Background & aims: To be clinically efficient, beta-cell replacement therapies such as pig islet xenotransplantation must ensure sufficient insulin secretion from grafted islets. While protection from host immune reactions is essential for islet engraftment and their subsequent functioning, intrinsic physiological properties of used cells are also a key factor. We have previously shown that islets from transgenic pigs expressing a dipeptidyl peptidase-resistant form of glucagon-like-peptide-1 (GLP-1) and a constitutively activated form of type 3 muscarinic receptor (M3R) in their beta-cells have greatly improved insulin secretory response to glucose stimulation that is otherwise 4-10 times lower than human islets. Our aim in the current study was to use such modified islets to treat insulin-dependent diabetes in a xenotransplantation model.

Material & methods: Pancreatic islets were isolated from transgenic and wild-type neonatal piglets and kept in culture in a maturation medium for 8 days. Glucose-stimulated insulin secretion from mature islets was evaluated in vitro during dynamic perifusion experiments. Streptozotocin-induced diabetic immunodeficient NSG mice received 2500 IEQ transgenic or control islets under the left kidney capsule. Blood glucose was monitored weekly and porcine c-peptide was measured monthly to evaluate graft function. Normoglycemic mice had their graft-bearing kidney removed 9 months post-implantation and follow-up continued for hyperglycemia to occur. Sections of graft bearing kidneys and of  recipient micee pancreas were stained for insulin and glucagon to evaluate graft maturation and exclude endogenous beta-cell regeneration.

Results: Islets isolated from transgenic InsGLP-1M3R piglets secreted 5.5 to 7.5-fold more insulin compared to controls upon stimulation with 15 mM glucose and had G15/G1 stimulation ratio of 3.7 to 7.5 compared to 2.6 for wild type islets. 80% of InsGLP-1M3R islet mice recipients became normoglycemic within 4 weeks vs. 11% of wild-type islet recipients. Porcine c-peptide could be detected in mice blood by the 3rd week post-implantation and gradually increased throughout the 9-month follow-up period with the highest values recorded in recipients of transgenic islets. Graft removal resulted in hyperglycemia in all normoglycemic mice. Immunohistological analysis showed a significant increase of insulin-positive cells within grafted islets whereas no insulin could be detected in the pancreas 9 months following streptozotocin treatment.     

Conclusion & perspectives: InsGLP-1M3R neonatal pig islets showed significantly improved insulin secretion both in vitro and in vivo. They were found to be more efficient in treating insulin-dependent diabetes in a xenotransplantation immunodeficient model compared to non-modified islets. These islets sourced from a specific pathogen-free herd represent a very promising beta-cell replacement therapy alternative for use in research and in the clinic.