Oxidant stress and a potential role for antioxidant therapy
Infertility affects up to 15% of Canadian couples and may be attributed to a number of diverse factors. Recent evidence suggests that while oxidative processes play an essential role in human reproduction, a state of oxidative stress may contribute significantly to the inability to conceive.Oxidative stress has been implicated in endometriosis, recurrent pregnancy loss and poor embryo quality. Human studies into the effect of antioxidants upon reproductive outcomes have shown some promising results. Interventions including vitamin A, vitamin E, n-acetyl cysteine, and melatonin may quench oxidative stress, while observational data suggests a possible role for CoQ10. Antioxidants may offer novel therapeutic options in the management of female infertility.
Infertility is defined as a “failure to achieve a successful pregnancy after 12 months or more of regular unprotected intercourse” (PCASRM 2008). Recent surveys suggest that between 11.5% and 15.7% of Canadian couples attempting to get pregnant are dealing with an inability to conceive, with prevalence rates increasing with advancing maternal age (Bushnik 2012). It is estimated that approximately 30% of infertility causes may be attributed to male factors and 40% to female factors. In the remainder of cases, a combination of influences or an undetermined cause are deemed to be responsible (AHRC 2010).
Through diagnostic imaging and laboratory assessment, many causes of female infertility may be identified and treated. Common causes include disturbances to ovulation caused by polycystic ovarian syndrome (PCOS) and its accompanying insulin resistance and androgen dominance, and disruptions to the physical structure of the reproductive tract as a result of fibroids, endometriosis and pelvic inflammatory disease. Factors external to the reproductive tract such as age, genetics, smoking status and toxin exposure history can also influence the viability of reproductive cells and the success of attempts at pregnancy.
Endocrine functioning, involving the complex interplay of reproductive hormones (estrogen, progesterone, luteinizing hormone and follicle stimulating hormone), thyroid hormones (thyroid stimulating hormone, T3, T4, reverse T3), prolactin, melatonin, insulin and cortisol, has an equally important role in determining the success or failure of any attempts at conception. This component presents perhaps the greatest challenge to those working to support conception. Factors such as body composition, dietary choices, exercise and exposure to stress may have significant impacts on the functioning of this elegant, dynamic system.
In spite of our ability to image, measure and quantify so many aspects of the reproductive tract, a clear cause of infertility cannot be identified in a number of cases (Ledger 2009). These unexplained cases may be attributed in the future to other pathological processes that are the subject of current research including immune functioning (Siam 2011), genetic enzymatic variants (Eloualid 2012) and signaling peptides (Sadeu 2012). A role of oxidative stress in infertility has also been proposed by numerous authors (Agarwal 2005, Ruder 2009, Visioli 2011) and forms the focus of this present analysis.
Free radicals and oxidative stress
Free radicals, comprising two main classes of reactive oxygen species (ROS) and reactive nitrogen species (RNS) are unstable compounds that are produced through many physiological processes. While they are essential to some functions of the body such as infection control (Valko 2007), free radicals have the potential to cause significant tissue damage and disease. For this reason, mechanisms within the body work to stabilize free radicals and to neutralize the damage they may cause. Antioxidant compounds and enzymes accomplish this function on an ongoing basis (Agarwal 2005). When the burden of reactive species overwhelms the compensatory mechanisms of the body, oxidative stress occurs, causing damage to cellular structures and DNA (Valko 2007).
Oxidative processes in reproduction
As in the rest of the body, oxidative processes are integral to the proper functioning of the reproductive system. Key functions such as follicle and oocyte follicular development, embryonic development and implantation (Agarwal 2005, Wiener-Megnazi 2011) involve ROS. However, as oxidative stress has also been implicated as a causative factor in age-related fertility decline (Keefe 2009), it is clear that oxidative processes are not wholly supportive of reproductive processes.
Oxidative stress and endometriosis
Oxidative stress has been put forth as a contributing factor in women with endometriosis, a significant cause of female infertility (Augoulea 2009). Women with this presentation have been found to have a lower antioxidant capacity, as evidenced by decreased levels of plasma superoxide dismustase in one recent investigation (Prieto 2012). Oxidative processes, originating in the peritoneum, are thought to contribute to the development of endometriosis (Gupta 2005, Lousse 2012) and affect not only the structure of the reproductive tract, but oocyte quality in these women (Saito 2002). It has also been suggested that more advanced cases are associated with more evidence of systemic oxidative stress (Andrade 2010).
The administration of antioxidant therapies may provide a novel strategy for the management of this reproductive concern. One recent trial demonstrated that levels of malondialdehyde (MDA), a marker of oxidative stress, could be significantly attenuated by low doses of vitamins A and E (343mg and 84mg respectively) over a six-month period (Mier-Cabrera 2008). At the end of the study, pregnancy rates were slightly higher in the treatment group but results did not reach significance. Future trials with higher doses of targeted antioxidant compounds may hold promise in the treatment of this common cause of female infertility.
Recurrent pregnancy loss (RPL)
Recurrent pregnancy loss, the spontaneous termination of three or more pregnancies under 20 weeks gestation (Gupta 2007), may also prove to have an association with oxidative mechanisms. One study of 45 women that had experienced recurrent pregnancy loss found significantly decreased total antioxidant capacity (TAC) and increased total oxidative status (TOS) among these women compared to healthy pregnant controls (Toy 2010). This evidence is supported by an earlier study identifying low activity of the paraoxonase-1 enzymatic system that prevents lipid oxidation compared to healthy controls (p<0.01). High levels of lipid hydroperoxide in these same women demonstrated increased levels of oxidative stress in these individuals (p<0.01). It should be noted here that other studies have identified oxidative stress as a result rather than a cause of RPL (Baban 2010).
N-acetyl cysteine (NAC) is a mucolytic compound that has been discussed previously in reference to its application in polycystic ovarian syndrome (PCOS) (Flower 2011), where it appears to improve insulin sensitivity and reduce resistance to clomiphene therapy (Abu Hashim 2010). A recent prospective study (Amin 2008) assessed the suitability of NAC in the treatment of RPL, reporting a significant decrease in the risk of pregnancy loss when treatment with a combination of NAC (0.6g) and folic acid (500mcg) was initiated at the time of pregnancy confirmation. In comparison to folic acid therapy alone, the addition of NAC greatly improved the rate of pregnancy maintenance up to 20 weeks (RR 2.9, 95% confidence interval (CI) 1.5-5.6). Perhaps most importantly, the so-called take home baby rate was significantly higher in the NAC-treated group (RR 1.98, 95%CI 1.3-4.0).
Given the established antioxidant activity of NAC (Amin 2008), coupled with the evidence of increased oxidative stress in women suffering RPL, it is reasonable to postulate that this intervention improved pregnancy outcomes by attenuating the predominance of oxidative reactions in the body. It is not yet known whether this effect is achieved through direct action of NAC as a scavenger of free radicals or whether it counters oxidation more indirectly by increasing endogenous glutathione levels (Amin 2008). NAC and other antioxidants may have an important role to play in the management of this devastating condition.
Oxidative factors and in-vitro fertilization (IVF)
IVF, with or without intracytoplasmic sperm injection (ICSI), provides hope for many couples struggling with fertility challenges. A review of the literature pertaining to the oxidative status of women undergoing IVF indicates that this population may be another arena where intervention with antioxidant therapies may be appropriate.
Observational data from a series of prospective trials have examined markers of oxidative activity in women undergoing both IVF and ICSI (Bedaiwy 2010, Bedaiwy 2011, Liu 2010) and their relationship to pregnant cycles. All reviewed trials report similar findings. In their assessment of follicular fluid, Bedaiwy et al. (2011) report significant associations between both lower levels of ROS and higher TAC and pregnant cycles. Non-pregnant patients in the study by Liu et al. had higher MDA measurements (p<0.05) and lower SOD levels (p<0.05), indicating higher amounts of oxidative activity. An earlier study of ROS levels in embryo culture dishes found that lower levels on day three were significantly associated with pregnant cycles (Bedaiwy 2010). A milieu dominated by oxidative processes does not appear to favour conception.
Some researchers have taken this concept of hindrance by oxidation a step further and have declared a cut point for ROS levels in follicular fluid (Jana 2010). After assessing ROS in women with a range of fertility concerns (PCOS, endometriosis, tubal factor infertility), Jana et al. propose that ROS levels over 107cps/400micromol follicular fluid do not favour the growth of viable embryos. One may speculate that these findings could eventually translate into improved clinical tools for predicting IVF success rates.
Melatonin and IVF
Melatonin is a hormone that is naturally secreted by the pineal gland to manage sleep/wake cycles in humans. This compound has also been studied extensively for its antioxidant properties (Tan 2007). In light of the evidence suggesting a detrimental effect of oxidation in IVF cycles, it is not surprising that this antioxidant hormone may promote favourable outcomes for individuals undergoing this therapy.
Three recent randomized controlled trials (RCTs) have evaluated the effect of 3mg of melatonin upon IVF-related parameters (Eryilmaz 2011, Rizzo 2010, Tamura 2008). A fourth trial did not specify dose in the abstract that was available for review (Batioğlu 2012). Trials reported significantly higher numbers of mature oocytes at pickup and higher quality embryos in treated patients (Batioğlu 2012, Rizzo 2010, Eryilmaz 2011). Trends towards higher pregnancy rates were also reported. When comparing current and previous IVF cycles, improved fertilization rates were seen among participants receiving melatonin (Tamura 2008).
The benefit of melatonin to parameters of successful IVF therapy may be attributed at least in part to its action as an antioxidant (Eryilmaz 2011, Rizzo 2010). This theory is supported and elaborated upon by a randomized trial (Taketani 2011) that demonstrates a protective effect of melatonin against reactive oxygen species (ROS) such as H2O2. The production of progesterone by luteinized granulosa cells was inhibited in vitro by H2O2 but this effect was reversed with the addition of melatonin. This effect was demonstrated in vivo by the same authors, through the restoration of deficient progesterone levels in some participants treated with 3mg of melatonin (see Figure 1).
Viewed from this perspective, melatonin may benefit women undergoing IVF, and presumably those trying to conceive naturally as well, through two related mechanisms. First, acting as a free-radical scavenger, melatonin may quench some of the oxidative stress that has been shown to be higher in women with diverse fertility challenges. Secondly, melatonin may reduce the activity of ROS such as H2O2 at the level of the ovary, preventing interference with endogenous progesterone production.
Future directions – CoQ10
While intervention studies have not yet been conducted, coenzyme Q10 (CoQ10) is another antioxidant that may also hold some promise in the treatment of infertility. A recent study investigated CoQ10 levels in the follicular fluid of women undergoing oocyte retrieval. Women with higher levels of CoQ10 had significantly increased numbers of mature oocytes and grade I-II embryos (Turi 2012), suggesting that the presence of this antioxidant compound may also contribute to positive fertility-associated outcomes.
Although oxidative processes are required for numerous essential physiological functions, a state of oxidative stress appears to be associated with conditions that may present barriers to successful conception. Markers of oxidative activity may be higher in women with endometriosis and a history of EPL. While antioxidant therapy has not been fully evaluated for either condition, treatment with 600mg of NAC may help women with EPL to prolong and preserve their pregnancies.
In women undergoing IVF treatments, increased levels of oxidative stress have been observed and may be associated with the success of individual IVF cycles. Melatonin administration at a dose of 3mg per night has been significantly associated with improved oocyte maturity and embryo quality. CoQ10 may also contribute positively to these outcomes but intervention studies are needed to support observational data. While the association between oxidation and female fertility is not entirely understood, this relationship offers some novel therapeutic options to care providers and may further our understanding of infertility that is otherwise unexplained.
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