Distribution of [14C]-trans-resveratrol,a cancer chemopreventive polyphenol,in mouse tissues after oral administration

Trans-resveratrol, a phenolic compound present in wine, has been reported to be a potential cancer chemopreventive agent. However, although it has numerous biological activities in vitro, there are few data about its bioavailability and tissue distribution in vivo. The objectives of this study were to investigate the absorption and tissue distribution of 14C-trans-resveratrol following oral administration to mice. Male Balb/c mice were given a single oral dose of 14C-trans-resveratrol and were sacrificed at 1.5, 3 or 6 h postdose. The distribution of radioactivity in tissues was evaluated using whole-body autoradiography, quantitative organ-level determination and microautoradiography. In addition, identification of radioactive compounds in kidney and liver was done with high-performance liquid chromatography. Autoradiographic survey of mice sections as well as radioactivity quantification in various organs revealed a preferential fixation of 14C-trans-resveratrol in the organs and biological liquids of absorption and elimination (stomach, liver, kidney, intestine, bile, urine). Moreover, we show that 14C-trans-resveratrol derived radioactivity is able to penetrate the tissues of liver and kidney, a finding supported by microautoradiography. The presence of intact 14C-trans-resveratrol together with glucurono- and/or sulfoconjugates in these tissues was also shown. This study demonstrates that trans-resveratrol is bioavailable following oral administration and remains mostly in intact form. The results also suggest a wide range of target organs for cancer chemoprevention by wine polyphenols in humans.     

Structured HCV nucleocapsids composed of P21 core protein assemble primary in the nucleus of Pichia pastoris yeast

The relationship between HCV core protein (HCcAg) processing and the structural composition and morphogenesis of nucleocapsid-like particles (NLPs) produced in Pichia pastoris cells was studied. At early stages of heterologous expression, data suggest that HCcAg (in the P21 form) was transported soon after its synthesis in the cytoplasm into the nucleus. HCcAg assembly into nucleocapsid-like particles with 20-30 nm in diameter took place primary in the cell nucleus. However, at later stages, when P21 and P23 forms were co-detected, data suggest that new assembly of nucleocapsid particles containing P21 possibly occurs at ER membranes and in the cytoplasm. This is the first report showing that structured HCV NLPs composed of P21 core protein assemble primary in the nucleus of P. pastoris yeast.     

DNA-protein cross-linking via guanine oxidation: dependence upon protein and photosensitizer

DNA-protein cross-links form when guanine undergoes a 1-electron oxidation in a flash-quench experiment, and the importance of reactive oxygen species, protein, and photosensitizer is examined here. In these experiments, a strong oxidant produced by oxidative quenching of a DNA-bound photosensitizer generates an oxidized guanine base that reacts with protein to form the covalent adduct. These cross-links are cleaved by hot piperidine and are not the result of reactive oxygen species, since neither a hydroxyl radical scavenger (mannitol) nor oxygen affects the yield of DNA-histone cross-linking, as determined via a chloroform extraction assay. The cross-linking yield depends on protein, decreasing as histone > cytochrome c > bovine serum albumin. The yield does not depend on the cytochrome oxidation state, suggesting that reduction of the guanine radical by ferrocytochrome c does not compete effectively with cross-linking. The photosensitizer strongly influences the cross-linking yield, which decreases in the order Ru(phen)(2)dppz(2+) [phen = 1,10-phenanthroline; dppz = dipyridophenazine] > Ru(bpy)(3)(2+) [bpy = 2,2'-bipyridine] > acridine orange > ethidium, in accordance with measured oxidation potentials. A long-lived transient absorption signal for ethidium dication in poly(dG-dC) confirms that guanine oxidation is inefficient for this photosensitizer. From a polyacrylamide sequencing gel of a (32)P-labeled 40-mer, all of these photosensitizers are shown to damage guanines preferentially at the 5' G of 5'-GG-3' steps, consistent with a 1-electron oxidation. Additional examination of ethidium shows that it can generate cross-links between histone and plasmid DNA (pUC19) and that the yield depends on the quencher. Altogether, these results illustrate the versatility of the flash-quench technique as a way to generate physiologically relevant DNA-protein adducts via the oxidation of guanine and expand the scope of such cross-linking reactions to include proteins that may associate only transiently with DNA.     

Estrogen receptor,cyclic adenosine monophosphate,and protein kinase A are involved in the nongenomic pathway by which estradiol accelerates oviductal oocyte transport in cyclic rats

This investigation examined the role of estrogen receptor (ER) on the stimulatory effect of estradiol (E2) on protein phosphorylation in the oviduct as well as on E2-induced acceleration of oviductal oocyte transport in cyclic rats. Estrous rats were injected with E2 s.c. and with the ER antagonist ICI 182 780 intrabursally (i.b.), and 6 h later, oviducts were excised and protein phosphorylation was determined by Western blot analysis. ICI 182 780 inhibited the E2-induced phosphorylation of some oviductal proteins. Other estrous rats were treated with E2 s.c. and ICI 182 780 i.b. The number of eggs in the oviduct, assessed 24 h later, showed that ICI 182 780 blocked the E2-induced egg transport acceleration. The possible involvement of adenylyl cyclase, protein kinase A (PK-A), protein kinase C (PK-C), or tyrosine kinases on egg transport acceleration induced by E2 was then examined. Selective inhibitors of adenylyl cyclase or PK-A inhibited the E2-induced egg transport acceleration, whereas PK-C or tyrosine kinase inhibitors had no effect. Furthermore, forskolin, an adenylyl cyclase activator, mimicked the effect of E2 on ovum transport and E2 increased the level of cAMP in the oviduct of cycling rats. Finally, we measured PK-A activity in vitro in the presence of E2 or E2-ER complex. Activity of PK-A in the presence of E2 or E2-ER was similar to PK-A alone, showing that E2 or E2-ER did not directly activate PK-A. We conclude that the nongenomic pathway by which E2 accelerates oviductal egg transport in the rat requires absolute participation of ER and cAMP and partial participation of PK-A signaling pathways in the oviduct.     

Bioavailability studies of a new iron source by means of the prophylactic-preventive method in rats

The bioavailability of iron from a new commercial source containing ferric gluconate stabilized with glycine sold under the trade name Bioferrico™ was studied in this work by means of the prophylactic-preventive test in rats. NaFeEDTA was also studied by the same methodology for comparative purposes and ferrous sulfate was used as the reference standard. The test was conducted for 4 wk with male weaned rats, which were randomized into four groups of at least eight animals each. A control group received a basal diet of low-iron content, whereas the other groups received the same diet with iron added at a dose of 20 mg/kg as FeSO₄·7H₂O, NaFeEDTA, and Bioferrico, respectively. Individual hemoglobin concentrations (HbC) and weights were determined at the beginning and at the end of the study and food intake was daily registered. The iron bioavailability (BioFe) of each source was calculated as the ratio between the amount of iron incorporated into hemoglobin during the treatment (HbFe) and the total iron intake per animal (ToFeIn). A relative biological value (RBV) was obtained for each iron source under study as the ratio between the BioFe of the tested compound and that of the reference standard. The RBVs were 98% and 86% for Bioferrico and NaFeEDTA, respectively. Bioferrico showed a high bioavailability and behaved inertly in relation to the sensorial properties of the fortified food when it was added to flour. These qualities emphasize Bioferrico as a promising source for iron fortification.     

Plasmodium falciparum: in vitro activity of sulfadoxine and dapsone in field isolates from Kenya: point mutations in dihydropteroate synthase may not be the only determinants in sulfa resistance

We have determined the relationship between point mutations in the gene that encodes the sulfa target, dihydropteroate synthase (DHPS) and the chemosensitivity profile to sulfadoxine and dapsone in 67 isolates from Kilifi, Kenya. We assessed the presence of mutations at codons 436, 437, 540, 581, and 613 of dhps. The results showed that the dhps genotype had a strong influence on the sensitivity to sulfadoxine and dapsone, but that the correlation was far from perfect. Eleven isolates carried a wild-type dhps allele, but were resistant to sulfadoxine (IC(50) values >10 microg/ml), and 4/28 isolates were classed as sensitive to sulfadoxine (IC(50) values <10 microg/ml), but carried a triple mutant (436/437/613) allele of dhps. These data show that in low folate medium in vitro, the dhps genotype alone did not account completely for sulfadoxine or dapsone resistance; other factors such as the utilisation of exogenous folate must also be considered.     

Temperature effects and genotype-temperature interactions on sex determination in the European sea bass (Dicentrarchus labrax L.)

The effect of temperature on sex-ratios in 27 families of sea bass reared in the same tank from the fertilization stage onward was investigated. An excess of males (68%) was found in the groups that were reared at high temperature (mean +/- standard deviation: 20+/-1 degrees C) until they reached the mean size of 8.1 cm (Standard Length, 149 days post-fertilization [p.f.]). Masculinization was higher (89% of males) in the groups maintained at low temperature (13 degrees C), from fertilization to a mean length of 6.5 cm (346 days p.f.). Shifts from high to low temperature at 8.1cm and from low to high temperature at 6.5 cm had no consequence on the sex-ratio. The percentage of males showing intratesticular oocytes was higher at low temperature (63%) than at high temperature (36%), suggesting that these males may be sensitive fish that have been masculinized by environmental factors. Fish sampled in the groups reared at high (2,200 fish) and low (500 fish) temperature were genotyped on three microsatellite loci. This allowed them to be assigned to the breeders used in the crossing design, thus permitting an analysis of parental influence on sex-ratios. In groups reared at high temperature, both parents had a significant additive effect on the percentage of females, and the interaction between sire and dam was not significant. Genotype temperature interactions were also detected and their existence suggests the interesting possibility of selecting nonsensitive genotypes in breeding programs.     

Hereditary spastic paraplegia SPG13 is associated with a mutation in the gene encoding the mitochondrial chaperonin Hsp60

SPG13, an autosomal dominant form of pure hereditary spastic paraplegia, was recently mapped to chromosome 2q24-34 in a French family. Here we present genetic data indicating that SPG13 is associated with a mutation, in the gene encoding the human mitochondrial chaperonin Hsp60, that results in the V72I substitution. A complementation assay showed that wild-type HSP60 (also known as "HSPD1"), but not HSP60 (V72I), together with the co-chaperonin HSP10 (also known as "HSPE1"), can support growth of Escherichia coli cells in which the homologous chromosomal groESgroEL chaperonin genes have been deleted. Taken together, our data strongly indicate that the V72I variation is the first disease-causing mutation that has been identified in HSP60.