Template-dependent biosynthesis of poly(G) x poly (C) and its antiviral activity in vitro and in vivo.

Experimental conditions for poly(G) synthesis from GTP on a poly(C) template with the aid of Escherichia coli DNA-dependent RNA polymerase were investigated. The reaction product was purified without the use of RNase. On the basis of spectral data, gel permeation chromatography, affinity adsorption and electron microscopic visualization, the poly(G) x poly(C) product was assumed to possess a high degree of structural regularity. Its in vitro and in vivo antiviral activities were compared with those of traditional poly(G) x poly(C) and poly(I) x poly(C). Template-dependent poly(G) x poly(C) was similar in its in vitro activity to poly(I) x poly(C) or even surpassed it, whereas the 'traditional' poly(G) x poly(C) was only slightly active in vitro. However, 'traditional' poly(G) x poly(C) and poly(I) x poly(C) had similar activity in vivo, whereas template-dependent poly(G) x poly(C) was much less active in vivo. The role of intramolecular structural regularity in the in vitro and in vivo antiviral activity of polyribonucleotide duplexes is discussed.

Evidence for human placental synthesis of 24,25-dihydroxyvitamin D3 and 23,25-dihydroxyvitamin D3.

The two principal dihydroxylated metabolites of the vitamin D prohormone 25-hydroxyvitamin D3 [25(OH)D3] are 1 alpha,25-dihydroxyvitamin D3 [1,25(OH)2D3, the active hormone] and 24R,25-dihydroxyvitamin D3 [24,25(OH)2D3, a putative regulator of developmental bone formation]. Although several studies have demonstrated placental synthesis of 1,25(OH)2D3 from 25(OH)D3, placental production of 24,25(OH)2D3 has not been thoroughly investigated. Therefore, we studied 25(OH)D3 metabolism in term human placenta using a villous explant model and cultures of isolated trophoblast and villous mesenchymal cells. We determined that both vitamin D-replete and vitamin D-deficient trophoblast metabolize 25(OH)D3 predominantly via 24-hydroxylation. Placental 24,25(OH)2D3 was identified by cochromatography with authentic standard on four different HPLC systems, scanning UV spectrophotometry profile of the metabolite, sensitivity to periodate cleavage, and mass spectrometry of the putative placental 24,25(OH)2D3 and its periodate cleavage product. We also identified for the first time placental synthesis of 23,25(OH)2D3 using cochromatography with authentic standard on two different HPLC systems, scanning UV spectrophotometry, resistance to periodate cleavage, and mass spectrometry. When trophoblast was incubated for up to 4 h with physiologic concentrations of [3H]25(OH)D3 (6 nM) significant amounts of [3H]24,25(OH)2D3 were produced, but [3H]1,25(OH)2D3 could not be consistently detected. In contrast, when we incubated trophoblast with supraphysiologic concentrations of 25(OH)D3 (6-10 microM), both 24,25(OH)2D3 and 1,25(OH)2D3 were synthesized. These results provide unequivocal evidence for placental synthesis of both 24,25(OH)2D3 and 23,25(OH)2D3.(ABSTRACT TRUNCATED AT 250 WORDS)

Biophysical studies of the complexes of polyriboguanylic acid with brominated and chlorinated polyribocytidylic acids.

Conformation of double-stranded complexes of polyriboguanylic acid with halogenated polyribocytidylic acid [poly(C)] was studied with the aid of differential pulse polarography, terbium fluorescence and circular dichroism spectrometry. It was shown that halogenation at C(5) of cytosine residues in poly(C) disturbed the ordered structure of the double-helical complex. In addition, this halogenation does not improve antiviral activity of the polynucleotide complex studied in the system of vesicular stomatitis virus and the cell culture of chicken embryos. It was concluded that the regularity of the secondary structure of synthetic RNAs might play an important role in the mechanism of biological activity of these biomacromolecules.