Production of biocompatible composite scaffolds shifts towards additive manufacturing where thermoplastic biodegradable polymers such as poly(lactic acid) (PLA) are used as matrices. Differences between industrial- and medical-grade polymers are often overlooked although they may affect properties and degradation behaviour as significantly as the filler addition. In the present research, composite films based on medical-grade PLA and biogenic hydroxyapatite (HAp) with 0, 10, and 20 wt.% of HAp were prepared by solvent casting technique. The degradation of composites incubated in phosphate-buffered saline solution (PBS) at 37 °C after 10 weeks showed that the higher HAp content slowed down the hydrolytic PLA degradation and improved its thermal stability. Morphological nonuniformity after degradation was indicated by the different glass transition temperatures (T_g) throughout the film. The T_g of the inner part of the sample decreased significantly faster compared with the outer part. The decrease was observed prior to the weight loss of composite samples.

Ionic substitutions within the hydroxyapatite lattice are a widely used approach to mimic the chemical composition of the bone mineral. In this work, Sr-substituted and Mg- and Sr-co-substituted calcium phosphate (CaP) scaffolds, with various levels of strontium and magnesium substitution, were prepared using the hydrothermal method at 200 °C. Calcium carbonate skeletons of cuttlefish bone, ammonium dihydrogenphosphate (NH₄H₂PO₄), strontium nitrate (Sr(NO₃)₂), and magnesium perchlorate (Mg(ClO₄)₂) were used as reagents. Materials were characterized by means of X-ray diffraction (XRD), Fourier transform infrared spectroscopy (FTIR) and scanning electron microscopy (SEM). Whole powder pattern decomposition refinements of XRD data indicated that increased magnesium content in the Mg- and Sr-co-substituted scaffolds was related to an increased proportion of the whitlockite (WH) phase in the biphasic hydroxyapatite (HAp)/WH scaffolds. In addition, refinements indicate that Sr²⁺ ions have replaced Ca²⁺ sites in the WH phase. Furthermore, PCL-coated Mg-substituted and Sr- and Mg-co-substituted scaffolds, with the HAp:WH wt. ratio of 90:10 were prepared by vacuum impregnation. Results of compression tests showed a positive impact of the WH phase and PCL coating on the mechanical properties of scaffolds. Human mesenchymal stem cells (hMSCs) were cultured on composite scaffolds in an osteogenic medium for 21 days. Immunohistochemical staining showed that Mg-Sr-CaP/PCL scaffold exhibited higher expression of collagen type I than the Mg-CaP/PCL scaffold, indicating the positive effect of Sr²⁺ ions on the differentiation of hMSCs, in concordance with histology results. Reverse transcription-quantitative polymerase chain reaction (RT-qPCR) analysis confirmed an early stage of osteogenic differentiation.

Increasing attention is focused on developing biomaterials as temporary scaffolds that provide a specific environment and microstructure for bone tissue regeneration. The aim of the present work was to synthesize silicon-doped biomimetic multi-phase composite scaffolds based on bioactive inorganic phases and biocompatible polymers (poly(ε-caprolactone), PCL) using simple and inexpensive methods. Porous multi-phase composite scaffolds from cuttlefish bone were synthesized using a hydrothermal method and were further impregnated with (3-aminopropyl)triethoxysilane 1–4 times, heat-treated (1000 °C) and coated with PCL. The effect of silicon doping and the PCL coating on the microstructure and mechanical and biological properties of the scaffolds has been investigated. Multi-phase scaffolds based on calcium phosphate (hydroxyapatite, α-tricalcium phosphate, β-tricalcium phosphate) and calcium silicate (wollastonite, larnite, dicalcium silicate) phases were obtained. Elemental mapping revealed homogeneously dispersed silicon throughout the scaffolds, whereas silicon doping increased bovine serum albumin protein adsorption. The highly porous structure of cuttlefish bone was preserved with a composite scaffold porosity of ~78%. A compressive strength of ~1.4 MPa makes the obtained composite scaffolds appropriate for non-load-bearing applications. Cytocompatibility assessment by an MTT assay of human mesenchymal stem cells revealed the non-cytotoxicity of the obtained scaffolds.

The application of divalent bioactive metal ions, such as Cu²⁺, Zn²⁺, and Mn²⁺, emerges as a growth factor-free approach for bone defect regeneration. Delivery of those ions can be achieved by organic or inorganic phases through desirable rapid or sustainable release in order to stimulate specific cell responses. In this work, bioactive ions were incorporated into both phases, chitosan (Cht), via chelation reactions, and mesoporous bioactive glass nanoparticles (MBGNs), by doping. The BG/Cht composites with undoped and Cu-, Zn-, or Mn-doped MBGNs were produced as spherical microparticles with a narrow size distribution and an average size of 42-45 μm via electrohydrodynamic atomization. Swelling studies showed enhanced water uptake in the complete cell culture medium with values between 2.5 and 3.1 compared to phosphate-buffered saline (2.2-2.5). Ion release experiments in phosphate-buffered saline revealed a pronounced release of silicon and calcium up to 7 days for all samples. A sustained release of manganese ions from the MnBG/Cht sample was detected for up to 14 days. A precipitated layer of calcium phosphates on all composites, except on the MnBG/Cht samples, confirmed the materials' bioactivity after 21 days in simulated body fluid. Indirect cytotoxicity tests indicated that the materials were generally nontoxic to human osteosarcoma (MG-63) cells at concentrations below 1 mg/mL. However, direct contact assays with MG-63 and human dermal fibroblast (HDFa) cells revealed concentration-dependent cytotoxic effects, particularly for MnBG/Cht microparticles at a concentration of 0.5 mg/mL. Vascular endothelial growth factor (VEGF) expression analysis on MG-63 and HDFa cells demonstrated that only a higher concentration of MnBG/Cht significantly enhanced the angiogenic response in MG-63 cells, likely due to the decreased cell viability and oxidative stress generated by the redox activity of Mn²⁺ ions. Our results show that composite microparticles have good potential in the design of microparticulate systems with tailored properties through the combination of bioactive metal ions.