Human islet isolation in 104 consecutive cases. Factors affecting isolation success.

One of the major steps toward successful islet transplantation for the treatment of type diabetes is to obtain islets of sufficient number and viability. Using a standardized method of isolating islets, the goal of this study was to analyze the factors influencing the outcome of islet isolation. A total of 104 cadaveric human pancreata were processed for islets by the same team. Data from the islet-processing charts were reviewed retrospectively. The two endpoints were the recovery of islets, viable after 2 days of culture (group V = viable, group NV = nonviable) and the islet yield. Viable islets were recovered in 61% of cases (n = 63). Minimal blood glucose recorded during hospitalization was very significantly lower in group V (124 +/- 5 vs. 148 +/- 9, P = 0.01). Lack of significant medical history in the donor was associated with better viability as compared with various donor predispositions (chi-2 4.21, P = 0.04). Cold ischemia time (8.1 +/- 0.5 hr in group V vs. 9.8 +/- 0.9 hr in group NV, P = 0.07) and collagenase lot (5 lots tested, chi-2 13.1, P = 0.01) also affected the recovery of viable islets. Hospital time was shorter in group V (65.3 +/- 6.8 vs. 80.9 +/- 17.9 hr, P = 0.35). Multivariate logistic regression analyses of viable islet recovery identified minimal blood glucose (P = 0.03) and collagenase lot (P = 0.06) as the most significant risk factors. However, the best multivariate predictive model--which includes blood glucose, collagenase lot, donor age and surgical procurement team--correctly predicted 66.2% of cases only. Multivariate analysis of final islet yield designed hospitalization length, cardiorespiratory arrest, surgical procurement team, and collagenase lot as the best predictors. These data obtained in a large series of pancreata emphasized several donor and technical factors that should target the attention of islet transplant researchers in order to improve islet yield and viability.

Relation between intragastric bile acid concentration and mucosal abnormality in the stomach after vagotomy and gastroenterostomy for duodenal ulcer.

A high incidence of possible premalignant conditions of the gastric mucosa has been reported in the postoperative stomach and it has been suggested that bile plays a role in their pathogenesis. Fifty eight patients who had had a vagotomy and gastrojejunostomy between 1957 and 1967 underwent endoscopy and biopsy. Subsequently, samples of fasting and postprandial gastric juice were taken from all patients for analysis of total bile acid. The patients were divided into three histological groups: group 1 (24 patients) had gastritis or mucosal atrophy but no more than mild dysplasia; group 2 (23 patients) had intestinal metaplasia in one or more biopsies; group 3 (11 patients) had moderate or severe dysplasia in one or more biopsies. Group 2 and group 3 patients had higher fasting intragastric bile acid concentrations than group 1 (p less than 0.01 in both cases). There was no difference between fasting bile acid concentrations in groups 2 and 3. In the postprandial phase groups 2 and 3 also had higher peak intragastric concentrations than group 1 (p less than 0.01 in both cases). Again, there was no significant difference between groups 2 and 3. It is concluded that there is a relation between mucosal abnormality in the postoperative stomach and intragastric bile acid concentration. The possible aetiological link between bile acid and these mucosal abnormalities is discussed.

Measurement of total bile acids in gastric juice.

An established method for the assay of total bile acids was validated for use in fasting and post-prandial gastric juice samples. Fasting and post-prandial intragastric bile acid concentrations were measured in 29 healthy volunteers, 15 patients after vagotomy and gastrojejunostomy (V and GJ) and 15 patients after vagotomy and pyloroplasty (V and P). Healthy female volunteers had higher post-prandial bile acid concentrations than age matched healthy males (p less than 0.02). Patients with V and GJ had higher fasting and post-prandial bile acid concentrations than age and sex matched control subjects (p less than 0.01). Patients with V and P had higher bile acid concentrations than control subjects only in post-prandial samples (p less than 0.05).

Relation between gastric histology and gastric juice pH and nitrite and N-nitroso compound concentrations in the stomach after surgery for duodenal ulcer.

Formation of N-nitroso compounds in gastric juice has been implicated in the pathogenesis of cancer in the stomach after operation. Gastric juice was aspirated from 85 subjects: 23 were controls, 51 had previously undergone vagotomy and gastrojejunostomy, and 11 had previously undergone vagotomy and pyloroplasty. The gastric juice samples were analysed for pH, nitrite, and total N-nitroso compounds. A significant correlation was found between pH and nitrite concentration (p less than 0.01). No significant correlation was found between pH and total N-nitroso compound concentration or between nitrite and N-nitroso compound concentration. The vagotomy and gastrojejunostomy patients had higher pH values and higher concentrations of nitrites and N-nitroso compounds than controls (p = 0.01 in all cases). The 51 vagotomy and gastrojejunostomy patients also underwent endoscopy and biopsy. They were divided into three groups: group 1 (21 patients) had no intestinal metaplasia and no more than mild dysplasia; group 2 (20 patients) had intestinal metaplasia; and group 3 (10 patients) had moderate or severe dysplasia. Groups 2 and 3 both had higher pH values and higher nitrite concentrations than group 1 (p = 0.01 in all cases). There was no significant difference, however, between either group 2 or 3 and group 1 for total N-nitroso compound concentration. Since there was no simple linear relation between pH and N-nitroso compound concentration, it was concluded that formation of N-nitroso compounds at high pH was unlikely to be involved in the pathogenesis of gastric cancer in the hypochlorhydric stomach after operation. The relation between nitrite and histological abnormality was not associated with a similar relation between N-nitroso compounds and histological abnormality. It therefore appears that there is no simple relation between N-nitroso compounds and the pathogenesis of premalignant gastric mucosal changes.

Clinical islet transplantation experience of the University of California Islet Transplant Consortium.

BACKGROUND: The University of California Islet Transplant Consortium was formed to evaluate the feasibility of performing clinical islet transplantation at different transplant centers by using a single centralized islet isolation laboratory. METHODS: From July 1992 through February 1995 seven adult islet transplantations were performed, six allografts and one autograft. Once procured, human pancreata were brought to the UCLA-VA Islet Core Laboratory for islet isolation and purification, which were then transported to different centers for transplantation. Patients 1 through 3 received their transplants in Los Angeles, patient 4 received her islet transplant in Torrance, and patients 5 through 7 received their transplants in San Francisco. RESULTS: Although none of these patients achieved insulin independence, four of seven had functioning grafts longer than 6 months as indicated by circulating C-peptide level greater than 0.7 ng/ml. Furthermore, improved glucose control as shown by a decreased insulin requirement was seen in 57% (four of seven patients) of these patients. The ability to isolate islets at a single laboratory and transport them long distances to different centers was shown in patients 4 through 7. CONCLUSIONS: Islet transplantation can be performed with improvements in blood glucose control, and islets can be isolated at a centralized location and successfully transported to different centers for transplantation.

Immunoneutralization of somatostatin, insulin, and glucagon causes alterations in islet cell secretion in the isolated perfused human pancreas.

INTRODUCTION: In this study, immunoneutralization of endogenous insulin, glucagon, and somatostatin with specific antibodies was used in an isolated perfused human pancreas (IPHP) model. AIMS: To study intrapancreatic cellular interactions and pancreatic hormonal secretion. METHODOLOGY: Randomized, sequential 10-minute test intervals of single-pass perfusion with each antibody were performed at 3.9 mM or 11.5 mM steady-state glucose concentrations. Somatostatin, insulin, and glucagon levels were measured in the effluent during basal and immunoneutralization intervals. RESULTS: At 3.9 mM glucose concentration, somatostatin antibody (SS-Ab) stimulated insulin and glucagon secretion, insulin antibody (IN-Ab) inhibited glucagon secretion, and glucagon antibody (GN-Ab) stimulated insulin secretion. At 11.5 mM glucose concentration, SS-Ab stimulated insulin secretion, IN-Ab stimulated glucagon and inhibited somatostatin secretion, and GN-Ab stimulated insulin secretion. CONCLUSION: The variation in hormonal responses to immunoneutralization during stimulated and nonstimulated glucose conditions suggests that a dynamic association exists between the pancreatic cells.

A new culture method for human pancreatic islets using a biopore membrane insert.

The availability of highly purified human islets has increased with the progressive improvement of isolation methods. This has provided opportunities to perform various in vitro studies on human islets. However, when islets are maintained in culture, the overgrowth of fibroblasts results in a reduced islet purity and often has an adverse effect on islet function. To reduce fibroblast growth and to maintain normal islet function, we have investigated a new three-dimensional culture technique using a noncoated transparent Biopore membrane insert (Millicell CM, Millipore). Islets were isolated from seven human pancreata and cultured for 2 months using this membrane insert. At various time intervals, the functional viability of islets was assessed by measurements of insulin released into the culture medium, static incubation assays of basal and stimulated insulin release, islet insulin contents, and insulin biosynthesis. Results were compared to those of islets cultured in hydrophobic plastic petri dishes, our standard procedure. We found that the non-coated membrane does not allow islet attachment to the surface and prevents fibroblast growth, so that islets maintain a three-dimensional structure and remain in a free-floating form. Islets cultured in a membrane insert showed a function similar to or better than that of islets cultured in plastic petri dishes.

The prognostic significance of marrow micrometastases in women with early breast cancer.

Twenty-five patients with early breast cancer (T0-T2, N0-N1, M0) have been studied prospectively to determine the relationship between marrow micrometastases, disease-free interval and survival. Marrow specimens were aspirated from three sites immediately prior to breast surgery. An immunocytochemical technique using monoclonal antibody LICR.LON.M8.4 was employed to detect micrometastases. The minimum follow-up was 38 months. Twelve of the 25 patients (48%) had micrometastatic lesions in their marrow at presentation. Four of these patients developed distal recurrence during follow-up, causing death in two of them. Five of the 13 patients with no evidence of micrometastases developed distant recurrence and four of them have died. There was no correlation between the state of the marrow and the development of metastatic disease, although axillary lymph node status, disease stage and tumour volume correlated significantly with outcome (all P less than 0.025). Micrometastatic lesions appear to be common in the marrow of patients with early breast cancer. We have been unable to demonstrate that they have prognostic significance.

A review of gene transfer techniques.

The ability to transfer genetic material from one cell to another provides a novel approach for the investigation and potential treatment of a variety of diseases. Several gene transfer techniques have been developed. Of these, the use of retroviruses as vectors has allowed highly efficient and stable introduction of foreign genes into target mammalian cells, thus making gene therapy possible. The current prospects are that some unusual lethal diseases can be corrected and that various forms of cancer will be treated in this manner.

Biologic behavior of sporadic gastrinoma located to the right and left of the superior mesenteric artery.

Among a series of 107 closely followed patients with gastrinoma, 60 patients with sporadic type tumors were identified and evaluated. There were 44 patients (73%) with tumors to the right of the superior mesenteric artery (SMA). Of these, 16 (36%) had extrapancreatic tumors, 28 (64%) had tumor within lymph nodes, and 9 (20%) had multiple tumors. In this group of patients, there were 19 (43%) cures, and only 9 (20%) patients had hepatic metastases. In contrast, in 16 patients (27%) with tumors to the left of the SMA, there were no extrapancreatic tumors, only 3 patients (19%) had tumor within lymph nodes, and 7 (44%) had multiple tumors. In this group, there was only one cure (6%), and nine (56%) patients had hepatic metastases. These findings suggest two distinct populations of sporadic gastrinoma, one to the right (gastrinoma triangle) and the other to the left (outside triangle) of the SMA, which appear to have different biologic behaviors. These differences may reflect divergent etiologies for these two groups of tumors.

A comparison of symptoms between non-ulcer dyspepsia patients positive and negative for Helicobacter pylori.

The role of Helicobacter pylori infection in the symptom complex associated with non-ulcer dyspepsia is uncertain, despite the presence of the organism in a high proportion of these patients. In order to exclude physician bias in history taking, 18 patients (9 female) diagnosed as non-ulcer dyspepsia, after endoscopy and gallbladder ultrasonography, underwent computer interrogation using the Glasgow Diagnostic System for Dyspepsia (GLADYS). Five antral and 3 fundal endoscopic biopsies from these patients were also histologically examined for the presence of Helicobacter pylori and quantitatively analysed for polymorph and chronic inflammatory cell densities per mm2 of lamina propria using computer-linked image analysis. In the group of 9/18 patients who were positive for Helicobacter pylori, there were significantly higher antral and fundal inflammatory cell counts than in negative patients. However, analysis of the GLADYS interrogation data showed no significant positive relationships between Helicobacter pylori positivity and any gastrointestinal symptoms. These results confirm a significant association between Helicobacter pylori and superficial gastritis but suggest that non-ulcer dyspepsia in patients with Helicobacter pylori colonisation is probably not a clinically identifiable and distinct syndrome.