We present a characterization of the single-copy gene, mIGFBP-2, encoding the murine insulin-like growth factor-binding protein-2 (mIGFBP-2). It consists of four exons with sizes of 470 +/- 2, 227, ...141 and > 475 nucleotides (nt). The first intron spans 23 kb of genomic sequence, and the complete gene extends to more than 28 kb. Two kb of the 5'-flanking region were sequenced. This region has no TATA or CAAT boxes but is G+G-rich and contains several potential regulatory sequence motifs. A total of five GC boxes, which may serve as potential binding sites for a transcription factor, Sp1, are present immediately upstream of the transcription start point (tsp). By primer extension, we identified a single tsp at nt position -85 +/- 2. The murine IGFBP-2 locus was mapped to the proximal region of mouse chromosome 1, to a region of conserved synteny with human chromosome 2q. A comparison of the deduced amino acid sequences of mouse, rat and human IGFBP-2 reveals a high degree of homology between all three species.
It is well documented that 1 alpha,25-dihydroxyvitamin D3 (1 alpha,25OH2D3), the most active vitamin D metabolite, inhibits epidermal keratinocyte proliferation and promotes differentiation. 1 ...alpha,25(OH)2D3 can be produced in keratinocytes from 25-hydroxyvitamin D3 by the enzyme 25-hydroxyvitamin D3-1 alpha-hydroxylase (1-OHase). Hydroxylation of 1 alpha,25(OH)2D3 by 25-hydroxyvitamin D3-24-hydroxylase (24-OHase), the first step in the catabolic pathway of 1 alpha,25(OH)2D3 could significantly reduce the intracellular concentration of 1 alpha,25(OH)2D3. Therefore, the expression of 24-OHase could have a critical regulatory role in 1 alpha,25(OH)2D3-dependent gene expression. As a first step to examine this possibility, the steady state level of 24-OHase mRNA in cultured human keratinocytes (CHK) was investigated. 24-OHase mRNA was not detected in control CHK. 1 alpha,25(OH)2D3 caused a dose- and time-dependent increase in 24-OHase mRNA level. The highest accumulation of 24-OHase mRNA was observed in CHK treated with 0.1-1 microM 1 alpha,25(OH)2D3. The level of 24-OHase mRNA reached a plateau 12-24 hr after 1 alpha,25(OH)2D3 treatment. 1 beta,25-dihydroxyvitamin D3, the stereoisomer of 1 alpha,25(OH)2D3, failed to induce 24-OHase mRNA expression significantly. In addition to 24-OHase mRNA, a 1.0-kb mRNA hybridized strongly with both rat and human 24-OHase cDNA probes. The origin of this 1.0-kb message is unknown at present, however, it was regulated by 1 alpha,25(OH)2D3. These results demonstrate that 1 alpha,25(OH)2D3 up-regulates the expression of 24-OHase mRNA, and this may be an important first step in the initiation of catabolism of 1 alpha,25(OH)2D3 in human keratinocytes.
In contrast to their excellent mechanical properties, titanium alloys possess poor wear characteristics. Diamond coatings appearto be a promising solution for the wear problem. Using standard ...deposition parameters for silicon (diamond scratched surface; microwave chemical vapour deposition, 500 W, 800 °C for 4 h, 50 mbar with an atmosphere of 1% CH4 + 99%H2) good coatings on pure titanium were obtained. The mechanical properties of the titanium are strongly influenced by the processing temperature and the gas atmosphere. As a result of the hydrogen adsorption, grain coarsening and formation of titanium hydride occur and subsequently the low cycle fatigue strength decreases by an order of magnitude. However, by annealing in vacuum (800°C for 2 h, furnace cooled) the hydrogen can be removed again and the initial mechanical properties can almost completely be restored. Based on scanning electron microscopy micrographs, Auger analysis, X-ray diffraction and microhardness measurements a basic understanding of the microstructural changes was developed. The surface structure may be visualized as consisting of five zones, created during the deposition process (diamond layer/nucleation zone/reaction layer/gradient layer/affected substrate).
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