title: Enzymatic cross-linking of human recombinant elastin (HELP) as biomimetic approach in vascular tissue engineering.
abstract: The use of polymers naturally occurring in the extracellular matrix (ECM) is a promising strategy in regenerative medicine. If compared to natural ECM proteins, proteins obtained by recombinant DNA technology have intrinsic advantages including reproducible macromolecular composition, sequence and molecular mass, and overcoming the potential pathogens transmission related to polymers of animal origin. Among ECM-mimicking materials, the family of recombinant elastin-like polymers is proposed for drug delivery applications and for the repair of damaged elastic tissues. This work aims to evaluate the potentiality of a recombinant human elastin-like polypeptide (HELP) as a base material of cross-linked matrices for regenerative medicine. The cross-linking of HELP was accomplished by the insertion of cross-linking sites, glutamine and lysine, in the recombinant polymer and generating ε-(γ-glutamyl) lysine links through the enzyme transglutaminase. The cross-linking efficacy was estimated by infrared spectroscopy. Freeze-dried cross-linked matrices showed swelling ratios in deionized water (≈2500%) with good structural stability up to 24 h. Mechanical compression tests, performed at 37°C in wet conditions, in a frequency sweep mode, indicated a storage modulus of 2/3 kPa, with no significant changes when increasing number of cycles or frequency. These results demonstrate the possibility to obtain mechanically resistant hydrogels via enzymatic crosslinking of HELP. Cytotoxicity tests of cross-linked HELP were performed with human umbilical vein endothelial cells, by use of transwell filter chambers for 1-7 days, or with its extracts in the opportune culture medium for 24 h. In both cases no cytotoxic effects were observed in comparison with the control cultures. On the whole, the results suggest the potentiality of this genetically engineered HELP for regenerative medicine applications, particularly for vascular tissue regeneration.
authors: Bioengineering Department, Biomaterials Laboratory, Politecnico di Milano, Milan, Italy. sabrina.bozzini@polimi.it
authors: Sabrina
authors: S
authors: Bozzini
authors: Liliana
authors: L
authors: Giuliano
authors: Lina
authors: L
authors: Altomare
authors: Paola
authors: P
authors: Petrini
authors: Antonella
authors: A
authors: Bandiera
authors: Maria Teresa
authors: MT
authors: Conconi
authors: Silvia
authors: S
authors: Farè
authors: Maria Cristina
authors: MC
authors: Tanzi
mesh_terms: D000595:Amino Acid Sequence; D001672:Biocompatible Materials / Q000737:chemistry; D032701:Biomimetics / Q000379:methods*; D001808:Blood Vessels / Q000473:pathology*; D002478:Cells, Cultured; D019245:Compressive Strength; D003432:Cross-Linking Reagents / Q000737:chemistry; D004337:Drug Carriers; D016503:Drug Delivery Systems; D004549:Elastin / Q000737:chemistry*; D005973:Glutamine / Q000737:chemistry; D061307:Human Umbilical Vein Endothelial Cells; D006801:Humans; D020100:Hydrogels / Q000737:chemistry; D008239:Lysine / Q000737:chemistry; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D011108:Polymers / Q000737:chemistry; D011312:Pressure; D011994:Recombinant Proteins / Q000737:chemistry*; D044968:Regenerative Medicine / Q000379:methods; D013055:Spectrophotometry, Infrared; D013696:Temperature; D023822:Tissue Engineering / Q000379:methods*; D011503:Transglutaminases / Q000378:metabolism
chemical_list: D001672:Biocompatible Materials; D003432:Cross-Linking Reagents; D004337:Drug Carriers; D020100:Hydrogels; D046911:Macromolecular Substances; D011108:Polymers; D011994:Recombinant Proteins; D005973:Glutamine; D004549:Elastin; D011503:Transglutaminases; D008239:Lysine