Engineering Pancreatic Islet Organoids with a Hierarchical Vascular Network

Background: One of the major challenges in subcutaneous islet transplantation is achieving adequate vascularization and a supportive scaffold for islet engraftment. Recent advances in cell engineering have enabled the fabrication of tissues containing pancreatic islets and vascular networks. However, reconstruction of a hierarchical vascular network comparable to that in native tissues has not yet been achieved. In this study, we aimed to develop pancreatic islet organoids incorporating a hierarchical vascular network capable of providing sufficient blood supply together with a structural scaffold for islet engraftment. Our goal was to establish a novel cell transplantation platform suitable for subcutaneous islet transplantation.
Methods: An organoid scaffold was fabricated using a 3D bioprinter with a 300-μm stainless steel needle inserted as a template for a large vessel. BALB/c mouse–derived pancreatic islets were combined with human umbilical vein endothelial cells (HUVECs) and mesenchymal stem cells (MSCs) and seeded into the scaffold.  After orbital shaking culture, the needle was removed and additional HUVECs were seeded into the lumen to form a large vessel structure.
Subsequent spontaneous microvascular formation was induced to establish a hierarchical vascular network within the islet organoid. Vascularization was quantified using AngioTool (National Cancer Institute). Cell viability was evaluated by Calcein-AM/PI staining, and insulin secretion was assessed by a glucose-stimulated insulin secretion (GSIS) assay.
Results: A stable vascular network including both large vessels and capillaries was successfully generated in vitro. The number and total length of vessels significantly increased from Day 0 to Day 2 (vessel number: 69 to 162, p < 0.0001; vessel length: 0.0572 mm to 0.1009 mm, p = 0.0009) and reached a plateau thereafter. Immunostaining for CD31 and insulin confirmed well-organized vascular networks surrounding the islets. Viability analysis demonstrated that islets within the organoids showed significantly higher survival rates than free islets under the same conditions (99.127% vs. 90.988%, p < 0.0001). GSIS assays showed clear glucose responsiveness (3.882 vs. 15.173, p < 0.0001), and the stimulation index was higher than that of the free islet group (4.571 vs. 3.142).In vivo transplantation into STZ-induced diabetic mice demonstrated that the organoid group achieved faster glycemic control compared with free islets. Blood glucose levels decreased from >600 mg/dL to <200 mg/dL within 6 weeks after transplantation.

Conclusion: This study demonstrated the successful construction of pancreatic islet organoids with a hierarchical vascular network, which improved the viability and functional performance of islet cells. These organoids may serve as a new transplantation platform that helps overcome key barriers in subcutaneous islet transplantation, including limited vascularization, insufficient engraftment support, and inflammation. This strategy may be extended to multiple cell sources, including stem cell–derived islets (SC-islets) and xenogeneic islets, potentially advancing next-generation islet transplantation and regenerative medicine.