ERα regulates lipid metabolism in bone through ATGL and perilipin

A decrease in bone mineral density during menopause is accompanied by an increase in adipocytes in the bone marrow space. Ovariectomy also leads to accumulation of fat in the bone marrow. Herein we show increased lipid accumulation in bone marrow from estrogen receptor alpha (ERα) knockout (ERαKO) mice compared to wild‐type (WT) mice or estrogen receptor beta (ERβ) knockout (ERβKO) mice. Similarly, bone marrow cells from ERαKO mice differentiated to adipocytes in culture also have increased lipid accumulation compared to cells from WT mice or ERβKO mice. Analysis of individual adipocytes shows that WT mice have fewer, but larger, lipid droplets per cell than adipocytes from ERαKO or ERβKO animals. Furthermore, higher levels of adipose triglyceride lipase (ATGL) protein in WT adipocytes correlate with increased lipolysis and fewer lipid droplets per cell and treatment with 17β‐estradiol (E2) potentiates this response. In contrast, cells from ERαKO mice display higher perilipin protein levels, promoting lipogenesis. Together these results demonstrate that E2 signals via ERα to regulate lipid droplet size and total lipid accumulation in the bone marrow space in vivo. J. Cell. Biochem. 114: 1306–1314, 2013. © 2012 Wiley Periodicals, Inc.     

Effects of estradiol and estradiol sulfamate on the uterus of ovariectomized or ovariectomized and hypophysectomized rats

Estradiol sulfamate (J995), estradiol-17beta with a substituted sulfamate group in position 3, has much higher systemic estrogenic activity after oral administration than 17beta-estradiol (E2) due to reduced hepatic metabolism of the drug. The lower dose necessary for achievement of adequate systemic estrogenic effects results in a substantial reduction of otherwise commonly observed hepatic side-effects. This makes J995 a strong candidate as an estrogen suitable for oral administration. The present study was performed to examine and compare the effects of J995 and E2 on the uterus after oral or subcutaneous administration to ovariectomized or ovariectomized+hypophysectomized female rats, in particular on the levels of the estrogen receptor (ER) (alpha+beta), ERalpha mRNA and insulin-like growth factor-I (IGF-I) mRNA. The ER levels were determined using a ligand binding assay and the mRNA levels using solution hybridization. The doses of J995 or E2 were chosen to achieve comparable uterotrophic activity. The rats were treated with hormones for 7 days and the treatment was initiated 14 days after surgery. We conclude that there are no major differences in the uterine response to treatment with J995 or E2 with respect to the effects on ER and ERalpha mRNA levels. The IGF-I mRNA level though, is more affected by J995 than by E2 after 7 days of treatment, indicating a prolonged effect of J995.     

Estrogen regulation of ion transporter messenger RNA levels in mouse efferent ductules are mediated differentially through estrogen receptor (ER) alpha and ER beta.

Earlier studies have shown that the efferent ductules (ED) of the male mouse are a target for estrogen. The loss of estrogen receptor (ER) function through either knockout technology (alpha ERKO mouse) or chemical interference (pure antagonist, ICI 182 780) results in a failure of a major function of the ED, the reabsorption of testicular fluids. The purpose of this study was to test the hypothesis that estrogen controls fluid (water) reabsorption in the ED by modulating ion transporters important for passive water movement through a leaky epithelium such as the ED. Northern blot analysis was used to detect the mRNA levels for key ion transporters in the following experimental groups: 1) wild-type (WT) control for the 14-day experiment, 2) ER alpha knockout (alpha ERKO) control for the 14-day experiment, 3) WT treated with ICI 182 780 (ICI) for 14 days, 4) alpha ERKO treated with ICI for 14 days, 5) WT control for the 35-day experiment, and 6) WT treated with ICI for 35 days. Estrogen differentially modulated the mRNA levels of key ion transporters. ER alpha mediated carbonic anhydrase II mRNA abundance, and there was a decrease in Na(+)/H(+) exchanger 3 mRNA levels in the alpha ERKO that appeared to be a cellular effect and not a direct estrogen effect. The loss of ER alpha control resulted in an increase in mRNA abundance for the catalytic subunit of Na(+)-K(+) ATPase alpha 1, whereas an increase in the mRNA abundance of the Cl(-)/HCO(3)(-) exchanger and the chloride channel cystic fibrosis transmembrane regulator was significantly ER beta mediated. Our results indicate for the first time that estrogen acting directly and indirectly through both ER alpha and ER beta probably modulates fluid reabsorption in the adult mouse ED by regulating the expression of ion transporters involved in the movement of Na(+) and Cl(-).     

Loss of ERE binding activity by estrogen receptor-alpha alters basal and estrogen-stimulated bone-related gene expression by osteoblastic cells

Estrogen receptor (ER)-alpha can signal either via estrogen response element (ERE)-mediated pathways or via alternate pathways involving protein-protein or membrane signaling. We previously demonstrated that, as compared to wild type (WT) controls, mice expressing a mutant ER-alpha lacking the ability to bind EREs (non-classical estrogen receptor knock-in (NERKI)) display significant impairments in the skeletal response to estrogen. To elucidate the mechanism(s) underlying these in vivo deficits, we generated U2OS cells stably expressing either WT ER-alpha or the NERKI receptor. Compared to cells transfected with the control vector, stable expression of ER-alpha, even in the absence of E2, resulted in an increase in mRNA levels for alkaline phosphatase (AP, by 400%, P < 0.01) and a decrease in mRNA levels for insulin growth factor-I (IGF-I) (by 65%, P < 0.001), with no effects on collagen I (col I) or osteocalcin (OCN) mRNA levels. By contrast, stable expression of the NERKI receptor resulted in the suppression of mRNA levels for AP, col I, OCN, and IGF-I (by 62, 89, 60, and 70%, P < 0.001). While E2 increased mRNA levels of AP, OCN, col I, and IGF-I in ER-alpha cells, E2 effects in the NERKI cells on AP and OCN mRNA levels were attenuated, with a trend for E2 to inhibit col I mRNA levels. In addition, E2 had no effects on IGF-I mRNA levels in NERKI cells. Collectively, these findings indicate that ERE signaling plays a significant role in mediating effects of estrogen on osteoblastic differentiation markers and on IGF-I mRNA levels.