PYCNOGENOL chewing gum minimizes gingival bleeding and plaque formation.

PYCNOGENOL is an antioxidant phytochemical shown to have antiinflammatory activity in both the in vitro and in vivo models. This study compared the effects of chewing gums with and without PYCNOGENOL on gingival bleeding and plaque formation in 40 human subjects. In this double-blind study, subjects were assigned randomly to receive either control gums without PYCNOGENOL or experimental gums containng 5 mg PYCNOGENOL. Subjects used chewing gums for 14 days. Gingival bleeding and plaque scores were taken before and after the experiment. PYCNOGENOL chewing gums significantly reduced gingival bleeding, while no changes were noted in bleeding indexes in control subjects who used regular chewing gums. Subjects using regular control gums had significant increases of dental plaque accumulation during the two-week period. No increases in plaque accumulation were noted in subjects using PYCNOGENOL chewing gums. The data of this study suggest that the use of Pycnogenol chewing gums can minimize gingival bleeding and plaque accumulation.     

Simple, reliable and fast spectrofluorometric method for determination of plasma Verteporfin (Visudyne) levels during photodynamic therapy for choroidal neovascularization.

Photodynamic therapy with Verteporfin, a potent photosensitizer dye, is a very effective treatment for age related macular degeneration due to choroidal neovascularization. Photodynamic therapy offers the potential for selective tissue injury in part attributable to preferential localization of Verteporfin, administrated by intravenous infusion, to the choroidal neovascularization complex and irradiation of the complex with non-laser thermal light at 690 nm resulting in at least temporary thrombosis and vessel closure. Verteporfin is a benzoporphyrin derivative monoacid ring A formulated as a unilamellar liposome. In the blood Verteporfin is associated with lipoprotein fractions and is rapidly cleared via a receptor-mediated uptake mechanism due the high expression of LDL receptors in neovascular tissues. Verteporfin was undetectable in plasma 24 hr after infusion of the recommended dose: 6 mg/m2 of body surface area. The main side effect is photosensitivity of skin which is usually short-lived (24-48 hr) with a low incidence (2.3%). As skin photosensitivity depends on circulating rather than tissue drug levels, we investigate the possibility of developing a simple, fast and reliable spectrofluorometric method to measure plasma Verteporfin levels. Fluorescence emission spectrum (550-750 nm) of 1:10 saline diluted plasma with lambda exc=430 nm showed a characteristic emission peak at 692 nm, the height being proportional to the Verteporfin levels. The sensitivity is around 100 ng/ml and the pharmacokinetics of Verteporfin has been studied from 0 to 5 hr after infusion in six patients older than 65 years with age-related macular degeneration.     

Improving solubility of catalytic domain of human beta-1,4-galactosyltransferase 1 through rationally designed amino acid replacements.

beta-1,4-galactosyltransferase 1 (beta4gal-T1, EC 2.4.1.38) transfers galactose from UDP-galactose to free N-acetyl-D-glucosamine or bound N-acetyl-D-glucosamine-R. Soluble beta4gal-T1, purified from human milk has been refractory to structural studies by X-ray or NMR. In a previous study (Malissard et al. 1996, Eur. J. Biochem. 239, 340-348) we produced in the yeast Saccaromyces cerevisiae an N-deglycosylated form of soluble beta4gal-T1 that was much more homogeneous than the human enzyme, as it displayed only two isoforms when analysed by IEF as compared to 13 isoforms for the native beta4gal-T1. The propensity of recombinant beta4gal-T1 to aggregate at concentrations > 1 mg.mL(-1) prevented structural and biophysical studies. In an attempt to produce a beta4gal-T1 form suitable for structural studies, we combined site-directed mutagenesis and heterologous expression in Escherichia coli. We produced a mutated form of the catalytic domain of beta4gal-T1 (sfbeta4gal-T1mut) in which seven mutations were introduced at nonconserved sites (A155E, N160K, M163T, A168T, T242N, N255D and A259T). Sfbeta4gal-T1mut was shown to be much more soluble than beta4gal-T1 expressed in S. cerevisiae (8.5 mg.mL(-1) vs. 1 mg.mL(-1)). Catalytic activity and kinetic parameters of sfbeta4gal-T1mut produced in E. coli were shown not to differ to any significant extent from those of the native enzyme.     

Effect of adrenalin, insulin and contractions on the content of the free fatty acid fraction in skeletal muscle.

The fraction of free fatty acids (FFA) is present in skeletal muscles. However, there is almost no data regarding regulation in the content of this intramuscular lipid pool. We took advantage of the isolated muscle preparation to examine whether: a) increasing exogenous concentration of FFA (500microM or 700microM, 30min) b) insulin (10.00 I.U./L, 30min), c) adrenalin (4.4 nM, 30 min), or d) contractions (200ms, tetani, 1Hz, 30min), affect the FFA content inside myocytes. Incubation of soleus (S) and extensor digitorum longus (EDL) with increasing concentrations of exogenous FFA (from 500microM to 700microM) resulted in an increase in the total FFA fraction in both muscles studied (by 280.2% and 259.1%, respectively). In contracting muscles FFA pool was significantly reduced both in S (by 73.1%) and in EDL (by 31.1%). Neither stimulation by adrenalin nor insulin affected the total content of FFA fraction in the muscles examined. We conclude that a) increased availability of exogenous FFA at the sarcolemma level results in an increase in the size of intramuscular FFA fraction b) the intracellular FFA fraction is utilized by contracting muscles with regard to the fiber composition and to a greater extent in more oxidative muscles, c) FFA fraction remains stable upon stimulation by insulin or adrenalin.     

Induction and Maintenance of Maternal Behaviour in the Domestic Hen: Influence of Testosterone and Oestradiol Treatments

Confining an adult hen with two newly-hatched chicks induced a gradual emergence of maternal behaviour in the hen. The aim of the experiments presented here was to analyse the effects of testosterone or oestradiol treatments on the induction and maintenance of maternal behaviour in hens with no previous maternal experience. The ability of a hen to positively respond towards chicks was not altered by either testosterone or oestradiol injections. However, testosterone therapy prevented both the appearance and the maintenance of the most typical call, clucking. Testosterone injected hens responded to chicks with a contact-type call, but oestradiol treated ones did not. These results indicate that some aspects of maternal behaviour can be modulated by peripheral levels of hormones.     

Active compounds in Populus nigra L. wilted leaves responsible for attracting Helicoverpa armigera (Hübner) (Lep., Noctuidae) and new agaropectin formulation

Abstract:  The leaf extracts of Populus nigra were collected and identified by steam distillation, air entrainment and gas chromatographic–mass spectrometric analysis. Electroantennograms were recorded from Helicoverpa armigera adults in response to the chemicals identified. Both aromatic compounds and green-leaf volatiles elicited strong responses. Field experiments revealed that the active compounds responsible for attracting H. armigera moths are mainly short-side-chain aromatic alcohols and aldehydes. We, for the first time, used agaropectin as the controlled-release matrix of insect attractants. A five-component lure containing all the aromatics without phenolics, mixed in the proportions as found in the steam distillate of the leaves collected in August, produced the best trap catch. The results showed that the volatiles of wilted leaves of P. nigra can attract H. armigera adults by feeding attraction.     

Silent mutations in secondary Shine-Dalgarno sequences in the cDNA of human serum amyloid A4 promotes expression of recombinant protein in Escherichia coli.

The serum amyloid A (SAA) superfamily comprises a number of differentially expressed genes with a high degree of homology in mammalian species. SAA4, an apolipoprotein constitutively expressed only in humans and mice, is associated almost entirely with lipoproteins of the high-density range. The presence of SAA4 mRNA and protein in macrophage-derived foam cells of coronary and carotid arteries suggested a specific role of human SAA4 during inflammation including atherosclerosis. Here we underline the importance of ribosome binding site (rbs)-like sequences (also known as Shine-Dalgarno sequences) in the SAA4 cDNA for expression of recombinant SAA4 protein in Escherichia coli. In contrast to rbs sequences coded by the expression vectors, rbs-like sequences in the cDNA of target protein(s) are known to interfere with protein translation via binding to the small 16S ribosome subunit, yielding low or even no expression. Here we show that PCR mutations of two rbs-like sequences in the human SAA4 cDNA promote expression of considerable amounts of recombinant SAA4 in E.coli.