Chapter 8

Premature Rupture of Membranes

and Oxidative Stress

Anamar Aponte and Ashok Agarwal

Abstract Premature rupture of membranes (PROM) is rupture of the chor-

ioamniotic membranes before the onset of labor. The chorioamniotic sac requires a
balance between collagen formation and enzymatic collagenolytic activity
expressed in the fetal membrane. The amniotic membrane is a complex tissue in
which collagen is fundamental for mechanical integrity and stress tolerance.
Membrane rupture is associated with biochemical disturbances between collagen
produced by fibroblasts and fetal membranes. An association exists between
PROM and oxidative stress (OS) in pregnancy. Reactive oxygen species (ROS),
which are unstable molecules produced in the body, may be the reason for the
collagen damage. OS produced by augmented ROS production debilitates the
collagens elasticity and strength, leading to PROM. Although antioxidants act as
defense agents against oxidation, they have not been found to be protective against
ROS and subsequent OS damage in PROM.

Keywords Premature rupture of membranes

Chorioamniotic membranes




Oxidative stress Enzymatic collagenolytic activity expressed Fetal membrane

A. Aponte
A. Agarwal (&)

Center for Reproductive Medicine, Lerner College of Medicine, Cleveland Clinic,
9500 Euclid Avenue, Cleveland, OH 44195, USA
e-mail: agarwaa@ccf.org
A. Aponte
e-mail: anamar06@hotmail.com

A. Agarwal et al. (eds.), Studies on Womens Health, Oxidative Stress in Applied 143
Basic Research and Clinical Practice, DOI: 10.1007/978-1-62703-041-0_8,
Springer Science+Business Media New York 2013
144 A. Aponte and A. Agarwal

8.1 Introduction

Parturition is determined by ripening of the cervix, augmented contractility of the

myometrium, and positive signaling of the maternal decidua and the chorioamn-
iotic membranes. Activation of genes involved in inflammatory activities promotes
the chorioamniotic membrane to undergo complex biochemical changes [1].
Premature rupture of membranes (PROM) is defined as rupture of membranes
before the onset of labor and membrane rupture that takes place before 37 weeks
of gestation is known as preterm PROM [2]. At term, weakening of the membranes
occurs from physiologic changes in association with shearing forces created by
uterine contractions [3, 4]. Normal rupture of membranes at term is thought to
begin at a specific weakened para-cervical region which forms late in gestation due
to collagen disruption [5]. Intraamniotic infection is commonly associated with
preterm PROM, especially if it occurs at an earlier gestational age[2]. Low
socioecononomic status, second- and third-trimester bleeding, low body mass
index less than 19.8, nutritional deficiencies of copper and ascorbic acid, con-
nective tissue disorders, maternal cigarette smoking, pulmonary disease in preg-
nancy, uterine overdistention, and amniocentesis are some of the risk factors
associated with PROM occurrence [6, 7].
The amnion is an avascular entity that depends on the amniotic fluid for its
stability and may be involved in the pathogenesis of PROM [8]. The chor-
ioamniotic sac requires a balance between collagen formation by fibroblasts and
the collagenolytic activity by enzymes expressed in the fetal membrane [9].
Membrane rupture is associated with biochemical disturbances between collagen
produced by fibroblasts and fetal membranes [9]. The elasticity and strength of
fetal membranes is primary determined by collagen. Reduced concentrations of it
or altered cross-linked organization may contribute to PROM [10].
MnSOD formation. MnSOD is a member of an iron/manganese superoxide
dismutase family which is located in the mitochondria and inhibits reactive oxygen
species (ROS) produced by oxidative phosphorylation [1]. A moderate increase in
ROS can stimulate cell growth and proliferation, while an excessive ROS accu-
mulation will lead to cellular injury, such as damage to DNA, protein, and lipid
membrane.ROS enhance NF-jB and promote the expression of cyclooxygenase-2
(COX-2), which upregulates the inflammatory pathway and the propagation of
labor [1]. These inflammatory processes may lead to increased formation of ROS
and subsequent oxidative stress (OS). OS occurs when the production of ROS
exceeds the endogenous antioxidant defense. On the contrary, MnSOD may be
activated by inflammatory processes, which in turn downregulates OS and
inflammation by inhibiting NF-jB and mitogen activated protein pathways
(MAPK) [1].
8 Premature Rupture of Membranes and Oxidative Stress 145

8.2 Chorioamniotic Membrane: What is it Made of?

The amnion is a single layer of non-ciliated cuboidal cells with its outermost part near
the amniotic fluid. Beneath the amnion there is a layer that consists of loosely packed
fibrils and where the chorion, which composed of reticular fibers, a basement
membrane, and the trophoblast cells, is found. Under the chorion is the maternal
decidual layer, which consists of epithelial cells that allow the separation of the
chorioamnion from the endometrial and myometrial layers of the uterus [11].
The amnion originates its strength from collagen. At least five (types I, III, IV,
V, and VI) types of collagen are encountered in the chorioamnion which are
organized in triple helices [10]. Collagen strength, which is primarily attributed to
collagen I, is derived from hydroxyproline and hydroxylysine bridges around the
helix. The reticular layers contain collagen I, III, IV, V, and VI. The chorionic
basement is composed of collagen IV and the chorion has types IV and V [12].
The chorioamnion is a biologically active membrane whose collagenolytic
enzyme is susceptible to ROS proliferation [13].

8.3 Premature Rupture of Membranes, Oxidative

Stress, and Antioxidants

In the normal physiological capacity, a delicate balance exists at the molecular

level between oxidants and antioxidants. ROS may play a role in the collagen
damage in the chorioamniotic sac leading to tearing [9]. OS occurs when the
production of reactive oxygen species exceeds the antioxidant defense. OS may
altercate the elasticity and strength of collagen and promote PROM [10, 14].
Isoprostanes (F2-IP) can be used to a marker of OS [15], especially of lipid
peroxidation [9]. They are prostaglandin-like products produced by free radical
catalyzed nonenzymatic peroxidation of arachidonic acid [9]. Peroxidation
decreases, disrupts membrane barrier function and lowers its fluidity [16]. F2-IP
levels in the amniotic fluid at 1518 weeks gestations are a predictor of PROM in
preterm deliveries [9].
The antioxidants play a role in the protection of the chorioamniotic sac from
oxidant damage [1720]. Vitamin C (ascorbic acid) is a known redox catalyst,
which has the ability to reduce and neutralize ROS. It plays a primordial role in the
formation of collagen triple helix and the fortification of collagen cross-links[5].
Vitamin E is a lipid soluble vitamin with antioxidant activity. It constitutes of eight
tocopherols and tocotrienols. a-Tocopherol is considered to play a central role due
to its capability to react with lipid radicals produced during lipid peroxidation
reaction. In vitro, Vitamins C and E have been successful in the protection of the
amnion and chorion from damage induced by ROS [8]. Mechanisms have pro-
posed that Vitamin C may salvage oxygen species in the amniotic fluid and
Vitamin E may prevent lipid peroxidation [8]. In contrast to the reports implying a
146 A. Aponte and A. Agarwal

protective action, Bolisetty et al. [20] found that antioxidant vitamins, especially
vitamin E, diminish markers of OS at birth. Mathews et al. [8] found no evidence
to support the hypothesis that antioxidants against PROM. Finally, Mercer et al.
[5] postulated that antioxidant treatment with Vitamin C and E may inhibit ROS
formation and resultant fetal membrane weakening. Two studies were conducted,
one in vitro and the other in vivo, and concluded that neither antioxidants were
effective in the prevention of PROM.
Superoxide dismutases are antioxidant enzymes that protect cells against ROS
and damage [2123]. There are four classes that differ by, their protein configuration,
the intra- or extracellular localization and the active metal ions (Cu/Zn, Mn, Fe, or
Ni) in their catalytic centers [21]. In humans, MnSOD is encoded by SOD2, which is
synthesized in the cytosol and imported into the mitochondrial matrix [1]. Than et al.
[1] showed MnSOD mRNA expression in the fetal membranes and in chorioamni-
onitis of PROM suggesting an antioxidant mechanism to counteract the inflamma-
tory process in the chorioamniotic membranes.

8.4 Conclusion

PROM is characterized by reduced collagen concentrations, altered collagen cross-

link profiles, and increased concentrations of biomarkers of oxidative damage. In
the amniotic membrane, collagen is primordial for mechanical integrity and stress
tolerance. The OS that occurs during pregnancy increases the risk for PROM that
is caused by changes in collagen integrity. In the future, trials should focus on
determining if supplementation with antioxidants could protect the fetal mem-
branes from premature rupture.

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