The outcome of islet isolation is considered uncertain because of the large variability of islet and insulin yield, but comparison of the isolated and native islet population has not been attempted. We therefore addressed the efficacy of collagenase digestion, and density gradient purification of islets from the splenic dog pancreas (n = 31) by morphometry of the islet volume and size distribution, and by extraction of insulin and amylase, in samples from the pancreas, the digest, and gradient fractions. In contrast to a ~90% recovery of pancreatic insulin and amylase after digestion, islet yield amounted to 50% of the islet content of the pancreas. After density separation, islets were mainly found in the purified fractions, while half of the recovered insulin was located in the acinar fraction of the gradients — indicating a substantial proportion of islets entrapped in acinar fragments. The islet and insulin content of the pancreas correlated well with islet and insulin yield after digestion (r = 0.7, p < .0001). The insulin content of digest suspensions did neither correlate with islet nor insulin recovery in the purified fraction of the gradients (r = 0.4) as opposed to the islet content of digest suspensions, which correlated with both (r = 0.7, p < .0001). After density separation near 100% purity was obtained, and no loss of insulin from isolated islets was demonstrated by extraction and microscopy. Size distributions of native and isolated islets demonstrated no fragmentation. We conclude that the variability of isolation outcome may be attributed to a large extent to the variability of the native endocrine pancreas. Isolation efficacy was best documented by morphometry, because insulin extraction did not discriminate between free and entrapped islets. However, assessment by both morphometry and extraction allowed the quantitation of entrapped islets, and demonstrated preservation of β-cell granulation. Similar studies should facilitate the analysis of other factors affecting islet isolation in man.

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Clinical human islet transplantation programmes are considerably hampered by the variability of islet isolation outcome. The effects of the islet content of the pancreas and other donor-related variables on isolation outcome have not been evaluated systematically so far-either in large animals, or in man. We studied the impact of interindividual differences in age, body weight and pancreatic islet content on the outcome of collagenase isolation of islets from the splenic pancreas of beagle dogs (n =31). The islet volume of the splenic pancreas amounted to a mean (+ SEM) 15.7 + 0.9 gl per gramme pancreas, and varied threefold (from 8.4 to 27.3 gl). Isolated islet yield was 7.6 +0.7 btl/g and varied nine-fold (1.8-16.3 pl). Animals also varied in age eight-fold (8-67 months) and body weight twofold (8.6-18.3 kg). Differences in body weight and age explained 60 % of variance in the fractional islet volume of the pancreas and 50 % of the variance in islet yield (p < 0.001). Fractional islet volume of the splenic pancreas also explained 50 % of the variance in islet yield (p < 0.001). We conclude that the outcome of islet isolation may be predictable after controlling for the variable islet content of pancreases, and other donor-related variables, and suggest that similar studies should be done in man. [Diabetologia (1994) 37: 111-114]

Adequate metabolic control is central to the concept of islet transplantation, but has received limited attention. We studied metabolic control in 8 dogs at 6-9 months after intrasplenic autografting of approximately 25% of the normal mass islets--as compared to 30 controls. A similar posttransplant reduction to approximately 25% of the insulin secretory capacity as assessed by intravenous arginine stimulation during 35 mM glucose clamps, mirrored the reduction of the islet mass. Postprandially, in contrast, the insulin response had increased to 140% in the islet recipients--with a concomitant rise of glycemia to approximately 8.5 mM. Posttransplant, the insulin secretory capacity correlated both with the index of insulin action (which averaged 55% of the normal value) as assessed by euglycemic hyperinsulinemic clamps, and--inverse--with the postprandial glucose excursions. Because insulin action did not correlate with postprandial glucose, the insulin secretory capacity appears to be the primary determinant of the impaired glucose tolerance. Marked postprandial hyperglucagonemia, and a virtually absent pancreatic polypeptide response in the grafted animals, may also have contributed to the impaired glucose tolerance. Posttransplant, infusion of a physiological dose of the gut hormone glucagon-like peptide-1 during 8.5 mM glucose clamps--mimicking the postprandial glycemia--potentiated glucose-stimulated insulin 175%. Thus, after transplantation of a suboptimal islet mass, postprandial glucose excursions are restrained by hyperglycemic potentiation of the entero-insular axis, which may account for the difference in the insulin response to the intravenous and oral challenges. Because, the insulin secretory capacity reflects the islet mass and appears to be the major determinant of glucoregulation, transplantation of a larger islet mass may allow near-normal glycemic control.

Our aim was to isolate and determine the contribution of partial pancreatectomy, systemic delivery of pancreatic hormones, and duct obliteration to glucose regulation after segmental pancreas transplantation in dogs. Fasting, postprandial, and intravenous glucose-stimulated glucose, insulin, glucagon, pancreatic polypeptide (PP), and cholecystokinin (CCK) and intravenous bombesin-stimulated PP levels were studied in beagles at three successive intervals in a crossover design. The first was 6 wk after partial (∼70%) pancreatectomy with intact regular enteric exocrine drainage from the duodenal pancreatic remnant, the next was 2 wk after venous transposition with systemic delivery of pancreatic hormones, and the third was 6 wk after in situ duct obliteration of the remnant. With partial pancreatectomy, K values were modestly diminished (30%), and a concomitant reduction of second-phase intravenous glucose-stimulated insulin release was observed. Other parameters were not significantly affected. Venous transposition doubled peripheral plasma levels of insulin under all conditions. Fasting glucose, PP, and CCK levels decreased slightly. Other parameters were not affected. Duct obliteration of the systemic draining pancreatic remnants seriously impaired glucose sensitivity, resulting in a 50% reduction of K values and fasting and sustained postprandial hyperglycemia (∼8 mM) and a 70–50% reduction (acute and overall responses, respectively) of intravenous glucose-stimulated insulin. Fasting hormone and postprandial insulin, glucagon, and CCK levels were not affected. The postprandial PP response was severely reduced, and bombesin-stimulated PP release was abolished by duct obliteration. We conclude that histological changes associated with duct obliteration are the major determinants of glucose regulation in segmental pancreas transplantation. The defective acute insulin response to intravenous glucose is probably secondary to obliteration-induced intrinsic denervation of the islets, as supported by an abolished PP response to bombesin. The difference in the effect of duct obliteration on the insulin response to intravenous glucose and a meal may be related to the postprandial activation of the enteropancreatic axis.