Epigenetic signals associated with delirium replicated across four independent cohorts.

Delirium is risky and indicates poor outcomes for patients. Therefore, it is crucial to create an effective delirium detection method. However, the epigenetic pathophysiology of delirium remains largely unknown. We aimed to discover reliable and replicable epigenetic (DNA methylation: DNAm) markers that are associated with delirium including post-operative delirium (POD) in blood obtained from patients among four independent cohorts. Blood DNA from four independent cohorts (two inpatient cohorts and two surgery cohorts; 16 to 88 patients each) were analyzed using the Illumina EPIC array platform for genome-wide DNAm analysis. We examined DNAm differences in blood between patients with and without delirium including POD. When we compared top CpG sites previously identified from the initial inpatient cohort with three additional cohorts (one inpatient and two surgery cohorts), 11 of the top 13 CpG sites showed statistically significant differences in DNAm values between the delirium group and non-delirium group in the same directions as found in the initial cohort. This study demonstrated the potential value of epigenetic biomarkers as future diagnostic tools. Furthermore, our findings provide additional evidence of the potential role of epigenetics in the pathophysiology of delirium including POD.

The bispectral electroencephalography (BSEEG) method quantifies post-operative delirium-like states in young and aged male mice after head mount implantation surgery.

Delirium, a syndrome characterized by an acute change in attention, awareness, and cognition, is commonly observed in older adults, although there are few quantitative monitoring methods in the clinical setting. We developed a bispectral electroencephalography (BSEEG) method capable of detecting delirium and can quantify the severity of delirium using a novel algorithm. Pre-clinical application of this novel BSEEG method can capture a delirium-like state in mice following LPS administration. However, its application to postoperative delirium (POD) has not yet been validated in animal experiments. This study aimed to create a POD model in mice with the BSEEG method by monitoring BSEEG scores following EEG head-mount implantation surgery and throughout the recovery. We compared the BSEEG scores of C57BL/6J young (2-3 months old) with aged (18-19 months old) male mice for quantitative evaluation of POD-like states. Postoperatively, both groups displayed increased BSEEG scores and a loss of regular diurnal changes in BSEEG scores. In young mice, BSEEG scores and regular diurnal changes recovered relatively quickly to baseline by postoperative day 3. Conversely, aged mice exhibited prolonged increases in postoperative BSEEG scores and it reached steady states only after postoperative day 8. This study suggests that the BSEEG method can be utilized as a quantitative measure of POD and assess the effect of aging on recovery from POD in the pre-clinical model.

Genome-wide DNA methylation analysis in female veterans with military sexual trauma and comorbid PTSD/MDD.

BACKGROUND: Military sexual trauma (MST) is a prevalent issue within the U.S. military. Victims are more likely to develop comorbid diseases such as posttraumatic stress disorder (PTSD) and major depressive disorder (MDD). Nonetheless, not everyone who suffers from MST develops PTSD and/or MDD. DNA methylation, which can regulate gene expression, might give us insight into the molecular mechanisms behind this discrepancy. Therefore, we sought to identify genomic loci and enriched biological pathways that differ between patients with and without MST, PTSD, and MDD. METHODS: Saliva samples were collected from 113 female veterans. Following DNA extraction and processing, DNA methylation levels were measured through the Infinium HumanMethylationEPIC BeadChip array. We used limma and bump hunting methods to generate the differentially methylated positions and differentially methylated regions (DMRs), respectively. Concurrently, we used Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genome to find enriched pathways. RESULTS: A DMR close to the transcription start site of ZFP57 was differentially methylated between subjects with and without PTSD, replicating previous findings and emphasizing the potential role of ZFP57 in PTSD susceptibility. In the pathway analyses, none survived multiple correction, although top GO terms included some potentially relevant to MST, PTSD, and MDD etiology. CONCLUSION: We conducted one of the first DNA methylation analyses investigating MST along with PTSD and MDD. In addition, we found one DMR near ZFP57 to be associated with PTSD. The replication of this finding indicates further investigation of ZFP57 in PTSD may be warranted.

NSAIDs use history: impact on the genome-wide DNA methylation profile and possible mechanisms of action.

BACKGROUND AND OBJECTIVE: NSAIDs inhibit cyclooxygenase, but their role in aging and other diseases is not well understood. Our group previously showed the potential benefit of NSAIDs in decreasing the risk of delirium and mortality. Concurrently, epigenetics signals have also been associated with delirium. Therefore, we sought to find differentially methylated genes and biological pathways related to exposure with NSAIDs by comparing the genome-wide DNA methylation profiles of patients with and without a history of NSAIDs use. METHODS: Whole blood samples were collected from 171 patients at the University of Iowa Hospital and Clinics from November 2017 to March 2020. History of NSAIDs use was assessed through a word-search function in the subjects' electronic medical records. DNA was extracted from the blood samples, processed with bisulfite conversion, and analyzed using Illumina's EPIC array. The analysis of top differentially methylated CpG sites and subsequent enrichment analysis were conducted using an established pipeline using R statistical software. RESULTS: Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genome (KEGG) showed several biological pathways relevant to NSAIDs' function. The identified GO terms included "arachidonic acid metabolic process," while KEGG results included "linoleic acid metabolism," "cellular senescence," and "circadian rhythm." Nonetheless, none of the top GO and KEGG pathways and the top differentially methylated CpG sites reached statistical significance. CONCLUSION: Our results suggest a potential role of epigenetics in the mechanisms of the action of NSAIDs. However, the results should be viewed with caution as exploratory and hypothesis-generating given the lack of statistically significant findings.

Bispectral EEG (BSEEG) Algorithm Captures High Mortality Risk Among 1,077 Patients: Its Relationship to Delirium Motor Subtype.

OBJECTIVE: Delirium is dangerous and a predictor of poor patient outcomes. We have previously reported the utility of the bispectral EEG (BSEEG) with a novel algorithm for the detection of delirium and prediction of patient outcomes including mortality. The present study employed a normalized BSEEG (nBSEEG) score to integrate the previous cohorts to combine their data to investigate the prediction of patient outcomes. We also aimed to test if the BSEEG method can be applicable regardless of age, and independent of delirium motor subtypes. METHODS: We calculated nBSEEG score from raw BSEEG data in each cohort and classified patients into BSEEG-positive and BSEEG-negative groups. We used log-rank test and Cox proportional hazards models to predict 90-day and 1-year outcomes for the BSEEG-positive and -negative groups in all subjects and motor subgroups. RESULTS: A total of 1,077 subjects, the BSEEG-positive group showed significantly higher 90-day (hazard ratio 1.33 [95% CI 1.16-1.52] and 1-year (hazard ratio 1.22 [95% CI 1.06-1.40] mortality rates than the negative group after adjustment for covariates such as age, sex, CCI, and delirium status. Among patients with different motor subtypes of delirium, the hypoactive group showed significantly higher 90-day (hazard ratio 1.41 [95% CI 1.12-1.76] and 1-year mortality rates (hazard ratio 1.32 [95% CI 1.05-1.67], which remained significant after adjustment for the same covariates. CONCLUSION: We found that the BSEEG method is capable of capturing patients at high mortality risk.

Genome-wide DNA methylation analysis of post-operative delirium with brain, blood, saliva, and buccal samples from neurosurgery patients.

OBJECTIVE: No previous study demonstrates the difference in the genome-wide DNA methylation status of post-operative delirium (POD) using human brain tissue obtained from neurosurgery and multiple peripheral tissues such as blood, saliva, and buccal samples from the same individuals. We aimed to identify epigenetic marks of DNA methylation in the brain and peripheral tissues to elucidate the potential pathophysiological mechanism of POD. METHODS: The four tissue types (brain, blood, saliva, buccal) of DNA samples from up to 40 patients, including 11 POD cases, were analyzed using Illumina EPIC array. DNAm differences between patients with and without POD were examined. We also conducted enrichment analysis based on the top DNAm signals. RESULTS: The most different CpG site between control and POD was found at cg16526133 near the ADAMTS9 gene from the brain tissue(p = 8.66E-08). However, there are no CpG sites to reach the genome-wide significant level. The enrichment analysis based on the 1000 top hit CpG site (p < 0.05) on the four tissues showed several intriguing pathways. In the brain, there are pathways including "positive regulation of glial cell differentiation". Blood samples showed also pathways related to immune function. Besides, both saliva and the buccal sample showed pathways related to circadian rhythm, although these findings were not FDR significant. CONCLUSION: Enrichment analysis found several intriguing pathways related to potential delirium pathophysiology. Present data may further support the role of epigenetics, especially DNA methylation, in the molecular mechanisms of delirium pathogenesis.

Super-Resolution Defocusing Nanoparticle Image Velocimetry Utilizing Spherical Aberration for Nanochannel Flows.

Understanding fluid flows and mass transport in nanospaces is becoming important with recent advances in nanofluidic analytical devices utilizing nanopores and nanochannels. In the present study, we developed a super-resolution and fast particle tracking method utilizing defocusing images with spherical aberration and demonstrated the measurement of nanochannel flow. Since the spherical aberration generates the defocusing nanoparticle image with diffraction rings, the position of fluorescent nanoparticles was determined from the radius of the diffraction ring. Effects of components of an optical system on the diffraction ring of the defocusing image were investigated and optimized to achieve the spatial resolution exceeding the optical diffraction limit. We found that there is an optimal magnitude of spherical aberration to enhance the spatial resolution. Furthermore, we confirmed that nanoparticles with diameters in the order of 101 nm, which is much smaller than the light wavelength, do not affect the defocusing images and the spatial resolution because such nanoparticles can be regarded as point light sources. At optimized conditions, we achieved a spatial resolution of 19 nm and a temporal resolution of 160 µs, which are sufficient for the nanochannel flow measurements. We succeeded in the measurement of pressure-driven flow in a nanochannel with a depth of 370 nm using 67 nm fluorescent nanoparticles. The measured nanoparticle velocities exhibited a parabolic flow profile with a slip velocity even at the hydrophilic glass surface but with an average velocity similar to the Hagen-Poiseuille law. The method will accelerate researches in the nanofluidics and other related fields.

Exercise Reverses Behavioral and Synaptic Abnormalities after Maternal Inflammation.

Abnormal behaviors in individuals with neurodevelopmental disorders are generally believed to be irreversible. Here, we show that voluntary wheel running ameliorates the abnormalities in sociability, repetitiveness, and anxiety observed in a mouse model of a neurodevelopmental disorder induced by maternal immune activation (MIA). Exercise activates a portion of dentate granule cells, normalizing the density of hippocampal CA3 synapses, which is excessive in the MIA-affected offspring. The synaptic surplus in the MIA offspring is induced by deficits in synapse engulfment by microglia, which is normalized by exercise through microglial activation. Finally, chemogenetically induced activation of granule cells promotes the engulfment of CA3 synapses. Thus, our study proposes a role of voluntary exercise in the modulation of behavioral and synaptic abnormalities in neurodevelopmental disorders.

Nitric Oxide-induced Activation of the Type 1 Ryanodine Receptor Is Critical for Epileptic Seizure-induced Neuronal Cell Death.

Status epilepticus (SE) is a life-threatening emergency that can cause neurodegeneration with debilitating neurological disorders. However, the mechanism by which convulsive SE results in neurodegeneration is not fully understood. It has been shown that epileptic seizures produce markedly increased levels of nitric oxide (NO) in the brain, and that NO induces Ca2+ release from the endoplasmic reticulum via the type 1 ryanodine receptor (RyR1), which occurs through S-nitrosylation of the intracellular Ca2+ release channel. Here, we show that through genetic silencing of NO-induced activation of the RyR1 intracellular Ca2+ release channel, neurons were rescued from seizure-dependent cell death. Furthermore, dantrolene, an inhibitor of RyR1, was protective against neurodegeneration caused by SE. These results demonstrate that NO-induced Ca2+ release via RyR is involved in SE-induced neurodegeneration, and provide a rationale for the use of RyR1 inhibitors for the prevention of brain damage following SE.

Repulsive guidance molecule a regulates hippocampal mossy fiber branching in vitro.

Repulsive guidance molecule a (RGMa), which binds to its receptor neogenin, has been well determined as a repulsive axon guidance molecule. However, whether RGMa affects the growth of hippocampal mossy fibers, the axons of dentate granule cells, has been unknown. In the present study, we found that the primary neurons in the hippocampus express both RGMa and neogenin in the postnatal rats. To examine the role of RGMa in the mossy fiber growth, the morphology of granule cells was clearly visualized by transfecting membrane-targeted green fluorescent protein using a single-cell electroporation method in cultured hippocampal slices. In the slice cultures, we found that intrinsic RGMa is required to inhibit excess branching of the hippocampal mossy fibers. Furthermore, hyperexcitability-induced aberrant branching of the mossy fibers in the cultured slices was blocked by applying the recombinant RGMa protein. Therefore, this study suggests that RGMa regulates the proper axonal branching of hippocampal mossy fibers.