Scaffolds are artificial extracellular matrices, capable of supporting cell growth and three-dimensional tissue formation. Along with cells and growth factors, scaffolds are one of the three key components for tissue engineering. The chemical composition and structure of the scaffolds, together with their surface properties, may affect their biocompatibility, limiting their range of clinical applications. Interfacial free energy, surface hydrophilicity, presence of functional groups, density of charges and surface topography are among the characteristics of the scaffolds that affect their biological performance. Hemocompatibility and cytotoxicity are also important aspects of biocompatibility that need to be evaluated prior to scaffold implantation. In this work, different aspects of the biocompatibility of scaffolds constituted of nature-derived polymers were evaluated. In this sense, surface properties as roughness and hydrophilicity, as well as thrombogenicity and in vitro
cytotoxicity were assessed for chitosan-alginate (Ch-A) and chitosan-pectin (Ch-P) scaffolds produced in the presence or not of the porogenic agent KolliphorÂ® P188 (K). Chitosan is a linear polysaccharide obtained by deacetylation of chitin, a polymer vastly found in the exoskeleton of crustaceans, consisting of units of N-acetyl-D-glucosamine and D-glucosamine. At low pH values, chitosan can behave as a polycation and interact with negatively charged polymers, forming polyelectrolyte complexes. Alginate is an anionic linear polysaccharide extracted from brown algae, and its structure is composed of two types of repeating subunits, Î²-(1,4)-D-mannuronate and Î±-(1,4)-L-guluronate. Pectin is an essentially linear anionic polysaccharide extracted from citrus fruits consisting mainly of D-galacturonic acid units joined by Î±-(1-4) glycosidic linkages. The use of alginate in combination with other compounds, such as natural and synthetic polymers, is widespread in the literature for the production of scaffolds for tissue engineering applications. Pectin, on the other hand, is less explored for this use. Combining chitosan with alginate, or pectin can provide matrices with improved properties when compared to those constituted of the isolated polymers, e.g. mechanical properties, pH stability and fluid uptake. Micro-scale roughness was observed for all formulations, with porous samples presenting more irregular surfaces. The surfaces of alginate-containing samples are more hydrophilic than those of the pectin-containing samples and the addition of K increases the hydrophilicity in both cases. For the hemocompatibility assessment, the free hemoglobin test was performed, in which red blood cells not entrapped in a thrombus formed in the sample surface are hemolyzed and free hemoglobin molecules are colorimetrically measured. The scaffolds presented between 68% and 95% of free hemoglobin when compared to the control Teflon®, commonly used in implantable devices. Ch-A formulations were found to be less thrombogenic than Ch-P, what can be attributed to their surface wettability, as the adsorption of blood proteins would be less intense over their more hydrophilic surface. In vitro
direct cytotoxicity of scaffolds to fibroblasts varied from 15% and 30%. No trend for the influence of the addition of K to the scaffolds was observed on cytotoxicity. Considering the studied properties of the materials, all formulations presented satisfactory performance and each of them may be directed to particular applications in tissue engineering. Complementary studies on other aspects of the biomaterials biocompatibility should be performed to assure the feasibility of the use of these devices as implantable materials.
The authors are grateful for the finantial support of this work by the Coordination for the Improvement of Higher Education Personnel (CAPES) and the São Paulo Research Foundation (FAPESP), both from Brazil, and from the Natural Sciences and Engineering Research Council (NSERC) of Canada.