Mechanisms involved in PGE2-induced transactivation of the epidermal growth factor receptor in MH1C1 hepatocarcinoma cells.

Background It is important to understand the mechanisms by which the cells integrate signals from different receptors. Several lines of evidence implicate epidermal growth factor (EGF) receptor (EGFR) in the pathophysiology of hepatocarcinomas. Data also suggest a role of prostaglandins in some of these tumours, through their receptors of the G protein-coupled receptor (GPCR) family. In this study we have investigated mechanisms of interaction between signalling from prostaglandin receptors and EGFR in hepatocarcinoma cells. Methods The rat hepatocarcinoma cell line MH1C1 and normal rat hepatocytes in primary culture were stimulated with EGF or prostaglandin E2 (PGE2) and in some experiments also PGF2α. DNA synthesis was determined by incorporation of radiolabelled thymidine into DNA, phosphorylation of proteins in signalling pathways was assessed by Western blotting, mRNA expression of prostaglandin receptors was determined using qRT-PCR, accumulation of inositol phosphates was measured by incorporation of radiolabelled inositol, and cAMP was determined by radioimmunoassay. Results In the MH1C1 hepatocarcinoma cells, stimulation with PGE2 or PGF2α caused phosphorylation of the EGFR, Akt, and ERK, which could be blocked by the EGFR tyrosine kinase inhibitor gefitinib. This did not occur in primary hepatocytes. qRT-PCR revealed expression of EP1, EP4, and FP receptor mRNA in MH1C1 cells. PGE2 stimulated accumulation of inositol phosphates but not cAMP in these cells, suggesting signalling via PLCβ. While pretreatment with EP1 and EP4 receptor antagonists did not inhibit the effect of PGE2, pretreatment with an FP receptor antagonist blocked the phosphorylation of EGFR, Akt and ERK. Further studies suggested that the PGE2-induced signal was mediated via Ca2+ release and not PKC activation, and that it proceeded through Src and shedding of membrane-bound EGFR ligand precursors by proteinases of the ADAM family. Conclusion The results indicate that in MH1C1 cells, unlike normal hepatocytes, PGE2 activates the MEK/ERK and PI3K/Akt pathways by transactivation of the EGFR, thus diversifying the GPCR-mediated signal. The data also suggest that the underlying mechanisms in these cells involve FP receptors, PLCβ, Ca2+, Src, and proteinase-mediated release of membrane-associated EGFR ligand(s)

Role of protein kinase C and epidermal growth factor receptor signalling in growth stimulation by neurotensin in colon carcinoma cells

<p>Abstract</p> <p>Background</p> <p>Neurotensin has been found to promote colon carcinogenesis in rats and mice, and proliferation of human colon carcinoma cell lines, but the mechanisms involved are not clear. We have examined signalling pathways activated by neurotensin in colorectal and pancreatic carcinoma cells.</p> <p>Methods</p> <p>Colon carcinoma cell lines HCT116 and HT29 and pancreatic adenocarcinoma cell line Panc-1 were cultured and stimulated with neurotensin or epidermal growth factor (EGF). DNA synthesis was determined by incorporation of radiolabelled thymidine into DNA. Levels and phosphorylation of proteins in signalling pathways were assessed by Western blotting.</p> <p>Results</p> <p>Neurotensin stimulated the phosphorylation of both extracellular signal-regulated kinase (ERK) and Akt in all three cell lines, but apparently did so through different pathways. In Panc-1 cells, neurotensin-induced phosphorylation of ERK, but not Akt, was dependent on protein kinase C (PKC), whereas an inhibitor of the β-isoform of phosphoinositide 3-kinase (PI3K), TGX221, abolished neurotensin-induced Akt phosphorylation in these cells, and there was no evidence of EGF receptor (EGFR) transactivation. In HT29 cells, in contrast, the EGFR tyrosine kinase inhibitor gefitinib blocked neurotensin-stimulated phosphorylation of both ERK and Akt, indicating transactivation of EGFR, independently of PKC. In HCT116 cells, neurotensin induced both a PKC-dependent phosphorylation of ERK and a metalloproteinase-mediated transactivation of EGFR that was associated with a gefitinib-sensitive phosphorylation of the downstream adaptor protein Shc. The activation of Akt was also inhibited by gefitinib, but only partly, suggesting a mechanism in addition to EGFR transactivation. Inhibition of PKC blocked neurotensin-induced DNA synthesis in HCT116 cells.</p> <p>Conclusions</p> <p>While acting predominantly through PKC in Panc-1 cells and via EGFR transactivation in HT29 cells, neurotensin used both these pathways in HCT116 cells. In these cells, neurotensin-induced activation of ERK and stimulation of DNA synthesis was PKC-dependent, whereas activation of the PI3K/Akt pathway was mediated by stimulation of metalloproteinases and subsequent transactivation of the EGFR. Thus, the data show that the signalling mechanisms mediating the effects of neurotensin involve multiple pathways and are cell-dependent.</p

Role of LPAR3, PKC and EGFR in LPA-induced cell migration in oral squamous carcinoma cells

Background Oral squamous cell carcinoma is an aggressive neoplasm with serious morbidity and mortality, which typically spreads through local invasive growth. Lysophosphatidic acid (LPA) is involved in a number of biological processes, and may have a role in cancer cell migration and invasiveness. LPA is present in most tissues and can activate cells through six different LPA receptors (LPAR1-6). Although LPA is predominantly promigratory, some of the receptors may have antimigratory effects in certain cells. The signalling mechanisms of LPA are not fully understood, and in oral carcinoma cells the specific receptors and pathways involved in LPA-stimulated migration are unknown. Methods The oral carcinoma cell lines E10, SCC-9, and D2 were investigated. Cell migration was studied in a scratch wound assay, and invasion was demonstrated in organotypic three dimensional co-cultures. Protein and mRNA expression of LPA receptors was studied with Western blotting and qRT-PCR. Activation of signalling proteins was examined with Western blotting and isoelectric focusing, and signalling mechanisms were further explored using pharmacological agents and siRNA directed at specific receptors and pathways. Results LPA stimulated cell migration in the two oral carcinoma cell lines E10 and SCC-9, but was slightly inhibitory in D2. The receptor expression profile and the effects of specific pharmacological antagonist and agonists indicated that LPA-stimulated cell migration was mediated through LPAR3 in E10 and SCC-9. Furthermore, in both these cell lines, the stimulation by LPA was dependent on PKC activity. However, while LPA induced transactivation of EGFR and the stimulated migration was blocked by EGFR inhibitors in E10 cells, LPA did not induce EGFR transactivation in SCC-9 cells. In D2 cells, LPA induced EGFR transactivation, but this was associated with slowing of a very high inherent migration rate in these cells. Conclusion The results demonstrate LPA-stimulated migration in oral carcinoma cells through LPAR3, mediated further by PKC, which acts either in concert with or independently of EGFR transactivation

En ett-års merke-gjenfangst studie av gjedde (Esox lucius) i Borrevannet, i sørøst Norge: Estimater av vitale demografiske og populasjonsdynamiske rater

Northern pike (Esox lucius) is distributed through Europe, Asia and North-America and is an important species in both commercial and recreational fisheries. It is a top predator that has an important role in regulation fish communities through predation and cannibalism. Borrevannet is a shallow and eutrophic lake located in Vestfold County, SE Norway. Cyprinids (mainly roach (Rutilus rutilus) and bleach (Alburnus alburnus)) dominate the fish community and pike along with large individuals of perch (Perca fluviatilis) constitute top predators. There is a latent conflict related to the way pike is supposed to be managed in the system. Anglers want to favor a large proportion of large pike individuals in the population, whereas regional water management authorities would rather favor a large proportion of medium-sized pike as this will improve the water quality due to medium-sized pike being the more effective roach predators. In order to arrive at a sustainable and justifiable pike management policy of the lake, detailed information about the pike population is a prerequisite. How the pike would be managed in Borrevannet could be a conflict of interest for the landowners whether deciding to focus on sports fishing or to improve the water quality. It is on that basis that I have done my study on the pike population in Borrevannet. The objective of this study was to estimate the population size, survival and migration rate of Borrevannet pike through capture-mark-recapture (CMR) analysis over 5 mark-recapture occasions covering a year. From scale samples individual back-calculated growth trajectories could be read and size-at age data was used for analysis of ambient temperature on growth. The population size of Borrevannet pike (larger than 40 cm) was estimated to be around 500 individuals. First-year individual growth in male pike responded positively to increasing ambient temperature, whereas females showed the opposite effect. Second- and third-year growths were either negatively or insignificantly affected by monthly growth-season temperatures. Cormack-Jolly-Seber (CJS) models were fitted the CMR-data used and the most supported model (AIC based) predicted that recapture probability for pike 40-65 cm was positively size-dependent during spring and negatively during fall. This finding can be related to differential activity levels between small and larger individuals during the two seasons. Borrevannet pike had an estimated monthly survival of 0.97, corresponding to an annual survival rate of 0.61. Most of the pike did not disperse far from the original marking point when recaptured (50 % dispersed less than 163 m), but there were a few individuals who dispersed up to more than 2000 m. Borrevannet landowners association want to utilize the fishing resource for recreational- and sports fishing, but also improve the water quality in the lake. This may come in conflict with keeping a portion of large pike in the population for sports fishing. Estimated density of pike was low compared to other lakes. This may be because large pike, through cannibalism, forage on smaller individuals. Since the mark-recapture analysis did not show any size dependent survival, predation on smaller pike is uncertain. The mark-recapture study needs to continue to determine the relationship between harvest mortality and natural mortality via cannibalism.M-N

Inhibitory effects of prostaglandin E2 on collagen synthesis and cell proliferation in human stellate cells from pancreatic head adenocarcinoma

Background Several studies have described an increased cyclooxygenase-2 (COX-2) expression in pancreatic cancer, but the role of COX-2 in tumour development and progression is not clear. The aim of the present study was to examine expression of COX-2 in cancer cells and stromal cells in pancreatic cancer specimens, and to explore the role of PGE2 in pancreatic stellate cell proliferation and collagen synthesis. Methods Immunohistochemistry and immunofluorescence was performed on slides from whole sections of tissue blocks using antibodies against COX-2 and α-smooth muscle actin (αSMA). Pancreatic stellate cells (PSC) were isolated from surgically resected tumour tissue by the outgrowth method. Cells were used between passages 4 and 8. Collagen synthesis was determined by [3H]-proline incorporation, or by enzyme immunoassay measurement of collagen C-peptide. DNA synthesis was measured by incorporation of [3H]-thymidine in DNA. Cyclic AMP (cAMP) was determined by radioimmunoassay. Collagen 1A1 mRNA was determined by RT-qPCR. Results Immunohistochemistry staining showed COX-2 in pancreatic carcinoma cells, but not in stromal cells. All tumours showed positive staining for αSMA in the fibrotic stroma. Cultured PSC expressed COX-2, which could be further induced by interleukin-1β (IL-1β), epidermal growth factor (EGF), thrombin, and PGE2, but not by transforming growth factor-β1 (TGFβ). Indirect coculture with the adenocarcinoma cell line BxPC-3, but not HPAFII or Panc-1, induced COX-2 expression in PSC. Treatment of PSC with PGE2 strongly stimulated cAMP accumulation, mediated by EP2 receptors, and also stimulated phosphorylation of extracellular signal-regulated kinase (ERK). Treatment of PSC with PGE2 or forskolin suppressed both TGFβ-stimulated collagen synthesis and PDGF-stimulated DNA synthesis. Conclusions The present results show that COX-2 is mainly produced in carcinoma cells and suggest that the cancer cells are the main source of PGE2 in pancreatic tumours. PGE2 exerts a suppressive effect on proliferation and fibrogenesis in pancreatic stellate cells. These effects of PGE2 are mediated by the cAMP pathway and suggest a role of EP2 receptors

HiF-Notat 2013:1 Snefjordvassdraget. Driftsplan

&lt;span style="font-size: 11.5pt; font-family: 'Times New Roman',serif; color: black; line-height: 107%; mso-fareast-font-family: Calibri; mso-fareast-theme-font: minor-latin; mso-ansi-language: NO-BOK; mso-fareast-language: EN-US; mso-bidi-language: AR-SA;"&gt;Høgskolen i Finnmark og Naturtjenester i Nord har fått oppdrag fra Snefjord jeger- og fiskeforening om utarbeiding av driftsplan for Snefjordvassdraget. Det ble utført et prøvefiske med multigarn, videoovervåkning i utløpselv og et el-fiske. Resultatene viser en god bestand av røye og ørret. Det finnes også en liten bestand av laks. Hovedvandringstidspunkt for anadrom røye er i juli og anadrom ørret vandrer i september/oktober (kameraovervåkning).&lt;/span&gt;</jats:p