Hypoxia-mimicking 3D bioglass-nanoclay scaffolds promote endogenous bone regeneration

Abstract

Large bone defect repair requires biomaterials that promote angiogenesis and osteogenesis. In present work, a nanoclay (Laponite, XLS)-functionalized 3D bioglass (BG) scaffold with hypoxia mimicking property was prepared by foam replication coupled with UV photopolymerization methods. Our data revealed that the incorporation of XLS can significantly promote the mechanical property of the scaffold and the osteogenic differentiation of human adipose mesenchymal stem cells (ADSCs) compared to the properties of the neat BG scaffold. Desferoxamine, a hypoxia mimicking agent, encourages bone regeneration via activating hypoxia-inducible factor-1 alpha (HIF-1α)-mediated angiogenesis. GelMA-DFO immobilization onto BG-XLS scaffold achieved sustained DFO release and inhibited DFO degradation. Furthermore, in vitro data demonstrated increased HIF-1α and vascular endothelial growth factor (VEGF) expressions on human adipose mesenchymal stem cells (ADSCs). Moreover, BG-XLS/GelMA-DFO scaffolds also significantly promoted the osteogenic differentiation of ADSCs. Most importantly, our in vivo data indicated BG-XLS/GelMA-DFO scaffolds strongly increased bone healing in a critical-sized mouse cranial bone defect model. Therefore, we developed a novel BG-XLS/GelMA-DFO scaffold which can not only induce the expression of VEGF, but also promote osteogenic differentiation of ADSCs to promote endogenous bone regeneration.

Keywords: 3D bioglass scaffold; Angiogenesis; Endogenous bone regeneration; Hypoxia; Osteogenesis.

Graphical abstract

Fig. 1

ALP staining of ADSCs after treated with BG extracts and sintered XLS extracts (left panel), ALP activities of ADSCs after treated with BG extracts and sintered XLS extracts (right panel).

Fig. 2

SEM images of (a, e) BG scaffold, (b, f) BG-XLS1 scaffold, (c, g) BG-XLS3 scaffold, (d, h) BG-XLS5 scaffold; (i, j) Porosity and shrinkage of BG, BG-XLS1, BG-XLS3 and BG-XLS5 scaffolds, respectively; (k) Compressive strength of BG, BG-XLS1, BG-XLS3 and BG-XLS5 scaffolds. Data are expressed as mean ± SD (n = 3). (*P < 0.05, **P < 0.05).

Fig. 3

SEM images (a, b) BG-XLS3 and (d, e) BG-XLS3/GelMA; EDS spectra of (c) BG-XLS3 and (f) BG-XLS3/GelMA scaffolds; (g) Compressive strength of BG-XLS3 and BG-XLS3/GelMA scaffolds; (h) In vitro weight loss of BG-XLS and BG-XLS/GelMA scaffolds after immersion in PBS solution for 14 days.

Fig. 4

SEM images and EDS spectra of (a, d, g) BG scaffolds, (b, e, h) BG-XLS1 scaffolds and (c, f, i) BG-XLS3 scaffolds after 1 d of SBF immersion; (j) FTIR spectra and (k) XRD spectra of BG-XLS3 scaffolds and BG-XLS3 scaffolds after immersed in SBF for 1 d.

Fig. 5

(a) Relative cell viabilities of MC3T3-E1 cells cultured on BG, BG-XLS1 and BG-XLS3 scaffolds after 1 and 4 days; MC3T3-E1 cells morphologies on (b, e) BG, (c, f) BG-XLS1 and (d, g) BG-XLS3 scaffolds after 1 and 4 days, respectively. Data are expressed as mean ± SD (n = 3). (*P < 0.05).

Fig. 6

Effects of BG, BG-XLS1 and BG-XLS3 scaffolds extracts on calcium contents of ADSCs after 21 days of culture. Data are expressed as mean ± SD (n = 3). (**P < 0.01, ***P < 0.001).

Fig. 7

DFO released from BG-XLS-DFO and BG-XLS/GelMA-DFO scaffolds.

Fig. 8

HIF-1α and VEGF expressions in ADSCs were measured after the cells were cultured in different culture medium for 24 h and 48 h. Data are expressed as mean ± SD (n = 3). (*P < 0.05, **P < 0.01, ***P < 0.001).

Fig. 9

Relative cell viabilities of ADSCs cultured on control, DFO (100 μM), BG-XLS scaffolds and BG-XLS/GelMA-DFO scaffolds for 1 and 3 days.

Fig. 10

ALP activity of ADSCs on C, DFO, BG-XLS scaffolds and BG-XLS/GelMA-DFO scaffolds after 7 days of culture. Calcium content of ADSCs on C, DFO, BG-XLS scaffolds and BG-XLS/GelMA-DFO scaffolds after 21 days of culture. *P < 0.05, **P < 0.01(n = 3).

Fig. 11

RUNX2 and OCN expressions after 7 days of culture. *P < 0.05, **P < 0.01 (n = 3).

Fig. 12

Micro-CT images of: (a) BG-XLS, (b) BG-XLS/GelMA-DFO, (c) BG-XLS-BMP2 scaffolds at 8 weeks post-operation; (d) BV/TV and (e) BMD values for corresponding groups at 8 weeks post-operation; H&E staining of: (f, i) BG-XLS, (g, j) BG-XLS/GelMA-DFO, (h, k) BG-XLS-BMP2 scaffolds at 8 weeks post-operation. *P < 0.05, **P < 0.01 (n = 3–4).