We have previously reported epithelial cellular hyperplasia in ventral prostates (VP) of mice lacking estrogen receptor β (ERβ). To investigate the causes of this phenomenon, we measured cellular proliferation and apoptosis in VP of ERβ -/- and WT mice. With BrdUrd labeling, the number of proliferating cells was 3.6-fold higher in ERβ -/- mice. There was also a decrease in apoptosis as measured by terminal deoxynucleotidyltransferase-mediated dUTP nick end labeling assay and an increase in expression of the anti-apoptotic bcl-2. The state of differentiation of the epithelial cells of the VP was studied by immunohistochemical staining, Western blotting, and fluorescence-activated cell sorting (FACS). In ERβ -/- mouse VP, the number of p63-positive cells (basal phenotype) was 2.6-fold higher, and expression level of cytokeratin (CK) 8, a luminal cell marker, was lower. FACS analysis with p63 showed that in WT mice the ratio of basal to intermediate/luminal cell populations expressing p63 was 1:2.5, whereas in ERβ -/- mice it was 1:9. The expression of basal/intermediate marker CK 19 in three FACS areas, g1, g2, and g3, gated according to cellular size and granularity, was 1:0.6:2 in WT and 1:4:6.7 in ERβ -/- mice, showing a shift of CK 19-positive cells toward a cell population of intermediate size and granularity. We conclude that, in ERβ -/- mouse VP, there is increased epithelial proliferation, decreased apoptosis, and accumulation of incompletely differentiated cells in an intermediate pool. The continued proliferation of intermediate cells leads to the prostatic epithelial hyperplasia observed in the absence of ERβ signaling.

Proliferation of pluripotent, bone marrow stem cells, which develop to lymphoid and myeloid progenitors, is negatively regulated by estrogen. Although in estrogen deficiency and in estrogen receptor knockout mice there is significant alteration in bone marrow hematopoiesis, the effects of aging on estrogen receptor deficiencies in mice have not been investigated yet. In this study we show that by 1.5 years of age, estrogen receptor ␤ knockout (ER␤ ؊/؊ ) mice develop pronounced splenomegaly that is much more severe in females than in males. Further characterization of these mice revealed myelogenous hyperplasia in bone marrow, an increase in the number of granulocytes and B lymphocytes in blood, lymphadenopathy, and infiltration of leukocytes in the liver and lung. Analysis by flow cytometry of the bone marrow cells revealed that the percentage and total number of Gr-1 hi ͞Mac-1 hipositive granulocytes were increased by 15-30% and 100%, respectively. The numbers of B cells in the bone marrow and spleen were significantly higher in ER␤ ؊/؊ mice than in WT littermates. Some of the ER␤ ؊/؊ mice also had a severe lymphoproliferative phenotype. Thus the absence of ER␤ results in a myeloproliferative disease resembling human chronic myeloid leukemia with lymphoid blast crisis. Our results indicate a previously unknown role for ER␤ in regulating the differentiation of pluripotent hematopoietic progenitor cells and suggest that the ER␤ ؊/؊ mouse is a potential model for myeloid and lymphoid leukemia. Furthermore, we suggest that ER␤ agonists might have clinical value in the treatment of leukemia. O ver 100 years ago, it was noted that women were more affected by systemic lupus erythematosus than men. In fact, the incidence of many autoimmune diseases is higher in women (1). For this reason it has been speculated for a long time that estrogen plays important roles in the immune system. Loss of estrogen in ovariectomized mice results in splenomegaly (2) and increased production of colony-forming units-granulocyte͞ erythroid͞macrophage͞megakaryocytes, burst-forming unitserythroid cells (3-5), and B lymphocytes in mouse bone marrow (6). Conversely, pregnancy or administration of exogenous estrogen decreases bone marrow B lymphocyte population in mice (7,8). Recently, it was shown that estrogen represses the differentiation of multipotent hematopoietic stem cells into both lymphoid and myeloid cells (9, 10). Thus, estrogen is directly implicated in the proliferation and differentiation of various cell lineages in normal hematopoietic tissue.Estrogen exerts its effects through two distinct receptors, estrogen receptor (ER)␣ and ER␤. ER␣ has been detected in nonhematopoietic cells in bone marrow and in B lymphocyte precursors in mouse (8). ER␤ has also been found in nonhematopoietic cells in mouse bone marrow (11) and human spleen (12). Recently, the roles of ER in lymphopoiesis have been investigated in ER␣ These results seemed to indicate that negative effects of estrogen on the immune system might be mediated by ER␤.We have ...

Sjögren's syndrome (SS) is an incurable, autoimmune exocrinopathy that predominantly affects females and whose pathogenesis remains unknown. Like rheumatoid arthritis, its severity increases after menopause, and estrogen deficiency has been implicated. We have reported that estrogen receptor-α and -β-knockout mice develop autoimmune nephritis and myeloid leukemia, respectively, but neither develops SS. One model of estrogen deficiency in rodents is the aromatase-knockout (ArKO) mouse. In these animals, there is elevated B lymphopoiesis in bone marrow. We now report that ArKO mice develop severe autoimmune exocrinopathy resembling SS. By 1 year of age, there is B cell hyperplasia in the bone marrow, spleen, and blood of ArKO mice and spontaneous autoimmune manifestations such as proteinuria and severe leukocyte infiltration in the salivary glands and kidney. Also, as is typically found in human SS, there were proteolytic fragments of α-fodrin in the salivary glands and anti-α-fodrin antibodies in the serum of both female and male ArKO mice. When mice were raised on a phytoestrogen-free diet, there was a mild but significant incidence of infiltration of B lymphocytes in WT mice and severe destructive autoimmune lesions in ArKO mice. In age-matched WT mice fed a diet containing normal levels of phytoestrogen, there were no autoimmune lesions. These results reveal that estrogen deficiency results in a lymphoproliferative autoimmune disease resembling SS and suggest that estrogen might have clinical value in the prevention or treatment of this disease.

Estrogens, acting through its two receptors, ESR1 (hereafter designated ER alpha) and ESR2 (hereafter designated ER beta), have diverse physiological effects in the reproductive system, bone, cardiovascular system, hematopoiesis, and central and peripheral nervous systems. Mice with inactivated ER alpha, ER beta, or both show a number of interesting phenotypes, including incompletely differentiated epithelium in tissues under steroidal control (prostate, ovary, mammary, and salivary glands) and defective ovulation reminiscent of polycystic ovarian syndrome in humans (in ER beta-/- mice), and obesity, insulin resistance, and complete infertility (both in male and female ER alpha-/- mice). Estrogen agonists and antagonists are frequently prescribed drugs with indications that include postmenopausal syndrome (agonists) and breast cancer (antagonists). Because the two estrogen receptors (ERs) have different physiological functions and have ligand binding pockets that differ enough to be selective in their ligand binding, opportunities now exist for development of novel ER subtype-specific selective-ER modulators.

In mice, ovariectomy accelerates the progression of the end-stage renal disease glomerulosclerosis. In women, the incidence of this disease increases after menopause, and estrogen alters its progression. Polymorphisms in the human estrogen receptor α (ERα) gene have been suggested to constitute a genetic predisposition for lupus nephritis. Here we show that by 1 year of age, mice lacking ERα (ERα -/- ) but not those lacking ERβ (ERβ -/- ) exhibit immune complex-type glomerulonephritis, proteinuria, and destruction of tubular cells with severe infiltration of B lymphocytes in the kidney and the presence of anti-DNA antibodies in serum. No gender difference occurred in the incidence or severity of these symptoms. However, in female but not in male ERα -/- mice there were elevated serum levels of IgG3. Other prominent features of these mice were ( i ) spontaneous formation of germinal centers in the spleen in the absence of antigen challenge and ( ii ) infiltration of plasma cells in the kidney and plasmacytosis in the spleen. Immunohistochemistry indicated a selective expression of ERα protein in the germinal centers but not in the follicular mantle zone of murine spleens and human tonsils. Our results indicate that ERα has indispensable functions in the kidney and in germinal centers, and that defective ERα signaling results in glomerulonephritis.