Processing of Biopolymers-based, Efficiently Integrated Bilayer Membranes for Use in Guided Tissue Regeneration Procedures

Abstract

Periodontitis is a chronic inflammatory disease affecting the tooth supporting structures - the alveolar bone in particular, to extent of eventual tooth loss. Among others, the guided tissue regeneration (GTR) procedure including placement of occlusive barrier membrane is very promising approach. Herein we develop chitosan (CHT) / gelatin (GEL) bilayer membranes via successive solvent- and freeze-casting procedures and genipin (GEN)-mediated cross-linking chemistry. By utilizing the auto-fluorescence signal from GEN cross-linking products (i.e. the secondary CHT (GEL) amines and GEN esters), the Confocal Fluorescent Microscopy (CFM) identifies the chemical as well as physical integration between layers' interfaces, supported also by Scanning Electron Microscopy (SEM) data. The presence of non- and highly μ-porous and pore-interconnecting regions is demonstrated within cross-sections of GEL-prevalent membranes in contrast to the sheet-like organization in membrane with equal components presence. The constant processing conditions onto variable compositions did not significantly affect the pore size distributions (in 1-230 μm range), while pore wall thickness increases up to 220 μm with GEL increase, which also improves the yield stress at compression from 10 to 19 kPa and elastic modulus from 26 to 34 kPa. The further applied rapid mineralization resulted in deposition of non-regular to spherical minerals, containing nonstoichiometric carbonated apatite with Ca/P ration in 1.7-2 range, which demonstrates formation of osseointegrative interface. The intensive, composition-dependent swelling (up to 580%), as well as 67% to 100% weigh loss in 4 weeks in vitro degradation experiment imply on bilayer membranes GTR relevance.