Evidence of estrogen receptors in normal human osteoblast-like cells

In seven strains of cultured normal human osteoblast-like cells, a mean of 1615 molecules of tritium-labeled 17 beta-estradiol per cell nucleus could be bound to specific nuclear sites. The nuclear binding of the labeled steroid was temperature-dependent, steroid-specific, saturable, and cell type-specific. These are characteristics of biologically active estrogen receptors. Pretreatment with 10 nanomolar estradiol in vitro increased the specific nuclear binding of progesterone in four of six cell strains, indicating an induction of functional progesterone receptors. RNA blot analysis demonstrated the presence of messenger RNA for the human estrogen receptor. The data suggest that estrogen acts directly on human bone cells through a classical estrogen receptor-mediated mechanism.     

Sequence and expression of human estrogen receptor complementary DNA

The mechanism by which the estrogen receptor and other steroid hormone receptors regulate gene expression in eukaryotic cells is not well understood. In this study, a complementary DNA clone containing the entire translated portion of the messenger RNA for the estrogen receptor from MCF-7 human breast cancer cells was sequenced and then expressed in Chinese hamster ovary (CHO-K1) cells to give a functional protein. An open reading frame of 1785 nucleotides in the complementary DNA corresponded to a polypeptide of 595 amino acids and a molecular weight of 66,200, which is in good agreement with published molecular weight values of 65,000 to 70,000 for the estrogen receptor. Homogenates of transformed Chinese hamster ovary cells containing a protein that bound [3H]estradiol and sedimented as a 4S complex in salt-containing sucrose gradients and as an 8 to 9S complex in the absence of salt. Interaction of this receptor-[3H]estradiol complex with a monoclonal antibody that is specific for primate ER confirms the identity of the expressed complementary DNA as human estrogen receptor. Amino acid sequence comparisons revealed significant regional homology among the human estrogen receptor, the human glucocorticoid receptor, and the putative v-erbA oncogene product. This suggests that steroid receptor genes and the avian erythroblastosis viral oncogene are derived from a common primordial gene. The homologous region, which is rich in cysteine, lysine, and arginine, may represent the DNA-binding domain of these proteins.     

Estrogen receptor analysis by flow cytometry

A fluorescently labeled estradiol, N'-fluoresceino-N'-(17 beta-estradiol hemisuccinamide) thiourea (FE) was used for measuring estrogen receptor content per cell in tumor cells. The cellular content of FE was measured quantitatively by flow cytometry. Binding of FE occurs in the nanomolar concentration range, an indication of the high affinity of the labeled estradiol. Competition of FE for binding sites is observed with estrogens, but not with progestins, androgens, or glucocorticosteroids, indicating the specificity of FE binding. In contrast to other estrogen receptor assays, this new technique requires a small sample size (about 5000 cells) and permits the assessment of heterogeneity in estrogen receptor expression among tumor cells.     

雌二醇及其受体在北方山溪鲵精巢中的周期性分布

用免疫组织化学方法检测雌二醇(estradiol)及其受体(estrogen receptor)在北方山溪鲵(Batrachuperus tibetanus)精巢发育周期中的表达变化。结果表明,在4~5月,雌二醇及受体在精原细胞的胞质中强表达;6月,雌二醇在精母细胞的胞质中表达较弱,其受体在精母细胞的核质中有较强表达;7月,雌二醇及受体在精子细胞的胞质中表达;8月,雌二醇及受体在精原细胞的胞质中和变态后的精子头部有弱表达;9-11月,两者均在精原细胞的胞质中及精子头部有较强表达。说明雌二醇及其受体存在于整个精巢周期中,二者在生精细胞中的强弱变化基本一致,其变化规律与精子发育周期密切相关;雌激素受体多存在于生精细胞的胞质中,雌激素可能以非基因组效应调控细胞的增殖,并与精子的发育成熟密切相关。     

Estrogen-receptor interaction

The interaction of estradiol with uterine cells involves the association of the hormone with an extranuclear receptor protein, followed by temperature dependent translocation of the resulting complex to the nucleus. During this process, the steroid binding unit of the protein undergoes an alteration, called "receptor transformation," that can be recognized by an increase in its sedimentation rate from 3.8S to 5.2S, and by its acquisition of the ability to bind to isolated uterine nuclei and to alleviate a tissue specific deficiency in the RNA synthesizing capacity of such nuclei. Receptor transformation can be effected in the absence of nuclei by warming uterine cytosol with estradiol. This preparation of transformed complex resembles that extracted from nuclei both in its sedimentation rate (5.3S) and in its ability to bind to uterine nuclei and augment RNA synthesis, properties that are not shown by the native complex. It is proposed that receptor transformation is an important step in estrogen action and that a principal role of the hormone is to induce conversion of the receptor protein to a biochemically functional form.     

Saccharomyces cerevisiae produces a yeast substance that exhibits estrogenic activity in mammalian systems

Partially purified lipid extracts of Saccharomyces cerevisiae contain a substance that displaces tritiated estradiol from rat uterine cytosol estrogen receptors. The yeast product induces estrogenic bioresponses in mammalian systems as measured by induction of progesterone receptors in cultured MCF-7 human breast cancer cells and by a uterotrophic response and progesterone receptor induction after administration to ovariectomized mice. The findings raise the possibility that bakers' yeast may be a source of environmental estrogens.     

Transfection of v-rasH DNA into MCF-7 human breast cancer cells bypasses dependence on estrogen for tumorigenicity

The natural history of estrogen-responsive breast cancers often involves a phenotypic change to an estrogen-unresponsive, more aggressive tumor. The human breast cancer cell line, MCF-7, which requires estradiol for tumor formation in vivo and shows growth stimulation in response to estradiol in vitro, is a model for hormone-responsive tumors. The v-rasH onc gene was transfected into MCF-7 cells. The cloned MCF-7ras transfectants, which expressed the v-rasH messenger RNA and v-rasH p21 protein (21,000 daltons), were characterized. In contrast to the parental cell line, MCF-7ras cells no longer responded to exogenous estrogen in culture and their growth was minimally inhibited by exogenous antiestrogens. When tested in the nude mouse, the MCF-7ras cells were fully tumorigenic in the absence of estrogen supplementation. Thus, cells acquiring an activated onc gene can bypass the hormonal regulatory signals that trigger the neoplastic growth of a human breast cancer cell line.     

雌激素对老年大鼠小脑ER、ChAT、NGF表达的影响

 【目的】 探讨雌激素对大鼠小脑内雌激素受体（ER）、神经生长因子（NGF）和胆碱乙酰转移酶（ChAT）表达的影响。【方法】运用超敏感的免疫组织化学SP法，以老年SD大鼠小脑为研究对象，通过补充17β－雌二醇对ER、NGF和ChAT在小脑中的表达和分布变化进行研究。【结果】ER、NGF和ChAT免疫阳性反应物分布于小脑的蒲肯野氏细胞层、小脑齿状核、小脑间位核和小脑室顶核，ER阳性产物主要定位于细胞胞浆和突起中，也存在于胞膜和胞核中。老年大鼠小脑皮质及小脑核中ER、NGF和ChAT的表达强度及阳性细胞数量总趋势是显著降低，而补充17β－雌二醇后三种阳性产物的强度和阳性细胞数目显著回升，蒲肯野氏细胞的阳性突起长度和数量也呈此变化趋势。【结论】雌激素可促进NGF和ChAT的表达，在维持和保护小脑神经元的结构和功能中发挥了重要作用；另外ER、NGF和ChAT表达变化的相似性提示三者在雌激素对小脑的作用中是相互调节和影响的，同时表明雌激素在小脑发挥作用可能既通过基因组机制，也通过非基因组机制途径。     

Gene transfer and molecular cloning of the human NGF receptor

Nerve growth factor (NGF) and its receptor are important in the development of cells derived from the neural crest. Mouse L cell transformants have been generated that stably express the human NGF receptor gene transfer with total human DNA. Affinity cross-linking, metabolic labeling and immunoprecipitation, and equilibrium binding with 125I-labeled NGF revealed that this NGF receptor had the same size and binding characteristics as the receptor from human melanoma cells and rat PC12 cells. The sequences encoding the NGF receptor were molecularly cloned using the human Alu repetitive sequence as a probe. A cosmid clone that contained the human NGF receptor gene allowed efficient transfection and expression of the receptor.     

雌激素和雄激素对鸡成骨细胞增殖、凋亡、细胞周期及其受体mRNA转录的影响

为探讨雌激素和雄激素对鸡胚额骨成骨细胞(OB)增殖、凋亡、细胞周期及其受体mRNA转录的影响,用雌激素(17β雌二醇)和雄激素(十一酸睾酮)单独以及联合处理鸡胚额骨成骨细胞,MTT法测定细胞增殖率,流式细胞仪检测细胞凋亡和细胞周期的变化,荧光定量PCR检测成骨细胞中雌激素受体(ER)及雄激素受体(AR)mRNA的转录。结果显示:一定浓度的雌激素或雄激素在24h内均能促进成骨细胞的增殖,促进成骨细胞的细胞周期进程,但对细胞凋亡无明显作用。雌激素单独或联合雄激素作用能上调ERmRNA的转录,对ARmRNA的转录无明显作用。     

Identification of a novel thyroid hormone receptor expressed in the mammalian central nervous system

A complementary DNA clone derived from rat brain messenger RNA has been isolated on the basis of homology to the human thyroid hormone receptor gene. Expression of this complementary DNA produces a high-affinity binding protein for thyroid hormones. Sequence analysis and the mapping of this gene to a distinct human genetic locus indicate the existence of multiple human thyroid hormone receptors. Messenger RNA from this gene is expressed in a tissue-specific fashion with highest levels in the central nervous system.     

Sulfhydryl groups and estradiol-receptor interaction

The characteristic ability of rat uteri to take up tritiated estradiol in vitro or to retain estradiol previously incorporated either in vivo or in vitro is destroyed by treating the tissue with various sulfhydryl-blocking reagents. The two radioactive estradiol-receptor complexes, observed in uterine homogenates in the supernatant fraction and in an extract of the nuclear fraction, respectively, are disrupted by brief exposure to organic mercurials in the cold. Sulfhydryl groups of uterine receptor substances apparently play a vital role in estradiol binding, perhaps indirectly through contribution to receptor conformation.     

益母草总黄酮对去卵巢大鼠下丘脑和海马雌激素受体α及β mRNA表达的影响

目的：观察益母草总黄酮对去卵巢大鼠下丘脑和海马雌激素受体α及β mRNA表达的影响,探讨其对绝经后骨质疏松的作用机制。方法选取10~11月龄SD雌性大鼠48只,随机分为对照组、去势组、益母草总黄酮组和雌二醇组,以双侧卵巢切除法建立大鼠骨质疏松模型。益母草总黄酮组、雌二醇组分别用益母草总黄酮、雌二醇给药4个月,采用RT-PCR法检测各组下丘脑和海马雌激素受体α及β mRNA的表达。结果大鼠卵巢切除后,血清中雌二醇水平、椎骨骨密度、子宫湿重、下丘脑和海马雌激素受体α及βmRNA的表达均明显下降。雌二醇治疗后血清中雌二醇水平、椎骨骨密度、子宫湿重升高,下丘脑和海马β mRNA的表达均明显下降。益母草总黄酮治疗后,血清中雌二醇水平、椎骨骨密度、下丘脑和海马雌激素受体α及β mRNA的表达均明显升高,子宫湿重无明显改变。结论益母草总黄酮对切除卵巢所致的骨质疏松大鼠有防治作用,对子宫无不良影响。     

Expression of high-affinity binding of human immunoglobulin E by transfected cells

The receptor with high affinity for immunoglobulin E (IgE) on mast cells and basophils is critical in initiating allergic reactions. It is composed of an IgE-binding alpha subunit, a beta subunit, and two gamma subunits. The human alpha subunit was expressed on transfected cells in the presence of rat beta and gamma subunits or in the presence of the gamma subunit alone. The IgE binding properties of the expressed human alpha were characteristic of receptors on normal human cells. These results now permit a systematic analysis of human IgE binding and a search for therapeutically useful inhibitors of that binding.     

Cloning of an interleukin-3 receptor gene: a member of a distinct receptor gene family

Interleukin-3 (IL-3) binds to its receptor with high and low affinities, induces tyrosine phosphorylation, and promotes the proliferation and differentiation of hematopoietic cells. A binding component of the IL-3 receptor was cloned. Fibroblasts transfected with the complementary DNA bound IL-3 with a low affinity [dissociation constant (Kd) of 17.9 +/- 3.6 nM]. No consensus sequence for a tyrosine kinase was present in the cytoplasmic domain. Thus, additional components are required for a functional high affinity IL-3 receptor. A sequence comparison of the IL-3 receptor with other cytokine receptors (erythropoietin, IL-4, IL-6, and the beta chain IL-2 receptor) revealed a common motif of a distinct receptor gene family.     

Prolactin receptors in estrogen receptor-deficient mammary carcinoma

We demonstrate for the first time the presence of specific high-affinity receptors for prolactin in rat mammayy carcinoma. There appear to be no significant differences between normal rat mammary tissue and the transplantable, estrogen receptor-deficient R3230AC tumor with regard to the number of binding sites, the affinity of the receptor for prolactin, or the specificity of binding.     

Chimeric NGF-EGF receptors define domains responsible for neuronal differentiation

To determine the domains of the low-affinity nerve growth factor (NGF) receptor required for appropriate signal transduction, a series of hybrid receptors were constructed that consisted of the extracellular ligand-binding domain of the human epidermal growth factor (EGF) receptor (EGFR) fused to the transmembrane and cytoplasmic domains of the human low-affinity NGF receptor (NGFR). Transfection of these chimeric receptors into rat pheochromocytoma PC12 cells resulted in appropriate cell surface expression. Biological activity mediated by the EGF-NGF chimeric receptor was assayed by the induction of neurite outgrowth in response to EGF in stably transfected cells. Furthermore, the chimeric receptor mediated nuclear signaling, as evidenced by the specific induction of transin messenger RNA, an NGF-responsive gene. Neurite outgrowth was not observed with chimeric receptors that contained the transmembrane domain from the EGFR, suggesting that the membrane-spanning region and cytoplasmic domain of the low-affinity NGFR are necessary for signal transduction.