A Blood Vessel Organoid Model Recapitulating Aspects of Vasculogenesis, Angiogenesis and Vessel Wall Maturation

Abstract

Blood vessel organoids are an important in vitro model to understand the underlying mechanisms of human blood vessel development and for toxicity testing or high throughput drug screening. Here we present a novel, cost-effective, and easy to manufacture vascular organoid model. To engineer the organoids, a defined number of human induced pluripotent stem cells are seeded in non-adhesive agarose coated wells of a 96-well plate and directed towards a lateral plate mesoderm fate by activation of Wnt and BMP4 signaling. We observe the formation of a circular layer of angioblasts around days 5–6. Induced by VEGF application, CD31+ vascular endothelial cells appear within this vasculogenic zone at approximately day 7 of organoid culture. These cells arrange to form a primitive vascular plexus from which angiogenic sprouting is observed after 10 days of culture. The differentiation outcome is highly reproducible, and the size of organoids is scalable depending on the number of starting cells. We observe that the initial vascular ring forms at the interface between two cell populations. The inner cellular compartment can be distinguished from the outer by the expression of GATA6, a marker of lateral plate mesoderm. Finally, 14-days-old organoids were transplanted on the chorioallantois membrane of chicken embryos resulting in a functional connection of the human vascular network to the chicken circulation. Perfusion of the vessels leads to vessel wall maturation and remodeling as indicated by the formation of a continuous layer of smooth muscle actin expressing cells enwrapping the endothelium. In summary, our organoid model recapitulates human vasculogenesis, angiogenesis as well as vessel wall maturation and therefore represents an easy and cost-effective tool to study all steps of blood vessel development and maturation directly in the human setting without animal experimentation.

Neuro-mesodermal assembloids recapitulate aspects of peripheral nervous system development in vitro

“…The generation of mesodermal aggregates takes 4 days and is directed by CHIR99021 and BMP4. At the day of assembloid fusion, mesodermal aggregates consist of undifferentiated mesodermal progenitor cells ( Schmidt et al., 2022 ). In the assembloid, the mesodermal part is characterized by loose connective tissue and the presence of CD31-positive blood vessel-like structures that form a perineural vascular plexus at the neuro-mesodermal interface ( Figures 1 C and 1D).…”

In ovo model in cancer research and tumor immunology

“…A major advantage relies in the CAM’s easy accessibility and low rejection rate to facilitate increasing the model’s complexity by engrafting 3D spheroids ( 118 , 119 ) or multicellular organoids. For instance, mature organoids derived from human-induced pluripotent stem cells (PSCs) rapidly connected to the vascular network of the chick embryo after transferring them on the CAM ( 120 ), and PSC–derived inner ear and kidney organoids demonstrated the model’s potential to optimize developmental maturity and functionality of organoids based on vascularization ( 121 , 122 ). Along similar lines, grafting of tumor organoids, also combined with immune cells and/or fibroblasts, reflecting tissue heterogeneity in cancer to a greater extent, can be employed ( 123 ).…”

Advances in the differentiation of pluripotent stem cells into vascular cells

“…However, the exact composition can vary between different production batches. To solve this problem, Schmidt et al [ 54 ] proposed a vascular organoid induction protocol without Matrigel. They used a 96-well plate coated with conical agarose to aggregate iPSCs for subsequent organoid culture.…”