Elsevier

Food and Chemical Toxicology

Volume 118, August 2018, Pages 111-118
Food and Chemical Toxicology

Postnatal exposure to a glyphosate-based herbicide modifies mammary gland growth and development in Wistar male rats

https://doi.org/10.1016/j.fct.2018.05.011Get rights and content

Highlights

  • Postnatal subacute exposure to GBH accelerates mammary gland development in male rats.

  • GBH augments mast cell infiltration in the mammary stroma of post-pubertal males.

  • GBH increases mammary gland proliferation and estrogen receptor alpha expression in post-pubertal males.

Abstract

Our aim was to evaluate whether postnatal exposure to a glyphosate-based herbicide (GBH) modifies mammary gland development in pre- and post-pubertal male rats. From postnatal day 1 (PND1) to PND7, male rats were injected subcutaneously every 48 h with either saline solution (vehicle) or 2 mg GBH/kg·bw. On PND21 and PND60, mammary gland and blood samples were collected. Estradiol (E2) and testosterone (T) serum levels, mammary gland histology, collagen fiber organization, mast cell infiltration, proliferation index, and estrogen (ESR1) and androgen receptor (AR) expression levels were evaluated. At PND21, GBH-exposed male rats exhibited greater development of the mammary gland with increased stromal collagen organization and terminal end buds (TEBs) compared to control rats. At PND60, the number of TEBs remained high and was accompanied by an increase in mast cell infiltration, proliferation and ESR1 expression in GBH-exposed male rats. In contrast, no effects were observed in E2 and T serum levels and AR expression in both days studied. Our results showed that a postnatal subacute treatment with GBH induces endocrine-disrupting effects in the male mammary gland in vivo, altering its normal development.

Introduction

Glyphosate (N-phosphonomethyl glycine) is an active ingredient in many commercially available broad-spectrum herbicides. Over the past decades, glyphosate-based herbicide (GBH) use has diversified and expanded significantly. Benbrook (2016) reported that the application of these herbicides increased ∼ 100-fold worldwide from 1974 to 2014. In Argentina, increasing use of these chemicals has been associated with agricultural expansion due to the ongoing adoption of glyphosate tolerant genetically modified soybeans (CASAFE, 2012). In recent years, glyphosate and its major metabolite, aminomethylphosphonic acid (AMPA), have been detected in surface water, sediments and soil that surround horticultural production areas of different regions of Argentina (Aparicio et al., 2013; Bonansea et al., 2017; Lupi et al., 2015; Mac Loughlin et al., 2017; Primost et al., 2017; Ronco et al., 2016). These pesticide residues have also been detected in foodstuff (EFSA, 2017) and human urine (Connolly et al., 2017; Goen et al., 2017; Mills et al., 2017; Parvez et al., 2018) and serum samples (Kongtip et al., 2017). In addition, it has been demonstrated in rats that exposure to a mixture containing glyphosate is able to induce hepatoxicity (Docea et al., 2018) and that low doses of GBH provokes kidney and especially liver oxidative damage and non-alcoholic fatty liver disease (Mesnage et al., 2015a, 2017b). These findings suggest that there is a risk of environmental exposure to GBH and raise concern of its possible effects on the environment and human health.

Several studies have reported the adverse effects of GBH exposure on both female and male reproductive systems at low and environmentally relevant doses (Cai et al., 2017; Guerrero Schimpf et al., 2017; Ingaramo et al., 2016, 2017; Nardi et al., 2017; Varayoud et al., 2017). In male rats, glyphosate exposure modifies testicular function and morphology, decreases testosterone serum levels, increases aromatase expression level, and alters sperm production, suggesting that GBH could act as an endocrine disruptor in vivo (Cassault-Meyer et al., 2014; Dallegrave et al., 2007; Owagboriaye et al., 2017; Romano et al., 2010). In addition, glyphosate was shown to reduce aromatase enzyme activity in human placental cells (Richard et al., 2005) and induce human breast cancer cell proliferation by directly activating the estrogen receptor alpha (ESR1) in vitro (Thongprakaisang et al., 2013). In contrast, Mesnage et al. (2017a) showed that glyphosate activates ESR1 through a ligand-independent mechanism in hormone-dependent human cancer cells. Therefore, the estrogenic potential effect of glyphosate remains under investigation. In general, new epidemiological and toxicological studies as well as human biomonitoring are urgently needed to determinate whether GBH could be considered an endocrine disruptor and to improve safety standard (Myers et al., 2016; Vandenberg et al., 2017).

Given the effects on male reproductive organs and the suggested estrogenic properties of this compound, male mammary gland development could also be affected. The male mammary gland of rats has been used by several researchers as a useful model to study the effects of potential endocrine disruptors that may affect the risk of breast cancer in humans (Filgo et al., 2016; Mandrup et al., 2015, 2016). Recently, we showed that prenatal exposure to the endocrine disruptor bisphenol A induces a growth delay and decreases the expression of androgen receptor (AR) in the pre-pubertal male rat mammary gland (Kass et al., 2015) and that early postnatal exposure to endosulfan induces premalignant lesions in the mammary gland of post-pubertal male rats (Altamirano et al., 2017). There is evidence that chronic exposure to GBH increases mammary tumor incidence in adults rats (Seralini et al., 2014). However, to date, the effects of postnatal GBH exposure on the male mammary gland in vivo remain mostly unknown. Therefore, the aim of the present study was to evaluate whether early postnatal exposure to GBH affects mammary gland growth and development in pre- and post-pubertal male rats.

Section snippets

Animals

Sexually mature female rats (90 days old) of a Wistar-derived strain bred at the Department of Human Physiology (Facultad de Bioquímica y Ciencias Biológicas, Universidad Nacional del Litoral (UNL), Santa Fe, Argentina) were used. The animals were maintained in a controlled environment (22 ± 2 °C; 14 h of light) and had free access to pellet laboratory chow (16-014007 Rat-Mouse diet, Nutrición Animal, Santa Fe, Argentina). All the experimental protocols were approved by the Ethical Committee of

Results

No signs of acute or chronic toxicity were observed in the litters, and no differences were found in the weight gain between treated and control pups during the treatment (PND1-PND7). At PND21, the pup body weight was similar between the experimental groups (Control: 34.39 ± 0.96 g vs GBH: 35.85 ± 0.78 g), whereas in post-pubertal (PND60) male rats exposed to GBH, a significant increase in the body weight was observed (Control: 218.10 ± 3.82 g vs GBH: 230.70 ± 2.98 g; p < 0.05).

Discussion

In recent years, the effects of endocrine-disrupting chemicals on the development of the male mammary gland of rodents has received increased attention due to its sensitive response to estrogenic and/or androgenic compounds (Altamirano et al., 2017; Filgo et al., 2016; Kass et al., 2015; Kolla et al., 2017; Mandrup et al., 2015; Vandenberg et al., 2013). In the current study, postnatal subacute exposure of male rats to GBH during a critical period of development resulted in alterations in the

Declaration of interest

The authors declare that there are no conflicts of interest that could be perceived as prejudicing the impartiality of the research reported.

Funding

This work was supported by grants from the Universidad Nacional del Litoral (CAI + D program 2016 50420150100088L) and the Agencia Nacional de Promoción Científica y Tecnológica (ANPCyT PICT 2014 Nª1348). These funding sources were not involved in the study design, the collection, analysis or interpretation of the data, the writing of the report, or the decision to submit the article for publication.

Acknowledgments

We would like to thank Dr. Pablo M. Beldomenico (Laboratorio de Ecología de Enfermedades, Instituto de Ciencias Veterinarias del Litoral (ICiVet-Litoral), UNL-CONICET) for advice on statistics and Juan Grant and Laura Bergero (ISAL; UNL-CONICET) for technical assistance and animal care. G.A.A. and A.L.G. are fellows and L.K., P·I., V.L.B. and E.H.L. are Career Investigators of CONICET.

References (64)

  • M. Kwiatkowska et al.

    DNA damage and methylation induced by glyphosate in human peripheral blood mononuclear cells (in vitro study)

    Food Chem. Toxicol.

    (2017)
  • K.R. Levental et al.

    Matrix crosslinking forces tumor progression by enhancing integrin signaling

    Cell

    (2009)
  • J.N. Lilla et al.

    Mast cells contribute to the stromal microenvironment in mammary gland branching morphogenesis

    Dev. Biol.

    (2010)
  • L. Lupi et al.

    Occurrence of glyphosate and AMPA in an agricultural watershed from the southeastern region of Argentina

    Sci. Total Environ.

    (2015)
  • T.M. Mac Loughlin et al.

    Pesticide impact study in the peri-urban horticultural area of Gran La Plata, Argentina

    Sci. Total Environ.

    (2017)
  • K.R. Mandrup et al.

    Mixtures of environmentally relevant endocrine disrupting chemicals affect mammary gland development in female and male rats

    Reprod. Toxicol.

    (2015)
  • R. Mesnage et al.

    Ethoxylated adjuvants of glyphosate-based herbicides are active principles of human cell toxicity

    Toxicology

    (2013)
  • R. Mesnage et al.

    Potential toxic effects of glyphosate and its commercial formulations below regulatory limits

    Food Chem. Toxicol.

    (2015)
  • R. Mesnage et al.

    Evaluation of estrogen receptor alpha activation by glyphosate-based herbicide constituents

    Food Chem. Toxicol.

    (2017)
  • M.M. Milesi et al.

    Neonatal exposure to low doses of endosulfan induces implantation failure and disrupts uterine functional differentiation at the pre-implantation period in rats

    Mol. Cell. Endocrinol.

    (2015)
  • M.M. Milesi et al.

    Uterine ERalpha epigenetic modifications are induced by the endocrine disruptor endosulfan in female rats with impaired fertility

    Mol. Cell. Endocrinol.

    (2017)
  • G.S. Montes

    Structural biology of the fibres of the collagenous and elastic systems

    Cell Biol. Int.

    (1996)
  • J. Nardi et al.

    Prepubertal subchronic exposure to soy milk and glyphosate leads to endocrine disruption

    Food Chem. Toxicol.

    (2017)
  • F.O. Owagboriaye et al.

    Reproductive toxicity of Roundup herbicide exposure in male albino rat

    Exp. Toxicol. Pathol.

    (2017)
  • J.E. Primost et al.

    Glyphosate and AMPA, “pseudo-persistent” pollutants under real-world agricultural management practices in the Mesopotamic Pampas agroecosystem, Argentina

    Environ. Pollut.

    (2017)
  • S. Thongprakaisang et al.

    Glyphosate induces human breast cancer cells growth via estrogen receptors

    Food Chem. Toxicol.

    (2013)
  • L.N. Vandenberg et al.

    The male mammary gland: a target for the xenoestrogen bisphenol A

    Reprod. Toxicol.

    (2013)
  • A. Aponte-Lopez et al.

    Mast cell, the neglected member of the tumor microenvironment: role in breast cancer

    J. Immunol. Res.

    (2018)
  • C.M. Benbrook

    Trends in glyphosate herbicide use in the United States and globally

    Environ. Sci. Eur.

    (2016)
  • R.I. Bonansea et al.

    The fate of glyphosate and AMPA in a freshwater endorheic basin: an ecotoxicological risk assessment

    Toxics

    (2017)
  • R.H. Cardy

    Sexual dimorphism of the normal rat mammary gland

    Vet. Pathol.

    (1991)
  • CASAFE

    Mercado Argentino 2012 de productos fitosanitarios. Camara Argentina de Sanidad Agropecuaria y Fertilizantes

    (2012)
  • Cited by (0)

    View full text