Engineering Islet-Specific Microvascular Network within hPSC Derived Pancreatic Islet Organoids

An emerging area in tissue engineering is the development of three dimensional engineered constructs, organoids, which are comprised of multiple organ-specific cell populations capable of recapitulating an in-vivo organ system’s structure and function in an in-vitro setting. Engineering tissue specific organoids from human pluripotent stem cells (hPSCs) has resulted in successful reproduction of similar organ functionality from renewable cell source for a variety of target organ systems. Such systems have included derivation of intestinal, brain, and renal organ models. Development of organoid systems requires organ-specific parenchyma cell source and a platform for self-organization and lineage specific induction of chosen cell types. Additionally, reproducing the organ-specific functional vasculature during reproduction of the complex organ structure is extremely vital for maintaining nutrient supply and appropriate organ function. This is particularly crucial while engineering pancreatic islet organoids, since a dense fenestrated intra islet vasculature is vital for supporting glucose delivery and insulin response.

We have investigated multiple 3D platforms which promote spontaneous aggregation of pre- differentiated hPSC derived pancreatic progenitor cells (hESC-PPs) into a 3D organoid which demonstrated functional insulin production in vitro as well as in vivo mouse model. The resulting spheroids are readily recoverable and amenable for size and cellular composition tuning. This hydrogel mediated aggregation allowed direct integration of isolated microvessel elements within the hESC-PP organoids. Continued in vitro culture of the multicellular islet organoids promoted microvascular expansion and formation of vascular networks, especially with inclusion of supporting stromal cell populations. Pancreatic phenotype of the vascularized organoids was strengthened, demonstrated by the gene expression of key pancreatic maturation markers (NKX6.1, PDX1, and INS). Moreover, the intra-organoid vasculature demonstrated an increase in islet endothelial specific API gene expression and PLVAP, an indicator of increased endothelial diaphragm and fenestration, key elements of islet vascular development. In vivo vascular development within organoids was further characterized in mouse implantation models. In conclusion, we believe the results present a major step in the in-vitro production of hPSC islet organoids with microvasculature, likely to enhance function and foster faster in-vivo integration, while also being conducive for organ-on-a-chip applications.