The estrogen-injected female mouse: new insight into the etiology of PCOS

<p>Abstract</p> <p>Background</p> <p>Female mice and rats injected with estrogen perinatally become anovulatory and develop follicular cysts. The current consensus is that this adverse response to estrogen involves the hypothalamus and occurs because of an estrogen-induced alteration in the GnRH delivery system. Whether or not this is true has yet to be firmly established. The present study examined an alternate possibility in which anovulation and cyst development occurs through an estrogen-induced disruption in the immune system, achieved through the intermediation of the thymus gland.</p> <p>Methods, Results and Conclusion</p> <p>A putative role for the thymus in estrogen-induced anovulation and follicular cyst formation (a model of PCOS) was examined in female mice by removing the gland prior to estrogen injection. Whereas all intact, female mice injected with 20 ug estrogen at 5–7 days of age had ovaries with follicular cysts, no cysts were observed in animals in which thymectomy at 3 days of age preceded estrogen injection. In fact, after restoring immune function by thymocyte replacement, the majority of thymectomized, estrogen-injected mice had ovaries with corpora lutea. Thus, when estrogen is unable to act on the thymus, ovulation occurs and follicular cysts do not develop. This implicates the thymus in the cysts' genesis and discounts the role of the hypothalamus. Subsequent research established that the disease is transferable by lymphocyte infusion. Transfer took place between 100-day-old estrogen-injected and 15-day-old naïve mice only when recipients were thymectomized at 3 days of age. Thus, a prerequisite for cyst formation is the absence of regulatory T cells. Their absence in donor mice was judged to be the result of an estrogen-induced increase in the thymus' vascular permeability, causing de facto circumvention of the final stages of regulatory T cell development. The human thymus has a similar vulnerability to steroid action during the fetal stage. We propose that in utero exposure to excessive levels of steroids such as estrogen has a long-term effect on the ability of the thymus to produce regulatory T cells. In female offspring this can lead to PCOS.</p

Murine pregnancy leads to reduced proliferation of maternal thymocytes and decreased thymic emigration

During mammalian pregnancy the maternal thymus undergoes significant involution, and then recovers in size after birth. The mechanism behind this involution is not known, but it has been suggested that elevated levels of hormones during pregnancy induce the involution. We have recently shown that injection of 17β-oestradiol into mice causes loss of early thymocyte precursors and inhibits proliferation of developing thymocytes. This suggests that elevated oestrogen in pregnancy may contribute to thymic involution. We have investigated this idea by examining the fate of thymocytes during mouse pregnancy in much greater detail than has been previously reported. Looking over a broad time–course, we find that pregnancy does not affect thymocyte precursor populations in the bone marrow, but induces a profound loss of early thymic progenitors in the thymus as early as day 12·5 of pregnancy. This loss is accompanied by decreased thymocyte proliferation, which returns to normal 2–4 days postpartum. No enhancement of apoptosis is detectable at any stage of pregnancy. We also find that there is a reduction in recent thymic emigrants after oestrogen treatment and at day 17·5 of pregnancy, suggesting that thymic involution during pregnancy influences the peripheral T-cell repertoire. The similarities between oestrogen-mediated involution and pregnancy-mediated involution suggest that oestrogen is a significant contributor to loss of thymocyte cellularity during pregnancy, and probably functions primarily by reducing thymocyte proliferation

Immunomodulation by estrogen and estren

Estrogen affects the development and regulation of the immune system. Treatment of gonadectomized mice with estrogen results in suppression of T and B lymphopoiesis, as well as decreased delayed type hypersensitivity reaction, granulocyte mediated inflammation and levels of IL-6 in serum. Conversely, immunoglobulin production is stimulated by estrogen. The effects of estrogen are mediated through the estrogen receptors (ER), ERa and ERb, which are ligand activated transcription factors that induce expression of specific estrogen responsive genes. The aims of this thesis were to investigate the role of ERs on B lymphopoiesis and immunoglobulin production, as well as on the aged immune system. Furthermore, the ER specific effects of the synthetic molecule estren on T and B lymphopoiesis, T cell-mediated inflammation and submandibular glands were studied. ER knock-out mice lacking ERa, ERb or both ERa and ERb, were gonadectomized and treated with 17b-estradiol-3-benzoate (E2) or 4-estren-3a,17b-diol (estren). We found that both ERa and ERb are required for the estrogen-induced decreased frequency of B lymphopoietic cells in the bone marrow. ERa alone is necessary for the estrogen-mediated, as well as for the age-induced, increased frequency of immunoglobulin producing B cells. We could also show that estren inhibits inflammation through ER-mediated pathways, while the inhibitory effects on T and B lymphopoiesis are not dependent on ERs. Furthermore, estren promotes an androgen phenotype in submandibular glands that is independent of ERs. In conclusion, our results show that the effects of estrogen on the immune system are mainly mediated via ERa, but signalling through ERb is necessary for complete inhibitory effect on B lymphopoiesis. Furthermore, estren treatment induces effects on lymphopoiesis and submandibular glands that are not mediated through ERs, but instead possibly through the androgen receptor

17 beta-estradiol expands IgA-Producing b cells in mice deficient for the mu chain

Oestrogen is not only a sex hormone but also an important regulator of the immune system. Expression of the heavy chain of IgM (mu) is essential for B-cell differentiation. However, a small number of IgA-positive B cells can be found in mice lacking the mu chain (mu MT-/-). The aim of this study was to investigate the effects of oestrogen on this alternative B-cell pathway in mu MT-/- mice. Our results clearly demonstrate that oestrogen increases the frequency of IgA-producing B cells in mu MT-/- mice in both bone marrow and spleen cells. We also show that mature IgM-producing B cells are not required for oestrogen-mediated suppression of granulocyte-mediated inflammation or thymic involution. In conclusion, we demonstrate that 17 beta-estradiol benzoate increases the frequency of IgA-producing B cells in mu MT-/- mice, suggesting that oestrogen can influence the alternative B-cell pathway found in mu MT-/- mice

Influence of oestrogen receptor α and β on the immune system in aged female mice

Oestrogen has a dichotomous effect on the immune system. T and B lymphopoiesis in thymus and bone marrow is suppressed, whereas antibody production is stimulated by oestrogen. In this study the importance of the oestrogen receptors (ER) ER-α and ER-β in the aged immune system was investigated in 18 months old-wild type (WT), ER-α (ERKO), ER-β (BERKO) and double ER-α and ER-β (DERKO) knock-out mice, and compared with 4 months old WT mice. Cell phenotypes in bone marrow, spleen and thymus, and the frequency of immunoglobulin (Ig) spot forming cells (SFC) were determined. We show here that the 17-β-oestradiol (E2)-induced downregulation of B lymphopoietic cells in bone marrow of young ovariectomized mice can be mediated through both ER-α and ER-β. However, only ER-α is required for the age-related increased frequency of immunoglobulin M (IgM) SFC in the bone marrow, as well as for the increased production of interleukin-10 (IL-10) from cultured splenocytes in aged mice. Furthermore, increased age in WT mice resulted in lower levels of both pro- and pre-B cells but increased frequency of IgM SFC in the bone marrow, as well as increased frequency of both IgM and IgA SFC in the spleen. Results from this study provide valuable information regarding the specific functions of ER-α and ER-β in the aged immune system

T lymphocytes are not the target for estradiol-mediated suppression of DTH in reconstituted female severe combined immunodeficient (SCID) mice

Oestrogen has the capacity to suppress T cell-dependent DTH. To explore the mechanisms whereby oestrogen exerts its effects on the immune system we have used SCID mice which are largely devoid of functional T and B lymphocytes, hence being unable to raise DTH, but display intact antigen-presenting capacity. Transfer of lymphocytes to SCID mice restores the DTH capacity. In order to analyse if oestrogen down-regulates DTH by a direct action on T cells we reconstituted SCID mice with either splenocytes or thymocytes from congenic C.B-17 or allogeneic B6 donor mice. Either donor or recipient mice were exposed to estradiol before cell transfer. DTH response was registered in recipient SCID mice 1 and 3 weeks after challenge with oxazolone (OXA). SCID mice receiving estradiol-exposed spleen cells from congenic or allogeneic donor mice displayed lower DTH responses compared with control mice. In contrast, SCID mice receiving estradiol-exposed thymocytes from congenic donor mice showed no significant difference in DTH response compared with control mice. Estradiol-treated SCID mice, transferred with either spleen cells or thymocytes from congenic, hormonally non-treated donors, displayed a significantly lower DTH response compared with control mice. In contrast, estradiol-treated SCID mice receiving hormonally non-treated allogeneic spleen cells showed no difference in DTH response compared with control mice. The results show that T lymphocytes are not the target cell population for estradiol-mediated suppression of DTH in reconstituted female SCID mice