Effects of fungal pancreatic enzymes on the function of islet cells in Syrian golden hamsters.

CONTEXT: Our previous studies showed that porcine pancreatic enzymes in Syrian golden hamsters with peripheral insulin resistance normalizes the plasma insulin level, reduces the size of enlarged islets and inhibits the increased DNA synthesis in the beta-cell of islets. OBJECTIVE: In order to exclude the possibility that these effects was attributed to some contaminants of this crude material, we tested the effect of purified fungal pancreatic enzyme (FPE) that contains primarily amylase and lipase without (FPE) and with addition of chymotrypsin (FPE+chy). MATERIAL AND METHODS: In a pilot study we tested the effect of different doses of FPE given in drinking water on insulin level, islet size and DNA synthesis of islet cells in hamsters with induced peripheral insulin resistance by a high fat diet. The most effective dose of FPE on these parameters was used in a long-term experiment with FPE and FPE+chy in hamsters fed a high-fat diet for 36 or 40 weeks. RESULTS: In the pilot study a dose of 2 g/kg body weight was found to be optimal for controlling the body weight, normalizing plasma insulin level, the size of islets, the DNA synthesis and the number of insulin cells in the islets. These data were produced in the long-term study, where steatorrhea was also inhibited. Addition of chymotrypsin had no effects on these parameters. CONCLUSION: Pancreatic lipase and amylase appear to be responsible for the observed effects and offer a safe and effective natural product for the treatment of pancreatic diseases, including acute pancreatitis, chronic pancreatic, cystic fibrosis and any conditions associated with peripheral insulin resistance, including obesity and type 2 diabetes. The possible mechanism of the action is discussed.

Effects of porcine pancreatic enzymes on the pancreas of hamsters. Part 1: basic studies.

CONTEXT: Porcine pancreatic enzymes (PPE) extracted from glandular stomach has been used for the treatment of pancreatic cancer patients. Unfortunately, no information is available on the in vitro and in vivo effect on the pancreas and other tissues. OBJECTIVE: We used Syrian Golden hamsters, a unique pancreatic cancer model, to obtain basic information on PPE for its eventual use for the treatment of pancreatic cancer. DESIGN: PPE was used in different concentrations in vitro and in vivo. The stability of the enzyme in the water solution was investigated. It was given to the hamsters by gavage in concentrations of 1g/kg and 400 mg/kg for short periods and in aqueous solution for 65 days. Plasma enzyme and insulin, the size of islets and the number of the insulin cells per islet were examined. RESULTS: The enzyme activity of PPE was maintained in water solution for at least 24 hours. Due to its content of calcium chloride it showed a high toxicity to normal and malignant hamster pancreatic cancer cells and human pancreatic cancer cell lines in vitro. PPE did not alter the plasma pancreatic enzyme levels regardless of the dose, duration and application route. On the contrary, PPE reduced their levels significantly. Remarkably, it also reduced the level of insulin, the size of the islets and the number of insulin cells in the islets significantly. CONCLUSION: The results imply that PPE does not enter the blood circulation but it appears to slow down the function of both the exocrine and endocrine pancreas.

Effects of porcine pancreatic enzymes on the pancreas of hamsters. Part 2: carcinogenesis studies.

CONTEXT: Our previous study suggested that porcine pancreatic extract in hamsters with peripheral insulin resistance, normalizes insulin output, islet size and pancreatic DNA synthetic rate. It also inhibited the growth of human pancreatic cancer cells in nude mice. OBJECTIVE: To examine the potential value of the porcine pancreatic extract in controlling pancreatic carcinogenesis in this model, the present experiment was performed. DESIGN: Hamsters were fed a high fat diet and four weeks later when insulin resistance emerges, they were divided into two groups. One group received 1 g/kg BW of porcine pancreatic extract in drinking water and the other group received tap water. One week later, when insulin output normalizes in porcine pancreatic extract-treated hamsters, a single subcutaneous injection of N-nitrosobis-(2-oxopropyl) amine (BOP) at a dose of 40 mg/kg BW was given to all hamsters. The experiment was terminated at 43 weeks after the porcine pancreatic extract treatment. The number and size of pancreatic tumors, blood glucose, insulin, amylase and lipase levels, the average size of islets and the number of insulin cells/islets were determined. RESULTS: The incidence of pancreatic cancer was significantly lower in the porcine pancreatic extract group (P=0.043), as well as the plasma insulin level and the size of the islets in the porcine pancreatic extract group were significantly lower (P<0.001) than in the control group. No significantly differences were found in the glucose level between the groups. CONCLUSION: These results show that porcine pancreatic extract has a potential to inhibit pancreatic cancer growth.

Analysis of the invasion-metastasis mechanism in pancreatic cancer: involvement of plasmin(ogen) cascade proteins in the invasion of pancreatic cancer cells.

To clarify the potential involvement of plasmin(ogen) cascade proteins in the cell dissociation and subsequent invasion of pancreatic cancer cells, western blot analysis, immunocytochemistry, immunohistochemistry, and in vitro invasion assay were performed in the cell lines or tissue of pancreatic cancer. The strong expression of plasmin(ogen), urokinase type plasminogen activator (uPA) and uPA receptor (uPAR), and apparently weak expression of the relevant proteins were found in the conditioned medium of dissociated (PC-1.0) and non-dissociated (PC-1) pancreatic cancer cells, respectively. Furthermore, uPA-treatment significantly induced the expression of plasmin(ogen) and uPAR in the conditioned medium of non-dissociated (PC-1) pancreatic cancer cells. Moreover, the expression of plasmin(ogen) and uPAR was stronger at the invasive front than at the center of human pancreatic cancer tissue. On the other hand, plasmin-treatment induced the expression of matrix metalloproteinase-2 (MMP-2), MMP-7 and MMP-9 in PC-1 cells. Simultaneously, plasmin- or uPA-treatments obviously induced the dissociation of cell colonies and in vitro invasiveness in PC-1 cells. The plasmin(ogen) cascade is closely involved in the invasion of pancreatic cancer cells and, especially in its early stage, cell dissociation. Targeting the plasmin(ogen) cascade may provide a new insight into molecular target therapy based on anti-invasion and anti-metastasis for pancreatic cancer.

Involvement of matrix metalloproteinase-7 in invasion-metastasis through induction of cell dissociation in pancreatic cancer.

Epidermal growth factor receptor (EGFR) mediated mitogen-activated protein kinase kinase (MEK)/extracellular signal-regulated kinase (ERK) signaling pathway was isolated as invasion-metastasis related factor in pancreatic cancer in our previous studies. Matrix metalloproteinase-7 (MMP-7) and tight junction (TJ) proteins are indicated to be involved in cancer invasion-metastasis. To clarify the underlying mechanism of involvement of MMP-7 in cancer invasion, western blotting, invasion assay and immunohistochemistry were performed in dissociated (PC-1.0 and AsPC-1) and non-dissociated (PC-1 and Capan-2) pancreatic cancer cells, as well as pancreatic cancer tissues. Intracellular MMP-7 protein presented as pre-proenzyme and its expression was decreased by AG1478 (EGFR inhibitor) or U0126 (MEK inhibitor) treatment in pancreatic cancer cells. Activated MMP-7 protein was only detected in the medium of PC-1.0 and AsPC-1 cells, but not detected in the medium of PC-1 and Capan-2 cells. Moreover, MMP-7 treatment significant induced the dissociation of cell colonies in PC-1 and Capan-2 cells. Synchronously, TJ structure was apparently disrupted and translocation of TJ proteins to cytoplasm or extracellular medium was induced in PC-1 and Capan-2 cells. Furthermore, MMP-7 treatment markedly increased the in vitro invasion of PC-1 and Capan-2 cells. In addition, MMP-7 expression at the invasive front was obviously stronger than that at the center of pancreatic cancer tissues. Activation of MMP-7 protein is closely involved in disruption of TJ structure and consequent induction of cell dissociation as well as invasion in pancreatic cancer. EGFR mediated MEK/ERK signaling pathway is implied to be involved in regulation of MMP-7 expression in pancreatic cancer cells.

ErbB2 oncogene expression supports the acute pancreatitis-chronic pancreatitis sequence.

The pathogenesis of chronic pancreatitis remains controversial. According to the general opinion, chronic pancreatitis is a de novo disease with a silent but progressive restructure of the pancreas in response to environmental, nutritional or genetic factors. The necrosis-fibrosis sequence hypothesis, on the other hand, postulates that relapsing attacks of acute pancreatitis with subsequent development of fibrosis leads to chronic pancreatitis. Since in our previous studies the expression of two anti-ErbB2 growth factor receptor (ErbB2) antibodies was shown to discriminate between primary chronic pancreatitis, normal tissue, and secondary chronic pancreatitis caused by pancreatic cancer, we studied the ErbB2 expression in tissues obtained from acute, recurrent acute, and chronic pancreatitis to investigate a possible evolution of the ErbB2 expression pattern during the course of the disease. We subjected 14 normal pancreas, 15 chronic pancreatitis, and 12 acute pancreatitis (three with recurrent acute pancreatitis) specimens to immunohistochemical studies using polyclonal anti-ErbB2 antibodies from Santa Cruz and Dako. The immunoreactivity of islet cells in acute pancreatitis cases with the Santa Cruz antibody was less than that in normal pancreas in relation to the degree of tissue damage and fibrosis, and was negative in recurrent acute and chronic pancreatitis tissues. The Dako antibody, on the other hand, revealed a membrane staining of ductal and ductular cells only in chronic pancreatitis specimens and in some areas of recurrent acute pancreatitis. In conclusion, the similarities in the immunoreactivity of anti-ErbB2 antibodies in recurrent acute pancreatitis and chronic pancreatitis support the hypothesis that acute pancreatitis can be a forerunner of chronic pancreatitis.

Indications for EMR/ESD in cases of early gastric cancer: relationship between histological type, depth of wall invasion, and lymph node metastasis.

BACKGROUND: Limited surgery by endoscopic mucosal resection (EMR) or endoscopic submucosal dissection (ESD) for gastric cancer is frequently performed in many institutions. These techniques do preserve gastric function and maintain a high quality of life but may compromise survival. The treatment strategy for early tumors should therefore be based on a complete cure, and limited surgery must thus have clear indications. METHODS: D2 gastric resection was performed in 278 early gastric adenocarcinomas, and a retrospective histological review of the specimens was made. The extended indications for EMR or ESD, according to the Japanese Gastric Cancer Association Treatment guidelines for gastric cancer in Japan, were also assessed. RESULTS: Of the 278 early gastric cancers, 115 were mucosal (M) cancers without ulcer. No lymph node metastases were seen in these specimens. Six of the 41 specimens of M cancer with ulcers had lymph node metastases at the N1 level only. One of these had lymph node metastases from a tumor measuring less than 3 cm in size. Twenty-eight of 122 submucosal cancers had lymph node metastases (23%). Twenty of these were SM1 tumors and 5 had lymph node metastases; 4 of these 5 had lymph node metastases despite the absence of vascular invasion. CONCLUSION: Three cases had lymph node metastases that met the extended criteria for EMR/ESD. EMR and/or ESD should be limited to M cancers without ulcer or differentiated-type M cancer with ulcers smaller than 2 cm. When the depth of tumor invasion is deeper than M, then a gastric resection with lymph node dissection is necessary.

Pancreatic hepatocellular tumor.

BACKGROUND: Hepatocellular differentiation of pancreatic cells has been observed under certain conditions in several species, including humans. Their cell of origin and biology has remained controversial. Generally, these lesions have been considered a degenerative process. The present study describes a neoplastic hepatocellular lesion in Syrian hamsters. METHODS AND RESULTS: Syrian hamsters were treated with a high dose of pancreatic carcinogen, N-nitrosobis(2-oxopropyl)amine. The lesion was confined within a single islet and expressed albumin and HSA (hepatocyte-specific antigen). The pleomorphic tumor cells exhibited numerous mitotic figures and were intermingled with insulin and glucagon cells. The hamster had multicentric premalignant and malignant ductal-type lesions, most of which appeared to arise from within the islets. This is the first demonstration of pancreatic hepatoma. CONCLUSION: Pancreatic islet cells appear to have the potential to transdifferentiate into neoplastic hepatocytes.

Pancreatic enzyme extract improves survival in murine pancreatic cancer.

OBJECTIVES: The disappointing current therapeutic approaches for pancreatic cancer (PC) represent an urgent need for the development of novel methods to control the disease. Based on a recent report on the effectiveness of pancreatic enzyme therapy, we examined the effect of porcine pancreatic enzyme extracts (PPE) on human PC xenografts in nude mice. METHODS: The malignant human PC cell line AsPC1 was transplanted into the pancreas of male beige XID nude mice that were treated or not with PPE in drinking water. The survival, size, and volume of tumors, plasma pancreatic enzyme levels, fecal fat, and urine were examined as were the expression of transforming growth factor alpha, insulinlike growth factor-I, epidermal growth factor, epidermal growth factor receptor, apoptosis, and proliferation rate of tumor cells. RESULTS: PPE-treated mice survived significantly longer than the control group (P < 0.002). Tumors in the PPE-treated group were significantly smaller than in the control group. All mice in the control group showed steatorrhea, hyperglucosuria, hyperbilirubinuria, and ketonuria at early stages of tumor growth, whereas only a few in the treated group showed some of these abnormalities at the final stage. There were no differences in the expression of growth factors, epidermal growth factor receptor, or the apoptotic rate between the tumors of treated and control mice. CONCLUSIONS: The treatment with PPE significantly prolongs the survival of mice with human PC xenografts and slows the tumor growth. The data indicate that the beneficial effect of PPE on survival is primarily related to the nutritional advantage of the treated mice.

The combination of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL/Apo2L) and Genistein is effective in inhibiting pancreatic cancer growth.

OBJECTIVES: Our previous studies have shown that, contrary to many other human pancreatic adenocarcinoma cell lines, AsPC1 cells are resistant to the apoptotic effect of the tumor necrosis factor-related apoptosis-inducing ligand, also called Apo2L (TRAIL/Apo2L). In our in vitro studies, the combination of TRAIL/Apo2L and protein synthesis inhibitor, genistein, but not genistein alone, was, however, effective in inducing apoptosis in AsPC1 cells. In the present study, we examined the effect of TRAIL/Apo2L with genistein on the growth of AsPC1 cells in vitro and in vivo. METHODS: Mice with orthotopically transplanted AsPC1 cells were treated either with TRAIL/Apo2L, Genistein (Gen) or a combination of both (TRAIL/Apo2L + Gen) for 14 days. After 14 days, the size and weight of the tumors were registered and the apoptosis of the tumor cells were determined by the TUNEL method. In vitro, the effect of combination treatment on cytotoxicity was assessed by MTT assay and apoptosis was assessed by DAPI staining. FADD, caspase 3, and PARP proteins were determined by Western blot. RESULTS: No toxic side effects were observed in either group. The tumor volume was significantly smaller and the apoptotic ratio was higher in the TRAIL + Gen group than in the other 2 groups. The apoptotic effect was associated with the caspase-3 activation. Z-VAD-FMK partially inhibited apoptosis by TRAIL + Gen. CONCLUSIONS: These results indicate that the combination of TRAIL/Apo2L with genistein presents a promising therapeutic approach for the treatment of pancreatic cancer. Further detail investigations are needed, however, to verify the mechanisms of this combination therapy.