Stefan Spycher

A Novel Human Tocopherol-associated Protein CLONING, IN VITRO EXPRESSION, AND CHARACTERIZATION

Vitamin E (α-tocopherol) is an essential dietary nutrient for humans and animals. The mechanisms involved in cellular regulation as well as in the preferential cellular and tissue accumulation of α-tocopherol are not yet well established. We previously reported (Stocker, A., Zimmer, S., Spycher, S. E., and Azzi, A. (1999)  IUBMB Life 48, 49–55) the identification of a novel 46-kDa tocopherol-associated protein (TAP) in the cytosol of bovine liver. Here, we describe the identification, the molecular cloning into Escherichia coli, and the in vitroexpression of the human homologue of bovine TAP,hTAP. This protein appears to belong to a family of hydrophobic ligand binding proteins, which have the CRAL (cis-retinal binding motif) sequence in common. By using a biotinylated α-tocopherol derivative and the IASys resonant mirror biosensor, the purified recombinant protein was shown to bind tocopherol at a specific binding site with K d 4.6 × 10−7 m. Northern analyses showed that hTAP mRNA has a size of approximately 2800 base pairs and is ubiquitously expressed. The highest amounts of hTAP message are found in liver, brain, and prostate. In conclusion, hTAP has sequence homology to proteins containing the CRAL_TRIO structural motif. TAP binds to α-tocopherol and biotinylated tocopherol, suggesting the existence of a hydrophobic pocket, possibly analogous to that of SEC14.

Modulation of α-tropomyosin expression by α-tocopherol in rat vascular smooth muscle cells

The effect of α‐tocopherol (vitamin E) on gene expression in rat vascular smooth muscle cells was studied by the differential display technique. One gene out of about 1000 genes analyzed, identified as α‐tropomyosin, showed an increased transcription level caused by α‐tocopherol treatment. Northern and Western blot analysis revealed a time‐dependent transient up‐regulation of the amount of mRNA (peak between 2 and 3 h) and protein (peak at 5 h) in α‐tocopherol‐treated cells. No effect was observed in cells treated with β‐tocopherol.

Identification of a Novel Cytosolic Tocopherol‐Binding Protein: Structure, Specificity, and Tissue Distribution

alpha‐Tocopherol plays an important role as a lipid‐soluble antioxidant. It is present in all major mammalian cell types and shows tissue‐specific distribution. This suggests the presence of specific proteins involved in intracellular distribution or metabolism of alpha‐tocopherol. A diminution of tocopherol plasma concentrations contributes to the development of diseases such as vitamin E deficiency (AVED), atherosclerosis, and prostate cancer. Further evidence has been obtained for the existence of sites in cellular metabolism and signal transduction where alpha‐tocopherol potentially plays a regulatory role. A signal transduction modulation specific for alpha‐tocopherol has been described in several model systems. Using radioactively labeled alpha‐tocopherol as tracer, we have isolated a new alpha‐tocopherol‐associated protein (TAP) from bovine liver. This protein has a molecular mass of 46 kDa and an isoelectric point of 8.1. From its partial amino acid sequence, a human gene has been identified with high homology to the newly described protein. Sequence analysis has established that the new TAP has structural motifs suggesting its belonging to a family of hydrophobic ligand‐binding proteins (RALBP, CRALBP,alpha‐TTP, SEC 14, PTN 9, RSEC 45). Human TAP has been cloned into Escherichia coli, and its tissue‐specific expression has been assessed by Northern blot analysis.

The validity of the MIMIC health index — some empirical evidence

The rise of medical care expenditure over the last few decades has made the economic evaluation of medical services increasingly important. However, economic appraisal of specific medical therapies is a complex issue, particularly with respect to the benefit side of the analysis. Previous studies often have ignored treatment benefits, restricting analysis to cost differentials between treatment alternatives or have focused exclusively on improvements in patients’ earnings capacity through reductions of morbidity or mortality. Both approaches are clearly unsatisfactory if (i) a new therapy constitutes significant therapeutic progress without reducing treatment costs, (ii) the major effect of a new therapy is to improve patients’ well-being, enjoyment of life, or life quality rather than to increase longevity or earnings capacity and (iii) the benefits of a new therapy accrue mainly to patients who are not part of the labor force, i.e. persons working in the household or retired people. In all these situations, economic evaluation requires assessment of the direct and immediate treatment benefits: the improvement in patients’ well-being (life quality).

Modulation by Oxidants and Antioxidants of Signal Transduction and Smooth Muscle Cell Proliferation

Arteriosclerosis, restenosis, and hypertension as well as a number of other vascular diseases are characterized by vascular smooth muscle cell proliferation.1-4 Smooth muscle cell growth is regulated by specific factors released from blood cells,1-5 by the vessel wall cells3,6 and by natural compounds, such as tocopherols. Active oxygen species (i.e., O2, H2O2, and OH) may act as vascular smooth muscle growth factors by inducing proto-oncogene expression and DNA synthesis,7,8 and antioxidants can retard atherosclerosis and foam cell formation as shown in animal model,9,10 and human studies.11-14 Tocopherols, well-established natural antioxidants, may in some cases stimulate cell proliferation by preventing the formation of growth-inhibitory lipid peroxides.15-19 In addition to having antioxidant properties, RRR-α-tocopherol is a cell growth inhibitor, this effect being unrelated with its radical scavenger properties.20,21 Parallel to the inhibition of cell proliferation, RRR-α-tocopherol has been shown to inhibit protein kinase C activity in smooth muscle cells.20,21 The mechanism of this inhibition appears to be related with a diminished phosphorylation of protein kinase C-α.