Animal-free alternatives for Matrigel in human iPSC-derived blood vessel organoid culture.

The use of blood vessel organoids (BVOs), or vascular organoids derived from human induced pluripotent stem cells (hiPSC), provides a valuable tool for research on vascular regeneration. Vascular organoid culture relies on the use of Matrigel, a murine sarcoma-derived matrix, which limits its translational potential and reproducibility. This study evaluated the potential alternatives to Matrigel for both 2D hiPSC and 3D vascular organoid culture to enhance the clinical applicability of hiPSC-derived vascular organoids. Vitronectin is a suitable replacement for Matrigel in hiPSC culture and expansion, maintaining pluripotency and facilitating subsequent differentiation into vascular organoids. Additionally, we demonstrated that fibrin-based hydrogels effectively support vascular organoid differentiation, promoting vascular network formation and endothelial cell sprouting comparable to Matrigel-based cultures. Through gene expression analysis, surface area quantification, and immunohistochemistry, we validated the efficacy of Vitronectin and fibrin in supporting hiPSC-derived vascular organoid differentiation. This Matrigel-free protocol not only enhances reproducibility but is a step forward in xeno-free culture conditions, offering a versatile platform for disease modelling, vascular tissue engineering, and patient-specific therapeutic screening.