Downregulation of the Expression of Steroidogenic Acute Regulatory Protein and Aromatase in Steroidogenic KGN Human Granulosa Cells after Exposure to Bisphenol A

다국어 초록 (Multilingual Abstract)

Although increasing evidence of cause-and-effect relationship between BPA exposure and female reproductive disorders have been suggested through many studies, the precise biochemical and molecular mechanism(s) by which BPA interferes with steroidogenesis in the ovarian cells still remain unclear. Therefore, the purpose of this study was to discover the steroidogenic biomarker(s) associated with BPA treatment in human granulosa cell line, KGN. In this study, our results obtained via the analysis of steroidogenesis-related protein expression in KGN cells using quantitative polymerase chain reaction (qPCR) and western blot analyses revealed that the expression levels of steroidogenic acute regulatory (StAR) and aromatase decreased considerably and gradually after BPA treatment in a dosedependent manner under BPA treatment. Further, remarkable decreases in their expression levels at the cellular levels were also confirmed via immunocytochemistry, and subsequent StAR and aromatase mRNA expression levels showed profiles similar to those observed for their proteins, i.e., both StAR and aromatase mRNA expression levels were significantly decreased under BPA treatment at concentrations ≥0.1 μM. We observed that follicle stimulating hormone upregulated StAR and aromatase protein expression levels; however, this effect was suppressed in the presence of BPA. Regarding the steroidogenic effects of BPA on KGN cells, controversies remain regarding the ultimate outcomes. Nevertheless, we believe that the results here presented imply that KGN cells have a good cellular and steroidogenic machinery for evaluating endocrine disruption. Therefore, StAR and aromatase could be stable and sensitive biomarkers in KGN cells for the cellular screening of the potential risk posed by exogenous and environmental chemicals to female reproductive (endocrine) function.

참고문헌 (Reference)

1 Kelleher AM, "Uterine glands: Developmental biology and functional roles in pregnancy" 40 : 1424-1445, 2019

2 Dilaver N, "The regulation and signalling of anti-Müllerian hormone in human granulosa cells: Relevance to polycystic ovary syndrome" 34 : 2467-2479, 2019

3 Stocco C, "The molecular control of corpus luteum formation, function, and regression" 28 : 117-149, 2007

4 Miller WL, "The molecular biology, biochemistry, and physiology of human steroidogenesis and its disorders" 32 : 81-151, 2011

5 Shi J, "The interference effects of bisphenol A on the synthesis of steroid hormones in human ovarian granulosa cells" 36 : 665-674, 2021

6 Ernst J, "The endocannabinoid system in the human granulosa cell line KGN" 423 : 67-76, 2016

7 Miller WL, "Steroidogenic enzymes" 13 : 1-18, 2008

8 Miller WL, "Steroidogenic acute regulatory protein (StAR), a novel mitochondrial cholesterol transporter" 1771 : 663-676, 2007

9 Takeuchi T, "Serum bisphenol a concentrations showed gender differences, possibly linked to androgen levels" 291 : 76-78, 2002

10 Pierre P, "Role of adiponectin receptors, AdipoR1 and AdipoR2, in the steroidogenesis of the human granulosa tumor cell line, KGN" 24 : 2890-2901, 2009

11 Palioura E, "Polycystic ovary syndrome (PCOS) and endocrine disrupting chemicals (EDCs)" 16 : 365-371, 2015

12 Kwintkiewicz J, "Peroxisome proliferator–activated receptor-γmediates bisphenol A inhibition of FSH-stimulated IGF-1, aromatase, and estradiol in human granulosa cells" 118 : 400-406, 2010

13 Schönfelder G, "Parent bisphenol A accumulation in the human maternal-fetal-placental unit" 110 : A703-A707, 2002

14 Channing CP, "Ovarian follicular and luteal physiology" 22 : 117-201, 1980

15 Ye X, "Measuring environmental phenols and chlorinated organic chemicals in breast milk using automated on-line column-switching–high performance liquid chromatography–isotope dilution tandem mass spectrometry" 831 : 110-115, 2006

16 Kuroda N, "Measurement of bisphenol A levels in human blood serum and ascitic fluid by HPLC using a fluorescent labeling reagent" 30 : 1743-1749, 2003

17 Yamada H, "Maternal serum and amniotic fluid bisphenol A concentrations in the early second trimester" 16 : 735-739, 2002

18 Leung PCK, "Interactions of steroids and gonadotropins in the control of steroidogenesis in the ovarian follicle" 42 : 71-82, 1980

19 Tremblay PG, "Gene analysis of major signaling pathways regulated by gonadotropins in human ovarian granulosa tumor cells (KGN)" 103 : 583-598, 2020

20 Hillier SG, "Follicular oestrogen synthesis: The ‘two-cell, twogonadotrophin’ model revisited" 100 : 51-54, 1994

21 Calafat AM, "Exposure of the U.S. population to bisphenol A and 4-tertiary-octylphenol: 2003–2004" 116 : 39-44, 2008

22 Ziv-Gal A, "Evidence for bisphenol A-induced female infertility: a review (2007–2016)" 106 : 827-856, 2016

23 Wallach EE, "Estrogen biosynthesis—regulation, action, remote effects, and value of monitoring in ovarian stimulation cycles" 65 : 687-701, 1996

24 Nishi Y, "Establishment and characterization of a steroidogenic human granulosa-like tumor cell line, KGN, that expresses functional folliclestimulating hormone receptor" 142 : 437-445, 2001

25 Adashi EY, "Endocrinology of the ovary" 9 : 815-827, 1994

26 Watanabe M, "Effects of bisphenol A on the expression of cytochrome P450 aromatase (CYP19) in human fetal osteoblastic and granulosa cell-like cell lines" 210 : 95-99, 2012

27 Sun Y, "Determination of bisphenol A in rat brain by microdialysis and column switching high-performance liquid chromatography with fluorescence detection" 16 : 319-326, 2002

28 Ikezuki Y, "Determination of bisphenol A concentrations in human biological fluids reveals significant early prenatal exposure" 17 : 2839-2841, 2002

29 Devoto L, "Control of human luteal steroidogenesis" 186 : 137-141, 2002

30 Pivonello C, "Bisphenol A: An emerging threat to female fertility" 18 : 22-, 2020

31 Nelson W, "Bisphenol A-induced mechanistic impairment of decidualization" 87 : 837-842, 2020

32 Qi J, "Bisphenol A decreases progesterone synthesis in human ovarian granulosa cells" 112 : 1843-1849, 2020

33 Bloom MS, "Bisphenol A and ovarian steroidogenesis" 106 : 857-863, 2016

34 Huang M, "Bisphenol A and its analogues bisphenol S, bisphenol F and bisphenol AF induce oxidative stress and biomacromolecular damage in human granulosa KGN cells" 253 : 126707-, 2020

35 Liu H, "A low dose of bisphenol A stimulates estradiol production by regulating β-catenin-FOXL2-CYP19A1 pathway in human ovarian granulosa cells" 583 : 192-198, 2021