Effects of continuous ethinylestradiol and drospirenone administration on body mass and mammary gland in female mice fed a standard or high-fat diet

Authors

DOI:

https://doi.org/10.52521/nutrivisa.v12i1.15667

Keywords:

Mammary gland, Obesity, Oral contraceptives.

Abstract

Combined oral contraception (COC) is used by millions of women worldwide, but some studies suggest it may increase the risk of breast cancer. Overweight and obesity have reached epidemic levels and are also risk factors for mammary gland neoplasms. This study evaluated the effects of a COC containing ethinylestradiol (EE2) and drospirenone (DRSP) on reproductive organs, particularly the histopathology of inguinal mammary glands in reproductively active female mice fed a standard diet (SD) or high-fat diet (HFD). Adult Swiss female mice received either SD or HFD and were treated daily by gavage with distilled water [control (CTL)-SD and CTL-HFD] or 0.6 µg EE2 and 60 µg DRSP (COC-SD and COC-HFD) for 65 days. COC treatment disrupted the estrous cycle, with vaginal smears showing fewer squamous cornified cells but more leukocytes, deep cells, and mucus, characteristic of metestrus and proestrus. At the end of the experiment, COC-SD females showed body weight (BW), inguinal white adipose tissue (WAT) mass, and uterus and ovary weights similar to CTL-SD. HFD increased BW and WAT mass without altering uterus and ovary weights. COC reduced HFD-induced BW and WAT gain but increased uterus weight in COC-HFD females. Morphological analysis of mammary glands revealed no pathological signs or structural changes in acini, ducts, or lumens due to COC or diet. Notably, COC-HFD females had smaller white adipocytes than CTL-HFD, while beige adipocyte levels remained unchanged across groups.  These findings suggest that EE2 and DRSP may regulate HFD-induced obesity without adversely affecting mammary gland morphology.

References

AGUSTIN, S.A.; BAROKAH, L. Correlation between Obesity and Contraceptive Method on Estrogen and Progesterone Receptors and Human Epidermal Growth Factor Receptor–2 Expression among Breast Cancer Patients in Dr. Moewardi Hospital, Surakarta. Indonesian Journal of Medicine, v.4, n.3, p.259–265, 2019. doi: 10.26911/theijmed.2019.4.3.204

ALIMKHODJAEVA, L.T.; NISHANOV, D.A.; BOZAROVA, L.M.; NORBEKOVA, M.K.H. Immunohistochemical Aspects of the Expression of Markers of Cell Cycle Regulation, Proliferation and Apoptosis (p53, Ki-67, bcl-2, cyclin D1) in Breast Neoplasia. Research In Cancer and Tumor, v.11, n.1, p.1–5, 2023. doi:10.5923/j.rct.20231101.01

AL-QAHTANI, S.M.; BRYZGALOVA, G.; VALLADOLID-ACEBES, I.; KORACH-ANDRÉ, M.; DAHLMAN-WRIGHT, K.; EFENDIĆ, S.; BERGGREN, P.O.; PORTWOOD, N. 17β-Estradiol suppresses visceral adipogenesis and activates brown adipose tissue-specific gene expression. Hormone Molecular Biology and Clinical Investigation, v.29, n.1, p.13–26, 2017. doi: 10.1515/hmbci-2016-0031

ARENDT, L.M.; KUPERWASSER, C. Form and Function: how Estrogen and Progesterone Regulate the Mammary Epithelial Hierarchy. Journal of Mammary Gland Biology and Neoplasia, v.20, n.1–2, p.9–25, 2015. doi: 10.1007/s10911-015-9337-0

ARMANI, A.; CINTI, F; MARZOLLA, V.; MORGAN, J.; CRANSTON, G.A.; ANTELMI, A.; CARPINELLI, G.; CANESE, R.; PAGOTTO, U.; QUARTA, C.; MALORNI, W.; MATARRESE, P.; MARCONI, M.; FABBRI, A.; ROSANO, G.; CINTI, S.; YOUNG, M.J.; CAPRIO, M. Mineralocorticoid receptor antagonism induces browning of white adipose tissue through impairment of autophagy and prevents adipocyte dysfunction in high‐fat‐diet‐fed mice. The FASEB Journal, v.28, n.8, p.3745–3757, 2014. doi: 10.1096/fj.13-245415

BARAŃSKA, A. Oral Contraceptive Use and Assessment of Breast Cancer Risk among Premenopausal Women via Molecular Characteristics: Systematic Review with Meta-Analysis. International Journal of Environmental Research and Public Health, v.19, n.22, p.15363, 2022. doi: 10.3390/ijerph192215363

BATES, P.; FISHER, R.; WARD, A.; RICHARDSON, L.; HILL, D.; GRAHAM, C. Mammary cancer in transgenic mice expressing insulin-like growth factor II (IGF-II). British Journal of Cancer, v.72, n.5, p.1189–1193, 1995. doi: 10002E1038/bjc.1995.484

BOUTINAUD, M.; SHAND, J.; PARK, M.; PHILLIPS, K.; BEATTIE, J.; FLINT, D.; ALLAN, G. A quantitative RT-PCR study of the mRNA expression profile of the IGF axis during mammary gland development. Journal of Molecular Endocrinology, v.33, n.1, p.195–207, 2004. doi: 10.1038/bjc.1995.484

CAO, J.; LI, J.; ZHANG, Z.; QIN, G.; PANG, Y.; WU, M.; GU, K.; XU, H. Interaction between body mass index and family history of cancer on the risk of female breast cancer. Scientific Reports, v.14, n.1, p.4927, 2024. doi: 10.1038/s41598-024-54762-x

CASIMIRO, M.C.; WANG, C.; LI, Z.; SANTE, G. DI; WILLMART, N.E.; ADDYA, S.; CHEN, L.; LIU, Y.; LISANTI, M.P.; PESTELL, R.G. Cyclin D1 Determines Estrogen Signaling in the Mammary Gland In Vivo. Molecular Endocrinology, v.27, n.9, p.1415–1428, 2013. doi: 10.1210/me.2013-1065

CLEUREN, A.C.A.; LINDEN, I.K. VAN DER; VISSER, Y.P. DE; WAGENAAR, G.T. M.; REITSMA, P.H.; VLIJMEN, B.J.M. VAN. 17α‐Ethinylestradiol rapidly alters transcript levels of murine coagulation genes via estrogen receptor α. Journal of Thrombosis and Haemostasis, v.8, n.8, p.1838–1846, 2010. doi: 10.1111/j.1538-7836.2010.03930.x

CONWAY, J.R.W.; DINÇ, D.D.; FOLLAIN, G.; PAAVOLAINEN, O.; KAIVOLA, J.; BOSTRÖM, P.; HARTIALA, P.; PEUHU, E.; IVASKA, J. IGFBP2 secretion by mammary adipocytes limits breast cancer invasion. Science Advances, v.9, n.28, 2023. doi: 10.1126/sciadv.adg1840

DEROSSI, D.R.; ITO, K.; COUTO FILHO, J.D.; BACCHI, C.E. Avaliação da expressão da proteína bcl-2 no carcinoma de mama: estudo em punção aspirativa por agulha fina; correlação com grau histológico em espécimes cirúrgicos correspondentes. Jornal Brasileiro de Patologia e Medicina Laboratorial, v39, n. 3, 2003. doi: 10.1590/S1676-24442003000300010

DIETZ OSTERGAARD, S.; BUTLER, K.; RITTER, J. M.; JOHNSON, R.; SANDERS, J.; POWELL, N.; LATHROP, G.; ZAKI, S. R.; MCNICHOLL, J.M.; KERSH, E.N. A combined oral contraceptive affects mucosal SHIV susceptibility factors in a pigtail macaque (Macaca nemestrina) model. Journal of Medical Primatology, v.44, n.2, p.97–107, 2015. doi: 10.1111/jmp.12157

DIXON, A.; ROWAN, E.G.; YACKLEY, A.N.; HINES, E.P. PFAS Modulate Osmotic Signaling Independent of Gravimetric Changes in the Rat Uterus. Toxics, v.12, n.3, p.170, 2024. doi: 10.3390/toxics12030170

FIGUEIREDO, L.S.; OLIVEIRA, K.M.; FREITAS, I.N.; SILVA, J.A.; SILVA, J.N.; FAVERO-SANTOS, B.C.; BONFLEUR, M.L.; CARNEIRO, E.M.; RIBEIRO, R.A. Bisphenol-A exposure worsens hepatic steatosis in ovariectomized mice fed on a high-fat diet: Role of endoplasmic reticulum stress and fibrogenic pathways. Life Sciences, v.256, p.118012, 2020. doi: 10.1016/j.lfs.2020.118012

FITZPATRICK, D.; PIRIE, K.; REEVES, G.; GREEN, J.; BERAL, V. Combined and progestagen-only hormonal contraceptives and breast cancer risk: A UK nested case–control study and meta-analysis. PLOS Medicine, v.20, n.3, p.e1004188, 2023. doi: 10.1371/journal.pmed.1004188

FREITAS, G.C.; CARREGARO, A.B. Aplicabilidade da extrapolação alométrica em protocolos terapêuticos para animais selvagens. Ciência Rural, v.43, n.2, p.297–304, 2013. doi: 10.1590/S0103-84782013000200017

FUHRMANN, U.; KRATTENMACHER, R.; SLATER, E.P.; FRITZEMEIER, K.H. The novel progestin drospirenone and its natural counterpart progesterone: Biochemical profile and antiandrogenic potential. Contraception, v.54, n.4, p.243–251, 1996. doi: 10.1016/s0010-7824(96)00195-3

FULLER, K.N.Z.; MCCOIN, C.S.; STIERWALT, H.; ALLEN, J.; GANDHI, S.; PERRY, C.G.R.; JAMBAL, P.; SHANKAR, K.; THYFAULT, J.P. Oral combined contraceptives induce liver mitochondrial reactive oxygen species and whole‐body metabolic adaptations in female mice. The Journal of Physiology, v.600, n.24, p.5215–5245, 2022. doi: 10.1113/JP283733

GAO, M.; MA, Y.; LIU, D. High-Fat Diet-Induced Adiposity, Adipose Inflammation, Hepatic Steatosis and Hyperinsulinemia in Outbred CD-1 Mice. PLOS ONE, v.10, n.3, p.e0119784, 2015. doi: 10.1371/journal.pone.0119784

GOUVEIA, T.V.C.; AGUIAR, G.S.; CHAVES, J.O.; NASCIMENTO, D.S.C.; OLIVEIRA, C.A.R.; RIBEIRO, R.A.; LATINI, J.T.P.; BLANC, H.N.H. Efeito do uso ininterrupto de contraceptivo oral combinado na vagina de camundongos. Em: Tecnologia e Inovação para o Cuidar em Enfermagem. Atena Editora, p.99–109. 2020. doi: 10.22533/at.ed.94820261010

GRAAFLAND, L.; ABBOTT, M.; ACCORDINO, M. Breast Cancer Risk Related to Combined Oral Contraceptive Use. The Journal for Nurse Practitioners, v.16, n.2, p.116–120, 2020. doi: 10.1016/j.nurpra.2019.11.018

HENRIQUES, H.N. Efeito do uso contínuo de hormônios esteroides sexuais na mama de ratas Wistar. [Tese de Doutorado]. Niterói: Universidade Federal Fluminense, 2013.

HILLERS-ZIEMER, L.E.; ARENDT, L.M. Weighing the Risk: effects of Obesity on the Mammary Gland and Breast Cancer Risk. Journal of Mammary Gland Biology and Neoplasia, v.25, n.2, p.115–131, 2020. doi: 10.1007/s10911-020-09452-5

HIRSCHBERG, A.L.; TANI, E.; BRISMAR, K.; LUNDSTRÖM, E. Effects of drospirenone and norethisterone acetate combined with estradiol on mammographic density and proliferation of breast epithelial cells—A prospective randomized trial. Maturitas, v.126, p.18–24, 2019. doi: 10.1016/j.maturitas.2019.04.205

IYENGAR, N.M.; GUCALP, A.; DANNENBERG, A.J.; HUDIS, C.A. Obesity and Cancer Mechanisms: Tumor Microenvironment and Inflammation. Journal of Clinical Oncology, v.34, n.35, p.4270–4276, 2016. doi: 10.1200/JCO.2016.67.4283

KIRAN, H.; TOK, A.; YÜKSEL, M.; ARIKAN, D.C.; EKERBICER, H.C. Estradiol plus drospirenone therapy increases mammographic breast density in perimenopausal women. European Journal of Obstetrics & Gynecology and Reproductive Biology, v.159, n.2, p.384–387, 2011. doi: 10.1016/j.ejogrb.2011.09.023

KOTSIFAKI, A.; MAROULAKI, S.; KARALEXIS, E.; STATHAKI, M.; ARMAKOLAS, A. Decoding the Role of Insulin-like Growth Factor 1 and Its Isoforms in Breast Cancer. International Journal of Molecular Sciences, v.25, n.17, p.9302, 2024. doi: 10.3390/ijms25179302

KRATTENMACHER, R. Drospirenone: pharmacology and pharmacokinetics of a unique progestogen. Contraception, v.62, n.1, p.29–38, 2000. doi: 10.1016/s0010-7824(00)00133-5

KUBOTA, N.; TERAUCHI, Y.; MIKI, H.; TAMEMOTO, H.; YAMAUCHI, T.; KOMEDA, K.; SATOH, S.; NAKANO, R.; ISHII, C.; SUGIYAMA, T.; ETO, K.; TSUBAMOTO, Y.; OKUNO, A.; MURAKAMI, K.; SEKIHARA, H.; HASEGAWA, G.; NAITO, M.; TOYOSHIMA, Y.; TANAKA, S.; SHIOTA, K.; KITAMURA, T.; FUJITA, T.; EZAKI, O.; AIZAWA, S.; NAGAI, R.; TOBE1, K.; KIMURA, S.; KADOWAKI, T. PPARγ Mediates High-Fat Diet–Induced Adipocyte Hypertrophy and Insulin Resistance. Molecular Cell, v.4, n.4, p.597–609, 1999. doi: 10.1016/s1097-2765(00)80210-5

LAUBY-SECRETAN, B.; SCOCCIANTI, C.; LOOMIS, D.; GROSSE, Y.; BIANCHINI, F.; STRAIF, K. Body Fatness and Cancer — Viewpoint of the IARC Working Group. New England Journal of Medicine, v.375, n.8, p.794–798, 2016. doi: 10.1056/NEJMsr1606602

LEE, J.S.; TOCHENY, C.E.; SHAW, L.M. The Insulin-like Growth Factor Signaling Pathway in Breast Cancer: An Elusive Therapeutic Target. Life, v.12, n.12, p.1992, 2022. doi: 10.3390/life12121992

LIU, N.Q.; CAO, W.H.; WANG, X.; CHEN, J.; NIE, J. Cyclin genes as potential novel prognostic biomarkers and therapeutic targets in breast cancer. Oncology Letters, v.24, n.4, p.374, 2022.

MENEZES, C.A.; OLIVEIRA, V.S.; BARRETO, R.F. Estudo da correlação entre obesidade e câncer de mama no período pré e pós-menopausa / Study of the correlation between obesity and breast cancer in the pre and post-menopause period. Brazilian Journal of Health Review, v.4, n.1, p.1487–1501, 2021. doi: 10.3892/ol.2022.13494

MØRCH, L.S.; SKOVLUND, C.W.; HANNAFORD, P.C.; IVERSEN, L.; FIELDING, S.; LIDEGAARD, Ø. Contemporary Hormonal Contraception and the Risk of Breast Cancer. New England Journal of Medicine, v.377, n.23, p.2228–2239, 2017. doi: 10.1056/NEJMoa1700732

MUHN, P.; KRATTENMACHER, R.; BEIER, S.; ELGER, W.; SCHILLINGER, E. Drospirenone: A novel progestogen with antimineralocorticoid and antiandrogenic activity. Contraception, v.51, n.2, p.99–110, 1995. doi: 10.1016/0010-7824(94)00015-o

NERY, L.C.D.E.S.; BRAZ, L.C.S.; FERREIRA, L.L.D.M.; VIEIRA, F.P.; SILVA, L.L.; BLANC, H.N.H.; RAIMUNDO, J.M. A combined injectable contraceptive improves plasma redox status and does not induce vascular changes in female rats. Anais da Academia Brasileira de Ciências, v.93, n.3, 2021. doi: 10.1590/0001-3765202120201924

NGUYEN, H.L.; GEUKENS, T.; MAETENS, M.; APARICIO, S.; BASSEZ, A.; BORG, A.; BROCK, J.; BROEKS, A.; CALDAS, C.; CARDOSO, F.; SCHEPPER, M.; DELORENZI, M.; DRUKKER, C.A.; GLAS, A.M.; GREEN, A.R.; ISNALDI, E.; EYFJÖRÐ, J.; KHOUT, H.; KNAPPSKOG, S.; KRISHNAMURTHY, S.; LAKHANI, S.R.; LANGEROD, A.; MARTENS, J.W.M.; MCCART REED, A.E.; MURPHY, L.; NAULAERTS, S.; NIK-ZAINAL, S.; NEVELSTEEN, I.; NEVEN, P.; PICCART, M.; PONCET, C.; PUNIE, K.; PURDIE, C.; RAKHA, E.A.; RICHARDSON, A.; RUTGERS, E.; VINCENT-SALOMON, A.; SIMPSON, P.T.; SCHMIDT, M.K.; SOTIRIOU, C.; SPAN, P.N.; TAN, K.T.B.; THOMPSON, A.; TOMMASI, S.; BAELEN, K.V.; VIJVER, M.V.; LAERE, S.V.; VAN’T VEER, L.; VIALE, G.; VIARI, A.; VOS, H.; WITTEVEEN, A.T.; WILDIERS, H.; FLORIS,G.; GARG, A.D.; SMEETS, A. LAMBRECHTS, D.; BIGANZOLI, E.; RICHARD, F.; DESMEDT, C. Obesity-associated changes in molecular biology of primary breast cancer. Nature Communications, v.14, n.1, p.4418, 2023. doi: 10.1038/s41467-023-39996-z

OLIVEIRA, C.A.R.; ARAUJO, T.R.; AGUIAR, G.S.; DA SILVA JUNIOR, J.A.; VETTORAZZI, J.F.; FREITAS, I.N.; OLIVEIRA, K.M.; BOSCHERO, A.C.; BONFLEUR, M.L.; CLARKE, J.R.; HENRIQUES, H.N.; RIBEIRO, R.A. Combined oral contraceptive in female mice causes hyperinsulinemia due to β-cell hypersecretion and reduction in insulin clearance. Journal of Steroid Biochemistry and Molecular Biology, v.190, p.54–63, 2019. doi: 10.1016/j.jsbmb.2019.03.018

OLIVEIRA, K.M.; FIGUEIREDO, L.S.; ARAUJO, T.R.; FREITAS, I.N.; SILVA, J.N.; BOSCHERO, A.C.; RIBEIRO, R.A. Prolonged bisphenol-A exposure decreases endocrine pancreatic proliferation in response to obesogenic diet in ovariectomized mice. Steroids, v.160, p.108658, 2020. doi: 10.1016/j.steroids.2020.108658

OTTO, C.; FUCHS, I.; ALTMANN, H.; KLEWER, M.; WALTER, A.; PRELLE, K.; VONK, R.; FRITZEMEIER, K.H. Comparative Analysis of the Uterine and Mammary Gland Effects of Drospirenone and Medroxyprogesterone Acetate. Endocrinology, v.149, n.8, p.3952–3959, 2008. doi: 10.1210/en.2007-1612

POMPEI, L.M.; CARVALHO, F.M.; ORTIZ, S.C.B.C.; MOTTA, M.C.; CRUZ, R.J.; MELO, N.R. Morphometric evaluation of effects of two sex steroids on mammary gland of female rats. Maturitas, v.51, n.4, p.370–379, 2005. doi: 10.1016/j.maturitas.2004.09.007

RAIMONDI, G.M.; ENG, A.K.; KENNY, M.P.; BRITTING, M.A.; OSTROFF, L.E. Track-by-Day: A standardized approach to estrous cycle monitoring in biobehavioral research. Behavioural Brain Research, v.461, p.114860, 2024. doi: 10.1016/j.bbr.2024.114860

ROSENBAUM, P.; SCHMIDT, W.; HELMERHORST, F.M.; WUTTKE, W.; ROSSMANITH, W.; FREUNDL, F.; THOMAS, K.; GRILLO, M.; WOLF, A.; HEITHECKER, R. Inhibition of ovulation by a novel progestogen (drospirenone) alone or in combination with ethinylestradiol. The European Journal of Contraception & Reproductive Health Care, v.5, n.1, p.16–24, 2000. doi: 10.1080/13625180008500376

SKOVLUND, C.W.; MØRCH, L.S.; KESSING, L.V.; LIDEGAARD, Ø. Association of Hormonal Contraception With Depression. JAMA Psychiatry, v.73, n.11, p.1154, 2016. doi: 10.1001/jamapsychiatry.2016.2387

SMITH, M.S.; FREEMAN, M.E.; NEILL, J.D. The control of progesterone secretion during the estrous cycle and early pseudopregnancy in the rat: prolactin, gonadotropin and steroid levels associated with rescue of the corpus luteum of pseudopregnancy. Endocrinology, v.96, n.1, p.219–226, 1975. doi: 10.1210/endo-96-1-219

SMOLAREK, A.K.; SO, J.Y.; THOMAS, P.E.; LEE, H.J.; PAUL, S.; DOMBROWSKI, A.; WANG, C.X.; SAW, C.L.L.; KHOR, T.O.; KONG, A.N.T.; REUHL, K.; LEE, M.J.; YANG, C.S.; SUH, N. Dietary tocopherols inhibit cell proliferation, regulate expression of ERα, PPARγ, and Nrf2, and decrease serum inflammatory markers during the development of mammary hyperplasia. Molecular Carcinogenesis, v.52, n.7, p.514–525, 2013. doi: 10.1002/mc.21886

TIWARI, S.; KAUR, K. Clinico-pathological association of BCL2 in invasive breast carcinoma: A study from tertiary care health centre in Northern India. Indian Journal of Pathology and Microbiology, v.68, n.2, p.324–327, 2025. doi: 10.4103/ijpm.ijpm_259_24

TOLG, C.; COWMAN, M.; TURLEY, E. Mouse Mammary Gland Whole Mount Preparation and Analysis. BIO-PROTOCOL, v.8, n.13, 2018. doi: 10.21769/BioProtoc.2915

UNITED NATIONS. United Nations Department of Economy and Social Affairs. Population Division 2024. World Contraceptive Use 2024.

VALENTINE, J.M.; AHMADIAN, M.; KEINAN, O.; ABU-ODEH, M.; ZHAO, P.; ZHOU, X.; KELLER, M.P.; GAO, H.; YU, R.T.; LIDDLE, C.; DOWNES, M.; ZHANG, J.; LUSIS, A.J.; ATTIE, A.D.; EVANS, R.M.; RYDÉN, M.; SALTIEL, A.R. β3-Adrenergic receptor downregulation leads to adipocyte catecholamine resistance in obesity. Journal of Clinical Investigation, v.132, n.2, 2022. doi: 10.1172/JCI153357

WAWRZKIEWICZ-JAŁOWIECKA, A.; LALIK, A.; SOVERAL, G. Recent Update on the Molecular Mechanisms of Gonadal Steroids Action in Adipose Tissue. International Journal of Molecular Sciences, v.22, n.10, p.5226, 2021. doi: 10.3390/ijms22105226

WORLD OBESITY FEDERATION [Internet]. Prevalence of adult overweight & obesity. Acesso em: 24 ago. 2024. Disponível em: <https://data.worldobesity.org/tables/prevalence-of-adult-overweight-obesity-2/>.

Published

2025-07-24

How to Cite

AGUIAR , G. de S.; CHAVES (IN MEMORIAN), J. de O.; FREITAS, I. N.; OLIVEIRA, K. M. de; SILVA, A. B. B. da; SILVA, S. C. da; RIBEIRO , R. A.; BLANC, H. N. H. Effects of continuous ethinylestradiol and drospirenone administration on body mass and mammary gland in female mice fed a standard or high-fat diet. Journal of Nutrition and Health Surveillance, Fortaleza, v. 12, n. 1, p. e15667, 2025. DOI: 10.52521/nutrivisa.v12i1.15667. Disponível em: https://revistas.uece.br/index.php/nutrivisa/article/view/15667. Acesso em: 5 dec. 2025.

Issue

Section

Artigos originais