preliminary basis. Direct contact tests were performed using a human osteosarcoma cell line (SaOs-2). Cell morphology (optical microscopy and SEM) and cell viability (MTS test) were evaluated for untreated and surface modified membranes. The results revealed that both plasma treatment, and the presence of sulfonic groups on the surface of chitosan membranes, improve SaOs-2 adhesion and proliferation when compared to untreated or AA-grafted membranes. This effect was strongly related to the polar and Lewis basic components of the total surface energy.
Chitosan biocompatibility is often associated with the structural similarities with glycosaminoglycans (GAGs). Although all of the GAGs are built from repeating disaccharide units and some of them contain N-glucosamine (the main hexosamine in the chitosan backbone), all of them also contain negatively charged functional groups. These charged units are believed to have a crucial role for the formation of proteoglycans and hence for key biochemical processes/signaling related to cell functionality and survival. Lack of these groups in chitosan structure could be the reason for the previously observed poor cell adhesion to this material. Herein, we report that plasma induced grafting of negatively charged phosphonic groups can induce remarkably distinguishable cell response and significantly improve the adhesion, proliferation and viability of osteoblast cells. The proposed plasma induced polymerization is a very simple and versatile method and can be easily adapted to other materials and different negatively charged units.
A commonly applied strategy in the field of tissue engineering (TE) is the use of temporary three-dimensional scaffolds for supporting and guiding tissue formation in various in vitro strategies and in vivo regeneration approaches. The interactions of these scaffolds with highly sensitive bioentities such as living cells and tissues primarily occur through the material surface. Hence, surface chemistry and topological features have principal roles in coordinating biological events at the molecular, cellular and tissue levels on timescales ranging from seconds to weeks. However, tailoring the surface properties of scaffolds with a complex shape and architecture remains a challenge in materials science. Commonly applied wet chemical treatments often involve the use of toxic solvents whose oddments in the construct could be fatal in the subsequent application. Aiming to shorten the culture time in vitro (i.e. prior the implantation of the construct), in this work we propose a modification of previously described bone TE scaffolds made from a blend of starch with polycaprolactone (SPCL). The modification method involves surface grafting of sulfonic or phosphonic groups via plasma-induced polymerization of vinyl sulfonic and vinyl phosphonic acid, respectively. We demonstrate herein that the presence of these anionic functional groups can modulate cell adhesion mediated through the adsorbed proteins (from the culture medium). Under the conditions studied, both vitronectin adsorption and osteoblast proliferation and viability increased in the order SPCL << sulfonic-grafted SPCL < phosphonic-grafted SPCL. The results revealed that plasma-induced polymerization is an excellent alternative route, when compared to the commonly used wet chemical treatments, for the surface functionalization of biodevices with complex shape and porosity.
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