Abstract
Standard cell cultures may not predict the proliferation and differentiation potential of human mesenchymal stromal cells (MSCs) after seeding on a scaffold and implanting this construct in a bone defect. We aimed to develop a more biologically relevant in vitro 3D-model for preclinical studies on the bone regeneration potential of MSCs. Human adipose tissue-derived mesenchymal stromal cells (hASCs; five donors) were seeded on biphasic calcium phosphate (BCP) granules and cultured under hypoxia (1% O2) for 14 days with pro-inflammatory TNFα, IL4, IL6, and IL17F (10 mg/mL each) added during the first three days, simulating the early stages of repair (bone construct model). Alternatively, hASCs were cultured on plastic, under 20% O2 and without cytokines for 14 days (standard cell culture). After two days, the bone construct model decreased total DNA (3.9-fold), COL1 (9.8-fold), and RUNX2 expression (19.6-fold) and metabolic activity (4.6-fold), but increased VEGF165 expression (38.6-fold) in hASCs compared to standard cultures. After seven days, the bone construct model decreased RUNX2 expression (64-fold) and metabolic activity (2.3-fold), but increased VEGF165 (54.5-fold) and KI67 expression (5.7-fold) in hASCs compared to standard cultures. The effect of the bone construct model on hASC proliferation and metabolic activity could be largely mimicked by culturing on BCP alone (20% O2, no cytokines). The effect of the bone construct model on VEGF165 expression could be mimicked by culturing hASCs under hypoxia alone (plastic, no cytokines). In conclusion, we developed a new, biologically relevant in vitro 3D-model to study the bone regeneration potential of MSCs. Our model is likely more suitable for the screening of novel factors to enhance bone regeneration than standard cell cultures.