Drug-induced chromatin accessibility changes associate with sensitivity to liver tumor promotion

Liver cancer susceptibility varies amongst humans and between experimental animal models due to multiple genetic and epigenetic factors. The molecular characterization of such susceptibilities has the potential to enhance cancer risk assessment of xenobiotic exposures and disease prevention strategies. Here, using DNase I hypersensitivity mapping coupled with transcriptomic profiling, we investigate perturbations in cis-acting gene regulatory elements associated with the early stages of phenobarbital (PB)- mediated liver tumor promotion in susceptible versus resistant mouse strains (B6C3F1 versus C57BL/6J). Integrated computational analyses of strain-selective changes in liver chromatin accessibility underlying PB-response reveal differential epigenetic regulation of molecular pathways associated with PB-mediated tumor promotion, including Wnt/-catenin signalling. Complementary transcription factor motif analyses reveal mouse strain-selective gene regulatory networks and a novel role for Stat, Smad and Fox transcription factors in the early stages of PB-mediated tumor promotion. Mapping perturbations in cis-acting gene regulatory elements provides novel insights into the molecular basis for susceptibility to xenobiotic-induced rodent liver tumor promotion and has the potential to enhance mechanism-based cancer risk assessments of xenobiotic exposures

Usefulness of Parasternal Intercostal Muscle Ultrasound during Weaning from Mechanical Ventilation

International audienceAbstract Background The assessment of diaphragm function with diaphragm ultrasound seems to bring important clinical information to describe diaphragm work and weakness. When the diaphragm is weak, extradiaphragmatic muscles may play an important role, but whether ultrasound can also assess their activity and function is unknown. This study aimed to (1) evaluate the feasibility of measuring the thickening of the parasternal intercostal and investigate the responsiveness of this muscle to assisted ventilation; and (2) evaluate whether a combined evaluation of the parasternal and the diaphragm could predict failure of a spontaneous breathing trial. Methods First, an exploratory evaluation of the parasternal in 23 healthy subjects. Second, the responsiveness of parasternal to several pressure support levels were studied in 16 patients. Last, parasternal activity was compared in presence or absence of diaphragm dysfunction (assessed by magnetic stimulation of the phrenic nerves and ultrasound) and in case of success/failure of a spontaneous breathing trial in 54 patients. Results The parasternal was easily accessible in all patients. The interobserver reproducibility was good (intraclass correlation coefficient, 0.77 (95% CI, 0.53 to 0.89). There was a progressive decrease in parasternal muscle thickening fraction with increasing levels of pressure support (Spearman ρ = −0.61 [95% CI, −0.74 to −0.44]; P < 0.0001) and an inverse correlation between parasternal muscle thickening fraction and the pressure generating capacity of the diaphragm (Spearman ρ = −0.79 [95% CI, −0.87 to −0.66]; P < 0.0001). The parasternal muscle thickening fraction was higher in patients with diaphragm dysfunction: 17% (10 to 25) versus 5% (3 to 8), P < 0.0001. The pressure generating capacity of the diaphragm, the diaphragm thickening fraction and the parasternal thickening fraction similarly predicted failure or the spontaneous breathing trial. Conclusions Ultrasound assessment of the parasternal intercostal muscle is feasible in the intensive care unit and provides novel information regarding the respiratory capacity load balance. Editor’s Perspective What We Already Know about This Topic What This Article Tells Us That Is Ne

Phenobarbital mediates an epigenetic switch at the constitutive androstane receptor (CAR) target gene Cyp2b10 in the liver of B6C3F1 mice.

Evidence suggests that epigenetic perturbations are involved in the adverse effects associated with some drugs and toxicants, including certain classes of non-genotoxic carcinogens. Such epigenetic changes (altered DNA methylation and covalent histone modifications) may take place at the earliest stages of carcinogenesis and their identification holds great promise for biomedical research. Here, we evaluate the sensitivity and specificity of genome-wide epigenomic and transcriptomic profiling in phenobarbital (PB)-treated B6C3F1 mice, a well-characterized rodent model of non-genotoxic liver carcinogenesis. Methylated DNA Immunoprecipitation (MeDIP)-coupled microarray profiling of 17,967 promoter regions and 4,566 intergenic CpG islands was combined with genome-wide mRNA expression profiling to identify liver tissue-specific PB-mediated DNA methylation and transcriptional alterations. Only a limited number of significant anti-correlations were observed between PB-induced transcriptional and promoter-based DNA methylation perturbations. However, the constitutive androstane receptor (CAR) target gene Cyp2b10 was found to be concomitantly hypomethylated and transcriptionally activated in a liver tissue-specific manner following PB treatment. Furthermore, analysis of active and repressive histone modifications using chromatin immunoprecipitation revealed a strong PB-mediated epigenetic switch at the Cyp2b10 promoter. Our data reveal that PB-induced transcriptional perturbations are not generally associated with broad changes in the DNA methylation status at proximal promoters and suggest that the drug-inducible CAR pathway regulates an epigenetic switch from repressive to active chromatin at the target gene Cyp2b10. This study demonstrates the utility of integrated epigenomic and transcriptomic profiling for elucidating early mechanisms and biomarkers of non-genotoxic carcinogenesis

Pharmacological BACE1 and BACE2 inhibition induces hair hypopigmentation by inhibiting PMEL17 processing in mice

Melanocytes of the hair follicle produce melanin and are essential in determining the differences in hair color. Pigment cell-specific MELanocyte Protein (PMEL17) plays a crucial role in melanogenesis. One of the critical steps is the amyloid-like functional oligomerization of PMEL17. Beta Site APP Cleaving Enzyme-2 (BACE2) and γ-secretase have been shown to be key players in generating the proteolytic fragments of PMEL17. The β-secretase (BACE1) is responsible for the generation of amyloid-β (Aβ) fragments in the brain and is therefore proposed as a therapeutic target for Alzheimer’s disease (AD). Currently BACE1 inhibitors, most of which lack selectivity over BACE2, have demonstrated efficacious reduction of amyloid-β peptides in animals and the CSF of humans. BACE2 knock-out mice have a deficiency in PMEL17 proteolytic processing leading to impaired melanin storage and hair depigmentation. Here, we confirm BACE2-mediated inhibition of PMEL17 proteolytic processing in vitro in mouse and human melanocytes. Furthermore, we show that wildtype as well as bace2+/− and bace2−/− mice treated with a potent dual BACE1/BACE2 inhibitor NB-360 display dose-dependent appearance of irreversibly depigmented hair. Retinal pigmented epithelium showed no morphological changes. Our data demonstrates that BACE2 as well as additional BACE1 inhibition affects melanosome maturation and induces hair depigmentation in mice

High-flow oxygen through nasal cannula in acute hypoxemic respiratory failure

BACKGROUND: Whether noninvasive ventilation should be administered in patients with acute hypoxemic respiratory failure is debated. Therapy with high-flow oxygen through a nasal cannula may offer an alternative in patients with hypoxemia. METHODS: We performed a multicenter, open-label trial in which we randomly assigned patients without hypercapnia who had acute hypoxemic respiratory failure and a ratio of the partial pressure of arterial oxygen to the fraction of inspired oxygen of 300 mm Hg or less to high-flow oxygen therapy, standard oxygen therapy delivered through a face mask, or noninvasive positive-pressure ventilation. The primary outcome was the proportion of patients intubated at day 28; secondary outcomes included all-cause mortality in the intensive care unit and at 90 days and the number of ventilator-free days at day 28. RESULTS: A total of 310 patients were included in the analyses. The intubation rate (primary outcome) was 38% (40 of 106 patients) in the high-flow-oxygen group, 47% (44 of 94) in the standard group, and 50% (55 of 110) in the noninvasive-ventilation group (P = 0.18 for all comparisons). The number of ventilator-free days at day 28 was significantly higher in the high-flow-oxygen group (24±8 days, vs. 22±10 in the standard-oxygen group and 19±12 in the noninvasive-ventilation group; P = 0.02 for all comparisons). The hazard ratio for death at 90 days was 2.01 (95% confidence interval [CI], 1.01 to 3.99) with standard oxygen versus high-flow oxygen (P = 0.046) and 2.50 (95% CI, 1.31 to 4.78) with noninvasive ventilation versus high-flow oxygen (P = 0.006). CONCLUSIONS: In patients with nonhypercapnic acute hypoxemic respiratory failure, treatment with high-flow oxygen, standard oxygen, or noninvasive ventilation did not result in significantly different intubation rates. There was a significant difference in favor of high-flow oxygen in 90-day mortality. (Funded by the Programme Hospitalier de Recherche Clinique Interrégional 2010 of the French Ministry of Health; FLORALI ClinicalTrials.gov number, NCT01320384.

High-flow oxygen through nasal cannula in acute hypoxemic respiratory failure

International audienceBACKGROUND: Whether noninvasive ventilation should be administered in patients with acute hypoxemic respiratory failure is debated. Therapy with high-flow oxygen through a nasal cannula may offer an alternative in patients with hypoxemia. METHODS: We performed a multicenter, open-label trial in which we randomly assigned patients without hypercapnia who had acute hypoxemic respiratory failure and a ratio of the partial pressure of arterial oxygen to the fraction of inspired oxygen of 300 mm Hg or less to high-flow oxygen therapy, standard oxygen therapy delivered through a face mask, or noninvasive positive-pressure ventilation. The primary outcome was the proportion of patients intubated at day 28; secondary outcomes included all-cause mortality in the intensive care unit and at 90 days and the number of ventilator-free days at day 28. RESULTS: A total of 310 patients were included in the analyses. The intubation rate (primary outcome) was 38% (40 of 106 patients) in the high-flow-oxygen group, 47% (44 of 94) in the standard group, and 50% (55 of 110) in the noninvasive-ventilation group (P=0.18 for all comparisons). The number of ventilator-free days at day 28 was significantly higher in the high-flow-oxygen group (24+/-8 days, vs. 22+/-10 in the standard-oxygen group and 19+/-12 in the noninvasive-ventilation group; P=0.02 for all comparisons). The hazard ratio for death at 90 days was 2.01 (95% confidence interval [CI], 1.01 to 3.99) with standard oxygen versus high-flow oxygen (P=0.046) and 2.50 (95% CI, 1.31 to 4.78) with noninvasive ventilation versus high-flow oxygen (P=0.006). CONCLUSIONS: In patients with nonhypercapnic acute hypoxemic respiratory failure, treatment with high-flow oxygen, standard oxygen, or noninvasive ventilation did not result in significantly different intubation rates. There was a significant difference in favor of high-flow oxygen in 90-day mortality. (Funded by the Programme Hospitalier de Recherche Clinique Interregional 2010 of the French Ministry of Health; FLORALI ClinicalTrials.gov number, NCT01320384.)

Identification of Dlk1-Dio3 Imprinted Gene Cluster Noncoding RNAs as Novel Candidate Biomarkers for Liver Tumor Promotion

The molecular events during non-genotoxic carcinogenesis and their temporal order are poorly understood but thought to include long-lasting perturbations of gene expression. Here, we have investigated the temporal sequence of molecular and pathological perturbations at early stages of phenobarbital-mediated liver tumor promotion in vivo. Molecular profiling (mRNA, miRNA, DNA methylation & proteins) of mouse liver during 13 weeks of phenobarbital treatment revealed progressive increases in hepatic expression of long non-coding RNAs and microRNAs originating from the Dlk1-Dio3 imprinted gene cluster, a locus that has recently been associated with stem cell pluripotency in mice and various neoplasms in humans. Phenobarbital-induction of the Dlk1-Dio3 cluster non-coding RNA Meg3 was localised to glutamine synthetase positive hypertrophic perivenous hepatocytes suggesting a role for β-catenin signaling in the dysregulation of Dlk1-Dio3 non-coding RNAs. The carcinogenic relevance of Dlk1-Dio3 locus non-coding RNA induction was further supported by in vivo genetic dependence on Constitutive Androstane Receptor (CAR) and β-catenin pathways. Our data identify Dlk1-Dio3 non-coding RNAs as novel candidate early biomarkers for mouse liver tumor promotion and provide new opportunities for assessing the carcinogenic potential of novel compounds

Identification of Dlk1-Dio3 imprinted gene cluster non-coding RNAs as novel candidate biomarkers for liver tumor promotion

The molecular events during non-genotoxic carcinogenesis and their temporal order are poorly understood but thought to include long-lasting perturbations of gene expression. Here, we have investigated the temporal sequence of molecular and pathological perturbations at early stages of phenobarbital-mediated liver tumor promotion in vivo. Molecular profiling (mRNA, miRNA, DNA methylation & proteins) of mouse liver during 13 weeks of phenobarbital treatment revealed progressive increases in hepatic expression of long non-coding RNAs and microRNAs originating from the Dlk1-Dio3 imprinted gene cluster, a locus that has recently been associated with stem cell pluripotency in mice and various neoplasms in humans. Phenobarbital-induction of the Dlk1-Dio3 cluster non-coding RNA Meg3 was localised to glutamine synthetase positive hypertrophic perivenous hepatocytes suggesting a role for β-catenin signaling in the dysregulation of Dlk1-Dio3 non-coding RNAs. The carcinogenic relevance of Dlk1-Dio3 locus non-coding RNA induction was further supported by in vivo genetic dependence on Constitutive Androstane Receptor (CAR) and β-catenin pathways. Our data identify Dlk1-Dio3 non-coding RNAs as novel candidate early biomarkers for mouse liver tumor promotion and provide new opportunities for assessing the carcinogenic potential of novel compounds