Unexpected immunoresponse to Gal and APA antigens in diabetic type 1 patients receiving neonatal pig islets after 6 years.

Cotransplantation of porcine islets and Sertoli cells into preimplanted subcutaneous devices improve metabolic control in type 1 diabetic patients, and survive grafted for more than 4 years. We report here, further assessment of the endocrine and porcine nature of the surviving cells and the immune responses elicited toward Gal alpha(1,3)-Gal beta(1,4)-GlcNAc (Gal) antigen in patients who received a second and third transplants. No immunosuppressive drugs were administered. We were able to immunostain insulin- and glucagon-positive cells in all biopsies of patients and Sertoli cell markers in 60.9% of biopsies. Additionally, all biopsies tested, amplified the porcine COII gene. Patients demonstrated an increase in antipig antibodies in response to the first transplant with a decreasing response toward the second and third transplants. In all transplants, the IgG levels promptly returned to basal values after 3-4 months. The long-term survival of porcine cells and the reduced humoral immune response to multiple transplants indicate a form of tolerance. We have not been able to find CD25-positive cells, indicating that it is probably an immune accommodation of the graft.

Xenotransplantation of porcine neonatal islets of Langerhans and Sertoli cells: a 4-year study.

OBJECTIVE: Porcine islets of Langerhans for xenotransplantation into humans have been proposed as a solution to the shortage of human donors. Rejection is one of the main constraints. This study presents the results of a clinical trial using a novel method for transplanting and immunoprotecting porcine islets in type 1 diabetic patients. DESIGN: A 4-year follow up of a clinical trial involving 12 patients, with no immunosuppressive drugs at any point. Eleven age matched untransplanted diabetics served as controls. METHODS: We have developed a procedure for protecting neonatal porcine islets by combining them with Sertoli cells and placing them in a novel subcutaneous autologous collagen-covered device. RESULTS: In the patients in the treatment group, no complications arose and no porcine endogenous retrovirus infection was detected. Half of the patients showed a significant reduction in insulin requirements compared with both their pre transplant levels and controls, and this reduction was maintained for up to 4 years. Two patients became insulin-independent for several months. Porcine insulin was detected in three patients' sera following glucose stimulation up to 4 years post transplant. Three years post transplant, one of four devices was removed from four patients, and the presence of insulin-positive cells in the transplant was demonstrated by immunohistology in all 4 patients. CONCLUSIONS: Long-term cell survival with concurrent positive effects on metabolic control are possible by this technique.

Improved method for isolation of porcine neonatal pancreatic cell clusters.

UNLABELLED: Based on the described methods for the isolation of neonatal pancreatic cell clusters (NPCCs), we have developed modifications in order to improve their quality, functionality, and process reproducibility in the isolation technique, for potential use in research. In addition, we indicate techniques for describing yield, functionality, viability and purity of the NPCCs. METHODS: Purity of the NPCCs was determined through dithizone staining and subjected to image analysis. Viability and apoptosis was measured through flow cytometry with propidium iodide and annexin, respectively. NPCC functionality was measured through a static glucose stimulation test. RESULTS: We developed a high-yield reproducible technique that had 81 279.55 +/- 18 257.05 IEQ/g of pancreas at 4 days of culture, with a 94% viability and an 88 +/- 2.73% purity. Stimulation index from the glucose stimulation test was >10. CONCLUSION: The technique allowed us to obtain NPCC with optimal viability, functionality, purity, and endurance for use in research.