ADULT PORCINE ISLET TRANSPLANTATION FROM N-ACETYLGLUCOSAMINYL-TRANSFERASE-III (GNT-III) TRANSGENIC PIGS IN CYNOMOLGUS MONKEYS

Abstract

P1074 Introduction: Because of the severe shortage of human donor pancreases, pig islets has been considered as an attractive donor source. The findings of our previous in vitro study indicated that the level of expression of alpha-Gal by adult pig islets is very low, and that adult pig islets have non-alpha-Gal antigenicity, including H-D antigen, especially in terms of N-linked sugars. In addition, pig islets can be injured by both the classical and the alternative complement pathways in human serum. In the present study, we examined the issue of whether these humoral factors are involved in the early graft loss of islets in the monkey, using N-acetylglucosaminyltransferase-III (GnT-III) transgenic (Tg) pigs which revealed a significant down-regulation in xenoantigenicity and in cynomolgus monkeys.. Material and Methods: Preparation of pig islets. The pancreatic glands were removed from adult pigs. After removed, the gland was infused with a collagenase solution and distended. The gland was digested at 37 degrees C with static incubation. After digestion, purification was carried out by centrifugation on a Ficoll discontinuous gradient. The purified islets were cultured in Medium 199 supplemented with 10% porcine serum and antibiotics. GnT-III enzyme activity. GnT-III enzyme activity in pig islets was measured by high performance liquid chromatography (HPLC). Transplantation procedure. Recipient cynomolgus monkeys were injected with streptozotocin (STZ) at a dose of 150 mg/Kg/body weight (BW) 3 days before transplantation. Diabetes was then confirmed by checking the endogenous insulin production in the monkeys. After laparotomy, pelleted pig islets were transplanted under the left kidney capsule of the monkeys. Three monkeys were transplanted with islets from naive pigs (Group 1: naive), and five from GnT-III Tg pigs (Group 2: Tg). After transplantation, blood was sampled daily and plasma insulin was monitored to evaluate the graft function. Results: GnT-III was expressed at high levels in islets from Tg pigs (1628 684 p mol/h/mg protein) and xenoantigenicity, was reduced, while it was barely expressed in naive islets (10 p mol/h/mg protein). The survival time of the islets was estimated by the detection of insulin production in the recipient monkeys. Islets isolated from GnT-III Tg pigs survived longer than those from naive pigs in cynomolgus monkey, as follows: Group 1: < 1, < 1, and 3 day; Group 2: 1, 3, 4 and 5 day. Changes in anti-pig antibodies in the monkeys and histological analyses of the rejected islets are now under investigation. Conclusion: The data suggest that a reduction in xenoantigenicity by GnT-III can prolong the survival time of porcine islets, and that a humoral response is involved in the early rejection of porcine islets in the monkey.