**6. Endometriosis and oxidative stress**

Some authors have suggested that endometriosis might be associated with oxidative stress (Agarwal et al., 2003; Szczepanska et al., 2003; Gupta et al., 2006). In pelvic endometriosis there might be an activation of macrophages in the peritoneal environment leading to increased production of reactive oxygen and nitrogen species, cytokines, prostaglandins, growth factors and, therefore, oxidative stress generating lipid peroxidation and its degradation products and other products formed by its interactions with low density lipoproteins and other proteins. Peroxidized lipids, when decomposed, generate products such as malondialdehyde (MDA) and could be recognized as foreign bodies, leading to an antigenic response with consequent production of antibodies (Halliwell, 1994; Murphy et al., 1998). This process would induce oxidative damage to red blood cells and to endometrial and peritoneal cells which would stimulate recruitment and activation of a larger number of mononuclear phagocytes, maintaining oxidative damage to the pelvic environment (Van Langendonckt et al., 2002). Oxidative stress compromises mesothelial cells and might induce adhesion sites for endometrial cells, contributing to development and progression of the endometriosis focus (Alpay et al., 2006).

In a recent study by our group, blood samples were collected during the early follicular phase of the menstrual cycle for the analysis of serum MDA, GSH and total hydroxyperoxide levels by spectrophotometry and of vitamin E by high performance liquid chromatography. A positive association between infertility related to endometriosis, advanced disease stage and increased serum hydroxyperoxide levels was demonstrated, suggesting an increased production of reactive species in women with endometriosis. These data, taken together with the reduction of serum vitamin E and GSH levels, suggest the

• GPX2 encodes a gastrointestinal form, and no specific function for it is known in

• GPX4 encodes an isoform that specifically detoxifies phospholipid hydroperoxide and is thus referred to as PhGPx, and is expressed at high levels in the testis. A defect in GPX4 has been suspected to be a cause of male infertility triggered by selenium deficiency, although direct evidence for its requirement is missing (Hansen & Deguchi,

In a reaction promoted by peroxidase, GSH is oxidized to GSSG. Regeneration of GSH is, therefore, crucial for the ability of cells to fight exposure to oxidant metabolites. GSH levels are maintained by de novo synthesis that is catalyzed by two enzymes, γ- glutamylcysteine synthetase (γ-GCS) and glutathione synthetase (GS). The reduction of GSSG is catalyzed by **glutathione reductase** (GR) using NADPH as an electron donor **(2 GSSG + NADPH + H+ → 2GSH + NADP+)**. GR is also inhibited by compounds produced in response to nitrosative stress, such as nitrosoglutathione. In the female reproductive system, GSH is assumed to play a role by reducing oxidative stress either by direct interaction with ROS, by the

High levels of SeGPx were found in follicles that held oocytes with the potential to be fertilized and lower levels were related to fertilization failure (Paszkowski et al., 1995).

Some authors have suggested that endometriosis might be associated with oxidative stress (Agarwal et al., 2003; Szczepanska et al., 2003; Gupta et al., 2006). In pelvic endometriosis there might be an activation of macrophages in the peritoneal environment leading to increased production of reactive oxygen and nitrogen species, cytokines, prostaglandins, growth factors and, therefore, oxidative stress generating lipid peroxidation and its degradation products and other products formed by its interactions with low density lipoproteins and other proteins. Peroxidized lipids, when decomposed, generate products such as malondialdehyde (MDA) and could be recognized as foreign bodies, leading to an antigenic response with consequent production of antibodies (Halliwell, 1994; Murphy et al., 1998). This process would induce oxidative damage to red blood cells and to endometrial and peritoneal cells which would stimulate recruitment and activation of a larger number of mononuclear phagocytes, maintaining oxidative damage to the pelvic environment (Van Langendonckt et al., 2002). Oxidative stress compromises mesothelial cells and might induce adhesion sites for endometrial cells, contributing to development and progression of the

In a recent study by our group, blood samples were collected during the early follicular phase of the menstrual cycle for the analysis of serum MDA, GSH and total hydroxyperoxide levels by spectrophotometry and of vitamin E by high performance liquid chromatography. A positive association between infertility related to endometriosis, advanced disease stage and increased serum hydroxyperoxide levels was demonstrated, suggesting an increased production of reactive species in women with endometriosis. These data, taken together with the reduction of serum vitamin E and GSH levels, suggest the

**glutathione redox system**, or by donating an electron to GPx (Fujii et al., 2005).

reproduction

1996).

• GPX3 is present in plasma and in epididymal fluid

**6. Endometriosis and oxidative stress** 

endometriosis focus (Alpay et al., 2006).

occurrence of systemic oxidative stress in women with infertility associated with endometriosis (Andrade et al., 2010).

The activation of polymorphonuclear leucocytes and macrophages observed in endometriosis patients might be induced by several factors, including damaged red blood cells, apoptotic endometrial cells, cellular debris and some other inflammatory cells. In endometriosis these actions of peritoneal macrophages appear to be stimulated *in vitro* by the immune response or by agents such as α and γ-interferon, increasing inducible nitric oxide synthase (NOS) expression, producing more nitric oxide and nitrite and nitrate compounds (Agarwal et al., 2005). However, we obtained no conclusive data concerning nitric oxide, peroxidized lipids and ROS levels in the peritoneal fluid of patients with and without endometriosis (Agarwal et al., 2003; Amaral et al., 2005).

In women with endometriosis and adenomyosis, we also observe a greater expression of Mn-SOD and CuZn-SOD in the endometrium throughout the menstrual cycle, as well as aberrant expression of GPx and xanthine peroxidase (XO), in topic and ectopic endometrium. SOD activity seems to be significantly higher in the ectopic endometrium of endometriomas than in the topic endometrium (Alpay et al., 2006). However, this increase in the expression of antioxidant enzymes in the topic and ectopic endometrium of endometriosis patients could be a primary event or secondary to an increase of ROS, which needs to be evaluated. If, on the one hand, we have no conclusive data concerning the pattern of expression of the most important oxidant and antioxidant enzymes in topic and ectopic endometrium, on the other hand, we have not found, so far, any studies that have evaluated the expression of these enzymes in granulosa cells of patients with endometriosis, whose anomalies could contribute to the impairment of folliculogenesis and of the acquisition of oocyte competence to permit fertilization and support embryo development.

The above data suggest a trend to a greater production of free radicals in endometriosis patients associated with a potential alteration of antioxidant capacity. This may contribute to oxidative stress which could be related to the pathogenesis and progression of endometriosis.

Another very interesting aspect of endometriosis is its enigmatic association with infertility, observed in 25 to 30% of women with this affection. Until now, little is known about the mechanisms involved in the pathogenesis of infertility, especially in minimal and mild endometriosis, where there is no significant alteration of pelvic anatomy.

New approaches to the treatment of infertility related to this disorder have included the increasingly more common application of ART. The introduction of *in vitro* fertilization (IVF) for the treatment of infertility secondary to endometriosis has become an important tool for the study of the potential effects of endometriosis on specific stages of the reproductive process, including folliculogenesis, fertilization, embryo development and implantation. Contradictory data have been reported for IVF outcomes in patients with endometriosis (García-Velasco & Arici, 1999; Garrido et al., 2000). This discrepancy seems to be multifactorial since IVF outcomes might be affected by different variables, such as ovulation induction protocol, patient selection criteria, laboratory procedures, and embryo transfer technique, among other factors.

Endometriosis and Infertility: The Role of Oxidative Stress 405

might be involved in the etiopathogenesis of poor oocyte quality in patients with this disease. In some recent studies, sperm incubated with peritoneal fluid of endometriosis patients showed increased DNA fragmentation and the extent of fragmentation increased according to endometriosis stage and infertility duration. Similarly, oocytes incubated with peritoneal fluid of endometriosis patients presented increased DNA damage and the extent of damage was proportional to the period of exposure. As expected, embryos incubated with peritoneal fluid also showed DNA fragmentation as indicated by an increase of apoptosis. The increase of DNA damage in spermatozoa, oocytes and embryos seems to be responsible for the numerous abortions and for fertilization and implantation failure among

Our group was the first to assess the meiotic spindle and chromosome distribution of in vitro–matured (IVM) oocytes obtained from stimulated cycles of endometriosis patients and to compare them with a control group consisting of couples with male or tubal factors of infertility. We showed that, although IVM rates were similar for the two groups evaluated, a higher proportion of telophase I oocytes tended to occur in the endometriosis group. The number of oocytes was too low to detect statistically significant differences. However, this finding suggests a potential delay or impairment of meiosis I during IVM in the context of endometriosis. The mechanisms underlying this finding remain unclear. Recent studies demonstrated significant DNA damage and increased anomalies in the microtubules and chromosomes of oocytes incubated with PF from endometriosis patients (Mansour et al, 2009; Carbone et al., 2003), which were prevented by supplementation of the culture medium with the antioxidant L-carnitine, suggesting that impaired oocyte quality in endometriosis may be mediated by oxidative stress (Carbone et al., 2003). Although the data were obtained from frozen/thawed MII mouse oocytes and may not necessarily be extrapolated to human oocytes, they support our hypothesis that oxidative stress might be involved in the delay or impairment of meiosis I in oocytes of women with endometriosis (Barcelos et al., 2009), a possibility that requires more in-depth evaluation

Unpublished data from our group suggest that this finding is also confirmed in in vivo matured oocytes of patients with moderate and severe endometriosis. However, we did not find well designed studies evaluating different pro and antioxidants markers in this group of patients, co-relating them with ART outcome as indirect predictors of oocyte quality.

If we have very little evidence correlating endometriosis and meiotic oocyte anomalies, data about the potential association between endometriosis and oocyte cytoplasmic maturation markers are even rarer. The gene expression of the antioxidant enzymatic system is one of the markers of oocyte cytoplasmic maturation, playing an important role by minimizing the hazardous effects of oxidative stress (Cetica et al., 2001). It has already been demonstrated that catalase, SOD and GPx are found in oocytes and COCs. GSH is one of the oocyte cytoplasmic maturation markers that have been intensely investigated. Some studies show that an adequate expansion of COCs, which is considered to be an oocyte maturation marker, is partially dependent on the intracellular concentration of GSH (Furnus et al., 1998). Intracellular GSH levels increase as the oocyte develops from germinal vesicle to metaphase II (Ali et al., 2003). After fertilization, the total amount of intracellular GSH correlates with spermatic chromatin decondensation, with consequent oocyte activation and

endometriosis patients (Mansour et al., 2009).

in future studies.

As previously said, contradictory data have been reported for IVF outcomes in patients with endometriosis (Garrido et al., 2002; Garcia-Velasco & Arici, 1999; Kumbak et al., 2008; Fernando et al., 2008). Some studies suggest lower fertilization, implantation, and pregnancy rates in women with endometriosis (Barnhart et al., 2002; Al-Fadhli et al., 2006), possibly owing to impaired oocyte quality with consequent poor embryo quality, or to endometrial defects or defective interactions between the endometrium and the embryo (Kumbak et al., 2008, Brizek et al., 1995; Pellicer et al., 1995). Conflicting findings of some alterations in topic endometrium of endometriosis patients could explain, at least partially, the disturbance of the interaction between embryo and endometrium, generating anomalies in the implantation process (García-Velasco & Arici, 1999; Garrido et al., 2000). However, similar implantation rates in oocyte donation cycles have been recorded for women with endometriosis and control subjects, suggesting the crucial role of oocyte quality in impaired implantation processes (Pellicer et al., 1995; 2001; Díaz et al., 2000; Garrido et al., 2000; Katsoff et al., 2006). According to some authors, impaired oocyte quality would be responsible for compromising (Brizek et al., 1995) or completely blocking embryo development (Pellicer et al., 1995) in women with endometriosis, reinforcing the role of poor oocyte quality in the outcome of ART procedures in this group of patients.

Studies that intended to evaluate indirectly oocyte quality in patients with endometriosis analyzed multiple paracrine factors present in FF, such as interleukins, vascular endothelial growth factor (VEGF), and tumor necrosis factor (TNF), as well as granulosa cells apoptosis, leucocyte number and activity, among other indirect predictors of oocyte quality (Garrido et al., 2000, 2002). However, few studies have evaluated oocyte quality in patients with endometriosis by more objective morphological criteria.

Oocyte quality depends on factors related to the acquisition of nuclear and cytoplasmic competence. Although involving different processes, nuclear and cytoplasmic maturation are connected events that occur simultaneously in determined situations, although cytoplasmic molecular programming starts in the oocyte growth phase (Ferreira et al., 2009).

Nuclear competence depends on the anatomic and functional integrity of the meiotic spindle, a temporary and dynamic structure responsible for chromosomal segregation during meiosis (Wang & Keefe, 2002; Navarro et al., 2005). Meiotic anomalies might contribute to cell development failure by different paths, such as the inability of the oocyte to complete the maturation process in order to be fertilized, or the occurrence of variable errors of the meiotic maturation process that do not stop fertilization but might compromise embryo development pre or post implantation, as well as the future viability of the fetus (Armstrong, 2001; Chaube et al., 2005; Mansour et al., 2009). On the other hand, there is evidence that oxidative stress might promote meiotic anomalies and pre-implantation embryo development (Liu et al., 2003; Navarro et al., 2004, 2006; Agarwal et al., 2006; Mansour et al., 2009). Oxidative stress also seems to induce genomic and mitochondrial DNA damage (Aitken et al., 2001), which leads directly to reduced fertility (Guerin et al., 2001). Recently it was demonstrated that the peritoneal fluid of endometriosis patients promotes anomalies in oocyte cytoskeleton and increases embryo apoptosis, preventable by antioxidant supplementation (L-carnitine) in the culture medium, as shown in a study using mice as the experimental model (Mansour et al., 2009), suggesting that oxidative stress

As previously said, contradictory data have been reported for IVF outcomes in patients with endometriosis (Garrido et al., 2002; Garcia-Velasco & Arici, 1999; Kumbak et al., 2008; Fernando et al., 2008). Some studies suggest lower fertilization, implantation, and pregnancy rates in women with endometriosis (Barnhart et al., 2002; Al-Fadhli et al., 2006), possibly owing to impaired oocyte quality with consequent poor embryo quality, or to endometrial defects or defective interactions between the endometrium and the embryo (Kumbak et al., 2008, Brizek et al., 1995; Pellicer et al., 1995). Conflicting findings of some alterations in topic endometrium of endometriosis patients could explain, at least partially, the disturbance of the interaction between embryo and endometrium, generating anomalies in the implantation process (García-Velasco & Arici, 1999; Garrido et al., 2000). However, similar implantation rates in oocyte donation cycles have been recorded for women with endometriosis and control subjects, suggesting the crucial role of oocyte quality in impaired implantation processes (Pellicer et al., 1995; 2001; Díaz et al., 2000; Garrido et al., 2000; Katsoff et al., 2006). According to some authors, impaired oocyte quality would be responsible for compromising (Brizek et al., 1995) or completely blocking embryo development (Pellicer et al., 1995) in women with endometriosis, reinforcing the role of poor

oocyte quality in the outcome of ART procedures in this group of patients.

endometriosis by more objective morphological criteria.

2009).

Studies that intended to evaluate indirectly oocyte quality in patients with endometriosis analyzed multiple paracrine factors present in FF, such as interleukins, vascular endothelial growth factor (VEGF), and tumor necrosis factor (TNF), as well as granulosa cells apoptosis, leucocyte number and activity, among other indirect predictors of oocyte quality (Garrido et al., 2000, 2002). However, few studies have evaluated oocyte quality in patients with

Oocyte quality depends on factors related to the acquisition of nuclear and cytoplasmic competence. Although involving different processes, nuclear and cytoplasmic maturation are connected events that occur simultaneously in determined situations, although cytoplasmic molecular programming starts in the oocyte growth phase (Ferreira et al.,

Nuclear competence depends on the anatomic and functional integrity of the meiotic spindle, a temporary and dynamic structure responsible for chromosomal segregation during meiosis (Wang & Keefe, 2002; Navarro et al., 2005). Meiotic anomalies might contribute to cell development failure by different paths, such as the inability of the oocyte to complete the maturation process in order to be fertilized, or the occurrence of variable errors of the meiotic maturation process that do not stop fertilization but might compromise embryo development pre or post implantation, as well as the future viability of the fetus (Armstrong, 2001; Chaube et al., 2005; Mansour et al., 2009). On the other hand, there is evidence that oxidative stress might promote meiotic anomalies and pre-implantation embryo development (Liu et al., 2003; Navarro et al., 2004, 2006; Agarwal et al., 2006; Mansour et al., 2009). Oxidative stress also seems to induce genomic and mitochondrial DNA damage (Aitken et al., 2001), which leads directly to reduced fertility (Guerin et al., 2001). Recently it was demonstrated that the peritoneal fluid of endometriosis patients promotes anomalies in oocyte cytoskeleton and increases embryo apoptosis, preventable by antioxidant supplementation (L-carnitine) in the culture medium, as shown in a study using mice as the experimental model (Mansour et al., 2009), suggesting that oxidative stress might be involved in the etiopathogenesis of poor oocyte quality in patients with this disease. In some recent studies, sperm incubated with peritoneal fluid of endometriosis patients showed increased DNA fragmentation and the extent of fragmentation increased according to endometriosis stage and infertility duration. Similarly, oocytes incubated with peritoneal fluid of endometriosis patients presented increased DNA damage and the extent of damage was proportional to the period of exposure. As expected, embryos incubated with peritoneal fluid also showed DNA fragmentation as indicated by an increase of apoptosis. The increase of DNA damage in spermatozoa, oocytes and embryos seems to be responsible for the numerous abortions and for fertilization and implantation failure among endometriosis patients (Mansour et al., 2009).

Our group was the first to assess the meiotic spindle and chromosome distribution of in vitro–matured (IVM) oocytes obtained from stimulated cycles of endometriosis patients and to compare them with a control group consisting of couples with male or tubal factors of infertility. We showed that, although IVM rates were similar for the two groups evaluated, a higher proportion of telophase I oocytes tended to occur in the endometriosis group. The number of oocytes was too low to detect statistically significant differences. However, this finding suggests a potential delay or impairment of meiosis I during IVM in the context of endometriosis. The mechanisms underlying this finding remain unclear. Recent studies demonstrated significant DNA damage and increased anomalies in the microtubules and chromosomes of oocytes incubated with PF from endometriosis patients (Mansour et al, 2009; Carbone et al., 2003), which were prevented by supplementation of the culture medium with the antioxidant L-carnitine, suggesting that impaired oocyte quality in endometriosis may be mediated by oxidative stress (Carbone et al., 2003). Although the data were obtained from frozen/thawed MII mouse oocytes and may not necessarily be extrapolated to human oocytes, they support our hypothesis that oxidative stress might be involved in the delay or impairment of meiosis I in oocytes of women with endometriosis (Barcelos et al., 2009), a possibility that requires more in-depth evaluation in future studies.

Unpublished data from our group suggest that this finding is also confirmed in in vivo matured oocytes of patients with moderate and severe endometriosis. However, we did not find well designed studies evaluating different pro and antioxidants markers in this group of patients, co-relating them with ART outcome as indirect predictors of oocyte quality.

If we have very little evidence correlating endometriosis and meiotic oocyte anomalies, data about the potential association between endometriosis and oocyte cytoplasmic maturation markers are even rarer. The gene expression of the antioxidant enzymatic system is one of the markers of oocyte cytoplasmic maturation, playing an important role by minimizing the hazardous effects of oxidative stress (Cetica et al., 2001). It has already been demonstrated that catalase, SOD and GPx are found in oocytes and COCs. GSH is one of the oocyte cytoplasmic maturation markers that have been intensely investigated. Some studies show that an adequate expansion of COCs, which is considered to be an oocyte maturation marker, is partially dependent on the intracellular concentration of GSH (Furnus et al., 1998). Intracellular GSH levels increase as the oocyte develops from germinal vesicle to metaphase II (Ali et al., 2003). After fertilization, the total amount of intracellular GSH correlates with spermatic chromatin decondensation, with consequent oocyte activation and

Endometriosis and Infertility: The Role of Oxidative Stress 407

3β-Hydroxysteroide dehydrogenase/delta 5-delta 4-isomerase (3β-HSD) is an important enzyme associated with the biosynthesis of progesterone. Bar Ami (1994) evaluated the fertilization capacity related to the competence of granulosa cells and COCs to secrete progesterone. COCs from fertilized oocytes presented a 1.9 times higher progesterone level (p<0.001) on days 0-3 and a 1.6 times higher level (p<0.02) on days 3-5 of culture when compared to the levels in COCs of non-fertilized oocytes. Nevertheless, in COCs of fertilized oocytes, the activity of 3β-hydroxysteroid dehydrogenase was significantly higher after oocyte aspiration and also 3 to 5 days later compared to non-fertilized oocytes. These results suggest that, in stimulated cycles, in follicles that hold mature COCs there is a synchrony and correlation between competence to perform progesterone secretion by COCs as well as by granulosa cells and the potential of these oocytes to be fertilized. Such correlation suggests and supports the intimate relation of enzymatic activity of 3β-hydroxysteroid dehydrogenase and progesterone production with oocyte fertilization capacity, which may suggest the important role of this enzyme as coadjuvant in the acquisition of oocyte competence. The reduction of the gene expression and/or activity of this enzyme could lead

to a lower production of progesterone and impairment of the lutheal phase.

lower gene expression.

between granulosa cells and the oocyte.

Aromatase is present in granulosa cells and actually plays a fundamental role in follicle maturation and in the establishment of oocyte quality (Erickson et al., 1989; Foldesi et al., 1998; Speroff & Fritz, 2005). But, if on the one hand we find evidence of increased aromatase expression in ectopic endometrium, on the other, there are poor and inconclusive data concerning the expression of this enzyme by luteinized granulosa cells, suggesting a lower activity of this enzyme, but with no confirmation of an associated

It is known that oocyte quality results from a complex and synchronized process that lasts several months, from primordial follicle to pre-ovulatory follicle. This process starts in a gonadotropin-independent way and later becomes gonadotropin dependent. In this last phase, oocyte, granulosa cells and FSH interact synergically. Granulosa cell multiplication and the specific way they respond first to FSH and later to LH in order to produce intrafollicle steroids are crucial events in this process (Speroff & Fritz, 2005). We know that there are gap junctions between granulosa cells, which is evidence that there are molecular interactions between them and possibly with the oocyte itself, through signaling molecules such as growth and differentiation factor-9 (GDF-9) and bone morphogenetic protein-15 (BMP 15) (Albertini & Barrett, 2003; Combelles et al., 2004; Thomas & Vanderhyden, 2006; Hutt & Albertini, 2007). However, little information is available about the communication

Granulosa cells differentiate into mural and cumulus cells during folliculogenesis, a fact that has stimulated the study of their potential as mesenchymal stem cells. To date there are no studies comparing the gene expression of mural granulosa cells and COCs and, possibly, since they are cells with distinct function and differentiation, there might be genes with different patterns of expression. When they reach the pre-antral follicle stage, granulosa cells can synthetize all three types of steroids (androgens, progestagens and estrogens) (Speroff & Fritz, 2005). However, the proportions and timing of their production are crucial. It is known that FSH and also LH have hormonal receptors on granulosa cells and there is a synergism between these receptors and intra-follicle hormonal production to permit the

also with the transformation of the sperm head to male pronucleus (De Matos & Furnus, 2000). However, no studies have evaluated the expression of this enzyme or of the entire GSH redox system in COCs of patients with infertility related to endometriosis. Matos et al. (2009) suggested a positive correlation between the SOD activity of COCs of infertile women submitted o ovarian stimulation for ART due to male factor and ART outcomes. In this same study an increase in SOD activity was observed in in vitro culture of COCs from infertile women with endometriosis. However, the authors analyzed the COCs of only six patients.

Some authors have associated minimal endometriosis with impaired steroidogenesis in granulosa cells, represented not only by a reduced baseline activity of aromatase, but also by a lower production of progesterone in non-stimulated and stimulated cycles (Harlow et al., 1996; Gomes et al., 2008). A functional failure of oocytes due to abnormal follicular function could be a result of this disease (Wardle et al., 1985). The antioxidants not only have an antiapoptotic effect on preovulatory in vitro cultured follicles (Tsai-Turton & Luderer, 2006), but are also involved in the regulation of steroidogenic enzyme function dependent on cytochrome P450 (Verit et al., 2007). Some studies have suggested that ascorbic acid (Murray et al., 2001), as well as SOD (Lapolt & Hong, 1995) may have inhibitory effects on aromatase, an enzyme responsible for the conversion of androgens to estrogens, which could induce storage of androgens in the follicular fluid, leading to follicular atresia (Verit et al., 2007). As mentioned earlier, some recent data have demonstrated an increase of SOD activity in COCs (in vitro culture) of infertile women with endometriosis (Matos et al., 2009). Since no studies on endometriosis patients have evaluated antioxidant enzyme expression in luteinized granulosa cells and their correlation with steroidogenic enzymes dependent on cytochrome P450 expression, involved in ovarian steroidogenesis, our group has performed studies evaluating these possible associations.
