Replacement of dietary saturated fat with unsaturated fats increases numbers of circulating endothelial progenitor cells and decreases number of microparticles: findings from the randomized, controlled DIVAS study

Background Endothelial progenitor cells (EPC) and microparticles (MP) are emerging novel markers of cardiovascular disease (CVD) risk, which could potentially be modified by dietary fat. We have previously shown that replacing dietary saturated fat (SFA) with monounsaturated (MUFA) or n-6 polyunsaturated fat (PUFA) improved lipid biomarkers, blood pressure and markers of endothelial activation, but their effects on circulating EPCs and MPs are unclear. Objective The Dietary Intervention and VAScular function (DIVAS) study investigated the replacement of 9.5-9.6% total energy (%TE) SFA with MUFA or n-6 PUFA for 16 weeks on EPC and MP numbers in UK adults with moderate CVD risk. Design In this randomized, controlled, single-blind, parallel group dietary intervention, men and women aged 21-60 y (n=190) with moderate CVD risk (≥50% above the population mean) consumed one of three 16-week isoenergetic diets. Target compositions for total fat, SFA, MUFA and n-6 PUFA (%TE) were: SFA-rich diet (36:17:11:4, n=64), MUFA-rich diet (36:9:19:4, n=62) and n-6 PUFA-rich diet (36:9:13:10, n=66). Circulating EPC, endothelial MP (EMP) and platelet MP (PMP) numbers were analysed by flow cytometry. Dietary intake, vascular function and other cardio-metabolic risk factors were determined at baseline. Results Relative to the SFA-rich diet, MUFA and n-6 PUFA-rich diets decreased EMP (-47.3%, -44.9%) and PMP numbers (-36.8%, -39.1%) (overall diet effects P<0.01). The MUFA-rich diet increased EPC numbers (+28.4%; P=0.023). Additional analyses using stepwise regression models identified the augmentation index (measuring arterial stiffness determined by pulse wave analysis) as an independent predictor of baseline EPC and MP numbers. Conclusions Replacing 9.5-9.6%TE dietary SFA with MUFA increased EPC numbers and replacement with either MUFA or n-6 PUFA decreased MP numbers, suggesting beneficial effects on endothelial repair and maintenance. Further studies are warranted to determine the mechanisms underlying the favourable effects on EPC and MP numbers following SFA replacement

Loss of DPP6 in neurodegenerative dementia : a genetic player in the dysfunction of neuronal excitability

Emerging evidence suggested a converging mechanism in neurodegenerative brain diseases (NBD) involving early neuronal network dysfunctions and alterations in the homeostasis of neuronal firing as culprits of neurodegeneration. In this study, we used paired-end short-read and direct long-read whole genome sequencing to investigate an unresolved autosomal dominant dementia family significantly linked to 7q36. We identified and validated a chromosomal inversion of ca. 4Mb, segregating on the disease haplotype and disrupting the coding sequence of dipeptidyl-peptidase 6 gene (DPP6). DPP6 resequencing identified significantly more rare variants-nonsense, frame-shift, and missense-in early-onset Alzheimer's disease (EOAD, p value = 0.03, OR = 2.21 95% CI 1.05-4.82) and frontotemporal dementia (FTD, p = 0.006, OR = 2.59, 95% CI 1.28-5.49) patient cohorts. DPP6 is a type II transmembrane protein with a highly structured extracellular domain and is mainly expressed in brain, where it binds to the potassium channel K(v)4.2 enhancing its expression, regulating its gating properties and controlling the dendritic excitability of hippocampal neurons. Using in vitro modeling, we showed that the missense variants found in patients destabilize DPP6 and reduce its membrane expression (p < 0.001 and p < 0.0001) leading to a loss of protein. Reduced DPP6 and/or K(v)4.2 expression was also detected in brain tissue of missense variant carriers. Loss of DPP6 is known to cause neuronal hyperexcitability and behavioral alterations in Dpp6-KO mice. Taken together, the results of our genomic, genetic, expression and modeling analyses, provided direct evidence supporting the involvement of DPP6 loss in dementia. We propose that loss of function variants have a higher penetrance and disease impact, whereas the missense variants have a variable risk contribution to disease that can vary from high to low penetrance. Our findings of DPP6, as novel gene in dementia, strengthen the involvement of neuronal hyperexcitability and alteration in the homeostasis of neuronal firing as a disease mechanism to further investigate

Investigation of the role of miRNA variants in neurodegenerative brain diseases

IntroductionmiRNAs are small noncoding elements known to regulate different molecular processes, including developmental and executive functions in the brain. Dysregulation of miRNAs could contribute to brain neurodegeneration, as suggested by miRNA profiling studies of individuals suffering from neurodegenerative brain diseases (NBDs). Here, we report rare miRNA variants in patients with Alzheimer’s dementia (AD) and frontotemporal dementia (FTD).MethodsWe initially used whole exome sequencing data in a subset of FTD patients (n = 209) from Flanders-Belgium. We then performed targeted resequencing of variant-harboring miRNAs in an additional subset of FTD patients (n = 126) and control individuals (n = 426). Lastly, we sequenced the MIR885 locus in a Flanders-Belgian AD cohort (n = 947) and a total number of n = 755 controls.ResultsWES identified rare seed variants in MIR656, MIR423, MIR122 and MIR885 in FTD patients. Most of these miRNAs bind to FTD-associated genes, implicated in different biological pathways. Additionally, some miRNA variants create novel binding sites for genes associated with FTD. Sequencing of the MIR885 locus in the AD cohort initially showed a significant enrichment of MIR885 variants in AD patients compared to controls (SKAT-O, p-value = 0.026). Genetic association was not maintained when we included sex and APOE status as covariates. Using the miRVaS prediction tool, variants rs897551430 and rs993255773 appeared to evoke significant structural changes in the primary miRNA. These variants are also predicted to strongly downregulate mature miR885 levels, in line with what is reported for MIR885 in the context of AD.DiscussionFunctional investigation of miRNAs/variants described in this study could propose novel miRNA-mediated molecular cascades in FTD and AD pathogenicity. Furthermore, we believe that the genetic evidence presented here suggests a role for MIR885 in molecular mechanisms involved in AD and warrants genetic follow-up in larger cohorts to explore this hypothesis

Rare mutations in SQSTM1 modify susceptibility to frontotemporal lobar degeneration

Mutations in the gene coding for Sequestosome 1 (SQSTM1) have been genetically associated with amyotrophic lateral sclerosis (ALS) and Paget disease of bone. In the present study, we analyzed the SQSTM1 coding sequence for mutations in an extended cohort of 1,808 patients with frontotemporal lobar degeneration (FTLD), ascertained within the European Early-Onset Dementia consortium. As control dataset, we sequenced 1,625 European control individuals and analyzed whole-exome sequence data of 2,274 German individuals (total n = 3,899). Association of rare SQSTM1 mutations was calculated in a meta-analysis of 4,332 FTLD and 10,240 control alleles. We identified 25 coding variants in FTLD patients of which 10 have not been described. Fifteen mutations were absent in the control individuals (carrier frequency < 0.00026) whilst the others were rare in both patients and control individuals. When pooling all variants with a minor allele frequency < 0.01, an overall frequency of 3.2 % was calculated in patients. Rare variant association analysis between patients and controls showed no difference over the whole protein, but suggested that rare mutations clustering in the UBA domain of SQSTM1 may influence disease susceptibility by doubling the risk for FTLD (RR = 2.18 [95 % CI 1.24-3.85]; corrected p value = 0.042). Detailed histopathology demonstrated that mutations in SQSTM1 associate with widespread neuronal and glial phospho-TDP-43 pathology. With this study, we provide further evidence for a putative role of rare mutations in SQSTM1 in the genetic etiology of FTLD and showed that, comparable to other FTLD/ALS genes, SQSTM1 mutations are associated with TDP-43 pathology

TMEM106B is a genetic modifier of frontotemporal lobar degeneration with C9orf72 hexanucleotide repeat expansions

Hexanucleotide repeat expansions in chromosome 9 open reading frame 72 (C9orf72) have recently been linked to frontotemporal lobar degeneration (FTLD) and amyotrophic lateral sclerosis, and may be the most common genetic cause of both neurodegenerative diseases. Genetic variants at TMEM106B influence risk for the most common neuropathological subtype of FTLD, characterized by inclusions of TAR DNA-binding protein of 43 kDa (FTLD-TDP). Previous reports have shown that TMEM106B is a genetic modifier of FTLD-TDP caused by progranulin (GRN) mutations, with the major (risk) allele of rs1990622 associating with earlier age at onset of disease. Here, we report that rs1990622 genotype affects age at death in a single-site discovery cohort of FTLD patients with C9orf72 expansions (n = 14), with the major allele correlated with later age at death (p = 0.024). We replicate this modifier effect in a 30-site international neuropathological cohort of FTLD-TDP patients with C9orf72 expansions (n = 75), again finding that the major allele associates with later age at death (p = 0.016), as well as later age at onset (p = 0.019). In contrast, TMEM106B genotype does not affect age at onset or death in 241 FTLD-TDP cases negative for GRN mutations or C9orf72 expansions. Thus, TMEM106B is a genetic modifier of FTLD with C9orf72 expansions. Intriguingly, the genotype that confers increased risk for developing FTLD-TDP (major, or T, allele of rs1990622) is associated with later age at onset and death in C9orf72 expansion carriers, providing an example of sign epistasis in human neurodegenerative disease

Protein interaction network analysis reveals genetic enrichment of immune system genes in frontotemporal dementia

To further unravel the complex genetic etiology of frontotemporal dementia (FTD), we hypothesized that interactors of the protein products of known FTD genes might be involved in the molecular pathways towards disease. We therefore applied protein interaction network (PIN) analysis to prioritize candidate genes for rare variant association. We created an FTD-PIN starting from known FTD genes downloading their physical interactors and performed functional enrichment analyses. We identified overrepresented processes in FTD and selected genes (n=440) belonging to the FTD processes for rare variant analysis in a Belgian cohort of 228 FTD patients and 345 controls. SKAT-O analysis suggested TNFAIP3 as the top gene (P = 0.7 × 10−3) reaching near test-wide significance (P = 2.5 × 10−4). We then analyzed the TNFAIP3-subnetwork within the FTD-PIN which indicated enrichment of several immune signaling networks, suggesting that disrupted immune signaling may be implicated in TNFAIP3-related FTD. Our study demonstrates that integration of PINs with genetic data is a useful approach to increase the power for rare variant association analysis. Furthermore, we present a computational pipeline for identifying potential novel therapeutic targets and risk-modifying variants

Promoter DNA methylation regulates progranulin expression and is altered in FTLD

BACKGROUND: Frontotemporal lobar degeneration (FTLD) is a heterogeneous group of neurodegenerative diseases associated with personality changes and progressive dementia. Loss-of-function mutations in the growth factor progranulin (GRN) cause autosomal dominant FTLD, but so far the pathomechanism of sporadic FTLD is unclear. RESULTS: We analyzed whether DNA methylation in the GRN core promoter restricts GRN expression and, thus, might promote FTLD in the absence of GRN mutations. GRN expression in human lymphoblast cell lines is negatively correlated with methylation at several CpG units within the GRN promoter. Chronic treatment with the DNA methyltransferase inhibitor 5-aza-2(′)-deoxycytidine (DAC) strongly induces GRN mRNA and protein levels. In a reporter assay, CpG methylation blocks transcriptional activity of the GRN core promoter. In brains of FTLD patients several CpG units in the GRN promoter are significantly hypermethylated compared to age-matched healthy controls, Alzheimer and Parkinson patients. These CpG motifs are critical for GRN promoter activity in reporter assays. Furthermore, DNA methyltransferase 3a (DNMT3a) is upregulated in FTLD patients and overexpression of DNMT3a reduces GRN promoter activity and expression. CONCLUSION: These data suggest that altered DNA methylation is a novel pathomechanism for FTLD that is potentially amenable to targeted pharmacotherapy

Frontotemporal dementia and its subtypes: a genome-wide association study

SummaryBackground Frontotemporal dementia (FTD) is a complex disorder characterised by a broad range of clinical manifestations, differential pathological signatures, and genetic variability. Mutations in three genes—MAPT, GRN, and C9orf72—have been associated with FTD. We sought to identify novel genetic risk loci associated with the disorder. Methods We did a two-stage genome-wide association study on clinical FTD, analysing samples from 3526 patients with {FTD} and 9402 healthy controls. To reduce genetic heterogeneity, all participants were of European ancestry. In the discovery phase (samples from 2154 patients with {FTD} and 4308 controls), we did separate association analyses for each {FTD} subtype (behavioural variant FTD, semantic dementia, progressive non-fluent aphasia, and {FTD} overlapping with motor neuron disease FTD-MND), followed by a meta-analysis of the entire dataset. We carried forward replication of the novel suggestive loci in an independent sample series (samples from 1372 patients and 5094 controls) and then did joint phase and brain expression and methylation quantitative trait loci analyses for the associated (p&lt;5 × 10−8) single-nucleotide polymorphisms. Findings We identified novel associations exceeding the genome-wide significance threshold (p&lt;5 × 10−8). Combined (joint) analyses of discovery and replication phases showed genome-wide significant association at 6p21.3, \{HLA\} locus (immune system), for rs9268877 (p=1·05 × 10−8; odds ratio=1·204 95% \{CI\} 1·11–1·30), rs9268856 (p=5·51 × 10−9; 0·809 0·76–0·86) and rs1980493 (p value=1·57 × 10−8, 0·775 0·69–0·86) in the entire cohort. We also identified a potential novel locus at 11q14, encompassing RAB38/CTSC (the transcripts of which are related to lysosomal biology), for the behavioural \{FTD\} subtype for which joint analyses showed suggestive association for rs302668 (p=2·44 × 10−7; 0·814 0·71–0·92). Analysis of expression and methylation quantitative trait loci data suggested that these loci might affect expression and methylation in cis. Interpretation Our findings suggest that immune system processes (link to 6p21.3) and possibly lysosomal and autophagy pathways (link to 11q14) are potentially involved in FTD. Our findings need to be replicated to better define the association of the newly identified loci with disease and to shed light on the pathomechanisms contributing to FTD. Funding The National Institute of Neurological Disorders and Stroke and National Institute on Aging, the Wellcome/MRC Centre on Parkinson's disease, Alzheimer's Research UK, and Texas Tech University Health Sciences Center