Infiltration of silica inside fibrillar collagen.

Diatom frustules (the hard and porous cell wall of diatoms) are created under the control of biomolecules (silaffins, silacidins, and long-chain polyamines) at close to physiologic conditions. The mechanism of biosilica formation was traditionally based on the ability of zwitterionic water-soluble proteins to create macromolecular assemblies for silica polymerization. Recent discoveries of water-insoluble collagen matrices within certain sponge biosilica spicules, chitin-based scaffolds in sponge and diatom biosilica formations, as well as cingulins within diatom girdle bands, revive the use of insoluble biomimetic organic templates for morphogenesis of nonporous silica structures. The use of fibrillar collagen as templates for biosilica synthesis was unsuccessful in the past as only extrafibrillar silica deposition was observed. Intrafibrillar mineralization of collagen has important implications from a biophysical perspective. Herein, we report a collagen biosilicification scheme based on the fusion of stabilized polysilicic acid into a fluidic precursor phase upon their infiltration into polyamine-enriched collagen. The latter serves as a template and catalyst for polymerization of the precursor phase into silica that faithfully reproduces the collagen tertiary architecture. Our findings provide a new concept in biosilica materials synthesis, which does not require phosphate supplements. Type I collagen has been widely used for hybrid biomaterials synthesis because of its biocompatibility. Nevertheless, the lack of mechanical resistance of highly porous, nonmineralized collagen matrices limits their application as stress-bearing scaffolds for bone repair. The potential stimulating effect of silicic acid on osteogenesis provides the incentive to produce silicified collagen matrices for bone regeneration applications. Previous studies have explored methods to produce silica–collagen hybrid materials by simultaneous self-assembly of collagen and silica. However, the kinetics of collagen self-assembly and silica polymerization are not fully compatible. Other studies that utilized fibrillar collagen as templates produced silica particles in close vicinity of the collagen fibrils but not within the fibrils, which limits the strength of the biohybrid materials. Silica– collagen hybrids may also be potentially used for preparation of nonbiological functional materials; however, the fundamental prerequisite is that collagen fibrils should be fully infiltrated with a high intrafibrillar silica content. During diatom cell wall synthesis, silicic acid is stabilized and prevented from spontaneous polymerization into silica. Silicic acid gels rapidly in vitro at pH 5.5 without stabilization (see the Supporting Information S2, Figure S2A and S3, Figure S3A). Thus, we used choline chloride to stabilize silicic acid hydrolyzed from tetraethyl orthosilicate. Based on the assessment of gelling times, 72 mm choline chloride is required to stabilize 1.5% silicic acid (pH 5.5) for 72 h before gelling (see the Supporting Information S2, Figure S2A). This method provided sufficient time for the choline-stabilized silicic acid (Ch–SA) to infiltrate type I collagen sponges. The mean particle diameter of Ch–SA was approximately 9.7 nm based on dynamic light scattering (DLS) measurements (see the Supporting Information S2, Figure S2B and S2C). A series of control experiments was initially performed using unstained sections prepared for transmission electron microscopy (TEM) to determine the optimal conditions for silicification of collagen sponges with Ch–SA (see the Supporting Information 3, Figure S3A-E). By embracing the model of cingulin proteins as water-insoluble templates in the silicification of diatom girdle bands, successful intrafibrillar silicification of collagen was achieved in 4 days after immersing polyallylamine(PAH)-enriched collagen sponges [*] Dr. D. H. Pashley, Dr. F. R. Tay School of Graduate Studies, Georgia Health Sciences University Augusta, Georgia, 30912-1129 (USA) E-mail: [email protected] Dr. L.-n. Niu, Dr. K. Jiao, Dr. J.-h. Chen School of Stomatology, Fourth Military Medical University Xi’an, 710032 (PR China) E-mail: [email protected]