Type 1 diabetes is caused by the autoimmune destruction of insulin-producing β-cells, resulting in a chronic metabolic disorder typically treated with exogenous insulin. Even with the aid of advanced insulin pumps and real time feedback systems, blood glucose levels still deviate outside of the range maintained by native islets, which places the individual at risk for vascular complications and life-threatening hypoglycemic events. Cell replacement therapies have demonstrated the capacity to tightly control blood glucose levels. The wide adoption of cell replacement therapy is hindered by limited availability of donor islets, and the lack of effective methods to support the long-term function of these cells within a clinically accessible site. The results presented in this thesis address these limitations: through studying maturation of human pluripotent progenitor cell (hPPC) derived β cells within a transplantable biomaterial platform, and evaluating novel approaches to the implantation and support of these cells during their continued maturation in vivo.

First, I present a study that examined delivery of hPPC-derived pancreatic progenitors within microporous PLG scaffolds into the epididymal fat pad, the murine surrogate for the clinically relevant omental pouch. Kidney capsule injection, the site most commonly utilized to test stem cell-derived β cell function in murine models, was the comparison condition. We observed that the microporous scaffolds supported cell engraftment, however the levels of circulating C-peptide were lower when compared to the kidney capsule condition. The scaffolds were subsequently modified to provide sustained release of exendin-4, which led to significantly increased C-peptide production. Image analysis revealed that exendin-4 releasing scaffolds enhanced the proportion of pancreatic progenitors that matured to monohormonal insulin producing cells.

Next I present my findings from studying how hPPC-derived β cells mature and function within three transplantation sites: the i) scaffold delivery into the epididymal fat pad, ii) scaffold delivery into the subcutaneous space, and iii) the kidney capsule injection (control). Additionally, we investigated the impact of blood glucose levels on maturation of the hPPC-derived β cells by transplanting mice with pre- or post-engraftment diabetes induction. Hyperglycemia was ameliorated in the cohorts of mice that received scaffolds into the epididymal fat pad, following a period of in vivo maturation. The function of these cells was demonstrated by the reduction in blood glucose levels, healthy increase in weight, therapeutic levels of circulating human insulin, and healthy responses to glucose challenge tests. The function from the epididymal fat pad was superior to the subcutaneous space and was observed to be comparable to the kidney capsule. No differences were observed in graft function between the cohorts whose grafts matured in a diabetic or non-diabetic environment, yet several differences in gene expression were observed.

Many of the current differentiation protocols culture the cells above a feeder layer in monolayer, or in suspension within a bioreactor. Typically, these protocols require the disruption of the cell niche during key differentiation stages or pre-transplantation handling. Biomaterial scaffolds maintain the integrity of cell-to-cell and cell-to-matrix connections by providing both a space for cell niche development as well as a vehicle for transplantation into the body. Herein, I present results from testing the developmental stage in which progenitors are seeded into the 3D niche, and two differentiation strategies prior to seeding: monolayer and suspension culture. Maturation was characterized via gene expression analysis, glucose stimulated insulin secretion assay, and nondestructive microscopy utilizing a sfGFP-C-peptide cell line that reports C-peptide production and secretion. We observed that seeding clusters during the key transition phase from pancreatic progenitor to pancreatic endocrine enhanced commitment to the final beta cell fate.

This work enhances our understanding of hPPC-derived beta cell manufacturing within scaffolds, and delivery to an extrahepatic site to achieve normoglycemic blood glucose levels.

Date: Wednesday, August 4, 2021
Time: 10:00 AM
Zoom: https://umich.zoom.us/j/98009467275
Password: betacell
Chair: Dr. Lonnie Shea