Journal of Translational Medicine and Developmental Disorders, 2015, Vol. 2, No. 1, 1-9
Available online at http://pubs.sciepub.com/jtmdd/2/1/1© Science and Education Publishing
DOI:10.12691/jtmdd-2-1-1

Down-regulation of Signaling Mediator in Related to
Increased Ratio of Docosahexaenoic Acid/Arachidonic
Acid in Individuals with Autism Spectrum Disorders
Kunio Yui1,*, George Imataka2, Yohei Kawasaki3, Tsutomu Yamada3
1

Research Institute of Pervasive Developmental Disorders, Ashiya University, Rokurokusocho, Ashiya, Hyogo, Japan
Department of Pediatrics, Dokkyo Medical University School of Medicine, 880 Kitakobayashi, Mibu, Tochigi, Japan3
Department of Drug Evaluation and Information, School of Pharmaceutical Science University of Shizuoka, Shizuoka, Japan*Corresponding author: yui16@bell.ocn.ne.jp
2

Received September 10, 2015; Revised September 29, 2015; Accepted September 29, 2015

Abstract Background and aim: Autism spectrum disorder (ASD) is a highly prevalent neurodevelopmental
disorder characterized by abnormal social interactions, communication deficits and stereotyped or repetitive
behaviors. Converging lines of research indicate that the altered composition of polyunsaturated fatty acids (PUFAs)
may contribute to the pathophysiology of ASD. Methods: We examined the relationships between the plasma ratiosof omega-3PUFs/arachidonic acid (AA), plasma levels of 21 fatty acid fractions, and biomarkers of AA-related
signaling mediators (ceruloplasmin, transferrin and superoxide dismutase) with the behavioral symptoms of 32
individuals with ASD (mean age, 13.5 ± 4.3 years old) and 20 age- and gender-matched normal controls (mean age,
13.2 ± 5.4 years old). Behavioral symptoms were assessed using the Aberrant Behavior Checklists (ABC). Results:
Plasma levels of EPA, DPA and DHA, and the plasma ratios of docosahesaenoic acid (DHA)/AA were significantly
higher while plasma levels of AA, 5,8,11,14-eicosatetraenoic acid and Cp were significantly lower in the 32
individuals with ASD compared with the 20 normal controls. The ABC scores were significantly increased in the
ASD group compared to those of the control group. Discussion and Conclusion: High plasma DHA/AA ratio
related to the increased plasma DHA levels and reduced plasma AA level may down-regulate mediators of AA
signaling, such as Cp. Additionally, as 5,8,11,14-eicosatetraenoic acid is an arachidonate metabolite, the reduced
plasma AA revels might have a pathophysiological factor in the reduced plasma Cp levels. Subsequently, reduced
Cp levels may reduce the protective capacity of the brain against damage, which may cause in the pathophysiology
underlying behavioral symptoms observed in individuals with ASD.

Keywords: signaling mediators, ceruloplasmin, competitive interaction, arachidonic acid, docosahexaenoic acid
Cite This Article: Kunio Yui, George Imataka, Yohei Kawasaki, and Tsutomu Yamada, “Down-regulation of
Signaling Mediator in Related to Increased Ratio of Docosahexaenoic Acid/Arachidonic Acid in Individuals with
Autism Spectrum Disorders.” Journal of Translational Medicine and Developmental Disorders, vol. 2, no. 1(2015): 1-9. doi: 10.12691/jtmdd-2-1-1.

1. Introduction
Autism spectrum disorder (ASD) comprises a group of
pervasive developmental disorders characterized by social
deficits, language impairment, and repetitive/restrictedbehaviors. Even though pathogenesis of ASD remains
unclear, it appears to be associated with systemic
metabolic abnormalities [1]. There is increasing evidence
that altered metabolism of polyunsaturated fatty acids(PUFAs) may be involved in the pathophysiology of ASD[2]. PUFAs are essential components of cell membranes,
where the majority of the cell signaling machinery resides[3]. The omega-3 PUFA docosahexaenoic acid (DHA,
22:6ω-3) and eicosapentaenoic acid (EPA, 20:5ω-3), andthe omega-6 PUFA arachidonic acid (AA, 20:4ω-6) play
important roles in the functions of the nervous system [4],neuroplasticity [5]], and signal transduction [6]. AA acts

as a second messenger and regulates the activity of othersignaling cascades [7]. Therefore, ASD may be attributed
to altered PUFA composition and related signaling
mediators. Previous studies have shown that there is amarked reduction in plasma DHA levels in various
neuronal disorders [8] and ASD [9], with changes inplasma omega-3/omega-6 ratio [10]. Importantly, thebalance between omega-3 and omega-6 PUFAs, which is
critical for normal brain function and development, isachieved through both synergistic and antagonistic
mechanisms [11]. The relationship between omega-3
PUFAs and AA is antagonistic at high EPA and DHA
concentrations for maintaining homeostasis [12]. Bothsynergism and antagonism may aim at a balance between
omega-3 and omega-6 PUFAs to maintain homeostasis[12]. Dietary omega-6 PUFAs lowering increases
bioavailability of omega-3 PUFAs in human plasma [13].
This competitive interaction between omega-3 PUFAs and
AA reflects increased plasma omega-3/omega-6 PUFA

2

Journal of Translational Medicine and Developmental Disorders

ratios [12,15,16]. Omega-3 PUFAs may attenuate bothtissue AA levels and eicosanoid formation [16]. The
antagonism of AA-signaling has been reported [17,18],
and this antagonism has important effects upon cellsignaling in the CNS [17], modulating the activity ofmany neurotransmitters in affective disorders [17]. The
omega-3 PUFAs are incorporated at existent of AA, andEPA and DHA inhibit AA metabolism, decreasing
production of AA-derived eicosanoids [19,20]. Thus,
reduced AA levels associated with the competitive
interaction between omega-3 PUFAs and AA may
downregulate AA-related signaling mediators. However,
the relationship between this competitive interaction andrelated AA-related signaling mediators is unclear inneuropsychiatric disorders including ASD.
With respect to interactions between PUFAs andsignaling mediators, ceruloplasmin (Cp) [21], superoxide
dismutase (SOD) [22] and transferrin (Tf) [23] have been
known to act as signaling mediators in ASD. Cp is an
important copper signaling biomarker of neuronal function[24]. Cp has a natural neuroprotective mechanism [25,26],
and may contribute to pathophysiological conditions in thecentral nervous systems [27]. SOD is a biomarker ofcopper signaling [28]. Tf is an iron signaling mediator[29]. Changes in the blood levels of SOD, TF and CP
were shown to indicate altered antioxidant status [30-34]
and copper dyshomeostasis [35,36] in ASD patients. As
described above, the relationship between imbalance ofomega-3/omega-6 PUFA ratios, which is indicative of thecompetitive interaction between increased omega-3
PUFAs and AA, and AA-related signaling mediators (i.e.,
Cp, Tf and SOD) remains unclear in ASD. Moreover,there is no information regarding the mechanisms that
underlie alterations of these signaling mediators in
psychiatric disorders including ASD.
We hypothesized that higher plasma ratios of omega3PUFAs/AA would correlate with a downregulation of
AA-related signaling mediators in individuals with ASD.
In investigating this hypothesis, we measured the plasma
levels (%) of the compositions of PUFAs, monosaturated
and saturated fatty acids, and DHA/AA and EPA/AA
ratios. Since, multiple subfamilies of AA-derived
eicosanoid signaling mediators exist [37] and since these
mediators are affected by multiple factors [38], we
examined plasma levels of Cp, SOD and Tf as known AArelated signaling mediators. Furthermore, because plasma
fatty acid levels can be affected by prior dietary intake, we
conducted controlling for dietary intake and assessed theconsumption of nutrients using the self- administered diethistory questionnaire system.

2. Methods

criteria, and additionally confirmed by the Autism
Diagnostic Interview-Revised (ADI-R)[39]. Among the 52
individuals, 32 had an independent clinical diagnosis of
ASD (22 males and 10 females, mean age: 13.5 ± 4.3
years old, range: 6-21 years old), and the remaining 20
were normal healthy controls (14 males and 6 females,mean age: 13.2 ± 5.4 years old, range: 5-21 years old).
The ASD and control groups were matched for homeenvironment, feeding habits, age, gender and full
intelligent quotient (IQ) scores (Table 1). All 32
individuals with ASD were diagnosed by two psychiatrists
specializing in ASD and developmental disorders. The 32
individuals with ASD had the core symptoms of the DSMIV-R diagnostic criteria for ASD, without any abnormal
neurological symptoms (e.g., seizure or movement
disorders). The 20 individuals in the control group wereconsidered to be physically and mentally healthy based on
initial physical and mental screening tests. Moreover, the32 individuals with ASD and the 20 normal controls has
not any gastrointestinal dysfunction including dysphagia,
gastroparesis and bowel dysfunction. At the initial
screening, physical (resting blood pressure and heart rate)and clinical laboratory examinations (hematology andplasma chemistry including plasma fatty acids, cholesterol
and triglycerides) were performed on all 52 participants.
These 52 participants did not have any abnormalities on
physical examinations and laboratory findings. Comorbid
psychiatric disorders were evaluated by the Structured
Clinical Interview for DSM-IV Axis I Disorders (SCID).
None of the ASD or control individuals had any historiesof neurological conditions, including seizure, movement
disorders or a history of head injury, Attention-Deficit
Hyperactivity Disorder or learning disorders. The
additional inclusion criteria were as follows: (a) the
absence of any other medical or comorbid psychiatricdisorders; (b) a baseline verbal or full IQ greater than 70
as calculated by the WISC-III [40] or the respective scale
for adults (WAIS-R) [41] because subjects with highfunctioning pervasive developmental disorders wererequired to have a total IQ of at least 70 [42]; (c) notreatment with antidepressants, anxiolytic medications orneuroleptics within the three months prior to the study (thetreatment of ADHD symptoms with stimulants wasallowed during this study provided that the patient’s
dosage had remained stable for at least 3 months before
and during the study).
This study complied with the Declaration of Helsinki
and was performed with the approval of the ethics
committee of the Fujimoto Medical Clinic in Kobe City,
Japan. This ethics committee was registered with thePharmaceuticals and Medical Devices Agency of Japanfor registration of IRB information (http://www.info.
pmda.go.jp/). Written informed consent was obtained
from the participants and/or their parents.

2.1. Subjects
A total of 52 young, physically healthy individuals whoin the Kansai area (Hyogo and Osaka Prefectures) ofJapan participated in this study. They were recruited frommedical care facilities of the Research Institute ofPervasive Developmental Disorders, Ashiya University by
the order of their submission to medical consultation in
the Research Institute between January 2012 and July
2014. Diagnosis was performed based on DSM-IV-TR

2.2. Controlling for Dietary Intake andAssessment of Nutrient Intake
Because plasma fatty acid levels can be affected byprior dietary intake, many prior studies have used the 2010
dietary Guidelines for Americans [43]). In this study, all
52 participants received “Japanese Food Guide SpinningTOP” [44], which was characterized by favorable dietaryintakes of foods and nutrients, based the “Dietary

Journal of Translational Medicine and Developmental Disorders

Reference Intakes for Japanese. -2010-”[45]. All 52
individuals or their parents were provided a diet plansamplemenus(KAWASAKI
FOODMODEL)(http://item.rakuten.co.jp/foodmodel/751741), which wasedited according to “Japanese Food Guide”. Moreover, toassess daily food and nutrients intake, a semi-constructive
questionnaire for assessment of dietary food and nutrient
intake for Japanese (DHQ support center, Tokyo, Japan;
http://www.ebnjapan.org/) was performed using the juniorhigh school version (DHQ15) (DHQ support center,
Tokyo, Japan). DHQ15 is a structured questionnaire thatasks about the consumption of frequency and portion sizeof selected foods commonly consumed in Japan [46], andusual cooking methods. The validity of the junior highschool version of the DHQ 15 has been verified [47].
Energy and selected nutrients for 150 various food andbeverage items were calculated using a custom-designed
computer algorism for DHQ based on the Standard Tablesof Food Consumption in Japan (DHA support center).
DHQ15 was administered during one month before thisstudy in 17 individuals with ASD and 10 normal controls
by the order of their submission to medical
consultation during January 2014 and December 2014.
Their mothers completed the DHQ15 at home andreturned the questionnaires to the Fujimoto clinic at Kobe.
If missing or unclear information was recorded on the
sheet by the guardian, the researcher (KY) questioned theguardian by phone or e-mail.

2.3. Assessment of Behavioral Symptoms
The ABC was used to assess the behavioral symptomsof the 32 individuals with ASD and the 20 normal controls.
The ABC is primarily intended to evaluate treatment
responses in psychopharmacological and behavioral
intervention trials for children and adolescents withmental retardation [48] and normal IQ levels [49]. The
subscales are as follows: (1) irritability (15 items); (2)social withdrawal (16 items); (3) stereotypic behavior(seven items); (4) hyperactivity (16 items); and (5)inappropriate speech (four items). The ABC is a broad
assessment that captures a wide variety of behavioral
problems [50].

2.4. Assays of Plasma Levels of PUFAs, Cp,
SOD and Tf
2.4.1. Methods of Blood Sampling
Whole-blood samples were collected in EDTA tubes byvenipuncture and immediately placed on ice in arefrigerator. Plasma was obtained by centrifugation for 20
minutes at 3000 x g at room temperature (22 °C). To
decrease the effects of circadian variation, the bloodcollection performed at 11:00-12:30 in a quiet laboratory
room after supine rest for 20 minutes. The samples werefrozen at –80°C until analysis. The plasma levels of
PUFAs, Cp, Tf and SOD were measured by specialists at
SRL, Inc (Tokyo, Japan).
2.4.2. Plasma Levels of PUFAs.
The fatty acid composition of the PUFA fraction fromeach patient’s plasma was determined as previously
described [51]. In summary, total lipids were extracted

3

from the plasma according to [52]. After transmethylation
with HCl-methanol, PUFA composition was analyzed
using gas chromatography (GC2010 Shimadzu Co.,
Japan). A total of 24 long-chain fatty acids were identified.
The intra- and inter-assay coefficients of AA were 110.14μg/ml (standard deviation (SD), 3.87; coefficient ofvariation (CV), 5.28%) and 100.63 μg/ml (SD, 5.51; CV,
5.48%), respectively, and those of DHA were 73.87 μg/ml(SD, 2.30; CV, 3.11%) and 68.07 μg/ml (SD, 2.30; CV,
3.33%), respectively. The plasma level of each PUFA isexpressed as the mean ± SD weight (percentage) of thetotal fatty acids.
2.4.3. Plasma Levels of Cp.
A Bering BN Ⅱ Nephelometer (Siemens Healthcare
Diagnostics K.K., USA) was utilized to estimate plasma
CP levels. The assay sensitivity was 3.0 mg/dl. The intraand inter-assay coefficients were 10.2 mg/dl and 10.1
mg/dl, respectively.
2.4.4. Plasma Levels of SOD
Plasma SOD levels were estimated from the rate ofdecrease in nitrite produced by hydroxylamine and thesuperoxide anions based on the nitrite method, using a
Versa max instrument (Molecular Devices Co, Tokyo,
Japan). Human plasma was assayed using an SOD Assay
Kit (Takara Bio, Tokyo) according to the cytochrome cmethod. The plasma SOD levels are expressed as units per
milliliter. The assay sensitivity was 0.3 U/ml. The intraassay and inter-assay coefficients were 2.11 and 2.10 U/ml,
respectively.
2.4.5. Plasma Levels of Tf
A standard turbidimetric assay and an automatedbiochemical analyzer (JCA-BM8000 series, JEOL Ltd.,Tokyo, Japan) were utilized to estimate plasma Tf levels.
The intra- and inter-assay coefficients were 108.1 mg/dl
and 107.4 mg/dl, respectively.

2.5. Statistical Analyses
We calculated the ratios of DHA/AA and EPA/AA toevaluate the balance between omega-3 and omega-6
PUFAs. Because the data were not normally distributed,
the non-parametric Mann-Whitney U test was used for
multiple comparisons in order to find out the statisticaldifferences in estimates of daily intake for nutrients,
plasma levels of the 23 composition of fatty acids, plasma
ratios of DHA/AA and EPA/AA, and the plasma levels of
the three biomarkers (Cp, Tf and SOD), and the fivesubscale and total ABC scores between the 32 individuals
with ASD and the 20 normal controls. Spearman’s rankcorrelation coefficients (r) were used to determine thecorrelations between the plasma DHA/AA and EPA/AA
ratios, plasma signaling levels of biomarkers (Cp, Tf and
SOD), and the five subscale and total ABC scores in theentire population. In addition, Spearman’s rank correlation
coefficients were used to examine the relationshipbetween dietary intake of main nutrients (protein, fat ,omega-3 PUFA, omega 6-PUFAs, AA, EPA and DHA)
affecting these plasma PUFA levels and ratios and plasma
levels of EPA, DHA and AA, and plasma ratios of
DHA/AA and EPA/AA.

4

Journal of Translational Medicine and Developmental Disorders

Moreover, multiple linear regression analysis was used
to confirm the relationships between plasma DHA/AA andEPA/AA ratios, and the other main variables, adjusting fortwo subject groups, the three signaling biomarkers and thetotal ABC scores in order to confirm the relationshipbetween the ratios of omega-3 PUFAs/AA and AA-related
signaling mediators (Table 4). We conducted statisticalanalysis with SPSS version 18.0 (IBM Tokyo, 2009).

3. Results
3.1. Characteristics of Subjects

The mean total ABC score was 66.00 ± 29.22. Anearlier study reported a total ABC score of 60.14 forchildren and adolescents with moderate to severe ASD[53]. Thus, our patients were suffering from moderate tosevere ASD symptoms including restricted repetitive andstereotyped patterns of interests and behaviors. The age
did not differ significantly between the two groups (p =0.90). The ABC subscales scores for irritability (P =0.000), social withdrawal (p = 0.000), stereotypic behavior
p = 0.000), hyperactivity (p = 0.000) and inappropriate
speech (p = 0.000) and total ABC scores (p = 0.000) weresignificantly higher in the ASD group than in the control
group (Table 1).

Table 1. Subject characteristics, plasma levels of signaling mediators and the ABC subscale scores in the 32 individuals with ASD and 20
normal controls
Parameters
ASD (n=32)
Controls (n=20)
U
p valueAge (Year)13.5 ± 4.7
13.9 ± 5.7
313.000.90
Sex (male/female)
10/22
6/14
χ2=0.00
1.00
Scores of Autism Diagnostic Interview-Revised
Domain A (social)20.7 ± 6.9
N/A
Domain B (communication)
13.9 ± 5.0
N/A
Domain C (stereotyped)
7.0 ± 5.8
N/A
Plasma biomarkers levels (%)
Cp
24.90 ± 4.59
28.20 ± 4.97
209.500.037*
Tf
274.28 ± 47.77
270.50 ± 36.75
305.500.78
SOD
3.42 ± 2.58
3.84 ± 3.00
273.500.38
Scores of the ABC
Irritability
13.66 ± 9.23
0.50 ± 0.76
9.50
0.000***
Social withdrawal
22.81 ± 9.23
0.85 ± 2.06
1.50
0.000***
Stereotypy
6.00 ± 5.72
0.15 ± 0.49
68.50
0.000***
Hyperactivity
18.59 ± 10.67
0.60 ±1.60
10.00
0.000***
Inappropriate speech
4.91 ± 4.19
0.20 ± 0.52
50.00
0.000***
Total
66.00 ± 29.22
2.30 ± 4.44
1.00
0.000***
Data are means ± SD. ABC, Aberrant Behavior Cheklist; Cp, ceruloplasmin; Tf, transferrin, SOD, superoxide dismutase.* p < 0.05, *** p < 0.001:
individuals with ASD vs. controls, using Mann-Whitney-U test.

3.2. Assessment of Nutrient Intake
All 52 participants received “Japanese Food Guide
Spinning TOP” [44]. As shown in the Table 2, there wereno significant differences in weight, high, energy, andintake of cholesterol, protein, carbohydrate, fat, animal fat,saturated fatty acids and unsaturated fatty acids.
Importantly, intake of omega-3 (p = 0.54) and omega-6
PUFAs (p = 0.26), AA (p = 0.46), EPA (p = 0.71), DHA(p =0.81), iron (p = 1.00) and cupper (p = 0.81) did not
significantly differ between the ASD and control groups(Table 2).

Table 2. The intake of nutrients in the random subsamples of 17 individuals with ASD and 10 normal controls
ASD (n = 17)
Control (n = 10)U
p valueAge (years)
12.3 ± 4.1
13.3 ± 5.1
23.5
0.90
Heigh (cm)146.0 ± 23.0
151.6 ±1 9.1
21.5
0.71
Weight (kg)42.6± 23.6
45.6 ± 15.6
19.5
0.54
Energy (kcal)
2223.5 ± 563.3
2421.0 ± 503.3
19.0
0.54
Fat (g/day)
75.2 ± 28.5
87.3 ± 22.2
17.0
0.38
Saturated fatty acid (g/day)
25.8 ± 14.5
28.0 ± 10.2
20.0
0.62
Unsaturated fatty acid (g/day)
15.3 ± 4.2
18.4 ± 4.5
14.0
0.21
Omega-3 PUFAs (g/day)
2.8 ± 0.8
3.1 ± 0.5
19.0
0.54
Omega-6 PUFAs (g/day)
12.7 ± 3.7
15.6 ± 4.2
15.0
0.26
EPA (mg/day)
250.6 ± 168.1
197.8 ± 103.7
21.5
0.71
DPA (mg/day)
80.2 ± 47.0
72.3 ± 26.5
23.5
0.90
DHA (mg/day)
436.1 ± 237.0
380.8 ± 115.8
22.5
0.81
AA (mg/day)
172.6 ± 11.9
209.1 ± 75.0
18.5
0.34
Iron (mg/day)
9.7 ± 2.99.5 ± 3.0
22.0
1.00
Copper (g/day)
1.4 ± 0.4
1.3 ± 0.4
28.0
0.81
Zinc (mg/day)
10.3 ± 2.6
11.5 ± 3.3
16.0
0.32
Calcium (mg/day)
993.2 ± 633.1
876.9 ± 709.8
20.0
0.62
Protein (g/day)
84.2 ± 23.9
90.5 ± 24.1
19.0
0.54
Cholesterol (mg/day)
140.2 ± 195.8
95.6 ± 169.6
21.0
0.71
Carbohydrates (g/day)
298.9 ± 61.6
309.1 ± 59.7
24.0
1.00
Vitamin B6 (mg/day)
1.5 ± 0.5
1.4 ± 0.4
24.0
1.00
Vitamin D (mg/day)
10.6 ± 5.4
9.1 ± 2.4
21.5
0.71
Data are mean ± SD.

Journal of Translational Medicine and Developmental Disorders

3.3. Plasma levels of PUFAs and Biomarkers
The Mann-Whitney U test revealed that plasma levelsof EPA, DPA, DHA, and plasma DHA/AA and EPA/AA
ratios were significantly higher while plasma levels of AA,

5

5,8,11,14-eicosatetraenoic acid and Cp were significantly
lower in the 32 individuals with ASD than in the 20
normal controls (Table 3).

Table 3. Plasma levels of fatty acid profiles of the 32 individuals with ASD and the 20 normal controls
Parameters
ASD (n=32)
Controls (n=20)
U
Plasma PUFA levels (%)
Omega-3 series
C18:3ω3 (ALA)
0.74 ± 0.37
0.59 ± 0.20
234.00p value

0.11

C20:5ω3 (EPA)
C22:5ω3 (DPA)
C22:6ω3 (DHA)

1.36 ± 0.96
0.53 ± 0.17
3.67 ± 1.40

0.77 ± 0.40
0.45 ± 0.99
2.96 ± 0.89

160.00214.00205.000.003**
0046*
0.03*

Omega-6 series
C18:2ω6 (LA)C18:3ω6 (GLA)

29.08 ± 3.64
0.32 ± 0.16

29.58 ± 3.66
0.33 ± 0.03

316.50300.500.95
0.71

C20:2ω6 (DGLA)
C20:2ω6

1.16 ± 0.40
0.19 ± 0.04

1.36 ± 0.27
0.19 ± 0.04

221.00309.000.06
0.84

C20:4ω6 (ARA)

5.64 ± 1.36

6.77 ± 1.29

187.500.01*

Ratios of plasma levels of PUFAs
DHA/AA

0.66±0.22

0.45 ±0.15118.500.000***EPA/AA

0.24 ± 0.16

0.12 ± 0.07

109.500.000***Monounsaturated fatty acids
C14:1ω5

0.02 ± 0.03

0.04 ± 0.04

194.000.07

C16:1ω7
C18:1ω9

1.17 ± 0.77
20.44 ± 3.50

1.77 ± 0.69
21.01 ± 2.79

208.00260.000.82
0.26

C20:1ω9
C20:3ω9(5-8-11-14 eicosatetraenoic acid)

0.16 ± 0.06
0.08 ± 0.04

0.14 ± 0.03
0.08 ± 0.02

274.50191.500.39
0.020*

C22:1ω9
C24:1ω90.04 ± 0.03
1.17 ± 0.32

0.03 ± 0.02
1.16 ± 0.25

292.50305.500.59
0.79

Saturated fatty acids
C12
C14

0.14 ± 0.13
0.92 ± 0.44

0.16 ± 0.25
0.83 ± 0.33

282.50295.500.48
0.65

270.00259.50260.00

0.43
0.26
0.26

C16
23.27 ± 2.38
21.11 ± 5.09
C22
0.70 ± 0.17
0.67 ± 0.08
C24
0.60 ± 0.16
0.58 ± 0.09
Data are mean ± SD.
* p < 0.05. ** p < 0.01, *** p < 0.001: individuals with ASD vs. controls using Mann-Whitney-U test.

The Spearman’s rank correlation coefficients indicatedthat plasma DHA/AA (Figure 1a) and EPA/AA (Figure 1b)
ratios were significantly correlated with all five ABC
subscale scores and total scores (all r values were greaterthan 0.32, p < 0.05, p < 0.01 or p < 0.001) for the whole
population. Plasma levels of PUFAs (AA, EPA and DHA)
and signaling biomarkers (Cp, Tf and SOD) were notsignificantly correlated with the ABC scores.
A multiple linear regression demonstrated that theplasma ratios of DHA/AA ratio (R2 = 0.324, P = 0.002)

and AA (R2 = 0.260, P = 0.01) significantly associatedwith the variables adjusting for the two subject groups, thethree signaling biomarkers and the total ABC scores(Table 4). Using a group as a dependent variable
significantly contributed to the plasma DHA/AA ratio(unstandardized coefficients, B = -0.229 ± 0.091, β = 0.513, and p = 0.023). These findings indicated that theplasma DHA/AA ratio allowed for the prediction of thesevariables in the ASD and control groups.

Table 4. Results of the multiple linear regression
Model

Model R2

Model P-value

DHA/AA

0.32

0.002**

Coefficients
B

Beta coefficiences

p valueCp

0.011 ± 0.006

0.249

0.077†

Tf-0.001 ± 0.001-0.2350.079†

SOD-0.012 ± 0.010-0.145

0.248

ABC total score0.001 ± 0.001

0.053

0.800

Group (1 = ASD,2 = control)-0.229 ± 0.097-0.5130.023#
R2, R-squared values; Cp, ceruloplasmin; Tf, transferrin; SOD, superoxide dismutase; B = unstandardized coefficients; ABC, Aberrant Behavior
Checklist.
** p < 0.01: significance of R square. #p < 0.05, ## p <0.01: significant contribution., †p < 0.07 or p < 0.08: a trend towards significance.

6

Journal of Translational Medicine and Developmental Disorders

Figure 1. Correlation between plasma ratios of DHA/AA (Figure 1a) and EPA/AA (Figure 1b) and the ABC five subscale and total scores. Solid lines
indicated Spearman’s rank correlation coefficients in the 30 individuals with ASD and dotted lines indicated the 20 normal controls. Plasma DHA/AA
and EPA/AA ratios were significantly correlated with all five ABC subscale scores and total scores (all r values were greater than 0.32, p < 0.05, p <0.01 or p < 0.001) (Spearman’s rank correlation coefficients)

3.4. The Relationship between Main
Nutrients and Plasma Levels of EPA, DHA
and AA, and Plasma DHA/AA and EPA/AA
RatiosAlthough plasma EPA levels were significantly
negatively correlated to dietary intake of AA (r = -0.63. p= 0.015) in the random subsamples of 17 individuals with
ASD and the 10 normal controls, there was no significant
correlation between dietary intake of the main nutrientsand the plasma variables (r = 0.02 -0.59, p = 0.192 0.969).

4. Discussion
Plasma PUFA levels have been shown to reflectchanges in brain PUFA levels [54]. In this study, plasma

levels of EPA, DPA, and DHA, and plasma DHA/AA andEPA/AA ratios were significantly higher, while plasma
levels of AA, 5,8,11,14-eicosatetraenoic acid and Cp weresignificantly lower in the 32 individuals with ASD than in
the 20 normal controls (Table 1). We conducted
controlling for dietary intake in all 52 participants.
Moreover, the assessment of daily nutrients revealed nosignificant differences in intake of fat, omega-3 andomega-6 PUFAs, AA, EPA, DHA, iron, copper, zinc,Vitamin B6 and Vitamin D between the random
subsamples of 17 individuals with ASD and the 10 normal
controls. Although plasma EPA levels were significantly
negatively correlated to intake of AA in the random
subsamples of 17 individuals with ASD and the 10 normal
controls, there was no significant correlation between themain nutrients and the plasma variables. Thus, dietaryintake of nutrients may not directly affect the altered

Journal of Translational Medicine and Developmental Disorders

composition of fatty acids detected in this study may not
due to dietary food and nutrient intake.
Plasma DHA/AA and EPA/AA ratio were significantly
correlated with all five of the ABC subscales and total
scores. Notably, the multiple linear regression identified
that plasma DHA/AA ratio was fitting models for
distinguishing the ASD group from the control group andpredicting the adjusted variables (Table 4).
Cp is an important copper signaling biomarker ofneuronal function [55] with neuroprotective properties[25,26]. Cp is involved in the etiologies of several central
nervous system diseases [56]. Collectively, the reduced
plasma Cp levels may contribute to abnormal behavioral
symptoms in the 32 individuals with ASD.
Considering that plasma 5,8,11,14-eicosatetraenoic acidis an omega-6 PUFA, known as a trivial name of AA [57],the reduced plasma concentrations of AA may constitute apathophysiological factor that can lead to reduction inplasma Cp. As described above, Cp is closely related to
AA-derived eicosanoid mediator [58]. The omega-3
PUFAs are incorporated at existent of AA, and EPA and
DHA inhibit AA metabolism, decreasing production of
AA-derived
eicosanoids[16,19,20].
AA-derived
eicosanoids levels is decreased by these omega-3 PUFAs[20]. The antagonism of AA-signaling has been proposed
and plays important effects upon cell signaling in the CNS[17,18]. These previously reported strands therefore
support the present finding that the reduced plasma AA
levels were related to lowered plasma Cp levels.
In this study, increased plasma levels of omega-3
PUFAs such as DHA, DPA and EPA may be reflective of
increased biosynthesis of these omega-3 PUFAs. In
human cells, PUFAs are converted into elongated anddesaturated forms by elongase and desaturase enzymes[59], particularly delta-5 and delta-5 desaturases. The
activity of these enzymes is primarily regulated at thetranscriptional level [59]. Moreover, PUFAs effect on themodulation of genes related to fatty acid oxidation [60].
Therefore, the biosynthesis of DHA, DPA and EPA may
depend on the presence of transcriptional factors.
However, effects of altered omega-3 and omega-6 PUFA
biosynthesis in psychiatric disorders remains unclear.
However, dietary intake of AA might affect the alteredcomposition of fatty acids. Further studies are needed to
study the effects of altered omega-3 and omega-6 PUFA
biosynthesis and dietary intake of nutrients on plasma
variables
Previous studies have shown altered PUFA
compositions in autistic children aged 3-17 years old [9],3-15years old [10] and 4-12 years old [61], and in thoseyounger than 5 years of age [62]. In this study, an
imbalance between plasma levels of omega-3 PUFAs andplasma AA levels was found in ASD individuals with anaverage age of 13.5. At higher ages, omega-3 PUFAs may
protect against neurodegeneration [63]. By protecting thebrain from oxidative stress, omega-3 PUFAs minimize
brain damage and deterioration that occur with aging [64].
Therefore, the difference in the results between theseprevious studies and the present study might be due to anage-dependent metabolic mechanism.
This study had some limitations: a) the concentrationsof eicosanoid family members (e.g., cleukotriene [64])were not measured. However, AA-derived eicosanoid
signaling mediators includes many prostaglandin families

7[37], and these mediators are affected by multiple factors[38]. The regulation of iron and copper homeostasis isessential for life and may be related to thepathophysiology of several neurodegenerative disorders[65]. We therefore measured Cp, Tf and SOD. Further
studies should measure plasma levels of PUFA-related
eicosanoid family members such as prosdtaglandin E2 [66]or 18 eicosanoids [67]; b) the higher number of maleparticipants with ASD may have influenced the data
presented here because ASD is strongly biased towards
males with ratios of 4:1 (male : female) [68]; however, in
this study, ASD and normal control groups were age- andgender-matched. Further studies should evaluate the effectof gender on PUFAs compositions.
In conclusion, the present study reveals that a highplasma DHA/AA ratio related to the reduced plasma AA
levels may down-regulate mediators of AA signaling, suchas Cp, and dampen copper homeostasis. The plasma
concentrations of AA related fatty acid fractions may
constitute a pathophysiological factor that can lead to
reduction in plasma Cp. Subsequently, copper
dyshomeostasis may reduce the protective capacity of thebrain against damage, which may result in thepathophysiology of behavioral symptoms observed inindividuals with ASD.

Acknowledgements
Kunio Yui received a Grant-in-Aid for Scientific
Research on Innovative Areas (Grant No 21200017)(2010-2012) and a Grant-in-Aid for Scientific Research (C)(2014-2016) from the Ministry of Education, Culture,
Sports, Science and Technology, Japan.

Conflict of Interest
The authors have no conflict of interests to report.

References
[1]

[2]

[3]

[4]

[5]
[6]
[7]

Frye, R.E., Delatorre, R., Taylor, H., Slattery, J., Melnyk, S.,
Chowdhury, N., James, S.J. “Redox metabolism abnormalities in
autistic children associated with mitochondrial disease.” Transl
Psychiatry, 3, e273, 2013.
Wong, C., Crawford, D.A. Lipid signaling in the pathology ofautism spectrum disorders. In: V.B. Patel, V.R. Preedy, C.R.
Martin (Eds.), Comprehensive guide to autism, Springer
Publishing, New York, 2014, 1259-1283, 2014.
Holowka, D., Korzeniowski,.K., Bryant, K.L., Baird, B.
“Polyunsaturated fatty acids inhibit stimulated coupling between
the ER Ca2+ sensor STIM1 and the Ca2+ channel protein Orai1 in aprocess that correlates with inhibition of stimulated STIM1
oligomerization.” Biochim Biophys, 1841, 1210-1216, 2014.
Vrablik, T.L., Watts, J.L. “Polyunsaturated fatty acid derived
signaling in reproduction and development: insights from
Caenorhabditis elegans and Drosophila melanogaster.” Mol
Reprod Dev, 80, 244-259, 2013.
R. Crupi, A. Marino, S. Cuzzocrea, n-3 fatty acids: role inneurogenesis and neuroplasticity. Curr. Med. Chem. 20 (2013)2953-2963.
Tamiji, J., Crawford, D.A. “The neurobiology of lipid metabolism
in autism spectrum disorders.” Neurosignals, 18, 98-112, 2010.
Kurlak, L.O., Stephenson, T.J. “Plausible explanations for effectsof long chain polyunsaturated fatty acids (LCPUFA) on neonates.”
ArchDis Child Fetal Neonatal Ed, 80, F148-154, 1999.

8
[8]
[9]

[10]

[11]
[12]

[13]

[14]
[15]

[16]
[17]

[18]
[19]

[20]
[21]
[22]
[23]
[24]

[25]
[26]

[27]

[28]
[29]

Journal of Translational Medicine and Developmental Disorders
Bazinet, R.P., Layé, S. “Polyunsaturated fatty acids and their
metabolites in brain function and disease.” Nat Rev Neurosci, 15,771-785. 2014.
Vancassel, S., Durand, G., Barthélémy, C. Lejeune, B., Martineau,J., Guilloteau, D., Andrès, C., Chalon, S. “Plasma fatty acid levels
in autistic children.” Prostaglandins Leukot Essent Fatty Acids, 65,
1-7. 2001.
El-Anary, A.K., Bacha, A,G., Al-Ayahdi, L.Y. “Impaired plasma
phospholipids and relative amounts of essential polyunsaturated
fatty acids in autistic patients from Saudi Arabia.” Lipids HealthDis, 10 63, 2011a.
http://www.ncbi.nlm.nih. gov/pmc/articles/PMC3107801/.
Simopoulos, A.P. “Evolutionary aspects of diet: the omega6/omega-3 ratio and the brain.” Mol Neurobiol, 44, 203-215, 2011.
Luxwolda, M.F., Kuipers, R.S., Smit, E.N., Velzing-Aarts, F.V.,
Dijck-Brouwer, D.A., Muskiet, F.A. “The relation between theomega-3 index and arachidonic acid is bell shaped: synergistic at
low EPA+DHA status and antagonistic at high EPA+DHA status.”
Prostaglandins Leukot. Essent. Fatty Acids, 8, 171-178, 2011.
Taha, A.Y., Cheon, Y., Faurot, K.F., Macintosh,B., MajchrzakHong, S.F., Mann, J.D., Hibbeln, J.R., Ringel, A., Ramsden, C.E.
“Dietary omega-6 fatty acid lowering increases bioavailability ofomega-3 polyunsaturated fatty acids in human plasma lipid pools.”
Prostaglandins Leukot Essent Fatty Acids, 90, 151-157, 2014.
Schmitz, G., Ecker, J. “The opposing effects of n-3 and n-6 fatty
acid.” Prog Lipid Res, 47, 147-155, 2008.
Janssen, C.L., Kiliaan, A.J. “Long-chain polyunsaturated fatty
acids (LCPUFA) from genesis to senescence: The influence of
LCPUFA on neural development, aging, and neurodegeneration.”
Prog Lipid Res, 53, 1-17, 2014.
Whelan J. “Antagonistic effects of dietary arachidonic acid and n3 polyunsaturated fatty acids.” J Nutr, 126, 1086S-10891S, 1996.
Obajimi, O., Black, K.D., MacDonald, D.J., Boyle, R.M., Glen, I.,
Ross, B.M. “Differential effects of eicosapentaenoic anddocosahexaenoic acids upon oxidant-stimulated release and uptakeof arachidonic acid in human lymphoma U937 cells.” Pharmacol
Res, 52, 183-191, 2005.
Mori, T.A., Chem, C.P., Beilin, L.J. “Omega-3 Fatty Acids andInflammation.” Curr Atheroscler Rep, 6, 461-467, 2004.
Morbtaten, K., Haug, T.M., Kleiveland, C.R., Lea,T.”Omega-3
and omega-6 PUFAs induce the same GPR120-mediated signaling
events, but with different kinetics and intensity in Caco-2 cells.”
Lipids Health Dis, 12, 101, 2013.
Calder, C.P. “Polyunsaturated fatty acids and inflammatory
processes: new twists in an old tale.” Biochimie, 91, 791-795,
2009.
Sidhu, A., Miller, P.J., Hollenbach, A.D. “FOXO1 stimulates
ceruloplasmin promoter activity in human hepatoma cells treatedwith IL-6. Biochem. Biophys.” Res Commun, 404, 963-967, 2011.
Tsang, C.K., Liu, Y., Thoma, J., Zhang, Y., Zheng, X.F.
“Superoxide dismutase 1 acts as a nuclear transcription factor to
regulate oxidative stress resistance.” Nat Commun, 5, 3446, 2014.
Jian, J., Yang, Q., Huang, X. “Src regulates Tyr (20)phosphorylation of transferrin receptor-1 and potentiates breast
cancer cell survival.” J Biol Chem, 286, 35708-35715, 2011.
Barbariga, M., Curnis, F., Spitaleri, A., Andolfo, A., Zucchell, C.,
Lazzaro, M., Magnani, G., Musco, G., Corti, A., Alessio, M.
“Oxidation-induced structural changes of ceruloplasmin foster
NGR motif deamidation that promotes integrin binding andsignaling.” J Biol Chem, 289, 3736-3748, 2014.
Glezer, I., Chernomoretz, A., David, S., Plante, M.M., Rivest, S.
“Genes involved in the balance between neuronal survival anddeath during inflammation.” PLoS One, 2(3), e310, 2007.
Ayton, S., Zhang, M., Roberts, B.R., Lam, L.Q., Lind, M.,
McLean, C., Bush, A.I., Frugier, T., Crack, P.J., Duce, J.A.
“Ceruloplasmin and β-amyloid precursor protein confer
neuroprotection in traumatic brain injury and lower neuronaliron.” Free Radic Biol Med, 69, 331-337, 2014.
Lazzaro, M., Bettegazzi, B., Barbariga, M., Codazzi, F., Zacchetti,
D., Alessio, M. “Ceruloplasmin potentiates nitric oxide synthaseactivity and cytokine secretion in activated microglia.” J
Neuroinflammation, 11, 164, 2014.
Fukai, T., Ushio-Fukai, M. “Superoxide dismutases: role in redox
signaling, vascular function, and diseases.” Antioxid Redox Signal,
15, 1583-1606, 2011.
Moura, I.C., Hermine, O., Lacombe, C., Mayeux, P.
“Erythropoiesis and transferrin receptors.” Curr Opin Hematol,
193-198, 2015.

[30] Yorbik, O.Sayal, A., Akay, C., Akbiyik, D.I., Sohmen, T.

[31]

[32]

[33]

[34]

[35]

[36]

[37]

[38]
[39]
[40]
[41]
[42]

[43]

[44]

[45]
[46]
[47]

[48]

[49]

“Investigation of antioxidant enzymes in children with autistic
disorder.” Prostagland Leukot Essent Fatty Acids, 67, 341-343,
2002.
Söğüt, S., Zoroğlu, S.S., Ozyurt, H., Yilmaz, H.R., Ozuğurlu, F.,
Sivasli, E., Yetkin, O., Yanik, M., Tutkun, H., Savaş, H.A.,
Tarakçioğlu, M., Akyol, O. “Changes in nitric oxide levels andantioxidant enzyme activities may have a role in thepathophysiolgical mechanisms involved in autism.” Clin Chim
Acta, 331, 111-117, 2006.
Zoroglu, S.S., Armutcu, F., Ozen, S., Gurel, A., Sivasli, E.,
Yetkin,O., Meram, I. “Increased oxidative stress and alteredactivities of erythrocyte free radical scavenging enzymes in
autism.” Eur Arch Psychiatry Clin Neurosci, 254, 143-147, 2004.
Meguid, N.A., Dardir, A.A., Abdel-Raouf, E.R., Hashish, A.
“Evaluation of oxidative stress in autism: defective antioxidant
enzymes and increased lipid peroxidation.” Biol Trace Elem Res,
143, 58-65, 2011.
Parellada, M., Moreno, C., Mac-Dowell, K., Leza, J.C., Giraldez,
M., Bailón, C., Castro, C., Miranda-Azpiazu, P., Fraguas, D.,
Arango, C. “Plasma antioxidant capacity is reduced in Asperger
syndrome.” J Psychiat Res, 46, 394-401, 2012.
Chauhan, A., Chauhan, V., Brown, W.T., Cohen, I. “Oxidative
stress in autism: increased lipid peroxidation and reduced serum
levels of ceruloplasmin and transferrin-the antioxidant proteins.”
Life Sci, 75, 2539-2549, 2004.
Tórsdóttir, G., Hreidarsson, S., Kristinsson, J., Snaedal, J.,
Jóhannesson, T. “Ceruloplasmin, superoxide dismutase and copperin autistic patients.” Basic Clin Pharmacol Toxicol, 96, 146-148,
2005.
F.H. Tessaro, F.H., T.S. Ayala, T.S., J.Q. Martins, J.Q. “Lipid
mediators are critical in resolving inflammation: a review of theemerging roles of eicosanoids in diabetes mellitus.” Biomed Res
Int, 2015:568408.
http://www.ncbi.nlm.nih. gov/ pmc/articles/PMC4383369/.
Hata, A.N., Breyer, R.M. “Pharmacology and signaling of
prostaglandin receptors: multiple roles in inflammation andimmune modulation.” Pharmacol Ther, 103, 147-166, 2004.
Rutter, M., Le Couteur, A., Lord, C. “ADI-R Autism Diagnostic
Interview Revised Manual.” Western Psychological Services, Los
Angeles, 2003, UAS.
D. Wechsler, Wechsler Intelligence Scale for Children-Revised
Manual. The Psychological Corporation, New York, NY, USA(1974).
D. Wechsler. Wechsler Adult Intelligence Scale-Revised Manual.
The Psychological Corporation, San Antonio, TX, USA (1981).
Koyama, T., Kamio, Y., Inada, N., Kurita, H. “Sex differences in
WISC-III profiles of children with high-functioning pervasive
developmental disorders.” J Autism Dev Disord 39, 135-141,
2009.
Schroeder, N., Park, Y.H., Kang, M.S., Kim, Y., Ha, G.K., Kim,
H.R., Yates, A.A., Caballero. B. “A Randomized Trial on the
Effects of 2010 Dietary Guidelines for Americans and Korean
Diet Patterns on Cardiovascular Risk Factors in Overweight andObese Adults.” J Acad Nutr Diet, 115, 1083-1092, 2015.
Ministry of Health, Labour and Welfare, and Ministry ofAgriculture, Forestry and Fishers, “Japanese Food Guide.”
Ministry of Health, Labour and Welfare, and Ministry ofAgriculture, Forestry and Fishers, Tokyo, Japan, 2012.
Ministry of Health, Labour and Welfare, “Dietary Reference
Intakes for Japanese. -2010-.” Ministry of Health, Labour andWelfare, Tokyo, Japan, 2010.
Ministry of Health, Labour, and Welfare. “Overview of dietaryreference Intake for Japanese (2015).” Ministry of Health, Labour,and Welfare, Tokyo, Japan, 2015.
Okuda, M., Sasaki, S., Bando, N., Hashimoto, M., Kunitsugu, I.,
Sugiyama, S., Terao, J., Hobara, T. “Carotenoid, tocopherol, andfatty acid biomarkers and dietary intake estimated by using a brief
self-administered diet history questionnaire for older Japanesechildren and adolescents.” J Nutr Sci Vitaminol (Tokyo), 55, 231241, 2009.
Rojahn, J., Aman, M.G., Matson, J.L., Mayville, E. “The Aberrant
Behavior Checklist and the Behavior Problems Inventory:
convergent and divergent validity.” Res Dev Disabil, 24 (2003)
391-404, 2003.
Hollander, E., Chaplin, W., Soorya, L., Wasserman, S., Novotny,
S., Rusoff, J., Feirsen, N., Pepa, L., Anagnostou, E. “Divalproex
sodium vs. placebo for the treatment of irritability in children andJournal of Translational Medicine and Developmental Disorders

[50]
[51]

[52]
[53]

[54]

[55]

[56]

[57]

[58]
[59]

adolescentswithautismspectrum
disorders.”
Neuropsychopharmacology, 35, 990-998, 2009.
Karabekiroglu, K., Aman, M.G., “Validity of the aberrant
behavior checklist in a clinical sample of toddlers.” Child
Psychiatry Hum Dev, 40, 99-110, 2009.
Hamazaki, H., Itomura, M., Hamazaki, T., Sawazaki, S. “Effectsof cooking plant oils on recurrent aphthous stomatitis: arandomized, placebo-controlled, double-blind trial.” Nutrition, 22,534-536, 2006.
Bligh, E.G., Dyer, W.J. “A rapid method of total lipid extraction
and purification.” Can J Med Sci, 37, 911-917, 1959.
Singh, K., Connors, S.L., Macklin, E.A., Smith, K.D., Fahey, J.W.,
Talalay, P., Zimmerman, A,W. “Sulforaphane treatment of autism
spectrum disorder (ASD).” Proc Natl Acad Sci U S A, 111, 1555015555, 2014.
Sublette, M.E., Bosetti, F., DeMar, J.C., Ma, K., Bell, J.M., FaginJones, S., Russ, M.J., Rapoport, S.I. “Plasma free polyunsaturated
fatty acid levels are associated with symptom severity in acutemania.” Bipolar Disord, 9, 759-765, 2007.
Barbariga, M., Curnis, F., Spitaleri, A., Andolfo, A., Zucchelli, C.,
Lazzaro, M., Magnani, G., Musco, G., Corti, A., M. Alessio, M.
“Oxidation-induced structural changes of ceruloplasmin foster
NGR motif deamidation that promotes integrin binding andsignaling.” J Biol Chem, 289, 3736-3748, 2014.
Squitti, R., Ghidoni, R., Siott, M., Ventriglia, M., Benussi, L.,
Paterlini, A., Magri, M. Binetti, G., Cassetta, E., Caprara, D.,
Vernieri, F., Rossini, P.M., Pasqualetti, P. Value of serum
Nonceruloplasmin copper for prediction of mild cognitive
impairment conversion to Alzheimer disease. Ann Neurol, 75,
574-580, 2014.
Dalvi, S., Nguyen, H.H., On, N., Mitchell, R.W., Aukema, H.M.,
Miller, D.W., Hatch, G.M. “Exogenous arachidonic acid mediates
permeability of human brain microvessel endothelial cells throughprostaglandin E2 activation of EP3 and EP4 receptors.” J
Neurochem, 2015.
Sokov, A.V., Golenkina, E.A., Kostevich, V.A., Vasilyev, V.B.,
Sud’ina, G.F. “Interaction of ceruloplasmin and 5-lipoxygenase.”
Biochemistry (Mosc), 75, 1464-1469, 2010.
Seegmiller. A.C. “Abnormal unsaturated fatty acid metabolism incystic fibrosis: biochemical mechanisms and clinicalimplications.” Int J Mol Sci, 15, 16083-16099, 2014.

9[60] Jacometo, C.B., Schmitt, E., Pfeifer, L.F., Schneider, A., Bado, F.,

[61]
[62]

[63]
[64]
[65]

[66]
[67]

[68]

[69]

da Rosa, F.T., Halfen, S., Del Pino, F.A., Loor, J.J., Corrêa, M.N.,
Dionello, N.J. “Linoleic and α-linolenic fatty acid consumption
over three generations exert cumulative regulation of hepatic
expression of genes related to lipid metabolism.” Genes Nutr, 9,405, 2014.
El-Ansary, A.K., Bacha, A.G., Al-Ayahdi. L.Y. “Plasma fatty
acids as diagnostic markers in autistic patients from Saudi
Arabia.” Lipids Health Dis, 10, 62, 2011b.
Al-Farsi, Y.M., Waly, M.I., Deth, R.C., Al-Sharbati, M.M., AlShafaee, M., Al-Farsi, O., Al-Khaduri, M.M., Al-Adawi, S.,
Hodgson, N.W., Gupta, I., Ouhtit, A, “Impact of nutrition on
serum levels of docosahexaenoic acid among Omani children with
autism.” Nutrition, 29, 1142-1146, 2013.
Karr, J.E., Alexander, R.G. Winningham, R.G. “Omega-3
polyunsaturated fatty acids and cognition throughout the lifespan:a review.” Nutr Neurosci, 14, 216-225, 2011.
Innis SM. Dietary (n-3) fatty acids and brain development. J Nutr,
137, 855-859, 2007.
Adel, S., Kakularam, K.R., Horn, T., Reddanna, P., Kuhn, H.,
Heydeck, D. “Leukotriene signaling in the extinct humansubspecies Homo denisovan and Homo neanderthalensis.
Structural and functional comparison with Homo sapiens.” Arch
Biochem Biophys, 565, 17-24, 2015.
Zheng, W., Monnot, A.D. “Regulation of brain iron and copper
homeostasis by brain barrier systems: implication inneurodegenerative diseases.” Pharmacol Ther, 133,177-188, 2012.
J.H. Chen C.J. Perry, Y.C. Tsui, M.M. Staron I.A. Parish, C.X.
Dominguez, D.W. Rosenberg, S.M. Kaech, Prostaglandin E2 andprogrammed cell death 1 signaling coordinately impair CTL
function and survival during chronic viral infection. Nat. Med.
21(2015) 327- 334
Rago, B., Fu, C. “Development of a high-throughput ultra
performance liquid chromatography-mass spectrometry assay to
profile 18 eicosanoids as exploratory biomarkers foratherosclerotic diseases.” J Chromatogr B Analyt Technol Biomed
Life Sci, 936, 25-32, 2013.
Baron-Cohen, S., Lombardo, M.V., Auyeung, B., Ashwin, E.,
Chakrabarti, B., Knickmeyer, R. “Why are autism spectrum
conditions more prevalent in males?” PLoS Biol. 9, e1001081,
2012.