Localized Tolerance and Development of an Alternative Transplant Site to Treat Type 1 Diabetes

[摘要] Islet transplantation is an attractive treatment for type 1 diabetes (T1D) to restore the body’s ability to endogenously produce insulin and rapidly respond to changes in blood glucose levels. Current clinical strategies where donor islets were intrahepatically transplanted have demonstrated success in a small number of patients. However, the widespread use of this approach is limited due the generation of allo- and autoimmune responses, which contributes to significant islet loss and eventual graft failure. This dissertation presents the development of an extrahepatic biomaterial scaffold that creates an alternative transplant site for the localized delivery of soluble factors and immunoregulatory proteins to enhance long-term transplant function. Two scaffold designs were employed to improve islet cell transplantation and explore the effects of different scaffold architectures on islet engraftment in the form of encapsulating and microporous polyethylene glycol (PEG)-based hydrogels to support islet function in the fat pad transplantation site of mice using syngeneic and allogeneic models. This allowed for the unique comparison of encapsulation and microporous techniques with the same material. Microporous hydrogels demonstrated rapid response to glucose challenge and were quickly infiltrated by host tissue. In contrast, islet-encapsulating PEG hydrogels both engraft and respond to fluctuations in blood glucose slower than microporous scaffolds. To modulate the local inflammatory environment, transforming growth factor β 1 (TGF-β1) was delivered from the PEG hydrogels and delayed rejection of allogeneic islets. Methods for sustained delivery of soluble factors were further explored by utilizing affinity peptides to localize lentiviral vectors for viral gene delivery. Poly-L-lysine (PLL), a cationic polypeptide, was covalently attached to PEG hydrogels and demonstrated the ability to modulate the extent of virus adsorption and increase the half-life of the adsorbed virus by 20%. An alternative to PLL was discovered through phage display technology, with peptide sequences specific for the glycoprotein of the vesicular stomatitis virus (VSV-G) ectodomain, an envelope protein pseudotyped on the virus. These short, 12 amino acid affinity peptides were easily incorporated into the hydrogel, and reporter protein expression was increased 20-fold relative to control peptide, comparable to levels observed with the high molecular weight PLL. Finally, the modification of biomaterials scaffolds with Fas ligand (FasL) was explored to create an immunoprivileged microenvironment that is translatable to the clinic. Poly(lactide-co-glycolide) (PLGA) was conjugated with biotin and fabricated into particles and microporous scaffolds to allow for rapid and efficient conjugation with the chimera protein streptavidin-FasL (SA-FasL). PLGA particles and microporous scaffolds coated with FasL demonstrated the ability to induce apoptosis in a mouse B lymphoma cell line. Scaffolds were functionalized with FasL, seeded with islets from BALB/c donors, and implanted in epididymal fat pad C57BL/6 recipients. Scaffolds with FasL and a short course treatment of rapamycin restored euglycemia and showed robust tolerance indefinitely without the sustained use of immunosuppressive drugs. Together, this dissertation presents work that will further our understanding of allogeneic transplants and create tools that can be applied to treat T1D.

[发布机构] University of Michigan

[效力级别] Scaffold

[关键词] PEG hydrogel;Scaffold;Localized gene delivery;Immunoprivilege;Type 1 diabetes;Biomedical Engineering;Chemical Engineering;Engineering;Science;Chemical Engineering