Title: EVALUATION OF THREE DIFFERENT PERFUSION ROUTES FOR WHOLE RAT PANCREAS DECELLULARIZATION
Co-Author(s): Hendrik Napierala, Benjamin Strücker, Karl-Herbert Hillebrandt, Peter Tang, Dietrich Polenz, Anja Reutzel-Selke, Steffen Lippert, Nathanael Raschzok, Johann Pratschke, Igor-Maximilian Sauer
INTRODUCTION: A cell based treatment for type 1 diabetes can significantly reduce the appearance of life-threatening complications, like hypoglycemic episodes, and progression of diseaseassociated co-morbidities. Islet cell transplantation is able to achieve insulin independence in about 40-60% of the cases. But unfortunately graft-site-specific issues still remain a major obstacle to achieve long-term functionality of transplanted islets. The aim of this study was to address this problem by engineering a decellularized pancreas graft, which could be used for the generation of an endocrine Neo-Pancreas.
METHODS: Pancreata from 32 Fisher DPP-IV- rats (m/f, 150 – 550 g) were explanted. 8 organs served a controls. The rest was cannulated either via the A) abdominal aorta, B) the portal vein or the C) bile duct. All organs were then perfused with a protocol using the detergents Triton-X100 and SDS. Matrices were evaluated by histology (H/E, Sirius-Red, Alcian-Blue), immunohistochemistry (Collagen-IV, Laminin, Fibronectin) and evaluation of the remaining DNA and sGAG content. Statistical significance was defined as p < 0.05.
RESULTS: All three decellularization protocols were effective in removing cellular components as evaluated by histology. Macro- and microscopic conservation of the extracellular matrix could be observed. The extracellular matrix (ECM) was conserved with an intact pancreas micro-anatomy in all cases. Total DNA content significantly declined in experimental groups (p = 0.004). sGAG content appeared significantly higher in experimental groups (p = 0.027). No statistical difference was observed between the three experimental groups (DNA: p = 0.812; sGAG: p = 0.8980).
CONCLUSION: We present an effective protocol for perfusion decellularization of rat pancreas via three different routes. Our protocol is quick, reproducible and can be used for different settings. Further studies have to evaluate if the scaffold can be repopulated with cells to generate a functional and transplantable endocrine pancreas in-vitro.