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[email protected] Current Alzheimer Research, 2014, 11, 97-106
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Memory Impairment is Associated with Serum Methylarginines in Older Adults Mark McEvoy1,*, Peter Schofield2, Wayne Smith3, Kingsley Agho4, Arduino A. Mangoni5,6, Roy L. Soiza5,6, Roseanne Peel1 and John Attia1,7,* 1
Centre for Clinical Epidemiology & Biostatistics, Hunter Medical Research Institute, School of Medicine & Public Health, University of Newcastle, Australia, 2Centre for Translational Neuroscience and Mental Health, University of Newcastle, Newcastle, NSW, Australia, 3Environmental Health Branch, New South Wales Ministry of Health, Sydney, NSW, Australia, 4Science of Mental Health & Adversity unit, School of Medicine, University of Western Sydney, Parramatta, New South Wales, Australia, 5Division of Applied Medicine, School of Medicine and Dentistry, University of Aberdeen, Aberdeen, United Kingdom, 6Department of Medicine for the Elderly, Woodend Hospital, NHS Grampian, Aberdeen, United Kingdom, 7Department of General Medicine, John Hunter Hospital, Newcastle, New South Wales, Australia. Abstract: Background: This study measured serum concentrations of vascular risk factors, asymmetric dimethylarginine (ADMA) and symmetric dimethylarginine (SDMA) in a representative sample of older community-dwelling adults and determined their associations with objective and subjective memory impairment. Methods: Data on clinical, lifestyle, and demographic characteristics, serum ADMA, SDMA, and L-arginine (measured using LC-MS/MS) were collected from a population-based sample of older Australian adults from the Hunter Community Study. Objective memory was measured with the Audio Recorded Cognitive Screen (ARCS) neuropsychological battery and subjective memory impairment was measured using the Memory Complaint Questionnaire (MAC-Q). Results: Multivariate analysis revealed that SDMA and diabetes were significantly associated with objective memory impairment (Adjusted Odd ratio (AOR) = 3.90; 95% CI. 1.21 – 12.52 for fourth quartile (Q4) of SDMA. ADMA, SDMA, education, number of general practitioner visits and atrial fibrillation were all significantly associated with subjective memory impairment. (AOR = 1.82; 95% CI. 1.04 – 3.18 for Q4 ADMA. Conclusions: Higher serum SDMA was associated with objective and subjective memory impairment while higher serum ADMA was associated with subjective memory impairment.
Keywords: ADMA, Alzheimer’s disease, memory impairment, methylarginines, nitric oxide, older adults, SDMA. 1. INTRODUCTION Dementia is a devastating syndrome affecting more than 35.6 million people world-wide approximately 60-70% of whom will have Alzheimer’s disease (AD) as an underlying cause [1]. With the worldwide incidence expected to increase at a rate of 7.7 million cases each year, without an effective intervention to cure or halt progression of the disease, dementia will become the most burdensome disease by 2050 [1]. The possible role of Nitric oxide (NO) in the development of dementia syndromes and the role of agents which may modulate concentrations of NO is attracting growing attention. There are a large number of studies implicating excess CNS synthesis of NO in the pathophysiology of neurodegenerative disease including AD [2]. Nitric oxide plays an important role in synaptic events responsible for memory formation and learning [3]. In neuronal cells the synthesis of NO from the amino acid L-arginine is primarily accomp*Address correspondence to these authors at the Level 4 Hunter Medical Research Institute, Centre for Clinical Epidemiology and Biostatistics, School of Medicine and Public Health, University of Newcastle, Callaghan, NSW 2308 Australia; Tel: +61 2 40420518; Fax: +61 2 40420044; Emails:
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lished by neuronal NO synthase (nNOS). Research has established nNOS as the principle source of NO during longterm potentiation (LTP) [4], a key process responsible for memory formation and learning [5]. In human endothelial cells, NO is synthesised from L-arginine by the constitutive enzyme, endothelial NO synthase (eNOS) [6]. Endothelial NOS is expressed by vascular endothelial cells in the brain where it regulates cerebral blood flow. An inducible nitric oxide synthase (iNOS) is also expressed in large amounts (relative to eNOS and nNOS) in nervous tissue in response to inflammation, viral infection, or trauma [7]. Activation of iNOS produces micromolar amounts of potentially damaging NO compared with the constitutive nanomolar amounts produced by eNOS and nNOS. In existing AD, the increased synthesis of NO is primarily due to activation of iNOS, however eNOS and nNOS also contribute to the aberrant synthesis of NO [8, 9]. There is extensive epidemiologic evidence demonstrating that vascular disease and vascular risk factors are associated with an increased risk of developing AD in addition to vascular dementia [10-12]. Endothelial dysfunction, vasoconstriction, vascular remodelling, atherosclerosis and decreased cerebral blood flow may play a causal role in mediating this association. Indeed chronic brain hypoperfusion is associated with oxidative stress, energy metabolic deficits and impaired © 2014 Bentham Science Publishers
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memory long before signs of dementia are seen [13]. Furthermore, decreased cerebral perfusion is a powerful predictor for the onset of dementia [14]. Given that endothelialderived NO contributes to the maintenance of vascular haemostasis a disturbed synthesis and/or availability of NO early in the disease continuum might contribute to the development of AD. The methylarginine asymmetric dimethylarginine (ADMA) is an endogenous NOS inhibitor [15] that inhibits the activity of all three NO synthase isoforms, and has been shown to reduce NO concentrations in numerous cell types [16-18]. Elevated serum ADMA concentrations have been consistently demonstrated in individuals with traditional vascular risk factors [19-21] and those with existing cardiovascular disease [19, 20, 22, 23] and these risk factors have been shown to independently predict cerebrovascular disease and dementia [10-12]. Furthermore, higher serum methylarginine concentrations have been associated with increased mortality and cardiovascular endpoints in numerous longitudinal studies involving healthy [20, 24] and patient populations [19, 22, 23]. Symmetric dimethylarginine (SDMA) was originally not considered as a potential risk factor for cardiovascular disease as it does not directly influence NOS isoforms. However, SDMA has been reported to competitively inhibit arginine uptake In vitro [19, 25] and recent studies suggest that it too may be implicated in vascular disease [26, 27]. Endogenous methylarginines are important potentially modifiable molecules that may be associated with poor cognitive performance/memory impairment and the development of dementia. This view is supported by the following observations. First, ADMA is responsible for endothelial dysfunction, vasoconstriction, atherosclerosis and decreased cerebral blood flow and these factors are consistently associated with the development of mild cognitive impairment (MCI) and dementia [28-32]. Secondly, ADMA impairs endothelial-derived NO synthesis and this has been shown to be associated with the development of A plaques in the rat brain [33]. Third, ADMA is a NOS inhibitor and there are a large number of studies demonstrating that naturally occurring and synthetic NOS inhibitors impair cognitive performance in animals and this is mediated by inhibition of NOS [3]. Fourth, SDMA has been reported to competitively inhibit arginine uptake In vitro [19, 25] and recent studies suggest that it too may be implicated in vascular disease [26, 27]. Finally, five studies have examined the association between serum ADMA and AD with most showing an increase in serum ADMA concentrations [34-38]. Hence an increase in endogenous ADMA and/or SDMA, may contribute to the underlying pathology of cognitive impairment and AD through a direct influence on brain NOS isoforms and/or depletion of the L-arginine pool which is required for adequate synthesis of NO. This research further supports the need for an investigation of the role of the endogenous methylarginines, ADMA and SDMA with cognitive impairment/decline in humans. It is important to determine if increased serum ADMA concentrations are associated with cognitive impairment/decline because modulation of ADMA metabolism, by acting on DDAH activity, is a potential target for pharmacological manipulation [39]. Given that no previous study has examined the association of endogenous methylarginines with cognitive perform-
McEvoy et al.
ance/memory impairment, the primary aim of this research is to measure serum concentrations of the methylarginines ADMA and SDMA in a representative sample of older community-dwelling adults and determine their association with cognition/memory impairment. Since L-arginine is a precursor of NO synthesis and L-arginine to ADMA ratio is considered a marker of vascular function a secondary aim was to determine if serum L-arginine and arginine/ADMA ratio are associated with cognition/memory impairment. We hypothesize that in a representative sample of older community-dwelling adults, increased serum ADMA and SDMA will be associated with reduced measures of cognitive function and greater memory impairment as measured by the memory domain subscale of the Audio Recorded Cognitive Screen (ARCS) neuropsychological battery [40] and the Memory Complaint Questionnaire (MAC-Q) [41]. 2. MATERIALS AND METHODS Data for this study was obtained from the Hunter Community Study (HCS), a cohort of community-dwelling men and women aged 55 to 85 years of age in Newcastle, New South Wales (NSW), Australia. Approval to conduct the research was granted by the University of Newcastle Human Research Ethics Committees. This study has been described in detail elsewhere [42]. In brief, participants were randomly selected from the New South Wales State Electoral roll and 9,784 individuals were contacted between December 2004 and May 2007. Of these, a total of 3,253 actually participated (response rate 44.5% after removing incorrect addresses and non-contacts). Participants completed a series of self-reported questionnaires, attended a clinic visit, and consented to linkage of health records. The sample for this investigation (n=500) was derived from the initial cohort by simple random sampling. Of the 500 subjects randomly selected there were complete exposure and outcome data for 483 subjects. No a priori sample size was determined, however assuming that at least 10-15 subjects are needed for each independent variable included in the multivariate analysis the sample size was more than sufficient to accommodate the number of co-variables examined in this investigation (see Statistical analysis paragraph). A comparison of this sample with the entire cohort showed no significant difference for a range of clinical, biochemical, socioeconomic, and behavioural factors (data not shown). For full details of this cohort and sample characteristics see McEvoy M. et al., 2010 [42]. 2.1. Main Exposure Variables: L-Arginine and Methylarginines The primary explanatory variables in this study were ADMA and SDMA. Secondary predictors were L-arginine, and L-arginine/ADMA ratio. Blood was collected in EDTA tubes and centrifuged at 4oC and 3000g for 10 minutes to separate serum, which was stored for three years at -80oC before analysis. L-Arginine and its di-methylated forms (ADMA and SDMA) were measured in serum by hydrophilic- interaction liquid chromatography and isotope dilution tandem mass spectrometry [43]. The intra and interassay coefficients of variation for arginine, ADMA and SDMA were all less than 15%.
Methylarginines and Memory Impairment
2.2. Primary Outcomes Cognitive screening was performed using the ARCS neuropsychological battery [44]. The ARCS assesses five key domains: memory, verbal fluency, visuospatial functioning, language and attention/executive function, and speed-ofwriting. The ARCS is adjusted for age, gender and educational status and is scaled to a mean of 100 and SD of 15. Subjective memory was measured using the MACQ(41).The MAC-Q is a six item self-report tool developed with the intent of screening for Age-Associated Memory Impairment (AAMI) and for research that requires an index of memory complaint. It is a measure of subjective memory complaint where an individual reports their current memory ability relative to the past (scores range from 7-35). The MAC-Q has good internal consistency ( = 0.57, P
