Resistin aggravates the expression of proinflammatory cytokines in cerulein­stimulated AR42J pancreatic acinar cells.

Resistin, an adipocytokine secreted by fat tissues, has been shown to be associated with increased local and systemic complications in acute pancreatitis (AP). However, the mechanism underlying the effect of resistin in the aggravation of AP remains to be elucidated. The aim of the present study was to investigate the functional consequences of exposing rat pancreatic acinar cells to resistin and to determine whether it amplifies proinflammatory signaling in an in vitro AP model. AR42J cells pretreated with recombinant resistin were activated by cerulein as an in vitro model of AP. The secretion of amylase was measured to evaluate the cytotoxic effect. The mRNA expression levels of tumor necrosis factor (TNF)­α and interleukin (IL)­6 were determined using reverse transcription­quantitative polymerase chain reaction analysis. The nuclear protein expression levels of the nuclear factor (NF)­κB p65 subunit were determined using western blot analysis. Resistin treatment significantly increased the secretion of amylase, and the mRNA expression levels of TNF­α and IL­6 in the cerulein­induced in vitro AP model. High protein levels of the NF­κB p65 subunit were observed in the nuclei of cells in the resistin­treated AP model, compared with the untreated AP model. Pretreatment of the in vitro resistin­treated AP model with the NF­κB inhibitor, pyrrolidine dithiocarbamate decreased the protein expression of the NF­κB p65 subunit in nuclei, and significantly attenuated the increased mRNA expression levels of TNF­α and IL­6 induced by resistin. The results of the present study showed that resistin increased the production of the TNF­α and IL­6 proinflammatory cytokines via the NF­κB­dependent pathway during AP. Thus, the overproduction of obesity­associated resistin and the associated amplification of the inflammatory response may result in the aggravation of AP severity.

Sensitization to and Challenge with Gliadin Induce Pancreatitis and Extrapancreatic Inflammation in HLA-DQ8 Mice: An Animal Model of Type 1 Autoimmune Pancreatitis.

BACKGROUND/AIMS: The aim of this study was to establish a pathogenetic mechanism of pancreatitis in celiac disease and IgG4-related disease using gluten-sensitive human leukocyte antigen (HLA)-DQ8 transgenic mice. METHODS: Transgenic mice expressing HLA-DQ8 genes were utilized. Control mice were not sensitized but were fed gliadin-free rice cereal. Experimental groups consisted of gliadin-sensitized and gliadin-challenged mice; nonsensitized mice with cerulein hyperstimulation; and gliadin-sensitized and gliadinchallenged mice with cerulein hyperstimulation. RESULTS: Gliadin-sensitized and gliadin-challenged mice with cerulein hyperstimulation showed significant inflammatory cell infiltrates, fibrosis and acinar atrophy compared with the control mice and the other experimental groups. The immunohistochemical analysis showed greater IgG1-positive plasma cells in the inflammatory infiltrates of gliadin-sensitized and gliadin-challenged mice with cerulein hyperstimulation compared with the control mice and the other experimental groups. Gliadin-sensitized and gliadin-challenged mice with cerulein hyperstimulation or gliadin-sensitized and gliadinchallenged mice showed IgG1-stained inflammatory cell infiltrates in the extrapancreatic organs, including the bile ducts, salivary glands, kidneys, and lungs. CONCLUSIONS: Gliadinsensitization and cerulein hyperstimulation of gluten-sensitive HLA-DQ8 transgenic mice resulted in pancreatitis and extrapancreatic inflammation. This animal model suggests that chronic gliadin ingestion in a susceptible individual with the HLA-DQ8 molecule may be associated with pancreatitis and extrapancreatic inflammation.

SiRNA-mediated PIAS1 silencing promotes inflammatory response and leads to injury of cerulein-stimulated pancreatic acinar cells via regulation of the P38MAPK signaling pathway.

Our aim in this study was to investigate the changes of inflammatory response by protein inhibitor of activated signal transducer and activator of transcription 1 (PIAS1) gene silencing treatment in cerulein-stimulated AR42J cells, and relate them to changes in cell injury, thus providing evidence for developing clinical therapies. This study examined the effects of cerulein on the activity of P38 mitogen activated protein kinase (P38MAPK), c-jun NH2-terminal kinase/stress-activated protein kinase and the inflammatory mediators released by PIAS1 gene-silenced AR42J cells. Consequently, the markers including DNA ladder, cell apoptotic rat, cell cycles, levels of cell cycle and apoptotic related factors were used to determine the effects of PIAS1 gene silencing on the cerulein-induced cell injury. The results indicated that in the cerulein-stimulated PIASI silencing cells, the activity of P38MAPK was enhanced, while at the same time, the levels of inflammatory mediators such as the tumor necrosis factor-alpha, interleukin (IL)-1beta, IL-6 and matrix metallopeptidase-9, were markedly higher than those of other cerulein-stimulated cells. Thus, the cerulein-stimulated PIASI gene-silenced cells obviously increased cell arrest in the G1/M phase by increasing P21 and P27 expression, and also induced apoptosis by regulating the P53 signaling pathway. This study suggests that the down-regulation of PIAS1 is efficacious at enhancing the expression of inflammatory mediators and inducing cell injury in acute pancreatitis (AP), thus deteriorating the severity of disease. It provides evidence that PIAS1 is a potential therapeutic target for AP.

The role of pro-inflammatory molecules and pharmacological agents in acute pancreatitis and sepsis.

Acute pancreatitis (AP) and sepsis are inflammatory disorder varying in magnitude of response to infection or inflammatory stimuli. The specific role of various causative factors in AP, septic shock, current pharmacological treatments, animal models, role of infiltrating cells and novel molecules that play an important role in the disease progression to sepsis are explored. AP is an inflammatory disease of the pancreas. Over the years accumulating evidence suggests numerous molecules as key regulators of the inflammatory signaling cascade such as selectins, chemokine signaling and expression of intergrins on leukocytes facilitate adhesion to vessel walls. Inhibition of any of these molecules has proven to be effective in animal models of AP. Recently, the biochemical role of hydrogen sulfide (H(2)S) and substance P in caerulein induced AP and in cecal ligation and puncture induced sepsis and their role in the pathogenesis of the disease have highlighted the importance of novel molecules as therapeutic targets in addition to the known pro-inflammatory molecules, cytokines and chemoattractant chemokines and their receptors upregulated in AP and sepsis. This review aims to give an overview of the multifaceted complex interactions in a prearranged fashion and their functional role in the inflammatory process that afflict AP and sepsis. The interlinking molecules in AP and sepsis emphasize the similarities in the inflammatory response and the importance of pharmacological agents that reduce or inhibit the progression to chronic stage.

Cellular immune reaction in the pancreas is induced by constitutively active IkappaB kinase-2.

BACKGROUND: Activation of the nuclear factor kappaB (NF-kappaB) system is a major event in acute and chronic inflammatory processes. NF-kappaB cascades are comprised of IkappaB kinases, IkappaBs and NF-kappaB dimers. Little is known of the individual roles of these proteins in organ specific inflammation. The aim of the present study was to analyse the consequences of ectopic IkappaB kinase-2 (IKK2) activation in the pancreas of mice. METHODS: Transgenic mice were generated using an inducible genetic system (tet system) to conditionally overexpress a gain of function mutant of IKK2 (tetO-IKK2-EE) in the pancreas. To achieve transgene expression in the pancreas, these animals were crossed with CMV-rtTA mice that are known to express the rtTA protein in the pancreas. RESULTS: In these double transgenic animals, doxycycline treatment induced expression of IKK2-EE (IKK2(CA)) in pancreatic acinar cells resulting in moderate activation of the IkappaB kinase complex, as measured by the immune complex kinase assay, and up to 200-fold activation of the transgene expression cassette, as detected by luciferase assay. IKK2(CA) expression in the pancreas had a mosaic appearance. Ectopic IKK2(CA) mostly activated the classical NF-kappaB pathway. The activation level of the NF-kappaB cascade induced by IKK2(CA) was considerably lower compared with that observed after supramaximal caerulein stimulation but still led to the formation of leucocyte infiltrates first observed after 4 weeks of doxycycline stimulation with a maximum after 8-12 weeks. The infiltrates were mainly composed of B lymphocytes and macrophages. Increased mRNA levels of tumour necrosis factor alpha and RANTES were detected in pancreatic acinar cells. However, only minor damage to pancreatic tissue was observed. A combination of supramaximal caerulein stimulation with induction of IKK2(CA) caused increased tissue damage compared with either IKK2(CA) or caerulein alone. CONCLUSIONS: Our observations suggest that the role of IKK2 activation in pancreatic acini is to induce leucocyte infiltration, but at a moderate level of activation it is not sufficient to induce pancreatic damage in mice. The IKK2(CA) induced infiltrations resemble those observed in autoimmune pancreatitis, indicating a role for IKK2/NF-kappaB in this disease. IKK2(CA) in pancreatic acinar cells increases tissue damage of secretagogue induced experimental pancreatitis underlining the proinflammatory role of the IKK/NF-kappaB pathway in this disease.

Characterization of antral gastrin cells with region-specific antisera.

A number of gastrin antisera, which in radioimmunoassay systems showed no or negligible cross-reactivity towards the structurally and functionally related peptide cholecystokinin were found to react with both gastrin and cholecystokinin cells when used for immunocytochemistry. This discrepancy was shown to be due either to reactivity against a COOH-terminal region common to gastrin and cholecystokinin or to the occurrence of heterogenous antibody populations in the antisera. By differential absorptions the latter type of antisera could be rendered specific for gastrin. Antisera reactive against the NH2-terminal, middle or COOH-terminal regions of human heptadecapeptide gastrin were prepared and together with a specific cholecystokinin antiserum used for the characterization of antral gastrin cells of different species. The results indicate that only the COOH-terminal region of gastrin is conserved during evolution.

Radioimmunoassay of the gastrointestinal hormones.

The principle and types of saturation analysis are discussed on the basis of data in the literature. This is followed by summarizing the main steps of radioimmunoassay (RIA) of gastrointestinal hormones. The normal fasting blood levels of these hormones are also given. RIA measurement of the individual hormones is discussed critically, and attention is called to the theoretical importance of immune cross-reactions. The RIA model developed by the author for measuring the C-terminal pentapeptide of gastrin is demonstrated.

Cytochemical and immunofluorescence investigations of insulin-like producing cells in the intestine of Mytilus edulis L. (Bivalvia).

Insulin-like immunoreactivity can be localized to cells of the intestine in the area of the hepatopancreas of Mytilus edulis L. No cross-reactivity can be obtained with anti-glucagon, anti-gastrin, anti-pentagastrin or anti-caerulin. The cells containing the substance immunoreactive to mammalian anti-insulin, can be restrained with paraldehyde-fuchsin.