Mendelian randomization analysis using GWAS and eQTL data to investigate the relationship between chronotype and neuropsychiatric disorders and their molecular basis.

Chronotype is a proxy sleep measure that has been associated with neuropsychiatric disorders. By investigating how chronotype influences risk for neuropsychiatric disorders and vice versa, we may identify modifiable risk factors for each phenotype. Here we used Mendelian randomization (MR), to explore causal effects by (1) studying the causal relationships between neuropsychiatric disorders and chronotype and (2) characterizing the genetic components of these phenotypes. Firstly, we investigated if a causal role exists between five neuropsychiatric disorders and chronotype using the largest genome-wide association studies (GWAS) available. Secondly, we integrated data from expression quantitative trait loci (eQTLs) to investigate the role of gene expression alterations on these phenotypes. Evening chronotype was causal for increased risk of schizophrenia and autism spectrum disorder and schizophrenia was causal for a tendency toward evening chronotype. We identified 12 eQTLs where gene expression changes in brain or blood were causal for one of the phenotypes, including two eQTLs for SNX19 in hippocampus and hypothalamus that were causal for schizophrenia. These findings provide important evidence for the complex, bidirectional relationship that exists between a sleep-based phenotype and neuropsychiatric disorders, and use gene expression data to identify causal roles for genes at associated loci.

Circadian Variation in the Response to Vaccination: A Systematic Review and Evidence Appraisal.

Molecular timing mechanisms known as circadian clocks drive endogenous 24-h rhythmicity in most physiological functions, including innate and adaptive immunity. Consequently, the response to immune challenge such as vaccination might depend on the time of day of exposure. This study assessed whether the time of day of vaccination (TODV) is associated with the subsequent immune and clinical response by conducting a systematic review of previous studies. The Cochrane Library, PubMed, Google, Medline, and Embase were searched for studies that reported TODV and immune and clinical outcomes, yielding 3114 studies, 23 of which met the inclusion criteria. The global severe acute respiratory syndrome coronavirus 2 vaccination program facilitated investigation of TODV and almost half of the studies included reported data collected during the COVID-19 pandemic. There was considerable heterogeneity in the demography of participants and type of vaccine, and most studies were biased by failure to account for immune status prior to vaccination, self-selection of vaccination time, or confounding factors such as sleep, chronotype, and shiftwork. The optimum TODV was concluded to be afternoon (5 studies), morning (5 studies), morning and afternoon (1 study), midday (1 study), and morning or late afternoon (1 study), with the remaining 10 studies reporting no effect. Further research is required to understand the relationship between TODV and subsequent immune outcome and whether any clinical benefit outweighs the potential effect of this intervention on vaccine uptake.

Susceptibility to the common cold virus is associated with day length.

Seasonal rhythms are endogenous timing mechanisms that allow animals living at temperate latitudes to synchronize their physiology to the seasons. Human viral respiratory disease is prevalent in the winter at temperate latitudes, but the role of endogenous mechanisms in these recurring annual patterns is unclear. The Common Cold Project is a repository of data describing the experimental viral challenge of 1,337 participants across the seasons of the year. We report a secondary analysis of these data to investigate if susceptibility to the common cold is associated with day length. The majority of the participants (78%) showed signs of infection but only 32% developed clinical signs of disease, and the probability of infection was significantly higher in longer day lengths (summer), but the disease was more likely in short (winter) day lengths. The persistence of winter disease patterns in experimental conditions supports the role of endogenous seasonality in human susceptibility to viral infection.

Circadian clock protein BMAL1 regulates IL-1ß in macrophages via NRF2.

A variety of innate immune responses and functions are dependent on time of day, and many inflammatory conditions are associated with dysfunctional molecular clocks within immune cells. However, the functional importance of these innate immune clocks has yet to be fully characterized. NRF2 plays a critical role in the innate immune system, limiting inflammation via reactive oxygen species (ROS) suppression and direct repression of the proinflammatory cytokines, IL-1ß and IL-6. Here we reveal that the core molecular clock protein, BMAL1, controls the mRNA expression of Nrf2 via direct E-box binding to its promoter to regulate its activity. Deletion of Bmal1 decreased the response of NRF2 to LPS challenge, resulting in a blunted antioxidant response and reduced synthesis of glutathione. ROS accumulation was increased in Bmal1-/- macrophages, facilitating accumulation of the hypoxic response protein, HIF-1α. Increased ROS and HIF-1α levels, as well as decreased activity of NRF2 in cells lacking BMAL1, resulted in increased production of the proinflammatory cytokine, IL-1ß. The excessive prooxidant and proinflammatory phenotype of Bmal1-/- macrophages was rescued by genetic and pharmacological activation of NRF2, or through addition of antioxidants. Our findings uncover a clear role for the molecular clock in regulating NRF2 in innate immune cells to control the inflammatory response. These findings provide insights into the pathology of inflammatory conditions, in which the molecular clock, oxidative stress, and IL-1ß are known to play a role.

Genome-Wide Association Study of Circadian Rhythmicity in 71,500 UK Biobank Participants and Polygenic Association with Mood Instability.

BACKGROUND: Circadian rhythms are fundamental to health and are particularly important for mental wellbeing. Disrupted rhythms of rest and activity are recognised as risk factors for major depressive disorder and bipolar disorder. METHODS: We conducted a genome-wide association study (GWAS) of low relative amplitude (RA), an objective measure of rest-activity cycles derived from the accelerometer data of 71,500 UK Biobank participants. Polygenic risk scores (PRS) for low RA were used to investigate potential associations with psychiatric phenotypes. OUTCOMES: Two independent genetic loci were associated with low RA, within genomic regions for Neurofascin (NFASC) and Solute Carrier Family 25 Member 17 (SLC25A17). A secondary GWAS of RA as a continuous measure identified a locus within Meis Homeobox 1 (MEIS1). There were no significant genetic correlations between low RA and any of the psychiatric phenotypes assessed. However, PRS for low RA was significantly associated with mood instability across multiple PRS thresholds (at PRS threshold 0·05: OR = 1·02, 95% CI = 1·01-1·02, p = 9·6 × 10-5), and with major depressive disorder (at PRS threshold 0·1: OR = 1·03, 95% CI = 1·01-1·05, p = 0·025) and neuroticism (at PRS threshold 0·5: Beta = 0·02, 95% CI = 0·007-0·04, p = 0·021). INTERPRETATION: Overall, our findings contribute new knowledge on the complex genetic architecture of circadian rhythmicity and suggest a putative biological link between disrupted circadian function and mood disorder phenotypes, particularly mood instability, but also major depressive disorder and neuroticism. FUNDING: Medical Research Council (MR/K501335/1).

Association of disrupted circadian rhythmicity with mood disorders, subjective wellbeing, and cognitive function: a cross-sectional study of 91 105 participants from the UK Biobank.

BACKGROUND: Disruption of sleep and circadian rhythmicity is a core feature of mood disorders and might be associated with increased susceptibility to such disorders. Previous studies in this area have used subjective reports of activity and sleep patterns, but the availability of accelerometer-based data from UK Biobank participants permits the derivation and analysis of new, objectively ascertained circadian rhythmicity parameters. We examined associations between objectively assessed circadian rhythmicity and mental health and wellbeing phenotypes, including lifetime history of mood disorder. METHODS: UK residents aged 37-73 years were recruited into the UK Biobank general population cohort from 2006 to 2010. We used data from a subset of participants whose activity levels were recorded by wearing a wrist-worn accelerometer for 7 days. From these data, we derived a circadian relative amplitude variable, which is a measure of the extent to which circadian rhythmicity of rest-activity cycles is disrupted. In the same sample, we examined cross-sectional associations between low relative amplitude and mood disorder, wellbeing, and cognitive variables using a series of regression models. Our final model adjusted for age and season at the time that accelerometry started, sex, ethnic origin, Townsend deprivation score, smoking status, alcohol intake, educational attainment, overall mean acceleration recorded by accelerometry, body-mass index, and a binary measure of childhood trauma. FINDINGS: We included 91 105 participants with accelerometery data collected between 2013 and 2015 in our analyses. A one-quintile reduction in relative amplitude was associated with increased risk of lifetime major depressive disorder (odds ratio [OR] 1·06, 95% CI 1·04-1·08) and lifetime bipolar disorder (1·11, 1·03-1·20), as well as with greater mood instability (1·02, 1·01-1·04), higher neuroticism scores (incident rate ratio 1·01, 1·01-1·02), more subjective loneliness (OR 1·09, 1·07-1·11), lower happiness (0·91, 0·90-0·93), lower health satisfaction (0·90, 0·89-0·91), and slower reaction times (linear regression coefficient 1·75, 1·05-2·45). These associations were independent of demographic, lifestyle, education, and overall activity confounders. INTERPRETATION: Circadian disruption is reliably associated with various adverse mental health and wellbeing outcomes, including major depressive disorder and bipolar disorder. Lower relative amplitude might be linked to increased susceptibility to mood disorders. FUNDING: Lister Institute of Preventive Medicine.

Population-level seasonality in cardiovascular mortality, blood pressure, BMI and inflammatory cells in UK biobank.

INTRODUCTION: The risk of mortality from cardiovascular disease (CVD) is higher in wintertime throughout the world, but it is not known if this reflects annual changes in diet or lifestyle, or an endogenous photoperiodic mechanism that is sensitive to changes in day length. METHODS: Phenotypic data on cardiometabolic and lifestyle factors were collected throughout a 4 year time period from 502,642 middle-aged participants in UK Biobank. To assess the impact of seasonal environmental changes on cardiovascular risk factors, we linked these data to the outdoor temperature and day length at the time of assessment. Self-reported information on physical activity, diet and disease status were used to adjust for confounding factors related to health and lifestyle. RESULTS: Mortality related to CVD was higher in winter, as were risk factors for this condition including blood pressure, markers of inflammation and body mass index (BMI). These seasonal rhythms were significantly related to day length after adjustment for other factors that might affect seasonality including physical activity, diet and outdoor temperature. CONCLUSIONS: The risk of CVD may be modulated by day length at temperate latitudes, and the implications of seasonality should be considered in all studies of human cardiometabolic health. Key messages In this cross-sectional study in UK Biobank, we report annual variations in cardiovascular risk factors and mortality that were associated with day length independent of environmental and lifestyle factors. These seasonal changes in day length might contribute to annual patterns in cardiovascular disease and mortality.

Seasonality of depressive symptoms in women but not in men: A cross-sectional study in the UK Biobank cohort.

BACKGROUND: We examined whether seasonal variations in depressive symptoms occurred independently of demographic and lifestyle factors, and were related to change in day length and/or outdoor temperature. METHODS: In a cross-sectional analysis of >150,000 participants of the UK Biobank cohort, we used the cosinor method to assess evidence of seasonality of a total depressive symptoms score and of low mood, anhedonia, tenseness and tiredness scores in women and men. Associations of depressive symptoms with day length and mean outdoor temperature were then examined. RESULTS: Seasonality of total depressive symptom scores, anhedonia and tiredness scores was observed in women but not men, with peaks in winter. In women, increased day length was associated with reduced reporting of low mood and anhedonia, but with increased reporting of tiredness, independent of demographic and lifestyle factors. Associations with day length were not independent of the average outdoor temperature preceding assessment. LIMITATIONS: This was a cross-sectional investigation - longitudinal studies of within-subject seasonal variation in mood are necessary. Outcome measures relied on self-report and measured only a subset of depressive symptoms. CONCLUSION: This large, population-based study provides evidence of seasonal variation in depressive symptoms in women. Shorter days were associated with increased feelings of low mood and anhedonia in women. Clinicians should be aware of these population-level sex differences in seasonal mood variations in order to aid recognition and treatment of depression and subclinical depressive symptoms.

Adverse metabolic and mental health outcomes associated with shiftwork in a population-based study of 277,168 workers in UK biobank.

BACKGROUND: Reported associations between shiftwork and health have largely been based on occupation-specific, or single sex studies that might not be generalizable to the entire working population. The objective of this study was to investigate whether shiftwork was independently associated with obesity, diabetes, poor sleep, and well-being in a large, UK general population cohort. METHODS: Participants of the UK Biobank study who were employed at the time of assessment were included. Exposure variables were self-reported shiftwork (any shiftwork and night shiftwork); and outcomes were objectively measured obesity, inflammation and physical activity and self-reported lifestyle, sleep and well-being variables, including mental health. RESULTS: Shiftwork was reported by 17% of the 277,168 employed participants. Shiftworkers were more likely to be male, socioeconomically deprived and smokers, and to have higher levels of physical activity. Univariately, and following adjustment for lifestyle and work-related confounders, shiftworkers were more likely to be obese, depressed, to report disturbed sleep, and to have neurotic traits. CONCLUSIONS: Shiftwork was independently associated with multiple indicators of poor health and wellbeing, despite higher physical activity, and even in shiftworkers that did not work nights. Shiftwork is an emerging social factor that contributes to disease in the urban environment across the working population. Key messages Studies have linked shiftwork to obesity and diabetes in nurses and industry workers, but little is known about the implications of shiftwork for the general workforce In this large cross sectional study of UK workers, shiftwork was associated with obesity, depression and sleep disturbance, despite higher levels of physical activity. Shiftwork was associated with multiple indicators of compromised health and wellbeing and were more likely to report neurotic traits and evening preference.

Effects of Photoperiod Extension on Clock Gene and Neuropeptide RNA Expression in the SCN of the Soay Sheep.

In mammals, changing daylength (photoperiod) is the main synchronizer of seasonal functions. The photoperiodic information is transmitted through the retino-hypothalamic tract to the suprachiasmatic nuclei (SCN), site of the master circadian clock. To investigate effects of day length change on the sheep SCN, we used in-situ hybridization to assess the daily temporal organization of expression of circadian clock genes (Per1, Per2, Bmal1 and Fbxl21) and neuropeptides (Vip, Grp and Avp) in animals acclimated to a short photoperiod (SP; 8h of light) and at 3 or 15 days following transfer to a long photoperiod (LP3, LP15, respectively; 16h of light), achieved by an acute 8-h delay of lights off. We found that waveforms of SCN gene expression conformed to those previously seen in LP acclimated animals within 3 days of transfer to LP. Mean levels of expression for Per1-2 and Fbxl21 were nearly 2-fold higher in the LP15 than in the SP group. The expression of Vip was arrhythmic and unaffected by photoperiod, while, in contrast to rodents, Grp expression was not detectable within the sheep SCN. Expression of the circadian output gene Avp cycled robustly in all photoperiod groups with no detectable change in phasing. Overall these data suggest that synchronizing effects of light on SCN circadian organisation proceed similarly in ungulates and in rodents, despite differences in neuropeptide gene expression.

The bright-nights and dim-days of the urban photoperiod: implications for circadian rhythmicity, metabolism and obesity.

Artificial light decreases the amplitude of daily rhythms in human lifestyle principally by permitting activity and food intake to occur during hours of darkness, and allowing day-time activity to occur in dim light, indoors. Endogenous circadian timing mechanisms that oscillate with a period of 24 h have evolved to ensure physiology is synchronized with the daily variations in light, food, and social cues of the environment. Artificial light affects the synchronization between these oscillators, and metabolic disruption may be one consequence of this. By dampening the amplitude of environmental timing cues and disrupting circadian rhythmicity, artificial lighting might initiate metabolic disruption and contribute to the association between global urbanization and obesity. The aim of this review is to explore the historical, physiological, and epidemiological relationships between artificial light and circadian and metabolic dysfunction.

Does human evolution in different latitudes influence susceptibility to obesity via the circadian pacemaker?: migration and survival of the fittest in the modern age of lifestyle-induced circadian desynchrony.

The variable photoperiods of Northern latitudes challenge the entrainment capacity of the circadian pacemaker, which evolved under constant photoperiods in Equatorial regions. Entrainment to the erratic photoperiods facilitated by artificial light presents an additional challenge. Metabolic dysfunction and obesity are potential consequences of such desynchronization of circadian and environmental rhythms.

A molecular switch for photoperiod responsiveness in mammals.

Seasonal synchronization based on day length (photoperiod) allows organisms to anticipate environmental change. Photoperiodic decoding relies on circadian clocks, but the underlying molecular pathways have remained elusive [1]. In mammals and birds, photoperiodic responses depend crucially on expression of thyrotrophin ß subunit RNA (TSHß) in the pars tuberalis (PT) of the pituitary gland [2-4]. Now, using our well-characterized Soay sheep model [2], we describe a molecular switch governing TSHß transcription through the circadian clock. Central to this is a conserved D element in the TSHß promoter, controlled by the circadian transcription factor thyrotroph embryonic factor (Tef). In the PT, long-day exposure rapidly induces expression of the coactivator eyes absent 3 (Eya3), which synergizes with Tef to maximize TSHß transcription. The pineal hormone melatonin, secreted nocturnally, sets the phase of rhythmic Eya3 expression in the PT to peak 12 hr after nightfall. Additionally, nocturnal melatonin levels directly suppress Eya3 expression. Together, these effects form a switch triggering a strong morning peak of Eya3 expression under long days. Species variability in the TSHß D element influences sensitivity to TEF, reflecting species variability in photoperiodic responsiveness. Our findings define a molecular pathway linking the circadian clock to the evolution of seasonal timing in mammals.

Impact of aging on diurnal expression patterns of CLOCK and BMAL1 in the mouse brain.

Mammalian circadian rhythms are generated by a network of transcriptional and translational loops in the expression of a panel of clock genes in various brain and peripheral sites. Many of the output rhythms controlled by this system are significantly affected by ageing, although the mechanisms of age-related circadian dysfunction remain opaque. The aim of this study was to investigate the effect of aging on the daily oscillation of two clock gene proteins (CLOCK, BMAL1) in the mouse brain. Clock gene protein expression in the brain was measured by means of immunohistochemistry in groups of young (4 months) and older (16 months) mice sampled every 4h over a 24-h cycle. CLOCK and BMAL1 were constitutively expressed in the suprachiasmatic nucleus (SCN; the master circadian pacemaker) in young adult animals. We report novel rhythmic expression of CLOCK and BMAL1 in a number of extra-SCN sites in the young mouse brain, including the hippocampus, amygdala and the paraventricular, arcuate and dorsomedial nuclei of the hypothalamus. Aging altered the amplitude and/or phase of expression in these regions. These results indicate hitherto unreported expression patterns of CLOCK and BMAL1 in non-SCN brain circadian oscillators, and suggest that alterations of these patterns may contribute to age-related circadian dysfunction.

Neuroimmunology of the circadian clock.

Circadian timekeeping is a ubiquitous feature of all eukaryotes which allows for the imposition of a biologically appropriate temporal architecture on an animal's physiology, behavior and metabolism. There is growing evidence that in mammals the processes of circadian timing are under the influence of the immune system. Such a role for the neuroimmune regulation of the circadian clock has inferences for phenomena such as sickness behavior. Conversely, there is also accumulating evidence for a circadian influence on immune function, raising the likelihood that there is a bidirectional communication between the circadian and immune systems. In this review, we examine the evidence for these interactions, including circadian rhythmicity in models of disease and immune challenge, distribution of cytokines and their receptors in the suprachiasmatic nucleus of the hypothalamus, the site of the master circadian pacemaker, and the evidence for endogenous circadian timekeeping in immune cells.

Retention of acetaminophen in an in vitro model of solid-phase gastric emptying of animals.

OBJECTIVE: To apply an in vitro model for assessment of the solid-phase binding capacity of acetaminophen and thus assess the reliability of this marker for evaluation of solid-phase gastric emptying in vivo in animals. SAMPLE POPULATION: 4 test meals. PROCEDURES: A spectrophotometric method for detection of acetaminophen was validated and applied for assessment of the percentage retention of acetaminophen in the solid phase of 4 test meals. The gastric milieu was simulated by incubating each meal in artificial gastric juice for 2 hours in a shaking water bath maintained at 37 degrees C. Solid-phase retention was then assessed 3 times by measuring the amount of acetaminophen that had leached into the liquid phase. RESULTS: Acetaminophen was poorly retained in the solid phase of all the test meals examined in the study. There was also a large degree of variability in the percentage retention for each meal when the experiment was repeated 3 times. CONCLUSIONS AND CLINICAL RELEVANCE: Analysis of the results of this in vitro study confirmed that acetaminophen may not be an appropriate marker of solid-phase gastric emptying. The acetaminophen gastric emptying test should be applied only for the assessment of liquid-phase emptying in animals.

Development of high-resolution real-time sub-ppb ethane spectroscopy and some pilot studies in life science.

We describe a high-resolution real-time spectroscopy system targeted to ethane gas with sensitivity > or = 70 ppt and response time from > or = 0.7 s. The measurement technique is based on a mid-IR lead-salt laser passing through a Herriott cell through which a gas sample flows. We compare wavelength scanning and locked configurations and discuss their relative merits. The technology has been motivated by clinical breath testing applications, ethane being widely regarded as the most important breath biomarker for cell damage via free-radical-mediated oxidative attack. We discuss preliminary human and animal studies in which ultrasensitive real-time ethane detection offers new diagnostic and monitoring potential.

Comparison of the carbon 13-labeled octanoic acid breath test and ultrasonography for assessment of gastric emptying of a semisolid meal in dogs.

OBJECTIVE: To compare the rate of gastric emptying of a semisolid meal by use of the carbon 13-labeled octanoic acid breath test (13C-OBT) and gastric emptying ultrasonography (GEU) in dogs. ANIMALS: 10 healthy dogs. PROCEDURE: Food was withheld from dogs for 12 hours before ingestion of a test meal (bread, egg, and skimmed milk) containing 13C-octanoic acid. The gastric antrum was visualized by use of a 6.5-MHz microconvex transducer, and the area of the ellipse defined by the craniocaudal and ventrodorsal diameters of the stomach was measured. Samples of expired air and antral images were obtained 30 minutes before ingestion of the test meal and then every 15 minutes for 4 hours and every 30 minutes for a further 2 hours. The half-dose recovery time with the 13C-OBT (t1/2[BT]) and the gastric half emtying time with GEU (t50%[GEU]) was calculated. RESULTS: Mean +/- SD values for the t1/2(BT) and t50%(GEU) were 3.44 +/- 0.48 hours and 1.89 +/- 0.78 hours, respectively. A significant correlation was detected between the t1/2(BT) and t50%(GEU), although there was a large (1.55 hours) mean difference between these indices. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicated that there was a correlation between the rate of solid-phase gastric emptying assessed by use of GEU and the 13C-OBT in dogs. Gastric emptying ultrasonography may be a useful, noninvasive method for assessment of the rate of solid-phase gastric emptying in dogs.