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Endocannabinoids and pregnancy

Authors:
  • Leicester University and Nottingham Trent University

Abstract and Figures

Acylethanolamides such as anandamide (AEA), and monoacylglycerols like 2-arachidonoylglycerol are endocannabinoids that bind to cannabinoid, vanilloid and peroxisome proliferator-activated receptors. These compounds, their various receptors, the purported membrane transporter(s), and related enzymes that synthesize and degrade them are collectively referred to as the "endocannabinoid system (ECS)". Poorly defined cellular and molecular mechanisms control the biological actions of the ECS. Over the last decade evidence has been emerging to suggest that the ECS plays a significant role in various aspects of human reproduction. In this review, we summarize our current understanding of this role especially the involvement of AEA and related ECS elements in regulating oogenesis, embryo oviductal transport, blastocyst implantation, placental development and pregnancy outcomes, and sperm survival, motility, capacitation and acrosome reaction. Additionally, the possibility that plasma and tissue AEA and other cannabinoids may represent reliable diagnostic markers of natural and assisted reproduction and pregnancy outcomes in women will be discussed.
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Invited critical review
Endocannabinoids and pregnancy
Anthony H. Taylor, Akwasi A. Amoako, Katerina Bambang, Tulay Karasu, Alpha Gebeh, Patricia M.W. Lam,
Timothy H. Marzcylo, Justin C. Konje
Endocannabinoid Research Group, Reproductive Sciences Section, University of Leicester, Leicester, United Kingdom
abstractarticle info
Article history:
Received 18 December 2009
Received in revised form 9 March 2010
Accepted 9 March 2010
Available online 17 March 2010
Keywords:
Anandamide
Endocannabinoids
Pregnancy
Reproduction
Acylethanolamides such as anandamide (AEA), and monoacylglycerols like 2-arachidonoylglycerol are
endocannabinoids that bind to cannabinoid, vanilloid and peroxisome proliferator-activated receptors. These
compounds, their various receptors, the purported membrane transporter(s), and related enzymes that
synthesize and degrade them are collectively referred to as the endocannabinoid system (ECS). Poorly
dened cellular and molecular mechanisms control the biological actions of the ECS. Over the last decade
evidence has been emerging to suggest that the ECS plays a signicant role in various aspects of human
reproduction. In this review, we summarize our current understanding of this role especially the
involvement of AEA and related ECS elements in regulating oogenesis, embryo oviductal transport,
blastocyst implantation, placental development and pregnancy outcomes, and sperm survival, motility,
capacitation and acrosome reaction. Additionally, the possibility that plasma and tissue AEA and other
cannabinoids may represent reliable diagnostic markers of natural and assisted reproduction and pregnancy
outcomes in women will be discussed.
© 2010 Elsevier B.V. All rights reserved.
Contents
1. Introduction . . . . . . . . . . .................................................. 921
2. The endocannabinoid system (ECS).................................................. 922
3. Endocannabinoids and gametogenesis . . . . . ........................................... 923
3.1. Oogenesis . . . . . . . . .................................................. 923
3.2. Spermatogenesis . . . . . .................................................. 924
4. Endocannabinoids, early embryo-blastocyst development and oviductal transport . . . . . . . ..................... 924
4.1. Endocannabinoids and early embryo development . . . . . . . . . . . . . . . . . . ..................... 924
4.2. Endocannabinoids and transport through the Oviduct . . . . . . . . . . . . . . . . . ..................... 924
5. Endocannabinoids and implantation ................................................. 925
6. Endocannabinoids in early pregnancy................................................. 925
7. Endocannabinoids in the maintenance of pregnancy and labour. . . . . . . . . . . . . . . . ..................... 925
8. Feto-placental changes in endocannabinoid levels ........................................... 926
9. Possible immunocannabinoidsystem actions in pregnancy . . . . . . . . . . . . . . . . . ..................... 926
10. Conclusions . . . . . . . . . . .................................................. 927
Conicts of interest . . . . . . . . . . .................................................. 927
References . . .............................................................. 927
1. Introduction
Endocannabinoids are endogenous ligands that bind to the same
receptors as the most psychoactive agent in marijuana, delta-9-
tetrahydrocannabinol (Δ
9
-THC). The use of marijuana, an illicit drug
in the UK and USA [1,2], appears to be increasing [3,4]. Marijuana use
is associated with infertility in males and females, spontaneous
miscarriages, placental abruption, preterm birth, stillbirths [57] and
Clinica Chimica Acta 411 (2010) 921930
Corresponding author. Endocannabinoid Research Group, Reproductive Sciences
Section, Department of Cancer Studies and Molecular Medicine, University of Leicester,
Robert Kilpatrick Clinical Sciences Building, Leicester Royal Inrmary, Leicester,
Leicestershire, LE2 7LX UnitedKingdom. Tel.: +44 116 252 5826;fax: + 44 116252 5824.
E-mail address: jck4@le.ac.uk (J.C. Konje).
0009-8981/$ see front matter © 2010 Elsevier B.V. All rights reserved.
doi:10.1016/j.cca.2010.03.012
Contents lists available at ScienceDirect
Clinica Chimica Acta
journal homepage: www.elsevier.com/locate/clinchim
Author's personal copy
delivery of low birth weight babies [8]. The fact that marijuana has
severe consequences on pregnancy, would suggest that the endoge-
nous ligands of the receptors are involved in the modulation of
reproduction. Recently, comprehensive studies by us and others have
indicated that endocannabinoids (especially the prototypical endo-
cannabinoid, N-acyl arachidonoylethanolamide [anandamide, AEA];
Fig. 1), are actively involved in the process of reproduction and
pregnancy. This review will examine the role of cannabinoids in
pregnancy, from gametogenesis through to birth with special focus
upon the effects of endocannabinoids and the endocannabinoid
system on these processes.
2. The endocannabinoid system (ECS)
The isolation of a specic cannabinoid receptor in the porcine brain
(termed CB
1
), that is responsible for mediating the pharmacological
effects of Δ
9
-THC [9], initiated the search for the endogenous ligands.
The rst ligand identied that could imitate the pharmacology of Δ
9
-
THC at the CB
1
receptor was anandamide [10] (Fig. 1). Subsequently,
isolation of a second cannabinoid receptor (CB
2
)associatedwiththe
immune system [11] preceded the identication of a second major
endocannabinoid, 2-arachidonoylglycerol (2-AG) (Fig. 1).Both AEA and
2-AG have afnity for CB
1
and CB
2
[12], although with differentavidities.
More recently, other bioactive endocannabinoids have been identied
including O-arachidonoylethanolamine (virodhamine) [13],N-arachi-
donoyl dopamine [14] and most recently N-acyl taurines, such as N-
arachidonoyl taurine [15]. In addition, several endocannabinoid
congeners have been identied which have greatly reduced afnity
for CB
1
and CB
2
yet elicit endocannabinoid-like activities. These include
oleoylethanolamide (OEA) and palmitoylethanolamide (PEA) (Fig. 1),
which are thought to elicit their endocannabinoid-like activities
through the so-called entourage effect, by either inhibition of
endocannabinoid catabolism [16], or reducing cellular uptake of AEA
thus leading to increased AEA concentrations [17,18]. Of all the possible
endocannabinoids, 2-AG and AEA are the most frequently studied, with
quantied levels in human tissues for 2-AG being consistently 10100
times higher than those of AEA [1921]. AEA, the entourage lipids OEA
and PEA, and to a lesser extent 2-AG have been quantied in human
biological uids including plasma and serum, seminal plasma, cerebro-
spinal uid, follicular uid, oviductal uid, amniotic uid, ovarian cyst
uid, milk and peritoneal uid, but seem to be absent from urine and
saliva [2228].
The ligands only comprise part of the endocannabinoid system;
the enzymes concerned with endocannabinoid synthesis and degra-
dation, a putative membrane transport system and the receptors
through which they elicit their physiological effects complete the
various components (Fig. 2). Endocannabinoids are synthesised on
demand from membrane phospholipids precursors and are not stored
[29].
In most tissues, the main rate-limiting step in AEA synthesis is the
conversion of N-arachidonoyl phosphatidylethanolamine (NAPE) into
AEA by a specic phospholipase D enzyme (NAPE-PLD) [30,31].
However, the continued synthesis of AEA in NAPE-PLD knockout mice
highlighted alternative pathways for the formation of AEA [32]. These
pathways involve either hydrolysis of NAPE by phospholipase C to
yield phosphoAEA which is dephosphorylated by a protein tyrosine
phosphatase [33] or sequential acyl hydrolysis of NAPE and lysoNAPE
by abhydrolase domain containing 4 (Abh4) to form glycerophospho-
NAPE, which is then hydrolysed to AEA by a phosphodiesterase [34].
The synthesis of other endocannabinoids, especially the monoacyl
glycerol, 2-AG, is less well-dened, but also thought to occur on
demand. For example, 2-AG is produced via diacylglycerol from
arachidonic acid-rich phospholipid precursors by the sequential
actions of phospholipase C and diacylglycerol lipase [35].
Degradation of AEA and 2-AG to free arachidonic acid is primarily
by the serine hydrolases fatty acid amide hydrolase (FAAH) [36] and
monoacylglycerol lipase (MAGL) [3739], respectively. In addition to
hydrolysis, AEA has been shown to be metabolised by cyclooxygenase
2 to prostamides, ethanolamide derivatives of prostaglandins [40],
and by the cytochrome P450 isozyme CYP2D6 to several epoxide
metabolites [41], which are thought to have their own physiological
actions. Endocannabinoid degradation requires transport of the
endocannabinoids into cells and how these cross the plasma
membrane is still open to debate [42,43]. Current theories include
carrier protein-mediated transport [44], diffusion driven by metab-
olism by FAAH [45], diffusion driven by compartmentalization of
intracellular AEA [46] and endocytosis [47].
Endocannabinoids predominantly exert their effects through bind-
ing to the cannabinoid receptors (CB
1,
CB
2
and GPR55). These seven-
transmembrane G-protein coupled receptors differ signicantly in their
distributions. Although CB
1
was initially localized in the nervous system
and CB
2
in the spleen,these receptors are now knownto be more widely
distributed throughoutthe body [4858]. These receptors seem to signal
ligand occupancy by coupling inhibitory Gα
i/o
proteins which inhibit
adenylate cyclase activity, thus reducing intracellular cAMP concentra-
tions [59]. In addition, agonist binding to CB receptors also leads to
activation of MAP kinases, which may partly explain the extensive
pathophysiological roles ascribed to endocannabinoids such as cell cycle
progression, apoptosis, cell differentiation and oncogenesis [60,61].
Stimulation of CB receptors can also cause rapid, transient elevation of
intracellular Ca
2+
concentrations via phospholipase C, the inhibition of
voltage-gated Ca
2+
channelsand activation of K
+
channels[31].Butthis
may not be the only effects endocannabinoids exert, as they bind to and
activate other types of receptors. For example, the transient receptor
potential vanilloid type 1 (TRPV1), a ligand-gated non-selective cation
channel that can be activated by a number of stimuli including heat and
pH and is involved in the transmission and modulation of pain, also
binds endocannabinoids, especially AEA and N-arachidonoyl dopamine
[62]. Similarly, both endocannabinoids and entourage compounds bind
to and activate the G-protein coupled receptor GPR55, which is
considered a putative cannabinoid receptor [63] because compounds
Fig. 1. Chemical structures of the most psychoactive component of marijuana (Δ
9
-
tetrahydrocannabinol, the major endocannabinoids, and the entourage compounds.
922 A.H. Taylor et al. / Clinica Chimica Acta 411 (2010) 921930
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other than endocannabinoids also bind to this receptor. The so-called,
entourage endocannabinoids,OEA and PEA, also bind to peroxisome
proliferator-activated receptors (PPAR). In particular, they activate the
PPARαisoform to modulate cell differentiation, lipid metabolism and
regulate food intake [64].
3. Endocannabinoids and gametogenesis
3.1. Oogenesis
The recent demonstration of the endocannabinoid system in the
human ovary has attracted a growing interest into the exact role of the
endocannabinoids in mammalian folliculogenesis [65].CB
1
and CB
2
receptors have been localized in oocytes at all stages of maturation
and the AEA synthetic enzyme NAPE-PLD and the hydrolytic enzyme
FAAH have also been localized in the growing secondary and tertiary
follicles and corpora lutea and albicantes [65]. In addition, AEA has
been quantied in human follicular uid [28] and its concentration
found to be strongly correlated to oocyte maturation and quality in
women undergoing infertility treatment [66]. The exact molecular
mechanism by which endocannabinoids modulate ovarian folliculo-
genesis remains speculative, but it is believed that they act through a
combination of central and peripheral mechanisms related to the
actions of gonadotrophins. Evidence for this comes from the now
dated studies showing that Δ
9
-THC exerted an adverse effect on
oocyte development and ovulation [67] in which Δ
9
-THC was shown
to interrupt the hypothalamic-pituitary-ovarian axis [6870] and
suppress plasma follicle-stimulating hormone (FSH) and pre-ovula-
tory luteinizing hormone (LH) surge [7173], causing an ovulation in
humans [74],rats[75], rabbits and rhesus monkeys [76].The
reduction in gonadotrophin production results in poor quality oocytes
being retrieved during in-vitro fertilization [77]. Since it is known that
cAMP accumulation is required for normal follicular development,
maturation and ovulation [78] and Δ
9
-THC acting through the CB
1
receptor normally has an inhibitory effect on cAMP levels, then the
inhibitory effect of Δ
9
-THC on folliculogenesis may be mediated via its
inhibition of adenylate cyclase, thereby reducing tissue concentra-
tions of cAMP, as has recently been shown in cultured rat granulosa
cells [79,80].
Other evidence also suggests that the endocannabinoid system,
which regulates energy balance by modulating appetite, food intake
and glucose metabolism [8184] could interact with gametogenesis
through nutritional control. Obesity is commonly associated with
menstrual irregularities, chronic oligo-anovulation and infertility [85]
and regular ovulation is restored after simple management strategies
aimed at weight reduction leading to improved natural conception
[86,87]. Simultaneously, insulin resistance is a cardinal feature of
obesity-associated ovulatory dysfunction in the presence or absence
of polycystic ovary syndrome [88] and androgen hypersecretion, as a
result of hyperinsulinaemia, leads to altered ovarian physiology and
ovulatory dysfunction [89]. Activation of CB
1
receptors in pancreatic
beta cells by AEA induces glucose intolerance, insulin hypersecretion
and insulin resistance in rats [90,91], suggesting that activation of the
ECS in the pancreas might explain some forms of female infertility.
The use of CB
1
antagonists (such as Accomplia®; rimonabant) for the
treatment of obesity, and its metabolic consequences, has been shown
to be very effective in reducing body weight or controlling leptin
concentrations [92,93] although with some serious adverse side
effects that have led to its discontinuance [94]. It is therefore clear that
oogenesis could be disrupted through perturbations of the complex
interplay that exists between the endocannabinoids, leptin produc-
tion and obesity. For example, sustained stimulation of CB
1
in the
hypothalamus by AEA or 2-AG has been observed in leptin-decient
mice, with elevation of hypothalamic levels of AEA and 2-AG which
Fig. 2. Components of the endocannabinoid system: The ligands 2-acylglycerol (2AG) and anandamide (AEA) are show as blue and red spheres respectively. The putative, but not yet
identied endocannabinoid membrane transporter (EMT) is shown as a bi-directional regulator. The enzymes responsible for the synthesis and degradation of AEA (N-acyl
transferase (NAT) and N-arachidonoyl phosphatidylethanolamine phospholipase D (NAPE-PLD) and fatty acid amide hydrolase (FAAH-1/2)) and of 2-AG (Di-acyl glycerol lipase
(DAG) and monoacyl glycerol lipase (MAGL)), convert these two endocannabinoids into arachidonic acid (AA; green spheres) and ethanolamine (silver spheres) or AA and glycerol
(yellow spheres), respectively. Cyclooxygenase 2 (COX-2) converts AEA into prostaglandin ethanolamines. Glycerol is acted upon by phospholipase C to produce diacylglycerols that
are reconverted into 2AG by DAGL. AEA once internalised may interact with the cytoplasmic domain of the transient vanilloid receptor type 1 (TRPV1) channel protein to alter
intracellular calcium levels. To aid clarity the possible signalling pathways of cannabinoid receptor (CBR) signalling have been omitted.
923A.H. Taylor et al. / Clinica Chimica Acta 411 (2010) 921930
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was reversed after leptin treatment [95]. Since FAAH is regulated in
some tissues by leptin [96101] then these data are highly suggestive
of regulatory networks between leptin, FAAH and endocannabinoids
existing in the hypothalamus that are responsible for the exquisite
control of folliculogenesis. Nevertheless, this area of research is at an
early stage and further work is required to understand the complex
network of central and peripheral actions of endocannabinoids with
regard to ovarian function.
3.2. Spermatogenesis
Accumulating evidence suggest that endocannabinoids may also
modulate male reproductive function [102]. The rst indication for
this comes from studies of chronic marijuana users, where transient
adverse reproductive toxicity occurs and Δ
9
-THC directly inhibits
sperm motility and viability in vitro [103]. In experimental animals,
Δ
9
-THC induces impotence in rats [104], decreases rat testicular
weight [105,106], decreases testosterone production which impairs
spermatogenesis and sperm motility [107,108] and in mice similarly
reduces the production of spermatozoa that tend to have abnormal
morphology [109].
The exact mechanism by which Δ
9
-THC inhibits spermatogenesis
remains speculative but may involve a complex cross-talk between
FSH, LH and cannabinoids [110].Δ
9
-THC inhibits the release of FSH
and LH from the pituitary gland and consequently [111114] thereby
inhibits spermatogenesis and testosterone production. By contrast, 2-
AG and to a greater extent AEA, mimic Δ
9
-THC by inhibiting FSH, LH
and prolactin release from the pituitary gland [115120] suggesting a
common mechanism of action in both the hypothalamus and
pituitary. CB
1
and CB
2
receptors have been demonstrated in
gonadotrophs within the hypothalamus [121] whilst the anterior
pituitary gland expresses CB
1
and TRPV1 receptors [115117]
supporting a direct endocannabinoid effect on gonadotrophin
production in the male, similar to that observed in the female.
There is also some evidence of a direct effect of cannabinoids on
the testis [122126]. Recent interest in the localization of the
endocannabinoid system within male reproductive tissues and
measurement of endocannabinoids in uids of several mammalian
species has been the subject of intense interest, as well as provoking
controversy. For example, spermatogenic output depends on a
balance between cell proliferation and apoptosis and although,
functional CB
1
has been identied in rodent Sertoli and Leydig cells,
suggesting that endocannabinoids are produced by the testis and may
play a crucial role in spermatogenesis and testosterone production
[127,128], the predominant cannabinoid receptor found in mamma-
lian Sertoli cells is CB
2
, which has anti-apoptotic effects and counter-
acts the pro-apoptotic effects that occur through the TRPV1 receptor
(also present in the Sertoli cell), thereby protecting the germinal
epithelium and germ cells from apoptosis, promoting meiotic
progression [127] and ensuring increased spermatogenic output
[126,128131]. The recent demonstration and cloning of testis-
specic functional isoforms of the CB
1
and CB
2
receptor that have
differential ligand binding properties [132,133] may help to resolve
some of these controversies.
4. Endocannabinoids, early embryo-blastocyst development and
oviductal transport
The oviduct is central to the process of human reproduction and
plays a critical role in oocyte fertilization and pre-implantation
embryo development during its passage to the uterus [134]. Events
occurring during oviductal transport must be co-ordinated such that
the blastocyst is ready for implantation when it arrives in the uterine
cavity and the latter is ready to receive the blastocyst, usually 6 days
post-fertilization [134]. These critical events are synchronised and
regulated by several endocrine, paracrine and autocrine factors
[96,135] of which the endocannabinoid system has been identied
as one of the key hormonecytokine/receptor signalling systems
[79,136140].
4.1. Endocannabinoids and early embryo development
In the mouse, FAAH is expressed at the 2-cell stage of the early pre-
implantation embryo [136,141,142] while NAPE-PLD, CB
1
,CB
2
and are
expressed at later stages. As development proceeds to the blastocyst
stage FAAH levels are signicantly up regulated. In homozygous FAAH
knockout mice, embryos recovered from oviducts on day 3 of
pregnancy showed delayed and asynchronous development com-
pared to wild type mice [142] suggesting that low levels of AEA are
benecial and high levels detrimental to blastocyst development
[136,141]. Furthermore, in vitro culture of mouse blastocysts with AEA
indicated that 7nM concentrations of AEA allowed blastocyst
hatching, and 28nM concentrations prevented blastocyst hatching
and induced apoptosis [138]. These data suggest that AEA synthesis by
NAPE-PLD and its degradation by FAAH in embryos and within
oviducts has to be well synchronised such that a locally appropriate
anandamide toneis created to permit normal embryo development
and oviductal transport [138]. Furthermore, studies with CB receptor
knockout mice suggest that CB receptor signalling is important in
oviductal transport and blastocyst development. For example, on day
4 of pregnancy, only 6070% of CB
1
knockout, CB
2
knockout and CB
1
/
CB
2
double knockout embryos were blastocysts, whereas most of the
wild type embryos were blastocysts [143]. Data on the effects of
endocannabinoids on human embryo/blastocyst development are,
however, lacking. Nevertheless, the observation that low plasma
levels of AEA and high levels of FAAH in peripheral mononuclear cells
observed during the putative implantation windowin humans
[66,144,145] suggests that the ECS also plays a similar role in human
embryo development.
4.2. Endocannabinoids and transport through the Oviduct
CB
1
signalling has been shown to play a crucial role in oviductal
transport of embryos. Studies on CB
1
,CB
2
and CB
1
/CB
2
double
knockout mice showed that only those lacking CB
1
protein had a
large number of embryos in the morula or blastocyst stage retained in
their oviducts on day 4 of pregnancy whereas CB2 knockout and wild
type mice had none retained, providing evidence that oviductal
transport is delayed in CB
1
decient mice [140,143] and that CB
2
is
probably not involved in oviductal transport. This was conrmed by
studies using wild type mice treated with the CB
1
receptor antagonist
(SR141716) with retention of embryos in the oviducts but not when
the CB
2
antagonist (SR144528) was used [140].
The mechanism whereby endocannabinoids might affect normal
transport is currently unclear. Transport of embryos through the
oviduct is aided by a peristaltic wave of oviductal smooth muscle
movement regulated by the sympathetic nervous system [146]. The
demonstration of CB
1
co-localization with adrenergic receptors in
rodent oviduct, suggests that endocannabinoids could affect cate-
cholamine function [140]. Indeed, excess sympathetic stimulation has
been shown to lead to increased smooth muscle contraction and
impaired embryo transport at the isthmus-uterine junction [140]. The
overstimulation of the oviductal smooth muscle with potassium
chloride led to a 33% overow of radio-labelled noradrenaline into the
oviducts of CB
1
knockout mice, an effect that was signicantly higher
compared to that in CB
2
knockout and wild-type oviducts [140],
suggesting that CB
1
signalling negatively regulates neuronal nor-
adrenaline release to directly modulate tubal motility. The authors
suggest that aberrations in the endocannabinoid system could
therefore form a mechanistic basis for ectopic pregnancy, where
there is no obvious tubal damage.
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In humans, CB
1
has been localized in the Fallopian tube smooth
muscle wall and CB
1
transcript levels were found to be signicantly
lower in the Fallopian tubes of patients with ectopic pregnancy
compared to non pregnant controls [147]. Furthermore, that study
examined 2 polymorphisms of the CB
1
gene and found a possible
association between the 1359G/A polymorphism and ectopic preg-
nancy. The small numbers involved make a denitive association
difcult, but if larger studies support a possible genetic predisposition
to ectopic pregnancy due to CB
1
dysfunction, then genetic screening of
women could lead to the identication or directed therapies for
women at risk of this life-threatening condition [147].
5. Endocannabinoids and implantation
It is now acknowledged that the endocannabinoid system is
critical in the process of implantation [66,79,110,139,147154].
Successful implantation depends on an accurate interaction between
the endometrium and the implanting blastocyst. Firstly, it requires a
receptive uterine environment and secondly, an activated blastocyst,
capable of invasion and subsequent implantation [155157].
Implantation occurs about six to seven days after fertilization and
in order for the process to begin, the blastocyst needs to emerge from
the zona pellucida (blastocyst hatching) thereby exposing a
trophoblast capable of invasion [152,155], followed by apposition
with the endometrium, adhesion and nally, trophoblast invasion
[155]. Animal studies have established that AEA regulates the
windowduring which the uterus is receptive [29,79,139,157].As
the blastocyst enters the uterus, it is differentially affected by local
physiological AEA concentrations. At low concentrations, AEA acti-
vates ERK signalling in dormant mouse blastocysts, whereas high
levels of AEA inhibit calcium mobilisation [141]. Furthermore, low
AEA levels promote blastocyst growth. Studies by Paria et al.have
shown that trophoblasts exposed to low levels of endocannabinoids
show accelerated growth and differentiation as well as increased rates
of zona hatching [137,158,159]. These effects are mediated via CB
1
receptors. Trophoblasts also expresses CB
2
receptors whose function
in this tissue is currently unknown [158,160]. At the same time, lower
AEA levels and elevated levels of FAAH occur in the uterus at
implantation sites [159] when compared to the inter-implantation
sites, which have much higher levels of AEA and lower levels of FAAH.
There is indirect evidence that during apposition, the mouse
blastocyst produces an unidentied lipid [161] that induces the
changes in FAAH and NAPE-PLD enzyme expression conducive for
attachment and initial invasion and thus the changes in localized AEA
concentrations at the implantation site [157,162]. Evidence that local
AEA concentrations are critical to implantation comes from studies
demonstrating that embryos exposed to high levels of AEA in the
uterus suffer embryotoxicity, reduced trophoblast proliferation and
implantation failure [158,160,162].
Similarly, women in an in vitro fertilization programme who have
successful implantation, have low serum AEA levels associated with
elevated levels of FAAH in their peripheral lymphocytes at 6 weeks
gestation [163,164]. The process is likely being controlled by the
expression and activity of lymphocytic FAAH. Indeed, it has been
suggested that the decreased activity and expression of FAAH in
peripheral lymphocytes could be used as an early marker for rst
trimester miscarriage [164]. These data are mirrored by the studies of
Trabucco et al.who showed that placental tissue from women
following spontaneous miscarriage had very low levels of FAAH and
increased expression of the CB
1
receptor [151]. These ndings are
corroborated by observations that higher plasma AEA levels in women
with threatened miscarriage are associated with a subsequent
spontaneous miscarriage [165].
Plasma AEA levels uctuate during the menstrual cycle
[23,24,144,145]. Recently our group showed that plasma AEA levels
declined during the implantation window in women with a successful
pregnancy after IVF treatment and in women with normal menstrual
cycles [66,144,145]. During this period, FAAH has been shown to have
the highest activity, whereas there is no change in CB
1
binding, or
AMT and NAPE-PLD activity [145]. Interestingly, FAAH and proges-
terone show the same uctuations during the menstrual cycle [145].
This is conrmed by the nding that progesterone up-regulates the
FAAH gene [166,167], but contradicted by the nding that serum
progesterone was not correlated with plasma AEA levels during the
menstrual cycle or in women with positive implantation in an IVF
treatment programme [144]. There were, however, positive correla-
tions between the levels of 17β-estradiol, FSH and LH, suggesting that
implantation could be regulated by changes in the levels of these
hormones and their inuence of AEA levels in the uterus [144].
Research so far indicates that the endocannabinoid system is an
important regulator for implantation and successful pregnancy.
Endocannabinoid signalling is under tight control and any disruption
of this balance may lead to reproductive failure. The delicate
intricacies related to how endocannabinoids regulate implantation
and whether this is a direct inuence or via T lymphocyte function
remains to be elucidated.
6. Endocannabinoids in early pregnancy
Although Δ
9
-THC is known to cross the placenta and affect human
pregnancies, it was not known if the effect was via actions on
trophoblast activity. Recently, however, we demonstrated that Δ
9
-
THC acts directly on the human BeWo cytotrophoblast cell line to
inhibit cell growth and the transcription of genes involved in growth
and apoptosis [168], via the relatively unstudied CB
2
receptor [169].
Indeed, using human cell line models of the 1st and 3rd trimester
trophoblast and the pregnant rat model, we have shown that in
human cells AEA inhibits cell proliferation via the CB
2
receptor by the
differential modulation of histone deactylase 3 (HDAC3) expres-
sion [170]. AEA at concentrations in excess of 15 μM stimulate HDAC3
expression in rst trimester trophoblast cells and inhibit HDAC3
expression in third trimester trophoblasts. These data support the
observations that Δ
9
-THC has differential effects on the rst and third
trimester trophoblast [171,172], and that differential CB receptor
expression in the rat placenta occurs during gestation [173]. Indeed,
treatment of rats with varying doses of AEA in excess of 10 μM induces
trophoblast cell apoptosis via the CB
1
receptor [173]. These data
suggest that there may be subtle differences in the actions of exo- and
endo-cannabinoids in the placenta in the human and rat and that a
complex interplay between AEA concentrations and implantation of
the blastocyst and subsequent placentation is important during early
pregnancy. Indeed, in studies of IVF-ET women, where the introduc-
tion of the embryo can precisely be timed and implantation can be
closely followed [66], plasma AEA levels initially increase, presumably
to aid blastocyst hatching and implantation, but then subsequently
fall, presumably to prevent trophoblast apoptosis and so aid retention
of the developing embryo. It therefore seems that a critical balance in
endocannabinoid levels, which has been shown to be vital to
successful pregnancy, is required for early trophoblast development
and subsequently that of the placenta.
7. Endocannabinoids in the maintenance of pregnancy and labour
The fact that the use of marijuana is associated with preterm
labour, prolonged pregnancy, fetal growth restriction, stillbirth and
placental abruption suggest that the endocannabinoid system is
involved in late pregnancy and labour. We have quantied AEA levels
during pregnancy and found that the levels are low in the rst
trimester (similar to the levels measured in the luteal phase of the
menstrual cycle) and are maintained at these low levels until the
beginning of the 3rd trimester when they start to rise. They then rise
by almost 2- to 4-fold with the onset of labour (Table 1;[23,24,174]).
925A.H. Taylor et al. / Clinica Chimica Acta 411 (2010) 921930
Author's personal copy
In the rat, CB1, CB
2
and TRPV1 expression uctuates in the decidua
in a predictable pattern from a high level on days 1012 of pregnancy
to a lower level towards the end of pregnancy [173]. Simultaneously,
plasma AEA levels are not signicantly altered, but local tissue levels
of AEA and PEA, but not OEA are increased at two points through
pregnancy; on day 10, when the placenta and decidua regress to make
room for the developing embryo and on day 19 when the activation of
the myometrium is initiated in preparation for parturition on day 21
[175]. These observations suggest that local production of endocan-
nabinoids may be associated with the apoptosis-induced regression of
the decidua in the rat and the preparation of parturition events. These
rodent data together with the aforementioned increase of plasma AEA
levels indicated in the 3rd trimester of pregnancy suggest that plasma
AEA levels could be elevated in women with incipient preterm labour,
preterm premature rupture of membranes or cervical insufciency,
obstetrical problems that are almost impossible to predict. Data
emerging from our laboratory support this suggestion, as women at
high risk of preterm birth, or presenting with abdominal pain
consistent with the Braxton Hicks myometrial contractions indicative
of impending preterm labour, that subsequently went on to deliver
preterm have higher levels of plasma AEA than those that continue to
term [174]. Indeed, those who delivered quicker (less than 6 days)
have even higher plasma AEA levels than those that took longer to
deliver but still delivered prematurely. These data are consistent with
studies performed in the CB
1
knockout mouse, in which delivery
occurs preterm (i.e. 1 day early) and could be prevented by changing
cortisol and progesterone levels, suggesting that the entire endocan-
nabinoid system, (i.e. not only endocannabinoid levels), is involved in
the timing of parturition [176]. The alternative observation that other
endocannabinoids are present in the uterus at this important time
during pregnancy, and the apparent increase in plasma AEA levels in
women in labour, even if they had been induced [177], suggest that
the entourage effectis in action and that FAAH activity is maximised
at this point and that FAAH is dealing with the other endocannabi-
noids that exist at higher levels than AEA [1921,28]. This might
explain the apparent discrepancy between increasing lymphocyte
numbers during the latter stages of pregnancy [178] and the increased
levels of plasma AEA during labour. The obvious extension of these
observations is that CB
1
receptor mutations [147] and CB
1
signalling
or FAAH mutations or activities may be important in the timing of
birth and hence an important area of study for perinatologists. In our
opinion, this understudied area of endocannabinoid research needs
urgent investigation.
It is not clear what elevated plasma AEA levels mean in relation to
parturition. Three main tissue-specic events occur during the
parturition process; myometrial activation, cervical ripening and
fetal membrane rupture. The actions of endocannabinoids in
myometrial activation, cervical ripening and fetal membrane are
unclear due to the paucity of data. Studies on oxytocin-stimulated
myometrial muscle strips demonstrated a relaxant effect of both Δ
9
-
THC and AEA through CB
1
[179]. Recently, we demonstrated that AEA
increased the production of phosphorylated ERK in primary human
myometrial cells and a human myometrial cell line, without changes
in intracellular calcium levels [180]. Such changes in the myometrial
cell should result in myometrial relaxation as was suggested by
Dennedy and colleagues [179]. The data suggest that the myometrium
is a possible endocannabinoid target during the latter stages of
pregnancy. The fetal membranes and placenta at term are also obvious
targets as these tissues express the cannabinoid receptors [181] and
contain signicant amounts of AEA (Table 1).
8. Feto-placental changes in endocannabinoid levels
Levels of FAAH in both the human placenta and the maternal
circulation increase towards the end of the rst trimester of
pregnancy, before declining by the early second trimester [163,182].
Furthermore, high levels of FAAH have been observed in the villous
cytotrophoblast [183]. Its expression in the syncytiotrophoblast
suggest that FAAH in these cells help prevent the transfer of AEA
from maternal blood [182], which, to some degree, has been
suggested as a protective mechanism for the developing fetus [182].
Previous studies have suggested that circulating FAAH and AEA levels
may be critical to the outcome of early pregnancy [163,164]. As stated
earlier, decreased expression and activity of FAAH in peripheral blood
lymphocytes maybe an early marker of spontaneous miscarriage
[163]. The importance of FAAH in early placental development has
been supported by a recent study that demonstrated the expression of
FAAH and CB
2
(rather than CB
1
) receptors in the human rst trimester
placenta. This study also provided evidence that the endocannabinoid
regulation within placental tissue is independent of the maternal
immune system [182]. We recently demonstrated the presence of
both CB
1
and CB
2
receptors and FAAH immunoreactive protein in rst
trimester trophoblast which was supported by the presence of
transcripts for these genes in the tissue [183]. The apparent
discrepancy between these two studies could be related to the
antibodies used, as has been documented [184]. Nevertheless,
cannabinoid receptors are expressed by placental tissue from the
early rst trimester right through to term [181183]. Interestingly,
levels of AEA fall progressively during pregnancy [23,24], supporting
other evidence that low systemic levels are required for normal
pregnancy progression [163]. Therefore, exposure to the exocanna-
binoid Δ
9
-THC could lead to inappropriate activation of the CB
mediated pathways in the placental trophoblast.
9. Possible immunocannabinoidsystem actions in pregnancy
Although the involvement of the endocannabinoid system in
reproduction may be direct, it may also be through modulation of the
immune system. Despite a great interest in the role of cannabinoids
within the immune system [57,185189], the possible relationship
between endocannabinoids and immunity, especially in relation to
reproductive events, remains to be clearly determined. For successful
implantation and maintenance of pregnancy, the embryonic/fetal
allograft [190,191] has to be tolerated by the maternal immune
Table 1
Anandamide concentrations in different human biomatrices related to reproduction
and the reproductive tract.
Biomatrix/Tissue AEA (nM)
1
Ref
Seminal Plasma
3
12.1 [28]
Follicular uid Immature oocyte 0.361.96 [65]
Mature oocyte 0.252.78 [65]
Oviductal uid
3
10.7 [28]
Endometrium ?
Myometrium ?
Cervix ?
Vagina ?
Hypothalamus
2
86.387.6 [21]
Pituitary
2
?
Plasma (menstrual cycle) Follicular Phase 1.451.68 [23,24,143]
Ovulation 1.264.04 [143]
Luteal Phase 0.770.87 [23,24,143]
Plasma (pregnancy) 1st trimester 0.890.91 [23,24]
2nd trimester 0.440.91 [23,24]
3rd trimester 0.441.14 [23,24]
Term not in labour 0.681.27 [23,24]
Term in active labour 2.302.50 [23,24]
Placenta
2
0.984.54 [215]
Fetal Membranes
2
0.422.83 [215]
Amniotic uid 0.010.18 [26,215]
Umbilical artery plasma 0.321.53 [26,215]
Umbilical vein plasma 0.381.84 [26,215]
Maternal Plasma at term 0.591.91 [26,215]
1
AEA = anandamide;
2
in pmol/g wet weight;
3
only a single human study performed;
? = data unknown.
926 A.H. Taylor et al. / Clinica Chimica Acta 411 (2010) 921930
Author's personal copy
system through mechanisms that might involve the endocannabinoid
system [169]. For example, arachidonic acid (AA), a local immuno-
modulatory molecule [192], is released from cultured mouse
macrophage cells by Δ
9
-THC and AEA through a CB
2
-dependent
mechanism and by nitric oxide (without raising AEA levels) indicating
that both CB receptor-dependent and -independent mechanisms are
involved in this possible immunomodulatory pathway [193,194].
Maccarrone et al. [195] also demonstrated that bacterial lipopolysac-
charide down regulates FAAH expression in human peripheral
lymphyocytes, an effect that was not inuence by AEA-induced
activation of the cannabinoid receptors, whilst mast cells absorb AEA
in a saturable process that activates intracellular FAAH and AEA
degradation [196]. Additionally, the physiological protection of the
fetus from rejection is believed to be dependent on a type 2 T-helper
(Th2) cell immune response at the fetalmaternal interface [197]. The
initial state of pregnancy is nevertheless characterised by immuno-
suppression after an early inammatory state during implantation
[198]. Indeed, the demonstration that monocytes [199] and leuko-
cyte/macrophages [200,201] numbers increase towards the end of
pregnancy suggest an inammationimmunosuppressioninamma-
tion phenotype during pregnancy [202]. The production of plasma
AEA mimics this inammationimmunosuppressioninammation
pattern during implantation, the 2nd and 3rd trimesters of pregnancy
and just before labour [23,24,66]. Such data suggest that immune cells
may take part in regulating the peripheral endocannabinoid system or
local endocannabinoid homeostasis and vice versa.
Many studies have demonstrated that, in the main, endocannabi-
noids suppress the production of inammatory cytokines in innate
and adaptive immune responses both in animal models and in human
cell cultures [186,203,204]. Immunosuppression is one of the most
important steps involved in early pregnancy [205] and evidence of a
reduction in interleukin-1β, -10, and -12 levels in animal models of
inammatory disease [206208] suggests that similar reductions in
the levels of these cytokines in reproductive tissues may occur in
response to local endocannabinoid production, although this is yet to
be directly determined. However, cannabinoids have also been
demonstrated to increase the production of IL-1, -4, -6 and -10
cytokines either in the presence of bacteria [204,207,209] or alone
[210,211], suggesting that in vivo, cannabinoids may either suppress
or enhance the production of pro-inammatory agents, depending on
either the nature of the pro-inammatory stimulus or the type of
cannabinoid present. These changes may arise from the predominant
effect of cannabinoids on helper T-cell (Th) biasing [186,203] whereby
Th-1 cell activity is suppressed and Th-2 cell activity is increased
[208,212]. This may change the levels of leukaemia inhibitory factor,
which is known to be essential for successful invasion and migration
during placental development [213,214].
Taken together these data suggest that endocannabinoids have a
key role, not only on the cells and tissues of the fetalmaternal
interface but also on their interaction with the innate and adaptive
immune system. This under-explored area of possible interaction
between the immunocannabinoid system [187] and reproduction is
likely to be a focus of intense research in the near future.
10. Conclusions
The endocannabinoid system (ECS) is involved in various aspects
of human reproduction. It appears to be an important cog in the
complex mechanisms (wheel) of gametogenesis. With respect to
spermatogenesis, 2-AG appears to promote meiotic progression in the
germ cell by activating type-2 cannabinoid receptors [126]. Levels of
AEA also appear to be related to follicular size and quality of the ova
produced during oogenesis. This system is involved in the regulation
of fertilization through actions that result in spermatozoa activation
and thereafter inuence hatching of the embryo, blastocyst matura-
tion, oviductal transport, implantation and early pregnancy mainte-
nance. Changes in plasma levels of some of the cannabinoid ligands
(e.g. anandamide) provide evidence for a tight regulation of early
pregnancy by this group of compounds. Changes in pregnancy and
labour also suggest an involvement in labour onset and maintenance.
Although this review has highlighted areas where the evidence is
coherent and strong, so many questions remain unanswered and only
through collaborative and international studies will these be
adequately addressed. Such collaboration may initially focus on the
changes of the cannabinoid receptors, FAAH, NAPE-PLD, and AEA
which have been thoroughly investigated in animals and partly in
humans, to determine the translational potential of the ECS in
establishing and maintaining a healthy pregnancy.
Conicts of interest
The authors afrm that there are no conicts of interest.
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... Here, they play roles that include oocyte production [45,59,[96][97][98], oviductal transport [98][99][100], and blastocyst maturity and implantation [101], as well as in preparing the endometrium for implantation [47,[102][103][104][105]. When the ECS is dysfunctional or interfered with by, for example, cannabinoids [106,107], human fertility may be impaired (reviewed in [12,101,108,109]) and there may be associated reproductivetissue-dependent pathologies, such as endometriosis, miscarriage, ectopic pregnancy, or pre-eclampsia [51,101,108,[110][111][112]. Two recent reviews [12,38] on the ECS in the female reproductive tract summarise what is known on this topic, but crucially, these omit some important details on the main ECS components in gynaecological cancers, which we hope to address in this review (see Table 1). ...
... Here, they play roles that include oocyte production [45,59,[96][97][98], oviductal transport [98][99][100], and blastocyst maturity and implantation [101], as well as in preparing the endometrium for implantation [47,[102][103][104][105]. When the ECS is dysfunctional or interfered with by, for example, cannabinoids [106,107], human fertility may be impaired (reviewed in [12,101,108,109]) and there may be associated reproductivetissue-dependent pathologies, such as endometriosis, miscarriage, ectopic pregnancy, or pre-eclampsia [51,101,108,[110][111][112]. Two recent reviews [12,38] on the ECS in the female reproductive tract summarise what is known on this topic, but crucially, these omit some important details on the main ECS components in gynaecological cancers, which we hope to address in this review (see Table 1). ...
... In this context, the ECS has been studied the most in the human ovary [45,[59][60][61][63][64][65]113], cervix [80,[84][85][86][87], and endometrium [47,50,55,[68][69][70][72][73][74][75]81], the most common gynaecological cancers ( [95,114,115]); however, other female cancers have not been studied, as shown in Figure 2. Although the presence and actions of the most commonly studied endogenous ligands (AEA, 2-AG, OEA, and PEA) in human reproductive tissues have been demonstrated, the presence and actions of others, such as SEA, virodhamine, stearamide, and monoolein [12,30,38,45,49,51,54,57,69,78,95,101,105,108,112,[116][117][118][119][120][121][122][123] have not. Studies on receptor expression and function are few, and although there are some published studies on NAPE-PLD and FAAH expression and action in the female reproductive tract, many other (endo)cannabinoid metabolising enzymes have yet to be investigated, either in the normal female reproductive tract or in their related neoplasms (Figures 1 and 2; Table 1). ...
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Gynaecological cancers can be primary neoplasms, originating either from the reproductive tract or the products of conception, or secondary neoplasms, representative of metastatic disease. For some of these cancers, the exact causes are unknown; however, it is recognised that the precise aetiopathogeneses for most are multifactorial and include exogenous (such as diet) and endogenous factors (such as genetic predisposition), which mutually interact in a complex manner. One factor that has been recognised to be involved in the pathogenesis and progression of gynaecological cancers is the endocannabinoid system (ECS). The ECS consists of endocannabinoids (bioactive lipids), their receptors, and metabolic enzymes responsible for their synthesis and degradation. In this review, the impact of plant-derived (Cannabis species) cannabinoids and endocannabinoids on gynaecological cancers will be discussed within the context of the complexity of the proteins that bind, transport, and metabolise these compounds in reproductive and other tissues. In particular, the potential of endocannabinoids, their receptors, and metabolic enzymes as biomarkers of specific cancers, such as those of the endometrium, will be addressed. Additionally, the therapeutic potential of targeting selected elements of the ECS as new action points for the development of innovative drugs will be presented.
... ECB system has been shown to play critical role at all stages of such synchronization, starting from the development of the preimplantation embryo, its movement through oviduct, throughout implantation and placenta development. AEA synthesis and degradation enzymes NAPE-PLD and FAAH, respectively, are expressed at the two-cell stage of embryonic development Taylor et al., 2010). While CB1 receptor is detected from the four-cell stage, CB2 is found as early as the one-cell stage of embryonic development Taylor et al., 2010). ...
... AEA synthesis and degradation enzymes NAPE-PLD and FAAH, respectively, are expressed at the two-cell stage of embryonic development Taylor et al., 2010). While CB1 receptor is detected from the four-cell stage, CB2 is found as early as the one-cell stage of embryonic development Taylor et al., 2010). Systematic studies point at CB1 receptor and CB1-mediated signaling as critical players in early development. ...
Chapter
The endocannabinoid (eCB) system comprises endogenously produced cannabinoids (CBs), enzymes of their production and degradation, and CB-sensing receptors and transporters. The eCB system plays a critical role in virtually all stages of animal development. Studies on eCB system components and their physiological role have gained increasing attention with the rising legalization and medical use of marijuana products. The latter represent exogenous interventions that target the eCB system. This chapter summarizes knowledge in the field of CB contribution to gametogenesis, fertilization, embryo implantation, fetal development, birth, and adolescence-equivalent periods of ontogenesis. The material is complemented by the overview of data from our laboratory documenting the functional presence of the eCB system within cerebral arteries of baboons at different stages of development.
... ECS modulates the endometrium plasticity and the uterus receptivity. AEA is considered to be the main factor enabling the synchronization between the embryo development and the endometrium, thus allowing its implantation during the so-called "implantation window" (Fig. 2) (46)(47)(48)(49)(50)(51). AEA levels are high in non-receptive uteri and in the interimplantary areas in the receptive uterus, and low in receptive uteri and in implantary areas (low levels of NAPE-PLD and high FAAH activity and expression), thus leading to the idea that a balanced endocannabinoid system is essential for the synchronization between embryo implantation and endometrium preparation for it (37). ...
Article
The discovery of the cannabinoid receptors CB1 and CB2 in 1990 and 1993, respectively, as well as of the two main endocannabinoids, anandamide in 1992 and 2-arachidonylglycerol in 1995, was an important step in identifying the strongest homeostatic system in the human body, namely the endocannabinoid system. Ever since, research has highlighted the crucial part played by this system in all the reproduction stages: folliculogenesis, spermatogenesis, oogenesis, fecundation, transport of the egg through the fallopian tubes, blastocyte implantation and pregnancy progression, as well as its implications in the physiopathology of the reproductive system: in endometriosis, ectopic pregnancy, miscarriage, preeclampsia, endometrial cancer, polycystic ovary syndrome, ovarian cancer. A special attention must be paid to the phytocannabinoids, natural components originating especially from the Cannabis plant inflorescences, whose medical effects are well-established nowadays with also acting on the receptors of the endocannabinoid system. The most recent research mainly focuses on the reproductive dysfunctions and disorders of the reproductive tissues, respectively, through its action upon the endocannabinoid system. Medical cannabis is nowadays legalized in more and more countries all over the world. At the same time, recreational cannabis remains one of the most consumed drugs (in Romania the most consumed one by young adults). Therefore, it is mandatory for specialists in obstetrics and gynecology, endocrinology, public health, hygiene or for general practitioners, to permanently update their information on this subject.
... The presence of ECS elements in several reproductive fluids, cells and tissues like human serum and follicular fluid, ovary, uterus and placenta, have been documented by several studies in animal models and humans [45][46][47][48][49][50]. Cannabis extracts, THC and modulators of the ECS clearly have a major impact on virtually all steps of female reproduction [41,51,52]. ...
Article
Cannabis extracts like marijuana have the highest consumption rate worldwide. Yet, their societal acceptance as recreational and therapeutic drugs could represent a serious hazard to female human reproduction, because cannabis ingredients [termed (phyto)cannabinoids] can perturb an endogenous system of lipid signals known as endocannabinoids. Accumulated evidence on animal models and humans has demonstrated a crucial role of these endogenous signals on different aspects of female reproduction, where they act through an ensamble of proteins that synthesize, transport, degrade and traffic them. Several reports have recently evidenced the potential role of endocannabinoids as biomarkers of female infertility for disease treatment and prevention, as well as their possible epigenetic effects on pregnancy. The purpose of this review is to provide an update of data collected in the last decade on the effects of cannabinoids and endocannabinoids on female reproductive events, from development and maturation of follicles and oocytes, to fertilization, oviductal transport, implantation and labor. In this context, a particular attention has ben devoted to the ovary and the production of fertilizable oocytes, because recent studies have addressed this hot topic with conflicting results among species.
... Recently, a relationship between the endocannabinoid system and other forms of cancer has emerged (8)(9)(10). This system consists of a family of signalling lipids called N-acylethanolamines (NAE), of which the endocannabinoid N-arachidonoylethanolamine (anandamide; AEA) and the endocannabinoid-like compounds, N-oleoylethanolamine (OEA) and N-palmitoylethanolamine (PEA), are the best characterised (10) with respect to their receptors and the enzymes responsible for their synthesis and degradation (11,12). Where this relationship has been investigated, it has provided an improved understanding of the biology of various tumours (13,14). ...
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Objective: To identify new biochemical markers for endometrial cancer (EC). Recent evidence suggests that members of the endocannabinoid system (N-acylethanolamines) that bind to and activate receptors that are dysregulated in EC are involved in this tumour's biology. These observations suggest increased N-acylethanolamine levels in the tissue that might appear in plasma and could be used as disease biomarkers. Methods:N-arachidonoylethanolamine (anandamide, AEA) and the N-acylethanolamine substances, N-oleoylethanolamine (OEA), and N-palmitoylethanolamine (PEA) were quantified in plasma and endometrial tissue collected from 31 EC and seven atrophic controls using UHPLC-MS/MS. Receiver-operating characteristics (ROC) and logistic regression were used to determine diagnostic accuracy. Cannabinoid receptor 1 (CB1) and 2 (CB2) protein levels were determined by specific immunohistochemistry and histomorphometric analyses. Correlations between plasma and tissue levels of the three N-acylethanolamines and tissue levels of the three N-acylethanolamines and CB1 and CB2 receptor expression levels were determined using correlation analysis. Results: Plasma and tissue AEA and PEA levels were significantly (p < 0.05) higher in EC than controls whilst OEA levels were significantly elevated in type 1 EC tissues but not in plasma. There were significant positive correlations between plasma and tissue levels of AEA (R2 = 0.302, p = 0.008) and PEA (R2 = 0.182, p = 0.047), but not for OEA (R2 = 0.022, p = 0.506). The diagnostic accuracies for EC were: sensitivity of 53.3%, specificity of 100% for plasma AEA (>1.36 nM); sensitivity of 73.3%, specificity of 100% for plasma PEA (>27.5 nM); and sensitivity of 93.3%, specificity of 28.6% for plasma OEA (>4.97 nM). Logistic regression increased the area under the ROC curve (AUC) from 0.781 for AEA, 0.857 for PEA, and 0.543 for OEA to a combined AUC of 0.933 for EC diagnosis. Significant inverse correlations between tissue AEA (R2 = 0.343, p = 0.003) and PEA (R2 = 0.384, p < 0.0001) levels and CB1 expression were observed. No correlation between tissue levels of OEA and CB1 and tissue levels of any of the three N-acylethanolamines and CB2 protein expression were observed, except in the type 1 EC patients. Conclusion: Since plasma AEA and PEA are significantly elevated in patients with EC and a reflection of production by the endometrial tumour, then these lipids have the potential to be useful biomarkers for the early diagnosis of EC.
Chapter
s Sociability is foundational to individual and species success and is regulated by a complex neural circuitry that integrates reward, threat, context, and experience. Many of these aspects differ for males and females and across the life span. The principles guiding the formation of the social circuit are in most cases poorly defined but a clear role for endocannabinoids has been identified in the amygdala and preoptic area where they drive the establishment of sex differences in brain and behavior. These findings help inform the risk of gestational cannabis exposure as well as fundamental principles of brain development.
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The endocannabinoid system (ECS) is a complex of neurotransmitters in the central nervous system and plays a key role in regulating cognitive and physiological processes. 2-Arachidonoylglycerol (2-AG) is one of the imperative endocannabinoids that play key roles in the central nervous system. It acts as a signaling lipid and activates the cannabinoid CB1 receptor. In addition, 2-AG is involved in a variety of physiological functions such as energy balance, emotion, pain sensation, cognition, and neuroinflammation. So, a rapid and specific diagnosis of 2-AG is of great importance in medical neuroscience. The development of new methods in this area has been one of the most important research areas in recent years. Herein, an innovative immunosensor is developed for quantification of 2- AG. For this means, gold nanostars (GNS) were synthesized and conjugated with a specific biotinylated antibody against 2-AG. The resultant bioconjugate, a bioreceptor with GNS, was immobilized on the surface of a gold electrode and used for the detection of the antigen based on the immunocomplex formation followed by analysis using different electrochemical techniques. For the first time, the 2-AG protein was measured with an excellent linear range of 0.48–1 ng mL�1 and a lower limit of quantification of 0.48 ng L�1 by the electroanalysis method. The engineered immunosensor showed high sensitivity and specificity in the presence of interfering antigens, proving its utility in neurological disorder detection. This immunosensor is the first sandwich-type immunoassay for the detection of 2-AG in real samples and the first innovation in designing a novel sandwich-type immunosensor for this analyte. Also, excellent analytical results are other advantages of this biosensor for the detection of 2-AG in human plasma samples and serum samples of rats under sleep deprivation. So, this is the first report of an immunosensor of 2-AG using a sandwich-type immunosensor.
Chapter
The global demand for innovative consumer products containing cannabis- and hemp-derived constituents has placed safety at the forefront of consumers, retailers, and regulatory agencies. Safety studies have not kept pace with the diverse number of consumer “health” products, containing the main constituents of cannabis and hemp (e.g., Δ⁹-tetrahydrocannabinol (THC), cannabidiol, (CBD), and terpenes), entering the marketplace. In particular, the reproductive and developmental effects of cannabis, hemp, and their isolated constituents on vulnerable populations including pregnant women, newborns, and youth remain to be fully understood. Discrepancies in existing preclinical and clinical data do not project confidence in the safety concerns concluded from investigators. This chapter focuses on the potential reproductive and developmental toxicities of both cannabis- and hemp-derived constituents with a focus on the interpretation and translation of nonclinical data to humans given ethical constraints over conducting intervention-type clinical trials in sensitive populations.
Article
Globally, the availability and formulations for the administration of cannabis are changing with decriminalization or legalization of recreational use in some jurisdictions, and the prescription of cannabis also occurring. These changes are likely to affect the prevalence of use, including by women of childbearing age. The effects of in utero and infant alcohol and tobacco exposure are well-documented, but the outcomes of cannabis exposure are less certain. The content of delta-9-tetrahydrocannabinol (THC), the psychoactive component of cannabis has progressively increased over several decades. This review explores the limited knowledge surrounding the epidemiology of gestational and postnatal cannabis exposure and implications for the mother–placenta–fetus/neonate triad. We examine cannabis’ effects from antenatal and lactation exposure on (a) pregnancy and perinatal outcomes, (b) placental health, and (c) longer term cardiometabolic and neurodevelopmental risks and outcomes. Though definitive outcomes are lacking, gestational cannabis has been associated with increased risk of other substance use during pregnancy; impaired placental blood flow; increased risk of small for gestational age births; and associated complications. Childhood and adolescent outcomes are sparsely assessed, with suggested outcomes including increased risk of depression and attention-deficit hyperactivity disorder. Cardiometabolic implications of gestational cannabis use may include maternal fatty liver, obesity, insulin resistance, and increased risk of gestational diabetes mellitus (GDM), with potential consequences for the fetus. Clinical implications for pediatric practice were explored in a bid to understand any potential risk or impact on child health and development.
Article
Background People with psychotic disorders have reduced life expectancy largely because of physical health problems, especially cardiovascular disease, that are complicated by the use of tobacco and cannabis. Objectives We set out to (1) chart lifestyle and substance use choices and the emergence of cardiometabolic risk from the earliest presentation with psychosis, (2) develop a pragmatic health promotion intervention integrated within the clinical teams to improve the lifestyle choices and health outcomes of people with psychosis and (3) evaluate the clinical effectiveness and cost-effectiveness of that health promotion intervention. Design We performed a longitudinal cohort study of people presenting with their first episode of psychosis in three mental health trusts and followed up participants for 1 year [work package 1, physical health and substance use measures in first episode of psychosis (PUMP)]. We used an iterative Delphi methodology to develop and refine a modular health promotion intervention, improving physical health and reducing substance use in psychosis (IMPaCT) therapy, which was to be delivered by the patient’s usual care co-ordinator and used motivational interviewing techniques and cognitive–behavioural therapy to improve health choices of people with psychosis (work package 2). We then conducted a multicentre, two-arm, parallel-cluster, randomised controlled trial to determine the clinical effectiveness and cost-effectiveness of using the intervention with people with established psychosis (work package 3: IMPaCT randomised controlled trial) in five UK mental health trusts. The work took place between 2008 and 2014. Participants All people aged between 16 and 65 years within 6 months of their first presentation with a non-organic psychosis and who were proficient in English were eligible for inclusion in the PUMP study. Participants in the work package 2 training development were staff selected from a range of settings, working with psychosis. Participants in the phase 3 Delphi consensus and manual development comprised three expert groups of (1) therapists/researchers recruited from the local and national community, (2) clinicians and (3) service users, each of whom took part in two iterative review and feedback sessions. For work package 3, IMPaCT randomised controlled trial, care co-ordinators in participating community mental health teams who were permanently employed and had a minimum of four eligible patients (i.e. aged between 18 and 65 years with a diagnosis of a psychotic disorder) on their caseload were eligible to participate. In studies 1 and 3, patient participants were ineligible if they were pregnant or had a major illness that would have had an impact on their metabolic status or if they had a significant learning disability. All participants were included in the study only after giving written confirmed consent. Main outcome measures Cardiometabolic risk markers, including rates of obesity and central obesity, and levels of glycated haemoglobin (HbA 1c ) and lipids, were the main outcomes in work package 1 (PUMP), with descriptive data presented on substance use. Our primary outcome measure for the IMPaCT randomised controlled trial was the physical or mental health component Short Form questionnaire-36 items quality-of-life scores at 12 months. Results Obesity rates rose from 18% at first presentation with psychosis to 24% by 1 year, but cardiometabolic risk was not associated with baseline lifestyle and substance use choices. Patterns of increase in the levels of HbA 1c over the year following first presentation showed variation by ethnic group. We recruited 104 care co-ordinators, of whom 52 (with 213 patients) were randomised to deliver IMPaCT therapy and 52 (with 193 patients) were randomised to deliver treatment as usual, in keeping with our power calculations. Of these 406 participants with established psychosis, 318 (78%) and 301 (74%) participants, respectively, attended the 12- and 15-month follow-ups. We found no significant effect of IMPaCT therapy compared with treatment as usual on the physical or mental health component Short Form questionnaire-36 items scores at either time point in an intention-to-treat analysis [physical health score (‘d’) –0.17 at 12 months and –0.09 at 15 months; mental health score (‘d’) 0.03 at 12 months and –0.05 at 15 months] or on costs. Nor did we find an effect on other cardiovascular risk indicators, including diabetes, except in the case of high-density lipoprotein cholesterol, which showed a trend for greater benefit with IMPaCT therapy than with treatment as usual (treatment effect 0.085, 95% confidence interval 0.007 to 0.16; p = 0.034). Limitations Follow-up in work package 1 was challenging, with 127 out of 293 participants attending; however, there was no difference in cardiometabolic measures or demographic factors at baseline between those who attended for follow-up and those who did not. In work package 3, the IMPaCT randomised controlled trial, care co-ordinators struggled to provide additional time to their patients that was devoted to the health promotion intervention on top of their usual clinical care contact with them. Conclusions Cardiometabolic risk is prominent even soon after first presentation with psychosis and increases over time. Lifestyle choices and substance use habits at first presentation do not predict those who will be most cardiometabolically compromised 1 year later. Training and supervising care co-ordinators to deliver a health promotion intervention to their own patients on top of routine care is not effective in the NHS for improving quality of life or reducing cardiometabolic risk. Future work Further work is needed to develop and evaluate effective, cost-effective and affordable ways of preventing the emergence of and reversing existing cardiometabolic risk indicators in people with psychosis. Trial registration Current Controlled Trials ISRCTN58667926. Funding This project was funded by the National Institute for Health Research (NIHR) Programme Grants for Applied Research programme and will be published in full in Programme Grants for Applied Research ; Vol. 8, No. 1. See the NIHR Journals Library website for further project information.
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Human reproduction is a rather inefficient process, yet the molecular reasons for this inefficiency remain unknown. IVF and embryo transfer (IVF–embryo transfer) also results in a high frequency of implantation failures and early spontaneous abortions. Here we show that the anandamide (AEA)-degrading enzyme, fatty acid amide hydrolase (FAAH), had significantly lower activity (46 17 versus 161 74 pmol/min per mg protein) and protein content (0.10 0.03 versus 0.23 0.06 units) in lymphocytes of IVF–embryo transfer patients who failed to achieve an ongoing pregnancy than in those who become pregnant, and this was paralleled by a significant increase in blood AEA (4.0 2.2 pmol/ml and 0.9 1.0 pmol/ml respectively). The blood levels of the other endocannabinoid, 2-arachidonoylglycerol, or of the AEA congener, N-palmitoylethanolamine, which are metabolized by enzymes different from FAAH, was not different between the pregnant and nonpregnant women, nor was there any difference in the activity of the AEA membrane transporter or the amounts of cannabinoid receptors in lymphocytes. Taken together with the reported negative effects of AEA on embryo implantation, this study indicates that low FAAH activity and subsequent increased AEA levels in blood might be one of the causes of implantation failure or pregnancy loss, thereby leading to a better understanding of the pathophysiological and therapeutic implications of endocannabinoids in human fertility.
Article
In a prospective study of 3,857 pregnancies ending in singleton live births at Yale-New Haven Hospital, New Haven, Connecticut, in 1980–1982,9.5% of mothers reported using marijuana (4.1% occasionally and 5.4% at least 2–3 times monthly). Among whtte women, regular use was associated with an increased risk of delivering a low birth weight (<2,500 gm) infant (odds ratio (OR) = 2.6, 95% confidence interval (CI) = 1.1–6.2) and small for gestational age infant (OR = 2.3, 95% CI = 1.3–4.1) after adjustment for other risk factors. Nonwhite marijuana users were not at further increased risk for delivering a low birth weight or small for gestational age infant beyond the elevated rates of these conditions already experienced by nonwhites in general. Marijuana use was also related to preterm delivery (gestational age <37 weeks from last menstrual period) in white women (OR = 1.9, 95% CI = 1.0–3.9) but not nonwhtte women. Occasional use was unrelated to the risk of low birth weight, small for gestational age, or preterm delivery.
Article
Single doses of ∆ 9 -tetrahydrocannabinol (THC) (5.0, 2.5, 1.25, or 0.625 mg/kg) can decrease the levels of both luteinizing hormone (LH) and follicle-stimulating hormone (FSH) in the ovariectomized rhesus monkey. The inhibition of gonadotropins (50% to 88%) lasts for 6 to 24 hours depending upon the dose of THC. There are no great differences in the responses of the two gonadotropins to THC. The inhibition of gonadotropin levels by THC appears to be at the level of the hypothalamus, since both LH and FSH are released from the pituitary gland in response to LH-releasing factor in the presence of THC.
Article
Platelets and leukocytes in venous blood were counted with automated methodology in 23 women studied longitudinally throughout pregnancy and at six weeks after delivery. The average platelet count declined monotonically in a manner consistent with the gestational increase in blood volume. Total leukocyte count rose early in gestation and remained elevated, with neutrophils accounting for the changes. Absolute neutrophil counts during gestation were nearly twice those of the same women postpartum (5.76 vs 3.13çu mm ×10-3.) The monocyte count also tended to increase, whereas lymphocytes, eosinophils, and basophils declined in number. (JAMA 242:2696-2698, 1979)