Reduced Expression of the PP2A Methylesterase, PME-1, or the PP2A Methyltransferase, LCMT-1, Alters Sensitivity to Beta-Amyloid-Induced Cognitive and Electrophysiological Impairments in Mice

Beta-amyloid (Ab) is thought to play a critical role in Alzheimer’s disease (AD), and application of soluble oligomeric forms of Ab produces AD-like impairments in cognition and synaptic plasticity in experimental systems. We found previously that transgenic overexpression of the PP2A methylesterase, PME-1, or the PP2A methyltransferase, LCMT-1, altered the sensitivity of mice to Ab-induced impairments, suggesting that PME-1 inhibition may be an effective approach for preventing or treating these impairments. To explore this possibility, we examined the behavioral and electrophysiological effects of acutely applied synthetic Ab oligomers in male and female mice heterozygous for either a PME-1 KO or an LCMT-1 gene-trap mutation. We found that heterozygous PME-1 KO mice were resistant to Ab-induced impairments in cognition and synaptic plasticity, whereas LCMT-1 gene-trap mice showed increased sensitivity to Ab-induced impairments. The heterozygous PME-1 KO mice produced normal levels of endogenous Ab and exhibited normal electrophysiological responses to picomolar concentrations of Ab, suggesting that reduced PME-1 expression in these animals protects against Ab-induced impairments without impacting normal physiological Ab functions. Together, these data provide additional support for roles for PME-1 and LCMT-1 in regulating sensitivity to Ab-induced impairments, and suggest that inhibition of PME-1 may constitute a viable therapeutic approach for selectively protecting against the pathologic actions of Ab in AD

The REASON score: an epigenetic and clinicopathologic score to predict risk of poor survival in patients with early stage oral squamous cell carcinoma

Background Oral squamous cell carcinoma (OSCC) is a capricious cancer with poor survival rates, even for early-stage patients. There is a pressing need to develop more precise risk assessment methods to appropriately tailor clinical treatment. Genome-wide association studies have not produced a viable biomarker. However, these studies are limited by using heterogeneous cohorts, not focusing on methylation although OSCC is a heavily epigenetically-regulated cancer, and not combining molecular data with clinicopathologic data for risk prediction. In this study we focused on early-stage (I/II) OSCC and created a risk score called the REASON score, which combines clinicopathologic characteristics with a 12-gene methylation signature, to predict the risk of 5-year mortality. Methods We combined data from an internal cohort (n = 515) and The Cancer Genome Atlas (TCGA) cohort (n = 58). We collected clinicopathologic data from both cohorts to derive the non-molecular portion of the REASON score. We then analyzed the TCGA cohort DNA methylation data to derive the molecular portion of the risk score. Results 5-year disease specific survival was 63% for the internal cohort and 86% for the TCGA cohort. The clinicopathologic features with the highest predictive ability among the two the cohorts were age, race, sex, tobacco use, alcohol use, histologic grade, stage, perineural invasion (PNI), lymphovascular invasion (LVI), and margin status. This panel of 10 non-molecular features predicted 5-year mortality risk with a concordance (c)-index = 0.67. Our molecular panel consisted of a 12-gene methylation signature (i.e., HORMAD2, MYLK, GPR133, SOX8, TRPA1, ABCA2, HGFAC, MCPH1, WDR86, CACNA1H, RNF216, CCNJL), which had the most significant differential methylation between patients who survived vs. died by 5 years. All 12 genes have already been linked to survival in other cancers. Of the genes, only SOX8 was previously associated with OSCC; our study was the first to link the remaining 11 genes to OSCC survival. The combined molecular and non-molecular panel formed the REASON score, which predicted risk of death with a c-index = 0.915. Conclusions The REASON score is a promising biomarker to predict risk of mortality in early-stage OSCC patients. Validation of the REASON score in a larger independent cohort is warranted

Co‑expressed microRNAs, target genes and pathways related to metabolism, inflammation and endocrine function in individuals at risk for type 2 diabetes.

MicroRNAs (miRNAs) may be considered important regulators of risk for type 2 diabetes (T2D). The aim of the present study was to identify novel sets of miRNAs associated with T2D risk, as well as their gene and pathway targets. Circulating miRNAs (n=59) were measured in plasma from participants in a previously completed clinical trial (n=82). An agnostic statistical approach was applied to identify novel sets of miRNAs with optimal co‑expression patterns. In silico analyses were used to identify the messenger RNA and biological pathway targets of the miRNAs within each factor. A total of three factors of miRNAs were identified, containing 18, seven and two miRNAs each. Eight biological pathways were revealed to contain genes targeted by the miRNAs in all three factors, 38 pathways contained genes targeted by the miRNAs in two factors, and 55, 18 and two pathways were targeted by the miRNAs in a single factor, respectively (all q<0.05). The pathways containing genes targeted by miRNAs in the largest factor shared a common theme of biological processes related to metabolism and inflammation. By contrast, the pathways containing genes targeted by miRNAs in the second largest factor were related to endocrine function and hormone activity. The present study focused on the pathways uniquely targeted by each factor of miRNAs in order to identify unique mechanisms that may be associated with a subset of individuals. Further exploration of the genes and pathways related to these biological themes may provide insights about the subtypes of T2D and lead to the identification of novel therapeutic targets

Review of databases for experimentally validated human microRNA–mRNA interactions

MicroRNAs (miRs) may contribute to disease etiology by influencing gene expression. Numerous databases are available for miR target prediction and validation, but their functionality is varied, and outputs are not standardized. The purpose of this review is to identify and describe databases for cataloging validated miR targets. Using Tools4miRs and PubMed, we identified databases with experimentally validated targets, human data, and a focus on miR-messenger RNA (mRNA) interactions. Data were extracted about the number of times each database was cited, the number of miRs, the target genes, the interactions per database, experimental methodology and key features of each database. The search yielded 10 databases, which in order of most cited to least were: miRTarBase, starBase/The Encyclopedia of RNA Interactomes, DIANA-TarBase, miRWalk, miRecords, miRGator, miRSystem, miRGate, miRSel and targetHub. Findings from this review suggest that the information presented within miR target validation databases can be enhanced by adding features such as flexibility in performing queries in multiple ways, downloadable data, ongoing updates and integrating tools for further miR-mRNA target interaction analysis. This review is designed to aid researchers, especially those new to miR bioinformatics tools, in database selection and to offer considerations for future development and upkeep of validation tools. Database URL http://mirtarbase.cuhk.edu.cn/

Multi-Tiered Assessment of Gene Expression Provides Evidence for Mechanisms That Underlie Risk for Type 2 Diabetes

IntroductionIntegrated transcriptome and microRNA differential gene expression (DEG) analyses may help to explain type 2 diabetes (T2D) pathogenesis in at-risk populations. The purpose of this study was to characterize DEG in banked biospecimens from underactive adult participants who responded to a randomized clinical trial measuring the effects of lifestyle interventions on T2D risk factors. DEGs were further examined within the context of annotated biological pathways.MethodsParticipants (n = 52) in a previously completed clinical trial that assessed a 12-week behavioural intervention for T2D risk reduction were included. Participants who showed >6mg/dL decrease in fasting blood glucose were identified as responders. Gene expression was measured by RNASeq, and overrepresentation analysis within KEGG pathways and weighted gene correlation network analysis (WGCNA) were performed.ResultsNo genes remained significantly differentially expressed after correction for multiple comparisons. One module derived by WGCNA related to body mass index was identified, which contained genes located in KEGG pathways related to known mechanisms underlying risk for T2D as well as pathways related to neurodegeneration and protein misfolding. A network analysis showed indirect connections between genes in this module and islet amyloid polypeptide (IAPP), which has previously been hypothesized as a mechanism for T2D.DiscussionWe validated prior studies that showed pathways related to metabolism, inflammation/immunity, and endocrine/hormone function are related to risk for T2D. We identified evidence for new potential mechanisms that include protein misfolding. Additional studies are needed to determine whether these are potential therapeutic targets to decrease risk for T2D

Eicosanoyl-5-hydroxytryptamide (EHT) prevents Alzheimer's disease-related cognitive and electrophysiological impairments in mice exposed to elevated concentrations of oligomeric beta-amyloid.

Soluble forms of oligomeric beta-amyloid (Aβ) are thought to play a central role in Alzheimer's disease (AD). Transgenic manipulation of methylation of the serine/threonine protein phosphatase, PP2A, was recently shown to alter the sensitivity of mice to AD-related impairments resulting from acute exposure to elevated levels of Aβ. In addition, eicosanoyl-5-hydroxytryptamide (EHT), a naturally occurring component from coffee beans that modulates PP2A methylation, was shown to confer therapeutic benefits in rodent models of AD and Parkinson's disease. Here, we tested the hypothesis that EHT protects animals from the pathological effects of exposure to elevated levels of soluble oligomeric Aβ. We treated mice with EHT-containing food at two different doses and assessed the sensitivity of these animals to Aβ-induced behavioral and electrophysiological impairments. We found that EHT administration protected animals from Aβ-induced cognitive impairments in both a radial-arm water maze and contextual fear conditioning task. We also found that both chronic and acute EHT administration prevented Aβ-induced impairments in long-term potentiation. These data add to the accumulating evidence suggesting that interventions with pharmacological agents, such as EHT, that target PP2A activity may be therapeutically beneficial for AD and other neurological conditions

A positive affect intervention alters leukocyte DNA methylation in sexual minority men with HIV who use methamphetamine

•Intervention-related decreases in methylation of β2 and oxytocin receptor genes.•Intervention effects on pathways for neural function and neurotransmitter release.•Accompanied by significant reductions in the self-reported frequency of stimulant use.This epigenomics sub-study embedded within a randomized controlled trial examined whether an evidenced-based behavioral intervention model that decreased stimulant use altered leukocyte DNA methylation (DNAm).Sexual minority men with HIV who use methamphetamine were randomized to a five-session positive affect intervention (n = 32) or an attention-control condition (n = 21), both delivered during three months of contingency management for stimulant abstinence. All participants exhibited sustained HIV virologic control – an HIV viral load less than 40 copies/mL at baseline and six months post-randomization. The Illumina EPIC BeadChip measured leukocyte methylation of cytosine-phosphate-guanosine (CpG) sites mapping onto five a priori candidate genes of interest (i.e., ADRB2, BDNF, FKBP5, NR3C1, OXTR). Functional DNAm pathways and soluble markers of immune dysfunction were secondary outcomes.Compared to the attention-control condition, the positive affect intervention significantly decreased methylation of CpG sites on genes that regulate β2 adrenergic and oxytocin receptors. There was an inconsistent pattern for the direction of the intervention effects on methylation of CpG sites on genes for glucocorticoid receptors and brain-derived neurotrophic factor. Pathway analyses adjusting for the false discovery rate (padj < 0.05) revealed significant intervention-related alterations in DNAm of Reactome pathways corresponding to neural function as well as dopamine, glutamate, and serotonin release. Positive affect intervention effects on DNAm were accompanied by significant reductions in the self-reported frequency of stimulant use.There is an epigenetic signature of an evidence-based behavioral intervention model that reduced stimulant use, which will guide the identification of biomarkers for treatment responses

Chronic EHT treatment prevents Aβ-induced impairment of long-term potentiation.

<p><b>(</b>A-B) Time course of averaged Schaffer collateral fEPSP responses (± SEM) in hippocampal slices prepared from animals fed control or EHT-containing diets and treated with either vehicle or 100 nM Aβ (horizontal bar) 20 min prior to delivery of theta-burst stimulation (arrow). (A) Aβ treatment significantly reduces potentiated responses following TBS in slices prepared from animals on control diets (2-way RM-ANOVA for treatment with time and treatment as factors: F(1,29) = 8.913, P = 0.0057). (B) Mice fed diets containing 0.01% EHT are resistant to Aβ-induced LTP impairment (2-way RM-ANOVA for treatment with time and treatment as factors: F(1,26) = 0.0943, P = 0.7612). C) Input/output (2-way RM-ANOVA for treatment with stimulus and treatment as factors: F(1,57) = 0.5466, P = 0.4628) (N = 31 control, 28 0.01% EHT).</p

EHT treatment reduces tau phosphorylation.

<p><b>(</b>A) Representative western blots for the indicated proteins and methylated PP2A/C performed on hippocampal homogenates prepared from animals fed control diet, or diets containing 0.01 or 0.1% EHT. (B) Histogram showing average ± SEM for tubulin-normalized band intensities expressed as average percent of control band intensity from replicate western blots in A show no significant differences in expression levels for any of the indicated proteins or for methylated PP2A/C (ANOVA for: PP2A/C: F(2,22) = 0.2533, P = 0.7784; PP2A/A: F(2,22) = 0.2588, P = 0.7743; B55α: F(2,22) = 0.06221, P = 0.9399; PME-1: F(2,22) = 0.4942, P = 0.6167; LCMT-1: F(2,22) = 0.2498, P = 0.7812; methyl-PP2A/C: F(2,22) = 0.1666, P = 0.8476). (C) Representative western blots for demethylated PP2A, and total PP2A/C performed on the homogenates described in A either treated (+) or mock treated (-) with 0.5 M sodium hydroxide. (D) Histogram of average demethylated PP2A/C (± SEM) in hippocampal homogenates prepared from animals fed control, or 0.01 or 0.1% EHT containing-diets show no significant differences in demethylated PP2A/C levels (ANOVA: F(2,22) = 0.1436, P = 0.8670). Values were calculated as ratios of demethyl-PP2A/C to total PP2A/C band intensities for -NaOH treated samples from replicate western blots shown in C and expressed as percent of the average of control. (E) Representative western blots performed on hippocampal homogenates prepared from animals fed control diet, or diets containing 0.01 or 0.1% EHT for phospho-Ser396/404 (PHF1), phospho-Ser202 (CP13) together with their corresponding total tau loading controls, as well as and total tau together with its corresponding β-actin loading control. (F) Histogram showing average band intensities ± SEM for phospho-Ser396/404-tau (PHF1), phospho-Ser202-tau (CP13) normalized to corresponding total tau loading control, and total tau normalized to corresponding β-actin loading control for replicate western blots shown in H show a trend for reduced phosphorylation at these sites in EHT-treated animals (ANOVA for: PHF1: F(2,22) = 5.147, P = 0.147, Bonferroni post-hoc for PHF1 0.1% EHT vs. Control: t = 3.154, P = 0.0092; CP13: F(2,22) = 1.433, P = 0.2599; total tau: F(2,22) = 0.1268, P = 0.8815). (G and H) Representative western blots for phospho-Ser9 and total GSK3B and phospho-Ser133 and total Creb performed on hippocampal homogenates prepared from animals fed control diet, or diets containing 0.01 or 0.1% EHT. (I) Histogram showing average ± SEM of phospho-GSK3β and phospho-Creb band intensities normalized to corresponding total GSK3β and Creb respectively for replicate western blots shown in G&H show no significant effect of EHT treatment on phosphorylation at these sites (ANOVA for: P-GSK3β F(2,22) = 1.761, P = 0.1952; P-Creb: F(2,22) = 0.1776, P = 0.8385). (N = 8 control, 8 0.01% EHT and 9 0.1% EHT treated animals for each measure).</p