Pharmacological activation of the pyruvate dehydrogenase complex reduces statin-mediated upregulation of FOXO gene targets and protects against statin myopathy in rodents.

We previously reported that statin myopathy is associated with impaired carbohydrate (CHO) oxidation in fast-twitch rodent skeletal muscle, which we hypothesised occurred as a result of forkhead box protein O1 (FOXO1) mediated upregulation of pyruvate dehydrogenase kinase-4 (PDK4) gene transcription. Upregulation of FOXO gene targets known to regulate proteasomal and lysosomal muscle protein breakdown was also evident. We hypothesised that increasing CHO oxidation in vivo, using the pyruvate dehydrogenase complex (PDC) activator, dichloroacetate (DCA), would blunt activation of FOXO gene targets and reduce statin myopathy. Female Wistar Hanover rats were dosed daily for 12 days (oral gavage) with either vehicle (control, 0.5% w/v hydroxypropyl-methylcellulose 0.1% w/v polysorbate-80; n = 9), 88 mg( )kg(-1) day(-1) simvastatin (n = 8), 88 mg( )kg(-1) day(-1) simvastatin + 30 mg kg(-1) day(-1) DCA (n = 9) or 88 mg kg(-1) day(-1) simvastatin + 40 mg kg(-1) day(-1) DCA (n = 9). Compared with control, simvastatin reduced body mass gain and food intake, increased muscle fibre necrosis, plasma creatine kinase levels, muscle PDK4, muscle atrophy F-box (MAFbx) and cathepsin-L mRNA expression, increased PDK4 protein expression, and proteasome and cathepsin-L activity, and reduced muscle PDC activity. Simvastatin with DCA maintained body mass gain and food intake, abrogated the myopathy, decreased muscle PDK4 mRNA and protein, MAFbx and cathepsin-L mRNA, increased activity of PDC and reduced proteasome activity compared with simvastatin. PDC activation abolished statin myopathy in rodent skeletal muscle, which occurred at least in part via inhibition of FOXO-mediated transcription of genes regulating muscle CHO utilisation and protein breakdown.

A role for the pregnane X receptor in flucloxacillin-induced liver injury.

UNLABELLED: Drug-induced liver injury (DILI) due to flucloxacillin is a rare but serious complication of treatment. There is some evidence that flucloxacillin is a human pregnane X receptor (PXR) agonist. This study was designed to investigate the relevance of PXR to flucloxacillin toxicity and to identify genes changing in expression in response to flucloxacillin. Changes in gene expression in human hepatocytes after treatment with 500 microM flucloxacillin for 72 hours were examined by expression microarray analysis. The ability of flucloxacillin to act as a PXR agonist was investigated with reporter gene experiments. Flucloxacillin DILI cases (n = 51), drug-exposed controls without toxicity (n = 64), and community controls (n = 90) were genotyped for three common PXR polymorphisms. Luciferase reporter assays were used to assess the significance of a promoter region PXR polymorphism. Seventy-two probe sets representing 50 different genes showed significant changes in expression of 1.2-fold or higher. Most genes showing changes greater than 3-fold were known to be rifampicin-responsive, and this suggested a PXR-dependent mode of regulation. Using a luciferase-everted repeat separated by 6 base pairs element construct, we confirmed that flucloxacillin was a PXR agonist. We found a difference in the distribution of a PXR polymorphism (rs3814055; C-25385T) between flucloxacillin DILI cases and controls with the CC genotype associated with an increased risk of disease (odds ratio = 3.37, 95% confidence interval = 1.55-7.30, P = 0.0023). Reporter gene experiments showed lower promoter activity for the C allele than the T allele. CONCLUSION: Flucloxacillin is a PXR agonist at pharmacologically relevant concentrations, and a functionally significant upstream PXR polymorphism is a risk factor for flucloxacillin-induced DILI.

The incidence of sexually dimorphic gene expression varies greatly between tissues in the rat.

The sexually dimorphic expression of genes across 26 somatic rat tissues was using Affymetrix RAE-230 genechips. We considered probesets to be sexually dimorphically expressed (SDE) if they were measurably expressed above background in at least one sex, there was at least a two-fold difference in expression (dimorphism) between the sexes, and the differences were statistically significant after correcting for false discovery. 14.5% of expressed probesets were SDE in at least one tissue, with higher expression nearly twice as prevalent in males compared to females. Most were SDE in a single tissue. Surprisingly, nearly half of the probesets that were (SDE) in multiple tissues were oppositely sex biased in different tissues, and most SDE probesets were also expressed without sex bias in other tissues. Two genes were widely SDE: Xist (female-only) and Eif2s3y (male-only). The frequency of SDE probesets varied widely between tissues, and was highest in the duodenum (6.2%), whilst less than 0.05% in over half of the surveyed tissues. The occurrence of SDE probesets was not strongly correlated between tissues. Within individual tissues, however, relational networks of SDE genes were identified. In the liver, networks relating to differential metabolism between the sexes were seen. The estrogen receptor was implicated in differential gene expression in the duodenum. To conclude, sexually dimorphic gene expression is common, but highly tissue-dependent. Sexually dimorphic gene expression may provide insights into mechanisms underlying phenotypic sex differences. Online data are provided as a resource for further analyses (GEO reference GSE63362).

Ticagrelor inhibits adenosine uptake in vitro and enhances adenosine-mediated hyperemia responses in a canine model.

AIMS: A routine secondary pharmacology screen indicated that reversibly binding oral P2Y(12) receptor antagonist ticagrelor could inhibit adenosine uptake in human erythrocytes, suggesting that ticagrelor may potentiate adenosine-mediated responses in vivo. The aim of this study was to further characterize the adenosine uptake inhibition in vitro and study possible physiological consequences of adenosine uptake inhibition by ticagrelor in an anesthetized dog model of coronary blood flow compared to dipyridamole. METHODS AND RESULTS: We measured [2-3H]adenosine uptake in purified human erythrocytes and several cell lines in the presence of ticagrelor or the known uptake inhibitor dipyridamole as a comparator. Using an open-chest dog model (beagles), we measured the left anterior descending (LAD) coronary artery blood flow during reactive hyperemia after 1 minute occlusion or intracoronary infusion of adenosine before and after administration of vehicle, ticagrelor, or dipyridamole (each n = 8). Ticagrelor concentration-dependently inhibited adenosine uptake in human erythrocytes and in cell lines of rat, canine, or human origin. In the dog model, ticagrelor and dipyridamole dose-dependently augmented reactive hyperemia after LAD occlusion, as assessed by percentage repayment of flow debt relative to control (both Ps < .05). Ticagrelor and dipyridamole also dose-dependently augmented intracoronary adenosine-induced increases in LAD blood flow relative to control (both Ps < .05). CONCLUSION: Ticagrelor inhibits adenosine uptake in vitro and subsequently augments cardiac blood flow in a canine model of reactive hypoxia- or adenosine-induced blood flow increases. These findings suggest that ticagrelor may have additional benefits in patients with acute coronary syndrome beyond inhibition of platelet aggregation.

Generation and analysis of transcriptomics data.

Transcript profiling ("Transcriptomics") is a widely used technique that obtains information on the abundance of multiple mRNA transcripts within a biological sample simultaneously. Therefore, when a number of such samples are analysed, as in a scientific experiment, large and complex data sets are gene-rated. Here, we describe the use of one method commonly used to generate transcriptomics data, namely the use of Affymetrix GeneChip microarrays. Data generated in transcriptomics experiments can be analysed using a multitude of approaches, but a common goal is to identify those transcripts whose abundance is altered by the experimental conditions, or which differ between sets of samples. Here, we describe a simple approach, the calculation of the volcano score, which identifies transcripts with altered abundance, taking into account both the magnitude of the alteration and its statistical significance.