Excessive Daytime Sleepiness Is Associated with Changes in Salivary Inflammatory Genes Transcripts

Excessive daytime sleepiness (EDS) is a ubiquitous problem that affects public health and safety. A test that can reliably identify individuals that suffer from EDS is needed. In contrast to other methods, salivary biomarkers are an objective, inexpensive, and noninvasive method to identify individuals with inadequate sleep. Although we have previously shown that inflammatory genes are elevated in saliva samples taken from sleep deprived individuals, it is unclear if inflammatory genes will be elevated in clinical populations with EDS. In this study, salivary samples from individuals with sleep apnea were evaluated using the Taqman low density inflammation array. Transcript levels for 3 genes, including prostaglandin-endoperoxide synthase 2 (PTGS2), were elevated in patients with sleep apnea. Interestingly, PTGS2 was also elevated in patients with EDS but who did not have sleep apnea. These data demonstrate the feasibility of using salivary transcript levels to identify individuals that self-report excessive daytime sleepiness

Measuring the impact of apnea and obesity on circadian activity patterns using functional linear modeling of actigraphy data

<p>Abstract</p> <p>Background</p> <p>Actigraphy provides a way to objectively measure activity in human subjects. This paper describes a novel family of statistical methods that can be used to analyze this data in a more comprehensive way.</p> <p>Methods</p> <p>A statistical method for testing differences in activity patterns measured by actigraphy across subgroups using functional data analysis is described. For illustration this method is used to statistically assess the impact of apnea-hypopnea index (apnea) and body mass index (BMI) on circadian activity patterns measured using actigraphy in 395 participants from 18 to 80 years old, referred to the Washington University Sleep Medicine Center for general sleep medicine care. Mathematical descriptions of the methods and results from their application to real data are presented.</p> <p>Results</p> <p>Activity patterns were recorded by an Actical device (Philips Respironics Inc.) every minute for at least seven days. Functional linear modeling was used to detect the association between circadian activity patterns and apnea and BMI. Results indicate that participants in high apnea group have statistically lower activity during the day, and that BMI in our study population does not significantly impact circadian patterns.</p> <p>Conclusions</p> <p>Compared with analysis using summary measures (e.g., average activity over 24 hours, total sleep time), Functional Data Analysis (FDA) is a novel statistical framework that more efficiently analyzes information from actigraphy data. FDA has the potential to reposition the focus of actigraphy data from general sleep assessment to rigorous analyses of circadian activity rhythms.</p

Comparison of single-channel EEG, actigraphy, and sleep diary in cognitively normal and mildly impaired older adults

STUDY OBJECTIVES: Multiple methods for monitoring sleep-wake activity have identified sleep disturbances as risk factors for Alzheimer disease (AD). In order to identify the level of agreement between different methods, we compared sleep parameters derived from single-channel EEG (scEEG), actigraphy, and sleep diaries in cognitively normal and mildly impaired older adults. METHODS: Two hundred ninety-three participants were monitored at home for up to six nights with scEEG, actigraphy, and sleep diaries. Total sleep time (TST), sleep efficiency (SE), sleep onset latency (SOL), and wake after sleep onset (WASO) were calculated using each of these methods. In 109 of the 293 participants, the ratio of cerebrospinal fluid concentrations of phosphorylated tau (p-tau) and amyloid-β-42 (Aβ42) was used as a biomarker for AD pathology. RESULTS: Agreement was highest for TST across instruments, especially in cognitively normal older adults. Overall, scEEG and actigraphy appeared to have greater agreement for multiple sleep parameters than for scEEG and diary or actigraphy and diary. Levels of agreement between scEEG and actigraphy overall decreased in mildly impaired participants and those with biomarker evidence of AD pathology, especially for measurements of TST. CONCLUSIONS: Caution should be exercised when comparing scEEG and actigraphy in individuals with mild cognitive impairment or with AD pathology. Sleep diaries may capture different aspects of sleep compared to scEEG and actigraphy. Additional studies comparing different methods of measuring sleep-wake activity in older adults are necessary to allow for comparison between studies using different methods

Cross-translational studies in human and Drosophila identify markers of sleep loss

Inadequate sleep has become endemic, which imposes a substantial burden for public health and safety. At present, there are no objective tests to determine if an individual has gone without sleep for an extended period of time. Here we describe a novel approach that takes advantage of the evolutionary conservation of sleep to identify markers of sleep loss. To begin, we demonstrate that IL-6 is increased in rats following chronic total sleep deprivation and in humans following 30 h of waking. Discovery experiments were then conducted on saliva taken from sleep-deprived human subjects to identify candidate markers. Given the relationship between sleep and immunity, we used Human Inflammation Low Density Arrays to screen saliva for novel markers of sleep deprivation. Integrin αM (ITGAM) and Anaxin A3 (AnxA3) were significantly elevated following 30 h of sleep loss. To confirm these results, we used QPCR to evaluate ITGAM and AnxA3 in independent samples collected after 24 h of waking; both transcripts were increased. The behavior of these markers was then evaluated further using the power of Drosophila genetics as a cost-effective means to determine whether the marker is associated with vulnerability to sleep loss or other confounding factors (e.g., stress). Transcript profiling in flies indicated that the Drosophila homologues of ITGAM were not predictive of sleep loss. Thus, we examined transcript levels of additional members of the integrin family in flies. Only transcript levels of scab, the Drosophila homologue of Integrin α5 (ITGA5), were associated with vulnerability to extended waking. Since ITGA5 was not included on the Low Density Array, we returned to human samples and found that ITGA5 transcript levels were increased following sleep deprivation. These cross-translational data indicate that fly and human discovery experiments are mutually reinforcing and can be used interchangeably to identify candidate biomarkers of sleep loss

Glioma-associated microglia/macrophages augment tumorigenicity in canine astrocytoma, a naturally occurring model of human glioma.

BackgroundGlioma-associated microglia/macrophages (GAMs) markedly influence glioma progression. Under the influence of transforming growth factor beta (TGFB), GAMs are polarized toward a tumor-supportive phenotype. However, neither therapeutic targeting of GAM recruitment nor TGFB signaling demonstrated efficacy in glioma patients despite efficacy in preclinical models, underscoring the need for a comprehensive understanding of the TGFB/GAM axis. Spontaneously occurring canine gliomas share many features with human glioma and provide a complementary translational animal model for further study. Given the importance of GAM and TGFB in human glioma, the aims of this study were to further define the GAM-associated molecular profile and the relevance of TGFB signaling in canine glioma that may serve as the basis for future translational studies.MethodsGAM morphometry, levels of GAM-associated molecules, and the canonical TGFB signaling axis were compared in archived samples of canine astrocytomas versus normal canine brain. Furthermore, the effect of TGFB on the malignant phenotype of canine astrocytoma cells was evaluated.ResultsGAMs diffusely infiltrated canine astrocytomas. GAM density was increased in high-grade tumors that correlated with a pro-tumorigenic molecular signature and upregulation of the canonical TGFB signaling axis. Moreover, TGFB1 enhanced the migration of canine astrocytoma cells in vitro.ConclusionsCanine astrocytomas share a similar GAM-associated immune landscape with human adult glioma. Our data also support a contributing role for TGFB1 signaling in the malignant phenotype of canine astrocytoma. These data further support naturally occurring canine glioma as a valid model for the investigation of GAM-associated therapeutic strategies for human malignant glioma

Cross-Translational Studies in Human and <i>Drosophila</i> Identify Markers of Sleep Loss

Inadequate sleep has become endemic, which imposes a substantial burden for public health and safety. At present, there are no objective tests to determine if an individual has gone without sleep for an extended period of time. Here we describe a novel approach that takes advantage of the evolutionary conservation of sleep to identify markers of sleep loss. To begin, we demonstrate that IL-6 is increased in rats following chronic total sleep deprivation and in humans following 30 h of waking. Discovery experiments were then conducted on saliva taken from sleep-deprived human subjects to identify candidate markers. Given the relationship between sleep and immunity, we used Human Inflammation Low Density Arrays to screen saliva for novel markers of sleep deprivation. Integrin αM (ITGAM) and Anaxin A3 (AnxA3) were significantly elevated following 30 h of sleep loss. To confirm these results, we used QPCR to evaluate ITGAM and AnxA3 in independent samples collected after 24 h of waking; both transcripts were increased. The behavior of these markers was then evaluated further using the power of Drosophila genetics as a cost-effective means to determine whether the marker is associated with vulnerability to sleep loss or other confounding factors (e.g., stress). Transcript profiling in flies indicated that the Drosophila homologues of ITGAM were not predictive of sleep loss. Thus, we examined transcript levels of additional members of the integrin family in flies. Only transcript levels of scab, the Drosophila homologue of Integrin α5 (ITGA5), were associated with vulnerability to extended waking. Since ITGA5 was not included on the Low Density Array, we returned to human samples and found that ITGA5 transcript levels were increased following sleep deprivation. These cross-translational data indicate that fly and human discovery experiments are mutually reinforcing and can be used interchangeably to identify candidate biomarkers of sleep loss.</p

Human subjects show an increase in salivary IL-6.

<p>(<b>A</b>) Subjective sleepiness as assessed using the Stanford Sleepiness Scale was increased following 30 h of wakefulness. Each subject served as their untreated circadian-matched control; t-test p = 0.0008. (<b>B</b>–<b>D</b>) Cognitive performance, assessed using the psychomotor vigilance task (PVT), was impaired following 30 h of wakefulness: (<b>B</b>) Lapses increased during waking compared to controls; t-test, p = 0.03. (<b>C</b>) Mean reaction time, expressed as 1/RT) slowed; t-test p = 0.001. (<b>D</b>) Reaction times in the slowest 10% also slowed t-test p = 0002. (<b>E</b>) Levels of IL-6 protein from human saliva after 30 hours of waking. Levels in sleep deprived subjects (n = 16) are expressed as a % of that their own untreated circadian-matched sample (con); t-test p = 0.042.</p

The <i>Drosophila</i> integrin <i>scb</i> and Human <i>ITGA5</i> are modified by sleep loss.

<p>(<b>A</b>) <i>scb</i> is elevated following caffeine administration but not after waking induced by methamphetamine. (<b>B</b>) <i>scb</i> is elevated in sleep deprived <i>cyc⁰¹</i> mutants but is not increased when waking is induced by starvation. (<b>C</b>) <i>scb</i> mRNA levels remain low following deprivations that do not activate homeostatic mechanisms (3 and 6 h SD), but are elevated following deprivations that activate homeostatic mechanisms (9 and 12 h SD). (<b>D</b>–<b>E</b>) Salivary <i>ITGA5</i> transcripts are increased in humans following 30 h of wakefulness while 24 h of wakefulness do not quite reach significance; t-test and p = 0.04 and p = 0.056 respectively. Each subject serves as their own circadian matched untreated control; data are expressed as a percent change from control.</p

IL-6 is elevated in serum of chronic total sleep deprived rats.

<p>(<b>A</b>) Energy Expenditure expressed as % of home cage controls (HCC, n = 7) is significantly increased in total sleep deprived (TSD, n = 7) rats compared to yoked controls (TSC, n = 6); One way ANOVA F_(2,11) = 6.79, p = .012, * modified Bonferroni test p<0.05 (<b>B</b>) Serum concentrations of IL-6 protein from individual TSD, TSC, and HCC rats. Arrows denote which animals tested positive for bacteria. (<b>C</b>) Mean concentration of IL-6 from individual rats in (B). One way ANOVA F_(2,17) = 5.22, p = .017 * p<0.05 modified Bonferroni test <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0061016#pone.0061016-Keppel1" target="_blank">[58]</a>. Data are presented as mean±SEM.</p