Estrogen Resistance Research Articles

The effects of oestrogens on linear bone growth.

Regulation of linear bone growth in children and adolescents comprises a complex interaction of hormones and growth factors. Growth hormone (GH) is considered to be the key hormone regulator of linear growth in childhood. The pubertal increase in growth velocity associated with GH has traditionally been attributed to testicular androgen secretion in boys, and to oestrogens or adrenal androgen secretion in girls. Research data indicating that oestrogen may be the principal hormone stimulating the pubertal growth spurt in boys as well as girls is reviewed. Such an action is mediated by oestrogen receptors (ER-alpha and ER-beta) in the human growth plate, and polymorphisms in the ER gene may influence adult height in healthy subjects. Prepubertal oestradiol concentrations are significantly higher in girls than in boys, explaining sex-related differences in pubertal onset. Men with a disruptive mutation in the ER gene (oestrogen resistance) or in the CYP19 gene (aromatase deficiency) who have no pubertal growth spurt and continue to grow into adulthood due to lack of epiphyseal fusion supports this notion. Furthermore, phenotypic females with complete androgen insensitivity syndrome have a normal female growth spurt despite lack of androgen action. Oestrogens may also influence linear bone growth indirectly via modulation of the GH-insulin-like growth factor-I (IGF-I) axis. Thus, ER blockade diminishes endogenous GH secretion, androgen receptor (AR) blockade increases GH secretion in peripubertal boys, and non-aromatizable androgens [oxandrolone or dihydrotestosterone (DHT)] have no effect on GH secretion. Treatment with aromatase inhibitors reduces circulating IGF-I concentrations in healthy males, and reduces growth in boys with testotoxicosis. Taken together, these findings suggest that oestrogens may, in addition to their direct effects, stimulate GH secretion and thereby increase circulating IGF-I, which in turn may stimulate growth. Thus, oestrogens have important biphasic actions on longitudinal growth in boys as well as in girls. Very low levels of oestrogens may stimulate bone growth without affecting sexual maturation directly at the growth plate as well as through stimulation of the GH-IGF axis, which in turn may stimulate growth. Conversely, higher levels of oestrogens stimulate secondary sexual characteristics and epiphyseal fusion.

Loss of growth inhibitory effects of retinoic acid in human breast cancer cells following long-term exposure to retinoic acid.

Although retinoids are known to be inhibitory to breast cancer cell growth, a key remaining question is whether they would remain effective if administered long-term. We describe here the long-term effects of all-trans retinoic acid on two oestrogen-dependent human breast cancer cell lines MCF7 and ZR-75-1. Although both cell lines were growth inhibited by retinoic acid in the short-term in either the absence or the presence of oestradiol, prolonged culture with 1 μM all-trans retinoic acid resulted in the cells acquiring resistance to the growth inhibitory effects of retinoic acid. Time courses showed that oestrogen deprivation of the cell lines resulted in upregulation of the basal non-oestrogen stimulated growth rate such that cells learned to grow at the same rate without as with oestradiol, but the cells remained growth inhibited by retinoic acid throughout. Addition of 1 μM all-trans retinoic acid to steroid deprivation conditions resulted in reproducible loss of growth response to both retinoic acid and oestradiol, although the time courses were separable in that loss of growth response to retinoic acid preceded that of oestradiol. Loss of growth response to retinoic acid did not involve loss of receptors, ER as measured by steroid binding assay or RARα as measured by Northern blotting. Function of the receptors was retained in terms of the ability of both oestradiol and retinoic acid to upregulate pS2 gene expression, but there was reduced ability to upregulate transiently transfected ERE- and RRE-linked reporter genes. Despite the accepted role of IGFBP3 in retinoic acid-mediated growth inhibition, progression to retinoic acid resistance occurred irrespective of level of IGFBP3, which remained high in the resistant MCF7 cells. Measurement of AP1 activity showed that the two cell lines had markedly different basal AP1 activities, but that progression to resistance was accompanied in both cases by a lost ability of retinoic acid to reduce AP1 activity. These results warn of potential resistance which could arise on long-term treatment with retinoic acid in a clinical situation and echo the problems of progression to endocrine resistance. It seems that whatever the constraints imposed on growth, these cells have a remarkable ability to escape from growth inhibition. However, the ability of retinoic acid to delay progression to oestrogen resistance is encouraging for endocrine therapy, and the concentration-dependence of retinoic acid resistance suggests that progression is not absolute but could be manipulated by dose. © 2000 Cancer Research Campaign

Aromatase deficiency and estrogen resistance: from molecular genetics to clinic.

Our knowledge of the physiologic roles of estrogen in women and men has been advanced by recent descriptions of mutations disrupting estrogen biosynthesis and action. Aromatase deficiency results from autosomal recessive inheritance of mutations in the CYP19 gene. It gives rise to ambiguous genitalia in 46,XX individuals. At puberty, affected girls have hypergonadotropic hypogonadism, fail to develop secondary sexual characteristics, and exhibit progressive virilization. The affected 46,XY individuals have normal male sexual differentiation and pubertal maturation. These men are extremely tall and have eunuchoid proportions with continued linear growth into adulthood, lack of epiphyseal closure, and osteoporosis due to estrogen deficiency. Although estrogen was shown to be essential for normal sperm production and function in mice, its role in fertility is not clear in men. Thus far, one estrogen-resistant human, a man with a mutant estrogen receptor-alpha gene, has been described. His clinical presentation was similar to that of aromatase-deficient men.

Estrogen: consequences and implications of human mutations in synthesis and action.

Recent developments have advanced our knowledge of the role of estrogen in the male. Studies of the mutations in CYP19, the gene encoding aromatase, in six females and two males and a mutant estrogen receptor alpha in a man are described. These observations provide illuminating new insights into the critical role of estrogen in the male (as well as female) in the pubertal growth spurt and skeletal maturation, and in the importance of estrogen sufficiency in the accrual and maintenance of bone mass. The weight of evidence supports an effect of androgens on the latter processes, but this effect has not been quantitated. There is a discordance in the estrogen-deficient male between skeletal growth and skeletal maturation and the accrual of bone mass and density. Estrogen synthesis by the testis is limited before puberty, and estrogen deficiency does not affect the age of pubertal onset. Estrogen deficiency in men leads to hypergonadotropism, macroorchidism, and increased testosterone levels. Estrogen lack has a significant effect on carbohydrate and lipid metabolism, and estrogen resistance was associated with evidence of premature coronary atherosclerosis in a man. These observations have highlighted the role of extraglandular estrogen synthesis and intracrine and paracrine actions. In the human, in contrast to nonprimate vertebrates, aromatase deficiency and estrogen resistance (alpha) does not seem to affect gender identity or psychosexual development. The clinical repercussions of mutations in CYP19 on the fetal-placental unit have highlighted the major role of placental aromatase in the protection of the female fetus from androgen excess, thus preventing androgen-induced pseudohermaphrodism and virilization of the mother. These features are compared with the virilization that occurs in utero in the female spotted hyena. The novel features of the aromatase deficiency syndrome in the affected female--in the fetus, during childhood, and at puberty--are discussed, including virilization at puberty and development of polycystic ovaries. The severity of the syndrome correlates with the severity of impairment of aromatase formation in expression systems. Finally, the structural consequences of missense mutations in CYP19 are described in accordance with a model of the structure of human aromatase.

Disproportional Body Growth in Female Estrogen Receptor-α-Inactivated Mice

Estrogens play an important role in the regulation of longitudinal bone growth in man, as demonstrated by recent descriptions of individuals with estrogen insensitivity or aromatase deficiency. Two estrogen receptors, ERα and ERβ, have been cloned. The aim of the present study was to investigate the function of ERα in the regulation of body growth and skeletal growth. Adult female mice with inactivated ERα (ERα−/−) demonstrated an increased body weight compared with wild-type mice (114% of control). However, the length of the appendicular skeleton was decreased in adult ERα−/− mice (femur 93% of control). In contrast, the axial skeleton was normal (crown–rump length 98% of control). The decreased growth of the appendicular skeleton was associated with decreased serum levels of IGF-I (77% of control), indicating that the GH/IGF-I axis may be involved in the decreased longitudinal bone growth seen in female ERα−/− mice.

Estrogen and vascular resistance

Male sex and menopause are independent risk factors for cardiovascular disease. Estrogens bind to vascular receptors, including the novel estrogen receptor-β and membrane-bound receptors, and they contribute to vascular resistance through several mechanisms. Estrogens modulate gene expression, membrane potentials, and expression of receptors; modify activity of vasoactive peptides and vascular enzymes; and act as antioxidants, favoring a state of vasodilatation and inhibiting vascular growth. Accordingly, estrogens have been shown to improve vascular function in animals and humans and to inhibit the response to injury after balloon angioplasty and the progression of atherosclerosis. New mechanisms of estrogen action, the role of estrogen receptor-β in the vasculature, and recent evidence for synthetic progestogens as potential antagonists of vascular estrogen actions are discussed.

Alternative Initiation of Translation Accounts for a 67/45 kDa Dimorphism of the Human Estrogen Receptor ERα

The estrogen receptor protein, in the nuclear receptor superfamily, carries two transactivator domains designated AF1 and AF2. The activity of AF2, localized in the carboxy-terminal region, is ligand-dependent, whereas AF1 (amino-terminal) seems to be activated via the MAPKkinase pathway. Uterine and mammary cells exhibiting large amounts of ERα were the first estrogen target organs demonstrated. The response intensity in these tissues is related to the affinity of the receptor and to the number of sites occupied by its ligand. Certain physiological and pharmacological phenomena of estrogen resistance associated with a truncated form of ERα (deleted in the AF1 domain) would seem however to challenge this assertion. The 45 kDa truncated form is unable to induce cell proliferation but can still increase the expression of certain genes. In this work we suggest that this 45 kDa ERα form may originate from differential regulation of translation of the mRNA encoding the ERα.In vitrotranslation studies and transient expression in COS-7 cellsin vivodemonstrated a mechanism of translation regulation that produced from a given mRNA either the wild type ER 67 kDa form or the AF1 deleted ER 45 kDa isoform. Bicistronic vectors were used to demonstrate that the 45 kDa protein originates from translation initiation at AUG 174 induced by an internal ribosome entry.

The effects of oestrogen exposure on bone mass in male to female transsexuals

The importance of oestrogen on bone mineral density (BMD) in males was suggested by reports of patients with oestrogen resistance and aromatase deficiency who demonstrated osteoporosis and epiphyseal plate maturation defect despite high testosterone levels. In the present study, we examined the effects of oestrogen exposure on BMD in transsexual men. Cross-sectional study of BMD in male to female transsexuals. Subjects consisted of two groups of transsexual male dancers aged 16-34 years who did not receive transsexual operations (n = 28). Group 1 (n = 11) and group 2 (n = 17) had used oestrogen for 2 years or less and more than 2 years, respectively. Twenty-four healthy adult males served as controls. Signs of feminization were presented in both group 1 and group 2, with Tanner's stage II-III breast development. BMD at various sites were correlated only to body weight and not to smoking or milk consumption. After controlling for body weight, it was found that group 2 had significantly higher BMD at L2-4 than controls (1.22 +/- 0.03 vs. 1.14 +/- 0.03 g/cm2, P < 0.05) and group 1 (1.22 +/- 0.03 vs. 1.08 +/- 0.04 g/cm2, P < 0.05). BMD at femoral neck was also higher in group 2 compared to controls (1.10 +/- 0.03 vs. 1.01 +/- 0.03 g/cm2, P < 0.05) and group 1 (1.10 +/- 0.03 vs. 0.95 +/- 0.04 g/cm2, P < 0.05). Group 1 subjects had lower BMD compared to controls at femoral trochanter (0.70 +/- 0.04 vs. 0.83 +/- 0.03 g/cm2, P < 0.05) and total femur (0.96 +/- 0.05 vs. 1.07 +/- 0.03 g/cm2, P < 0.05). Long-term oestrogen exposure transsexual men result in an increase in bone mineral density despite signs of feminization. This suggests that oestrogen has positive effects on bone density in males. The finding of the trend towards reduced bone density in group 1 remains unexplained.

Serum oestradiol and oestrogen-receptor gene polymorphism are associated with bone mineral density independently of serum testosterone in normal males.

The physiological effects of oestrogens on bone in men were largely unanticipated until recently, when oestrogen deficiency in males with aromatase deficiency and oestrogen resistance was found to cause osteoporosis and delayed fusion of epiphyses despite sufficient serum testosterone. This raises the possibility that in normal men oestrogens rather than androgens are of physiological importance in bone maturation. In the present study, we examined the association of serum oestradiol (E2) compared to that of free testosterone (FT) with bone mineral density (BMD) in normal men. The effect of oestrogen receptor (ER) gene polymorphism on BMD in men was also addressed. Eighty-one Thai men aged 20-79 years. All were healthy and did not take medication which may affect calcium and bone metabolism. BMD was assessed by DEXA. Dietary calcium was assessed by a 3-day dietary record. Serum E2 and FT concentrations were measured by radioimmunoassay. Polymorphism at intron 1 of the alpha isoform of ER gene was determined by PCR-RFLP. Small p represents the presence of the restriction site while capital P indicates the absence of the restriction site. Serum FT decreased with increasing age (r = -0.58, P < 0.0001) while E2 did not. However, there was a positive association between E2 and FT (r = 0.28, P < 0.05). Serum FT was related to BMD at femoral neck (r = 0.26, P < 0.05) and Ward's triangle (r = 0.30, P < 0.01) while E2 was related to BMD at anteroposterior (AP) lumbar spine (r = 0.29, P < 0.05), femoral neck (r = 0.23, P < 0.05) and femoral trochanter (r = 0.27, P < 0.05). Besides FT and E2, age, body weight, fat mass and fat-free mass were also correlated to BMD at various skeletal sites. Using stepwise multiple linear regression to control for the confounding effects among these factors, fat-free mass was found to be strongly associated with BMD at most skeletal sites. Serum E2 was related to BMD independently of other factors including FT at AP lumbar spine (r = 0.22, P < 0.05), femoral neck (r = 0.26, P < 0.01), femoral trochanter (r = 0.22, P < 0.05) and Ward's triangle (r = 0.26, P < 0.01) while serum FT was not associated with BMD at any site after controlling for E2 and other related factors. Concerning ER alpha gene polymorphism, 27 (33.3%) of the subjects had pp genotype, while 42 (51.9%) and 12 (14.8%) Pp and PP genotypes, respectively. After controlling for age, body weight, fat mass, fat-free mass, calcium intake, FT and E2, the presence of P allele was associated with higher BMD at AP L2-L4 (P < 0.05). Serum oestradiol is more related to bone mass than free testosterone in normal men. Oestrogen-receptor gene polymorphism is also associated with bone mass in men independently of oestradiol levels. Serum oestradiol together with oestrogen-receptor genotype may partly determine bone mass in males.

Cloning and Expression of a Novel Dominant-Negative-acting Estrogen Response Element-binding Protein in the Heterogeneous Nuclear Ribonucleoprotein Family

Most genera of New World primates exhibit a compensated form of resistance to steroid hormones produced by the adrenal gland, gonads, and kidney. Estrogen resistance in New World primate cells is associated with the relative overexpression of a nonreceptor-related estrogen response element-binding protein (ERE-BP) that competes with estrogen receptor for ERE binding. Using the concatamerized ERE half-site (AGGTCAcag) in DNA affinity chromatography, we purified to homogeneity a 40-42-kDa ERE-BP. The affinity-purified ERE-BP bound specifically to either single- or double-stranded DNA bearing the consensus ERE half-site motif AGGTCA. Four distinct internal tryptic peptides from this protein were generated and shown to exhibit sequence similarity to proteins in the heterogeneous nuclear ribonucleoprotein family. These tryptic peptide fragments were used to generate a series of degenerate oligonucleotides that were successfully employed in isolating a full-length ERE-BP cDNA by polymerase chain reaction. Although a member of a family of proteins generally recognized for their ability to bind single strand RNA, the estrogen resistance-associated protein ERE-BP can effectively bind double strand DNA and competitively squelch estrogen receptor-directed transactivation.

Genetic mutations resulting in estrogen insufficiency in the male

The biosynthesis of estrogens is catalyzed by an enzyme known as aromatase (aromatase cytochrome P450; P450 arom; the product of the CYP19 gene). In recent years a number of patients have been described suffering from aromatase deficiency due to mutations in the CYP19 gene, resulting in the synthesis of a non-functional gene product and a resulting failure to synthesize estrogens. Males with this condition have sustained linear growth into adulthood resulting from failure of epiphyseal closure. Osteopenia and reduced bone mineral density and bone age are also characteristic. Lack of circulating estrogens is accompanied by elevated testosterone and gonadotropins. One of the men had macroorchidism with testicular volumes in excess of 25 ml (Morishima et al. J. Clin. Endocrinol. Metab. 80, 3689, 1995). Semen analysis was not performed on this patient, but it is of note that the one patient described with estrogen insensitivity due to a mutation in the estrogen receptor had a normal sperm count, although motility was decreased (Smith et al., New England J. Med. 331. 1056, 1994). By contrast, the other man with aromatase deficiency had testicular volumes of only 8 ml per testes, and was infertile. Sperm analysis revealed a count of 1 million/ml with 100% immotile sperm (Carani et al. New England J. Med. 337, 91, 1997). However, his clinical picture is confused by the fact that another male sibling has azoospermia, but has no CYP19 mutation, suggesting that the infertility problem may be due to a second genetic condition in this consanguineous family. Recently mice have been generated in which the aromatase (CYP19) gene and the gene encoding the estrogen receptor- α have been inactivated by targeted disruption (ArKO and ERKO mice, respectively). Male ERKO mice are infertile with atrophy of the testes and seminiferous tubule dysmorphogenesis resulting in decreased spermatogenesis and inacive sperm. By contrast the ArKO mice are initially fertile and sire litters of normal size ad frequency, however with advancing age the number of litters sired decreases relative to those of wild type litter ates. In contrast to the ERKO mice, light microscopic analysis of the testes of the ArKO mice reveals no gross morphological abnormalties and the testes are of normal size. Following recent observations that the estrogen receptor- β isoform is highly expressed in seminiferous epthelium, spermatids and spermatocytes, it is conceivable that the relatively high levels of estrogens present in the ERKO mice can act through the ER- β to cause infertility by a direct action on the testes. In this context it is well known that administration of high levels of estrogen to men results in infertility. It is apparent that studies of human and mouse models with disruptions of aromatase and the estrogen receptor have as yet failed to clarify the role of estrogens in male fertility and testicular function. Development of an ER- β knockout mouse, or else a double, or even triple, knockout model, may be required in order to resolve these issues.

The influence of estrogen on growth.

The greatest advances in our understanding of the role of estrogens in growth and skeletal maturity have come from male and female patients who are devoid of estrogen or have estrogen insensitivity. Among males, the absence of an estrogen effect results in lack of completion of skeletal maturity and continued growth into adult life, resulting in very tall stature. This has been exemplified in one male with an estrogen-receptor defect and one male with aromatase deficiency. In females with aromatase deficiency, androgen levels are excessive, causing inappropriate virilization but failing to induce skeletal maturity. Thus, estrogen is the hormone necessary for normal skeletal growth and maturity. Estradiol concentrations measured using an ultrasensitive assay are similar in males and females at the time of the pubertal growth spurt; these data are consistent with the premise that estrogen has a role in growth.

Importance of estrogens in human males' fertility and bone pathophysiology

Since estrogens play a crucial role in several processes such as embryo implantation and fetal development it has been thought that mutation of the estrogenreceptor (ER) gene would be lethal. This hypothesis was supported by the fact that no mutations of the ER gene leading to estrogen insensitivity had been observed in mammalians. In contrast, conditions of resistance to other hormones (tri-iodothyronine, vitamin D, cortisol, androgens), which also act via receptors belonging to the nuclear receptor superfamily, have been reported with increasing frequency. However, this notion changed in 1994 when Korach (1) created a mutant mouse line without a functional ER, obtained by the gene knock-out technique (ER knock-out; ERKO). These animals provided an attractive experimental model which could be used to gain a better understanding of the role of estrogens in a variety of tissues during early developmental stages. The main finding was that crosses of the heterozygous ERKO mice resulted in live birth offspring containing the expected Mendelian distribution of the three possible genotypes. This result suggested that, in contrast to what was previously thought, disruption of the ER was not crucial for survival. Furthermore, the homozygous ERKO mice showed no bias in sex ratio, indicating that sex determination was not affected by the absence of the estrogen receptor. As far as the phenotype was concerned, both sexes of the homozygous ERKO mice were infertile, as demonstrated by mating studies, and exhibited a variety of changes, some of which were associated with the gonads, mammary glands, reproductive tracts and skeletal tissues. In particular, homozygous mutant female mice showed the presence of hypoplastic reproductive tract structures, hemorrhagic cystic ovaries with no functional corpora lutea and no uterine response to estrogen treatment. The mammary glands were undeveloped with only vestigial ducts present at the nipples. Of particular relevance was the observation that the bone density of the ERKO male and female mice was 20 to 25% lower than in wild-type mice, suggesting a direct role for estrogen in bone physiology. An unexpected finding was the infertility of the sexually mature male adult ERKO mice, despite the presence of anatomically normal reproductive apparatus. In order to assess the essential role for ER-mediated processes in the regulation of male reproduction, further studies on ERKO male mice were recently carried out by Eddy and colleagues (2). These authors demonstrated that the absence of ER was detrimental to spermatogenesis, sperm function and mating performance. Although the findings in ERKO mice were exciting, their relevance to human physiology appeared to be questionable, due to the species differences in genetic background. Nevertheless, the first case report of estrogen resistance in man due to mutation in the ER gene (3) provided the evidence that the ERKO mouse could be an acceptable model for the evaluation of a variety of estrogen responses and accompanying mechanisms related to human population. In the above report, the patient was a 28-year-old white man, fully masculinized, who had osteoporosis, unfused epiphyses and continuing linear growth in adulthood. He also had elevated serum estrogen, steroid and gonadotropin levels and no target-tissue responses to estrogen therapy. Genetic analysis showed that the patient was homozygous for a mutation in the second exon (R157X) of the ER gene, generating a premature stop codon. Similar to the observations in the ERKO mice, these findings suggested that estrogens in human males have a pivotal role in bone maturation and mineralization and are also important for complete epiphyseal closure. Therefore, normal androgen levels alone, as in this case, are not sufficient to promote skeletal maturation and retain bone mass, in contrast with the idea that androgens are important for the maintenance of bone mass in men. Data on the fertility of this patient were not available.

Improving Hormonal Therapy for Breast Cancer

Abstract: Since Beatson's observation of breast cancer regression after oophorectomy one hundred years ago (1), therapies directed toward the estrogen-estrogen receptor (ER) endocrine axis have become a cornerstone of breast cancer treatment. Surgical ablative techniques, such as oophorectomy, adrenalectomy, and hypophysectomy have been largely replaced by pharmacological approaches that inhibit estrogen-dependent breast cancer growth, either by reducing estrogen synthesis (LHRH agonists, aromatase inhibitors, and progestins), by blocking ER function (tamoxifen) or by interfering with the cellular response to ER activation (androgens). Indeed, one could argue that hormone therapy for breast cancer represents one of the most successful areas of logical drug design in cancer treatment. Recent drug developments have led to improved side-effect profiles for hormonal agents. In addition, the remarkable organ-selective agonist/antagonist properties of tamoxifen-related compounds are the basis for an effort to develop hormone replacement therapies that combine breast cancer treatment/prevention with reduced osteoporosis and heart disease risk. While the ER axis will remain a primary focus for the further development of hormonal therapies (Table 1), the value of ER as a therapeutic target will always be limited by the primary or acquired estrogen insensitivity that nearly all breast cancers ultimately display. Current hopes for circumventing this problem lie in the vast therapeutic potential afforded by recent molecular insights into growth regulatory pathways. Logical choices for novel therapeutic targets for “estrogen insensitive” breast cancer includes alternative nuclear hormone receptors, growth factor receptors, and signal transduction molecules in “ER-independent” growth-regulatory pathways. Inhibitors of pathophysiological processes intrinsic to tumor progression, such as angiogenesis, invasion, and metastasis are also promising noncytotoxic alternatives in breast cancer therapy (Table 2). Studies of growth regulators in breast cancer are also likely to lead to new tumor markers to accurately guide choices from the increasing array of therapeutic options available.

Interferon alfa-2c and proteinuria in a patient with focal and segmental glomerulosclerosis

Vol 349 • April 19, 1997 1147 women. JAMA 1991; 265: 1861–67. 2 MacDonald PC, Madden JD, Brenner PF, Wilson JD, Siiteri PK. Origin of estrogen in normal men and in women with testicular feminization. J Clin Endocrinol Metab 1979; 49: 905–16. 3 Bagatell CJ, Knopp RH, Rivier JE, Bremner WJ. Physiological levels of estradiol stimulate plasma high density lipoprotein2 cholesterol levels in normal men. J Clin Endocrinol Metab 1994; 78: 855–61. 4 Smith EP, Boyd J, Frank GR, et al. Estrogen resistance caused by a mutation in the estrogen-receptor gene in a man. N Engl J Med 1994; 331: 1056–61. 5 Celermajer DS, Sorensen KE, Gooch VM, et al. Non-invasive detection of endothelial dysfunction in children and adults at risk of atherosclerosis. Lancet 1992; 340: 1111–15.

Endothelial dysfunction in a man with disruptive mutation in oestrogen-receptor gene

Oestrogens protect against cardiovascular disease in women;1 in men, oestrogens are produced by local tissue aromatisation of androgenic precursors from the testes and adrenal glands.2 Physiological concentrations of oestrogen have been reported to influence plasma high-density lipoprotein cholesterol in men.3 It is unclear whether oestrogens influence vascular function in men. We studied the peripheral vasculature of a 31-year-old man, with oestrogen resistance caused by a disruptive mutation in the oestrogen-receptor gene, as previously described.

Nonassociation of estrogen receptor genotypes with bone mineral density and estrogen responsiveness to hormone replacement therapy in Korean postmenopausal women.

Hormone replacement therapy (HRT) prevents bone loss in postmenopausal women, but some women are resistant to therapy. A recently reported case of severe estrogen resistance caused by a germline mutation at the estrogen receptor (ER) gene locus suggests the possibility that other variants of the ER gene could be responsible for resistance to HRT and could also be an answer to the heritable components of bone density. Three restriction fragment length polymorphisms (RFLPs) at the ER gene locus, represented as BstUI (or B variant), PvuII, and XbaI, and their relationship to bone mineral density (BMD) and estrogen responsiveness to HRT were examined in 248 healthy postmenopausal women, aged 41-68 yr (mean +/- SD, 52.0 +/- 4.6 yr) in Korea. The BstUI restriction site was not found in Korean women. The distribution of the PvuII and XbaI RFLPs was as follows: PP, 35 (14.1%); Pp, 136 (54.8%); pp, 77 (31.1%) and XX, 18 (7.3%); Xx, 72 (29.0%); and xx, 158 (63.7%), respectively (capital letters signify the absence of and lower case letters signify the presence of the restriction site of each RFLP). There was no significant relation between ER genotypes and z score values of lumbar spine BMD. Also, no significant genotypic differences were found in the change in lumbar spine BMD and those in biochemical markers before and after 1 yr of HRT. These data indicate no significant effects of ER genotypes on BMD and estrogen responsiveness after HRT.

Nonassociation of Estrogen Receptor Genotypes with Bone Mineral Density and Estrogen Responsiveness to Hormone Replacement Therapy in Korean Postmenopausal Women

Hormone replacement therapy (HRT) prevents bone loss in postmenopausal women, but some women are resistant to therapy. A recently reported case of severe estrogen resistance caused by a germ-line mutation at the estrogen receptor (ER) gene locus suggests the possibility that other variants of the ER gene could be responsible for resistance to HRT and could also be an answer to the heritable components of bone density. Three restriction fragment length polymorphisms (RFLPs) at the ER gene locus, represented as BstUI (or B variant), PvuII, and XbaI, and their relationship to bone mineral density (BMD) and estrogen responsiveness to HRT were examined in 248 healthy postmenopausal women, aged 41–68 yr (mean± sd, 52.0 ± 4.6 yr) in Korea. The BstUI restriction site was not found in Korean women. The distribution of the PvuII and XbaI RFLPs was as follows: PP, 35 (14.1%); Pp, 136 (54.8%); pp, 77 (31.1%); and XX, 18 (7.3%); Xx, 72 (29.0%); and xx, 158 (63.7%), respectively (capital letters signify the absence of and lower case letters signify the presence of the restriction site of each RFLP). There was no significant relation between ER genotypes and z score values of lumbar spine BMD. Also, no significant genotypic differences were found in the change in lumbar spine BMD and those in biochemical markers before and after 1 yr of HRT. These data indicate no significant effects of ER genotypes on BMD and estrogen responsiveness after HRT.

Sex steroids and the acquisition of bone mass.

Bone mineralization increases with age, height and weight through childhood, with a significant gain during pubertal development. Approximately 37% of bone mass is accumulated between pubertal stages 2-5. The factors driving the pubertal increase of bone mass is not fully known, but a crucial role is played by the sex steroids. Patients with central precocious puberty show an increased bone mineral density (BMD) for their chronological age, but appropriate for their bone age and pubertal stage. In girls and boys with hypogonadism, as well as in constitutional delay of puberty, the 'tempo' of puberty seems to be another important determinant of peak bone mass. Hypogonadal patients increase their BMD when treated in adulthood with sex steroid substitutive therapy, but normal adult BMD values are not reached. If these patients start sex steroid therapy at the appropriate age for the onset of pubertal development, however, a greater increase in BMD occurs. Both oestrogen and androgen receptors have been demonstrated in bone cells. Patients with oestrogen resistance or aromatase deficiency had a severely undermineralized skeleton. Reduced area and volume BMD are also present in patients with complete androgen resistant syndrome compared to normal. The pubertal increase of BMD may be the result of the coordinated activation of oestrogen and androgen receptors at the bone level in both sexes. In conclusion, the gonadal secretion of sex steroids, the tempo of puberty, and the normal function of both oestrogen and androgen receptors seem to be important factors in the acquisition of bone mass.

SKOV3 ovarian carcinoma cells have functional estrogen receptor but are growth-resistant to estrogen and antiestrogens

Estrogen receptor positive ovarian cancer is often refractile to antiestrogen therapy. Here we describe the SKOV3 human ovarian carcinoma cell line as an in vitro model for estrogen and antiestrogen resistant ovarian cancer. While SKOV3 cells expressed estrogen receptor (ER) mRNA and protein at a similar level as the estrogen responsive T47D breast carcinoma cell line, their growth was not responsive to estradiol (E 2) and was not inhibited by the antiestrogens OH-tamoxifen and ICI 164,384. The ER in SKOV3 cells was normal with respect to apparent K d for binding with E 2, E 2 regulation of a transiently transfected ERE driven reporter gene, and E 2 stimulation of expression of the early growth response genes c-myc and c-fos. However, the SKOV3 cells exhibited no expression of the progesterone receptor gene (PR) even after addition of E 2, and the protein products of the estrogen responsive genes HER-2/ neu and cathepsin D were expressed at constitutive levels that were not regulated by E 2. Therefore, estrogen resistance in these cells may be a result of constitutive expression and loss of E 2 regulation of selected growth regulatory gene products rather than a defect in estrogen activation of ER as a transcriptional regulator.