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P.169 Decellularized cotyledons isolated from human placenta used like a scaffold for the generation of a human bioartificial pancreas

Kevin Bellofatto, Switzerland

Post-doctoral assistant
Dpt of Surgery
Université de Genève

Abstract

Decellularized cotyledons isolated from human placenta used like a scaffold for the generation of a human bioartificial pancreas

Kevin Bellofatto1,2, Fanny Lebreton1, Charles-Henri Wassmer2, Masoud Hasany1,2, Reine Hanna1, Rahul Khatri1,2, Laura do Mar Fonseca1,2, Lisa Perez2, Caroline Rouget2, David Cottet-Dumoulin1, Géraldine Parnaud2, Domenico Bosco1, Thierry Berney2, Ekaterine Berishvili1,3.

1Surgery, Université de Genève, Geneva, Switzerland; 2Surgery, Hopitaux Universitaires de Genève, Geneva, Switzerland; 3Institute of Medical Research, Ilia State University, Tbilisi, Georgia

Background: Replacing damaged organs with biological substitutes capable of protecting the islets and facilitating vascularization is a great objective in the field of islet transplantation. A decellularized placenta includes a large number of cotyledons with a conserved vessel structure of the native organ. Our goal is to obtain a perfect decellularization protocol to generate pre-vascularized organoids by recellularizing this ECM with HUVECs and pancreatic islets.
Methods: After blood removal, cotyledons were dissected from the placentas and decellularized using a bioreactor. Cell removal was assessed by histology and quantification of residual DNA. Presence of structural proteins and ECM structure were analyzed using SEM, CT scan and mass spectrometry. Recellularization protocols were conducted, with HUVECs or BOECs as endothelial cell sources, and with Ins-1E cells or rat islets as insulin secreting cell sources. Function of recellularized cotyledons was assessed in vitro with glucose stimulated insulin secretion tests (GSIS). To assess in vivo biocompatibility and function of the scaffolds, we transplanted in diabetic NSG mice. Glycaemia was measured every day to monitor normalization of blood glucose levels.
Results: Our protocol led to successful decellularization, as evidenced by the absence of cells and the preserved ECM structure. Moreover, DNA quantification did not reveal any residual DNA. Quantification of GAG and hydroxyproline, and mass spectrometry analysis show that structural proteins are conserved. SEM and CT scan images revealed that the ECM structure was preserved after the decellularization protocol. Cells after recellularization showed a good vascularization after 7 days. The GSIS test shows a perfect organ response in the production of insulin.
Conclusions: The decellularized cotyledon is the perfect scaffold to reproduce a prevascularized insulin-producing organ, which allows transplanted cells to survive during the peri-transplantation period.