Formulation of Different Chitosan Hydrogels for Cartilage Tissue Repair

Abstract

Different formulations of Chitosan/sulphate and Chitosan/PEGDE were produced by physical and chemical reticulation to obtain hydrogels with better physiochemical properties. The hydrogels were analyzed – in terms of their non-toxicity, proliferative, differentiative, inflammatory and immunology responses. Commercial grade Chitosan (Sigma) was solubilized and purified by progressive filtrations. Then, the polymer was freeze-dried in a water soluble cationic form. A physical hydrogel was prepared by mixing a 3 % w/w water solution of the a.m. polymer with different stoichiometric ratios of (SO4=) 1/0.5;1/0.75;1/1 respectively. The hydrogels were cross-linked in multiwells. The chemical hydrogel was synthesized mixing a 3 % w/w water solution of the a.m. polymer, with PEGDE. The hydrogel was purified by water diffusion, then loaded into multiwells. All gels were freeze-dried and sterilized by EtO. Primary bovine chondrocytes were extracted by collagenases (ABIEL) and Pronase (Sigma) from articular hoof tissue. MTS and Acrydin Orange assays were performed to quantify cell viability and apoptosis. CD11b, CD69 and ROS activation were evaluated in inflammatory response and CD11 expression in immune-response. Biocompatibility analyses showed that formulated hydrogels were not toxic and didn’t induce apoptosis. Chondrocyte were alive but not proliferative, inflammation and immune responses were observed but under certain acceptability threshold. The results of this preliminary work suggest that the formulated Chitosan hydrogels are cyto-compatible and induce chondrocyte differentiation. The here tested synthesis process generated hydrogels, which are potentially useful for cartilage regenerative processes. Further mechanical characterizations are in progress.