Analysing migrations of Atlantic cod Gadus morhua in the north-east Atlantic Ocean: then, now and the future.

The application of data storage tags bears the potential for a quantum leap in the research on fish migrations, because not only first-capture and recapture positions are known, but at least theoretically, the migration path during the period at large can be reconstructed. Position, however, cannot be measured directly but has to be estimated using the available data on light, temperature, pressure and salinity. The reconstructed locations based on advanced estimation techniques have been termed geolocations. Examples are discussed which illustrate the applicability of geolocations in individual path descriptions, separation of reproductively isolated populations, timing and areas of spawning, tidal transport and use of protected areas. The examples are based on archival tag data from the North Sea, the Baltic Sea, the Barents Sea and Faroese and Icelandic Waters. Besides presenting the state-of-the-art geolocations for cod Gadus morhua in the north-east Atlantic Ocean, the major aim of this review is to raise awareness of gaps in knowledge and to identify ideas for new research.

The insulinoma-associated 1: a novel promoter for targeted cancer gene therapy for small-cell lung cancer.

The insulinoma-associated 1 (INSM1) gene is expressed exclusively during early embryonal development, but has been found re-expressed at high levels in neuroendocrine tumors. The regulatory region of the INSM1 gene is therefore a potential candidate for regulating expression of a therapeutic gene in transcriptionally targeted cancer gene therapy against neuroendocrine tumors. We analyzed expression of a reporter gene from a 1.7 kb region of the INSM1 promoter in a large number of small-cell lung cancer (SCLC) cell lines. This INSM1 promoter region showed very high levels of expression in most of the SCLC cell lines and expression was absent in cell lines of non-neuroendocrine origin. Inclusion of the general transcriptional enhancer from SV40 compromised the specificity of the promoter and did not enhance transcription in most of the SCLC cell lines. For comparison, the region of the gastrin releasing peptide (GRP) previously suggested for SCLC gene therapy was analyzed in a similar manner. High expression was observed for a number of cell lines, but unlike for the INSM1 promoter, reporter gene expression from the GRP promoter did not correlate to the relative GRP mRNA levels, demonstrating that this region may not contain all necessary regulatory elements. Expression of the suicide gene herpes simplex virus thymidine kinase (HSV-TK) from the INSM1 promoter in combination with treatment with the prodrug ganciclovir (GCV) caused a significant increase in GCV sensitivity specifically in INSM1-expressing cell lines. The INSM1 promoter is therefore a potential novel tool for transcriptionally targeted gene therapy for neuroendocrine tumors.

Differential response to gefitinib of cells expressing normal EGFR and the mutant EGFRvIII.

Epidermal growth factor receptor (EGFR) is frequently amplified and/or mutated in a number of human tumours and abnormal signalling from this receptor is believed to contribute to the malignant phenotype seen in these tumours. Gefitinib is a small molecule inhibitor that specifically binds and inhibits the EGFR tyrosine kinase and has been shown to inhibit the growth, proliferation, survival and invasion of a range of tumour cells overexpressing EGFR. However, clinical response to gefitinib has failed to correlate with EGFR levels and activity, indicating that other molecular mechanisms such as downstream signalling and mutations could be of importance in predicting clinical response. We therefore investigated the effect of the specific EGFR inhibitor gefitinib on the phosphorylation level, signalling and growth of cells expressing the naturally occurring constitutively active EGFR variant EGFRvIII, a low nontransforming level of EGFR and a high transforming level of EGFR. Results show that levels of gefitinib sufficient to suppress EGFR phosphorylations, EGFR-mediated proliferation and EGFR-mediated anchorage-independent growth are not sufficient to inhibit these features in cells expressing EGFRvIII. Furthermore, the data indicate that long-term exposure of EGFRvIII-expressing cells to low concentrations of gefitinib (0.01-0.1 microM) result in increased phosphotyrosine load of the receptor, increased signalling to ERK and stimulation of proliferation and anchorage-independent growth, presumably by inducing EGFRvIII dimerisation. Higher concentrations of gefitinib (1-2 microM), on the other hand, significantly decreased EGFRvIII phosphotyrosine load, EGFRvIII-mediated proliferation and anchorage-independent growth. Further studies are needed to investigate the implications of these important findings in the clinical setting.

Profile of differentially expressed genes mediated by the type III epidermal growth factor receptor mutation expressed in a small-cell lung cancer cell line.

Previous studies have shown a correlation between expression of the EGF receptor type III mutation (EGFRvIII) and a more malignant phenotype of various cancers including: non-small-cell lung cancer, glioblastoma multiforme, prostate cancer and breast cancer. Thus, a detailed molecular genetic understanding of how the EGFRvIII contributes to the malignant phenotype is of major importance for future therapy. The GeneChip Hu6800Set developed by Affymetrix was used to identify changes in gene expression caused by the expression of EGFRvIII. The cell line selected for the study was an EGF receptor negative small-cell-lung cancer cell line, GLC3, stably transfected with the EGFRvIII gene in a Tet-On system. By comparison of mRNA levels in EGFRvIII-GLC3 with those of Tet-On-GLC3, it was found that the levels of mRNAs encoding several transcription factors (ATF-3, JunD, and c-Myb), cell adhesion molecules (CD36, CD24), signal transduction related molecules (MKP-1) and other molecules related to cancer (CD98, thymosin beta-10) were altered in the EGFRvIII transfected cell line. Northern hybridisations and Western blot analyses were used to verify selected results. The results indicate that expression of EGFRvIII alters expression of genes involved in the control of cell growth, survival and motility.

The type III epidermal growth factor receptor mutation. Biological significance and potential target for anti-cancer therapy.

Mutations in the epidermal growth factor receptor occur frequently in a number of human tumours including gliomas, non-small-cell lung carcinomas, ovarian carcinomas and prostate carcinomas. The type III epidermal growth factor receptor mutation (variously named EGFRvIII, de2-7 EGFR or AEGFR), which lacks a portion of the extracellular ligand binding domain, is the most common. Here, we review the current status with regard to the role of EGFRvIII in human cancers. A detailed discussion of the formation of EGFRvIII and its structure at the protein level are likewise included along with a discussion of its more functional roles. The design and use (preclinical and clinical) of small molecule inhibitors, antibodies, and antisense oligonucleotides against wild-type EGFR are considered in detail as these strategies can be directly adapted to target EGFRvIII. Finally, the status of EGFRvIII targeted therapy is reviewed.

Coregulation of glucose uptake and vascular endothelial growth factor (VEGF) in two small-cell lung cancer (SCLC) sublines in vivo and in vitro.

We examined the relationship between (18)F- labeled 2-fluro-2-deoxy-d-glucose (FDG) uptake, and expression of glucose transporters (GLUTs) in two human small-cell lung cancer (SCLC) lines CPH 54A and CPH 54B. Changes in the expression of GLUTs and vascular endothelial growth factor (VEGF) during 12-, 18-, and 24 hours of severe hypoxia in vivo (xenografts) and in vitro (cell cultures) were recorded for both tumor lines. The two SCLC lines are subpopulations of the same patient tumor. In spite of their common genomic origin they represent consistently different metabolic and microenvironmental phenotypes as well as treatment sensitivities. There were higher levels of Glut-1 protein in 54B and a correspondingly higher FDG uptake in this tumor line (P<.001). During hypoxia a significant upregulation of in VEGF mRNA, GLUT-1 mRNA, and Glut-1 and -3 protein occurred with a distinctly different time course in the two cell lines. A similar co-upregulation of GLUT and VEGF was seen in hypoxic tumors of both lines. There were no significant changes of HIF-1alpha mRNA during hypoxia in either of the cell lines. A more detailed understanding of such correlations between glucose metabolism, angiogenesis, and microenvironmental phenotype of tumors, by positron emission tomography (PET) and molecular techniques might further sophisticate our interpretation of glycolytic predominance in tumors as seen by 18FFDG PET.

Relationship between vessel density and expression of vascular endothelial growth factor and basic fibroblast growth factor in small cell lung cancer in vivo and in vitro.

In 21 human small cell lung cancer (SCLC) cell lines, we determined the expression of mRNA and secreted protein levels of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF). The VEGF expression was highly variable between cell lines, with a > 100-fold variation, under identical in vitro conditions. The bFGF expression in cell lines was generally very low. Nine of the cell lines were further analyzed during growth as solid tumor xenografts in nude mice (in vivo). A more uniform VEGF protein expression was present in vivo. Compared with the variable in vitro expression, VEGF was relatively up-regulated in the tumor lines CPH 54A and CPH 54B and down-regulated in GLC 3. One line, DMS 79, had a high VEGF expression in vivo as well as in vitro. The vessel density was determined by Chalkley point counting on CD31 immunostained cryosections of tumors of each of the nine SCLC lines. We found a strong positive correlation between vessel density and tissue VEGF protein expression (r(s) = 0.75; P = 0.02) and a comparatively strong negative correlation (r(s) = -0.80; P = 0.01) between vessel density and tissue bFGF expression. No significant correlation was present between vessel density and in vitro VEGF expression. We conclude that VEGF and bFGF expression is dependent on microenvironmental conditions, as well as cell line-specific factors, and that a strong positive correlation exists between in vivo VEGF expression and vessel density, whereas high tissue levels of bFGF are not correlated with higher vessel densities in SCLC xenografts.

Detection and strain differentiation of infectious bronchitis virus in tracheal tissues from experimentally infected chickens by reverse transcription-polymerase chain reaction. Comparison with an immunohistochemical technique.

Oligonucleotide pairs were constructed for priming the amplification of fragments of nucleocapsid (N) protein and spike glycoprotein (S) genes of avian infectious bronchitis virus (IBV) by reverse transcriptionpolymerase chain reaction (RT-PCR). One oligonucleotide pair amplified a common segment of the N-gene and could detect various strains of IBV in allantoic fluid from inoculated chicken embryos, and in tracheal tissue preparations from experimentally infected chickens. Four pairs of oligonucleotides selectively primed the amplification of the S1 gene of Massachusetts/Connecticut, D1466, D274/D3896 and 793B strains of IBV, respectively. Groups of specific pathogen free chickens were experimentally inoculated with the Massachusetts (H120, M41), the D1466 and the 793B strains of IBV, and tracheal tissue preparations were made from each bird for RT-PCR and for immunohistochemistry (IHC) up to 3 days post-inoculation. The N-gene RT-PCR detected IBV in 82% of the chickens, while IHC only detected IBV in 60%. This difference was significant (P<0.02). The detection rate by N-gene RT-PCR varied from 67 to 100% for the various strains of IBV inoculated. The S1 gene oligonucleotide pairs were applied to the same tissue preparations and they detected specifically the Massachusetts (M41 and H120), the D1466 and the 793B strains of IBV at rates varying between 58 and 92%. When the mixtures of the primers were applied, the detection rate in tissue preparations was reduced to the level of 50 to 67%. It is concluded that the direct detection of IBV in tracheal tissues by RT-PCR is more sensitive than IHC and that the RT-PCR technique is able to distinguish between types of IBV.

The effect of whisky and wine consumption on total phenol content and antioxidant capacity of plasma from healthy volunteers.

OBJECTIVE: To assess whether consumption of 100 ml of whisky or red wine by healthy male subjects increased plasma total phenol content and antioxidant capacity. DESIGN: A Latin square arrangement to eliminate ordering effects whereby, after an overnight fast, nine volunteers consumed 100 ml of red wine, malt whisky or unmatured 'new make' spirit. Each volunteer participated on three occasions one week apart, consuming one of the beverages each time. Blood samples were obtained from the anticubital vein at intervals up to 4h after consumption of the beverages when a urine sample was also obtained. RESULTS: Within 30 min of consumption of the wine and whisky, there was a similar and significant increase in plasma total phenol content and antioxidant capacity as determined by the ferric reducing capacity of plasma (FRAP). No changes were observed following consumption of 'new make' spirit. CONCLUSIONS: Consumption of phenolic-containing alcoholic beverages transiently raises total phenol concentration and enhances the antioxidant capacity of plasma. This is compatible with suggestions that moderate alcohol usage and increased antioxidant intake decrease the risk of coronary heart disease.

Mycoplasma hyopneumoniae infection in pigs: duration of the disease and evaluation of four diagnostic assays.

200 SPF pigs were infected by aerosol with Mycoplasma hyopneumoniae and the development of clinical signs, serological and pathological reactions were studied. Mean time to onset of coughing was 13 days. A mean delay of 9 days was observed from onset of coughing until seroconversion against M. hyopneumoniae as measured by ELISA. At an individual level, the sensitivity for this ELISA was estimated to 98-100% and the specificity to 93-100%. Pasteurella multocida was isolated from the majority of the lungs 4 weeks post inoculation with M. hyopneumoniae and the lung lesions in pigs were significantly larger when P. multocida was present as compared to pigs with M. hyopneumoniae alone. An evaluation of cultivation, immunofluorescence, ELISA and polymerase chain reaction for demonstration of M. hyopneumoniae in lungs showed that all four methods have a high sensitivity in the acute stages of pneumonia. In the later stages the sensitivity of cultivation was superior to the other methods. No differences in specificity were observed between the methods. The antigen-ELISA OD values and the immunofluorescence scores revealed a strong positive correlation. Nasal swabs were additionally used for demonstration of M. hyopneumoniae and the polymerase chain reaction was found superior to the other methods.