Fabrication and Characterization of a Resorbable Barrier Membrane Using Silk Fibroin-Fish Collagen Materials

Abstract

Background: A collagen membrane had been the most widely used resorbable barrier membrane in guided bone regeneration, however, a membrane was collapsible and had fast degradation. Silk fibroin had been introduced for improving the properties of a membrane due to its high mechanical strength, low immunogenic responses, and economical advantage. This study aimed to fabricate a resorbable barrier membrane using the composite of silk fibroin and fish collagen materials. Physical and mechanical properties, and in vitro biocompatibilities were evaluated. Materials and Methods: A silk fibroin film was made of Bombyx Mori. silkworm and immobilized with collagen from brown-banded bamboo shark skin by chemical cross-linked. The various ratio of the silk fibroin film was hydrated prior tensile test to select suitable formula before immobilization with collagen. Then, physical and mechanical properties were evaluated to verify the characteristics of a barrier membrane compared with a commercial collagen membrane by using Scanning Electron Microscopy (SEM), Fourier Transform Infrared Spectroscopy (FTIR) and Atomic Force Microscopy (AFM), water contact angles, percentage of elongation, stiffness, swelling degree and degradation degree. In vitro biocompatibility was evaluated in terms of cell attachment and morphology on the membrane, cell viability, cell proliferation and differentiation using MC3T3-E1 cells. The statistical analysis was performed using SPSS software (Version 16.0, SPSS Inc., Chicago, IL, USA). An analysis of variance (ANOVA) was used to compare among experimental groups. The statistical significance was defined as a p-value less than 0.05. Results: The SEM examination demonstrated collagen fibril structure covering silk fibroin-collagen film on the front side and differed from silk fibroin film that showed a non- homogeneous smooth surface on both sides. The AFM demonstrated that silk fibroin-collagen film showed rougher surface and had a higher surface roughness (0.2155 μm) than silk fibroin film (0.1424 um). The FTIR of both silk fibroin film and silk fibroin-collagen film showed the spectrum of peptide bonds of Amide I, II, III and A. The silk fibroin film demonstrated fewer contact angles (76.75° ± 3.07) than silk fibroin-collagen film (112.67° ± 1.94), referred to the more hydrophilic surface. The average percentage of elongation of the silk fibroin film (28.93 ± 15.56) was less than the silk fibroin-collagen film (42.10 ± 11.46) and Bio-GideR collagen membrane (54.79 ± 13.44) but showed no statistical significance among groups (p-value> 0.05). The commercial collagen membrane exhibited significantly higher swelling degree and had slower degradation than other groups (p-value < 0.05). All study groups showed a similar pattern of cell proliferation that continuously increased with time in all groups. The silk fibroin film had significantly highest cell numbers (36,784.17 ± 2,681.73), which was higher than silk fibroin-collagen film (15,905.00 + 2,148.88) and Bio-Gide collagen membrane (12,317.50 ± 1,267.68) and showed statistical significance (p-value < 0.05) on culture Day 7. The result of mineralization on culture Day 21, collagen membrane was higher than other groups. On culture Day 28, collagen membrane (0.78 + 0.19) were in the same level as silk fibroin film (0.77 ± 0.13) and higher than silk fibroin-collagen film (0.56 +0.04) but showed no statistical significance (p-value> 0.05). Conclusion: The in-house silk fibroin film and silk fibroin-collagen film had been developed at an economic cost and possessed physical, mechanical and in vitro biocompatibility properties of a barrier membrane used for GBR. Further study on biocompatibility and the barrier efficacy in vivo should be performed.