Chitosan/POSS Hybrid Hydrogels for Bone Tissue Engineering

Abstract

Hybrid hydrogels composed of chitosan (CS) have shown great potential in bone tissue engineering and regeneration. The introduction of polyhedral oligomeric silsesquioxanes (POSS) in the biopolymeric matrix has been demonstrated to improve the rheological and biological properties of the hybrid composites. In this work, we have integrated the favourable features of chitosan (CS) and POSS nanoparticles to design new nanocomposites for bone tissue regeneration, focusing our attention on the effect of POSS concentration within the CS matrix (0.5, 1, and 1.5 equivalents in weight of POSS with respect to CS) on the chemical, physical, rheological, and in vitro biological properties of the final composites. The drug release ability of the synthesized hydrogel scaffolds were also investigated using, as the model drug, ketoprofen, that was included in the scaffold during the gelling procedure, showing a more controlled release for the hybrids with respect to CS (86-91% of drug released after two weeks). The results of the in vitro biological tests performed on human fetal osteoblastic cells (hFOB 1.19) culture demonstrated the great biocompatibility of the hybrid materials. The hybrids, at the different POSS concentrations, showed values of cell mortality superimposable with control cells (11.1 vs. 9.8%), thus revealing the CS/POSS hydrogels as possible candidates for bone tissue engineering applications.

Keywords: biomaterials; biopolymers; hybrid materials; hydrogels; polyhedral oligomeric silsesquioxanes; tissue engineering.

Figure 1

Structure of CS-POSS.

Scheme 1

Synthesis of CS-POSS hybrid hydrogels. Reagents and conditions: (a) acetic acid 2%, 50 °C, 15 h, 50 °C, overnight, then NaHCO₃ until pH 7. (b) genipin, acetic acid 2%, 45 °C, 30 min.

Figure 2

FTIR spectra of samples CS, POSS, CS-POSS 1, CS-POSS 2, and CS-POSS 3.

Figure 3

TGA curves for CS-, POSS-, and CS-POSS-based samples, CS-POSS 1, CS-POSS 2, and CS-POSS 3. All experiments were performed in triplicate under argon atmosphere.

Figure 4

Complex viscosity (η*) (a) and G′ modulus (b) vs. frequency for CS and CS-POSS samples, at different POSS concentrations.

Figure 5

Ketoprofen release from CS and CS-POSS hydrogels at different POSS concentrations at 37 °C in PBS (pH 7.4). Data in the inset represent mean and standard deviation of three experiments in the first 12 h (SD ≤ 0.45).

Figure 6

Results of MTT test for the assessment of the assayed hydrogel biocompatibility in hFOB 1.19. Each value represents the mean (±SD) of the percentage of dead cells in the experiments made in triplicate.

Figure 7

CLSM images of hFOB 1.19 cells labelled with the metachromatic fluorophore AO. (A) Control cells; (B) osteoblastic cells grown in the presence of CS hydrogel; (C) osteoblastic cells grown in presence of CS hydrogel with 1.5 wt% of POSS. In acidic compartment of cytosol (intact lysosomes and mature endosomes; i.e., phagolysosomes), the fluorophore is sequestered thanks to the proton pump and emits a red fluorescence, while it emits green florescence in the absence of a low pH. The cells are morphologically analogous in (A–C).