Bioengineering methods for vascularizing organoids

Abstract

Organoids, self-organizing three-dimensional (3D) structures derived from stem cells, offer unique advantages for studying organ development, modeling diseases, and screening potential therapeutics. However, their translational potential and ability to mimic complex in vivo functions are often hindered by the lack of an integrated vascular network. To address this critical limitation, bioengineering strategies are rapidly advancing to enable efficient vascularization of organoids. These methods encompass co-culturing organoids with various vascular cell types, co-culturing lineage-specific organoids with vascular organoids, co-differentiating stem cells into organ-specific and vascular lineages, using organoid-on-a-chip technology to integrate perfusable vasculature within organoids, and using 3D bioprinting to also create perfusable organoids. This review explores the field of organoid vascularization, examining the biological principles that inform bioengineering approaches. Additionally, this review envisions how the converging disciplines of stem cell biology, biomaterials, and advanced fabrication technologies will propel the creation of increasingly sophisticated organoid models, ultimately accelerating biomedical discoveries and innovations.

Keywords: CP: Biotechnology; CP: Stem cell; bioengineering methods; human pluripotent stem cells; organoid-on-a-chip; organoids; vascularization.

Figure 1

Schematic of the bioengineering methods for vascularizing organoids Organoids can be vascularized through co-culture with vascular cells, co-culture with vascular organoids, organoid co-differentiation, OOC platforms, and organoid 3D bioprinting. Created with BioRender.com.

Figure 2

VEGF gradient-driven angiogenesis and Piezo1-mediated EC signaling (A) VEGF gradient-driven angiogenesis in response to hypoxia. Angiogenic stimuli: ECs lining the blood vessels are exposed to gradients of VEGF, with concentrations inversely proportional to oxygen tension in the tissue. Tip cell competition: in response to the angiogenic gradient, VEGFR2 is expressed on one cell, stimulating Notch signaling in the neighboring cell. This intercellular communication results in the downregulation of VEGFR2 and upregulation of sVEGFR1, which binds to and sequesters free VEGF. The communication between the adjacent tip cells determines which cell becomes the tip cell and which remains a stalk cell. Sprouting: the tip cell leads the sprouting angiogenesis, extending over the elongating stalk cells toward the hypoxic region. Reprinted from “The Process of Sprouting Angiogenesis in a Healthy Blood Vessel,” by BioRender.com (2024). (B) Piezo1-mediated EC signaling in response to flow conditions. Under laminar flow (left), Piezo1 channel activation results in calcium influx, which initiates a cascade of downstream effectors, including calpain, MT1-MMP, and Akt. These pathways effect several physiologic changes in the ECs. In turbulent conditions (right), Piezo1 activation results in activation of α5-integrin, promoting atherogenic inflammation of vessels over time. Adapted from “Piezo1 Endothelium Signaling,” by BioRender.com (2024). Retrieved from https://app.biorender.com/biorender-templates.

Figure 3

Various scaffolds for engineering vascularization (A) Comparison of animal-derived (left) and synthetic (right) scaffolding materials for organoid engineering. Animal-derived Matrigel is characterized by complex and variable matrix components in undefined ratios. Moreover, the presence of xenogeneic contaminants and proteins can result in undesirable effects and batch variability. The synthetic polymeric scaffold (right) has components in well-defined ratios with highly tunable physicochemical properties, offering a controlled cellular response. (B) Decellularization process of native tissue. The progression from left to right illustrates the transition from a native tissue, characterized by abundant cellular components, to a partially decellularized state with reduced intracellular material, and subsequently to a fully decellularized matrix, rich in extracellular matrix (ECM) components. The intricate ECM architecture is preserved for use in organoid vascularization. Reprinted from “Comparison of Matrigel and Synthetic Scaffolds” and “The Decellularization Effect on the Extracellular Matrix (ECM)” by BioRender.com (2024). Retrieved from https://app.biorender.com/biorender-templates.