Characterisation of the NRF2 transcriptional network and its response to chemical insult in primary human hepatocytes: implications for prediction of drug-induced liver injury

The transcription factor NRF2, governed by its repressor KEAP1, protects cells against oxidative stress. There is interest in modelling the NRF2 response to improve the prediction of clinical toxicities such as drug-induced liver injury (DILI). However, very little is known about the makeup of the NRF2 transcriptional network and its response to chemical perturbation in primary human hepatocytes (PHH), which are often used as a translational model for investigating DILI. Here, microarray analysis identified 108 transcripts (including several putative novel NRF2-regulated genes) that were both downregulated by siRNA targeting NRF2 and upregulated by siRNA targeting KEAP1 in PHH. Applying weighted gene co-expression network analysis (WGCNA) to transcriptomic data from the Open TG-GATES toxicogenomics repository (representing PHH exposed to 158 compounds) revealed four co-expressed gene sets or 'modules' enriched for these and other NRF2-associated genes. By classifying the 158 TG-GATES compounds based on published evidence, and employing the four modules as network perturbation metrics, we found that the activation of NRF2 is a very good indicator of the intrinsic biochemical reactivity of a compound (i.e. its propensity to cause direct chemical stress), with relatively high sensitivity, specificity, accuracy and positive/negative predictive values. We also found that NRF2 activation has lower sensitivity for the prediction of clinical DILI risk, although relatively high specificity and positive predictive values indicate that false positive detection rates are likely to be low in this setting. Underpinned by our comprehensive analysis, activation of the NRF2 network is one of several mechanism-based components that can be incorporated into holistic systems toxicology models to improve mechanistic understanding and preclinical prediction of DILI in man.Medicinal Chemistr

Rise and fall of sleep quantity and quality with student experiences across the first year of university

Covariations of self-reported sleep quantity (duration) and quality (disturbances) with affective, stressful, academic, and social experiences across the first year of university in 187 Canadian students (M age=18.4) were examined with multilevel models. Female students reported sleeping fewer hours on average than did male students. In months when negative affect and general levels of stress were higher, sleep quantity was lower. Poorer sleep quality was seen in students living away from home and reporting more financial stress at baseline. In addition, sleep quality was poorer in months when negative affect and general levels of stress were higher (attenuating the effect of financial stress) and better in months when students spent more days with friends. Three themes are presented to explore the mechanisms by which sleep quantity and quality rise and fall in tandem with experiences of the first year of university

Losing sleep over it: Daily variation in sleep quantity and quality in canadian students' first seme

Daily covariation of sleep quantity and quality with affective, stressful, academic, and social experiences were observed in a sample of Canadian 17-19-year-olds in their first year of university. Participants (N=191) completed web-based checklists for 14 consecutive days during their first semester. Multilevel models predicting sleep quantity and quality from daily experiences indicated that more time on schoolwork, expecting a test, and alcohol use predicted less sleep whereas socializing predicted more sleep. More positive affect and no alcohol use predicted better sleep quality. Models predicting daily experiences from sleep the night before indicated that less sleep preceded increases in negative affect, decreases in schoolwork time, and a higher likelihood of socializing. Better sleep quality preceded increased positive affect, decreased negative affect and stress, and less time on schoolwork. These data are informative for understanding relations between sleep and daily experiences as they occur naturally in first-year university students

College Student Affect and Heavy Drinking: Variable Associations Across Days, Semesters, and People

This study tested associations between positive and negative affect and heavy drinking in 734 college students who completed daily diaries in 14-day bursts once per semester over 7 semesters (98 days per person). Three-level multilevel models tested whether affect and heavy drinking were linked across days, semesters, and persons. Higher daily, between-semester, and between-person positive affect were each associated with greater odds of heavy drinking on weekdays and on weekend days. A significant interaction with semester in college showed that the association between daily positive affect and heavy drinking on weekend days became stronger over time. That is, heavy drinking on a weekend day with higher positive affect was more likely in later years of college (OR = 2.93, Fall of 4th year), compared to earlier in college (OR = 1.80, Fall of 1st year). A similar interaction was found for between-semester positive affect and heavy drinking on weekdays. Higher daily negative affect was associated with a greater odds of heavy drinking on weekdays only for students who first began drinking in 7th grade or earlier (OR = 2.36). Results of this study highlight the importance of varied time spans in studying the etiology, consequences, and prevention of heavy drinking. Harm-reduction strategies that target positive affect-related drinking by encouraging protective behaviors during celebratory events may become increasingly important as students transition to later years of college

Formation and protein binding of the acyl glucuronide of a leukotriene b4 antagonist (sb-209247) relation to species differences in hepatotoxicity

SB-209247 [(E)-3-[6-[[(2,6-dichlorophenyl)-thio]methyl]-3-(2-phenylethoxy)-2-pyridinyl]-2-propenoic acid], an anti-inflammatory leukotriene B4 receptor antagonist, was associated in beagle dogs but not male rats with an inflammatory hepatopathy. It also produced a concentration-dependent (10-1000 μM) but equal leakage of enzymes from dog and rat precision-cut liver slices. The hepatic metabolism of SB-209247 was investigated with reference to the formation of reactive acyl glucuronides. [14C]SB-209247 (100 μmol/kg) administered i.v. to anesthetized male rats was eliminated by biliary excretion of the acyl glucuronides of the drug and its sulfoxide. After 5 h, 1.03 ± 0.14% (mean ± S.E.M., n = 4) of the dose was bound irreversibly to liver tissue. The sulfoxide glucuronide underwent pH-dependent rearrangement in bile more rapidly than did the SB-209247 conjugate. [14C]SB-209247 was metabolized by sulfoxidation and glucuronidation in rat and dog hepatocytes, and approximately 1 to 2% of [14C]SB-209247 (100 μM) became irreversibly bound to cellular material. [14C]SB-209247 sulfoxide and glucuronide were the only metabolites produced by dog, rat, and human liver microsomes in the presence of NADPH and UDP-glucuronic acid (UDPGA), respectively. Vmax values for [14C]SB-209247 glucuronidation by dog, rat, and human microsomes were 2.6 ± 0.1, 1.2 ± 0.1, and 0.4 ± 0.0 nmol/min/mg protein, respectively. Hepatic microsomes from all three species catalyzed UDPGA-dependent but not NADPH-dependent irreversible binding of [14C]SB-209247 (100-250 μM) to microsomal protein. Although a reactive acyl glucuronide was formed by microsomes from every species, the binding did not differ between species. Therefore, neither the acute cellular injury nor glucuronidation-driven irreversible protein binding in vitro is predictive of the drug-induced hepatopathy

Who Sleeps Best? Longitudinal Patterns and Covariates of Change in Sleep Quantity, Quality, and Timing Across Four University Years

This study tracked change over time in sleep quantity, disturbance, and timing, and sleep's covariations with living situation, stress, social support, alcohol use, and grade point average (GPA) across four years of university in 186 Canadian students. Women slept longer as they moved through university, and men slept less; rise times were later each year. Students reported sleeping fewer hours, more sleep disturbances, and later rise times during years with higher stress. In years when students lived away from home, they reported more sleep disturbances, later bedtimes, and later rise times. Living on campus was associated with later bedtimes and rise times. Alcohol use was higher and GPA was lower when bedtimes were later. The implications of these observed patterns for understanding the correlates and consequences of university students' sleep are discussed