Personalized Medicine Universe xxx (2013) 1e4

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Personalized Medicine Universe
journal homepage: www.elsevier.com/locate/pmu

Original article

DLe/POephosphatidylcholine serves as a memory enhancer for normal healthy
subjects
Takaaki Nishimoto a, Takeshi Kanno a, Akinobu Gotoh b, Tomoyuki Nishizaki a, *abDivision of Bioinformation, Department of Physiology, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya 663-8501, Japan
Laboratory of Cell and Gene Therapy, Institute for Advanced Medical Sciences, Hyogo College of Medicine, 1-1 Mukogawa-cho, Nishinomiya 663-8501, Japana r t i c l e i n f oa b s t r a c t

Article history:
Received 11 December 2012
Accepted 18 March 2013

Purpose: The present study investigated the effect of 1,2-dilinoleoyl-sn-glycero-3-phosphocholine (DLePC) and 1-palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine (POePC) on learning and memory innormal healthy mice and humans.
Study section and results: In the water maze test, normal mice orally co-administered DLePC (1 mg/kg)
and POePC (1 mg/kg) showed signiﬁcantly shortened retention latency compared to mice administeredpolyethylene glycol; however, the acquisition latency was not affected. For humans, a memory test wascarried out on 34 subjects; they had 5 min to learn 3 different kinds of 2 Chinese compound words, aJapanese poem consisting of 17 syllables, and a sentence chosen from an old Japanese essay. One minute(short-term memory test) and 1 week (long-term memory test) later, they wrote what they had learned.
For subjects taking DLePC (50 mg/day) and POePC (45 mg/day), the percentage of correct answers forboth the short- and long-term memory tests increased in a trial-number-dependent manner. The percentage of correct answers in the long-term memory test at the third trial for subjects taking DLe/POePC
was nearly 3 times higher than that of control subjects, even though there was no signiﬁcant differencein the percentage of correct answers in the short-term memory test between the 2 groups.
Conclusion: The results of the present study indicate that DLe/POePC could enhance long-term memory
potential for normal healthy subjects.
Copyright Ó 2013, International Society of Personalized Medicine. Published by Elsevier B.V. All rights
reserved.

Keywords:Phosphatidylcholine
Healthy subject
MemoryEnhancer

1. Introduction
Phospholipids such as phosphatidylcholine, phosphatidylethanolamine, phosphatidylserine, and phosphatidylinositol are implicated in the regulation of a wide variety of bioreactions. In ourprevious study, 1,2-dilinoleoyl-sn-glycero-3-phosphocholine (DLePC) potentiated a7 nicotinic acetylcholine (nACh) receptor responses and facilitated hippocampal synaptic transmission in an a7nACh receptor-dependent manner [1]. The a7 nACh receptor playsa critical role in the expression of long-term potentiation (LTP), acellular model of learning and memory [2e9]. DLePC, therefore,
could improve learning impairments by targeting the a7 nACh

Abbreviations: DLePC, 1,2-dilinoleoyl-sn-glycero-3-phosphocholine; POePC, 1palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine; ACh, Acetylcholine; nACh, nicotinic ACh; LTP, long-term potentiation; LTD, long-term depression; AMPA, a-amino3-hydroxy-5-methyl-4-isoxazole propionic acid; AChEIs, acetylcholinesterase inhibitors; PEG, polyethylene glycol; PLSD, Protected Least Signiﬁcant Difference.
* Corresponding author. Tel.: þ81 798 45 6397; fax: þ81 798 45 6649.
E-mail address: tomoyuki@hyo-med.ac.jp (T. Nishizaki).

receptor.
Incontrast,
1-palmitoyl-2-oleoyl-sn-glycero-3phosphocholine (POePC) enhanced long-term depression (LTD),
another cellular model of learning and memory, in concert withdecreased expression of the a-amino-3-hydroxy-5-methyl-4isoxazole propionic acid (AMPA) receptor subunit GluR1 on theplasma membrane [10]. Consequently, POePC may also improve
memory disorders. Indeed, DLe and/or POePC ameliorates cognitive decline in humans [10,11]. Interestingly, taking a combinationof DLe and POePC had an additive effect that was more beneﬁcial
in treating cognitive impairment than was the intake of either DLe
PC or POePC alone [11].
Alzheimer disease is characterized by severe dementia in association with very rapidly progressive brain atrophy. Although nopromising drug has been identiﬁed, ACh esterase inhibitors(AChEIs), such as donepezil, galantamine, and rivastigmine, havebeen clinically used in Alzheimer disease to alleviate symptoms[12]. We recently found that DLe/POePC has the potential toameliorate Alzheimer dementia, and is more beneﬁcial with ahigher efﬁcacy than donepezil (unpublished data). Thus, DLe/POe
PC could be useful for the treatment of cognitive decline and2186-4950/$ e see front matter Copyright Ó 2013, International Society of Personalized Medicine. Published by Elsevier B.V. All rights reserved.
http://dx.doi.org/10.1016/j.pmu.2013.04.002

Please cite this article in press as: Nishimoto T, et al., DLe/POephosphatidylcholine serves as a memory enhancer for normal healthy subjects,
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2T. Nishimoto et al. / Personalized Medicine Universe xxx (2013) 1e4

dementia. The present study was conducted to understand the effect of DLe/POePC on learning and memory abilities of normal,
healthy subjects.
To address this question, we carried out a water maze test byusing normal mice and a memory test that we devised for healthy
humans. We show here that DLe/POePC serves as a memoryenhancer for normal healthy subjects.
2. Materials and methods
2.1. Animal care
All procedures were approved by the Animal Care and Use
Committee at Hyogo College of Medicine and were in compliancewith the National Institutes of Health Guide for the Care and Use ofLaboratory Animals.
2.2. Participants
The study with human subjects was approved by the Institutional Review Board at Hyogo College of Medicine, and informed
consent was obtained from all the participants.
2.3. Water maze test
A circular plastic water tank 100 cm in diameter and 35 cm deep
was used for the water maze test. The entire inside of the pool waspainted white, and the pool was ﬁlled to a depth of 25 cm withmuddy water containing India ink at 22  C. A platform (11 cm indiameter) painted white was placed into the pool, such that the top
was 1 cm below the water surface. The pool was put in a test room
containing several marks that mice were able to see from the pool.
The position of the marks remained unchanged throughout testing.
A platform was located in a constant position, i.e., in the middle ofone quadrant, equidistant from the center and edge of the pool.
Male C57BL/6 mice (7 weeks old) facing the wall of the pool wereplaced into water at 1 of 5 positions selected at random, and thetime from start to escape onto the platform (acquisition latency)
was measured. DLe and POePC dissolved in polyethylene glycol(PEG) (ﬁnal volume, 0.1 ml) or PEG alone (ﬁnal volume, 0.1 ml) wereorally administered to mice for 16 days from the beginning to theend of all phases of the water maze test. There were 2 trials of thewater maze task per day, and the second trial began 1 min after theend of the ﬁrst trial. Mice participated in the task for 8 consecutive
days and the acquisition latency (time from start to arrival onto theplatform) was measured.
Eight days later, the platform was removed and the retention
latency (time from start to arrival to the place where the platform
had been set, 90 s maximum) was measured.

Fig. 1. Effect of DLe/POePC on spatial learning and memory. Mice were orally
administered DLePC (1 mg/kg) plus POePC (1 mg/kg) (DLe/POePC) or PEG everyday
from the beginning to the end of all phases of the water maze test. (A) Each point
represents the mean (SEM) acquisition latency from 2 consecutive days (n ¼ 10). (B)Each column represents the mean (SEM) retention latency (n ¼ 10). P value, unpaired
t-test.

2.5. Statistical analysis
Statistical analysis was carried out using Fisher’s Protected Least
Signiﬁcant Difference (PLSD) test and unpaired t-test.

2.4. Memory test
The memory test that we devised was carried out in 34 healthy
humans, who were divided into 2 groups, i.e., 20 subjects aged 24e
56 years (average, 38  2 years; male:female ¼ 6:14) took both DLe
PC (50 mg/day) and POePC (45 mg/day) orally everyday for 1
month prior to the test (DLe/POePC-intake group), and 14 subjects
aged 23e49 years (average, 35  3 years; male:female ¼ 2:12) took
nothing (control group). All subjects had 5 min to learn 3 different
kinds of 2 Chinese compound words, a Japanese poem consisting of
17 syllables, and a sentence chosen from an old Japanese essay, asshown in Fig. 2. They were then asked to recall the words, thepoem, and the sentence that they remembered 1 min (short-termmemory test) and 1 week (long-term memory test) after the test,
and the percentage of correct answers was calculated.

Fig. 2. An example of the memory test including 2 Chinese compound words, a Japanese poem consisting of 17 syllables, and a sentence chosen from an old Japanese
essay.

Please cite this article in press as: Nishimoto T, et al., DLe/POephosphatidylcholine serves as a memory enhancer for normal healthy subjects,
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3. Results and discussion
The water maze test was carried out in mice with oral administration of DLePC (1 mg/kg) and POePC (1 mg/kg) (DLe/POePCgroup) or PEG (PEG group). There was no signiﬁcant difference inthe swimming speed between the PEG (63.0  13.4 mm/s) and DLe

3/POePC groups (82.9  20.5 mm/s) (P ¼ 0.0550, unpaired t-test),
indicating that DLe/POePC has no effect on swimming speed.
Oral co-administration of DLePC (1 mg/kg) and POePC (1 mg/kg) did not affect the acquisition latency compared to the latencyfrom the PEG group (P ¼ 0.1651, Fisher’s PLSD test) (Fig. 1A). In
contrast, the retention latency for the DLe/POePC group wassigniﬁcantly shorter than the latency for the PEG group (Fig. 1B).
Taken together, these results indicate that DLe/POePC is capable ofenhancing memory ability rather than learning ability for normalmice.
We next carried out a memory test on normal healthy humans
who were untreated (control group) or were taking DLePC (50 mg/day) and POePC (45 mg/day) orally (DLe/POePC-intake group). For
the DLe/POePC-intake group, the percentage of correct answers on
both the short- and long-term memory tests increased in a trialnumber-dependent manner, but this effect was not found withthe control group (Fig. 3A,B). The percentage of correct answers in
the long-term memory test at the third trial was signiﬁcantly
higher for the DLe/POePC-intake group than for the control group,with the DLe/POePC-intake group having approximately 3 times
as many correct answers (Fig. 3C). In contrast, there was no signiﬁcant difference in the percentage of correct answer in the shortterm memory test between the 2 groups (Fig. 3C). This indicatesthat DLe/POePC has the potential to enhance long-term memory
ability in healthy humans.
DLePC induces LTP by targeting a7 nACh receptor [1], suggesting that DLePC is a learning enhancer. In contrast, POePC by itself
potentiates LTD by decreasing cell surface localization of the GluR1
AMPA receptor [10], suggesting that POePC may be a memoryenhancer. We have provided evidence that DLe and/or POePC
actually ameliorates cognitive decline and dementia in humans[10,11]. The results of the present study clearly demonstrate thatDLe/POePC serves as a memory enhancer even in normal, healthy
subjects. Thus, DLe/POePC may not only avert a variety of dementias including Alzheimer disease in humans, but also enhancecognitive functions. DLe/POePC has the advantage that it is anatural substance in living cells and exhibits few side effects.
Therefore, many humans could safely and easily take DLe/POePCfor improving brain health.
ReferencesFig. 3. Effect of DLe/POePC on memory in normal, healthy humans. A memory test
was carried out on 14 untreated subjects [DLe/POePC ()] and 20 subjects with taking
both DLePC (50 mg/day) and POePC (45 mg/day) orally [DLe/POePC (þ)]. (A) The
percentages of correct answers on the short-term memory test (1 min after the trial) at
each trial for individual subjects from the control group [DLe/POePC ()] and thegroup taking DLe and POePC [DL/POePC (þ)] were plotted and lines ﬁtted to eachpoint were drawn. (B) The percentages of correct answer for the long-term memory
test (1 week after the trial) at each trial for individual subjects from the control group[DLe/POePC ()] and the group taking DLe and POePC [DLe/POePC (þ)] were plotted
and the lines ﬁtted to each point were drawn. (C) Each column represents the mean(SEM) percentage of correct answers in the short- and long-term memory tests at thethird trial for untreated subjects [DLe/POePC ()] and subjects taking DLe and POePC[DLe/POePC (þ)]. P value, unpaired t-test. NS, not signiﬁcant.

[1] Yaguchi T, Nagata T, Nishizaki T. Dilinoleoylphosphatidylcholine ameliorates
scopolamine-induced impairment of spatial learning and memory by targeting a7 nicotinic ACh receptors. Life Sci 2009;84:263e6.
[2] Matsuyama S, Matsumoto A, Enomoto T, et al. Activation of nicotinic acetylcholine receptors induces long-term potentiation in vivo in the intact mousedentate gyrus. Eur J Neurosci 2000;12:3741e7.
[3] Nishizaki T, Matsuoka T, Nomura T, et al. Presynaptic nicotinic acetylcholine
receptors as a functional target of neﬁracetam in inducing a long-lasting
facilitation of hippocampal neurotransmission. Alzheimer Dis Assoc Disord
2000;14(Suppl. 1):S82e94.
[4] Nishizaki T, Matsuoka T, Nomura T, et al. A ‘long-term-potentiation-like’
facilitation of hippocampal synaptic transmission induced by the nootropic
neﬁracetam. Brain Res 1999;826:281e8.
[5] Nishizaki T, Nomura T, Matsuoka T, et al. Arachidonic acid induces a longlasting facilitation of hippocampal synaptic transmission by modulating PKC
activity and nicotinic ACh receptors. Mol Brain Res 1999;69:263e72.
[6] Nishizaki T, Nomura T, Matsuyama S, et al. Critical role of nicotinic ACh receptor in the formation of long-term potentiation: implications for development of anti-dementia drugs. Psychogeriatrics 2001;1:209e17.
[7] Nishizaki T, Nomura T, Matuoka T, et al. The anti-dementia drug neﬁracetam
facilitates hippocampal synaptic transmission by functionally targeting presynaptic nicotinic ACh receptors. Mol Brain Res 2000;80:53e62.
[8] Tanaka A, Nishizaki T. The newly synthesized linoleic acid derivative
FR236924 induces a long-lasting facilitation of hippocampal neurotransmission by targeting nicotinic acetylcholine receptors. Bioorg Med Chem Lett
2003;13:1037e40.
[9] Yamamoto S, Kanno T, Nagata T, et al. The linoleic acid derivative FR236924
facilitates hippocampal synaptic transmission by enhancing activity of presynaptic a7 acetylcholine receptors on the glutamatergic terminals. Neuroscience 2005;130:207e13.

Please cite this article in press as: Nishimoto T, et al., DLe/POephosphatidylcholine serves as a memory enhancer for normal healthy subjects,
Personalized Medicine Universe (2013), http://dx.doi.org/10.1016/j.pmu.2013.04.002

4T. Nishimoto et al. / Personalized Medicine Universe xxx (2013) 1e4[10] Yaguchi T, Nagata T, Nishizkai T. 1-Palmitoyl-2-oleoyl-sn-glycero-3-phosphocholine improves cognitive decline by enhancing long-term depression.
Behav Brain Res 2009;204:129e32.
[11] Nagata T, Yaguchi T, Nishizaki T. DL- and PO-phosphatidylcholines as apromising learning and memory enhancer. Lipids Health Dis 2011;10:25.

[12] Bond M, Rogers G, Peters J, et al. The effectiveness and cost-effectivenessof donepezil, galantamine, rivastigmine and memantine for the treatmentof Alzheimer’s disease (review of Technology Appraisal No. 111): asystematic review and economic model. Health Technol Assess 2012;16:
1e470.

Please cite this article in press as: Nishimoto T, et al., DLe/POephosphatidylcholine serves as a memory enhancer for normal healthy subjects,
Personalized Medicine Universe (2013), http://dx.doi.org/10.1016/j.pmu.2013.04.002