Biomedical subjects

P M Pour

Publications and source records attributed to P M Pour.

At least 19 recordsLinked to original sources

PubMed2003

Immortalization with telomerase of the Nestin-positive cells of the human pancreas.

Cells expressing the neuronal stem cell marker Nestin are present in the human pancreas but the biological role of these cells has yet to be resolved. We report here the establishment with the catalytic subunit of human telomerase (hTERT) of a line of normal human cells representing this cell type. Primary human cells derived from the ducts of the pancreas were transduced with an hTERT cDNA. The infected cells became positive for telomerase, failed to senesce, and were still proliferating after more than 150 doublings. The immortalized cells were positive for the expression of Nestin (at both the mRNA and protein levels) and were found to be free of cancer-associated changes: diploid and expressing wild type p16(INK4a), p53, and K-Ras. An established line of normal human cells representing this cell type should be of great value to help define the biological properties of this novel cell type.

Base Sequence↗

PubMed2001

Identification and sequencing of the Syrian Golden hamster (Mesocricetus auratus) p16(INK4a) and p15(INK4b) cDNAs and their homozygous gene deletion in cheek pouch and pancreatic tumor cells.

Previous studies have shown that the p16(INK4a) tumor suppressor gene is inactivated in up to 98% of human pancreatic cancer specimens and 83% of oral squamous cell carcinomas. Inactivation of the related p15(INK4b) gene has also been identified in a number of tumors and cell lines, however, its role as an independent tumor suppressor remains to be elucidated. Chemically-induced tumors in the Syrian Golden hamster (Mesocricetus auratus) have been shown to be excellent representative models for the comparative development and progression of a number of human malignancies. The purpose of this study was to determine the importance of the p16(INK4a) and p15(INK4b) genes in two experimental hamster models for human pancreatic and oral carcinogenesis. First, hamster p16(INK4a) and p15(INK4b) cDNAs were cloned and sequenced. The hamster p16(INK4a) cDNA open reading frame (ORF) shares 78%, 80%, and 81% identity with the human, mouse, and rat p16(INK4a) sequences, respectively. Similarly, the hamster p15(INK4b) cDNA ORF shares 82% and 89% sequence identity with human and mouse p15(INK4b), respectively. Second, a deletion analysis of hamster p16(INK4a) and p15(INK4b) genes was performed for several tumorigenic and non-tumorigenic hamster cell lines and revealed that both p16(INK4a) and p15(INK4b) were homozygously deleted in a cheek pouch carcinoma cell line (HCPC) and two pancreatic adenocarcinoma cell lines (KL5B, H2T), but not in tissue matched, non-tumorigenic cheek pouch (POT2) or pancreatic (KL5N) cell lines. These data strongly suggest that homozygous deletion of the p16(INK4a) and p15(INK4b) genes plays a prominent role in hamster pancreatic and oral tumorigenesis, as has been well established in correlative studies in comparable human tumors. Furthermore, this study supports the comparative importance of the hamster pancreatic and cheek pouch models of carcinogenesis in subsequent mechanistic-, therapeutic-, and preventive-based studies aimed at providing important translational data applicable to pancreatic adenocarcinoma and oral squamous cell carcinoma in humans.

Amino Acid Sequence↗

PubMed2001

Introduction.

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Animals↗

PubMed2001

The patterns of extrainsular endocrine cells in pancreatic cancer.

Abnormal glucose tolerance and frank diabetes mellitus develop in up to 80% of pancreatic cancer patients. Islets within these tumors show a decreased number of beta cells and increased number of alpha cells. The reduced number of beta cells could induce beta cell neogenesis in extrainsular tissue to compensate for the loss of insulin in islets. On the other hand, because the beta cell depletion in pancreatic cancer seems to be the effect of substances released by cancer cells, suppression of extrainsular endocrine cells is expected. We compared the pattern of extrainsular endocrine cells in pancreatic cancer patients with normal pancreas as well as chronic pancreatitis, which is known to be associated with impaired glucose tolerance or frank diabetes. As in the normal tissue, extrainsular endocrine cells were found in chronic pancreatitis and pancreatic cancer. However, in the chronic pancreatitis specimens insulin cells were the predominant cell type, whereas in pancreatic cancer specimens more glucagon than insulin cells were found, although the differences were statistically insignificant. Thus, our results indicate that the alteration of beta cells in pancreatic cancer patients is mainly restricted to the endocrine cells within the islets and that there is no compensatory proliferation of beta cells.

Adenocarcinoma↗

PubMed2001

Prevention of pancreatic cancer induction in hamsters by metformin.

BACKGROUND AND AIMS: Our previous study suggested that the known promotional effect of a high fat diet, which in hamsters induces peripheral insulin resistance, is related to a compensatory proliferation of islet cells. The present study was to examine whether the prevention of islet cell proliferation can inhibit the promotional effect of a high-fat diet in pancreatic carcinogenesis. METHODS: Two groups of high fat-fed hamsters were used. One group received Metformin in drinking water for life (HF+Met group), and the other group served as a control (HF group). At the time when the normalization of the plasma insulin level was expected, all hamsters were treated with the pancreatic carcinogen, N-nitrosobis-(2-oxopropyl)amine, and the experiment was terminated 42 weeks later. RESULTS: Although 50% of the hamsters in the high-fat group developed malignant lesions, none was found in the HF+Met group (P < 0.05). Also, significantly more hyperplastic and premalignant lesions, most of which were found within the islets, were detected in the high-fat group (8.6 lesions/hamster) than in the HF+Met group (1.8 lesions/hamster). CONCLUSIONS: The results lend further support on the significant role of islet cells in pancreatic carcinogenesis and may explain the association between pancreatic cancer and obesity, which is usually associated with peripheral insulin resistance.

Adenocarcinoma↗

PubMed2001

Increased expression of glutathione S-transferase-pi in the islets of patients with primary chronic pancreatitis but not secondary chronic pancreatitis.

The mechanism of tissue alteration in chronic pancreatitis (CP) is still unclear. Different hypotheses have been discussed, including increasing oxidant stress in the acinar cells, often as a result of exposure to xenobiotics. To evaluate the role of oxidative stress in CP, the authors investigated the expression of the drug-metabolizing phase II enzyme, glutathione S-transferase-pi (GST-pi), in the pancreatic tissue of patients with CP and compared it with the healthy pancreatic tissue from age-matched donors. Pancreatic tissue from patients with secondary CP resulting from ductal obstruction by pancreatic cancer (PC) was also examined. The percentage of cells immunoreacting with anti-GST-pi was counted within 15 randomly selected islets in each slide of the three groups. In all specimens, ductal and ductular cells, and in PC, cancer cells, expressed GST-pi in a moderate intensity. Acinar cells did not stain. Various numbers of islet cells in each of the three groups were stained strongly. More islet cells expressed GST-pi in CP (42%) than in healthy pancreatic tissue (16%, p < 0.001) or PC (17%, p < 0.001). Our results imply an important role of islet cells in the metabolism of substances, which are the substrate for GST-pi, and lend support to the hypothesis of oxidative stress as the cause of CP.

Antibodies, Monoclonal↗

PubMed2001

Pacinian corpuscle in the human pancreas.

During our systematic examination of the distribution of cytochrome P450 enzymes in the normal and diseased human pancreas, we observed a Pacinian corpuscle in a serial section of a tissue from a pancreatic cancer patient. We report the histologic and immunohistochemical patterns in this corpuscle and review the literature. The Pacinian corpuscle was situated within the pancreas of a 76-year-old woman with cancer in the head of the pancreas. We could demonstrate immunoreactivity within the corpuscle for the neurofilament protein. neuron-specific enolase, S-100 Protein, and for four cytochrome P450-isozymes. The possible function of Pacinian corpuscles in the mammalian and human pancreas is discussed.

Aged↗

PubMed2001

Transdifferentiation of human islet cells in a long-term culture.

It has been established that ductal cells or precursor cells within the ductal tree of the pancreas can differentiate into islet cells. Although islet cells can also form exocrine cells, it is unclear whether they arise from precursor (stem) cells or from mature endocrine cells by transdifferentiation. Using a defined culture medium and technique for islet purification, for the first time we were able to maintain human islets in culture for more than a year. Multilabeling immunohistochemical and immunoelectron microscopic examination of the islets at different days of culture using islet cell markers (antibodies to hormones, neuron-specific enolase, chromogranin A) and ductal cell markers (cytokeratins 7 and 19, carbonic anhydrase II, DU-PAN2, CA 19-9, and MUC1) revealed that endocrine cells gradually transdifferentiate to ductal, acinar, and intermediary cells. Although islet hormone secretion ceased after day 28 in culture, endocrine cells were still detectable at day 60. However, later, all endocrine and exocrine cells were replaced by undifferentiated cells that expressed neuron-specific enolase, chromogranin A, laminin, vimentin, cytokeratin 7 and 19, alpha-1-antitrypsin, transforming growth factor-alpha, and epidermal growth factor receptor. Our data thus show that, under proper conditions, human islets can be maintained in vitro over a long period and that, in the culture condition, islet cells seem to transdifferentiate to exocrine cells and undifferentiated cells, which may be considered pancreatic precursor (stem) cells.

Biomarkers↗

PubMed2001

Experimental animal models in pancreatic carcinogenesis: lessons for human pancreatic cancer.

The silent course of pancreatic cancer and its explosive fatal outcome have hindered studies of tumor histogenesis and the identification of early biochemical and genetic alterations that could help to diagnose the disease at a curable stage and develop therapeutic strategies. Experimental animal models provide important tools to assess risk factors, as well as preventive and therapeutic possibilities. Although several pancreatic cancer models presently exist, only models that closely resemble human tumors in morphological, clinical, and biological aspects present useful media for preclinical studies. Because an estimated 70% of human tumors are induced by carcinogens and because a significant association has been found between cigarette smoking and pancreatic cancer, chemically induced models are of particular value. Moreover, in such models the etiology, modifying factors, effects of diets, and naturally occurring products can be studied and early diagnostic, preventive, and therapeutic possibilities sought out. Many of the existing models are described in this review, and the advantages and shortcomings of each model and their clinical implications are discussed.

Animals↗

PubMed2001

Abnormal differentiation of islet cells in pancreatic cancer.

Pancreatic cancer in many patients is associated with altered glucose metabolism and abnormalities in pancreatic islet hormones at serum and tissue levels. Our previous studies have indicated a tendency of islet cells to differentiate toward ductal cell lineage, but the specificity of these findings for pancreatic cancer was not investigated. In the present study, we examined the immunoreactivity of pancreatic islets to antibodies against tumor-associated antigens DU-PAN-2, TAG-72 and CA19-9 in tissues from the normal pancreas, chronic pancreatitis and pancreatic cancer. Although no immunoreactive islet cells were found in the 12 normal pancreases and 20 chronic pancreatitis patients, 25 of 37 pancreatic cancer tissues showed the expression of these antigens, primarily CA19-9 and TAG-72, where the number of immunoreactive cells varied considerably from case to case. In 4 cases over 50% and in 2 of them more than 75% of the islets showed positive staining of 60-70% of islet cells within each islet. The presence of intrainsular ductular structures expressing the same antigen as the surrounding islet cells suggested transformation of antigen expressing islet cells to ductal cells. All but four islets were within or around the cancer favoring the notion that factors produced by cancer cells are responsible for the altered islet cell differentiation.

Antigens, Neoplasm↗

PubMed2001

Expression of nerve growth factors in pancreatic neural tissue and pancreatic cancer.

One of the characteristics of pancreatic cancer is its tendency to invade neural tissue. We hypothesized that the affinity of cancer cells for nerve tissue is related to the presence of growth factors in neural tissue and their receptors in cancer cells. Sections of pancreatic cancer and normal pancreatic tissue were examined by immunohistochemistry for the expression of the neurotrophins NGF, BDNF, NT-3, NT-4, and their receptors TrkA, TrkB, and TrkC, as well as the low-affinity receptor, p75NTR. TrkA expression was found in duct, islet, and cancer cells; TrkB was found in the alpha-cells of the islet only. The anti-pan-Trk antibody (TrkB3), which is presumed to recognize all three receptors, immunoreacted with duct and acinar cells in normal tissue and with cancer cells. The staining with TrkC was similar to that of TrkA. The low-affinity receptor p75NTR was expressed in the neural tissue and in scattered duct cells of the normal tissue only. Duct and acinar cells, as well as neural tissue and cancer cells, showed weak to strong immunoreactivity with NGF. NT-3 expression was noted in capillary endothelia and erythrocytes. NT-4 showed specific staining for ductule cells. The expression and distribution of neurotrophins and their receptors suggest their role in the potential of pancreatic cancer cells for neural invasion.

Humans↗

PubMed2001

Early changes in islet hormone secretion in the hamster pancreatic cancer model.

The diabetic state that is seen at a high frequency in association with pancreatic cancer is characterized by elevated plasma levels of several islet hormones and by marked insulin resistance. Both the diabetic state and insulin sensitivity improve after tumor removal by sub-total pancreatectomy. Impaired glucose tolerance has also been found in the hamster pancreatic cancer model, but conflicting data regarding islet function have been reported. In order to further investigate islet function and secretion during early development of pancreatic cancer, we measured the concentrations of insulin, glucagon, somatostatin, and islet amyloid polypeptide (IAPP) in plasma, pancreatic tissue, and secretin-stimulated pancreatic juice at 12 and 27 weeks after the ductal-cell-specific carcinogen, BOP had been used to induce tumors in Syrian golden hamsters. At 12 weeks after BOP, plasma glucagon levels were significantly increased. An exaggerated plasma-glucose response and concomitant hyperinsulinemia were observed at 27 but not 12 weeks after BOP. Plasma IAPP concentrations, but not glucagon or somatostatin, were elevated at 27 weeks. Tissue concentrations of IAPP were substantially reduced in BOP-treated hamsters at 27 weeks. No differences in hormone concentrations were seen in pancreatic juice from the two groups at either of the two time points investigated. The study showed that islet hormone changes accompany the early development of pancreatic tumors in the hamster pancreatic model. The hormone changes and apparent insulin resistance resemble the metabolic changes found in humans with pancreatic cancer.

Adenocarcinoma↗

PubMed2001

Combination of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and actinomycin D induces apoptosis even in TRAIL-resistant human pancreatic cancer cells.

Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a novel member of the tumor necrosis factor superfamily of cytokines that induces cell death by apoptosis. TRAIL has been shown to be effective in almost two-thirds of solid tumors tested thus far, but its effect on pancreatic cancer cells is unknown. We tested the effect of TRAIL on seven human pancreatic cancer cell lines (HPAF, Panc1, Miapaca2, Bxpc3, Panc89, SW979, and Aspc1) in vitro. Of these cell lines, all but Aspc1 showed a significant dose-dependent increase in apoptosis. The apoptotic rate, as detected by a terminal deoxynucleotidyl transferase-mediated nick end labeling assay, was highest in Bxpc3 (71.5%), followed by HPAF (38.0%), Miapaca2 (24.9%), Panc1 (16.1%), Panc89 (15.8%), SW979 (13.9%), and Aspc1 (5.2%). Multiple treatments were more effective than a single treatment and caused a sustained and profound cell death in all but Aspc1 cells. There was no correlation between the effect of TRAIL and the differentiation grade of the cell lines, p53 mutation, or bcl-2 or bax expression. The resistance of Aspc1 cells to TRAIL was not related to the lack of TRAIL receptors. The combination of actinomycin D and TRAIL induced an almost complete lysis of Aspc1 cells, whereas actinomycin D alone had no effect on cell survival but inhibited the expression of the Flice inhibitory protein, which is assumed to play a role in the apoptotic pathway of TRAIL. Thus, the combination of actinomycin D and TRAIL appears to be a promising approach for the therapy of pancreatic cancers resistant to TRAIL.

Antibiotics, Antineoplastic↗

PubMed2000

Inhibition of growth and metastatic progression of pancreatic carcinoma in hamster after somatostatin receptor subtype 2 (sst2) gene expression and administration of cytotoxic somatostatin analog AN-238.

The sst2 somatostatin receptor mediates the antiproliferative effects of somatostatin analogs. The present study demonstrates that stable expression of sst2 in the hamster pancreatic cancer cells PC-1 and PC-1.0 activates an autocrine negative loop leading to an in vitro inhibition of cell proliferation. In vivo studies conducted in Syrian golden hamsters after orthotopic implantation of PC-1.0 cells showed that both tumor growth and metastatic progression of allografts containing 100% of sst2-expressing cells were significantly inhibited for up to 20 days after implantation, as compared with control allografts that did not express sst2. A local antitumor bystander effect was observed after induction of mixed tumors containing a 1:3 ratio of sst2-expressing cells to control cells. Tumor volume and incidence of metastases of mixed tumors were significantly reduced at day 13 post implantation. This effect decreased with time as at day 20, growth of mixed tumors was similar to that of control tumors. After administration of the cytotoxic somatostatin conjugate AN-238 on day 13, antitumor bystander effect observed in mixed tumors was significantly extended to day 20. We also observed that in vitro invasiveness of sst2-expressing PC-1.0 cells was significantly reduced. Tyrosine dephosphorylation of E-cadherin may participate in restoring the E-cadherin function, reducing in turn pancreatic cancer cell motility and invasiveness. This dephosphorylation depends on the tyrosine phosphatase src homology 2-containing tyrosine phosphatase 1 (SHP-1) positively coupled to sst2 receptor. The inhibitory effect of sst2 gene expression on pancreatic cancer growth and invasion combined with chemotherapy with targeted cytotoxic somatostatin analog administration provides a rationale for a therapeutic approach to gene therapy based on in vivo sst2 gene transfer.

Animals↗

PubMed2000

In vitro induction of giant cell tumors from cultured hamster islets treated with N-Nitrosobis(2-Oxopropyl)amine.

Giant cell carcinoma of the pancreas is a rare tumor. Its histogenesis is still controversial. In a Syrian hamster pancreatic cancer model, tumors similar to human giant cell carcinomas have been induced at an extremely low rate of incidence and after the use of high doses of pancreatic carcinogens. Thus far no tumors of giant cell type have been induced by the in vitro treatment of hamster pancreatic ductal cells with the potent pancreatic carcinogen N-nitrosobis(2-oxopropyl)amine (BOP). In the present study we report the induction of giant cell carcinoma from hamster islets treated with BOP in vitro. The results suggest that in hamsters some component of islet cells, probably stem cells, are the origin of giant cell carcinoma.

Animals↗

PubMed2000

Maintenance of human islets in long-term culture.

The long-term maintenance of human islets in culture has remained a challenge. Despite advancements in culture techniques, human islets proved to have a short life span in vitro. For the first time, we have succeeded in maintaining human islets in a defined culture medium for more than 12 months. Freshly isolated islets from a 38-year-old donor were cultured in M3:5 medium and placed on a rocker for 14 days to remove contaminated exocrine and mesenchymal cells which attached to the bottom. The floating islets were purified by daily hand-picking and transfer into fresh medium. After 14 days, purified islets were allowed to attach to the bottom of the flasks and to expand. At various time points, islets were examined immunohistochemically and electron microscopically, and the secretion of islet hormones and their mRNA were determined by radioimmunoassay and reverse transcriptase polymerase chain reaction, respectively. Within seven days of culture, ductular and acinar cells developed within the initially normal islets. With time, exocrine cell types expanded while the number of the endocrine cells and their secretion decreased. At day 60, only a few endocrine cells were identifiable, whereas most of the cells appeared undifferentiated and expressed cytokeratin 7 and 19, neuron specific enolase, tomato lectin, phaseolus leucoagglutinin, laminin, and vimentin. After 60 days, the culture consisted entirely of undifferentiated cells which could be maintained in culture for 270 days before they became senescent. This is the first report on the long-term maintenance of human islet cells in culture and allows an insight into the complex process of endocrine cell differentiation.

Adult↗

PubMed2000

Differentiation of islet cells in long-term culture.

Our previous studies in the hamster pancreatic cancer model have shown that exocrine pancreatic cancer arises from ductal/ductular cells, as well as from within the islets, most probably from islet precursor (stem) cells. To identify and characterize these cells, we established a long-term culture from isolated hamster islets and investigated their growth, differentiation, and expression of biomarkers. Islets maintained their original form and structure within the first 14 days in culture. However, beginning at day 7, ductular structures began to form within the islets. At day 21 in culture, acinar cells, intermediary cells, oncocytes, and cells comparable to pancreatic hepatocytes also appeared between ductular and endocrine cells. The number of duct-like cells gradually increased, whereas the number of hormone-producing cells decreased. After 35 days in culture, the exocrine cells disappeared, and undifferentiated cells formed a monolayer. These cells expressed cytokeratins, alpha1-antitrypsin, transforming growth factor-alpha, epidermal growth factor receptor, carbonic anhydrase II, vimentin, laminin, and showed binding to tomato lectin and Phaseolus vulgaris leukoagglutinin. They did not express the regulatory transcriptional factors, insulin-promoting factor 1, NKx6.1, Pax6, and NeuroD. The results thus indicate that islet cells have potential to form exocrine cells. At present, it is not clear whether these cells originate from preexisting stem cells or from transdifferentiated islet cells.

Animals↗

PubMed2000

Establishment of human pancreatic ductal cells in a long-term culture.

Cultivation and preservation of human pancreatic ductal cells have remained a challenge. With a defined culture medium and refinement of culturing techniques, we have been able to maintain human pancreatic ductal cells without any genetic manipulation in culture for more than 16 months. Freshly isolated ductal fragments were placed on a rocker in M3:5 medium free of collagen for 14 days to remove fibroblasts and endocrine cells before allowing them to attach. The cells produced an excessive amount of mucin and expressed the duct specific cytokeratins (CK) 7 and 19, DU-PAN2, CA19-9, carbonic anhydrase II (CA II), and secretin receptors. During the course of the culture, however, the cells gradually lost the expression of CA II, secretin receptors, DU-PAN2, and CA 19-9 and assumed an undifferentiated phenotype, which showed an upregulation of transforming growth factor alpha (TGFalpha) and epidermal growth factor receptor (EGFR), an increase in the expression of Ki-67, and an increased binding to Phaseolus vulgaris leucoagglutinin (PHA-L) and tomato lectin. These ductal cells present a useful source with which to study physiologic aspects of ductal cells including differentiation.

Adult↗