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demethylation, PPARg promoter leads to reduction in ovarian granulosa cells, Aromatase (Cyp19a1)
Key Event Relationship Overview
AOPs Referencing Relationship
Life Stage Applicability
Key Event Relationship Description
This KER establishes the link between PPARγ activation and reduced levels of aromatase in ovarian granulosa cells. Aromatase is a key enzyme in steroidogenesis, catalysing the conversion of androgens to estrogens.
Evidence Collection Strategy
Evidence Supporting this KER
Peroxisome proliferator-activated receptor γ (PPARγ) is master switch of lipid metabolism and cell differentiation, their role has also been acknowledged in regulation of reproductive function and development [reviewed by (P Froment et al., 2006), (Minge, Robker, & Norman, 2008)]. The PPARs are implicated in regulation of steroidogenesis from in vitro data [reviewed by (Carolyn M Komar, 2005)].
PPARγ involvement in aromatase regulation in granulosa cells
The PPARγ is activated upon the ligand binding in granulosa cells, and then indirectly alters the expression of aromatase, the rate-limiting enzyme in conversion of androgens to estrogens (Kwintkiewicz, Nishi, Yanase, & Giudice, 2010), (Lovekamp-Swan, Jetten, & Davis, 2003), (Mu et al., 2000). The ligands of PPARγ were also shown to regulate other enzymes involved in steroidogenesis (Dupont, Chabrolle, Ramé, Tosca, & Coyral-Castel, 2008). All PPAR isoforms have been detected in both human and rodent ovary [reviewed by (Carolyn M Komar, 2005)]. In female rats the PPARγ have been detected in granulosa cells.
• PPARγ is primarily expressed in the granulosa cells and pre-ovulatory follicles, less strongly in the theca cells and corpus luteum where its expression increases after ovulation and falls after the LH surge, (C M Komar, Braissant, Wahli, & Curry, 2001). In the absence of fertilization or embryo implantation, PPARγ expression decreases as a result of corpus luteum regression (Viergutz, Loehrke, Poehland, Becker, & Kanitz, 2000).
• PPARγ is directly involved in oocyte maturation and ovulation [reviewed by (P Froment et al., 2006)]
Additional studies have shown that PPARγ is active in the ovary (P Froment et al. 2003).
The precise molecular mechanism by which PPARγ regulates aromatase is unclear given the fact that the proximal promoter regulating aromatase expression in the rat ovary does not contain an obvious peroxisome proliferator response element (PPRE) (Young and McPhaul 1997). There are plausible ways by which the PPARγ (as transcriptionally active PPAR:RXR heterodimer) could modify the transcription of aromatase including activation of RXR competition for binding sites on DNA and competition for limiting co-activators required for gene transcription. A new insight in the mechanism of regulation of the aromatase gene and activation of PPAR gamma and RXR was brought by Fan et al proposing disruption of NF-κB interaction with the aromatase promoter (Fan et al. 2005). The authors showed that activation of PPARγ and RXR impaired the interaction between NF-κB and aromatase promoter II and the p65 based transcription in both ovarian and fibroblast cells in a PPARγ-dependent manner (Fan et al. 2005). Studies supporting that hypothesis show that both PPARγ ligand (Troglitazone) and RXR ligand (LG100268) suppress aromatase activity in human granulosa cells (Mu et al. 2000), (Mu et al. 2001) and together causing a greater reduction than either compound alone (Mu et al. 2000).
Another possibility is that PPARγ is able to modify protein–protein interactions involved in the transcription of aromatase. Activation of PPARγ may recruit cofactors away from aromatase to inhibit normal transcription. Further studies are necessary to determine how PPARγ transcriptionally repress aromatase.
Uncertainties and Inconsistencies
There is substantial evidence in literature supporting the KER, however the underlying mechanism are to be investigated, together with other pathways involved in aromatase down regulation. The pattern of the PPARγ expression in ovarian follicles is not steady, unlike expression of PPARs α and δ. This fact adds to the complexity to the interpretation of mechanisms involved in the pathway. The PPARγ is down-regulated in response to the LH surge (C M Komar, Braissant, Wahli, & Curry, 2001), but only in follicles that have responded to the LH surge (Carolyn M Komar & Curry, 2003). Because PPARγ is primarily expressed in granulosa cells, it may influence development of these cells and their ability to support normal oocyte maturation. PPARγ could also potentially affect somatic cell/oocyte communication not only by impacting granulosa cell development, but by direct effects on the oocyte. Modulation of the PPARγ activity/expression in the ovary therefore, could potentially affect oocyte developmental competence. Some evidence implies that the regulatory role of PPARγ might be connected to the other events in estradiol synthesis like the impairment of cholesterol transport to mitochondria (Cui et al., 2002).
PPARα The experimental data supports the parallel involvement of another member of PPAR superfamily of nuclear receptors, PPARα. PPARα was shown to be implicated in the down regulation of aromatase in rat: in vitro (Lovekamp-Swan et al., 2003); in vivo (Xu et al., 2010) and in mice in vivo (Toda, Okada, Miyaura, & Saibara, 2003). The ovarian aromatase promoter contains one half of a PPRE (peroxisome proliferator response element), which is the binding site for steroidogenic factor 1 (SF-1) (Young & McPhaul, 1997). While it is unknown whether PPARα can compete for binding on an incomplete response element, disruption of SF-1 binding to this half site would disrupt normal aromatase transcription. Studies by S. Plummer et al showed that PPARα and SF1 share a common coactivator (S. Plummer, Sharpe, Hallmark, Mahood, & Elcombe, 2007), (S. M. Plummer et al., 2013), CREB-binding protein (CBP), which is present in limiting concentrations (McCampbell, 2000). Binding of CBP to PPARα could therefore starve SF1 a cofactor essential for its transactivation functions. Surprisingly, aromatase levels were increased in ovaries of PPARα-null mice upon treatment with PPARα ligand (Toda et al., 2003).
PPARα was also reported to regulate other enzymes involved in steroidogenesis like: 17 beta-hydroxysteroid dehydrogenase type IV (HSD IV) (Corton et al., 1996), 3 beta-hydroxysteroid dehydrogenase (Wong, Ye, Muhlenkamp, & Gill, 2002) or 11beta-hydroxysteroid dehydrogenase type (Hermanowski-Vosatka et al., 2000). While PPARα/γ activators (like MEHP ) suppress aromatase, they showed no effect on Cholesterol side-chain cleavage enzyme (P450scc) in granulosa cells, demonstrating a more specific effect on steroidogenesis (Lovekamp-Swan et al., 2003). Experiments with PPARα-null mice indicate involvement of the receptor in reproductive toxicity, however cannot be entirely explained by the activation of PPARα mediated pathway as PPARα-null mice remain sensitive to DEHP-mediated reproductive toxicity (Ward et al. 1998), which implies other players including PPARγ. The above evidence supports the involvement of PPARα in regulation of steroidogenesis on its different steps. As PPARα is found primarily in the theca and stroma and the expression of PPARα in granulosa cells is very low (Carolyn M Komar, 2005) therefore it might be involved in steps in steroidogenesis upstream of aromatase.
Retinoid X Receptor (RXR)
Chemicals are able to activate RXR–PPARγ through RXR because this heterodimer interacts poorly with co-repressors in vivo and belongs to the group of so-called ‘permissive’ heterodimers, which can be stimulated by RXR ligands on their own (Germain, Iyer, Zechel, & Gronemeyer, 2002). Studies demonstrated that a PPARγ ligand and/or a RXR ligand decreased the aromatase activity in both cultured human ovarian granulosa cells (Mu et al., 2000), (Mu et al., 2001) and human granulosa-like tumor KGN cells (Kwintkiewicz et al., 2010) and combined treatment causes a greater reduction than either compound alone (Mu et al., 2000), (Mu et al., 2001).
No effect on aromatase protein expression was observed after PPARγ ligand (rosiglitazone) treatment in porcine ovarian follicles (Rak-Mardyła & Karpeta, 2014).
Known modulating factors
Known Feedforward/Feedback loops influencing this KER
Domain of Applicability
See the Table 1.
Barak, Y., Nelson, M. C., Ong, E. S., Jones, Y. Z., Ruiz-Lozano, P., Chien, K. R., … Evans, R. M. (1999). PPAR gamma is required for placental, cardiac, and adipose tissue development. Molecular Cell, 4(4), 585–95.
Bhattacharya, N., Dufour, J. M., Vo, M.-N., Okita, J., Okita, R., & Kim, K. H. (2005). Differential effects of phthalates on the testis and the liver. Biology of Reproduction, 72(3), 745–54. doi:10.1095/biolreprod.104.031583
Corton, J., Bocos, C., Moreno, E., Merritt, A., Marsman, D., Sausen, P., … Gustafsson, J. (1996). Rat 17 beta-hydroxysteroid dehydrogenase type IV is a novel peroxisome proliferator-inducible gene. Mol. Pharmacol., 50(5), 1157–1166.
Cui, Y., Miyoshi, K., Claudio, E., Siebenlist, U. K., Gonzalez, F. J., Flaws, J., … Hennighausen, L. (2002). Loss of the peroxisome proliferation-activated receptor gamma (PPARgamma ) does not affect mammary development and propensity for tumor formation but leads to reduced fertility. The Journal of Biological Chemistry, 277(20), 17830–5. doi:10.1074/jbc.M200186200
Dufour, J. M., Vo, M.-N., Bhattacharya, N., Okita, J., Okita, R., & Kim, K. H. (2003). Peroxisome proliferators disrupt retinoic acid receptor alpha signaling in the testis. Biology of Reproduction, 68(4), 1215–24. doi:10.1095/biolreprod.102.010488
Dupont, J., Chabrolle, C., Ramé, C., Tosca, L., & Coyral-Castel, S. (2008). Role of the peroxisome proliferator-activated receptors, adenosine monophosphate-activated kinase, and adiponectin in the ovary. PPAR Research, 2008, 176275. doi:10.1155/2008/176275
Dupont, J., Reverchon, M., Cloix, L., Froment, P., & Ramé, C. (2012). Involvement of adipokines, AMPK, PI3K and the PPAR signaling pathways in ovarian follicle development and cancer. The International Journal of Developmental Biology, 56(10-12), 959–67. doi:10.1387/ijdb.120134jd
Fan, W., Yanase, T., Morinaga, H., Mu, Y.-M., Nomura, M., Okabe, T., … Nawata, H. (2005). Activation of peroxisome proliferator-activated receptor-gamma and retinoid X receptor inhibits aromatase transcription via nuclear factor-kappaB. Endocrinology, 146(1), 85–92. doi:10.1210/en.2004-1046
Feige, J. N., Gelman, L., Rossi, D., Zoete, V., Métivier, R., Tudor, C., … Desvergne, B. (2007). The endocrine disruptor monoethyl-hexyl-phthalate is a selective peroxisome proliferator-activated receptor gamma modulator that promotes adipogenesis. The Journal of Biological Chemistry, 282(26), 19152–66. doi:10.1074/jbc.M702724200
Froment, P., Fabre, S., Dupont, J., Pisselet, C., Chesneau, D., Staels, B., & Monget, P. (2003). Expression and functional role of peroxisome proliferator-activated receptor-gamma in ovarian folliculogenesis in the sheep. Biology of Reproduction, 69(5), 1665–74. doi:10.1095/biolreprod.103.017244
Froment, P., Gizard, F., Defever, D., Staels, B., Dupont, J., & Monget, P. (2006). Peroxisome proliferator-activated receptors in reproductive tissues: from gametogenesis to parturition. The Journal of Endocrinology, 189(2), 199–209. doi:10.1677/joe.1.06667
Germain, P., Iyer, J., Zechel, C., & Gronemeyer, H. (2002). Co-regulator recruitment and the mechanism of retinoic acid receptor synergy. Nature, 415(6868), 187–92. doi:10.1038/415187a Hermanowski-Vosatka, A., Gerhold, D., Mundt, S. S., Loving, V. A., Lu, M., Chen, Y., … Thieringer, R. (2000). PPARalpha agonists reduce 11beta-hydroxysteroid dehydrogenase type 1 in the liver. Biochemical and Biophysical Research Communications, 279(2), 330–6. doi:10.1006/bbrc.2000.3966
Hurst, C. H., & Waxman, D. J. (2003). Activation of PPAR ␣ and PPAR ␥ by Environmental Phthalate Monoesters, 308, 297–308. doi:10.1093/toxsci/kfg145
Kaya, T., Mohr, S. C., Waxman, D. J., & Vajda, S. (2006). Computational screening of phthalate monoesters for binding to PPARgamma. Chemical Research in Toxicology, 19(8), 999–1009. doi:10.1021/tx050301s
Komar, C. M. (2005). Peroxisome proliferator-activated receptors (PPARs) and ovarian function--implications for regulating steroidogenesis, differentiation, and tissue remodeling. Reproductive Biology and Endocrinology : RB&E, 3, 41. doi:10.1186/1477-7827-3-41
Komar, C. M., Braissant, O., Wahli, W., & Curry, T. E. (2001). Expression and localization of PPARs in the rat ovary during follicular development and the periovulatory period. Endocrinology, 142(11), 4831–8. doi:10.1210/endo.142.11.8429
Kwintkiewicz, J., Nishi, Y., Yanase, T., & Giudice, L. C. (2010). Peroxisome proliferator-activated receptor-gamma mediates bisphenol A inhibition of FSH-stimulated IGF-1, aromatase, and estradiol in human granulosa cells. Environmental Health Perspectives, 118(3), 400–6. doi:10.1289/ehp.0901161
Lapinskas, P. J., Brown, S., Leesnitzer, L. M., Blanchard, S., Swanson, C., Cattley, R. C., & Corton, J. C. (2005). Role of PPARα in mediating the effects of phthalates and metabolites in the liver. Toxicology, 207(1), 149–163.
Lovekamp-Swan, T., Jetten, A. M., & Davis, B. J. (2003). Dual activation of PPARalpha and PPARgamma by mono-(2-ethylhexyl) phthalate in rat ovarian granulosa cells. Molecular and Cellular Endocrinology, 201(1-2), 133–41.
Luebker, D. J., Hansen, K. J., Bass, N. M., Butenhoff, J. L., & Seacat, A. M. (2002). Interactions of fluorochemicals with rat liver fatty acid-binding protein. Toxicology, 176(3), 175–85.
Maloney, E. K., & Waxman, D. J. (1999). trans-Activation of PPARα and PPARγ by Structurally Diverse Environmental Chemicals. Toxicology and Applied Pharmacology, 161(2), 209–218.
McCampbell, A. (2000). CREB-binding protein sequestration by expanded polyglutamine. Human Molecular Genetics, 9(14), 2197–2202. doi:10.1093/hmg/9.14.2197 Minge, C. E., Robker, R. L., & Norman, R. J. (2008). PPAR Gamma: Coordinating Metabolic and Immune Contributions to Female Fertility. PPAR Research, 2008, 243791. doi:10.1155/2008/243791
Mu, Y. M., Yanase, T., Nishi, Y., Takayanagi, R., Goto, K., & Nawata, H. (2001). Combined treatment with specific ligands for PPARgamma:RXR nuclear receptor system markedly inhibits the expression of cytochrome P450arom in human granulosa cancer cells. Molecular and Cellular Endocrinology, 181(1-2), 239–48.
Mu, Y. M., Yanase, T., Nishi, Y., Waseda, N., Oda, T., Tanaka, A., … Nawata, H. (2000). Insulin sensitizer, troglitazone, directly inhibits aromatase activity in human ovarian granulosa cells. Biochemical and Biophysical Research Communications, 271(3), 710–3. doi:10.1006/bbrc.2000.2701
Plummer, S. M., Dan, D., Quinney, J., Hallmark, N., Phillips, R. D., Millar, M., … Elcombe, C. R. (2013). Identification of transcription factors and coactivators affected by dibutylphthalate interactions in fetal rat testes. Toxicological Sciences : An Official Journal of the Society of Toxicology, 132(2), 443–57. doi:10.1093/toxsci/kft016
Plummer, S., Sharpe, R. M., Hallmark, N., Mahood, I. K., & Elcombe, C. (2007). Time-dependent and compartment-specific effects of in utero exposure to Di(n-butyl) phthalate on gene/protein expression in the fetal rat testis as revealed by transcription profiling and laser capture microdissection. Toxicological Sciences : An Official Journal of the Society of Toxicology, 97(2), 520–32. doi:10.1093/toxsci/kfm062
Rak-Mardyła, A., & Karpeta, A. (2014). Rosiglitazone stimulates peroxisome proliferator-activated receptor gamma expression and directly affects in vitro steroidogenesis in porcine ovarian follicles. Theriogenology, 82(1), 1–9. doi:10.1016/j.theriogenology.2014.02.016
Reinsberg, J., Wegener-Toper, P., van der Ven, K., van der Ven, H., & Klingmueller, D. (2009). Effect of mono-(2-ethylhexyl) phthalate on steroid production of human granulosa cells. Toxicology and Applied Pharmacology, 239(1), 116–23. doi:10.1016/j.taap.2009.05.022
Rotman, N., Haftek-Terreau, Z., Lücke, S., Feige, J., Gelman, L., Desvergne, B., & Wahli, W. (2008). PPAR Disruption: Cellular Mechanisms and Physiological Consequences. CHIMIA International Journal for Chemistry, 62(5), 340–344. doi:10.2533/chimia.2008.340
Saitoh, M., Yanase, T., Morinaga, H., Tanabe, M., Mu, Y. M., Nishi, Y., Nomura, M., Okabe, T., Goto, K., Takayanagi, R., and Nawata, H. (2001). Tributyltin or triphenyltin inhibits aromatase activity in the human granulosa-like tumor cell line KGN. Biochem. Biophys. Res. Commun., 289, 198–204.
Toda, K., Okada, T., Miyaura, C., & Saibara, T. (2003). Fenofibrate, a ligand for PPARalpha, inhibits aromatase cytochrome P450 expression in the ovary of mouse. Journal of Lipid Research, 44(2), 265–70. doi:10.1194/jlr.M200327-JLR200
ToxCastTM Data. “ToxCastTM Data.” US Environmental Protection Agency. http://www.epa.gov/ncct/toxcast/data.html.
Venkata, N. G., Robinson, J. a, Cabot, P. J., Davis, B., Monteith, G. R., & Roberts-Thomson, S. J. (2006). Mono(2-ethylhexyl)phthalate and mono-n-butyl phthalate activation of peroxisome proliferator activated-receptors alpha and gamma in breast. Toxicology Letters, 163(3), 224–34. doi:10.1016/j.toxlet.2005.11.001
Viergutz, T., Loehrke, B., Poehland, R., Becker, F., & Kanitz, W. (2000). Relationship between different stages of the corpus luteum and the expression of the peroxisome proliferator-activated receptor gamma protein in bovine large lutein cells. Journal of Reproduction and Fertility, 118(1), 153–61.
Willson, T. M., Brown, P. J., Sternbach, D. D., & Henke, B. R. (2000). The PPARs: from orphan receptors to drug discovery. Journal of Medicinal Chemistry, 43(4), 527–50.
Wong, J. S., Ye, X., Muhlenkamp, C. R., & Gill, S. S. (2002). Effect of a peroxisome proliferator on 3 beta-hydroxysteroid dehydrogenase. Biochemical and Biophysical Research Communications, 293(1), 549–53. doi:10.1016/S0006-291X(02)00235-8
Xu, C., Chen, J.-A., Qiu, Z., Zhao, Q., Luo, J., Yang, L., … Shu, W. (2010). Ovotoxicity and PPAR-mediated aromatase downregulation in female Sprague-Dawley rats following combined oral exposure to benzo[a]pyrene and di-(2-ethylhexyl) phthalate. Toxicology Letters, 199(3), 323–32. doi:10.1016/j.toxlet.2010.09.015
Young, M., & McPhaul, M. J. (1997). Definition of the elements required for the activity of the rat aromatase promoter in steroidogenic cell lines. The Journal of Steroid Biochemistry and Molecular Biology, 61(3-6), 341–8.
Synthetic ligand, rosiglitazone stimulates AMP-activated protein kinase (AMPK) and enhances the meiotic resumption of mouse oocytes (Dupont, Reverchon, Cloix, Froment, & Ramé, 2012).