Pre-receptorial regulation of steroid hormones in bone cells: insights on glucocorticoid-induced osteoporosis

In the past decades, concern on glucocorticoid-induced osteoporosis has increased with the widespread use of exogenous glucocorticoids (GC). Mature bone-forming cells (osteoblasts) are considered to be the principal site of action of GC in the skeleton. More likely, it is the entire cellular and molecular network surrounding these cells that is targeted by pharmacological doses of GC. Not only osteoblast and osteocyte metabolism, but the whole differentiation of mesenchymal stem cell toward the osteoblast lineage has been proven to be sensitive to GC. The effects of GC on this process are different according to the stage of differentiation of bone cell precursors. The presence of intact GC signalling is crucial for normal bone development and physiology, as opposed to the detrimental effect of high dose exposure. Both the physiological and pharmacological effects of GC are locally modulated by the activity of the 11β-hydroxysteroid dehydrogenase 1 (HSD1) that acts primarily as a glucocorticoid activator converting the inactive glucocorticoid (cortisone) into the active hormone (cortisol). We reviewed the metabolic and differentiation pathways controlled by GC signalling. These data have been merged with the recent evidences that 11β-HSD1 exert an important role by regulating the vulnerability of bone cells to GC. The different kinetics of 11β-HSD1 at various stage of differentiation and the GC-dependency of enzymatic activity have been presented.     

Applications of forensic chemistry to environmental work

Recent trends in environmental remediation have increasingly employed the use of environmental chemistry techniques to decipher the source(s) and fate of the contaminants and, in some cases, to determine their age or apportion them to sources. An extensive database of pyrogenic and petrogenic 'chemical fingerprints' has been constructed by the Gas Technology Institute (GTI) and META Environmental, Inc. using gas chromatography coupled with a flame ionization detector (GC/FID) or with a mass spectrometer (GC/MS). The use of these chemical fingerprinting techniques have been highly successful in discerning wastes from wholly different sources as well as among Manufactured Gas Plant (MGP)-type wastes from different plant operations. However, these techniques have been limited when low-level polycyclic aromatic hydrocarbon (PAH) discernment is required. Specifically, these techniques often do not provide data with sufficient conclusive discriminating power between the 'urban background'PAH sources and those from MGP-operations, which is pertinent for meeting low-level, stringent site-cleanup standards. GTI has been developing a new analytical method for the measurement of 'urban background' PAH contamination. This method measures the compound-specific isotope ratio (CSIR) carbon with a GC/IRMS (isotope ratio mass spectrometer). The GC/IRMS technique is a relatively new analytical tool that has great potential as an environmental forensic method at former MGP sites. This paper focuses on the applications of both chemical and isotopic analysis of samples to discern PAH contamination in the environment.     

Functions and dysfunctions of receptors for adrenal corticoids in the central nervous system

Glucocorticoids (GC) have several known effects on the function of the nervous system, and GC receptors have been identified in regions responding to hormonal action. In the spinal cord, GC receptors have been characterized in vitro, which share several biochemical properties in common with receptors in better studied areas such as the hippocampus. Moreover, enzymes which are induced by GC in the hippocampus, such as glycerolphosphate dehydrogenase and ornithine decarboxylase, are also under specific GC control in the spinal cord. Yet GC receptors in the latter tissue divert from those in hippocampus during some in vivo as well as in vitro studies. In vivo, uptake of [3H]corticosterone by purified cell nuclei was 5-8-fold higher in the hippocampus as compared to the cord. In vitro, a higher percentage of GC receptors previously transformed by heating, showed affinity towards DNA-cellulose in the spinal cord than in the hippocampus. The enzyme RNAse A effectively increased receptor binding to DNA-cellulose in hippocampus, whereas the cord was insensitive to its action. These results suggest that there is a "receptor dysfunction" in the spinal cord, the significance of which is poorly understood in terms of the accepted model of steroid hormone action.     

Preferential recombination between GC clusters in yeast mitochondrial DNA.

Yeast mitochondrial DNA molecules have long, AT-rich intergenic spacers punctuated by short GC clusters. GC-rich elements have previously been characterized by others as preferred sites for intramolecular recombination leading to the formation of subgenomic petite molecules. In the present study we show that GC clusters are favored sites for intermolecular recombination between a petite and the wild-type grande genome. The petite studied retains 6.5 kb of mitochondrial DNA reiterated tandemly to form molecules consisting of repeated units. Genetic selection for integration of tandem 6.5 kb repeats of the petite into the grande genome yielded a novel recombination event. One of two crossovers in a double exchange event occurred as expected in the 6.5 kb of matching sequence between the genomes, whereas the second exchange involved a 44 bp GC cluster in the petite and another 44 bp GC cluster in the grande genome 700 bp proximal to the region of homology. Creation of a mitochondrial DNA molecule with a repetitive region led to secondary recombination events that generated a family of molecules with zero to several petite units. The finding that 44 bp GC clusters are preferred as sites for intermolecular exchange adds to the data on petite excision implicating these elements as recombinational hotspots in the yeast mitochondrial genome.     

Applications of forensic chemistry to environmental work

Recent trends in environmental remediation have increasingly employed the use of environmental chemistry techniques to decipher the source(s) and fate of the contaminants and, in some cases, to determine their age or apportion them to sources. An extensive database of pyrogenic and petrogenic ‘chemical fingerprints’ has been constructed by the Gas Technology Institute (GTI) and META Environmental, Inc. using gas chromatography coupled with a flame ionization detector (GC/FID) or with a mass spectrometer (GC/MS). The use of these chemical fingerprinting techniques have been highly successful in discerning wastes from wholly different sources as well as among Manufactured Gas Plant (MGP)-type wastes from different plant operations. However, these techniques have been limited when low-level polycyclic aromatic hydrocarbon (PAH) discernment is required. Specifically, these techniques often do not provide data with sufficient conclusive discriminating power between the ‘urban background’PAH sources and those from MGP-operations, which is pertinent for meeting low-level, stringent site-cleanup standards. GTI has been developing a new analytical method for the measurement of ‘urban background’ PAH contamination. This method measures the compound-specific isotope ratio (CSIR) carbon with a GC/IRMS (isotope ratio mass spectrometer). The GC/IRMS technique is a relatively new analytical tool that has great potential as an environmental forensic method at former MGP sites. This paper focuses on the applications of both chemical and isotopic analysis of samples to discern PAH contamination in the environment.

     

DNA asymmetric strand bias affects the amino acid composition of mitochondrial proteins.

Variations in GC content between genomes have been extensively documented. Genomes with comparable GC contents can, however, still differ in the apportionment of the G and C nucleotides between the two DNA strands. This asymmetric strand bias is known as GC skew. Here, we have investigated the impact of differences in nucleotide skew on the amino acid composition of the encoded proteins. We compared orthologous genes between animal mitochondrial genomes that show large differences in GC and AT skews. Specifically, we compared the mitochondrial genomes of mammals, which are characterized by a negative GC skew and a positive AT skew, to those of flatworms, which show the opposite skews for both GC and AT base pairs. We found that the mammalian proteins are highly enriched in amino acids encoded by CA-rich codons (as predicted by their negative GC and positive AT skews), whereas their flatworm orthologs were enriched in amino acids encoded by GT-rich codons (also as predicted from their skews). We found that these differences in mitochondrial strand asymmetry (measured as GC and AT skews) can have very large, predictable effects on the composition of the encoded proteins.     

The response of the Golgi complex to microtubule alterations: the roles of metabolic energy and membrane traffic in Golgi complex organization

A striking example of the interrelation between the Golgi complex (GC) and microtubules is the reversible fragmentation and dispersal of the GC which occurs upon microtubule depolymerization. We have characterized dispersal of the GC after nocodazole treatment as well as its recovery from the dispersed state by immunofluorescent localization of beta 1, 4-galactosyltransferase in Madin-Darby bovine kidney cells. Immunofluorescent anti-tubulin staining allowed simultaneous examination of the microtubule array. Based on our results, dispersal can be divided into a three-step process: microtubule depolymerization, GC fragmentation, and fragment dispersal. In cells treated with metabolic inhibitors after microtubule depolymerization, neither fragmentation nor dispersal occur, despite the absence of assembled microtubules. Thus, fragmentation is energy dependent and not tightly linked to microtubule depolymerization. The slowing of fragmentation and dispersal by monensin or ammonium chloride, as well as progressive inhibition at less than 34 degrees C, suggest that ongoing membrane traffic is required for these processes. Similarly, recovery may be separated into four steps: microtubule depolymerization, GC fragment centralization, fragment coalescence, and polarization of the reticular GC network. Fragment centralization and coalescence were arrested by metabolic inhibitors, despite the presence of microtubules. Neither monensin nor ammonium choride inhibited GC recovery. Partial inhibition of recovery at reduced temperatures paralleled the extent of microtubule assembly. These data demonstrate that dispersal and recovery are multi-step operations, and that the individual steps differ in temperature dependence, energy dependence, and sensitivity to ionic perturbation. GC distribution and microtubule status have also been clearly dissociate, thereby proving that organization of the GC is an active process that is not simply determined by microtubule binding. Furthermore, the results indicate that ongoing intra-GC membrane traffic may participate in fragmentation and dispersal.     

Failed up-regulation of the inhibitory IgG Fc receptor Fc gamma RIIB on germinal center B cells in autoimmune-prone mice is not associated with deletion polymorphisms in the promoter region of the Fc gamma RIIB gene

FcgammaRIIB, a low-affinity FcR for IgG, inhibits BCR-mediated activation when these two receptors are co-cross-linked by Ags and IgG-containing immune complexes. Although a role for FcgammaRIIB in the germinal center (GC) reaction has been proposed, conflicting results have been published regarding the levels of FcgammaRIIB expressed on GC B cells in normal and autoimmune-prone mice and humans. In the present study, we investigate this issue in detail in mice by using multiple GC B cell markers, two different antigenic systems, primary and secondary GC responses, and by excluding the influence of splenic influx of immature B cells and passive acquisition of FcgammaRIIB from follicular dendritic cells. Our results are in concordance with previous data indicating that FcgammaRIIB expression is up-regulated on GC B cells in normal mice. In contrast, we observe comparable levels of FcgammaRIIB on GC and non-GC B cells in New Zealand White, New Zealand Black, and B6.Sle1 autoimmune-prone strains. Therefore, we suggest that these strains exhibit failed up-regulation of FcgammaRIIB on GC B cells, rather than down-regulation, as previously suggested. Also, in contrast to previous indications, this perturbed regulation is not uniquely associated with deletion polymorphisms in the promoter region of the FcgammaRIIB gene but does appear to be independent of genetic background. Finally, we present evidence indicating that FcgammaRIII, a low-affinity activating IgG FcR, is expressed on the GC B cells of normal but not autoimmune-prone mice.     

Bacteriologic and gas chromatographic diagnosis of nonclostridial anaerobic infection in suppurative-inflammatory diseases of the maxillofacial area and of the ENT organs

The etiological role of nonsporulating anaerobic microorganisms as the causative agents of inflammatory purulent processes in the E. N. T. and maxillofacial regions has been studied. The rapid diagnosis has been made by the method of gas and liquid chromatography (GC and LC), the results of chromatographic and bacteriological studies have been compared and the main causes of false results obtained in the GC/LC analysis have been considered. The information content of the GC/LC analysis has been shown to exceed that of the main clinico-laboratory results.     

Fish genomes: Sequencing trends,taxonomy and influence of taxonomy on genome attributes

594 fish genomes have been sequenced in past two decades, this represents 1.85% of the total reported fish species (32,000). Despite this no study represents the trends and only some studies have delved into how the genome size (GS) of the genomes are shaped by species taxonomy. However, all these studies have used data obtained by traditional cytometric methods and also have largely disregarded other genome attributes namely GC, number of chromosomes (CR), number of genes (GE), and protein count (PC). The present study used the most current data on genome attributes of fishes as generated by the whole genome sequencing projects to understand the trends, effect of taxonomy on the genome attributes (GS, GC, CR, GE, and PC) and the interrelation of genome attributes. The trends states that maximum number of fish genomes were sequenced in year 2020, order Cichliformes represents the highest number of published genomes, Illumina is the most used technology for sequencing fish genomes, etc. Our analyses exhibit some concrete trends for fishes as a whole and indicated a strong selection for smaller genomes among all vertebrates and a strong effect of taxonomy on all genome attributes. It also provides clear insights that the fish GS is significantly different from birds, amphibians, reptiles, mammals and insects while the GC only varied from insects. An inverse relation was observed between the GS and GC, and a direct relation was observed between the GS and CR, GE and PC. The results also signify that the per MB value of all the genome attributes decline with increasing GS.     

GC-biased gene conversion and selection affect GC content in the Oryza genus (rice)

Base composition varies among and within eukaryote genomes. Although mutational bias and selection have initially been invoked, more recently GC-biased gene conversion (gBGC) has been proposed to play a central role in shaping nucleotide landscapes, especially in yeast, mammals, and birds. gBGC is a kind of meiotic drive in favor of G and C alleles, associated with recombination. Previous studies have also suggested that gBGC could be at work in grass genomes. However, these studies were carried on third codon positions that can undergo selection on codon usage. As most preferred codons end in G or C in grasses, gBGC and selection can be confounded. Here we investigated further the forces that might drive GC content evolution in the rice genus using both coding and noncoding sequences. We found that recombination rates correlate positively with equilibrium GC content and that selfing species (Oryza sativa and O. glaberrima) have significantly lower equilibrium GC content compared with more outcrossing species. As recombination is less efficient in selfing species, these results suggest that recombination drives GC content. We also detected a positive relationship between expression levels and GC content in third codon positions, suggesting that selection favors codons ending with G or C bases. However, the correlation between GC content and recombination cannot be explained by selection on codon usage alone as it was also observed in noncoding positions. Finally, analyses of polymorphism data ruled out the hypothesis that genomic variation in GC content is due to mutational processes. Our results suggest that both gBGC and selection on codon usage affect GC content in the Oryza genus and likely in other grass species.     

Use of Electric Linear Dichroism and Competition Experiments with Intercalating Drugs to Investigate the Mode of Binding of Hoechst 33258, Berenil and DAPI to GC Sequences

Abstract

The drugs Hoechst 33258, berenil and DAPI bind preferentially to the minor groove of AT sequences in DNA Despite a strong selectivity for AT sites, they can interact with GC sequences by a mechanism which remains so far controversial. The 2-amino group of guanosine represents a steric hindrance to the entry of the drugs in the minor groove of GC sequences. Intercalation and major groove binding to GC sites of GC-rich DNA and polynucleotides have been proposed for these drugs. To investigate further the mode of binding of Hoechst 33258, berenil and DAPI to GC sequences, we studied by electric linear dichroism the mutual interference in the DNA binding reaction between these compounds and a classical intercalator, proflavine, or a DNA-threading intercalating drug, the amsacrine-4-carboxamide derivative SN16713. The results of the competition experiments show that the two acridine intercalators markedly affect the binding of Hoechst 33258, berenil and DAPI to GC polynucleotides but not to DNA containing AT/GC mixed sequences such as calf thymus DNA Proflavine and SN16713 exert dissimilar effects on the binding of Hoechst 33258, berenil and DAPI to GC sites. The structural changes in DNA induced upon intercalation of the acridine drugs into GC sites are not identically perceived by the test compounds. The electric linear dichroism data support the hypothesis that Hoechst 33258, berenil and DAPI interact with GC sites via a non-classical intercalation process.     

Base compositions of genes encoding alpha-actin and lactate dehydrogenase-A from differently adapted vertebrates show no temperature-adaptive variation in G + C content

There is a long-standing debate in molecular evolution concerning the putative importance of GC content in adapting the thermal stabilities of DNA and RNA. Most studies of this relationship have examined broad-scale compositional patterns, for example, total GC percentages in genomes and occurrence of GC-rich isochores. Few studies have systematically examined the GC contents of individual orthologous genes from differently thermally adapted species. When this has been done, the emphasis has been on comparing large numbers of genes in only a few species. We have approached the GC-adaptation temperature hypothesis in a different manner by examining patterns of base composition of genes encoding lactate dehydrogenase-A (ldh-a) and alpha-actin (alpha-actin) from 51 species of vertebrates whose adaptation temperatures ranged from -1.86 degrees C (Antarctic fishes) to approximately 45 degrees C (desert reptile). No significant positive correlation was found between any index of GC content (GC content of the entire sequence, GC content of the third codon position [GC(3)], and GC content at fourfold degenerate sites [GC(4)]) and any index of adaptation temperature (maximal, mean, or minimal body temperature). For alpha-actin, slopes of regression lines for all comparisons did not differ significantly from zero. For ldh-a, negative correlations between adaptation temperature and total GC content, GC(3), and GC(4) were observed but were shown to be due entirely to phylogenetic influences (as revealed by independent contrast analyses). This comparison of GC content across a wide range of ectothermic ("cold-blooded") and endothermic ("warm-blooded") vertebrates revealed that frogs of the genus Xenopus, which have commonly been used as a representative cold-blooded species, in fact are outliers among ectotherms for the alpha-actin analyses, raising concern about the appropriateness of choosing these amphibians as representative of ectothermic vertebrates in general. Our study indicates that, whereas GC contents of isochores may show variation among different classes of vertebrates, there is no consistent relationship between adaptation temperature and the percentage of thermal stability-enhancing G + C base pairs in protein-coding genes.     

Comparison of Headspace and Distillation Techniques for Soy Sauce Aroma in Relation to Analysis by Silica Capillary Gas Chromatography and Sensory Evaluation

A method has been applied to the collection of headspace volatiles from soy sauce at room temperature, using Tenax GC adsorption polymer. Concentrated volatiles obtained from fifty soy sauce samples consisting of genuine- and semi-fermented types using Tenax GC and distillation-extraction methods were analyzed by silica capillary GC analysis in order to compare the suitability to sensory data. From a multiple regression analysis, close relationships between two variously transformed GC data sets and sensory data were found, the square root transformation of peak area from the headspace GC data being the best amongst them. The precise extraction of characteristics contained in the sample volatiles, using either of the two aroma collecting methods, was confirmed by cluster analysis.     

Granger causality analysis of steady-state electroencephalographic signals during propofol-induced anaesthesia

Changes in conscious level have been associated with changes in dynamical integration and segregation among distributed brain regions. Recent theoretical developments emphasize changes in directed functional (i.e., causal) connectivity as reflected in quantities such as 'integrated information' and 'causal density'. Here we develop and illustrate a rigorous methodology for assessing causal connectivity from electroencephalographic (EEG) signals using Granger causality (GC). Our method addresses the challenges of non-stationarity and bias by dividing data into short segments and applying permutation analysis. We apply the method to EEG data obtained from subjects undergoing propofol-induced anaesthesia, with signals source-localized to the anterior and posterior cingulate cortices. We found significant increases in bidirectional GC in most subjects during loss-of-consciousness, especially in the beta and gamma frequency ranges. Corroborating a previous analysis we also found increases in synchrony in these ranges; importantly, the Granger causality analysis showed higher inter-subject consistency than the synchrony analysis. Finally, we validate our method using simulated data generated from a model for which GC values can be analytically derived. In summary, our findings advance the methodology of Granger causality analysis of EEG data and carry implications for integrated information and causal density theories of consciousness.     

Glucocorticoid resistance as a major drive in sepsis pathology

Sepsis is an acute systemic inflammatory disease. Glucocorticoids (GCs), which function by binding to the GC receptor GR have very powerful anti-inflammatory activities, yet they are hardly useful in sepsis. We can thus consider sepsis as a GC resistant disease. We here review the literature which has investigated this GC resistance, and summarize the mechanisms of GC resistance that have been observed in other diseases and in experimental models. We also discuss the importance of GC resistance in sepsis, in terms of the contribution of this phenomenon to the pathogenesis of sepsis.     

Population genetics of the vitamin D binding protein (GC) subtypes in the Asian-Pacific area: Description of new alleles at the GC locus

Isoelectric focussing (IEF) in thin layer polyacrylamide gels pH range 4-6.5 has been used to analyse the GC phenotypes of 4233 individuals from 28 different population groups in the Asian, Pacific, and Australian area. Because this technique reveals subtypes of the common GC*1 allele, there is almost a two-fold increase in the mean heterozygosity at the GC locus using IEF compared with conventional electrophoresis. The highest frequency (above 50%) of the GC*1S allele was encountered in Indian populations, reflecting genetic affinities with Europeans. By comparison, east and south east Asians are unique offing maximum values of the GC*1F allele (50%). With the exception of a few Pacific populations which show similar frequencies to east Asians, all other groups in the Pacific area, including Australia, have values of GC*1F similar to GC*1S ranging from 27% to 40%. The GC*2 frequency in most populations varies from 20% to 30%. However, some Polynesian groups have values up to 40% and Australian Aborigines less than 10%. Among other alleles, GC*1A1 is found to be widely distributed among Australian Aborigines and Melanesians and occurs sporadically in Polynesians, Micronesians, and in the Lesser Sunda Islands. Four new alleles, GC*1C24, GC*1C35 Aborigine, GC*1A21, and GC*1A22 are described. The gene frequency data at the GC locus has been used to calculate Nei genetic distances between the populations studied.