IECM (EA4644 University/Oniris, USC1383 INRA)
Characterization of Neu5Gc and α1-3 Gal double knockout pig pancreatic islets for type 1 diabetes cell therapy.
Apolline SALAMA2, Xavier LEVEQUE1, Mathilde MOSSER1, Sylvie POGU1, Jérôme ABADIE3, Olivier GAUTHIER6, Ludmilla LEBERRE2, Sophie COCHON2, Andrea PEROTA4, Irina LAGUTINA4, Roberto DUCHI4, Giovanna LAZZARI4, Jérémy HERVOUET2, David MINAULT2, David H. SACHS7, Sophie BROUARD2, Gilles BLANCHO2, Cesare GALLI4,5, Jean-Paul SOULILLOU2, Jean-Marie BACH1.
1IECM, EA 4644 University/Oniris, USC1383 INRA, University/Oniris/INRA, Nantes, France; 2ITUN, UMR 1064, University/INSERM, Nantes, France; 3AMaROC, Oniris, Nantes, France; 4Laboratory of Reproductive Technologies, Avantea, Cremona, Italy; 5Department of Veterinary Medical Sciences, University, Bologna, Italy; 6Centre de Recherche et d'Investigation Préclinique, Oniris, Nantes, France; 7Center for Transplantation Sciences, Massachusetts General Hospital, Boston, MA, United States
Type 1 diabetes is a serious chronic autoimmune disease in children and young adults. The allotransplantation of pancreatic islets is an alternative to insulin therapy. Today, xenotransplantation of neonate or adult pig pancreatic islets remains the most promising alternative to allotransplantation in order to address the shortage of human donor organs. Evidence has been established that islet encapsulation can diminish the need for immunosuppressive treatments and their associated deleterious side effects. However, strong humoral immune responses to pig antigens have been demonstrated to be responsible for graft rejection, especially against porcine galactose-α(1,3)-galactose (α1-3 Gal) and the glycolyl form of neuraminic acid Neu5Gc, which are major pig xenoantigens recognized by human (natural) antibodies. We obtained double knockout pigs (DKO) for the cytidine monophosphate-N-acetylneuraminic acid hydroxylase (CMAH) and galactosyltransferase1 (GT1) enzymes, thus lacking Neu5Gc and α1-3 Gal determinants [S. Conchon et al. 2013]. As CMAH knockout mice showed a pancreatic β-cell dysfunction and a strong decrease in islet number [S. Kavaler et al. 2011], we evaluated the potential of pancreatic islets from these DKO pigs for transplantation. At necropsy, the pancreas of DKO (n=3 vs 2 wt controls) adult pigs showed neither macroscopic nor histological modifications. Changes in blood glucose levels after intravenous administration of glucose (glucose tolerance test) showed no difference in glycaemic regulation in DKO pigs (n = 4) compared to wt controls (n = 2). Isolation of islet cells from neonate piglets (less than 2 weeks of age, n=5) yielded IEQ/g pancreas similar to those obtained for wt control pigs. In vitro, pancreatic islets isolated from neonate piglets were functional and exhibited insulin secretion stimulated by glucose and theophylline. Non-GAL and non-Neu-5GC anti-pig antibody responses will be analyzed following a subcutaneous graft of cellulose-encapsulated neonate pig pancreatic islets in mice. Antibody responses specific for Neuc-5GC will be tested post-graft in CMAH KO mice.
In conclusion, our work provides preliminary ex vivo and in vitro evidence that pancreatic islets from DKO pigs are functional and should be tested in preclinical assays of cell therapy for type 1 diabetes.
Pays de la Loire Region, France. European Center for Transpl. and Immunotherapy Sc. (ECTIS IHU), Nantes, France. French National Research Agency (ANR), France.
 S. Conchon et al., Generation of CMAH-/- piglets on GAL-/- genetic background, IXA2013, Xenotranspl.
 S. Kavaler et al., FASEB J. 2011 Jun; 25(6): 1887–1893.
11:00 - 12:30
|Alternative Sources of Beta Cells: Xenoislet, Stem Cells, Tissue Engineering||Characterization of Neu5Gc and ?1-3 Gal double knockout pig pancreatic islets for type 1 diabetes cell therapy.||Room 110|