An encapsulation system for the immunoisolation of pancreatic islets.

Over a thousand combinations of polyanions and polycations were tested to search for new polymer candidates that would be suitable for encapsulation of living cells. The combination of sodium alginate, cellulose sulfate, poly (methylene-co-guanidine) hydrochloride, calcium chloride, and sodium chloride was most promising. In parallel, a novel multiloop chamber reactor was developed to control the time of complex formation and to negate gravitational effects such as pancreatic islet sedimentation and droplet deformation during the encapsulation process. Encapsulated rat islets demonstrated glucose-stimulated insulin secretion in vitro, and reversed diabetes in mice. This new capsule formulation and encapsulation system allows independent adjustments of capsule size, wall thickness, mechanical strength, and permeability, which may offer distinct advantages for immunoisolating cells.

Permeability assessment of capsules for islet transplantation.

Despite considerable progress in the development of immunoisolation devices, the optimal permeability of such devices is not known. This limitation stems partly from deficits in knowledge about which molecules should be allowed to traverse the semipermeable membrane and which molecules should be excluded, and also partly from experimental obstacles that have prevented a systematic study of permeability. To determine the optimal permeability of immunoisolation devices, we have created a series of microcapsules (800 microM diameter) that span a broad range of molecular exclusion limits yet are identical in wall thickness and chemical composition. Capsule permeability was precisely defined by two complementary methods--size exclusion chromatography (SEC) and a newly developed methodology to assess permeability of biologically relevant proteins. The entry of interleukin-1 beta-125I was significantly delayed, but not prevented, when the capsule exclusion limit was decreased from 230 kD to 3.2 kD, as determined by SEC with dextran standards. The influx of IgG was as predicted, based on the viscosity radius R eta of IgG and the capsule exclusion limit defined by SEC. Glucose-stimulated insulin secretion by encapsulated pancreatic islets did not differ as capsule permeability was decreased from a molecular exclusion limit of 230 kD to 120 kD. These studies should assist in the design of immunoisolation devices by defining the permeability optimal for cell function and also should be applicable to any cell type or immunoisolation device.

Time course of adaptation to a high-fat diet in obesity-resistant and obesity-prone rats.

The purpose of the present study was to characterize the time course of adaptation (i.e., circulating metabolites and hormones, fat pad mass, lipoprotein lipase) to a high-fat diet in obesity-prone (OP) and obesity-resistant (OR) male Wistar rats. Delineation of OP and OR was based on body weight gain (upper tertile for OP; lower tertile for OR) after 1 wk on a high-fat diet (60% of kcal from corn oil). Rats were killed after 1, 2, or 5 wk of the dietary period. Increased body weight and percent body fat in OP rats at 1 wk could not be accounted for by increased retroperitoneal or epididymal fat pad weight. Plasma nonesterified fatty acids and triglycerides, as well as blood concentrations of glucose, lactate, and glycerol, were similar throughout the study. Plasma insulin was significantly greater in OP vs. OR rats and low-fat diet (LFD; 20% of kcal from corn oil) controls at 5 wk only, and blood beta-hydroxybutyrate (mM) was significantly higher in OR compared with OP and LFD rats at 1, 2, and 5 wk. Lipoprotein lipase mRNA and activity were significantly greater in epididymal fat pad and significantly lower in gastrocnemius muscle of OP vs. OR rats at 1 wk. Results suggest that early (i.e., 1 wk) differences in body weight and fat weight between OP and OR rats are not due to fat deposition in retroperitoneal or epididymal fat depots, and tissue-specific changes in LPL (increase in epididymal fat pad and decrease in gastrocnemius muscle) that occur in OP compared with OR rats after 1 wk on a high-fat diet provide a metabolic environment favoring fat storage.

Involvement of liver and skeletal muscle in sucrose-induced insulin resistance: dose-response studies.

The ability of dietary sucrose to induce insulin resistance independent of changes in body weight is controversial. In the present study male rats were fed a high-starch (ST) diet (starch 68% of total kcal) ad libitum for 2 wk and then were fed equicalorically either the ST diet or a high-sucrose (SU) diet (sucrose 68% of total kcal) for 8 wk. Euglycemic, hyperinsulinemic (0, 1.2, 4.1, 8, 15 mU.kg-1.min-1, n = 6-8/group per dose) clamps were then used to establish dose-response relationships for glucose kinetics and metabolism. Body weight (513 +/- 3 g) and composition were similar between groups after the 8-wk dietary period. Glucose infusion rates (GIR; mg.kg-1.min-1) were significantly less in SU (0.9 +/- 5.8 +/- 0.6, 14.8 +/- 1.3, and 18 +/- 1.1) than in ST rats (4.1 +/- 0.9, 12.3 +/- 1.2, 22.6 +/- 1.5, and 25.9 +/- 1.8) at 1.2, 4.1, 8, and 15 mU.kg-1.min-1, respectively. Impaired suppression of endogenous glucose production accounted for 46, 43, 23, and 0% of the reduction in GIR in SU rats at 1.2, 4.1, 8, and 15 mU.kg-1.min-1, respectively. Despite basal hyperinsulinemia (38 +/- 2 microU/ml in SU vs. 26 +/- 2 microU/ml in ST rats), liver phosphoenolpyruvate carboxykinase (PEPCK) activity was 50% higher in SU than in ST rats and remained elevated in SU rats (by 30-40%) at the two lower insulin doses. No skeletal muscle glycogen accumulation occurred in SU rats at any of the insulin doses, and glycogen synthase I activity was significantly lower in SU rats at the two highest insulin doses.(ABSTRACT TRUNCATED AT 250 WORDS)

Skeletal muscle glucose metabolism in obesity-prone and obesity-resistant rats.

Ad libitum access to a high-fat (HF) diet produces a wide range of weight gain in rats. Rats most susceptible to weight gain on such a diet (obesity prone; OP) are more insulin resistant after 4-5 wk of diet exposure than are those most resistant (obesity resistant; OR) to weight gain. To investigate whether skeletal muscle glucose metabolism contributes to insulin resistance in this model, insulin-stimulated glucose metabolism was assessed in the perfused hindquarter of rats exposed to either a low-fat (LF, n = 6) or HF diet for 5 wk. Delineation of OP (n = 6) and OR (n = 6) rats was based on body weight gain. OP rats gained 60% more body weight while eating only 10% more energy than OR rats. Single-pass perfusions were carried out for 2 h in the presence of glucose, insulin, and [U-14C]glucose. Insulin-stimulated glucose uptake (mumol.100 g-1.min-1) was 14.2 +/- 0.9 in LF, 11.1 +/- 0.8 in OR, and 6.2 +/- 0.6 in OP. Glucose oxidation (mumol.100 g-1.min-1) was 1.7 +/- 0.3 and 1.2 +/- 0.3 in LF and OR, respectively, but was 0.2 +/- 0.1 in OP. Net glycogen synthesis was significantly reduced in OP compared with OR and LF despite similar glycogen synthase I activity. Muscle triglyceride concentration was not significantly different in OR and OP rats. These results demonstrate significant defects in skeletal muscle glucose uptake and disposal in rats most susceptible to HF diet-induced obesity. Clearly, the heterogeneous response to a HF diet involves not only body weight gain but also skeletal muscle fuel metabolism.