The fabrication of self-assembling biomineral-related complexes for cell growth and gene/biomolecule delivery offers important opportunities for skeletal repair. In this work, 5–30 µm-sized calcium carbonate (vaterite) spheroids with elaborate sponge-like macroporous architectures were prepared by passive evaporation of water-in-oil microemulsions, and investigated as potential vehicles for the delivery of skeletal growth factors to human bone marrow stromal cells maintained in culture. The spheroids spontaneously form within dispersed droplets of supersaturated calcium bicarbonate, and become patterned in situ by entrapped micro-bubbles of carbon dioxide. Water-soluble biomolecules such as proteins (collagen type I, haemoglobin), cell growth factors (dexamethasone, pleiotrophin), bone morphogenetic proteins (rhBMP-2, SaOS ‘retentate’), and plasmid DNA were incorporated into the microspheres during synthesis or by soaking pre-washed spheroids. Primary human bone marrow stromal cells labelled with cell tracker green and ethidium homodimer-1 or transfected with an adenoviral vector expressing green fluorescent protein were co-cultured with the vaterite microspheres over three weeks without loss of function and viability. At three weeks, microspheres were encapsulated and integrated with the human bone marrow stromal cells. Histological analysis confirmed the expression of alkaline phosphatase, synthesis of extracellular matrix, and capacity for extensive mineralization. Use of vaterite microspheres to adsorb and deliver active growth factors such as pleiotrophin and an admixture of bone morphogenetic proteins derived from an osteosarcoma (SaOS ‘retentate’) was demonstrated using human bone marrow stromal cells. Ex vivo chorioallantoic membrane culture of pelleted vaterite spheres and human bone marrow cells placed into a chick femur defect resulted in the secretion of an organised collagen matrix. Vaterite spheres containing bioactive proteins induced osteogenesis in promyoblast C2C12 cells. These studies demonstrate the development of facile techniques for the generation of inorganic scaffolds constructed from porous microspheres that are biocompatible, aid mineralization, and offer potential for growth factor delivery through entrapment of functional biomolecules.
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