Endocrine Pancreas based on Decellularized Pancreatic Matrix with Mesenchymal Stem Cells and Islet Cells Coculture
Claudius Conrad, MD, PhD, Christian Schuetz, MD, Ben Clippinger, MD, Joseph P. Vacanti, MD, James F. Markmann, MD, Harold C. Ott, MD.
Massachusetts General Hospital, Boston, MA
PURPOSE OF STUDY
Pancreas transplantation is the only treatment of Type I diabetes that can consistently establish insulin independence. Donor organ shortage and the need for immunosuppressive therapy limit number and outcomes of pancreas transplantation. To overcome this dilemma we attempted to engineer functional endocrine pancreas tissue by seeding pancreatic extracellular matrix (ECM) scaffolds with supportive mesenchymal stem cells (MSC) human islet cells.
Cadaveric pancreases were isolated from adult, heparinzed SD rats (n=8) and decellularized via detergent perfusion. The resulting matrices were analyzed for integrity of the vascular tree using methylen blue injections. The matrices were then seeded with human donor islet cells (200*106) and supportive MSC (10*106) via intravascular and intraparenchymal delivery. Endocrine function of the resulting constructs in response to high and low dose glucose stimulation was measured at three different timepoints over the course of seven days in culture (n=3). In parallel, islet cells were cultured under standard conditions and served as controls (n=3). Matrix constructs were analyzed using histology, and immunofluorescence.
SUMMARY OF RESULTS
Pancreatic ECM scaffolds with intact vascular and matrix architecture could be generated using perfusion decellularization. Decellularized pancreatic ECM can be seeded with MSCs and islet cells and maintained in a bioreactor under perfused isolated organ conditions to form insulin producing tissue. Insulin response assay showed preserved insulin response to glucose challenge on day 2, 5 and 7 after seeding. Histology and immunofluorescence showed preserved islet cell viability, the formation of three dimensional tissue and a close relationship between islet cells and MSCs.
Cadaveric pancreas can be decellularized to isolate intact pancreatic ECM scaffolds. Seeding of these scaffolds with islet cells, and MSCs yields perfusable, functional tissue constructs with preserved islet cell survival and function. The potential of pancreatic ECM in providing a niche for islet progenitors warrants further investigation.
TABLES AND CHARTS
Progressively decellularized bioartifical pancreas scaffold from a nude rat. Staining of the microvasculature with Tryphan blue.