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This paper reports on the rapid development of porous hydroxyapatite (HA) microspheres with large external pores and fully interconnected porosity. These porous microspheres were produced by converting borates glasses (namely 45B5, B53P4 and 13-93B) into HA by immersing them in potassium phosphate media and simulated body fluid (SBF). Solid (SGMS) non-porous and highly porous (PGMS) microspheres were prepared from borate glasses via a novel flame spheroidisation process and their physicochemical properties including in vitro biological response were investigated. Morphological and physical characterisation of the PGMS showed interconnected porosity (up to 75 ± 5%) with average external pore sizes of 50 ± 5 μm. Mass loss, ion release, X-ray diffraction (XRD) and Scanning electron microscopy (SEM) analysis confirmed complete conversion to HA in 0.02 M K2HPO4 solution for the PGMS (with exception of 13-93B glass) and at significantly faster rates compared to their SGMS counterparts. However, 13-93B microspheres only converted to HA in Na2HPO4 solution. The in vitro SBF bioactivity studies for all the borate compositions showed HA formation and much earlier for PGMS compared to SGMS. Direct cell culture studies using hMSCs revealed that the converted porous HA microspheres showed enhanced pro-osteogenic properties compared to their unconverted counterparts and such are considered as highly promising candidate materials for bone repair (and orthobiological) applications.

A major challenge in orthopedics is the repair of large non-union bone fractures. A promising therapy for this indication is the use of biodegradable bioinspired biomaterials that stabilize the fracture site, relieve pain and initiate bone formation and healing. This study uses a multidisciplinary evaluation strategy to assess immunogenicity, allergenicity, bone responses and physicochemical properties of a novel biomaterial scaffold. Two-photon stereolithography generated personalized custom-built scaffolds with a repeating 3D structure of Schwarz Primitive minimal surface unit cell with a specific pore size of ∼400 μm from three different methacrylated poly(d,l-lactide-co-ε-caprolactone) copolymers with lactide to caprolactone monomer ratios of 16 : 4, 18 : 2 and 9 : 1. Using in vitro and in vivo assays for bone responses, immunological reactions and degradation dynamics, we found that copolymer composition influenced the scaffold physicochemical and biological properties. The scaffolds with the fastest degradation rate correlated with adverse cellular effects and mechanical stiffness correlated with in vitro osteoblast mineralization. The physicochemical properties also correlated with in vivo bone healing and immune responses. Overall these observations provide compelling support for these scaffolds for bone repair and illustrate the effectiveness of a promising multidisciplinary strategy with great potential for the preclinical evaluation of biomaterials.

Silica-coated superparamagnetic iron nanoparticles (SiMAGs) are an exciting biomedical technology capable of targeted delivery of cell-based therapeutics and disease diagnosis. However, in order to realise their full clinical potential, their intracellular fate must be determined. The analytical techniques of super-resolution fluorescence microscopy, particle counting flow cytometry and pH-sensitive nanosensors were applied to elucidate mechanisms of intracellular SiMAG processing in human mesenchymal stem cell (hMSCs). Super-resolution microscopy showed SiMAG fluorescently-tagged nanoparticles are endocytosed and co-localised within lysosomes. When exposed to simulated lysosomal conditions SiMAGs were solubilised and exhibited diminishing fluorescence emission over 7 days. The in vitro intracellular metabolism of SiMAGs was monitored in hMSCs using flow cytometry and co-localised pH-sensitive nanosensors. A decrease in SiMAG fluorescence emission, which corresponded to a decrease in lysosomal pH was observed, mirroring ex vivo observations, suggesting SiMAG lysosomal exposure degrades fluorescent silica-coatings and iron cores. These findings indicate although there is a significant decrease in intracellular SiMAG loading, sufficient particles remain internalised (>50%) to render SiMAG treated cells amenable to long-term magnetic cell manipulation. Our analytical approach provides important insights into the understanding of the intracellular fate of SiMAG processing, which could be readily applied to other particle therapeutics, to advance their clinical translation.

Bone regeneration is a complex biological process where major cellular changes take place to support the osteogenic differentiation of mesenchymal bone progenitors. To characterise these biological changes and better understand the pathways regulating the formation of mature bone cells, the metabolic profile of mesenchymal stem cell (MSC) differentiation in vitro has been assessed non-invasively during osteogenic (OS) treatment using a footprinting technique. Liquid chromatography (LC)-mass spectrometry (MS)-based metabolite profiling of the culture medium was carried out in parallel to mineral deposition and alkaline phosphatase activity which are two hallmarks of osteogenesis in vitro. Metabolic profiles of spent culture media with a combination of univariate and multivariate analyses investigated concentration changes of extracellular metabolites and nutrients linked to the presence of MSCs in culture media. This non-invasive LC-MS-based analytical approach revealed significant metabolic changes between the media from control and OS-treated cells showing distinct effects of MSC differentiation on the environmental footprint of the cells in different conditions (control vs. OS treatment). A subset of compounds was directly linked to the osteogenic time-course of differentiation, and represent interesting metabolite candidates as non-invasive biomarkers for characterising the differentiation of MSCs in a culture medium.

Raman microscopy was used as a label-free method to study the mineralisation of bone nodules formed by mesenchymal stem cells cultured in osteogenic medium in vitro. Monitoring individual bone nodules over 28 days revealed temporal and spatial changes in the crystalline phase of the hydroxyapatite components of the nodules.