Hydroxyl Groups Induce Bioactivity in Silica/Chitosan Aerogels Designed for Bone Tissue Engineering. In Vitro Model for the Assessment of Osteoblasts Behavior

Abstract

Silica (SiO2)/chitosan (CS) composite aerogels are bioactive when they are submerged in simulated body fluid (SBF), causing the formation of bone-like hydroxyapatite (HAp) layer. Silica-based hybrid aerogels improve the elastic behavior, and the combined CS modifies the network entanglement as a crosslinking biopolymer. Tetraethoxysilane (TEOS)/CS is used as network precursors by employing a sol-gel method assisted with high power ultrasound (600 W). Upon gelation and aging, gels are dried in supercritical CO2 to obtain monoliths. Thermograms provide information about the condensation of the remaining hydroxyl groups (400–700 °C). This step permits the evaluation of the hydroxyl group’s content of 2 to 5 OH nm−2. The formed Si-OH groups act as the inductor of apatite crystal nucleation in SBF. The N2 physisorption isotherms show a hysteresis loop of type H3, characteristic to good interconnected porosity, which facilitates both the bioactivity and the adhesion of osteoblasts cells. After two weeks of immersion in SBF, a layer of HAp microcrystals develops on the surface with a stoichiometric Ca/P molar ratio of 1.67 with spherulite morphology and uniform sizes of 6 μm. This fact asserts the bioactive behavior of these hybrid aerogels. Osteoblasts are cultured on the selected samples and immunolabeled for cytoskeletal and focal adhesion expression related to scaffold nanostructure and composition. The initial osteoconductive response observes points to a great potential of tissue engineering for the designed composite aerogels.

Effect of Washing Treatment on the Textural Properties and Bioactivity of Silica/Chitosan/TCP Xerogels for Bone Regeneration

“…When HOB ® cells were seeded on the xerogels, an initial polarization could be seen from the first 24 h, followed by cell adhesion and morphological changes identified as initial markers of osteoblast differentiation and due to the presence of the biomaterial as described previously by us and others [ 44 , 53 , 75 , 76 , 77 ]. Adhesion, attachment, cell growth, and morphological changes in osteoblasts appeared to be substantially better in silica–chitosan xerogels’ experimental samples, mainly in those including TCP, than in cells grown on the bare substrata and revealed a successful cell attachment with marked morphological changes, such as filopodial and lamellipodial emission and an improved cell spreading.…”

“…In all cases, an initial weight loss of about 1–2 wt.% from 50 °C to 100 °C takes place, associated with the evaporation of physically trapped water, thus confirming the hydrophilic character of the xerogel surfaces. Additionally, the thermal decomposition profiles of U, E1, and E7 in Figure 4 a–c, respectively, were very similar, and the same thermal events should be implicated, e.g., SCS8 and SCS8T10 thermal profiles of these washing series show two main step losses after the evaporation of water: the first one in the range 120–250 °C, accounting for the removal of silanols and combustion of CS, and furthermore, a gradual weight loss from 260 to 700 °C, accounting for dehydroxilation of isolated -OH groups [ 53 ]. Otherwise, the thermal profile of SCS8T20_U indicates the influence of TCP, giving an explanation for the step loss (ca.1.3 wt.%) occurring in the range 160–210 °C, presumed to be due to a dehydration of a crystalline phase of TCP, according to Jinlong et al [ 64 ].…”

“…The ten major peaks observed in Figure 5 a corresponding to the following wavenumbers, 470, 540, 800, 965, 1070, 1190, 1510, 1650, 3500, and 3650 cm −1 , are associated with the following absorption bands: 470 cm −1 and 540 cm −1 , related to bending vibrations and Si–O− rocking mode, respectively [ 53 ]. At a higher frequency, absorption bands at 800 cm −1 are related to symmetric Si–O–Si stretching vibrations, and bands at 965 cm −1 to surface silanol Si–O(H) bond stretching [ 66 ].…”

“…In this region, a characteristic band between 3400 and 3200 cm −1 due to hydrogen bonding interaction between the carbonyl group of chitosan and the silica network occurs for U samples. The resolution of this broad band into two peaks at 3410 and 3480 cm −1 , exclusively observed in W30 xerogels, ( Figure 5 b) is assigned to surface –OH isolated, germinal, and vicinal silanol groups [ 53 ]. This Si–OH surface film accounts for the increase in the hydrophilic character of the surface of these materials, as well as for the siloxane ring structure, regarding the micro-mesoporosity [ 69 , 70 ].…”

“…The composition of silica xerogels can be tuned so that they elicit bone bioactive characteristics [ 44 , 53 ]. As it is widely accepted, the bone-bonding ability of a biomaterial depends on ion release and exchange to develop a biological apatite layer when these materials are in the presence of body fluids.…”