Impact of Oxidative stress on Infertility, with emphasis on infertility management strategies

Oxidative stress (OS) is a state of imbalance between pro-oxidants (Reactive Oxygen species and Reactive Nitrogen species) and antioxidant defences. It plays a pivotal role in infertility issues (paternal as well as maternal). As such different reproductive diseases such as Polycystic Ovary Syndrome (PCOS), Endometriosis, unexplained infertility, Impairment of Spermatozoa, Sperm Dysfunction and Sperm DNA Fragmentation (SDF), are caused, which destroys the process of reproduction. Several pregnancy complications such as miscarriage, recurrent respiratory loss (RPL), preeclampsia and intrauterine growth restriction (IUGR), are also associated to OS. It is well established that factors such as obesity, malnutrition, life style factors – smoking, alcohol and recreational drugs, environmental and occupational exposures – pesticides and endocrine disrupting chemicals, EDCs, hampers the reproductive process by setting up OS. Thus, as an infertility management strategy, procedures such as antioxidant supplementation and Assisted Reproductive Techniques (ART), are to be undertaken. There is also a wide scope for research in the fi eld of EDCs linked to oxidative stress, which can be a hope to further elucidate the path of reproduction. Review Article Impact of Oxidative stress on Infertility, with emphasis on infertility management strategies Pamela Banerjee1* and Jayashree Bhattacharya2 1Senior Research Scientist, A.H IVF & Infertility Research Centre 853, Kalikapur Road, Kolkata 78, India 2MD, MRCOG, FRCOG, London, UK, Director, A.H IVF & Infertility Research Centre, 853, Kalikapur Road, Kolkata 78, India Received: 05 May, 2019 Accepted: 31 July, 2019 Published: 02 August, 2019 *Corresponding author: Pamela Banerjee M.Sc PhD, Senior Research Scientist, A.H IVF & Infertility Research Centre 853, Kalikapur Road, Kolkata 78, India,Tel: +919830915163; Email:


Introduction
Oxidative stress is the root for many disease conditions such as Infertility. It is caused due to an imbalance between prooxidants and antioxidants [1]. This imbalance is created due to enhanced levels of ROS (Reactive Oxygen Species) and / or RNS (Reactive Nitrogen Species) in plasma and blood of patients.
In addition, there is often a decreased level of antioxidant mechanism, thereby deteriorating the balance ratio.
As oxidative stress is one of the major causes for infertility, excessive generation of ROS causes an environment unsuitable for normal female physiological reactions [1]. This leads to reproductive diseases such as PCOS, Endometriosis and unexplained infertility in women [2]. Pregnancy complications such as miscarriage, recurrent respiratory loss (RPL), preeclampsia and intrauterine growth restriction, IUGR, are all linked to oxidative stress [3].
Not only maternal, but paternal reproductive disease conditions are also due to oxidative stress. In this review, some of them such as Impairment of Spermatozoa, Sperm Dysfunction, Sperm DNA Fragmentation (SDF), are discussed briefl y.
Various factors are also responsible for infertility, such as obesity, malnutrition, life style factors -smoking, alcohol and recreational drugs [4], environmental and occupational exposures -pesticides and endocrine disrupting chemicals, EDCs [5][6][7]. These factors are linked to oxidative stress.
This problem of infertility due to oxidative stress or other reasons, can be solved by Antioxidant supplementation such as glutathione and/ or Assisted Reproductive Techniques, ART [2], fi nally to bring a smile on the couple's face. Measurement of oxidation-reduction potential by MiOXSYS ® system, is yet another approach to solve the problem of oxidative stress [8].

Male Infertility
i) Impairment of Spermatozoa in humans: Impaired sperms are one of the major causes of infertility. It has been documented that the nucleohistone compartment of sperms which contains histone-bound DNA sequences like telomeres and promoters of genes for embryonic development, are often at a high risk of oxidative stress, leading to infertility [8,9]. Also, the relative decrease in protamine 2 levels, in the ratio of protamine1: protamine2, at the level of m-RNA and protein, also results in infertility. Again, gene mutations encoding protamines lead to structural changes in sperm chromatin structure, thereby leading to infertility [10,11].
ii) Sperm dysfunction : The mechanism of sperm dysfunction is related to the oxidative stress, which actually causes distortion of the sperm DNA integrity, by damaging proteins and lipids present in sperm cell plasma membrane. This has a negative effect on sperm cell membrane fl uidity and permeability, leading to infertility [12]. Also, it has been established that sperm motility gets reduced as ROS becomes directly proportional to the level of lipid peroxidation [13]. The sperm dysfunction is also related to the lipid peroxidation cascade activation, initiated by ROS-mediated oxidation of lipids in sperm cell membrane. Figure 1, explains the same.
iii) Sperm DNA Fragmentation (SDF): According to the researchers, SDF is mediated by free-radicals [14,15]. Again, the mechanisms underlying SDF include single-strand and double-strand breaks, DNA fragmentation, the introduction of abasic sites, purine, pyrimidine and deoxyribose modifi cations and DNA crosslinking, which can result in arrest or induction of gene transcription, induction of signal transduction pathways, accelerated telomeric DNA attrition, repli cation errors, genomic instability and GC to TA trans versions [16][17][18]. As such mechanisms are also known to be the causes of carcinogenesis, these may explain a link between infertility and cancer. Also, male infertility is closely associated with ROS-induced DNA damage, which in turn accelerates the germ-cell apoptosis process, leading to decline in sperm count [19]. Figure 2 explains the same.

Female Infertility
i) Polycystic Ovary Syndrome (PCOS): The word "polycystic" means "many cysts. PCOS is said to develop when within the ovaries, numerous minute fl uid-fi lled sacs appear. Each of these sacs is a follicle which holds an immature egg. These eggs do not develop to complete the ovulation process. Again, due to this lack of ovulation, there is a change in the hormonal levels of the female body. For eg. estrogen and progesterone levels get reduced while androgen levels are higher. The menstrual cycle in the females therefore, gets disturbed due to hyperandrogenism. So, women with fewer periods than normal, often lead to PCOS. Figure 3

Oxidative stress (OS) and PCOS
Oxidative stress leads to PCOS. It is well established, that ROS (Reactive Oxygen Species) is more in PCOS patients, especially for insulin resistance patients [22]. Also, a decrease in GSH content and GSH to GSSG ratio in leukocytes suggests a deterioration of mitochondrial function and an increase in  oxidative stress leading to PCOS. As Protein carbonyl (PC) is an important biomarker of oxidative stress, it is observed that PC content is higher in PCOS patients compared to control. Further, PC content is known to have a positive correlation with fasting insulin, suggesting a strong association between insulin resistance and protein oxidation in PCOS. Malonal dialdehyde or MDA (produced during decomposition of polyunsaturated fatty acids) is one of the biomarkers of OS. It is seen that blood MDA levels are signifi cantly (P= 0.01) higher in young, lean, non-IR PCOS patients, in comparison to the control [23]. Again, SOD (Superoxide dismutase) levels are higher in PCOS patients, as compared to control. As IL-6 (Interleukin-6) and TNF- (Tumor necrosis factor alpha) are pro-infl ammatory markers of oxidative stress, it is well established that IL-6 is more in PCOS patients and shoots up in the case of PCOS-IR patients [22]. TNF-, on the other hand is higher in both PCOS-IR as well as PCOS-non IR patients.
ii) Endometriosis: Endometriosis occurs when the tissue that forms the uterine lining (the lining of the womb) is present on various organs within the body. Endometriosis is usually found in the lower abdomen, or pelvis, but can also appear elsewhere in the body. Endometriosis is explained by Figure 4.
Women with endometriosis often suffer from lower abdominal pain, pain with periods, or pain with sexual intercourse, and may report having a hard time getting pregnant. However, some women with Endometriosis, may be asymptomatic.

Endometriosis and Fertility
Between 20 and 40% of women with infertility, normally suffer from endometriosis. Endometriosis seems to impair fertility in 2 ways: fi rstly, by causing deformation in the fallopian tubes so that they are unable to pick up the egg after ovulation, and secondly, by causing infl ammations that in turn causes impairment of the functioning of the ovary, egg, fallopian tubes or uterus. Figure 5 shows the endometriosis tissue on the bladder wall and left ovary.

Pregnancy complications
i) The Placenta: In cases of incomplete placentation, oxidative stress occurs at an excessive level and leads to an elevated release of placental debris and vesicles into the maternal circulation. These extracellular vesicles carry active molecules (proteins of microRNA) generated by stressed placental cells. Once in the blood, they meet maternal endothelial cells and potentially transfer their contents, leading to transcriptome alterations and infl ammation. Eventually, the endothelium of maternal organs is affected along with the deterioration of synciotrophoblast [24]. Figure 6 explains the effect of oxidative stress on placental function and pathological events in pregnancy.    ii) Miscarriage or Spontaneous Abortion: Miscarriage or spontaneous abortion is the unintentional termination of pregnancy when the fetal weight is < 500g. Placental oxidative stress is one of the major cause of miscarriage. Due to premature intraplacental circulation at 8 to 9 weeks of pregnancy, instead of 10-12 weeks of gestation, high oxidative stress is set up [25,26] which distorts the syncytiotrophoblast layer of the placenta, thereby causing impairment of the placenta [27]. Apoptosis of the placental tissues may also occur due to oxidative stress-induced infl ammatory processes. Hence, due to this degeneration of the placenta, miscarriage often occurs [2]. Figure 7 depicts the types of spontaneous abortion.

iii) Recurrent Pregnancy Loss (RPL):
Recurrent pregnancy loss refers to more than three consecutive pregnancy losses.
The reason behind this situation is the enhancement of Natural Killer cells (NK cells) in placental blood circulation, thereby increasing pre-implantation angiogenesis. This in turn causes premature intra-placental maternal circulation, leading to very high oxidative stress [28]. It has been well established, that in patients with RPL, there is an enhancement of lipid peroxides and GSH in plasma [29], in addition to the reduction in levels of vitamin E and ß-carotene, indicating en elevated level of oxidative stress [30]. Another study, reveals that there is signifi cantly decreased levels of antioxidant enzymes like GPx, SOD and catalase, in addition to increased MDA levels, in patients with RPL, again indicating augmented oxidative stress [31]. Figure 8, explains the etiology of Recurrent Pregnancy loss, that is, Primary versus Secondary. iv) Preeclampsia: Preeclampsia refers to a complex multisystem disorder that is characterized by high blood pressure pregnancy complication. This is a leading cause of maternal and fetal morbidity, accounting to 3% to 14% of pregnancies worldwide [32,33]. In this context, Placental ischemia /hypoxia plays a very important role in augmenting the oxidative stress, which in turn leads to endothelial cell dysfunction [33,34] and systemic vasoconstriction [35]. Also, it is seen that the oxidative stress has contributed to the apoptosis of villous trophoblasts, in patients with Preeclampsia [2]. Figure 9 explains Preeclampsia and Placental Abruption, diagrammatically.

Factors governing Infertility, associated with Oxidative
Stress :

Age:
Age is a vital factor for infertility, as fertility declines with advancing mother's age. Paternal age is also important in terms of fertility, as retrogression in gamete quality, increased levels of oxidative DNA damage (ODD) and decline in semen quality. Hence, sperm DNA is often exposed to oxidative stress because of advancement in paternal age [9,36].

Body weight
i) Obesity: The pathogenesis of obesity is closely associated to the excessive generation of ROS, thereby setting up of Oxidative stress [32]. Hence, overweight can complicate pregnancy issues, both in terms of maternal as well as fetal health. Also, dysfunctional hormonal system and irregular menses, are known to be closely associated with overweight or obesity [38].

Environmental and Occupational exposures
i) Pesticides: It has been well documented that the spouses of agricultural male workers who are in direct contact with pesticide chemicals like DDT (Organochlorine pesticides), are victims of miscarriage or spontaneous abortion [50].
Again, polychlorinated biphenyls or PCBs (pesticide) are known to enhance free radical generation by setting up oxidative stress, through induction of endothelial dysfunction [51] and cell membrane destruction [2]. Interestingly, PCB exposure causes suppression of Vitamin E levels [51].
Organophosphate pesticides on the other hand, causes depletion of GSH along with increase in ROS, triggering oxidative stress [52][53][54]. Figure 12, explains the role of pesticides in infertility.

ii) Endocrine Disrupting Chemicals (EDCs): Endocrine
Disrupting Chemicals (EDCs) like the phthalates acts as an anti-androgenic compound, deteriorating the male reproductive system and is associated with the setting up of the oxidative stress [55]. Also, it has been documented, that there is an association between urinary phthalate metabolites and polyvinyl chloride workers [55]. After taking into consideration, the age, smoking status and coffee consumption, it has been observed that the urinary phthalate metabolites causes sperm   As oxidative stress is one of the main causes of infertility, antioxidant supplementation is required to scavenge the free radicals, thereby creating a balance between ROS and antioxidants. Antioxidants such as vitamin C, vitamin E, selenium, zinc and glutathione are important for male infertility [56,57]. Table 1, depicts the antioxidants, their characteristics and effects on semen parameters.

Glutathione -the principal antioxidant for male infertility
Glutathione prevents male infertility by scavenging lipid peroxides in the plasma membrane of spermatozoa, thereby reducing the oxidative stress. It also scavenges hydrogen peroxide, thereby arresting lipid peroxidation and reducing oxidative stress [20].

Glutathione -the principal antioxidant for female infertility
During folliculogenesis, Glutathione shields eggs from oxidative stress [20]. Researches, in this context have thus proved that oocytes with elevated levels of intracellular glutathione produce healthier and stronger embryos [58]. Figure 14, explains the antioxidant supplementation important for female fertility.

Glutathione -impact on autoimmune issues
Glutathione is indeed a boon for those with immunological miscarriages or if the body is rejecting one's mate's sperm [20].

ii) Assisted Reproductive techniques
Assisted Reproductive techniques are the treatments of choice for the infertile men or women. In cases of endometriosis, tubal factor infertility, male factor infertility and unexplained infertility, these techniques can bring a new born baby to the affected couple [59].
These techniques include intrauterine insemination, IVF and intracytoplasmic sperm injection (ICSI). The same have been explained by

Conclusion
As oxidative stress is one of the major causes of infertility ( Figure 18), couples especially must include antioxidants in Figure 12: Explains role of pesticides in infertility [72].  and endometriosis [60] need to be carried out, in order to elucidate the path of human fertility. Advancement in science and technology have also gifted techniques such as Assisted Reproductive Techniques (ART), which aims to bring a smile on the couple's face.