In Vitro Evaluation of a Bioartificial Pancreas Under Various Hemodynamic Conditions

A bioartificial pancreas in which isolated islets of Langerhans are placed between two polyacrylonitrile membranes, blood circulating successively above the upper and below the lower membranes following a U-shaped circuit, has been developed. The two parts are connected by an outer loop consisting of a thin tubing. The length of this tubing determines the magnitude of the flow rate of blood through the device. The aim of this work was to determine experimentally the optimal configuration of the system containing isolated rat islets and a Krebs buffer circulating through the device. The amount of insulin released by the bioartificial pancreas was determined during a 20-mM square-wave glucose stimulation. First, the inlet pressure was set at 100 mm Hg, and the effect of the length of the tubing was investigated with two devices perfused simultaneously. For a short tubing (flow rate, 20 ml/min), a sharp increase in insulin release in response to glucose was observed; it increased within 4 min from 217 +/- 50 to 761 +/- 237 microU/500 islets/min (p less than 0.05), the peak value being reached at 11 +/- 2 min following the beginning of the stimulation. For a long tubing (flow rate, 3 ml/min), the increase in insulin release was more sluggish. It increased from 133 +/- 53 to 222 +/- 43 microU/500 islets/min at 4 min, the peak value being reached only at 20 +/- 3 min. These data are consistent with a more efficient diffusional transfer of insulin in the case of the high circulating flow.(ABSTRACT TRUNCATED AT 250 WORDS)     

Glucose-Insulin Kinetics of a Bioartificial Pancreas Made of an AN69 Hydrogel Hollow Fiber Containing Porcine Islets and Implanted in Diabetic Mice

The goal of this study was to determine whether porcine islets encapsulated in hollow fibers made of AN69 copolymer can correct hyperglycemia in diabetic mice and provide normal tolerance to a glucose challenge. In vitro perifusion of hollow fibers demonstrated the rapid kinetics of insulin release in response to glucose. Two fibers containing islets were transplanted into the peritoneal cavity of each of 17 streptozotocin induced diabetic mice. In 11 mice, diabetes was reversed within 3 days with plasma glucose levels decreasing from 19.7 ± 0.9 (mean ± SEM) before implantation to 10.9 ± 0.8 mmol/L. Intraperitoneal glucose tolerance tests were performed in transplanted (n = 7), nondiabetic (n = 15), and diabetic mice (n = 6). A normal glucose pattern was observed in the transplanted diabetic mice. This was achieved in the presence of plasma insulin levels lower than those observed in control nondiabetic mice, suggesting the presence of a state of hypersensitivity to insulin, which was demonstrated in this model by exogenous insulin tolerance tests. In conclusion, encapsulation of islets suspended in ultraculture medium in biocompatible membranes of AN69 can provide xenograft survival, and complete normalization of glucose tolerance can be achieved.     

Absence of Effect of Heparin on Insulin Secretion

Bioartificial pancreatic devices containing isolated islets of Langerhans have been designed, in which the blood of the recipient circulates in contact with an artificial membrane, protecting the islets against immune rejection. This system assumes that heparin, required to prevent blood clotting, does not alter insulin secretion. However, heparin has been reported to inhibit in vitro insulin secretion by rat islets and to suppress in vivo insulin secretion in dogs. Therefore, the following evaluation was made on the effect of different heparin preparations on insulin secretion. (a) Isolated rat islets of Langerhans were perfused or incubated in the absence or presence of 20 micrograms/ml heparin; insulin secretion in response to a stimulation by glucose 20 mM was not altered by the presence of heparin. (b) Insulin secretion by an insulin-secreting cell line (RINm5F) in response to leucine and theophylline was not suppressed by heparin up to 100 micrograms/ml concentration. However, an inhibitory effect was observed at 200 micrograms/ml, which is 100 times higher than the heparin concentration commonly used for therapeutic use. (c) Neither in normal rats nor in dogs did heparin alter portal plasma insulin levels and the increase in plasma insulin following an intravenous injection of glucose. In conclusion, these data do not confirm the formerly observed inhibitory effect of heparin, which can therefore be used for the in vivo evaluation of a bioartificial pancreas.     

Evaluation of a Modified HTK Solution Containing the New Iron Chelator LK 614 in an Isolated Rat Liver Perfusion Model

Background: Cold storage-induced injury to donor organs remains a persistent problem in transplantation medicine. We here evaluated a modified HTK solution including, among others, a new, membrane-permeable iron chelator, LK 614, in the isolated perfused rat liver model. Methods: Rat livers were stored at 4°C for 24 hr in modified HTK solution, in modified HTK solution with LK 614 or in traditional HTK solution, and reperfused for 60 min at 37°C with Krebs-Henseleit buffer (KH buffer). Bile secretion and lactate dehydrogenase (LDH) release into the perfusate were measured, and hepatic microcirculation evaluated optically after the addition of trypan blue to the perfusate. Biopsies were evaluated by a pathologist blinded to the experimental conditions. Results: Compared with HTK-preserved livers, LDH leakage was significantly lower and bile secretion significantly higher in the modified HTK solution with the iron chelator (both p <. 05), while values for the modified solution without the iron chelator were in between (no significant difference to either of the two other solutions). The hepatic microcirculation was also preserved best in livers stored in the modified HTK solution with LK 614. Histomorphological investigation confirmed the improved preservation in the modified HTK solution with LK 614. Conclusions: These results suggest that livers might be better preserved in modified HTK solution containing the iron chelator LK 614 (and also, but less so, in the modified solution without the iron chelator) than in the original HTK solution, and such a modified solution should now be evaluated in an animal model of liver transplantation.