Biofunctional Ionic-Doped Calcium Phosphates: Silk Fibroin Composites for Bone Tissue Engineering Scaffolding

Abstract

The treatment and regeneration of bone defects caused by traumatism or diseases have not been completely addressed by current therapies. Lately, advanced tools and technologies have been successfully developed for bone tissue regeneration. Functional scaffolding materials such as biopolymers and bioresorbable fillers have gained particular attention, owing to their ability to promote cell adhesion, proliferation, and extracellular matrix production, which promote new bone growth. Here, we present novel biofunctional scaffolds for bone regeneration composed of silk fibroin (SF) and β-tricalcium phosphate (β-TCP) and incorporating Sr, Zn, and Mn, which were successfully developed using salt-leaching followed by a freeze-drying technique. The scaffolds presented a suitable pore size, porosity, and high interconnectivity, adequate for promoting cell attachment and proliferation. The degradation behavior and compressive mechanical strengths showed that SF/ionic-doped TCP scaffolds exhibit improved characteristics for bone tissue engineering when compared with SF scaffolds alone. The in vitro bioactivity assays using a simulated body fluid showed the growth of an apatite layer. Furthermore, in vitro assays using human adipose-derived stem cells presented different effects on cell proliferation/differentiation when varying the doping agents in the biofunctional scaffolds. The incorporation of Zn into the scaffolds led to improved proliferation, while the Sr- and Mn-doped scaffolds presented higher osteogenic potential as demonstrated by DNA quantification and alkaline phosphatase activity. The combination of Sr with Zn led to an influence on cell proliferation and osteogenesis when compared with single ions. Our results indicate that biofunctional ionic-doped composite scaffolds are good candidates for further in vivo studies on bone tissue regeneration.

In Vivo Performance of Hierarchical HRP-Crosslinked Silk Fibroin/β-TCP Scaffolds for Osteochondral Tissue Regeneration

“…The good outcomes of the ionic presence into the hierarchical scaffolds were confirmed by the similar porosity ( Figure 2B), trabecular thickness ( Figure 2C), and pore size distribution ( Figure 2D) of the BdTCP scaffolds to that obtained for the healthy controls. Moreover, in the subchondral bone region the BdTCP scaffolds were filled by hard and soft tissue at similar levels ( Figure 3A), which indicates the strong impact of Zn and Sr elements on bone formation and mineralization, by means of improving the microstructure and crystallinity of β-TCP [34,46].…”

“…We hypothesize that the spatially limited incorporation of ZnSr-βTCP bioresorbable powder will affect the new bone ingrowth's and mineralization, in comparison to the same structures with no ion incorporation. precipitation, followed by milling and sieving, with an average particle size of 1-10 µm, as previously described [34]. The ZnSr-β-TCP and β-TCP powders were mixed with the HRP-SF solution and transferred into cylindrical silicone molds (9 mm inner diameter; Deltalab, Barcelona, Spain), followed by adding 2 g of granular sodium chloride low in endotoxins (EMPROVE, VWR BDH Prolabo, Briare, France) sieved at 500-1000 μm range (Analytic Sieve Shaker; AS 200 Digit, Retsch, Germany) ( Figure 1A).…”

Hierarchical HRP-Crosslinked Silk Fibroin/ZnSr-TCP Scaffolds for Osteochondral Tissue Regeneration: Assessment of the Mechanical and Antibacterial Properties

“…In order to address the OC regeneration challenges, SF combined with ZnSr-doped β-TCP scaffolds has been recently reported (Pina et al, 2017;Ribeiro et al, 2019b). As a first approach, SF were mixed with β-TCP incorporating Sr, Zn, and Mn, to produce biofunctional scaffolds with adequate microand macro-porosity for bone tissue regeneration (Pina et al, 2017). In vitro assays using human adipose-derived stem cells (hASCs) demonstrated different cellular responses by varying the ionic doping elements in the scaffolds, namely when combined with Sr and Zn in comparison to the single ions.…”

Functionalization of Silk Fibers by PDGF and Bioceramics for Bone Tissue Regeneration

“…Several studies have successfully used silk scaffolds for bone tissue engineering applications. Besides pure silk scaffolds [10][11][12][13][14], the combination of silk fibroin and ceramic materials such as β-tricalcium phosphate (β-TCP) [15,16] or hydroxyapatite (HA) [17][18][19][20] have been investigated extensively. HA represents the largest part of the inorganic component of the human bone [21] and has an osteoconductive effect [22].…”