Spherical dense monodispersed bioactive glass nanoparticles (BGNPs) of 90 ± 10 nm in diameter have therapeutic potential due to their ability to be internalised by cellsand to perform the sustained intracellular delivery of therapeutic cations. BGNPs with 25% strontium (Sr) substitution for calcium (Ca) (25%Sr-BGNPs: 90.6 mol% SiO2, 5.0 mol% CaO, and 4.4% mol% SrO) and 75% strontium substitution for calcium (75%Sr-BGNPs: 88.8 mol% SiO2, 1.8 mol% CaO, and 9.4% mol% SrO) were synthesised in the diameter range of 80 – 100 nm through the modified Stӧber process and were physically and biologically characterised. The dense silica nanoparticles (Si-NPs) were fabricated prior to incorporating with calcium nitrate and strontium nitrate as precursor sources of Ca and Sr through calcination at 680 °C. The amount of Ca and Sr incorporated into the silica network of Si-NPs is the key factor for maintaining monodispersity and sustainable release during subsequent degradation in aqueous environments. The incorporation of Ca and Sr were homogenously distributed without affecting the particles’ size, morphology and dispersity. Not all of the nominal Ca and Sr added incorporated into the silica network. Excess Ca and Sr components on the particles’ surface were removed.
The in vitro cytotoxicity was evaluated in which cells were in contact with the nanoparticles or their dissolution products. The cell viability of murine pre-osteoblast cell line (MC3T3-E1), human mesenchymal stromal cells (hMSCs), mice murine bone marrow stromal stem cells (BMSCs), and primary macrophages was not affected up to particle concentrations of 250 μg/mL. There was no significant effect on DNA quantity of treated cells with Sr-BGNPs compared to the control (untreated cells). Sr-BGNPs stimulated osteogenic differentiation of MC3T3-E1, hMSCs and BMSCs in the absence of osteogenic supplements, which was confirmed by detecting early-, mid- and late-osteogenic marker expressions and was attributed to their ionic release products. Sr-BGNPs affected both mineralisation and extracellular matrix formation. These particles also showed the potential to reduce osteoclastogenesis, indicated by the reduction in TRAP activity and multinucleated osteoclasts when cultures were supplemented with RANKL.Confocal fluorescent microscopy presented cellular uptake and localisation of the Sr-BGNPs in the cytoplasm of the cells. Transmission electron microscopy (TEM) confirmed internalisation and localisation of Sr-BGNPs inside the endosome/lysosome-like vesicles bordered by a membrane inside the cells via a mixed-uptake mechanism in which the clathrin-dependent endocytosis was the main pathway. In a co-culture system of osteoblasts-osteoclasts, Sr-BGNPs had either the ability to stimulate osteogenic differentiation of MC3T3-E1 in the absence of osteogenic supplements or to inhibit osteoclastogenesis of RAW264.7 in the presence of RANKL and this effect could be attributed to their ionic release products. Sr-BGNPs present great potential for bone regeneration applications. These particles may be used as an alternative injectable or incorporated into nano-composites.
To extend the applications of BGNPs, quaternary BGNP systems, in which zinc (Zn) (Sr-ZnBGNPs) or Cerium (Ce) (Sr-CeBGNPs) were incorporated into Sr-BGNPs, were developed through the modified two-step post-functionalisation. Sr-ZnBGNPs had the high capacity to kill cancer cells preferentially compared to normal cells because the released Zn ions that cause toxicity to the cancer cells was more present under an acidic environment. The differences in incorporation between these four cations, including Ca, Sr, Zn, and Ce, might be attributed to the different roles as the network modifier or the network former in the silica network.