J Tissue Eng Regen Med. 2019 Jan 16. doi: 10.1002/term.2807. [Epub ahead of print]

Effect of Donor Variation on Osteogenesis and Vasculogenesis in Hydrogel Co-cultures.

Pennings I¹, van Dijk LA¹, van Huuksloot J¹, Fledderus JO², Schepers K³, Braat AK³, Hsiao EC⁴, Barruet E⁴, Morales BM⁴, Verhaar MC², Rosenberg AJWP¹, Gawlitta D¹.

Abstract

To introduce a functional vascular network into tissue-engineered bone equivalents, human endothelial colony forming cells (ECFCs) and multipotent mesenchymal stromal cells (MSCs) can be co-cultured. Here, we studied the impact of donor variation on human bone marrow-derived MSCs and cord blood-derived ECFCs on vasculogenesis and osteogenesis using a 3D in vitro co-culture model. Further, to make the step towards co-cultures consisting of cells derived from a single donor, we tested how induced pluripotent stem cell (iPSC)-derived human endothelial cells (iECs) perform in co-culture models. Co-cultures with varying combinations of human donors of MSCs, ECFCs, or iECs were prepared in Matrigel. The constructs were cultured in an osteogenic differentiation medium. Following a 10-day culture period, the length of the pre-vascular structures and osteogenic differentiation were evaluated for up to 21 days of culture. The particular combination of MSC and ECFC donors influenced the vasculogenic properties significantly and induced variation in osteogenic potential. In addition, the use of iECs in the co-culture model resulted in pre-vascular structure formation in osteogenically differentiated constructs. Together, these results showed that close attention to the source of primary cells such as ECFCs and MSCs is critical to address variability in vasculogenic and osteogenic potential. The 3D co-culture model appeared to be successful in generation of pre-vascularized constructs and has shown to be sufficient in exceeding the 200 μm diffusion limit. In addition, iPSC-derived cell lineages may decrease variability by providing a larger and potentially more uniform source of cells for future pre-clinical and clinical applications.