Bone tissue engineering with premineralized silk scaffolds

Abstract

Silks fibroin biomaterials are being explored as novel protein-based systems for cell and tissue culture. In the present study, biomimetic growth of calcium phosphate on porous silk fibroin polymeric scaffolds was explored to generate organic/inorganic composites as scaffolds for bone tissue engineering. Aqueous-derived silk fibroin scaffolds were prepared with the addition of polyaspartic acid during processing, followed by the controlled deposition of calcium phosphate by exposure to CaCl 2 and Na 2 HPO 4 . These mineralized protein-composite scaffolds were subsequently seeded with human bone marrow stem cells (hMSC) and cultured in vitro for 6 weeks under osteogenic conditions with or without BMP-2. The extent of osteoconductivity was assessed by cell numbers, alkaline phosphatase and calcium deposition, along with immunohistochemistry for bone related outcomes. The results suggest increased osteoconductive outcomes with an increase in initial content of apatite and BMP-2 in the silk fibroin porous scaffolds. The premineralization of these highly porous silk fibroin protein scaffolds provided enhanced outcomes for the bone tissue engineering.

Effect of freezing methods on the properties of lyophilized porous silk fibroin membranes

“…Figure 5 exhibits the SEM micrographs of surface and cross-sectional areas of the membranes. Both samples presented similar aspects of rough surface and porous cross-sectional area, important characteristics observed in biomaterials suitable for tissue engineering 22 . Comparing the results from XRD, FTIR, TGA and DSC analyses of wet and frozen/lyophilized membranes, the main difference observed was in the presence of water.…”

Elucidation of differential mineralisation on native and regenerated silk matrices

“…The peaks became more intense upon pre-treatment with CaCl 2 , indicating enhanced nucleation and deposition of HA ( Figure 6) in these groups. The broad halo at 2è of 20.2° with a shoulder peak located at 24.6°, attributed to the â-sheet crystalline structure (silk II) of silk fibroin, which was detected in all the samples [13]. Apart from the peaks corresponding to silk fibroin protein, no other peak was detected in the case of 2D planar silk films in both treatment conditions, suggesting no traces of apatite deposition on films.…”

“…XRD pattern of both lyophilised matrices and braids exhibited characteristic peaks at 20.9° and 24.6° indicating silk II conformation [13]. Broadening of the peaks was observed in all the cases indicating poor crystallinity.…”

“…Silk fibroin, another fibrous protein that closely mimics collagen type I of bone has gained popularity in tissue engineering due to interesting intrinsic properties such as resorbability [5], toughness [6], minimal immunogenicity [7,8], cytocompatibility with abundant polar, hydrophilic groups [9], strong chemical bonding with HA [10][11][12][13] and versatility in processing depending upon the target application.…”

“…Also, the reason as to why mineralisation could not be observed on methanol treated pure fibroin films (without calcium chloride) which contained at least some amount of â-sheet content remained unexplained. Nevertheless, most studies have reported an additional requirement of either pre-exposure of fibroin or its association with calcium-and phosphate-based solutions to promote nucleation and growth of HA nanocrystals [12,13,16,22]. Whether it is the processing parameters during dissolution and reconstitution that disrupt the native structure of the fibroin chain; firstly by dissociating the specific amino acid chains and secondly by hampering their tendency to re-assemble the necessary â-sheet conformation, or if it is the inherent inert nature of fibroin chain that modulates the extent of mineralisation is still under wraps.…”