The Sleep Quality- and Myopia-Linked PDE11A-Y727C Variant Impacts Neural Physiology by Reducing Catalytic Activity and Altering Subcellular Compartmentalization of the Enzyme.

Recently, a Y727C variant in the dual-specific 3',5'-cyclic nucleotide phosphodiesterase 11A (PDE11A-Y727C) was linked to increased sleep quality and reduced myopia risk in humans. Given the well-established role that the PDE11 substrates cAMP and cGMP play in eye physiology and sleep, we determined if (1) PDE11A protein is expressed in the retina or other eye segments in mice, (2) PDE11A-Y7272C affects catalytic activity and/or subcellular compartmentalization more so than the nearby suicide-associated PDE11A-M878V variant, and (3) Pde11a deletion alters eye growth or sleep quality in male and female mice. Western blots show distinct protein expression of PDE11A4, but not PDE11A1-3, in eyes of Pde11a WT, but not KO mice, that vary by eye segment and age. In HT22 and COS-1 cells, PDE11A4-Y727C reduces PDE11A4 catalytic activity far more than PDE11A4-M878V, with both variants reducing PDE11A4-cAMP more so than PDE11A4-cGMP activity. Despite this, Pde11a deletion does not alter age-related changes in retinal or lens thickness or axial length, nor vitreous or anterior chamber depth. Further, Pde11a deletion only minimally changes refractive error and sleep quality. That said, both variants also dramatically alter the subcellular compartmentalization of human and mouse PDE11A4, an effect occurring independently of dephosphorylating PDE11A4-S117/S124 or phosphorylating PDE11A4-S162. Rather, re-compartmentalization of PDE11A4-Y727C is due to the loss of the tyrosine changing how PDE11A4 is packaged/repackaged via the trans-Golgi network. Therefore, the protective impact of the Y727C variant may reflect a gain-of-function (e.g., PDE11A4 displacing another PDE) that warrants further investigation in the context of reversing/preventing sleep disturbances or myopia.

The sleep quality- and myopia-linked PDE11A-Y727C variant impacts neural physiology by reducing catalytic activity and altering subcellular compartmentalization of the enzyme.

Recently, a Y727C variant in the dual-specific 3',5'-cyclic nucleotide phosphodiesterase 11A (PDE11A-Y727C) was linked to increased sleep quality and reduced myopia risk in humans. Given the well-established role that the PDE11 substrates cAMP and cGMP play in eye physiology and sleep, we determined if 1) PDE11A protein is expressed in the retina or other eye segments in mouse, 2) PDE11A-Y7272C affects catalytic activity and/or subcellular compartmentalization more so than the nearby suicide-associated PDE11A-M878V variant, and 3) Pde11a deletion alters eye growth or sleep quality in male and female mice. Western blots show distinct protein expression of PDE11A4, but not PDE11A1-3, in eyes of Pde11a WT-but not KO mice-that vary by eye segment and age. In HT22 and COS-1 cells, PDE11A4-Y727C reduces PDE11A4 catalytic activity far more than PDE11A4-M878V, with both variants reducing PDE11A4-cAMP more so than PDE11A4-cGMP activity. Despite this, Pde11a deletion does not alter age-related changes in retinal or lens thickness, axial length, nor vitreous or anterior chamber depth. Further, Pde11a deletion only minimally changes refractive error and sleep quality. That said, both variants also dramatically alter the subcellular compartmentalization of human and mouse PDE11A4, an effect occurring independently of dephosphorylating PDE11A4-S117/S124 or phosphorylating PDE11A4-S162. Rather, re-compartmentalization of PDE11A4-Y727C is due to the loss of the tyrosine changing how PDE11A4 is packaged/repackaged via the trans-Golgi network. Therefore, the protective impact of the Y727C variant may reflect a gain-of-function (e.g., PDE11A4 displacing another PDE) that warrants further investigation in the context of reversing/preventing sleep disturbances or myopia.

Reducing brain kynurenic acid synthesis precludes kynurenine-induced sleep disturbances.

Patients with neurocognitive disorders often battle sleep disturbances. Kynurenic acid is a tryptophan metabolite of the kynurenine pathway implicated in the pathology of these illnesses. Modest increases in kynurenic acid, an antagonist at glutamatergic and cholinergic receptors, result in cognitive impairments and sleep dysfunction. We explored the hypothesis that inhibition of the kynurenic acid synthesising enzyme, kynurenine aminotransferase II, may alleviate sleep disturbances. At the start of the light phase, adult male and female Wistar rats received systemic injections of either: (i) vehicle; (ii) kynurenine (100 mg kg-1 ; i.p.); (iii) the kynurenine aminotransferase II inhibitor, PF-04859989 (30 mg kg-1 ; s.c.); or (iv) PF-04859989 and kynurenine in combination. Kynurenine and kynurenic acid levels were evaluated in the plasma and brain. Separate animals were implanted with electroencephalogram and electromyogram telemetry devices to record polysomnography, and evaluate the vigilance states wake, rapid eye movement sleep and non-rapid eye movement sleep following each treatment. Kynurenine challenge increased brain kynurenic acid and resulted in reduced rapid eye movement sleep duration, non-rapid eye movement sleep delta power and sleep spindles. PF-04859989 reduced brain kynurenic acid formation when given prior to kynurenine, prevented disturbances in rapid eye movement sleep and sleep spindles, and enhanced non-rapid eye movement sleep. Our findings suggest that reducing kynurenic acid in conditions where the kynurenine pathway is activated may serve as a potential strategy for improving sleep dynamics.

The stress of losing sleep: Sex-specific neurobiological outcomes.

Sleep is a vital and evolutionarily conserved process, critical to daily functioning and homeostatic balance. Losing sleep is inherently stressful and leads to numerous detrimental physiological outcomes. Despite sleep disturbances affecting everyone, women and female rodents are often excluded or underrepresented in clinical and pre-clinical studies. Advancing our understanding of the role of biological sex in the responses to sleep loss stands to greatly improve our ability to understand and treat health consequences of insufficient sleep. As such, this review discusses sex differences in response to sleep deprivation, with a focus on the sympathetic nervous system stress response and activation of the hypothalamic-pituitary-adrenal (HPA) axis. We review sex differences in several stress-related consequences of sleep loss, including inflammation, learning and memory deficits, and mood related changes. Focusing on women's health, we discuss the effects of sleep deprivation during the peripartum period. In closing, we present neurobiological mechanisms, including the contribution of sex hormones, orexins, circadian timing systems, and astrocytic neuromodulation, that may underlie potential sex differences in sleep deprivation responses.

Kynurenine aminotransferase II inhibition promotes sleep and rescues impairments induced by neurodevelopmental insult.

Dysregulated sleep is commonly reported in individuals with neuropsychiatric disorders, including schizophrenia (SCZ) and bipolar disorder (BPD). Physiology and pathogenesis of these disorders points to aberrant metabolism, during neurodevelopment and adulthood, of tryptophan via the kynurenine pathway (KP). Kynurenic acid (KYNA), a neuroactive KP metabolite derived from its precursor kynurenine by kynurenine aminotransferase II (KAT II), is increased in the brains of individuals with SCZ and BPD. We hypothesize that elevated KYNA, an inhibitor of glutamatergic and cholinergic neurotransmission, contributes to sleep dysfunction. Employing the embryonic kynurenine (EKyn) paradigm to elevate fetal brain KYNA, we presently examined pharmacological inhibition of KAT II to reduce KYNA in adulthood to improve sleep quality. Pregnant Wistar rats were fed either kynurenine (100 mg/day)(EKyn) or control (ECon) diet from embryonic day (ED) 15 to ED 22. Adult male (N = 24) and female (N = 23) offspring were implanted with devices to record electroencephalogram (EEG) and electromyogram (EMG) telemetrically for sleep-wake data acquisition. Each subject was treated with either vehicle or PF-04859989 (30 mg/kg, s.c.), an irreversible KAT II inhibitor, at zeitgeber time (ZT) 0 or ZT 12. KAT II inhibitor improved sleep architecture maintaining entrainment of the light-dark cycle; ZT 0 treatment with PF-04859989 induced transient improvements in rapid eye movement (REM) and non-REM (NREM) sleep during the immediate light phase, while the impact of ZT 12 treatment was delayed until the subsequent light phase. PF-04859989 administration at ZT 0 enhanced NREM delta spectral power and reduced activity and body temperature. In conclusion, reducing de novo KYNA production alleviated sleep disturbances and increased sleep quality in EKyn, while also improving sleep outcomes in ECon offspring. Our findings place attention on KAT II inhibition as a novel mechanistic approach to treating disrupted sleep behavior with potential translational implications for patients with neurodevelopmental and neuropsychiatric disorders.

Prenatal Kynurenine Elevation Elicits Sex-Dependent Changes in Sleep and Arousal During Adulthood: Implications for Psychotic Disorders.

Dysregulation of the kynurenine pathway (KP) of tryptophan catabolism has been implicated in psychotic disorders, including schizophrenia and bipolar disorder. Kynurenic acid (KYNA) is a KP metabolite synthesized by kynurenine aminotransferases (KATs) from its biological precursor kynurenine and acts as an endogenous antagonist of N-methyl-D-aspartate and α7-nicotinic acetylcholine receptors. Elevated KYNA levels found in postmortem brain tissue and cerebrospinal fluid of patients are hypothesized to play a key role in the etiology of cognitive symptoms observed in psychotic disorders. Sleep plays an important role in memory consolidation, and sleep disturbances are common among patients. Yet, little is known about the effect of altered KP metabolism on sleep-wake behavior. We presently utilized a well-established experimental paradigm of embryonic kynurenine (EKyn) exposure wherein pregnant dams are fed a diet laced with kynurenine the last week of gestation and hypothesized disrupted sleep-wake behavior in adult offspring. We examined sleep behavior in adult male and female offspring using electroencephalogram and electromyogram telemetry and determined sex differences in sleep and arousal in EKyn offspring. EKyn males displayed reduced rapid eye movement sleep, while female EKyn offspring were hyperaroused compared to controls. We determined that EKyn males maintain elevated brain KYNA levels, while KYNA levels were unchanged in EKyn females, yet the activity levels of KAT I and KAT II were reduced. Our findings indicate that elevated prenatal kynurenine exposure elicits sex-specific changes in sleep-wake behavior, arousal, and KP metabolism.

Application of Machine Learning to Sleep Stage Classification.

Sleep studies are imperative to recapitulate phenotypes associated with sleep loss and uncover mechanisms contributing to psychopathology. Most often, investigators manually classify the polysomnography into vigilance states, which is time-consuming, requires extensive training, and is prone to inter-scorer variability. While many works have successfully developed automated vigilance state classifiers based on multiple EEG channels, we aim to produce an automated and openaccess classifier that can reliably predict vigilance state based on a single cortical electroencephalogram (EEG) from rodents to minimize the disadvantages that accompany tethering small animals via wires to computer programs. Approximately 427 hours of continuously monitored EEG, electromyogram (EMG), and activity were labeled by a domain expert out of 571 hours of total data. Here we evaluate the performance of various machine learning techniques on classifying 10-second epochs into one of three discrete classes: paradoxical, slow-wave, or wake. Our investigations include Decision Trees, Random Forests, Naive Bayes Classifiers, Logistic Regression Classifiers, and Artificial Neural Networks. These methodologies have achieved accuracies ranging from approximately 74% to approximately 96%. Most notably, the Random Forest and the ANN achieved remarkable accuracies of 95.78% and 93.31%, respectively. Here we have shown the potential of various machine learning classifiers to automatically, accurately, and reliably classify vigilance states based on a single EEG reading and a single EMG reading.

Neutrophil-to-Lymphocyte Ratio, Monocyte-to-Lymphocyte Ratio, Platelet-to-Lymphocyte Ratio, and Mean Platelet Volume-to-Platelet Count Ratio as Biomarkers in Critically Ill and Injured Patients: Which Ratio to Choose to Predict Outcome and Nature of Bacteremia?

BACKGROUND: Neutrophil-to-lymphocyte ratio (NLR), monocyte-to-lymphocyte ratio (MLR), platelet-to-lymphocyte ratio (PLR), and mean platelet volume-to-platelet count (MPV/PC) ratio are readily available parameters that might have discriminative power regarding outcome. The aim of our study was to assess prognostic value of these biomarkers regarding outcome in critically ill patients with secondary sepsis and/or trauma. METHODS: A total of 392 critically ill and injured patients, admitted to surgical ICU, were enrolled in a prospective observational study. Leukocyte and platelet counts were recorded upon fulfilling Sepsis-3 criteria and for traumatized Injury Severity Score > 25 points. Patients were divided into four subgroups: peritonitis, pancreatitis, trauma with sepsis, and trauma without sepsis. RESULTS: NLR and MPV/PC levels were significantly higher in nonsurvivors (AUC/ROC of 0.681 and 0.592, resp., in the peritonitis subgroup; 0.717 and 0.753, resp., in the pancreatitis subgroup); MLR and PLR did not differ significantly. There was no significant difference of investigated biomarkers between survivors and nonsurvivors in trauma patients with and without sepsis except for PLR in the trauma without sepsis subgroup (significantly higher in nonsurvivors, AUC/ROC of 0.719). Independent predictor of lethal outcome was NLR in the whole cohort and in the peritonitis subgroup as well as MPV in the pancreatitis subgroup. Also, there were statistically significant differences in MPV/PC, MLR, and PLR values regarding nature of bacteremia. In general, the lowest levels had been found in patients with Gram-positive blood cultures. CONCLUSIONS: NLR and MPV were very good independent predictors of lethal outcome. For the first time, we demonstrate that nature of bacteremia influences MPV/PC, MLR, and PLR. In heterogeneous cohort subgroup, analysis is essential.

Methylation-dependent regulation of hypoxia inducible factor-1 alpha gene expression by the transcription factor Kaiso.

Low oxygen tension (hypoxia) is a common characteristic of solid tumors and strongly correlates with poor prognosis and resistance to treatment. In response to hypoxia, cells initiate a cascade of transcriptional events regulated by the hypoxia inducible factor-1 (HIF-1) heterodimer. Since the oxygen-sensitive HIF-1α subunit is stabilized during hypoxia, it functions as the regulatory subunit of the protein. To date, while the mechanisms governing HIF-1α protein stabilization and function have been well studied, those governing HIF1A gene expression are not fully understood. However, recent studies have suggested that methylation of a HIF-1 binding site in the HIF1A promoter prevents its autoregulation. Here we report that the POZ-ZF transcription factor Kaiso modulates HIF1A gene expression by binding to the methylated HIF1A promoter in a region proximal to the autoregulatory HIF-1 binding site. Interestingly, Kaiso's regulation of HIF1A occurs primarily during hypoxia, which is consistent with the finding that Kaiso protein levels peak after 4 h of hypoxic incubation and return to normoxic levels after 24 h. Our data thus support a role for Kaiso in fine-tuning HIF1A gene expression after extended periods of hypoxia.

Kaiso overexpression promotes intestinal inflammation and potentiates intestinal tumorigenesis in Apc(Min/+) mice.

Constitutive Wnt/ß-catenin signaling is a key contributor to colorectal cancer (CRC). Although inactivation of the tumor suppressor adenomatous polyposis coli (APC) is recognized as an early event in CRC development, it is the accumulation of multiple subsequent oncogenic insults facilitates malignant transformation. One potential contributor to colorectal carcinogenesis is the POZ-ZF transcription factor Kaiso, whose depletion extends lifespan and delays polyp onset in the widely used Apc(Min/+) mouse model of intestinal cancer. These findings suggested that Kaiso potentiates intestinal tumorigenesis, but this was paradoxical as Kaiso was previously implicated as a negative regulator of Wnt/ß-catenin signaling. To resolve Kaiso's role in intestinal tumorigenesis and canonical Wnt signaling, we generated a transgenic mouse model (Kaiso(Tg/+)) expressing an intestinal-specific myc-tagged Kaiso transgene. We then mated Kaiso(Tg/+) and Apc(Min/+) mice to generate Kaiso(Tg/+):Apc(Min/+) mice for further characterization. Kaiso(Tg/+):Apc(Min/+) mice exhibited reduced lifespan and increased polyp multiplicity compared to Apc(Min/+) mice. Consistent with this murine phenotype, we found increased Kaiso expression in human CRC tissue, supporting a role for Kaiso in human CRC. Interestingly, Wnt target gene expression was increased in Kaiso(Tg/+):Apc(Min/+) mice, suggesting that Kaiso's function as a negative regulator of canonical Wnt signaling, as seen in Xenopus, is not maintained in this context. Notably, Kaiso(Tg/+):Apc(Min/+) mice exhibited increased inflammation and activation of NFκB signaling compared to their Apc(Min/+) counterparts. This phenotype was consistent with our previous report that Kaiso(Tg/+) mice exhibit chronic intestinal inflammation. Together our findings highlight a role for Kaiso in promoting Wnt signaling, inflammation and tumorigenesis in the mammalian intestine.

Influence of spinal and general anesthesia on the metabolic, hormonal, and hemodynamic response in elective surgical patients.

BACKGROUND: The aim of the study was to determine the significance of spinal anesthesia in the suppression of the metabolic, hormonal, and hemodynamic response to surgical stress in elective surgical patients compared to general anesthesia. MATERIAL AND METHODS: The study was clinical, prospective, and controlled and it involved 2 groups of patients (the spinal and the general anesthesia group) who underwent the same surgery. We monitored the metabolic and hormonal response to perioperative stress based on serum cortisol level and glycemia. We also examined how the different techniques of anesthesia affect these hemodynamic parameters: systolic arterial pressure (AP), diastolic AP, heart rate (HR), and arterial oxygen saturation (SpO2). These parameters were measured before induction on anesthesia (T1), 30 min after the surgical incisions (T2), 1 h postoperatively (T3) and 24 h after surgery (T4). RESULTS: Serum cortisol levels were significantly higher in the general anesthesia group compared to the spinal anesthesia group (p<0.01). Glycemia was significantly higher in the general anesthesia group (p<0.05). There was a statistically significant, positive correlation between serum cortisol levels and glycemia at all times observed (p<0.01). Systolic and diastolic AP did not differ significantly between the groups (p=0.191, p=0.101). The HR was significantly higher in the general anesthesia group (p<0.01). SpO2 values did not differ significantly between the groups (p=0.081). CONCLUSIONS: Based on metabolic, hormonal, and hemodynamic responses, spinal anesthesia proved more effective than general anesthesia in suppressing stress response in elective surgical patients.