Vascular endothelial (VE)-cadherin is exclusively expressed at interendothelial junctions of normal and tumour vessels. In this report, we characterized the transcriptional activity of the human VE-cadherin promoter. Transient transfection assays revealed that sequences at positions À1135/À744 and À166/À5 base pairs are critical for promoter activity in endothelial cells. We show that specific sequences in the proximal region interact with Ets and Sp1 family members. Transgenic mice were created and the human VE-cadherin promoter was able to confer correct temporal and spatial expression on the LacZ gene in embryos. In adults, the transgene was specifically and strongly expressed in the lung, heart, ovary, spleen and kidney glomeruli, whereas expression was weak or absent in the vasculature of other organs, including the brain, thymus, liver and skeletal muscle. Neovessels in tumour grafts and Matrigel implants harboured strong stainings, indicating that promoter activity is enhanced in angiogenic situations. Furthermore, Matrigel and transfection assays showed that VE-cadherin promoter is subjected to bFGF induction. Transgene expression was also noticed in extravascular sites of the central nervous system, suggesting that silencer elements may be located elsewhere in the gene. These results are a first step towards addressing the organ-and tumour-specific regulation of the VE-cadherin gene.

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We isolated an 18-kilobase (kb) genomic selenoprotein P clone from a human placenta library and cloned, sequenced, and characterized the 5-flanking region of the human selenoprotein P gene. Sequence analysis revealed an intron between base pairs (bp) ؊13 and ؊14 upstream of the ATG codon and another one between bp 534 and 535 of the coding region. The major transcription start site of selenoprotein P in human HepG2 hepatocarcinoma cells was mapped to bp ؊70 by 5-rapid amplification of cDNA ends and by primer extension. 1.8 kb of the 5-flanking sequence were fused to a luciferase reporter gene. They exhibited functional promoter activity in HepG2 hepatocarcinoma and Caco2 colon carcinoma cells in transient transfection experiments. Treatment of transfected HepG2 cells with the cytokines interleukin 1␤, tumor necrosis factor ␣, and interferon ␥ repressed promoter activity. Nuclear extracts of interferon ␥-treated cells bound to a signal transducer and activator of transcription response element of the promoter in gel retardation experiments. By transfection of promoter-deletion constructs, a TATA box and a putative SP1 site were identified to be necessary for selenoprotein P transcription. These data indicate that the human selenoprotein P gene contains a strong promoter that is cytokine responsive. Furthermore, selenoprotein P, secreted by the liver, might react as a negative acute phase protein.

Objective: The sick euthyroid syndrome in critically ill patients without primary disease of the thyroid gland is characterised by low serum total triiodothyronine (T 3 ), normal to elevated thyroxine (T 4 ), elevated reverse T 3 (rT 3 ) and normal TSH levels. The aim of this work was to clarify if impaired T 4 and rT 3 5 0 -deiodination is an underlying mechanism. Design and Methods: We analysed the effect of the human recombinant proinflammatory cytokines interleukin (IL)-6 and IL-1b, tumour necrosis factor-a (TNF-a) and interferon-g (IFN-g) on human type I 5 0 -iodothyronine deiodinase (5 0 DI) enzyme activity in the human hepatocarcinoma cell line HepG2, i.e. in a homologous human system. Furthermore, we analysed transcriptional effects of the cytokines by transient transfection assays using the luciferase or chloramphenicol acetyltransferase (CAT) reporter genes under the control of 1480 nucleotides of the human 5 0 DI promoter. Results: IL-6 at 500 pg/ml and TNF-a at 25 ng/ml had no significant effect, whereas 100 ng/ml IFN-g or 10 ng/ml IL-1b reduced 5 0 DI enzyme activity to 77.9 and 59.5% of control values. IFN-g did not alter, IL-6 and TNF-a moderately decreased (in the case of IL-6 only in the CAT system), and IL-1b (0.01 -10 ng/ml) dose-dependently inhibited 5 0 DI promoter activity to a minimum of 38.1%. Conclusion: IL-1b inhibited both 5 0 DI enzyme and promoter activity and, thus, may exert its effect on thyroid hormone metabolism at least partially through direct inhibition of hepatic 5 0 DI gene transcription.

There is experimental and epidemiological evidence for an association between low selenium levels and gastrointestinal cancer incidence, prevalence, and mortality. To identify targets for selenium supplementation in the human digestive tract, we examined mRNA expression of various selenocysteine-containing proteins in normal mucosa biopsy specimens. Tissue samples from the esophagus and from different sites of the stomach, small bowel, and colon were obtained during endoscopies of the upper and lower gastrointestinal tract. Northern blot analyses revealed a lack of cytosolic glutathione peroxidase mRNA but a differential mRNA expression pattern of gastrointestinal and plasma glutathione peroxidase, selenoprotein P, and thioredoxin reductase. Glutathione peroxidase and thioredoxin reductase activities were detected in the mucosa of all biopsies, but the differential pattern did not reflect the differential mRNA steady-state levels. In addition to gastrointestinal glutathione peroxidase, which was found to play a role in colon cancer resistance, we identified further gastrointestinal selenoproteins, which may be involved in gastrointestinal cell defense and cell differentiation.