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Background: Healthy eating habits at a young age are crucial to support growth and development and good general health. In this context, monitoring youth dietary intakes adequately with valid tools is important to develop efficient interventions and identify groups that are more at risk of inadequate intakes. This study aimed to assess the relative validity of the self-administered web-based 24-h dietary recall (R24W) for evaluating energy and nutrient intakes among active adolescents., Methods: Participants were invited to complete one interviewer-administered 24-h dietary dietary recall and the R24W on up to three occasions within one month. A total of 272 French-speaking active adolescents aged 12 to 17 years from the province of Québec were invited to complete three R24W and one interview-administered 24-h recall. Student's t-test and correlations were conducted on sex-adjusted data. Percent differences, cross-classification (percentage of agreement), weighted Kappa and Bland-Altman plots were calculated., Results: Mean (SD) energy intake from the R24W was 8.8% higher than from the interview-administered 24-h dietary recall (2558 kcal ± 1128 vs. 2444 kcal ± 998, p < 0.05). Significant differences in mean nutrient intake between the R24W and the interview-administered 24-h dietary recall ranged from 6.5% for % E from fat (p < 0.05) to 25.2% for saturated fat (p < 0.001), i.e., higher values with R24W. Sex-adjusted correlations were significant for all nutrients except for % E from proteins and thiamin (range: 0.24 to 0.52, p < 0.01). Cross-classification demonstrated that 36.6% of the participants were classified in the same fourth with both methods, 39.6% in the adjacent fourth, and 5.7% misclassified. Bland-Atman plots revealed proportional bias between the two methods for 7/25 nutrients. Completing at least two recalls with the R24W increased the precision of intake estimates., Conclusion: These data suggest that the R24W presents an acceptable relative validity compared to a standard interview-administered 24-h recall for estimating energy and most nutrients in a cohort of French-speaking adolescents from the province of Québec., (© 2024. Crown.)

Impediments in replication fork progression cause genomic instability, mutagenesis, and severe pathologies. At stalled forks, RPA-coated single-stranded DNA (ssDNA) activates the ATR kinase and directs fork remodeling, 2 key early events of the replication stress response. RFWD3, a recently described Fanconi anemia (FA) ubiquitin ligase, associates with RPA and promotes its ubiquitylation, facilitating late steps of homologous recombination (HR). Intriguingly, RFWD3 also regulates fork progression, restart and stability via poorly understood mechanisms. Here, we used proteomics to identify putative RFWD3 substrates during replication stress in human cells. We show that RFWD3 interacts with and ubiquitylates the SMARCAL1 DNA translocase directly in vitro and following DNA damage in vivo. SMARCAL1 ubiquitylation does not trigger its subsequent proteasomal degradation but instead disengages it from RPA thereby regulating its function at replication forks. Proper regulation of SMARCAL1 by RFWD3 at stalled forks protects them from excessive MUS81-mediated cleavage in response to UV irradiation, thereby limiting DNA replication stress. Collectively, our results identify RFWD3-mediated SMARCAL1 ubiquitylation as a novel mechanism that modulates fork remodeling to avoid genome instability triggered by aberrant fork processing., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Yates et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Growing numbers of researchers have investigated how training programmes influence coaches' and teachers' ability to promote life skills development, and concurrently, athlete-related outcomes. This study aimed to examine high school student-athletes' development of life skills through a three-year programme called Winner for Life (Gagnant pour la vie). Delivered online to high school coaches and teachers, the programme targeted five life skills: (a) Goal Setting and Concentration (Year 1), (b) Healthy Eating Habits and Safety Behaviours (Year 2), and Physical and Mental Recovery (Year 3). In all, 148 student-athletes participated in the programme and completed questionnaires at five time points to assess life skills development. A general linear repeated measures model was used to assess changes over time. Missing data were handled using multiple imputations. Student-athletes reported higher scores on Goal Setting subscale at time 2 (vs. time 1) and time 3 (vs. time 2) and on Social Recovery subscale at time 2 (vs. time 1). They also reported lower scores on Sleep Quality subscale at time 3 (vs. time 2) and time 5 (vs. time 4). Additionally, a significant interaction was observed between student-athletes' age (mean = 13.5 years) at study start, and improvements on certain life skills subscales over time. Overall, the fact that student-athletes did not improve on all life skills could be explained by the COVID-19 pandemic lockdowns, which limited the programme's influence from time 3. Results should be considered in light of limitations regarding gender distribution and ceiling effects on student-athletes' scores. Recommendations to improve future life skills programmes are discussed, such as involving parents in delivery and encouraging life skills teaching over time., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2023 Elsevier Ltd. All rights reserved.)

Online learning environments are challenging for deaf and hard of hearing (DHH) individuals. A major concern is split attention, which occurs when one simultaneously attends to multiple stimuli, a situation that characterizes most multimedia presentations and instruction that combines sound, text, images, graphs or charts, and video. Needing to take notes adds another stimulus. DHH learners face this issue when auditory content is accommodated visually, in combination with visually presented content. No one can view multiple visual signals simultaneously. Learners must choose to view one signal and miss others; view signals sequentially, which requires additional time and hinders class participation; or switch between visual signals, losing elements of each. This process increases auditory and visual fatigue and cognitive load, ultimately compromising learning. Several ways are suggested to achieve more equitable access to instructional content for learners challenged by the problem of split attention.

Ancient DNA is transforming our ability to reconstruct historical patterns and mechanisms shaping modern diversity and distributions. In particular, molecular data from extinct Holocene island faunas have revealed surprising biogeographic scenarios. Here, we recovered partial mitochondrial (mt) genomes for 1300-1400 year old specimens (n = 2) of the extinct "horned" crocodile, Voay robustus, collected from Holocene deposits in southwestern Madagascar. Phylogenetic analyses of partial mt genomes and tip-dated timetrees based on molecular, fossil, and stratigraphic data favor a sister group relationship between Voay and Crocodylus (true crocodiles). These well supported trees conflict with recent morphological systematic work that has consistently placed Voay within Osteolaeminae (dwarf crocodiles and kin) and provide evidence for likely homoplasy in crocodylian cranial anatomy and snout shape. The close relationship between Voay and Crocodylus lends additional context for understanding the biogeographic origins of these genera and refines competing hypotheses for the recent extinction of Voay from Madagascar.

Recently, there have been increasing indications that the endocannabinoid (eCB) system is involved in vision. Multiple research teams studied the cannabinoid receptor type 2 (CB2R) expression and function in the mouse retina. Here, we examined the consequence of CB2R modulation on visual acuity using genetic and pharmacologic tools. We found that Cnr2 knockout mice show an enhanced visual acuity, CB2R activation decreased visual acuity while CB2R blockade with the inverse agonist AM630 increased it. The inhibition of 2-arachidonylglycerol (2-AG) synthesis and degradation also greatly increased and decreased visual acuity, respectively. No differences were seen when the cannabinoid receptor type 1 (CB1R) was deleted, blocked or activated implying that CB2R exclusively mediates cannabinoid modulation of the visual acuity. We also investigated the role of cannabinoids in retinal function using electroretinography (ERG). We found that modulating 2-AG levels affected many ERG components, such as the a-wave and oscillatory potentials (OPs), suggesting an impact on cones and amacrine cells. Taken together, these results reveal that CB2R modulates visual acuity and that eCBs such as 2-AG can modulate both visual acuity and retinal sensitivity. Finally, these findings establish that CB2R is present in visual areas and regulates vision-related functions.

Previous quantitative assessments of the crocodylians' dermal bone ornamentation (this ornamentation consists of pits and ridges) has shown that bone sculpture results in a gain in area that differs between anatomical regions: it tends to be higher on the skull table than on the snout. Therefore, a comparative phylogenetic analysis within 17 adult crocodylian specimens representative of the morphological diversity of the 24 extant species has been performed, in order to test if the gain in area due to ornamentation depends on the skull morphology, i.e. shape and size. Quantitative assessment of skull size and shape through geometric morphometrics, and of skull ornamentation through surface analyses, produced a dataset that was analyzed using phylogenetic least-squares regression. The analyses reveal that none of the variables that quantify ornamentation, be they on the snout or the skull table, is correlated with the size of the specimens. Conversely, there is more disparity in the relationships between skull conformations (longirostrine vs. brevirostrine) and ornamentation. Indeed, both parameters GApit (i.e. pit depth and shape) and OArelat (i.e. relative area of the pit set) are negatively correlated with snout elongation, whereas none of the values quantifying ornamentation on the skull table is correlated with skull conformation. It can be concluded that bone sculpture on the snout is influenced by different developmental constrains than on the skull table and is sensible to differences in the local growth 'context' (allometric processes) prevailing in distinct skull parts. Whatever the functional role of bone ornamentation on the skull, if any, it seems to be restricted to some anatomical regions at least for the longirostrine forms that tend to lose ornamentation on the snout., (© 2016 Anatomical Society.)

The development of interferon-free regimens for the treatment of chronic HCV infection constitutes a preferred option that is expected in the future to provide patients with improved efficacy, better tolerability, and reduced risk for emergence of drug-resistant virus. We have pursued non-nucleoside NS5B polymerase allosteric inhibitors as combination partners with other direct acting antivirals (DAAs) having a complementary mechanism of action. Herein, we describe the discovery of a potent follow-up compound (BI 207524, 27) to the first thumb pocket 1 NS5B inhibitor to demonstrate antiviral activity in genotype 1 HCV infected patients, BILB 1941 (1). Cell-based replicon potency was significantly improved through electronic modulation of the pKa of the carboxylic acid function of the lead molecule. Subsequent ADME-PK optimization lead to 27, a predicted low clearance compound in man. The preclinical profile of inhibitor 27 is discussed, as well as the identification of a genotoxic metabolite that led to the discontinuation of the development of this compound.

The design and preliminary SAR of a new series of 1H-quinazolin-4-one (QAZ) allosteric HCV NS5B thumb pocket 2 (TP-2) inhibitors was recently reported. To support optimization efforts, a molecular dynamics (MD) based modeling workflow was implemented, providing information on QAZ binding interactions with NS5B. This approach predicted a small but critical ligand-binding induced movement of a protein backbone region which increases the pocket size and improves access to the backbone carbonyl groups of Val 494 and Pro 495. This localized backbone shift was consistent with key SAR results and was subsequently confirmed by X-ray crystallography. The MD protocol guided the design of inhibitors, exploiting novel H-bond interactions with the two backbone carbonyl groups, leading to the first thumb pocket 2 NS5B inhibitor with picomolar antiviral potency in genotype (gt) 1a and 1b replicons (EC50 = 120 and 110 pM, respectively) and with EC50 ≤ 80 nM against gt 2-6.

Conformational restrictions of flexible torsion angles were used to guide the identification of new chemotypes of HCV NS5B inhibitors. Sites for rigidification were based on an acquired conformational understanding of compound binding requirements and the roles of substituents in the free and bound states. Chemical bioisosteres of amide bonds were explored to improve cell-based potency. Examples are shown, including the design concept that led to the discovery of the phase III clinical candidate deleobuvir (BI 207127). The structure-based strategies employed have general utility in drug design.

An assay recapitulating the 3' processing activity of HIV-1 integrase (IN) was used to screen the Boehringer Ingelheim compound collection. Hit-to-lead and lead optimization beginning with compound 1 established the importance of the C3 and C4 substituent to antiviral potency against viruses with different aa124/aa125 variants of IN. The importance of the C7 position on the serum shifted potency was established. Introduction of a quinoline substituent at the C4 position provided a balance of potency and metabolic stability. Combination of these findings ultimately led to the discovery of compound 26 (BI 224436), the first NCINI to advance into a phase Ia clinical trial.

We describe here the design, synthesis and biological evaluation of antiviral compounds acting against human rhinovirus (HRV). A series of aminothiazoles demonstrated pan-activity against the HRV genotypes screened and productive structure-activity relationships. A comprehensive investigational library was designed and performed allowing the identification of potent compounds with lower molecular weight and improved ADME profile. 31d-1, 31d-2, 31f showed good exposures in CD-1 mice. The mechanism of action was discovered to be a host target: the lipid kinase phosphatidylinositol 4-kinase III beta (PI4KIIIß). The identification of the pan-HRV active compound 31f combined with a structurally distinct literature compound T-00127-HEV1 allowed the assessment of target related tolerability of inhibiting this kinase for a short period of time in order to prevent HRV replication., (Copyright © 2013 Elsevier Ltd. All rights reserved.)

The DNA damage-binding protein 2 (DDB2) is an adapter protein that can direct a modular Cul4-DDB1-RING E3 Ligase complex to sites of ultraviolet light-induced DNA damage to ubiquitinate substrates during nucleotide excision repair. The DDB2 transcript is ultraviolet-inducible; therefore, its regulation is likely important for its function. Curiously, the DDB2 mRNA is reportedly short-lived, but the transcript does not contain any previously characterized cis-acting determinants of mRNA stability in its 3' untranslated region (3'UTR). Here, we used a tetracycline regulated d2EGFP reporter construct containing specific 3'UTR sequences from DDB2 to identify novel cis-acting elements that regulate mRNA stability. Synthetic 3'UTRs corresponding to sequences as short as 25 nucleotides from the central region of the 3'UTR of DDB2 were sufficient to accelerate decay of the heterologous reporter mRNA. Conversely, these same 3'UTRs led to more rapid induction of the reporter mRNA, export of the message to the cytoplasm and the subsequent accumulation of the encoded reporter protein, indicating that this newly identified cis-acting element affects transcriptional and post-transciptional processes. These results provide clear evidence that nuclear and cytoplasmic processing of the DDB2 mRNA is inextricably linked.

The cold inducible RNA binding protein (CIRBP) responds to a wide array of cellular stresses, including short wavelength ultraviolet light (UVC), at the transcriptional and post-translational level. CIRBP can bind the 3'untranslated region of specific transcripts to stabilize them and facilitate their transport to ribosomes for translation. Here we used RNA interference and oligonucleotide microarrays to identify potential downstream targets of CIRBP induced in response to UVC. Twenty eight transcripts were statistically increased in response to UVC and these exhibited a typical UVC response. Only 5 of the 28 UVC-induced transcripts exhibited a CIRBP-dependent pattern of expression. Surprisingly, 3 of the 5 transcripts (IL1B, IL8 and TNFAIP6) encoded proteins important in inflammation with IL-1β apparently contributing to IL8 and TNFAIP6 expression in an autocrine fashion. UVC-induced IL1B expression could be inhibited by pharmacological inhibition of NFκB suggesting that CIRBP was affecting NF-κB signaling as opposed to IL1B mRNA stability directly. Bacterial lipopolysaccharide (LPS) was used as an activator of NF-κB to further study the potential link between CIRBP and NFκB. Transfection of siRNAs against CIRBP reduced the extent of the LPS-induced phosphorylation of IκBα, NF-κB DNA binding activity and IL-1β expression. The present work firmly establishes a novel link between CIRBP and NF-κB signaling in response to agents with diverse modes of action. These results have potential implications for disease states associated with inflammation.

Significant advances have led to receptor induced-fit and conformational selection models for describing bimolecular recognition, but a more comprehensive view must evolve to also include ligand shape and conformational changes. Here, we describe an example where a ligand's "structural hinge" influences potency by inducing an "L-shape" bioactive conformation, and due to its solvent exposure in the complex, reasonable conformation-activity-relationships can be qualitatively attributed. From a ligand design perspective, this feature was exploited by successful linker hopping to an alternate "structural hinge" that led to a new and promising chemical series which matched the ligand bioactive conformation and the pocket bioactive space. Using a combination of X-ray crystallography, NMR and modeling with support from binding-site resistance mutant studies and photoaffinity labeling experiments, we were able to derive inhibitor-polymerase complexes for various chemical series.

SAR studies at the N(1)-position of allosteric indole-based HCV NS5B inhibitors has led to the discovery of acetamide derivatives with good cellular potency in subgenomic replicons (EC(50) <200 nM). This class of inhibitors displayed improved physicochemical properties and favorable ADME-PK profiles over previously described analogs in this class., (Copyright (c) 2010 Elsevier Ltd. All rights reserved.)

Study Design: Case report., Objectives: To report a case where a monoplegic patient received L-DOPA and/or buspirone. These compounds, normally used in the treatment of Parkinson's disease and anxiety respectively, were recently shown to induce spinal locomotor network activity and reflex stepping-like movements in animal models of spinal cord injury (SCI). However, the safety of these drugs as potential treatments for Central Pattern Generator (CPG) activation in paralyzed individuals remain unclear., Setting: St-Jean-Chrysostome, Quebec, Canada., Method: The acute effects induced by these compounds were qualitatively assessed by the patient, a 38-year-old man who underwent surgery 17 years ago to remove an intracavernous angioma located at the mid-thoracic level (T5-T6) of the spinal cord., Results: Self-administration every 2 days of L-DOPA (200, 400 and 600 mg, p.o.) and buspirone (5, 10 and 15 mg, p.o.) either separately or combined led to no atypical side effects (that is, occasional sleepiness, nervousness, insomnia or mild headaches). No movement was induced ipsilaterally although some sensations referred to by the patient as an increased blood flow in the lower back and upper leg regions were reported shortly after administration of the combined treatment., Conclusion: The results show no significant side effects following acute administration of L-DOPA and/or buspirone. This constitutes the first report providing preliminary evidence of safety following administration of these drugs in incompletely paralyzed individuals. The sensations of increased blood flow ipsilaterally with the combined treatment may also suggest that the dose regimen was not optimal or sub-threshold for inducing detectable CPG-mediated leg movements.

Benzimidazole-based allosteric inhibitors of the hepatitis C virus (HCV) NS5B polymerase were diversified to a variety of topologically related scaffolds. Replacement of the polar benzimidazole core by lipophilic indoles led to inhibitors with improved potency in the cell-based subgenomic HCV replicon system. Transposing the indole scaffold into a previously described series of benzimidazole-tryptophan amides generated the most potent inhibitors of HCV RNA replication in cell culture reported to date in this series (EC(50) approximately 50 nM).

The virally encoded NS3 protease is essential to the life cycle of the hepatitis C virus (HCV), an important human pathogen causing chronic hepatitis, cirrhosis of the liver, and hepatocellular carcinoma. The design and synthesis of 15-membered ring beta-strand mimics which are capable of inhibiting the interactions between the HCV NS3 protease enzyme and its polyprotein substrate will be described. The binding interactions between a macrocyclic ligand and the enzyme were explored by NMR and molecular dynamics, and a model of the ligand/enzyme complex was developed.

Antimony-containing drugs are still the drugs of choice in the treatment of infections caused by the parasite Leishmania. Resistance to antimony is now common in some parts of the world, and several mechanisms of resistance have been described. By transfecting cosmid banks and selecting with potassium antimonyl tartrate (SbIII), we have isolated a cosmid associated with resistance. This cosmid contains 2 copies of the heat shock protein 70 (HSP70) and 1 copy of the heat shock cognate protein 70 (HSC70). Several data linked HSP70 to antimony response and resistance. First, several Leishmania species, both as promastigotes and amastigotes, increased the expression of their HSP70 proteins when grown in the presence of 1 or 2 times the Effect Concentration 50% of SbIII. In several mutants selected for resistance to either SbIII or to the related metal arsenite, the HSP70 proteins were found to be overexpressed. This increase was also observed in revertant cells grown for several passages in the absence of SbIII, suggesting that this increased production of HSP70 is stable. Transfection of HSP70 or HSC70 in Leishmania cells does not confer resistance directly, though these transfectants were better able to tolerate a shock with SbIII. Our results are consistent with HSP70 and HSC70 being a first line of defense against SbIII until more specific and efficient resistance mechanisms take over.

The synthesis of BILN 2061, an NS3 protease inhibitor with proven antiviral effect in humans, was accomplished in a convergent manner from four building blocks. The procedure described here was suitable for the preparation of multigram quantities of BILN 2061 for preclinical pharmacological evaluation., (Copyright 2004 American Chemical Society)

From the discovery of competitive hexapeptide inhibitors, potent and selective HCV NS3 protease macrocyclic inhibitors have been identified. Structure-activity relationship studies were performed focusing on optimizing the N-terminal carbamate and the aromatic substituent on the (4R)-hydroxyproline moiety. Inhibitors meeting the potency criteria in the cell-based assay and with improved oral bioavailability in rats were identified. BILN 2061 was selected as the best compound, the first NS3 protease inhibitor reported with antiviral activity in man.

The Boehringer Ingelheim compound collection was screened for inhibitors of the ATPase activity of human papillomavirus E1 helicase to develop antiviral agents that inhibit human papillomavirus (HPV) DNA replication. This screen led to the discovery of (biphenyl-4-sulfonyl)acetic acid 1, which inhibits the ATPase activity of HPV type 6 E1 helicase with a low micromolar IC(50) value. A hit-to-lead exercise rapidly converted 1 into a low nanomolar lead series.

In the protozoan parasite Leishmania, drug resistance can be a complex phenomenon. Several metabolic pathways and membrane transporters are implicated in the resistance phenotype. To monitor the expression of these genes, we generated custom DNA microarrays with PCR fragments corresponding to 44 genes involved with drug resistance. Transcript profiling of arsenite and antimony resistant mutants with these arrays pinpointed a number of genes overexpressed in mutants, including the ABC transporter PGPA, the glutathione biosynthesis genes gamma-glutamylcysteine synthetase (GSH1) and the glutathione synthetase (GSH2). Competitive hybridisations with total RNA derived from sensitive and methotrexate resistant cells revealed the overexpression of genes coding for dihydrofolate reductase (DHFR-TS), pteridine reductase (PTR1) and S-adenosylmethionine synthase (MAT2) and a down regulation of one gene of the folate transporter (FT) family. By labelling the DNA of sensitive and resistant parasites we could also detect several gene amplification events using DNA microarrays including the amplification of the S-adenosyl homocysteine hydrolase gene (SAHH). Alteration in gene expression detected by microarrays was validated by northern blot analysis, while Southern blots indicated that most genes overexpressed were also amplified, although other mechanisms were also present. The microarrays were useful in the study of resistant parasites to pinpoint several genes linked to drug resistance.

The first line drug against leishmaniasis consists of pentavalent antimony [Sb(V)], but there is general belief that the active form of the metal is the trivalent form [Sb(III)]. In this study, we have quantified the accumulation of Sb(V) and Sb(III) in Leishmania by using inductively coupled plasma mass spectrometry. The accumulation was studied in three Leishmania species at various life stages, sensitive or resistant to antimony. Both Sb(III) and Sb(V) are accumulated in promastigote and amastigote parasites, but through competition experiments with arsenite, we found that the routes of entry of Sb(V) and Sb(III) are likely to differ in Leishmania. The level of accumulation of either Sb(III) or Sb(V), however, was not correlated with the susceptibility of wild-type Leishmania cells to antimony. This suggests that other factors may also be implicated in the mode of action of the drugs. In contrast to metal susceptibility, resistance to Sb(III) correlated well with decreased antimony accumulation. This phenotype was energy dependent and highlights the importance of transport systems in drug resistance of this protozoan parasite.

Gluthathione (GSH) has been previously shown to promote the reduction of pentavalent antimony (Sb(V)) into the more toxic trivalent antimony (Sb(II)) in the antimonial drug, meglumine antimonate. However, this reaction occurred at acidic pH (pH 5) but not at the pH of the cytosol (pH 7.2) in which GSH is encountered. The aim of the present study was to further characterize the reaction between thiols and antimonial drugs, addressing the following aspects: (i) the reducing activity of cysteine (Cys) and cysteinyl-glycine (Cys-Gly), expected to be the predominant thiols in the acidic compartments of mammalian cells; (ii) the reducing activity of trypanothione (T(SH)2), the main intracellular thiol in Leishmania parasites; (iii) the influence of the state of complexation of Sb(V) on the rate of Sb(V) reduction. We report here that Cys, Cys-Gly and T(SH)2 did promote the reduction of Sb(V) into Sb(III) at 37 degrees C. Strikingly, the initial rates of reduction of Sb(V) were much greater in the presence of Cys-Gly, Cys and T(SH)2 than in the presence of GSH. These reactions occurred at both pH 5 and pH 7 but were favored at acidic pH. Moreover, our data shows that Sb(V) is reduced more slowly in the form of meglumine antimonate than in its non-complexed form, indicating that the complexation of Sb(V) tends to slow down the rate of its reduction. In conclusion, our data supports the hypothesis that Sb(V) is reduced in vivo by T(SH)2 within Leishmania parasites and by Cys or Cys-Gly within the acidic compartments of mammalian cells.

In many African countries, the economy largely depends on agriculture. Pesticides are therefore likely to represent an important source of xenoestrogens in contaminated rivers and lagoons. The largely uncontrolled use of diverse pesticides led us to hypothesize that these agents, and particularly organochlorine compounds, may pose a serious problem in the Republic of Benin. To verify our hypothesis, tilapia (Sarotherodon melanotheron) from five sites in the southern part of the main Ouémé River were analyzed. Ouémé River drains the southern region of the country. Vitellogenin (Vtg) was used as an indicator of contaminated sites. This approach has its limitations, because there are a wide variety of man-made chemicals present in the aquatic environment likely to induce Vtg in male fish. Therefore, in this study this approach allows us to define potential contaminated target sites. In order to determine whether the presence of Vtg could be attributable to pesticides, organochlorine pesticides in the flesh of tilapia were also analyzed. Significant amounts of Vtg in fish from contaminated sites were detected, and were correlated with organochlorine pesticide levels in tissue. These results indicate that organochlorine pesticides are present in the Ouémé River and that these compounds can act as endocrine modulators in this ecosystem. Eating fish from contaminated rivers, such as the Ouémé River, may contribute to the accumulation of high concentrations of these pesticides in the body, leading to exposure to their negative effects.

The copper redhorse (Catostomidae: Moxostoma hubbsi) is an endangered fish species whose worldwide distribution is limited to the St. Lawrence River and three of its tributaries, in Canada. Severe reproductive impairment and lack of successful recruitment reported in this species have been hypothetically associated with water pollution. In order to obtain an initial description of contamination levels in copper redhorse, seven accidentally-killed specimens from the Richelieu River were analyzed for trace metals, organochlorine pesticides, chlorobenzenes, PAHs, PCBs, dioxins and furans. Fish varied between 9 and 33 years of age, which corresponds to mature individuals. The levels of contaminants analyzed in different body tissues were close to and often lower than levels reported in other catostomid fish species from nearby locations within the St. Lawrence River basin. Concentrations of total mercury, cadmium and co-planar PCBs increased with fish age. The types and concentrations of contaminants found suggested that the Richelieu River spawning population of copper redhorse would migrate and spend time in the St. Lawrence River. Concentrations of many contaminants were often highest in gonadal tissues, but levels were much lower than reported in the literature as causing reproductive impairment or egg and fry mortality in fish. Further research is needed to assess the potential link between contaminants and reproductive failure in this endangered fish species.

A substoichiometric amount of titanium-TADDOLate complex was effective at catalyzing the cyclopropanation reaction of allylic alcohols in the presence 1 equiv of bis(iodomethyl)zinc. After initial optimization of the catalyst structure, excellent yields and enantiomeric ratios were obtained for 3-aryl- or 3-heteroaryl-substituted allylic alcohols (up to 97:3). Alkyl-substituted allylic alcohols gave modest yields and enantiomeric ratios (up to 87:13) but these compare favorably with those observed with other substoichiometric chiral ligands. The full synthetic scope of the reaction is presented in this paper.

In this paper, we report our findings regarding the development of a Lewis acid-catalyzed cyclopropanation of allylic alcohols with bis(iodomethyl)zinc. Iodomethylzinc alkoxides can be formed by treatment of an alcohol with bis(iodomethyl)zinc. These species are not prone to undergo cyclopropanation at low temperature but the addition of a Lewis acid in catalytic amounts induces the cyclopropanation reaction. Using this procedure, we demonstrated that the Lewis acid-catalyzed pathway significantly overwhelms the uncatalyzed one. This paper describes fundamental issues regarding the preparation and stability of halomethyl zinc alkoxides in solution as well as their aggregation state in solution and solid-state structures. Furthermore, the competition reaction between the inter- vs intramolecular cyclopropanation will be studied. Finally, we will discuss the possible activation pathways to explain the Lewis acid activation of halomethylzinc alkoxides. These findings provided new insights on the reactivity of ROZnCH(2)I and established the groundwork for the elaboration of an enantioselective version of the reaction.

Patients with type 2 diabetes often have impaired exercise capacity compared with nondiabetic subjects. Left ventricular (LV) diastolic dysfunction has been shown to limit exercise performance in nondiabetic subjects. Men with well-controlled type 2 diabetes were divided into 2 groups: normal LV diastolic function (group 1, n = 9) or LV diastolic dysfunction (group 2, n = 10) based on standard echocardiographic criteria using pulmonary veins and transmitral flow recordings. They were matched for age and had no evidence of systemic hypertension, macroalbuminuria, coronary artery disease, congestive heart failure, clinical diabetic complications, and thyroid disease. Good metabolic control was demonstrated by glycated hemoglobin levels of 6.7+/-1.6% and 6.6+/-2.5% (means +/- SD) in patients with LV diastolic dysfunction and in controls, respectively. Each subject performed a symptom-limited modified Bruce protocol treadmill exercise test. Maximal treadmill performance was higher in subjects with normal diastolic function compared with subjects with LV diastolic dysfunction when expressed in time (803+/-29 vs. 662+/-44 seconds, respectively, p<0.02) or in METs (11.4+/-1.2 vs. 9.5+/-1.9 METs, respectively, p<0.02). Moreover, there was a correlation between E/A ratio and exercise duration (r = 0.64, p = 0.004) or E/A ratio and METs (r = 0.658, p = 0.003). There were no significant differences in maximal heart rate, maximal systolic and diastolic blood pressure, or maximal rate-pressure product attained during the exercise test. In conclusion, this study demonstrated that LV diastolic dysfunction influences maximal treadmill performance and could explain lower maximal performance observed in patients with type 2 diabetes.

Rat splenocyte mixed leukocyte reaction (MLR), splenic natural killer (NK) cell activity, and phagocytic activities of splenic, peritoneal, and peripheral blood leukocytes (PBLs) were evaluated in vitro to determine the immunotoxicity of mixtures containing low levels of methylmercury (MeHg), polychlorinated dibenzo-p-dioxins (PCDDs), polychlorinated dibenzofurans (PCDFs), and Aroclor polychlorinated biphenyls (PCBs). The mixtures were based on the concentrations of the chemicals in fish flesh. Leukocytes from male Fischer rats were exposed to MeHg (0.1-2 microg/ml), PCDD/PCDF mixtures (1-15 pg/ml) of three PCDDs (2,3,7,8-tetrachlorodibenzo-p-dioxin, 1,2,3,7,8-pentachlorodibenzo-p-dioxin, and 1,2,3,4,7,8-hexachlorodibenzo-p-dioxin) and two PCDFs (2,3,7,8-tetrachlorodibenzofuran and 1,2,3,7,8-pentachlorodibenzofuran), three Aroclor PCB (Aroclor 1242, 1254, and 1260) mixtures (0.01-0.5 microg/ml), or combinations of MeHg/PCB/PCDD/PCDF mixtures for 24 or 72 h before immunological assays. Phagocytosis and NK cell cytotoxicity were evaluated with a flow cytometer, and MLR of Fischer rat responder splenocytes cultured with mitomycin C-treated Long-Evans splenocytes by [3H]thymidine uptake. Exposure to MeHg (2 microg/ml) alone or with PCB/ PCDD/PCDF resulted in significant cytolethality in rat splenocytes, peritoneal leukocytes, and PBLs at 24 h exposure. Treatment with Aroclor PCB mixtures, PCDD/PCDF mixtures, 0.1 microg MeHg/ml (noncytolethal), or PCB/PCDD/PCDF mixtures with 0.1 microg MeHg/ml caused no suppression of splenocyte MLR response, splenic NK cell-mediated lysis of Yac-l cells, or phagocytosis of fluorescent beads by splenic, peritoneal, and peripheral blood phagocytic cells. The results indicate that in vitro exposure of rat leukocytes to low levels of MeHg, Aroclor PCB mixtures, PCDD/PCDF mixtures, or MeHg/PCB/PCDD/PCDF mixtures had no suppressive effects on the immune functions assayed, and thus produced no additive immunotoxicity. However, in order to predict the potential risk of these chemical mixtures to the human immune system, in vivo animal studies with blood (tissue) levels compatible with the levels of MeHg, PCBs, and PCDDs/PCDFs in exposed human populations should be evaluated.

ABC transporters have been found in several parasitic protozoa including Leishmania. At least two Leishmania ABC transporters are involved in drug resistance. One is PgpA, which is involved in resistance to arsenic and antimony-containing compounds. Antimonials are the drug of choice against Leishmania infections. Transfection and biochemical studies suggest that PgpA recognizes metals conjugated to thiols. The second ABC transporter is closely related to mammalian P-glycoproteins and confers resistance to anticancer drugs by a mechanism that remains to be elucidated. Additional ABC transporters are likely to be present in Leishmania and these are discussed in relation to the phenomenon of antimony resistance.

The mechanism of resistance to the metal arsenite has been studied and compared in L. mexicana, L. tropica, and L. tarentolae selected in a step by step manner for arsenite resistance. Amplification of the ABC transporter gene pgpA was found to be a frequent resistance mechanism in all species. Transfection of pgpA genes into different species indicated that both the origin of the pgpA gene and the recipient strain into which the gene is transfected seem important for resistance. An increase in the levels of trypanothione was also correlated with metal resistance in different Leishmania species. The mechanism used to increase the levels of trypanothione seems to differ, however, between the different species. This study points to a key role of transporters and thiol levels in metal resistance in Leishmania.

The effect of environmental stress on the accuracy of protein synthesis in an Escherichia coli and a rat brain cell-free system was investigated. Poly-U was translated in a rat brain and an E. coli cell-free extract under identical ionic conditions. The fidelity of translation, both in the E. coli and the rat brain extracts, was commensurate with what is known about the accuracy of translation in vivo. The incorporation of phenylalanine (code: UUU) and leucine (code: CUU, UUG or A) was measured at various Mg2+ concentrations (3 to 22 mM), various pH's (6.6 to 8.6), various temperatures (23 to 42 degrees C), and in the presence or absence of 2.4% (v/v) ethanol. It was observed that (i) the accuracy of translation was generally higher in extracts from E. coli than from rat brain, and (ii) relative to that in E. coli, the translation fidelity in rat brain extracts was about 2 times more sensitive to ethanol, at least 5 times more sensitive to temperature, and at least 50 times more sensitive to pH. It was found that this differential sensitivity was not due to a differential behavior of the bacterial and the mammalian aminoacyl-tRNA synthetases under stress, but rather to the process of chain elongation itself. It is concluded that the accuracy of protein synthesis is more resistant to environmental stress in E. coli extracts than in extracts from at least one mammalian tissue.