Perfluoroalkyl acid contamination of follicular fluid and its consequence for in vitro oocyte developmental competence☆
Introduction
Over the past century, the awareness of the harmful effects of environmental pollution on humans and wildlife has increased enormously, mainly due to the physiological abnormalities observed in species living in highly polluted areas (Bernanke and Kohler, 2009, Hamlin and Guillette, 2010). Thousands of new chemicals are being introduced onto the market, some of which have proven to induce negative effects on laboratory animals and in in vitro experiments (Zhao et al., 2012).
From this group of emerging toxicants, perfluoroalkyl acids (PFAAs), considered to be biologically inactive for a long time, are detected in humans at concentrations exceeding the levels found in many other species (Jensen and Leffers, 2008). PFAAs consist of a carbon backbone occupied by fluoride atoms, where the CF bond is the strongest in organic chemistry, which make them exceedingly stable and almost non bio-degradable. Perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA), used in fire fighting foams (Paul et al., 2009) and as processing aid (Trudel et al., 2008) respectively, and both end-metabolites of several PFAAs (Lau et al., 2004), are most frequently detected, whereby PFOS concentrations are in general the highest in biota (Lau et al., 2007). Due to their combined unique hydrophilic and hydrophobic properties, PFAAs have broad applications in consumer and industrial fields (‘Baygard’, ‘Scotchgard’, ‘Gore-Tex’) (Jensen and Leffers, 2008). In humans, PFAAs are readily absorbed, poorly eliminated and distributed primarily in serum and liver (Lau et al., 2004), with long half-lives of around 4 and 5 years for PFOA and PFOS, respectively (Olsen et al., 2007). Humans are mainly exposed to PFAAs through food, drinking water, indoor air and dust particles contaminated with PFAAs, via the anti-adhesive surface layer on cookware and a whole range of water- and/or oil-resistant PFAA-coated consumer products (e.g. clothes, carpets) (D'Hollander et al., 2010a, D'Hollander et al., 2010b, Haug et al., 2011, Jensen and Leffers, 2008).
Nowadays, epidemiological data are available suggesting associations between the presence of PFAAs and a range of human health outcomes (Fisher et al., 2013, Fletcher et al., 2013, Saikat et al., 2013). It has for example been demonstrated that in vivo exposure to PFOA and PFOS is associated with endometriosis diagnosis (Louis et al., 2012) and that it can influence the expression of genes involved in cholesterol metabolism (Fletcher et al., 2013). Conflicting results are reported when investigating the relationship between the presence of PFAAs in human serum and subfecundity (Fei et al., 2009, Whitworth et al., 2012). In rat and mice, prenatal exposure to high PFOS concentrations (5–20 mg/kg) led to subsequent sudden neonatal mortality, postnatal growth and developmental retardation among offspring (Lau et al., 2004). In addition, female mice neonates showed increased body weight and elevated serum insulin and leptin concentrations after in utero exposure to low (0.01 mg/kg) and medium (0.3 mg/kg) PFOA levels (Lau et al., 2009). While the ability of PFAAs to interact with the nuclear Peroxisome Proliferator-Activated Receptor family (PPARs) has been put forward as an explanation for the observed metabolic disturbances, mainly through PPARα-activation (Rees et al., 2008, Wolf et al., 2012), PPARs have also been suggested to be involved in gamete function and embryo development (Huang, 2008, Minge et al., 2008). For example, PPARγ-expression increases in granulosa cells during folliculogenesis and drops following the LH-surge (Froment et al., 2006), thereby possibly playing an important role in cell steroidogenesis (Huang, 2008). Within the ovarian follicular micro-environment, PFAA-binding to PPARs could thus possibly interfere with steroidogenesis (i.e. estradiol, testosterone and progesterone) and subsequently influence oocyte and embryo development. In adult male rats, PFAAs are shown to affect the endocrine system by decreasing testosterone and increasing estradiol levels (Jensen and Leffers, 2008). These estrogenic effects of PFAAs were also observed in PFAA-exposed MCF-7 cell cultures (Maras et al., 2006).
Thus, it can be hypothesized that PFAAs are capable of interfering with the tightly regulated endocrine processes taking place in the ovarian follicle, where well-balanced interactions between hormones, growth factors, the oocyte and its surrounding somatic cells are essential to generate a fully competent oocyte. Subsequently, it is important to demonstrate the presence of PFAAs within the follicular fluid, as it may further substantiate their potential direct effects on oocyte, cumulus and granulosa cells. While lipophilic chemicals, such as polychlorinated biphenyls (PCBs) and dichlorodiphenyldichloroethylene (DDE), have already been detected in human follicular fluid (Jarrell et al., 1993, Jirsova et al., 2010, Meeker et al., 2009, Pauwels et al., 1999, Petro et al., 2012, Trapp et al., 1984, Weiss et al., 2006, Younglai et al., 2002), to the best of our knowledge, no detailed information is currently available regarding the presence and distribution of PFAAs in human follicular fluid. Only one study mentions the presence of PFAAs in human follicular fluid, but no exact information regarding the identity and concentrations of the detected PFAAs is given (Governini et al., 2011).
Therefore, the main aim of the current study is to investigate whether the (possible) presence of PFAAs could be a risk factor for in vivo oocyte and embryo development. Therefore, we intended: 1) to detect and quantify individual PFAAs in human follicular fluid obtained through transvaginal follicular aspiration in the course of assisted reproductive therapies; 2) to detect and quantify the same set of PFAAs in human blood serum samples obtained at the moment of transvaginal oocyte retrieval; 3) to assess the distribution characteristics of the detected PFAAs in follicular fluid and serum; and 4) to link follicular fluid PFAA-concentrations to the final reproductive outcome in terms of fertilization rate and the amount of in vitro produced high quality embryos.
Section snippets
Patient information, cycle characteristics and sample collection
Patients undergoing an assisted reproductive technology (ART) treatment were invited to participate after Ethical Committee approvals of the University of Antwerp and ZNA Middelheim Hospital were obtained. All patients signed informed consent papers. During two periods, human follicular fluid samples (n = 18, March 2008) and paired serum and follicular fluid samples (n = 20, March–May 2009) were collected in the fertility unit of ZNA Middelheim Hospital, Antwerp, Belgium.
Data were collected on
Results
Patients (n = 38) were on average 34.6 ± 4.4 years old, had an average BMI of 23.2 ± 4.9 kg/m2 (Table 1) and all lived in urbanized areas. Four PFAAs, i.e. PFOS, PFOA, PFNA and PFHxS, were detected in each serum sample. For follicular fluid samples, PFOS was detected in every sample, except one and PFOA, PFNA and PFHxS showed detection frequencies above 75% (Table 2). PFOS was the compound found in the highest concentrations in follicular fluid (median 7.5, range 0.1–30.4 ng/mL) and serum samples
Discussion
This study was designed to investigate if the presence of PFAAs in the ovarian follicular micro-environment could be considered a risk factor for oocyte development. Firstly, we demonstrated that PFAAs are indeed present in the ovarian follicles of women undergoing ART. Moreover, we also report that PFAAs display a different distribution pattern in serum and follicular fluid compared to already detected POPs in follicular fluid. Finally, a positive association could be observed between
Conclusion
Our study provides the first, detailed overview of the contamination status of follicular fluid with PFAAs. It became clear that the different characteristics of PFAAs compared to POPs are being reflected by a different distribution pattern in follicular fluid and serum. Also opposite findings were found between the presence of PFAAs and POPs in relation to the ART-outcomes: in our small patient group, higher PFAA-contaminated follicular fluid samples were associated with a better fertilization
Acknowledgments
Petro EML acknowledges a scholarship BOF-UA from the University of Antwerp and D'Hollander W an EU-scholarship (‘PERFOOD’, KBBE-227525). Covaci A acknowledges financial support from the Research Scientific Foundation of Flanders (FWO) and the University of Antwerp. Ellen P.A. Jorssen acknowledges support from a Belgian Government research grant (Federale Overheidsdienst Volksgezondheid, Veiligheid van de Voedselketen en Leefmilieu, cel Contractueel Onderzoek) ‘Embryoscreen RF6222’. Tim Willems
References (69)
- et al.
Persistent organic pollutants (POPs) in human milk: a biomonitoring study in rural areas of Flanders (Belgium)
Chemosphere
(2012) - et al.
Brominated flame retardants and perfluorinated compounds in indoor dust from homes and offices in Flanders, Belgium
Chemosphere
(2010) - et al.
Do perfluoroalkyl substances affect metabolic function and plasma lipids?—analysis of the 2007–2009, Canadian Health Measures Survey (CHMS) cycle 1
Environ Res
(2013) - et al.
Associations between PFOA, PFOS and changes in the expression of genes involved in cholesterol metabolism in humans
Environ Int
(2013) - et al.
Characterisation of human exposure pathways to perfluorinated compounds — comparing exposure estimates with biomarkers of exposure
Environ Int
(2011) - et al.
Effect of polychlorinated biphenyls (PCBs) and 1,1,1-trichloro-2,2,-bis (4-chlorophenyl)-ethane (DDT) in follicular fluid on the results of in vitro fertilization embryo transfer (IVF-ET) programs
Fertil Steril
(2010) - et al.
Perfluorinated chemicals in relation to other persistent organic pollutants in human blood
Chemosphere
(2006) - et al.
The developmental toxicity of perfluoroalkyl acids and their derivatives
Toxicol Appl Pharmacol
(2004) - et al.
Perfluorinated chemicals 2008: PFAA days II meeting report and highlights
Reprod Toxicol
(2009) - et al.
The peroxisome proliferator-activated receptor gamma ligand troglitazone induces apoptosis and p53 in rat granulosa cells
Mol Cell Endocrinol
(2005)
Exposure to environmental endocrine disrupting compounds and men's health
Maturitas
Serum levels of perfluoroalkyl compounds in human maternal and umbilical cord blood samples
Environ Res
The relation between levels of selected PCB congeners in human serum and follicular fluid
Chemosphere
Evidence for the involvement of xenobiotic-responsive nuclear receptors in transcriptional effects upon perfluoroalkyl acid exposure in diverse species
Reprod Toxicol
Brominated flame retardants and perfluorinated chemicals, two groups of persistent contaminants in Belgian human blood and milk
Environ Pollut
Comparison of polyfluoroalkyl compound concentrations in maternal serum and amniotic fluid: a pilot study
Reprod Toxicol
Analysis of fluorotelomer alcohols, fluorotelorner acids, and short- and long-chain perfluorinated acids in water and biota
Journal of Chromatography A
Pollutants in human follicular fluid
Fertil Steril
Polyfluoroalkyl chemicals in the serum and milk of breastfeeding women
Reprod Toxicol
Activation of mouse and human peroxisome proliferator-activated receptor-alpha (PPAR alpha) by perfluoroalkyl acids (PFAAs): further investigation of C4–C12 compounds
Reprod Toxicol
Environmental contamination, human exposure and body loadings of perfluorooctane sulfonate (PFOS), focusing on Asian countries
Chemosphere
AMAP ring test for persistent organic pollutants in human serum
Report of the college of physicians for assisted reproduction therapy, Belgium 2008
Report of the college of physicians for assisted reproduction therapy, Belgium 2009
The impact of environmental chemicals on wildlife vertebrates
Rev Environ Contam Toxicol
Structure–activity relationships and human relevance for perfluoroalkyl acid-induced transcriptional activation of peroxisome proliferation in liver cell cultures
Toxicol Sci
Perfluorinated substances in human food and other sources of human exposure
Role of the peroxisome proliferator-activated receptors, adenosine monophosphate-activated kinase, and adiponectin in the ovary
PPAR Research
Directive 2006/122/EC of the European Parliament and of the Council of 12 December 2006 amending for the 30th time Council Directive 76/769/EEC on the approximation of the laws, regulations and administrative provisions of the member states relating to restrictions on the marketing and use of certain dangerous substances and preparations (perfluorooctane sulfonates)
Off J Eur Union
EPA and 3M announce phase out of PFOS
2010/2015 PFOA Stewardship Program
Maternal levels of perfluorinated chemicals and subfecundity
Hum Reprod
Peroxisome proliferator-activated receptors in reproductive tissues: from gametogenesis to parturition
J Endocrinol
Global distribution of perfluorooctane sulfonate in wildlife
Environ Sci Technol
Cited by (46)
In vitro exposure to triazoles used as fungicides impairs human granulosa cells steroidogenesis
2023, Environmental Toxicology and Pharmacology
- ☆
Financial interest declaration: There is no conflict of interest for any of the authors.