(10b) Targeting α5β1 On Pig Islets of Langerhans in Culture with PR_b, a Fibronectin-Mimetic Peptide, to Increase Islet Yield and Viability

Atchison, N., University of Minnesota
Fan, W., University of Massachusetts Amherst
Papas, K. K., University of Minnesota
Hering, B. J., University of Minnesota
Tsapatsis, M., University of Minnesota
Kokkoli, E., University of Minnesota

Islet transplantation is a promising treatment for type 1 diabetes but the limited supply of human donor pancreases dictates the need for alternative islet cell sources. Pig islets could provide an unlimited supply for xenotransplantation and have shown promise as an alternative to human islet allografts. However, stresses imposed during islet isolation and transplantation decrease islet yield and viability, leading to loss of graft function. Extracellular matrix (ECM) is lost during the islet isolation process and has been shown to cause islet cell apoptosis, decreasing the viability and number of islets recovered per pancreas. Additionally, transplanted islets suffer from hypoxic and other stresses that affect cell viability, leading to eventual loss of graft function. Therefore, to increase islet graft survival, the islets must be protected from apoptosis occurring from both isolation and transplantation.

This study investigated the ability of a fibronectin-mimetic peptide, PR_b (KSSPHSRN(SG)5RGDSP)), which specifically binds to the α5β1 integrin, to reestablish the ECM around pig islets that is destroyed during isolation. Additionally, PR_b functionalized liposomes were investigated as possible drug-delivery vehicles to pig islet cells. Firstly, we demonstrated the presence of the α5β1 integrin on the pig islet cells using confocal microscopy and Western blotting. We then found that islets cultured in medium supplemented with varying concentrations of free PR_b peptide had dose-dependent, increased levels of ECM fibronectin secretion compared to control islets. This technology could potentially lead to increased viable islet yields after isolation, decreasing the number of donors necessary per xenotransplantation. Using confocal microscopy and flow cytometry we found that PR_b functionalized liposomes delivered to the pig islets internalized into the cells in a PR_b concentration dependent manner, and non-functionalized liposomes showed virtually no internalization. PR_b functionalized liposomes targeted to the cells ex vivo could increase delivery of therapeutics or imaging molecules that could protect the cells from stresses encountered after transplantation, increasing graft function and survival, and allow for post transplant monitoring. These studies demonstrated that the fibronectin-mimetic peptide, PR_b, is an appropriate peptide bullet for applications involving α5β1 expressing pig islet cells. PR_b could provide a multi-faceted approach to protecting and maintaining islet viability, increasing the success of pig islet xenotransplantation.