Discovery of novel Cyclophilin D inhibitors starting from three dimensional fragments with millimolar potencies

Fragment-based screening by SPR enabled the discovery of chemical diverse fragment hits with millimolar binding affinities to the peptidyl-prolyl isomerase Cyclophilin D (CypD). The CypD protein crystal structures of 6 fragment hits provided the basis for subsequent medicinal chemistry optimization by fragment merging and linking yielding three different chemical series with either urea, oxalyl or amide linkers connecting millimolar fragments in the S1′ and S2 pockets. We successfully improved the in vitro CypD potencies in the biochemical FP and PPIase assays and in the biophysical SPR binding assay from millimolar towards the low micromolar and submicromolar range by >1000-fold for some fragment derivatives. The initial SAR together with the protein crystal structures of our novel CypD inhibitors provide a suitable basis for further hit-to-lead optimization.     

酵母菌促进铅和镉胁迫下麻疯树生长的研究

为了筛选对铅和镉具有抗性和吸附性的酵母菌,构建麻疯树根系-酵母菌联合修复体系,促进高浓度铅和镉胁迫下麻疯树的生长。该研究分别从麻疯树的根段、珙桐的茎段、珙桐的根段分离到3株具有铅、镉抗性的酵母菌,分别命名为Jc、Di1、Di2,测定了三者对铅、镉的抗性和吸附性,并将筛选出的2株能吸附铅、镉的酵母菌菌株接种到麻疯树幼苗,研究接种两种酵母菌的麻疯树植株对铅、镉胁迫的响应。结果表明:经形态学和生理生化特征观察,Jc初步鉴定红酵母属(Rhodotorula sp.),Di1为假丝酵母属(Candida sp.),Di2为德巴利酵母属(Debaryomyces sp.)。三种酵母菌对铅、镉都有一定的抗性,其抗性能力的大小为JcDi2Di1。Di1和Jc对铅和镉都具有一定的吸附性将其用于接种麻疯树幼苗。与不接种酵母菌(CK)的麻疯树植株相比,接种Di1和Jc的麻疯树植株在根、茎、叶、全株干重方面显著增加,叶绿素、全株氮、全株磷浓度显著增加,SOD、POD、CAT的活性提高,丙二醛(MDA)浓度显著下降。从综合接种效应来看,Jc、Di1作为铅、镉的钝化剂,是铅、镉胁迫下促进麻疯树生长的备选菌株,这对于提高麻疯树对铅、镉污染土壤修复效率具有重要的意义。     

大豆蛋白、苹果果胶及维生素C 排铅效果的探究

通过对铅中毒小鼠灌胃三种不同剂量的食物材料,探究大豆蛋白、苹果果胶及维生素C的排铅效果。实验通过每日对铅中毒小鼠喂食高、中、低三种不同剂量的苹果果胶、大豆蛋白和维生素C,连续28 d。然后收集小鼠的血液、肝脏和股骨,用原子吸收光谱仪进行铅检测。结果表明重金属铅的摄入会影响小鼠的健康和生长状况;大豆蛋白、苹果果胶和维生素C均具有一定的排铅效果,在最佳的条件下,血铅含量降低了65%~70%,肝脏铅含量下降了63%~85%,股骨铅含量下降了18%~26%。本实验证明大豆蛋白、苹果果胶及维生素C三种食品材料具有较好的降血铅效果,维生素C被认为是最适合用于开发排铅功能食品的原料。本研究为排铅功能食品的研发提供了重要的理论依据。     

Marshall to the rescue in cardiac resynchronization therapy: Left ventricular lead placement in coronary sinus ostial atresia

This case highlights the importance of proper identification of congenital anomalies of the coronary sinus for the successful placement of left ventricular lead during cardiac resynchronization therapy device implantation. We discuss an alternate route for left ventricular lead placement via the vein of Marshall when the coronary sinus ostium in the right atrium was atretic and was facing difficulty initially in detecting the anomaly.     

Quality control and statistical modeling for environmental epigenetics: A study on in utero lead exposure and DNA methylation at birth

DNA methylation data assayed using pyrosequencing techniques are increasingly being used in human cohort studies to investigate associations between epigenetic modifications at candidate genes and exposures to environmental toxicants and to examine environmentally-induced epigenetic alterations as a mechanism underlying observed toxicant-health outcome associations. For instance, in utero lead (Pb) exposure is a neurodevelopmental toxicant of global concern that has also been linked to altered growth in human epidemiological cohorts; a potential mechanism of this association is through alteration of DNA methylation (e.g., at growth-related genes). However, because the associations between toxicants and DNA methylation might be weak, using appropriate quality control and statistical methods is important to increase reliability and power of such studies. Using a simulation study, we compared potential approaches to estimate toxicant-DNA methylation associations that varied by how methylation data were analyzed (repeated measures vs. averaging all CpG sites) and by method to adjust for batch effects (batch controls vs. random effects). We demonstrate that correcting for batch effects using plate controls yields unbiased associations, and that explicitly modeling the CpG site-specific variances and correlations among CpG sites increases statistical power. Using the recommended approaches, we examined the association between DNA methylation (in LINE-1 and growth related genes IGF2, H19 and HSD11B2) and 3 biomarkers of Pb exposure (Pb concentrations in umbilical cord blood, maternal tibia, and maternal patella), among mother-infant pairs of the Early Life Exposures in Mexico to Environmental Toxicants (ELEMENT) cohort (n = 247). Those with 10 μg/g higher patella Pb had, on average, 0.61% higher IGF2 methylation (P = 0.05). Sex-specific trends between Pb and DNA methylation (P < 0.1) were observed among girls including a 0.23% increase in HSD11B2 methylation with 10 μg/g higher patella Pb.     

Lead exposure induces changes in 5-hydroxymethylcytosine clusters in CpG islands in human embryonic stem cells and umbilical cord blood

Prenatal exposure to neurotoxicants such as lead (Pb) may cause stable changes in the DNA methylation (5mC) profile of the fetal genome. However, few studies have examined its effect on the DNA de-methylation pathway, specifically the dynamic changes of the 5-hydroxymethylcytosine (5hmC) profile. Therefore, in this study, we investigate the relationship between Pb exposure and 5mC and 5hmC modifications during early development. To study the changes in the 5hmC profile, we use a novel modification of the Infinium™ HumanMethylation450 assay (Illumina, Inc.), which we named HMeDIP-450K assay, in an in vitro human embryonic stem cell model of Pb exposure. We model Pb exposure-associated 5hmC changes as clusters of correlated, adjacent CpG sites, which are co-responding to Pb. We further extend our study to look at Pb-dependent changes in high density 5hmC regions in umbilical cord blood DNA from 48 mother-infant pairs from the Early Life Exposure in Mexico to Environmental Toxicants (ELEMENT) cohort. For our study, we randomly selected umbilical cord blood from 24 male and 24 female children from the 1st and 4th quartiles of Pb levels. Our data show that Pb-associated changes in the 5hmC and 5mC profiles can be divided into sex-dependent and sex-independent categories. Interestingly, differential 5mC sites are better markers of Pb-associated sex-dependent changes compared to differential 5hmC sites. In this study we identified several 5hmC and 5mC genomic loci, which we believe might have some potential as early biomarkers of prenatal Pb exposure.     

Relationship of lead and essential elements in whole blood from school-age children in Nanning,China

ObjectivesThe aim of this study was to investigate blood lead level and its relationship to essential elements (zinc, copper, iron, calcium and magnesium) in school-age children from Nanning, China.MethodsA total of 2457 children aged from 6 to 14 years were enrolled in Nanning, China. The levels of lead (Pb), zinc (Zn), copper (Cu), iron (Fe), calcium (Ca) and magnesium (Mg) were determined by an atomic absorption spectrometer.ResultsThe mean blood lead level (BLL) was 57.21 ± 35.00 μg/L. 188 (7.65%) asymptomatic children had toxic lead level higher than 100 μg/L. The school-age boys had similar lead level among different age groups, while the elder girls had less BLL. The blood Zn and Fe were found to be increased in the boys with elevated BLL, but similar trends were not observed in the girls. Positive correlations between Pb and Fe or Mg (r = 0.112, 0.062, respectively, p < 0.01) and a negative correlation between Pb and Ca (r = −0.047, p < 0.05) were further established in the studied children.ConclusionsLead exposure in school-age children was still prevalent in Nanning. The boys and girls differed in blood levels of lead and other metallic elements. Lead exposure may induce metabolic disorder of other metallic elements in body.     

Cribra orbitalia and trace element content in human teeth from neolithic and early bronze age graves in southern Poland

Determination of element levels in bones and teeth can complement knowledge of the diagnostics and etiology of various diseases in prehistoric populations. Calcium (Ca), cadmium (Cd), copper (Cu), iron (Fe), and lead (Pb) content were analyzed in teeth from human skeletons dated to 3,000–1,400 BC from Malopolska Upland loess. Levels of iron and calcium were determined using atomic absorption spectroscopy (AAS), and lead, cadmium, and copper levels were measured using anodic stripping voltametry (ASV). Molar teeth from specimens with cribra orbitalia were selected for analyses, and teeth from specimens with no pathological changes were used as a control. No significant correlations between the content of particular elements and the tooth class, specimen age, or depth of burial pit were observed. The Fe content in specimens with cribra orbitalia is not the best measure for this disease's etiology. Thus, interelement correlations and proportions might give a better picture of the biological condition of the specimen and of the investigated groups. Am J Phys Anthropol 103:201–207, 1997. © 1997 Wiley-Liss, Inc.     

Lead and catechol hematotoxicity in vitro using human and murine hematopoietic progenitor cells

In vitro cloning assays for hematopoietic myeloid and erythroid precursor cells have been used as screening systems to investigate the hematotoxic potential of environmental chemicals in humans and mice. Granulocyte-monocyte progenitors (CFU-GM) from human umbilical cord blood and from mouse bone marrow (Balb/c and B6C3F1) were cultured in the presence of lead and the benzene metabolite catechol. Erythroid precursors (BFU-E) from human umbilical cord blood were cultured in the presence of lead. The in vitro exposure of the human and murine cells resulted in a dose-dependent depression of the colony numbers. The concentration–effect relationship was studied. Results showed that: (1) Based on calculated IC50 values, human progenitors are more sensitive to lead and catechol than are murine progenitors. The dose that caused a 50% decrease in colony formation after catechol exposure was 6 times higher for murine cells (IC50 = 24 μmol/L) than for human cord blood cells (IC50 = 4 μmol/L). Lead was 10–15 times more toxic to human hematopoietic cells (IC50 = 61 μmol/L) than to murine bone marrow cells from both mice strains tested (Balb/c, IC50 = 1060 μmol/L; B6C3F1, IC50 = 536 μmol/L). (2) A lineage specificity was observed after exposure to lead. Human erythroid progenitors (hBFU-E) (IC50 = 3.31 μmol/L) were found to be 20 times more sensitive to the inhibitory effect of lead than were myeloid precursors (hCFU-GM) (IC50 = 63.58 μmol/L). (3) Individual differences in the susceptibility to the harmful effect of lead were seen among cord blood samples. (4) Toxicity of lead to progenitor cells occurred at environmentally relevant concentrations. This revised version was published online in July 2006 with corrections to the Cover Date.     

Metabolic responses of Lemna minor to lead ions I. Growth, chlorophyll level and activity of fermentative enzymes

Duckweed Lemna minor L. was grown on Wang culture medium supplemented with lead ions for 24 hours. Metal was tested at 1.5, 3 and 6 mg·dm⁻³ concentrations. The growth of Lemna minor was inhibited by lead ions, but the dry to fresh weight ratio increased as the concentration of Pb²⁺ in the medium increased. With increased concentrations of Pb ions, the contents of chlorophyll a and chlorophyll b in roots and fronds were correspondingly lower in comparision with the control. The effect of lead upon activities of some glycolitic and fermentative enzymes in roots of duckweed was examined. The activity of pyruvate kinase decreased with increasing lead concentrations, but cytosolic malate dehydrogenase behaved in an opposite manner. The lowest concentration of Pb stimulated alcohol dehydrogenase; phosphoenolopyruvate carboxylase activity was maintained at relatively constant values at all tested lead concentrations.