# Placental Implications of Peroxisome Proliferator-Activated Receptors in Gestation and Parturition

Authors: Valerie Borel; Denis Gallot; Geoffroy Marceau; Vincent Sapin; Loïc Blanchon
Journal: PPAR Research
 (2008)

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## Abstract

The placenta is a transitory structure indispensable for the proper development of the embryo and fetus during mammalian gestation. Like other members of the nuclear receptor family, the peroxisome proliferator-activated receptors
(PPARs) are known to be involved in the physiological and pathological events occurring during the placentation.
This placental involvement has been recently reviewed focusing on the early stages of placental development (implantation and invasion, etc.), mouse PPARs knockout phenotypes, and cytotrophoblast physiology. In this
review, we describe the placental involvement of PPARs (e.g., fat transport and metabolism, etc.) during the late stages of
gestation and in the amniotic membranes, highlighting their roles in the inflammation process (e.g., chorioamnionitis), metabolic disorders (e.g., diabetes), and parturition.


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## Body

## 1. The Peroxisome Proliferator-Activated  Receptors (PPARs)

### 1.1. Nomenclature and Structure

Discovered in 1990,
PPARs are known for their biological role in inducing the proliferation of
peroxisomes in rodents [1]. They are transcription factors
belonging to the ligand-activated nuclear hormone receptor superfamily [2] and have been identified in
different species such as the xenopus, mouse, rat, and humans. In all these
species, PPARs present three isotypes encoded by distinct single-copy genes:
PPARα (NR1C1), PPARβ/δ (also called NUC1 or NR1C2), and PPARγ (NR1C3), located
on chromosomes 15, 17, 6 in the mouse and chromosomes 22, 6, 3
in humans, respectively. The PPARγ gene alternative
promoters give rise to three different isoforms named γ1, γ2, and γ3 which
differ at their 5′ends (see Figure 1(a)) [3]. PPAR α, β, γ1/γ3, γ2 translation produces proteins of 468, 441, 475, and 505 amino acids,
respectively, with a molecular weight  of
49 to 56 kDa [4]. By performing multiple PPAR nucleotide/protein
alignments of PPARs in different species, a strong interspecies identity
(human, mouse, rat, bovine, ≈90%) has been
established, illustrating a strong evolutionary conservation among species by
derivation from a common ancestor (Table 1). PPARγ shows the highest
conservation in terms of cDNA and proteins.Table 1
Percentage of nucleotide and amino acid identity between the human, mouse, rat,
and bovine PPAR sequences. No PPARγ3 alignment was carried out owing to lack of
data on different species. The different sequences came from Ensembl and were
aligned with Genomatix software.
cDNA homology (%)Protein homology (%)MouseRatBovineMouseRatBovineHuman relative identity percentPPARα446472929294PPARβ606975929195PPARγ1798478989797PPARγ2868688969595(a) Schematic representation of PPARγ genes, mRNA, and proteins. The 5′exons A1, A2, B can be alternatively spliced to give rise to the different PPARγ isoforms. The boxes 1 to 6 correspond to exons which are common to PPARγ1, γ2, γ3 genes. ATG is the initiation transcription site. The molecular weight of these isoforms ranges from 49 to 56 kDa. (b) Schematic representation of typical nuclear receptor structure. AF1: activating function 1 (ligand-independent function), AF2: activating function 2 (ligand-dependent function), NLS: nuclear signal localization.
(a)(b)Like several other
members of the nuclear receptor superfamily, PPARs possess the typical
structure organised in six domains named A to F (see Figure1(b)) [5]. Domain C (DBD: DNA binding domain)
contains two zinc fingers and allows promoter target gene interaction and
dimerization with its preferential nuclear receptor: retinoid X receptor (RXR).
The PPAR/RXR heterodimer binds to the target gene promoter response element
named peroxisome proliferator response element (PPRE) which is made up of two
half site AGGTCA separated by one or two nucleotides (also called DR1 or DR2
for direct repeat 1 or 2) and a 5′ extension A (A/T) CT. Domain E/F allows
ligand binding and contains a ligand-dependent transactivation function called
AF2 (activating function 2). It is involved in dimerization and interaction
with cofactors.


### 1.2. PPAR Ligands

As with the other nuclear receptors, the binding of the ligand is a key step in
the control of PPAR transcriptional activity. In the absence of a ligand,
corepressors and histone deacetylases (HDAC) bind to PPARs and inhibit the transcription
activation of target genes. PPAR ligands have the ability to dissociate the
corepressor complexes from the PPAR/RXR heterodimer, allowing the binding of
the coactivators in order to initiate and activate transcription.There are two kinds of
ligands for the PPARs: natural and synthetic. Among the natural ligands the
monounsaturated fatty acids (FA) (e.g., oleic acid) and the polyunsaturated
fatty acids (PUFA) (e.g., linoleic acid, linolenic acid, and arachidonic acid) are
described as ligands for PPARα, PPARβ, and PPARγ. They act with concentrations
consistent with those found in human serum [6]. The different
PUFA metabolites: 8(S)- and 15-hydroxyeicosatetraenoic acid (8(S)- and 15-HETE), leukotriene B4
(LTB4), 9- and 13-hydroxyoctadedienoic acid (9-HODE and 13-HODE) and 15-deoxy-Δ12,
14-prostaglandin J2 (PGJ2) are potent selective activators of PPARα and
PPARγ. Some oxidized low-density lipoproteins (LDLs), oxidized
alkyphospholipids, nitrolinolenic acid, and prostaglandin metabolites can also
activate PPARγ [7]. Recently, it has been demonstrated
that P450 eicosanoids are potent PPARα and PPARγ ligands [8]. Indeed,
Ng et al. [8] have shown that P450 catalysed arachidonic acid metabolites like
20-hydroxyeicosatetraenoic acid (20-HETE) or 11, 12-epoxyeicosatrienoic acid
(11, 12-EET) can activate PPARα and PPARγ. These ligands induce PPAR binding to
PPRE and can modify the expression of PPARα responsive genes like apoA-I or
apoA-II in the same way than synthetic ligands. Thus the finely regulated
conversion of PUFAs to eicosanoids through either the lipoxygenase, cyclooxygenase,
or cytochrome P450 monooxygenase pathways may provide a mechanism for the
differential regulation of PPARα and PPARγ and their respective target genes.
PPARβ can be activated by different types of eicosanoids including prostaglandinA1
(PGA1) and prostaglandin D2 (PGD2). Many synthetic ligands exist and have been
used in PPAR work. These ligands include prostaglandin 12 analogs, pirinixic
acid (Wy-14643) for PPARα, hypolipidemic and hypoglycemic agents
(nonthiazolidinedione) for PPARβ, and thiazolidinediones (e.g., rosiglitasone,
troglitazone) for PPARγ [2].


## 1.1. Nomenclature and Structure

Discovered in 1990,
PPARs are known for their biological role in inducing the proliferation of
peroxisomes in rodents [1]. They are transcription factors
belonging to the ligand-activated nuclear hormone receptor superfamily [2] and have been identified in
different species such as the xenopus, mouse, rat, and humans. In all these
species, PPARs present three isotypes encoded by distinct single-copy genes:
PPARα (NR1C1), PPARβ/δ (also called NUC1 or NR1C2), and PPARγ (NR1C3), located
on chromosomes 15, 17, 6 in the mouse and chromosomes 22, 6, 3
in humans, respectively. The PPARγ gene alternative
promoters give rise to three different isoforms named γ1, γ2, and γ3 which
differ at their 5′ends (see Figure 1(a)) [3]. PPAR α, β, γ1/γ3, γ2 translation produces proteins of 468, 441, 475, and 505 amino acids,
respectively, with a molecular weight  of
49 to 56 kDa [4]. By performing multiple PPAR nucleotide/protein
alignments of PPARs in different species, a strong interspecies identity
(human, mouse, rat, bovine, ≈90%) has been
established, illustrating a strong evolutionary conservation among species by
derivation from a common ancestor (Table 1). PPARγ shows the highest
conservation in terms of cDNA and proteins.Table 1
Percentage of nucleotide and amino acid identity between the human, mouse, rat,
and bovine PPAR sequences. No PPARγ3 alignment was carried out owing to lack of
data on different species. The different sequences came from Ensembl and were
aligned with Genomatix software.
cDNA homology (%)Protein homology (%)MouseRatBovineMouseRatBovineHuman relative identity percentPPARα446472929294PPARβ606975929195PPARγ1798478989797PPARγ2868688969595(a) Schematic representation of PPARγ genes, mRNA, and proteins. The 5′exons A1, A2, B can be alternatively spliced to give rise to the different PPARγ isoforms. The boxes 1 to 6 correspond to exons which are common to PPARγ1, γ2, γ3 genes. ATG is the initiation transcription site. The molecular weight of these isoforms ranges from 49 to 56 kDa. (b) Schematic representation of typical nuclear receptor structure. AF1: activating function 1 (ligand-independent function), AF2: activating function 2 (ligand-dependent function), NLS: nuclear signal localization.
(a)(b)Like several other
members of the nuclear receptor superfamily, PPARs possess the typical
structure organised in six domains named A to F (see Figure1(b)) [5]. Domain C (DBD: DNA binding domain)
contains two zinc fingers and allows promoter target gene interaction and
dimerization with its preferential nuclear receptor: retinoid X receptor (RXR).
The PPAR/RXR heterodimer binds to the target gene promoter response element
named peroxisome proliferator response element (PPRE) which is made up of two
half site AGGTCA separated by one or two nucleotides (also called DR1 or DR2
for direct repeat 1 or 2) and a 5′ extension A (A/T) CT. Domain E/F allows
ligand binding and contains a ligand-dependent transactivation function called
AF2 (activating function 2). It is involved in dimerization and interaction
with cofactors.


## 1.2. PPAR Ligands

As with the other nuclear receptors, the binding of the ligand is a key step in
the control of PPAR transcriptional activity. In the absence of a ligand,
corepressors and histone deacetylases (HDAC) bind to PPARs and inhibit the transcription
activation of target genes. PPAR ligands have the ability to dissociate the
corepressor complexes from the PPAR/RXR heterodimer, allowing the binding of
the coactivators in order to initiate and activate transcription.There are two kinds of
ligands for the PPARs: natural and synthetic. Among the natural ligands the
monounsaturated fatty acids (FA) (e.g., oleic acid) and the polyunsaturated
fatty acids (PUFA) (e.g., linoleic acid, linolenic acid, and arachidonic acid) are
described as ligands for PPARα, PPARβ, and PPARγ. They act with concentrations
consistent with those found in human serum [6]. The different
PUFA metabolites: 8(S)- and 15-hydroxyeicosatetraenoic acid (8(S)- and 15-HETE), leukotriene B4
(LTB4), 9- and 13-hydroxyoctadedienoic acid (9-HODE and 13-HODE) and 15-deoxy-Δ12,
14-prostaglandin J2 (PGJ2) are potent selective activators of PPARα and
PPARγ. Some oxidized low-density lipoproteins (LDLs), oxidized
alkyphospholipids, nitrolinolenic acid, and prostaglandin metabolites can also
activate PPARγ [7]. Recently, it has been demonstrated
that P450 eicosanoids are potent PPARα and PPARγ ligands [8]. Indeed,
Ng et al. [8] have shown that P450 catalysed arachidonic acid metabolites like
20-hydroxyeicosatetraenoic acid (20-HETE) or 11, 12-epoxyeicosatrienoic acid
(11, 12-EET) can activate PPARα and PPARγ. These ligands induce PPAR binding to
PPRE and can modify the expression of PPARα responsive genes like apoA-I or
apoA-II in the same way than synthetic ligands. Thus the finely regulated
conversion of PUFAs to eicosanoids through either the lipoxygenase, cyclooxygenase,
or cytochrome P450 monooxygenase pathways may provide a mechanism for the
differential regulation of PPARα and PPARγ and their respective target genes.
PPARβ can be activated by different types of eicosanoids including prostaglandinA1
(PGA1) and prostaglandin D2 (PGD2). Many synthetic ligands exist and have been
used in PPAR work. These ligands include prostaglandin 12 analogs, pirinixic
acid (Wy-14643) for PPARα, hypolipidemic and hypoglycemic agents
(nonthiazolidinedione) for PPARβ, and thiazolidinediones (e.g., rosiglitasone,
troglitazone) for PPARγ [2].


## 2. PPAR Expression Patterns

The adult PPAR expression patterns have been extensively established at the mRNA
and protein levels in several species (Table2) [8, 9]. Several studies conducted during
mammalian gestation have established the placenta as an important expression
site of the different PPARs isoforms. Our review will focus only on term
placental expression and on the amniotic/fetal membranes. The placental dynamic
expression of the 3 PPARs during early and midgestation (of mouse, rat, and
human) is well described in Fournier et
al., 2007 [4]. In rat placenta, all three PPAR
isoforms are ubiquitously expressed from 11 days postcoitum (dpc) [10]. Both PPARβ/δ and PPARγ are expressed after 8.5 dpc in mouse placenta. By immunohistochemistry
and RT-PCR, the three PPAR isoforms are been shown to be expressed in the
villous trophoblastic cells and syncytiotrophoblasts of the human term placenta
[4]. To extend the previously published
results [11] and to assess the potential
importance of PPAR proteins in fetal membranes, RT-PCR and immunohistochemistry
experiments were performed on human term placental samples. The three PPARs are
present in total placenta, amnion, chorion, and in amnion-derived WISH
epithelial cell line at the mRNA (see Figure 2(a)) and protein levels (see Figure
2(b)). The expression of PPARα and PPARγ seems to be weaker than that observed
for PPARβ/δ. In addition, a greater amplification of the PPARγ cDNA is obtained
in chorion than in amnion, where PPARγ is almost undetectable.Table 2
Summary of PPAR expression patterns.
(a) Global expression patternGeneSpeciesExpression localizationReferencesPPARαRodentsCardiomyocytes[6, 8]HepatocytesHumanHeart[6, 8]KidneyLarge intestineLeydig and seminiferous tubule cells[12]Liver[6, 8]Skeletal muscleUterus[12]Ovary (Theca and stroma cells)PPARβRodentsUbiquitous[6, 8]HumanUbiquitous[6, 8]PPARγRodentsBrown and white adipose tissue[6, 8]Lymphoid organsRetinaSkeletal muscleUterus[12]Granulosa cells, corpus luteumHumanColon[6, 8]KidneyLiverSkeltal muscleVascular endotheliumWhite adipose tissueSertoli cells[12]UterusGranulosa cells(B) Placental expression patternPPARα, β, γRodentsTerm placenta[4, 9]HumanVillous trophoblastic cells and syncytiotrophoblasts[4, 9, 10]Amnion, chorion, and amnion derived-WISH cell linePPARs expression in term placenta and amniotic membrances. (a) RT-PCR assays of PPARα, PPARβ, and PPARγ mRNA in amnion, chorion, placenta, and WISH cells. PCR products were analyzed on 1.8% agarose gel and stained with ethidium bromide. 36B4 corresponds to the housekeeping gene. A: Amnion, C: Chorion, A+C: Amnino+Chorion, P: Total placenta, W: WISH cells. (b) PPARs immunostaining of amnion and chorion. Note that all PPARs are expressed in nucleus. Magnification: x200.
(a)(b)


## 3. Implications of PPARs in Placenta and Fetal Membranes

### 3.1. Placental and Amniotic Presence of PPARs Ligands

The lipids of human amnion and chorion are enriched in the essential fatty acid
arachidonic acid, which is the precursor of all the prostaglandins of the 2 series
[13]. Sixty-six percent of the
arachidonic acid of the human fetal membranes are available in
the glycerophospholipids of these tissues and can easily be converted into PGD2 [14]. The placenta produces considerable
amounts of PGD2 [15]. The enzymes necessary to convert PGD2 into prostaglandin J2 (PGJ2) are present and coexpressed with PPARγ in placenta. 15-Deoxy-Δ12, 14-PGJ2 (15dPGJ2) and its precursor PGD2 are present in amniotic fluid at concentrations that do
not exceed 3 nM [16]. However,
this amniotic fluid concentration cannot be an exact representation of the
physiological placental reality for PPARs ligands because the nuclear
concentration is not measured. The maternal blood may also be a source of PPAR
ligands for the human placenta and the fetal membranes. It has been established
that a heat-stable compound (not a protein, but rather a prostanoid or a fatty
acid) is detected in maternal blood serum and is able to activate the PPARγ [17]. The presence of classical and new PPARs
ligands (e.g., P450 eicosanoids, PUFA metabolites) in placenta and fetal
membranes suggests that they could activate PPAR, induce PPAR binding to PPRE,
and modify the expression of PPAR target genes; but this hypothesis has to be
confirmed by further analysis, based on PPARs activation in other organs. For
example, PUFAs, such as and eicosapentaenoic acid (EPA) and docosahexaenoic
acid (DHA), increased PPARγ mRNA expression and binding to PPRE in renal
tubular epithelial cell line
(HK-2). Furthermore, they downregulate LPS-induced activation of NF-κB via a
PPARγ-dependent pathway
in HK-2 cells [18]. Another example showed that PGD2
is among the most abundantly produced prostaglandins in synovial fluid by
synovial fibroblasts [19]. It can be converted into PGJ2. It has been demonstrated that PPARγ
ligands (15dPGJ2) inhibit IL-1β–induced
production of nitric oxide (NO) and matrix metalloproteinase-13 (MMP-13) in
chondrocytes. This inhibition was PPARγ-dependent and occurred at the
transcriptional level, through repression of NF-κB signalling [20]. These two examples support a role
of PPAR ligands in fetal membranes.


### 3.2. Fundamental Implications of PPARs during Early Placentation

As a determining result, the knockout of the PPARγ in mice [21] yielded the first findings indicating
the importance of this factor in early embryonic and perinatal development.
These results are concomitant with those obtained by the generation of RXRα or
β null mice (PPARγ partner in the functional heterodimer), also showing an
embryonic lethality explained by the lack of generation of a functional
labyrinthine zone [22]. Furthermore, complementary studies
conducted by the inactivation of PPARγ coactivators or coregulators, such as peroxisome
proliferators activator receptor-binding protein (PBP) and peroxisome proliferator-activated receptor-interacting protein (PRIP),
also lead to severe placental dysfunction, such as inadequate vascularisation
of the structure [23–25]. Recently, Barak et al. also
demonstrated that the inactivation of PPARβ/δ led to the formation of abnormal
gaps and a thinner but fully differentiated vascular structure in the placentodecidual
interface [26]. These results establish the nonredundant
roles of PPARγ and PPARβ/δ in early mouse placental
development. By contrast, the inactivation of PPARα has no
effect on placental formation or on the developing foetus and by the way theirs
possible roles during pregnancy had to be clarified [2]. In humans, the studies are almost
exclusively focused on the PPARγ roles during early placentation. It
has been clearly established that all three PPARs can stimulate or inhibit the
differentiation and/or proliferation of the villous cytotrophoblasts into
syncytiotrophoblasts and the synthesis of chorionic gonadotrophic hormone, and
may hamper extravillous trophoblastic cell invasion (for more details, see
Fournier et al., 2007 [4]).


### 3.3. Roles of PPARs in the Uptake and Transport of Trophoblastic Lipids

As one of the first
functions described for PPARγ in other tissues, trophoblastic lipid uptake and
accumulation are also regulated in part by this factor [27]. The PPARγ ligands seem to increase the uptake and accumulation of the fatty acids
in human placenta [28]. This regulation is associated with
an enhanced expression of adipophilin (fat droplet-associated protein) and fatty acid transport
proteins (1 and 4) in human trophoblasts [28–30]. These results were confirmed
recently by the in vivo activation of PPARγ by its agonist rosiglitazone in
mice, which also leads to the enhancement of the previous described genes plus
two new ones involved in the lipid transport: S3-12 (plasma associated protein)
and myocardial lipid droplet protein/MLDP [27]. Taken together, these results
confirm the results obtained on PPARγ-null mutants: the absence of the
lipid droplets normally present around the fetal vessels in the wild-type
placenta [21].


### 3.4. PPARs in Placental Inflammatory Response and in the Parturition Signalling

At this stage of our knowledge of PPARs, the most interesting results have been
obtained with the study of their involvement in the inflammation process,
which may be linked to labor at term and also to the premature rupture of fetal
membranes (see Figure3). Term labor is associated with an increase in proinflammatory
proteins and cytokines such as IL1β, IL6, IL8, IL10, and TNF-α. This increase in
proinflammatory proteins and cytokines induces uterine contractions. PPARγ ligands have been demonstrated to inhibit the secretion of IL6, IL8,
and TNF-α in amnion and chorion [31], highlighting the role of PPARs in
the regulation of the inflammatory response in human gestational tissues and
cells [32–35]. The parathyroid hormone-related
protein (presenting a cytokine-like action) is involved in many processes
during normal and pathological pregnancies, and is decreased by PPARγ stimulation [36], which also blocks proinflammatory
cytokine release by adiponectin and leptin [37]. The production of prostaglandins
by the endometrium, the myometrium, and the fetal membranes induces the
contraction of the myometrium during labor. This generation of uterotonic
prostaglandins correlates with the increased prostaglandin-endoperoxide
synthase type 2/cyclooxygenase type 2 (COX-2) activity and the increased secretory phospholipase A2-IIA (sPLA2) mRNA, proteins and activities. By inhibiting the production of the COX-2 and
sPLA2 in fetal membranes, PPARγ promotes the quiescence of the
uterus during gestation [34]. The molecular action of 15dPGJ2
seems to involve interactions of the NF-Kappa B signaling pathway, inducing
reduction of PGF2α, PGE2, and MMP9 release in the placental
environment [31]. This suppressive action of PPARγ
on inflammation is aPPARently time-dependent during pregnancy. The PPARγ level
of expression remains stable throughout gestation, except for the period just
before labor, when its expression in fetal membranes declines. This reduction
is coincidental with a relative increase in COX-2 expression [38]. Further work has shown this simple
scheme to be more complex. While the expression of PPARα does not change at
term in amnion, it decreases in chorion. An increase was also demonstrated for
PPARβ/δ in chorionic and amniotic zones [11]. These last two findings raise the
question of the involvement of the α and β isoforms in this process. The
absence of a real link between COX-2 and PPARγ is presented by Lindstrom and Bennett [39]. Finally, the PPAR action seems to
be concentration-dependent. A small amount of 15dPGJ2 (<0.1 μM) acts through
the PPARγ signaling pathway, where at high
concentration (1 μM) its actions are most probably mediated through
other pathways: PPARβ/δ and/or an inhibition of NF-κB
independent of PPARs [35]. Furthermore, 15dPGJ2 and troglitazone were also demonstrated to have some antiinflammatory or apoptosis-induction
specific effects by PPARγ-independent pathways. This was suggested by the work of Lappas et
al. on human gestational tissues, demonstrating that this effect could
passed by antagonist effect of 15dPGJ2 on the NF-κB pathways and
antioxidant effects of the troglitazone, a synthetic ligand of PPARγ [31].Figure 3
Schematic representation of PPARγ implication in pregnancy maintenance and labor.  IL1β: Interleukin 1β; IL6: Interleukin 6; IL8: Interleukin 8; IL10: Interleukin 10, TNFα: Tumor Necrosis Factor α; COX2: Cyclo-oxygenase type 2; PLA2: Phospholipase A2; NF-κB: Nuclear Factor-Kappa B; MMP9: Matrix Metalloproteinase 9; 15dPGJ2: 15-Deoxy-Δ12, 14-prostaglandin J2.


### 3.5. PPARs in Placental and Amniotic Membranes Pathologies

In contrast to the
different roles described for PPARs during human placentation, only a few
studies on PPARs and placental pathologies have been conducted. In choriocarcinoma
and hydatiform moles, a downregulation of the PPARγ expression is observed but
this real influence needs to be elucidated [40]. The potential involvement of PPARγ on preeclampsia is suggested by the fact that this pathology is
associated with an increased peroxidation in trophoblasts [41, 42]. An overproduction of 15-HETE has
also been noted, suggesting a deregulation of PPARγ [43]. This can cause a strong
transactivation of PPARγ during early pregnancy, resulting
in a reduction of extravillous trophoblastic invasion, one cellular explanation
often cited in the physiopathology of preeclampsia [44, 45]. Other abnormal transactivation of
PPARs may be hypothesized to explain placental pathologies. The 15dPGJ2 has
been shown to induce apoptosis of the placental (JEG-3) and amniotic (WISH) established cell
line, [46, 47]. An excess of 15dPGJ2 production can
be a source of placental dysfunction linked to an increase in trophoblastic
death. It is also established that deletion of PPARγ, PPARβ/δ, and some of their coactivators (PBP, PRIP, and
RAP250) induce abnormal placental phenotypes (abruption,
reduction of fetomaternal exchanges, and alterations of trophoblastic
differentiation) in null mutants [21, 23, 24, 26, 48, 49]. Chromosomal and/or genetic alterations
(point mutation or deletion) may occur for these genes, inducing human
placental alterations. The placental 11β hydroxysteroid dehydrogenase type
2 is a target gene of PPARs [50]. This enzyme plays a key role in
fetal development by controlling fetal exposure to maternal glucocorticoids. An
abnormal regulation by PPARs may result in an absence of fetal protection. In
the rat placental HRP-1 established cell line, the phthalate and derivatives
transactivate PPARs (α and γ) induced an increase in uptake rates of fetal essential fatty acid and
the transport of arachidonic and docosahexaenoic acid [51]. If such a mechanism can be induced
by the phthalates during human placentation, this may strongly affect the fetal
essential fatty acid content during growth.Gestational diabetes is
linked to impaired lipids metabolism [52]. Decreased 15dPGJ2 in blood of
diabetic mothers is also linked to a decrease in
placental PPARγ expression. The inhibition of PPARγ results in an induction of a placental proinflammatory environment associated with an
increase in nitrogen monoxide production and release, which can impair fetoplacental
development [53, 54].The PPAR regulation of
inflammation may be very important in another obstetrical pathology of the
amniotic membranes: the chorioamnionitis. This pathology, usually due to an
ascendant colonization of pathogenic microorganisms from the vagina to the
uterus, is closely associated with preterm labor and premature rupture of
membranes (chorion and amnion). These
ruptures of membranes seem to arise from deregulated proinflammatory factor
synthesis. It has already been reported in this pathology that IL1β, IL6, IL8,
TNF-α, and prostaglandinE(2) show
inadequate concentrations in placental membrane and in amniotic fluid [55–58]. As PPARs may be involved in the occurrence
and control of this inflammatory response, further studies are needed to assess
their importance in this process and to find new possible therapeutic strategies
to prevent this damaging pathology.More generally, the use of natural and
synthetic PPAR ligands looks to be a promising way in preventing placental
pathologies such as endometriosis or preeclampsia. An interesting study also
demonstrates that the reduction of LPS induction of cytokines is reduced by
PPARγ ligands in fetal membranes. Nevertheless, the few studies already
conducted were done practically only on animal (rodent) models and looks to
have positive effects on the pathologies (for review see Toth et al. [59]). Till now, the major problem using,
for example, TZD (thiazolidinedionzes) linking to the PPARγ pathways still the numerous adverse effects of this kind of treatment (e.g.,
weight gain, anemia, leukopenia, etc.). These facts and the potential placental
impacts raised also the question of the use of these medical drugs to treat the
gestational diabetes. Perhaps, at the level of clinician actual knowledge, PPARγ and its ligands could be used  in
a first time, only as good early marker candidates for the diagnosis of
pregnancy pathologies like, for example, preeclampsia.


## 3.1. Placental and Amniotic Presence of PPARs Ligands

The lipids of human amnion and chorion are enriched in the essential fatty acid
arachidonic acid, which is the precursor of all the prostaglandins of the 2 series
[13]. Sixty-six percent of the
arachidonic acid of the human fetal membranes are available in
the glycerophospholipids of these tissues and can easily be converted into PGD2 [14]. The placenta produces considerable
amounts of PGD2 [15]. The enzymes necessary to convert PGD2 into prostaglandin J2 (PGJ2) are present and coexpressed with PPARγ in placenta. 15-Deoxy-Δ12, 14-PGJ2 (15dPGJ2) and its precursor PGD2 are present in amniotic fluid at concentrations that do
not exceed 3 nM [16]. However,
this amniotic fluid concentration cannot be an exact representation of the
physiological placental reality for PPARs ligands because the nuclear
concentration is not measured. The maternal blood may also be a source of PPAR
ligands for the human placenta and the fetal membranes. It has been established
that a heat-stable compound (not a protein, but rather a prostanoid or a fatty
acid) is detected in maternal blood serum and is able to activate the PPARγ [17]. The presence of classical and new PPARs
ligands (e.g., P450 eicosanoids, PUFA metabolites) in placenta and fetal
membranes suggests that they could activate PPAR, induce PPAR binding to PPRE,
and modify the expression of PPAR target genes; but this hypothesis has to be
confirmed by further analysis, based on PPARs activation in other organs. For
example, PUFAs, such as and eicosapentaenoic acid (EPA) and docosahexaenoic
acid (DHA), increased PPARγ mRNA expression and binding to PPRE in renal
tubular epithelial cell line
(HK-2). Furthermore, they downregulate LPS-induced activation of NF-κB via a
PPARγ-dependent pathway
in HK-2 cells [18]. Another example showed that PGD2
is among the most abundantly produced prostaglandins in synovial fluid by
synovial fibroblasts [19]. It can be converted into PGJ2. It has been demonstrated that PPARγ
ligands (15dPGJ2) inhibit IL-1β–induced
production of nitric oxide (NO) and matrix metalloproteinase-13 (MMP-13) in
chondrocytes. This inhibition was PPARγ-dependent and occurred at the
transcriptional level, through repression of NF-κB signalling [20]. These two examples support a role
of PPAR ligands in fetal membranes.


## 3.2. Fundamental Implications of PPARs during Early Placentation

As a determining result, the knockout of the PPARγ in mice [21] yielded the first findings indicating
the importance of this factor in early embryonic and perinatal development.
These results are concomitant with those obtained by the generation of RXRα or
β null mice (PPARγ partner in the functional heterodimer), also showing an
embryonic lethality explained by the lack of generation of a functional
labyrinthine zone [22]. Furthermore, complementary studies
conducted by the inactivation of PPARγ coactivators or coregulators, such as peroxisome
proliferators activator receptor-binding protein (PBP) and peroxisome proliferator-activated receptor-interacting protein (PRIP),
also lead to severe placental dysfunction, such as inadequate vascularisation
of the structure [23–25]. Recently, Barak et al. also
demonstrated that the inactivation of PPARβ/δ led to the formation of abnormal
gaps and a thinner but fully differentiated vascular structure in the placentodecidual
interface [26]. These results establish the nonredundant
roles of PPARγ and PPARβ/δ in early mouse placental
development. By contrast, the inactivation of PPARα has no
effect on placental formation or on the developing foetus and by the way theirs
possible roles during pregnancy had to be clarified [2]. In humans, the studies are almost
exclusively focused on the PPARγ roles during early placentation. It
has been clearly established that all three PPARs can stimulate or inhibit the
differentiation and/or proliferation of the villous cytotrophoblasts into
syncytiotrophoblasts and the synthesis of chorionic gonadotrophic hormone, and
may hamper extravillous trophoblastic cell invasion (for more details, see
Fournier et al., 2007 [4]).


## 3.3. Roles of PPARs in the Uptake and Transport of Trophoblastic Lipids

As one of the first
functions described for PPARγ in other tissues, trophoblastic lipid uptake and
accumulation are also regulated in part by this factor [27]. The PPARγ ligands seem to increase the uptake and accumulation of the fatty acids
in human placenta [28]. This regulation is associated with
an enhanced expression of adipophilin (fat droplet-associated protein) and fatty acid transport
proteins (1 and 4) in human trophoblasts [28–30]. These results were confirmed
recently by the in vivo activation of PPARγ by its agonist rosiglitazone in
mice, which also leads to the enhancement of the previous described genes plus
two new ones involved in the lipid transport: S3-12 (plasma associated protein)
and myocardial lipid droplet protein/MLDP [27]. Taken together, these results
confirm the results obtained on PPARγ-null mutants: the absence of the
lipid droplets normally present around the fetal vessels in the wild-type
placenta [21].


## 3.4. PPARs in Placental Inflammatory Response and in the Parturition Signalling

At this stage of our knowledge of PPARs, the most interesting results have been
obtained with the study of their involvement in the inflammation process,
which may be linked to labor at term and also to the premature rupture of fetal
membranes (see Figure3). Term labor is associated with an increase in proinflammatory
proteins and cytokines such as IL1β, IL6, IL8, IL10, and TNF-α. This increase in
proinflammatory proteins and cytokines induces uterine contractions. PPARγ ligands have been demonstrated to inhibit the secretion of IL6, IL8,
and TNF-α in amnion and chorion [31], highlighting the role of PPARs in
the regulation of the inflammatory response in human gestational tissues and
cells [32–35]. The parathyroid hormone-related
protein (presenting a cytokine-like action) is involved in many processes
during normal and pathological pregnancies, and is decreased by PPARγ stimulation [36], which also blocks proinflammatory
cytokine release by adiponectin and leptin [37]. The production of prostaglandins
by the endometrium, the myometrium, and the fetal membranes induces the
contraction of the myometrium during labor. This generation of uterotonic
prostaglandins correlates with the increased prostaglandin-endoperoxide
synthase type 2/cyclooxygenase type 2 (COX-2) activity and the increased secretory phospholipase A2-IIA (sPLA2) mRNA, proteins and activities. By inhibiting the production of the COX-2 and
sPLA2 in fetal membranes, PPARγ promotes the quiescence of the
uterus during gestation [34]. The molecular action of 15dPGJ2
seems to involve interactions of the NF-Kappa B signaling pathway, inducing
reduction of PGF2α, PGE2, and MMP9 release in the placental
environment [31]. This suppressive action of PPARγ
on inflammation is aPPARently time-dependent during pregnancy. The PPARγ level
of expression remains stable throughout gestation, except for the period just
before labor, when its expression in fetal membranes declines. This reduction
is coincidental with a relative increase in COX-2 expression [38]. Further work has shown this simple
scheme to be more complex. While the expression of PPARα does not change at
term in amnion, it decreases in chorion. An increase was also demonstrated for
PPARβ/δ in chorionic and amniotic zones [11]. These last two findings raise the
question of the involvement of the α and β isoforms in this process. The
absence of a real link between COX-2 and PPARγ is presented by Lindstrom and Bennett [39]. Finally, the PPAR action seems to
be concentration-dependent. A small amount of 15dPGJ2 (<0.1 μM) acts through
the PPARγ signaling pathway, where at high
concentration (1 μM) its actions are most probably mediated through
other pathways: PPARβ/δ and/or an inhibition of NF-κB
independent of PPARs [35]. Furthermore, 15dPGJ2 and troglitazone were also demonstrated to have some antiinflammatory or apoptosis-induction
specific effects by PPARγ-independent pathways. This was suggested by the work of Lappas et
al. on human gestational tissues, demonstrating that this effect could
passed by antagonist effect of 15dPGJ2 on the NF-κB pathways and
antioxidant effects of the troglitazone, a synthetic ligand of PPARγ [31].Figure 3
Schematic representation of PPARγ implication in pregnancy maintenance and labor.  IL1β: Interleukin 1β; IL6: Interleukin 6; IL8: Interleukin 8; IL10: Interleukin 10, TNFα: Tumor Necrosis Factor α; COX2: Cyclo-oxygenase type 2; PLA2: Phospholipase A2; NF-κB: Nuclear Factor-Kappa B; MMP9: Matrix Metalloproteinase 9; 15dPGJ2: 15-Deoxy-Δ12, 14-prostaglandin J2.


## 3.5. PPARs in Placental and Amniotic Membranes Pathologies

In contrast to the
different roles described for PPARs during human placentation, only a few
studies on PPARs and placental pathologies have been conducted. In choriocarcinoma
and hydatiform moles, a downregulation of the PPARγ expression is observed but
this real influence needs to be elucidated [40]. The potential involvement of PPARγ on preeclampsia is suggested by the fact that this pathology is
associated with an increased peroxidation in trophoblasts [41, 42]. An overproduction of 15-HETE has
also been noted, suggesting a deregulation of PPARγ [43]. This can cause a strong
transactivation of PPARγ during early pregnancy, resulting
in a reduction of extravillous trophoblastic invasion, one cellular explanation
often cited in the physiopathology of preeclampsia [44, 45]. Other abnormal transactivation of
PPARs may be hypothesized to explain placental pathologies. The 15dPGJ2 has
been shown to induce apoptosis of the placental (JEG-3) and amniotic (WISH) established cell
line, [46, 47]. An excess of 15dPGJ2 production can
be a source of placental dysfunction linked to an increase in trophoblastic
death. It is also established that deletion of PPARγ, PPARβ/δ, and some of their coactivators (PBP, PRIP, and
RAP250) induce abnormal placental phenotypes (abruption,
reduction of fetomaternal exchanges, and alterations of trophoblastic
differentiation) in null mutants [21, 23, 24, 26, 48, 49]. Chromosomal and/or genetic alterations
(point mutation or deletion) may occur for these genes, inducing human
placental alterations. The placental 11β hydroxysteroid dehydrogenase type
2 is a target gene of PPARs [50]. This enzyme plays a key role in
fetal development by controlling fetal exposure to maternal glucocorticoids. An
abnormal regulation by PPARs may result in an absence of fetal protection. In
the rat placental HRP-1 established cell line, the phthalate and derivatives
transactivate PPARs (α and γ) induced an increase in uptake rates of fetal essential fatty acid and
the transport of arachidonic and docosahexaenoic acid [51]. If such a mechanism can be induced
by the phthalates during human placentation, this may strongly affect the fetal
essential fatty acid content during growth.Gestational diabetes is
linked to impaired lipids metabolism [52]. Decreased 15dPGJ2 in blood of
diabetic mothers is also linked to a decrease in
placental PPARγ expression. The inhibition of PPARγ results in an induction of a placental proinflammatory environment associated with an
increase in nitrogen monoxide production and release, which can impair fetoplacental
development [53, 54].The PPAR regulation of
inflammation may be very important in another obstetrical pathology of the
amniotic membranes: the chorioamnionitis. This pathology, usually due to an
ascendant colonization of pathogenic microorganisms from the vagina to the
uterus, is closely associated with preterm labor and premature rupture of
membranes (chorion and amnion). These
ruptures of membranes seem to arise from deregulated proinflammatory factor
synthesis. It has already been reported in this pathology that IL1β, IL6, IL8,
TNF-α, and prostaglandinE(2) show
inadequate concentrations in placental membrane and in amniotic fluid [55–58]. As PPARs may be involved in the occurrence
and control of this inflammatory response, further studies are needed to assess
their importance in this process and to find new possible therapeutic strategies
to prevent this damaging pathology.More generally, the use of natural and
synthetic PPAR ligands looks to be a promising way in preventing placental
pathologies such as endometriosis or preeclampsia. An interesting study also
demonstrates that the reduction of LPS induction of cytokines is reduced by
PPARγ ligands in fetal membranes. Nevertheless, the few studies already
conducted were done practically only on animal (rodent) models and looks to
have positive effects on the pathologies (for review see Toth et al. [59]). Till now, the major problem using,
for example, TZD (thiazolidinedionzes) linking to the PPARγ pathways still the numerous adverse effects of this kind of treatment (e.g.,
weight gain, anemia, leukopenia, etc.). These facts and the potential placental
impacts raised also the question of the use of these medical drugs to treat the
gestational diabetes. Perhaps, at the level of clinician actual knowledge, PPARγ and its ligands could be used  in
a first time, only as good early marker candidates for the diagnosis of
pregnancy pathologies like, for example, preeclampsia.


## 4. Conclusion

Since the discovery of the PPARs, there
has been a marked increase in available data on their involvement in mammalian
development. Concerning the placenta, all PPARs, but particularly PPARγ, are
essential for multiple physiological functions of the trophoblastic and
amniotic parts, leading to major involvement of PPARs in the pathophysiology of
gestational diseases. However, special care must be taken when this particular PPAR
signaling cascade is involved, because part of the regulation may involve PPAR
ligand signalling (by the natural 15dPGJ2 ligand or the troglitazone
synthetic ligand) but may be transduced by independent nuclear receptor
pathways (as, e.g., by antagonizing effects on NF-κB pathway for 15dPGJ2 and by acting as an antioxidant for troglitazone). This last point
introduces a new level of complexity in PPAR biology. It does not close preclusion of the
eventual use of PPARs for therapeutic treatment during pregnancy, but future
medical applications seem still to be a long way off. We can reasonably expect
to see some obstetrical use of PPARs in diagnosis (detection of PPARs mutations
in intrauterine growth retardation, predisposition of preeclampsia) and
therapeutics (tocolysis or treatment of chorioamniotis).



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*Source: 758562-2008-01-20.xml*