Pancreatic islet transplantation for patients with diabetes mellitus has been hindered by the problem of donor shortage, as is the case for transplantation of other organs. Among several measures to overcome this problem, cell transplantation using xenogenic cell lines has been considered. For the treatment of diabetic patients, a murine pancreatic β-cell line MIN6 is a potential source of cell transplant. In order to restrict otherwise unlimited proliferation of transplanted MIN6 cells, cells are rendered to form spheroidal aggregates (SMIN6) on nonadherent culture dishes. SMIN6 stopped its growth around day 7 with a diameter of 220 ± 40 μm and kept its size almost constant at least until day 28. SMIN6 cells, however, had reduced responsiveness of insulin secretion to glucose concentration compared with MIN6 cells cultured in a monolayer. On the other hand, spheroid MIN6 cells formed in the presence of extracellular matrix gel (SMIN6E) possessed the capacity for glucose-dependent insulin secretion comparable with conventional MIN6 cells. SMIN6E encapsulated in agarose beads (SMIN6E-B) was also viable for at least 1 month in vitro with a constant diameter and favorable glucose responsiveness. The development of spheroid-type MIN6 may contribute to the future clinical application of MIN6 or other β-cell lines for treatment of diabetes mellitus.

stents, respectively (mean follow-up periods, 224 _+ 280 and 154 _+ 114 days, p > 0.05).The rates of stent patency (Figure ) and of survival derived from lifetable analysis (Kaplan-Meyer method, log-rank test) were similar in both groups.E1oo~ 80 ~ 2o 0 '~ Diamond stentJ

We examined the effectiveness of an improved version of a three-layer agarose microcapsule in islet xenotransplantation. The microcapsule is composed of a mixture of 5% agarose and 5% polystyrene sulfonic acid. The other two outer layers are polybrene and carboxymethyl cellulose. The agarose/polystyrene sulfonic acid membrane is for the purpose of immunoisolation, suppression of complement activity and reinforcement of the microcapsule. The polybrene layer suppresses the polystyrene sulfonic acid leakage by forming a polyionic complex at the surface of the agarose/polystyrene sulfonic acid membrane. The outermost layer, a carboxymethyl cellulose coating, improves the biocompatibility of the microcapsule. In vitro static incubation study showed that the insulin secretion from rat islets in microcapsules in response to 16.7 mM glucose stimulation was more than four times higher than that on 3.3 mM glucose stimulation (n = 8). In an in vivo study, 500 rat islets in microcapsules were xenogenically implanted in the abdominal cavity of mice with streptozotocin-induced diabetes. The graft survival times ranged from 2 to 5 mo, the average being 75 days (n = 5). Our results demonstrate that the improved version of the three-layer agarose microcapsule can effectively prolong the xenograft survival time without employing immunosuppressants, suggesting that this microcapsule could provide a promising biohybrid artificial pancreas for future clinical applications.

We describe a simple stationary digestion method of islet isolation and separation by various density gradients from monkey pancreas (Macaca radiata radiata). Effective method, different types and concentrations of collagenase were standardized. Sigma type XI collagenase yielded >1000 islets/gram pancreas at the concentration of 4 mg/ml and 3 ml Hank’s/gram pancreas. Slow digestion with less concentration of collagenase was suitable for monkey islet isolation. Discontinuous density gradients of bovine serum albumin (BSA) and dextran were compared with standard Ficoll for separation of islets. Islet yield (1038±81), insulin secretory response (stimulation index, S.I.11) and histological examination revealed dextran gradients were more appropriate for monkey islets when compared to BSA and Ficoll. Insulin secretory characteristics of monkey islets were studied by exposing them to low and high concentrations glucose (S.I.11.5), arginine (S.I.4.2), leucine (S.I.2.3) and tolbutamide (S.I.1.7). The results indicated that the magnitude of glucose induced insulin secretion of monkey islet is about half as that of rat and mouse islets. However, it is higher than that of porcine and bovine islets. In conclusion, the knowledge of insulin secretory ability of Indian bonnet monkey islets together with the techniques of isolation and separation are useful tool for diabetic research especially islet transplantation.

Endothelin-1, a 21-residue peptide isolated from vascular endothelial cells, has a broad spectrum of actions. To clarify the involvement of endothelin-1 in acute pancreatitis, we examined the effects of endothelin-1 and its receptor antagonist BQ-123 on cerulein-induced pancreatitis in rats. Rats were infused intravenously with heparin-saline (control), endothelin-1 (100 pmol/kg/hr), cerulein (5 µg/kg/hr), or cerulein plus endothelin-1 for 3.5 hr. In another experiment, cerulein or cerulein plus BQ-123 (3 mg/kg/hr) was infused. Infusion of cerulein caused hyperamylasemia and pancreatic edema. Endothelin-1, when infused with cerulein, decreased the extent of pancreatic edema with a significant increase in the pancreatic dry- to wet-weight ratio. Histological changes induced by cerulein were markedly attenuated when endothelin-1 was given with cerulein. In contrast, endothelin-receptor blockade with BQ-123 further augmented pancreatic edema caused by cerulein. The extent of inflammatory cell infiltration was greater when BQ-123 was given with cerulein. Endothelin-1 or BQ-123 had no influence on hyperamylasemia. This study suggests that endothelin-1 has protective effects on experimental acute pancreatitis.

Objective: To use mice to examine the effects of cyclosporine and tacrolimus (FK 506) on two forms of acute pancreatitis often seen after clinical organ transplantation.Methods and Design: In the first experiment, male CD-1 mice received cyclosporine (10 mg/kg), tacrolimus (0.32 mg/kg), or saline solution (control) subcutaneously once a day for 10 days. On the 11th day, acute edematous pancreatitis was induced by ceruletide (cerulein). In the second experiment, female ICR mice were fed with a choline-deficient, ethionine-supplemented (CDE) diet for 72 hours to induce necrotizing pancreatitis. After 30 hours on the CDE diet, the mice received cyclosporine (10 mg/kg), tacrolimus (0.32 mg/kg), or saline solution (control) subcutaneously twice daily for 3 days.Results: The pancreatic dry-to-wet weight ratios after ceruletide injections significantly decreased in mice treated with cyclosporine but did not with tacrolimus. Cyclosporine also significantly increased serum amylase levels, but tacrolimus did not. Cyclosporine or tacrolimus alone did not produce pancreatitis. In the CDE diet groups there was a significant difference in survival among the cyclosporine-treated, the tacrolimus-treated, and the control groups.Conclusions: Cyclosporine or tacrolimus given alone does not induce acute pancreatitis. In contrast, cyclosporine can adversely affect the course of acute edematous pancreatitis, and both immunosuppressants may worsen the survival of mice with acute hemorrhagic necrotizing pancreatitis. This study also demonstrated that the deteriorating effect of tacrolimus is less potent than that of cyclosporine.

This study purposed to examine the effects of cyclosporin A (CsA) and FK506 on edematous pancreatitis induced by caerulein (CER), and it demonstrated that CsA alone accelerated the progress of pancreatic edema, and enhanced CER-induced hyperamylasemia. Moreover, CsA made the reduction of pancreatic dry/wet weight ratio and the increase of serum amylase levels more pronounced. Neither CsA nor FK506 when administered singly produced edema or hyperamylasemia. We concluded that neither CsA nor FK506 given alone will induce acute pancreatitis. By contrast, both immunosuppressants can adversely affect the course of acute edematous pancreatitis. The results imply that the deteriorating effect of FK506 is less potent than that of CsA.

Cionin is a protochordean octapeptide similar to CCK and gastrin, and has two sulfated tyrosyl residues in its CCK- and gastrin-like positions: the 7th and 6th amino acids from the C-terminus, respectively. Cionin and three kinds of derivatives that lack either or both sulfations were synthesized to investigate the differential roles of these sulfations in the CCK-like effect of cionin. We examined the effect of these peptides and synthetic CCK-8 on exocrine pancreas (protein secretion and tissue blood flow) in anesthetized dogs (n=5) under continuous secretin infusion (0.5 mu g/kg/hr). Cionin-A, in its original form, and cionin-B, with CCK-like sulfation alone, were found to be as potent as CCK-8. Cionin-C with gastrin-like sulfation alone proved to be less potent than these three peptides but more potent than cionin-D minus either sulfation. The increment of protein secretion (mg/10 min/kg body weight) and the percent increase in tissue blood flow in response to 200 pmol/kg of the peptides were: (1) cionin-A: 8.2 (1.0)** and 107(22)%*; (2)cionin-B: 9.1 (1.0)** and 163 (60)%*; (3) cionin-C: 2.8 (0.6)* and 84 (29)%; (4) cionin-D: 0.7 (0.2) and 23(5)%; and (5) CCK-8: 8.5 (1.0)** and 110 (13)%*. Values are mean (SEM). ** and * denote significant defferences from cionin-C and cionin-D, respectively. Accordingly, CCK-like sulfation is definitively important, while gastrin-like sulfation is less important but partly contributory to the CCK-like effect of cionin.