Apo Phenotypes Research Articles

High-Throughput Microbore LC-MS Lipidomics to Investigate APOE Phenotypes.

Microflow liquid chromatography interfaced with mass spectrometry (μLC-MS/MS) is increasingly applied for high-throughput profiling of biological samples and has been proven to have an acceptable trade-off between sensitivity and reproducibility. However, lipidomics applications are scarce. We optimized a μLC-MS/MS system utilizing a 1 mm inner diameter × 100 mm column coupled to a triple quadrupole mass spectrometer to establish a sensitive, high-throughput, and robust single-shot lipidomics workflow. Compared to conventional lipidomics methods, we achieve a ∼4-fold increase in response, facilitating quantification of 351 lipid species from a single iPSC-derived cerebral organoid during a 15 min LC-MS analysis. Consecutively, we injected 303 samples over ∼75 h to prove the robustness and reproducibility of the microflow separation. As a proof of concept, μLC-MS/MS analysis of Alzheimer's disease patient-derived iPSC cerebral organoid reveals differential lipid metabolism depending on APOE phenotype (E3/3 vs E4/4). Microflow separation proves to be an environmentally friendly and cost-effective method as it reduces the consumption of harmful solvents. Also, the data demonstrate robust, in-depth, high-throughput performance to enable routine clinical or biomedical applications.

Manipulation of Dietary DHA does not Alter DHA Brain Uptake Kinetics in Mice Despite Changing Brain and Plasma DHA Levelsvs

AbstractBackgroundIncreased brain docosahexaenoic acid (DHA) uptake reflects a compensatory mechanism for maintaining brain DHA homeostasis and can be captured through PET‐MRI scan uptake studies using radiolabeled DHA. It has been previously shown that APOE4 status and alcohol intake alter brain DHA uptake in mice. Whether a change in DHA intake affects the brain DHA incorporation coefficient (K*) however is not known.MethodFour‐month‐old C57BL/6J mice were fed either a diet rich in DHA (15 kcal% Fat, 1.7 g/kg ALA, and 7g/kg DHA) or deficient in DHA (15 kcal% Fat and 0.9 mg/kg ALA) for 4 months. At 8 months, anesthetized mice were subjected to dynamic PET‐MRI for 30 min following i.v. injection of 22‐[18F]fluorodocosahexaenoic acid ([18F]FDHA), prepared in‐house through radiosynthesis. Using an image‐derived cardiac input function, brain uptake was assessed by estimating [18F]FDHA K* values using both the Patlak model approach and the irreversible two‐tissue compartmental model (Irr2TCM) with and without correction of the brain radioactivity for radioactive spillover from the skull. Mice were then sacrificed, and the plasma and brains were analyzed using liquid chromatography‐mass spectrometry (LC‐MS) to evaluate differences in the unlabeled concentrations of fatty acids into phospholipids, triglycerides, and other lipids.ResultThere was no significant difference in the [18F]FDHA K* estimates (with either Patlak or Irr2TCM) between mice fed a DHA‐rich or a DHA‐deficient diet. LC‐MS based lipidomic analysis showed that the DHA‐rich diet significantly increased DHA concentration in triglycerides in plasma and in phosphatidylethanolamine in the brain.ConclusionThese results suggest that under physiological conditions, increased dietary intake of DHA in wild‐type mice increases DHA in brain phosphatidylethanolamine and in plasma triglycerides without changing the DHA K*. The net effect likely is increased brain DHA uptake, equal to the product of unesterified DHA plasma concentration and K*. Imaging brain DHA with dynamic [18F]FDHA PET‐MRI is a potentially useful tool not only to investigate the APOE4 phenotype, which promotes DHA dysregulation, but also to assess dietary and pharmacological interventions that affect brain DHA homeostasis as well as to guide future treatments.

EEG‐based neural biomarker for diagnosis and assessment of mild cognitive impairment and Alzheimer’s disease

AbstractBackgroundElectroencephalogram (EEG) has emerged as a non‐invasive tool to detect the aberrant neuronal activity related to different stages of Alzheimer’s disease (AD). However, the effectiveness of EEG in the precise diagnosis and prediction of AD and its preclinical stage, mild cognitive impairment (MCI), has yet to be fully elucidated. In this study, we aimed to identify key EEG biomarkers that are effective in distinguishing patients at the early stage of AD and monitoring the progression of AD.MethodA total of 890 participants, including 189 patients with MCI, 330 patients with AD, 125 patients with other dementias (frontotemporal dementia, dementia with Lewy bodies, and vascular cognitive impairment), and 246 healthy controls (HC) were enrolled. Biomarkers were extracted from resting‐state EEG recordings for a three‐level classification of HC, MCI, and AD. The optimal EEG biomarkers were then identified based on the classification performance. Random forest regression was used to train a series of predictive models by combining participants’ EEG biomarkers, demographic information (i.e., sex, age), CSF biomarkers, and APOE phenotype for predicting the disease progression and individual’s cognitive function.ResultThe identified EEG biomarkers achieved over 70% accuracy in the three‐level classification of HC, MCI, and AD. Among all six groups, the most prominent effects of AD‐linked neurodegeneration on EEG metrics were localized at parieto‐occipital regions. Noteworthy, the absolute theta power of channel O2 was negatively correlated with Aβ42 (r = ‐0.358, p < 0.001) and positively correlated with p‐tau (r = 0.442, p < 0.001). In terms of cognitive assessment, the Hjorth mobility of channel O1, O2 and P4 were found to be positively associated with MMSE and MoCA scores (r = 0.416‐0.464, all p < 0.001). In the predictive analyses, the optimal EEG features were more effective than the CSF + APOE biomarkers in predicting the age of onset and disease course, whereas the combination of EEG + CSF + APOE measures achieved the best performance for all targets of prediction.ConclusionOur study indicates that EEG can be used as a useful screening tool for the diagnosis and disease progression evaluation of MCI and AD.

Clinical heterogeneity of neuro-inflammatory PET profiles in early Alzheimer's disease.

The relationship between neuroinflammation and cognition remains uncertain in early Alzheimer's disease (AD). We performed a cross-sectional study to assess how neuroinflammation is related to cognition using TSPO PET imaging and a multi-domain neuropsychological assessment. A standard uptake value ratio (SUVR) analysis was performed to measure [18F]-DPA-714 binding using the cerebellar cortex or the whole brain as a (pseudo)reference region. Among 29 patients with early AD, the pattern of neuroinflammation was heterogeneous and exhibited no correlation with cognition at voxel-wise, regional or whole-brain level. The distribution of the SUVR values was independent of sex, APOE phenotype, early and late onset of symptoms and the presence of cerebral amyloid angiopathy. However, we were able to demonstrate a complex dissociation as some patients with similar PET pattern had opposed neuropsychological profiles while other patients with opposite PET profiles had similar neuropsychological presentation. Further studies are needed to explore how this heterogeneity impacts disease progression.

Neural biomarker diagnosis and prediction to mild cognitive impairment and Alzheimer’s disease using EEG technology

BackgroundElectroencephalogram (EEG) has emerged as a non-invasive tool to detect the aberrant neuronal activity related to different stages of Alzheimer’s disease (AD). However, the effectiveness of EEG in the precise diagnosis and assessment of AD and its preclinical stage, amnestic mild cognitive impairment (MCI), has yet to be fully elucidated. In this study, we aimed to identify key EEG biomarkers that are effective in distinguishing patients at the early stage of AD and monitoring the progression of AD.MethodsA total of 890 participants, including 189 patients with MCI, 330 patients with AD, 125 patients with other dementias (frontotemporal dementia, dementia with Lewy bodies, and vascular cognitive impairment), and 246 healthy controls (HC) were enrolled. Biomarkers were extracted from resting-state EEG recordings for a three-level classification of HC, MCI, and AD. The optimal EEG biomarkers were then identified based on the classification performance. Random forest regression was used to train a series of models by combining participants’ EEG biomarkers, demographic information (i.e., sex, age), CSF biomarkers, and APOE phenotype for assessing the disease progression and individual’s cognitive function.ResultsThe identified EEG biomarkers achieved over 70% accuracy in the three-level classification of HC, MCI, and AD. Among all six groups, the most prominent effects of AD-linked neurodegeneration on EEG metrics were localized at parieto-occipital regions. In the cross-validation predictive analyses, the optimal EEG features were more effective than the CSF + APOE biomarkers in predicting the age of onset and disease course, whereas the combination of EEG + CSF + APOE measures achieved the best performance for all targets of prediction.ConclusionsOur study indicates that EEG can be used as a useful screening tool for the diagnosis and disease progression evaluation of MCI and AD.

Walking pace, handgrip strength, age, APOE genotypes, and new-onset dementia: the UK Biobank prospective cohort study

BackgroundThe independent and additive associations of walking pace and grip strength on dementia risk and the potential modifying effects of age, APOE phenotypes, and other dementia risk factors on the walking pace and dementia relationships demand further clarification. We aimed to investigate the independent and additive relationships of walking pace and handgrip strength on the risk of new-onset dementia and examine the potentially modifying effects of age, APOE phenotypes, lifestyle factors, and family history of dementia in the relationships.MethodsA total of 495,700 participants from the UK Biobank, who were free of dementia at baseline, were included in this study. Walking pace was self-defined as slow, average, or brisk. Handgrip strength was assessed by dynamometer and was divided into sex-specific quartiles. The APOE genotypes were determined by a combination variant of rs429358 and rs7412. Other dementia risk factors, including education, physical activity, hypertension, depression, diabetes, and family history of dementia, were also collected. The primary outcome was new-onset all-cause dementia.ResultsOver a median follow-up duration of 12.0 years, 3986 (0.8%) participants developed new-onset all-cause dementia. Compared with those with slow walking pace, participants with average (HR, 0.61; 95%CI: 0.55–0.68) or brisk (HR, 0.59; 95%CI: 0.52–0.67) walking pace had a significantly lower risk of new-onset all-cause dementia. Moreover, compared with those with both slow walking pace and lower handgrip strength (the first quartile), the lowest risk of new-onset all-cause dementia was observed in participants with both average or brisk walking pace and higher handgrip strength (the 2–4 quartiles) (HR, 0.45; 95%CI: 0.40–0.52). Notably, the negative relationship between walking pace and the risk of new-onset all-cause dementia was significantly reduced as APOE ε4 dosage increased (APOE ε4 dosages = 0 or 1: brisk vs. slow: HR, 0.55; 95%CI: 0.48–0.63; vs. APOE ε4 dosages = 2: brisk vs. slow: HR, 1.14; 95%CI: 0.77–1.68; P for interaction = 0.001) or age increased (< 58 [median]: brisk vs. slow: HR, 0.27; 95%CI: 0.18–0.41; vs. ≥ 58 years: brisk vs. slow: HR, 0.55; 95%CI: 0.48–0.63; P for interaction = 0.007).ConclusionsWalking pace was inversely associated with new-onset dementia in the general population, especially in younger participants and those with lower APOE ε4 dosage. Participants with both faster walking pace and higher handgrip strength had the lowest risk of dementia, suggesting that maintaining both high handgrip strength and fast walking pace may be a more comprehensive strategy for preventing dementia risk.

Simultaneous Mass Spectrometry-Based Apolipoprotein Profiling and Apolipoprotein E Phenotyping in Patients with ASCVD and Mild Cognitive Impairment.

Apolipoprotein E (apoE) occurs on the majority of plasma lipoproteins and plays a major role in the lipid metabolism in the periphery and in the central nervous system. ApoE is a polymorphic protein with three common isoforms, apoE2, apoE3 and apoE4, derived from respective alleles ε2, ε3 and ε4. The aim of this study was to develop a sample pretreatment protocol combined with rapid mass spectrometry (MS)-based assay for simultaneous apolipoprotein profiling and apoE phenotype identification. This assay was validated in 481 samples from patients with stable atherosclerotic cardiovascular disease (ASCVD) and applied to study association with mild cognitive impairment (MCI) in the LIFE Adult study, including overall 690 study subjects. Simultaneous quantification of 8–12 major apolipoproteins including apoA-I, apoB-100 and apoE could be performed within 6.5 min. Phenotyping determined with the developed MS assay had good agreement with the genotyping by real-time fluorescence PCR (97.5%). ApoE2 isoform was associated with the highest total apoE concentration compared to apoE3 and apoE4 (p < 0.001). In the subgroup of diabetic atherosclerotic cardiovascular disease (ASCVD) patients, apoE2 isoform was related to higher apoC-I levels (apoE2 vs. apoE3, p < 0.05), while in the subgroup of ASCVD patients under statin therapy apoE2 was related to lower apoB-100 levels (apoE2 vs. apoE3/apoE4, p < 0.05). A significant difference in apoE concentration observed between mild cognitive impairment (MCI) subjects and controls was confirmed for each apoE phenotype. In conclusion, this study provides evidence for the successful implementation of an MS-based apoE phenotyping assay, which can be used to assess phenotype effects on plasma lipid and apolipoprotein levels.

Egg and cholesterol intake, apolipoprotein E4 phenotype and risk of venous thromboembolism: findings from a prospective cohort study.

The impact of egg consumption, a major source of dietary cholesterol, with the risk of atherosclerotic cardiovascular diseases (ASCVDs) is controversial. Venous thromboembolism (VTE) is a CVD which shares common risk factors and mechanistic pathways with ASCVD. However, there is no data on the relationship between egg or cholesterol intake and VTE risk. Therefore, we evaluated the prospective associations of egg and cholesterol intakes with VTE risk and whether the apolipoprotein E4 (apoE4) phenotype, which influences cholesterol metabolism, could modify the associations. Data involving 1,852 men aged 42-61 years at baseline without a history of VTE or coronary heart disease in the population-based Kuopio Ischaemic Heart Disease Risk Factor Study were analysed. Dietary intakes were assessed with 4-day food records. Incident VTE events were identified by record linkage to hospital discharge registries. Hazard ratios (95% confidence intervals [CIs]) for incident VTE were estimated using Cox regression. During a median follow-up of 28.8 years, 132 VTE events occurred. Comparing the top (>38 g/d) versus bottom (<20 g/d) tertiles of egg consumption, the HR (95% CI) for VTE was 0.99 (0.64-1.53) in analysis adjusted for several established risk factors and other dietary factors. There was also no evidence of an association between cholesterol intake and VTE risk. Imputed results were consistent with the observed results. The apoE4 phenotype did not modify the associations. In middle-aged and older Finnish men, egg or cholesterol intakes were not associated with future VTE risk. Other large-scale prospective studies are needed to confirm or refute these findings.

Low Molecular Weight Apolipoprotein(a) Phenotype Rather Than Lipoprotein(a) Is Associated With Coronary Atherosclerosis and Myocardial Infarction.

Background and AimsCurrent evidence suggests that lipoprotein(a) [Lp(a)] level above 50 mg/dL is associated with increased cardiovascular risk. Our study aim was to determine the relationship of apolipoprotein(a) [apo(a)] phenotypes and Lp(a) concentration below and above 50 mg/dL with coronary atherosclerosis severity and myocardial infarction (MI).Material and MethodsThe study population consisted of 540 patients (mean age 54.0 ± 8.8 years, 82% men) who passed through coronary angiography. The number of diseased major coronary arteries assessed atherosclerosis severity. Lipids, glucose, Lp(a) levels and apo(a) phenotypes were determined in all patients. All patients were divided into four groups: with Lp(a) <50 mg/dL [ “normal” Lp(a)] or ≥50 mg/dL [hyperLp(a)], and with low-molecular (LMW) or high-molecular weight (HMW) apo(a) phenotypes.ResultsBaseline clinical and biochemical characteristics were similar between the groups. In groups with LMW apo(a) phenotypes, the odds ratio (OR; 95% confidence interval) of multivessel disease was higher [10.1; 3.1–33.5, p < 0.005 for hyperLp(a) and 2.2; 1.0–4.9, p = 0.056 for normal Lp(a)], but not in the group with HMW apo(a) and hyperLp(a) [1.1; 0.3–3.3, p = 0.92] compared with the reference group with HMW apo(a) and normal Lp(a). Similarly, MI was observed more often in patients with LMW apo(a) phenotype and hyperLp(a) and normal Lp(a) than in groups with HMW apo(a) phenotype.ConclusionThe LMW apo(a) phenotype is associated with the severity of coronary atherosclerosis and MI even when Lp(a) level is below 50 mg/dL. The combination of Lp(a) level above 50 mg/dL and LMW apo(a) phenotype increases the risk of severe coronary atherosclerosis, regardless of other risk factors.

Polymorphism of apolipoprotein E in children with metabolic diseases of the biliary tract

Background: cholelithiasis in children is associated with a violation of cholesterol metabolism and insulin resistance. Objective: to study the correlation between Apo-E subclasses and lipid spectrum parameters in children with cholelithiasis. Methods: The study included patients with cholelithiasis: stage I (n = 23) and stage II (n = 21). The lipid profile was assessed in patients with different Apo-E phenotypes. Results: most patients had E3 / E3 (n = 32) and E3 allele (n = 36), rarely E3 / E4 (n = 8) and E4 / E4 (n = 4) and allele E4 (n = 8). LDL levels were significantly different in patients with phenotypes E3 / E4 and E4 / E4, pk-w <0.01. Higher LDL values were typical for children with the E4 / E4 phenotype (3.42 ± 0.34 mmol / L) compared to the E3 / E4 phenotype (2.0 ± 0.25 mmol / L, pu <0.05). HDL levels in children of both groups were within the reference values, the lowest values were recorded in patients with the E4 allele. At different stages of cholelithiasis in children, the presence of the E4 / E4 phenotype in combination with dyslipidemia can be considered a significant factor in connection with an increase in the proportion of atherogenic lipids. The data obtained require further in-depth research. Conclusion: Analysis of the lipid spectrum in children with cholelithiasis and various phenotypes of Apo-E indicates a genetic predisposition to lipid metabolism disorders. The marker of an increased risk of developing cholelithiasis is the E4 / E4 phenotype.

Associations of dairy, meat, and fish intakes with risk of incident dementia and with cognitive performance: the Kuopio Ischaemic Heart Disease Risk Factor Study (KIHD)

PurposeTo investigate if dairy, meat, and fish intakes associate with dementia and cognitive performance.MethodsWe included 2497 dementia-free men from Eastern Finland, aged 42–60 years in 1984–1989 at the baseline examinations. Data on cognitive tests [Mini Mental State Exam (MMSE), trail making test (TMT), verbal fluency test (VFL), selective reminding test (SRT), and Russell’s adaptation of the visual reproduction test (VRT)] at the 4-year re-examinations were available for 482 men and on the ApoE phenotype for 1259 men. Data on dementia events were obtained by linkage to national health registers. Diet was assessed with baseline 4-day food records. Cox regression and analysis of covariance were used for analyses.ResultsDuring a mean 22-year follow-up, 337 men had a dementia diagnosis. Among the foods, only cheese intake associated with dementia risk (hazard ratio in the highest vs. the lowest quartile = 0.72, 95% confidence interval = 0.52–0.99, P-trend = 0.05). In the cognitive tests, higher non-fermented dairy and milk intakes associated with worse verbal fluency (VFT). Higher processed red meat intake associated with worse verbal (SRT) and visual memory (VRT), whereas higher unprocessed red meat intake associated with better general cognitive functioning (MMSE) and processing speed and executive functioning (TMT). Higher fish intake associated with better verbal memory (SRT). Among APOE-ε4 carriers, especially non-fermented dairy intake associated with higher risk of dementia outcomes, and higher fish intake indicated better cognitive performance.ConclusionAlthough higher intake of some food groups associated with cognitive performance, we found little evidence for associations with dementia risk.

Dysbetalipoproteinemia: Differentiating Multifactorial Remnant Cholesterol Disease From Genetic ApoE Deficiency.

Dysbetalipoproteinemia (DBL) is characterized by the accumulation of remnant lipoprotein particles and associated with an increased risk of cardiovascular and peripheral vascular disease (PVD). DBL is thought to be mainly caused by the presence of an E2/E2 genotype of the apolipoprotein E (APOE) gene, in addition to environmental factors. However, there exists considerable phenotypic variability among DBL patients. The objectives were to verify the proportion of DBL subjects, diagnosed using the gold standard Fredrickson criteria, who did not carry E2/E2 and to compare the clinical characteristics of DBL patients with and without E2/E2. A total of 12 432 patients with lipoprotein ultracentrifugation as well as APOE genotype or apoE phenotype data were included in this retrospective study. Among the 12 432 patients, 4% (n = 524) were positive for Fredrickson criteria (F+), and only 38% (n = 197) of the F+ individuals were E2/E2. The F+ E2/E2 group had significantly higher remnant cholesterol concentration (3.44 vs 1.89 mmol/L) and had higher frequency of DBL-related xanthomas (24% vs 2%) and floating beta (95% vs 11%) than the F+ non-E2/E2 group (P < 0.0001). The F+ E2/E2 group had an independent higher risk of PVD (OR 11.12 [95% CI 1.87-66.05]; P = 0.008) events compared with the F+ non-E2/E2 group. In the largest cohort of DBL worldwide, we demonstrated that the presence of E2/E2 was associated with a more severe DBL phenotype. We suggest that 2 DBL phenotypes should be distinguished: the multifactorial remnant cholesterol disease and the genetic apoE deficiency disease.

Identification of APOE phenotypes in microglia and neurons derived from isogenic hips cells: A study of the IMI ADAPTED consortium

AbstractBackgroundOne of the most prominent genetic risk factors for Alzheimer’s Disease (AD) is the Apolipoprotein E (APOE) ε4 allele. Unlike the “neutral” ε3 variant, or the protective ε2variant, the ε4 variant is strongly associated with late‐onset AD, albeit by mechanisms that are not well‐understood.MethodsTo shed light on the biology of APOE in the development of AD, as part of the IMI (Innovative Medicines Initiative) consortium “ADAPTED”, we acquired two human induced pluripotent stem cell (hiPSC) lines, that were isogenically modified to carry the alleles ε2/ε2, ε3/ε3, ε3/ε4, ε4/ε4, as well as a knockout line. Neurons and the phagocytic cells of the CNS, the microglia were differentiated from all ten lines and subjected to thorough phenotyping, including RNA‐seq and proteomics analysis.ResultsCortical neurons do not secrete apoE under normal conditions and treatment of these cells with microglia‐enriched factors and apoE increased their overall Abeta secretion, without significant differences between the various genotypes. Microglia, on the other hand, secrete large amounts of apoE in varying quantities depending on the APOE genotype. Additionally, microglia carrying the ε4/ε4 genotype have a significantly reduced phagocytic capacity compared to their ε3/ε3 counterparts. These observations were supported by whole transcriptome sequencing, which revealed major differences in inflammatory and phagocytic activity‐related gene expression between the different APOE genotypes.ConclusionsThis unique set of ten isogenic hiPSC lines enabled us to assess the cellular phenotypes in functional human neuronal cells. Microglia omics show changes in an array of AD‐relevant pathways, and being the main producers of apoE, these changes need to be understood in depth. Follow‐up experiments will validate candidate genes that might help design new therapies for APOE ε4 AD patients. Disclosure: This project has received funding from the Innovative Medicines Initiative 2 Joint Undertaking under grant agreement No 115975. This Joint Undertaking receives support from the European Union’s Horizon 2020 research and innovation programme and EFPIA. This study was sponsored by AbbVie Deutschland GmbH &amp; Co. KG. AbbVie contributed to the study design, research, and interpretation of data, writing, reviewing, and approving the abstract. Some authors are currently employed by AbbVie and may own AbbVie stock.

Insights in the pathophysiology of haemorrhagic strokes in a sub-Sahara African country, an epidemiological and MRI study.

Intra-cerebral Haemorrhage (ICH) seems more prevalent in sub-Saharan Africa (SSA) than in High-Income Countries (HIC) with poorer clinical outcome. Higher impact of hypertension and/or amyloid angiopathy could account for this disproportion. Here, we sought to (i) retrospectively compare ICH clinical and imaging patterns in Belgium and Guinea and in a subsequent cohort (ii) prospectively compare brain MRI characteristics to seek evidence for a different proportion of amyloid angiopathy patterns. Ninety one consecutive patients admitted for spontaneous ICH at Brussels Erasme-ULB Hospital and at Conakry Ignace Deen-UGANC were retrospectively compared in terms of ICH volume estimated with the ABC/2 method, clinical characteristics and modified ranking (mRS) score at 30days. mRS was dichotomised as good outcomes (≤3) and poor outcomes (>3). A prospective cohort of 30 consecutive patients with ICH admitted at CHU Conakry Ignace Deen-UGANC was prospectively included to undergo brain MRI. Results of the Guinean MRI were compared to 30 patients randomly selected from Brussels' initial cohort. Paired Student's t-test and Mann-Whitney u-test were used for group comparisons. Age of ICH onset was higher in Belgium (68±17years vs. 56±14years, P<0.01) while ICH volume and 30-day mortality rate were higher in Guinea (20ml vs. 11ml, P<0.01 and mortality 33% vs. 10 %, P<0.01). ICH burden in survivors in Conakry and Brussels showed respectively good outcomes in 56.7% and 60.4% (P=0.09) and poor outcomes in 10.3% vs. 29.6% (P<0.001). MRI analysis of the prospective cohort failed to disclose significant differences regarding brain MRI characteristics. Intra-cerebral Haemorrhage affected patients 15years younger in Guinea with larger haematoma volumes and higher mortality than in Belgium. MRI findings did not show more prevalent amyloid angiopathy pathology suggesting that better primary prevention of hypertension could positively impact ICH epidemiology in Guinea.

A high-throughput mass spectrometry-based assay for large-scale profiling of circulating human apolipoproteins

Apolipoproteins govern lipoprotein metabolism and are promising biomarkers of metabolic and cardiovascular diseases. Unlike immunoassays, MS enables the quantification and phenotyping of multiple apolipoproteins. Hence, here, we aimed to develop a LC-MS/MS assay that can simultaneously quantitate 18 human apolipoproteins [A-I, A-II, A-IV, A-V, B48, B100, C-I, C-II, C-III, C-IV, D, E, F, H, J, L1, M, and (a)] and determined apoE, apoL1, and apo(a) phenotypes in human plasma and serum samples. The plasma and serum apolipoproteins were trypsin digested through an optimized procedure and peptides were extracted and analyzed by LC-MS/MS. The method was validated according to standard guidelines in samples spiked with known peptide amounts. The LC-MS/MS results were compared with those obtained with other techniques, and reproducibility, dilution effects, and stabilities were also assessed. Peptide markers were successfully selected for targeted apolipoprotein quantification and phenotyping. After optimization, the assay was validated for linearity, lower limits of quantification, accuracy (biases: -14.8% to 12.1%), intra-assay variability [coefficients of variation (CVs): 1.5-14.2%], and inter-assay repeatability (CVs: 4.1-14.3%). Bland-Altman plots indicated no major statistically significant differences between LC-MS/MS and other techniques. The LC-MS/MS results were reproducible over five repeated experiments (CVs: 1.8-13.7%), and we identified marked differences among the plasma and serum samples. The LC-MS/MS assay developed here is rapid, requires only small sampling volumes, and incurs reasonable costs, thus making it amenable for a wide range of studies of apolipoprotein metabolism. We also highlight how this assay can be implemented in laboratories.

A simplified diagnosis algorithm for dysbetalipoproteinemia

Dysbetalipoproteinemia (DBL) is a disease of remnant lipoprotein accumulation caused by a defective apolipoprotein (apo) E and is associated with a considerable atherogenic burden. However, there exists confusion concerning the diagnosis of this disorder, and as a consequence, misdiagnosis is frequent. The objective of the present study is to propose an algorithm for the diagnosis of DBL using simple clinical variables. In a large cohort of 12,434 dyslipidemic patients, 4891 patients presented with mixed dyslipidemia (total cholesterol≥5.2mmol/L [200mg/dL] and triglycerides≥2.0mmol/L [175mg/dL]), and 188 DBL patients were identified based on the presence of an elevated very-low-density lipoprotein cholesterol/triglyceride ratio and were carriers of apoE2/E2. The APOE genotype or phenotype as well as the lipoprotein ultracentrifugation results were available for all patients. Among the laboratory variables associated with the lipid profile, the non-high-density lipoprotein cholesterol (HDL-C)/apoB ratio was the best predictor of DBL diagnosis based on the C-statistic. Previous proposed criteria had either low sensitivity or low specificity for the diagnosis of DBL. Using a non-HDL-C/apoB cut point of 3.69mmol/g (1.43 in conventional units) followed by the presence of apoE2/E2 resulted in a good sensitivity (94.8%), negative predictive value (99.8%), specificity (99.6%), positive predictive value (88.5%), accuracy (99.4%), and area under the curve (0.97 [0.95-0.99]) for the prediction of DBL. We therefore propose a 3-step algorithm for the diagnosis of DBL using total cholesterol and triglycerides as a first step, the non-HDL-C/apoB ratio as a second screening criterion and finally the APOE genotype, lipoprotein ultracentrifugation, or electrophoresis as a confirmatory test.

Sex-dependent calcium hyperactivity due to lysosomal-related dysfunction in astrocytes from APOE4 versus APOE3 gene targeted replacement mice

BackgroundThe apolipoprotein E (APOE) gene exists in three isoforms in humans: APOE2, APOE3 and APOE4. APOE4 causes structural and functional alterations in normal brains, and is the strongest genetic risk factor of the sporadic form of Alzheimer’s disease (LOAD). Research on APOE4 has mainly focused on the neuronal damage caused by defective cholesterol transport and exacerbated amyloid-β and Tau pathology. The impact of APOE4 on non-neuronal cell functions has been overlooked. Astrocytes, the main producers of ApoE in the healthy brain, are building blocks of neural circuits, and Ca2+ signaling is the basis of their excitability. Because APOE4 modifies membrane-lipid composition, and lipids regulate Ca2+ channels, we determined whether APOE4 dysregulates Ca2+signaling in astrocytes.MethodsCa2+ signals were recorded in astrocytes in hippocampal slices from APOE3 and APOE4 gene targeted replacement male and female mice using Ca2+ imaging. Mechanistic analyses were performed in immortalized astrocytes. Ca2+ fluxes were examined with pharmacological tools and Ca2+ probes. APOE3 and APOE4 expression was manipulated with GFP-APOE vectors and APOE siRNA. Lipidomics of lysosomal and whole-membranes were also performed.ResultsWe found potentiation of ATP-elicited Ca2+responses in APOE4 versus APOE3 astrocytes in male, but not female, mice. The immortalized astrocytes modeled the male response, and showed that Ca2+ hyperactivity associated with APOE4 is caused by dysregulation of Ca2+ handling in lysosomal-enriched acidic stores, and is reversed by the expression of APOE3, but not of APOE4, pointing to loss of function due to APOE4 malfunction. Moreover, immortalized APOE4 astrocytes are refractory to control of Ca2+ fluxes by extracellular lipids, and present distinct lipid composition in lysosomal and plasma membranes.ConclusionsImmortalized APOE4 versus APOE3 astrocytes present: increased Ca2+ excitability due to lysosome dysregulation, altered membrane lipidomes and intracellular cholesterol distribution, and impaired modulation of Ca2+ responses upon changes in extracellular lipids. Ca2+ hyperactivity associated with APOE4 is found in astrocytes from male, but not female, targeted replacement mice. The study suggests that, independently of Aβ and Tau pathologies, altered astrocyte excitability might contribute to neural-circuit hyperactivity depending on APOE allele, sex and lipids, and supports lysosome-targeted therapies to rescue APOE4 phenotypes in LOAD.

Associations of dietary choline intake with risk of incident dementia and with cognitive performance: the Kuopio Ischaemic Heart Disease Risk Factor Study

Moderate egg intake has been associated with better cognitive performance in observational studies. This association may be due to the rich content of choline, especially phosphatidylcholine, in eggs because choline has been suggested to have a role in the prevention of cognitive decline. We investigated the associations of dietary choline intake with the risk of incident dementia and with cognitive performance in middle-aged and older men in the prospective, population-based Kuopio Ischaemic Heart Disease Risk Factor Study. A population-based sample of 2497 dementia-free men aged 42-60 y was examined in 1984-1989. A subset of 482 men completed 5 different cognitive performance tests 4 y later. Dementia and Alzheimer disease diagnoses were retrieved from Finnish health registers. Dietary intakes were assessed with the use of 4-d food records at baseline. Cox regression and ANCOVA were used for the analyses. All analyses were also stratified by the apolipoprotein E phenotype (APOE-ε4 compared with other phenotypes). These data were available for 1259 men. The mean±SD total choline intake was 431±88 mg/d, of which 188±63 mg/d was phosphatidylcholine. During a 21.9-y follow-up, 337 men were diagnosed with dementia. Those in the highest compared with the lowest phosphatidylcholine intake quartile had 28% (95% CI: 1%, 48%; P-trend=0.02 across quartiles) lower multivariable-adjusted risk of incident dementia. Total choline intake had no association with the risk of incident dementia. However, both total choline and phosphatidylcholine intakes were associated with better performance in cognitive tests assessing frontal and temporal lobe functioning. For example, higher intakes were associated with better performance in verbal fluency and memory functions. The APOE phenotype had little or no impact on the associations. Higher phosphatidylcholine intake was associated with lower risk of incident dementia and better cognitive performance in men in eastern Finland. This trial was registered at clinicaltrials.gov as NCT03221127.

Egg consumption, cholesterol intake, and risk of incident stroke in men: the Kuopio Ischaemic Heart Disease Risk Factor Study

Epidemiologic studies suggest inverse associations between consumption of egg, a major source of dietary cholesterol, and stroke. However, the evidence of the relation remains limited, especially among carriers of apolipoprotein E4 (apoE4), which influences cholesterol metabolism. The aim of this study was to investigate associations of egg and cholesterol intakes with risk of stroke and with the major stroke risk factor, blood pressure, in middle-aged and older men from eastern Finland and whether apoE phenotype could modify these associations. A total of 1950 men aged 42-60 y in 1984-1989 were included at the baseline examinations of the prospective population-based Kuopio Ischaemic Heart Disease Risk Factor Study. Data on apoE phenotype were available for 1015 men. Dietary intakes were assessed with 4-d food records at baseline and incident stroke events were assessed by record linkage to hospital discharge registries. Cox proportional hazards regression analyses were used to estimate associations with stroke risk. Associations with baseline blood pressure were evaluated with ANCOVA. During the mean±SD follow-up of 21.2±7.2 y, there were 217 incidences of any stroke: 166 of ischemic stroke and 55 of hemorrhagic stroke. Comparing the highest egg intake quartile with the lowest, the multivariable-adjusted HRs were 0.81 for total stroke (95% CI: 0.54, 1.23; P-trend=0.32), 0.84 for ischemic stroke (95% CI: 0.53, 1.34; P-trend=0.44), and 0.75 for hemorrhagic stroke (95% CI: 0.32, 1.77; P-trend=0.40). The respective HRs for the highest cholesterol intake quartile compared with the lowest were 0.86 (95% CI: 0.57, 1.32; P-trend=0.42), 0.74 (95% CI: 0.46, 1.20; P-trend=0.32), and 1.10 (95% CI: 0.45, 2.66; P-trend=0.75). Diastolic blood pressure was 1.6 mm Hg (P-trend=0.04) lower in the highest egg intake quartile compared with the lowest, but there were no associations with systolic blood pressure or with cholesterol intake. ApoE phenotype (32% had apoE4 phenotype) did not modify the associations. Neither egg nor cholesterol intakes were associated with stroke risk in this cohort, regardless of apoE phenotype.This trial was registered at www.clinicaltrials.gov as NCT03221127.

APOE genetic variants and apoE, miR-107 and miR-650 levels in Alzheimer's disease.

Alzheimer's disease (AD) is a progressive neurodegenerative dementia in adults. Pathogenesis of AD depends on various factors, including APOE genetic variants, apolipoprotein E (apoE) phenotype and oxidative stress, which may promote both DNA and RNA damage, including non-coding RNA (ncRNA). Among ncRNAs, microRNA (miRNA) is known to contribute to pathologic processes in AD. The aim of the study was to analyse the plasma concentration of apoE by ELISA as well as the plasma levels of miR-107 and miR-650 by qPCR in relation to APOE genetic variants and clinical features including the age of onset and dementia severity in 64 AD patients and 132 controls. Our data showed that a low apoE plasma concentration was a risk factor for developing AD (OR = 5.18, p = 6.58E-06) and was particularly pronounced in severe dementia (p < 0.001) and correlated with cognitive functions (R = 0.295, p = 0.020), similarly as the level of miR-650 (R = 0.385, p = 0.033). The presence of APOE E4 allele in both AD patients and controls led to a reduction in apoE, while APOE E3/E3 genotype was associated with an increased apoE concentration and level of miR-107 in AD (p < 0.05) which was inversely correlated with the number of APOE E4 alleles (R = -0.448, p = 0.009). Additionally, patients with the onset at 60-69 years of age showed a reduced level of miR-107 (p < 0.05, as compared to AD above 80 years of age). Changed levels of plasma apoE, miR-107 and miR-650 may be a marker of the neurodegenerative process in the course of AD, associated with amyloid β metabolism and inordinate cell cycle.