Generating Mature Islet Cells In Vitro Using Decellularized Murine Pancreas

Abstract

Generating new islets from a patient's own induced pluripotent stem cells (iPSCs) could address the shortcomings of islet transplantations for the treatment of type 1 diabetes. The maturation process, however, can take months in vivo and does not occur efficiently in vitro indicating that factors are missing from traditional two-dimensional (2D) cultures. The interaction between cells and the extracellular matrix (ECM) is one such factor. Three-dimensional (3D) culturing using ECM can address this deficit. The removal of cells from an organ, through decellularization, can be performed to obtain the ECM. Our aim is to develop a 3D culture system using decellularized murine pancreas to better support the growth and differentiation of iPSC derived islets. Specifically, two decellularization protocols were tested to determine the optimal detergent for removal of all cellular material without loss of ECM structure and associated proteins. Two detergents were tested, Triton X-100 or 3-[(3-Cholamidopropyl) dimethylammonio]-1-Propanesulfonate (CHAPS)). Results showed that both protocols produced a visibly decellularized ECM. Immunohistochemistry (IHC) was performed for key ECM proteins as well as the nuclear stain 4,6-diamidino-2-phenylindole (DAPI). Results of the IHC for the Triton X-100 protocol showed retention of the ECM proteins without the presence of nuclear material. Results for CHAPS are currently being analyzed. Our preliminary results indicate that the generation of decellularized murine pancreas using Triton X-100 and CHAPS is a promising method for 3D islet culturing.