Tuomas Rauramaa

BDNF is a novel marker of cognitive function in ageing women: The DR’s EXTRA Study☆

Brain-derived neurotrophic factor (BDNF) is one of the key molecules modulating brain plasticity. While low circulating levels of BDNF have been suggested to predispose to Alzheimers disease, very little data are available on its association with cognitive function in general population. We evaluated the association between plasma BDNF levels and cognition in a representative population sample of ageing men and women. The subjects (n=1389) were participants of the Dose-Responses to Exercise Training (DRs EXTRA) Study and represent a random sample of Eastern Finnish people (684 men and 705 women), 57-79 years of age at baseline of the study. Plasma BDNF levels were measured by enzyme-linked immunosorbent assay (ELISA). Cognitive function was evaluated using the Consortium to Establish a Registry for Alzheimers Disease (CERAD) neuropsychological test battery. Women had a higher mean (+/-SEM) plasma BDNF level than men (1721+/-55vs. 1495+/-54pg/ml, P<0.001). In women, 1 SD decrease in BDNF increased the risk for a low score in Naming Test by 53% (95% CI 1.21-1.92, P<0.001), in Mini-Mental State Examination by 63% (95% CI 1.21-2.20, P=0.001), in Word List Memory by 56% (95% CI 1.08-2.26, P=0.019), in Word List Recall by 50% (95% CI 1.10-2.05, P=0.010), in Word List Saving by 49% (95% CI 1.12-1.99, P=0.007), and in Word List Recognition by 64% (95% CI 1.19-2.25, P=0.002). Data were adjusted for age, education, depression, impaired glucose metabolism, cardiovascular disease, antihypertensive medication, lipid lowering medication, use of sex hormones, smoking, alcohol consumption, storing time of plasma in the freezer and platelet count. BDNF was not associated with cognition in men. Present data suggest that plasma BDNF is a biomarker of impaired memory and general cognitive function in ageing women.

Chronotropic incompetence and mortality in middle-aged men with known or suspected coronary heart disease

AIMS The objective is to study whether a heart rate (HR) increase from 40 to 100% of maximal work capacity in the exercise test (HR40-100) independently predicts mortality in men with known or suspected coronary heart disease (CHD). METHODS AND RESULTS The subjects were 294 men, 42-61 years of age, from eastern Finland with known or suspected CHD but without use of HR-lowering medication at baseline. HR was measured at rest and during a maximal, symptom-limited exercise test. During an average follow-up of 11.0 years, there were 61 all-cause deaths. In Cox-multivariable model, mortality increased by 41% (95% confidence interval, 12-79%) with a 1-SD (15 b.p.m.) decrement in HR40-100. HR increase from rest to 40% of maximal work capacity was not associated with an increased risk of death. Synergism was observed between HR40-100 and workload achieved at HR of 100 b.p.m. so that men having low values for both these HR variables had a particularly adverse prognosis compared with men with high values for these variables. CONCLUSION An attenuated HR increase particularly during the latter half of a maximal exercise test is an independent predictor of death in men with known or suspected CHD.

TAR-DNA binding protein-43 and alterations in the hippocampus

Immunocytochemistry for transactive response binding protein-43 (TDP43) was assessed in the granular cell layer of the dentate gyrus in 250 cases displaying hippocampal pathology identified by haematoxylin–eosin staining. 18%, nearly one in five displayed TDP43 immunoreactive pathology in the granular cell layer of hippocampus. This percentage increased to 43% when only subjects with hippocampal pathology other than vascular in origin were included. When only subjects with severe Alzheimer’s disease-related pathology were included, 42% displayed TDP43-immunoreactive pathology, increasing to 60% when concomitant Alzheimer’s disease and α-synuclein pathology were present. Within this setting, TDP43-immunoreactive pathology was observed to be present in 6% of subjects with hippocampal pathology but without any cognitive impairment. Our findings justify assessment of TDP43 pathology in every case where a pathological alteration is observed in the hippocampus using a routine stain.

Usefulness of Chronotropic Incompetence in Response to Exercise as a Predictor of Myocardial Infarction in Middle-Aged Men Without Cardiovascular Disease

An attenuated heart rate (HR) response to exercise, or chronotropic incompetence, has been shown to predict adverse cardiac events in subjects without known cardiovascular disease (CVD). The aim of the present study was to investigate whether chronotropic incompetence independently predicts acute myocardial infarction (AMI) in middle-aged men. In addition to previously established chronotropic incompetence variables, we focused on a new chronotropic incompetence variable, the HR increase from 40% to 100% of maximal work capacity (HR40-100), as a predictor of AMI. The subjects were a representative sample of 1,176 middle-aged men who did not have CVD and did not use HR-lowering medication at baseline. The association of chronotropic incompetence variables with the risk of AMI was examined by Cox regression models including numerous known risk factors for AMI. During an average follow-up of 11.0 years, there were 106 AMIs (9.0%). In Cox multivariable model, the risk of AMI increased by 33% for each SD decrement of 13 beats/min in HR40-100 (95% confidence interval [CI] 9 to 64). When considered concurrently, HR40-100 was the only chronotropic incompetence variable that improved the predictive value of the model containing other risk factors for AMI. Men with a low HR40-100 (<46 beats/min) and a heightened increase in systolic blood pressure (SBP) (>67 mm Hg) were at particularly high risk, with a 3.1-times higher incidence of AMI than those with a normal HR40-100 and SBP increase (95% CI 1.7 to 5.7). In conclusion, a low HR40-100 predicted AMI in men without previous CVD independent of other exercise test or clinical variables.

Transcriptomics and mechanistic elucidation of Alzheimer's disease risk genes in the brain and in vitro models.

In this study, we have assessed the expression and splicing status of genes involved in the pathogenesis or affecting the risk of Alzheimers disease (AD) in the postmortem inferior temporal cortex samples obtained from 60 subjects with varying degree of AD-related neurofibrillary pathology. These subjects were grouped based on neurofibrillary pathology into 3 groups: Braak stages 0-II, Braak stages III-IV, and Braak stages V-VI. We also examined the right frontal cortical biopsies obtained during life from 22 patients with idiopathic shunt-responding normal pressure hydrocephalus, a disease that displays similar pathologic alterations as seen in AD. These 22 patients were categorized according to dichotomized amyloid-β positive or negative pathology in the biopsies. We observed that the expression of FRMD4A significantly decreased, and the expression of MS4A6A significantly increased in relation to increasing AD-related neurofibrillary pathology. Moreover, the expression of 2 exons in both CLU and TREM2 significantly increased with increase in AD-related neurofibrillary pathology. However, a similar trend toward increased expression in CLU and TREM2 was observed with most of the studied exons, suggesting a global change in the expression rather than altered splicing. Correlation of gene expression with well-established AD-related factors, such as α-, β-, and γ-secretase activities, brain amyloid-β42 levels, and cerebrospinal fluid biomarkers, revealed a positive correlation between β-secretase activity and the expression of TREM2 and BIN1. In expression quantitative trait loci analysis, we did not detect significant effects of the risk alleles on gene expression or splicing. Analysis of the normal pressure hydrocephalus biopsies revealed no differences in the expression or splicing profiles of the studied genes between amyloid-β positive and negative patients. Using the protein-protein interaction-based in vitro pathway analysis tools, we found that downregulation of FRMD4A associated with increased APP-β-secretase interaction, increased amyloid-β40 secretion, and altered phosphorylation of tau. Taken together, our results suggest that the expression of FRMD4A, MS4A6A, CLU, and TREM2 is altered in relation to increasing AD-related neurofibrillary pathology, and that FRMD4A may play a role in amyloidogenic and tau-related pathways in AD. Therefore, investigation of gene expression changes in the brain and effects of the identified genes on disease-associated pathways in vitro may provide mechanistic insights on how alterations in these genes may contribute to AD pathogenesis.

Cardiovascular Diseases and Hippocampal Infarcts

The prevalence of hippocampal lesions such as hippocampal infarcts have not been studied in detail even though hippocampal alterations are known to be associated with various clinical conditions such as age‐related degenerative disorders and epilepsy. Methods: Here we defined the hippocampal infarcts and assessed the prevalence of this lesion in large unselected population of 1,245 subjects age ranging from 1 to 99 years (mean age 79 ± 1 S.E.M). Furthermore, we assessed the association of these lesions with various cardio‐ and cerebro‐vascular disorders and other neurodegenerative lesions. The prevalence of hippocampal infarct in the study population of 1,245 subjects was 12%, increasing to 13% when only those with a clinically diagnosed cognitive impairment (n = 311) were analyzed. Large hemispheric brain infarcts were seen in 31% of the study subjects and these lesions were strongly associated with cardiovascular risk factors such as hypertension (43%), coronary disease (32%), myocardial infarct (22%), atrial fibrillation (20%), and heart failure (20%). In contrast, hippocampal infarcts displayed a significant association only with large hemispheric brain infarct, heart failure, and cardiovascular index as assessed postmortem. It is noteworthy that only widespread hippocampal infarcts were associated with clinical symptoms of cognitive impairment or epilepsy. The surprisingly low prevalence of 12% of hippocampal infarcts in aged population found here and the failure to detect an association between this lesion and various cerebro‐ cardio‐vascular lesions is intriguing. Whether susceptibility to ischemia in line with susceptibility to neuronal degeneration in this region is influenced by still undetermined risk‐ factors need further investigation. Furthermore it should be noted that the size of the hippocampal tissue damage, i.e., small vs. large cystic infarcts is of significance regarding clinical alterations.

Somatic Activating KRAS Mutations in Arteriovenous Malformations of the Brain

BACKGROUND Sporadic arteriovenous malformations of the brain, which are morphologically abnormal connections between arteries and veins in the brain vasculature, are a leading cause of hemorrhagic stroke in young adults and children. The genetic cause of this rare focal disorder is unknown. METHODS We analyzed tissue and blood samples from patients with arteriovenous malformations of the brain to detect somatic mutations. We performed exome DNA sequencing of tissue samples of arteriovenous malformations of the brain from 26 patients in the main study group and of paired blood samples from 17 of those patients. To confirm our findings, we performed droplet digital polymerase‐chain‐reaction (PCR) analysis of tissue samples from 39 patients in the main study group (21 with matching blood samples) and from 33 patients in an independent validation group. We interrogated the downstream signaling pathways, changes in gene expression, and cellular phenotype that were induced by activating KRAS mutations, which we had discovered in tissue samples. RESULTS We detected somatic activating KRAS mutations in tissue samples from 45 of the 72 patients and in none of the 21 paired blood samples. In endothelial cell–enriched cultures derived from arteriovenous malformations of the brain, we detected KRAS mutations and observed that expression of mutant KRAS (KRASG12V) in endothelial cells in vitro induced increased ERK (extracellular signal‐regulated kinase) activity, increased expression of genes related to angiogenesis and Notch signaling, and enhanced migratory behavior. These processes were reversed by inhibition of MAPK (mitogen‐activated protein kinase)–ERK signaling. CONCLUSIONS We identified activating KRAS mutations in the majority of tissue samples of arteriovenous malformations of the brain that we analyzed. We propose that these malformations develop as a result of KRAS‐induced activation of the MAPK–ERK signaling pathway in brain endothelial cells. (Funded by the Swiss Cancer League and others.)

Relationship between ubiquilin-1 and BACE1 in human Alzheimer's disease and APdE9 transgenic mouse brain and cell-based models.

Accumulation of β-amyloid (Aβ) and phosphorylated tau in the brain are central events underlying Alzheimers disease (AD) pathogenesis. Aβ is generated from amyloid precursor protein (APP) by β-site APP-cleaving enzyme 1 (BACE1) and γ-secretase-mediated cleavages. Ubiquilin-1, a ubiquitin-like protein, genetically associates with AD and affects APP trafficking, processing and degradation. Here, we have investigated ubiquilin-1 expression in human brain in relation to AD-related neurofibrillary pathology and the effects of ubiquilin-1 overexpression on BACE1, tau, neuroinflammation, and neuronal viability in vitro in co-cultures of mouse embryonic primary cortical neurons and microglial cells under acute neuroinflammation as well as neuronal cell lines, and in vivo in the brain of APdE9 transgenic mice at the early phase of the development of Aβ pathology. Ubiquilin-1 expression was decreased in human temporal cortex in relation to the early stages of AD-related neurofibrillary pathology (Braak stages 0-II vs. III-IV). There was a trend towards a positive correlation between ubiquilin-1 and BACE1 protein levels. Consistent with this, ubiquilin-1 overexpression in the neuron-microglia co-cultures with or without the induction of neuroinflammation resulted in a significant increase in endogenously expressed BACE1 levels. Sustained ubiquilin-1 overexpression in the brain of APdE9 mice resulted in a moderate, but insignificant increase in endogenous BACE1 levels and activity, coinciding with increased levels of soluble Aβ40 and Aβ42. BACE1 levels were also significantly increased in neuronal cells co-overexpressing ubiquilin-1 and BACE1. Ubiquilin-1 overexpression led to the stabilization of BACE1 protein levels, potentially through a mechanism involving decreased degradation in the lysosomal compartment. Ubiquilin-1 overexpression did not significantly affect the neuroinflammation response, but decreased neuronal viability in the neuron-microglia co-cultures under neuroinflammation. Taken together, these results suggest that ubiquilin-1 may mechanistically participate in AD molecular pathogenesis by affecting BACE1 and thereby APP processing and Aβ accumulation.

Effects of NR1H3 genetic variation on the expression of liver X receptor α and the progression of Alzheimer's disease.

Alzheimers disease (AD) has been postulated to involve defects in the clearance of amyloid-β (Aβ). Activation of liver X receptor α (LXRα) increases the expression of apolipoprotein E (ApoE) as well as cholesterol transporters ABCA1 and ABCG1, leading to augmented clearance of Aβ. We have previously shown that the C allele of rs7120118 in the NR1H3 gene encoding LXRα reduces the risk of AD. Here, we wanted to assess whether the rs7120118 variation affects the progression of AD and modulates the expression of NR1H3 and its downstream targets APOE, ABCA1 and ABCG1.We utilized tissue samples from the inferior temporal cortex of 87 subjects, which were subdivided according to Braak staging into mild, moderate and severe AD groups on the basis of AD-related neurofibrillary pathology. APOE ε4 allele increased soluble Aβ42 levels in the tissue samples in a dose-dependent manner, but did not affect the expression status of APOE. In contrast, the CC genotype of rs7120118 was underrepresented in the severe group, although this result did not reach statistical significance. Also, patients with the CC genotype of rs7120118 showed significantly decreased soluble Aβ42 levels as compared to the patients with TT genotype. Although the severity of AD did not affect NR1H3 expression, the mRNA levels of NR1H3 among the patients with CT genotype of rs7120118 were significantly increased as compared to the patients with TT genotype. These results suggest that genetic variation in NR1H3 modulates the expression of LXRα and the levels of soluble Aβ42.

The Expression of Transthyretin and Amyloid-β Protein Precursor is Altered in the Brain of Idiopathic Normal Pressure Hydrocephalus Patients

BACKGROUND Idiopathic normal pressure hydrocephalus (iNPH) is a dementing condition in which Alzheimers disease (AD)-related amyloid-β (Aβ) plaques are frequently observed in the neocortex. iNPH patients with prominent Aβ pathology show AD-related alterations in amyloid-β protein precursor (AβPP) processing resulting from increased γ-secretase activity. OBJECTIVES Our goal was to assess potential alterations in the global gene expression profile in the brain of iNPH patients as compared to non-demented controls and to evaluate the levels of the identified targets in the cerebrospinal fluid (CSF) of iNPH patients. METHODS The genome-wide expression profile of ~35,000 probes was assessed in the RNA samples obtained from 22 iNPH patients and eight non-demented control subjects using a microarray chip. The soluble levels of sAβPPα, sAβPPβ, and transthyretin (TTR) were measured from the CSF of 102 iNPH patients using ELISA. RESULTS After correcting the results for multiple testing, significant differences in the expression of TTR and A βPP were observed between iNPH and control subjects. The mRNA levels of TTR were on average 17-fold lower in iNPH samples compared to control samples. Conversely, the expression level of A βPP was on average three times higher in iNPH samples as compared to control samples. Interestingly, the expression of α-secretase (ADAM10) was also increased in iNPH patients. In the lumbar CSF samples, soluble TTR levels showed a significant positive correlation with sAβPPα and sAβPPβ, but TTR levels did not predict the brain pathology or the shunt response. CONCLUSIONS These findings suggest differences in the expression profile of key factors involved in AD-related cellular events in the brain of iNPH patients.