Osteoblast-induced biodegradation of poly-(e-caprolactone)-carbonated apatite composites for bone tissue engineering.

Poly(-caprolactone)-apatite (PCL-AP) composites are promising devices as long-term biodegradable scaffolds for bone repair. To evaluate the influence of the PCL/AP ratio on in vitro degradation and bioactivity, the PCL-AP scaffolds were immersed in different aqueous media and analysed at different degradation times by vibrational spectroscopy and thermal analysis. The same techniques were used to comparatively characterise the composites incubated with human osteoblasts (HOB), in order to evaluate the cell role in degradation. The samples were synthesised using phase-inversion and a salt-leaching technique. The composites had different PCL/AP ratios: 50/50, 60/40 and 75/25. Pure PCL samples were analysed as control. The in vitro biodegradation was investigated under sterile conditions at 37°C in different media: saline phosphate buffer at pH 7.4 (SPB), 0.01 M NaOH solution, simulated body fluid at pH 7.4 (SBF). The samples were analysed before and after biodegradation for different degradation times (up to 2 months) by Raman spectroscopy, thermogravimetry (TG) and differential scanning calorimetry (DSC). HOB were isolated from bone fragments obtained during surgery for total hip replacement and cultivated in -MEM with 10% FBS on the PCL-AP samples for 4 weeks. Thermal analysis of the undegraded samples showed that in the 50/50 sample the PCL component was the most crystalline. Raman analysis confirmed this result as well as the relative composition of the scaffolds. The NaOH degradation medium was found to be the most aggressive towards both pure PCL and composites, confirming the catalytic effect of the OH- ion on degradation. Accordingly, the pure PCL samples degraded in NaOH solution underwent the most pronounced morphology changes: crystallinity increased as well as the intensity of the Raman bands due to crystalline PCL and the crystallisable fraction. These findings indicate that degradation preferentially involved the amorphous domains of the polymer with a consequent chain fragmentation and enrichment in crystalline PCL. Among the PCL-AP composites, the PCL component of the 50/50 sample showed the least degradation extent upon degradation in NaOH solution, as expected on the basis of its initially highest crystallinity. In the SPB medium, a certain degree of chain fragmentation was revealed. Upon immersion in SBF solution all the composites showed deposition of an apatitic phase; gravimetric measurements coupled to Raman and TG analyses indicated that the 50/50 sample was the most bioactive. The thermal analysis of the cell-incubated samples indicated a significant degradation of the polymeric component present in the composites, especially in the 75/25 sample, while the pure PCL samples appeared the least degraded. Raman spectroscopy coupled to thermal analysis proved a valid method to characterise the samples under study and their behavior in different media and after osteoblast-incubation.