ReviewGender differences in autoimmunity: molecular basis for estrogen effects in systemic lupus erythematosus☆
Introduction
Systemic lupus erythematosus (SLE) is an autoimmune disease that occurs primarily in women (9:1 compared to men) during their childbearing years. In the United States, the prevalence of SLE ranges from 14.6 to 50.8 cases per 100,000 people [1]. The effects of SLE are physically and emotionally debilitating and can be life threatening due to the involvement of a variety of organs including the renal and central nervous systems. SLE is characterized as a chronic inflammatory autoimmune disease that is accompanied by abnormal immunoregulation [2]. The disease is activated by genetic and environmental factors [3], [4], [5]. However, the strongest risk factor for the development of SLE is female gender [1]. Sex hormone influence on the gender bias in SLE is evident after puberty since disease activity fluctuates in some patients with the menstrual cycle [6], [7], [8], [9], [10] and pregnancy can flare the disease [11]. In lupus mouse models, NZB×NZW F1 female mice develop lupus and die at an earlier age than their male counterparts. Administration of exogenous estrogen to these mice enhanced lupus disease pathology and increased anti-DNA antibody levels [12].
Estrogens generally have been implicated as enhancers of the immune response, while androgens and progesterone are considered natural immune suppressors. Estrogen serves as a ligand for two specific receptor proteins termed estrogen receptor-α (ERα) and estrogen receptor-β (ERβ). Estrogen receptors (ERs) are ligand-activated transcription factors that bind to specific DNA sequences of target genes and alter the rates of transcription. These receptors belong to a super gene family of nuclear hormone receptors that share a centrally located domain responsible for binding to DNA [13] and separate protein domains that are necessary for transcriptional activation. While a considerable degree of homology between ERα and ERβ exists [14], the amino terminus of the two proteins is poorly conserved (see Ref. [14] for details). Moreover, cell- and promoter-specific differences in transcriptional activity between the receptor subtypes have been reported owing to differences in the activation function modules of the proteins [15], [16], [17]. Differences in transcriptional regulation between ERα and ERβ may involve interactions with unique coregulators at the receptor sites that are necessary for transcriptional modulation, thereby expanding the potential repertoire for estrogen-dependent gene transcription, particularly, in cells that express both receptor subtypes [14], [15], [16], [17], [18].
While a considerable body of anecdotal evidence implicated estrogens as one factor contributing to the pathology in SLE [19], [20], [21], [22], there has been no specific marker for estrogen action that consistently correlates with immunodysregulation in lupus. Since SLE is suggested to be a disease with aberrant T cell regulatory functions [23], [24], [25], our initial studies investigated the effect of estrogen on human T cells. We compared mRNA populations between T cells cultured without and with estrogen using the technique of differential mRNA display [26]. One cDNA uniquely expressed in T cells cultured with estradiol-17β showed on both DNA strands sequence identity with the published sequence of calcineurin A2 extending from nucleotides 2896–3079 [27]. Identification of calcineurin as a putative target for estrogen action in T cells provided us the necessary link with aberrant immune regulation in SLE because activated T cells provide help for the production of autoantibodies by B cells [23].
In human T cells, the binding of antigen to the T cell receptor (TCR) complex stimulates a signal transduction cascade that transmits signals from the cell surface to the nucleus [28], [29]. This signal transduction pathway is calcium-dependent and includes protein kinase C (PKC) activation, changes in the phosphorylation status of regulatory proteins, and the synthesis and secretion of interleukin-2 (IL-2) [30]. Calcineurin is a calcium and calmodulin-dependent protein phosphatase of the PP2B class [31], [32], [33], [34]. The protein is a heterodimeric serine–threonine phosphatase comprised of a catalytic subunit that has a calmodulin binding domain, and a regulatory subunit containing four calcium binding sites [33]. Dephosphorylation by calcineurin stimulates the translocation of nuclear factor of activated T cells (NFAT) from the cytoplasm to the nucleus where it binds promoter sequences of target genes [35]. Overexpression of calcineurin in Jurkat cells augments NFAT-dependent transcription of IL-2 and promotes resistance to the immunosuppressive drugs tacrolimus (FK506) and cyclosporin A (CsA) [31], [32], [34]. The mechanism of transcriptional inhibition is exerted, in part, by attenuating the phosphatase activity of calcineurin [36]. Lupus T cells release more intracellular calcium in response to TCR activation than normal healthy T cells, although the amount of inositol 3-phosphate is similar between lupus and normal cells [37]. Phosphatase activity increases in the lymphoid tissues and, particularly, in T lymphocytes of MRL/lpr lupus-prone mice [38]. Treatment of these mice with the calcineurin inhibitor FK506 and cyclophosphamide prolongs their survival [39]. We, therefore, had a basis for testing the hypothesis that estrogen, acting through the ER, increases calcineurin and NFAT activity in the T cells from female lupus patients. The downstream consequences of estrogen action could contribute to lupus pathogenesis as indicated in our hypothetical model shown in Fig. 1. This model proposes a molecular basis for estrogen effects in SLE T cells that is postulated to result in aberrant T–B cell interactions. We suggest that the gender bias in SLE may arise, in part, from an inherent estrogen sensitivity that is characteristic of female lupus T cells. This review focuses on the data that provide evidence for estrogen effects specifically in the T cells from with women with lupus.
Section snippets
Study participants
Participants in our studies include 24 female patients with lupus and 13 age-matched normal control women. The women enrolled in this study had regular menstrual cycles and were between the ages of 16 and 48. Male participants included five patients with lupus and six age-matched control males. The males enrolled in this study were between 19 and 55 years of age. Lupus patients met at least four of the criteria of the American College of Rheumatology for classification of SLE [40]. The disease
Dose-dependent response of SLE T cells to estradiol
To determine if physiological concentrations of estradiol increased calcineurin expression, T cells from lupus patients and normal control women were cultured in serum-free medium without and with estradiol (10−9 to 10−7 M) for 16 h (Fig. 2). These concentrations represent the physiological levels of estradiol in circulation during the follicular phase of the menstrual cycle and early and late pregnancy, respectively [26], [50]. In the female lupus patients, estradiol increased calcineurin mRNA
Discussion
Studies in our laboratory [26], [41] show that T cells from female lupus patients exhibit a sensitivity to estradiol that is not shared by T cells from normal women and men nor from males with SLE. The effects of estradiol on T cells from women with lupus are dose-dependent and hormone-specific. Estrogen control of calcineurin is temporally regulated with increased steady-state mRNA at 6 h followed by increased phosphatase activity by 8 h after exposure of SLE T cells to estradiol. This
Acknowledgements
The authors thank the patients and normal volunteers who donated blood for this study. We recognize the excellent research efforts of our colleagues (M. Evans, S. Jones and Dr. R. Suenaga) whose research contributed to this review. We thank J. Swafford (UMKC) for help with the figures. The authors are grateful to Dr. G.C. Tsokos (Walter Reed Army Institute of Research, Washington, DC) for intellectual stimulation.
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This research was supported in part by grants from the Sarah Morrison Fund, the Evans Endowment, and St. Luke's Foundation.