369Fabrication of 3D gel-printed β-tricalcium phosphate/titanium dioxide porous scaffolds for cancellous bone tissue engineering

Abstract

Human bone is composed of cortical bone and cancellous bone. The interior portion of natural bone is cancellous with a porosity of 50%-90%, but the outer layer is made of dense cortical bone, of which porosity was not higher than 10%. Porous ceramics were expected to be research hotspot in bone tissue engineering by virtue of their similarity to the mineral constituent and physiological structure of human bone. However, it is challenging to utilize conventional manufacturing methods to fabricate porous structures with precise shapes and pore sizes. Three-dimensional (3D) printing of ceramics is currently the latest research trend because it has many advantages in the fabrication of porous scaffolds, which can meet the requirements of cancellous bone strength, arbitrarily complex shapes, and individualized design. In this study, β-tricalcium phosphate (β-TCP)/titanium dioxide (TiO₂) porous ceramics scaffolds were fabricated by 3D gel-printing sintering for the first time. The chemical constituent, microstructure, and mechanical properties of the 3D-printed scaffolds were characterized. After sintering, a uniform porous structure with appropriate porosity and pore sizes was observed. Besides, biological mineralization activity and biocompatibility were evaluated by in vitro cell assay. The results demonstrated that the incorporation of TiO₂ (5 wt%) significantly improved the compressive strength of the scaffolds, with an increase of 283%. Additionally, the in vitro results showed that the β-TCP/TiO₂ scaffold had no toxicity. Meanwhile, the adhesion and proliferation of MC3T3-E1 cells on scaffolds were desirable, revealing that the β-TCP/TiO₂ scaffolds can be used as a promising candidate for repair scaffolding in orthopedics and traumatology.

Keywords: 3D printing; Bone tissue engineering; Porous scaffolds; Titanium dioxide; β-tricalcium phosphate.

Figure 1

(A) Ceramic samples of β-TCP (a), β-TCP/1-TiO₂ (b), β-TCP/3-TiO₂ (c), and β-TCP/5-TiO₂ (d). (B) XRD diagram of TiO₂, β-TCP, and β-TCP/5- TiO₂ ceramic and details of the XRD diagram of the figure at 24.5°-26.5°. (C) Infrared spectrum of TiO₂, β-TCP, and β-TCP/5-TiO₂ ceramic. (D) Micro-morphology of β-TCP/TiO₂ ceramics detected by SEmml: β-TCP/TiO₂ (a), β-TCP/1-TiO₂ (b), β-TCP/3-TiO₂ (c), and β-TCP/5-TiO₂ (d). (E) The shrinkage of β-TCP/TiO₂ ceramic samples with 0% TiO₂, 1% TiO₂, 3% TiO₂ and 5% TiO₂. (F) Average diameter of micropores of β-TCP and β-TCP/TiO₂ ceramics. (G) The compressive strength of β-TCP and β-TCP/TiO₂ceramics with different TiO₂ content.

Figure 2

Printability in the fabrication of ß-TCP/TiO₂ scaffolds

Figure 3

(A) Samples of β-TCP/3-TiO₂ ceramics scaffolds in different filling rates (al and a2: stents with a filling rate of 40%; bl and b2: stents with a filling rate of 30%; cl and c2: stents with a filling rate of 20%). (B) Effect of different filling rates on the shrinkage of β-TCP/3-TiO₂ scaffolds. (C) Effect of different filling rates on the porosity of β-TCP/3-TiO₂ scaffolds. (D) Average macropore diameter of β-TCP/3-TiO₂ ceramics scaffolds in different filling rate. (E) Effects of different filling rates on the compressive strength of β-TCP/3-TiO₂ceramics scaffolds.

Figure 4

(A) Appearance of β-TCP/TiO₂ ceramics scaffolds with different TiO₂ content. (B) Mean micropores diameter of β-TCP and β-TCP/TiO₂ ceramics scaffolds. (C) The compressive strength of β-TCP and β-TCP/TÌO2 ceramics scaffolds with different TiO₂ content. (D) Effect of different TiO₂ content on the porosity of β-TCP/ TiO₂ ceramics scaffolds with a filling rate of 30%. (E) Effect of different TiO₂ content on the shrinkage of β-TCP/TiO₂ ceramics.

Figure 5

Micromorphology of β-TCP/TiO₂ ceramics scaffolds detected by SEM. (A) Outward surface. (B) Fracture surface. Groups a, b, c, and d are ceramic scaffolds with 0% TiO₂, 1% TiO₂, 3% TiO₂, and 5% TiO₂ components, respectively. Groups al, bl, cl, and dl are visualized by SEM with 300× magnification; scale bar: 200 μm. Groups a2, b2, c2 and d2 are visualized by SEM with 600× magnification; scale bar: 100 |im. Groups a3, b3, c3, and d3 are visualized by SEM with 6000× magnification; scale bar: 10 μm.

Figure 6

Scanning energy spectrum of β-TCP/TiO₂ ceramic scaffolds after 28 days (A) and 14 days (B) of biomineralization. Groups a, b, c, and d show the mineralization of ceramic scaffolds with 0% TiO₂, 1% TiO₂, 3% TiO₂, and 5% TiO₂ components, respectively. Groups a1, b1, c1, and d1 are visualized by SEM with 75× magnification; scale bar: 200 μm. Groups a2, bl, c2, and d2 are visualized by SEM with 300× magnification; scale bar: 50 μm. Groups a3, b3, c3, and d3 are visualized by SEM with 1500× magnification; scale bar: 10 μm.

Figure 7

(A) The MTT assay results showing cell viability rate of MC3T3-E1 osteoblasts cultured on β-TCP/TiO₂ ceramic scaffolds with different content of TiO₂ (CK was the blank control group). (B) Osteoblast fluorescence detection of MC3T3-E1 cells on β-TCP/TiO₂ ceramic scaffolds for 1, 3, and 7 days. (C) Osteoblast fluorescence detection of dead MC3T3-E1 cells on β-TCP/TiO₂ ceramic scaffolds for 1, 3, and 7 days. (D) Corresponding osteoblast fluorescence detection of living/dead cells of MC3T3-E1 cells on β-TCP/TiO₂ ceramic scaffolds after 7 days of incubation. (E) Cell morphologies of MC3T3-E1 osteoblast cells cultured on β-TCP/TiO₂ ceramic scaffolds: β-TCP/TiO₂ (a), β-TCP/1-TiO₂ (b), β-TCP/3-TiO₂ (c), and β-TCP/5-TiO₂(d).