(436c) Transition Metal-Based Tyrosine Crosslinking of ELP Hydrogels for Tissue Engineering Applications.

Elastin like polypeptides (ELPs) are genetically modifiable synthetic polymers that are unique because of their lower critical solution temperature (LCST) properties and biocompatibility. Crosslinking of linear ELPs of general form (VPGXG)_n with guest residue ‘X’, results in gelation to form hydrogels. Gelation of these ELPs aids in designing biocompatible and genetically modifiable biomaterials that includes ability to match the mechanical properties of target tissues, in addition to providing biophysical and biochemical cues that direct cell responses and physiological development. Tyrosine based crosslinking in the synthesis of ELP hydrogels can result in biomaterials that have higher level of mechanical and structural characteristics compared to use of other residues for crosslinking. Dityrosine crosslinks possess mechanical properties such as elastic and Young’s moduli that can be increased by controlling the number of dityrosine crosslinks as shown by studies with recombinant resilin and silk proteins. Formation of di-tyrosine bonds with enzymes is temperature and pH sensitive and results in inefficient crosslinking with crosslinks that are not very stable. But on the other hand, transition metal based crosslinkers have resulted in mechanically and structurally stable covalent bonds, which are easy to form and are stable. Utilizing metal crosslinkers as catalysts for tyrosine polymerization of ELPs can make biocompatibility of formed ELP hydrogels questionable. In this work, we evaluated chemical reactions using transition metals to crosslink ELPs with tyrosine residues [(VPGYG)₃₆] and also developed a continuous cleaning process to demonstrate the ability of removing the trapped transition metals. Ruthenium complex [Ru (II)bpy₃ ²⁺] with initiator such as sodium per sulphate (SPS) in the presence of blue light was successful in forming ELP hydrogels. One factor at a time (OFAT) trails were performed to optimize the hydrogel preparation conditions. Optimized hydrogels were tested for their biological, mechanical, and structural characteristics. Washing the hydrogels improved the in-vitro biocompatibility of the hydrogels. The effect of pH on the washing solvent and the mode of washing (batch vs continuous) were analyzed. The mechanical and structural properties on as prepared and washed hydrogels were found to be equivalent. In- vitro biocompatibility of the washing method was shown to be effective and it was found that the hydrogels post-washing can be used directly in experimental studies as a biomaterial for various tissue engineering applications. The method of washing and rigorous testing shown in this work can be utilized as a protocol for improving and establishing biocompatibility of metal catalyzed hydrogels.