Dispersion of ceramic granules within human fractionated adipose tissue to enhance endochondral bone formation

Engineering of materials consisting of hypertrophic cartilage, as physiological template for de novo bone formation through endochondral ossification (ECO), holds promise as a new class of biological bone substitutes. Here, we assessed the efficiency and reproducibility of bone formation induced by the combination of ceramic granules with fractionated human adipose tissue ("nanofat"), followed by in vitro priming to hypertrophic cartilage. Human nanofat was mixed with different volumetric ratios of ceramic granules (0.2-1 mm) and cultured to sequentially induce proliferation (3 weeks), chondrogenesis (4 weeks), and hypertrophy (2 weeks). The resulting engineered constructs were implanted ectopically in nude mouse. The presence of ceramic granules regulated tissue formation, both in vitro and in vivo. In particular, their dispersion in nanofat at a ratio of 1:16 led to significantly increased cell number and glycosaminoglycan accumulation in vitro, as well as amount and inter-donor reproducibility of bone formation in vivo. Our findings outline a strategy for efficient utilization of nanofat for bone regeneration in an autologous setting, which should now be tested at an orthotopic site. STATEMENT OF SIGNIFICANCE: In this study, we assessed the efficiency and reproducibility of bone formation by a combination of ceramic granules and fractionated human adipose tissue, also known as nanofat, in vitro primed into hypertrophic cartilage. The resulting engineered cartilaginous constructs, when implanted ectopically in nude mouse, resulted in bone and bone marrow formation, more reproducibly and strongly that nanofat alone. This project evaluates the impact of ceramic granules on the functionality and chondrogenic differentiation of mesenchymal progenitors inside their native adipose tissue niche and outlines a novel strategy for an efficient application of nanofat for bone regeneration in an autologous setting.