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Yasuharu Otsubo (1) Takao Ashikaga (1) Tomoki Fukuyama (1) Ken Goto (1) Shinko Hata (1) Shigeru Hisada (1) Shiho Ito (1) Hiroyuki Komatsu (1) Sumie Konishi (1) Tadashi Kosaka (1) Kiyoshi Kushima (1) Shogo Matsumura (1) Takumi Ohishi (1) Junichiro Sugimoto (1) Yasuhiro Yoshida (1)
(1) AOP Working Group, Testing Methodology Committee, The Japanese Society of Immunotoxicology
Corresponding author: Yasuharu Otsubo (email@example.com)
Point of Contact
Yasuharu Otsubo (email point of contact)
- Takumi Ohishi
- Yasuharu Otsubo
|Author status||OECD status||OECD project||SAAOP status|
|Under development: Not open for comment. Do not cite||Under Development||1.73||Included in OECD Work Plan|
This AOP was last modified on September 08, 2020 19:29
|Binding to estrogen receptor (ER)-α in immune cells||August 14, 2020 20:59|
|Induction of GATA3 expression||August 14, 2020 21:16|
|Increase of Th2 cells producing IL-4||August 14, 2020 21:26|
|Increase of anti-DNA antibody from autoreactive B cell||August 14, 2020 21:57|
|Exacerbation of systemic lupus erythematosus (SLE)||August 14, 2020 22:11|
|Binding to estrogen receptor (ER)-α leads to Induction of GATA3 expression||August 14, 2020 22:17|
|Induction of GATA3 expression leads to Increase of Th2 cells producing IL-4||August 14, 2020 22:22|
|Increase of Th2 cells producing IL-4 leads to Increase of autoantibody production||August 14, 2020 22:29|
|Increase of autoantibody production leads to Exacerbation of SLE||August 14, 2020 22:35|
|Bisphenol A||December 29, 2019 18:38|
|17beta-Estradiol||November 29, 2016 18:42|
This AOP describes the linkage between the binding to estrogen receptor (ER) α in immune cells with the exacerbation of the autoimmune disease systemic lupus erythematosus (SLE).
Estrogen receptors (ERs), ERα and ERβ, are a group of proteins that are activated by the steroid hormone estrogen and are widely expressed in most tissue types, including most immune cells. ERα can be activated with exogenous and endogenous estrogens. Also, there are numerous xenoestrogens that exist in the environment and imitate estrogen. Bisphenol A (BPA) is an example of a xenoestrogen that is considered an endocrine disrupting (ED) compound. SLE is an autoimmune disease characterized by overproduction of a variety of anti-cell nuclear and other pathogenic autoantibodies. It is characterized by B-cell hyperactivity, polyclonal hypergammaglobulinemia, and immune complex deposition.
Binding to ERα in immune cells by a xenoestrogen or endogenous estrogen marks the molecular initiating event (MIE), which results in induction of GATA3 expression (KE1). One theory of immune regulation involves homeostasis between T-helper 1 (Th1) and T-helper2 (Th2) activity, however GATA3 expression induce increase of Th2 cells producing cytokine interleukin-4 (IL-4) (KE2), which results in increase of anti-DNA antibody from autoreactive B cell (KE3). This sequence of pathway means that the immune system skew from a Th1 to a Th2 profile, which results in the adverse outcome (AO) of exacerbated SLE.
We have identified a number of key events along this pathway and determined the key event relationships, based on which we have created an AOP for binding to ERα in immune cells leading to exacerbated SLE.
It is well recognized that allergic diseases and autoimmune diseases are markedly increased the last several decades. About the same time, increasing scientific and social attention had been paid to environmentally dispersed chemicals that can enter the body by ingestion or adsorption and that mimic the actions of estrogens. These chemicals are termed endocrine disruptors (EDs) or environmental estrogens and are found in plastics (bisphenol-A, phthalates), pesticides (DDT, hexachlorobenzene, and dieldrin) and the like. Some of these estrogenic chemicals have also been shown to influence the immune system. Endocrine disruptors mimic hormones, block or alter hormone binding to receptors, or alter the metabolism of natural estrogens. It has been widely noted that females have stronger immune capabilities than males, as evidenced by their better immune responses to a variety of self-antigens and non-self-antigens, or vaccination. Paradoxically, the stronger immune response comes at a steep price, which is the high incidence of autoimmune diseases in females. This phenomenon of gender-based immune capability is largely attributed to the effects of sex hormones. Estrogens regulate the level of serum and uterine IgM, IgA, and IgG, and they augment antibody production to several nonself- antigens and self-antigens. It is possible that endocrine disruptors that mimic estrogenic activity may be involved in the increased incidence of autoimmune diseases such as SLE (Yurino H. 2004, Vaishali RM. 2018).
Summary of the AOP
Events: Molecular Initiating Events (MIE)
|Sequence||Type||Event ID||Title||Short name|
|MIE||1710||Binding to estrogen receptor (ER)-α in immune cells||Binding to estrogen receptor (ER)-α|
|KE||1711||Induction of GATA3 expression||Induction of GATA3 expression|
|KE||1712||Increase of Th2 cells producing IL-4||Increase of Th2 cells producing IL-4|
|KE||1713||Increase of anti-DNA antibody from autoreactive B cell||Increase of autoantibody production|
|AO||1714||Exacerbation of systemic lupus erythematosus (SLE)||Exacerbation of SLE|
Relationships Between Two Key Events
(Including MIEs and AOs)
|Binding to estrogen receptor (ER)-α leads to Induction of GATA3 expression||adjacent||Moderate||Moderate|
|Induction of GATA3 expression leads to Increase of Th2 cells producing IL-4||adjacent||Moderate||Moderate|
|Increase of Th2 cells producing IL-4 leads to Increase of autoantibody production||adjacent||Moderate||Moderate|
|Increase of autoantibody production leads to Exacerbation of SLE||adjacent||Moderate||Moderate|
Life Stage Applicability
|All life stages||Moderate|
|Homo sapiens||Homo sapiens||Moderate||NCBI|
Overall Assessment of the AOP
Domain of Applicability
It has long been appreciated that most autoimmune disorders are characterized by increased prevalence in females, suggesting a potential role for sex hormones (estrogen) in the etiology of autoimmunity. Females generally exhibit a stronger response to a variety of antigens including ERα ligands than males, which is perhaps one reason that they are more prone to develop autoimmune and allergic diseases such as SLE in greater severity than males. Therefore, this AOP is applicable to females and is dependent on the levels of estrogen, which means it varies with life stage, and age.
SLE frequently develop and progress in setting in which sympathoadrenomedullary and gonadal hormone levels are changing, e.g., during pregnancy, the postpartum period, or estrogen administration in menopause (Wilder RL. 1999). Women using oral contraceptives that contain estrogen or undergoing hormone replacement therapy are susceptible to major flare ups and exacerbation of the disease (Whitelaw DA. 2007).
The mechanisms described in this AOP are applicable to rodents and humans, and then the findings of this AOP are not found in any other species. However, Th2 dominant conditions induced by binding to ERα is considered likely to occur in a variety of mammalian species since ERα are expressed in all vertebrates (Eick GN. 2011).
Essentiality of the Key Events
Stressor , MIE and later events:
The NZB/W F1 mouse is the oldest classical model of lupus generated by the F1 hybrid between the NZB and NZW strains. The administration of the estrogen antagonist tamoxifen diminishes immune complex deposition in the kidneys and increases survival in NZB/W F1 strain. Renal disease was evaluated by the development of albuminuria and histological changes in the kidney (Wu WM. 2000). In females of the NZB/NZW F1 strain, disruption of ERα attenuated glomerulonephritis and increased survival and reduced anti-dsDNA antibodies (Bynote KK. 2008, Isenberg DA. 2007) and ovariectomy of NZB/W F1 mice not only delayed onset of the disease but also decreased autoantibody titer Meanwhile, restoration of estradiol in ovariectomized NZB/W F1 mice reestablished high numbers of autoantibody-producing (DNA-specific) B cells, and thereby suggests a pathogenic role of estrogen in lupus (Daniel P. 2011). Both NZB and NZW display limited autoimmunity, while NZB/W F1 hybrids develop severe lupus-like phenotypes comparable to that of lupus patients. In NZM female mice, ERα inactivation markedly prolonged life-span, lowered proteinuria, and ameliorated glomerulonephritis but resulted in higher serum anti-dsDNA antibody levels (Svenson JL. 2008).
KE1 and later events:
GATA3 mRNA expression has potential to induced IL-4 production in CD4+T cell (Lambert KC. 2005). The differentiation of activated CD4+T cells into the T helper type 1 (Th1) or Th2 fate is regulated by cytokines and the transcription factors T-bet and GATA-3. Early GATA-3 expression, required for Th2 differentiation, was induced by T cell factor 1 (TCF-1) and its cofactor β-catenin, mainly from the proximal Gata3 promoter upstream of exon 1b. TCF-1 blocked Th1 fate by negatively regulating interferon-γ (IFN-γ) expression independently of β-catenin. Thus, TCF-1 initiates Th2 differentiation of activated CD4+T cells by promoting GATA-3 expression and suppressing IFN-γ expression. Higher GATA-3 expression promotes IL-4 production and initiates Th2 differentiation (Qing Y. 2009). GATA-3 mRNA expression also increased in patients with SLE, compared with the healthy control groups (Zheng H. 2015, Sonia GR. 2012).
KE2 and later events:
Administration of mAb against IL-4 before the onset of lupus was effective in preventing the onset of lupus nephritis (Nakajima A. 1997).
KE3 and later events:
In a study to investigate a novel subpopulation of B-1 cells and its roles in murine lupus, anti-double-stranded DNA (anti-dsDNA) autoantibodies were preferentially secreted by a subpopulation of CD5+ B-1 cells that expressed programmed death ligand 2 (L2pB1 cells) (Xuemei Z. 2009). A substantial proportion of hybridoma clones generated from L2pB1 cells reacted to dsDNA. L2pB1 cells are potent antigen-presenting cells and a dramatic increase of circulating L2pB1 cells in lupus-prone BXSB mice correlates with elevated serum titers of anti-dsDNA antibodies (Xuemei Z. 2009).
|KER||KEup-KEdown||Plausibility||Rationales supported by literatures|
|KER 1||Binding, Estrogen receptor α in immune cells - Induction, GATA3 expression||Weak||In immune cells, this event is confirmed indirectly; using artificial STAT6-ER fusion protein.|
|KER 2||Induction, GATA3 expression - Increase, Th2 cells producing IL-4||Strong||XXXX|
|KER 3||Increase, Th2 cells producing IL-4 - Increase, anti-DNA antibody production from autoreactive B cell||Weak||XXXX|
|KER 4||Increase, anti-DNA antibody production from autoreactive B cell -||Strong||XXXX|
|KER||Empirical support of KERs|
|MIE=>KE 1：Binding, Estrogen receptor α in immune cells leads to Induction, GATA3 expression||
Empirical support of the MIE => KE1 is weak.
MIE: XXXKE XX: XXXX
|KE 1=> KE 2: Induction, GATA3 expression leads to Increase, Th2 cells producing IL-4||
Empirical support of the KE 1=> KE 2 is strong.
KE XX: XXXXAO: XXXX
|KE 2=> KE 3: Increase, Th2 cells producing IL-4 leads to Increase, anti-DNA antibody production from autoreactive B cell||
Empirical support of the KE 2=> KE 3 is weak.
KE XX: XXXXAO: XXXX
|KE 3=>AO: Increase, antibody production from anti-DNA antibody production from autoreactive B cell leads to Exacerbation, systemic lupus erythematosus (SLE)||
Empirical support of the KE 3 => AO is strong.
KE XX: XXXXAO: XXXX
CD4+T cell expressed GATA3 mRNA cultured with 10-9 M (272.4 pg/mL) concentrations of 17β-estradiol for 12-16 hr (Lambert KC. 2005).
BPA (0.1 mM) also indirectly induced GATA3 expression of Th cells, and this effect is mediated by dendritic cells exposed to BPA for 24 hr (Guo H. 2010). Naïve Th cells increased GATA3 expression cultured with dendritic cells exposure of BPA (0.1 mM) for 7 days.
Pre-stimulation 16 hr of 17β-estradiol (the concentration 10-9 M = 272.4 pg/mL) increased IL-4 secretion from CD4+T cell (Lambert KC. 2005).
PBMCs or B cells were cultured for 7 days with 17β-estradiol (10–8 mol/L) and then, IgG and IgM production were increased up to about 150% (PBMC) and 200% (B cells) (Kanda N. 1999).
Considerations for Potential Applications of the AOP (optional)
- Yurino, H., Ishikawa, S., Sato, T., Akadegawa, K., Ito, T., Ueha, S., Inadera, H. and Matsushima, K. (2004). Endocrine disruptors (environmental estrogens) enhance autoantibody production by B1 cells. Toxicological Sciences 81(1): 139-147.
- Vaishali RM. Sex Hormones in Acquired Immunity and Autoimmune Disease. Frontiers in Immunology 2018. 9: 2279; 1-21.
- Wilder RL, Elenkov IJ, Hormonal regulation of tumor necrosis factor-alpha, interleukin-12 and interleukin-10 production by activated macrophages. A disease-modifying mechanism in rheumatoid arthritis and systemic lupus erythematosus? Ann N Y Acad Sci. 1999. 22; 876:14-31.
- Whitelaw DA, Jessop SJ. Major flares in women with SLE on combined oral contraception. Clin Rheumatol. 2007; 26(12):2163-2165.
- Eick GN, Thornton JW. Evolution of steroid receptors from an estrogen-sensitive ancestral receptor. Molecular and cellular endocrinology. 2011; 334: 31-38.
- Wu WM, Lin BF, Su YC, et al. (2000). Tamoxifen decreases renal inflammation and alleviates disease severity in autoimmune NZB/W F1 mice. Scandinavian Journal of Immunology 52(4): 393-400.
- Bynote, KK, Hackenberg, JM., Korach, K.S., Lubahn, D. B., Lane, P. H. and Gould, K. A. (2008). Estrogen receptor-alpha deficiency attenuates autoimmune disease in (NZB xNZW) F1 mice. Genes and Immunity. 9: 137-152.
- Isenberg, DA., Manson, JJ., Ehrenstein, MR. and Rahman, A. (2007). Fifty years of anti-ds DNA antibodies: are we approaching journey’s end? Rheumatology 46:1052-6.
- Daniel, P., Allison, S., Yiming, Y., Ying-Yi, Z. and Laurence, M. Murine Models of Systemic Lupus erythematosus. Journal of Biomedicine and Biotechnology 2011: ArticleID 271694
- Svenson JL, EuDaly J, Ruiz P, Korach KS, Gilkeson GS. Impact of estrogen receptor deficiency on disease expression in the NZM2410 lupus prone mouse. Clin Immunol. 2008;128(2):259-68.
- Lambert KC, Curran EM, et al. Estrogen receptor alpha (ERalpha) deficiency in macrophages results in increased stimulation of CD4+ T cells while 17beta-estradiol acts through ERalpha to increase IL-4 and GATA-3 expression in CD4+ T cells independent of antigen presentation. J Immunol. 2005; 175(9): 5716-23.
- Qing Y., Archna S., Sun Y. O., Hyung-Geun M., M Zulfiquer H., Theresa M. S., Karen E. L., Hansen D., Beibei W., Marian L. W., Zhou Z. and Jyoti M. S., T cell factor 1 initiates the T helper type 2 fate by inducing the transcription factor GATA-3 and repressing interferon-γ. Nat Immunol. 2009; 10(9): 992–999.
- Zheng H, Guo X, Zhu Y, et al., Distinct role of Tim-3 in systemic lupus erythematosus and clear cell renal cell carcinoma. Int J Clin Exp Med 2015;8(5):7029-7038.
- Sonia GR, et al. Altered AKT1 and MAPK1 Gene Expression on Peripheral Blood Mononuclear Cells and Correlation with T-Helper-Transcription Factors in Systemic Lupus Erythematosus Patients. Mediators of Inflammation 2012, Article ID 495934
- Nakajima A, Hirose S, Yagita H and Okumura K, Roles of IL-4 and IL-12 in the development of lupus in NZB/W F1 mice. J Immunol 1997; 158 (3) 1466-1472.
- Xuemei, Z., Stanley, L., et al. (2009). A Novel Subpopulation of B-1 Cells Is Enriched with Autoreactivity in Normal and Lupus-Prone Mice. Arthritis & Rheumatology 60 (12):3734-3743.
- Guo H, Liu T, Ling F, et al. Bisphenol A in combination with TNF-alpha selectively induces Th2 cell-promoting dendritic cells in vitro with an estrogen-like activity. Cell Mol Immunol. 2010;7(3):227-34.
- Kanda N. and Tamaki, K. (1999). Estrogen enhances immunoglobulin production by human PBMCs. The Journal of Allergy and Clinical Immunology 103(2): 282-288.