Transcriptional analysis in peripheral blood cells of individuals with elevated phthalate exposure - Results of the EuroMix study.

Exposure to phthalates is widespread in Europe. Phthalates are considered endocrine disrupting compounds and are classified as toxic for reproduction. However how phthalates affect the transcriptome in humans remains largely unknown. To investigate the effects of phthalate exposure on the transcriptomic profile we conducted RNA sequencing on peripheral blood samples from the Norwegian EuroMix cohort. We compared gene expression changes between participants with high, medium, and low exposure of six phthalates and 1,2-cyclohexane dicarboxylic acid diisononyl ester (DINCH). Comparing high and low exposure groups, DINCH was the compound that showed the highest number of differentially expressed genes (126 genes) followed by mono-n-butyl phthalate (MnBP; 89 genes) and mono-iso-nonyl phthalate (MiBP; 70 genes). Distributions between up- or down-regulated genes were similar across the different phthalates and DINCH. All phthalates including DINCH shared common differentially expressed genes ranging from 3 to 37 overlaps. Enriched Gene Ontology (GO) and biological pathway analysis revealed that most of the differentially expressed genes were associated with general cellular metabolism GO terms. MnBP and DINCH, particularly, showed a marked enrichment in various immunological function pathways including neutrophil degranulation, adaptive immune system and signaling by interleukins. Furthermore, the association between genes involved in the peroxisome proliferator activated receptor (PPAR) signaling pathway and phthalates, including DINCH, was evaluated. In total, 15 genes showed positive or negative associations across 5 phthalates and DINCH. MnBP and MiBP were the phthalate metabolites with the highest number of associations: 8 and 4 PPAR signaling pathway genes, respectively. Overall, we have performed an association study between phthalate exposure levels and modulation of transcriptomic profiles in human peripheral blood cells. DINCH, which is often mentioned as a substitute for phthalates, had comparable effects on differential gene expression in peripheral blood cells as phthalates.

The bovine alveolar macrophage DNA methylome is resilient to infection with Mycobacterium bovis.

DNA methylation is pivotal in orchestrating gene expression patterns in various mammalian biological processes. Perturbation of the bovine alveolar macrophage (bAM) transcriptome, due to Mycobacterium bovis (M. bovis) infection, has been well documented; however, the impact of this intracellular pathogen on the bAM epigenome has not been determined. Here, whole genome bisulfite sequencing (WGBS) was used to assess the effect of M. bovis infection on the bAM DNA methylome. The methylomes of bAM infected with M. bovis were compared to those of non-infected bAM 24 hours post-infection (hpi). No differences in DNA methylation (CpG or non-CpG) were observed. Analysis of DNA methylation at proximal promoter regions uncovered >250 genes harbouring intermediately methylated (IM) promoters (average methylation of 33-66%). Gene ontology analysis, focusing on genes with low, intermediate or highly methylated promoters, revealed that genes with IM promoters were enriched for immune-related GO categories; this enrichment was not observed for genes in the high or low methylation groups. Targeted analysis of genes in the IM category confirmed the WGBS observation. This study is the first in cattle examining genome-wide DNA methylation at single nucleotide resolution in an important bovine cellular host-pathogen interaction model, providing evidence for IM promoter methylation in bAM.

Atorvastatin up-regulate toxicologically relevant genes in rainbow trout gills.

There are large and increasing discharges of statins into the aquatic environment. Statins are cholesterol-lowering pharmaceuticals, inhibitors of 3-hydroxy-3-methylglutaryl coenzyme A reductase, an enzyme in the cholesterol synthesis pathway. Earlier studies have shown that statins will affect the expression of a range of genes in mammalian tissues and this group of pharmaceuticals has also been shown to affect membrane transporters. Changes in gene expression and ion transport in aquatic organisms may have dramatic consequences for the individual. The aim of the present study was to clarify whether waterborne exposure to a selected statin, atorvastatin, would affect gene expression in rainbow trout (Oncorhynchus mykiss) gill or liver or ion regulation in gills. Juvenile rainbow trout were exposed to two atorvastatin acid and atorvastatin lactone concentrations for 7 days (nominal concentrations 200 ng L(-1) and 10 µg L(-1)). The exposures caused up-regulated gene expression in gill, not liver, and only at the lowest concentration. Genes involved in membrane transport (pgp, mrp1), oxidative stress response (sod, mt), apoptosis (bax) and biotransformation (sult2b) were differentially expressed whereas the expression of genes involved in cholesterol biosynthesis (hmgr, fdps) or peroxisomal proliferation (ppar) were not affected. There were no significant changes in gill Na(+)/K(+) ATPase activity following exposure to atorvastatin. The pattern of differentially expressed genes in rainbow trout gills differ from responses previously observed in mammalian tissues following statin exposure.

Increased risk of phosphorus limitation at higher temperatures for Daphnia magna.

Invertebrate herbivores frequently face growth rate constraints due to their high demands for phosphorus (P) and nitrogen (N). Temperature is a key modulator of growth rate, yet the interaction between temperature and P limitation on somatic growth rate is scarcely known. To investigate this interaction, we conducted a study on the somatic growth rate (SGR) of the cladoceran Daphnia magna, known to be susceptible to P-limitation. We determined the SGR across a broad range of dietary P content of algae (carbon (C):P ratios (125-790), and at different temperatures (10-25°C). There was a strong impact of both temperature and C:P ratio on the SGR of D. magna, and also a significant interaction between both factors was revealed. The negative effect of dietary C:P on growth rate was reduced with decreased temperature. We found no evidence of P limitation at lowest temperature, suggesting that enzyme kinetics or other measures of food quality overrides the demands for P to RNA and protein synthesis at low temperatures. These findings also indicate an increased risk of P limitation and thus reduced growth efficiency at high temperatures.

Enhancing glutamate transport: mechanism of action of Parawixin1, a neuroprotective compound from Parawixia bistriata spider venom.

Previous studies have shown that a compound purified from the spider Parawixia bistriata venom stimulates the activity of glial glutamate transporters and can protect retinal tissue from ischemic damage. To understand the mechanism by which this compound enhances transport, we examined its effects on the functional properties of glutamate transporters after solubilization and reconstitution in liposomes and in transfected COS-7 cells. Here, we demonstrate in both systems that Parawixin1 promotes a direct and selective enhancement of glutamate influx by the EAAT2 transporter subtype through a mechanism that does not alter the apparent affinities for the cosubstrates glutamate or sodium. In liposomes, we observed maximal enhancement by Parawixin1 when extracellular sodium and intracellular potassium concentrations are within physiological ranges. Moreover, the compound does not enhance the reverse transport of glutamate under ionic conditions that favor efflux, when extracellular potassium is elevated and the sodium gradient is reduced, nor does it alter the exchange of glutamate in the absence of internal potassium. These observations suggest that Parawixin1 facilitates the reorientation of the potassium-bound transporter, the rate-limiting step in the transport cycle, a conclusion further supported by experiments showing that Parawixin1 does not stimulate uptake by an EAAT2 transport mutant (E405D) defective in the potassium-dependent reorientation step. Thus, Parawixin1 enhances transport through a novel mechanism targeting a step in the transport cycle distinct from substrate influx or efflux and provides a basis for the design of new drugs that act allosterically on transporters to increase glutamate clearance.