International Journal of Reproductive BioMedicine

Published date: May 31 2025

Journal Title: International Journal of Reproductive BioMedicine

Issue title: International Journal of Reproductive BioMedicine (IJRM): Volume 23, Issue No. 4

Pages: 303 – 312

DOI: 10.18502/ijrm.v23i4.18782

Alifia Candra Puriastuti alifia.candra.fik@um.ac.idMidwifery Department, Faculty of Medicine, State University of Malang, Jalan Semarang No. 5, Malang

Margarita Maria MaramisDivision of Biological Psychiatry and Neuroscience, Faculty of Medicine Airlangga University, Medical Faculty, Jalan Prof. Dr. Moestopo No. 47, Universitas Airlangga, Surabaya

Jimmy Yanuar AnnasDepartment of Obstetrics and Gynecology, Medical Faculty, Universitas Airlangga, Jalan Prof. Dr. Moestopo No. 47, Surabaya

Reny I’tishomDepartment of Biomedical Science, Medical Faculty, Universitas Airlangga, Jalan Prof. Dr. Moestopo No. 47, Surabaya

Purwo SrirejekiDepartment of Medical Physiology, Medical Faculty, Universitas Airlangga, Jalan Prof. Dr. Moestopo No. 47, Surabaya

Sulistiawati ‎ Department of of IKM-KP, Medical Faculty, Universitas Airlangga, Jalan Prof. Dr. Moestopo No. 47, Surabaya

Background: The chronic unpredictable mild stress (CUMS) model involves applying mild stressors over a prolonged period, that can induce oxidative stress. Oxidative stress disrupts normal cellular functions, leading to reduced survival of antral follicles through increased glucocorticoids.

Objective: This study aims to identify the effect of stress on the regulation of glucocorticoid hormones and its impact on impaired antral follicles. Specifically, it focuses on the number of antral follicles using the CUMS model in female rats.

Materials and Methods: 16 female Wistar rats (5–6 months, 300–350 gr) were divided into 2 groups (n = 8/each), the control and the CUMS model. 24 hr after the last treatment, they were eter euthanized, a blood sample was taken from the intracardial to measure corticosteroid levels using the enzyme-linked immunoassay (ELISA) method, ovarian preparations were made, and then the histological sections were observed.

Results: After 22 days of CUMS, a significant difference was observed in corticosterone levels (p = 0.03), but no significant difference was observed in the number of antral follicles between the 2 groups (p = 0.57). However, histological analysis indicated substantial differences. The control group’s ovaries exhibited a higher proportion of healthy antral follicles.

Conclusion: CUMS increases glucocorticoids, which in turn causes a decrease in the number of antral follicles. This happens through 2 mechanisms: suppression of gonadotrophin-releasing hormone and direct effects on the ovaries that elevate granulosa cell apoptosis and follicular atresia, ultimately leading to a reduction in antral follicles.

Keywords: Chronic stress, Corticosterone, Oocyte

[1] Chang X. Transcriptome analysis of rats with chronic unpredictable mild stress treated with electroacupuncture. Brain Behav 2024; 14: e70045.

[2] López-López AL, Bonilla HJ, Villanueva MDCE, Brianza Padilla M, Vázquez Palacios G, Alarcón Aguilar FJ. Chronic unpredictable mild stress generates oxidative stress and systemic inflammation in rats. Physiol Behav 2016; 161: 15–23.

[3] Shirokova OM, Kuzmina DM, Zaborskaya OG, Shchelchkova NA, Kozliaeva EV, Korotchenko SA, et al. The long-term effects of chronic unpredictable mild stress experienced during adolescence could vary depending on biological sex. Int J Mol Sci 2025; 26: 1251.

[4] Kurt S, Koca RH, Hürkul MM, Şeker U, Köroğlu A. The antioxidant effect of Michauxia Campanuloides on rat ovaries. J Hellenic Vet Med Soc 2021; 72: 3163–3170.

[5] Ma J, Wang L, Yang D, Luo J, Gao J, Wang J, et al. Chronic stress causes ovarian fibrosis to impair female fertility in mice. Cell Signal 2024; 122: 111334.

[6] Scarlet D, Ille N, Ertl R, Alves BG, Gastal GDA, Paiva SO, et al. Glucocorticoid metabolism in equine follicles and oocytes. Domest Anim Endocrinol 2017; 59: 11–22.

[7] Fritz MA, Speroff L. Clinical gynecologic endocrinology and infertility. 9th Ed. Philadelphia: Lippincott Williams and Wilkins; 2019.

[8] Willy R. [The effect of frequency of exposure to light stressors during pregnancy on the number and apoptosis index of brain neuron cells in newborn mice (Mus musculus)]. [M.Sc. thesis]. Indonesia: University of Airlangga; 2016. (In Bahasa)

[9] Dwiningsih RS. [Effect of bone marrow msenchimal stem cell (BMSCs) transplantation on TNF-a receptor 1, mRNA BCL-2, mRNA p21, mRNA Survivin, mRNA VEGF, granulosa cells apoptosis and folliculogenesis in mice model endometriosis]. [Doctoral thesis]. Indonesia: University of Airlangga; 2017. (In Bahasa)

[10] da Costa NN, Brito KNL, di Paula Bessa Santana P, da Silva Cordeiro M, Silva TVG, Santos AX, et al. Effect of cortisol on bovine oocyte maturation and embryo development in vitro. Theriogenology 2016; 85: 323–329.

[11] Alberts B, Johnson A, Lewis J, Morgan D, Raff M, Roberts K, et al. Molecular biology of the cell. 7th Ed. Canada: W.W. Norton & Company; 2022.

[12] Whirledge S, Cidlowski JA. Glucocorticoids, stress, and fertility. Minerva Endocrinol 2010; 35: 109–125.

[13] Lewinski A, Brzozowska M. Female infertility as a result of stress-related hormonal changes. Gynecol Reprod Endocrinol Metab 2022; 3: 94–98.

[14] Li M, Jan SZ, Hamer G, van Pelt AM, van der Stelt I, Keijer J, et al. Preantral follicular atresia occurs mainly through autophagy, while antral follicles degenerate mostly through apoptosis. Biol Reprod 2018; 99: 853–863.

[15] Scudieri A, Valbonetti L, Peric T, Cotticelli A, Ramal-Sánchez M, Loi P, et al. Autophagy is involved in granulosa cell death and follicular atresia in ewe ovaries. Theriogenology 2024; 226: 236–242.

[16] Knapczyk-Stwora K, Grzesiak M, Witek P, Duda M, Koziorowski M, Słomczyńska M. Neonatal exposure to agonists and antagonists of sex steroid receptors affects AMH and FSH plasma level and their receptors expression in the adult pig ovary. Animals 2019; 10: 12.

[17] Wu H, Zhu R, Zheng B, Liao G, Wang F, Ding J, et al. Single-cell sequencing reveals an intrinsic heterogeneity of the preovulatory follicular microenvironment. Biomolecules 2022; 12: 231.

[18] Seo S, Jeon S, Ha JK. Guidelines for experimental design and statistical analyses in animal studies submitted for publication in the Asian-Australasian Journal of Animal Sciences. Asian-Australas J Anim Sci 2018; 31: 1381–1386.

[19] Ghasemi A, Jeddi S, Kashfi K. The laboratory rat: Age and body weight matter. EXCLI J 2021; 20: 1431–1445.

[20] Maramis MM, Mahajudin MS, Khotib J. Impaired cognitive flexibility and working memory precedes depression: A rat model to study depression. Neuropsychobiology 2021; 80: 225–233.

[21] Puriastuti AC, Maramis MM, Januar Annas J. [Chronic stress decreases the thickness of the granulosa cell layer of the antral follicle of rattus norvegicus]. J Biosains Pascasarjana 2017; 19: 1818. (In Indonesia)

[22] Rosdianto AM. [Reproductive performance of female white rats (Rattus Norvegicus Sp.) after administration of fertility herbs]. [Ph.D. thesis]. Indonesia: Bogor Argicultural University; 2016. (In Indonesia)

[23] Byers SL, Wiles MV, Dunn SL, Taft RA. Mouse estrous cycle identification tool and images. PLoS One 2012; 7: e35538.

[24] Strauss L, Junnila A, Wärri A, Manti M, Jiang Y, Löyttyniemi E, et al. Consistent and effective method to define the mouse estrous cycle stage by a deep learning-based model. J Endocrinol 2024; 261: e230204.

[25] Reece JB, Urry LA, Cain ML, Wasserman SA, Minorsky PV, Jackson RB. Campbell biology. 10th Ed. UK: Pearson Education; 2021.

[26] Botía M, Escribano D, Martínez-Subiela S, Tvarijonaviciute A, Tecles F, López-Arjona M, et al. Different types of glucocorticoids to evaluate stress and welfare in animals and humans: General concepts and examples of combined use. Metabolites 2023; 13: 106.

[27] Hall JE. Guyton and Hall : Textbook of medical physiology. 12th Ed. Philadelphia: Elsevier; 2020.

[28] Whirledge S, Cidlowski JA. Glucocorticoids and reproduction : Traffic control on the road to reproduction. Trends Endocrinol Metab 2017; 28: 399–415.

[29] Wei F, Fan X, del Valle JS, Asseler JD, van der Meeren LE, Cheng H, et al. Classification of atretic small antral follicles in the human ovary. Int J Mol Sci 2023; 24: 16846.

[30] Vidal JD, Dixon D. Boorman’s pathology of the rat: Reference and Atlas. 2nd Ed. USA: Academic Press; 2018.

[31] Baumgarten SC, Stocco C. Granulosa cells. In: Skinner MK. Encyclopedia of reproduction. Cham: Elsevier; 2018. 8–13.

[32] Regan SLP, Knight PG, Yovich JL, Leung Y, Arfuso F, Dharmarajan A. Granulosa cell apoptosis in the ovarian follicle-A changing view. Front Endocrinol 2018; 9: 61.