Significant expression of interleukin 15 in rat experimental severe acute pancreatitis.

PURPOSE: In severe acute pancreatitis (SAP), multiple organ dysfunction syndrome is a contributor to high mortality. We recently demonstrated that the serum interleukin (IL)-15 level is a predictor of the complications and mortality in clinical SAP. The aim was to investigate the role of IL-15 in experimental SAP. MATERIALS AND METHODS: SAP was induced by retrograde injection of 3 and 20% sodium deoxycholate (DCA) into biliopancreatic ducts in rats (DCA pancreatitis). Expressions of IL-15 were evaluated by Western blotting and immunohistochemical staining. Recombinant IL-15 protein was administered intraperitoneally, and the effects were investigated. RESULTS: Western blotting revealed the expressions of IL-15 in the pancreas, liver, lung and intestine in 3% DCA pancreatitis. Immunohistochemical staining showed the expression of IL-15 in the cytoplasm of each organ. In 3% DCA pancreatitis, administration of recombinant IL-15 protein attenuated the elevation of serum alanine aminotransferase (ALT) levels and improved the morphological change of the lung 18 h after the induction of SAP. Moreover, in 20% DCA pancreatitis, IL-15 improved the elevation of serum amylase and ALT levels 6 h after the induction. CONCLUSIONS: These results suggest that IL-15 is related to organ dysfunction during SAP, and that IL-15 functions as a protective factor against the organ injuries.

Enhancement of secretagogue-induced phosphoinositide turnover and amylase secretion by bile acids in isolated rat pancreatic acini.

Small amounts (0.1-0.5 mM) of deoxycholate enhanced amylase secretion, which had been induced by submaximal doses of carbachol or cholecystokinin octapeptide, without affecting the maximal levels of these reactions from isolated rat pancreatic acini. Deoxycholate alone did not induce these reactions. The other bile acids such as cholate, chenodeoxycholate, ursodeoxycholate, and taurocholate were also active. Under the similar conditions, deoxycholate enhanced the secretagogue-induced diacylglycerol formation that was derived mainly from the phospholipase C-mediated hydrolysis of phosphatidylinositol and phosphatidylinositol-4-monophosphate. Deoxycholate did not enhance the secretagogue-induced hydrolysis of phosphatidylinositol-4,5-bisphosphate or Ca2+ mobilization. Deoxycholate did not affect amylase secretion, which was induced by the simultaneous addition of protein kinase C-activating 12-O-tetradecanoylphorbol-13-acetate and Ca2+ ionophore ionomycin. Since diacylglycerol and Ca2+ may be responsible for the secretagogue-induced amylase secretion, our results indicate that small amounts of bile acids increase the sensitivity to the secretagogue of diacylglycerol formation and subsequent activation of protein kinase C, and thereby enhance amylase secretion from pancreatic acini.

Protective effect of a microtubule stabilizer taxol on caerulein-induced acute pancreatitis in rat.

The effect of taxol, which is a microtubule stabilizer, was examined in a model of acute edematous pancreatitis induced in rat by the administration of caerulein. Prophylactic administration of taxol ameliorated inhibition of pancreatic secretion, increased level of serum amylase, pancreatic edema, and histological alterations in this model. Immunofluorescence studies revealed that taxol stabilized the arrangement of microtubules by the action of promoting tubulin polymerization and prevented inhibition of pancreatic digestive enzyme secretion. In isolated rat pancreatic acini, taxol reversed the inhibition of amylase secretion induced by supramaximal concentrations of cholecystokinin octapeptide and did not affect the binding of cholecystokinin octapeptide to its receptor. The results obtained in this study suggest that microtubule disorganization is the initiating event in caerulein-induced pancreatitis and that the inhibition of pancreatic digestive enzyme secretion by interfering with intracellular vesicular transport due to microtubule disorganization causes caerulein-induced pancreatitis.

Presence of human immunoglobulin G anti serum pancreatic elastase 1 autoantibodies and their influence on elastase 1 radioimmunoassay.

Human immunoglobulin G (IgG) anti human pancreatic elastase 1 autoantibodies were detected in sera of patients with pancreatic disorders. The characteristics of these anti elastase 1 autoantibodies and their influence on radioimmunoassay (RIA) for elastase 1 were investigated. They were placed in the IgG class by the double antibody method, and most were assumed to be of a monoclonal type from their elution profiles in gel filtration analysis. The presence of autoantibodies in serum caused an increase in apparent elastase 1 values and a decrease in the recovery of elastase 1 exogenously added to the serum. These results suggest that elastase 1 immunoassay data for autoantibody positive sera can cause misjudgement of clinical stages of patients.

Enhancement of growth factor-induced DNA synthesis by colon tumor-promoting bile acids in Swiss 3T3 cells. Their different mode of action from that of phorbol esters.

In Swiss 3T3 cells, colon tumor-promoting deoxycholate (DOC) enhanced DNA synthesis which was induced by fibroblast growth factor (FGF) in the presence of insulin. This effect was observed only when DOC was added within 10 h after the addition of FGF. DOC by itself did not induce DNA synthesis irrespective of the presence or absence of insulin. Similar results were obtained with other colon tumor-promoting bile acids such as cholate, chenodeoxycholate and taurocholate. In contrast to these bile acids, 12-O-tetradecanoylphorbol-13-acetate (TPA) induced DNA synthesis fully without FGF in the presence of insulin. DOC did not affect TPA-induced DNA synthesis. Prolonged treatment of the cells with phorbol-12,13-dibutyrate caused the down-regulation of the phorbol ester receptor and rendered the cells unresponsive to TPA. In these cells, FGF still induced DNA synthesis in the presence of insulin, but the maximal level was reduced to about one third of that in the control cells. DOC did not enhance this DNA synthesis any more. DOC did not alter the binding of FGF to the cells. These results indicate that colon tumor-promoting bile acids enhance the mitogenic action of FGF and thereby stimulate DNA synthesis, although the phorbol ester substitutes for the mitogenic action of FGF.

Enhancement of fibroblast growth factor-induced diacylglycerol formation and protein kinase C activation by colon tumor-promoting bile acid in Swiss 3T3 cells. Different modes of action between bile acid and phorbol ester.

A small amount (50-200 microM) of deoxycholate (DOC), a colon tumor-promoting bile acid, did not show a direct effect on protein kinase C activity in a cell-free system, but enhanced fibroblast growth factor (FGF)-induced diacylglycerol formation and protein kinase C activation in Swiss 3T3 cells. DOC potentiated both reactions induced by submaximal doses of FGF but showed little effect on the maximal levels of the reactions. DOC alone was inactive in eliciting both reactions in the absence of FGF. DOC did not affect the binding of FGF to the cells. Since it has been described that diacylglycerol serves as a messenger for the activation of protein kinase C in the action of FGF in Swiss 3T3 cells [(1985) FEBS Lett. 191, 205-210], these results suggest that a small amount of DOC increases the sensitivity to FGF of diacylglycerol formation and thereby potentiates protein kinase C activation in this cell line. This action of DOC was in marked contrast to that of 12-O-tetradecanoylphorbol-13-acetate, a potent tumor-promoting phorbol ester, which directly activated protein kinase C in cell-free and intact cell systems.

Papillary hyperplasia of the pancreas.

We report the first case of surgically resected pure papillary hyperplasia of the pancreas. Interestingly, it was not associated with chronic pancreatitis or pancreatic cancer. Histologic and immunohistochemical features are described, with a review of the literature on papillary hyperplasia of the pancreas.

Extracorporeal removal of anticancer drugs in hepatic artery infusion: the effect of direct hemoperfusion combined with venovenous bypass.

The effect of a new extracorporeal system combining direct hemoperfusion (DHP) with venovenous bypass was evaluated in the elimination of anticancer drugs in hepatic artery infusion. Adriamycin (3 mg/kg) and mitomycin C (1 mg/kg) were given to mongrel dogs through the hepatic artery with three different durations of 1, 10, and 20 minutes. Plasma drug levels were determined at the inlet and outlet of DHP and right external jugular vein (systemic level). Blood flow through DHP averaged 200 ml/min. In dogs without DHP (group I; n = 4), systemic levels of adriamycin and mitomycin C increased rapidly with 1-minute infusion, reaching the peak values of 6.61 +/- 2.44 (mean +/- SD) and 2.20 +/- 1.05 micrograms/ml, respectively. With DHP under single venous bypass (group II; n = 5), the peak values were reduced to 1.25 +/- 1.02 and 0.79 +/- 0.52 microgram/ml. Moreover, the peak levels were markedly reduced by DHP under hepatic venous isolation (group III; n = 6), the values being 0.41 +/- 0.15 and 0.13 +/- 0.07 microgram/ml with 1-minute infusion. The drug-removal rates were improved substantially in group III compared with group II. The longer the duration of infusion, the higher the removal rates tended to be in group III. These results indicate that effective elimination of anticancer drugs can be accomplished by this system during intraarterial chemotherapy of the liver.

Effectiveness of acidic oxidative potential water in preventing bacterial infection in islet transplantation.

At a number of points in the current procedures of islet isolation and islet culture after the harvesting of donor pancreata, microorganisms could potentially infect the islet preparation. Furthermore, the use of islets from multiple donors can compound the risks of contamination of individual recipients. Acidic oxidative potential water (also termed electrolyzed strong acid solution, function water, or acqua oxidation water), which was developed in Japan, is a strong acid formed on the anode in the electrolysis of water containing a small amount of sodium chloride. It has these physical properties: pH, from 2.3 to 2.7; oxidative-reduction potential, from 1,000 to 1,100 mV; dissolved chlorine, from 30 to 40 ppm; and dissolved oxygen, from 10 to 30 ppm. Because of these properties, acidic oxidative potential water has strong bactericidal effects on all bacteria including methicillin-resistant Staphylococcus aureus (MRSA), viruses including HIV, HBV, HCV, CMV, and fungi as a result of the action of the active oxygen and active chlorine that it contains. We conducted this study to evaluate the effect of acidic oxidative potential water irrigation on bacterial contamination on the harvesting of porcine pancreata from slaughterhouses for islet xenotransplantation by counting the number of pancreatic surface bacteria using the Dip-slide method, and on the results of islet culture; and to evaluate the direct effect on isolated islets when it is used to prevent bacterial contamination by the static incubation test and by morphological examination. Direct irrigation of the pancreas by acidic oxidative potential water was found to be very effective in preventing bacterial contamination, but direct irrigation of isolated islets slightly decreased their viability and function.

Improved large-scale isolation of breeder porcine islets: possibility of harvesting from nonheart-beating donor.

To establish a large-scale isolation procedure for adult porcine islets usable as a donor source for xenotransplantation and as a model of human islet isolation, we improved several characteristics of the conventional isolation procedure. At a slaughterhouse we first selected a breeder pig over 1.5 years old (and over 200 kg in weight) with warm ischemic time (WIT) of 15 +/- 2 minutes as nonheart-beating donors. Then, we made a special enzymic mixture that consisted of collagenase S-1 (260 U/mg, NittaZelatin, Japan), collagenase P (1.86 U/ml Lyo Boehringer-Mannheim, USA), DNase (Sigma, St. Louis, Mo), Disparse (NittaZelatin, Japan), and protease inhibitor (Sigma). Third, this mixture was injected very gently into the pancreatic duct at the time of pancreatic harvesting. To prevent overdigestion of the pancreas, the mixture was first cooled to less than 10 degrees C. Fourth, during the warm digestion of pancreas, the pancreas with the enzymic mixture was quietly put in a water bath at 37 degrees C without mechanical shaking. Fifth, we purified the islets with a COBE 2991 cell processor by the Dextran 70 gradient method, because Dextran 70 is very cheap and has the same purification effect as the Ficoll gradient. The results of 10 consecutive breeder porcine islet isolations are reported. The total yield of isolations of islets over 50 microm in the longest diameter after staining with Dithizone (DTZ) was 85,900 +/- 19,954 islets, 291,667 +/- 240,452 IEQ (2,900 +/- 2,324 IEQ/g). The purity of the isolated islets was very high: 90.2 +/- 3.8%. Glucose stimulation during in vitro incubation induced significant insulin release from isolated breeder porcine islets. In two of the diabetic rats receiving encapsulated islets grafts using a mesh-reinforced polyvinyl alcohol hydrogel bag (MRPB), a prominent reduction in serum glucose levels (less than 200 mg/dL) persisted for 13 and 19 days, respectively, after intraperitoneal xenotransplantation islets without immunosuppression. In conclusion, we succeeded in a more efficient and less-expensive isolation of a large amount of adult porcine islets from a nonheart-beating donor.

Radiolocalization of pancreatic carcinoma xenografts in nude mice with radiolabeled chimeric Fab fragments of anti-carcinoembryonic antigen monoclonal antibody A10.

Recombinant mouse-human chimeric Fab fragments of anti-carcinoembryonic antigen monoclonal antibody (MAb) A10 react with various GI carcinomas. We tested radiolocalization of pancreatic carcinoma xenografts in nude mice using radiolabeled chimeric A10 Fab fragments, comparing them with murine Fab fragments and parental MAb. For mice injected with chimeric A10 Fab fragments, we obtained significantly higher uptake in tumors than in normal tissues at 24 and 48 h after injection. In addition, tumor/normal tissues labeling ratios for chimeric A10 Fab fragment were significantly greater than those for murine MAb at 24 h postinfusion. However, no significant difference in biodistribution was observed between chimeric and murine Fab fragments. In autoradiography imaging studies, we obtained clearer tumor detection without visible uptake in normal organs for chimeric Fab fragments than for murine MAb. These results suggest that chimeric Fab fragments of A10 could be a potentially useful candidate for radioimmunodetection of pancreatic carcinomas.

The preparation of the chronic hyper-endotoxemia experimental animal model by means of a drug delivery system.

A drug delivery system (DDS) consisting of lipopolysaccharide (LPS) as a drug and 2-hydroxyethyl methacrylate (HEMA)-diethylene glycol dimethacrylate (2G) or -polyethylene glycol dimethacrylate (4G, 9G) copolymer was prepared, and used for the efficient preparation of an experimental animal model of chronic hyper-endotoxemia. The release profiles of LPS in the in-vitro test were greatly influenced by the composition of HEMA-2G, 4G, 9G in the copolymer. It was found that LPS release from the DDS continued gradually and constantly throughout 2 weeks. In the in-vivo experiment with rats, the DDS maintained a high blood concentration level of LPS for 3 days. These results strongly suggest the possibility of convenient and reproducible preparation of a chronic hyper-endotoxemia animal model.

Expression of manganese superoxide dismutase in esophageal and gastric cancers.

The tumor-killing activity of radiotherapy and chemotherapy for cancer is closely associated with the production of active oxygen, and the relation between therapeutic resistance and active oxygen scavengers produced by the tumor itself is gaining more attention. It is considered that manganese superoxide dismutase (MnSOD) protects host cells from oxidative stress, in synergy with other antioxidant enzymes. In this study, we used a quantitative polymerase chain reaction assay to measure MnSOD mRNA in resected specimens from patients with esophageal and gastric cancers. In both esophageal and gastric cancers, the level of MnSOD mRNA was significantly elevated in cancer tissue compared to non-cancer tissue (P < 0.01). In gastric cancer tissue, the MnSOD mRNA level was significantly higher than in esophageal cancer tissue (P < 0.01). The significance of MnSOD in cancer tissue was investigated further by measuring MnSOD content in resected specimens using an enzyme-linked immunosorbent assay, and by examining its location by an immunohistochemical method. Upregulation of MnSOD in cancer tissue most likely serves as a protective mechanism against anti-cancer therapies known to produce superoxide radicals as a key component of their tumor-killing activity.

A radioimmunoassay for pancreatic elastase 1: use of alpha 1-antitrypsin-elastase 1 conjugate as both standard and tracer to eliminate the influence of anti-elastase 1 autoantibodies.

We have recently reported the occurrence of anti-elastase 1 autoantibodies in human sera (Asada et al., Biochim Biophys Acta, 1991;1080:34-39). The usual radioimmunoassay of elastase 1 in autoantibody-positive sera gave abnormally high levels of elastase 1 and low recoveries of elastase 1 added to the sera. Now we have established a new radioimmunoassay system for determining pancreatic elastase 1 in serum, in which alpha 1-antitrypsin elastase 1 conjugate, the major conjugation form of elastase 1 in serum, is used as standard and alpha 1-antitrypsin-[125I]elastase 1 as tracer, because we found that the conjugate could bind to the elastase 1 specific antiserum but not to the autoantibodies. Thus the new assay system completely eliminate the unfavorable effects of the autoantibodies. The elastase 1 levels determined by the new assay method exhibited a good correlation with those obtained by a commercial assay kit in the autoantibody-negative sera, while no correlation was observed in the autoantibody-positive sera.

Effects of nucleosides and a nucleotide on DNA and RNA syntheses by the salvage and de novo pathway in primary monolayer cultures of hepatocytes and hepatoma cells.

Studies were made on the effects of inosine, guanosine 5'-monophosphate (GMP), cytidine, uridine, thymidine, and their mixture (4:4:4:3:1, OG-VI) on DNA and RNA syntheses in primary monolayer cultures of normal hepatocytes and cultures of hepatoma cells, AH130, to use these compounds for total parenteral nutrition. Addition of an appropriate amount of inosine, GMP, uridine, or thymidine to primary cultures of hepatocytes enhanced both DNA and RNA syntheses by the salvage and de novo pathways. Cytidine appeared to have lower optimal concentration for enhancing these pathways. The OG-VI mixture also enhanced the syntheses of DNA and RNA, but the composition of the mixture was not optimal. Additions of inosine, GMP, uridine, and thymidine to cultured hepatoma cells also enhanced their DNA and RNA syntheses, but the cells consumed more of the added nucleic acid compounds than hepatocytes did. Addition of cytidine had no effect on proliferation of the cells. The OG-VI mixture at relatively higher concentration inhibited the syntheses of DNA and RNA by hepatoma cells. Addition of high concentrations of nucleic acid compounds was found to suppress the proliferation of both hepatocytes and hepatoma cells. These results suggest that addition of optimal amounts of nucleic acid compounds such as nucleosides and nucleotides would enhance growth of hepatocytes, particularly during liver regeneration, but that they may also enhance proliferation of tumor cells in the liver.

Effect of methionine-deprived nutrition on cell growth and cell kinetics in cell cultures and experimental tumors.

The effect of methionine-deprived nutrition on cell growth and cell kinetics was investigated in cell cultures and in tumor-bearing rats using the total parenteral nutrition (TPN) technique. A simultaneous flow cytometric measurement of the cellular DNA content and the amount of 5-bromodeoxyuridine incorporated into cellular DNA was performed for analysis of cell kinetics. The methionine-free medium demonstrated a cytocidal effect on the growth of SLC cells after 6 hours of culturing. It decreased viability from 80% in the control medium to 23%, and it decreased the S phase and increased the G0/G1 phase of the cell cycles. The methionine-deprived medium showed a concentration-dependent inhibition in cellular growth. Methionine-deprived TPN was seen to inhibit AH109A and SLC tumor growth compared with conventional TPN and decreased the S phase and increased the G0/G1 phase of cell cycles. These results confirm that methionine deprivation blocks cells from processing into the G1 phase and recycling, and that it is effective in inhibiting tumor growth in cultures and in vivo.

Laparoscopic intervention for intrathoracic stomach in infants.

BACKGROUND: Intrathoracic stomach is an uncommon condition in infants. We report our experience managing such a condition successfully by laparoscopy in four patients. METHODS: Patients' ages at the time of operation ranged from 30 days to 14 months. In all cases, the intrathoracic stomach was easily pulled down into the abdominal cavity. The phrenoesophageal ligament was completely resected, and the enlarged hiatus was narrowed by intraabdominal suturing technique. The esophagus was wrapped with the mobilized fundus in a floppy fundoplication. Anchoring sutures were placed between the wrapping cuff and crura. RESULT: One patient had paraesophageal hernia (type 2), whereas the other had combined hiatal hernia (type 3). No adverse complications were observed in any of the cases. Symptomatic gastroesophageal reflux and radiographic recurrence of hernia were not seen in any case. The cosmesis was excellent in all cases. CONCLUSIONS: We conclude that laparoscopic repair for intrathoracic stomach is a safe and feasible method when preoperative evaluation is conducted adequately.

Immunohistochemical staining of pancreatic cancer with CA19-9, KM01, unabsorbed CEA, and absorbed CEA. A comparison with normal pancreas and chronic pancreatitis.

In order to test the effectiveness of CA19-9, KM01, unabsorbed CEA (carcinoembryonic antigen) and absorbed CEA by immunoperoxidase staining, we evaluated the staining distribution, intensity, and cellular localization in pancreatic cancer. The results were then compared with those of normal pancreas and chronic pancreatitis. The positive staining rate of the pancreatic cancer with any of the four tumor markers was higher than that of the normal pancreas. However, all markers except absorbed CEA showed a higher positive staining rate for chronic pancreatitis than for pancreatic cancer. There was no stromal type in normal pancreatic or chronic pancreatitis tissues with any of the four tumor markers. Our findings, therefore, indicate that absorbed CEA is useful in differentiating pancreatic cancer from normal pancreatic tissues. It is not useful in distinguishing chronic pancreatitis, however, unless a specific staining pattern is observed.

Metabolic effects of systemic venous drainage in small bowel transplantation.

Small bowel transplantation has proved feasible in rats and in larger animals, but several important questions remain to be addressed before it becomes routine therapy in humans. One consideration is the site of venous outflow of the allograft. While portal drainage reestablishes the physiologic route of venous outflow, systemic drainage creates a partial mesocaval shunt, the metabolic consequences of which have not been studied in detail. Using a canine model of orthotopic small bowel autotransplantation, we compared the metabolic changes following transplantation with portal versus systemic venous drainage. Changes in blood ammonia levels, plasma amino acid composition, and hepatic blood flow were studied, since the Eck procedure produces metabolic changes of hyperammonemia and amino acid imbalances, while portocaval venous drainage of small bowel transplant produces a profile similar to that in controls. These data suggest that there is no metabolic disadvantage of systemic venous drainage as compared with controls. Because of its technical simplicity, systemic venous drainage may be preferable for small bowel transplantation.

Hepatobiliary scintigraphy after biliary reconstruction--a comparative study on Roux-Y and ESCD.

BACKGROUND/AIMS: Reconstruction of extrahepatic biliary tract with benign lesion still has some unsettled problems, such as postoperative cholangitis. This study was conducted to compare bile through the remnant alimentary tract in patients undergoing end-to-side choledochoduodenostomy, and those undergoing Roux-Y choledochojejunostomy, using hepatobiliary scintigraphy. METHODOLOGY: Five normal human volunteers and 13 patients underwent end-to-side choledochoduodenostomy (n = 5), Roux-Y choledochojejunostomy (n = 8), using hepatobiliary scintigraphy. RESULTS: Postoperative acute cholangitis developed in 1 patient (12%) with Roux-Y choledochojejunostomy and none with end-to-side choledochoduodenostomy. Hepatobiliary scintigraphy showed prominent stasis of 99mTc in the proximal jejunum loop of the patients who underwent the Roux-Y choledochojejunostomy procedure, which was not found in the upper jejunum of the patients with end-to-side choledochoduodenostomy. The time taken before visualization of 99mTC at the upper jejunum in the patient who underwent Roux-Y choledochojejunostomy (66.5 +/- 5 min) was significantly longer than that in the healthy controls (40 +/- 5 min). On the other hand, the time taken before visualization of 99mTc at the upper jejunum in end-to-side choledochoduodenostomy (35 +/- 5 min) was similar to that of healthy controls. CONCLUSIONS: This data suggested that end-to-side choledochoduodenostomy procedure for reconstructing the extrahepatic biliary tract was more physiological with less postoperative complication than Roux-Y choledochojejunostomy procedure.