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The aim of this study was the assessment of the Regional Myocardial Work (RMW) by measuring the stress-strain relationships in the left myocardium, under basic and ischemic conditions. In 11 swines, 4 Sonometrics crystals were placed in the region perfused by the left anterior descending artery (LAD) circumscribing a tetrahedral myocardial volume (TMV). The 1st crystal was situated 45mm below the pulmonary artery, beside the LAD. Under control of the acquisition device, the 2nd was placed 10mm below the 1st following the LAD, the 3rd 10mm at the left side of the 1st and the 4th in equidistant point between the 1st and the 2nd, 5mm deep within the myocardium. A Millar, omnidirectional pressure catheter was placed within the TMV. The 6 distances and intramyocardial pressure measured were digitized and mathematically synthesized. The variation in TMV strain (µv) and stress (µp) was calculated throughout the cardiac cycle, resulting in 2 equations. The relationships between µp and µv result in a stress-strain loop, from which RMW was calculated. Simultaneously, Pulsed Doppler Tissue Imaging (DTI) assessed systolic (VS), early-diastolic (VE) and latediastolic (VA) myocardial velocities. Measurements were performed under basic (B), during ischemia by occlusion LDA for 40 seconds, after 1 and 15 minutes of reperfusion (Occlusion (O), Reperfusion1 (R1) and 2 (R2) periods). The average of RMW, under basic conditions, wa

PPARα transcriptionally induces AhR expression in Caco-2, but represses AhR pro-inflammatory effects

International audienceIn this work we demonstrate that Caco-2 cell treatment with WY-14643 (a potent PPARa agonist) causes an increase in AhR expression. Luciferase assays and directed mutagenesis experiments showed that induction mainly occurred at transcriptional level and involved a PPRE site located within the AhR promoter. These results were further confirmed by the use of PPARa knockout mice in which AhR induction by WY14643 was abrogated. In addition to CYP1 regulation, AhR has been described as being involved in inflammation , so we also studied the effect of AhR regulation by PPARa on the expression of some inflammation target genes. 3-Methylcho-lanthrene (a potent AhR agonist) increased the expression (mRNA) of the major inflammatory targets IL-1b and MMP9. WY-14643 co-treatment abrogated the 3-methylcholanthrene pro-inflammatory effect. Hence the anti-inflammatory effect of PPARa overrides the pro-inflammatory effect of AhR

Continuous Quinacrine Treatment Results in the Formation of Drug-Resistant Prions

Quinacrine is a potent antiprion compound in cell culture models of prion disease but has failed to show efficacy in animal bioassays and human clinical trials. Previous studies demonstrated that quinacrine inefficiently penetrates the blood-brain barrier (BBB), which could contribute to its lack of efficacy in vivo. As quinacrine is known to be a substrate for P-glycoprotein multi-drug resistance (MDR) transporters, we circumvented its poor BBB permeability by utilizing MDR0/0 mice that are deficient in mdr1a and mdr1b genes. Mice treated with 40 mg/kg/day of quinacrine accumulated up to 100 µM of quinacrine in their brains without acute toxicity. PrPSc levels in the brains of prion-inoculated MDR0/0 mice diminished upon the initiation of quinacrine treatment. However, this reduction was transient and PrPSc levels recovered despite the continuous administration of quinacrine. Treatment with quinacrine did not prolong the survival times of prion-inoculated, wild-type or MDR0/0 mice compared to untreated mice. A similar phenomenon was observed in cultured differentiated prion-infected neuroblastoma cells: PrPSc levels initially decreased after quinacrine treatment then rapidly recovered after 3 d of continuous treatment. Biochemical characterization of PrPSc that persisted in the brains of quinacrine-treated mice had a lower conformational stability and different immunoaffinities compared to that found in the brains of untreated controls. These physical properties were not maintained upon passage in MDR0/0 mice. From these data, we propose that quinacrine eliminates a specific subset of PrPSc conformers, resulting in the survival of drug-resistant prion conformations. Transient accumulation of this drug-resistant prion population provides a possible explanation for the lack of in vivo efficacy of quinacrine and other antiprion drugs

Comparative Analysis of Gene Regulation by the Transcription Factor PPARα between Mouse and Human

Studies in mice have shown that PPARalpha is an important regulator of hepatic lipid metabolism and the acute phase response. However, little information is available on the role of PPARalpha in human liver. Here we set out to compare the function of PPARalpha in mouse and human hepatocytes via analysis of target gene regulation

ATP-binding cassette (ABC) transporters in normal and pathological lung

ATP-binding cassette (ABC) transporters are a family of transmembrane proteins that can transport a wide variety of substrates across biological membranes in an energy-dependent manner. Many ABC transporters such as P-glycoprotein (P-gp), multidrug resistance-associated protein 1 (MRP1) and breast cancer resistance protein (BCRP) are highly expressed in bronchial epithelium. This review aims to give new insights in the possible functions of ABC molecules in the lung in view of their expression in different cell types. Furthermore, their role in protection against noxious compounds, e.g. air pollutants and cigarette smoke components, will be discussed as well as the (mal)function in normal and pathological lung. Several pulmonary drugs are substrates for ABC transporters and therefore, the delivery of these drugs to the site of action may be highly dependent on the presence and activity of many ABC transporters in several cell types. Three ABC transporters are known to play an important role in lung functioning. Mutations in the cystic fibrosis transmembrane conductance regulator (CFTR) gene can cause cystic fibrosis, and mutations in ABCA1 and ABCA3 are responsible for respectively Tangier disease and fatal surfactant deficiency. The role of altered function of ABC transporters in highly prevalent pulmonary diseases such as asthma or chronic obstructive pulmonary disease (COPD) have hardly been investigated so far. We especially focused on polymorphisms, knock-out mice models and in vitro results of pulmonary research. Insight in the function of ABC transporters in the lung may open new ways to facilitate treatment of lung diseases

Activation of the NFκB Pathway Enhances AhR Expression in Intestinal Caco-2 Cells

International audienceRecent data suggest that apart from its well-known role in the regulation of xenobiotic metabolizing enzymes, AhR is also involved in inflammation. However, the influence of inflammation on AhR expression remains unknown. Here, we demonstrated that proinflammatory conditions induced by either PMA or IL-1 beta enhance AhR expression in Caco-2 cells. This was associated with an increase in AhR promoter activity. By means of directed mutagenesis experiments and the use of proteasome inhibitors, we demonstrated that inflammation-induced AhR expression involved the NF kappa B pathway but not AP-1. Moreover, conditioned media from PMA-treated Caco-2 cells were also able to induce AhR expression, and this induction was repressed by anti-IL-1 beta blocking antibodies. Similar results were obtained with conditioned media from PMA-treated THP-1 cells. Taken together, these data suggest that AhR could be involved in vivo in an inflammatory loop. AhR was recently suspected to be implicated in inflammatory bowel disease. Our results support this hypothesis and suggest that AhR could be a new target for inflammatory bowel disease patient management

Evidence for a new human CYP1A1 regulation pathway involving PPAR-α and 2 PPRE sites

International audienceBackground & Aims: Cytochrome P450 1A1 catalyzes the degradation of endobiotics (estradiol, fatty acids, and so on) and the bioactivation of numerous environmental procarcinogens, such as arylamines and polycyclic aromatic hydrocarbons, that are found in food. Several peroxisome proliferators and arachidonic acid derivatives enhance cytochrome P450 1A1 activity, but the mechanisms involved remain unknown. The aim of this work was to study the role of peroxisome proliferator—activated receptors in cytochrome P450 1A1 gene induction.Methods: The role of peroxisome proliferator—activated receptor transcription factors in cytochrome P450 1A1 induction was assessed by means of enzymatic activities, quantitative real-time polymerase chain reaction, gene reporter assays, mutagenesis, and electrophoretic mobility shift assay.Results: We show that peroxisome proliferator—activated receptor-α agonists (WY-14643, bezafibrate, clofibrate, and phthalate) induce human cytochrome P450 1A1 gene expression, whereas 2,4-thiazolidinedione, a specific peroxisome proliferator—activated receptor-γ agonist, represses it. The induction of cytochrome P450 1A1 transcripts by WY-14643 was associated with a marked increase of ethoxyresorufin O-deethylase activity (10-fold at 200 μmol/L). Transfection of peroxisome proliferator—activated receptor-α complementary DNA enhanced cytochrome P450 1A1 messenger RNA induction by WY-14643, although WY-14643 failed to activate xenobiotic responsive element sequences. Two peroxisome proliferator response element sites were located at positions −931/−919 and −531/−519 of the cytochrome P450 1A1 promoter. Their inactivation by directed mutagenesis suppressed the inductive effect of WY-14643 on cytochrome P450 1A1 promoter activation. Electrophoretic mobility shift assay and chromatin immunoprecipitation assay experiments showed that the 2 cytochrome P450 1A1 peroxisome proliferator response element sites bind the peroxisome proliferator—activated receptor-α/retinoid X receptor-α heterodimer.Conclusions: We describe here a new cytochrome P450 1A1 induction pathway involving peroxisome proliferator—activated receptor-α and 2 peroxisome proliferator response element sites, indicating that peroxisome proliferator—activated receptor-α ligands, which are common environmental compounds, may be involved in carcinogenesis