Carriers of an apolipoprotein E epsilon 4 allele are more vulnerable to a dietary deficiency in omega-3 fatty acids and cognitive decline

    Carriers of an apolipoprotein E epsilon 4 allele are more vulnerable to a dietary deficiency in omega-3 fatty acids and cognitive decline Tanya Gwendolyn Nock, Rapha¨el Chouinard-Watkins, M´elanie Plourde PII: DOI: Reference:

S1388-1981(17)30135-X doi:10.1016/j.bbalip.2017.07.004 BBAMCB 58177

To appear in:

BBA - Molecular and Cell Biology of Lipids

Received date: Revised date: Accepted date:

26 March 2017 5 July 2017 15 July 2017

Please cite this article as: Tanya Gwendolyn Nock, Rapha¨el Chouinard-Watkins, M´elanie Plourde, Carriers of an apolipoprotein E epsilon 4 allele are more vulnerable to a dietary deficiency in omega-3 fatty acids and cognitive decline, BBA - Molecular and Cell Biology of Lipids (2017), doi:10.1016/j.bbalip.2017.07.004

This is a PDF file of an unedited manuscript that has been accepted for publication. As a service to our customers we are providing this early version of the manuscript. The manuscript will undergo copyediting, typesetting, and review of the resulting proof before it is published in its final form. Please note that during the production process errors may be discovered which could affect the content, and all legal disclaimers that apply to the journal pertain.

ACCEPTED MANUSCRIPT Carriers of an apolipoprotein E epsilon 4 allele are more vulnerable to a dietary deficiency in omega-3 fatty acids and cognitive decline.

PT

Tanya Gwendolyn Nock, Raphaël Chouinard-Watkins and Mélanie Plourde.

RI

Research Center on Aging, Centre Intégré Universitaire de Santé et Services Sociaux de l’Estrie-Centre Hospitalier Universitaire de Sherbrooke, Canada

SC

Faculté de médecine et des sciences de la santé, Université de Sherbrooke, Sherbrooke, Canada

To whom correspondence should be addressed:

MA

Mélanie Plourde, Ph.D. Research Center on Aging Sherbrooke, Canada, J1H 4C4

TE

Tel : 819-780-2220 extension 45664

D

1036 Belvédère Sud

Fax : 819-829-7141

AC CE P

Mail : [email protected] Running title:

Abbreviations used:

AA: arachidonic acid

ALA: alpha-linolenic acid APOE: apolipoprotein E Aβ : β-amyloid BBB: blood-brain-barrier

CNS: central nervous system DHA: docosahexaenoic acid, 22:6 n-3 DHAEE: DHA ethyl ester E4: epsilon 4 allele of apolipoprotein E EPA: eicosapentaenoic acid FA: fatty acid F2: second generation

NU

Institute of Nutrition and Functional Foods, Quebec City, Canada

ACCEPTED MANUSCRIPT GLUT: glucose transporter LA: linoleic acid MRI: magnetic resonance imaging n-3: omega-3

PT

PBD: peroxisome digenesis disorder

RI

PEX: peroxisomal biogenesis factor

SC

Abstract (170 words):

Carriers of an epsilon 4 allele (E4) of apolipoprotein E (APOE) develop Alzheimer’s disease (AD)

NU

earlier than carriers of other APOE alleles. The metabolism of plasma docosahexaenoic acid (DHA, 22:6n-3), an omega-3 fatty acid (n-3 FA), taken up by the brain and concentrated in neurons, is

MA

disrupted in E4 carriers, resulting in lower levels of brain DHA. Behavioural and cognitive impairments have been observed in animals with lower brain DHA levels, with emphasis on loss of spatial memory and increased anxiety. E4 mice provided a diet deficient in n-3 FA had a greater

D

depletion of n-3 FA levels in organs and tissues than mice carrying other APOE alleles. However,

TE

providing n-3 FA can restore levels of brain DHA in E4 animals and in other models of n-3 FA deficiency. In E4 carriers, supplementation with DHA as early as possible might help to prevent the

AC CE P

onset of AD and could halt the progression of, and reverse some of the neurological and behavioural consequences of their higher vulnerability to n-3 FA deficiency.

ACCEPTED MANUSCRIPT Introduction Today, there are around 44 million men and women worldwide with Alzheimer’s disease (AD). The onset of AD in carriers of an epsilon 4 allele of apolipoprotein E (E4) is 8-15 years earlier than in carriers of other apolipoprotein E (APOE) alleles [1]. It is unclear as to how E4 modulates AD

PT

pathology but neuropathological changes associated with AD such as β-amyloid (Aβ) plaque

RI

deposition occur as early as 30 years of age in E4 carriers [2]. However, one intriguing aspect is that

factors to modulate the clinical manifestation of AD.

SC

not all E4 carriers will develop AD suggesting that there is a window of action for environmental risk

To be healthy and function well, the brain needs a constant supply of blood to deliver oxygen

NU

and nutrients. The blood-brain-barrier (BBB) is an obligatory interface between the cardiovascular system and the brain. Recent evidence suggests that in humans, there is an age-dependent loss of BBB

MA

integrity in the hippocampus [3]. In 2014, two papers published in Nature reported a surprising dual role for the BBB transporter Mfsd2a both in establishing BBB integrity and the uptake of

D

docosahexaenoic acid (DHA) [4, 5], a long chain omega-3 fatty acid (n-3 FA) concentrated in brain

TE

membranes. Mutations in conserved residues of the MFSD2A gene lead to lethal microcephaly syndrome linked to inadequate brain uptake of FAs such as DHA [6]. Mice knockdown for MFSD2A

AC CE P

have significantly lower DHA in their brains, neuronal cell loss in the hippocampus and the cerebellum, neurological and behavioural deficits and reduced brain size [5]. The brain has a limited capacity to produce DHA de novo and obtains DHA from the plasma. The level of DHA in the plasma reflects a balance between uptake and release of DHA from organs and tissues. For the past 7 years, we have shown that plasma DHA metabolism is disrupted in E4 carriers. Hence, the following sections will describe evidence supporting the role of n-3 FA deficiency in neurological and behavioural consequences and explain the rationale behind our hypothesis of a connection between the dysregulated DHA metabolism occurring in E4 carriers, the neurological consequences observed thereafter, and the increase in risk of developing AD in E4 carriers. N-3 fatty acid deficiency and neurological consequences Consumption of n-3 FA and n-6 FA in the western diet has changed markedly over the 20th century largely due to the increased production of soybean oil and new food production methodologies. One study attempted to reconstruct consumption patterns and the availability of essential fatty acids in traditional methods (1909) and in current practices (1999) to predict the net effects of altered intakes on the concentration of FA in tissues. A discussion and comparison of reconstructed estimates to available data from other studies in the prediction of tissue content have been discussed [7]. The study found the

ACCEPTED MANUSCRIPT availability of linoleic acid (LA), an n-6 FA most commonly found in soybean oil, increased from 2.79% to 7.21% of energy and found substantial declines in the dietary availability of n-6 arachidonic acid (AA), and n-3 FA eicosapentaenoic acid (EPA) and DHA. The dietary increase in n-6 FA and decrease in n-3 FA results in n-6 FA replacing n-3 FA in the tissue membrane prevents DHA from

PT

carrying out its multiple roles in the neuronal membrane. Predicted net effects of these dietary changes

RI

included declines in tissue n-3 highly unsaturated fatty acid status by one-third and declines in the estimated omega-3 index, a direct measure of erythrocyte EPA + DHA as a percentage of total fatty

SC

acids by half. The apparent increased consumption of LA, has likely decreased tissue concentrations of EPA and DHA during the 20th century and provides evidence that the western diet is deficient in DHA

NU

[8]. This has negative consequences because an excess of n-6 FA and a deficient amount of n-3 FA is stimulating thrombogenesis, lowering immune response, increasing inflammation and decreasing

MA

neuronal membrane fluidity and function [9-11]. In fact, many experimental protocols in animals now commonly use a diet with a composition of 0% DHA as a typical western diet in their methodologies without citing a rationale for its composition [12, 13]. The ratio of n-6:n-3 FA in the US western diet is

D

around 15:1. This ratio is lower in most European countries however, and in Nordic countries the ratio

TE

is below 5:1. Ratios less than 5:1 were associated with decreased mortality and a reduced risk of chronic diseases such as colorectal cancer and rheumatoid arthritis by reducing rectal cell proliferation

AC CE P

and suppressing inflammation, respectively [14]. Further to this point, association studies have linked the dietary n-6:n-3 ratio to cognitive decline and the incidence of dementia [15]. Since there is no clear definition of a deficient diet in n-3 FA in humans, the Western diet is therefore categorized as a diet low in n-3 FA because of its n-6:n-3 FA ratio of around 15:1. Cumulative evidence in animal models beginning in the late 1970’s supports that diets devoid of n-3 FA produce observable pathologies in the brain which will be discussed herein [16]. Studies of n-3 fatty acid deficiency span a wide range of animal models including rat, monkey, mouse, guinea pig, frog, honey bee as well as humans with certain genetic disorders predisposing them to endogenous n-3 FA deficiency [17-20]. Deficiency in n-3 FAs, namely DHA or α-linolenic acid (ALA), modifies the FA composition of several organs in the body with the greatest changes in FA composition found in the highly metabolically active tissues of the brain, heart, muscle, retina and liver [21]. Modifications to FA in n-3 deficiency have been studied in the neuronal membranes throughout the brain with the greatest loss of n-3 FA at 90% of DHA from control found in the hippocampus of the n-3 deficient rat brain [22] and are unique to the region of the brain studied [23-26]. Furthermore, an alteration in fatty acid composition was observed in the synaptosomes and microsomes generated from the

ACCEPTED MANUSCRIPT neuronal membrane [27] as well as a 3-fold reduction in the capillaries of the adult rat brain deficient in n-3 [28]. In addition to the change in lipid composition of neuronal membranes, a couple of groups found the neurons of the DHA-deficient rat smaller in size (with no change in density, cell number, or cell volume) in the hypothalamus and parietal cortex in weaning, in the piriform cortex in maturity, and

PT

in the substantia nigra with corresponding decreases in cell number and total protein quantity [22, 29-

RI

31]. In addition to characterizing the neuron, a couple of groups looked at neuronal plasticity in the n-3 FA deficient developing frog and rat. In the frog, a decrease in dendritic branching with a smaller

SC

dendritic arbor was observed and in the rat, an increase in axonal sprouting and a delay in their pruning was observed in one study and decreased growth cone formation in another [20, 32, 33]. Altered

NU

development of the neuron may be the result of a concomitant decrease in nerve growth factor, known to maintain survival and stimulate differentiation of cells [34].

MA

The modifications in lipid composition from n-3 FA deficiency are also dependent on diet [35-37], stage of development [38-40], age [41], sex [42], generational n-3 FA status and any combination of these factors. Generational n-3 FA status indicates whether the diet of the previous

D

generation(s) was also deficient in n-3 FA, for example, mice pups fed an n-3 FA–deficient diet,

TE

whose mothers were also fed the same diet, had a 21% reduction in DHA brain content [43] in comparison to studies using a multigenerational model or chronic studies of n-3 FA deficiency which

AC CE P

generate a larger decrease of ~75% in brain DHA content [44]. In general, the reduction in n-3 FA of the lipid membrane is met by a compensatory increase in docosapentaenoic acid (22:5 n-6) to maintain 22-C levels and consistent brain volume in white and gray matter during aging [31]. Overall, levels of DHA in the brain are lower in dietary n-3 FA deficient animals compared to controls in the frontal cortex, striatum and cerebellum. Human syndromes with inherent n-3 FA deficiency In humans, certain genetic disorders result in a deficiency of n-3 FA in afflicted individuals. Most notably, peroxisome biogenesis disorders (PBD) are characterized by biochemical abnormalities, including low DHA in the brain, retina and other tissues [45]. Peroxisomes perform the biogenesis of DHA and in Zellweger syndrome, the most severe PBD initially referred to as cerebro-hepato-renal syndrome [46], there is an absence of all peroxisomes from all organs. Children with Zellweger syndrome have a profound deficiency in DHA and the syndrome is usually fatal within the first year of life. Neuropathology of the disease has been studied using mouse models of Zellweger syndrome with brain-restricted deficiency of the peroxisome biogenesis factor (PEX) 13, a cytosolic docking factor in the peroxisome translocation machinery [47]. Pex13(-/-) mice reproduce many of the features of

ACCEPTED MANUSCRIPT Zellweger syndrome and PEX13 deficiency in humans [48]. The brains of these animals showed disordered lamination in the cerebral cortex, consistent with an observed developmental delay in neuronal migration leading to neuronal dysfunction. Mice homozygous for the targeted deletion of the peroxisome integral membrane protein, PEX2, developed the same delay in migration resulting in the

PT

development of mice with such severe cerebellar defects as abnormal Purkinje cell development and an

RI

altered folial pattern. Inbred mice with a PEX2 mutation show significant embryonic lethality and widespread neuronal lipidosis throughout the brain. Biochemical analysis of PEX2 mutant mice shows

SC

the characteristic accumulation of very long chain fatty acids as well as a deficiency in plasmalogen, a phospholipid essential for proper brain function. DHA levels were reduced in the brain of mutant mice

NU

[49]. Attempts to explain the neuropathogenesis have implicated peroxisomal metabolic dysfunction, and more specifically the loss of peroxisomal products, such as plasmalogens and DHA, and the

MA

accumulation of peroxisomal substrates, such as very-long-chain-fatty acids [50]. In a particular case of

D

human, LA deficiency of a 6 year old, similar neurological abnormalities were observed [51, 52].

TE

Behavioural Consequences of n-3 FA deficiency One of the most important outcomes of n-3 deficiency is that it negatively affects certain animal

AC CE P

behaviours presumably the result of structural and functional changes in the brain that manifest in learning and locomotor defects [37, 53-55]. These behaviours can be measured using established tests with measurable outcomes. Impaired learning performance has been observed in several experimental paradigms of n-3 deficiency using a number of behavioural measures including the Morris water maze, the Barnes Maze, elevated plus maze and the learning of olfactory or visual discrimination tasks [26, 37, 44, 56-58]. Impairments in learning were supported by the expression pattern of genes involved in cerebral activity. The expression of c-fos mRNA was induced in the olfactory bulb, piriform and neocortex, the areas of the brain depleted of n-3 fatty acids, whereas glucocorticoid-induced receptor and GLUT1 mRNA were absent in the piriform cortex, indicating a lack of cerebral activity in the area processing olfactory cues not detecting the smells [26]. The second generation (F2), n-3-deficient adult male rats had 82% decrease in DHA and showed impaired learning- and memory-related behaviors. This was observed by significantly more total errors in a 7-problem, 2-odor discrimination task compared to the n-3-adequate group [44]. F2 generation, n-3 deficient young adult mice had 50% decrease in DHA and had learning impairments via passive avoidance test [17] supporting evidence for learning impairment on habituation [54, 55] and

ACCEPTED MANUSCRIPT the elevated plus maze learning protocol [53]. Despite a 76% lower brain DHA, n-3-deficient rats were able to acquire most simple 2-odor discrimination tasks, but were deficient in the acquisition of a 20problem olfactory learning set. This deficit could not be attributed to changes in sensory capacity and

PT

rather instead, appeared to represent a deficit in higher order learning [59]. A neuroimaging study found that resting-state functional connectivity among prefrontal cortical

RI

networks was impaired in monkeys raised on an n-3 FA deficient diet compared with monkeys raised on fish oil-fortified diet [60]. Specifically, n-3 FA deficiency during perinatal development was

SC

associated with reduced resting-state connectivity between multiple regions in the brain. Altogether, impairment of learning discrimination tasks as well as MRI studies support the hypothesis that DHA is

NU

involved in the communication with structures involved in higher order learning involving multiple

MA

brain regions.

One study also reported that ALA deficiency reduced the appetite, which can thereafter lead to other nutrient deficiencies [61]. Mice pups from n-3 FA deficient mothers fed an n-3 FA-deficient diet had

D

reduced sensitivity to sucrose and a reduced ability to feel pleasure which can also lead to reduced

TE

appetite and further nutrient deficiencies [61, 62]. Due to the fact that multigenerational studies of n-3 FA deficiency have shown a larger decrease in n-3

AC CE P

FA accrual in the brain, a number of groups looked at the resultant behavioural outcomes. For instance, in a second generation n-3 FA deficient rat model, a model made to replicate the dietary deficiency of current adolescents today, cognitive and motivated behaviour impairments distinct from the deficits observed in adults were observed and these include behavioural measures that reflected hyperactivity, increased anxiety, increased goal-irrelevant and decreased goal-directed activity, and reduced behavioural flexibility [41]. These behavioral deficits in animals fed a diet deficient in n-3 FA could be because DHA deficiency impairs communication between neurons and glia of the brain and between the brain and the vascular system [63, 64]. Indeed, cortical DHA deficiency during development is also associated with neuroinflammation [65]. It was previously shown that n-3 FA-deficient rats have impaired communication between neurons and glia during the reduction of electrical brain activity following stimulation in addition to cortical spreading depression, something that is more pronounced in second generation n-3 FA deficiency in rats [66]. Deficiency in n-3 FA was also shown to affect brain energy metabolism by modulating glucose through the expression of brain glucose transporters, Glut1 and Glut3 [67-69] as well as mRNA of GLUT1 [26]. The level of Glut1 may be responsible for learning and memory deficits since neuronal activation requires an enhanced glucose demand [70].

ACCEPTED MANUSCRIPT Glucose transport was decreased by 30% and utilization via cytochrome oxidase activity was decreased by 20-40% in the three brain regions that share a high rate of energy metabolism [69]. Therefore, there are a number of studies supporting that n-3 FA deficiency leads to deficits in behavior and function of

PT

the brain.

RI

Effect of dietary n-3 FA deficiency on anxiety-like and depressive-like behaviours

SC

Nutritional n-3 FA deficiency produces anxiety and depressive-like symptoms [71-74]. Behavioural studies suggest that development deficits in brain DHA accrual are associated with elevated

NU

behavioural indices of depression that emerge after puberty [39, 75]. In contrast, dietary fish oil fortification significantly decreases depression-like behavior similar to antidepressant medications [76,

MA

77]. Perinatal n-3 FA deficiency is associated with increased home cage stereotypy and locomotion bouts [78] which is consistent with dysregulated mesolimbic dopamine activity. Recent neuroimaging findings suggest that low n-3 FA intake may impair cortical structural and

D

functional maturation in corticolimbic regions repeatedly implicated in psychopathology [79]. Indeed,

TE

rats subjected to n-3 FA deficiency during 15 weeks from weaning display higher depressive-like

AC CE P

symptoms in the forced swimming test [75, 80]. A few groups have reported that n-3 FA deficiency appears to abolish the role of endocannabinoid signaling in long-term potentiation in different areas of the brain required for the consolidation of memories [81-83] and behaviourally, these mice display increased anxiety-like behaviour [71]. Organisms such as the honey bee use spatial and time-dependent cues for disseminating information of food location to the hive. A diet deficient in n-3 FA delayed proper learning by conventional behavioural tests. In addition, the hypopharyngeal glands, used for collecting pollen, were smaller in the n-3 FA deficient bees which further hampers proper nutrient intake in this model [19]. Altogether, there is evidence in animal models suggesting that dietary n-3 FA deficiency contributes to anxietylike- and depressive-like behaviours.

N-3 deficiency and dysrupted Dopaminergic Pathway There are some studies that looked at dietary deficiency in n-3 FA and the dopaminergic system. Dopamine is a major neurotransmitter in the central nervous system (CNS) with three major projections arising from the dopamine perikarya in the substantia nigra and ventral tegmental area. The substantia

ACCEPTED MANUSCRIPT nigra plays an important role in reward and movement and the ventral tegmental area is critical to an organism's capacity to detect rewards and novelty and to enlist appropriate behavioural responses. A decrease in dopamine availability was observed in the frontal cortex of n-3 FA deficient rats [62, 84] as well as other areas of the brain [85] and is accompanied by a prolonged compensatory increase in

PT

dopaminergic receptor levels [86]. The increase in receptor levels continues to rise up to 4 weeks

RI

suggesting that the dopaminergic system is adaptive to n-3 FA deficiency although inadequate in the absence of supplementation. There is also evidence supporting the idea that n-3 FA deficiency in rats

SC

leads to a 40–75% lower level of endogenous dopamine in the frontal cortex according to age and a 10% reduction in density of dopaminergic D2 receptors [62]. Reduced central dopaminergic function

NU

could be the result of a structural change at the synapse. One group showed lower levels of dopamine release in the frontal cortex and nucleus accumbens because of a depletion of the dopamine vesicular

MA

storage pool in animals fed a deficient diet in n-3 FA [87]. It is noteworthy that not all areas of the brain, such as the corpus striatum and the striatum, have a dopaminergic vulnerability to n-3 FA deficiency [23, 88]. Impaired rat spatial learning in the Barnes maze [57] may be the behavioural

D

manifestation of altered dopamine release as well as impaired neurogenesis in the dentate gyrus of the

TE

hippocampus [89]. The mechanism leading to this modification is yet unknown and could involve vesicle turnover, membrane fluidity, or vesicular monoamine transporter. Another group conducted

AC CE P

studies in a second generation n-3 FA deficiency rat model due to its similarities in the trajectory of n-3 FA deficiency to that of the adolescents of today. The interest in adolescents stems from the reality that they are the second generation of humans that would have been born and first been exposed to n-3 FA deficiencies in the 1960s and 1970s (4-5 decades ago) when dietary trends toward decreased consumption of these fats began. The group found an elevation of dopamine synthesis specific to the dorsal striatum and unique to the adolescent stage. Surprisingly, in this model, a dietary n-3 FA deficiency across consecutive generations increased dopamine availability [41]. These findings are in agreement with recent reports of enhanced dopamine availability in young individuals at high risk of developing schizophrenia [90]. Hence, it is not clear if today’s increased rate of neurological disorders such as attention deficit hyperactivity disorder, autism and schizophrenia are linked to the current multigenerational n-3 FA deficiency or to better diagnosis at earlier stages.

E4 allele and DHA deficiency: E4 carriers might be particularly vulnerable to a dietary deficiency in n-3 FAs as demonstrated by an experiment we conducted on 14C-DHA uptake in the brain in which E4 mice were fed a diet deficient in n-3 FAs. The plasma DHA pool is critical to bring DHA to the brain

ACCEPTED MANUSCRIPT and it is dynamic, constantly exchanging FA with organs and tissues. Under conditions of chronic low dietary n-3 FA, the liver usually upregulates its ability to synthesize DHA and presumably receives ALA from the adipose tissue. Hence, it is suggested that there is an adipose-liver-brain FA axis [91]. We have shown that under an n-3 FA deficient diet, E4 mice had similar plasma DHA levels to that of

PT

E3 mice, but the liver and the adipose tissue DHA levels were ~46% lower in E4 mice than in E3 mice

RI

fed the same diet, suggesting that to maintain plasma DHA levels, E4 mice had to pull DHA from, or prevent DHA getting into, the liver and the adipose tissue (Figure 1) [92]. In terms of lipid

SC

metabolism, there is evidence for both decreased tissue uptake as well as a global metabolic shift towards lipid oxidation and enhanced thermogenesis in E4 mice. Specifically, one study showed E4

NU

mice displayed elevated markers of insulin resistance, a reduced respiratory quotient during the postprandial period (0.95±0.03 versus 1.06±0.03, P<0.001), indicating increased usage of lipids as

MA

opposed to carbohydrates as a fuel source. Finally, E4 mice showed increased body temperature (37.30±0.68°C versus 36.9±0.58°C, P=0.039), augmented cold tolerance and more metabolically active brown adipose tissue compared with E3 mice [93]. Furthermore, our group showed that E4 mice had

D

higher levels of liver carnitine palmitoyltransferase 1, a protein needed for the entrance of FA in the

TE

mitochondria and their oxidation thereafter [94].

AC CE P

In support of decreased uptake, another group found that a difference in a single amino acid between apoE4 and apoE3 changes the organization and stability of both the N-terminal helix bundle domain and separately folded C-terminal domain conferring an enhanced lipid binding ability of apoE4 over apoE3 to the surface of very low density lipoprotein particles. This impairs the lipolytic processing in the circulation so that apoE4 is associated with a more pro-atherogenic lipoprotein-cholesterol distribution (higher very low density lipoprotein-cholesterol/high density lipoprotein-cholesterol ratio) [95]. Therefore, in the long term, E4 carriers will have a flat reservoir of ALA and DHA, hence accentuating their vulnerability to n-3 FA deficiency. We have data in 8-12 months E4 mice showing that ALA levels in the liver and the adipose tissue were >80% and >40% lower, respectively, compared to the levels in E3 mice when fed a deficient diet in n-3 FA (Figure 1). Moreover, E4 mice fed the n-3 FA-deficient diet had ~80% lower liver ALA/LA ratio, than in E3 mice (control). This ratio gives an indication of Δ6 desaturase activity which is an indicator of the capacity to convert ALA in DHA. Hence, carrying the E4 allele tended to amplifies the DHA deficiency in the liver, adipose tissue and brain, and this process might be accentuated during aging because there is a loss of Δ6 desaturase activity [96].

ACCEPTED MANUSCRIPT Brain function in E4 carriers in cognitive decline: In E4 carriers, brain atrophy is more pronounced compared to that of the non-carriers [97]. In E4 mice, it was shown that vascular defects precede neuronal dysfunctions and can initiate neurodegeneration [1, 98]. In E4 mice, cerebral vascularization is reduced and vascular walls are thinner compare to mice carrying other APOE alleles [99]. This result

PT

is consistent with human post-mortem brain observations where the basement membrane surface area

RI

was lower in E4 carriers compared to the non-carriers [100]. One recent study conducted on postmortem human brains reported that pericyte degradation was accelerated in E4 carriers with AD > non

SC

carriers and with AD > non-AD control [101]. Pericytes maintain the integrity of the BBB and degenerate in AD. To our knowledge, it is not clear when disruption of BBB permeability and cerebral

NU

vascularization including the number, size and length of veins and arteries are initiated in humans and mice with E4 allele. Previous studies reported lower integrity of the BBB in E4 mice compared to E3

MA

mice [99].

D

Brain function in cognitively intact E4 carriers: Data from MRI studies seems to provide strong

TE

evidence of lower brain mass in AD-afflicted areas of healthy E4 carriers (not known to have Aβ plaques or phosphorylated tau tangles) compared to non-carriers. These brain-based studies have been

AC CE P

done with similar findings in neonates [102], infants (2-25 months), [103], and children and adolescents (aged 8– 20) [104]. The evidence on cognitive effects of the E4 allele in midlife however, has been mixed [105]. One study demonstrated that the E4 allele disrupts the resting state connectivity in MRI in the healthy older individuals (mean age = 62) [106]. In another study, there were no structural differences in hippocampal or frontal volumes with respect to E4 carriers in healthy professionals without any presenting memory problems and without selection for a family history of AD. The data does show that late-middle age and older E4 carriers have lower memory recall performance than non-carriers and that memory recall was correlated positively with hippocampal volume. Their findings point to basic memory testing as a sensitive tool for detecting E4-related influences on memory in aging workers [107]. However, most of those studies used standardized neuropsychological tests, which may not be sensitive to subtle cognitive change (e.g., Kozauer et al., [108] used the Mini-Mental State Exam) since deficits in working memory and semantic memory have been shown to be the earliest functions to undergo decline during the AD prodrome [109]. Greenwood et al., [110], have also previously validated an information-processing working memory task sensitive to APOE genotype in middle-age [111, 112].

ACCEPTED MANUSCRIPT Using an advanced dynamic contrast-enhanced magnetic resonance imaging protocol with high spatial and temporal resolution to quantify regional BBB permeability in humans, the team of Zlokovic reported age-dependent BBB permeability damage in the hippocampus and its CA1 and dentate gyrus subdivisions [3]. Cerebrospinal fluid markers of BBB permeability damages such as albumin quotient,

PT

fibrinogen and CypA seems to be greater in late-onset AD patients and E4 carriers compared with

RI

neurologically healthy human [113]. More importantly, young cognitively normal E4 carriers have impaired cerebrovascular reactivity in response to memory task and CO2 inhalation [114]. However,

SC

the age at which BBB permeability is damaged in E4 carriers remains elusive but having this information could help us to design intervention to limit progression of these damages. One potential

NU

intervention is to provide DHA in the diet of E4 carriers but this may be conditional to DHA efficiency. Providing DHA in the diet has recently been shown to improve cerebrovascular parameters in several

MA

different scenarios. In a rat model of transient focal cerebral ischemia, the integrity of the BBB was improved by a reduction in the expression of the intercellular adhesion molecule-1, an increase in the

D

expression of collagen IV and a decrease in the extravasation of fluid from the vessels into the tissue in

TE

DHA-treated rats [115]. Another group, used transgenic Mfsd2a-/- mice [4] and showed that genetic deletion of Mfsd2a leads to BBB defects in the developing CNS. Furthermore, Nguyen et al. [116]

AC CE P

showed that Mfsd2a-/- mice have significantly reduced levels of DHA in brain accompanied with neuronal cell loss in the hippocampus and cerebellum, neurological and behavioral deficits, and reduced brain size. Mfsd2a is responsible for establishing a unique lipid environment that inhibits caveolae vesicle formation in CNS endothelial cells to suppress transcytosis and ensure BBB integrity [117]. Addullah et al., [118], observed an E4 dependent difference in mfsd2a expression and DHAcontaining phospholipids. Among AD patients, E4 homozygotes had lower expression of mfsd2a than E4 heterozygotes and non-carriers and lower of expression of mfsd2a correlated to lower quantity of DHA and a higher quantity of AA in the phospholipids of the membrane [119]. We observed a shorter latency in the Barnes maze test in E4 mice fed a DHA diet as compared to E4 carriers fed a control diet [120]. One could hypothesize the improvement in cognitive decline may be the result of an interaction with DHA and the BBB but this has yet to be investigated. One of the primary events in cognitive decline is a breakdown of the BBB and neuroinflammation so potentially, providing DHA in the diet to E4 carriers could restore the integrity of the BBB if provided prior to the irreversible stage of decline [121].

ACCEPTED MANUSCRIPT E4 allele modifies brain DHA levels? APOE regulates lipid transport and metabolism [122]. The brain has its own pool of apoE [123] that plays critical roles in lipid transport to neurons. Using in situ intracerebral perfusion, we showed that brain uptake of 14C-DHA was 24% lower in 4- and 13-monthold E4 mice compared to uptake in E2 mice but that cortex DHA was significantly lower only in 13-

PT

month-old E4 mice. This suggests that in E4 mice, lower cortex DHA levels are age-dependent but

RI

plasma DHA relative % was higher in E4 mice [124]. This pattern i.e., lower brain DHA uptake, was observed in mice knocked-down for the Mfsd2a transporter [6]. In the study using

14

C-DHA brain

SC

uptake, mice were fed a diet deficient in n-3 FA. Therefore, DHA imbalance in E4 mice might contribute to the loss of BBB integrity in E4 carriers. Yassine et al also proposes two other mechanisms

NU

of how the E4 allele can be involved in brain DHA levels. One of these mechanisms is an increased level of liver catabolism of DHA in E4 carriers compared to non-carriers as it was outlined by

MA

Chouinard-Watkins et al [125]. The other mechanism suggested by this group is a hypolipidated, or decreased number of, apoE particles leading to less efficient transport of DHA in the brain [126]. There is also an association between DHA and A deposition in pre-dementia [127] but not during or after

TE

D

the disease has expressed suggesting that timing is important to intervene to delay onset of the disease.

AC CE P

Is it possible to rebalance n-3 FA deficiencies and to reverse neurological consequences? There is ample evidence in the literature that n-3 FA supplementation following deficiency allows for the rebalancing of the fatty acid composition in the cell membranes and capillaries of the brain in animal models and human syndromes of n-3 deficiency when administered in the therapeutic window. Studies of DHA repletion include but are not limited to, the monkey [128, 129], maternal rats [130], F2 generation rats of DHA deficiency [131] and Zellweger syndrome [45, 132]. The rebalancing of DHA in the brain after a period of n-3 FA deficiency occurs over time in the young adult F2 generation rats studied with partial recovery of 20% of the n-3 adequate group at 1 week rising to 35% at 2 weeks and completely resolving at 8 weeks after implementation of the DHA repletion diet [131]. The slow reversal of DHA accrual in the brain is in contrast to the repletion levels of DHA in the serum and liver were approximately 90% and 100% replaced, respectively, within 2 weeks of diet repletion. Similar results were obtained in intrauterine n-3 FA deficient monkeys, depleted of 70-90% of n-3 FA at birth, requiring 15 weeks for total recovery of DHA in the cerebral cortex in contrast to the required 4 weeks and 8 weeks for plasma and erythrocytes, respectively [128]. The difference in the amount and time course of DHA accrual in the nervous system compared to the liver and circulation

ACCEPTED MANUSCRIPT suggests that transport-related processes may limit the rate of DHA repletion in the brain. There was a recovery of n-3 FA in the capillaries of an adult rat after substitution with a non-deficient diet however, the recovery was slow and resolved over 2.5 months in contrast to a relatively immediate recovery in the choroid plexus [28]. Switching to a fish oil-supplemented diet induced a recovery in DHA content

PT

in the frog embryos within 20 weeks and diminished the deprivation effects observed on neurons

RI

involved in vision [20]. In maternal rats, fish oil supplementation on an n-3 FA deficient diet during pregnancy and lactation, prevents depletion of maternal brain regional DHA levels, and reduces anxiety

SC

as observed in the elevated plus maze test [133]. In aged mice, there were no significant differences between those fed an n-3 FA deficient diet throughout their lifespan and those supplemented with ALA

NU

after an n-3 FA deficient diet in the open field test, used to assess capability and drive in exploratory activity [134]. However, the impaired learning performance observed in Morris water maze in mice fed

MA

the n-3 deficient diet was completely restored in old n-3 FA deficient mice supplemented with DHA [56]. In human studies of patients afflicted with Zellweger Syndrome, normalization of DHA levels in the brain was achieved by a quantity of 100-500 mg/day of DHA ethyl ester (DHAEE) for variable

D

periods of time. The daily doses varied between 100 mg and 500 mg of DHAEE, depending on age and

TE

degree of DHA deficiency, but never on a body-weight basis. This was done because the brain has the largest DHA requirements during early development; so that the youngest patients with a marked DHA

AC CE P

deficiency after regular plasma fatty acid analysis received the highest doses. Improvement was observed in myelination via MRI and this translated into a marked improvement in liver function, growth, muscle tone, vision, and social contact in those patients, most significantly when the treatment was started before 6 months of age [45]. In contrast, DHA supplementation of 100 mg/kg/day did not improve the growth of treated individuals with PBD with an inherent n-3 deficiency during an average of 1 year of follow-up in patients aged 1 to 144 months [135]. A similar result occurred in a patient with classic Zellweger syndrome treated with DHAEE for three months during which the patient showed no clear clinical improvement over the short period of treatment. Five other patients with Zellweger variants (four of them under one-year-old and a five-year-old) were also treated with DHAEE until normalization of the DHA levels in erythrocytes and in general, had accelerated developmental curves with the infants becoming more alert and acquiring better social contact [136]. Follow-up to these patients, MRI has shown advances in brain myelination [137]. In the case of the 6 year old child with human ALA deficiency, when the diet was supplemented to an emulsion containing linolenic acid, the neurological symptoms disappeared [51].

ACCEPTED MANUSCRIPT At the molecular level, dysregulation of multiple neurotransmitter systems in cortical DHA deficiency is reversible with early, but not later, postnatal n-3 FA supplementation [38, 138]. For example, n-3 FA supplementation before weaning to n-3 deficient rats, is able to reverse changes in serotonergic transmission however, increased basal serotonergic transmission and decreased stimulated release

PT

persisted if supplementation was introduced after weaning [138]. Therefore, there is evidence

RI

supporting the idea that tissue n-3 FA deficiency can be rebalanced if n-3 FA are provided in the diet at early stages and doing so, may reverse some of the neurological consequences caused by the n-3

SC

deficiency. In humans, there is evidence that an increase in consumption of DHA via supplementation could improve cognitive decline by decreasing the level of inflammation in the brain. One study

NU

demonstrated the ability of DHA to transfer from the plasma to the CSF in AD patients and to modify the level of inflammation in the brain. The increase of DHA in CSF, and therefore uptake in the brain,

MA

was inversely correlated with CSF levels of total and phosphorylated tau markers of AD and directly correlated with soluble interleukin-1 receptor type II, an anti-inflammatory marker. [139]. Thus, as DHA increased in CSF, markers of AD decreased and anti-inflammatory markers increased [139].

D

Another group demonstrated the ability of EPA and DHA to reduce inflammation by increasing

TE

phagocytosis of Alzheimer’s disease related-Aβ42 by human microglia, shown by decreasing proinflammatory markers CD40 and CD36 and increasing production of brain-derived neurotrophic factor

AC CE P

[140]. However, the direct link between plasma DHA, brain DHA and risk of cognitive decline is not crystal-clear. For instance, vegetarians have 50% lower plasma levels of n-3 FA compared to that of omnivores but the overall mortality in vegetarians does not differ significantly from that of omnivores [141]. Therefore, blood DHA levels may not be linearly related to tissue DHA levels. Moreover, one would have thought that the n-6:n-3 ratio of the vegetarian diet would be lower than that of the Western diet, which would reduce competition for Δ6 desaturase and improve conversion of ALA to EPA and DHA. However, Kornsteiner et al [142] reported that in Austria, the vegetarian and vegan diet have an n-6:n-3 ratio of 10:1. Therefore, the n-6:n-3 ratio is not tremendously lower in vegetarians and vegans compared to those eating the Western diet hence suggesting that there is probably something else than dietary fatty acids in the diet somehow protecting vegetarians and vegans from chronic diseases and risk of cognitive decline. Also important, long-term dietary restriction of n-3 FA such as following a vegetarian diet, could upregulate the expression and activity of the enzymes elongase and desaturase increasing its sensitivity, as found in studies of cats and rodents in order for the liver to make DHA and transport it to the brain thereby maintaining cerebral levels [143, 144]. The relation of plasma DHA to brain DHA concentration and metabolism in humans has been previously investigated. There was a slight correlation between plasma and brain n-3 FA with DHA and one area of the brain but only in

ACCEPTED MANUSCRIPT non-cognitively impaired indicating a tenuous link between plasma and tissue levels in the healthy human brain [146]. Despite apparently low DHA intake in AD, brain DHA levels are frequently the same as in controls, suggesting that low DHA intake results in low plasma DHA but does not necessarily reduce brain DHA in humans [145, 146]. One study determined the change in whole-body

PT

DHA synthesis rates from varied amounts of dietary ALA [147]. Rats fed a diet with adequate ALA

RI

had a 2-fold greater capacity to synthesize DHA than did rats fed either a diet rich, or deficient, in ALA and a DHA synthesis rate that was similar to that of rats fed the diet high in ALA. However, rats fed

SC

the diet deficient in ALA had a 750% lower DHA synthesis rate than rats fed the diet adequate or rich in ALA. Therefore, increasing dietary ALA from 3% to 10% of FA did not increase DHA synthesis

NU

rates, because of a decreased capacity to synthesize DHA in rats fed the diet high in ALA. Another study using intakes, food sources, and status of n−3 FAs according to dietary habit (fish-eaters and

MA

non-fish-eating meat-eaters, vegetarians, or vegans), estimated the conversion between dietary ALA and circulating n−3 FAs [148]. They found n-3 FA intakes in non-fish-eaters were 57-80% of those in fish-eaters, but tissue differences were considerably smaller potentially due to a higher conversion rate

TE

D

of ALA with low intake rates.

AC CE P

Is it possible to rebalance n-3 FA metabolic disturbances in E4 mice? In a recent experiment, we fed E4 mice a control diet containing ALA (0.14%) or a diet rich in DHA (0.7%). In mice fed the control diet, E4 mice had twice as much DHA in the liver compared to E3 mice (Figure 2). ALA levels in the liver were lower, but Δ6 desaturase activity was also estimated to be lower in E4 mice than in E3 mice, suggesting that a greater conversion of ALA to DHA did not explain the higher levels of DHA in the liver of E4 mice fed the control diet. The only possible explanation we have to date is that the FA binding protein 1 was upregulated in the livers of E4 mice fed the control diet. FA binding protein 1 has a critical role in FA uptake and intracellular transport together with regulating lipid metabolism and cellular signaling pathways [149]. However, E4 mice fed the control diet had plasma and cortex DHA levels similar to those of E3 mice fed the same diet suggesting that the E4-specific disruption in liver DHA did not contribute to lower cortical DHA levels. When fed the DHA diet, E4 mice had or tended to have lower plasma, liver and cortex DHA levels than did E3 mice (Figure 2). Despite the fact that E4 mice fed the DHA diet had more than twice the level of plasma DHA than when receiving the control diet, cortex DHA levels barely increased. We propose that this was because E4 carriers have poor brain protection, poor brain repair mechanisms, lower neurogenesis and lower vascular function as well as lower limitations. Hence, providing DHA in mice vulnerable to DHA deficiency helps to minimize

ACCEPTED MANUSCRIPT these detrimental processes. In support of this hypothesis, we tested the behaviour of 12-month old E3 (n = 32) and E4 (n = 38) fed a control or a DHA diet. Our results suggest that feeding DHA to E4 mice prevented spatial memory deficits due to the observation that escape latency of E4 mice in the Barnes maze was equal to E3 mice in the DHA supplemented group in contrast to a longer latency in E4 mice

PT

compared to E3 mice fed a control diet [120]. These results were independent of anxiety, as tested by

RI

the light and dark test, but were accompanied by modifications of hippocampal and liver AA content. Indeed, we reported a significant genotype x diet interaction for the levels of AA in the liver and the

SC

hippocampus where the DHA diet suppressed the E4-specific excess of AA in the liver and the hippocampus [120, 150]. The results of these studies suggest that AA metabolic disturbances might

NU

play a crucial role in the cognitive deficits reported in E4 mice, which in turn, could contribute to disrupt DHA metabolism since both AA and DHA compete for the same enzymatic pathways [151]. In

MA

another study, E4 mice, receiving a dietary treatment of Fortasyn (a supplementation including docosahexaenoic acid, eicosapentaenoic acid, uridine, choline, phospholipids, folic acid, vitamins B12, B6, C, and E, and selenium) ameliorated the reduced cerebral blood flow, accelerated synaptic loss,

D

improved white matter integrity and functional connectivity in both aging E4 and wild type mice as

TE

well as illustrating enhanced protective mechanisms on vascular and synapse health [36]. The results were found to be independent of APOE genotype however, genotype effects could have been masked

AC CE P

by the inclusion of coconut oil into the control diet. Coconut oil contains medium chain triglycerides and 5% PUFA. Medium chain triglycerides produce ketone bodies which is an alternative fuel to glucose for the brain [152]. Therefore, we believe that the use of coconut oil has largely biased this study design. Recently, our group also provided evidence that plasma and liver cholesterol levels are modified by the APOE genotype and diet and these findings should be considered when designing animal trials with E4 mice [153].

E4 carriers and DHA kinetics: Our group has consistently reported that DHA metabolism is disrupted in E4 carriers. In humans not supplemented with n-3 FA, E4 carriers have approximately twice as much DHA in the plasma triglycerides than the non-carriers. In a trial in which DHA uniformly labelled with carbon 13 (13C-DHA) and provided as a single oral dose of 40mg, the mean plasma 13C-DHA level in E4 carriers was 31% lower than in the non-carriers but oxidation in E4 carriers. Plasma

13

13

C-DHA was 50% more degraded via -

C-DHA half-life was similar between E4 carriers and non-carriers,

but the whole-body half-life in E4 carriers was 32 days vs. 140 days in the non-carriers [125]. The discrepancy between plasma

13

C-DHA half-life vs whole-body half-life suggests that, in E4 carriers

ACCEPTED MANUSCRIPT and compared to the non-carriers,

13

C-DHA might be cleared preferentially from non-plasma stores

such as the liver or the adipose, but this hypothesis needs to be confirmed. Then, 13C-DHA kinetics in E4 carriers and non-carriers was evaluated again after they were supplemented with 3 g/d of n-3 FAs for 5 months to assess if a dietary supplementation with n-3 FAs could prevent DHA metabolic 13

C-DHA in the plasma than the non-carriers, suggesting a slower clearance of DHA

RI

accumulation of

PT

disturbances in E4 carrier. In that context, we showed that E4 carriers (n = 4) had a greater

to and/or use by tissues, and this was opposite to what we reported in E4 participants not supplemented

SC

with n-3 FA [154]. Hence, when given a high dose of DHA, 13C-DHA kinetics were rebalanced in E4 carriers. In a more recent trial, we outlined that disturbances in DHA metabolism in E4 carriers might

NU

be dependent on body mass index [155]. In that study, E4 carriers were lower plasma responders to a DHA supplement compared to the non-carriers, but only if they were overweight.

MA

The brain has a limited capacity to produce DHA de novo and obtains DHA from the plasma. Recently, the orphan receptor, Mfsd2a was identified as a transporter of DHA esterified to lysophosphatidylcholine from the plasma and a receptor involved in BBB integrity. One group reported

D

lower Mfsd2a transporter levels in E4 homozygotes AD patients compared to control and

TE

heterozygotes E4 AD patients [119]. This group has also performed experiments in an in vitro BBB

AC CE P

model in the presence of different apoE isoforms. They showed that the brain apical-to-basolateral transit of AA and DHA is lower when using human serum of homozygotes for E4 allele compared to homozygotes for E3 allele [118]. In addition, another group found Msfd2a-deficient mice exhibit a substantial reduction in brain DHA levels due to deficiency in a membrane transporter involved in endothelial cell uptake of esterified DHA [116]. Therefore, providing a DHA supplement rebalances plasma DHA and might contribute to the prevention in the loss of BBB integrity in aging E4 carriers. However, the plasma pool into which DHA is incorporated might be key to this success since the nonesterified FA and the lysophosphatidylcoline pool, the latter being the preferred pool for Mfsd2a transporter, might be more important than the other plasma pools [156]. Recently, the team of Yassine and collaborators suggested that a high dose of DHA supplementation in E4 carriers might be a promising strategy to prevent development of AD but this treatment has to be provided before onset of any clinical manifestation of AD [126]. Our efforts for designing better clinical trials and combining our trials with state-of-the art technology will certainly help our team to understand the timespecificities where we have more chances to succeed by providing DHA supplementation at optimal doses for this at-risk population of developing late-onset AD.

ACCEPTED MANUSCRIPT Conclusion Altogether, this review provided evidence that dietary n-3 FA deficiencies lead to behavioural and functional deficits. E4 carriers may be more vulnerable to these dietary n-3 FA deficits because they

PT

catabolise more DHA and DHA is acquired mainly through the diet. It seems that to maintain a certain level of DHA in the plasma of E4 carriers, DHA and ALA are pulled from other organs and tissues

RI

such as the liver and the adipose. However, providing exogenous DHA in the diet seems to rebalance some of the behavioural and functional deficits in E4 mice and improve clinical outcomes of patients

SC

with inherent DHA deficiencies early in brain development when DHA accretion is maximal. DHA may have multiple roles beyond its incorporation into brain membranes and a need for understanding

NU

these diverse roles and their involvement in the prevention of AD is critical. Since there are no efficient drugs for treating the symptoms of AD or limiting the progression of the disease, we believe that

AC CE P

TE

D

MA

preventative strategies are urgently needed in this area.

ACCEPTED MANUSCRIPT Acknowledgments The authors declare no conflict of interest. Supported by the Canadian Institutes of Health Research (MOP119454) and the Natural Sciences and Engineering Research Council of Canada for the project funding, a Fonds de la recherche du Québec-Santé for a Junior 2 salary award to MP. MP also

PT

holds a New Investigator salary award from the Canadian Institutes for Health Research and a Chair on

AC CE P

TE

D

MA

NU

SC

RI

lipid metabolism during aging donated by Center of Medical Research of University of Sherbrooke.

ACCEPTED MANUSCRIPT References [1] B.V. Zlokovic, Cerebrovascular effects of apolipoprotein E: implications for Alzheimer disease, JAMA Neurol, 70 (2013) 440-444. [2] E. Kok, S. Haikonen, T. Luoto, H. Huhtala, S. Goebeler, H. Haapasalo, P.J. Karhunen, Apolipoprotein E-

PT

dependent accumulation of Alzheimer disease-related lesions begins in middle age, Ann Neurol, 65 (2009) 650657.

RI

[3] A. Montagne, S.R. Barnes, M.D. Sweeney, M.R. Halliday, A.P. Sagare, Z. Zhao, A.W. Toga, R.E. Jacobs, C.Y.

SC

Liu, L. Amezcua, M.G. Harrington, H.C. Chui, M. Law, B.V. Zlokovic, Blood-brain barrier breakdown in the aging human hippocampus, Neuron, 85 (2015) 296-302.

NU

[4] A. Ben-Zvi, B. Lacoste, E. Kur, B.J. Andreone, Y. Mayshar, H. Yan, C. Gu, Mfsd2a is critical for the formation and function of the blood-brain barrier, Nature, 509 (2014) 507-511.

MA

[5] L.N. Nguyen, D. Ma, G. Shui, P. Wong, A. Cazenave-Gassiot, X. Zhang, M.R. Wenk, E.L. Goh, D.L. Silver, Mfsd2a is a transporter for the essential omega-3 fatty acid docosahexaenoic acid, Nature, 509 (2014) 503-506. [6] A. Guemez-Gamboa, L.N. Nguyen, H. Yang, M.S. Zaki, M. Kara, T. Ben-Omran, N. Akizu, R.O. Rosti, B. Rosti, E.

D

Scott, J. Schroth, B. Copeland, K.K. Vaux, A. Cazenave-Gassiot, D.Q. Quek, B.H. Wong, B.C. Tan, M.R. Wenk, M.

TE

Gunel, S. Gabriel, N.C. Chi, D.L. Silver, J.G. Gleeson, Inactivating mutations in MFSD2A, required for omega-3 fatty acid transport in brain, cause a lethal microcephaly syndrome, Nat Genet, 47 (2015) 809-813.

AC CE P

[7] W.S. Harris, D.M. Klurfeld, Twentieth-century trends in essential fatty acid intakes and the predicted omega3 index: evidence versus estimates, The American Journal of Clinical Nutrition, 93 (2011) 907-908. [8] T.L. Blasbalg, J.R. Hibbeln, C.E. Ramsden, S.F. Majchrzak, R.R. Rawlings, Changes in consumption of omega-3 and omega-6 fatty acids in the United States during the 20th century, The American Journal of Clinical Nutrition, 93 (2011) 950-962.

[9] M.D. Andriamampandry, C. Leray, M. Freund, J.-P. Cazenave, C. Gachet, Antithrombotic Effects of (n-3) Polyunsaturated Fatty Acids in Rat Models of Arterial and Venous Thrombosis, Thrombosis Research, 93 (1999) 9-16. [10] E. Anil, The impact of EPA and DHA on blood lipids and lipoprotein metabolism: Influence of apoE genotype. , Proceedings of the Nutrition Society 66 (2007) 9. [11] P.C. Calder, R.F. Grimble, Polyunsaturated fatty acids, inflammation and immunity., Eur J Clin Nutr., ;56 (2002) 6. [12] C.R. Hooijmans, F. Rutters, P.J. Dederen, G. Gambarota, A. Veltien, T. van Groen, L.M. Broersen, D. Lutjohann, A. Heerschap, H. Tanila, A.J. Kiliaan, Changes in cerebral blood volume and amyloid pathology in aged Alzheimer APP/PS1 mice on a docosahexaenoic acid (DHA) diet or cholesterol enriched Typical Western Diet (TWD), Neurobiol Dis, 28 (2007) 16-29.

ACCEPTED MANUSCRIPT [13] M. Oksman, H. Iivonen, E. Hogyes, Z. Amtul, B. Penke, I. Leenders, L. Broersen, D. Lütjohann, T. Hartmann, H. Tanila, Impact of different saturated fatty acid, polyunsaturated fatty acid and cholesterol containing diets on beta-amyloid accumulation in APP/PS1 transgenic mice, Neurobiology of Disease, 23 (2006) 563-572. [14] A.P. Simopoulos, Evolutionary aspects of diet, the omega-6/omega-3 ratio and genetic variation:

PT

nutritional implications for chronic diseases, Biomedicine & Pharmacotherapy, 60 (2006) 502-507. [15] M. Loef, H. Walach, The Omega-6/Omega-3 Ratio and Dementia or Cognitive Decline: A Systematic Review

RI

on Human Studies and Biological Evidence, Journal of Nutrition in Gerontology and Geriatrics, 32 (2013) 1-23.

SC

[16] Neuringer M, Anderson GJ, C. WE., The essentiality of n-3 fatty acids for the development and function of the retina and brain., Annu Rev Nutr., 8 (1988) 25.

NU

[17] I. Carrié, M. Clément, D. De Javel, H. Francès, J.M. Bourre, Learning deficits in first generation OF1 mice deficient in (n-3) polyunsaturated fatty acids do not result from visual alteration, Neuroscience Letters, 266 (1999) 69-72.

MA

[18] H.S. Weisinger, A.J. Vingrys, L. Abedin, A.J. Sinclair, Effect of diet on the rate of depletion of n-3 fatty acids in the retina of the guinea pig.

D

[19] Y. Arien, A. Dag, S. Zarchin, T. Masci, S. Shafir, Omega-3 deficiency impairs honey bee learning, PNAS

TE

Proceedings of the National Academy of Sciences of the United States of America, 112 (2015) 15761-15766. [20] M. Igarashi, R.A. Santos, S. Cohen-Cory, Impact of Maternal n-3 Polyunsaturated Fatty Acid Deficiency on

AC CE P

Dendritic Arbor Morphology and Connectivity of Developing Xenopus laevis Central Neurons In Vivo, The Journal of Neuroscience, 35 (2015) 6079-6092. [21] Tinoco J, Babcock R, Hincenbergs I, Medwadowski B, M. P., Linolenic acid deficiency: changes in fatty acid patterns in female and male rats raised on a linolenic acid-deficient diet for two generations., Lipids, 13 (1978) 12.

[22] A. Ahmad, M. Murthy, R.S. Greiner, T. Moriguchi, N. Salem, A Decrease in Cell Size Accompanies a Loss of Docosahexaenoate in the Rat Hippocampus, Nutritional Neuroscience, 5 (2002) 103-113. [23] H.D. Cardoso, P.P. Passos, E.F. Santos Jr, R.S. Oliveira, P.E.L. Oliveira, R.C.F. de Santos, D.F. Santana, M.C.A. Rodrigues, B.L.S. Andrade da Costa, C.J. Lagranha, A.C. Ferraz, J.M.C. Borba, A.P. Rocha-de-Melo, R.C.A. Guedes, D.M.A.F. Navarro, G.K.N. Santos, R. Borner, C.W. Picanço-Diniz, E.I. Beltrão, J.F. Silva, Differential vulnerability of substantia nigra and corpus striatum to oxidative insult induced by reduced dietary levels of essential fatty acids, Frontiers in Human Neuroscience, (2012). [24] M.C. Soares, M.L. Aléssio, C.L. Léger, M.T. Bluet-Pajot, H. Clauser, A. Enjalbert, C. Kordon, D.E. Wandscheer, Effect of essential fatty acid deficiency on membrane fatty acid content and growth hormone stimulation of rat pituitaries during postnatal development, Journal of Lipid Research, 36 (1995) 1401-1406.

ACCEPTED MANUSCRIPT [25] S. Favrelière, L. Barrier, G. Durand, S. Chalon, C. Tallineau, Chronic dietary n-3 polyunsaturated fatty acids deficiency affects the fatty acid composition of plasmenylethanolamine and phosphatidylethanolamine differently in rat frontal cortex, striatum, and cerebellum, Lipids, 33 (1998) 401-407. [26] A. Hichami, F. Datiche, S. Ullah, F. Liénard, J.-M. Chardigny, M. Cattarelli, N.A. Khan, Olfactory

PT

discrimination ability and brain expression of c-fos, Gir and Glut1 mRNA are altered in n − 3 fatty acid-depleted rats, Behavioural Brain Research, 184 (2007) 1-10.

RI

[27] M. Foot, T.F. Cruz, M.T. Clandinin, Influence of dietary fat on the lipid composition of rat brain

SC

synaptosomal and microsomal membranes, Biochemical Journal, 208 (1982) 631-640. [28] P. Homayoun, G. Durand, G. Pascal, J.M. Bourre, Alteration in Fatty Acid Composition of Adult Rat Brain

NU

Capillaries and Choroid Plexus Induced by a Diet Deficient in n-3 Fatty Acids: Slow Recovery After Substitution with a Nondeficient Diet, Journal of Neurochemistry, 51 (1988) 45-48. [29] A. Ahmad, T. Moriguchi, J.N. Salem, Original article: Decrease in neuron size in docosahexaenoic acid-

MA

deficient brain, Pediatric Neurology, 26 (2002) 210-218.

[30] P.P. Passos, J.M.C. Borba, A.P. Rocha-de-Melo, R.C.A. Guedes, R.P. da Silva, W.T.M. Filho, K.M.M. Gouveia,

D

D.M.d.A.F. Navarro, G.K.N. Santos, R. Borner, C.W. Picanço-Diniz, J.A. Pereira, M.S.M. de Oliveira Costa, M.C.A.

TE

Rodrigues, B.L.d.S. Andrade-da-Costa, Dopaminergic cell populations of the rat substantia nigra are differentially affected by essential fatty acid dietary restriction over two generations, Journal of Chemical

AC CE P

Neuroanatomy, 44 (2012) 66-75.

[31] A. Ahmad, R. Momenan, P. van Gelderen, T. Moriguchi, R.S. Greiner, N. Salem Jr, Gray and White Matter Brain Volume in Aged Rats Raised on n -3 Fatty Acid Deficient Diets, Nutritional Neuroscience, 7 (2004) 13-20. [32] P.C. de Velasco, H.R. Mendonça, J.M.C. Borba, B.L.d.S. Andrade da Costa, R.C.A. Guedes, D.M.d.A.F. Navarro, G.K.N. Santos, A.d.C. Faria-Melibeu, P. Campello Costa, C.A. Serfaty, Nutritional restriction of omega-3 fatty acids alters topographical fine tuning and leads to a delay in the critical period in the rodent visual system, Experimental Neurology, 234 (2012) 220-229. [33] S.M. Innis, S. Presa Owens, Dietary fatty acid composition in pregnancy alters neurite membrane fatty acids and dopamine in newborn rat brain, J Nutr., 131 (2001). [34] A. Ikemoto, A. Nitta, S. Furukawa, M. Ohishi, A. Nakamura, Y. Fujii, H. Okuyama, Dietary n-3 fatty acid deficiency decreases nerve growth factor content in rat hippocampus, Neuroscience Letters, 285 (2000) 99102. [35] H. Tsukada, T. Kakiuchi, D. Fukumoto, S. Nishiyama, K. Koga, Docosahexaenoic acid (DHA) improves the age-related impairment of the coupling mechanism between neuronal activation and functional cerebral blood flow response: a PET study in conscious monkeys, Brain Research, 862 (2000) 180-186.

ACCEPTED MANUSCRIPT [36] M. Wiesmann, V. Zerbi, D. Jansen, R. Haast, D. Lütjohann, L.M. Broersen, A. Heerschap, A.J. Kiliaan, A Dietary Treatment Improves Cerebral Blood Flow and Brain Connectivity in Aging apoE4 Mice, Neural Plasticity, 2016 (2016) 6846721. [37] N. Yamamoto, A. Hashimoto, Y. Takemoto, H. Okuyama, M. Nomura, R. Kitajima, T. Togashi, Y. Tamai,

PT

Effect of the dietary alpha-linolenate/linoleate balance on lipid compositions and learning ability of rats. II. Discrimination process, extinction process, and glycolipid compositions, Journal of Lipid Research, 29 (1988)

RI

1013-1021.

SC

[38] E. Kodas, S. Vancassel, B. Lejeune, D. Guilloteau, S. Chalon, Reversibility of n-3 fatty acid deficiencyinduced changes in dopaminergic neurotransmission in rats: critical role of developmental stage, Journal of

NU

Lipid Research, 43 (2002) 1209-1219.

[39] M.J. Weiser, K. Wynalda, N. Salem, C.M. Butt, Dietary DHA during development affects depression-like behaviors and biomarkers that emerge after puberty in adolescent rats, J Lipid Res, 56 (2015).

MA

[40] P.C. Velasco, H.R. Mendonça, J.M. Borba, B.L. Andrade da Costa, R.C. Guedes, D.M. Navarro, Nutritional restriction of omega-3 fatty acids alters topographical fine tuning and leads to a delay in the critical period in

D

the rodent visual system, Exp Neurol, 234 (2012).

TE

[41] C.O. Bondi, A.Y. Taha, J.L. Tock, N.K.B. Totah, C. Yewon, G.E. Torres, S.I. Rapoport, B. Moghaddam, Adolescent Behavior and Dopamine Availability Are Uniquely Sensitive to Dietary Omega-3 Fatty Acid

AC CE P

Deficiency (English), Biological psychiatry (1969), 75 (2014) 38-46. [42] B. Levant, M.K. Ozias, S.E. Carlson, Sex-specific effects of brain LC-PUFA composition on locomotor activity in rats, Physiology & Behavior, 89 (2006) 196-204. [43] B. Levant, J.D. Radel, S.E. Carlson, Reduced Brain DHA Content After a Single Reproductive Cycle in Female Rats Fed a Diet Deficient in N-3 Polyunsaturated Fatty Acids, Biological Psychiatry, 60 (2006) 987-990. [44] Greiner RS, Moriguchi T, Hutton A, Slotnick BM, S.N. Jr., Rats with low levels of brain docosahexaenoic acid show impaired performance in olfactory-based and spatial learning tasks. , Lipids, 34 Suppl (1999) 4. [45] M. Martinez, Restoring the DHA levels in the brains of zellweger patients, Journal of Molecular Neuroscience, 16 (2001) 309-316. [46] S. Goldfisher, C.L. Moore, A.B. Johnson, A.J. Spiro, M.P. Valsamis, H.K. Wisniewski, R.H. Ritch, W.T. Norton, I. Rapin, L.M. Gartner, Peroxisomal and mitochondrial defects in the cerebro-hepato-renal syndrome, Science, 182 (1973). [47] R.S. Rahim, A.C.B. Meedeniya, D.I. Crane, Central serotonergic neuron deficiency in a mouse model of Zellweger syndrome, Neuroscience, 274 (2014) 229-241.

ACCEPTED MANUSCRIPT [48] Maxwell M, Bjorkman J, Nguyen T, Sharp P, Finnie J, Paterson C, Tonks I, Paton BC, Kay GF, C. DI., Pex13 inactivation in the mouse disrupts peroxisome biogenesis and leads to a Zellweger syndrome phenotype., Mol. Cell. Biol., 23 (2003) 11. [49] P.L. Faust, H.-M. Su, A. Moser, H.W. Moser, The peroxisome deficient PEX2 zellweger mouse, Journal of

PT

Molecular Neuroscience, 16 (2001) 289-297.

[50] D.I. Crane, Revisiting the neuropathogenesis of Zellweger syndrome, Neurochemistry International, 69

RI

(2014) 1-8.

SC

[51] R.T. Holman, S.B. Johnson, T.F. Hatch, A case of human linolenic acid deficiency involving neurological abnormalities, Am J Clin Nutr, 35 (1982).

Journal of Clinical Nutrition, 37 (1983) 157-160.

NU

[52] H.C. Meng, A case of human linolenic acid deficiency involving neurological abnormalities, The American

[53] H. Francès, C. Moníer, J.-M. Bourre, Effects of dietary alpha-linolenic acid deficiency on neuromuscular and

MA

cognitive functions in mice, Life Sciences, 57 (1995) 1935-1947.

[54] H. Francès, J.-P. Coudereau, P. Sandouk, M. Clément, C. Monier, J.-M. Bourre, Influence of a dietary α-

D

linolenic acid deficiency on learning in the Morris water maze and on the effects of morphine, European

TE

Journal of Pharmacology, 298 (1996) 217-225.

[55] H. Frances, C. Monier, M. Clement, A. Lecorsier, M. Debray, J.-M. Bourre, Effect of dietary α-linolenic acid

AC CE P

deficiency on habituation, Life Sciences, 58 (1996) 1805-1816. [56] I. Carrié, M. Smirnova, M. Clément, D. De Javel, H. Francès, J.M. Bourre, Docosahexaenoic Acid-rich Phospholipid Supplementation: Effect on Behavior, Learning Ability, and Retinal Function in Control and n-3 Polyunsaturated Fatty Acid Deficient Old Mice, Nutritional Neuroscience, 5 (2002) 43-52. [57] I. Fedorova, N. Hussein, M.H. Baumann, C. Di Martino, N. Salem, Jr., An n-3 fatty acid deficiency impairs rat spatial learning in the Barnes maze, Behavioral Neuroscience, 123 (2009) 196-205. [58] M. Neuringer, W.E. Connor, D.S. Lin, L. Barstad, S. Luck, Biochemical and functional effects of prenatal and postnatal omega 3 fatty acid deficiency on retina and brain in rhesus monkeys, Proc Natl Acad Sci U S A, 83 (1986). [59] J. Catalan, M. Toru, B. Slotnick, M. Murthy, R.S. Greiner, N. Salem, Jr., Cognitive deficits in docosahexaenoic acid-deficient rats, Behavioral Neuroscience, 116 (2002) 1022-1031. [60] D.S. Grayson, C.D. Kroenke, M. Neuringer, D.A. Fair, Dietary omega-3 fatty acids modulate large-scale systems organization in the rhesus macaque brain, J Neurosci, 34 (2014). 61

. ranc s, . ra , M. mirno a, . arri , M. e ray, .M. Bourre, Nutritional n-3) polyunsaturated fatty

acids influence the behavioral responses to positive events in mice, Neuroscience Letters, 285 (2000) 223-227.

ACCEPTED MANUSCRIPT [62] S. Delion, S. Chalon, D. Guilloteau, J.- . Besnard, G. urand, α-Linolenic Acid Dietary Deficiency Alters AgeRelated Changes of Dopaminergic and Serotoninergic Neurotransmission in the Rat Frontal Cortex, Journal of Neurochemistry, 66 (1996) 1582-1591. [63] R.K. McNamara, R. Jandacek, T. Rider, P. Tso, A. Cole-Strauss, J.W. Lipton, Omega-3 fatty acid deficiency

PT

increases constitutive pro-inflammatory cytokine production in rats: Relationship with central serotonin turnover, Prostaglandins, Leukotrienes and Essential Fatty Acids (PLEFA), 83 (2010) 185-191.

RI

[64] A. Latour, B. Grintal, G. Champeil-Potokar, M. Hennebelle, M. Lavialle, P. Dutar, B. Potier, J.-M. Billard, S.

SC

Vancassel, I. Denis, Omega-3 fatty acids deficiency aggravates glutamatergic synapse and astroglial aging in the rat hippocampal CA1, Aging Cell, 12 (2013) 76-84.

NU

[65] C. Madore, A. Nadjar, J.-C. Delpech, A. Sere, A. Aubert, C. Portal, C. Joffre, S. Layé, Nutritional n-3 PUFAs deficiency during perinatal periods alters brain innate immune system and neuronal plasticity-associated genes, Brain, Behavior, and Immunity, 41 (2014) 22-31.

MA

[66] J.M.C. Borba, A.P. Rocha-de-Melo, Â.A. dos Santos, B.L.d.S. Andrade da Costa, R.P. da Silva, P.P. Passos, R.C.A. Guedes, Essential fatty acid deficiency reduces cortical spreading depression propagation in rats: a two-

D

generation study, Nutritional Neuroscience, 13 (2010) 144-150.

TE

[67] Harbeby E, Jouin M, Alessandri JM, Lallemand MS, Linard A, Lavialle M, Huertas A, Cunnane SC, G. P, n-3 PUFA status affects expression of genes involved in neuroenergetics differently in the fronto-parietal cortex

AC CE P

compared to the CA1 area of the hippocampus: effect of rest and neuronal activation in the rat., Prostaglandins, Leukotrienes and Essential Fatty Acids., 86 (2012) 10. [68] F. Pifferi, F. Roux, B. Langelier, J.-M. Alessandri, S. Vancassel, M. Jouin, M. Lavialle, P. Guesnet, (n-3) Polyunsaturated Fatty Acid Deficiency Reduces the Expression of Both Isoforms of the Brain Glucose Transporter GLUT1 in Rats, The Journal of Nutrition, 135 (2005) 2241-2246. [69] A. Ximenes da Silva, F. Lavialle, G. Gendrot, P. Guesnet, J.M. Alessandri, M. Lavialle, Glucose transport and utilization are altered in the brain of rats deficient in n-3 polyunsaturated fatty acids, Journal of Neurochemistry, 81 (2002) 1328-1337. [70] C. Choeiri, W. Staines, T. Miki, S. Seino, C. Messier, Glucose transporter plasticity during memory processing, Neuroscience, 130 (2005) 591-600. [71] T. Larrieu, C. Madore, C. Joffre, S. Layé, Nutritional n-3 polyunsaturated fatty acids deficiency alters cannabinoid receptor signaling pathway in the brain and associated anxiety-like behavior in mice, Journal of Physiology and Biochemistry, 68 (2012) 671-681. [72] J.J. Liu, H.C. Galfalvy, T.B. Cooper, M.A. Oquendo, M.F. Grunebaum, J.J. Mann, Omega-3 polyunsaturated fatty acid (PUFA) status in major depressive disorder with comorbid anxiety disorders, J Clin Psychiatry, 74 (2013).

ACCEPTED MANUSCRIPT [73] V. Palsdottir, J.-E. Månsson, M. Blomqvist, E. Egecioglu, B. Olsson, Long-term effects of perinatal essential fatty acid deficiency on anxiety-related behavior in mice, Behavioral Neuroscience, 126 (2012) 361-369. [74] K.J. Ressler, C.B. Nemeroff, Role of serotonergic and noradrenergic systems in the pathophysiology of depression and anxiety disorders, Depression and Anxiety, 12 (2000) 2-19.

PT

[75] J.C. DeMar, Jr., K. Ma, J.M. Bell, M. Igarashi, D. Greenstein, S.I. Rapoport, One generation of n-3 polyunsaturated fatty acid deprivation increases depression and aggression test scores in rats, J Lipid Res, 47

RI

(2006) 172-180.

SC

[76] W.A. Carlezon, S.D. Mague, A.M. Parow, A.L. Stoll, B.M. Cohen, P.F. Renshaw, Antidepressant-like effects of uridine and omega-3 fatty acids are potentiated by combined treatment in rats, Biol Psychiatry, 57 (2005).

NU

[77] S.Y. Huang, H.T. Yang, C.C. Chiu, C.M. Pariante, K.P. Su, Omega-3 fatty acids on the forced-swimming test, J Psychiatr Res, 42 (2008).

[78] S. Reisbick, M. Neuringer, R. Hasnain, W.E. Connor, Home cage behavior of rhesus monkeys with long-term

MA

deficiency of omega-3 fatty acids, Physiol Behav, 55 (1994).

[79] S.M. Conklin, P.J. Gianaros, S.M. Brown, J.K. Yao, A.R. Hariri, S.B. Manuck, Long-chain omega-3 fatty acid

D

intake is associated positively with corticolimbic gray matter volume in healthy adults, Neurosci Lett, 421

TE

(2007).

[80] R.D. Porsolt, G. Brossard, C. Hautbois, S. Roux, Rodent Models of Depression: Forced Swimming and Tail

Sons, Inc., 2001.

AC CE P

Suspension Behavioral Despair Tests in Rats and Mice, in: Current Protocols in Neuroscience, John Wiley &

[81] Thomazeau A, C. Bosch-Bouju, O. Manzoni, L. S, Nutritional n-3 PUFA Deficiency Abolishes Endocannabinoid Gating of Hippocampal Long-Term Potentiation., Cereb Cortex, (2016). [82] M. Lafourcade, T. Larrieu, S. Mato, A. Duffaud, M. Sepers, I. Matias, V. De Smedt-Peyrusse, V.F. Labrousse, L. Bretillon, C. Matute, R. Rodriguez-Puertas, S. Laye, O.J. Manzoni, Nutritional omega-3 deficiency abolishes endocannabinoid-mediated neuronal functions, Nat Neurosci, 14 (2011) 345-350. [83] D. Yamada, K. Wada, M. Sekiguchi, Modulation of Long-Term Potentiation of Cortico-Amygdala Synaptic Responses and Auditory Fear Memory by Dietary Polyunsaturated Fatty Acid, Frontiers in Behavioral Neuroscience, 10 (2016). 84 . immer, . el al, . Guilloteau, . A oun, G. urand, .

alon,

ronic n-3 polyunsaturated fatty acid

deficiency alters dopamine vesicle density in the rat frontal cortex, Neuroscience Letters, 284 (2000) 25-28. [85] S.O. Ahmad, J.-H. Park, J.D. Radel, B. Levant, Reduced numbers of dopamine neurons in the substantia nigra pars compacta and ventral tegmental area of rats fed an n-3 polyunsaturated fatty acid-deficient diet: A stereological study, Neuroscience Letters, 438 (2008) 303-307.

ACCEPTED MANUSCRIPT [86] F. Kuperstein, R. Eilam, E. Yavin, Altered expression of key dopaminergic regulatory proteins in the postnatal brain following perinatal n-3 fatty acid dietary deficiency, Journal of Neurochemistry, 106 (2008) 662671. [87] L. Zimmer, S. Vancassel, S. Cantagrel, P. Breton, S. Delamanche, D. Guilloteau, G. Durand, S. Chalon, The

PT

do amine mesocorticolim ic at way is affected y deficiency in n−3 olyunsaturated fatty acids, T e American Journal of Clinical Nutrition, 75 (2002) 662-667.

RI

[88] E. Kodas, G. Page, L. Zimmer, S. Vancassel, D. Guilloteau, G. Durand, S. Chalon, Neither the density nor

SC

function of striatal dopamine transporters were influenced by chronic n-3 polyunsaturated fatty acid deficiency in rodents, Neuroscience Letters, 321 (2002) 95-99.

NU

[89] M. Tang, M. Zhang, H. Cai, H. Li, P. Jiang, R. Dang, Y. Liu, X. He, Y. Xue, L. Cao, Y. Wu, Maternal diet of polyunsaturated fatty acid altered the cell proliferation in the dentate gyrus of hippocampus and influenced glutamatergic and serotoninergic systems of neonatal female rats, Lipids in Health and Disease, 15 (2016) 71.

MA

[90] O. Howes, S. Bose, F. Turkheimer, I. Valli, A. Egerton, D. Stahl, L. Valmaggia, P. Allen, R. Murray, P. McGuire, Progressive increase in striatal dopamine synthesis capacity as patients develop psychosis: a PET

D

study, Mol Psychiatry, 16 (2011) 885-886.

Rev Neurosci, 15 (2014) 771-785.

TE

[91] R.P. Bazinet, S. Laye, Polyunsaturated fatty acids and their metabolites in brain function and disease, Nat

AC CE P

[92] V. Conway, A. Larouche, W. Alata, M. Vandal, F. Calon, M. Plourde, Apolipoprotein E isoforms disrupt longchain fatty acid distribution in the plasma, the liver and the adipose tissue of mice, Prostaglandins Leukot Essent Fatty Acids, 91 (2014) 261-267.

[93] J.M. Arbones-Mainar, L.A. Johnson, E. Torres-Perez, A.E. Garcia, S. Perez-Diaz, J. Raber, N. Maeda, Metabolic shifts toward fatty-acid usage and increased thermogenesis are associated with impaired adipogenesis in mice expressing human APOE4, Int J Obes, 40 (2016) 1574-1581. [94] V. Conway, A. Larouche, W. Alata, M. Vandal, F. Calon, M. Plourde, Apolipoprotein E isoforms disrupt longchain fatty acid distribution in the plasma, the liver and the adipose tissue of mice, Prostaglandins, Leukotrienes and Essential Fatty Acids (PLEFA), 91 (2014) 261-267. [95] M.C. Phillips, Apolipoprotein E isoforms and lipoprotein metabolism, IUBMB Life, 66 (2014) 616-623. [96] D.F. Horrobin, Loss of delta-6-desaturase activity as a key factor in aging, Med Hypotheses, 7 (1981) 12111220. [97] C.A. Hostage, K.R. Choudhury, P. Murali Doraiswamy, J.R. Petrella, Mapping the effect of the apolipoprotein E genotype on 4-year atrophy rates in an Alzheimer disease-related brain network, Radiology, 271 (2014) 211-219.

ACCEPTED MANUSCRIPT [98] R.D. Bell, E.A. Winkler, I. Singh, A.P. Sagare, R. Deane, Z. Wu, D.M. Holtzman, C. Betsholtz, A. Armulik, J. Sallstrom, B.C. Berk, B.V. Zlokovic, Apolipoprotein E controls cerebrovascular integrity via cyclophilin A, Nature, 485 (2012) 512-516. [99] W. Alata, Y. Ye, I. St-Amour, M. Vandal, F. Calon, Human apolipoprotein E varepsilon4 expression impairs

PT

cerebral vascularization and blood-brain barrier function in mice, J Cereb Blood Flow Metab, 35 (2015) 86-94. [100] S. Salloway, T. Gur, T. Berzin, R. Tavares, B. Zipser, S. Correia, V. Hovanesian, J. Fallon, V. Kuo-Leblanc, D.

RI

Glass, C. Hulette, C. Rosenberg, M. Vitek, E. Stopa, Effect of APOE genotype on microvascular basement

SC

membrane in Alzheimer's disease, Journal of the neurological sciences, 203-204 (2002) 183-187. [101] M.R. Halliday, S.V. Rege, Q. Ma, Z. Zhao, C.A. Miller, E.A. Winkler, B.V. Zlokovic, Accelerated pericyte

NU

degeneration and blood-brain barrier breakdown in apolipoprotein E4 carriers with Alzheimer's disease, J Cereb Blood Flow Metab, 36 (2016) 216-227.

[102] R.C. Knickmeyer, J. Wang, H. Zhu, X. Geng, S. Woolson, R.M. Hamer, T. Konneker, W. Lin, M. Styner, J.H.

MA

Gilmore, Common Variants in Psychiatric Risk Genes Predict Brain Structure at Birth, Cerebral Cortex, 24 (2014) 1230-1246. . . ean, B.A. erskey, K.

en, . rotas, . T iyyagura, A. Roonti a, . O’Muirc eartaig , . irks, N.

D

103

TE

Waskiewicz, K. Lehman, A.L. Siniard, M.N. Turk, X. Hua, S.K. Madsen, P.M. Thompson, A.S. Fleisher, M.J. Huentelman, S.C.L. Deoni, E.M. Reiman, Brain Differences in Infants at Differential Genetic Risk for Late-Onset

AC CE P

Alzheimer Disease A Cross-sectional Imaging Study, JAMA neurology, 71 (2014) 11-22. [104] P. Shaw, J.P. Lerch, J.C. Pruessner, K.N. Taylor, A.B. Rose, D. Greenstein, L. Clasen, A. Evans, J.L. Rapoport, J.N. Giedd, Cortical morphology in children and adolescents with different apolipoprotein E gene polymorphisms: an observational study, The Lancet Neurology, 6 (2007) 494-500. [105] G. Salvato, Does apolipoprotein E genotype influence cognition in middle-aged individuals?, Current Opinion in Neurology, 28 (2015) 612-617. [106] Y.I. Sheline, J.C. Morris, A.Z. Snyder, J.L. Price, Z. Yan, G. D'Angelo, C. Liu, S. Dixit, T. Benzinger, A. Fagan, A. Goate, M.A. Mintun, APOE4 Allele Disrupts Resting State fMRI Connectivity in the Absence of Amyloid laques or ecreased

Aβ42, T e ournal of Neuroscience, 30 2010) 17035-17040.

[107] M.M. Adamson, K.M. Landy, S. Duong, S. Fox-Bosetti, J.W. Ashford, G.M. Murphy, M. Weiner, J.L. Taylor, Apolipoprotein E ɛ4 influences on episodic recall and brain structures in aging pilots, Neurobiology of Aging, 31 (2010) 1059-1063. [108] N.A. Kozauer, M.M. Mielke, G.K. Chuen Chan, G.W. Rebok, C.G. Lyketsos, Apolipoprotein E genotype and lifetime cognitive decline, International Psychogeriatrics, 20 (2008) 109-123. [109] R.S. Wilson, C.T. Begeny, P.A. Boyle, J.A. Schneider, D.A. Bennett, Vulnerability to Stress, Anxiety, and Development of Dementia in Old Age, The American journal of geriatric psychiatry : official journal of the American Association for Geriatric Psychiatry, 19 (2011) 327-334.

ACCEPTED MANUSCRIPT [110] P.M. Greenwood, T. Espeseth, M.-K. Lin, I. Reinvang, R. Parasuraman, Longitudinal change in working memory as a function of APOE genotype in midlife and old age, Scandinavian Journal of Psychology, 55 (2014) 268-277. [111] P.M. Greenwood, T. Sunderland, J.L. Friz, R. Parasuraman, Genetics and visual attention: Selective deficits

PT

in healthy adult carriers of the ɛ4 allele of the apolipoprotein E gene, Proceedings of the National Academy of Sciences, 97 (2000) 11661-11666.

RI

[112] P.M. Greenwood, C. Lambert, T. Sunderland, R. Parasuraman, Effects of Apolipoprotein E Genotype on

SC

Spatial Attention, Working Memory, and Their Interaction in Healthy, Middle-Aged Adults: Results From the National Institute of Mental Health's BIOCARD Study, Neuropsychology, 19 (2005) 199-211.

NU

[113] B.D. Zipser, C.E. Johanson, L. Gonzalez, T.M. Berzin, R. Tavares, C.M. Hulette, M.P. Vitek, V. Hovanesian, E.G. Stopa, Microvascular injury and blood-brain barrier leakage in Alzheimer's disease, Neurobiol Aging, 28 (2007) 977-986.

MA

[114] S. Suri, C.E. Mackay, M.E. Kelly, M. Germuska, E.M. Tunbridge, G.B. Frisoni, P.M. Matthews, K.P. Ebmeier, D.P. Bulte, N. Filippini, Reduced cerebrovascular reactivity in young adults carrying the APOE epsilon4 allele,

D

Alzheimers Dement, (2014).

TE

[115] Y. Lin, M. Xu, J. Wan, S. Wen, J. Sun, H. Zhao, M. Lou, Docosahexaenoic acid attenuates hyperglycemiaenhanced hemorrhagic transformation after transient focal cerebral ischemia in rats, Neuroscience, 301 (2015)

AC CE P

471-479.

[116] L.N. Nguyen, D. Ma, G. Shui, P. Wong, A. Cazenave-Gassiot, X. Zhang, M.R. Wenk, E.L.K. Goh, D.L. Silver, Mfsd2a is a transporter for the essential omega-3 fatty acid docosahexaenoic acid, Nature, 509 (2014) 503-506. [117] B.J. Andreone, B.W. Chow, A. Tata, B. Lacoste, A. Ben-Zvi, K. Bullock, A.A. Deik, D.D. Ginty, C.B. Clish, C. Gu, Blood-Brain Barrier Permeability Is Regulated by Lipid Transport-Dependent Suppression of CaveolaeMediated Transcytosis, Neuron, 94 (2017) 581-594.e585. [118] L. Abdullah, J. Evans, T. Emmerick, T. NGguyen, G. Crynen, B. Shackleton, J. Reed, A. Keegan, C. Luis, L. Tai, et al., APOE4 specific imbalance in arachidonic acid and docosahexaenoic acid in serum phospholipids from individuals with preclinical MCI/AD., In Alzheimer's Association International Conference. Toronto (2016). [119] L. Abdullah, J. Evans, B. Shackleton, J. Ojo, T. Nguyen, J. Reed, M. Mullan, F. Crawford, C. Bachmeier, APOE4 genotype dependant deficits in DHA containing phospholipids and DHA transporters in the cerebrovasculature of Alzheimer's disease patients., In Alzheimer's Association International Conference. Toronto., (2016). [120] R. Chouinard-Watkins, M. Vandal, P. Léveillé, A. Pinçon, F. Calon, M. Plourde, Docosahexaenoic acid prevents cognitive deficits in human apolipoprotein E epsilon 4-targeted replacement mice, Neurobiology of Aging, 57 (2017) 28-35.

ACCEPTED MANUSCRIPT 121

. Yijun, K. anan, A.N. ose

, T e m act of ocosa exaenoic Acid on Alz eimer’s Disease: Is There a

Role of the Blood-Brain Barrier?, Current Clinical Pharmacology, 10 (2015) 222-241. [122] R.W. Mahley, Apolipoprotein E: cholesterol transport protein with expanding role in cell biology, Science, 240 (1988) 622-630.

PT

[123] R.E. Pitas, J.K. Boyles, S.H. Lee, D. Foss, R.W. Mahley, Astrocytes synthesize apolipoprotein E and metabolize apolipoprotein E-containing lipoproteins, Biochimica et biophysica acta, 917 (1987) 148-161.

RI

[124] M. Vandal, W. Alata, C. Tremblay, C. Rioux-Perreault, N. Salem, Jr., F. Calon, M. Plourde, Reduction in

SC

DHA transport to the brain of mice expressing human APOE4 compared to APOE2, J Neurochem, 129 (2014) 516-526.

NU

[125] R. Chouinard-Watkins, C. Rioux-Perreault, M. Fortier, J. Tremblay-Mercier, Y. Zhang, P. Lawrence, M.C. Vohl, P. Perron, D. Lorrain, J.T. Brenna, S.C. Cunnane, M. Plourde, Disturbance in uniformly 13C-labelled DHA metabolism in elderly human subjects carrying the apoE epsilon4 allele, Br J Nutr, 110 (2013) 1751-1759.

MA

[126] H.N. Yassine, M.N. Braskie, W.J. Mack, K.J. Castor, A.N. Fonteh, L.S. Schneider, M.G. Harrington, H.C. Chui, Association of Docosahexaenoic Acid Supplementation With Alzheimer Disease Stage in Apolipoprotein E

D

epsilon4 Carriers: A Review, JAMA Neurol, (2017).

TE

[127] H.N. Yassine, Q. Feng, I. Azizkhanian, V. Rawat, K. Castor, A.N. Fonteh, M.G. Harrington, L. Zheng, B.R. Reed, C. DeCarli, W.J. Jagust, H.C. Chui, Association of Serum Docosahexaenoic Acid With Cerebral Amyloidosis,

AC CE P

JAMA Neurol, 73 (2016) 1208-1216.

[128] G.J. Anderson, M. Neuringer, D.S. Lin, W.E. Connor, Can prenatal N-3 fatty acid deficiency be completely reversed after birth? Effects on retinal and brain biochemistry and visual function in rhesus monkeys, Pediatr Res, 58 (2005).

[129] W.E. Connor, M. Neuringer, D.S. Lin, Dietary effects on brain fatty acid composition: The reversibility of n3 fatty acid deficiency and turnover of docosahexaenoic acid in the brain, erythrocytes, and plasma of rhesus monkeys, Journal of Lipid Research, 31 (1990) 237-247. [130] H.F. Chen, H.M. Su, Exposure to a maternal n-3 fatty acid-deficient diet during brain development provokes excessive hypothalamic-pituitary-adrenal axis responses to stress and behavioral indices of depression and anxiety in male rat offspring later in life, J Nutr Biochem, 24 (2013). [131] T. Moriguchi, J. Loewke, M. Garrison, J.N. Catalan, N. Salem, Reversal of docosahexaenoic acid deficiency in the rat brain, retina, liver, and serum, Journal of Lipid Research, 42 (2001) 419-427. [132] K. Tanaka, T. Shimizu, Y. Ohtsuka, Y. Yamas iro, K. Os ida, Early dietary treatments wit

orenzo’s oil and

docosahexaenoic acid for neurological development in a case with Zellweger syndrome, Brain and Development, 29 (2007) 586-589.

ACCEPTED MANUSCRIPT [133] H.-F. Chen, H.-M. Su, Fish oil supplementation of maternal rats on an n-3 fatty acid-deficient diet prevents depletion of maternal brain regional docosahexaenoic acid levels and has a postpartum anxiolytic effect, The Journal of Nutritional Biochemistry, 23 (2012) 299-305. [134] I. Carrié, M. Clément, D. de Javel, H. Francès, J.-M. Bourre, Phospholipid supplementation reverses

PT

behavioral and biochemical alterations induced by n–3 polyunsaturated fatty acid deficiency in mice, Journal of Lipid Research, 41 (2000) 473-480.

RI

[135] Paker AM, Sunness JS, Brereton NH, Speedie LJ, Albanna L, Dharmaraj S, Moser AB, Jones RO, R. GV.,

SC

Docosahexanoic acid therapy in peroxisomal diseases: results of a double-blind, randomized trial. , Neurology. , 75 (2010) 5.

peroxisomal biogenesis., Lipids., 31 Suppl (1996) 8.

NU

[136] M. Martinez, Docosahexaenoic acid therapy in docosahexaenoic acid-deficient patients with disorders of

[137] Martínez M, Vázquez E, García-Silva MT, Beltrán JM, Castelló F, Pineda M, M. I., [Treatment of

MA

generalized peroxisomal disorders with docosahexaenoic acid ethyl ether]. [Article in Spanish], Rev Neurol., 28 Suppl 1 (1999) 6.

D

[138] E. Kodas, L. Galineau, S. Bodard, S. Vancassel, D. Guilloteau, J.-C. Besnard, S. Chalon, Serotoninergic

TE

neurotransmission is affected by n-3 polyunsaturated fatty acids in the rat, Journal of Neurochemistry, 89 (2004) 695-702.

AC CE P

[139] Y. Freund Levi, I. Vedin, T. Cederholm, H. Basun, G. Faxén Irving, M. Eriksdotter, E. Hjorth, M. Schultzberg, B. Vessby, L.O. Wahlund, N. Salem, J. Palmblad, Transfer of omega-3 fatty acids across the blood–brain barrier after dietary supplementation with a docosahexaenoic acid-rich omega-3 fatty acid preparation in patients with Alzheimer's disease: the OmegAD study, Journal of Internal Medicine, 275 (2014) 428-436. [140] E. Hjorth, M. Zhu, V.C. Toro, I. Vedin, J. Palmblad, T. Cederholm, Y. Freund-Levi, G. Faxen-Irving, L.-O. Wahlund, H. Basun, M. Eriksdotter, M. Schultzberg, Omega-3 atty Acids En ance

agocytosis of Alz eimer’s

Disease-Related Amyloid-42 by Human Microglia and ecrease nflammatory Markers, ournal of Alz eimer’s Disease 35 (2013) 17. [141] M.S. Rosell, Z. Lloyd-Wright, P.N. Appleby, T.A. Sanders, N.E. Allen, T.J. Key, Long-chain n–3 polyunsaturated fatty acids in plasma in British meat-eating, vegetarian, and vegan men, The American Journal of Clinical Nutrition, 82 (2005) 327-334. [142] M. Kornsteiner, I. Singer, I. Elmadfa, Very Low n–3 Long-Chain Polyunsaturated Fatty Acid Status in Austrian Vegetarians and Vegans, Annals of Nutrition and Metabolism, 52 (2008) 37-47. [143] S.I. Rapoport, M. Igarashi, Can the rat liver maintain normal brain DHA metabolism in the absence of dietary DHA?, Prostaglandins Leukot Essent Fatty Acids, 81 (2009).

ACCEPTED MANUSCRIPT [144] M. Igarashi, K. Ma, L. Chang, J.M. Bell, S.I. Rapoport, Dietary n-3 PUFA deprivation for 15 weeks upregulates elongase and desaturase expression in rat liver but not brain, Journal of Lipid Research, 48 (2007) 2463-2470. [145] S.C. Cunnane, R. Chouinard-Watkins, C.A. Castellano, P. Barberger-Gateau, Docosahexaenoic acid

PT

homeostasis, brain aging and Alzheimer's disease: Can we reconcile the evidence?, Prostaglandins, Leukotrienes and Essential Fatty Acids (PLEFA), 88 (2013) 61-70.

RI

[146] S.C. Cunnane, J.A. Schneider, C. Tangney, J. Tremblay-Mercier, M. Fortier, D.A. Bennett, M.C. Morris,

SC

lasma and Brain atty Acid rofiles in Mild ogniti e m airment and Alz eimer’s isease, ournal of Alzheimer's disease : JAD, 29 (2012) 691-697.

NU

[147] A.F. Domenichiello, A.P. Kitson, A.H. Metherel, C.T. Chen, K.E. Hopperton, P.M. Stavro, R.P. Bazinet, Whole-Body Docosahexaenoic Acid Synthesis-Secretion Rates in Rats Are Constant across a Large Range of ietary α-Linolenic Acid Intakes, The Journal of Nutrition, 147 (2017) 37-44.

MA

[148] A.A. Welch, S. Shakya-Shrestha, M.A. Lentjes, N.J. Wareham, K.-T. K aw, ietary intake and status of n−3 polyunsaturated fatty acids in a population of fish-eating and non-fish-eating meat-eaters, vegetarians, and

D

vegans and the precursor- roduct ratio of α-linolenic acid to long-c ain n−3 olyunsaturated fatty acids: results

TE

from the EPIC-Norfolk cohort, The American Journal of Clinical Nutrition, 92 (2010) 1040-1051. [149] G. Wang, H.L. Bonkovsky, A. de Lemos, F.J. Burczynski, Recent insights into the biological functions of liver

AC CE P

fatty acid binding protein 1, Journal of lipid research, 56 (2015) 2238-2247. [150] R. Chouinard-Watkins, A. Pinçon, J.-D. Coulombe, R. Spencer, L. Massenavette, M. Plourde, A Diet Rich in Docosahexaenoic Acid Restores Liver Arachidonic Acid and Docosahexaenoic Acid Concentrations in Mice omozygous for t e uman A oli o rotein E ε4 Allele, T e ournal of Nutrition, 146 2016) 1315-1321. [151] W. Zhang, R. Chen, T. Yang, N. Xu, J. Chen, Y. Gao, R.A. Stetler, Fatty acid transporting proteins: Roles in brain development, aging, and stroke, Prostaglandins, Leukotrienes and Essential Fatty Acids (PLEFA), (2017). [152] S.C. Cunnane, A. Courchesne-Loyer, V. St-Pierre, C. Vandenberghe, T. Pierotti, M. Fortier, E. Croteau, C.-A. Castellano, Can ketones compensate for deteriorating brain glucose uptake during aging? Implications for the risk and treatment of Alzheimer's disease, Annals of the New York Academy of Sciences, 1367 (2016) 12-20. [153] A. Pincon, J.D. Coulombe, R. Chouinard-Watkins, M. Plourde, Human apolipoprotein E allele and docosahexaenoic acid intake modulate peripheral cholesterol homeostasis in mice, J Nutr Biochem, 34 (2016) 83-88. [154] M. Hennebelle, M. Plourde, R. Chouinard-Watkins, C. Castellano, P. Barberger-Gateau, S.C. Cunnane, Ageing and apoE change DHA homeostasis: Relevance to age-related cognitive decline. , Proceedings of the Nutrition Society, 73 (2014) 7.

ACCEPTED MANUSCRIPT [155] R. Chouinard-Watkins, V. Conway, A.M. Minihane, K.G. Jackson, J.A. Lovegrove, M. Plourde, Interactive impact of BMI and APOE genotype on the plasma long chain polyunsaturated fatty acid response to a fish oil supplement in healthy participants Am J Clin Nutr, 102 (2015) 505-513. [156] C.T. Chen, A.P. Kitson, K.E. Hopperton, A.F. Domenichiello, M.-O. Trépanier, L.E. Lin, L. Ermini, M. Post, F.

PT

Thies, R.P. Bazinet, Plasma non-esterified docosahexaenoic acid is the major pool supplying the brain, Scientific

AC CE P

TE

D

MA

NU

SC

RI

Reports, 5 (2015) 15791.

ACCEPTED MANUSCRIPT Figure legends Figure 1: Plasma, liver, and adipose tissue concentrations of DHA (left panels) and ALA (right panels) in APOE3 and APOE4 mice fed a diet deficient in n-3 FA. To maintain plasma DHA and ALA levels,

PT

E4 mice had to pull DHA from or prevent DHA getting into the liver and the adipose tissue [92]

RI

Figure 2: Mean ± SEM of DHA levels in the plasma (left panel, n = 9-13 in each group), liver (middle, n = 9-13 in each group) and cortex (right, n = 4-5 in each group) in E3 and E4 mice. Mice consumed a

AC CE P

TE

D

MA

NU

SC

control or a diet enriched with DHA.

Figure 1

AC CE P

TE

D

MA

NU

SC

RI

PT

ACCEPTED MANUSCRIPT

SC

RI

PT

ACCEPTED MANUSCRIPT

AC CE P

TE

D

MA

NU

Figure 2

TE

AC CE P

Graphical abstract

D

MA

NU

SC

RI

PT

ACCEPTED MANUSCRIPT

ACCEPTED MANUSCRIPT Highlights

TE

D

MA

NU

SC

RI

PT

E4 carriers are more vulnerable to dietary n-3 FA deficits than other APOE alleles. DHA is pulled from the liver and adipose in E4 carriers to maintain plasma levels. Providing n-3 FA can restore levels of brain DHA in E4 mice. DHA supplementation in E4 carriers may help to prevent or halt cognitive decline.

AC CE P

   