Declining estrogen receptor-beta expression defines malignant progression of human breast neoplasia

It has been shown that the risk of breast cancer developing in certain morphologically identifiable benign breast lesions correlates with expression of estrogen receptor alpha (ER-alpha). Although ER-alpha and ER-beta genes share a large degree of homology, it is generally thought that their distribution and functions are substantially different in many tissues. Recent development of reliable antibodies to ER-beta has provided this first opportunity to test the hypothesis that the likelihood of malignant transformation in morphologically benign breast lesions can be accurately defined by the distribution and level of ER-beta expression relative to that of ER-alpha. Using a monoclonal antibody, ER-beta protein expression has been analyzed in 53 normal breasts and compared with a cohort of histologically distinct breast lesions of different prognostic risk (54 hyperplasia of usual type, 35 ductal carcinoma in situ, and 141 invasive cancers). All of these tissues were also assessed for ER-alpha. Expression of ER-beta protein was also analyzed in an additional spectrum of benign breast lesions with low or negligible risk of progression to malignancy. The median proportion of cells expressing ER-beta was highest in normal breast lobules (median 94.33%, interquartile range 78.25-99.00) but declined significantly through usual ductal hyperplasia (median 76.67, interquartile range 49.17-95.00, P = 0.002) and ductal carcinoma in situ (median 70.00, interquartile range 59.00-85.00, P = 0.009) to invasive cancer (median 60.00, interquartile range 50.00-80.00, P < 0.001). An appreciable proportion (33.81%) of ER-alpha-negative invasive cancers expressed ER-beta. A high but variable level of ER-beta expression occurred in the benign lesions. The data from the intact histologic tissues were evaluated with respect to the relative expression of ER-alpha and ER-beta in five mammary cell lines of different behavioral phenotype (MCF7, ZR-75, T47D, MDAMB231, HUMA121). The highly significant differences in expression and distinct tissue distributions of ER-alpha and ER-beta within the histologic lesions of defined risk, together with the data from the cell lines, support the original hypothesis that the tissue concentration, relative occurrence, and/or interaction of these two types of estrogen receptor may play an important role in modulating mammary tumorigenesis.     

人前列腺癌组织雌激素受体α、β亚单位基因mRNA的表达

目的　了解雌激素受体α(ERα)、β(ERβ)mRNA在人前列腺癌 (PCa)组织中的表达状况。方法　采用半定量逆转录 聚合酶链反应 (RT PCR)技术检测 32例PCa、32例良性前列腺增生患者、12例正常前列腺组织者中ERα、ERβmRNA的表达并进行基因测序。 结果　与良性前列腺增生组织相比 ,PCa组织ERαmRNA表达明显增强 ,而ERβmRNA表达明显减弱 (均P <0 0 1)。中晚期、低分化PCa组织ERαmRNA表达明显高于早期、中高分化PCa,而ERβmRNA则相反 (均P <0 0 1)。激素抵抗型PCaERαmRNA表达增强 ,而ERβmRNA表达明显降低 (均P <0 0 5 )。 结论　ERα、ERβ在PCa组织中的不同表达状况可能与PCa组织病理生物学特性有关。对ER特别是ERβ的研究 ,有助于对PCa特别是激素抵抗型PCa的生物学特性判断和治疗。     

Immunocytochemical localization of estrogen and progesterone receptors in human thyroid

The presence of steroid hormone receptors has previously been suggested in thyroid tissue by biochemical means. Our studies were designed to confirm these results and to localize the specific receptor-containing cell type using a novel immunocytochemical method. Monoclonal antibodies specific to estrogen receptors (ER) and progesterone receptors (PgR) were used to localize these steroid hormone receptors in the human thyroid gland. Frozen tissue sections from surgical specimens excised from 22 patients of both sexes with benign thyroid disease were studied. The sections were incubated with rat antiestrophilin and antiprogesterone receptor antibodies and were then exposed to rabbit anti-rat IgG and to rat peroxidase-antiperoxidase complex. The reaction product was visualized with diaminobenzidine tetrahydrochloride and hydrogen peroxide. Four specimens were positive for both ER and PgR, 16 were ER-positive and PgR-negative, and two were negative for both ER and PgR. Positive reactivity was limited to the follicular lining cell nuclei and varied from focal to diffuse. The immunohistochemical findings confirmed the presence of ER and PgR in the thyroid tissue and demonstrated for the first time that these receptors are present only in the nuclei of the lining cells of the thyroid follicle. The role of steroid hormone receptors in the thyroid in health and disease remains to be explained.     

Histological localization of estrogen receptors in normal and diseased human prostates by immunocytochemistry.

The role of estrogens and estrogen receptors (ER) in the human prostate remains unresolved. In this study we have used the monoclonal ER antibody H222 to investigate the histological localization of ER in normal and diseased human prostates by immunocytochemistry. Prostate tissue was obtained from 3 young organ donors (Group I-normal prostate), from 14 prostates removed by radical prostatectomy or radical cystoprostatectomy, which had caused no or only mild obstructive symptoms (Group II-non-obstructive prostate), and from 11 prostates removed by suprapubic prostatectomy, which had caused severe obstructive symptoms due to a large benign prostatic hyperplasia (BPH) (Group III-obstructive prostate). In prostates of all groups ER were found to be in nuclei of the prostatic urethra and of the periurethral prostatic duct. In striking contrast, ER in the interglandular prostatic stroma was not as homogeneous among the different groups. We observed a low concentration of ER in the stroma of normal prostates, the highest concentration in non-malignant stroma of non-obstructive prostates, and no ER at all in stroma of obstructive prostates. Based on the immunocytochemical localization of ER in normal and diseased human prostate, our results indicate that stromal growth in obstructive BPH may not be mediated via ER. However, we cannot exclude that an increase of stromal ER concentration (as observed in non-obstructive prostates) is directly involved in induction of BPH, leading further prostate growth thereafter into an estrogen independent state.     

Localization of estrogen receptor beta protein expression in adult human bone.

Evidence suggests that the newly described estrogen receptor beta (ER-beta) may be important for estrogen (17beta-estradiol) action on the skeleton, but its cellular localization in adult human bone requires clarification. We addressed this by using indirect immunoperoxidase with a novel affinity purified polyclonal antibody to human ER-beta, raised to hinge domain (D) sequences from the human receptor. Bone was demineralized in 20% EDTA and all biopsy specimens were formalin-fixed and wax-embedded. Vigorous retrieval was essential for ER-beta detection. In sections (5 microm) of benign prostate hyperplasia, used as positive control, clear nuclear immunoreactivity was seen in glandular epithelial cells, with a 1:500 dilution of ER-beta40. For bone sections, optimal antibody dilutions were 1:100-1:250. We found that in normal bone (from graft operations), in fracture callus from both men and women (>25 years old), pagetic bone, osteophytes, and secondary hyperparathyroid bone, all from older patients, ER-beta was expressed clearly in osteoclast nuclei, with little cytoplasmic immunoreactivity. Nuclear immunoreactivity was still prominent in osteoclasts, with antibody diluted 1:500, although it faded in other cells. Osteoblasts, in areas of active bone formation or bone remodeling, also expressed ER-beta, as did some osteocytes. However, hypertrophic chondrocytes were negative, unlike mesenchymal cells, adjacent to the osteogenesis. Megakaryocytes and some capillary blood vessels cells were receptor positive. All ER-beta expression was blocked totally by preincubation of antibody with antigen. We conclude that ER-beta is expressed in cells of osteoblast lineage and in osteoclasts. The latter appear relatively abundant in this receptor and this might provide a means for direct action of estrogen on osteoclasts.     

青少年特发性脊柱侧凸患者骨组织中雌激素受体的表达

目的:探讨青少年特发性脊柱侧凸(AIS)患者骨组织中雌激素受体的表达情况。方法:28例志愿者分为两组,其中AIS组患者18例,男3例,女15例,年龄12～18岁,平均(14.4±1.4)岁,Cobb角40°～125°,平均60.1°;先天性脊柱侧凸(CS)组患者10例,男6例,女4例,年龄11～19岁,平均(15.4±2.6)岁,Cobb角37°～120°,平均63.3°。术中取髂骨,用逆转录聚合酶链反应(RT-PCR)方法检测骨组织中雌激素受体α(ERα)和β(ERβ)的表达情况。结果:AIS组和CS组患者骨组织中ERα mRNA和ERβ mRNA的阳性表达率均无显著性差异(P>0.05);AIS组中ERα mRNA和ERβ mRNA表达阳性患者中的相对含量分别为1.24±0.08和1.07±0.10,CS组为1.24±0.09和1.08±0.10,两组比较无显著性差异(P>0.05);但两组患者ERβ mRNA的阳性表达率与其Cobb角大小呈明显负相关(AIS组r=-0.613,P=0.007;CS组r=-0.648,P=0.043)、与Risser分级无关,ERα mRNA的阳性表达率与Cobb角、Risser分级无关,AIS患者的ERα mRNA和ERβ mRNA的阳性表达率与其Lenke分型无关。结论:AIS患者骨组织中ERα mRNA和ERβ mRNA的表达可能不是引起AIS的主要原因,但ERβ mRNA的表达可能对骨生长的调控作用较大,值得进一步研究。     

Expression of glomerular heparan sulphate domains in murine and human lupus nephritis.

BACKGROUND: Recently, we identified specific N- and 6-O-sulphated heparan sulphate (HS) domains on activated glomerular endothelial cells. In this study, we evaluated in lupus nephritis the expression of different HS domains on glomerular endothelium and in the glomerular basement membrane (GBM). METHODS: The expression of specific glomerular HS domains and the presence of immunoglobulins (Ig) were determined by immunofluorescence staining of kidney sections of patients with nephritis due to systemic lupus erythematosus (SLE) and MRL/lpr lupus mice. The expression/presence of glomerular HS domains and Ig was also evaluated after eluting Ig from renal sections of lupus mice using two elution methods, and in renal sections of lupus mice treated with heparinoids. RESULTS: Both MRL/lpr mice and patients with lupus nephritis showed a decreased expression of HS in the GBM. The expression of N- and 6-O-sulphated HS domains on glomerular endothelium was decreased in MRL/lpr mice, but increased in SLE patients. MRL/lpr mice had more extensive glomerular Ig deposits than SLE patients. After elution of Ig, the glomerular endothelial expression of N- and 6-O-sulphated HS domains in MRL/lpr mice was recovered and even increased above normal levels, while the expression of HS in the GBM was restored to normal levels. Treatment with heparinoids prevented Ig deposition and preserved the expression of glomerular HS domains at normal levels in lupus mice. CONCLUSION: The expression of specific HS domains on glomerular endothelium and in the GBM is changed during lupus nephritis due to masking by Ig deposits and induction of inflammatory N- and 6-O-sulphated HS domains.     

Mechanical strain and estrogen activate estrogen receptor alpha in bone cells.

Bone cells' early responses to estrogen and mechanical strain were investigated in the ROS 17/2.8 cell line. Immunoblotting with antiphosphorylated estrogen receptor a (ER-alpha) antibody showed that when these cells were exposed for 10 minutes to estrogen (10(-8) M) or a single period of cyclic dynamic strain (peak 3400 microepsilon, 1 Hz, 600 cycles), there was an increase in the intensity of a 66-kDa band, indicating phosphorylation of ser122 in the amino terminus of ER-alpha. Increased phosphorylation was detected within 5 minutes of exposure to estrogen and 5 minutes after the end of the period of strain. Estrogen and strain also activated the mitogen-activated protein kinase (MAPK) family member extracellular regulated kinase-1 (ERK-1). Increases in ERK activation coincided with increased ER-alpha phosphorylation. Activation of ERK-1 and the phosphorylation of ER-alpha, by both estrogen and strain, were prevented by the MAP kinase kinase (MEK) inhibitor U0126 and the protein kinase A (PKA) inhibitor (PKI). These data support previous suggestions that resident bone cells' early responses to strain and estrogen share a common pathway, which involves ER-alpha. This pathway also appears to involve PKA and ERK-mediated phosphorylation of ser122 within the amino terminus of ER-alpha. Reduced availability of this pathway when estrogen levels are reduced could explain diminished effectiveness of mechanically related control of bone architecture after the menopause.     

Expression of estrogen receptor (ER-alpha and ER-beta) mRNA in human prostate cancer.

OBJECTIVE: The distribution of the two estrogen receptors (ER-alpha, ER-beta) in human prostate tissue have not been fully clarified, so the present study investigated the mRNA expression of the receptors to explain the broad spectrum of estrogen activity in prostate cancer. MATERIALS AND METHODS: Four human prostate cancer cell lines (LNCap, JCA-1, DU-145 and PC-3) and 24 pairs of untreated prostate cancer tissue and noncancerous tissue from resected prostate glands were subjected to RT-PCR testing. RESULTS: Both LNCap and JCA-1 expressed the mRNA of both receptors, but DU-145 and PC-3 only expressed ER-beta mRNA. In the human prostate tissue samples, 20 of the 24 prostate cancer tissues expressed ER-alpha, and 23 of the 24 expressed ER-beta. Of the 24 noncancer tissues, 14 expressed ER-alpha mRNA and 17 expressed ER-beta mRNA. The incidence of ER-beta mRNA expression between the paired cancer and noncancer tissues was statistically significantly different (p<0.05). CONCLUSIONS: A higher incidence of ER-beta mRNA expression in untreated prostate cancer tissues was observed. Furthermore, the absence of ER-alpha mRNA and the presence of ER-beta mRNA expression in hormone-independent and/or untreated prostate cancer cells leads to a tentative speculation of the mechanism of the hormone refractory feature of prostate cancer.     

Expression of smooth muscle actin in osteoblasts in human bone.

It is well known that certain connective tissue cells (viz., dermal fibroblasts) can express the gene for a muscle actin--alpha-smooth muscle actin--and can contract. This process contributes to skin wound closure and is responsible for Dupuytren's contracture. The objective of this study was to determine if human osteoblasts can also express the gene for alpha-smooth muscle actin. Immunohistochemistry using a monoclonal antibody for alpha-smooth muscle actin was performed on human cancellous bone samples obtained from 20 individuals at the time of total joint arthroplasty. The percentages of resting and active osteoblasts on the bone surfaces containing this muscle actin isoform were evaluated. Explants of human bone were also studied for the expression of alpha-smooth muscle actin in the tissue and in the outgrowing cells with time in culture. Western blot analysis was performed to quantify the alpha-smooth muscle actin content of the outgrowing cells relative to smooth muscle cell controls. Nine +/- 2% (mean +/- SEM; n = 20) of the cells classified as inactive osteoblasts and 69 +/- 3% (n = 19) of the cells identified as active osteoblasts on the bone surface contained alpha-smooth muscle actin. This difference was highly statistically significant (Student's t test, p < 0.0001). Similar profiles of alpha-smooth muscle actin-expressing cells were found in explants cultured for up to 12 weeks. Cells forming a layer on the surface of the explants and growing out from them in monolayer also contained alpha-smooth muscle actin by immunohistochemistry and Western blot analysis. Human osteoblasts can express the gene for alpha-smooth muscle actin. This expression should be considered a phenotypic characteristic of this cell type, conferred by its progenitor cells: bone marrow stromal-derived stem cells, and perhaps pericytes and smooth muscle cells.     

维甲酸β受体在结直肠癌组织中的表达及其意义

目的 探讨维甲酸β受体(RAR-β)在结直肠癌组织中的表达及其意义.方法 应用免疫组化法检测2006年1月至2007年1月收治的60例结直肠癌患者肿瘤组织、癌旁组织及正常组织标本中RAR-β的表达,采用化学发光免疫法检测该60例患者术前血清癌胚抗原(CEA)水平,结合其临床病理资料,分析人结直肠癌组织中RAR-β的表达情况及其意义.结果 60例结直肠癌患者的肿瘤组织中RAR-β阳性表达率(48%)显著低于正常组织(87%)及癌旁组织(87%)(P<0.05).有淋巴结转移患者的肿瘤组织中RAR-B阳性表达率(32%)显著低于无淋巴结转移患者的肿瘤组织(60.0%),且晚期肿瘤患者(TNM Ⅲ、Ⅳ期)的RAR-β阳性表达率(29%)显著低于早期肿瘤患者(Ⅰ、Ⅱ期)(69%)(P均<0.05).RAR-β阳性表达率在高、中、低分化肿瘤组织中无明显差异.本组患者中,CEA水平升高者20例,正常者40例,有淋巴结转移及晚期肿瘤患者中CEA水平升高者(48%、52%)分别高于无淋巴结转移及早期肿瘤患者(23%、14%)(P均<0.05).结论 RAR-β在结直肠癌组织中表达下调,这可能与结肠癌发生相关,且其表达下降程度与淋巴结转移情况及TNM分期呈明显负相关,与CEA类似,其表达可作为结直肠癌的预后指标.     

Autoradiographic localization of estrogen and androgen target cells in human and rat prostate carcinoma

The distribution of estrogen target cells within the Dunning R3327-H rat prostate tumor following intravenous injection of tritiated estradiol into rat hosts was compared to the distribution obtained following incubation of a 2 mm. sample of the tumor with tritiated estradiol in organ culture. No difference was observed, indicating that the in vitro method was an effective approach for autoradiographic analysis of tumor biopsy samples. Subsequently, tumor samples were excised from solid tumors of R3327-H and R3327-MAT LyLu tumors growing in Copenhagen rats. These tumor models were chosen as representatives of hormone sensitive (R3327-H) and hormone insensitive (R3327-MAT LyLu) tumors. Normal rat dorsal prostate and human tumor biopsy samples were also studied. Autoradiographic studies were performed in vitro utilizing tritiated estradiol and tritiated dihydrotestosterone to compare the distribution of estrogen and androgen target cells. The present research demonstrated that 1) similar patterns of nuclear uptake of steroids are obtained with in vivo and in vitro autoradiographic techniques, 2) estradiol receptors occur primarily in extra-acinar epitheloid cells in both rat and human prostate carcinomas, 3) these epithelioid cells are not characteristic of the normal rat dorsal prostate, 4) androgen receptors occur in both acinar and stromal epithelioid cells in rat and primarily in acinar epithelial cells in human tumors and 5) in vitro autoradiographic methods can provide insight into differences in sensitivity to steroids which may be of diagnostic importance in the treatment of cancer.     

Dietary intake of phytoestrogens, estrogen receptor-beta polymorphisms and the risk of prostate cancer

BACKGROUND: The causes of prostate cancer are poorly understood, but genetic factors may be more important than for many other malignancies, and dietary phytoestrogens may be protective. Because phytoestrogens bind tightly to the estrogen receptor-beta, we conducted an epidemiologic investigation of synergistic effects between phytoestrogen intake and estrogen receptor-beta gene polymorphisms. METHODS: We performed a population-based case-control study in Sweden. All participants reported their phytoestrogen intake and donated a blood sample. We identified four haplotype-tagging single nucleotide polymorphisms (htSNPs) and genotyped these htSNPs in 1314 prostate cancer patients and 782 controls. Odds ratios were estimated by multivariate logistic regression. Interactions between phytoestrogen intake and estrogen receptor-beta SNPs on prostate cancer risk were evaluated considering both multiplicative and additive effect scales. RESULTS: We found a significant multiplicative interaction (P = 0.04) between dietary intake of phytoestrogens and a promoter SNP in the estrogen receptor-beta gene (rs 2987983-13950), but not with any of the three other htSNPs (P = 0.11, 0.69, 0.85). Among carriers of the variant promoter alleles, we found strong inverse associations with increasing intake of total phytoestrogens (odds ratio for highest vs. lowest quartile = 0.43; P for trend <0.001), isoflavonoids (odds ratio = 0.63; P for trend = 0.05), and coumestrol (odds ratio = 0.57; P for trend = 0.003). We found no association between phytoestrogens and prostate cancer among carriers homozygous for the wild-type allele (TT). CONCLUSIONS: Our study provides strong evidence that high intake of phytoestrogens substantially reduce prostate cancer risk among men with specific polymorphic variation in the promoter region of the estrogen receptor-beta gene.     

Expression and regulation of chemokines in murine and human type 1 diabetes

More than one-half of the ~50 human chemokines have been associated with or implicated in the pathogenesis of type 1 diabetes, yet their actual expression patterns in the islet environment of type 1 diabetic patients remain, at present, poorly defined. Here, we have integrated a human islet culture system, murine models of virus-induced and spontaneous type 1 diabetes, and the histopathological examination of pancreata from diabetic organ donors with the goal of providing a foundation for the informed selection of potential therapeutic targets within the chemokine/receptor family. Chemokine (C-C motif) ligand (CCL) 5 (CCL5), CCL8, CCL22, chemokine (C-X-C motif) ligand (CXCL) 9 (CXCL9), CXCL10, and chemokine (C-X3-C motif) ligand (CX3CL) 1 (CX3CL1) were the major chemokines transcribed (in an inducible nitric oxide synthase-dependent but not nuclear factor-κB-dependent fashion) and translated by human islet cells in response to in vitro inflammatory stimuli. CXCL10 was identified as the dominant chemokine expressed in vivo in the islet environment of prediabetic animals and type 1 diabetic patients, whereas CCL5, CCL8, CXCL9, and CX3CL1 proteins were present at lower levels in the islets of both species. Of importance, additional expression of the same chemokines in human acinar tissues emphasizes an underappreciated involvement of the exocrine pancreas in the natural course of type 1 diabetes that will require consideration for additional type 1 diabetes pathogenesis and immune intervention studies.     

Low content of estrogen receptors in human giant cell tumors of bone

We tried to identify estrogen receptors (ERs) in human giant cell tumors of bone. Samples from eight patients were used for biochemical and immunohistochemical studies using [¹²⁵I]17-estradiol and monoclonal antibody against the human ER. Estradiol binding sites were detected in seven cases out of eight. The dissociation constant of the higher affinity binding site was 0.5 nM. Nonlabeled 17-estradiol and synthetic estrogen, diethylstilbestrol, inhibited the binding of labeled 17-estradiol to the tumor cytosol, while dexamethasone did not inhibit the binding. Using a monoclonal antibody, Western blotting identified bands ofMr 68000 and 59000. No ERs were observed in any case examined immunohistochemically. These results suggest that human giant cell tumors of bone have a low level of ERs that cannot be detected immunohistochemically.